Single-Dose Electrospun Nanoparticles-in-Nanofibers Wound Dressings with Enhanced Epithelialization, Collagen Deposition, and Granulation Properties.

PubMed

Ali, Isra H; Khalil, Islam A; El-Sherbiny, Ibrahim M

2016-06-15

Phenytoin (Ph), an antiepileptic drug, was reported to exhibit high wound healing activity. However, its limited solubility, bioavailability, and inefficient distribution during topical administration limit its use. Therefore, this study aims to develop new single-dose electrospun nanoparticles-in-nanofibers (NPs-in-NFs) wound dressings that allow a well-controlled release of Ph. These NPs-in-NFs systems are based on enhanced chitosan (CS)/poly(ethylene oxide) (PEO) electrospun nanofibers (NFs) incorporating optimized Ph-loaded nanocarriers. First, a study was conducted to investigate Ph loading efficiency into polymeric nanocarriers of different types; pluronic nanomicelles and poly(lactic-co-glycolic) acids nanoparticles (PLGA NPs). The drug release profile from the nanocarriers was further optimized via lecithin coating. Second, different electrospinning parameters were manipulated to fabricate beads-free homogeneous NFs with optimized polymer ratios. Plain and Ph-loaded nanocarriers were characterized using Fourier transform infrared (FTIR), differential scanning calorimetry (DSC), thermogravimetric analysis (TGA), dynamic light scattering (DLS), and scanning electron microscopy (SEM). Both entrapment efficiency of Ph (EE%) and its release profile in phosphate buffer saline (PBS; pH 5.5), simulating the wound environment, were studied. Biodegradability, swelling, vapor permeability, and porosity of the developed Ph-loaded NPs-in-NFs wound dressings were investigated. Morphology of the NPs-in-NFs was also studied using SEM and confocal laser microscopy (CLSM). Besides, the release profiles of Ph from the optimized NPs-in-NFs were assessed. The newly developed wound dressings were evaluated in vitro for their cytotoxicity using human fibroblasts and in vivo using a wound healing mice model. Nanocarriers with particle size ranging from 100 to 180 nm were successfully prepared. All nanocarriers attained a high drug entrapment efficiency exceeding 94% and showed promising sustained release profiles compared to free Ph. Results also demonstrated that NFs incorporating the optimized lecithin-coated Ph-loaded PLGA NPs could be the most promising candidate for efficient wound healing. These NPs-in-NFs systems conferred a well-controlled and sustained release of Ph over 9 days. Moreover, they showed the best re-epithelization and healing quality during the in vivo study with minimal inflammatory and necrotic cells formation.

Stability studies on florfenicol using developed derivative spectrophotometric methods.

PubMed

Elimam, M M; Shantier, S W; Gadkariem, E A; Mohamed, M A; Osman, Z

2017-01-01

This study aims to investigate the stability of florfenicol using previously developed derivative spectrophotometric methods (D 1 and D 2 ). The studied stability-indicating pararmeters included alkali (NaOH, 1M), acid (HCl, 1M), pH changes (buffer pH 2.2-11), temperature (80°C and 100°C at pH 10) and light. A constructed pH profile for the drug degradation rate revealed a significant effect of pH on the drug stability between pH ranges 8 and 11. The obtained profile indicated first order dependence of K obs on [OH - ]. Arrhenius plot at pH 10 was found linear at temperatures 80°C and 100°C with estimated activation energy of 19.35kcal/mol. The calculated rate constant (K obs ), t ½ and t 90 at 25°C were found to be 1.8×10 -3 h, 385h and 58.3h, respectively. The photostability of florfenicol was also studied by exposing the drug solution to direct sunlight during mid-day time. The obtained results reflected the instability of florfenicol under the study conditions. Copyright © 2016 Académie Nationale de Pharmacie. Published by Elsevier Masson SAS. All rights reserved.

A Study to Draw a Normative Database of Laryngopharynx pH Profile in Chinese

PubMed Central

Feng, Guijian; Wang, Junyao; Zhang, Lihong; Liu, Yulan

2014-01-01

Background/Aims To draw a normative database of laryngopharynx pH profile in Chinese subjects. Methods Normal volunteers were recruited from “www.Ganji.com” and People’s hospital between May 2008 and December 2009. The Restech pH Probes were calibrated in pH 7 and pH 4 buffer solutions according to the manufacturer's instructions. Each volunteer was asked to wear the device for a 24-hour period and was encouraged to participate in normal daily activities. Results The healthy volunteers consisted of 20 males and 9 females with a median age of 23 years (interquartile range, 21 to 32 years). The 95th percentiles for % total times at pH < 4, pH < 4.5, pH < 5.0 and pH < 5.5 for the oropharynx pH catheter were 0.06%, 1.01%, 7.23% and 27.34%, respectively. The 95th percentile for number of reflux events within the 24-hour period at pH < 4, pH < 4.5, pH < 5.0 and pH < 5.5 were 2.0, 18.0, 107.5 and 284.5, respectively. Conclusions This is the first study to systematically assess the degree of reflux detected by the new pH probe in healthy asymptomatic Chinese volunteers and to report normative values in Chinese people. Using an oropharyngeal pH catheter to monitor laryngopharyngeal reflux indicated that in healthy Chinese, reflux should be considered normal if the percent time at pH less than 4.5 is no more than 1%. PMID:24948130

Researchers Identify Genomic Alterations Associated with Drug-Targetable Kinase Activation in Ph-like Acute Lymphoblastic Leukemia | Office of Cancer Genomics

Cancer.gov

Acute lymphoblastic leukemia (ALL) is the most prevalent cancer among children and young adults, and standard treatments within this population generally result in favorable outcomes. By contrast, one particular subtype of this disease, Philadelphia chromosome-like ALL (Ph-like ALL), is associated with inferior outcomes. Ph-like ALL exhibits a gene expression profile similar to chromosome 9:22 translocation positive ALL, yet it lacks the characteristic BCR-ABL fusion protein.

Synthesis, characterization, and assessment of cytotoxic, antiproliferative, and antiangiogenic effects of a novel procainamide hydrochloride-poly(maleic anhydride-co-styrene) conjugate.

PubMed

Karakus, Gulderen; Akin Polat, Zubeyde; Sahin Yaglıoglu, Ayse; Karahan, Mesut; Yenidunya, Ali Fazil

2013-01-01

Poly(maleic anhydride-co-styrene) (MAST) was synthesized by a free-radical polymerization reaction. A bioactive molecule, procainamide hydrochloride (PH), was then conjugated to MAST. The conjugation product was named as MAST/PH. Structural characterization of MAST and MAST/PH was carried out by Fourier Transform Infrared and Nuclear Magnetic Resonance spectroscopy. Their molecular weights were determined by size-exclusion chromatography. A mechanism was then suggested for the conjugation reaction. The results of the cytotoxicity assay, employing a mouse fibroblast cell line (L929), indicated that MAST/PH had no cytotoxicity at concentrations [Formula: see text] 62 μg mL(-1) (p > 0.05). Antiproliferative activities of MAST/PH and PH were determined by the BrdU cell proliferation ELISA assay, using C6 and HeLa cell lines. In the experiment, two anticancer chemotherapy drugs, cisplatin and 5-fluorouracil, were included as positive control. Antiproliferative activity results demonstrated that MAST/PH yielded the highest suppression profile (approximately 42%) at 20 μg/ml, while free PH exerted the same activity at 100 μg/ml. Interestingly, both MAST/PH and PH suppressed the proliferation of only one of the cell lines, C6 cells. Both cisplatin and 5-fluorouracil yielded approximately 60% antiproliferative activity on C6 cells at 20 and 100 μg/ml concentrations. Antiangiogenic capacity of both MAST and MAST/PH was also investigated by using the chicken chorioallantoic membrane assay. Results obtained indicated that while MAST/PH could be included into the category of good antiangiogenic substances, the activity score of MAST was within the weak category.

Transcriptome Profiling of Shewanella oneidensis Gene Expression following Exposure to Acidic and Alkaline pH†

PubMed Central

Leaphart, Adam B.; Thompson, Dorothea K.; Huang, Katherine; Alm, Eric; Wan, Xiu-Feng; Arkin, Adam; Brown, Steven D.; Wu, Liyou; Yan, Tingfen; Liu, Xueduan; Wickham, Gene S.; Zhou, Jizhong

2006-01-01

The molecular response of Shewanella oneidensis MR-1 to variations in extracellular pH was investigated based on genomewide gene expression profiling. Microarray analysis revealed that cells elicited both general and specific transcriptome responses when challenged with environmental acid (pH 4) or base (pH 10) conditions over a 60-min period. Global responses included the differential expression of genes functionally linked to amino acid metabolism, transcriptional regulation and signal transduction, transport, cell membrane structure, and oxidative stress protection. Response to acid stress included the elevated expression of genes encoding glycogen biosynthetic enzymes, phosphate transporters, and the RNA polymerase sigma-38 factor (rpoS), whereas the molecular response to alkaline pH was characterized by upregulation of nhaA and nhaR, which are predicted to encode an Na+/H+ antiporter and transcriptional activator, respectively, as well as sulfate transport and sulfur metabolism genes. Collectively, these results suggest that S. oneidensis modulates multiple transporters, cell envelope components, and pathways of amino acid consumption and central intermediary metabolism as part of its transcriptome response to changing external pH conditions. PMID:16452448

Modelling and optimization of environmental conditions for kefiran production by Lactobacillus kefiranofaciens.

PubMed

Cheirsilp, B; Shimizu, H; Shioya, S

2001-12-01

A mathematical model for kefiran production by Lactobacillus kefiranofaciens was established, in which the effects of pH, substrate and product on cell growth, exopolysaccharide formation and substrate assimilation were considered. The model gave a good representation both of the formation of exopolysaccharides (which are not only attached to cells but also released into the medium) and of the time courses of the production of galactose and glucose in the medium (which are produced and consumed by the cells). Since pH and both lactose and lactic acid concentrations differently affected production and growth activity, the model included the effects of pH and the concentrations of lactose and lactic acid. Based on the mathematical model, an optimal pH profile for the maximum production of kefiran in batch culture was obtained. In this study, a simplified optimization method was developed, in which the optimal pH profile was determined at a particular final fermentation time. This was based on the principle that, at a certain time, switching from the maximum specific growth rate to the critical one (which yields the maximum specific production rate) results in maximum production. Maximum kefiran production was obtained, which was 20% higher than that obtained in the constant-pH control fermentation. A genetic algorithm (GA) was also applied to obtain the optimal pH profile; and it was found that practically the same solution was obtained using the GA.

Proton modulation of recombinant GABAA receptors: influence of GABA concentration and the β subunit TM2–TM3 domain

PubMed Central

Wilkins, Megan E; Hosie, Alastair M; Smart, Trevor G

2005-01-01

Regulation of GABAA receptors by extracellular pH exhibits a dependence on the receptor subunit composition. To date, the molecular mechanism responsible for the modulation of GABAA receptors at alkaline pH has remained elusive. We report here that the GABA-activated current can be potentiated at pH 8.4 for both αβ and αβγ subunit-containing receptors, but only at GABA concentrations below the EC40. Site-specific mutagenesis revealed that a single lysine residue, K279 in the β subunit TM2–TM3 linker, was critically important for alkaline pH to modulate the function of both α1β2 and α1β2γ2 receptors. The ability of low concentrations of GABA to reveal different pH titration profiles for GABAA receptors was also examined at acidic pH. At pH 6.4, GABA activation of αβγ receptors was enhanced at low GABA concentrations. This effect was ablated by the mutation H267A in the β subunit. Decreasing the pH further to 5.4 inhibited GABA responses via αβγ receptors, whereas those responses recorded from αβ receptors were potentiated. Inserting homologous β subunit residues into the γ2 subunit to recreate, in αβγ receptors, the proton modulatory profile of αβ receptors, established that in the presence of β2H267, the mutation γ2T294K was necessary to potentiate the GABA response at pH 5.4. This residue, T294, is homologous to K279 in the β subunit and suggests that a lysine at this position is an important residue for mediating the allosteric effects of both acidic and alkaline pH changes, rather than forming a direct site for protonation within the GABAA receptor. PMID:15946973

Computer Science | Classification | College of Engineering & Applied

Science.gov Websites

EMS 1011 profile photo Adrian Dumitrescu, Ph.D.ProfessorComputer Science(414) 229-4265Eng & Math @uwm.eduEng & Math Sciences 919 profile photo Hossein Hosseini, Ph.D.ProfessorComputer Science(414) 229 -5184hosseini@uwm.eduEng & Math Sciences 1091 profile photo Amol Mali, Ph.D.Associate ProfessorComputer

Substrate and pH-Dependent Kinetic Profile of 3-Mercaptopropionate Dioxygenase from Pseudomonas aeruginosa.

PubMed

Fellner, Matthias; Aloi, Sekotilani; Tchesnokov, Egor P; Wilbanks, Sigurd M; Jameson, Guy N L

2016-03-08

Thiol dioxygenases catalyze the synthesis of sulfinic acids in a range of organisms from bacteria to mammals. A thiol dioxygenase from the bacterium Pseudomonas aeruginosa oxidizes both 3-mercaptopropionic acid and cysteine, with a ∼70 fold preference for 3-mercaptopropionic acid over all pHs. This substrate reactivity is widened compared to other thiol dioxygenases and was exploited in this investigation of the residues important for activity. A simple model incorporating two protonation events was used to fit profiles of the Michaelis-Menten parameters determined at different pH values for both substrates. The pKs determined using plots of k(cat)/Km differ at low pH, but not in a way easily attributable to protonation of the substrate alone and share a common value at higher pH. Plots of k(cat) versus pH are also quite different at low pH showing the monoprotonated ES complexes with 3-mercaptopropionic acid and cysteine have different pKs. At higher pH, k(cat) decreases sigmoidally with a similar pK regardless of substrate. Loss of reactivity at high pH is attributed to deprotonation of tyrosine 159 and its influence on dioxygen binding. A mechanism is proposed by which deprotonation of tyrosine 159 both blocks oxygen binding and concomitantly promotes cystine formation. Finally, the role of tyrosine 159 was further probed by production of a G95C variant that is able to form a cysteine-tyrosine crosslink homologous to that found in mammalian cysteine dioxygenases. Activity of this variant is severely impaired. Crystallography shows that when un-crosslinked, the cysteine thiol excludes tyrosine 159 from its native position, while kinetic analysis shows that the thioether bond impairs reactivity of the crosslinked form.

Mechanistic characterization of the HDV genomic ribozyme: a mutant of the C41 motif provides insight into the positioning and thermodynamic linkage of metal ions and protons.

PubMed

Nakano, Shu-ichi; Bevilacqua, Philip C

2007-03-20

Binding of two Mg2+ and two H+ ions influences the self-cleavage activity of the genomic HDV ribozyme. The positioning of these four ligands and their thermodynamic linkage are not fully resolved. Protonated C41 engages in a base triple, whereas protonated C75 has been implicated as an acid-base catalyst in bond cleavage. Prior studies led to the identification of one structural inner-sphere ion and one catalytic outer-sphere ion. In the present study, the contributions of the C41 base triple to the metal ion- and pH-dependence of the reaction are examined. Experiments were conducted on a CG to UA double mutant (DM), which changes the base triple to one involving an unprotonated C41. Below pH 6, the DM has a steeper dependence on pH than the wild-type (WT), consistent with a single protonation misfolding the core; this conclusion is also supported by thermal denaturation studies. Between pH 6 and 8, the WT and DM display nearly identical catalytic metal ion and H+ binding profiles. In contrast, over the same pH range, the WT and DM have distinct structural ion binding profiles; for the WT, binding is favored at lower pH, whereas the DM shows no pH dependence. These data localize the structural ion to the vicinity of the C41 motif. An overall model is presented that accommodates binding affinity, coupling, and positioning of the two metal ions and the two protons within the ribozyme. The data suggest that a protonated base triple allows the WT ribozyme to maintain appreciable activity at acidic pH, which could play an important role in the life cycle of the virus.

Hypoglycemic and Hypolipidemic Effects of the Cracked-Cap Medicinal Mushroom Phellinus rimosus (Higher Basidiomycetes) in Streptozotocin-Induced Diabetic Rats.

PubMed

Rony, Kuttikkadan A; Ajith, Thekkuttuparambil A; Janardhanan, Kainoor K

2015-01-01

Phellinus rimosus is a parasitic host specific polypore mushroom with profound antioxidant, antihepatotoxic, anti-inflammatory, antitumor, and antimutagenic activities. This study investigated the hypoglycemic and hypolipidemic activities of the wood-inhabiting polypore mushroom Ph. Rimosus in streptozotocin (STZ)-induced diabetic rats. Diabetes was induced by single intraperitoneal injection of STZ (45 mg/kg) to Wistar rats. The effects of 30 days treatment with Ph. Rimosus (50 and 250 mg/ kg) and glibenclamide (0.65 mg/kg) on blood glucose level, serum insulin, serum lipid profile, liver glycogen, liver function enzymes, and non-enzymic and enzymic antioxidants activities in pancreas, liver, and kidney were evaluated in STZ-induced diabetic rats. Oral administration of Ph. Rimosus extract exhibited a significant reduction in blood glucose, triacylglycerol, total cholesterol, LDL-cholesterol, and liver function enzymes, and increased serum insulin, liver glycogen, and HDL-cholesterol levels in STZ-induced diabetic rats. Furthermore, Ph. Rimosus treatment increased antioxidant status in pancreas, liver, and kidney tissues with concomitant decreases in levels of thiobarbituric acid- reactive substances. Results of this study indicated that Ph. Rimosus possessed significant hypoglycemic and hypolipidemic activities and this effect may be related to its insulinogenic and antioxidant effect.

Development of an ultrahigh-temperature process for the enzymatic hydrolysis of lactose. IV. Immobilization of two thermostable beta-glycosidases and optimization of a packed-bed reactor for lactose conversion.

PubMed

Petzelbauer, Inge; Kuhn, Bernhard; Splechtna, Barbara; Kulbe, Klaus D; Nidetzky, Bernd

2002-03-20

Recombinant hyperthermostable beta-glycosidases from the archaea Sulfolobus solfataricus (Ss beta Gly) and Pyrococcus furiosus (CelB) were covalently attached onto the insoluble carriers chitosan, controlled pore glass (CPG), and Eupergit C. For each enzyme/carrier pair, the protein-binding capacity, the immobilization yield, the pH profiles for activity and stability, the activity/temperature profile, and the kinetic constants for lactose hydrolysis at 70 degrees C were determined. Eupergit C was best among the carriers in regard to retention of native-like activity and stability of Ss beta Gly and CelB over the pH range 3.0-7.5. Its protein binding capacity of approximately 0.003 (on a mass basis) was one-third times that of CPG, while immobilization yields were typically 80% in each case. Activation energies for lactose conversion by the immobilized enzymes at pH 5.5 were in the range 50-60 kJ/mol. This is compared to values of approximately 75 kJ/mol for the free enzymes. Immobilization expands the useful pH range for CelB and Ss beta Gly by approximately 1.5 pH units toward pH 3.5 and pH 4.5, respectively. A packed-bed enzyme reactor was developed for the continuous conversion of lactose in different media, including whey and milk, and operated over extended reaction times of up to 14 days. The productivities of the Eupergit C-immobilized enzyme reactor were determined at dilution rates between 1 and 12 h(-1), and using 45 and 170 g/L initial lactose. Results of kinetic modeling for the same reactor, assuming plug flow and steady state, suggest the presence of mass-transfer limitation of the reaction rate under the conditions used. Formation of galacto-oligosaccharides in the continuous packed-bed reactor and in the batch reactor using free enzyme was closely similar in regard to yield and individual saccharide components produced. Copyright 2002 John Wiley & Sons, Inc. Biotechnol Bioeng 77: 619-631, 2002; DOI 10.1002/bit.10110

Comparative Study on Different Expression Hosts for Alkaline Phytase Engineered in Escherichia coli.

PubMed

Chen, Weiwei; Yu, Hongwei; Ye, Lidan

2016-07-01

The application of alkaline phytase as a feed additive is restricted by the poor specific activity. Escherichia coli is a frequently used host for directed evolution of proteins including alkaline phytase towards improved activity. However, it is not suitable for production of food-grade products due to potential pathogenicity. To combine the advantages of different expression systems, mutants of the alkaline phytase originated from Bacillus subtilis 168 (phy168) were first generated via directed evolution in E. coli and then transformed to food-grade hosts B. subtilis and Pichia pastoris for secretory expression. In order to investigate the suitability of different expression systems, the phy168 mutants expressed in different hosts were characterized and compared in terms of specific activity, pH profile, pH stability, temperature profile, and thermostability. The specific activity of B. subtilis-expressed D24G/K70R/K111E/N121S mutant at pH 7.0 and 60 °C was 30.4 U/mg, obviously higher than those in P. pastoris (22.7 U/mg) and E. coli (19.7 U/mg). Moreover, after 10 min incubation at 80 °C, the B. subtilis-expressed D24G/K70R/K111E/N121S retained about 70 % of the activity at pH 7.0 and 37 °C, whereas the values were only about 25 and 50 % when expressed in P. pastoris and E. coli, respectively. These results suggested B. subtilis as an appropriate host for expression of phy168 mutants and that the strategy of creating mutants in one host and expressing them in another might be a new solution to industrial production of proteins with desired properties.

Effect of processing conditions on the organosulfides of shallot (Allium cepa L. Aggregatum group).

PubMed

Tocmo, Restituto; Lin, Yi; Huang, Dejian

2014-06-11

There is a growing account of the health benefits of H2S as an endogenous cell-signaling molecule. H2S from organic polysulfides, in particular, is increasingly gaining attention for their beneficial effects to cardiovascular health. Here, we studied shallot as a potential dietary source of organic polysulfides and examined the effects of processing conditions on its polysulfide profiles. Boiling, autoclaving, and freeze-drying were tested on whole and crushed shallot bulbs, analyzing their effect on the yield of organosulfides. Seventeen organosulfides, including disulfides, trisulfides, and cyclic polysulfides, were identified. Significant differences in the quantitative and qualitative profiles of organosulfides in the hydrodistilled and solvent extracted oils were observed. Freeze-drying retained the majority of the organosulfides, but the whole-autoclaved and whole-boiled shallots lost more than 95% of their organic polysulfides. Crushed-boiled and crushed-autoclaved shallot lost 76-80% of their organosulfides, likely due to the thermal sensitivity of these compounds. The organosulfide profiles are sensitive to the pH values of the processing media. In general, disulfides increased at basic pH (pH 9.0) while trisulfides and cyclic organosulfides are much higher at the acidic to neutral pH values (pH 3.0-5.0). Our results provide important information on the effects of processing conditions that are relevant for optimizing extraction of organosulfides from shallot for further studies evaluating their H2S-releasing activity.

pH Dependence of a Mammalian Polyamine Oxidase: Insights into Substrate Specificity and the Role of Lysine 315†

PubMed Central

Pozzi, Michelle Henderson; Gawandi, Vijay; Fitzpatrick, Paul F.

2009-01-01

Mammalian polyamine oxidases (PAO) catalyze the oxidation of N1-acetylspermine and N1-acetylspermidine to produce N-acetyl-3-aminopropanaldehyde and spermidine or putrescine. Structurally, PAO is a member of the monoamine oxidase family of flavoproteins. The effects of pH on kinetic parameters of mouse PAO have been determined to provide insight into the protonation state of the polyamine required for catalysis and the roles of ionizable residues in the active site in amine oxidation. For N1-acetylspermine, N1-acetylspermidine, and spermine, the kcat/Kamine-pH profiles are bell-shaped. In each case the profile agrees with that expected if the productive form of the substrate has a single positively charged nitrogen. The pKi-pH profiles for a series of polyamine analogs are most consistent with the nitrogen at the site of oxidation being neutral and one other nitrogen being positively charged in the reactive form of the substrate. With N1-acetylspermine as substrate, the value of kred, the limiting rate constant for flavin reduction, is pH dependent, decreasing below a pKa value of 7.3, again consistent with the requirement for an uncharged nitrogen for substrate oxidation. Lys315 in PAO corresponds to a conserved active site residue found throughout the monoamine oxidase family. Mutation of Lys315 to methionine has no effect on the kcat/Kamine profile for spermine, the kred value with N1-acetylspermine is only 1.8-fold lower in the mutant protein, and the pKa in the kred-pH profile with N1-acetylspermine shifts to 7.8. These results rule out Lys315 as a source of a pKa in the kcat/Kamine or kcat/kred profiles. They also establish that this residue does not play a critical role in amine oxidation by PAO. PMID:19199575

Smart tetrazole-based antibacterial nanoparticles as multifunctional drug carriers for cancer combination therapy.

PubMed

Zakerzadeh, Elham; Salehi, Roya; Mahkam, Mehrdad

2017-12-01

Due to multidrug resistance of cancer tissues and immune-suppression of cancerous patients during chemotherapy in one hand and the use of tetrazole derivatives in medicine because of its anticancer, antifungal, and antiviral properties, on the other, we were encouraged to design novel smart antibacterial nanocomposites-based polymer of tetrazole as dual anticancer drug delivery systems. The structures of nanocomposites characterized by FTIR, 1 H NMR, FESEM-EDX, and TGA analyzes and antibacterial activity of smart carriers were evaluated by determination of minimum inhibitory concentration (MIC) values against some bacteria and fungi. Then, the pH-responsive manner of both nanocomposites was proved by checking their release profiles at pH of the physiological environment (pH 7.4) and pH of tumor tissues (mildly acidic). Finally, the potential antitumoral activity of these nanocomposite systems against MCF7 cell lines was evaluated by MTT assay and cell cycle studies. The results demonstrated that the novel developed nanocomposites not only meet our expectations about simultaneous release of two anticancer drugs according to the predicted profile but also showed antibacterial and anticancer properties in vitro experimental. Moreover, it was proved that these carriers have tremendous potential in multifunctional drug delivery in cancer therapy.

Effect of prolonged refrigeration on the lipid profile, lipase activity, and oxidative status of human milk.

PubMed

Bertino, Enrico; Giribaldi, Marzia; Baro, Cristina; Giancotti, Valeria; Pazzi, Marco; Peila, Chiara; Tonetto, Paola; Arslanoglu, Sertac; Moro, Guido E; Cavallarin, Laura; Gastaldi, Daniela

2013-04-01

The study was aimed at evaluating the effect of prolonged refrigeration of fresh human milk (HM) on its fatty acid profile, free fatty acid content, lipase activities, and oxidative status. HM from mothers of preterm newborns was collected, pooled, and placed in the neonatal intensive care unit (NICU) refrigerator. Pooled milk was aliquoted and analyzed within 3 hours of collection, and after 24, 48, 72, and 96 hours of storage. The milk samples were analyzed for pH, total and free fatty acid profile, lipase activity at room temperature and at 4°C, lipase activity at room temperature in presence of sodium cholate (bile salt-dependent lipase), total antioxidant capacity, thiobarbituric acid reactive species, malondialdehyde, and conjugated diene concentration. The experiment was replicated in 3 independent trials. Prolonged refrigeration did not affect the fatty acid composition of breast milk, and preserved both its overall oxidative status and the activity of HM lipolytic enzymes. In particular, bile salt-dependent lipase activity, long-chain polyunsaturated fatty acids, and medium-chain saturated fatty acid concentrations were unaffected for up to 96 hours of refrigerated storage. Prolonged refrigeration of fresh HM for 96 hours maintained its overall lipid composition. The limited lipolysis during storage should be ascribed to the activity of lipoprotein lipase, responsible for the decrease in pH. Our study demonstrates that infants who receive expressed milk stored for up to 96 hours receive essentially the same supply of fatty acids and active lipases as do infants fed directly at the breast.

Enhancement of Biohydrogen Production via pH Variation using Molasses as Feedstock in an Attached Growth System

NASA Astrophysics Data System (ADS)

Che Zuhar, C. N. S.; Lutpi, N. A.; Idris, N.; Wong, Y. S.; Tengku Izhar, T. N.

2018-03-01

In this study, mesophilic biohydrogen production by a mixed culture, obtained from a continuous anaerobic reactor treating molasses effluent from sugarcane bagasse, was improved by using granular activated carbon (GAC) as the carrier material. A series of batch fermentation were performed at 37°C by feeding the anaerobic sludge bacteria with molasses to determine the effect of initial pH in the range of 5.5 to 7.5, and the effect of repeated batch cultivation on biohydrogen production. The enrichment of granular activated carbon (GAC) immobilised cells from the repeated batch cultivation were used as immobilised seed culture to obtain the optimal initial pH. The cumulative hydrogen production results from the optimal pH were fitted into modified Gompertz equation in order to obtained the batch profile of biohydrogen production. The optimal hydrogen production was obtained at an initial pH of 5.5 with the maximum hydrogen production (Hm) was found to be 84.14 ml, and maximum hydrogen production rate (Rm) was 3.63 mL/h with hydrogen concentration of 759 ppm. The results showed that the granular activated carbon was successfully enhanced the biohydrogen production by stabilizing the pH and therefore could be used as a carrier material for fermentative hydrogen production using industrial effluent.

An interfacial and comparative in vitro study of gastrointestinal lipases and Yarrowia lipolytica LIP2 lipase, a candidate for enzyme replacement therapy.

PubMed

Bénarouche, Anaïs; Point, Vanessa; Carrière, Frédéric; Cavalier, Jean-François

2014-07-01

Lipolytic activities of Yarrowia lipolytica LIP2 lipase (YLLIP2), human pancreatic (HPL) and dog gastric (DGL) lipases were first compared using lecithin-stabilized triacylglycerol (TAG) emulsions (Intralipid) at various pH and bile salt concentrations. Like DGL, YLLIP2 was able to hydrolyze TAG droplets covered by a lecithin monolayer, while HPL was not directly active on that substrate. These results were in good agreement with the respective kinetics of adsorption on phosphatidylcholine (PC) monomolecular films of the same three lipases, YLLIP2 being the most tensioactive lipase. YLLIP2 adsorption onto a PC monolayer spread at the air/water interface was influenced by pH-dependent changes in the enzyme/lipid interfacial association constant (KAds) which was optimum at pH 6.0 on long-chain egg PC monolayer, and at pH 5.0 on medium chain dilauroylphosphatidylcholine film. Using substrate monolayers (1,2-dicaprin, trioctanoin), YLLIP2 displayed the highest lipolytic activities on both substrates in the 25-35 mN m(-1) surface pressure range. YLLIP2 was active in a large pH range and displayed a pH-dependent activity profile combining DGL and HPL features at pH values found in the stomach (pH 3-5) and in the intestine (pH 6-7), respectively. The apparent maximum activity of YLLIP2 was observed at acidic pH 4-6 and was therefore well correlated with an efficient interfacial binding at these pH levels, whatever the type of interfaces (Intralipid emulsions, substrate or PC monolayers). All these findings support the use of YLLIP2 in enzyme replacement therapy for the treatment of pancreatic exocrine insufficiency, a pathological situation in which an acidification of intestinal contents occurs. Copyright © 2014 Elsevier Masson SAS. All rights reserved.

Anti-inflammation effects of Sophora flavescens nanoparticles.

PubMed

Han, Chun-Chao; Wang, Yingzi

2012-08-01

The roots of Sophora flavescens was reported to possess many pharmacological activities including anti-inflammatory, antiashmatic, antithelmintic, free radical scavenging and antimicrobial activities. However, the low saturated solubility and dissolution velocity of S. flavescens lead to poor bioavailability. The S. flavescens nanoparticles (SFNP) were prepared by a combination of ultrasound and hydrolysis developed by the authors. The drug dissolution profiles of SFNP in both pH 6.8 and pH 2 media showed complete dissolution within 30 min. The seropharmacology study showed that oral S. flavescens absorption in the SFNP was significantly increased. Anti-inflammation assay revealed the therapeutic efficiency of S. flavescens significantly enhanced upon nanoparticle formation.

Structural insights into the recovery of aldolase activity in N-acetylneuraminic acid lyase by replacement of the catalytically active lysine with γ-thialysine by using a chemical mutagenesis strategy.

PubMed

Timms, Nicole; Windle, Claire L; Polyakova, Anna; Ault, James R; Trinh, Chi H; Pearson, Arwen R; Nelson, Adam; Berry, Alan

2013-03-04

Chemical modification has been used to introduce the unnatural amino acid γ-thialysine in place of the catalytically important Lys165 in the enzyme N-acetylneuraminic acid lyase (NAL). The Staphylococcus aureus nanA gene, encoding NAL, was cloned and expressed in E. coli. The protein, purified in high yield, has all the properties expected of a class I NAL. The S. aureus NAL which contains no natural cysteine residues was subjected to site-directed mutagenesis to introduce a cysteine in place of Lys165 in the enzyme active site. Subsequently chemical mutagenesis completely converted the cysteine into γ-thialysine through dehydroalanine (Dha) as demonstrated by ESI-MS. Initial kinetic characterisation showed that the protein containing γ-thialysine regained 17 % of the wild-type activity. To understand the reason for this lower activity, we solved X-ray crystal structures of the wild-type S. aureus NAL, both in the absence of, and in complex with, pyruvate. We also report the structures of the K165C variant, and the K165-γ-thialysine enzyme in the presence, or absence, of pyruvate. These structures reveal that γ-thialysine in NAL is an excellent structural mimic of lysine. Measurement of the pH-activity profile of the thialysine modified enzyme revealed that its pH optimum is shifted from 7.4 to 6.8. At its optimum pH, the thialysine-containing enzyme showed almost 30 % of the activity of the wild-type enzyme at its pH optimum. The lowered activity and altered pH profile of the unnatural amino acid-containing enzyme can be rationalised by imbalances of the ionisation states of residues within the active site when the pK(a) of the residue at position 165 is perturbed by replacement with γ-thialysine. The results reveal the utility of chemical mutagenesis for the modification of enzyme active sites and the exquisite sensitivity of catalysis to the local structural and electrostatic environment in NAL. Copyright © 2013 WILEY-VCH Verlag GmbH & Co. KGaA, Weinheim.

A broad pH range indicator-based spectrophotometric assay for true lipases using tributyrin and tricaprylin[S

PubMed Central

Camacho-Ruiz, María de los Angeles; Mateos-Díaz, Juan Carlos; Carrière, Frédéric; Rodriguez, Jorge A.

2015-01-01

A continuous assay is proposed for the screening of acidic, neutral, or alkaline lipases using microtiter plates, emulsified short- and medium-chain TGs, and a pH indicator. The lipase activity measurement is based on the decrease of the pH indicator optical density due to protonation which is caused by the release of FFAs during the hydrolysis of TGs and thus acidification. Purified lipases with distinct pH optima and an esterase were used to validate the method. The rate of lipolysis was found to be linear with time and proportional to the amount of enzyme added in each case. Specific activities measured with this microplate assay method were lower than those obtained by the pH-stat technique. Nevertheless, the pH-dependent profiles of enzymatic activity were similar with both assays. In addition, the substrate preference of each enzyme tested was not modified and this allowed discriminating lipase and esterase activities using tributyrin (low water solubility) and tricaprylin (not water soluble) as substrates. This continuous lipase assay is compatible with a high sample throughput and can be applied for the screening of lipases and lipase inhibitors from biological samples. PMID:25748441

Potentiometric/turbidometric titration of antiperspirant actives.

PubMed

Johnston, Clifford T; Hem, Stanley L; Guenin, Eric; Mattai, Jairajh; Afflito, John

2003-01-01

A titration procedure that simultaneously monitors the pH and turbidity of an antiperspirant solution during neutralization with sodium hydroxide was developed to characterize antiperspirant actives. Aluminum chloride, aluminum chlorohydrate (ACH), and aluminum zirconium glycine complex (AZG) gave distinctive pH/turbidity profiles. The activated forms of aluminum chlorohydrate (ACH') and aluminum zirconium glycine complex (AZG') produced more turbidity than the non-activated forms. On an equimolar basis, AZG' produced more turbidity than any of the antiperspirant actives tested.

Gene encoding the group B streptococcal protein R4, its presence in clinical reference laboratory isolates & R4 protein pepsin sensitivity.

PubMed

Smith, B L; Flores, A; Dechaine, J; Krepela, J; Bergdall, A; Ferrieri, P

2004-05-01

R proteins were first identified by Lancefield in group B Streptococcus (GBS) as resistant to trypsin at pH8 and sensitive to pepsin at pH2. The R4 protein found predominantly in type III and some type II and V invasive isolates conforms to these criteria. The Rib protein, although structurally and epidemiologically similar to R4, was reported as resistant to both proteases. We report here the gene encoding the R4 protein from a type III group B streptococcal isolate (76-043) well characterized in our laboratory. Trypsin extracted GBS proteins were assayed for protease sensitivities by double-diffusion Ouchterlony using varying conditions for the enzyme pepsin. Standard haemoglobin assay was used to examine pepsin enzymatic activity. Thirty clinical isolates of varying protein profiles identified by double-diffusion from our reference strain laboratory were screened by PCR and Southern technique. SDS-PAGE gel purified R4 amino acid sequences were determined and used to design oligonucleotide primers for screening a 76-043 genomic library. R4 was sensitive to pepsin at pH2 but appeared resistant at pH4, the reported pH used for Rib. By standard haemoglobin assay and trypsin extract studies of R4 protein, pepsin was shown to be active at pH2, yet easily inactivated; assays of GBS surface proteins are critical at pH2. Of the amino acids initially sequenced from R4, 88 per cent (61/69) showed identity to Rib; the r4 nucleotide sequence was identical to that of rib. All isolates with strong positive protein reactions for R4 were positive in both PCR and Southern technique, whereas isolates expressing alpha, beta, R1/R4, and R5 (BPS) protein profiles were not. Sequenced PCR products aligned with identity to the R4 and Rib nucleotide sequences and confirmed the identity of these proteins and their molecular sequences.

Noninvasive Ph-telemetric Measurement of Gastrointestinal Function

NASA Technical Reports Server (NTRS)

Tietze, Karen J.

1991-01-01

The purpose of this study was to gain experience with and validate the Heidelberg pH-telemetric methodology in order to determine if the pH-telemetric methodology would be a useful noninvasive measure of gastrointestinal transit time for future ground-based and in-flight drug evaluation studies. The Heidelberg pH metering system is a noninvasive, nonradioactive telemetric system that, following oral ingestion, continuously measures intraluminal pH of the stomach, duodenum, small bowel, ileocecal junction, and large bowel. Gastrointestinal motility profiles were obtained in normal volunteers using the lactulose breath-hydrogen and Heidelberg pH metering techniques. All profiles were obtained in the morning after an overnight fast. Heidelberg pH profiles were obtained in the fasting and fed states; lactulose breath-hydrogen profiles were obtained after a standard breakfast. Mouth-to-cecum transit time was measured as the interval from administration of lactulose (30 ml; 20 g) to a sustained increase in breath-hydrogen of 10 ppm or more. Gastric emptying time was measured as the interval from the administration of the Heidelberg capsule to a sustained increase in pH of three units or more.

Studies on paclitaxel-loaded glyceryl monostearate nanoparticles.

PubMed

Shenoy, Vikram Subraya; Rajyaguru, Tushar Himmatlal; Gude, Rajiv Phondu; Murthy, Rayasa S Ramchandra

2009-09-01

Solid lipid nanoparticles (SLNs) of Paclitaxel were prepared by modified Hot homogenization method using Glyceryl monostearate (GMS). The SLNs were characterized for its physicochemical characteristics such as mean particle size, percentage entrapment efficiency and zeta potential, which were found to be 226 nm, 92.43% and -29.4 mV, respectively. The Transmission Electron Microscopy (TEM) studies showed that prepared SLNs were of spherical shape. The drug retarding efficiency of the lipid (GMS) was better in pH 7.4 compared to pH 3.5. The release profile showed a tendency to follow Higuchi diffusion pattern at pH 7.4 and Peppas-Korsenmeyer model at pH 3.5. Chemosensitivity assay carried out using B16F10 cell lines showed that anti-proliferative activity of Paclitaxel was not hindered due to encapsulation.

Ca2+-associated triphasic pH changes in mitochondria during brown adipocyte activation.

PubMed

Hou, Yanyan; Kitaguchi, Tetsuya; Kriszt, Rókus; Tseng, Yu-Hua; Raghunath, Michael; Suzuki, Madoka

2017-08-01

Brown adipocytes (BAs) are endowed with a high metabolic capacity for energy expenditure due to their high mitochondria content. While mitochondrial pH is dynamically regulated in response to stimulation and, in return, affects various metabolic processes, how mitochondrial pH is regulated during adrenergic stimulation-induced thermogenesis is unknown. We aimed to reveal the spatial and temporal dynamics of mitochondrial pH in stimulated BAs and the mechanisms behind the dynamic pH changes. A mitochondrial targeted pH-sensitive protein, mito-pHluorin, was constructed and transfected to BAs. Transfected BAs were stimulated by an adrenergic agonist, isoproterenol. The pH changes in mitochondria were characterized by dual-color imaging with indicators that monitor mitochondrial membrane potential and heat production. The mechanisms of pH changes were studied by examining the involvement of electron transport chain (ETC) activity and Ca 2+ profiles in mitochondria and the intracellular Ca 2+ store, the endoplasmic reticulum (ER). A triphasic mitochondrial pH change in BAs upon adrenergic stimulation was revealed. In comparison to a thermosensitive dye, we reveal that phases 1 and 2 of the pH increase precede thermogenesis, while phase 3, characterized by a pH decrease, occurs during thermogenesis. The mechanism of pH increase is partially related to ETC. In addition, the pH increase occurs concurrently with an increase in mitochondrial Ca 2+ . This Ca 2+ increase is contributed to by an influx from the ER, and it is further involved in mitochondrial pH regulation. We demonstrate that an increase in mitochondrial pH is implicated as an early event in adrenergically stimulated BAs. We further suggest that this pH increase may play a role in the potentiation of thermogenesis.

The acid tolerance response and pH adaptation of Enterococcus faecalis in extract of lime Citrus aurantiifolia from Aceh Indonesia.

PubMed

Mubarak, Zaki; Soraya, Cut

2018-01-01

Background:  The objective of the present study was to evaluate the acid tolerance response and pH adaptation when Enterococcus faecalis interacted with extract of lime ( Citrus aurant iifolia ). Methods : We used E. faecalis ATCC 29212 and lime extract from Aceh, Indonesia. The microbe was analyzed for its pH adaptation, acid tolerance response, and adhesion assay using a light microscope with a magnification of x1000. Further, statistical tests were performed to analyze both correlation and significance of the acid tolerance and pH adaptation as well as the interaction activity. Results : E. faecalis was able to adapt to a very acidic environment (pH 2.9), which was characterized by an increase in its pH (reaching 4.2) at all concentrations of the lime extract (p < 0.05). E. faecalis was also able to provide acid tolerance response to lime extract based on spectrophotometric data (595 nm) (p < 0.05). Also, the interaction activity of E. faecalis in different concentrations of lime extract was relatively stable within 6 up to 12 hours (p < 0.05), but it became unstable within 24-72 hours (p > 0.05) based on the mass profiles of its interaction activity. Conclusions : E. faecalis can adapt to acidic environments (pH 2.9-4.2); it is also able to tolerate acid generated by Citrus auranti ifolia extract, revealing a stable interaction in the first 6-12 hours.

pH-Dependent Binding of Chloride to a Marine Alkaline Phosphatase Affects the Catalysis, Active Site Stability, and Dimer Equilibrium.

PubMed

Hjörleifsson, Jens G; Ásgeirsson, Bjarni

2017-09-26

The effect of ionic strength on enzyme activity and stability varies considerably between enzymes. Ionic strength is known to affect the catalytic activity of some alkaline phosphatases (APs), such as Escherichia coli AP, but how ions affect APs is debated. Here, we studied the effect of various ions on a cold-adapted AP from Vibrio splendidus (VAP). Previously, we have found that the active form of VAP is extremely unstable at low ionic strengths. Here we show that NaCl increased the activity and stability of VAP and that the effect was pH-dependent in the range of pH 7-10. The activity profile as a function of pH formed two maxima, indicating a possible conformational change. Bringing the pH from the neutral to the alkaline range was accompanied by a large increase in both the K i for inorganic phosphate (product inhibition) and the K M for p-nitrophenyl phosphate. The activity transitions observed as the pH was varied correlated with structural changes as monitored by tryptophan fluorescence. Thermal and urea-induced inactivation was shown to be accompanied by neither dissociation of the active site metal ions nor dimer dissociation. This would suggest that the inactivation involved subtle changes in active site conformation. Furthermore, the VAP dimer equilibrium was studied for the first time and shown to highly favor dimerization, which was dependent on pH and NaCl concentration. Taken together, the data support a model in which anions bind to some specific acceptor in the active site of VAP, resulting in great stabilization and catalytic rate enhancement, presumably through a different mechanism.

Investigation of pH and Temperature Profiles in the GI Tract of Fasted Human Subjects Using the Intellicap(®) System.

PubMed

Koziolek, Mirko; Grimm, Michael; Becker, Dieter; Iordanov, Ventzeslav; Zou, Hans; Shimizu, Jeff; Wanke, Christoph; Garbacz, Grzegorz; Weitschies, Werner

2015-09-01

Gastrointestinal (GI) pH and temperature profiles under fasted-state conditions were investigated in two studies with each 10 healthy human subjects using the IntelliCap(®) system. This telemetric drug delivery device enabled the determination of gastric emptying time, small bowel transit time, and colon arrival time by significant pH and temperature changes. The study results revealed high variability of GI pH and transit times. The gastric transit of IntelliCap(®) was characterized by high fluctuations of the pH with mean values ranging from pH 1.7 to pH 4.7. Gastric emptying was observed after 7-202 min (median: 30 min). During small bowel transit, which had a duration of 67-532 min (median: 247 min), pH values increased slightly from pH 5.9-6.3 in proximal parts to pH 7.4-7.8 in distal parts. Colonic pH conditions were characterized by values fluctuating mainly between pH 5 and pH 8. The pH profiles and transit times described in this work are highly relevant for the comprehension of drug delivery of solid oral dosage forms comprising ionizable drugs and excipients with pH-dependent solubility. © 2014 Wiley Periodicals, Inc. and the American Pharmacists Association.

Identification and characterization of cysteine proteinases of Trypanosoma evansi.

PubMed

Yadav, S C; Kumar, R; Kumar, S; Tatu, U; Singh, R K; Gupta, A K

2011-09-01

Trypanosoma evansi is a causative agent of 'surra', a common haemoprotozoan disease of livestock in India causing high morbidity and mortality in disease endemic areas. The proteinases released by live and dead trypanosomes entail immunosuppression in the infected host, which immensely contribute in disease pathogenesis. Cysteine proteinases are identified in the infectious cycle of trypanosomes such as cruzain from Trypanosoma cruzi, rhodesain or brucipain from Trypanosoma brucei rhodesiense and congopain from Trypanosoma congelense. These enzymes localised in lysosome-like organelles, flagellar pocket and on cell surface, which play a critical role in the life cycle of protozoan parasites, viz. in host invasion, nutrition and alteration of the host immune response. The paper describes the identification of cysteine proteinases of T. evansi lysate, activity profile at different pH optima and inhibition pattern using a specific inhibitor, besides the polypeptide profile of an antigen. Eight proteinases of T. evansi were identified in the molecular weight (MW) ranges of 28-170 kDa using gelatin substrate-polyacrylamide gel electrophoresis (GS-PAGE), and of these proteinases, six were cysteine proteinases, as they were inhibited by L-3-carboxy-2,3-transepoxypropionyl-lecuylamido (4-guanidino)-butane (E-64), a specific inhibitor. These proteolytic enzymes were most reactive in acidic pH between 3.0 and 5.5 in the presence of dithiothreitol and completely inactive at alkaline pH 10.0. Similarly, the GS-PAGE profile of the serum samples of rats infected with T. evansi revealed strong proteolytic activity only at the 28-kDa zone at pH 5.5, while no proteolytic activity was observed in serum samples of uninfected rats. Further, the other zones of clearance, which were evident in T. evansi antigen zymogram, could not be observed in the serum samples of rats infected with T. evansi. The polypeptide pattern of the whole cell lysate antigen revealed 12-15 polypeptide bands ranging from 28 to 81 kDa along with five predominant polypeptides bands (MW of 81, 66, 62, 55 and 45 kDa), which were immunoreactive with hyperimmune serum (HIS) and serum of experimentally infected rabbits with T. evansi infection. The immunoblot recognised antibodies in experimentally infected rabbits and against HIS as well, corresponding to the zone of clearances at lower MW ranges (28-41 kDa), which may be attributed to the potential of these proteinases in the diagnosis of T. evansi infection. Since these thiol-dependent enzymes are most active in acidic pH and considering their inhibition characteristics, these data suggest that they resemble to the mammalian lysosomal cathepsin B and L.

Functional genomics of pH homeostasis in Corynebacterium glutamicum revealed novel links between pH response, oxidative stress, iron homeostasis and methionine synthesis

PubMed Central

2009-01-01

Background The maintenance of internal pH in bacterial cells is challenged by natural stress conditions, during host infection or in biotechnological production processes. Comprehensive transcriptomic and proteomic analyses has been conducted in several bacterial model systems, yet questions remain as to the mechanisms of pH homeostasis. Results Here we present the comprehensive analysis of pH homeostasis in C. glutamicum, a bacterium of industrial importance. At pH values between 6 and 9 effective maintenance of the internal pH at 7.5 ± 0.5 pH units was found. By DNA microarray analyses differential mRNA patterns were identified. The expression profiles were validated and extended by 1D-LC-ESI-MS/MS based quantification of soluble and membrane proteins. Regulators involved were identified and thereby participation of numerous signaling modules in pH response was found. The functional analysis revealed for the first time the occurrence of oxidative stress in C. glutamicum cells at neutral and low pH conditions accompanied by activation of the iron starvation response. Intracellular metabolite pool analysis unraveled inhibition of the TCA and other pathways at low pH. Methionine and cysteine synthesis were found to be activated via the McbR regulator, cysteine accumulation was observed and addition of cysteine was shown to be toxic under acidic conditions. Conclusions Novel limitations for C. glutamicum at non-optimal pH values were identified by a comprehensive analysis on the level of the transcriptome, proteome, and metabolome indicating a functional link between pH acclimatization, oxidative stress, iron homeostasis, and metabolic alterations. The results offer new insights into bacterial stress physiology and new starting points for bacterial strain design or pathogen defense. PMID:20025733

Purification and properties of adenosine kinase from rat brain.

PubMed

Yamada, Y; Goto, H; Ogasawara, N

1980-12-04

Adenosine kinase (ATP:adenosine 5'-phosphotransferase, EC 2.7.1.20) has been purified to apparent homogeneity from rat brain by (NH4)2SO4 fractionation, affinity chromatography on AMP-Sepharose 4B, gel filtration with Sephadex G-100, and DE-52 cellulose column chromatography. The yield was 56% of the initial activity with a final specific activity of 7.8 mumol/min per mg protein. The molecular weight was estimated as 38 000 by gel filtration with Sephadex G-100 and 41 000 by acrylamide gel electrophoresis in the presence of sodium dodecyl sulfate (SDS). The enzyme catalyzed the phosphorylation of adenosine, deoxyadenosine, arabinoadenosine, inosine and ribavirin. The activity of deoxyadenosine phosphorylation was 20% that of adenosine phosphorylation. The pH optimum profile was biphasic; a sharp pH optimum at pH 5.5 and a broad pH optimum at pH 7.5-8.5. The Km value for adenosine was 0.2 microM and the maximum activity was observed at 0.5 microM. At higher concentrations of adenosine, the activity was strongly inhibited. The Km value for ATP was 0.02 mM and that for Mg2+ was 0.1 mM. GTP, dGTP, dATP and UTP were also proved to be effective phosphate donors. Co2+ was as effective as Mg2+, and Ca2+, Mn2+ or Ni2+ showed about 50% of the activity for Mg2+. The kinase is quite unstable, but stable in the presence of a high concentration of salt; e.g., 0.15 M KCl.

A broad pH range indicator-based spectrophotometric assay for true lipases using tributyrin and tricaprylin.

PubMed

Camacho-Ruiz, María de Los Angeles; Mateos-Díaz, Juan Carlos; Carrière, Frédéric; Rodriguez, Jorge A

2015-05-01

A continuous assay is proposed for the screening of acidic, neutral, or alkaline lipases using microtiter plates, emulsified short- and medium-chain TGs, and a pH indicator. The lipase activity measurement is based on the decrease of the pH indicator optical density due to protonation which is caused by the release of FFAs during the hydrolysis of TGs and thus acidification. Purified lipases with distinct pH optima and an esterase were used to validate the method. The rate of lipolysis was found to be linear with time and proportional to the amount of enzyme added in each case. Specific activities measured with this microplate assay method were lower than those obtained by the pH-stat technique. Nevertheless, the pH-dependent profiles of enzymatic activity were similar with both assays. In addition, the substrate preference of each enzyme tested was not modified and this allowed discriminating lipase and esterase activities using tributyrin (low water solubility) and tricaprylin (not water soluble) as substrates. This continuous lipase assay is compatible with a high sample throughput and can be applied for the screening of lipases and lipase inhibitors from biological samples. Copyright © 2015 by the American Society for Biochemistry and Molecular Biology, Inc.

Cell function and viability in glucose polymer peritoneal dialysis fluids.

PubMed

Liberek, T; Topley, N; Mistry, C D; Coles, G A; Morgan, T; Quirk, R A; Williams, J D

1993-01-01

To investigate the biocompatibility profile of a new peritoneal dialysis fluid containing glucose polymer (GPF). Viability and function of peripheral neutrophils (PMN) from healthy donors and cultured human peritoneal mesothelial cells were assessed in vitro after exposure to dialysis fluids. Phagocytosis, leukotriene B4 synthesis, and respiratory burst activation were measured following stimulation with serum-treated zymosan (STZ) or opsonized Staphylococcus epidermidis (S. epidermidis). Bacterial growth in the fluids was also investigated. In vivo pH equilibration of GPF and subsequent respiratory burst activation following incubation in spent dialysate were studied. For all the host defense parameters measured, commercial dialysis fluids (Dianeal; 1.36% and 3.86% glucose) and GPF (pH 5.2) were significantly more inhibitory than the control buffer (pH 7.3). Mesothelial cell viability was reduced by all the fluids tested irrespective of pH. Glucose polymer fluid was significantly more inhibitory than Dianeal 1.36% for STZ phagocytosis and respiratory burst activation. In contrast, it was less suppressive than Dianeal 3.86% for LTB4 synthesis. For all parameters tested, except LTB4 generation, there was a marked effect of pH, with GPF being significantly more inhibitory at pH 5.2 than at pH 7.3. None of the fluids tested supported the growth of S. epidermidis, although the viable counts in GFP were significantly higher than in Dianeal. Fluid inhibition of PMN respiratory burst activation and cytotoxicity were reduced in a time-dependent manner following increasing dwell time in vivo. GPF does not appear to be significantly different from Dianeal as far as host defense parameters are concerned. However, the cell viability and bacterial survival data suggest some possibly negative aspects of this fluid formation.

Reactivity and operational stability of N-tailed TAMLs through kinetic studies of the catalyzed oxidation of orange II by H2 O2 : synthesis and X-ray structure of an N-phenyl TAML.

PubMed

Warner, Genoa R; Mills, Matthew R; Enslin, Clarissa; Pattanayak, Shantanu; Panda, Chakadola; Panda, Tamas Kumar; Gupta, Sayam Sen; Ryabov, Alexander D; Collins, Terrence J

2015-04-13

The catalytic activity of the N-tailed ("biuret") TAML (tetraamido macrocyclic ligand) activators [Fe{4-XC6 H3 -1,2-(NCOCMe2 NCO)2 NR}Cl](2-) (3; N atoms in boldface are coordinated to the central iron atom; the same nomenclature is used in for compounds 1 and 2 below), [X, R=H, Me (a); NO2 , Me (b); H, Ph (c)] in the oxidative bleaching of Orange II dye by H2 O2 in aqueous solution is mechanistically compared with the previously investigated activator [Fe{4-XC6 H3 -1,2-(NCOCMe2 NCO)2 CMe2 }OH2 ](-) (1) and the more aggressive analogue [Fe(Me2 C{CON(1,2-C6 H3 -4-X)NCO}2 )OH2 ](-) (2). Catalysis by 3 of the reaction between H2 O2 and Orange II (S) occurs according to the rate law found generally for TAML activators (v=kI kII [Fe(III) ][S][H2 O2 ]/(kI [H2 O2 ]+kII [S]) and the rate constants kI and kII at pH 7 both decrease within the series 3 b>3 a>3 c. The pH dependency of kI and kII was investigated for 3 a. As with all TAML activators studied to-date, bell-shaped profiles were found for both rate constants. For kI , the maximal activity was found at pH 10.7 marking it as having similar reactivity to 1 a. For kII , the broad bell pH profile exhibits a maximum at pH about 10.5. The condition kI ≪kII holds across the entire pH range studied. Activator 3 b exhibits pronounced activity in neutral to slightly basic aqueous solutions making it worthy of consideration on a technical performance basis for water treatment. The rate constants ki for suicidal inactivation of the active forms of complexes 3 a-c were calculated using the general formula ln([S0 ]/[S∞ ])=(kII /ki )[Fe(III) ]; here [Fe(III) ], [S0 ], and [S∞ ] are the total catalyst concentration and substrate concentration at time zero and infinity, respectively. The synthesis and X-ray characterization of 3 c are also described. © 2015 WILEY-VCH Verlag GmbH & Co. KGaA, Weinheim.

pH-dependent antitumor activity of proton pump inhibitors against human melanoma is mediated by inhibition of tumor acidity.

PubMed

De Milito, Angelo; Canese, Rossella; Marino, Maria Lucia; Borghi, Martina; Iero, Manuela; Villa, Antonello; Venturi, Giulietta; Lozupone, Francesco; Iessi, Elisabetta; Logozzi, Mariantonia; Della Mina, Pamela; Santinami, Mario; Rodolfo, Monica; Podo, Franca; Rivoltini, Licia; Fais, Stefano

2010-07-01

Metastatic melanoma is associated with poor prognosis and still limited therapeutic options. An innovative treatment approach for this disease is represented by targeting acidosis, a feature characterizing tumor microenvironment and playing an important role in cancer malignancy. Proton pump inhibitors (PPI), such as esomeprazole (ESOM) are prodrugs functionally activated by acidic environment, fostering pH neutralization by inhibiting proton extrusion. We used human melanoma cell lines and xeno-transplated SCID mice to provide preclinical evidence of ESOM antineoplastic activity. Human melanoma cell lines, characterized by different mutation and signaling profiles, were treated with ESOM in different pH conditions and evaluated for proliferation, viability and cell death. SCID mice engrafted with human melanoma were used to study ESOM administration effects on tumor growth and tumor pH by magnetic resonance spectroscopy (MRS). ESOM inhibited proliferation of melanoma cells in vitro and induced a cytotoxicity strongly boosted by low pH culture conditions. ESOM-induced tumor cell death occurred via rapid intracellular acidification and activation of several caspases. Inhibition of caspases activity by pan-caspase inhibitor z-vad-fmk completely abrogated the ESOM-induced cell death. ESOM administration (2.5 mg kg(-1)) to SCID mice engrafted with human melanoma reduced tumor growth, consistent with decrease of proliferating cells and clear reduction of pH gradients in tumor tissue. Moreover, systemic ESOM administration dramatically increased survival of human melanoma-bearing animals, in absence of any relevant toxicity. These data show preclinical evidence supporting the use of PPI as novel therapeutic strategy for melanoma, providing the proof of concept that PPI target human melanoma modifying tumor pH gradients.

Computer simulation of immobilized pH gradients at acidic and alkaline extremes - A quest for extended pH intervals

NASA Technical Reports Server (NTRS)

Mosher, Richard A.; Bier, Milan; Righetti, Pier Giorgio

1986-01-01

Computer simulations of the concentration profiles of simple biprotic ampholytes with Delta pKs 1, 2, and 3, on immobilized pH gradients (IPG) at extreme pH values (pH 3-4 and pH 10-11) show markedly skewed steady-state profiles with increasing kurtosis at higher Delta pK values. Across neutrality, all the peaks are symmetric irrespective of their Delta pK values, but they show very high contribution to the conductivity of the background gel and significant alteration of the local buffering capacity. The problems of skewness, due to the exponential conductivity profiles at low and high pHs, and of gel burning due to a strong electroosmotic flow generated by the net charges in the gel matrix, also at low and high pHs, are solved by incorporating in the IPG gel a strong viscosity gradient. This is generated by a gradient of linear polyacrylamide which is trapped in the gel by the polymerization process.

Teleseismic array analysis of upper mantle compressional velocity structure. Ph.D. Thesis

NASA Technical Reports Server (NTRS)

Walck, M. C.

1984-01-01

Relative array analysis of upper mantle lateral velocity variations in southern California, analysis techniques for dense data profiles, the P-wave upper mantle structure beneath an active spreading center: the Gulf of California, and the upper mantle under the Cascade ranges: a comparison with the Gulf of California are presented.

[Environmental Education Units.] Soil Sampling. Stream Profiles. Tree Watching. Plant Puzzles.

ERIC Educational Resources Information Center

Minneapolis Independent School District 275, Minn.

Five of these eleven units describe methods elementary school students can use when studying soil characteristics. Soil nitrogen and water holding capacity tests are included with two techniques for measuring soil pH. Survey methods for soil organisms are suggested. The remaining pamphlets describe diverse activities associated with field…

pH modulation differs during sunflower cotyledon colonization by the two closely related necrotrophic fungi Botrytis cinerea and Sclerotinia sclerotiorum.

PubMed

Billon-Grand, Geneviève; Rascle, Christine; Droux, Michel; Rollins, Jeffrey A; Poussereau, Nathalie

2012-08-01

During pathogenesis on sunflower cotyledons, Botrytis cinerea and Sclerotinia sclerotiorum show a striking resemblance in symptom development. Based on pH change profiles, the colonization process of both fungi can be divided into two stages. The first stage is associated with a pH decrease, resulting from an accumulation of citric and succinic acids. The second stage is correlated with a pH increase, resulting from an accumulation of ammonia. In this article, we also report that oxalic acid is produced at the late stage of the colonization process and that ammonia accumulation is concomitant with a decrease in free amino acids in decaying tissues. Sclerotinia sclerotiorum produces eight-fold more oxalic acid and two-fold less ammonia than B. cinerea. Consequently, during sunflower cotyledon colonization by B. cinerea, pH dynamics differ significantly from those of S. sclerotiorum. In vitro assays support the in planta results and show that decreases in pH are linked to glucose consumption. At different stages of the colonization process, expression profiles of genes encoding secreted proteases were investigated. This analysis highlights that the expression levels of the B. cinerea protease genes are higher than those of S. sclerotiorum. This work suggests that the overt similarities of S. sclerotiorum and B. cinerea symptom development have probably masked our recognition of the dynamic and potentially different metabolic pathways active during host colonization by these two necrotrophic fungi. © 2011 Bayer Crop Science. Molecular Plant Pathology © 2011 BSPP and Blackwell Publishing Ltd.

Portuguese Honeys from Different Geographical and Botanical Origins: A 4-Year Stability Study Regarding Quality Parameters and Antioxidant Activity.

PubMed

Soares, Sonia; Pinto, Diana; Rodrigues, Francisca; Alves, Rita C; Oliveira, M Beatriz P P

2017-08-11

Portuguese honeys (n = 15) from different botanical and geographical origins were analysed regarding their quality parameters (diastase activity, hydroxymethylfurfural content, moisture and pH), colour (L*, a*, b*) and antioxidant profile (total phenolics content, total flavonoids content, DPPH• scavenging activity, and ferric reducing power). The samples were analysed fresh and after 4-years of storage (at 25 °C and protected from light). The hydroxymethylfurfural content and diastase activity of the fresh samples were in accordance with the recommended values described in the legislation. In general, the antioxidant activity of the samples correlated more with the bioactive compounds content than with colour. The storage affected differently each individual sample, especially regarding the antioxidant profile. Nevertheless, although in general the lightness of the samples decreased (and the redness increased), after 4 years, 11 samples still presented acceptable diastase activity and hydroxymethylfurfural values.

Dynamic change of pH in acidogenic fermentation of cheese whey towards polyhydroxyalkanoates production: Impact on performance and microbial population.

PubMed

Gouveia, Ana R; Freitas, Elisabete B; Galinha, Cláudia F; Carvalho, Gilda; Duque, Anouk F; Reis, Maria A M

2017-07-25

Polyhydroxyalkanoates (PHA) are a sustainable alternative to conventional plastics that can be obtained from industrial wastes/by-products using mixed microbial cultures (MMC). MMC PHA production is commonly carried out in a 3-stage process of acidogenesis, PHA culture selection and accumulation. This research focused on the possibility of tailoring PHA by controlling the acidogenic reactor operating conditions, namely pH, using cheese whey as model feedstock. The objective was to investigate the impact that dynamically varying the acidogenic pH, when targeting different PHA monomer profiles, had on the performance and microbial community profile of the anaerobic reactor. To accomplish this, an anaerobic reactor was continuously operated under dynamic pH changes, ranging from pH 4 to 7, turning to pH 6 after each change of pH. At pH 6, lactate and acetate were the dominant products (41-48% gCOD basis and 22-44% gCOD basis, respectively). At low pH, lactate production was higher while at high pH acetate production was favoured. Despite the dynamic change of pH, the fermentation product composition at pH 6 was always similar, showing the resilience of the process, i.e. when the same pH value was imposed, the culture produced the same metabolic products independently of the history of changes occurring in the system. The different fermentation product fractions led to PHAs of different compositions. The microbial community, analysed by high throughput sequencing of bacterial 16S rRNA gene fragments, was dominated by Lactobacillus, but varied markedly when subjected to the highest and lowest pH values of the tested range (4 and 7), with increase in the abundance of Lactococcus and a member of the Candidate Division TM7. Different bacterial profiles obtained at pH 6 during this dynamic operation were able to produce a consistent profile of fermentation products (and consequently a constant PHA composition), demonstrating the community's functional redundancy. Copyright © 2016 Elsevier B.V. All rights reserved.

Spatial distribution and vertical migration of (137)Cs in soils of Belgrade (Serbia) 25 years after the Chernobyl accident.

PubMed

Petrović, Jelena; Ćujić, Mirjana; Đorđević, Milan; Dragović, Ranko; Gajić, Boško; Miljanić, Šćepan; Dragović, Snežana

2013-06-01

In this study, the specific activity of (137)Cs was determined by gamma-ray spectrometry in 72 surface soil samples and 11 soil profiles collected from the territory of Belgrade 25 years after the Chernobyl accident. Based on the data obtained the external effective gamma dose rates due to (137)Cs were assessed and geographically mapped. The influence of pedogenic factors (pH, specific electrical conductivity, cation exchange capacity, organic matter content, soil particle size and carbonate content) on the spatial and vertical distribution of (137)Cs in soil was estimated through Pearson correlations. The specific activity of (137)Cs in surface soil samples ranged from 1.00 to 180 Bq kg(-1), with a mean value of 29.9 Bq kg(-1), while in soil profiles they ranged from 0.90 to 58.0 Bq kg(-1), with a mean value of 15.3 Bq kg(-1). The mean external effective gamma dose at 1 m above the ground due to (137)Cs in the soil was calculated to be 1.96 nSv h(-1). Geographic mapping of the external effective gamma dose rates originating from (137)Cs revealed much higher dose rates in southern parts of Belgrade city and around the confluence of the Sava and Danube. Negative Pearson correlation coefficients were found between pH, cation exchange capacity and (137)Cs specific activity in surface soil. There were positive correlations between organic matter and (137)Cs specific activity in surface soil; and between specific electrical conductivity, organic matter, silt content and (137)Cs specific activity in soil profiles.

Helicobacter pylori colonization critically depends on postprandial gastric conditions

PubMed Central

Bücker, Roland; Azevedo-Vethacke, Marina; Groll, Claudia; Garten, Désirée; Josenhans, Christine; Suerbaum, Sebastian; Schreiber, Sören

2012-01-01

The risk of Helicobacter pylori infection is highest in childhood, but the colonization process of the stomach mucosa is poorly understood. We used anesthetized Mongolian gerbils to study the initial stages of H. pylori colonization. Prandial and postprandial gastric conditions characteristic of humans of different ages were simulated. The fraction of bacteria that reached the deep mucus layer varied strongly with the modelled postprandial conditions. Colonization success was weak with fast gastric reacidification typical of adults. The efficiency of deep mucus entry was also low with a slow pH decrease as seen in pH profiles simulating the situation in babies. Initial colonization was most efficient under conditions simulating the postprandial reacidification and pepsin activation profiles in young children. In conclusion, initial H. pylori colonization depends on age-related gastric physiology, providing evidence from an in vivo infection model that suggests an explanation why the bacterium is predominantly acquired in early childhood. PMID:23251780

Formulation and advantages of furazolidone in liposomal drug delivery systems.

PubMed

Alam, Muhammad Irfan; Paget, Timothy; Elkordy, Amal Ali

2016-03-10

Furazolidone has proven to have antiprotozoal and antibacterial activity. A number of literature supported its use against Helicobacter pylori. This potential application opens new prospects of its use in clinical settings in triple therapy. In order to avoid side effects associated with this drug, liposomal mucoadhesive drug delivery that can work locally in stomach is considered as an appropriate approach. This study is a focus on formulations and in vitro characterization of liposomes containing furazolidone. Therefore, the effects of variable amounts of drug and cholesterol on encapsulation efficacy and in vitro drug release were evaluated for different liposomal formulations. Mucoadhesive behavior of chitosan coated liposomal at two different pHs was also evaluated and increase in pH from 1.3 to 4.5 increased mucoadhesion from 42% to 60% respectively. Increasing the amount of drug from 4mg to 5mg increased encapsulation activity however, increasing the drug any further decreased encapsulation activity. In contrast, by increasing the amount of cholesterol decrease in encapsulation activity was observed. The optimized formulation with 5mg of drug and 53mg of cholesterol in formulation gave 57% drug release at pH 1.3 but release was increased up to 71% by increasing pH to 4.5 for same amount of drug. However, by using 10.6mg of cholesterol and 5mg of drug the overall release was increased at both pH conditions, at pH 1.3 release was 69% as compared to 77% at pH 4.5. This trend of drug release profile and mucoadhesion that favors pH 4.5 is documented as useful in targeting H. pylori as normal pH of stomach is expected to be higher by the influence of this microbe. Hence, the results of this research can be taken further into a future in vivo study. Copyright © 2016 Elsevier B.V. All rights reserved.

High Frequency and Poor Outcome of Philadelphia Chromosome–Like Acute Lymphoblastic Leukemia in Adults

PubMed Central

Roberts, Kathryn G.; Gu, Zhaohui; Payne-Turner, Debbie; McCastlain, Kelly; Harvey, Richard C.; Chen, I-Ming; Pei, Deqing; Iacobucci, Ilaria; Valentine, Marcus; Pounds, Stanley B.; Shi, Lei; Li, Yongjin; Zhang, Jinghui; Cheng, Cheng; Rambaldi, Alessandro; Tosi, Manuela; Spinelli, Orietta; Radich, Jerald P.; Minden, Mark D.; Rowe, Jacob M.; Luger, Selina; Litzow, Mark R.; Tallman, Martin S.; Wiernik, Peter H.; Bhatia, Ravi; Aldoss, Ibrahim; Kohlschmidt, Jessica; Mrózek, Krzysztof; Marcucci, Guido; Bloomfield, Clara D.; Stock, Wendy; Kornblau, Stephen; Kantarjian, Hagop M.; Konopleva, Marina; Paietta, Elisabeth; Willman, Cheryl L.

2017-01-01

Purpose Philadelphia chromosome (Ph) –like acute lymphoblastic leukemia (ALL) is a high-risk subtype of childhood ALL characterized by kinase-activating alterations that are amenable to treatment with tyrosine kinase inhibitors. We sought to define the prevalence and genomic landscape of Ph-like ALL in adults and assess response to conventional chemotherapy. Patients and Methods The frequency of Ph-like ALL was assessed by gene expression profiling of 798 patients with B-cell ALL age 21 to 86 years. Event-free survival and overall survival were determined for Ph-like ALL versus non–Ph-like ALL patients. Detailed genomic analysis was performed on 180 of 194 patients with Ph-like ALL. Results Patients with Ph-like ALL accounted for more than 20% of adults with ALL, including 27.9% of young adults (age 21 to 39 years), 20.4% of adults (age 40 to 59 years), and 24.0% of older adults (age 60 to 86 years). Overall, patients with Ph-like ALL had an inferior 5-year event-free survival compared with patients with non–Ph-like ALL (22.5% [95% CI, 14.9% to 29.3%; n = 155] v 49.3% [95% CI, 42.8% to 56.2%; n = 247], respectively; P < .001). We identified kinase-activating alterations in 88% of patients with Ph-like ALL, including CRLF2 rearrangements (51%), ABL class fusions (9.8%), JAK2 or EPOR rearrangements (12.4%), other JAK-STAT sequence mutations (7.2%), other kinase alterations (4.1%), and Ras pathway mutations (3.6%). Eleven new kinase rearrangements were identified, including four involving new kinase or cytokine receptor genes and seven involving new partners for previously identified genes. Conclusion Ph-like ALL is a highly prevalent subtype of ALL in adults and is associated with poor outcome. The diverse range of kinase-activating alterations in Ph-like ALL has important therapeutic implications. Trials comparing the addition of tyrosine kinase inhibitors to conventional therapy are required to evaluate the clinical utility of these agents in the treatment of Ph-like ALL. PMID:27870571

Mechanistic Studies of the Yeast Polyamine Oxidase Fms1: Kinetic Mechanism, Substrate Specificity, and pH Dependence†

PubMed Central

Adachi, Mariya S.; Torres, Jason M.; Fitzpatrick, Paul F.

2010-01-01

The flavoprotein oxidase Fms1 from Saccharomyces cerevisiae catalyzes the oxidation of spermine and N1-acetylspermine to yield spermidine and 3-aminopropanal or N-acetyl-3-aminopropanal. The kinetic mechanism of the enzyme has been determined with both substrates. The initial velocity patterns are ping-pong, consistent with reduction being kinetically irreversible. Reduction of Fms1 by either substrate is biphasic. The rate constant for the rapid phase varies with the substrate concentration, with limiting rates for reduction of the enzyme of 126 and 1410 s−1 and apparent Kd values of 24.3 and 484 μM for spermine and N1-acetylspermine, respectively. The rapid phase is followed by a concentration-independent phase that is slower than turnover. The reaction of the reduced enzyme with oxygen is monophasic, with a rate constant of 402 mM−1 s−1 with spermine at 25 °C, and 204 mM−1 s−1 with N1-acetylspermine at 4 °C, pH 9.0. This step is followed by rate-limiting product dissociation. The kcat/Kamine-pH profiles are bell-shaped, with an average pKa value of 9.3 with spermine and pKa values of 8.3 and 9.6 with N1-acetylspermine. Both profiles are consistent with the active forms of substrates having two charged nitrogens. The pH profiles for the rate constant for flavin reduction show pKa values of 8.3 and 7.2 for spermine and N1-acetylspermine, respectively, for groups that must be unprotonated; these pKa values are assigned to the substrate N4. The kcat/KO2-pH profiles show pKa values of 7.5 for spermine and 6.8 for N1-acetylspermine. With both substrates, the kcat value decreases when a single residue is protonated. PMID:21067138

Diversities and similarities in pH dependency among bacterial NhaB-like Na+/H+ antiporters.

PubMed

Kiriyama, Wakako; Honma, Kei; Hiratsuka, Tomoaki; Takahashi, Itsuka; Nomizu, Takahiro; Takashima, Yuta; Ohtsuka, Masataka; Takahashi, Daiki; Moriyama, Kazuya; Mori, Sayoko; Nishiyama, Shiho; Fukuhara, Masahiro; Nakamura, Tatsunosuke; Shigematsu, Toru; Yamaguchi, Toshio

2013-10-01

NhaB-like antiporters were the second described class of Na(+)/H(+) antiporters, identified in bacteria more than 20 years ago. While nhaB-like gene sequences have been found in a number of bacterial genomes, only a few of the NhaB-like antiporters have been functionally characterized to date. Although earlier studies have identified a few pH-sensitive and -insensitive NhaB-like antiporters, the mechanisms that determine their pH responses still remain elusive. In this study, we sought to investigate the diversities and similarities among bacterial NhaB-like antiporters, with particular emphasis on their pH responsiveness. Our phylogenetic analysis of NhaB-like antiporters, combined with pH profile analyses of activities for representative members of several phylogenetic groups, demonstrated that NhaB-like antiporters could be classified into three distinct types according to the degree of their pH dependencies. Interestingly, pH-insensitive NhaB-like antiporters were only found in a limited proportion of enterobacterial species, which constitute a subcluster that appears to have diverged relatively recently among enterobacterial NhaB-like antiporters. Furthermore, kinetic property analyses of NhaB-like antiporters at different pH values revealed that the degree of pH sensitivity of antiport activities was strongly correlated with the magnitude of pH-dependent change in apparent Km values, suggesting that the dramatic pH sensitivities observed for several NhaB-like antiporters might be mainly due to the significant increases of apparent Km at lower pH. These results strongly suggested the possibility that the loss of pH sensitivity of NhaB-like antiporters had occurred relatively recently, probably via accumulation of the mutations that impair pH-dependent change of Km in the course of molecular evolution.

A Novel Phytase Derived from an Acidic Peat-Soil Microbiome Showing High Stability under Acidic Plus Pepsin Conditions.

PubMed

Tan, Hao; Wu, Xiang; Xie, Liyuan; Huang, Zhongqian; Peng, Weihong; Gan, Bingcheng

2016-01-01

Four novel phytases of the histidine acid phosphatase family were identified in two publicly available metagenomic datasets of an acidic peat-soil microbiome in northeastern Bavaria, Germany. These enzymes have low similarity to all the reported phytases. They were overexpressed in Escherichia coli and purified. Catalytic efficacy in simulated gastric fluid was measured and compared among the four candidates. The phytase named rPhyPt4 was selected for its high activity. It is the first phytase identified from unculturable Acidobacteria. The phytase showed a longer half-life than all the gastric-stable phytases that have been reported to date, suggesting a strong resistance to low pH and pepsin. A wide pH profile was observed between pH 1.5 and 5.0. At the optimum pH (2.5) the activity was 2,790 μmol/min/mg at the physiological temperature of 37°C and 3,989 μmol/min/mg at the optimum temperature of 60°C. Due to the competent activity level as well as the high gastric stability, the phytase could be a potential candidate for practical use in livestock and poultry feeding. © 2016 S. Karger AG, Basel.

Modeling the dynamic volatile fatty acids profiles with pH and hydraulic retention time in an anaerobic baffled reactor during the startup period.

PubMed

Shi, En; Li, Jianzheng; Leu, Shao-Yuan; Antwi, Philip

2016-12-01

To predict the dynamic profiles in volatile fatty acids (VFAs) with pH and hydraulic retention time (HRT) during the startup of a 4-compartment ABR, a mathematical model was constructed by introducing pH and thermodynamic inhibition functions into the biochemical processes derived from the ADM1. The calibration of inhibition parameter for propionate uptake effectively improved the prediction accuracy of VFAs. The developed model could simulate the VFAs profiles very well no matter the observable change of pH or/and HRT. The simulation results indicated that both H 2 -producing acetogenesis and methanogenesis in the ABR would be inhibited with a pH less than 4.61, and the propionate oxidation could be thermodynamically restricted even with a neutral pH. A decreased HRT would enhanced the acidogenesis and H 2 -producing acetogenesis in the first 3 compartments, but no observable increase in effluent VFAs could be found due to the synchronously enhanced methanogenesis in the last compartment. Copyright © 2016 Elsevier Ltd. All rights reserved.

The Redundancy of Peptidoglycan Carboxypeptidases Ensures Robust Cell Shape Maintenance in Escherichia coli

PubMed Central

Peters, Katharina; Kannan, Suresh; Rao, Vincenzo A.; Biboy, Jacob; Vollmer, Daniela; Erickson, Stephen W.; Lewis, Richard J.

2016-01-01

ABSTRACT Peptidoglycan (PG) is an essential structural component of the bacterial cell wall and maintains the integrity and shape of the cell by forming a continuous layer around the cytoplasmic membrane. The thin PG layer of Escherichia coli resides in the periplasm, a unique compartment whose composition and pH can vary depending on the local environment of the cell. Hence, the growth of the PG layer must be sufficiently robust to allow cell growth and division under different conditions. We have analyzed the PG composition of 28 mutants lacking multiple PG enzymes (penicillin-binding proteins [PBPs]) after growth in acidic or near-neutral-pH media. Statistical analysis of the muropeptide profiles identified dd-carboxypeptidases (DD-CPases) that were more active in cells grown at acidic pH. In particular, the absence of the DD-CPase PBP6b caused a significant increase in the pentapeptide content of PG as well as morphological defects when the cells were grown at acidic pH. Other DD-CPases (PBP4, PBP4b, PBP5, PBP6a, PBP7, and AmpH) and the PG synthase PBP1B made a smaller or null contribution to the pentapeptide-trimming activity at acidic pH. We solved the crystal structure of PBP6b and also demonstrated that the enzyme is more stable and has a lower Km at acidic pH, explaining why PBP6b is more active at low pH. Hence, PBP6b is a specialized DD-CPase that contributes to cell shape maintenance at low pH, and E. coli appears to utilize redundant DD-CPases for normal growth under different conditions. PMID:27329754

[Influence of polymer type on the physical properties and the release study of papaverine hydrochloride from tablets].

PubMed

Kasperek, Regina; Polski, Andrzej; Sobótka-Polska, Karolina; Poleszak, Ewa

2014-01-01

Polymers are widely used in drug manufacturing. Researchers studied their impact on the bioavailability of active substances or on physical properties of tablets for many years. To study the influence of polymer excipients, such as microcrystalline cellulose (Avicel PH 101, Avicel PH 102), croscarmellose sodium, crospovidone or polyvinylpyrrolidone, on the release profile of papaverine hydrochloride from tablets and on the physical properties of tablets. Six series of uncoated tablets were prepared by indirect method, with previous wet granulation. Tablets contained papaverine hydrochloride and various excipients. The physical properties of the prepared granules, tablets and the release profile of papaverine hydrochloride from tablets were examined. The content of papaverine hydrochloride from the release study were determined spectrophotometrically. All tablets met the pharmacopoeia requirements during following tests: the disintegration time of tablets, uncoated tablets resistance to abrasion, the weight uniformity and dose formulations, their dimensions, the resistance to crushing of tablets and the drug substance content in the tablet. In four cases more than 80% of papaverine was released up to 2 min, in one formula it was up to 5 min, and in last one up to 10 min. Tablets containing crospovidone disintegrated faster than tablets with croscarmellose sodium. Adding gelatinized starch to the tablet composition increased the disintegration time, hardness and delayed the release of papaverine. During the wet granulation process, granules containing polyvinylpyrrolidone were characterized by a suitable flow properties and slightly prolonged disintegration time. Tablets containing Avicel PH 102 compared to tablets with Avicel PH 101 had less weight loss during the test of mechanical resistance, improved hardness and faster release profile of papaverine from tablets.

Geochemistry of Red Mountain Creek, Colorado, under low-flow conditions, August 2002

USGS Publications Warehouse

Runkel, Robert L.; Kimball, Briant A.; Walton-Day, Katherine; Verplanck, Philip L.

2005-01-01

Red Mountain Creek, an acid mine drainage stream in southwestern Colorado, was the subject of a synoptic study conducted in August 2002. During the synoptic study, a solution containing lithium chloride was injected continuously to allow for the calculation of streamflow using the tracer-dilution method. Synoptic water-quality samples were collected from 48 stream sites and 29 inflow locations along a 5.4-kilometer study reach. Data from the study provide profiles of pH, concentration, and mass load with a high degree of spatial resolution. Despite the presence of 10 circumneutral inflows, pH remained below 3.4 at all stream sites. Concentration profiles indicate that dissolved concentrations of aluminum, cadmium, copper, lead, and zinc exceed chronic aquatic-life standards established by the State of Colorado along the entire study reach. Comparison of total recoverable and dissolved concentrations suggests that most constituents were transported conservatively. Exceptions to this pattern include arsenic, iron, molybdenum, and vanadium, four constituents that were subject to precipitation and(or) sorption reactions as the addition of a circumneutral tributary resulted in a slight increase in instream pH. Evaluation of data from the 29 inflow locations indicates a sharp contrast between the east and west sides of the watershed; inflows from the east side have high constituent concentrations and acidic pH, whereas inflows from the west side have lower concentrations and generally higher pH. Loading profiles, the product of streamflow and concentration, are used to rank potential sources of metals and acidity within the watershed. Four sources account for 83, 72, 70, 69, 64, and 61 percent of the aluminum, iron, arsenic, zinc, copper, and cadmium loading within the study reach, respectively. All four sources appear to be the result of surface inflows that have been affected by mining activities. The relatively small number of major sources and the fact that they are attributable to surface inflows are two factors that may facilitate effective remediation.

Proteases and nucleases involved in the biphasic digestion process of the brown marmorated stink bug, Halyomorpha halys (Hemiptera: Pentatomidae).

PubMed

Lomate, Purushottam R; Bonning, Bryony C

2018-07-01

Management of the brown marmorated stink bug, Halyomorpha halys (Hemiptera: Pentatomidae), an invasive, agricultural pest in the United States, has presented significant challenges. This polyphagous insect uses both extra-oral and gut-based digestion thwarting protein- or nucleotide-based control strategies. The objective of this study was to biochemically characterize the digestive enzymes (proteases and nucleases) from the saliva, salivary gland and the gut of H. halys. Enzyme profiles for the two tissues and saliva radically differ: The pH optimum for proteases in the gut was six, with cysteine proteases predominant. In contrast, the alkaline pH optima for protease activity in the salivary gland (8-10) and saliva (7) reflected abundant serine protease and cathepsin activities. RNase enzymes were most abundant in saliva, while dsRNase and DNase activities were higher in the salivary gland and saliva compared to those in the gut. These very different enzyme profiles highlight the biphasic digestive system used by this invasive species for efficient processing of plant nutrients. Knowledge of H. halys digestive physiology will allow for counteractive measures targeting digestive enzymes or for appropriate protection of protein- or nucleotide-based management options targeting this pest. © 2018 Wiley Periodicals, Inc.

Chemical mechanism of D-amino acid oxidase from Rhodotorula gracilis: pH dependence of kinetic parameters.

PubMed Central

Ramón, F; Castillón, M; De La Mata, I; Acebal, C

1998-01-01

The variation of kinetic parameters of d-amino acid oxidase from Rhodotorula gracilis with pH was used to gain information about the chemical mechanism of the oxidation of D-amino acids catalysed by this flavoenzyme. d-Alanine was the substrate used. The pH dependence of Vmax and Vmax/Km for alanine as substrate showed that a group with a pK value of 6.26-7.95 (pK1) must be unprotonated and a group with a pK of 10.8-9.90 (pK2) must be protonated for activity. The lower pK value corresponded to a group on the enzyme involved in catalysis and whose protonation state was not important for binding. The higher pK value was assumed to be the amino group of the substrate. Profiles of pKi for D-aspartate as competitive inhibitor showed that binding is prevented when a group on the enzyme with a pK value of 8.4 becomes unprotonated; this basic group was not detected in Vmax/Km profiles suggesting its involvement in binding of the beta-carboxylic group of the inhibitor. PMID:9461524

Intragastric pH and pressure profiles after intake of the high-caloric, high-fat meal as used for food effect studies.

PubMed

Koziolek, M; Schneider, F; Grimm, M; Modeβ, Chr; Seekamp, A; Roustom, T; Siegmund, W; Weitschies, W

2015-12-28

The intraluminal conditions of the fed stomach are critical for drug release from solid oral dosage forms and thus, often associated with the occurrence of food effects on oral bioavailability. In this study, intragastric pH and pressure profiles present after the ingestion of the high-caloric, high-fat (964 kcal) FDA standard breakfast were investigated in 19 healthy human subjects by using the telemetric SmartPill® capsule system (26 × 13 mm). Since the gastric emptying of such large non-digestible objects is typically accomplished by the migrating motor complex phase III activity, the time required for recurrence of fasted state motility determined the gastric emptying time (GET). Following the diet recommendations of the FDA guidance on food effect studies, the mean GET of the telemetric motility capsule was 15.3 ± 4.7 h. Thus, the high caloric value of the standard breakfast impeded gastric emptying before lunch in 18 out of 19 subjects. During its gastric transit, the capsule was exposed to highly dynamic conditions in terms of pH and pressure, which were mainly dependent on further meal and liquid intake, as well as the intragastric capsule deposition behavior. Maximum pH values in the stomach were measured immediately after capsule intake. The median pH value of the 5 min period after capsule ingestion ranged between pH 3.3 and 5.3. Subsequently, the pH decreased relatively constantly and reached minimum values of pH 0-1 after approximately 4 h. The maximum pressure within the stomach amounted to 293 ± 109 mbar and was clearly higher than the maximum pressure measured at the ileocaecal junction (60 ± 35 mbar). The physiological data on the intraluminal conditions within the fed stomach generated in this study will hopefully contribute to a better understanding of food effects on oral drug product performance.

Purification of an Exopolygalacturonase from Penicillium viridicatum RFC3 Produced in Submerged Fermentation.

PubMed

Gomes, Eleni; Leite, Rodrigo Simões Ribeiro; da Silva, Roberto; Silva, Dênis

2009-01-01

An exo-PG obtained from Penicillium viridicatum in submerged fermentation was purified to homogeneity. The apparent molecular weight of the enzyme was 92 kDa, optimum pH and temperature for activity were pH 5 and 50-55 degrees C. The exo-PG showed a profile of an exo-polygalacturonase, releasing galacturonic acid by hydrolysis of pectin with a high degree of esterification (D.E.). Ions Ca(2+) enhanced the stability of enzyme and its activity by 30%. The K(m) was 1.30 in absence of Ca(2+) and 1.16 mg mL(-1) in presence of this ion. In relation to the V(max) the presence of this ion increased from 1.76 to 2.07 mumol min(-1)mg(-1).

Production of Pectate Lyase by Penicillium viridicatum RFC3 in Solid-State and Submerged Fermentation.

PubMed

Ferreira, Viviani; da Silva, Roberto; Silva, Dênis; Gomes, Eleni

2010-01-01

Pectate lyase (PL) was produced by the filamentous fungus Penicillium viridicatum RFC3 in solid-state cultures of a mixture of orange bagasse and wheat bran (1 : 1 w/w), or orange bagasse, wheat bran and sugarcane bagasse (1 : 1 : 0.5 w/w), and in a submerged liquid culture with orange bagasse and wheat bran (3%) as the carbon source. PL production was highest (1,500 U mL(-1) or 300 Ug(-1) of substrate) in solid-state fermentation (SSF) on wheat bran and orange bagasse at 96 hours. PL production in submerged fermentation (SmF) was influenced by the initial pH of the medium. With the initial pH adjusted to 4.5, 5.0, and 5.5, the peak activity was observed after 72, 48, and 24 hours of fermentation, respectively, when the pH of the medium reached the value 5.0. PL from SSF and SmF were loaded on Sephadex-G75 columns and six activity peaks were obtained from crude enzyme from SSF and designated PL I, II, III, IV, V, and VI, while five peaks were obtained from crude enzyme from SmF and labeled PL I', II', III', IV', and VII'. Crude enzyme and fraction III from each fermentative process were tested further. The optimum pH for crude PL from either process was 5.5, while that for PL III was 8.0. The maximum activity of enzymes from SSF was observed at 35 degrees C, but crude enzyme was more thermotolerant than PL III, maintaining its maximum activity up to 45 degrees C. Crude enzyme from SmF and PL III' showed thermophilic profiles of activity, with maximum activity at 60 and 55 degrees C, respectively. In the absence of substrate, the crude enzyme from SSF was stable over the pH range 3.0-10.0 and PL III was most stable in the pH range 4.0-7.0. Crude enzyme from SmF retained 70%-80% of its maximum activity in the acid-neutral pH range (4.0-7.0), but PIII showed high stability at alkaline pH (7.5-9.5). PL from SSF was more thermolabile than that from SmF. The latter maintained 60% of its initial activity after 1 h at 55 degrees C. The differing behavior of the enzymes with respect to pH and temperature suggests that they are different isozymes.

Whey-grape juice drink processed by supercritical carbon dioxide technology: Physicochemical characteristics, bioactive compounds and volatile profile.

PubMed

Amaral, Gabriela V; Silva, Eric Keven; Cavalcanti, Rodrigo N; Martins, Carolina P C; Andrade, Luiz Guilherme Z S; Moraes, Jeremias; Alvarenga, Verônica O; Guimarães, Jonas T; Esmerino, Erick A; Freitas, Mônica Q; Silva, Márcia C; Raices, Renata S L; Sant' Ana, Anderson S; Meireles, M Angela A; Cruz, Adriano G

2018-01-15

The effect of supercritical carbon dioxide technology (SCCD, 14, 16, and 18MPa at 35±2°C for 10min) on whey-grape juice drink characteristics was investigated. Physicochemical characterization (pH, titratable acidity, total soluble solids), bioactive compounds (phenolic compounds, anthocyanin, DPPH and ACE activity) and the volatile compounds were performed. Absence of differences were found among treatments for pH, titratable acidity, soluble solids, total anthocyanin and DPPH activity (p-value>0.05). A direct relationship between SCCD pressure and ACE inhibitory activity was observed, with 34.63, 38.75, and 44.31% (14, 16, and 18MPa, respectively). Regards the volatile compounds, it was noted few differences except by the presence of ketones. The findings confirm the SCCD processing as a potential promising technology to the conventional thermal treatment. Copyright © 2017 Elsevier Ltd. All rights reserved.

Effects of Low pH on Photosynthesis, Related Physiological Parameters, and Nutrient Profiles of Citrus

PubMed Central

Long, An; Zhang, Jiang; Yang, Lin-Tong; Ye, Xin; Lai, Ning-Wei; Tan, Ling-Ling; Lin, Dan; Chen, Li-Song

2017-01-01

Seedlings of “Xuegan” (Citrus sinensis) and “Sour pummelo” (Citrus grandis) were irrigated daily with a nutrient solution at a pH of 2.5, 3, 4, 5, or 6 for 9 months. Thereafter, the following responses were investigated: seedling growth; root, stem, and leaf concentrations of nutrient elements; leaf gas exchange, pigment concentration, ribulose-1,5-bisphosphate carboxylase/oxygenase activity and chlorophyll a fluorescence; relative water content, total soluble protein level, H2O2 production and electrolyte leakage in roots and leaves. This was done (a) to determine how low pH affects photosynthesis, related physiological parameters, and mineral nutrient profiles; and (b) to understand the mechanisms by which low pH may cause a decrease in leaf CO2 assimilation. The pH 2.5 greatly inhibited seedling growth, and many physiological parameters were altered only at pH 2.5; pH 3 slightly inhibited seedling growth; pH 4 had almost no influence on seedling growth; and seedling growth and many physiological parameters reached their maximum at pH 5. No seedlings died at any given pH. These results demonstrate that citrus survival is insensitive to low pH. H+-toxicity may directly damage citrus roots, thus affecting the uptake of mineral nutrients and water. H+-toxicity and a decreased uptake of nutrients (i.e., nitrogen, phosphorus, potassium, calcium, and magnesium) and water were likely responsible for the low pH-induced inhibition of growth. Leaf CO2 assimilation was inhibited only at pH 2.5. The combinations of an impaired photosynthetic electron transport chain, increased production of reactive oxygen species, and decreased uptake of nutrients and water might account for the pH 2.5-induced decrease in CO2 assimilation. Mottled bleached leaves only occurred in the pH 2.5-treated C. grandis seedlings. Furthermore, the pH 2.5-induced alterations of leaf CO2 assimilation, water-use efficiency, chlorophylls, polyphasic chlorophyll a fluorescence (OJIP) transients and many fluorescence parameters, root and leaf total soluble proteins, H2O2 production, and electrolyte leakage were all slightly greater in C. grandis than in C. sinensis seedlings. Hence, C. sinensis was slightly more tolerant to low pH than C. grandis. In conclusion, our findings provide novel insight into the causes of low pH-induced inhibition of seedling growth and leaf CO2 assimilation. PMID:28270819

Effects of Low pH on Photosynthesis, Related Physiological Parameters, and Nutrient Profiles of Citrus.

PubMed

Long, An; Zhang, Jiang; Yang, Lin-Tong; Ye, Xin; Lai, Ning-Wei; Tan, Ling-Ling; Lin, Dan; Chen, Li-Song

2017-01-01

Seedlings of "Xuegan" ( Citrus sinensis ) and "Sour pummelo" ( Citrus grandis ) were irrigated daily with a nutrient solution at a pH of 2.5, 3, 4, 5, or 6 for 9 months. Thereafter, the following responses were investigated: seedling growth; root, stem, and leaf concentrations of nutrient elements; leaf gas exchange, pigment concentration, ribulose-1,5-bisphosphate carboxylase/oxygenase activity and chlorophyll a fluorescence; relative water content, total soluble protein level, H 2 O 2 production and electrolyte leakage in roots and leaves. This was done ( a ) to determine how low pH affects photosynthesis, related physiological parameters, and mineral nutrient profiles; and ( b ) to understand the mechanisms by which low pH may cause a decrease in leaf CO 2 assimilation. The pH 2.5 greatly inhibited seedling growth, and many physiological parameters were altered only at pH 2.5; pH 3 slightly inhibited seedling growth; pH 4 had almost no influence on seedling growth; and seedling growth and many physiological parameters reached their maximum at pH 5. No seedlings died at any given pH. These results demonstrate that citrus survival is insensitive to low pH. H + -toxicity may directly damage citrus roots, thus affecting the uptake of mineral nutrients and water. H + -toxicity and a decreased uptake of nutrients (i.e., nitrogen, phosphorus, potassium, calcium, and magnesium) and water were likely responsible for the low pH-induced inhibition of growth. Leaf CO 2 assimilation was inhibited only at pH 2.5. The combinations of an impaired photosynthetic electron transport chain, increased production of reactive oxygen species, and decreased uptake of nutrients and water might account for the pH 2.5-induced decrease in CO 2 assimilation. Mottled bleached leaves only occurred in the pH 2.5-treated C. grandis seedlings. Furthermore, the pH 2.5-induced alterations of leaf CO 2 assimilation, water-use efficiency, chlorophylls, polyphasic chlorophyll a fluorescence (OJIP) transients and many fluorescence parameters, root and leaf total soluble proteins, H 2 O 2 production, and electrolyte leakage were all slightly greater in C. grandis than in C. sinensis seedlings. Hence, C. sinensis was slightly more tolerant to low pH than C. grandis . In conclusion, our findings provide novel insight into the causes of low pH-induced inhibition of seedling growth and leaf CO 2 assimilation.

Mechanistic and Structural Analyses of the Role of His67 in the Yeast Polyamine Oxidase Fms1†

PubMed Central

Adachi, Mariya S.; Taylor, Alexander B.; Hart, P. John; Fitzpatrick, Paul F.

2012-01-01

The flavoprotein oxidase Fms1 from Saccharomyces cerevisiae catalyzes the oxidation of spermine and N1-acetylspermine to spermidine and 3-aminopropanal or N-acetyl-3-aminopropanal. Within the active site of Fms1, His67 is positioned to form hydrogen bonds with the polyamine substrate. This residue is also conserved in other polyamine oxidases. The catalytic properties of H67Q, H67N, and H67A Fms1 have been characterized to evaluate the role of this residue in catalysis. With both spermine and N1-acetylspermine as the amine substrate, the value of the first-order rate constant for flavin reduction decreases 2–3 orders of magnitude, with the H67Q mutation having the smallest effect and H67N the largest. The kcat/KO2 value changes very little upon mutation with N1-acetylspermine as the amine substrate and decreases only an order of magnitude with spermine. The kcat/KM-pH profiles with N1-acetylspermine are bell-shaped for all the mutants; the similarity to the profile of the wild-type enzyme rules out His67 as being responsible for either of the pKa values. The pH profiles for the rate constant for flavin reduction for all the mutant enzymes similarly show the same pKa as wild-type Fms1, about ~7.4; this pKa is assigned to the substrate N4. The kcat/KO2-pH profiles for wild-type Fms1 and the H67A enzyme both show a pKa of about ~6.9; this suggests His67 is not responsible for this pH behaviour. With the H67Q, H67N, and H67A enzymes the kcat value decreases when a single residue is protonated, as is the case with the wild-type enzyme. The structure of H67Q Fms1 has been determined at a resolution of 2.4 Å. The structure shows that the mutation disrupts a hydrogen bond network in the active site, suggesting that His67 is important both for direct interactions with the substrate and to maintain the overall active site structure. PMID:22642831

Mechanistic and structural analyses of the role of His67 in the yeast polyamine oxidase Fms1.

PubMed

Adachi, Mariya S; Taylor, Alexander B; Hart, P John; Fitzpatrick, Paul F

2012-06-19

The flavoprotein oxidase Fms1 from Saccharomyces cerevisiae catalyzes the oxidation of spermine and N(1)-acetylspermine to spermidine and 3-aminopropanal or N-acetyl-3-aminopropanal. Within the active site of Fms1, His67 is positioned to form hydrogen bonds with the polyamine substrate. This residue is also conserved in other polyamine oxidases. The catalytic properties of H67Q, H67N, and H67A Fms1 have been characterized to evaluate the role of this residue in catalysis. With both spermine and N(1)-acetylspermine as the amine substrate, the value of the first-order rate constant for flavin reduction decreases 2-3 orders of magnitude, with the H67Q mutation having the smallest effect and H67N the largest. The k(cat)/K(O2) value changes very little upon mutation with N(1)-acetylspermine as the amine substrate and decreases only an order of magnitude with spermine. The k(cat)/K(M)-pH profiles with N(1)-acetylspermine are bell-shaped for all the mutants; the similarity to the profile of the wild-type enzyme rules out His67 as being responsible for either of the pK(a) values. The pH profiles for the rate constant for flavin reduction for all the mutant enzymes similarly show the same pK(a) as wild-type Fms1, about ∼7.4; this pK(a) is assigned to the substrate N4. The k(cat)/K(O2)-pH profiles for wild-type Fms1 and the H67A enzyme both show a pK(a) of about ∼6.9; this suggests His67 is not responsible for this pH behavior. With the H67Q, H67N, and H67A enzymes the k(cat) value decreases when a single residue is protonated, as is the case with the wild-type enzyme. The structure of H67Q Fms1 has been determined at a resolution of 2.4 Å. The structure shows that the mutation disrupts a hydrogen bond network in the active site, suggesting that His67 is important both for direct interactions with the substrate and to maintain the overall active site structure.

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) 229-6916bsra@uwm.eduEng & Math Sciences 995 profile photo Robert Cuzner, Ph.D.Assistant ChairDepartment Chair of Electrical Engineering(414) 229-3885george@uwm.eduEng & Math Sciences 1245 profile photo Hossein Hosseini, Ph.D.ProfessorComputer Science(414) 229-5184hosseini@uwm.eduEng & Math

Profile of Ph.Ds in Canada.

ERIC Educational Resources Information Center

von Zur-Muehlen, Max

A profile of the socio-economic characteristics of the permanent residents of Canada holding an earned doctorate is presented. In 1973, there were 27,410 Canadian residents who had obtained an earned doctorate. (Holders of such professional doctoral degrees as Doctor of Medicine are excluded from this study.) Only 9 percent of the Ph.Ds were…

Direct comparison of two different mesalamine formulations for the induction of remission in patients with ulcerative colitis: A double-blind, randomized study

PubMed Central

Ito, Hiroaki; Iida, Mitsuo; Matsumoto, Takayuki; Suzuki, Yasuo; Sasaki, Hidetaka; Yoshida, Toyomitsu; Takano, Yuichi; Hibi, Toshifumi

2010-01-01

Background: Mesalamine is the first-line drug for the treatment of ulcerative colitis (UC). We directly compared the efficacy and safety of two mesalamine formulations for the induction of remission in patients with UC. Methods: In a multicenter, double-blind, randomized study, 229 patients with mild-to-moderate active UC were assigned to 4 groups: 66 and 65 received a pH-dependent release formulation of 2.4 g/day (pH-2.4 g) or 3.6 g/day (pH-3.6 g), respectively; 65 received a time-dependent release formulation of 2.25 g/day (Time-2.25 g), and 33 received placebo (Placebo). The drugs were administered three times daily for eight weeks. The primary endpoint was a decrease in the UC disease activity index (UC-DAI). Results: In the full analysis set (n = 225) the decrease in UC-DAI in each group was 1.5 in pH-2.4 g, 2.9 in pH-3.6 g, 1.3 in Time-2.25 g and 0.3 in Placebo, respectively. These results demonstrate the superiority of pH-3.6 g over Time-2.25 g (P = 0.003) and the noninferiority of pH-2.4 g to Time-2.25 g. Among the patients with proctitis-type UC, a significant decrease in UC-DAI was observed in pH-2.4 g and pH-3.6 g as compared to Placebo, but not in Time-2.25 g. No differences were observed in the safety profiles. Conclusions: Higher dose of the pH-dependent release formulation was more effective for induction of remission in patients with mild-to-moderate active UC. Additionally, the pH-dependent release formulation was preferable to the time-dependent release formulation for patients with proctitis-type UC (UMIN Clinical Trials Registry, no. C000000288). (Inflamm Bowel Dis 2010) PMID:20049950

Innovative Approach for Interstitial Cystitis: Vaginal Pessaries Loaded Diazepam—A Preliminary Study

PubMed Central

Capra, P.; Perugini, P.; Bleve, M.; Pavanetto, P.; Musitelli, G.; Rovereto, B.; Porru, D.

2013-01-01

Bladder pain is a characteristic disorder of interstitial cystitis. Diazepam is well known for its antispasmodic activity in the treatment of muscular hypertonus. The aim of this work was to develop and characterize vaginal pessaries as an intravaginal delivery system of diazepam for the treatment of interstitial cystitis. In particular, the performance of two types of formulations, with and without beta-glucan, was compared. In particular, the preparation of pessaries, according to the modified Pharmacopeia protocol, the setup of the analytical method to determine diazepam, pH evaluation, dissolution profile, and photostability assay were reported. Results showed that the modified protocol permitted obtaining optimal vaginal pessaries, without air bubbles, with good consistency and handling and with good pH profiles. In order to determine the diazepam amount, calibration curves with good correlation coefficients were obtained, by the spectrophotometric method, using placebo pessaries as matrix with the addition of diazepam standard solution. This method was demonstrated sensible and accurate to determine the amount of drug in batches. Dissolution profiles showed a complete diazepam release just after 15 minutes, even if beta-glucan pessaries released drug more gradually. Finally, a possible drug photodegradation after exacerbated UV-visible exposition was evaluated. PMID:26555976

Enzyme electrokinetics: hydrogen evolution and oxidation by Allochromatium vinosum [NiFe]-hydrogenase.

PubMed

Léger, Christophe; Jones, Anne K; Roseboom, Winfried; Albracht, Simon P J; Armstrong, Fraser A

2002-12-31

The mechanism of catalytic hydrogen evolution and oxidation by Allochromatium vinosum [NiFe]-hydrogenase has been studied by protein film voltammetry (PFV) with the enzyme adsorbed at a pyrolytic graphite edge electrode. By analyzing the entire shapes of catalytic voltammograms, the energetics of the catalytic cycles (reduction potentials and acidity constants of the active states), including the detailed profiles of activity against pH and the sequences of proton and electron transfers, have been determined, and these are discussed with respect to the mechanism. PFV, which probes rates as a continuous function of the electrochemical potential (i.e., in the "potential domain"), is proven to be an invaluable tool for determining the redox properties of an active site in the presence of its substrate, at room temperature, and during turnover. This is especially relevant in the case of the active states of hydrogenase, since one of its substrates (the proton) is always present at significant levels in the titration medium at physiological pH values.

Recovery of soluble proteins from migratory locust (Locusta migratoria) and characterisation of their compositional and techno-functional properties.

PubMed

Purschke, Benedict; Tanzmeister, Helene; Meinlschmidt, Pia; Baumgartner, Sabine; Lauter, Kathrin; Jäger, Henry

2018-04-01

Edible insects emerged as an alternative source of high-quality proteins. Therefore, the effect of an extraction procedure for the recovery of migratory locust (Locusta migratoria) protein concentrate (MLPC) on the compositional characteristics and techno-functional properties was studied. The influence of pH value (2-10) and salt concentration (0, 1 and 3% w/v) on techno-functional properties was evaluated. Proteins were identified and characterized by RP-HPLC, SDS-PAGE and LC-MS/MS. The initial crude protein content of the whole locusts (65.9% on dry base) could be enhanced to 82.3% (MLPC). Solubility profiles of MLPC showed maximum solubility at pH9 (100%). Promising functionality comparable to egg white protein in terms of emulsifying activity at pH5, foamability at pH3 and 3% NaCl, and foam stability at pH9 were found. Consequently, MLPC offers a nutritious protein source with good functional properties at certain conditions, which could be used as food ingredient in a variety of food systems. Copyright © 2018 Elsevier Ltd. All rights reserved.

Mechanistic Studies of Human Spermine Oxidase: Kinetic Mechanism and pH Effects†

PubMed Central

Adachi, Maria S.; Juarez, Paul R.; Fitzpatrick, Paul F.

2009-01-01

In mammalian cells, the flavoprotein spermine oxidase (SMO) catalyzes the oxidation of spermine to spermidine and 3-aminopropanal. Mechanistic studies have been carried out with the recombinant human enzyme. The initial velocity pattern when the ratio between the concentrations of spermine and oxygen is kept constant establishes the steady-state kinetic pattern as ping-pong. Reduction of SMO by spermine in the absence of oxygen is biphasic. The rate constant for the rapid phase varies with the substrate concentration, with a limiting value (k3) of 49 s−1 and an apparent Kd value of 48 µM at pH 8.3. The rate constant for the slow step is independent of the spermine concentration, with a value of 5.5 s−1, comparable to the kcat value of 6.6 s−1. The kinetics of the oxidative half-reaction depend on the aging time after spermine and enzyme are mixed in a double mixing experiment. At an aging time of 6 s the reaction is monophasic with a second order rate constant of 4.2 mM−1 s−1. At an aging time of 0.3 s the reaction is biphasic with two second order constants equal to 4.0 and 40 mM−1 s−1. Neither is equal to the kcat/KO2 value of 13 mM−1s−1. These results establish the existence of more than one pathway for the reaction of the reduced flavin intermediate with oxygen. The kcat/KM value for spermine exhibits a bell-shaped pH-profile, with an average pKa value of 8.3. This profile is consistent with the active form of spermine having three charged nitrogens. The pH profile for k3 shows a pKa value of 7.4 for a group that must be unprotonated. The pKi-pH profiles for the competitive inhibitors N,N’-dibenzylbutane-1,4-diamine and spermidine show that the fully protonated forms of the inhibitors and the unprotonated form of an amino acid residue with a pKa of about 7.4 in the active site are preferred for binding. PMID:20000632

Surface charge engineering of a Bacillus gibsonii subtilisin protease.

PubMed

Jakob, Felix; Martinez, Ronny; Mandawe, John; Hellmuth, Hendrik; Siegert, Petra; Maurer, Karl-Heinz; Schwaneberg, Ulrich

2013-08-01

In proteins, a posttranslational deamidation process converts asparagine (Asn) and glutamine (Gln) residues into negatively charged aspartic (Asp) and glutamic acid (Glu), respectively. This process changes the protein net charge affecting enzyme activity, pH optimum, and stability. Understanding the principles which affect these enzyme properties would be valuable for protein engineering in general. In this work, three criteria for selecting amino acid substitutions of the deamidation type in the Bacillus gibsonii alkaline protease (BgAP) are proposed and systematically studied in their influence on pH-dependent activity and thermal resistance. Out of 113 possible surface amino acids, 18 (11 Asn and 7 Gln) residues of BgAP were selected and evaluated based on three proposed criteria: (1) The Asn or Gln residues should not be conserved, (2) should be surface exposed, and (3) neighbored by glycine. "Deamidation" in five (N97, N253, Q37, Q200, and Q256) out of eight (N97, N154, N250, N253, Q37, Q107, Q200, and Q256) amino acids meeting all criteria resulted in increased proteolytic activity. In addition, pH activity profiles of the variants N253D and Q256E and the combined variant N253DQ256E were dramatically shifted towards higher activity at lower pH (range of 8.5-10). Variant N253DQ256E showed twice the specific activity of wild-type BgAP and its thermal resistance increased by 2.4 °C at pH 8.5. These property changes suggest that mimicking surface deamidation by substituting Gln by Glu and/or Asn by Asp might be a simple and fast protein reengineering approach for modulating enzyme properties such as activity, pH optimum, and thermal resistance.

Enzymatic properties of the glycine D-alanine [corrected] aminopeptidase of Aspergillus oryzae and its activity profiles in liquid-cultured mycelia and solid-state rice culture (rice koji).

PubMed

Marui, Junichiro; Matsushita-Morita, Mayumi; Tada, Sawaki; Hattori, Ryota; Suzuki, Satoshi; Amano, Hitoshi; Ishida, Hiroki; Yamagata, Youhei; Takeuchi, Michio; Kusumoto, Ken-Ichi

2012-01-01

The gdaA gene encoding S12 family glycine-D-alanine aminopeptidase (GdaA) was found in the industrial fungus Aspergillus oryzae. GdaA shares 43% amino acid sequence identity with the D-aminopeptidase of the Gram-negative bacterium Ochrobactrum anthropi. GdaA purified from an A. oryzae gdaA-overexpressing strain exhibited high D-stereospecificity and efficiently released N-terminal glycine and D-alanine of substrates in a highly specific manner. The optimum pH and temperature were 8 to 9 and 40°C, respectively. This enzyme was stable under alkaline conditions at pH 8 to 11 and relatively resistant to acidic conditions until pH 5.0. The chelating reagent EDTA, serine protease inhibitors such as AEBSF, benzamidine, TPCK, and TLCK, and the thiol enzyme inhibitor PCMB inhibited the enzyme. The aminopeptidase inhibitor bestatin did not affect the activity. GdaA was largely responsible for intracellular glycine and D-alanine aminopeptidase activities in A. oryzae during stationary-phase growth in liquid media. In addition, the activity increased in response to the depletion of nitrogen or carbon sources in the growth media, although the GdaA-independent glycine aminopeptidase activity highly increased simultaneously. Aminopeptidases of A. oryzae attract attention because the enzymatic release of a variety of amino acids and peptides is important for the enhancement of the palatability of fermented foods. GdaA activity was found in extracts of a solid-state rice culture of A. oryzae (rice koji), which is widely used as a starter culture for Japanese traditional fermented foods, and was largely responsible for the glycine and D-alanine aminopeptidase activity detected at a pH range of 6 to 9.

Nanoparticle-stabilized liposomes for pH-responsive gastric drug delivery.

PubMed

Thamphiwatana, Soracha; Fu, Victoria; Zhu, Jingying; Lu, Diannan; Gao, Weiwei; Zhang, Liangfang

2013-10-01

We report a novel pH-responsive gold nanoparticle-stabilized liposome system for gastric antimicrobial delivery. By adsorbing small chitosan-modified gold nanoparticles (diameter ~10 nm) onto the outer surface of negatively charged phospholipid liposomes (diameter ~75 nm), we show that at gastric pH the liposomes have excellent stability with limited fusion ability and negligible cargo releases. However, when the stabilized liposomes are present in an environment with neutral pH, the gold stabilizers detach from the liposomes, resulting in free liposomes that can actively fuse with bacterial membranes. Using Helicobacter pylori as a model bacterium and doxycycline as a model antibiotic, we demonstrate such pH-responsive fusion activity and drug release profile of the nanoparticle-stabilized liposomes. Particularly, at neutral pH the gold nanoparticles detach, and thus the doxycycline-loaded liposomes rapidly fuse with bacteria and cause superior bactericidal efficacy as compared to the free doxycycline counterpart. Our results suggest that the reported liposome system holds a substantial potential for gastric drug delivery; it remains inactive (stable) in the stomach lumen but actively interacts with bacteria once it reaches the mucus layer of the stomach where the bacteria may reside.

Deep-Sea DuraFET: A Pressure Tolerant pH Sensor Designed for Global Sensor Networks.

PubMed

Johnson, Kenneth S; Jannasch, Hans W; Coletti, Luke J; Elrod, Virginia A; Martz, Todd R; Takeshita, Yuichiro; Carlson, Robert J; Connery, James G

2016-03-15

Increasing atmospheric carbon dioxide is driving a long-term decrease in ocean pH which is superimposed on daily to seasonal variability. These changes impact ecosystem processes, and they serve as a record of ecosystem metabolism. However, the temporal variability in pH is observed at only a few locations in the ocean because a ship is required to support pH observations of sufficient precision and accuracy. This paper describes a pressure tolerant Ion Sensitive Field Effect Transistor pH sensor that is based on the Honeywell Durafet ISFET die. When combined with a AgCl pseudoreference sensor that is immersed directly in seawater, the system is capable of operating for years at a time on platforms that cycle from depths of several km to the surface. The paper also describes the calibration scheme developed to allow calibrated pH measurements to be derived from the activity of HCl reported by the sensor system over the range of ocean pressure and temperature. Deployments on vertical profiling platforms enable self-calibration in deep waters where pH values are stable. Measurements with the sensor indicate that it is capable of reporting pH with an accuracy of 0.01 or better on the total proton scale and a precision over multiyear periods of 0.005. This system enables a global ocean observing system for ocean pH.

Spatial patterns of soil pH and the factors that influence them in plantation forests of northern China

NASA Astrophysics Data System (ADS)

Hong, Songbai; Liu, Yongwen; Piao, Shilong

2017-04-01

Climate and anthropogenic activities such as afforestation and nitrogen deposition all impact soil pH. Understanding the spatial pattern of soil pH and the factors that influence it can provide basic information for generating appropriate strategies for soil resource management and protection, especially in light of increasing anthropogenic influences and climate change. In this study, we investigated the spatial and vertical pattern of soil pH and evaluated the influence of climate and nitrogen deposition using 1647 soil profiles 1 meter in depth from 549 plots in plantation forests of northern China. We found that soil pH decreased from the southwest to the northeast in the study region and had a similar spatial pattern before and after afforestation. Furthermore, our results show that climate and nitrogen deposition fundamentally influence the pattern of soil pH. Specifically, increasing precipitation significantly decreased soil pH (with a mean rate of 0.3 for every 100 mm rainfall, p<0.001), whereas increasing temperature significantly increased soil pH (0.13 for every degree centigrade, p<0.001). Nitrogen deposition, especially nitrate nitrogen, significantly decreased soil pH (p<0.01). All these factors impact soil pH directly and indirectly through climate-plant-soil interactions. As the risks from both climate change and nitrogen deposition increase, there is an urgent need to further understanding of soil pH dynamics and to develop informed policies to protect soil resources.

Purification of an Exopolygalacturonase from Penicillium viridicatum RFC3 Produced in Submerged Fermentation

PubMed Central

Gomes, Eleni; Leite, Rodrigo Simões Ribeiro; da Silva, Roberto; Silva, Dênis

2009-01-01

An exo-PG obtained from Penicillium viridicatum in submerged fermentation was purified to homogeneity. The apparent molecular weight of the enzyme was 92 kDa, optimum pH and temperature for activity were pH 5 and 50–55°C. The exo-PG showed a profile of an exo-polygalacturonase, releasing galacturonic acid by hydrolysis of pectin with a high degree of esterification (D.E.). Ions Ca2+ enhanced the stability of enzyme and its activity by 30%. The K m was 1.30 in absence of Ca2+ and 1.16 mg mL−1 in presence of this ion. In relation to the V max the presence of this ion increased from 1.76 to 2.07 μmol min−1mg−1. PMID:20148174

Expression, purification, and kinetic characterization of full-length human fibroblast activation protein.

PubMed

Sun, Shaoxian; Albright, Charles F; Fish, Barbara H; George, Henry J; Selling, Bernard H; Hollis, Gregory F; Wynn, Richard

2002-03-01

Human fibroblast activation protein (FAP), an integral membrane serine protease, was produced in insect cells as a hexa-His-tagged protein using a recombinant baculovirus expression system. Two isoforms of FAP, glycosylated and nonglycosylated, were identified by Western blotting using an anti-His-tag antibody and separated by lectin chromatography. The glycosylated FAP was purified to near homogeneity using immobilized metal affinity chromatography and was shown to have both postprolyl dipeptidyl peptidase and postgelatinase activities. In contrast, the nonglycosylated isoform demonstrated no detectable gelatinase activity by either zymography or a fluorescence-based gelatinase activity assay. The kinetic parameters of the dipeptidyl peptidase activity for glycosylated FAP were determined using dipeptide Ala-Pro-7-amino-trifluoromethyl-coumarin as the substrate. The k(cat) is 2.0 s(-1) and k(cat)/K(m) is 1.0 x 10(4) M(-1) s(-1) at pH 8.5. The pH dependence of k(cat) reveals two ionization groups with pK(a1) of 7.0 and pK(a2) of 11.0. The pH profile of k(cat)/K(m) yields similar results with pK(a1) 6.2 and pK(a2) 11.0. The neutral pK(a1) is associated with His at the active site. The basic pK(a2) might be contributed from an ionization group that is not involved directly in catalysis, instead associated with the stability of the active site structure. Copyright 2002 Elsevier Science (USA).

Characterization and ontogenetic development of digestive enzymes in Pacific bluefin tuna Thunnus orientalis larvae.

PubMed

Murashita, Koji; Matsunari, Hiroyuki; Kumon, Kazunori; Tanaka, Yosuke; Shiozawa, Satoshi; Furuita, Hirofumi; Oku, Hiromi; Yamamoto, Takeshi

2014-12-01

The major digestive enzymes in Pacific bluefin tuna Thunnus orientalis larvae were characterized, and the physiological characteristics of the enzymes during early ontogeny were clarified using biochemical and molecular approaches. The maximum activity of trypsin (Try), chymotrypsin (Ct) and amylase (Amy) was observed at pH 6-11, 8-11 and 6-9, respectively. Maximum activity of Try, Ct and Amy occurred at 50 °C, that of lipase (Lip) was at 60 °C and that of pepsin (Pep) was at 40-50 °C. These pH and thermal profiles were similar to those for other fish species but differed from those previously reported for adult bluefin tuna. Enzyme activity for all enzymes assayed was found to decrease at high temperatures (Try, Ct, Amy and Pep: 50 °C; Lip: 40 °C), which is similar to findings for other fish species with one marked exception-increased Try activity was observed at 40 °C. Lip activity appeared to be dependent on bile salts under our assay conditions, resulting in a significant increase in activity in the presence of bile salts. Ontogenetic changes in pancreatic digestive enzymes showed similar gene expression patterns to those of other fish species, whereas marked temporal increases in enzyme activities were observed at 10-12 days post hatching (dph), coinciding with previously reported timing of the development of the pyloric caeca in bluefin tuna larvae. However, complete development of digestive function was indicated by the high pep gene expression from 19 dph, which contradicts the profile of Pep activity and previously reported development timing of the gastric gland. These findings contribute to the general knowledge of bluefin tuna larval digestive system development.

Intensive cytokine induction in pandemic H1N1 influenza virus infection accompanied by robust production of IL-10 and IL-6.

PubMed

Yu, Xuelian; Zhang, Xi; Zhao, Baihui; Wang, Jiayu; Zhu, Zhaokui; Teng, Zheng; Shao, Junjie; Shen, Jiaren; Gao, Ye; Yuan, Zhengan; Wu, Fan

2011-01-01

The innate immune system is the first line of defense against viruses by inducing expression of cytokines and chemokines. Many pandemic influenza H1N1 virus [P(H1N1)] infected severe cases occur in young adults under 18 years old who were rarely seriously affected by seasonal influenza. Results regarding host cytokine profiles of P(H1N1) are ambivalent. In the present study we investigated host cytokine profiles in P(H1N1) patients and identified cytokines related to disease severity. We retrieved 77, 59, 26 and 26 sera samples from P(H1N1) and non-flu influenza like illness (non-ILIs) cases with mild symptoms (mild patients), P(H1N1) vaccinees and healthy individuals, respectively. Nine and 16 sera were from hospitalized P(H1N1) and non-ILIs patients with severe symptoms (severe patients). Cytokines of IL-1, IL-2, IL-4, IL-5, IL-6, IL-8, IL-10, IL-12, IFN-γ and TNF-α were assayed by cytokine bead array, IL-17 and IL-23 measured with ELISA. Mild P(H1N1) patients produced significantly elevated IL-2, IL-12, IFN-γ, IL-6, TNF-α, IL-5, IL-10, IL-17 and IL-23 versus to healthy controls. While an overwhelming IL-6 and IL-10 production were observed in severe P(H1N1) patients. Higher IL-10 secretion in P(H1N1) vaccinees confirmed our observation that highly increased level of sera IL-6 and IL-10 in P(H1N1) patients may lead to disease progression. A comprehensive innate immune response was activated at the early stage of P(H1N1) infection with a combine Th1/Th2/Th3 cytokines production. As disease progression, a systemic production of IL-6 and IL-10 were observed in severe P(H1N1) patients. Further analysis found a strong correlation between IL-6 and IL-10 production in the severe P(H1N1) patients. IL-6 may be served as a mediator to induce IL-10 production. Highly elevated level of sera IL-6 and IL-10 in P(H1N1) patients may lead to disease progression, but the underlying mechanism awaits further detailed investigations.

Intensive Cytokine induction in Pandemic H1N1 Influenza Virus Infection Accompanied by Robust Production of IL-10 and IL-6

PubMed Central

Yu, Xuelian; Zhang, Xi; Zhao, Baihui; Wang, Jiayu; Zhu, Zhaokui; Teng, Zheng; Shao, Junjie; Shen, Jiaren; Gao, Ye; Yuan, Zhengan; Wu, Fan

2011-01-01

Background The innate immune system is the first line of defense against viruses by inducing expression of cytokines and chemokines. Many pandemic influenza H1N1 virus [P(H1N1)] infected severe cases occur in young adults under 18 years old who were rarely seriously affected by seasonal influenza. Results regarding host cytokine profiles of P(H1N1) are ambivalent. In the present study we investigated host cytokine profiles in P(H1N1) patients and identified cytokines related to disease severity. Methods and Principal Findings We retrieved 77, 59, 26 and 26 sera samples from P(H1N1) and non-flu influenza like illness (non-ILIs) cases with mild symptoms (mild patients), P(H1N1) vaccinees and healthy individuals, respectively. Nine and 16 sera were from hospitalized P(H1N1) and non-ILIs patients with severe symptoms (severe patients). Cytokines of IL-1, IL-2, IL-4, IL-5, IL-6, IL-8, IL-10, IL-12, IFN-γ and TNF-α were assayed by cytokine bead array, IL-17 and IL-23 measured with ELISA. Mild P(H1N1) patients produced significantly elevated IL-2, IL-12, IFN-γ, IL-6, TNF-α, IL-5, IL-10, IL-17 and IL-23 versus to healthy controls. While an overwhelming IL-6 and IL-10 production were observed in severe P(H1N1) patients. Higher IL-10 secretion in P(H1N1) vaccinees confirmed our observation that highly increased level of sera IL-6 and IL-10 in P(H1N1) patients may lead to disease progression. Conclusion and Significance A comprehensive innate immune response was activated at the early stage of P(H1N1) infection with a combine Th1/Th2/Th3 cytokines production. As disease progression, a systemic production of IL-6 and IL-10 were observed in severe P(H1N1) patients. Further analysis found a strong correlation between IL-6 and IL-10 production in the severe P(H1N1) patients. IL-6 may be served as a mediator to induce IL-10 production. Highly elevated level of sera IL-6 and IL-10 in P(H1N1) patients may lead to disease progression, but the underlying mechanism awaits further detailed investigations. PMID:22174866

Physicochemical profile of microbial-assisted composting on empty fruit bunches of oil palm trees.

PubMed

Lim, Li Yee; Bong, Cassendra Phun Chien; Chua, Lee Suan; Lee, Chew Tin

2015-12-01

This study was carried out to investigate the physicochemical properties of compost from oil palm empty fruit bunches (EFB) inoculated with effective microorganisms (EM∙1™). The duration of microbial-assisted composting was shorter (∼7 days) than control samples (2 months) in a laboratory scale (2 kg) experiment. The temperature profile of EFB compost fluctuated between 26 and 52 °C without the presence of consistent thermophilic phase. The pH of compost changed from weak acidic (pH ∼5) to mild alkaline (pH ∼8) because of the formation of nitrogenous ions such as ammonium (NH4 (+)), nitrite (NO2 (-)), and nitrate (NO3 (-)) from organic substances during mineralization. The pH of the microbial-treated compost was less than 8.5 which is important to prevent the loss of nitrogen as ammonia gas in a strong alkaline condition. Similarly, carbon mineralization could be determined by measuring CO2 emission. The microbial-treated compost could maintain longer period (∼13 days) of high CO2 emission resulted from high microbial activity and reached the threshold value (120 mg CO2-C kg(-1) day(-1)) for compost maturity earlier (7 days). Microbial-treated compost slightly improved the content of minerals such as Mg, K, Ca, and B, as well as key metabolite, 5-aminolevulinic acid for plant growth at the maturity stage of compost. Graphical Abstract Microbial-assisted composting on empty fruit bunches.

Interpretation of pH-activity profiles for acid-base catalysis from molecular simulations.

PubMed

Dissanayake, Thakshila; Swails, Jason M; Harris, Michael E; Roitberg, Adrian E; York, Darrin M

2015-02-17

The measurement of reaction rate as a function of pH provides essential information about mechanism. These rates are sensitive to the pK(a) values of amino acids directly involved in catalysis that are often shifted by the enzyme active site environment. Experimentally observed pH-rate profiles are usually interpreted using simple kinetic models that allow estimation of "apparent pK(a)" values of presumed general acid and base catalysts. One of the underlying assumptions in these models is that the protonation states are uncorrelated. In this work, we introduce the use of constant pH molecular dynamics simulations in explicit solvent (CpHMD) with replica exchange in the pH-dimension (pH-REMD) as a tool to aid in the interpretation of pH-activity data of enzymes and to test the validity of different kinetic models. We apply the methods to RNase A, a prototype acid-base catalyst, to predict the macroscopic and microscopic pK(a) values, as well as the shape of the pH-rate profile. Results for apo and cCMP-bound RNase A agree well with available experimental data and suggest that deprotonation of the general acid and protonation of the general base are not strongly coupled in transphosphorylation and hydrolysis steps. Stronger coupling, however, is predicted for the Lys41 and His119 protonation states in apo RNase A, leading to the requirement for a microscopic kinetic model. This type of analysis may be important for other catalytic systems where the active forms of the implicated general acid and base are oppositely charged and more highly correlated. These results suggest a new way for CpHMD/pH-REMD simulations to bridge the gap with experiments to provide a molecular-level interpretation of pH-activity data in studies of enzyme mechanisms.

Interpretation of pH-activity Profiles for Acid-Base Catalysis from Molecular Simulations

PubMed Central

Dissanayake, Thakshila; Swails, Jason; Harris, Michael E.; Roitberg, Adrian E.; York, Darrin M.

2015-01-01

The measurement of reaction rate as a function of pH provides essential information about mechanism. These rates are sensitive to the pKa values of amino acids directly involved in catalysis that are often shifted by the enzyme active site environment. Experimentally observed pH-rate profiles are usually interpreted using simple kinetic models that allow estimation of “apparent pKa” values of presumed general acid and base catalysts. One of the underlying assumptions in these models is that the protonation states are uncorrelated. In the present work, we introduce the use of constant pH molecular dynamics simulations in explicit solvent (CpHMD) with replica exchange in the pH-dimension (pH-REMD) as a tool to aid in the interpretation of pH-activity data of enzymes, and test the validity of different kinetic models. We apply the methods to RNase A, a prototype acid/base catalyst, to predict the macroscopic and microscopic pKa values, as well as the shape of the pH-rate profile. Results for apo and cCMP-bound RNase A agree well with available experimental data, and suggest that deprotonation of the general acid and protonation of the general base are not strongly coupled in transphosphorylation and hydrolysis steps. Stronger coupling, however, is predicted for the Lys41 and His119 protonation states in apo RNase A, leading to the requirement for a microscopic kinetic model. This type of analysis may be important for other catalytic systems where the active forms of implicated general acid and base are oppositely charged and more highly correlated. These results suggest a new way for CpHMD/pH-REMD simulations to bridge the gap with experiments to provide a molecular-level interpretation of pH-activity data in studies of enzyme mechanisms. PMID:25615525

Production of Pectate Lyase by Penicillium viridicatum RFC3 in Solid-State and Submerged Fermentation

PubMed Central

Ferreira, Viviani; da Silva, Roberto; Silva, Dênis; Gomes, Eleni

2010-01-01

Pectate lyase (PL) was produced by the filamentous fungus Penicillium viridicatum RFC3 in solid-state cultures of a mixture of orange bagasse and wheat bran (1 : 1 w/w), or orange bagasse, wheat bran and sugarcane bagasse (1 : 1 : 0.5 w/w), and in a submerged liquid culture with orange bagasse and wheat bran (3%) as the carbon source. PL production was highest (1,500 U  mL−1 or 300 Ug−1 of substrate) in solid-state fermentation (SSF) on wheat bran and orange bagasse at 96 hours. PL production in submerged fermentation (SmF) was influenced by the initial pH of the medium. With the initial pH adjusted to 4.5, 5.0, and 5.5, the peak activity was observed after 72, 48, and 24 hours of fermentation, respectively, when the pH of the medium reached the value 5.0. PL from SSF and SmF were loaded on Sephadex-G75 columns and six activity peaks were obtained from crude enzyme from SSF and designated PL I, II, III, IV, V, and VI, while five peaks were obtained from crude enzyme from SmF and labeled PL  I′, II′, III′, IV′, and VII′. Crude enzyme and fraction III from each fermentative process were tested further. The optimum pH for crude PL from either process was 5.5, while that for PL III was 8.0. The maximum activity of enzymes from SSF was observed at 35°C, but crude enzyme was more thermotolerant than PL III, maintaining its maximum activity up to 45°C. Crude enzyme from SmF and PL III′ showed thermophilic profiles of activity, with maximum activity at 60 and 55°C, respectively. In the absence of substrate, the crude enzyme from SSF was stable over the pH range 3.0–10.0 and PL III was most stable in the pH range 4.0–7.0. Crude enzyme from SmF retained 70%–80% of its maximum activity in the acid-neutral pH range (4.0–7.0), but PIII showed high stability at alkaline pH (7.5–9.5). PL from SSF was more thermolabile than that from SmF. The latter maintained 60% of its initial activity after 1 h at 55°C. The differing behavior of the enzymes with respect to pH and temperature suggests that they are different isozymes. PMID:20689719

Kinetics and equilibrium modelling of lead uptake by algae Gelidium and algal waste from agar extraction industry.

PubMed

Vilar, Vítor J P; Botelho, Cidália M S; Boaventura, Rui A R

2007-05-08

Pb(II) biosorption onto algae Gelidium, algal waste from agar extraction industry and a composite material was studied. Discrete and continuous site distribution models were used to describe the biosorption equilibrium at different pH (5.3, 4 and 3), considering competition among Pb(II) ions and protons. The affinity distribution function of Pb(II) on the active sites was calculated by the Sips distribution. The Langmuir equilibrium constant was compared with the apparent affinity calculated by the discrete model, showing higher affinity for lead ions at higher pH values. Kinetic experiments were conducted at initial Pb(II) concentrations of 29-104 mgl(-1) and data fitted to pseudo-first Lagergren and second-order models. The adsorptive behaviour of biosorbent particles was modelled using a batch mass transfer kinetic model, which successfully predicts Pb(II) concentration profiles at different initial lead concentration and pH, and provides significant insights on the biosorbents performance. Average values of homogeneous diffusivity, D(h), are 3.6 x 10(-8); 6.1 x 10(-8) and 2.4 x 10(-8)cm(2)s(-1), respectively, for Gelidium, algal waste and composite material. The concentration of lead inside biosorbent particles follows a parabolic profile that becomes linear near equilibrium.

Release Profile of Andrographis paniculata Leaf Extract Nanocapsule as α-Glucosidase Inhibitors

NASA Astrophysics Data System (ADS)

Zahrani, K.; Imansari, F.; Utami, T. S.; Arbianti, R.

2017-07-01

Andrographis paniculata is one of 13 leading commodities Indonesian medicinal plants through the Ditjen POM. Andrographolide as main active compound has been shown to have many pharmacological activities, one of which is as α-glucosidase enzyme inhibitors which has clinical potential as an antitumor, antiviral, antidiabetic, and immunoregulator agents. This study aims to do nanoencapsulation of Andrographis paniculatar leaf extract to increase its active compound bioavailability and get a release profile through synthetic fluids media simulation. Nanoencapsulation with ionic gelation method result the encapsulation efficiency and loading capacity values of 73.47% and 46.29% at 2%: 1% of chitosan: STPP ratio. The maximum α-glucosidase inhibition of 37.17% was obtained at 16% concentration. Burst release at gastric pH conditions indicate that most of the drug (in this study is an Andrographis paniculata leaf extract) adsorbed on the surface of the nanoparticles an indicates that the kind of nanoparticle formed is nanosphere.

Isolation, Characterization, Molecular Gene Cloning, and Sequencing of a Novel Phytase from Bacillus subtilis

PubMed Central

Kerovuo, Janne; Lauraeus, Marko; Nurminen, Päivi; Kalkkinen, Nisse; Apajalahti, Juha

1998-01-01

The Bacillus subtilis strain VTT E-68013 was chosen for purification and characterization of its excreted phytase. Purified enzyme had maximal phytase activity at pH 7 and 55°C. Isolated enzyme required calcium for its activity and/or stability and was readily inhibited by EDTA. The enzyme proved to be highly specific since, of the substrates tested, only phytate, ADP, and ATP were hydrolyzed (100, 75, and 50% of the relative activity, respectively). The phytase gene (phyC) was cloned from the B. subtilis VTT E-68013 genomic library. The deduced amino acid sequence (383 residues) showed no homology to the sequences of other phytases nor to those of any known phosphatases. PhyC did not have the conserved RHGXRXP sequence found in the active site of known phytases, and therefore PhyC appears not to be a member of the phytase subfamily of histidine acid phosphatases but a novel enzyme having phytase activity. Due to its pH profile and optimum, it could be an interesting candidate for feed applications. PMID:9603817

Differential Gene Expression of Longan Under Simulated Acid Rain Stress.

PubMed

Zheng, Shan; Pan, Tengfei; Ma, Cuilan; Qiu, Dongliang

2017-05-01

Differential gene expression profile was studied in Dimocarpus longan Lour. in response to treatments of simulated acid rain with pH 2.5, 3.5, and a control (pH 5.6) using differential display reverse transcription polymerase chain reaction (DDRT-PCR). Results showed that mRNA differential display conditions were optimized to find an expressed sequence tag (EST) related with acid rain stress. The potential encoding products had 80% similarity with a transcription initiation factor IIF of Gossypium raimondii and 81% similarity with a protein product of Theobroma cacao. This fragment is the transcription factor activated by second messenger substances in longan leaves after signal perception of acid rain.

Pathways for external alkalinization in intact and in microwounded Chara cells are differentially sensitive to wortmannin

PubMed Central

Bulychev, Alexander A.; Foissner, Ilse

2017-01-01

ABSTRACT Proton flows across the plant cell membranes play a major role in electrogenesis and regulation of photosynthesis and ion balance. The profiles of external pH along the illuminated internodal cells of characean algae consist of alternating high- and low-pH zones that are spatially coordinated with the distribution of photosynthetic activity of chloroplasts underlying these zones. The results based on confocal laser scanning fluorescence microscopy, pH microsensors, and pulse-amplitude-modulated chlorophyll microfluorometry revealed that the coordination of H+ transport and photosynthesis is disrupted by the 2 different environmental cues (low light and wounding) and by a chemical, wortmannin interfering with the inositol phospholipid metabolism. On the one hand, the transition from moderate to low irradiance diminished the peaks in the profiles of photosystem II (PSII) quantum efficiency but did not remove the pH bands. On the other hand, the microwounding of the internode with a glass micropipette, impacting primarily the cell wall, resulted in a rapid local alkalinization of the external medium (by 2–2.5 pH units) near the cell surface, thus mimicking the appearance of natural pH bands. Despite their seeming similarity, the alkaline bands of intact cells were eliminated by wortmannin, whereas the wound-induced alkalinization was insensitive to this drug. Furthermore, the attenuation of natural pH bands in wortmannin-treated cells was accompanied by the enhancement in spatial heterogeneity of PSII efficiency and electron transport rates, which indicates the complexity of chloroplast–plasma membrane interactions. The results suggest that the light- and wound-induced alkaline areas on the cell surface are associated with different ion-transport systems. PMID:28805493

Ammonia oxidisers in a non-nitrifying Brazilian savanna soil.

PubMed

Catão, Elisa C P; Thion, Cécile; Krüger, R H; Prosser, James I

2017-11-01

Low nitrification rates in Brazilian savanna (Cerrado) soils have puzzled researchers for decades. Potential mechanisms include biological inhibitors, low pH, low microbial abundance and low soil moisture content, which hinders microbial activity, including ammonia oxidation. Two approaches were used to evaluate these potential mechanisms: (i) manipulation of soil moisture and pH in microcosms containing Cerrado soil and (ii) assessment of nitrification inhibition in slurries containing mixtures of Cerrado soil and an actively nitrifying agricultural soil. Despite high ammonium concentration in Cerrado soil microcosms, little NO3- accumulation was observed with increasing moisture or pH, but in some Cerrado soil slurries, ammonia-oxidising archaea (AOA) amoA transcripts were detected after 14 days. In mixed soil slurries, the final NO3- concentration reflected the initial proportions of agricultural and Cerrado soils in the mixture, providing no evidence of nitrification inhibitors in Cerrado soil. AOA community denaturing gradient gel electrophoresis profiles were similar in the mixed and nitrifying soils. These results suggest that nitrification in Cerrado soils is not constrained by water availability, ammonium availability, low pH or biological inhibitors, and alternative potential explanations for low nitrification levels are discussed. © FEMS 2017. All rights reserved. For permissions, please e-mail: journals.permissions@oup.com.

Optimization and modeling of laccase production by Trametes versicolor in a bioreactor using statistical experimental design.

PubMed

Tavares, A P M; Coelho, M A Z; Agapito, M S M; Coutinho, J A P; Xavier, A M R B

2006-09-01

Experimental design and response surface methodologies were applied to optimize laccase production by Trametes versicolor in a bioreactor. The effects of three factors, initial glucose concentration (0 and 9 g/L), agitation (100 and 180 rpm), and pH (3.0 and 5.0), were evaluated to identify the significant effects and its interactions in the laccase production. The pH of the medium was found to be the most important factor, followed by initial glucose concentration and the interaction of both factors. Agitation did not seem to play an important role in laccase production, nor did the interaction agitation x medium pH and agitation x initial glucose concentration. Response surface analysis showed that an initial glucose concentration of 11 g/L and pH controlled at 5.2 were the optimal conditions for laccase production by T. versicolor. Under these conditions, the predicted value for laccase activity was >10,000 U/L, which is in good agreement with the laccase activity obtained experimentally (11,403 U/L). In addition, a mathematical model for the bioprocess was developed. It is shown that it provides a good description of the experimental profile observed, and that it is capable of predicting biomass growth based on secondary process variables.

Measurements of pH and redox potential distributions in TNT-contaminated plant-soil systems using microelectrode techniques

DOE Office of Scientific and Technical Information (OSTI.GOV)

Pang, H.; Zhang, T.C.

1997-12-31

The pH and redox potential profiles in TNT-contaminated soils with and without plants were investigated using microelectrode techniques. The new pH cocktail and double-barreled structure greatly improved the performance of the pH microelectrode. For soil without plants, there is almost no pH difference at different locations with different heights; while for the TNT-contaminated soils with plants there exist pH profiles. The soil immediately near the root of the plant has the lowest pH value. The pH value increases as the distance between the measuring point and the plant roots increases. The pH gradient (the increased pH value over the unitmore » distance) decreases with an increase of the distance between the measuring point and the plant roots. These results show that the plant presence can greatly affect the pH distribution. In vegetated soil, the redox potentials in the layer nearest the plant roots are higher than those in the bulk soil without plants. The redox potentials in the central part of the plant are lower than those in the soil around the plant and soil without the plant. The redox potentials in the soil without plants decrease with an increase of depth.« less

Functional expression of the catalytic domains of two cysteine proteinases from Trypanosoma congolense.

PubMed

Boulangé, A; Serveau, C; Brillard, M; Minet, C; Gauthier, F; Diallo, A; Lalmanach, G; Authié, E

2001-11-01

The catalytic domains of two closely related cysteine proteinases (CP1 and CP2) from Trypanosoma congolense, referred to as C1 and C2, were expressed as proforms in Escherichia coli (C1) and in the baculovirus system (C1 and C2). While the bacterial expression system did not allow recovery of active C1, the baculovirus system led to secretion of inactive zymogens which could be processed at acidic pH into mature enzymes. Active C1 and C2 were purified from serum-free culture supernatants by anion-exchange chromatography and characterised. Their kinetic parameters and pH activity profiles confirmed the relatedness between C2 and native CP2 (congopain). These properties also underline major functional differences between C1 and C2, that appear to relate to discrete but essential sequence differences. It is likely that these two enzymes perform distinct roles in vivo, in the parasite and/or in the host-parasite relationships.

[Characteristics of soil pH and exchangeable acidity in red soil profile under different vegetation types].

PubMed

Ji, Gang; Xu, Ming-gang; Wen, Shi-lin; Wang, Bo-ren; Zhang, Lu; Liu, Li-sheng

2015-09-01

The characteristics of soil pH and exchangeable acidity in soil profile under different vegetation types were studied in hilly red soil regions of southern Hunan Province, China. The soil samples from red soil profiles within 0-100 cm depth at fertilized plots and unfertilized plots were collected and analyzed to understand the profile distribution of soil pH and exchangeable acidity. The results showed that, pH in 0-60 cm soil from the fertilized plots decreased as the following sequence: citrus orchard > Arachis hypogaea field > tea garden. As for exchangeable acidity content, the sequence was A. hypogaea field ≤ citrus orchard < tea garden. After tea tree and A. hypogaea were planted for long time, acidification occurred in surface soil (0-40 cm), compared with the deep soil (60-100 cm), and soil pH decreased by 0.55 and 0.17 respectively, but such changes did not occur in citrus orchard. Soil pH in 0-40 cm soil from the natural recovery vegetation unfertilized plots decreased as the following sequence: Imperata cylindrica land > Castanea mollissima garden > Pinus elliottii forest ≥ Loropetalum chinensis forest. As for exchangeable acidity content, the sequence was L cylindrica land < C. mollissima garden < L. chinensis forest ≤ P. elliottii forest. Soil pH in surface soil (0-20 cm) from natural forest plots, secondary forest and Camellia oleifera forest were significantly lower than that from P. massoniana forest, decreased by 0.34 and 0.20 respectively. For exchangeable acidity content in 0-20 cm soil from natural forest plot, P. massoniana forest and secondary forest were significantly lower than C. oleifera forest. Compared with bare land, surface soil acidification in unfertilized plots except I. cylindrica land had been accelerated, and the natural secondary forest was the most serious among them, with surface soil pH decreasing by 0.52. However, the pH increased in deep soils from unfertilized plots except natural secondary forest, and I. cylindrica land was the most obvious among them, with soil pH increasing by 0.43. The effects of fertilization and vegetation type on pH and exchangeable acidity decreased with the increasing soil depth from all plots.

Digital Presence of Norwegian Scholars on Academic Network Sites—Where and Who Are They?

PubMed Central

Gjesdal, Øyvind Liland; Al Ruwehy, Hemed Ali

2015-01-01

The use of academic profiling sites is becoming more common, and emerging technologies boost researchers’ visibility and exchange of ideas. In our study we compared profiles at five different profiling sites. These five sites are ResearchGate, Academia.edu, Google Scholar Citations, ResearcherID and ORCID. The data set is enriched by demographic information including age, gender, position and affiliation, which are provided by the national CRIS-system in Norway. We find that approximately 37% of researchers at the University of Bergen have at least one profile, the prevalence being highest (> 40%) for members at the Faculty of Psychology and the Faculty of Social Sciences. Across all disciplines, ResearchGate is the most widely used platform. However, within Faculty of Humanities, Academia.edu is the preferred one. Researchers are reluctant to maintain multiple profiles, and there is little overlap between different services. Age turns out to be a poor indicator for presence in the investigated profiling sites, women are underrepresented and professors together with PhD students are the most likely profile holders. We next investigated the correlation between bibliometric measures, such as publications and citations, and user activities, such as downloads and followers. We find different bibliometric indicators to correlate strongly within individual platforms and across platforms. There is however less agreement between the traditional bibliometric and social activity indicators. PMID:26565408

Blood tolerant laccase by directed evolution.

PubMed

Mate, Diana M; Gonzalez-Perez, David; Falk, Magnus; Kittl, Roman; Pita, Marcos; De Lacey, Antonio L; Ludwig, Roland; Shleev, Sergey; Alcalde, Miguel

2013-02-21

High-redox potential laccases are powerful biocatalysts with a wide range of applications in biotechnology. We have converted a thermostable laccase from a white-rot fungus into a blood tolerant laccase. Adapting the fitness of this laccase to the specific composition of human blood (above neutral pH, high chloride concentration) required several generations of directed evolution in a surrogate complex blood medium. Our evolved laccase was tested in both human plasma and blood, displaying catalytic activity while retaining a high redox potential at the T1 copper site. Mutations introduced in the second coordination sphere of the T1 site shifted the pH activity profile and drastically reduced the inhibitory effect of chloride. This proof of concept that laccases can be adapted to function in extreme conditions opens an array of opportunities for implantable nanobiodevices, chemical syntheses, and detoxification. Copyright © 2013 Elsevier Ltd. All rights reserved.

Metabolomic Investigation of Rat Serum Following Oral Administration of the Willow Bracket Medicinal Mushroom, Phellinus igniarius (Agaricomycetes), by UPLC-HDMS.

PubMed

Dong, Yu; He, Ying; Yu, Zhongming; Zhang, Yang; Wang, Nani; Shou, Dan; Li, Changyu

2016-01-01

The medicinal willow bracket mushroom, Phellinus igniarius, is a species that has been reported to possess antibacterial, antioxidative, antitumor, antidiabetic, and antihyperlipidemia activities. The aim of this study was to elucidate the changes in endogenous metabolites after oral administration of a decoction of Ph. Igniarius. Ultraperformance liquid chromatography (UPLC)/electrospray ionization synapt high-definition mass spectrometry (ESI-HDMS) combined with pattern recognition approaches, including principal component analysis and orthogonal partial least squares discriminant analysis, were integrated to discover differentiating metabolites. The current metabolomics approach identified 16 ions (5 in the negative mode, 11 in the positive mode) as "differentiating metabolites". The results illustrated that Ph. Igniarius is likely to increase the biosynthesis and secretion of bile acids that provide hypolipidemic activity and showed that robust UPLC/ESI-HDMS techniques are promising for profiling analysis of medicinal mushroom metabolites.

The relationship between the electrochemical proton gradient and active transport in Escherichia coli membrane vesicles.

PubMed

Ramos, S; Kaback, H R

1977-03-08

In the previous paper [ramos, S., and Kaback, H.R. (1977), Biochemistry 16 (preceding paper in this issue)], it was demonstrated that Escherichia coli membrane vesicles generate a large electrochemical proton gradient (delta-muH+) under appropriate conditions, and some of the properties of delta-muH+ and its component forces [i.e., the membrane potential (delta psi) and the chemical gradient of protons (deltapH)] were described. In this paper, the relationship between delta-muH+, delta psi, and deltapH and the active transport of specific solutes is examined. Addition of lactose or glucose 6-phosphate to membrane vesicles containing the appropriate transport systems results in partial collapse of deltapH, providing direct evidence for the suggestion that respiratory energy can drive active transport via the pH gradient across the membrane. Titration studies with valinomycin and nigericin lead to the conclusion that, at pH 5.5, there are two general classes of transport systems: those that are driven primarily by delta-muH+ (lactose, proline, serine, glycine, tyrosine, glutamate, leucine, lysine, cysteine, and succinate) and those that are driven primarily by deltapH (glucose 6-phosphate, D-lactate, glucuronate, and gluconate). Importantly, however, it is also demonstrated that at pH 7.5, all of these transport systems are driven by delta psi which comprises the only component of delta-muH+ at this external pH. In addition, the effect of external pH on the steady-state levels of accumulation of different solutes is examined, and it is shown that none of the pH profiles correspond to those observed for delta-muH+, delta psi, or deltapH. Moreover, at external pH values above 6.0-6.5, delta-muH+ is insufficient to account for the concentration gradients established for each substrate unless the stoichiometry between protons and accumulated solutes is greater than unity. The results confirm many facets of the chemiosmotic hypothesis, but they also extend the concept in certain important respects and allow explanations for some earlier observations which seemed to preclude the involvement of chemiosmotic phenomena in active transport.

The effects of starch and rapidly degradable dry matter from concentrate on ruminal digestion in dairy cows fed corn silage-based diets with fixed forage proportion.

PubMed

Lechartier, C; Peyraud, J-L

2011-05-01

This study investigated the effects of the type (starch vs. nonstarch) and rate of ruminal degradation of carbohydrates from the concentrate on digestion in dairy cows fed corn silage-based diets. Six ruminally cannulated cows were assigned to 6 treatments in a 6 × 6 Latin square design. Treatments were arranged in a 3 × 2 factorial design. Two starch levels [25 and 41% dry matter (DM) for low starch (LS) and high starch (HS) diets, respectively] were obtained by replacing starch-rich feedstuffs by nonstarch feedstuffs. These starch levels were combined with slowly, moderately, and rapidly rumen-degradable feedstuffs to obtain 3 levels of rapidly degradable carbohydrates from concentrate (18, 23, and 28% DM). These levels were estimated from the DM disappearance of concentrate after 4h of in sacco incubation (CRDM). Wheat and corn grain were used as rapidly degradable and slowly degradable starch feedstuffs, respectively. Soybean hulls and citrus pulp were used as slowly degradable and rapidly degradable nonstarch feedstuffs, respectively. No interaction effect was found between dietary starch content and CRDM on pH range, volatile fatty acid (VFA) range, or VFA profile. Increasing CRDM led to a linear decrease in acetate-to-propionate ratio (from 2.7 to 2.1), and a linear increase in the pH and VFA ranges (from 0.86 to 1.12 pH units and from 34 to 56mM, respectively). Feeding HS diets decreased acetate-to-propionate ratio (2.6 vs. 2.0) and increased pH range (0.89 vs. 1.04 pH units), but had no effect on VFA range. Increasing CRDM linearly decreased mean ruminal pH in LS diets but linearly increased mean ruminal pH in HS diets. Fibrolytic activity was unaffected in LS diets but decreased strongly in HS diets (from 62 to 50%). These findings suggest that pH regulation differs on a short-term and on a longer-term basis. In the short-term, increasing CRDM increased the rate of VFA production, which may have been partly buffered under LS diets due to the higher cation exchange capacity of nonstarch feedstuffs compared with starch-rich feedstuffs. In the longer term, feeding starch reduced fibrolytic activity, which may have led to lower total VFA production and higher mean pH. The results of this experiment clearly show that both fermentative characteristics of the concentrate and dietary starch content should be taken into account when formulating diets to prevent subacute ruminal acidosis and to predict VFA profile. Copyright © 2011 American Dairy Science Association. Published by Elsevier Inc. All rights reserved.

Evidence of the Generation of Isosaccharinic Acids and Their Subsequent Degradation by Local Microbial Consortia within Hyper-Alkaline Contaminated Soils, with Relevance to Intermediate Level Radioactive Waste Disposal

PubMed Central

Rout, Simon P.; Charles, Christopher J.; Garratt, Eva J.; Laws, Andrew P.; Gunn, John; Humphreys, Paul N.

2015-01-01

The contamination of surface environments with hydroxide rich wastes leads to the formation of high pH (>11.0) soil profiles. One such site is a legacy lime works at Harpur Hill, Derbyshire where soil profile indicated in-situ pH values up to pH 12. Soil and porewater profiles around the site indicated clear evidence of the presence of the α and β stereoisomers of isosaccharinic acid (ISA) resulting from the anoxic, alkaline degradation of cellulosic material. ISAs are of particular interest with regards to the disposal of cellulosic materials contained within the intermediate level waste (ILW) inventory of the United Kingdom, where they may influence radionuclide mobility via complexation events occurring within a geological disposal facility (GDF) concept. The mixing of uncontaminated soils with the alkaline leachate of the site resulted in ISA generation, where the rate of generation in-situ is likely to be dependent upon the prevailing temperature of the soil. Microbial consortia present in the uncontaminated soil were capable of surviving conditions imposed by the alkaline leachate and demonstrated the ability to utilise ISAs as a carbon source. Leachate-contaminated soil was sub-cultured in a cellulose degradation product driven microcosm operating at pH 11, the consortia present were capable of the degradation of ISAs and the generation of methane from the resultant H2/CO2 produced from fermentation processes. Following microbial community analysis, fermentation processes appear to be predominated by Clostridia from the genus Alkaliphilus sp, with methanogenesis being attributed to Methanobacterium and Methanomassiliicoccus sp. The study is the first to identify the generation of ISA within an anthropogenic environment and advocates the notion that microbial activity within an ILW-GDF is likely to influence the impact of ISAs upon radionuclide migration. PMID:25748643

Evidence of the generation of isosaccharinic acids and their subsequent degradation by local microbial consortia within hyper-alkaline contaminated soils, with relevance to intermediate level radioactive waste disposal.

PubMed

Rout, Simon P; Charles, Christopher J; Garratt, Eva J; Laws, Andrew P; Gunn, John; Humphreys, Paul N

2015-01-01

The contamination of surface environments with hydroxide rich wastes leads to the formation of high pH (>11.0) soil profiles. One such site is a legacy lime works at Harpur Hill, Derbyshire where soil profile indicated in-situ pH values up to pH 12. Soil and porewater profiles around the site indicated clear evidence of the presence of the α and β stereoisomers of isosaccharinic acid (ISA) resulting from the anoxic, alkaline degradation of cellulosic material. ISAs are of particular interest with regards to the disposal of cellulosic materials contained within the intermediate level waste (ILW) inventory of the United Kingdom, where they may influence radionuclide mobility via complexation events occurring within a geological disposal facility (GDF) concept. The mixing of uncontaminated soils with the alkaline leachate of the site resulted in ISA generation, where the rate of generation in-situ is likely to be dependent upon the prevailing temperature of the soil. Microbial consortia present in the uncontaminated soil were capable of surviving conditions imposed by the alkaline leachate and demonstrated the ability to utilise ISAs as a carbon source. Leachate-contaminated soil was sub-cultured in a cellulose degradation product driven microcosm operating at pH 11, the consortia present were capable of the degradation of ISAs and the generation of methane from the resultant H2/CO2 produced from fermentation processes. Following microbial community analysis, fermentation processes appear to be predominated by Clostridia from the genus Alkaliphilus sp, with methanogenesis being attributed to Methanobacterium and Methanomassiliicoccus sp. The study is the first to identify the generation of ISA within an anthropogenic environment and advocates the notion that microbial activity within an ILW-GDF is likely to influence the impact of ISAs upon radionuclide migration.

Production and characterization of a novel yeast extracellular invertase activity towards improved dibenzothiophene biodesulfurization.

PubMed

Arez, Bruno F; Alves, Luís; Paixão, Susana M

2014-11-01

The main goal of this work was the production and characterization of a novel invertase activity from Zygosaccharomyces bailii strain Talf1 for further application to biodesulfurization (BDS) in order to expand the exploitable alternative carbon sources to renewable sucrose-rich feedstock. The maximum invertase activity (163 U ml(-1)) was achieved after 7 days of Z. bailii strain Talf1 cultivation at pH 5.5-6.0, 25 °C, and 150 rpm in Yeast Malt Broth with 25 % Jerusalem artichoke pulp as inducer substrate. The optimum pH and temperature for the crude enzyme activity were 5.5 and 50 °C, respectively, and moreover, high stability was observed at 30 °C for pH 5.5-6.5. The application of Talf1 crude invertase extract (1 %) to a BDS process by Gordonia alkanivorans strain 1B at 30 °C and pH 7.5 was carried out through a simultaneous saccharification and fermentation (SSF) approach in which 10 g l(-1) sucrose and 250 μM dibenzothiophene were used as sole carbon and sulfur sources, respectively. Growth and desulfurization profiles were evaluated and compared with those of BDS without invertase addition. Despite its lower stability at pH 7.5 (loss of activity within 24 h), Talf1 invertase was able to catalyze the full hydrolysis of 10 g l(-1) sucrose in culture medium into invert sugar, contributing to a faster uptake of the monosaccharides by strain 1B during BDS. In SSF approach, the desulfurizing bacterium increased its μmax from 0.035 to 0.070 h(-1) and attained a 2-hydroxybiphenyl productivity of 5.80 μM/h in about 3 days instead of 7 days, corresponding to an improvement of 2.6-fold in relation to the productivity obtained in BDS process without invertase addition.

Cathepsin K in Lymphangioleiomyomatosis: LAM Cell-Fibroblast Interactions Enhance Protease Activity by Extracellular Acidification.

PubMed

Dongre, Arundhati; Clements, Debbie; Fisher, Andrew J; Johnson, Simon R

2017-08-01

Lymphangioleiomyomatosis (LAM) is a rare disease in which LAM cells and fibroblasts form lung nodules and it is hypothesized that LAM nodule-derived proteases cause cyst formation and tissue damage. On protease gene expression profiling in whole lung tissue, cathepsin K gene expression was 40-fold overexpressed in LAM compared with control lung tissue (P ≤ 0.0001). Immunohistochemistry confirmed cathepsin K protein was expressed in LAM but not control lungs. Cathepsin K gene expression and protein and protease activity were detected in LAM-associated fibroblasts but not the LAM cell line 621-101. In lung nodules, cathepsin K immunoreactivity predominantly co-localized with LAM-associated fibroblasts. In vitro, fibroblast extracellular cathepsin K activity was minimal at pH 7.5 but significantly enhanced at pH 7 and 6. 621-101 cells reduced extracellular pH with acidification dependent on 621-101 mechanistic target of rapamycin activity and net hydrogen ion exporters, particularly sodium bicarbonate co-transporters and carbonic anhydrases, which were also expressed in LAM lung tissue. In LAM cell-fibroblast co-cultures, acidification paralleled cathepsin K activity, and both were reduced by sodium bicarbonate co-transporter (P ≤ 0.0001) and carbonic anhydrase inhibitors (P = 0.0021). Our findings suggest that cathepsin K activity is dependent on LAM cell-fibroblast interactions, and inhibitors of extracellular acidification may be potential therapies for LAM. Copyright © 2017 American Society for Investigative Pathology. Published by Elsevier Inc. All rights reserved.

Soil pH on mobility of imazaquin in oxisols with positive balance of charges.

PubMed

Regitano, Jussara B; da Rocha, Wadson S D; Alleoni, Luís R F

2005-05-18

The influence of soil pH on the leaching potential of the ionizable herbicide imazaquin was assessed on the profile of two highly weathered soils having a net positive charge in the B horizon, in contrast to a soil having a net negative charge in the whole profile, using packed soil column experiments. Imazaquin leached to a large extent and faster at Kd values lower than 1.0 L kg(-1), a much more lenient limit than usually proposed for pesticides in the literature (Kd < 5.0 L kg(-1)). The amount of imazaquin leached increased with soil pH. As the soil pH increased, the percentage of imazaquin in the anionic forms, the negative surface potential of the soils, as well as imazaquin water solubility also increased, thus reducing sorption because of repulsive electrostatic forces (hydrophilic interactions). For all surface samples (0-0.2 m), imazaquin did not leach at soil pH values lower than pKa (3.8) and more than 80% of the applied amount was leached at pH values higher than 5.5. For subsurface samples from the acric soils, imazaquin only began to leach at soil pH values > zero point of salt effects (ZPSE > 5.7). In conclusion, the use of surface K(oc) values to predict the amount of imazaquin leached within soil profiles having a positive balance of charges may greatly overestimate its actual leaching potential.

Impacts of Edaphic Factors on Communities of Ammonia-Oxidizing Archaea, Ammonia-Oxidizing Bacteria and Nitrification in Tropical Soils

PubMed Central

de Gannes, Vidya; Eudoxie, Gaius; Hickey, William J.

2014-01-01

Nitrification is a key process in soil nitrogen (N) dynamics, but relatively little is known about it in tropical soils. In this study, we examined soils from Trinidad to determine the edaphic drivers affecting nitrification levels and community structure of ammonia-oxidizing bacteria (AOB) and ammonia-oxidizing archaea (AOA) in non-managed soils. The soils were naturally vegetated, ranged in texture from sands to clays and spanned pH 4 to 8. The AOA were detected by qPCR in all soils (ca. 105 to 106 copies archaeal amoA g−1 soil), but AOB levels were low and bacterial amoA was infrequently detected. AOA abundance showed a significant negative correlation (p<0.001) with levels of soil organic carbon, clay and ammonium, but was not correlated to pH. Structures of AOA and AOB communities, as determined by amoA terminal restriction fragment (TRF) analysis, differed significantly between soils (p<0.001). Variation in AOA TRF profiles was best explained by ammonium-N and either Kjeldahl N or total N (p<0.001) while variation in AOB TRF profiles was best explained by phosphorus, bulk density and iron (p<0.01). In clone libraries, phylotypes of archaeal amoA (predominantly Nitrososphaera) and bacterial amoA (predominanatly Nitrosospira) differed between soils, but variation was not correlated with pH. Nitrification potential was positively correlated with clay content and pH (p<0.001), but not to AOA or AOB abundance or community structure. Collectively, the study showed that AOA and AOB communities were affected by differing sets of edaphic factors, notably that soil N characteristics were significant for AOA, but not AOB, and that pH was not a major driver for either community. Thus, the effect of pH on nitrification appeared to mainly reflect impacts on AOA or AOB activity, rather than selection for AOA or AOB phylotypes differing in nitrifying capacity. PMID:24586878

Biochemical characterization of recombinant cinnamoyl CoA reductase 1 (Ll-CCRH1) from Leucaena leucocephala.

PubMed

Sonawane, Prashant; Vishwakarma, Rishi Kishore; Khan, Bashir M

2013-07-01

Recombinant cinnamoyl CoA reductase 1 (Ll-CCRH1) protein from Leucaena leucocephala was overexpressed in Escherichia coli BL21 (DE3) strain and purified to apparent homogeneity. Optimum pH for forward and reverse reaction was found to be 6.5 and 7.8 respectively. The enzyme was most stable around pH 6.5 at 25°C for 90 min. The enzyme showed Kcat/Km for feruloyl, caffeoyl, sinapoyl, coumaroyl CoA, coniferaldehyde and sinapaldehyde as 4.6, 2.4, 2.3, 1.7, 1.9 and 1.2 (×10(6) M(-1) s(-1)), respectively, indicating affinity of enzyme for feruloyl CoA over other substrates and preference of reduction reaction over oxidation. Activation energy, Ea for various substrates was found to be in the range of 20-50 kJ/mol. Involvement of probable carboxylate ion, histidine, lysine or tyrosine at the active site of enzyme was predicted by pH activity profile. SAXS studies of protein showed radius 3.04 nm and volume 49.25 nm(3) with oblate ellipsoid shape. Finally, metal ion inhibition studies revealed that Ll-CCRH1 is a metal independent enzyme. Copyright © 2013 Elsevier B.V. All rights reserved.

Purification and characterization of paraoxon hydrolase from rat liver.

PubMed Central

Rodrigo, L; Gil, F; Hernandez, A F; Marina, A; Vazquez, J; Pla, A

1997-01-01

Paraoxonase (paraoxon hydrolase), an enzyme that hydrolyses paraoxon (O,O-diethyl O-p-nitrophenyl phosphate), is located in mammals primarily in the serum and liver. Although considerable information is available regarding serum paraoxonase, little is known about the hepatic form of this enzyme. The present work represents the first study on the purification of rat liver paraoxonase. This enzyme has been purified 415-fold to apparent homogeneity with a final specific activity of 1370 units/mg using a protocol consisting of five steps: solubilization of the microsomal fraction, hydroxyapatite adsorption, chromatography on DEAE-Sepharose CL-6B, non-specific affinity chromatography on Cibacron Blue 3GA and anion exchange on Mono Q HR 5/5. The presence of Ca2+ and Triton X-100 in the buffers throughout the purification procedure was essential for maintaining enzyme activity. SDS/PAGE of the final preparation indicated a single protein-staining band with an apparent Mr of 45 000. N-terminal and internal amino acid sequences were determined and compared with those of paraoxonases from human and rabbit serum and mouse liver, showing a high similarity. The pH profile showed optimum activity at pH 8.5. The pH stability and heat inactivation of the enzyme were also studied. The Km for liver paraoxonase was 1.69 mM. PMID:9032442

Mechanistic and Structural Analyses of the Role of His67 in the Yeast Polyamine Oxidase Fms1

DOE Office of Scientific and Technical Information (OSTI.GOV)

Adachi, Mariya S.; Taylor, Alexander B.; Hart, P. John

2012-07-25

The flavoprotein oxidase Fms1 from Saccharomyces cerevisiae catalyzes the oxidation of spermine and N1-acetylspermine to spermidine and 3-aminopropanal or N-acetyl-3-aminopropanal. Within the active site of Fms1, His67 is positioned to form hydrogen bonds with the polyamine substrate. This residue is also conserved in other polyamine oxidases. The catalytic properties of H67Q, H67N, and H67A Fms1 have been characterized to evaluate the role of this residue in catalysis. With both spermine and N1-acetylspermine as the amine substrate, the value of the first-order rate constant for flavin reduction decreases 2-3 orders of magnitude, with the H67Q mutation having the smallest effect andmore » H67N the largest. The k{sub cat}/K{sub O2} value changes very little upon mutation with N{sup 1}-acetylspermine as the amine substrate and decreases only an order of magnitude with spermine. The k{sub cat}/K{sub M}-pH profiles with N{sup 1}-acetylspermine are bell-shaped for all the mutants; the similarity to the profile of the wild-type enzyme rules out His67 as being responsible for either of the pK{sub a} values. The pH profiles for the rate constant for flavin reduction for all the mutant enzymes similarly show the same pK{sub a} as wild-type Fms1, about {approx}7.4; this pK{sub a} is assigned to the substrate N4. The k{sub cat}/K{sub O2}-pH profiles for wild-type Fms1 and the H67A enzyme both show a pK{sub a} of about {approx}6.9; this suggests His67 is not responsible for this pH behavior. With the H67Q, H67N, and H67A enzymes the k{sub cat} value decreases when a single residue is protonated, as is the case with the wild-type enzyme. The structure of H67Q Fms1 has been determined at a resolution of 2.4 {angstrom}. The structure shows that the mutation disrupts a hydrogen bond network in the active site, suggesting that His67 is important both for direct interactions with the substrate and to maintain the overall active site structure.« less

Expression, purification and characterization of a phyAm-phyCs fusion phytase*

PubMed Central

Zou, Li-kou; Wang, Hong-ning; Pan, Xin; Tian, Guo-bao; Xie, Zi-wen; Wu, Qi; Chen, Hui; Xie, Tao; Yang, Zhi-rong

2008-01-01

The phyAm gene encoding acid phytase and optimized neutral phytase phyCs gene were inserted into expression vector pPIC9K in correct orientation and transformed into Pichia pastoris in order to expand the pH profile of phytase and decrease the cost of production. The fusion phytase phyAm-phyCs gene was successfully overexpressed in P. pastoris as an active and extracellular phytase. The yield of total extracellular fusion phytase activity is (25.4±0.53) U/ml at the flask scale and (159.1±2.92) U/ml for high cell-density fermentation, respectively. Purified fusion phytase exhibits an optimal temperature at 55 °C and an optimal pH at 5.5~6.0 and its relative activity remains at a relatively high level of above 70% in the range of pH 2.0 to 7.0. About 51% to 63% of its original activity remains after incubation at 75 °C to 95 °C for 10 min. Due to heavy glycosylation, the expressed fusion phytase shows a broad and diffuse band in SDS-PAGE (sodium dodecyl sulfate-polyacrylamide gel electrophoresis). After deglycosylation by endoglycosidase H (EndoHf), the enzyme has an apparent molecular size of 95 kDa. The characterization of the fusion phytase was compared with those of phyCs and phyAm. PMID:18600783

Towards elucidation of the drug release mechanism from compressed hydrophilic matrices made of cellulose ethers. III. Critical use of thermodynamic parameters of activation for modeling the water penetration and drug release processes.

PubMed

Ferrero, Carmen; Massuelle, Danielle; Jeannerat, Damien; Doelker, Eric

2013-09-10

The two main purposes of this work were: (i) to critically consider the use of thermodynamic parameters of activation for elucidating the drug release mechanism from hydroxypropyl methylcellulose (HPMC) matrices, and (ii) to examine the effect of neutral (pH 6) and acidic (pH 2) media on the release mechanism. For this, caffeine was chosen as model drug and various processes were investigated for the effect of temperature and pH: caffeine diffusion in solution and HPMC gels, and drug release from and water penetration into the HPMC tablets. Generally, the kinetics of the processes was not significantly affected by pH. As for the temperature dependence, the activation energy (E(a)) values calculated from caffeine diffusivities were in the range of Fickian transport (20-40 kJ mol⁻¹). Regarding caffeine release from HPMC matrices, fitting the profiles using the Korsmeyer-Peppas model would indicate anomalous transport. However, the low apparent E(a) values obtained were not compatible with a swelling-controlled mechanism and can be assigned to the dimensional change of the system during drug release. Unexpectedly, negative apparent E(a) values were calculated for the water uptake process, which can be ascribed to the exothermic dissolution of water into the initially dry HPMC, the expansion of the matrix and the polymer dissolution. Taking these contributions into account, the true E(a) would fall into the range valid for Fickian diffusion. Consequently, a relaxation-controlled release mechanism can be dismissed. The apparent anomalous drug release from HPMC matrices results from a coupled Fickian diffusion-erosion mechanism, both at pH 6 and 2. Copyright © 2013 Elsevier B.V. All rights reserved.

Abundant Hydrothermal Venting in the Southern Ocean Near 62°S/159°E on the Australian-Antarctic Ridge

NASA Astrophysics Data System (ADS)

Baker, E. T.; Hahm, D.; Rhee, T. S.; Park, S. H.; Lupton, J. E.; Walker, S. L.; Choi, H.

2014-12-01

Circum-Antarctic Ridges (CARs) comprise almost one-third of the global Mid-Ocean Ridge, yet remain terra incognita for hydrothermal activity and chemosynthetic ecosystems. The InterRidge Vents Database lists only 3 confirmed (visualized) and 35 inferred (plume evidence) active sites along the ~21,000 km of CARs. Here, we report on a multi-year effort to locate and characterize hydrothermal activity on two 1st-order segments of the Australian-Antarctic Ridge that are perhaps more isolated from other known vent fields than any other vent site on the Mid-Ocean Ridge. KR1 is a 300-km-long segment near 62°S/159°E, and KR2 a 90-km-long segment near 60°S/152.5°E. We used profiles collected by Miniature Autonomous Plume Recorders (MAPRs) on rock corers in March and December of 2011 to survey each segment, and an intensive CTD survey in Jan/Feb 2013 to pinpoint sites and sample plumes on KR1. Optical and oxidation-reduction potential (ORP, aka Eh) anomalies indicate multiple active sites on both segments. Seven profiles on KR2 found 3 sites, each separated by ~25 km. Forty profiles on KR1 identified 13 sites, some within a few km of each other. The densest site concentration on KR1 occurred along a relatively inflated, 90-km-long section near the segment center. CTD tows covered 20 km of the eastern, most inflated portion of this area, finding two 6-km-long zones centered near 158.6°E and 158.8°E with multiple plume anomalies. Three ORP anomalies within 50 m of the seafloor indicate precise venting locations. We call this area the Mujin "Misty Harbor" vent field. Vent frequency sharply decreases away from Mujin. 3He/heat ratios determined from 20 plume samples in the Mujin field were mostly <0.015 fM/J, indicative of chronic venting, but 3 samples, 0.021-0.034 fM/J, are ratios typical of a recent eruption. The spatial density of hydrothermal activity along KR1 and KR2 is similar to other intermediate-rate spreading ridges. We calculate the plume incidence (ph) along KR1 and KR2 as the mean of the fraction of MAPR casts detecting a plume in each 2nd-order segment. For all 6 segments, ph=0.37±0.25, consistent with the prediction of 0.33 from the global trend of ph for a spreading rate of 68 mm/yr.

A low pH does not determine the community dynamics of spontaneously developed backslopped liquid wheat sourdoughs but does influence their metabolite kinetics.

PubMed

Van Kerrebroeck, Simon; Bastos, Francisca Casanova C; Harth, Henning; De Vuyst, Luc

2016-12-19

This study dealt with the influence of a crucial pH value of 4.0 on the microbiota of spontaneously fermented backslopped liquid wheat sourdoughs. Two spontaneously fermented wheat sourdough fermentation experiments were carried out, one without control of the pH and one with the pH kept constant at pH4.0, both during nine backslopping steps. In each case, two additional backslopping steps were carried out, with the pH kept constant at 4.0 and with free pH, respectively. Keeping the pH constant at 4.0 changed the microbial community dynamics and metabolite kinetics of the sourdough fermentations. A slower prevalence of sourdough-specific Kazachstania yeasts occurred. Nevertheless, in both experiments, Lactobacillus fermentum, Lb. plantarum/pentosus/paraplantarum, and Kazachstania exigua/bulderi/barnettii prevailed ultimately. The lactic acid and ethanol concentration profiles were affected positively by keeping the pH constant at a minimum of 4.0 as well as the l- and d-lactic acid ratio profile, a potential biological marker for sourdough stability and maturity. Also, the concentration and diversity of acetate esters and their precursors, in particular isoamyl acetate and isoamyl alcohol, were affected negatively by the pH control, indicating the role of pH stress in the sourdough aroma formation. Copyright © 2016 Elsevier B.V. All rights reserved.

Studies on the production of alkaline α-amylase from Bacillus subtilis CB-18.

PubMed

Nwokoro, Ogbonnaya; Anthonia, Odiase

2015-01-01

Amylases are among the main enzymes used in food and other industries. They hydrolyse starch molecules into polymers composing glucose units. Amylases have potential applications in a number of industrial processes including foods and pharmaceutical industries. Alkaline α-amylase has the potential of hydrolysing starch under alkaline pH and is useful in the starch and textile industries and as an ingredient of detergents. Amylases are produced from plants, however, microbial production processes have dominated applications in the industries. Optimization of microbial production processes can result in improved enzyme yields. Amylase activity was assayed by incubating the enzyme solution (0.5 ml) with 1% soluble starch (0.5 ml) in 0.1 M Tris/HCl buffer (pH 8.5). After 30 minutes, the reaction was stopped by the addition of 4 mL of 3,5-dinitrosalicylic acid (DNS) reagent then heated for 10 min in boiling water bath and cooled in a refrigerator. Absorbance readings were used to estimate the units of enzyme activity from glucose standard curve. Hydrolysed native starches from cassava, rice, corn, coco yam, maize and potato and soluble starch were adjusted to pH 8.5 prior to incubation with crude enzyme solution. Reducing sugars produced were therefore determined. The effect of pH on enzyme activity of the alkaline α-amylase was determined by using buffer solutions of different pH (potassium phosphate buffer, 6.0-7.0; Tris-HCl buffer 7.5 to 9.0 and carbonate/bicarbonate buffer, pH 9.5-11) for enzyme assay. The pH stability profile of the enzyme was determined by incubating 0.5 ml of α-amylase enzyme in 0.1 M Tris/HCl buffer (pH 8.5) and 0.5 ml of 1% (w/v) soluble starch (Merck) in 0.1 M Tris/HCl buffer (pH 8.5) for 3 h in various buffers. The effect of temperature on enzyme activity was studied by incubating 0.5 mL of the enzyme solution contained in the test tube and 0.5 mL of 1% soluble starch (Merck) solution prepared in 0.1 M Tris/HCl buffer (pH 8.5) for 3 h at various temperatures (25, 30, 35, 40, 45, 50, 55 and 60°C) in a thermo static water bath. The reactions were stopped by adding DNS reagent. The enzyme activity was therefore determined. Thermal stability was studied by incubating 0.5 ml of enzyme solution in 0.1 M Tris/HCl buffer (pH 8.5) and 0.5 ml of 1% (w/v) soluble starch (Merck) in 0.1 M Tris/HCl buffer (pH 8.5) for 3 h at various temperatures (20, 30, 40, 50, 60 and 70°C) for 60 min. The enzyme displayed optimal activity at pH 8.0 at which it produced maximum specific activity of 34.3 units/mg protein. Maximum stability was at pH 8.0 to 9.0. Maximum activity was observed at temperature of 50°C while thermo stability of the enzyme was observed at 40-50°C. The enzyme displayed a wide range of activities on starch and caused the release of 5.86, 4.75, 5.98, 3.44, 3.96, 8.84 mg/mL reducing sugar from cassava, potato, cocoyam, corn, rice and soluble starch respectively. This investigation reports some biochemical characterization of alkaline α-amylase from Bacillus subtilis CB-18. The substrate specificities of this enzyme on various starches suggested that the alkaline α-amylase enzyme had combined activities on raw and soluble starch.

Stability studies on diloxanide furoate: effect of pH, temperature, gastric and intestinal fluids.

PubMed

Gadkariem, E A; Belal, F; Abounassif, M A; El-Obeid, H A; E E Ibrahim, K

2004-04-01

The degradation of the amoebicide diloxanide furoate in alkaline medium at different temperatures was investigated using both a spectrophotometric and a developed HPLC method. In solutions, the drug was found to undergo decomposition, i.e., temperature and pH dependent. The pH-rate profile at pH between 7.6 and 9.6 indicated a first-order dependence of Kobs on [-OH]. Arrhenius plot obtained at pH 8 was linear between 40 and 63 degrees C. The estimated activation energy of hydrolysis was found to be 18.25 kcal degree.mol(-1). The effect of simulated gastric and intestinal fluids on the drug was also investigated. A new thin-layer chromatographic (TLC) procedure for the fractionation of the drug and its alkaline hydrolysis products has been developed and was found to compare favorably with that of the British Pharmacopoeia. Three hydrolysis products of a basic methanolic solution of the drug, namely furoic acid, diloxanide and methylfuroate could be identified by the use of TLC, HPLC, infrared and mass spectrometry.

Comparative Studies on the Dissolution Profiles of Oral Ibuprofen Suspension and Commercial Tablets using Biopharmaceutical Classification System Criteria

PubMed Central

Rivera-Leyva, J. C.; García-Flores, M.; Valladares-Méndez, A.; Orozco-Castellanos, L. M.; Martínez-Alfaro, M.

2012-01-01

In vitro dissolution studies for solid oral dosage forms have recently widened the scope to a variety of special dosage forms such as suspensions. For class II drugs, like Ibuprofen, it is very important to have discriminative methods for different formulations in physiological conditions of the gastrointestinal tract, which will identify different problems that compromise the drug bioavailability. In the present work, two agitation speeds have been performed in order to study ibuprofen suspension dissolution. The suspensions have been characterised relatively to particle size, density and solubility. The dissolution study was conducted using the following media: buffer pH 7.2, pH 6.8, 4.5 and 0.1 M HCl. For quantitative analysis, the UV/Vis spectrophotometry was used because this methodology had been adequately validated. The results show that 50 rpm was the adequate condition to discriminate the dissolution profile. The suspension kinetic release was found to be dependent on pH and was different compared to tablet release profile at the same experimental conditions. The ibuprofen release at pH 1.0 was the slowest. PMID:23626386

Predominant processing adaptability of Staphylococcus xylosus strains isolated from Chinese traditional low-salt fermented whole fish.

PubMed

Zeng, Xuefeng; He, Laping; Guo, Xu; Deng, Li; Yang, Wangen; Zhu, Qiujin; Duan, Zhenhua

2017-02-02

This study aimed to determine the predominant processing adaptability of 27 selected isolates of Staphylococcus xylosus in 'Suan yu', a traditional Chinese low-salt fermented whole-fish product. The isolates were screened for proteolytic, lipolytic, and enzymatic profiles; amino-acid decarboxylase content; antimicrobial activities; and tolerance to low temperatures, pH5.0, and salt. Two S. xylosus strains grew at 10°C in the presence of 10% NaCl and at pH5.0. Agar-plate assays and sodium dodecyl sulphate-polyacrylamide gel electrophoresis revealed that 21 and 8 of the strains exhibited appropriate proteolytic activities against myofibrillar and sarcoplasmic proteins, respectively. All S. xylosus strains also displayed different enzymatic profiles, and most strains showed negative decarboxylase activities. The results of this step were used as input data for a Principal Component Analysis; therefore, the most technologically relevant strain 3 and 8 were combined with L. plantarum 120 as MS1 and MS2, respectively, were further selected for the fermented fish surimi, and the fish surimi inoculated with mixed starter cultures (MS1, MS2) scored high for overall acceptability. Free amino acid contents of 1757 and 1765mg/100g sample were found in fish surimi inoculated with MS1 and MS2, respectively, after 72h of fermentation. Therefore, Sx-3 and Sx-8, which presented the best predominant processing adaptability, is an eligible starter culture for fermented fish production. Copyright © 2016 Elsevier B.V. All rights reserved.

The design of a low-cost Thomson Scattering system for use on the ORNL PhIX device

NASA Astrophysics Data System (ADS)

Biewer, T. M.; Lore, J.; Goulding, R. H.; Hillis, D. L.; Owen, L.; Rapp, J.

2012-10-01

Study of the plasma-material interface (PMI) under high power and particle flux on linear plasma devices is an active area of research that is relevant to fusion-grade toroidal devices such as ITER and DEMO. ORNL is assembling a 15 cm diameter, ˜3 m long linear machine, called the Physics Integration eXperiment (PhIX), which incorporates a helicon plasma source, electron heating, and a material target. The helicon source has demonstrated coupling of up to 100 kW of rf power, and produced ne >= 4 x 10^19 m-3 in D, and He fueled plasmas, measured with interferometry and Langmuir probes (LP). Optical emission spectroscopy was used to confirm LP measurements that Te is about 10 eV in helicon heated plasmas, which will presumably increase when electron heating is applied. Plasma parameters (ne, Te, n0) of the PhIX device will be measured with a novel, low-cost Thomson Scattering (TS) system. The data will be used to characterize the PMI regime with multiple profile measurements in front of the target. Profiles near the source and target will be used to determine the parallel transport regime via comparison to 2D fluid plasma simulations. This work was supported by the US. D.O.E. contract DE-AC05-00OR22725.

Cyclodextrin-insulin complex encapsulated polymethacrylic acid based nanoparticles for oral insulin delivery.

PubMed

Sajeesh, S; Sharma, Chandra P

2006-11-15

Present investigation was aimed at developing an oral insulin delivery system based on hydroxypropyl beta cyclodextrin-insulin (HPbetaCD-I) complex encapsulated polymethacrylic acid-chitosan-polyether (polyethylene glycol-polypropylene glycol copolymer) (PMCP) nanoparticles. Nanoparticles were prepared by the free radical polymerization of methacrylic acid in presence of chitosan and polyether in a solvent/surfactant free medium. Dynamic light scattering (DLS) experiment was conducted with particles dispersed in phosphate buffer (pH 7.4) and size distribution curve was observed in the range of 500-800 nm. HPbetaCD was used to prepare non-covalent inclusion complex with insulin and complex was analyzed by Fourier transform infrared (FTIR) and fluorescence spectroscopic studies. HPbetaCD complexed insulin was encapsulated into PMCP nanoparticles by diffusion filling method and their in vitro release profile was evaluated at acidic/alkaline pH. PMCP nanoparticles displayed good insulin encapsulation efficiency and release profile was largely dependent on the pH of the medium. Enzyme linked immunosorbent assay (ELISA) study demonstrated that insulin encapsulated inside the particles was biologically active. Trypsin inhibitory effect of PMCP nanoparticles was evaluated using N-alpha-benzoyl-L-arginine ethyl ester (BAEE) and casein as substrates. Mucoadhesive studies of PMCP nanoparticles were conducted using freshly excised rat intestinal mucosa and the particles were found fairly adhesive. From the preliminary studies, cyclodextrin complexed insulin encapsulated mucoadhesive nanoparticles appear to be a good candidate for oral insulin delivery.

Offline and online capillary electrophoresis enzyme assays of β-N-acetylhexosaminidase.

PubMed

Křížek, Tomáš; Doubnerová, Veronika; Ryšlavá, Helena; Coufal, Pavel; Bosáková, Zuzana

2013-03-01

Enzyme assays of β-N-acetylhexosaminidase from Aspergillus oryzae using capillary electrophoresis in the offline and online setup have been developed. The pH value and concentration of the borate-based background electrolyte were optimized in order to achieve baseline separation of N,N',N″-triacetylchitotriose, N,N'-diacetylchitobiose, and N-acetyl-D-glucosamine. The optimized method using 25 mM tetraborate buffer, pH 10.0, was evaluated in terms of repeatability, limits of detection, quantification, and linearity. The method was successfully applied to the offline enzyme assay of β-N-acetylhexosaminidase, which was demonstrated by monitoring the hydrolysis of N,N',N″-triacetylchitotriose. The presented method was also utilized to study the pH dependence of enzyme activity. An online assay with N,N'-diacetylchitobiose as a substrate was developed using the Transverse Diffusion of Laminar Flow Profiles model to optimize the injection sequence and in-capillary mixing of substrate and enzyme plugs. The experimental results were in good agreement with predictions of the model. The online assay was successfully used to observe the inhibition effect of N,N'-dimethylformamide on the activity of β-N-acetylhexosaminidase with nanoliter volumes of reagents used per run and a high degree of automation. After adjustment of background electrolyte pH, an online assay with N,N',N″-triacetylchitotriose as a substrate was also performed.

Production and Partial Characterization of α-Amylase Enzyme from Bacillus sp. BCC 01-50 and Potential Applications

PubMed Central

Qureshi, Abdul Sattar; Khushk, Imrana; Ali, Chaudhry Haider; Lashari, Safia; Bhutto, Muhammad Aqeel; Mangrio, Ghulam Sughra; Lu, Changrui

2017-01-01

Amylase is an industrially important enzyme and applied in many industrial processes such as saccharification of starchy materials, food, pharmaceutical, detergent, and textile industries. This research work deals with the optimization of fermentation conditions for α-amylase production from thermophilic bacterial strain Bacillus sp. BCC 01-50 and characterization of crude amylase. The time profile of bacterial growth and amylase production was investigated in synthetic medium and maximum enzyme titer was observed after 60 h. In addition, effects of different carbon sources were tested as a substrate for amylase production and molasses was found to be the best. Various organic and inorganic compounds, potassium nitrate, ammonium chloride, sodium nitrate, urea, yeast extract, tryptone, beef extract, and peptone, were used and beef extract was found to be the best among the nitrogen sources used. Temperature, pH, agitation speed, and size of inoculum were also optimized. Highest enzyme activity was obtained when the strain was cultured in molasses medium for 60 h in shaking incubator (150 rpm) at 50°C and pH 8. Crude amylase showed maximal activity at pH 9 and 65°C. Enzyme remained stable in alkaline pH range 9-10 and 60–70°C. Crude amylase showed great potential for its application in detergent industry and saccharification of starchy materials. PMID:28168200

Production and Partial Characterization of α-Amylase Enzyme from Bacillus sp. BCC 01-50 and Potential Applications.

PubMed

Simair, Altaf Ahmed; Qureshi, Abdul Sattar; Khushk, Imrana; Ali, Chaudhry Haider; Lashari, Safia; Bhutto, Muhammad Aqeel; Mangrio, Ghulam Sughra; Lu, Changrui

2017-01-01

Amylase is an industrially important enzyme and applied in many industrial processes such as saccharification of starchy materials, food, pharmaceutical, detergent, and textile industries. This research work deals with the optimization of fermentation conditions for α -amylase production from thermophilic bacterial strain Bacillus sp. BCC 01-50 and characterization of crude amylase. The time profile of bacterial growth and amylase production was investigated in synthetic medium and maximum enzyme titer was observed after 60 h. In addition, effects of different carbon sources were tested as a substrate for amylase production and molasses was found to be the best. Various organic and inorganic compounds, potassium nitrate, ammonium chloride, sodium nitrate, urea, yeast extract, tryptone, beef extract, and peptone, were used and beef extract was found to be the best among the nitrogen sources used. Temperature, pH, agitation speed, and size of inoculum were also optimized. Highest enzyme activity was obtained when the strain was cultured in molasses medium for 60 h in shaking incubator (150 rpm) at 50°C and pH 8. Crude amylase showed maximal activity at pH 9 and 65°C. Enzyme remained stable in alkaline pH range 9-10 and 60-70°C. Crude amylase showed great potential for its application in detergent industry and saccharification of starchy materials.

The AMPK-v-ATPase-pH axis: A key regulator of the pro-fibrogenic phenotype of human hepatic stellate cells.

PubMed

Marrone, Giusi; De Chiara, Francesco; Böttcher, Katrin; Levi, Ana; Dhar, Dipok; Longato, Lisa; Mazza, Giuseppe; Zhang, Zhenzhen; Marrali, Martina; Iglesias, Anabel Fernández-; Hall, Andrew; Luong, Tu Vinh; Viollet, Benoit; Pinzani, Massimo; Rombouts, Krista

2018-04-17

Liver fibrosis and cirrhosis are characterized by activation of hepatic stellate cells (HSC) which is associated with higher intracellular pH (pHi). The vacuolar H + adenosine-tri-phosphatase (v-ATPase) multi-subunit complex is a key regulator of intracellular pH homeostasis. The present work was aimed at investigating the functional role of v-ATPase in primary human HSC (hHSC) activation and its modulation by specific AMPK subunits. Here, we demonstrated that the expression of different v-ATPase subunits was increased in in vivo and in vitro activated hHSC, compared to non-activated hHSC. Specific inhibition of v-ATPase with Bafilomycin and KM91104 induced a down-regulation of the HSC fibrogenic gene profile, which coincided with increased lysosomal pH, decreased pHi, activation of AMPK, reduced proliferation, and a lower metabolic activity. Similarly, pharmacological activation of AMPK by treatment with Diflunisal, A769662 and ZLN024, reduced the expression of v-ATPase subunits and pro-fibrogenic markers. V-ATPase expression was differently regulated by AMPKα1 and AMPKα2, as demonstrated in mouse embryo fibroblasts (MEF) specific deficient for AMPKα subunits. In addition, the activation of v-ATPase in hHSC was shown to be AMPKα1 dependent. Accordingly, pharmacological activation of AMPK in AMPKα1-depleted hHSC prevented v-ATPase downregulation. Finally, we showed that v-ATPase expression was increased in fibrotic livers from Bile Duct Ligated mice and in human cirrhotic livers. The down-regulation of v-ATPase might represent a new promising target for the development of anti-fibrotic strategies. This article is protected by copyright. All rights reserved. © 2018 by the American Association for the Study of Liver Diseases.

Functional and rheological properties of proteins in frozen turkey breast meat with different ultimate pH.

PubMed

Chan, J T Y; Omana, D A; Betti, M

2011-05-01

Functional and rheological properties of proteins from frozen turkey breast meat with different ultimate pH at 24 h postmortem (pH(24)) have been studied. Sixteen breast fillets from Hybrid Tom turkeys were initially selected based on lightness (L*) values for each color group (pale, normal, and dark), with a total of 48 breast fillets. Further selection of 8 breast samples was made within each class of meat according to the pH(24). The average L* and pH values of the samples were within the following range: pale (L* >52; pH ≤5.7), normal (46 < L* < 52; 5.9 < pH <6.1), and dark (L* <46; pH ≥6.3), referred to as low, normal, and high pH meat, respectively. Ultimate pH did not cause major changes in the emulsifying and foaming properties of the extracted sarcoplasmic and myofibrillar proteins. An SDS-PAGE profile of proteins from low and normal pH meat was similar, which revealed that the extent of protein denaturation was the same. Low pH meat had the lowest water-holding capacity compared with normal and high pH meat as shown by the increase in cooking loss, which can be explained by factors other than protein denaturation. Gel strength analysis and folding test revealed that gel-forming ability was better for high pH meat compared with low and normal pH meat.Dynamic viscoelastic behavior showed that myosin denaturation temperature was independent of pH(24). Normal and high pH meat had similar hardness, springiness, and chewiness values as revealed by texture profile analysis. The results from this study indicate that high pH meat had similar or better functional properties than normal pH meat. Therefore, high pH meat is suitable for further processed products, whereas low pH meat may need additional treatment or ingredient formulations to improve its functionality.

The Role of the Dissertation in Scientific Careers.

ERIC Educational Resources Information Center

Porter, Alan L.; And Others

1982-01-01

Presents a profile of science Ph.D.'s and discusses the dissertation as research publishing patterns, predicting commitment to research, researchers versus nonresearchers, and the future of the Ph.D. degree. (JN)

A copper-methionine interaction controls the pH-dependent activation of peptidylglycine monooxygenase.

PubMed

Bauman, Andrew T; Broers, Brenda A; Kline, Chelsey D; Blackburn, Ninian J

2011-12-20

The pH dependence of native peptidylglycine monooxygenase (PHM) and its M314H variant has been studied in detail. For wild-type (WT) PHM, the intensity of the Cu-S interaction visible in the Cu(I) extended X-ray absorption fine structure (EXAFS) data is inversely proportional to catalytic activity over the pH range of 3-8. A previous model based on more limited data was interpreted in terms of two protein conformations involving an inactive Met-on form and an active flexible Met-off form [Bauman, A. T., et al. (2006) Biochemistry 45, 11140-11150] that derived its catalytic activity from the ability to couple into vibrational modes critical for proton tunneling. The new studies comparing the WT and M314H variant have led to the evolution of this model, in which the Met-on form has been found to be derived from coordination of an additional Met residue, rather than a more rigid conformer of M314 as previously proposed. The catalytic activity of the mutant decreased by 96% because of effects on both k(cat) and K(M), but it displayed the same activity-pH profile with a maximum around pH 6. At pH 8, the reduced Cu(I) form gave spectra that could be simulated by replacement of the Cu(M) Cu-S(Met) interaction with a Cu-N/O interaction, but the data did not unambiguously assign the ligand to the imidazole side chain of H314. At pH 3.5, the EXAFS still showed the presence of a strong Cu-S interaction, establishing that the Met-on form observed at low pH in WT cannot be due to a strengthening of the Cu(M)-methionine interaction but must arise from a different Cu-S interaction. Therefore, lowering the pH causes a conformational change at one of the Cu centers that brings a new S donor residue into a favorable orientation for coordination to copper and generates an inactive form. Cys coordination is unlikely because all Cys residues in PHM are engaged in disulfide cross-links. Sequence comparison with the PHM homologues tyramine β-monooxygenase and dopamine β-monooxygenase suggests that M109 (adjacent to H site ligands H107 and H108) is the most likely candidate. A model is presented in which H108 is protonated with a pK(a) of 4.6 to generate the inactive low-pH form with Cu(H) coordinated by M109, H107, and H172.

Sialidase activities of cultured human fibroblasts and the metabolism of GM3 ganglioside

DOE Office of Scientific and Technical Information (OSTI.GOV)

Usuki, S.; Lyu, S.C.; Sweeley, C.C.

1988-05-15

Free sialic acid has been found in the cell-conditioned medium of human foreskin fibroblasts. It is proposed that the accumulation of extracellular sialic acid may result from the hydrolysis of GM3 ganglioside on the cell surface of these fibroblasts. Sialidase activities with GM3 ganglioside and sialyllactitol as substrates were demonstrated in cell-conditioned medium, and the levels of their activities correlated positively with cell density. The GM3 sialidase activity at pH 4.5 was 4.1 and 38 pmol/h/ml of medium at sparse and confluent densities, respectively; the corresponding activities with sialyllactitol as the substrate were 12 and 75 pmol/h/ml of medium (pHmore » 4.5). The pH versus activity profiles with GM3 as the substrate suggested the presence of a second sialidase with an optimal activity at pH 6.5 in the conditioned medium of preconfluent cells. This activity was virtually absent in the medium of contact-inhibited cells and could not be assayed with sialyllactitol as the substrate. The turnover of cell surface GM3 was assessed by pulse labeling human foreskin fibroblasts with a radioactive precursor of sialic acid ((1-14C)N-acetylmannosamine) and a radioactive precursor of ceramide ((3,3-3H2)serine). During a chase period of 24 h turnover of the doubly labeled cellular GM3 was observed; there was a loss of about 35% of the 14C-labeled sialic acid without any measureable loss of 3H-labeled ceramide from GM3. We have speculated that the enzyme-catalyzed removal of sialic acid from the GM3 ganglioside on the extracellular aspect of the plasma membrane may be a necessary event involved in the modulation of cell growth.« less

Effect of formulation and processing variables on the characteristics of microspheres for water-soluble drugs prepared by w/o/o double emulsion solvent diffusion method.

PubMed

Lee, J; Park, T G; Choi, H

2000-02-25

80% except for acetaminophen, due to its lower solubility in water and higher solubility in corn oil. The release profile of the drug was pH dependent. In acidic medium, the release rate was much slower, however, the drug was released quickly at pH 7.4. Tacrine showed unexpected release profiles, probably due to ionic interaction with polymer matrix and the shell structure and the highest release rate was obtained at pH 2.0. The prepared microspheres had a sponge-like inner structure with or without central hollow core and the surface was dense with no apparent pores.

Development of a common priority list of pharmaceuticals relevant for the water cycle.

PubMed

de Voogt, P; Janex-Habibi, M-L; Sacher, F; Puijker, L; Mons, M

2009-01-01

Pharmaceutically active compounds (PhACs), including prescription drugs, over-the-counter medications, drugs used in hospitals and veterinary drugs, have been found throughout the water cycle. A desk study was initiated by the Global Water Research Coalition to consolidate a uniform selection of such compounds in order to judge risks of PhACs for the water cycle. By identifying major existing prioritization efforts and evaluating the criteria they use, this study yields a representative and qualitative profile ('umbrella view') of priority pharmaceuticals based on an extensive set of criteria. This can then be used for further studies on analytical methods, occurrence, treatability and potential risks associated with exposure to PhACs in water supply, identifying compounds most likely to be encountered and that may have significant impact on human health. For practical reasons, the present study excludes veterinary drugs. The pragmatic approach adopted provides an efficient tool to manage risks related to pharmaceuticals and provides assistance for selecting compounds for future studies.

A nanobuffer reporter library for fine-scale imaging and perturbation of endocytic organelles

PubMed Central

Wang, Chensu; Wang, Yiguang; Li, Yang; Bodemann, Brian; Zhao, Tian; Ma, Xinpeng; Huang, Gang; Hu, Zeping; DeBerardinis, Ralph J.; White, Michael A.; Gao, Jinming

2015-01-01

Endosomes, lysosomes and related catabolic organelles are a dynamic continuum of vacuolar structures that impact a number of cell physiological processes such as protein/lipid metabolism, nutrient sensing and cell survival. Here we develop a library of ultra-pH-sensitive fluorescent nanoparticles with chemical properties that allow fine-scale, multiplexed, spatio-temporal perturbation and quantification of catabolic organelle maturation at single organelle resolution to support quantitative investigation of these processes in living cells. Deployment in cells allows quantification of the proton accumulation rate in endosomes; illumination of previously unrecognized regulatory mechanisms coupling pH transitions to endosomal coat protein exchange; discovery of distinct pH thresholds required for mTORC1 activation by free amino acids versus proteins; broad-scale characterization of the consequence of endosomal pH transitions on cellular metabolomic profiles; and functionalization of a context-specific metabolic vulnerability in lung cancer cells. Together, these biological applications indicate the robustness and adaptability of this nanotechnology-enabled ‘detection and perturbation' strategy. PMID:26437053

Asparagine deamidation dependence on buffer type, pH, and temperature.

PubMed

Pace, Amanda L; Wong, Rita L; Zhang, Yonghua Taylor; Kao, Yung-Hsiang; Wang, Y John

2013-06-01

The deamidation of asparagine into aspartate and isoaspartate moieties is a major pathway for the chemical degradation of monoclonal antibodies (mAbs). It can affect the shelf life of a therapeutic antibody that is not formulated or stored appropriately. A new approach to detect deamidation using ion exchange chromatography was developed that separates papain-digested mAbs into Fc and Fab fragments. From this, deamidation rates of each fragment can be calculated. To generate kinetic parameters useful in setting shelf life, buffers prepared at room temperature and then placed at the appropriate stability temperatures. Solution pH was not adjusted to the same at different temperatures. Deamidation rate at 40°C was faster in acidic buffers than in basic buffers. However, this trend is reversed at 5°C, attributed to the change in hydroxide ion concentration influenced by buffer and temperature. The apparent activation energy was higher for rates generated in an acidic buffer than in a basic buffer. The rate-pH profile for mAb1 can be deconvoluted to Fc and Fab. The Fc deamidation showed a V-shaped profile: deamidation of PENNY peptide is responsible for the rate at high-pH, whereas deamidation of a new site, Asn323, may be responsible for the rate at low-pH. The profile for Fab is a straight line without curvature. Copyright © 2013 Wiley Periodicals, Inc.

Effects of infant cereals with different carbohydrate profiles on colonic function--randomised and double-blind clinical trial in infants aged between 6 and 12 months--pilot study.

PubMed

Bernal, María José; Periago, María Jesús; Martínez, Rosario; Ortuño, Inmaculada; Sánchez-Solís, Manuel; Ros, Gaspar; Romero, Fernando; Abellán, Pedro

2013-11-01

Infant cereals are often the elected foodstuff for beginning complementary feeding and provide carbohydrates which are different to those found in maternal milk. The objective of this preliminary study was to ascertain the colonic effects of two infant cereals, with different carbohydrate profiles, in a randomised and double-blind trial in healthy infants. Nineteen term infants between 6.3 and 9.8 months of age were enrolled, after written informed consent was obtained from parents. Ten subjects were allocated to take infant cereal A and nine, infant cereal B. An intervention period was 2 months, with five visits every 15 days, to take anthropometric measurements and faeces samples for the analysis of microbiota, short-chain fatty acids concentration (SCFA), pH value and secretory immunoglobulin A (sIgA). An adequate growth and stool frequency was registered in both intervention groups. Faecal counts of Bifidobacterium, Lactobacillus, Enterobacteriaceae, Enterococcus, Clostridium and Bacteroides did not show any statistical differences. However, a significantly (P < 0.05) higher butyric acid and sIgA, and lower faecal pH were observed in infants who had ingested infant cereal A, with a higher ratio complex/simple carbohydrates. In conclusion, small changes in the carbohydrate profile of infant cereals could lead to significant differences in parameters related to fermentative activity of intestinal microbiota.

[Co-adaptation of enzymatic systems of cells and blood supply in smooth muscle tumors of the corpus uteri].

PubMed

Lazaarev, A F; Avbalian, A M; Bobrov, I P; Klimachev, V V; Mischenko, E V

2008-01-01

We investigated co-adaptation of enzymatic systems of cells using data on activity of NAD(Ph)-dependent enzymes and AgNOR proteins of vascular endothelium vis-a-vis angiogenesis in benign and malignant smooth muscle tumors of the corpus uteri. Overall metabolic activity (NAD-H2 diaphorase) was found to directly correlate with angiogenesis and endothelial vessel proliferation (r = 0.76 and 0.84, respectively). SDH-regulated oxidation in the main metabolic succession of a tricarbonic acid cycle depended on blood supply and endothelial vessel proliferation (r = 0.84 and 0.92, respectively; p = 0.04). A similar relationship was shown for anaerobic glycolysis of SDH (LDH content), on the one hand, and blood supply and endothelial vessel proliferation(r = 0.57 and 0.70, respectively; p = 0.02), on the other. Hence, metabolic profile varied in unaltered myometrium and tumor with variable cellular density and peculiar extracellular matrix. The highest levels of metabolic activity with NAD(Ph)-dependent enzyme co-adaptation was observed in sarcomas which were also characterized by the highest vascular density for endothelial proliferation.

Multitaxon activity profiling reveals differential microbial response to reduced seawater pH and oil pollution.

PubMed

Coelho, Francisco J R C; Cleary, Daniel F R; Costa, Rodrigo; Ferreira, Marina; Polónia, Ana R M; Silva, Artur M S; Simões, Mário M Q; Oliveira, Vanessa; Gomes, Newton C M

2016-09-01

There is growing concern that predicted changes to global ocean chemistry will interact with anthropogenic pollution to significantly alter marine microbial composition and function. However, knowledge of the compounding effects of climate change stressors and anthropogenic pollution is limited. Here, we used 16S and 18S rRNA (cDNA)-based activity profiling to investigate the differential responses of selected microbial taxa to ocean acidification and oil hydrocarbon contamination under controlled laboratory conditions. Our results revealed that a lower relative abundance of sulphate-reducing bacteria (Desulfosarcina/Desulfococcus clade) due to an adverse effect of seawater acidification and oil hydrocarbon contamination (reduced pH-oil treatment) may be coupled to changes in sediment archaeal communities. In particular, we observed a pronounced compositional shift and marked reduction in the prevalence of otherwise abundant operational taxonomic units (OTUs) belonging to the archaeal Marine Benthic Group B and Marine Hydrothermal Vent Group (MHVG) in the reduced pH-oil treatment. Conversely, the abundance of several putative hydrocarbonoclastic fungal OTUs was higher in the reduced pH-oil treatment. Sediment hydrocarbon profiling, furthermore, revealed higher concentrations of several alkanes in the reduced pH-oil treatment, corroborating the functional implications of the structural changes to microbial community composition. Collectively, our results advance the understanding of the response of a complex microbial community to the interaction between reduced pH and anthropogenic pollution. In future acidified marine environments, oil hydrocarbon contamination may alter the typical mixotrophic and k-/r-strategist composition of surface sediment microbiomes towards a more heterotrophic state with lower doubling rates, thereby impairing the ability of the ecosystem to recover from acute oil contamination events. © 2016 John Wiley & Sons Ltd.

Dynamic Vertical Profiles of Peat Porewater Chemistry in a Northern Peatland

DOE Office of Scientific and Technical Information (OSTI.GOV)

Griffiths, Natalie A.; Sebestyen, Stephen D.

We measured pH, cations, nutrients, and total organic carbon (TOC) over 3 years to examine weekly to monthly variability in porewater chemistry depth profiles (0–3.0 m) in an ombrotrophic bog in Minnesota, USA. We also compared temporal variation at one location to spatial variation in depth profiles at 16 locations across the bog. Most solutes exhibited large gradients with depth. pH increased by two units and calcium concentrations increased over 20 fold with depth, and may reflect peatland development from minerotrophic to ombrotrophic conditions. Ammonium concentrations increased almost 20 fold and TOC concentrations decreased by half with depth, and thesemore » patterns likely reflect mineralization of peat or decomposition of TOC. There was also considerable temporal variation in the porewater chemistry depth profiles. Ammonium, soluble reactive phosphorus, and potassium showed greater temporal variation in near-surface porewater, while pH, calcium, and TOC varied more at depth. This variation demonstrates that deep peat porewater chemistry is not static. Lastly, temporal variation in solute chemistry depth profiles was greater than spatial variation in several instances, especially in shallow porewaters. In conclusion, characterizing both temporal and spatial variability is necessary to ensure representative sampling in peatlands, especially when calculating solute pools and fluxes and parameterizing process-based models.« less

Dynamic Vertical Profiles of Peat Porewater Chemistry in a Northern Peatland

DOE PAGES

Griffiths, Natalie A.; Sebestyen, Stephen D.

2016-10-14

We measured pH, cations, nutrients, and total organic carbon (TOC) over 3 years to examine weekly to monthly variability in porewater chemistry depth profiles (0–3.0 m) in an ombrotrophic bog in Minnesota, USA. We also compared temporal variation at one location to spatial variation in depth profiles at 16 locations across the bog. Most solutes exhibited large gradients with depth. pH increased by two units and calcium concentrations increased over 20 fold with depth, and may reflect peatland development from minerotrophic to ombrotrophic conditions. Ammonium concentrations increased almost 20 fold and TOC concentrations decreased by half with depth, and thesemore » patterns likely reflect mineralization of peat or decomposition of TOC. There was also considerable temporal variation in the porewater chemistry depth profiles. Ammonium, soluble reactive phosphorus, and potassium showed greater temporal variation in near-surface porewater, while pH, calcium, and TOC varied more at depth. This variation demonstrates that deep peat porewater chemistry is not static. Lastly, temporal variation in solute chemistry depth profiles was greater than spatial variation in several instances, especially in shallow porewaters. In conclusion, characterizing both temporal and spatial variability is necessary to ensure representative sampling in peatlands, especially when calculating solute pools and fluxes and parameterizing process-based models.« less

Biochemical characterization of recombinant mevalonate kinase from Bacopa monniera.

PubMed

Kumari, Uma; Vishwakarma, Rishi K; Sonawane, Prashant; Abbassi, Shakeel; Khan, Bashir M

2015-01-01

Mevalonate kinase (MK; ATP: mevalonate 5-phosphotransferase; EC 2.7.1.36) plays a key role in isoprenoid biosynthetic pathway in plants. MK catalyzes the phosphorylation of mevalonate to form mevalonate-5-phosphate. The recombinant BmMK was cloned and over-expressed in E. coli BL21 (DE3), and purified to homogeneity by affinity chromatography followed by gel filtration. Optimum pH and temperature for forward reaction was found to be 7.0 and 30 °C, respectively. The enzyme was most stable at pH 8 at 25 °C with deactivation rate constant (Kd*) 1.398 × 10(-4) and half life (t1/2) 49 h. pH activity profile of BmMK indicates the involvement of carboxylate ion, histidine, lysine, arginine or aspartic acid at the active site of enzyme. Activity of recombinant BmMK was confirmed by phosphorylation of RS-mevalonate in the presence of Mg(2+), having Km and Vmax 331.9 μM and 719.1 pKat μg(-1), respectively. The values of kcat and kcat/Km for RS-mevalonate were determined to be 143.82 s(-1) and 0.43332 M(-1) s(-1) and kcat and kcat/Km values for ATP were found 150.9 s(-1) and 1.023 M(-1) s(-1). The metal ion studies suggested that BmMK is a metal dependent enzyme and highly active in the presence of MgCl2. Copyright © 2014 Elsevier B.V. All rights reserved.

Bacopa monniera recombinant mevalonate diphosphate decarboxylase: Biochemical characterization.

PubMed

Abbassi, Shakeel J; Vishwakarma, Rishi K; Patel, Parth; Kumari, Uma; Khan, Bashir M

2015-08-01

Mevalonate diphosphate decarboxylase (MDD; EC 4.1.1.33) is an important enzyme in the mevalonic acid pathway catalyzing the Mg(2+)-ATP dependant decarboxylation of mevalonate 5-diphosphate (MVAPP) to isopentenyl diphosphate (IPP). Bacopa monniera recombinant MDD (BmMDD) protein was overexpressed in Escherichia coli BL21 (DE3) strain and purified to apparent homogeneity. Km and Vmax for MVAPP were 144 μM and 52 U mg(-1) respectively. The values of turnover (kcat) and kcat/Km for mevalonate 5-diphosphate were determined to be 40s(-1) and 2.77×10(5) M(-1) s(-1) and kcat and kcat/Km values for ATP were found to be 30 s(-1) and 2.20×10(4) M(-1) s(-1), respectively. pH activity profile indicated the involvement of carboxylate ion, lysine and arginine for the activity of enzyme. The apparent activation energy for the BmMDD catalyzed reaction was 12.7 kJ mol(-1). Optimum pH and temperature for the forward reaction was found to be 8.0 and 45 °C. The enzyme was most stable at pH 7 at 20 °C with the deactivation rate constant (Kd(*)) of 1.69×10(-4) and half life (t1/2) of 68 h. The cation studies suggested that BmMDD is a cation dependant enzyme and optimum activity was achieved in the presence of Mg(2+). Copyright © 2015 Elsevier B.V. All rights reserved.

A novel NhaD-type Na+/H+ antiporter from the moderate halophile and alkaliphile Halomonas alkaliphila.

PubMed

Wang, Yanhong; Song, Na; Yang, Lina; Abdel-Motaal, Heba; Zhang, Rui; Zhang, Zhenglai; Meng, Fankui; Jiang, Juquan

2017-07-01

In this study, a NhaD-type Na + /H + antiporter gene designated Ha-nhaD was obtained by selection of genomic DNA from the moderate halophile and alkaliphile Halomonas alkaliphila in Escherichia coli KNabc lacking 3 major Na + /H + antiporters. The presence of Ha-NhaD conferred tolerance of E. coli KNabc to NaCl up to 0.6 mol·L -1 and to LiCl up to 0.2 mol·L -1 and to an alkaline pH. pH-dependent Na + (Li + )/H + antiport activity was detected from everted membrane vesicles prepared from E. coli KNabc/pUC-nhaD but not those of KNabc/pUC18. Ha-NhaD exhibited Na + (Li + )/H + antiport activity over a wide pH range from 7.0 to 9.5, with the highest activity at pH 9.0. Protein sequence alignment and phylogenetic analysis revealed that Ha-NhaD is significantly different from the 7 known NhaD-type Na + /H + antiporters, including Dw-NhaD, Dl-NhaD, Vp-NhaD, Vc-NhaD, Aa-NhaD, He-NhaD, and Ha-NhaD1. Although Ha-NhaD showed a closer phylogenetic relationship with Ha-NhaD2, a significant difference in pH-dependent activity profile exists between Ha-NhaD and Ha-NhaD2. Taken together, Ha-nhaD encodes a novel pH-dependent NhaD-type Na + /H + antiporter.

Evaluation of fatty acid profile and biodiesel properties of microalga Scenedesmus abundans under the influence of phosphorus, pH and light intensities.

PubMed

Mandotra, S K; Kumar, Pankaj; Suseela, M R; Nayaka, S; Ramteke, P W

2016-02-01

The present study dealt with biomass, lipid concentration, fatty acid profile and biodiesel properties of microalga Scenedesmus abundans under different phosphate concentrations, pH and light intensities, one at a time. Among different phosphate concentrations, higher biomass (770.10±11.0mg/L) and lipid concentration (176.87±4.6mg/L) were at the concentration of 60mg/L. Light intensity at 6000lux yielded higher biomass and lipid concentration of 742.0±9.7 and 243.15±9.1mg/L, respectively. The biomass (769.0±12.3mg/L) and lipid (179.47±5.5mg/L) concentration were highest at pH 8 and pH 6, respectively. All the culture treatments showed marked effect on the fatty acid profile and biodiesel properties of the extracted oil. FAME derived biodiesel properties were compared with European biodiesel standards (EN 14214), Indian biodiesel standards (IS 15607) and American biodiesel standards (ASTM D 6751-08) to assess the suitability of algal oil as biodiesel feedstock. Copyright © 2015. Published by Elsevier Ltd.

Pharmaceutical Equivalence of Clarithromycin Oral Dosage Forms Marketed in Nairobi County, Kenya

PubMed Central

Manani, Rebecca O.; Abuga, Kennedy O.; Chepkwony, Hezekiah K.

2017-01-01

Clarithromycin is a broad-spectrum semi-synthetic macrolide indicated for treatment of pneumonias, Helicobacter pylori, and chlamydial and skin infections. The object of this study was to evaluate the pharmaceutical equivalence of 14 generic clarithromycin products marketed in Nairobi County, Kenya, to the innovator products, using in vitro dissolution profiles and similarity factors (f2). Further, dissolution profiles of four innovator formulations manufactured in different sites were compared. Fourteen clarithromycin tablets/capsules and four suspensions were subjected to assay and comparative dissolution runs at pH 1.2, 4.5 and 6.8, for 60 and 90 min, respectively. All products complied with pharmacopoeial assay specifications. However, significant differences were observed in their dissolution profiles. The non-compliance rates for tablets/capsules were 50% at pH 1.2, 33% at pH 4.5 and 50% at pH 6.8, while none of the four suspensions were compliant. Overall, only four (25%) products complied with the specifications for similarity factor. The results obtained indicate that a significant percentage of generic clarithromycin products are pharmaceutically non-equivalent to the innovator products, and that assay and single-point dissolution tests are insufficient demonstration of equivalence between the generic and innovator products. PMID:28445444

pH Regulation of Electrogenic Sugar/H+ Symport in MFS Sugar Permeases

PubMed Central

Bazzone, Andre; Madej, M. Gregor; Kaback, H. Ronald

2016-01-01

Bacterial sugar symporters in the Major Facilitator Superfamily (MFS) use the H+ (and in a few cases Na+) electrochemical gradients to achieve active transport of sugar into the cell. Because a number of structures of MFS sugar symporters have been solved recently, molecular insight into the transport mechanism is possible from detailed functional analysis. We present here a comparative electrophysiological study of the lactose permease (LacY), the fucose permease (FucP) and the xylose permease (XylE), which reveals common mechanistic principles and differences. In all three symporters energetically downhill electrogenic sugar/H+ symport is observed. Comparison of the pH dependence of symport at symmetrical pH exhibits broad bell-shaped pH profiles extending over 3 to 6 pH units and a decrease at extremely alkaline pH ≥ 9.4 and at acidic to neutral pH = 4.6–7.5. The pH dependence can be described by an acidic to neutral apparent pK (pKapp) and an alkaline pKapp. Experimental evidence suggests that the alkaline pKapp is due to H+ depletion at the protonation site, while the acidic pKapp is due to inhibition of deprotonation. Since previous studies suggest that a single carboxyl group in LacY (Glu325) may be the only side chain directly involved in H+ translocation and a carboxyl side chain with similar properties has been identified in FucP (Asp46) and XylE (Asp27), the present results imply that the pK of this residue is switched during H+/sugar symport in all three symporters. PMID:27227677

Effect of amino acid sequence and pH on nanofiber formation of self-assembling peptides EAK16-II and EAK16-IV.

PubMed

Hong, Yooseong; Legge, Raymond L; Zhang, S; Chen, P

2003-01-01

Atomic force microscopy (AFM) and axisymmetric drop shape analysis-profile (ASDA-P) were used to investigate the mechanism of self-assembly of peptides. The peptides chosen consisted of 16 alternating hydrophobic and hydrophilic amino acids, where the hydrophilic residues possess alternating negative and positive charges. Two types of peptides, AEAEAKAKAEAEAKAK (EAK16-II) and AEAEAEAEAKAKAKAK (EAK16-IV), were investigated in terms of nanostructure formation through self-assembly. The experimental results, which focused on the effects of the amino acid sequence and pH, show that the nanostructures formed by the peptides are dependent on the amino acid sequence and the pH of the solution. For pH conditions around neutrality, one of the peptides used in this study, EAK16-IV, forms globular assemblies and has lower surface tension at air-water interfaces than another peptide, EAK16-II, which forms fibrillar assemblies at the same pH. When the pH is lowered below 6.5 or raised above 7.5, there is a transition from globular to fibrillar structures for EAK16-IV, but EAK16-II does not show any structural transition. Surface tension measurements using ADSA-P showed different surface activities of peptides at air-water interfaces. EAK16-II does not show a significant difference in surface tension for the pH range between 4 and 9. However, EAK16-IV shows a noticeable decrease in surface tension at pH around neutrality, indicating that the formation of globular assemblies is related to the molecular hydrophobicity.

Application of arteether-loaded polyurethane nanomicelles to induce immune response in breast cancer model.

PubMed

Jabbarzadegan, Mona; Rajayi, Hajar; Mofazzal Jahromi, Mirza Ali; Yeganeh, Hamid; Yousefi, Mehdi; Muhammad Hassan, Zuhair; Majidi, Jafar

2017-06-01

To concentrate a potent anticancer drug (Arteether) in tumor microenvironment, we encapsulated it in biodegradable and pH sensitive polyurethane (PU) nanomicelles (NMs). The nanocomplex was characterized by Fourier transform infrared (FTIR), dynamic light scattering (DLS). The loading capacity and release profile in pH of 5.4 and 7.4 were considered. The cytotoxicity effect was evaluated in vitro and in vivo settings. The level of IFN-γ and IL-4 cytokines of mice splenocytes were assessed by enzyme-linked immunosorbent assay (ELISA). The nanocomplex showed negative zeta charge of -26.2 mV, size of 42.30 nm and high loading capacity (92%). Release profile showed a faster rate of drug liberation at pH 5.4 as compared to that of pH 7.4. It indicated significant inhibitory effect on the growth of 4T1 cell line and increased IFN-γ level.

Observing Physical and Biological Drivers of pH and O2 in a Seasonal Ice Zone in the Ross Sea Using Profiling Float Data

NASA Astrophysics Data System (ADS)

Briggs, E.; Martz, T. R.; Talley, L. D.; Mazloff, M. R.

2015-12-01

Ice cover has strong influence over gas exchange, vertical stability, and biological production which are critical to understanding the Southern Ocean's central role in oceanic biogeochemical cycling and heat and carbon uptake under a changing climate. However the relative influence of physical versus biological processes in this hard-to-study region is poorly understood due to limited observations. Here we present new findings from a profiling float equipped with biogeochemical sensors in the seasonal ice zone of the Ross Sea capturing, for the first time, under-ice pH profile data over a two year timespan from 2014 to the present. The relative influence of physical (e.g. vertical mixing and air-sea gas exchange) and biological (e.g. production and respiration) drivers of pH and O2 within the mixed layer are explored during the phases of ice formation, ice cover, and ice melt over the two seasonal cycles. During the austral fall just prior to and during ice formation, O2 increases as expected due to surface-layer undersaturation and enhanced gas exchange. A small increase in pH is also observed during this phase, but without a biological signal in accompanying profiling float chlorophyll data, which goes against common reasoning from both a biological and physical standpoint. During the phase of ice cover, gas exchange is inhibited and a clear respiration signal is observed in pH and O2 data from which respiration rates are calculated. In the austral spring, ice melt gives rise to substantial ice edge phytoplankton blooms indicated by O2 supersaturation and corresponding increase in pH and large chlorophyll signal. The influence of the duration of ice cover and mixed layer depth on the magnitude of the ice edge blooms is explored between the two seasonal cycles.

[Gene expression and activity regulation of two calmodulin binding protein kinases in tobacco seedling].

PubMed

Hua, Wei; Li, Rong-Jun; Liang, Shu-Ping; Lu, Ying-Tang

2005-06-01

Two different calmodulin-binding protein kinase cDNAs (NtCBK1/2) have been isolated from tobacco. To understand the CBK protein activity regulation, we compared the activity regulation of NtCBK1 and NtCBK2 by pH, Mg(2+) concentration and Na(+) concentration. We found the autophosphorylation of NtCBK1/2 reached the maximum in pH 7.5 and 8 respectively; Mg(2+) and Na(+) shown different effects on the activity of NtCBKs, high and low Mg(2+) concentrations both inhibited the activity of NtCBKs, but Na+ had little effect on the kinase activity. In addition, to obtain further insight about the physiological roles of individual NtCBKs, we detected the expression profiles of CBKs. The results revealed different patterns of expression of NtCBK1 and NtCBK2. Both are largely expressed in leaf and flower; but in stem and root, NtCBK1 gene had stronger expression than NtCBK2. NtCBK2 expression was induced by GA treatment, while NtCBK1 expression remained unchanged under GA treatment. Expression of both NtCBK1 and NtCBK2 increased in response to salt stress, the former to a greater extent, and both expressions did not change under high/low temperature, drought, NAA and ABA treatments.

[Preparation of coated tablets of glycyrrhetic acid-HP-beta-cyclodextrin tablets for colon-specific release].

PubMed

Cui, Qi-Hua; Cui, Jing-Hao; Zhang, Jin-Jin

2008-10-01

To prepare coated tablets of glycyrrhetinic acid and hydroxypropyl-beta-cyclodextrin (GTA-HP-beta-CYD) inclusion complex tablets for colon-specific release. In order to improve the solubility of GTA, the GTA-HP-beta-CYD inclusion complex was prepared by ultrasonic-lyophilization technique and its formation were characterized by X-ray powder diffraction profiles and infrared spectrometry. The effects of inclusion condition on the inclusion efficiency and stability coefficient of inclusion complex were investigated, respectively. After prepared GTA-HP-beta-CYD tablets by powder direct compression, the pH dependant polymer Eudragit III and/or mixed with Eudragit II were used for further coating materials in fluid-bed coater. The influences of coating weight on the GTA release in different pH conditions were evaluated to establish the method for prepering colon specific delivery tablets with pulsed release properties. The formation of inclusion complexes were proved by X-ray powder diffraction profile and phase solubility curve. The effect of pH value of solvent was played critical role on the preparation of GTA- HP-beta-CYD inclusion complex. And the inclusion efficiency of GTA was 9. 3% and the solubility was increased to 54. 6 times at optimized method. The Eudragit III coated GTA- HP-beta-CYD tablets with coating weight 10% and 16% were showed pH dependant colon specific release profiles with slow release rate. The release profile of tablets coated with the mixture of Eudragit II and Eudragit III (1:2) were indicated typical pH dependant colon specific and pulsed release properties while the coating weight was 17%. The preliminary method for preparation of colon specific release tablets containing glycyrrhetinic acid with improved solubility was established for further in vivo therapeutic experiment.

2H Kinetic Isotope Effects and pH Dependence of Catalysis as Mechanistic Probes of Rat Monoamine Oxidase A: Comparisons with the Human Enzyme‡

PubMed Central

Wang, Jin; Edmondson, Dale E.

2011-01-01

Monoamine oxidase A (MAO A) is a mitochondrial outer membrane-bound flavoenzyme important in the regulation of serotonin and dopamine levels. Since the rat is extensively used as an animal model in drug studies, it is important to understand how rat MAO A behaves in comparison with the more extensively studied human enzyme. For many reversible inhibitors, rat MAO A exhibits Ki values similar to those of human MAO A. The pH profile of kcat for rat MAO A shows a pKa of 8.2±0.1 for the benzylamine ES complex and pKa values of 7.5±0.1 and 7.6±0.1 for the respective ES complexes with p-CF3-1H and p-CF3-2H-benzylamine. In contrast to the human enzyme, the rat enzyme exhibits a single pKa value (8.3±0.1) with kcat/Km benzylamine vs. pH and pKa values of 7.8±0.1 and 8.1±0.2 are found for the ascending limbs, respectively, of kcat/Km vs. pH profiles for p-CF3-1H and p-CF3-2H-benzylamine and 9.3±0.1 and 9.1±0.2 for their respective descending limbs. The oxidation of para-substituted benzylamine substrate analogues by rat MAO A exhibit large deuterium kinetic isotope effects on kcat and on kcat/Km. These effects are pH-independent, and range from 7 to 14, demonstrating a rate-limiting α-C-H bond cleavage step in catalysis. Quantitative structure-activity correlations of log kcat with the electronic substituent parameter (σ) at pH 7.5 and at 9.0 show a dominant contribution with positive ρ values (+1.2 – 1.3) and a pH-independent negative contribution from the steric term. Quantitative structure-activity relationship analysis of the binding affinities of the para-substituted benzylamine analogues to rat MAO A show an increased van der Waals volumes (Vw) increases the affinity of the deprotonated amine for the enzyme. These results demonstrate that rat MAO A exhibits similar but not identical functional properties with the human enzyme and provide additional support for C-H bond cleavage via a polar nucleophilic mechanism. PMID:21819071

Design of sustained release tablet containing fucoidan.

PubMed

Tran, Thao Truong-Dinh; Ngo, Dai Kieu-Phuong; Vo, Toi Van; Tran, Phuong Ha-Lien

2015-01-01

The study introduced a new therapeutic agent, fucoidan, which can offer potential medical treatments including anti-inflammatory and anti-coagulant activities, as well as anti-proliferative effects on cancer cells. Fucoidan was included in sustained release formulations expected for an effective plasma drug concentration for approximately 24 h. The matrices based on the two polymers hydroxypropyl methycellulose (HPMC) and polyethylene oxide (PEO) were prepared with various ratios between the polymers and fucoidan. The dissolution profiles of various matrix tablets performed in enzyme-free simulated intestinal fluid (pH 6.8) for 24 h indicated a higher potential of PEO-based matrix tablets in sustaining release of fucoidan. The swelling and erosion of the tablets were also characterized to elucidate the difference among those dissolution profiles.

Comparative chemical profiling, cholinesterase inhibitions and anti-radicals properties of essential oils from Polygonum hydropiper L: a preliminary anti- Alzheimer's study.

PubMed

Ayaz, Muhammad; Junaid, Muhammad; Ullah, Farhat; Sadiq, Abdul; Khan, Mir Azam; Ahmad, Waqar; Shah, Muhammad Raza; Imran, Muhammad; Ahmad, Sajjad

2015-11-04

Cholinesterase inhibition is a vital target for the development of novel and mechanism based inhibitors, owing to their role in the breakdown of acetylcholine (ACh) neurotransmitter to treat various neurological disorders including Alzheimer's disease (AD). Similarly, free radicals are implicated in the progression of various diseases like neurodegenerative disorders. Due to lipid solubility and potential to easily cross blood brain barrier, this study was designed to investigate the anticholinesterase and antioxidant potentials of the standardized essential oils from the leaves and flowers of Polygonum hydropiper. Essential oils from the leaves (Ph.LO) and flowers (Ph.FO) of P. hdropiper were isolated using Clevenger apparatus. Oil samples were analyzed by GC-MS to identify major components and to attribute the antioxidant and anticholinesterase activity to specific components. Acetylcholinesterase (AChE) and butyrylcholinesterase (BChE) inhibitory potentials of the samples were determined following Ellman's assay. Antioxidant assays were performed using 1,1-diphenyl,2-picrylhydrazyl (DPPH), 2,2-azinobis[3-ethylbenzthiazoline]-6-sulfonic acid (ABTS) and hydrogen peroxide (H2O2) free radical scavenging assays. In the GC-MS analysis 141 and 122 compounds were indentified in Ph.LO and Ph.FO respectively. Caryophylene oxide (41.42 %) was the major component in Ph.FO while decahydronaphthalene (38.29 %) was prominent in Ph.LO. In AChE inhibition, Ph.LO and Ph.FO exhibited 87.00** and 79.66***% inhibitions at 1000 μg/ml with IC50 of 120 and 220 μg/ml respectively. The IC50 value for galanthamine was 15 μg/ml. In BChE inhibitory assay, Ph.LO and Ph.FO caused 82.66*** (IC50 130 μg/ml) and 77.50***% (IC50 225 μg/ml) inhibitions respectively at 1000 μg/ml concentration. In DPPH free radical scavenging assay, Ph.LO and Ph.FO exhibited IC50 of 20 and 200 μg/ml respectively. The calculated IC50s were 180 & 60 μg/ml for Ph.LO, and 45 & 50 μg/ml for Ph.FO in scavenging of ABTS and H2O2 free radicals respectively. In the current study, essential oils from leaves and flowers of P. hydropiper exhibited dose dependent anticholinesterase and antioxidant activities. Leaves essential oil were more effective and can be subjected to further in-vitro and in-vivo anti-Alzheimer's studies.

Monomer volume fraction profiles in pH responsive planar polyelectrolyte brushes

DOE PAGES

Mahalik, Jyoti P.; Yang, Yubo; Deodhar, Chaitra V.; ...

2016-03-06

Spatial dependencies of monomer volume fraction profiles of pH responsive polyelectrolyte brushes were investigated using field theories and neutron reflectivity experiments. In particular, planar polyelectrolyte brushes in good solvent were studied and direct comparisons between predictions of the theories and experimental measurements are presented. The comparisons between the theories and the experimental data reveal that solvent entropy and ion-pairs resulting from adsorption of counterions from the added salt play key roles in affecting the monomer distribution and must be taken into account in modeling polyelectrolyte brushes. Furthermore, the utility of this physics-based approach based on these theories for the predictionmore » and interpretation of neutron reflectivity profiles in the context of pH responsive planar polyelectrolyte brushes such as polybasic poly(2-(dimethylamino)ethyl methacrylate) (PDMAEMA) and polyacidic poly(methacrylic acid) (PMAA) brushes is demonstrated. The approach provides a quantitative way of estimating molecular weights of the polymers polymerized using surface-initiated atom transfer radical polymerization.« less

T-REX: education and formation

NASA Astrophysics Data System (ADS)

Marconi, A.; Marano, B.

An important goal of the T-REX project is the creation and the formation of new experts and professional profiles with extremely high specialisation to prepare tomorrow astronomers and instrument scientists that will take full advantage of E-ELT and that will make the Italian participation a success. The existing tight interaction and integration between INAF institutes and universities as allowed the T-REX Project to fund and support a total of 4 PhD grants. Here we briefly summarise their present and future activities.

Main features of anthropogenic inner-urban soils in Szeged, Hungary

NASA Astrophysics Data System (ADS)

Puskás, Irén.; Farsang, Andrea

2010-05-01

At the beginning of the 21st century, due to the intensive urbanization it is necessary to gather more and more information on altered physical, chemical and biological parameters of urban soils in order to ensure their suitable management and protection for appropriate living conditions. Nowadays, these measures are very relevant since negative environmental effects can modify the soil forming factors in cities. Szeged, the 4th largest city of Hungary, proved to be an ideal sampling area for the research of urban soils since its original surface has been altered by intensive anthropogenic activities. The main objectives of my research are the investigation, description and evaluation of the altered soils in Szeged. For the physical and chemical analysis (humus, nitrogen, carbonate content, heavy metals, pH, artefacts etc.) of soils 124 samples were taken from the horizons of 25 profiles in Szeged and its peripherals (as control samples). The profiles were sampled at sites affected by different extent of artificial infill according to infill maps (1. profiles fully made up of infill; 2. so-called mixed profiles consisting of considerable amount of infill material and buried soil horizons; 3. natural profiles located in the peripherals of the city). With the help of the above-mentioned parameters, the studied soils of Szeged were assigned into the classification system of WRB(2006), which classifies the soils of urban and industrial areas as an individual soil group (under the term Technosols) for the first time. In accordance with the WRB(2006) nomenclature three main soil types can be identified in Szeged with respect to the degree of human influence: profiles slightly influenced, strongly modified, completely altered by human activities. During this poster, we present the peculiarities of typical urban profiles strongly and completely altered by human influence. Most profiles were placed into the group of Technosols due to the considerable transformation of their diagnostic properties (e.g. coverage by artificial objects, intensive compaction, horizontal and vertical variability, abrupt colour and textural changes usually high amount of artefacts, irregular fluctuation of diagnostic properties along the profiles, anthropogenic parent material, high pH and carbonate content, poor humus quality, mainly sand, sandy loam texture etc.). Transformations were best reflected by suffixes such as Ekranic, Urbic, Linic. Among the suffix qualifiers Calcaric, Ruptic, Densic and Arenic were used the most frequently. Furthermore, we found that some of the studied profiles were not situated in the city centre. Consequently, the location of these profiles in the city centre is not necessary since local influences can overwhelm the effect of artificial infill. Considering all the profiles, two of them in city centre can be consider to be the most anthropogenic: profile No. 11 [Ekranic Technosol (Ruptic, Toxic, Endoclayic)] and profile No. 22 [Urbic Technosol (Calcaric, Ruptic, Densic, Arenic)]. It can be claimed that profile No. 11 with "technic hard rock" has the least chance to experience pedogenetic processes since the horizons are covered by thick, surface artificial object, and isolated from the outside world. However, in case of profile No. 22 with dense vegetation and without surface artificial object, the high amount of artefact inhibits pedogenesis.

The effect of pH control and 'hydraulic flush' on hydrolysis and Volatile Fatty Acids (VFA) production and profile in anaerobic leach bed reactors digesting a high solids content substrate.

PubMed

Cysneiros, Denise; Banks, Charles J; Heaven, Sonia; Karatzas, Kimon-Andreas G

2012-11-01

The effect of hydraulic flush and pH control on hydrolysis, Volatile Fatty Acids (VFA) production and profile in anaerobic leach bed reactors was investigated for the first time. Six reactors were operated under different regimes for two consecutive batches of 28days each. Buffering at pH ∼6.5 improved hydrolysis (Volatile Solid (VS) degradation) and VFA production by ∼50%. Butyric and acetic acid were dominant when reactors were buffered, while only butyric acid was produced at low pH. Hydraulic flush enhanced VS degradation and VFA production by ∼15% and ∼32%, respectively. Most Probable Number (MPN) of cellulolytic microorganisms indicated a wash out when hydraulic flush was applied, but pH control helped to counteract this. The highest VS degradation (∼89%), VFA yield (0.84kgCODkg(-1)VS(added)) and theoretical methane potential (0.37m(3)CH(4)kg(-1)VS(added)) were obtained when pH control and hydraulic flush were applied, and therefore, these conditions are recommended. Copyright © 2012 Elsevier Ltd. All rights reserved.

Analytical coupled-wave model for photonic crystal surface-emitting quantum cascade lasers.

PubMed

Wang, Zhixin; Liang, Yong; Yin, Xuefan; Peng, Chao; Hu, Weiwei; Faist, Jérôme

2017-05-15

An analytical coupled-wave model is developed for surface-emitting photonic-crystal quantum cascade lasers (PhC-QCLs). This model provides an accurate and efficient analysis of full three-dimensional device structure with large-area cavity size. Various laser properties of interest including the band structure, mode frequency, cavity loss, mode intensity profile, and far field pattern (FFP), as well as their dependence on PhC structures and cavity size, are investigated. Comparison with numerical simulations confirms the accuracy and validity of our model. The calculated FFP and polarization profile well explain the previously reported experimental results. In particular, we reveal the possibility of switching the lasing modes and generating single-lobed FFP by properly tuning PhC structures.

Modeling with the logistic regression of the growth/no growth interface of Saccharomyces cerevisiae in relation to 2 antimicrobial terpenes (citral and linalool), pH, and a(w).

PubMed

Tabanelli, Giulia; Montanari, Chiara; Patrignani, Francesca; Siroli, Lorenzo; Lanciotti, Rosalba; Gardini, Fausto

2014-03-01

The antimicrobial effects of 2 terpenes (citral and linalool) on a Saccharomyces cerevisiae strain isolated from spoiled soft drink have been evaluated, alone or in combination, in relation to pH and aw using in vitro assays. The obtained data were fitted with the logit model to find the growth/no growth boundary regions of the 2 terpenes, focusing the attention on the type of interaction exerted by citral and linalool. In particular, the results showed an increase of citral antimicrobial effect in growth media characterized by low aw value, as well as a higher linalool antimicrobial effect in media at low pH. Moreover, the interactive effects of the 2 terpenes were exploited. The results obtained with the model were validated in an independent experiment. The knowledge of the interactions of essential oil molecules with enhanced antimicrobial activity, in relation to some of the most important chemicophysical variables, can have important industrial applications, since these substances are able to assure the desired antimicrobial effect without negatively modifying the product flavor profile. The effects of the main chemicophysical parameters (such as aw and pH) on the antimicrobial activity of bioactive terpenes are necessary for the definition of an industrially applicable preservation strategy based on the use of essential oils as natural antimicrobials aimed to prolong shelf life of food products. © 2014 Institute of Food Technologists®

Assessment of the in vitro bioactive properties of lactic acid bacteria isolated from native ecological niches of Ecuador.

PubMed

Benavides, Ana B; Ulcuango, Mario; Yépez, Lucía; Tenea, Gabriela N

Lactic acid bacteria are known for their biotechnological potential. In various regions of Ecuador numerous indigenous biological resources are largely undocumented. In this study, we evaluated the potential probiotic characteristics and antagonistic in vitro properties of some lactic acid bacteria from native niches of the subtropical rain forests of Ecuador. These isolates were identified according to their morphological properties, standard API50CH fermentation profile and RAPD-DNA polymorphism pattern. The selected isolates were further evaluated for their probiotic potential. The isolates grew at 15°C and 45°C, survived at a pH ranging from 2.5 to 4.5 in the presence of 0.3% bile (>90%) and grew under sodium chloride conditions. All selected isolates were sensitive to ampicillin, amoxicillin and cefuroxime and some showed resistance to gentamicin, kanamycin and tetracycline. Moreover, the agar well diffusion assay showed that the supernatant of each strain at pH 3.0 and pH 4.0, but not at pH 7.0 exhibited increased antimicrobial activity (inhibition zone >15mm) against two foodborne pathogens, Escherichia coli and Salmonella spp. To our knowledge, this is the first report describing the antagonistic activity against two foodborne pathogens and the probiotic in vitro potential of lactic acid bacteria isolated from native biota of Ecuador. Copyright © 2016 Asociación Argentina de Microbiología. Publicado por Elsevier España, S.L.U. All rights reserved.

The pH profile for acid-induced elongation of coleoptile and epicotyl sections is consistent with the acid-growth theory

NASA Technical Reports Server (NTRS)

Cleland, R. E.; Buckley, G.; Nowbar, S.; Lew, N. M.; Stinemetz, C.; Evans, M. L.; Rayle, D. L.

1991-01-01

The acid-growth theory predicts that a solution with a pH identical to that of the apoplast of auxin-treated tissues (4.5.-5.0) should induce elongation at a rate comparable to that of auxin. Different pH profiles for elongation have been obtained, however, depending on the type of pretreatment between harvest of the sections and the start of the pH-incubations. To determine the acid sensitivity under in vivo conditions, oat (Avena sativa L.) coleoptile, maize (Zea mays L.) coleoptile and pea (Pisum sativum L.) epicotyl sections were abraded so that exogenous buffers could penetrate the free space, and placed in buffered solutions of pH 3.5-6.5 without any preincubation. The extension, without auxin, was measured over the first 3 h. Experiments conducted in three laboratories produced similar results. For all three species, sections placed in buffer without pretreatment elongated at least threefold faster at pH 5.0 than at 6.0 or 6.5, and the rate elongation at pH 5.0 was comparable to that induced by auxin. Pretreatment of abraded sections with pH-6.5 buffer or distilled water adjusted to pH 6.5 or above gave similar results. We conclude that the pH present in the apoplast of auxin-treated coleoptile and stems is sufficiently low to account for the initial growth response to auxin.

Isolation and characterization of acid-sensitive mutants of Pediococcus acidilactici.

PubMed

Kurdi, Peter; Smitinont, Thitapha; Valyasevi, Ruud

2009-02-01

Acid-sensitive mutants of Pediococcus acidilactici BCC 9545, a starter culture of the Thai fermented pork sausage nham, were isolated as spontaneous neomycin resistant mutants. The mutants generally produced less acid and acidified the culture media less than the parent strain in a 72 h culturing period. Interestingly, the ATPase activities of the mutants did not differ considerably from that of the parent strain in acidic conditions. It was also found that the internal pH values of the mutant strains were somewhat lower in neutral environment, while at pH 5.0 their internal pHs were significantly lower compared to the parent's. Inhibiting the H(+)-ATPase activities in energized cells by N,N'-dicyclohexyl carbodiimide also revealed that protons were leaking from the mutants at neutral pH, which increased under acidic conditions. In contrast, the parent strain exhibited a smaller proton leak and only under acidic conditions. The membrane fatty acid analysis of the mutants indicated that under acidic conditions the mutants had a significantly smaller major unsaturated/saturated fatty acids ratio ((C(18:1)+C(18:3n6))/(C(16:0)+C(18:0))) compared to the parent strain's membrane. Taken together, these observations suggest there is a reasonable possibility that the membrane fatty acid profile differences in the mutants resulted in their acid-sensitivity.

Studies of the electron density in the highest occupied molecular orbitals of PH 3, PF 3 and P(CH 3) 3 by electron momentum spectroscopy and Hartree-Fock, MRSD-CI and DFT calculations

NASA Astrophysics Data System (ADS)

Rolke, J.; Brion, C. E.

1996-06-01

The spherically averaged momentum profiles for the highest occupied molecular orbitals of PF 3 and P(CH 3) 3 have been obtained by electron momentum spectroscopy. The measurements provide a stringent test of basis set effects and the quality of ab-initio methods in the description of these larger molecular systems. As in previous work on the methyl-substituted amines, intuitive arguments fail to predict the correct amount of s- and p-type contributions to the momentum profile while delocalized molecular orbital concepts provide a more adequate description of the HOMOs. The experimental momentum profiles have been compared with theoretical momentum profiles calculated at the level of the target Hartree-Fock approximation with a range of basis sets. New Hartree-Fock calculations are also presented for the HOMO of PH 3 and compared to previously published experimental and theoretical momentum profiles. The experimental momentum profiles have further been compared to calculations at the level of the target Kohn-Sham approximation using density functional theory with the local density approximation and also with gradient corrected (non-local) exchange correlation potentials. In addition, total energies and dipole moments have been calculated for all three molecules by the various theoretical methods and compared to experimental values. Calculated 'density difference maps' show the regions where the HOMO momentum and position electron densities of PF 3 and P(CH 3) 3 change relative to the corresponding HOMO density of PH 3. The results suggest that methyl groups have an electron-attracting effect (relative to H) on the HOMO charge density in trimethyl phosphines. These conclusions are supported by a consideration of dipole moments and the 31P NMR chemical shifts for PH 3, PF 3 and P(CH 3) 3.

Evaluation of flocculating performance of a thermostable bioflocculant produced by marine Bacillus sp.

PubMed

Okaiyeto, Kunle; Nwodo, Uchechukwu U; Mabinya, Leonard V; Okoli, Arinze S; Okoh, Anthony I

2016-01-01

This study assessed the bioflocculant (named MBF-W7) production potential of a bacterial isolate obtained from Algoa Bay, Eastern Cape Province of South Africa. The 16S ribosomal deoxyribonucleic acids gene sequence analysis showed 98% sequence similarity to Bacillus licheniformis strain W7. Optimum culture conditions for MBF-W7 production include 5% (v/v) inoculum size, maltose and NH4NO3 as carbon and nitrogen sources of choice, medium pH of 6 as the initial pH of the growth medium. Under these optimal conditions, maximum flocculating activity of 94.9% was attained after 72 h of cultivation. Chemical composition analyses showed that the purified MBF-W7 was a glycoprotein which was predominantly composed of polysaccharides 73.7% (w/w) and protein 6.2% (w/w). Fourier transform infrared spectroscopy revealed the presence of hydroxyl, carboxyl and amino groups as the main functional groups identified in the bioflocculant molecules. Thermogravimetric analyses showed the thermal decomposition profile of MBF-W7. Scanning electron microscopy imaging revealed that bridging played an important role in flocculation. MBF-W7 exhibited excellent flocculating activity for kaolin clay suspension at 0.2 mg/ml over a wide pH range of 3-11; with the maximal flocculation rate of 85.8% observed at pH 3 in the presence of Mn(2+). It maintained and retained high flocculating activity of over 70% after heating at 100°C for 60 min. MBF-W7 showed good turbidity removal potential (86.9%) and chemical oxygen demand reduction efficiency (75.3%) in Tyume River. The high flocculating rate of MBF-W7 makes it an attractive candidate to replace chemical flocculants utilized in water treatment.

Identifying Potential Mechanisms Enabling Acidophily in the Ammonia-Oxidizing Archaeon “Candidatus Nitrosotalea devanaterra”

PubMed Central

Sayavedra-Soto, Luis A.; Gallois, Nicolas; Schouten, Stefan; Stein, Lisa Y.; Prosser, James I.; Nicol, Graeme W.

2016-01-01

Ammonia oxidation is the first and rate-limiting step in nitrification and is dominated by two distinct groups of microorganisms in soil: ammonia-oxidizing archaea (AOA) and ammonia-oxidizing bacteria (AOB). AOA are often more abundant than AOB and dominate activity in acid soils. The mechanism of ammonia oxidation under acidic conditions has been a long-standing paradox. While high rates of ammonia oxidation are frequently measured in acid soils, cultivated ammonia oxidizers grew only at near-neutral pH when grown in standard laboratory culture. Although a number of mechanisms have been demonstrated to enable neutrophilic AOB growth at low pH in the laboratory, these have not been demonstrated in soil, and the recent cultivation of the obligately acidophilic ammonia oxidizer “Candidatus Nitrosotalea devanaterra” provides a more parsimonious explanation for the observed high rates of activity. Analysis of the sequenced genome, transcriptional activity, and lipid content of “Ca. Nitrosotalea devanaterra” reveals that previously proposed mechanisms used by AOB for growth at low pH are not essential for archaeal ammonia oxidation in acidic environments. Instead, the genome indicates that “Ca. Nitrosotalea devanaterra” contains genes encoding both a predicted high-affinity substrate acquisition system and potential pH homeostasis mechanisms absent in neutrophilic AOA. Analysis of mRNA revealed that candidate genes encoding the proposed homeostasis mechanisms were all expressed during acidophilic growth, and lipid profiling by high-performance liquid chromatography–mass spectrometry (HPLC-MS) demonstrated that the membrane lipids of “Ca. Nitrosotalea devanaterra” were not dominated by crenarchaeol, as found in neutrophilic AOA. This study for the first time describes a genome of an obligately acidophilic ammonia oxidizer and identifies potential mechanisms enabling this unique phenotype for future biochemical characterization. PMID:26896134

Identifying Potential Mechanisms Enabling Acidophily in the Ammonia-Oxidizing Archaeon "Candidatus Nitrosotalea devanaterra".

PubMed

Lehtovirta-Morley, Laura E; Sayavedra-Soto, Luis A; Gallois, Nicolas; Schouten, Stefan; Stein, Lisa Y; Prosser, James I; Nicol, Graeme W

2016-05-01

Ammonia oxidation is the first and rate-limiting step in nitrification and is dominated by two distinct groups of microorganisms in soil: ammonia-oxidizing archaea (AOA) and ammonia-oxidizing bacteria (AOB). AOA are often more abundant than AOB and dominate activity in acid soils. The mechanism of ammonia oxidation under acidic conditions has been a long-standing paradox. While high rates of ammonia oxidation are frequently measured in acid soils, cultivated ammonia oxidizers grew only at near-neutral pH when grown in standard laboratory culture. Although a number of mechanisms have been demonstrated to enable neutrophilic AOB growth at low pH in the laboratory, these have not been demonstrated in soil, and the recent cultivation of the obligately acidophilic ammonia oxidizer "Candidatus Nitrosotalea devanaterra" provides a more parsimonious explanation for the observed high rates of activity. Analysis of the sequenced genome, transcriptional activity, and lipid content of "Ca Nitrosotalea devanaterra" reveals that previously proposed mechanisms used by AOB for growth at low pH are not essential for archaeal ammonia oxidation in acidic environments. Instead, the genome indicates that "Ca Nitrosotalea devanaterra" contains genes encoding both a predicted high-affinity substrate acquisition system and potential pH homeostasis mechanisms absent in neutrophilic AOA. Analysis of mRNA revealed that candidate genes encoding the proposed homeostasis mechanisms were all expressed during acidophilic growth, and lipid profiling by high-performance liquid chromatography-mass spectrometry (HPLC-MS) demonstrated that the membrane lipids of "Ca Nitrosotalea devanaterra" were not dominated by crenarchaeol, as found in neutrophilic AOA. This study for the first time describes a genome of an obligately acidophilic ammonia oxidizer and identifies potential mechanisms enabling this unique phenotype for future biochemical characterization. Copyright © 2016 Lehtovirta-Morley et al.

Development of an Integrated ISFET pH Sensor for High Pressure Applications in the Deep-Sea

DTIC Science & Technology

2012-09-30

Measurements in the upper ocean suggest that sensor precision is comparable to the annual pH change due to ocean acidification (Fig. 2). An array of...profiling floats equipped with pH sensors would be capable of directly monitoring the process of ocean acidification . Further refinement of the sensor...Quality of Life The high pressure pH sensor will have direct applications to our understanding of ocean acidification and the impacts on ecosystem

Morphology and pH changes in leached solidified/stabilized waste forms

DOE Office of Scientific and Technical Information (OSTI.GOV)

Cheng, K.Y.; Bishop, P.L.

1996-12-31

Leaching of cement-based waste forms in acetic acid solutions with different acidic strengths has been investigated in this work. The examination of the morphology and pH profile along the acid penetration route by an optical microscope and various pH color indicators is reported. A clear-cut leaching boundary, where the pH changes from below 6 in the leached surface layers to above 12 in the unleached waste form, was observed in every leached sample.

Growth, survival, and peptidolytic activity of Lactobacillus plantarum I91 in a hard-cheese model.

PubMed

Bergamini, C V; Peralta, G H; Milesi, M M; Hynes, E R

2013-09-01

In this work, we studied the growth, survival, and peptidolytic activity of Lactobacillus plantarum I91 in a hard-cheese model consisting of a sterile extract of Reggianito cheese. To assess the influence of the primary starter and initial proteolysis level on these parameters, we prepared the extracts with cheeses that were produced using 2 different starter strains of Lactobacillus helveticus 138 or 209 (Lh138 or Lh209) at 3 ripening times: 3, 90, and 180 d. The experimental extracts were inoculated with Lb. plantarum I91; the control extracts were not inoculated and the blank extracts were heat-treated to inactivate enzymes and were not inoculated. All extracts were incubated at 34°C for 21 d, and then the pH, microbiological counts, and proteolysis profiles were determined. The basal proteolysis profiles in the extracts of young cheeses made with either strain tested were similar, but many differences between the proteolysis profiles of the extracts of the Lh138 and Lh209 cheeses were found when riper cheeses were used. The pH values in the blank and control extracts did not change, and no microbial growth was detected. In contrast, the pH value in experimental extracts decreased, and this decrease was more pronounced in extracts obtained from either of the young cheeses and from the Lh209 cheese at any stage of ripening. Lactobacillus plantarum I91 grew up to 8 log during the first days of incubation in all of the extracts, but then the number of viable cells decreased, the extent of which depended on the starter strain and the age of the cheese used for the extract. The decrease in the counts of Lb. plantarum I91 was observed mainly in the extracts in which the pH had diminished the most. In addition, the extracts that best supported the viability of Lb. plantarum I91 during incubation had the highest free amino acids content. The effect of Lb. plantarum I91 on the proteolysis profile of the extracts was marginal. Significant changes in the content of free amino acids suggested that the catabolism of free amino acids by Lb. plantarum I91 prevailed in a weakly proteolyzed medium, whereas the release of amino acids due to peptidolysis overcame their catabolism in a medium with high levels of free amino acids. Lactobacillus plantarum I91 was able to use energy sources other than lactose to support its growth because equivalent numbers of cells were observed in extracts containing residual amounts of lactose and in lactose-depleted extracts. The contribution of Lb. plantarum I91 to hard-cooked cheese peptidolysis was negligible compared with that of the starter strain; however, its ability to transform amino acids is a promising feature of this strain. Copyright © 2013 American Dairy Science Association. Published by Elsevier Inc. All rights reserved.

Effect of micro-environment modification and polymer type on the in-vitro dissolution behavior and in-vivo performance of amorphous solid dispersions.

PubMed

Sun, Weiwei; Pan, Baoliang

2017-06-15

This study investigates the effects of micro-environment modification and polymer type on the in-vitro dissolution behavior and in-vivo performance of micro-environment pH modifying solid dispersions (pH M -SD) for the poorly water-soluble model drug Toltrazuril (TOL). Various pH M -SDs were prepared using Ca(OH) 2 as a pH-modifier in hydrophilic polymers, including polyethylene glycol 6000 (PEG6000), polyvinylpyrrolidone k30 (PVPk30) and hydroxypropyl methylcellulose (HPMC). Based on the results of physicochemical characterizations and in-vitro dissolution testing, the representative ternary (Ca(OH) 2 :TOL:PEG6000/HPMC/PVPk30=1:8:24, w/w/w) and binary (TOL:PVPk30=1:3, w/w) solid dispersions were selected and optimized to perform in-vivo pharmacokinetic study. The micro-environment pH modification improved the in-vitro water-solubility and in-vivo bioavailability of parent drug TOL. Furthermore, the addition of alkalizers not only enhanced the release and absorption of prototype drug, but also promoted the generation of active metabolites, including toltrazuril sulfoxide (TOLSO) and toltrazuril sulfone (TOLSO 2 ). The in-vitro dissolution profiles and in-vivo absorption, distribution and metabolism behaviors of the pH M -SDs varied with polymer type. Moreover, in-vivo bioavailability of three active pharmaceutical ingredients increased with an increase in in-vitro dissolution rates of the drug from the pH M -SDs prepared with various polymers. Therefore, a non-sink in-vitro dissolution method can be used to predict the in-vivo performance of pH M -SDs formulated with various polymers with trend consistency. In-vitro and in-vivo screening procedures revealed that the pH M -SD composed of Ca(OH) 2 , TOL and PVPk30 at a weight ratio of 1:8:24, of which the safety was adequately proved via histopathological examination, may be a promising candidate for providing better clinical outcomes. Copyright © 2017. Published by Elsevier B.V.

"Artificial lymphatic system": a new approach to reduce interstitial hypertension and increase blood flow, pH and pO2 in solid tumors.

PubMed

DiResta, G R; Lee, J; Healey, J H; Levchenko, A; Larson, S M; Arbit, E

2000-05-01

A mechanical drainage system, the "artificial lymphatic system" (ALS), consisting of a vacuum source and drain, is evaluated for its ability to aspirate the interstitial fluids responsible for the elevated interstitial fluid pressure (IFP) observed in solid tumors. IFP, pH, and pO2 radial profiles were measured before and after aspiration using wick-in-needle (WIN) probes, needle pH and oxygen electrodes, respectively. Laser Doppler flowmetry measured temporal changes in blood flow rate (BFR) at the tumor surface during aspiration. The WIN probe and IFP profile data were analyzed using numerical simulation and distributed mathematical models, respectively. The model parameter, P(E), reflecting central tumor IFP, was reduced from 15.3 to 5.7 mm Hg in neuroblastoma and from 13.3 to 12.1 mm Hg in Walker 256, respectively, following aspiration. The simulation demonstrated that spatial averaging inherent in WIN measurements reduced the calculated magnitude of the model parameter changes. IFP was significantly lower (p<0.05), especially in regions surrounding the drain, and BFR was significantly higher (p<0.05) following 25 and 45 min of aspiration, respectively; pH and pO2 profiles increased following aspiration. The experimental and mathematical findings suggest that ALS aspiration may be a viable way of reducing IFP and increasing BFR, pO2, and pH and should enhance solid tumor chemo and radiation therapy.

pH and bacterial profile of dental plaque in children and adults of a low caries population.

PubMed

Raner, Elisabeth; Lindqvist, Lina; Johansson, Sofia; Hassan, Haidar; Carlén, Anette; Suksu-art, Narong; Dahlén, Gunnar

2014-06-01

This study compares pH and microbiological profile of dental plaque in children and adults of a low caries population. Thirty-nine children, 12-14 years of age and 45 adults between 20 and 39 years of age in 5 Karen villages of the Tak province, Northern Thailand were examined for plaque, calculus, caries (DMFT) and pH measurements in resting plaque and after a sucrose rinse. Information on dietary and oral hygiene habits was obtained through interviews using a fixed questionnaire. Microbiological profile of plaque samples was analyzed with DNA-DNA checkerboard technique. Mean DMFT was 0.77 ± 1.56 and 87% of the adults and 67% of the children were caries free (p < 0.05). The mean resting pH was for both age groups in the range of 7.0-7.1 and significantly higher than a Swedish caries free reference group. Karen adult men had significantly lower pH minimum than females and children (p < 0.05). Supragingival plaque samples showed high levels of low acidogenic and anaerobic species, which dominated over strong acid producers such as streptococci. The study indicates that the Karen children and adults has a plaque physiology and microbiology predominating by low acidogenic anaerobes, which in addition to the low sucrose intake explains the low caries prevalence in this population. Copyright © 2014 Elsevier Ltd. All rights reserved.

pH-Dependent silica nanoparticle dissolution and cargo release.

PubMed

Giovaninni, Giorgia; Moore, Colin J; Hall, Andrew J; Byrne, Hugh J; Gubala, Vladimir

2018-05-16

The dissolution of microporous silica nanoparticles (NP) in aqueous environments of different biologically relevant pH was studied in order to assess their potential as drug delivery vehicles. Silica NPs, loaded with fluorescein, were prepared using different organosilane precursors (tetraethoxysilane, ethyl triethoxysilane or a 1:1 molar ratio of both) and NP dissolution was evaluated in aqueous conditions at pH 4, pH 6 and pH 7.4. These conditions correspond to the acidity of the intracellular environment (late endosome, early endosome, cytosol respectively) and gastrointestinal tract ('fed' stomach, duodenum and jejunum respectively). All NPs degraded at pH 6 and pH 7.4, while no dissolution was observed at pH 4. NP dissolution could be clearly visualised as mesoporous hollows and surface defects using electron microscopy, and was supported by UV-vis, fluorimetry and DLS data. The dissolution profiles of the NPs are particularly suited to the requirements of oral drug delivery, whereby NPs must resist degradation in the harsh acidic conditions of the stomach (pH 4), but dissolve and release their cargo in the small intestine (pH 6-7.4). Particle cores made solely of ethyl triethoxysilane exhibited a 'burst release' of encapsulated fluorescein at pH 6 and pH 7.4, whereas NPs synthesised with tetraethoxysilane released fluorescein in a more sustained fashion. Thus, by varying the organosilane precursor used in NP formation, it is possible to modify particle dissolution rates and tune the release profile of encapsulated fluorescein. The flexible synthesis afforded by silica NPs to achieve pH-responsive dissolution therefore makes this class of nanomaterial an adaptable platform that may be well suited to oral delivery applications. Copyright © 2018 Elsevier B.V. All rights reserved.

Student understanding of pH: "i don't know what the log actually is, i only know where the button is on my calculator".

PubMed

Watters, Dianne J; Watters, James J

2006-07-01

In foundation biochemistry and biological chemistry courses, a major problem area that has been identified is students' lack of understanding of pH, acids, bases, and buffers and their inability to apply their knowledge in solving acid/base problems. The aim of this study was to explore students' conceptions of pH and their ability to solve problems associated with the behavior of biological acids to understand the source of student difficulties. The responses given by most students are characteristic of an atomistic approach in which they pay no attention to the structure of the problem and concentrate only on juggling the elements together until they get a solution. Many students reported difficulty in understanding what the question was asking and were unable to interpret a simple graph showing the pH activity profile of an enzyme. The most startling finding was the lack of basic understanding of logarithms and the inability of all except one student to perform a simple calculation on logs without a calculator. This deficiency in high school mathematical skills severely hampered their understanding of pH. This study has highlighted a widespread deficiency in basic mathematical skills among first year undergraduates and a fragmented understanding of acids and bases. Implications for the way in which the concepts of pH and buffers are taught are discussed. Copyright © 2006 International Union of Biochemistry and Molecular Biology, Inc.

Effect of ionization and vehicle on skin absorption and penetration of azelaic acid.

PubMed

Li, Nan; Wu, Xiaohong; Jia, Weibu; Zhang, Michelle C; Tan, Fengping; Zhang, Jerry

2012-08-01

The aim of this study is to investigate the effect of ionization and vehicle of topical formulations on skin absorption and penetration of azelaic acid (AZA). In vitro transport of AZA was determined for two topical formulations containing AZA with pH values of 3.9 and 4.9, respectively. FINACEA(®) (15% AZA gel), a US Food and Drug Administration approved drug for treatment of acne and rosacea, was also used for comparison. Release profile and flux of AZA were determined in an in vitro hairless mouse skin model using Franz Diffusion Cell. The data have shown that a higher concentration of AZA is retained in the epidermis/dermis layer and the whole skin for the formulation with pH = 4.9 as compared to that with pH = 3.9 at an active loading level of 2.82 mg/cm(2). In addition, the flux of ionized species of AZA in the pH 4.9 formulation (128.4 ± 35.9 μg/cm(2)/h) is approximately five-fold greater than that in the pH 3.9 formulation (27.7 ± 4.0 μg/cm(2)/h). The results suggest that the ionized AZA penetrates through the skin and accounts for majority of the total flux. This study has demonstrated that the penetration and absorption of AZA show a strong pH- and vehicle-dependency. Solubilization is the rate-limiting step in percutaneous absorption of AZA.

Effect of sodium tripolyphosphate concentration and simulated gastrointestinal fluids on release profile of paracetamol from chitosan microsphere

NASA Astrophysics Data System (ADS)

Mulia, Kamarza; Andrie; Krisanti, Elsa A.

2018-03-01

The problem to overcome in oral drug administration is the significant pH changes present in the human digestive system. In this study, ionotropic gelation method employing 2-8% (w/v) tripolyphosphate solutions were used to crosslink chitosan microspheres for a controlled release of paracetamol as a model drug. The release profiles of paracetamol from chitosan microspheres were determined using simulated gastrointestinal fluids having pH values of 1.2, 6.8, and 7.4. The results showed that the paracetamol loading and the encapsulation efficiency values increased with increasing concentration of tripolyphosphate solutions used in the preparation step. Paracetamol released at pH 1.2 and 6.8 buffer solutions was significantly higher than that at pH 7.4; also, more paracetamol was released in the presence of α-amylase and β-glucosidase enzymes. The release profiles showed zero-order release behaviour up to 8 hours where the highest drug release was 39% of the paracetamol loaded in the chitosan microspheres, indicating a strong crosslinking between chitosan and TPP anions. The relatively low accumulated drug release could be compensated by employing suitable enzymes, lower TPP solution concentration, and addition of other biodegradable polymer to reduce the TPP crosslink.

Electron and proton transfer in chloroplasts in silico. 2: The effect of diffusion limitations on the process of photosynthesis in spatially inhomogeneous thylakoids

NASA Astrophysics Data System (ADS)

Vershubskii, A. V.; Tikhonov, A. N.

2017-07-01

The lateral mobility of protons and mobile electron carriers (plastoquinone and plastocyanin) is subjected to diffusion limitations; the effect of these limitations on the kinetics of photoinduced pH i changes has been investigated in the present work for metabolic states 3 (conditions of intensive ATP synthesis) and 4 (the state of photosynthetic control). Computer simulations were based on a mathematical model of electron and proton transport in chloroplasts developed earlier by the authors. Non-uniform distribution of electron carriers and ATP synthase complexes in the membranes of grana and intergranal thylakoids was taken into account in the model. The kinetics of intrathylakoid pH i changes and the lateral profiles of distribution of the mobile electron transporters in granal and intergranal thylakoids were studied. The formation of non-uniform pH i profiles (with lumen acidification in the central parts of the grana being substantially slower than in the stromal thylakoids) was shown to occur under the conditions of ATP synthesis. Variation of the diffusion coefficients of intrathylakoid hydrogen ions and mobile electron carriers (plastoquinone and plastocyanin) can have substantial effects on the lateral pH i profiles and the redox state of the mobile electron carriers.

Preparation and Characterization of Liquisolid Compacts for Improved Dissolution of Telmisartan

PubMed Central

Narra, Nataraj; Rama Rao, Tadikonda

2014-01-01

The objective of the present work was to obtain pH independent and improved dissolution profile for a poorly soluble drug, telmisartan using liquisolid compacts. Liquisolid compacts were prepared using Transcutol HP as vehicle, Avicel PH102 as carrier, and Aerosil 200 as a coating material. The formulations were evaluated for drug excipient interactions, change in crystallinity of drug, flow properties, and general quality control tests of tablets using Fourier transform infrared (FTIR) spectroscopy, differential scanning calorimetry (DSC), X-ray diffraction (XRD), angle of repose, and various pharmacopoeial tests. In vitro dissolution studies were performed at three pH conditions (1.2, 4.5 and 7.4). Stability studies were performed at 40°C and 75% RH for three months. The formulation was found to comply with Indian pharmacopoeial limits for tablets. FTIR studies confirmed no interaction between drug and excipients. XRD and DSC studies indicate change/reduction in crystallinity of drug. Dissolution media were selected based on the solubility studies. The optimized formulation showed pH independent release profile with significant improvement (P < 0.005) in dissolution compared to plain drug and conventional marketed formulation. No significant difference was seen in the tablet properties, and drug release profile after storage for 3 months. PMID:25371826

[19F]fluorine nuclear-magnetic-resonance study of the interaction of difluoro-oxaloacetate with aspartate transaminase.

PubMed Central

Briley, P A; Eisenthal, R; Harrison, R; Smith, G D

1977-01-01

Difluoro-oxaloacetate interacts with the aldimine form of aspartate transaminase to give a complex, the dissociation constant of which has been determined spectrophotometrically and by 19F n.m.r. (nuclear magnetic resonance). The 19F n.m.r. line-width-pH and chemical-shift-pH profiles of difluoro-oxaloacetate in the presence of the aldimine form of the enzyme both show inflexion points in the pH5 and pH8 regions, which may arise from variations in the binding of difluoro-oxaloacetate as specific groups on the enzyme are successively protonated. Difluoro-oxaloacetate also interacts with apoenzyme to form a complex, the dissociation constant of which was determined by 19F n.m.r. The 19F n.m.r. line-width-pH and chemical-shift-pH profiles of difluoro-oxaloacetate in the presence of apoenzyme show a single inflexion point in the region of pH8. The absence, in this case, of an inflexion in the pH5 region indicates that the latter, present in the corresponding profiles for the aldimine form of the enzyme, results from ionization of an enzyme group associated with the pyridoxal phosphate cofactor. PMID:17399

Water-quality data for selected sites on Reversed, Rush, and Alger Creeks and Gull and Silver Lakes, Mono County, California, April 1994 to March 1995

USGS Publications Warehouse

Wang, Bronwen; Rockwell, G.L.; Blodgett, J.C.

1995-01-01

Water-quality data for selected sites on Reversed, Rush, and Alger Creeks and Gull and Silver Lakes, Mono County, California, were collected from April 1994 to March 1995. Water samples were analyzed for major ions and trace elements, nutrients, methylene blue active substances, and oil and grease. Field measurements were made for discharge, specific conductance, pH, water temperature, barometric pressure, dissolved oxygen, and alkalinity. Additional data collected include vertical water profiles of specific conductance, pH, water temperature, and dissolved oxygen collected at 3.3-foot intervals for Gull and Silver Lakes; chlorophyll-a and -b concentrations and Secchi depth for Gull and Silver Lakes; sediment interstitial- water nutrient concentrations in cores from Gull Lake; and lake surface and volume of Gull and Silver Lakes.

Probiotic Properties of Lactobacillus fermentum Strains Isolated from Human Oral Samples and Description of their Antibacterial Activity.

PubMed

Fuochi, Virginia; Volti, Giovanni Li; Furneri, Pio M

2017-01-01

Gram positive bacteria produce peptides, defined bacteriocins which exhibit good antibacterial activity. We evaluated the ability of L. fermentum to produce bacteriocins having therefore, good probiotic features and finally, be safe towards microglial cells. Eight wild strains, identified using molecular techniques, were investigated for the evaluation of resistance to bile salts, low pH, H2O2 production, biofilm formation, antibacterial activity and safety on microglia cells (BV2). The determination of the susceptibility/resistance profile showed that the strains are sensitive to the antibiotics tested. All strains showed a good tolerability to extremely low pH as well as resisting in presence of bile salts. In addition, the strains showed excellent activity against pathogens and one of them (LAC 42) showed activity also against Pseudomonas aeruginosa and Klebsiella pneumoniae. Finally, LAC 42 and its active compound did not change microglia cell viability following 24h exposure. Our data on this antibacterial molecule suggest that it is a compound with low molecular weight and with highly hydrophilic component. These results describe the characteristics of Lactobacillus strains and provide evidences for their possible use as new potential probiotic. In addition, other studies are now warranted to exploit the antibacterial activity of the supernatant LAC 42 and for its complete chemical characterization. Copyright© Bentham Science Publishers; For any queries, please email at epub@benthamscience.org.

An acid phosphatase in the plasma membranes of human astrocytoma showing marked specificity toward phosphotyrosine protein.

PubMed

Leis, J F; Kaplan, N O

1982-11-01

The plasma membrane from the human tumor astrocytoma contains an active acid phosphatase activity based on hydrolysis of p-nitrophenyl phosphate. Other acid phosphatase substrates--beta-glycerophosphate, O-phosphorylcholine, and 5'-AMP--are not hydrolyzed significantly. The phosphatase activity is tartrate insensitive and is stimulated by Triton X-100 and EDTA. Of the three known phosphoamino acids, only free O-phosphotyrosine is hydrolyzed by the membrane phosphatase activity. Other acid phosphatases tested from potato, wheat germ, milk, and bovine prostate did not show this degree of specificity. The plasma membrane activity also dephosphorylated phosphotyrosine histone at a much greater rate than did the other acid phosphatases. pH profiles for free O-phosphotyrosine and phosphotyrosine histone showed a shift toward physiological pH, indicating possible physiological significance. Phosphotyrosine histone dephosphorylation activity was nearly 10 times greater than that seen for phosphoserine histone dephosphorylation, and Km values were much lower for phosphotyrosine histone dephosphorylation (0.5 microM vs. 10 microM). Fluoride and zinc significantly inhibited phosphoserine histone dephosphorylation. Vanadate, on the other hand, was a potent inhibitor of phosphotyrosine histone dephosphorylation (50% inhibition at 0.5 microM) but not of phosphoserine histone. ATP stimulated phosphotyrosine histone dephosphorylation (160-250%) but inhibited phosphoserine histone dephosphorylation (95%). These results suggest the existence of a highly specific phosphotyrosine protein phosphatase activity associated with the plasma membrane of human astrocytoma.

Variations of soil profile characteristics due to varying time spans since ice retreat in the inner Nordfjord, western Norway

NASA Astrophysics Data System (ADS)

Navas, A.; Laute, K.; Beylich, A. A.; Gaspar, L.

2014-06-01

In the Erdalen and Bødalen drainage basins located in the inner Nordfjord in western Norway the soils were formed after deglaciation. The climate in the uppermost valley areas is sub-arctic oceanic, and the lithology consists of Precambrian granitic orthogneisses on which Leptosols and Regosols are the most common soils. The Little Ice Age glacier advance affected parts of the valleys with the maximum glacier extent around AD 1750. In this study five sites on moraine and colluvium materials were selected to examine main soil properties, grain size distribution, soil organic carbon and pH to assess if soil profile characteristics and patterns of fallout radionuclides (FRNs) and environmental radionuclides (ERNs) are affected by different stages of ice retreat. The Leptosols on the moraines are shallow, poorly developed and vegetated with moss and small birches. The two selected profiles show different radionuclide activities and grain size distribution. The sampled soils on the colluviums outside the LIA glacier limit became ice-free during the Preboral. The Regosols present better-developed profiles, thicker organic horizons and are fully covered by grasses. Activity of 137Cs and 210Pbex concentrate at the topsoil and decrease sharply with depth. The grain size distribution of these soils also reflects the difference in geomorphic processes that have affected the colluvium sites. Significantly lower mass activities of FRNs were found in soils on the moraines than on colluviums. Variations of ERN activities in the valleys were related to characteristics of soil mineralogical composition. These results indicate differences in soil development that are consistent with the age of ice retreat. In addition, the pattern distribution of 137Cs and 210Pbex activities differs in the soils related to the LIA glacier limits in the drainage basins.

Starch-entrapped microspheres show a beneficial fermentation profile and decrease in potentially harmful bacteria during in vitro fermentation in faecal microbiota obtained from patients with inflammatory bowel disease.

PubMed

Rose, Devin J; Venema, Koen; Keshavarzian, Ali; Hamaker, Bruce R

2010-05-01

The purpose of this research was to test the hypothesis that starch-entrapped microspheres would produce favourable fermentation profiles and microbial shifts during in vitro fermentation with the faecal microbiota from patients with inflammatory bowel disease (IBD). In vitro fermentation was carried out using a validated, dynamic, computer-controlled model of the human colon (Toegepast Natuurwetenschappelijk Onderzoek gastro-intestinal model-2) after inoculation with pooled faeces from healthy individuals, patients with inactive IBD (Crohn's disease (CD)) or patients with active IBD (ulcerative colitis (UC)). Starch-entrapped microspheres fermented more slowly and produced more butyrate than fructo-oligosaccharides (FOS) when fermented with the faecal microbiota from patients with active UC. When fermented with the microbiota from patients with inactive CD, starch-entrapped microspheres also fermented more slowly but produced similar amounts of butyrate compared with FOS. Starch-entrapped microspheres showed a greater ability to maintain a low pH during simulated-distal colon conditions compared with FOS. After fermentation with the microbiota from inactive CD patients, starch-entrapped microspheres resulted in lower concentrations of some potentially harmful gut bacteria, included in Bacteroides, Enterococcus, Fusobacterium and Veillonella, compared with FOS. These findings suggest that slow fermenting starch-entrapped microspheres may induce a favourable colonic environment in patients with IBD through high butyrate production, maintenance of low pH in the distal colon and inhibition of the growth of potentially harmful bacteria.

Growth and recombinant protein expression with Escherichia coli in different batch cultivation media.

PubMed

Hortsch, Ralf; Weuster-Botz, Dirk

2011-04-01

Parallel operated milliliter-scale stirred tank bioreactors were applied for recombinant protein expression studies in simple batch experiments without pH titration. An enzymatic glucose release system (EnBase), a complex medium, and the frequently used LB and TB media were compared with regard to growth of Escherichia coli and recombinant protein expression (alcohol dehydrogenase (ADH) from Lactobacillus brevis and formate dehydrogenase (FDH) from Candida boidinii). Dissolved oxygen and pH were recorded online, optical densities were measured at-line, and the activities of ADH and FDH were analyzed offline. Best growth was observed in a complex medium with maximum dry cell weight concentrations of 14 g L(-1). EnBase cultivations enabled final dry cell weight concentrations between 6 and 8 g L(-1). The pH remained nearly constant in EnBase cultivations due to the continuous glucose release, showing the usefulness of this glucose release system especially for pH-sensitive bioprocesses. Cell-specific enzyme activities varied considerably depending on the different media used. Maximum specific ADH activities were measured with the complex medium, 6 h after induction with IPTG, whereas the highest specific FDH activities were achieved with the EnBase medium at low glucose release profiles 24 h after induction. Hence, depending on the recombinant protein, different medium compositions, times for induction, and times for cell harvest have to be evaluated to achieve efficient expression of recombinant proteins in E. coli. A rapid experimental evaluation can easily be performed with parallel batch operated small-scale stirred tank bioreactors.

Saccharification efficiencies of multi-enzyme complexes produced by aerobic fungi.

PubMed

Badhan, Ajay; Huang, Jiangli; Wang, Yuxi; Abbott, D Wade; Di Falco, Marcos; Tsang, Adrian; McAllister, Tim

2018-05-24

In the present study, we have characterized high molecular weight multi-enzyme complexes in two commercial enzymes produced by Trichoderma reesei (Spezyme CP) and Penicillium funiculosum (Accellerase XC). We successfully identified 146-1000 kDa complexes using Blue native polyacrylamide gel electrophoresis (BN-PAGE) to fractionate the protein profile in both preparations. Identified complexes dissociated into lower molecular weight constituents when loaded on SDS PAGE. Unfolding of the secondary structure of multi-enzyme complexes with trimethylamine (pH >10) suggested that they were not a result of unspecific protein aggregation. Cellulase (CMCase) profiles of extracts of BN-PAGE fractionated protein bands confirmed cellulase activity within the multi-enzyme complexes. A microassay was used to identify protein bands that promoted high levels of glucose release from barley straw. Those with high saccharification yield were subjected to LC-MS analysis to identify the principal enzymatic activities responsible. The results suggest that secretion of proteins by aerobic fungi leads to the formation of high molecular weight multi-enzyme complexes that display activity against carboxymethyl cellulose and barley straw. Copyright © 2018. Published by Elsevier B.V.

Isolation and in vitro partial characterization of hemolytic proteins from the nematocyst venom of the jellyfish Stomolophus meleagris.

PubMed

Li, Rongfeng; Yu, Huahua; Xing, Ronge; Liu, Song; Qing, Yukun; Li, Kecheng; Li, Bing; Meng, Xiangtao; Cui, Jinhui; Li, Pengcheng

2013-09-01

Jellyfish venom contains various toxins and can cause itching, edema, muscle aches, shortness of breath, blood pressure depression, shock or even death after being stung. Hemolytic protein is one of the most hazardous components in the venom. The present study investigated the hemolytic activity of the nematocyst venom from jellyfish Stomolophus meleagris. Anion exchange chromatography, DEAE Sepharose Fast Flow, and gel filtration chromatography, Superdex200 had been employed to isolate hemolytic proteins from the nematocyst venom of jellyfish S. meleagris. Hemolysis of chicken red blood cells was used to quantify hemolytic potency of crude nematocyst venom and chromatography fractions during the purification process. Native-PAGE profile displayed one protein band in the purified hemolytic protein (SmTX); however, two protein bands with apparent molecular weights of ≈ 45 kDa and 52 kDa were observed in the reducing SDS-PAGE analysis. Approximately 70 μg/mL of SmTX caused 50% hemolysis (HU50) of the erythrocyte suspension. The hemolytic activity of SmTX was shown to be temperature and pH dependent, with the optimum temperature and pH being 37°C and pH 5.0. The present study is the first report of isolation and partial characterization of hemolytic proteins from the nematocyst venom of the jellyfish S. meleagris. The mechanism of the hemolytic activity of SmTX is not clear and deserves further investigation. Copyright © 2013. Published by Elsevier Ltd.

Comparison of cardiovascular risk factors in maintenance hemodialysis patients based on phase angle of bioimpedance analysis

NASA Astrophysics Data System (ADS)

Muzasti, R. A.; Lubis, H. R.

2018-03-01

Mortality and morbidity rate, especially from cardiovascular disease in hemodialysis patients in Indonesia is still quite high. One of indicator to assess the predictive value of mortality is the phase angle (PhA) of bioimpedance analysis (BIA) scan examination. Determining the comparison of BMI and laboratory data as cardiovascular risk factors in hemodialysis patients based on PhA.A cross-sectional analytical study was done on 155 outpatientsin RasyidaRenal Hospital, Medan in 2016. Patients were two groups, namely PhA<4 group and ≥ 4 group. The comparison of BMI and laboratory data based on PhA were by analyzingthe independent T-test. A P-value <0.05 was considered statistically significant. Most of thepatients are male (56.7%), obese (39.4%), with age 40-59 years (56.1%). Based on PhA, 56.7% patients have PhA ≥4. There are differences in the profile of age (p: 0.01), BMI (p: 0.028) and hemoglobin (p: 0.00) between two groups, but not in the profile of albumin (p: 0.071), total cholesterol (p: 0.65), HDL (p: 0.06), LDL (p: 0.07), triglyceride (p: 0.87), calcium (p: 0.59) and phosphorus (p: 0.17).Based onPhA, the cardiovascular risk factors of hemodialysis patients were determined by age, BMI, and hemoglobin.

A Profile of Chemistry Degree Holders in Academic Jobs

ERIC Educational Resources Information Center

Robinson, G. W.

1973-01-01

Presents statistical data on the distribution of institutions' Bachelor and Ph.D. chemistry graduates in faculty positions at research oriented schools with a comparison between the institutions' Ph.D. degree rankings and the rankings given by the American Council on Education included. (CC)

Glucose bioconversion profile in the syngas-metabolizing species Clostridium carboxidivorans.

PubMed

Fernández-Naveira, Ánxela; Veiga, María C; Kennes, Christian

2017-11-01

Some clostridia produce alcohols (ethanol, butanol, hexanol) from gases (CO, CO 2 , H 2 ) and others from carbohydrates (e.g., glucose). C. carboxidivorans can metabolize both gases as well as glucose. However, its bioconversion profile on glucose had not been reported. It was observed that C. carboxidivorans does not follow a typical solventogenic stage when grown on glucose. Indeed, at pH 6.2, it produced first a broad range of acids (acetic, butyric, hexanoic, formic, and lactic acids), several of which are generally not found, under similar conditions, during gas fermentation. Medium acidification did not allow the conversion of fatty acids into solvents. Production of some alcohols from glucose was observed in C. carboxidivorans but at high pH rather than under acidic conditions, and the total concentration of those solvents was low. At high pH, formic acid was produced first and later converted to acetic acid, but organic acids were not metabolized at low pH. Copyright © 2017 Elsevier Ltd. All rights reserved.

Numerical analysis for electrokinetic soil processing enhanced by chemical conditioning of the electrode reservoirs.

PubMed

Park, Jin-Soo; Kim, Soon-Oh; Kim, Kyoung-Woong; Kim, Byung Ro; Moon, Seung-Hyeon

2003-04-04

A numerical analysis was undertaken for enhanced electrokinetic soil processing. To perform chemical conditioning of the electrode reservoirs, the electrokinetic soil process employed a membrane as a barrier between the electrode reservoirs and the contaminated soil. An alkaline solution was purged in the anode reservoir that was bounded by the membrane. A mathematical model was used for demonstration of pH change and phenol removal from a kaolinite soil bed, the prediction of pH variations in both electrode reservoirs, and the determination of an optimized injection time of the anode-purging solution. The time-dependent dispersion coefficient was employed in consideration of the averaging effect of the velocity profile on a one-dimensional transport. The estimation of pH and phenol profiles in the soil bed reasonably agreed with the experimental data. The simulation revealed that the removal efficiency of phenol from the kaolinite soil could be improved by maintaining pH of the anode solution.

Esophageal motility and 24-h pH profiles of patients with heterotopic gastric mucosa in the cervical esophagus.

PubMed

Korkut, Esin; Bektaş, Mehmet; Alkan, Murat; Ustün, Yusuf; Meco, Cem; Ozden, Ali; Soykan, Irfan

2010-02-01

Heterotopic gastric mucosa occurs as a flat island of red mucosa in the proximal third of the esophagus where it gives rise to the cervical inlet patch. The aims of this study were to investigate the esophageal motility pattern and 24-h pH profiles of patients with cervical inlet patch. Thirty patients (16 women, mean age: 44.9 years, range: 23-72) diagnosed as having heterotopic gastric mucosa in the cervical esophagus with upper gastrointestinal symptoms had undergone esophageal motility testing and 24-h pH monitorisation with a double-channel pH probe. Manometric investigation was abnormal in 7 patients (non-specific esophageal motor disorder in 4 patients, esophageal hypomotility in 1 patient, and hypotensive LES in 2 patients). Pathological acid reflux (pH<4) was found in 9 (30%) of 30 heterotopic gastric mucosa patients during pH monitorisation from the distal probe. Pathological acid reflux in the proximal esophagus (percentage of total time of pH<4) was seen in four of these nine patients. Only four of the 30 patients (13.3%) presented with "acid independent episodes" during the 24-h esophageal pH monitorisation. Manometric investigation and 24-h pH monitorisation revealed that some of the patients with HGM have signs of esophageal motor dysfunction and "acid independent episodes" from the patches. These abnormalities may be responsible for some of the symptoms of HGM patients. Copyright 2009 European Federation of Internal Medicine. Published by Elsevier B.V. All rights reserved.

Mechanistic characterization of the HDV genomic ribozyme: solvent isotope effects and proton inventories in the absence of divalent metal ions support C75 as the general acid.

PubMed

Cerrone-Szakal, Andrea L; Siegfried, Nathan A; Bevilacqua, Philip C

2008-11-05

The hepatitis delta virus (HDV) ribozyme uses the nucleobase C75 and a hydrated Mg(2+) ion as the general acid-base catalysts in phosphodiester bond cleavage at physiological salt. A mechanistic framework has been advanced that involves one Mg(2+)-independent and two Mg(2+)-dependent channels. The rate-pH profile for wild-type (WT) ribozyme in the Mg(2+)-free channel is inverted relative to the fully Mg(2+)-dependent channel, with each having a near-neutral pKa. Inversion of the rate-pH profile was used as the crux of a mechanistic argument that C75 serves as general acid both in the presence and absence of Mg(2+). However, subsequent studies on a double mutant (DM) ribozyme suggested that the pKa observed for WT in the absence of Mg(2+) arises from ionization of C41, a structural nucleobase. To investigate this further, we acquired rate-pH/pD profiles and proton inventories for WT and DM in the absence of Mg(2+). Corrections were made for effects of ionic strength on hydrogen ion activity and pH meter readings. Results are accommodated by a model wherein the Mg(2+)-free pKa observed for WT arises from ionization of C75, and DM reactivity is compromised by protonation of C41. The Brønsted base appears to be water or hydroxide ion depending on pH. The observed pKa's are related to salt-dependent pH titrations of a model oligonucleotide, as well as electrostatic calculations, which support the local environment for C75 in the absence of Mg(2+) being similar to that in the presence of Mg(2+) and impervious to bulk ions. Accordingly, the catalytic role of C75 as the general acid does not appear to depend on divalent ions or the identity of the Brønsted base.

Vertical Distribution of PH(sub 3) in Saturn from Observations of the 1-0 and 3-2 Rotational Lines

NASA Technical Reports Server (NTRS)

Orton, G.; Serabyn, E.; Lee, Y.

1999-01-01

Far-infrared Fourier-transform spectrometer measurements of the 1-0 and 3-2 PH(sub 3) transitions in Saturn's disk near 267 and 800 GHz (8.9 and 26.7 cm(sup -1)), respectively, were analyzed simultaneously to derive a global mean profile for the PH(sub 3) vertical mixing ratio between 100 and 800 mbar total pressure.

A De Novo Floral Transcriptome Reveals Clues into Phalaenopsis Orchid Flower Development

PubMed Central

Huang, Jian-Zhi; Lin, Chih-Peng; Cheng, Ting-Chi; Chang, Bill Chia-Han; Cheng, Shu-Yu; Chen, Yi-Wen; Lee, Chen-Yu; Chin, Shih-Wen; Chen, Fure-Chyi

2015-01-01

Phalaenopsis has a zygomorphic floral structure, including three outer tepals, two lateral inner tepals and a highly modified inner median tepal called labellum or lip; however, the regulation of its organ development remains unelucidated. We generated RNA-seq reads with the Illumina platform for floral organs of the Phalaenopsis wild-type and peloric mutant with a lip-like petal. A total of 43,552 contigs were obtained after de novo assembly. We used differentially expressed gene profiling to compare the transcriptional changes in floral organs for both the wild-type and peloric mutant. Pair-wise comparison of sepals, petals and labellum between peloric mutant and its wild-type revealed 1,838, 758 and 1,147 contigs, respectively, with significant differential expression. PhAGL6a (CUFF.17763), PhAGL6b (CUFF.17763.1), PhMADS1 (CUFF.36625.1), PhMADS4 (CUFF.25909) and PhMADS5 (CUFF.39479.1) were significantly upregulated in the lip-like petal of the peloric mutant. We used real-time PCR analysis of lip-like petals, lip-like sepals and the big lip of peloric mutants to confirm the five genes’ expression patterns. PhAGL6a, PhAGL6b and PhMADS4 were strongly expressed in the labellum and significantly upregulated in lip-like petals and lip-like sepals of peloric-mutant flowers. In addition, PhAGL6b was significantly downregulated in the labellum of the big lip mutant, with no change in expression of PhAGL6a. We provide a comprehensive transcript profile and functional analysis of Phalaenopsis floral organs. PhAGL6a PhAGL6b, and PhMADS4 might play crucial roles in the development of the labellum in Phalaenopsis. Our study provides new insights into how the orchid labellum differs and why the petal or sepal converts to a labellum in Phalaenopsis floral mutants. PMID:25970572

Release of 5-Aminosalicylic Acid (5-ASA) from Mesalamine Formulations at Various pH Levels.

PubMed

Abinusawa, Adeyinka; Tenjarla, Srini

2015-05-01

Oral formulations of 5-aminosalicylic acid (5-ASA) for treatment of ulcerative colitis have been developed to minimize absorption prior to the drug reaching the colon. In this study, we investigate the release of 5-ASA from available oral mesalamine formulations in physiologically relevant pH conditions. Release of 5-ASA from 6 mesalamine formulations (APRISO®, Salix Pharmaceuticals, Inc., USA; ASACOL® MR, Procter & Gamble Pharmaceuticals UK Ltd.; ASACOL® HD, Procter & Gamble Pharmaceuticals, USA; MEZAVANT XL®, Shire US Inc.; PENTASA®, Ferring Pharmaceuticals, Ltd., UK; SALOFALK®, Dr. Falk Pharma UK Ltd.) was evaluated using United States Pharmacopeia apparatus I and II at pH values of 1.0 (2 h), 6.0 (1 h), and 6.8 (8 h). Dissolution profiles were determined for each formulation, respectively. Of the tested formulations, only the PENTASA formulation demonstrated release of 5-ASA at pH 1.0 (48%), with 56% cumulative release after exposure to pH 6.0 and 92% 5-ASA release after 6-8 h at pH 6.8. No other mesalamine formulation showed >1% drug release at pH 1.0. The APRISO formulation revealed 36% 5-ASA release at pH 6.0, with 100% release after 3 h at pH 6.8. The SALOFALK formulation revealed 11% 5-ASA release at pH 6.0, with 100% release after 1 h at pH 6.8. No 5-ASA was released by the ASACOL MR, ASACOL HD, and MEZAVANT XL formulations at pH 6.0. At pH 6.8, the ASACOL MR and ASACOL HD formulations exhibited complete release of 5-ASA after 4 and 2 h, respectively, and the MEZAVANT XL formulation demonstrated complete 5-ASA release over 6-7 h. 5-Aminosalicylic acid release profiles were variable among various commercially available formulations. Shire Development LLC.

Ion Electrodiffusion Governs Silk Electrogelation.

PubMed

Kojic, Nikola; Panzer, Matthew J; Leisk, Gary G; Raja, Waseem K; Kojic, Milos; Kaplan, David L

2012-07-14

Silk electrogelation involves the transition of an aqueous silk fibroin solution to a gel state (E-gel) in the presence of an electric current. The process is based on local pH changes as a result of water electrolysis - generating H(+) and OH(-) ions at the (+) and (-) electrodes, respectively. Silk fibroin has a pI=4.2 and when local pH

Pharmaceutical Evaluation of Cefuroxime Axetil Tablets Available in Drug Market of Pakistan

PubMed Central

Israr, F.; Mahmood, Z. A.; Hassan, F.; Hasan, S. M. F.

2016-01-01

Cefuroxime is a second generation cephalosporin antibiotic with a broad spectrum activity against Gram positive and Gram negative bacteria. The purpose of this research work was to evaluate the pharmaceutical quality standards of four different brands of cefuroxime axetil 125 mg tablets with different price ranges purchased from retail pharmacies of Pakistan. The brands were tested for physicochemical evaluation and in vitro dissolution studies in different medium like 0.07N HCl, distilled water, 0.1N HCl of pH 1.2 and phosphate buffers of pH 4.5 and pH 6.8. Statistical analysis, model dependent (zero order, first order, Korsmeyer-Peppas, Hixson-Crowell, Weibull) and model independent (Difference f1, similarity f2) approaches were applied to multiple dissolution profile of all brands. All brands were found to be similar with reference and meeting the compendial quality standard. Inter brand variation was observed in disintegration time and assay which was resulted in significant differences (P<0.05) in drug release data and Weibull was observed as best fill model. PMID:27168677

Measurements of Submicron Particle Adsorption and Particle Film Elasticity at Oil-Water Interfaces.

PubMed

Manga, Mohamed S; Hunter, Timothy N; Cayre, Olivier J; York, David W; Reichert, Matthew D; Anna, Shelly L; Walker, Lynn M; Williams, Richard A; Biggs, Simon R

2016-05-03

The influence of particle adsorption on liquid/liquid interfacial tension is not well understood, and much previous research has suggested conflicting behaviors. In this paper we investigate the surface activity and adsorption kinetics of charge stabilized and pH-responsive polymer stabilized colloids at oil/water interfaces using two tensiometry techniques: (i) pendant drop and (ii) microtensiometer. We found, using both techniques, that charge stabilized particles had little or no influence on the (dynamic) interfacial tension, although dense silica particles affected the "apparent" measured tension in the pendent drop, due to gravity driven elongation of the droplet profile. Nevertheless, this apparent change additionally allowed the study of adsorption kinetics, which was related qualitatively between particle systems by estimated diffusion coefficients. Significant and real interfacial tension responses were measured using ∼53 nm core-shell latex particles with a pH-responsive polymer stabilizer of poly(methyl methacrylate)-b-poly(2-(dimethylamino)ethyl methacrylate) (pMMA-b-pDMAEMA) diblock copolymer. At pH 2, where the polymer is strongly charged, behavior was similar to that of the bare charge-stabilized particles, showing little change in the interfacial tension. At pH 10, where the polymer is discharged and poorly soluble in water, a significant decrease in the measured interfacial tension commensurate with strong adsorption at the oil-water interface was seen, which was similar in magnitude to the surface activity of the free polymer. These results were both confirmed through droplet profile and microtensiometry experiments. Dilational elasticity measurements were also performed by oscillation of the droplet; again, changes in interfacial tension with droplet oscillation were only seen with the responsive particles at pH 10. Frequency sweeps were performed to ascertain the dilational elasticity modulus, with measured values being significantly higher than previously reported for nanoparticle and surfactant systems, and similar in magnitude to protein stabilized droplets.

Effect of pH, water activity and gel micro-structure, including oxygen profiles and rheological characterization, on the growth kinetics of Salmonella Typhimurium.

PubMed

Theys, T E; Geeraerd, A H; Verhulst, A; Poot, K; Van Bree, I; Devlieghere, F; Moldenaers, P; Wilson, D; Brocklehurst, T; Van Impe, J F

2008-11-30

In this study, the growth of Salmonella Typhimurium in Tryptic Soy Broth was examined at different pH (4.50-5.50), water activity a(w) (0.970-0.992) and gelatin concentration (0%, 1% and 5% ) at 20 degrees C. Experiments in TSB with 0% gelatin were carried out in shaken erlenmeyers, in the weak 1% gelatin media in petri plates and in the firm 5% gelatin media in gel cassettes. A quantification of gel strength was performed by rheological measurements and the influence of oxygen supply on the growth of S. Typhimurium was investigated. pH, as well as a(w) as well as gelatin concentration had an influence on the growth rate. Both in broth and in gelatinized media, lowering pH or water activity caused a decrease of growth rate. In media with 1% gelatin a reduction of growth rate and maximal cell density was observed compared to broth at all conditions. However, the effects of decreasing pH and a(w) were less pronounced. A further increase in gelatin concentration to 5% gelatin caused a small or no additional drop of growth rate. The final oxygen concentration dropped from 5.5 ppm in stirred broth to anoxic values in petri plates, also when 0% and 5% gelatin media were tested in this recipient. Probably, not stirring the medium, which leads to anoxic conditions, has a more pronounced effect on the growth rate of S. Typhimurium then medium solidness. Finally, growth data were fitted with the primary model of Baranyi and Roberts [Baranyi, J. and Roberts, T. A., 1994. A dynamic approach to predicting bacterial growth in food. International Journal of Food Microbiology 23, 277-294]. An additional factor was introduced into the secondary model of Ross et al. [Ross, T. and Ratkowsky, D. A. and Mellefont, L. A. and McMeekin, T. A., 2003. Modelling the effects of temperature, water activity, pH and lactic acid concentration on the growth rate of Escherichia coli. International Journal of Food Microbiology 82, 33-43.] to incorporate the effect of gelatin concentration, next to the effect of pH and a(w). A two step and a global regression procedure were applied. Both procedures were able to fit the data well, but the global regression procedure led to smaller standard errors on the parameters.

In view of an optimal gut antifungal therapeutic strategy: an in vitro susceptibility and toxicity study testing a novel phyto-compound.

PubMed

Metugriachuk, Yussef; Kuroi, Olivia; Pavasuthipaisit, Kanok; Tsuchiya, Junji; Minelli, Emilio; Okura, Ruichi; Fesce, Edoardo; Marotta, F

2005-01-01

In view of the raising concern for gut fungal infection, the aim of the present research was to carry out a systematic in vitro study testing the antifungal activity and possible toxicity of a polygodyal-anethole compound (Kolorex) in several strains of Candida albicans and in other fungal pathogens. The in vitro susceptibility tests were carried out on 4 strains of C. albicans (C. krusei, C. lipolytica, C. tropicalis, C. utilis), Aspergillus flavus and A. fumigatus. Cultures were also analyzed by varying medium, pH and inoculum size, and a time-course killing test was carried out. In the present study the polygodyal-anethole compound showed remarkable in vitro activity against the most common fungi, which was significantly better than polygodyal alone. Moreover, such mixture compound was shown to exert its activity against a wide spectrum of fungi, including C. lipolytica and C. tropicalis, which required significantly higher MIC of polygodyal to be unfeasible in clinical application. The activity of the polygodyal-anethole compound was significantly better than polygodyal alone with high inoculum size and low pH. Moreover, it proved to exert a significantly faster biological activity against low inoculum. This study suggests that the mixture compound Kolorex has a very good profile of antifungal activity in terms of effectiveness and spectrum of action while being devoid of any significant toxicity.

Sequential optimization of methotrexate encapsulation in micellar nano-networks of polyethyleneimine ionomer containing redox-sensitive cross-links.

PubMed

Abolmaali, Samira Sadat; Tamaddon, Ali; Yousefi, Gholamhossein; Javidnia, Katayoun; Dinarvand, Rasoul

2014-01-01

A functional polycation nanonetwork was developed for delivery of water soluble chemotherapeutic agents. The complexes of polyethyleneimine grafted methoxy polyethylene glycol (PEI-g-mPEG) and Zn(2+) were utilized as the micellar template for cross-linking with dithiodipropionic acid, followed by an acidic pH dialysis to remove the metal ion from the micellar template. The synthesis method was optimized according to pH, the molar ratio of Zn(2+), and the cross-link ratio. The atomic force microscopy showed soft, discrete, and uniform nano-networks. They were sensitive to the simulated reductive environment as determined by Ellman's assay. They showed few positive ζ potential and an average hydrodynamic diameter of 162±10 nm, which decreased to 49±11 nm upon dehydration. The ionic character of the nano-networks allowed the achievement of a higher-loading capacity of methotrexate (MTX), approximately 57% weight per weight, depending on the cross-link and the drug feed ratios. The nano-networks actively loaded with MTX presented some suitable properties, such as the hydrodynamic size of 117±16 nm, polydispersity index of 0.22, and a prolonged swelling-controlled release profile over 24 hours that boosted following reductive activation of the nanonetwork biodegradation. Unlike the PEI ionomer, the nano-networks provided an acceptable cytotoxicity profile. The drug-loaded nano-networks exhibited more specific cytotoxicity against human hepatocellular carcinoma cells if compared to free MTX at concentrations above 1 μM. The enhanced antitumor activity in vitro might be attributed to endocytic entry of MTX-loaded nano-networks that was found in the epifluorescence microscopy experiment for the fluorophore-labeled nano-networks.

Sequential optimization of methotrexate encapsulation in micellar nano-networks of polyethyleneimine ionomer containing redox-sensitive cross-links

PubMed Central

Abolmaali, Samira Sadat; Tamaddon, Ali; Yousefi, Gholamhossein; Javidnia, Katayoun; Dinarvand, Rasoul

2014-01-01

A functional polycation nanonetwork was developed for delivery of water soluble chemotherapeutic agents. The complexes of polyethyleneimine grafted methoxy polyethylene glycol (PEI-g-mPEG) and Zn2+ were utilized as the micellar template for cross-linking with dithiodipropionic acid, followed by an acidic pH dialysis to remove the metal ion from the micellar template. The synthesis method was optimized according to pH, the molar ratio of Zn2+, and the cross-link ratio. The atomic force microscopy showed soft, discrete, and uniform nano-networks. They were sensitive to the simulated reductive environment as determined by Ellman’s assay. They showed few positive ζ potential and an average hydrodynamic diameter of 162±10 nm, which decreased to 49±11 nm upon dehydration. The ionic character of the nano-networks allowed the achievement of a higher-loading capacity of methotrexate (MTX), approximately 57% weight per weight, depending on the cross-link and the drug feed ratios. The nano-networks actively loaded with MTX presented some suitable properties, such as the hydrodynamic size of 117±16 nm, polydispersity index of 0.22, and a prolonged swelling-controlled release profile over 24 hours that boosted following reductive activation of the nanonetwork biodegradation. Unlike the PEI ionomer, the nano-networks provided an acceptable cytotoxicity profile. The drug-loaded nano-networks exhibited more specific cytotoxicity against human hepatocellular carcinoma cells if compared to free MTX at concentrations above 1 μM. The enhanced antitumor activity in vitro might be attributed to endocytic entry of MTX-loaded nano-networks that was found in the epifluorescence microscopy experiment for the fluorophore-labeled nano-networks. PMID:24944513

CHARACTERIZATION AND PH/EH-BASED LEACHING TESTS OF MINING WASTES CONTAINING MERCURY

EPA Science Inventory

This study was undertaken as a part of developing treatment alternatives for waste materials, primarily waste rock and roaster tailings, from sites contaminated with mercury (Hg) mining wastes. Leaching profiles of waste rock over a range of different pH and oxidation-reduction (...

CHARACTERIZATION AND PH/EH-BASED LEACHING TESTS OF MINING WASTES CONTAINING MERCURY

EPA Science Inventory

This study was undertaken as a part of developing treatment alternatives for waste materials, primarily waste rock and roaster tailings, from sites contaminated with Mercury (Hg) mining wastes. Leaching profiles of waste rock over a range of different pH and oxidation-reduction ...

Kinetic and crystallographic studies of Escherichia coli UDP-N-acetylmuramate:L-alanine ligase.

PubMed Central

Emanuele, J. J.; Jin, H.; Jacobson, B. L.; Chang, C. Y.; Einspahr, H. M.; Villafranca, J. J.

1996-01-01

Uridine diphosphate-N-acetylmuramate:L-alanine ligase (EC 6.3.2.8, UNAM:L-Ala ligase or MurC gene product) catalyzes the ATP-dependent ligation of the first amino acid to the sugar moiety of the peptidoglycan precursor. This is an essential step in cell wall biosynthesis for both gram-positive and gram-negative bacteria. Optimal assay conditions for initial velocity studies have been established. Steady-state assays were carried out to determine the effect of various parameters on enzyme activity. Factors studies included: cation specificity, ionic strength, buffer composition and pH. At 37 degrees C and pH 8.0, kcat was equal to 980 +/- 40 min-1, while K(m) values for ATP, UNAM, and L-alanine were, 130 +/- 10, 44 +/- 3, and 48 +/- 6 microM, respectively. Of the metals tested only Mn, Mg, and Co were able to support activity. Sodium chloride, potassium chloride, ammonium chloride, and ammonium sulfate had no effect on activity up to 75 mM levels. The enzyme, in appropriate buffer, was stable enough to be assayed over the pH range of 5.6 to 10.1. pH profiles of Vmax/K(m) for the three substrates and of Vmax were obtained. Crystallization experiments with the enzyme produced two crystal forms. One of these has been characterized by X-ray diffraction as monoclinic, space group C2, with cell dimensions a = 189.6, b = 92.1, c = 75.2 A, beta = 105 degrees, and two 54 kDa molecules per asymmetric unit. It was discovered that the enzyme will hydrolyze ATP in the absence of L-alanine. This L-alanine independent activity is dependent upon the concentrations of both ATP and UNAM; kcat for this activity is less than 4% of the biosynthetic activity measured in the presence of saturating levels of L-alanine. Numerous L-alanine analogs tested were shown to stimulate ATP hydrolysis. A number of these L-alanine analogs produced novel products as accessed by HPLC and mass spectral analysis. All of the L-alanine analogs tested as inhibitors were competitive versus L-alanine. PMID:8976565

Mechanical Engineering | Classification | College of Engineering & Applied

Science.gov Websites

ProfessorMechanical Engineering(414) 229-6949avdeev@uwm.eduEng & Math Sciences 975 profile photo Robert Balmer, Sc . D.Professor EmeritusMechanical Engineering(414) 229-3374balmer@uwm.eduEng & Math Sciences E260 profile -6614wjchang@uwm.eduEng & Math Sciences 1113 profile photo Junhong Chen Ph.D.UWM Distinguished

Materials Science & Engineering | Classification | College of Engineering &

Science.gov Websites

ChairMaterials Science and Engineering(414) 229-2668nidal@uwm.eduEng & Math Sciences E351 profile photo (414) 229-2615jhchen@uwm.eduEng & Math Sciences 1225 profile photo Benjamin Church, Ph.D.Associate ProfessorMaterials Science & Engineering(414) 229-2825church@uwm.eduEng & Math Sciences EMS 1175 profile

Inhibitory effect of antioxidant-rich marinades on the formation of heterocyclic aromatic amines in pan-fried beef.

PubMed

Viegas, Olga; Amaro, L Filipe; Ferreira, Isabel M P L V O; Pinho, Olívia

2012-06-20

The inhibitory effect of antioxidant-rich marinades containing beer and white wine (with/without alcohol) alone or mixed with herbs commonly used as meat flavoring (garlic, ginger, thyme, rosemary, and red chili pepper) on the formation of heterocyclic aromatic amines (HAs) in pan-fried beef was studied. Radical-scavenging activity was evaluated by DPPH assay, before the addition of meat to the marinade (T0) and after 4 h of meat marinating (T4). At T0, wine with herbs possessed the highest scavenging activity (73.5%), followed by wine (72.5%), dealcoholized wine with herbs (53.4%), beer and herbs (41.7%), dealcoholized wine (39.6%), and beer (25.9%). At T4, a decrease in the radical-scavenging activity of all marinades was observed, although with a similar radical-scavenging profile. All of the six marinades under the study reduced the total amount of HAs, keeping meat with good overall sensory quality. Beer marinades were more efficient than white wine marinades, and the addition of herbs provided a superior inhibitory effect, reducing around 90% of HAs. No correlation was observed between radical-scavenging activity of marinades and total or individual HAs formation. Herbs explained around 30% of inhibition of PhIP formation, whereas alcohol increased PhIP formation.

4-Chloro-7-nitrobenzo-2-oxa-1,3-diazole as a reactivity probe for the investigation of the thiol proteinases. evidence that ficin and bromelain may lack carboxyl groups conformationally equivalent to that of aspartic acid-158 of papain.

PubMed Central

Shipton, M; Stuchbury, T; Brocklehurst, K

1976-01-01

1. 4-Chloro-7-nitrobenzo-2-oxa-1,3-diazole (Nbd chloride) was used as a reactivity probe to characterize the active centres of papin (EC 3.4.22.2), ficin (EC 3.4.22.3) and bromelain (EC 3.4.22.4). 2. In the pH range 0-8 Nbd chloride probably exists mainly as a monocation, possibly with the proton located on N-1 of the oxadiazole ring. 3. Spectroscopic evidence is presented for the intermediacy of Meisenheimer-type adducts in the reaction of Nbd chloride with nucleophiles. 4. The pH-dependence of the second-order rate constants (k) of the reactions of the three enzymes with Nbd chloride was determined at 25 degrees C, I = 0.1 mol/litre in 6.7% (v/v) ethanol in the pH range 2.5-5, where, at least for papain and ficin, the reactions occur specifically with their active-centre thiol groups. The pH-k profile for the papain reaction is bell-shaped (pKaI = 3.24, pKaII = 3.44 and k = 86M(-1)-s(-1), whereas that for ficin is sigmoidal (pKa = 3.6, k = 0.36M(-1)-s(-1), the rate increasing with increasing pH. The profile for the bromelain reaction appears to resemble that for the ficin reaction, but is complicated by amino-group labelling. 5. The bell-shaped profile of the papain reaction is considered to arise from the reaction of the thiolate ion of cysteine-25, maintained in acidic media by interaction with the side chain of histidine-159, with the Nbd chloride monocation hydrogen-bonded at its nitro group to the un-ionized form of the carboxyl group of aspartic acid-158. The lack of acid catalysis in the corresponding reactions of ficin and probably of bromelain suggests that these enzymes may lack carboxyl groups conformationally equivalent to that of aspartic acid-158 of papain. The possible consequences of this for the catalytic sites of these enzymes is discussed. PMID:11778

USGS field activities 11BHM03 and 11BHM04 on the west Florida shelf, Gulf of Mexico, September and November 2011

USGS Publications Warehouse

Robbins, Lisa L.; Knorr, Paul O.; Daly, Kendra L.; Barrera, Kira E.

2014-01-01

During September and November 2011 the (USGS), in cooperation with (USF), conducted geochemical surveys on the west Florida Shelf to investigate the effects of climate change on ocean acidification within the northern Gulf of Mexico, specifically, the effect of ocean acidification on marine organisms and habitats. The first cruise was conducted from September 20 to 28 (11BHM03) and the second was from November 2 to 4 (11BHM04). To view each cruise's survey lines, please see the Trackline page. Each cruise took place aboard the Research Vessel (R/V) Weatherbird II, a ship of opportunity led by Dr. Kendra Daly (USF), which departed from and returned to Saint Petersburg, Florida. Data collection included sampling of the surface and water column with lab analysis of pH, dissolved inorganic carbon (DIC) or total carbon dioxide (TCO2), and total alkalinity (TA). lLb analysis was augmented with a continuous flow-through system (referred to as sonde data) with a conductivity-temperature-depth (CTD) sensor, which also recorded salinity and pH. Corroborating the USGS data are the vertical CTD profiles (referred to as station samples) collected by USF. The CTD casts measured continuous vertical profiles of oxygen, chlorophyll fluorescence and optical backscatter. Discrete samples for nutrients, chlorophyll, and particulate organic carbon/nitrogen were also collected during the CTD casts. Two autonomous flow-through (AFT) instruments recorded pH and CO2 every 3-5 minutes on each cruise (referred to as AFT data).

Soybean hull induced production of carbohydrases and protease among Aspergillus and their effectiveness in soy flour carbohydrate and protein separation.

PubMed

Li, Qian; Loman, Abdullah Al; Coffman, Anthony M; Ju, Lu-Kwang

2017-04-20

Soybean hull consists mainly of three major plant carbohydrates, i.e., cellulose, hemicellulose and pectin. It is inexpensive and a good potential substrate for carbohydrase production because it is capable of inducing a complete spectrum of activities to hydrolyze complex biomass. Aspergillus is known for carbohydrase production but no studies have evaluated and compared, among Aspergillus species and strains, the soybean hull induced production of various carbohydrases. In this study, A. aculeatus, A. cinnamomeus, A. foetidus, A. phoenicis and 11 A. niger strains were examined together with T. reesei Rut C30, another known carbohydrase producer. The carbohydrases evaluated included pectinase, polygalacturonase, xylanase, cellulase, α-galactosidase and sucrase. Growth morphology and pH profiles were also followed. Among Aspergillus strains, morphology was found to correlate with both carbohydrase production and pH decrease profile. Filamentous strains gave higher carbohydrase production while causing slower pH decrease. The enzyme broths produced were also tested for separation of soy flour carbohydrate and protein. Defatted soy flour contains about 53% protein and 32% carbohydrate. The enzymatic treatment can increase protein content and remove indigestible oligo-/poly-saccharides, and improve use of soy flour in feed and food. Protease production by different strains was therefore also compared for minimizing protein degradation. A. niger NRRL 322 and A. foetidus NRRL 341 were found to be the most potent strains that produced maximal carbohydrases and minimal protease under soybean hull induction. Copyright © 2017 Elsevier B.V. All rights reserved.

USGS field activities 11BHM01 and 11BHM02 on the west Florida shelf, Gulf of Mexico, May and June 2011

USGS Publications Warehouse

Robbins, Lisa L.; Knorr, Paul O.; Daly, Kendra L.; Taylor, Carl A.; Barrera, Kira E.

2014-01-01

During May and June 2011 the (USGS), in cooperation with (USF), conducted geochemical surveys on the west Florida Shelf to investigate the effects of climate change on ocean acidification within the northern Gulf of Mexico, specifically, the effect of ocean acidification on marine organisms and habitats. The first cruise was conducted from May 3 to 9 (11BHM01) and the second was from June 25 to 30 (11BHM02). To view each cruise's survey lines, please see the Trackline page. Each cruise took place aboard the Research Vessel (R/V) Weatherbird II, a ship of opportunity led by Dr. Kendra Daly (USF), which departed from and returned to Saint Petersburg, Florida. Data collection included sampling of the surface and water column with lab analysis of pH, dissolved inorganic carbon (DIC) or total carbon dioxide (TCO2), and total alkalinity (TA). lLb analysis was augmented with a continuous flow-through system (referred to as sonde data) with a conductivity-temperature-depth (CTD) sensor, which also recorded salinity and pH. Corroborating the USGS data are the vertical CTD profiles (referred to as station samples) collected by USF. The CTD casts measured continuous vertical profiles of oxygen, chlorophyll fluorescence and optical backscatter. Discrete samples for nutrients, chlorophyll, and particulate organic carbon/nitrogen were also collected during the CTD casts. Two autonomous flow-through (AFT) instruments recorded pH and CO2 every 3-5 minutes on each cruise (referred to as AFT data).

Preformulation studies of EFdA, a novel nucleoside reverse transcriptase inhibitor for HIV prevention

PubMed Central

Zhang, Wei; Parniak, Michael A.; Mitsuya, Hiroaki; Sarafianos, Stefan G.; Graebing, Phillip W.; Rohan, Lisa C.

2014-01-01

4′-Ethynyl-2-fluoro-2′-deoxyadenosine (EFdA) is a novel nucleoside analog of great interest because of its superior activity against wild-type and multidrug-resistant HIV-1 strains, and favorable safety profiles in vitro and in vivo. The aim of this work was to provide preformulation information of EFdA important for delivery system development. A simple, accurate and specific reverse-phase high performance liquid chromatographic (RP-HPLC) method with UV detection was developed for quantification of EFdA. In addition, physicochemical characterizations including pH solubility profile, octanol/water partition coefficient (Log Po/w), DSC analysis, field emission scanning electron microscopy, and stability studies under various conditions were conducted. EFdA existed in planar or flake shape, with a melting point of ~130 °C, and had a pH dependent solubility. The log Po/w value of EFdA was −1.19. The compound was stable upon exposure to pH levels from 3 to 9 and showed good stability at elevated temperature (65 °C). In vitro cytotoxicity assessments were performed in two different epithelial cell lines. In cell-based studies, the EFdA selectivity index (50% cytotoxic concentration [CC50] values/50% effective concentration [EC50]) was found to be greater than 1 × 103. Permeability studies using cell- and tissue-based models showed that EFdA had an apparent permeability coefficient (Papp) <1 × 10−6cm/s and that the paracelluar pathway was the dominant transport route for EFdA. Overall, EFdA possesses favorable characteristics for further formulation development. PMID:23841536

Correlating the Impact of Well-Defined Oligosaccharide Structures on Physical Stability Profiles of IgG1-Fc Glycoforms.

PubMed

More, Apurva S; Toprani, Vishal M; Okbazghi, Solomon Z; Kim, Jae H; Joshi, Sangeeta B; Middaugh, C Russell; Tolbert, Thomas J; Volkin, David B

2016-02-01

As part of a series of articles in this special issue describing 4 well-defined IgG1-Fc glycoforms as a model system for biosimilarity analysis (high mannose-Fc, Man5-Fc, GlcNAc-Fc and N297Q-Fc aglycosylated), the focus of this work is comparisons of their physical properties. A trend of decreasing apparent solubility (thermodynamic activity) by polyethylene glycol precipitation (pH 4.5, 6.0) and lower conformational stability by differential scanning calorimetry (pH 4.5) was observed with reducing size of the N297-linked oligosaccharide structures. Using multiple high-throughput biophysical techniques, the physical stability of the Fc glycoproteins was then measured in 2 formulations (NaCl and sucrose) across a wide range of temperatures (10°C-90°C) and pH (4.0-7.5) conditions. The data sets were used to construct 3-index empirical phase diagrams and radar charts to visualize the regions of protein structural stability. Each glycoform showed improved stability in the sucrose (vs. salt) formulation. The HM-Fc and Man5-Fc displayed the highest relative stability, followed by GlcNAc-Fc, with N297Q-Fc being the least stable. Thus, the overall physical stability profiles of the 4 IgG1-Fc glycoforms also show a correlation with oligosaccharide structure. These data sets are used to develop a mathematical model for biosimilarity analysis (as described in a companion article by Kim et al. in this issue). Copyright © 2016 American Pharmacists Association®. Published by Elsevier Inc. All rights reserved.

Preformulation studies of EFdA, a novel nucleoside reverse transcriptase inhibitor for HIV prevention.

PubMed

Zhang, Wei; Parniak, Michael A; Mitsuya, Hiroaki; Sarafianos, Stefan G; Graebing, Phillip W; Rohan, Lisa C

2014-08-01

4'-Ethynyl-2-fluoro-2'-deoxyadenosine (EFdA) is a novel nucleoside analog of great interest because of its superior activity against wild-type and multidrug-resistant HIV-1 strains, and favorable safety profiles in vitro and in vivo. The aim of this work was to provide preformulation information of EFdA important for delivery system development. A simple, accurate and specific reverse-phase high performance liquid chromatographic (RP-HPLC) method with UV detection was developed for quantification of EFdA. In addition, physicochemical characterizations including pH solubility profile, octanol/water partition coefficient (Log Po/w), DSC analysis, field emission scanning electron microscopy, and stability studies under various conditions were conducted. EFdA existed in planar or flake shape, with a melting point of ∼130 °C, and had a pH dependent solubility. The log Po/w value of EFdA was -1.19. The compound was stable upon exposure to pH levels from 3 to 9 and showed good stability at elevated temperature (65 °C). In vitro cytotoxicity assessments were performed in two different epithelial cell lines. In cell-based studies, the EFdA selectivity index (50% cytotoxic concentration [CC50] values/50% effective concentration [EC50]) was found to be greater than 1 × 10(3). Permeability studies using cell- and tissue-based models showed that EFdA had an apparent permeability coefficient (Papp) <1 × 10(-6)cm/s and that the paracelluar pathway was the dominant transport route for EFdA. Overall, EFdA possesses favorable characteristics for further formulation development.

Geochemical Data for Upper Mineral Creek, Colorado, Under Existing Ambient Conditions and During an Experimental pH Modification, August 2005

USGS Publications Warehouse

Runkel, Robert L.; Kimball, Briant A.; Steiger, Judy I.; Walton-Day, Katherine

2009-01-01

Mineral Creek, an acid mine drainage stream in south-western Colorado, was the subject of a water-quality study that employed a paired synoptic approach. Under the paired synoptic approach, two synoptic sampling campaigns were conducted on the same study reach. The initial synoptic campaign, conducted August 22, 2005, documented stream-water quality under existing ambient conditions. A second synoptic campaign, conducted August 24, 2005, documented stream-water quality during a pH-modification experiment that elevated the pH of Mineral Creek. The experimental pH modification was designed to determine the potential reductions in dissolved constituent concentrations that would result from the implementation of an active treatment system for acid mine drainage. During both synoptic sampling campaigns, a solution containing lithium bromide was injected continuously to allow for the calculation of streamflow using the tracer-dilution method. Synoptic water-quality samples were collected from 30 stream sites and 11 inflow locations along the 2-kilometer study reach. Data from the study provide spatial profiles of pH, concentration, and streamflow under both existing and experimentally-altered conditions. This report presents the data obtained August 21-24, 2005, as well as the methods used for sample collection and data analysis.

Differential Effects of Mutations on the Transport Properties of the Na+/H+ Antiporter NhaA from Escherichia coli*

PubMed Central

Mager, Thomas; Braner, Markus; Kubsch, Bastian; Hatahet, Lina; Alkoby, Dudu; Rimon, Abraham; Padan, Etana; Fendler, Klaus

2013-01-01

Na+/H+ antiporters show a marked pH dependence, which is important for their physiological function in eukaryotic and prokaryotic cells. In NhaA, the Escherichia coli Na+/H+ antiporter, specific single site mutations modulating the pH profile of the transporter have been described in the past. To clarify the mechanism by which these mutations influence the pH dependence of NhaA, the substrate dependence of the kinetics of selected NhaA variants was electrophysiologically investigated and analyzed with a kinetic model. It is shown that the mutations affect NhaA activity in quite different ways by changing the properties of the binding site or the dynamics of the transporter. In the first case, pK and/or KDNa are altered, and in the second case, the rate constants of the conformational transition between the inside and the outside open conformation are modified. It is shown that residues as far apart as 15–20 Å from the binding site can have a significant impact on the dynamics of the conformational transitions or on the binding properties of NhaA. The implications of these results for the pH regulation mechanism of NhaA are discussed. PMID:23836890

Basalt weathering rates on Earth and the duration of liquid water on the plains of Gusev Crater, Mars

DOE Office of Scientific and Technical Information (OSTI.GOV)

Steefel, Carl; Hausrath, E.M.; Navarre-Sitchler, A.K.

2008-03-15

Where Martian rocks have been exposed to liquid water, chemistry versus depth profiles could elucidate both Martian climate history and potential for life. The persistence of primary minerals in weathered profiles constrains the exposure time to liquid water: on Earth, mineral persistence times range from {approx}10 ka (olivine) to {approx}250 ka (glass) to {approx}1Ma (pyroxene) to {approx}5Ma (plagioclase). Such persistence times suggest mineral persistence minima on Mars. However, Martian solutions may have been more acidic than on Earth. Relative mineral weathering rates observed for basalt in Svalbard (Norway) and Costa Rica demonstrate that laboratory pH trends can be used tomore » estimate exposure to liquid water both qualitatively (mineral absence or presence) and quantitatively (using reactive transport models). Qualitatively, if the Martian solution pH > {approx}2, glass should persist longer than olivine; therefore, persistence of glass may be a pH-indicator. With evidence for the pH of weathering, the reactive transport code CrunchFlow can quantitatively calculate the minimum duration of exposure to liquid water consistent with a chemical profile. For the profile measured on the surface of Humphrey in Gusev Crater, the minimum exposure time is 22 ka. If correct, this estimate is consistent with short-term, episodic alteration accompanied by ongoing surface erosion. More of these depth profiles should be measured to illuminate the weathering history of Mars.« less

Alginate nanoparticles protect ferrous from oxidation: Potential iron delivery system.

PubMed

Katuwavila, Nuwanthi P; Perera, A D L C; Dahanayake, Damayanthi; Karunaratne, V; Amaratunga, Gehan A J; Karunaratne, D Nedra

2016-11-20

A novel, efficient delivery system for iron (Fe 2+ ) was developed using the alginate biopolymer. Iron loaded alginate nanoparticles were synthesized by a controlled ionic gelation method and was characterized with respect to particle size, zeta potential, morphology and encapsulation efficiency. Successful loading was confirmed with Fourier Transform Infrared spectroscopy and Thermogravimetric Analysis. Electron energy loss spectroscopy study corroborated the loading of ferrous into the alginate nanoparticles. Iron encapsulation (70%) was optimized at 0.06% Fe (w/v) leading to the formation of iron loaded alginate nanoparticles with a size range of 15-30nm and with a negative zeta potential (-38mV). The in vitro release studies showed a prolonged release profile for 96h. Release of iron was around 65-70% at pH of 6 and 7.4 whereas it was less than 20% at pH 2.The initial burst release upto 8h followed zero order kinetics at all three pH values. All the release profiles beyond 8h best fitted the Korsmeyer-Peppas model of diffusion. Non Fickian diffusion was observed at pH 6 and 7.4 while at pH 2 Fickian diffusion was observed. Copyright © 2016 Elsevier B.V. All rights reserved.

Effective genetic modification and differentiation of hMSCs upon controlled release of rAAV vectors using alginate/poloxamer composite systems.

PubMed

Díaz-Rodríguez, P; Rey-Rico, A; Madry, H; Landin, M; Cucchiarini, M

2015-12-30

Viral vectors are common tools in gene therapy to deliver foreign therapeutic sequences in a specific target population via their natural cellular entry mechanisms. Incorporating such vectors in implantable systems may provide strong alternatives to conventional gene transfer procedures. The goal of the present study was to generate different hydrogel structures based on alginate (AlgPH155) and poloxamer PF127 as new systems to encapsulate and release recombinant adeno-associated viral (rAAV) vectors. Inclusion of rAAV in such polymeric capsules revealed an influence of the hydrogel composition and crosslinking temperature upon the vector release profiles, with alginate (AlgPH155) structures showing the fastest release profiles early on while over time vector release was more effective from AlgPH155+PF127 [H] capsules crosslinked at a high temperature (50°C). Systems prepared at room temperature (AlgPH155+PF127 [C]) allowed instead to achieve a more controlled release profile. When tested for their ability to target human mesenchymal stem cells, the different systems led to high transduction efficiencies over time and to gene expression levels in the range of those achieved upon direct vector application, especially when using AlgPH155+PF127 [H]. No detrimental effects were reported on either cell viability or on the potential for chondrogenic differentiation. Inclusion of PF127 in the capsules was also capable of delaying undesirable hypertrophic cell differentiation. These findings are of promising value for the further development of viral vector controlled release strategies. Copyright © 2015 Elsevier B.V. All rights reserved.

Mechanistic modeling of thermo-hydrological processes and microbial interactions at pore to profile scales resolve methane emission dynamics from permafrost soil

NASA Astrophysics Data System (ADS)

Ebrahimi, Ali; Or, Dani

2017-04-01

The sensitivity of the Earth's polar regions to raising global temperatures is reflected in rapidly changing hydrological processes with pronounced seasonal thawing of permafrost soil and increased biological activity. Of particular concern is the potential release of large amounts of soil carbon and the stimulation of other soil-borne GHG emissions such as methane. Soil methanotrophic and methanogenic microbial communities rapidly adjust their activity and spatial organization in response to permafrost thawing and a host of other environmental factors. Soil structural elements such as aggregates and layering and hydration status affect oxygen and nutrient diffusion processes thereby contributing to methanogenic activity within temporal anoxic niches (hotspots or hot-layers). We developed a mechanistic individual based model to quantify microbial activity dynamics within soil pore networks considering, hydration, temperature, transport processes and enzymatic activity associated with methane production in soil. The model was the upscaled from single aggregates (or hotspots) to quantifying emissions from soil profiles in which freezing/thawing processes provide macroscopic boundary conditions for microbial activity at different soil depths. The model distinguishes microbial activity in aerate bulk soil from aggregates (or submerged parts of the profile) for resolving methane production and oxidation rates. Methane transport pathways through soil by diffusion and ebullition of bubbles vary with hydration dynamics and affect emission patterns. The model links seasonal thermal and hydrologic dynamics with evolution of microbial community composition and function affecting net methane emissions in good agreement with experimental data. The mechanistic model enables systematic evaluation of key controlling factors in thawing permafrost and microbial response (e.g., nutrient availability, enzyme activity, PH) on long term methane emissions and carbon decomposition rates in the rapidly changing polar regions.

Anti-cancerous efficacy and pharmacokinetics of 6-mercaptopurine loaded chitosan nanoparticles.

PubMed

Kumar, G Prem; Sanganal, Jagadeesh S; Phani, A R; Manohara, C; Tripathi, Syamantak M; Raghavendra, H L; Janardhana, P B; Amaresha, S; Swamy, K B; Prasad, R G S V

2015-10-01

6-Mercaptopurine is a cytotoxic and immunosuppressant drug. The use of this drug is limited due to its poor bioavailability and short plasma half-life. In order to nullify these drawbacks, 6-mercaptopurine-chitosan nanoparticles (6-MP-CNPs) were prepared and evaluated to study the influence of preparation conditions on the physicochemical properties by using DLS, SEM, XRD and FTIR. The in vitro drug release profile at pH 4.8 and 7.4 revealed sustained release patterns for a period of 2 days. The nanoformulations showed enhanced in vitro anti-cancer activities (MTT assay, apoptosis assay, cell cycle arrest and ROS indices) on HT-1080 and MCF-7 cells. In vivo pharmacokinetics profiles of 6-MP-CNPs showed improved bioavailability. Thus, the results of the present study revealed that, the prepared 6-MP-CNPs have a significant role in increasing anti-cancer efficacy, bioavailability and in vivo pharmacokinetics profiles. Copyright © 2015 Elsevier Ltd. All rights reserved.

Effect of alginate composition on profile release and characteristics of chitosan-alginate microparticles loaded with mangosteen extract

NASA Astrophysics Data System (ADS)

Mulia, Kamarza; Halimah, Nur; Krisanti, Elsa

2017-03-01

Preparation of mangostin-loaded chitosan-alginate microparticles, chemical and physical characterization of the particles, and mangostin release profiles, are described herein. Mangostin rich fraction was obtained from Garcinia mangostana L. pericarp by extraction followed by fractionation. Mangostin-loaded chitosan-alginate microparticles were prepared by ionic gelation method using tripolyphosphate as the linking agent and various concentration of alginate. Mangostin was effectively loaded in all microparticle formulations, resulting in ˜97% encapsulation efficiencies. The loading of mangostin and the in-vitro release profiles in simulated gastrointestinal fluids were affected by the chitosan to alginate ratios used in the preparation of the microparticles. Increased alginate concentration resulted in lowered release of mangostin from microparticles immersed in simulated gastric fluid (pH 1.2) up to two hours. Low release of mangostin in acidic fluid but high release in simulated colon fluid, indicated that the chitosan-alginate microparticles are prospective carrier for extended release of active compound in gastrointestinal system.

Production, purification and characterization of an exo-polygalacturonase from Penicillium janthinellum sw09.

PubMed

Ma, Yuping; Sun, Siwen; Hao, Hui; Xu, Chunping

2016-01-01

A soil isolate, Penicillium janthinellum sw09 has been found to produce significant amounts of an extracellular pectinase subsequently characterized as exo-polygalacturonase (exo-PG). By optimizing growth conditions, P. janthinellum sw09 produced high amount of exo-PG (16.54 units/mL). The crude enzyme was purified by gel filtration chromatography and two exo-PG activity peaks (designated as PGI and PGII) were revealed. On SDS-PAGE analysis, purified PGII using DEAE-Sepharose FF column, was found to be a single band with a molecular mass of 66.2 kDa. The purified PGII exhibited maximal activity at the temperature of 45 oC and pH 5.0. The stability profiles show that PGII is more stable in the pH range of 4.0-8.0 and below 60 oC. The Km and Vmax for the enzyme was 1.74 mg/mL and 18.08 μmol/ (mL•min), respectively. Due to this enzymatic characterization, this pectinase is an attractive candidate for applications in degradation of pectin.

Endophytic Streptomyces sp. AC35, a producer of bioactive isoflavone aglycones and antimycins.

PubMed

Ondrejíčková, P; Šturdíková, M; Hushegyi, A; Švajdlenka, E; Markošová, K; Čertík, M

2016-09-01

In this research, a microbial endophytic strain obtained from the rhizosphere of the conifer Taxus baccata and designated as Streptomyces sp. AC35 (FJ001754.1 Streptomyces, GenBank) was investigated. High 16S rDNA gene sequence similarity suggests that this strain is closely related to S. odorifer. The major fatty acid profile of intracellular lipids was also carried out to further identify this strain. Atomic force microscopy and scanning acoustic microscopy were used to image our strain. Its major excreted substances were extracted, evaluated for antimicrobial activity, purified, and identified by ultraviolet-visible spectroscopy (UV-vis), liquid chromatography-mass spectrometry (LC-MS/MS) and nuclear magnetic resonance as the bioactive isoflavone aglycones-daidzein, glycitein and genistein. Batch cultivation, performed under different pH conditions, revealed enhanced production of antimycin components when the pH was stable at 7.0. Antimycins were detected by HPLC and identified by UV-vis and LC-MS/MS combined with the multiple reaction monitoring. Our results demonstrate that Streptomyces sp. AC35 might be used as a potential source of effective, pharmaceutically active compounds.

Pharmacokinetic profiles contribute to the differences in behavioral pharmacology of 071031B enantiomers as novel serotonin and norepinephrine reuptake inhibitors.

PubMed

Xue, Rui; Li, Ying; He, Xin-Hua; Jin, Zeng-Liang; Fan, Shi-Yong; Zhang, Ting-Ting; Li, Nuo-Min; Yuan, Li; Zheng, Ai-Ping; Zhong, Bo-Hua; Li, Yun-Feng; Zhang, You-Zhi

2017-03-01

Our previous study indicated that a chiral compound 071031B was a novel serotonin and noradrenaline reuptake inhibitor with superior antidepressant activity compared to duloxetine. The present study aimed to investigate chiral pharmacology differences of 071031B enantiomers, S-071031B and R-071031B, and disclose mechanisms underlying the behavioral differences based on target profiles and pharmacokinetic profiles. In vivo behavioral tests indicated that S-071031B was more potent than R-071031B in two depression models (the forced swimming test in mice and rats) and two pain models (the acetic acid-induced writhing and formalin tests in mice). In vitro assays revealed that both S-071031B and R-071031B showed high affinity for human serotonin transporters and norepinephrine transporters with equal potency, and showed consistently equipotent inhibitory effects on serotonin and norepinephrine uptake. Pharmacokinetic studies demonstrated that oral availability and hepatic metabolism, rather than pH stability, intestinal transport, and plasma binding, contributed to enantiomers' behavioral differences. Based on these findings, it is suggested that S-071031B is a more active enantiomer, and the differential pharmacokinetic profiles, but not target affinity, contribute to differences of S-071031B and R-071031B in behavioral pharmacology. Moreover, current PK-PD study may provide positive exploration for chiral antidepressants development.

HPLC determination of olanzapine and carbamazepine in their nicotinamide cocrystals and investigation of the dissolution profiles of cocrystal tablet formulations.

PubMed

Renkoğlu, Pelin; Çelebier, Mustafa; Arıca-Yegin, Betül

2015-05-01

Cocrystals have recently gained importance in the pharmaceutical industry. In this study, olanzapine and carbamazepine cocrystals were synthesized by using nicotinamide as cocrystal forming agent to achieve improvements in the physicochemical characteristics of the active ingredients. An HPLC method was developed to determine the amount, thus, the stoichiometric ratios of olanzapine and carbamazepine in the synthesized cocrystals. Olanzapine:nicotinamide and olanzapine tablet formulations were prepared and the developed HPLC method was applied successfully in order to compare the dissolution profiles of these formulations. An ACE 5 CN, 25 cm × 4.6 mm, 5 µm column was used and a gradient elution program was performed for simultaneous determination of olanzapine, carbamazepine and nicotinamide. Phosphate buffer (pH 5.0, 25 mM) and methanol was used in a ratio from 80:20 to 70:30 while the flow rate was 1 mL min(-1) for the elution of the compounds within 12 min. In conclusion, two different aims were achieved, the first one was to indicate the stoichiometric ratios of the active ingredients olanzapine and carbamazepine with nicotinamide in their cocrystals, and the second one was the comparison of the dissolution profiles of the olanzapine and olanzapine:nicotinamide cocrystal formulations. It was found that the cocrystal formulation with nicotinamide improved the dissolution profile of olanzapine.

To Be, or Not to Be, Trained

ERIC Educational Resources Information Center

Bryant, Dennis; Richardson, Alice

2015-01-01

This paper compares the learning output from study units taught by PhD qualified lecturers against study units taught by PhD qualified lecturers who additionally have acquired formal postgraduate certification in higher education teaching. The findings reveal that the two teaching profiles exhibit different patterns of learning outputs.…

Nanoscale Investigation of the Impact of pH and Orthophosphate on the Corrosion of Copper Surfaces in Water

EPA Science Inventory

Advanced surface characterization techniques were used to systematically investigate the passivation of copper during corrosion in water as impacted by pH and orthophosphate. Atomic force microscopy, depth profiling with time-of-flight secondary ion mass spectrometry and X-ray d...

[Dependence of the pharmacokinetics of captopril on the type of adaptation reactions in the organism].

PubMed

Udut, V V; Khazanov, V A; Gurto, R V; Borodulina, E V; Postnikova, Iu E

2007-01-01

The dependence of the pharmacokinetic profiles (PhP) of captopril in the phase of adaptation reactions in the organism has been studied within the framework of randomized, comparative, double cross research of bioeqivalency of captopril (Aspharma Co, Anzhero-Sudzhensk) and capoten (Bristol Myers Squibb Co.; official Russian producer, Akrikhin KhimFarmKombinat). It is established that the maximum bioaccessibility and high concentration of captopril in the blood plasma is determined on the background of physiologically optimum reactions of training and in the zone of quiet activation. These characteristics decrease during the reactions of general adaptation syndrome according to the type of increased activation and reactivation.

Hyperforin: To Be or Not to Be an Activator of TRPC(6).

PubMed

Friedland, Kristina; Harteneck, Christian

2015-01-01

Meantime, it is well accepted that hyperforin, the chemical instable phloroglucinol derivative of Hypericum perforatum, St. John's wort, is the pharmacophore of St. John's wort extracts. With the decline of this scientific discussion, another controversial aspect has been arisen, the question regarding the underlying mechanism leading to the pharmacological profile of the plant extract used in therapy of depression. We will summarize the different concepts described for hyperforin's antidepressive activity. Starting with unspecific protein-independent mechanisms due to changes in pH, we will summarize data of protein-based concepts beginning with concepts based on involvement of a variety of proteins and will finally present concepts based on the modulation of a single protein.

Phrenic motoneuron discharge patterns following chronic cervical spinal cord injury

PubMed Central

Lee, Kun-Ze; Dougherty, Brendan J.; Sandhu, Milapjit S.; Lane, Michael A.; Reier, Paul J.; Fuller, David D.

2013-01-01

Cervical spinal cord injury (SCI) dramatically disrupts synaptic inputs and triggers biochemical, as well as morphological, plasticity in relation to the phrenic motor neuron (PhMN) pool. Accordingly, our primary purpose was to determine if chronic SCI induces fundamental changes in the recruitment profile and discharge patterns of PhMNs. Individual PhMN action potentials were recorded from the phrenic nerve ipsilateral to lateral cervical (C2) hemisection injury (C2Hx) in anesthetized adult male rats at 2, 4 or 8 wks post-injury and in uninjured controls. PhMNs were phenotypically classified as early (Early-I) or late inspiratory (Late-I), or silent according to discharge patterns. Following C2Hx, the distribution of PhMNs was dominated by Late-I and silent cells. Late-I burst parameters (e.g., spikes per breath, burst frequency and duration) were initially reduced but returned towards control values by 8 wks post-injury. In addition, a unique PhMN burst pattern emerged after C2Hx in which Early-I cells burst tonically during hypocapnic inspiratory apnea. We also quantified the impact of gradual reductions in end-tidal CO2 partial pressure (PETCO2) on bilateral phrenic nerve activity. Compared to control rats, as PETCO2 declined, the C2Hx animals had greater inspiratory frequencies (breaths*min−1) and more substantial decreases in ipsilateral phrenic burst amplitude. We conclude that the primary physiological impact of C2Hx on ipsilateral PhMN burst patterns is a persistent delay in burst onset, transient reductions in burst frequency, and the emergence of tonic burst patterns. The inspiratory frequency data suggest that plasticity in brainstem networks is likely to play an important role in phrenic motor output after cervical SCI. PMID:23954215

²H kinetic isotope effects and pH dependence of catalysis as mechanistic probes of rat monoamine oxidase A: comparisons with the human enzyme.

PubMed

Wang, Jin; Edmondson, Dale E

2011-09-06

Monoamine oxidase A (MAO A) is a mitochondrial outer membrane-bound flavoenzyme important in the regulation of serotonin and dopamine levels. Because the rat is extensively used as an animal model in drug studies, it is important to understand how rat MAO A behaves in comparison with the more extensively studied human enzyme. For many reversible inhibitors, rat MAO A exhibits K(i) values similar to those of human MAO A. The pH profile of k(cat) for rat MAO A shows a pK(a) of 8.2 ± 0.1 for the benzylamine ES complex and pK(a) values of 7.5 ± 0.1 and 7.6 ± 0.1 for the ES complexes with p-CF(3)-(1)H- and p-CF(3)-(2)H-benzylamine, respectively. In contrast to the human enzyme, the rat enzyme exhibits a single pK(a) value (8.3 ± 0.1) with k(cat)/K(m) for benzylamine versus pH and pK(a) values of 7.8 ± 0.1 and 8.1 ± 0.2 for the ascending limbs, respectively, of k(cat)/K(m) versus pH profiles for p-CF(3)-(1)H- and p-CF(3)-(2)H-benzylamine and 9.3 ± 0.1 and 9.1 ± 0.2 for the descending limbs, respectively. The oxidation of para-substituted benzylamine substrate analogues by rat MAO A has large deuterium kinetic isotope effects on k(cat) and on k(cat)/K(m). These effects are pH-independent and range from 7 to 14, demonstrating a rate-limiting α-C-H bond cleavage step in catalysis. Quantitative structure-activity correlations of log k(cat) with the electronic substituent parameter (σ) at pH 7.5 and 9.0 show a dominant contribution with positive ρ values (1.2-1.3) and a pH-independent negative contribution from the steric term. Quantitative structure-activity relationship analysis of the binding affinities of the para-substituted benzylamine analogues for rat MAO A shows an increased van der Waals volume (V(w)) increases the affinity of the deprotonated amine for the enzyme. These results demonstrate that rat MAO A exhibits functional properties similar but not identical with those of the human enzyme and provide additional support for C-H bond cleavage via a polar nucleophilic mechanism.

Production, optimization and characterization of fibrinolytic enzyme by Bacillus subtilis RJAS19.

PubMed

Kumar, D J Mukesh; Rakshitha, R; Vidhya, M Annu; Jennifer, P Sharon; Prasad, Sandip; Kumar, M Ravi; Kalaichelvan, P T

2014-04-01

The present study aimed at the production, purification and characterization of fibrinolytic nattokinase enzyme from the bacteria isolated from natto food. For the purpose, a fibrinolytic bacterium was isolated and identified as Bacillus subtilis based on 16S rDNA sequence analysis. The strain was employed for the production and optimization of fibrinolytic enzyme. The strain showed better enzyme production during 72nd h of incubation time with 50 degrees C at the pH 9. The lactose and peptone were found to be increasing the enzyme production rate. The enzyme produced was purified and also characterized with the help of SDS-PAGE analysis. The activity and stability profile of the purified enzyme was tested against different temperature and pH. The observations suggesting that the potential of fibrinolytic enzyme produced by Bacillus subtilis RJAS 19 for its applications in preventive medicines.

Studies on H-Translocating ATPases in Plants of Varying Resistance to Salinity : I. Salinity during Growth Modulates the Proton Pump in the Halophyte Atriplex nummularia.

PubMed

Braun, Y; Hassidim, M; Lerner, H R; Reinhold, L

1986-08-01

Membrane vesicles were isolated from the roots of the halophyte Atriplex nummularia Lindl. H(+)-translocating Mg(2+)-ATPase activity was manifested by the establishment of a positive membrane potential (measured as SCN(-) accumulation); and also by the establishment of a transmembrane pH gradient (measured by quinacrine fluorescence quenching). H(+)-translocation was highly specific to ATP and was stable to oligomycin. Growing the plants in the presence of 400 millimolar NaCl doubled the proton-translocating activity per milligram of membrane protein and otherwise modulated it in the following ways. First, the flat pH profile observed in non-salt-grown plants was transformed to one showing a peak at about pH 6.2. Second, the lag effect observed at low ATP concentration in curves relating SCN(-) accumulation to ATP concentration was abolished; the concave curvature shown in the double reciprocal plot was diminished. Third, sensitivity to K-2 (N-morpholino)ethanesulfonic acid stimulation was shown in salt-grown plants (about 40% stimulation) but was absent in non-salt-grown plants. Fourth, the KCl concentration bringing about 50% dissipation of ATP-dependent SCN(-) accumulation was 20 millimolar for salt-grown plants and 50 millimolar for non-salt-grown plants. Vanadate sensitivity was shown in both cases. No clear NO(3) (-) inhibition was observed.

Studies on H+-Translocating ATPases in Plants of Varying Resistance to Salinity 1

PubMed Central

Braun, Yael; Hassidim, Miriam; Lerner, Henri R.; Reinhold, Leonora

1986-01-01

Membrane vesicles were isolated from the roots of the halophyte Atriplex nummularia Lindl. H+-translocating Mg2+-ATPase activity was manifested by the establishment of a positive membrane potential (measured as SCN− accumulation); and also by the establishment of a transmembrane pH gradient (measured by quinacrine fluorescence quenching). H+-translocation was highly specific to ATP and was stable to oligomycin. Growing the plants in the presence of 400 millimolar NaCl doubled the proton-translocating activity per milligram of membrane protein and otherwise modulated it in the following ways. First, the flat pH profile observed in non-salt-grown plants was transformed to one showing a peak at about pH 6.2. Second, the lag effect observed at low ATP concentration in curves relating SCN− accumulation to ATP concentration was abolished; the concave curvature shown in the double reciprocal plot was diminished. Third, sensitivity to K-2 (N-morpholino)ethanesulfonic acid stimulation was shown in salt-grown plants (about 40% stimulation) but was absent in non-salt-grown plants. Fourth, the KCl concentration bringing about 50% dissipation of ATP-dependent SCN− accumulation was 20 millimolar for salt-grown plants and 50 millimolar for non-salt-grown plants. Vanadate sensitivity was shown in both cases. No clear NO3− inhibition was observed. Images Fig. 3 PMID:16664942

Differential Effects of Anaesthesia on the phMRI Response to Acute Ketamine Challenge

PubMed Central

Hodkinson, Duncan J.; de Groote, Carmen; McKie, Shane; Deakin, J. F. William; Williams, Steve R.

2012-01-01

Aims Pharmacological-challenge magnetic resonance imaging (phMRI) is powerful new tool enabling researchers to map the central effects of neuroactive drugs in vivo. To employ this technique pre-clinically, head movements and the stress of restraint are usually reduced by maintaining animals under general anaesthesia. However, interactions between the drug of interest and the anaesthetic employed may potentially confound data interpretation. NMDA receptor (NMDAR) antagonists used widely to mimic schizophrenia have recently been shown to interact with the anaesthetic halothane. It may be the case that neural and cerebrovascular responses to NMDAR antagonists are dependent on the types of anaesthetic used. Methodology We compared the phMRI response to NMDAR antagonist ketamine in rats maintained under α-chloralose to those under isoflurane anaesthesia. A randomized placebo/vehicle controlled design was used in each of the anaesthetic groups. Results Changes in the anaesthetic agent resulted in two very different profiles of activity. In the case of α-chloralose, positive activations in cortical and sub-cortical structures reflected a response which was similar to patterns seen in healthy human volunteers and metabolic maps of conscious rats. However, the use of isoflurane completely reversed such effects, causing widespread deactivations in the cortex and hippocampus. Conclusion This study provides initial evidence for a drug-anesthetic interaction between ketamine and isoflurane that is very different from responses to α-chloralose-ketamine. PMID:22737655

Controlled release of acidic drugs in compendial and physiological hydrogen carbonate buffer from polymer blend-coated oral solid dosage forms.

PubMed

Wulff, R; Rappen, G-M; Koziolek, M; Garbacz, G; Leopold, C S

2015-09-18

The objective of this study was to investigate the suitability of "Eudragit® RL/Eudragit® L55" (RL/L55) blend coatings for a pH-independent release of acidic drugs. A coating for ketoprofen and naproxen mini tablets was developed showing constant drug release rate under pharmacopeial two-stage test conditions for at least 300 min. To simulate drug release from the mini tablets coated with RL/L55 blends in the gastrointestinal (GI) tract, drug release profiles in Hanks buffer pH 6.8 were recorded and compared with drug release profiles in compendial media. RL/L55 blend coatings showed increased drug permeability in Hanks buffer pH 6.8 compared to phosphate buffer pH 6.8 due to its higher ion concentration. However, drug release rates of acidic drugs were lower in Hanks buffer pH 6.8 because of the lower buffer capacity resulting in reduced drug solubility. Further dissolution tests were performed in Hanks buffer using pH sequences simulating the physiological pH conditions in the GI tract. Drug release from mini tablets coated with an RL/L55 blend (8:1) was insensitive to pH changes of the medium within the pH range of 5.8-7.5. It was concluded that coatings of RL/L55 blends show a high potential for application in coated oral drug delivery systems with a special focus on pH-independent release of acidic drugs. Copyright © 2015 Elsevier B.V. All rights reserved.

Changes of the soil environment affected by fly ash dumping site of the electric power plant

NASA Astrophysics Data System (ADS)

Weber, Jerzy; Gwizdz, Marta; Jamroz, Elzbieta; Debicka, Magdalena; Kocowicz, Andrzej

2014-05-01

In this study the effect of fly ash dumping site of the electric power plant on the surrounding soil environment was investigated. The fly ash dumping site collect wastes form brown coal combustion of Belchatow electric power station, central Poland. The dumping site is surrounding by forest, where pine trees overgrow Podzols derived from loose quartz sands. The soil profiles under study were located at a distance of 50, 100, 400 and 500 m from the dumping site, while control profiles were located 8 km away from the landfill. In all horizons of soil profiles the mpain hysico-chemical and chemical properties were determined. The humic substances were extracted from ectohumus horizons by Shnitzer's method, purified using XAD resin and freeze-dried. The fulvic acids were passed through a cation exchange column and freeze-dried. Optical density, elemental composition and atomic ratios were determined in the humic and fulvic acids. Organic carbon by KMnO4 oxidation was also determined in the organic soil horizons. The fly ash from the landfill characterized by high salinity and strong alkaline reaction (pH=10), which contributed significantly to the changes of the pH values in soils horizons. The alkalization of soils adjacent to the landfill was found, which manifested in increasing of pH values in the upper soil horizons. The impact of the landfill was also noted in the changes of the soil morphology of Podzols analysed. As a result of the alkalization, Bhs horizons have been converted into a Bs horizons. Leaching of low molecular humus fraction - typical for podzolization - has been minimized as a result of pH changes caused by the impact of the landfill, and originally occurring humic substances in the Bhs horizon (present in the control profiles) have been probably transported out of the soil profile and then into the groundwater.

Monitoring of internal pH gradients within multi-layer tablets by optical methods and EPR imaging.

PubMed

Eisenächer, Friederike; Schädlich, Andreas; Mäder, Karsten

2011-09-30

The high variability of gastrointestinal pH is a general challenge regarding constant release from oral drug delivery systems, especially for ionisable drugs. These drugs often show a pH-dependent solubility and therewith associated intra- and inter-individual variability of emerging drug plasma levels. Several strategies have been investigated with the intention to influence the microenvironmental pH (pH(M)) within solid formulations and therefore achieve pH-independent release profiles. Because of the heterogeneity of solid systems, a precise prediction of the occurring pH(M) is rather difficult. It is therefore important to monitor the pH(M) within the formulations to achieve requested release as well as to minimise pH-dependent degradation processes of the active compound. The purpose of the current study was the analysis of pH(M) gradients within 2- and 3-layer tablets during hydration using 3 different techniques for comparison intensions, in particular a pH indicator dye, fluorescence imaging and EPR imaging. The influence of the presence or absence of pH modifying substances and of an additional lipophilic inter layer on the pH(M) was investigated as well as the variation of matrix forming excipient and buffer pH. The influence of the pH(M) on drug release was analysed as well. In addition, benchtop MRI was accomplished to gain a deeper insight on the hydration and erosion behaviour of 2- and 3-layer tablets. Copyright © 2010 Elsevier B.V. All rights reserved.

Intracellular pH Recovery Rates of Hemocytes from Estuarine and Open Ocean Bivalve Species Following In vitro Acid Challenge

NASA Astrophysics Data System (ADS)

Croxton, A.; Wikfors, G.

2013-12-01

Decreasing pH in estuarine systems is a growing concern for researchers studying mollusk species. Debates continue on whether estuarine bivalve species are more or less vulnerable to ocean acidification than marine species because estuaries can present multiple environmental stressors. The aim of this study is to understand the homeostatic mechanisms of bivalve hemocytes following exposure to extracellular acid treatment. Previous measurements using fluorescent SNARF probes and flow-cytometry have determined the intracellular pH of hemocytes from several bivalve species (eastern oyster, bay scallop, northern quahog, soft-shell clam, and blue mussel) to range between 7.0-7.4. In the present study of four bivalve species, recovery rate profiles were determined for intracellular hemocyte pH following addition of acid to hemolymph in vitro. These profiles indicate that soft-shell clams and bay scallops maintained homeostasis with very little change in intracellular pH. In contrast, an initial drop in intracellular pH in northern quahogs was followed by a steady recovery of intracellular pH. Contrasting results between species appear to be unrelated to mineral shell composition (aragonite vs. calcite) or habitat location (infaunal vs. epifaunal). The next phase of this study will be to determine if offshore species (surfclams and sea scallops) will have similar responses. Results from these studies will provide a better understanding of the physiological responses of estuarine and marine species exposed to acidified environments.

The distribution and origins of extremely acidic saline groundwaters in the south of Western Australia - Groundwater and digital mapping datasets provide new insights

NASA Astrophysics Data System (ADS)

Lillicrap, Adam M.; Biermann, Vera; George, Richard J.; Gray, David J.; Oldham, Carolyn E.

2018-01-01

Some of the largest extents of naturally occurring acidic waters are found across southern Australia. The origins of these systems remain poorly understood with many hypotheses for their genesis. Australian government agency groundwater datasets and mapping data (vegetation, geology, regolith and soils) for south-western Australia, unavailable to previous researchers, were statistically analysed to better understand the origins of acidic groundwater and guide additional fieldwork to study the origins of acidic saline groundwater. The groundwater data showed a distinct bimodal distribution in pH; the 'acid' population had a median pH of 3.5 and the larger 'non-acid' population had a median pH of 6.6. Acidic groundwater became progressively more common further from the coast towards the drier internally drained regions. Acidic groundwater was mostly confined to the lower slopes and valley floors with localised controls on distribution. Paradoxically, subsoil alkalinity within the internally drained inland regions had the strongest correlation with acidic groundwater (r2 = 0.85). Vegetation was also a strong predictor of acidic groundwater. Acidic groundwater had the highest occurrence under Eucalyptus woodlands and shrublands that grew on alkaline calcareous soils. Pre-clearing soil data in areas with acidic saline groundwater showed that the upper 1 m of the unsaturated zone had a pH around 8 while the pH at depths greater than 5 m decreased to <4. Based on the observations it is proposed that biogenic formation of calcareous soils occurs in the upper 1 m of the profile, calcium is sourced from the deeper profile where the root biota exchanges calcium for hydrogen ions to maintain charge balance. Iron is mobilised from the upper soil profile and concentrates lower in the profile at depths >1.5 m. There, the iron is reduced around roots and the alkalinity generated by microbial iron reduction is removed by biogenic calcification processes. The iron moves in solution further down the profile following roots where it comes in contact with the oxygenated unsaturated zone matrix and is oxidised generating acid. The resulting acidic recharging solution acidifies the unsaturated zone matrix. Saline groundwater moving through the matrix becomes acidified due to ion exchange or direct recharge. The main chemical processes were modelled in PHREEQC to test the plausibility of the hypothesis and acidic solutions with a pH of 3.8 or lower were obtained.

A two-state computational investigation of methane C--H and ethane C--C oxidative addition to [CpM(PH3)]n+ (M = Co, Rh, Ir; n = 0, 1).

PubMed

Petit, Alban; Richard, Philippe; Cacelli, Ivo; Poli, Rinaldo

2006-01-11

Reductive elimination of methane from methyl hydride half-sandwich phosphane complexes of the Group 9 metals has been investigated by DFT calculations on the model system [CpM(PH(3))(CH(3))(H)] (M = Co, Rh, Ir). For each metal, the unsaturated product has a triplet ground state; thus, spin crossover occurs during the reaction. All relevant stationary points on the two potential energy surfaces (PES) and the minimum energy crossing point (MECP) were optimized. Spin crossover occurs very near the sigma-CH(4) complex local minimum for the Co system, whereas the heavier Rh and Ir systems remain in the singlet state until the CH(4) molecule is almost completely expelled from the metal coordination sphere. No local sigma-CH(4) minimum was found for the Ir system. The energetic profiles agree with the nonexistence of the Co(III) methyl hydride complex and with the greater thermal stability of the Ir complex relative to the Rh complex. Reductive elimination of methane from the related oxidized complexes [CpM(PH(3))(CH(3))(H)](+) (M = Rh, Ir) proceeds entirely on the spin doublet PES, because the 15-electron [CpM(PH(3))](+) products have a doublet ground state. This process is thermodynamically favored by about 25 kcal mol(-1) relative to the corresponding neutral system. It is essentially barrierless for the Rh system and has a relatively small barrier (ca. 7.5 kcal mol(-1)) for the Ir system. In both cases, the reaction involves a sigma-CH(4) intermediate. Reductive elimination of ethane from [CpM(PH(3))(CH(3))(2)](+) (M = Rh, Ir) shows a similar thermodynamic profile, but is kinetically quite different from methane elimination from [CpM(PH(3))(CH(3))(H)](+): the reductive elimination barrier is much greater and does not involve a sigma-complex intermediate. The large difference in the calculated activation barriers (ca. 12.0 and ca. 30.5 kcal mol(-1) for the Rh and Ir systems, respectively) agrees with the experimental observation, for related systems, of oxidatively induced ethane elimination when M = Rh, whereas the related Ir systems prefer to decompose by alternative pathways.

Charge heterogeneity of rat pituitary prolactin in relation to the estrous cycle, gonadectomy, and estrogen treatment.

PubMed

Ishikawa, J; Wakabayashi, K; Igarashi, M

1985-10-01

Pituitary samples were obtained from female rats at various stages of the estrous cycle, and from intact male and gonadectomized rats with and without estradiol treatment. The pituitary extracts with 60% EtOH pH 9.5, were fractionated by preparative isoelectric focusing (IEF), and immunoreactive prolactin (IR-PRL) was measured by RIA. Three types of IR-PRL molecular species were found in these IEF profiles. The first type (species A) was consistently found in an area of pH 4.5-5.4, and consisted of two main subspecies with pls 5.0 (Al) and 5.25 (A2). Species A occupied most part of pituitary IR-PRL in males, gonadectomized animals, and in females in a basal state such as diestrus (D) II 17:00. Species A was also found exclusively in the serum at proestrus (PE) 19:00. The amounts of species A decreased notably when the secretion became active from PE 15:00 to 22:00, then increased at estrus (E) 6:00 and 10:00 when the second type (species B), which was found in the area of pH 5.4-6.8 only in trace amounts at basal states, increased markedly. Species B decreased again at E 17:00, while species A fully recovered. Species B also increased when PRL biosynthesis was stimulated by estradiol in intact male and gonadectomized rats. These findings indicate that species A must be the storage and secretory type of IR-PR, and that species B must be IR-PRL in the biosynthetic process which is to be finally converted into species A. A third type (species C) was found in a region of pH 3.5-4.5 in the IEF profiles of gonadectomized animals. This species is possibly IR-PRL molecules under degradation. When the pituitary was extracted serially with 0.25 M ammonium sulfate pH 5.5 (fraction AMS) first, then with 60% EtOH pH 9.5 (fraction ET), fraction AMS contained mostly species B and C, while fraction ET contained species A almost exclusively. The results obtained with this differential extraction roughly coincided with IEF data, though some disagreements were observed.

Phosphatase synthesis in Klebsiella (Aerobacter) aerogenes growing in continuous culture

PubMed Central

Bolton, P. G.; Dean, A. C. R.

1972-01-01

1. Phosphatase synthesis was studied in Klebsiella aerogenes grown in a wide range of continuous-culture systems. 2. Maximum acid phosphatase synthesis was associated with nutrient-limited, particularly carbohydrate-limited, growth at a relatively low rate, glucose-limited cells exhibiting the highest activity. Compared with glucose as the carbon-limiting growth material, other sugars not only altered the activity but also changed the pH–activity profile of the enzyme(s). 3. The affinity of the acid phosphatase in glucose-limited cells towards p-nitrophenyl phosphate (Km 0.25–0.43mm) was similar to that of staphylococcal acid phosphatase but was ten times greater than that of the Escherichia coli enzyme. 4. PO43−-limitation derepressed alkaline phosphatase synthesis but the amounts of activity were largely independent of the carbon source used for growth. 5. The enzymes were further differentiated by the effect of adding inhibitors (F−, PO43−) and sugars to the reaction mixture during the assays. In particular, it was shown that adding glucose, but not other sugars, stimulated the rate of hydrolysis of p-nitrophenyl phosphate by the acid phosphatase in carbohydrate-limited cells at low pH values (<4.6) but inhibited it at high pH values (>4.6). Alkaline phosphatase activity was unaffected. 6. The function of phosphatases in general is discussed and possible mechanisms for the glucose effect are outlined. PMID:4342213

Proteolysin, a Novel Highly Thermostable and Cosolvent-Compatible Protease from the Thermophilic Bacterium Coprothermobacter proteolyticus

PubMed Central

Toplak, Ana; Wu, Bian; Fusetti, Fabrizia; Quaedflieg, Peter J. L. M.

2013-01-01

Through genome mining, we identified a gene encoding a putative serine protease of the thermitase subgroup of subtilases (EC 3.4.21.66) in the thermophilic bacterium Coprothermobacter proteolyticus. The gene was functionally expressed in Escherichia coli, and the enzyme, which we called proteolysin, was purified to near homogeneity from crude cell lysate by a single heat treatment step. Proteolysin has a broad pH tolerance and is active at temperatures of up to 80°C. In addition, the enzyme shows good activity and stability in the presence of organic solvents, detergents, and dithiothreitol, and it remains active in 6 M guanidinium hydrochloride. Based on its stability and activity profile, proteolysin can be an excellent candidate for applications where resistance to harsh process conditions is required. PMID:23851086

pH regulates genes for flagellar motility, catabolism, and oxidative stress in Escherichia coli K-12.

PubMed

Maurer, Lisa M; Yohannes, Elizabeth; Bondurant, Sandra S; Radmacher, Michael; Slonczewski, Joan L

2005-01-01

Gene expression profiles of Escherichia coli K-12 W3110 were compared as a function of steady-state external pH. Cultures were grown to an optical density at 600 nm of 0.3 in potassium-modified Luria-Bertani medium buffered at pH 5.0, 7.0, and 8.7. For each of the three pH conditions, cDNA from RNA of five independent cultures was hybridized to Affymetrix E. coli arrays. Analysis of variance with an alpha level of 0.001 resulted in 98% power to detect genes showing a twofold difference in expression. Normalized expression indices were calculated for each gene and intergenic region (IG). Differential expression among the three pH classes was observed for 763 genes and 353 IGs. Hierarchical clustering yielded six well-defined clusters of pH profiles, designated Acid High (highest expression at pH 5.0), Acid Low (lowest expression at pH 5.0), Base High (highest at pH 8.7), Base Low (lowest at pH 8.7), Neutral High (highest at pH 7.0, lower in acid or base), and Neutral Low (lowest at pH 7.0, higher at both pH extremes). Flagellar and chemotaxis genes were repressed at pH 8.7 (Base Low cluster), where the cell's transmembrane proton potential is diminished by the maintenance of an inverted pH gradient. High pH also repressed the proton pumps cytochrome o (cyo) and NADH dehydrogenases I and II. By contrast, the proton-importing ATP synthase F1Fo and the microaerophilic cytochrome d (cyd), which minimizes proton export, were induced at pH 8.7. These observations are consistent with a model in which high pH represses synthesis of flagella, which expend proton motive force, while stepping up electron transport and ATPase components that keep protons inside the cell. Acid-induced genes, on the other hand, were coinduced by conditions associated with increased metabolic rate, such as oxidative stress. All six pH-dependent clusters included envelope and periplasmic proteins, which directly experience external pH. Overall, this study showed that (i) low pH accelerates acid consumption and proton export, while coinducing oxidative stress and heat shock regulons; (ii) high pH accelerates proton import, while repressing the energy-expensive flagellar and chemotaxis regulons; and (iii) pH differentially regulates a large number of periplasmic and envelope proteins.

Mechanism of Flavoprotein l-6-Hydroxynicotine Oxidase: pH and Solvent Isotope Effects and Identification of Key Active Site Residues.

PubMed

Fitzpatrick, Paul F; Chadegani, Fatemeh; Zhang, Shengnan; Dougherty, Vi

2017-02-14

The flavoenzyme l-6-hydroxynicotine oxidase is a member of the monoamine oxidase family that catalyzes the oxidation of (S)-6-hydroxynicotine to 6-hydroxypseudooxynicotine during microbial catabolism of nicotine. While the enzyme has long been understood to catalyze oxidation of the carbon-carbon bond, it has recently been shown to catalyze oxidation of a carbon-nitrogen bond [Fitzpatrick, P. F., et al. (2016) Biochemistry 55, 697-703]. The effects of pH and mutagenesis of active site residues have now been utilized to study the mechanism and roles of active site residues. Asn166 and Tyr311 bind the substrate, while Lys287 forms a water-mediated hydrogen bond with flavin N5. The N166A and Y311F mutations result in ∼30- and ∼4-fold decreases in k cat /K m and k red for (S)-6-hydroxynicotine, respectively, with larger effects on the k cat /K m value for (S)-6-hydroxynornicotine. The K287M mutation results in ∼10-fold decreases in these parameters and a 6000-fold decrease in the k cat /K m value for oxygen. The shapes of the pH profiles are not altered by the N166A and Y311F mutations. There is no solvent isotope effect on the k cat /K m value for amines. The results are consistent with a model in which both the charged and neutral forms of the amine can bind, with the former rapidly losing a proton to a hydrogen bond network of water and amino acids in the active site prior to the transfer of hydride to the flavin.

Phosphorylation of the amino terminus of maize sucrose synthase in relation to membrane association and enzyme activity.

PubMed

Hardin, Shane C; Winter, Heike; Huber, Steven C

2004-04-01

Sucrose synthase (SUS) is phosphorylated on a major, amino-terminal site located at Ser-15 (S15) in the maize (Zea mays) SUS1 protein. Site- and phospho-specific antibodies against a phosphorylated S15 (pS15) peptide allowed direct analysis of S15 phosphorylation in relation to membrane association. Immunoblots of the maize leaf elongation zone, divided into 4-cm segments, demonstrated that the abundance of soluble (s-SUS) and membrane (m-SUS) SUS protein showed distinct positional profiles. The content of m-SUS was maximal in the 4- to 8-cm segment where it represented 9% of total SUS and occurred as a peripheral membrane protein. In contrast, s-SUS was highest in the 12- to 16-cm segment. Relative to s-SUS, m-SUS was hypophosphorylated at S15 in the basal 4 cm but hyperphosphorylated in apical segments. Differing capabilities of the anti-pS15 and anti-S15 peptide antibodies to immunoprecipitate SUS suggested that phosphorylation of S15, or exposure of unphosphorylated SUS to slightly acidic pH, altered the structure of the amino terminus. These structural changes were generally coincident with the increased sucrose cleavage activity that occurs at pH values below 7.5. In vitro S15 phosphorylation of the S170A SUS protein by a maize calcium-dependent protein kinase (CDPK) significantly increased sucrose cleavage activity at low pH. Collectively, the results suggest that (1) SUS membrane binding is controlled in vivo; (2) relative pS15 content of m-SUS depends on the developmental state of the organ; and (3) phosphorylation of S15 affects amino-terminal conformation in a way that may stimulate the catalytic activity of SUS and influence membrane association.

Smart nanovehicles based on pH-triggered disassembly of supramolecular peptide-amphiphiles for efficient intracellular drug delivery.

PubMed

Xu, Xianghui; Li, Yunkun; Li, Haiping; Liu, Rong; Sheng, Mingming; He, Bin; Gu, Zhongwei

2014-03-26

A novel type of nanovehicle (NV) based on stimuli-responsive supramolecular peptide-amphiphiles (SPAs, dendritic poly (L-lysine) non-covalently linked poly (L-leucine)) is developed for intracellular drug delivery. To determine the pH-dependent mechanism, the supramolecular peptide-amphiphile system (SPAS) is investigated at different pH conditions using a variety of physical and chemical approaches. The pH-triggered disassembly of SPAS can be attributed to the disappearance of non-covalent interactions within SPAs around the isoelectric point of poly (L-leucine). SPAS is found to encapsulate guest molecules at pH 7.4 but release them at pH 6.2. In this way, SPAS is able to act as a smart NV to deliver its target to tumor cells using intracellular pH as a trigger. The DOX-loaded NVs are approximately 150 nm in size. In vitro release profiles and confocal laser scanning microscopy (CLSM) images of HepG2 cells confirm that lower pH conditions can trigger the disassembly of NVs and so achieve pH-dependent intracellular DOX delivery. In vitro cytotoxicity of the DOX-loaded NVs to HepG2 cells demonstrate that the smart NVs enhance the efficacy of hydrophobic DOX. Fluorescence-activated cell sorting (FACS) and CLSM results show that the NVs can enhance the endocytosis of DOX into HepG2 cells considerably and deliver DOX to the nuclei. © 2013 WILEY-VCH Verlag GmbH & Co. KGaA, Weinheim.

Isolated p.H62L Mutation in the CYP21A2 Gene in a Simple Virilizing 21-Hydroxylase Deficient Patient

PubMed Central

Fernández, Cecilia; Belli, Susana; Buzzalino, Noemi; Dain, Liliana

2013-01-01

Congenital adrenal hyperplasia due to 21-hydroxylase deficiency accounts for 90%–95% of cases. This autosomal recessive disorder has a broad spectrum of clinical forms, ranging from severe or classical, which includes the salt-wasting and simple virilizing forms, to the mild late onset or nonclassical form. Most of the disease-causing mutations described are likely to be the consequence of nonhomologous recombination or gene conversion events between the active CYP21A2 gene and its homologous CYP21A1P pseudogene. Nevertheless, an increasing number of naturally occurring mutations have been found. The change p.H62L is one of the most frequent rare mutations of the CYP21A2 gene. It was suggested that the p.H62L represents a mild mutation that may be responsible for a more severe enzymatic impairment when presented with another mild mutation on the same allele. In this report, a 20-year-old woman carrying an isolated p.H62L mutation in compound heterozygosity with c.283-13A/C>G mutation is described. Although a mildly nonclassical phenotype was expected, clinical signs and hormonal profile of the patient are consistent with a more severe simple virilizing form of 21-hydroxylase deficiency. The study of genotype-phenotype correlation in additional patients would help in defining the role of p.H62L in disease manifestation. PMID:23936690

Isolated p.H62L Mutation in the CYP21A2 Gene in a Simple Virilizing 21-Hydroxylase Deficient Patient.

PubMed

Taboas, Melisa; Fernández, Cecilia; Belli, Susana; Buzzalino, Noemi; Alba, Liliana; Dain, Liliana

2013-01-01

Congenital adrenal hyperplasia due to 21-hydroxylase deficiency accounts for 90%-95% of cases. This autosomal recessive disorder has a broad spectrum of clinical forms, ranging from severe or classical, which includes the salt-wasting and simple virilizing forms, to the mild late onset or nonclassical form. Most of the disease-causing mutations described are likely to be the consequence of nonhomologous recombination or gene conversion events between the active CYP21A2 gene and its homologous CYP21A1P pseudogene. Nevertheless, an increasing number of naturally occurring mutations have been found. The change p.H62L is one of the most frequent rare mutations of the CYP21A2 gene. It was suggested that the p.H62L represents a mild mutation that may be responsible for a more severe enzymatic impairment when presented with another mild mutation on the same allele. In this report, a 20-year-old woman carrying an isolated p.H62L mutation in compound heterozygosity with c.283-13A/C>G mutation is described. Although a mildly nonclassical phenotype was expected, clinical signs and hormonal profile of the patient are consistent with a more severe simple virilizing form of 21-hydroxylase deficiency. The study of genotype-phenotype correlation in additional patients would help in defining the role of p.H62L in disease manifestation.

Investigation of the retention/pH profile of zwitterionic fluoroquinolones in reversed-phase and ion-interaction high performance liquid chromatography.

PubMed

Pistos, C; Tsantili-Kakoulidou, A; Koupparis, M

2005-09-15

The retention/pH profiles of three fluoroquinolones, ofloxacin, norfloxacin and ciprofloxacin, was investigated by means of reversed-phase high performance liquid chromatography (RP-HPLC) and reversed-phase ion-interaction chromatography (RP-IIC), using an octadecylsilane stationary phase and acetonitrile as organic modifier. Sodium hexanesulphonate and tetrabutylammonium hydroxide were used as sources of counter ions in ion-interaction chromatography. The retention/pH profiles under in RP-HPLC were compared to the corresponding lipophilicity/pH profiles. Despite the rather hydrophilic nature of the three fluoroquinolones positive retention factors were obtained while there was a shift of the retention maximum towards more acidic pH values. This behavior was attributed mainly to non-hydrophobic silanophilic interactions with the silanized silica gel material of the stationary phase. In ion-interaction chromatography the effect of counter ions over a broad pH range was found to be ruled rather by the ion pair formation in the mobile phase which led to a drastic decrease in retention as a consequence of the disruption of the zwitterionic structure and thereupon the deliberation of a net charge in the molecules. At pH values at which zwitterionic structure was not favored both the ion-exchange and ion pair formation mechanisms were assumed to contribute to the retention.

Constructing an Institutional Identity in University Tea Rooms: The International PhD Student Experience

ERIC Educational Resources Information Center

Fotovatian, Sepideh; Miller, Jenny

2014-01-01

This case study profiles eight international PhD students and describes the process of the construction and negotiation of their social and institutional identities in an Australian university. Audio-recorded informal conversations of the students highlight the role of social membership, staffroom interactions and language in the construction of…

Nutritional Profile and Chemical Stability of Pasta Fortified with Tilapia (Oreochromis niloticus) Flour

PubMed Central

Monteiro, Maria Lúcia G.; Mársico, Eliane T.; Soares, Manoel S.; Magalhães, Amanda O.; Canto, Anna Carolina V. C. S.; Costa-Lima, Bruno R. C.; Alvares, Thiago S.; Conte, Carlos A.

2016-01-01

Physicochemical parameters of pasta enriched with tilapia (Oreochromis niloticus) flour were investigated. Five formulations were prepared with different concentrations of tilapia flour as partial substitute of wheat flour: pasta without tilapia flour (PTF0%), pasta with 6% (PTF6%), 12% (PTF12%), 17% (PTF17%), and 23% (PTF23%) of tilapia flour. The formulations were assessed for proximate composition, fatty acid and amino acid profile on day 1 whereas, instrumental color parameters (L*, a* and b* values), pH, water activity (aw), and lipid and protein oxidation were evaluated on days 1, 7, 14, and 21 of storage at 25°C. Fortification with tilapia flour increased (p < 0.05) protein, lipid, ash, total essential amino acids, and total polyunsaturated fatty acids contents. In addition, supplementation of pasta with tilapia flour decreased (p < 0.05) lightness and water activity while redness, yellowness, pH values, and lipid oxidation were increased (p < 0.05) in a level-dependent manner. Nevertheless, all formulations were exhibited storage stability at 25°C. In general, protein oxidation was greater (p < 0.05) in the pasta containing 12%, 17%, and 23% of tilapia flour than their counterparts, and the storage promoted an increase (p < 0.05) on the carbonyl content in all formulations. Thus, pasta with 6% of tilapia flour has the potential to be a technological alternative to food industry for the nutritional enrichment of traditional pasta with negligible negative effects on the chemical stability of the final product during 21 days at 25°C. PMID:27973565

Nutritional Profile and Chemical Stability of Pasta Fortified with Tilapia (Oreochromis niloticus) Flour.

PubMed

Monteiro, Maria Lúcia G; Mársico, Eliane T; Soares, Manoel S; Magalhães, Amanda O; Canto, Anna Carolina V C S; Costa-Lima, Bruno R C; Alvares, Thiago S; Conte, Carlos A

2016-01-01

Physicochemical parameters of pasta enriched with tilapia (Oreochromis niloticus) flour were investigated. Five formulations were prepared with different concentrations of tilapia flour as partial substitute of wheat flour: pasta without tilapia flour (PTF0%), pasta with 6% (PTF6%), 12% (PTF12%), 17% (PTF17%), and 23% (PTF23%) of tilapia flour. The formulations were assessed for proximate composition, fatty acid and amino acid profile on day 1 whereas, instrumental color parameters (L*, a* and b* values), pH, water activity (aw), and lipid and protein oxidation were evaluated on days 1, 7, 14, and 21 of storage at 25°C. Fortification with tilapia flour increased (p < 0.05) protein, lipid, ash, total essential amino acids, and total polyunsaturated fatty acids contents. In addition, supplementation of pasta with tilapia flour decreased (p < 0.05) lightness and water activity while redness, yellowness, pH values, and lipid oxidation were increased (p < 0.05) in a level-dependent manner. Nevertheless, all formulations were exhibited storage stability at 25°C. In general, protein oxidation was greater (p < 0.05) in the pasta containing 12%, 17%, and 23% of tilapia flour than their counterparts, and the storage promoted an increase (p < 0.05) on the carbonyl content in all formulations. Thus, pasta with 6% of tilapia flour has the potential to be a technological alternative to food industry for the nutritional enrichment of traditional pasta with negligible negative effects on the chemical stability of the final product during 21 days at 25°C.

Delivery of resveratrol, a red wine polyphenol, from solutions and hydrogels via the skin.

PubMed

Hung, Chi-Feng; Lin, Yin-Ku; Huang, Zih-Rou; Fang, Jia-You

2008-05-01

Resveratrol, the main active polyphenol in red wine, has been demonstrated to show benefits against skin disorders. The bioavailability of orally administered resveratrol is insufficient to permit high enough drug concentrations for systemic therapy. In this study, we examined the feasibility of the topical/transdermal delivery of resveratrol. The effects of vehicles on the in vitro permeation and skin deposition from saturated solutions such as aqueous buffers and soybean oil were investigated. The general trend for the delivery from solutions was: pH 6 buffer=pH 8 buffer>10% glycerol formal in pH 6 buffer>pH 9.9 buffer>pH 10.8 buffer>soybean oil. A linear relationship was established between the permeability coefficient (K(p)) and drug accumulation in the skin reservoir. Viable epidermis/dermis served as the predominant barrier for non-ionic resveratrol permeation. On the other hand, both the stratum corneum (SC) and viable skin acted as barriers to anionic resveratrol. Several prototype hydrogel systems were also studied as resveratrol vehicles. The viscosity but not the polarity of the hydrogels controlled resveratrol permeation/deposition. Piceatannol, a derivative of resveratrol with high pharmacological activity, showed 11.6-fold lower skin permeation compared to resveratrol. The safety profiles of resveratrol suggested that the hydrogel caused no SC disruption or skin erythema. It was concluded that delivery via a skin route may be a potent way to achieve the therapeutic effects of resveratrol. This is the first report to establish the permeation profiles for topically applied resveratrol.

Empirical algorithms to estimate water column pH in the Southern Ocean

NASA Astrophysics Data System (ADS)

Williams, N. L.; Juranek, L. W.; Johnson, K. S.; Feely, R. A.; Riser, S. C.; Talley, L. D.; Russell, J. L.; Sarmiento, J. L.; Wanninkhof, R.

2016-04-01

Empirical algorithms are developed using high-quality GO-SHIP hydrographic measurements of commonly measured parameters (temperature, salinity, pressure, nitrate, and oxygen) that estimate pH in the Pacific sector of the Southern Ocean. The coefficients of determination, R2, are 0.98 for pH from nitrate (pHN) and 0.97 for pH from oxygen (pHOx) with RMS errors of 0.010 and 0.008, respectively. These algorithms are applied to Southern Ocean Carbon and Climate Observations and Modeling (SOCCOM) biogeochemical profiling floats, which include novel sensors (pH, nitrate, oxygen, fluorescence, and backscatter). These algorithms are used to estimate pH on floats with no pH sensors and to validate and adjust pH sensor data from floats with pH sensors. The adjusted float data provide, for the first time, seasonal cycles in surface pH on weekly resolution that range from 0.05 to 0.08 on weekly resolution for the Pacific sector of the Southern Ocean.

Multiscale simulation reveals a multifaceted mechanism of proton permeation through the influenza A M2 proton channel

PubMed Central

Liang, Ruibin; Li, Hui; Swanson, Jessica M. J.; Voth, Gregory A.

2014-01-01

The influenza A virus M2 channel (AM2) is crucial in the viral life cycle. Despite many previous experimental and computational studies, the mechanism of the activating process in which proton permeation acidifies the virion to release the viral RNA and core proteins is not well understood. Herein the AM2 proton permeation process has been systematically characterized using multiscale computer simulations, including quantum, classical, and reactive molecular dynamics methods. We report, to our knowledge, the first complete free-energy profiles for proton transport through the entire AM2 transmembrane domain at various pH values, including explicit treatment of excess proton charge delocalization and shuttling through the His37 tetrad. The free-energy profiles reveal that the excess proton must overcome a large free-energy barrier to diffuse to the His37 tetrad, where it is stabilized in a deep minimum reflecting the delocalization of the excess charge among the histidines and the cost of shuttling the proton past them. At lower pH values the His37 tetrad has a larger total charge that increases the channel width, hydration, and solvent dynamics, in agreement with recent 2D-IR spectroscopic studies. The proton transport barrier becomes smaller, despite the increased charge repulsion, due to backbone expansion and the more dynamic pore water molecules. The calculated conductances are in quantitative agreement with recent experimental measurements. In addition, the free-energy profiles and conductances for proton transport in several mutants provide insights for explaining our findings and those of previous experimental mutagenesis studies. PMID:24979779

Multiscale simulation reveals a multifaceted mechanism of proton permeation through the influenza A M2 proton channel.

PubMed

Liang, Ruibin; Li, Hui; Swanson, Jessica M J; Voth, Gregory A

2014-07-01

The influenza A virus M2 channel (AM2) is crucial in the viral life cycle. Despite many previous experimental and computational studies, the mechanism of the activating process in which proton permeation acidifies the virion to release the viral RNA and core proteins is not well understood. Herein the AM2 proton permeation process has been systematically characterized using multiscale computer simulations, including quantum, classical, and reactive molecular dynamics methods. We report, to our knowledge, the first complete free-energy profiles for proton transport through the entire AM2 transmembrane domain at various pH values, including explicit treatment of excess proton charge delocalization and shuttling through the His37 tetrad. The free-energy profiles reveal that the excess proton must overcome a large free-energy barrier to diffuse to the His37 tetrad, where it is stabilized in a deep minimum reflecting the delocalization of the excess charge among the histidines and the cost of shuttling the proton past them. At lower pH values the His37 tetrad has a larger total charge that increases the channel width, hydration, and solvent dynamics, in agreement with recent 2D-IR spectroscopic studies. The proton transport barrier becomes smaller, despite the increased charge repulsion, due to backbone expansion and the more dynamic pore water molecules. The calculated conductances are in quantitative agreement with recent experimental measurements. In addition, the free-energy profiles and conductances for proton transport in several mutants provide insights for explaining our findings and those of previous experimental mutagenesis studies.

Genomic and Expression Profiling of Benign and Malignant Nerve Sheath Profiling of Benign and Malignant Nerve Sheath

DTIC Science & Technology

2007-05-01

Benign and Malignant Nerve Sheath Tumors in Neurofibromatosis Patients PRINCIPAL INVESTIGATOR: Matt van de Rijn, M.D., Ph.D. Torsten...Annual 3. DATES COVERED 1 May 2006 –30 Apr 2007 4. TITLE AND SUBTITLE 5a. CONTRACT NUMBER Genomic and Expression Profiling of Benign and Malignant Nerve...Award Number: DAMD17-03-1-0297 Title: Genomic and Expression Profiling of Benign and Malignant Nerve Sheath Tumors in Neurofibromatosis

Mixed-mode ion exchange-based integrated proteomics technology for fast and deep plasma proteome profiling.

PubMed

Xue, Lu; Lin, Lin; Zhou, Wenbin; Chen, Wendong; Tang, Jun; Sun, Xiujie; Huang, Peiwu; Tian, Ruijun

2018-06-09

Plasma proteome profiling by LC-MS based proteomics has drawn great attention recently for biomarker discovery from blood liquid biopsy. Due to standard multi-step sample preparation could potentially cause plasma protein degradation and analysis variation, integrated proteomics sample preparation technologies became promising solution towards this end. Here, we developed a fully integrated proteomics sample preparation technology for both fast and deep plasma proteome profiling under its native pH. All the sample preparation steps, including protein digestion and two-dimensional fractionation by both mixed-mode ion exchange and high-pH reversed phase mechanism were integrated into one spintip device for the first time. The mixed-mode ion exchange beads design achieved the sample loading at neutral pH and protein digestion within 30 min. Potential sample loss and protein degradation by pH changing could be voided. 1 μL of plasma sample with depletion of high abundant proteins was processed by the developed technology with 12 equally distributed fractions and analyzed with 12 h of LC-MS gradient time, resulting in the identification of 862 proteins. The combination of the Mixed-mode-SISPROT and data-independent MS method achieved fast plasma proteome profiling in 2 h with high identification overlap and quantification precision for a proof-of-concept study of plasma samples from 5 healthy donors. We expect that the Mixed-mode-SISPROT become a generally applicable sample preparation technology for clinical oriented plasma proteome profiling. Copyright © 2018 Elsevier B.V. All rights reserved.

Investigation of the Dissolution Profile of Gliclazide Modified-Release Tablets Using Different Apparatuses and Dissolution Conditions.

PubMed

Skripnik, K K S; Riekes, M K; Pezzini, B R; Cardoso, S G; Stulzer, H K

2017-07-01

In the absence of an official dissolution method for modified-release tablets of gliclazide, dissolution parameters, such as apparatuses (1, 2, and 3), rotation speeds, pH, and composition of the dissolution medium were investigated. The results show that although the drug presents a pH-mediated solubility (pH 7.0 > 6.8 > 6.4 > 6.0 > 5.5 > 4.5), the in vitro release of the studied tablets was not dependent on this parameter, despite of the apparatus tested. On the other hand, the rotation speed demonstrated a greater influence (100 rpm >50 rpm). Using similar hydrodynamic conditions, the three different apparatuses were compared in pH 6.8 and provided the following trend: apparatus 1 at 100 rpm >2 at 50 rpm ≈3 at 10 dpm. As a complete, but slow release is expected from modified-release formulations, apparatus 2, in phosphate buffer pH 6.8 and 100 rpm, were selected as the optimized dissolution method. In comparison to apparatus 1 under the same conditions, the paddle avoids the stickiness of formulation excipients at the mesh of the basket, which could prejudice the release of gliclazide. Results obtained with biorelevant medium through the developed dissolution method were similar to the buffer solution pH 6.8. The application of the optimized method as a quality control test between two different brands of gliclazide modified-release tablets showed that both dissolution profiles were considered similar by the similarity factor (f2 = 51.8). The investigation of these dissolution profiles indicated a dissolution kinetic following first-order model.

Dynamic vertical profiles of peat porewater chemistry in a northern peatland

Treesearch

Natalie A. Griffiths; Stephen D. Sebestyen

2016-01-01

We measured pH, cations, nutrients, and total organic carbon (TOC) over 3 years to examine weekly to monthly variability in porewater chemistry depth profiles (0–3.0 m) in an ombrotrophic bog in Minnesota, USA. We also compared temporal variation at one location to spatial variation in depth profiles at 16 locations across the bog. Most solutes exhibited large...

pH Control Enables Simultaneous Enhancement of Nitrogen Retention and N2O Reduction in Shewanella loihica Strain PV-4.

PubMed

Kim, Hayeon; Park, Doyoung; Yoon, Sukhwan

2017-01-01

pH has been recognized as one of the key environmental parameters with significant impacts on the nitrogen cycle in the environment. In this study, the effects of pH on NO 3 - /NO 2 - fate and N 2 O emission were examined with Shewanella loihica strain PV-4, an organism with complete denitrification and respiratory ammonification pathways. Strain PV-4 was incubated at varying pH with lactate as the electron donor and NO 3 - /NO 2 - and N 2 O as the electron acceptors. When incubated with NO 3 - and N 2 O at pH 6.0, transient accumulation of N 2 O was observed and no significant NH 4 + production was observed. At pH 7.0 and 8.0, strain PV-4 served as a N 2 O sink, as N 2 O concentration decreased consistently without accumulation. Respiratory ammonification was upregulated in the experiments performed at these higher pH values. When NO 2 - was used in place of NO 3 - , neither growth nor NO 2 - reduction was observed at pH 6.0. NH 4 + was the exclusive product from NO 2 - reduction at both pH 7.0 and 8.0 and neither production nor consumption of N 2 O was observed, suggesting that NO 2 - regulation superseded pH effects on the nitrogen-oxide dissimilation reactions. When NO 3 - was the electron acceptor, nirK transcription was significantly upregulated upon cultivation at pH 6.0, while nrfA transcription was significantly upregulated at pH 8.0. The highest level of nosZ transcription was observed at pH 6.0 and the lowest at pH 8.0. With NO 2 - as the electron acceptor, transcription profiles of nirK, nrfA , and nosZ were statistically indistinguishable between pH 7.0 and 8.0. The transcriptions of nirK and nosZ were severely downregulated regardless of pH. These observations suggested that the kinetic imbalance between N 2 O production and consumption, but neither decrease in expression nor activity of NosZ, was the major cause of N 2 O accumulation at pH 6.0. The findings also suggest that simultaneous enhancement of nitrogen retention and N 2 O emission reduction may be feasible through pH modulation, but only in environments where C:N or NO 2 - :NO 3 - ratio does not exhibit overarching control over the NO 3 - /NO 2 - reduction pathways.

In Silico Functional Networks Identified in Fish Nucleated Red Blood Cells by Means of Transcriptomic and Proteomic Profiling.

PubMed

Puente-Marin, Sara; Nombela, Iván; Ciordia, Sergio; Mena, María Carmen; Chico, Verónica; Coll, Julio; Ortega-Villaizan, María Del Mar

2018-04-09

Nucleated red blood cells (RBCs) of fish have, in the last decade, been implicated in several immune-related functions, such as antiviral response, phagocytosis or cytokine-mediated signaling. RNA-sequencing (RNA-seq) and label-free shotgun proteomic analyses were carried out for in silico functional pathway profiling of rainbow trout RBCs. For RNA-seq, a de novo assembly was conducted, in order to create a transcriptome database for RBCs. For proteome profiling, we developed a proteomic method that combined: (a) fractionation into cytosolic and membrane fractions, (b) hemoglobin removal of the cytosolic fraction, (c) protein digestion, and (d) a novel step with pH reversed-phase peptide fractionation and final Liquid Chromatography Electrospray Ionization Tandem Mass Spectrometric (LC ESI-MS/MS) analysis of each fraction. Combined transcriptome- and proteome- sequencing data identified, in silico, novel and striking immune functional networks for rainbow trout nucleated RBCs, which are mainly linked to innate and adaptive immunity. Functional pathways related to regulation of hematopoietic cell differentiation, antigen presentation via major histocompatibility complex class II (MHCII), leukocyte differentiation and regulation of leukocyte activation were identified. These preliminary findings further implicate nucleated RBCs in immune function, such as antigen presentation and leukocyte activation.

In Silico Functional Networks Identified in Fish Nucleated Red Blood Cells by Means of Transcriptomic and Proteomic Profiling

PubMed Central

Puente-Marin, Sara; Ciordia, Sergio; Mena, María Carmen; Chico, Verónica; Coll, Julio

2018-01-01

Nucleated red blood cells (RBCs) of fish have, in the last decade, been implicated in several immune-related functions, such as antiviral response, phagocytosis or cytokine-mediated signaling. RNA-sequencing (RNA-seq) and label-free shotgun proteomic analyses were carried out for in silico functional pathway profiling of rainbow trout RBCs. For RNA-seq, a de novo assembly was conducted, in order to create a transcriptome database for RBCs. For proteome profiling, we developed a proteomic method that combined: (a) fractionation into cytosolic and membrane fractions, (b) hemoglobin removal of the cytosolic fraction, (c) protein digestion, and (d) a novel step with pH reversed-phase peptide fractionation and final Liquid Chromatography Electrospray Ionization Tandem Mass Spectrometric (LC ESI-MS/MS) analysis of each fraction. Combined transcriptome- and proteome- sequencing data identified, in silico, novel and striking immune functional networks for rainbow trout nucleated RBCs, which are mainly linked to innate and adaptive immunity. Functional pathways related to regulation of hematopoietic cell differentiation, antigen presentation via major histocompatibility complex class II (MHCII), leukocyte differentiation and regulation of leukocyte activation were identified. These preliminary findings further implicate nucleated RBCs in immune function, such as antigen presentation and leukocyte activation. PMID:29642539

Sources and pathways of artificial radionuclides to soils at a High Arctic site.

PubMed

Lokas, E; Bartmiński, P; Wachniew, P; Mietelski, J W; Kawiak, T; Srodoń, J

2014-11-01

Activity concentrations, inventories and activity ratios of (137)Cs, (238)Pu, (239 + 240)Pu and (241)Am in soil profiles were surveyed in the dry tundra and the adjoining proglacial zones of glaciers at a High Arctic site on Svalbard. Vertical profiles of radionuclide activities were determined in up to 14-cm-thick soil sequences. Additionally, soil properties (pH, organic matter, texture, mineral composition and sorption capacity) were analyzed. Results obtained in this study revealed a large range of activity concentrations and inventories of the fallout radionuclides from the undetectable to the uncommonly high levels (inventories of 30,900 ± 940, 47 ± 6, 886 ± 80 and 296 ± 19 Bq/m(2) for (137)Cs, (238)Pu, (239 + 240)Pu and (241)Am, respectively) found in two profiles from the proglacial zone. Concentration of these initially airborne radionuclides in the proglacial zone soils is related to their accumulation in cryoconites that have a large ability to concentrate trace metals. The cryoconites develop on the surface of glaciers, and the material they accumulate is deposited on land surface after the glaciers retreat. The radionuclide inventories in the tundra soils, which effectively retain radionuclides due to high organic matter contents, were comparable to the global fallout deposition for this region of the world. The (238)Pu/(239 + 240)Pu activity ratios for tundra soils suggested global fallout as the dominant source of Pu. The (238)Pu/(239 + 240)Pu and (239 + 240)Pu/(137)Cs activity ratios in the proglacial soils pointed to possible contributions of these radionuclides from other, unidentified sources.

Single-walled carbon nanotubes-ciprofloxacin nanoantibiotic: strategy to improve ciprofloxacin antibacterial activity.

PubMed

Assali, Mohyeddin; Zaid, Abdel Naser; Abdallah, Farah; Almasri, Motasem; Khayyat, Rasha

2017-01-01

As infectious diseases continue to be one of the greatest health challenges worldwide, the demand toward alternative agents is continuously increasing. Recent advancement in nanotechnology has expanded our ability to design and construct nanomaterials to treat bacterial infections. Carbon nanotubes are one among these nanomaterials. Herein, we describe the covalent functionalization of the single-walled carbon nanotubes (SWCNTs) with multiple molecules of ciprofloxacin. The prepared nanoantibiotics were characterized using different techniques, including transmission electron microscopy, Raman spectroscopy, and thermogravimetric analysis. The characterization of the nanoantibiotics confirmed the successful covalent functionalization of the SWCNTs with 55% of functionalization as has been observed by thermogravimetric analysis. The release profile revealed that 90% of the loaded ciprofloxacin was released within 2.5 h at pH 7.4 showing a first-order release profile with R 2 >0.99. Interestingly, the results of the antibacterial activity indicated that the functionalized SWCNTs have significant increase in the antibacterial activity against the three strains of bacteria - by 16-fold for Staphylococcus aureus and Pseudomonas aeruginosa and by 8-fold for Escherichia coli - in comparison to the ciprofloxacin free drug. Moreover, the synthesized nanoantibiotic showed high hemocompatibility and cytocompatibility over a wide concentration range.

Single-walled carbon nanotubes-ciprofloxacin nanoantibiotic: strategy to improve ciprofloxacin antibacterial activity

PubMed Central

Assali, Mohyeddin; Zaid, Abdel Naser; Abdallah, Farah; Almasri, Motasem; Khayyat, Rasha

2017-01-01

As infectious diseases continue to be one of the greatest health challenges worldwide, the demand toward alternative agents is continuously increasing. Recent advancement in nanotechnology has expanded our ability to design and construct nanomaterials to treat bacterial infections. Carbon nanotubes are one among these nanomaterials. Herein, we describe the covalent functionalization of the single-walled carbon nanotubes (SWCNTs) with multiple molecules of ciprofloxacin. The prepared nanoantibiotics were characterized using different techniques, including transmission electron microscopy, Raman spectroscopy, and thermogravimetric analysis. The characterization of the nanoantibiotics confirmed the successful covalent functionalization of the SWCNTs with 55% of functionalization as has been observed by thermogravimetric analysis. The release profile revealed that 90% of the loaded ciprofloxacin was released within 2.5 h at pH 7.4 showing a first-order release profile with R2>0.99. Interestingly, the results of the antibacterial activity indicated that the functionalized SWCNTs have significant increase in the antibacterial activity against the three strains of bacteria – by 16-fold for Staphylococcus aureus and Pseudomonas aeruginosa and by 8-fold for Escherichia coli – in comparison to the ciprofloxacin free drug. Moreover, the synthesized nanoantibiotic showed high hemocompatibility and cytocompatibility over a wide concentration range. PMID:28924348

In vitro dissolution of proton-pump inhibitor products intended for paediatric and geriatric use in physiological bicarbonate buffer.

PubMed

Liu, Fang; Shokrollahi, Honaz

2015-05-15

Proton-pump inhibitor (PPI) products based on enteric coated multiparticulates are design to meet the needs of patients who cannot swallow tablets such as children and older adults. Enteric coated PPI preparations exhibit delays in in vivo absorption and onset of antisecretory effects, which is not reflected by the rapid in vitro dissolution in compendial pH 6.8 phosphate buffer commonly used for assessment of these products. A more representative and physiological medium, pH 6.8 mHanks bicarbonate buffer, was used in this study to evaluate the in vitro dissolution of enteric coated multiparticulate-based PPI products. Commercially available omeprazole, lansoprazole and esomeprazole products were subject to dissolution tests using USP-II apparatus in pH 4.5 phosphate buffer saline for 45 min (acid stage) followed by pH 6.8 phosphate buffer or pH 6.8 mHanks bicarbonate buffer. In pH 6.8 phosphate buffer, all nine tested products displayed rapid and comparable dissolution profiles meeting the pharmacopeia requirements for delayed release preparations. In pH 6.8 mHanks buffer, drug release was delayed and failed the pharmacopeia requirements from most enteric coated preparations. Despite that the same enteric polymer, methacrylic acid-ethyl acrylate copolymer (1:1), was applied to all commercial multiparticulate-based products, marked differences were observed between dissolution profiles of these preparations. The use of pH 6.8 physiological bicarbonate (mHanks) buffer can serve as a useful tool to provide realistic and discriminative in vitro release assessment of enteric coated PPI preparations and to assist rational formulation development of these products. Copyright © 2015 Elsevier B.V. All rights reserved.

Effect of Microenvironmental pH Modulation on the Dissolution Rate and Oral Absorption of the Salt of a Weak Acid - Case Study of GDC-0810.

PubMed

Hou, Hao Helen; Jia, Wei; Liu, Lichuan; Cheeti, Sravanthi; Li, Jane; Nauka, Ewa; Nagapudi, Karthik

2018-01-29

The purpose of this work is to investigate the effect of microenvironmental pH modulation on the in vitro dissolution rate and oral absorption of GDC-0810, an oral anti-cancer drug, in human. The pH-solubility profile of GDC-0810 free acid and pH max of its N-Methyl-D-glucamine (NMG) salt were determined. Precipitation studies were conducted for GDC-0810 NMG salt at different pH values. GDC-0810 200-mg dose NMG salt tablet formulations containing different levels of sodium bicarbonate as the pH modifier were tested for dissolution under the dual pH-dilution scheme. Three tablet formulations were evaluated in human as a part of a relative bioavailability study. A 200-mg dose of GDC-0810 was administered QD with low fat food. Intrinsic solubility of GDC-0810 free acid was found to be extremely low. The pH max of the NMG salt suggested a strong tendency for form conversion to the free acid under GI conditions. In vitro dissolution profiles showed that the dissolution rate and extent of GDC-0810 increased with increasing the level of sodium bicarbonate in the formulation. The human PK data showed a similar trend for the geometric mean of C max and AUC 0-t for formulations containing 5%, 10%, and 15% sodium bicarbonate, but the difference is not statistically significant. Incorporation of a basic pH modifier, sodium bicarbonate, in GDC-0810 NMG salt tablet formulations enhanced in vitro dissolution rate of GDC-0810 via microenvironmental pH modulation. The human PK data showed no statistically significant difference in drug exposure from tablets containing 5%, 10%, and 15% sodium bicarbonate.

Investigation of the adsorption mechanism of a peptide in reversed phase liquid chromatography, from pH controlled and uncontrolled solutions

DOE Office of Scientific and Technical Information (OSTI.GOV)

Gritti, Fabrice; Guiochon, Georges A

2009-01-01

The single-component equilibrium adsorption of the tripeptide Leucyl-Leucyl-Leucine (LLL) on a high-efficiency Jupiter Proteo column (C{sub 12}) was investigated experimentally and modeled theoretically. The experimental equilibrium isotherms of LLL for adsorption on a C{sub 12} packing material from an aqueous solution of methanol (48%) and trifluoroacetic acid (0.1%) were measured by frontal analysis (FA). The FA measurements were done with two solutions, one in which the pH was controlled, the other in which it was not. Two solutions of LLL in the mobile phase were prepared (4.3 and 5.4 g/L) and their pH measured (2.94 and 2.88), respectively. The firstmore » solution was titrated with TFA to match the pH of the mobile phase (2.03), so its pH was controlled. The pH of the other solution was left uncontrolled. In both cases the isotherms could be modeled by a bi-Langmuir equation, a choice consistent with the bimodal affinity energy distribution (AED) obtained for LLL. The isotherm parameters derived from the inverse method (IM) of isotherm determination under controlled pH conditions (by fitting calculated profiles to experimental breakthrough profiles) are in a good agreement with those derived from the FA data. Under uncontrolled pH conditions, the application of IM suggests the coexistence of two different adsorption mechanisms. According to the isotherm parameters found by these three methods (FA, AED and IM), the C{sub 12}-bonded silica can adsorb around 500 and 70 g/L of LLL under controlled and uncontrolled pH conditions, respectively. The adsorption of LLL on the C{sub 12} material strongly depends on the pH of the mobile phase and on the quantity of TFA added, which plays the role of an ion-pairing agent.« less

Applied PhD Research in a Work-Based Environment: An Activity Theory-Based Analysis

ERIC Educational Resources Information Center

Granata, S. N.; Dochy, F.

2016-01-01

Activity theory is used to compare PhD undertaken at university, that is, academic PhD, with PhD performed in collaboration with industry, that is, semi-industrial PhD. The research is divided into a literature review and a case study. Semi-industrial and academic PhD are modelled as activity systems, and differences are highlighted in terms of…

Design and Evaluation of Hydrophilic Matrix System for pH-Independent Sustained Release of Weakly Acidic Poorly Soluble Drug.

PubMed

Huang, Jinheng; Lin, Huaqing; Peng, Bingxin; Huang, Qianfeng; Shuai, Fangzhou; Xie, Yanxian

2018-04-30

The aim of this research was to design and evaluate a hydrophilic matrix system for sustained release of glipizide, a weakly acidic poor soluble drug. A combination of inclusion complexation and microenvironmental pH modification techniques was utilized to improve the dissolution and pH-independent release of glipizide. Hydroxypropyl-β-cyclodextrin (HP-β-CD) was used as the complexation agent while sodium citrate and magnesium oxide (MgO) were used as model pH modifiers. The hydrophilic matrix tablets were prepared by powder direct compression and evaluated by in vitro dissolution study respectively in pH 6.8 and pH 1.2 dissolution media. The formulations containing MgO exhibited increased cumulative drug release from less than 40% in the reference formulation to 90% within 24 h in acidic media (pH 1.2). The release profile in acidic media was similar to the alkaline media (pH 6.8) with a similarity factor (f 2 ) of 55.0, suggesting the weakening of the effect of pH on the dissolution efficiency of glipizide. The release profile fitted well into the Higuchi model and the dominant mechanism of drug release was Fickian diffusion while case II transport/polymer relaxation occurred. In conclusion, combining inclusion complexation agents and pH modifiers had improved the dissolution of glipizide as well as achieved the pH-independent release profile.

Effects of mouthrinses with chlorhexidine and zinc ions combined with fluoride on the viability and glycolytic activity of dental plaque.

PubMed

Giertsen, E; Scheie, A A

1995-10-01

Inhibition of plaque acidogenicity by a mouthrinse with chlorhexidine (CHX) or zinc ions has been ascribed to a prolonged bacteriostasis due to substantive properties of the agents. The present aim was to study the effects of mouthrinses with CHX and Zn ions combined with fluoride on the viability and glycolytic activity of dental plaque in order to assess the bacteriostatic versus possible bactericidal effects. Following 2 d of plaque accumulation, 4 groups of 10 students rinsed with either 12 mM NaF (F), 0.55 mM CHX diacetate+F (F-CHX), 10 mM Zn acetate+F (F-Zn), or with the three agents in combination (F-CHX-Zn). Plaque samples were collected before and 90 min after mouthrinsing. Thereafter, the in vivo plaque pH response to sucrose was monitored in each student using touch microelectrodes. F-CHX and F-CHX-Zn reduced the in vivo pH fall significantly as compared with F, whereas F-Zn exerted a non-significant inhibition. Pooled pre- and post-rinse plaque samples were used to measure the pH fall during fermentation of [14C]-glucose, and the glycolytic profiles were analyzed by HPLC. Bacterial viability was assessed by counting the colony-forming units (CFU). All mouthrinses except F reduced glucose consumption and acid formation and thus the pH fall. F-CHX reduced the CFU equal to the reduction of glucose consumption, indicating that inhibition of plaque acidogenicity was due to a bactericidal rather than a bacteriostatic effect. F and F-Zn did not reduce the CFU, thus F-Zn decreased glucose metabolism without affecting plaque viability. F-CHX-Zn reduced both the CFU and glucose metabolism of surviving plaque microorganisms.

DOE Office of Scientific and Technical Information (OSTI.GOV)

Solc, J.

The reclamation effort typically deals with consequences of mining activity instead of being planned well before the mining. Detailed assessment of principal hydro- and geochemical processes participating in pore and groundwater chemistry evolution was carried out at three surface mine localities in North Dakota-the Fritz mine, the Indian Head mine, and the Velva mine. The geochemical model MINTEQUA2 and advanced statistical analysis coupled with traditional interpretive techniques were used to determine site-specific environmental characteristics and to compare the differences between study sites. Multivariate statistical analysis indicates that sulfate, magnesium, calcium, the gypsum saturation index, and sodium contribute the most tomore » overall differences in groundwater chemistry between study sites. Soil paste extract pH and EC measurements performed on over 3700 samples document extremely acidic soils at the Fritz mine. The number of samples with pH <5.5 reaches 80%-90% of total samples from discrete depth near the top of the soil profile at the Fritz mine. Soil samples from Indian Head and Velva do not indicate the acidity below the pH of 5.5 limit. The percentage of samples with EC > 3 mS cm{sup -1} is between 20% and 40% at the Fritz mine and below 20% for samples from Indian Head and Velva. The results of geochemical modeling indicate an increased tendency for gypsum saturation within the vadose zone, particularly within the lands disturbed by mining activity. This trend is directly associated with increased concentrations of sulfate anions as a result of mineral oxidation. Geochemical modeling, statistical analysis, and soil extract pH and EC measurements proved to be reliable, fast, and relatively cost-effective tools for the assessment of soil acidity, the extent of the oxidation zone, and the potential for negative impact on pore and groundwater chemistry.« less

Glyphosate Inhibition of 5-Enolpyruvylshikimate 3-Phosphate Synthase from Suspension-Cultured Cells of Nicotiana silvestris.

PubMed

Rubin, J L; Gaines, C G; Jensen, R A

1984-07-01

Treatment of isogenic suspension-cultured cells of Nicotiana silvestris Speg. et Comes with glyphosate (N-[phosphonomethyl]glycine) led to elevated levels of intracellular shikimate (364-fold increase by 1.0 millimolar glyphosate). In the presence of glyphosate, it is likely that most molecules of shikimate originate from the action of 3-deoxy-d-arabino-heptulosonate 7-phosphate (DAHP) synthase-Mn since this isozyme, in contrast to the DAHP synthase-Co isozyme, is insensitive to inhibition by glyphosate. 5-Enolpyruvylshikimate 3-phosphate (EPSP) synthase (EC 2.5.1.19) from N. silvestris was sensitive to micromolar concentrations of glyphosate and possessed a single inhibitor binding site. Rigorous kinetic studies of EPSP synthase required resolution from the multiple phosphatase activities present in crude extracts, a result achieved by ion-exchange column chromatography. Although EPSP synthase exhibited a broad pH profile (50% of maximal activity between pH 6.2 and 8.5), sensitivity to glyphosate increased dramatically with increasing pH within this range. In accordance with these data and the pK(a) values of glyphosate, it is likely that the ionic form of glyphosate inhibiting EPSP synthase is COO(-)CH(2)NH(2) (+)CH(2)PO(3) (2-), and that a completely ionized phosphono group is essential for inhibition. At pH 7.0, inhibition was competitive with respect to phosphoenolpyruvate (K(i) = 1.25 micromolar) and uncompetitive with respect to shikimate-3-P (K(i)' = 18.3 micromolar). All data were consistent with a mechanism of inhibition in which glyphosate competes with PEP for binding to an [enzyme:shikimate-3-P] complex and ultimately forms the dead-end complex of [enzyme:shikimate-3-P:glyphosate].

Effects of feeding the herb Borreria latifolia on the meat quality of village chickens in Malaysia.

PubMed

Abbood, Ali A; Kassim, Azhar Bin; Jawad, Hasan S A; Manap, Yazid Abdul; Sazili, Awis Qurni

2017-06-01

An experiment was carried out to estimate the meat quality characteristics of village chickens (Gallus gallus) fed diets supplemented with dry leaves of Borreria latifolia (BL) used as a potential antioxidant source in chicken feed. In this study, 252 sexed 9-week-old village chickens with mean live body weight of 1,525.4 g for males and 1,254.1 g for females were divided into 7 groups (each group 18 birds) for each sex represented in 2 experiments. The first experiment was to evaluate the antioxidant activity of BL and the effect on meat quality through a comparison with Rosmarinus officinalis (RO); hence, 3 groups were conducted and included: T1 (control), basal diet without supplementation; T2, basal diet with 1% of BL; T3, basal diet with 1% of RO. T2 and T3 significantly affect pH value, lipid oxidation, cooking loss, and overall acceptability compared to T1, while no significant difference was observed between the dietary groups in respect of drip loss, color, tenderness, fatty acid profile, and meat composition. Furthermore, a significant effect of sex on lipid oxidation, pH, yellowness, and fatty acid profile was observed. There was no significant effect of sex on WHC, tenderness, lightness, redness, and sensory evaluation. A significant influence of postmortem aging period was detected on lipid oxidation, pH, tenderness, cooking loss, and redness. The obtained result in this study revealed a significance in the interaction of herb by sex in pH parameter and between herb and sex, herb by aging period, sex by aging period, and the herb by sex by aging period interactions with regard to lipid oxidation test. The second experiment was to estimate the effect of 3 different levels of BL on meat quality. Four groups were provided and involved: T1 (control), basal diet without supplementation; T2, basal diet with 1.5% of BL; T3, basal diet with 2% of BL; and T4, basal diet with 2.5% of BL. The result of this study showed a significant effect (P < 0.05) of the dietary groups on redness value, sensory evaluation, and lipid oxidation. A significant effect of sex on fatty acid profile and lipid oxidation was detected in addition a significant effect of postmortem aging period on redness, pH, tenderness, cooking loss, and lipid oxidation was noticed. There was no significance in the interaction between herb and sex through pH test while a significance in the interaction between herb and sex, herb  × aging period, sex × aging period, and herb × sex × aging period interactions with regard to lipid oxidation was found. It can be concluded that dietary supplementation of BL improved oxidative stability and enhanced meat quality. © 2017 Poultry Science Association Inc.

Screening for Indian isolates of egg-parasitic fungi for use in biological control of fascioliasis and amphistomiasis in ruminant livestock.

PubMed

De, S; Sanyal, P K; Sarkar, A K; Patel, N K; Pal, S; Mandal, S C

2008-09-01

Wild isolates of the egg-parasitic fungi Paecilomyces lilacinus and Verticillium chlamydosporium, obtained from the organic environment of Durg, Chhattisgarh, India, were subjected to screening for in vitro growth using different media types, range of incubation temperature and pH, and their predatory activity to the eggs of Fasciola gigantica and Gigantocotyle explanatum. Maximum growth of P. lilacinus was obtained in corn-meal agar compared to any other media types. The preferred medium for growth of V. chlamydosporium was corn-meal agar, followed by potato-dextrose agar. After initial growth for 16 h of incubation, no growth was observed in water agar for both the fungi. Six different temperatures--4 degrees C, 10 degrees C, 18 degrees C, 26 degrees C, 34 degrees C and 40 degrees C--were used to observe growth profiles of the fungi in corn-meal agar medium. While no and very little growth of P. lilacinus and V. chlamydosporium was observed at 4 degrees C and 10 degrees C, respectively, growth profiles of both the fungi were optimal at 26-40 degrees C. A range of pH (pH 4-8) supported growth of both P. lilacinus and V. chlamydosporium. Full-grown plates of the fungi baited with viable eggs of F. gigantica and G. explanatum revealed that V. chlamydosporium was more vigorous in its egg-parasitic ability compared to P. lilacinus. Distortion of the eggs started on day 2-3 of egg baiting in culture plates of V. chlamydosporium, with complete distortion by day 7. On the contrary, P. lilacinus exhibited very limited egg-parasitic ability and some of the baited eggs even showed development of miracidia.

Casein micelle dissociation in skim milk during high-pressure treatment: effects of pressure, pH, and temperature.

PubMed

Orlien, V; Boserup, L; Olsen, K

2010-01-01

The effect of pH (from 5.5 to 7.5) and temperature (from 5 to 40 degrees C) on the turbidity of reconstituted skim milk powder was investigated at ambient pressure and in situ under pressure (up to 500MPa) by measurement of light scattering. High-pressure treatment reduced the turbidity of milk for all combinations of pH and temperature due to micelle dissociation. The turbidity profiles had a characteristic sigmoidal shape in which almost no effect on turbidity was observed at low pressures (100MPa), followed by a stronger pressure dependency over a pressure range of 150MPa during which turbidity decreased extremely. From the turbidity profiles, the threshold pressure for disruption of micelle integrity was determined and ranged from 150MPa at low pH to 350-400MPa at high pH. The threshold pressure diagram clearly showed a relationship between the barostability of casein micelles and pH, whereas almost no effect of temperature was shown. This remarkable pH effect was a consequence of pressure-induced changes in the electrostatic interactions between colloidal calcium phosphate and the caseins responsible for maintaining micellar structure and was explained by a shift in the calcium phosphate balance in the micelle-serum system. Accordingly, a mechanism for high pressure-induced disruption of micelle integrity is suggested in which the state of calcium plays a crucial role in the micelle dissociation process. Copyright 2010 American Dairy Science Association. Published by Elsevier Inc. All rights reserved.

Comparative study on the in vitro performance of blister molded and conventional lornoxicam immediate release liquitablets: accelerated stability study and anti-inflammatory and ulcerogenic effects.

PubMed

El-Setouhy, Doaa Ahmed; Gamiel, Alaa Abdel-Rahman; Badawi, Alia Abd El-Latif; Osman, Afaf Sayed; Labib, Dina Ahmed

2017-03-01

Lornoxicam is a potent non-steroidal anti-inflammatory drug (NSAID). It shows limited solubility in the gastric pH, delayed bioavailability and pharmacodynamic effects with aggravated gastric side effects (due to longer residence in the stomach wall). To enhance dissolution of lornoxicam in the gastric fluid and expectedly absorption and pharmacological action, with less ulcerogenic effects. Formulation of immediate release (IR) lornoxicam liquitablets containing both liquid and solid release modulators (wetting agent, solubilizers and microenvironmental pH modifiers). Beside the traditional direct compression technique employed for the preparation of liquitablets a new technique, blister molding, was also used. The effect of the two different manufacturing methods on the fast release characteristics (rapid disintegration and dissolution) was studied. Stability and pharmacological activity of the optimum formula were also explored. Similarity factor pointed out the superiority of molding technique in enhancing dissolution of lornoxicam owing to significant crystallinity reduction (XRD). Optimum formula showed negligible change in drug content and dissolution profiles over 12 weeks, significantly improved anti-inflammatory activity and significantly reduced gastric ulcerative effect over pure lornoxicam and commercial formula. Blister molded lornoxicam liquitablet of improved dissolution and pharmacological activity and less gastric erosion was successfully prepared.

Role of alloy additions on strengthening in 17-4 PH stainless steel

NASA Astrophysics Data System (ADS)

Murthy, Arpana Sudershan

Alloy modifications by addition of niobium (Nb), vanadium (V), nitrogen (N) and cobalt (Co) to cast 17-4 PH steel were investigated to determine the effect on mechanical properties. Additions of Nb, V, and N increased the yield strength from 1120 MPa to 1310 MPa while decreased the room temperature charpy V notch (CVN) toughness from 20 J to four Joules. The addition of Co to cast 17-4 PH steel enhanced the yield strength and CVN toughness from 1140 MPa to 1290 MPa and from 3.7 J to 5.5 J, respectively. In the base 17-4 PH steel, an increase in block width from 2.27 ± 0.10 μm in the solution treated condition to 3.06 ± 0.17 μm upon aging at 755 K was measured using orientation image microscopy. Cobalt inhibited recrystallization and block boundary migration during aging resulting in a finer martensitic block structure. The influence of Co on copper (Cu) precipitation in steels was studied using atom probe tomography. A narrower precipitate size distribution was observed in the steels with Co addition. The concentration profile across the matrix / precipitate interface indicated rejection of Co atoms from the copper precipitates. This behavior was observed to be energetically favorable using first principle calculations. The activation energies for Cu precipitation increased from 205 kJ/ mol in the non-cobalt containing alloy, to 243 kJ/ mol, and 272 kJ/ mol in alloys with 3 wt. %Co, and 7 wt. % Co, respectively. The role of Co on Cu precipitation in cast 17-4 PH steel is proposed as follows: (i) Co is rejected out of the Cu precipitate and sets up a barrier to the growth of the Cu precipitate; (ii) results in Cu precipitates of smaller size and narrower distribution; (iii) the coarsening of Cu precipitates is inhibited; and (iv) the activation energy for Cu precipitation increases.

Release behavior and toxicity profiles towards leukemia (WEHI-3B) cell lines of 6-mercaptopurine-PEG-coated magnetite nanoparticles delivery system.

PubMed

Dorniani, Dena; Kura, Aminu Umar; Hussein-Al-Ali, Samer Hasan; bin Hussein, Mohd Zobir; Fakurazi, Sharida; Shaari, Abdul Halim; Ahmad, Zalinah

2014-01-01

The coating of an active drug, 6-mercaptopurine, into the iron oxide nanoparticles-polyethylene glycol (FNPs-PEG) in order to form a new nanocomposite, FPEGMP-2, was accomplished using coprecipitation technique. The resulting nanosized with a narrow size distribution magnetic polymeric particles show the superparamagnetic properties with 38.6 emu/g saturation magnetization at room temperature. Fourier transform infrared spectroscopy and the thermal analysis study supported the formation of the nanocomposite and the enhancement of thermal stability in the resulting nanocomposite comparing with its counterpart in free state. The loading of 6-mercaptopurine (MP) in the FPEGMP-2 nanocomposite was estimated to be about 5.6% and the kinetic experimental data properly correlated with the pseudo-second order model. Also, the release of MP from the FPEGMP-2 nanocomposite shows the sustained release manner which is remarkably lower in phosphate buffered solution at pH 7.4 than pH 4.8, due to different release mechanism. The maximum percentage release of MP from the nanocomposite reached about 60% and 97% within about 92 and 74 hours when exposed to pH 7.4 and 4.8, respectively.

Release Behavior and Toxicity Profiles towards Leukemia (WEHI-3B) Cell Lines of 6-Mercaptopurine-PEG-Coated Magnetite Nanoparticles Delivery System

PubMed Central

Kura, Aminu Umar; Hussein-Al-Ali, Samer Hasan; Hussein, Mohd Zobir bin; Fakurazi, Sharida; Shaari, Abdul Halim; Ahmad, Zalinah

2014-01-01

The coating of an active drug, 6-mercaptopurine, into the iron oxide nanoparticles-polyethylene glycol (FNPs-PEG) in order to form a new nanocomposite, FPEGMP-2, was accomplished using coprecipitation technique. The resulting nanosized with a narrow size distribution magnetic polymeric particles show the superparamagnetic properties with 38.6 emu/g saturation magnetization at room temperature. Fourier transform infrared spectroscopy and the thermal analysis study supported the formation of the nanocomposite and the enhancement of thermal stability in the resulting nanocomposite comparing with its counterpart in free state. The loading of 6-mercaptopurine (MP) in the FPEGMP-2 nanocomposite was estimated to be about 5.6% and the kinetic experimental data properly correlated with the pseudo-second order model. Also, the release of MP from the FPEGMP-2 nanocomposite shows the sustained release manner which is remarkably lower in phosphate buffered solution at pH 7.4 than pH 4.8, due to different release mechanism. The maximum percentage release of MP from the nanocomposite reached about 60% and 97% within about 92 and 74 hours when exposed to pH 7.4 and 4.8, respectively. PMID:24895684

INFLUENCE OF PH AND OXIDATION-REDUCTION POTENTIAL (EH) ON THE DISSOLUTION OF MERCURY-CONTAINING MINE WASTES FROM THE SULFUR BANK MERCURY MINE

EPA Science Inventory

This study was undertaken as a part of developing treatment alternatives for waste materials, primarily waste rock and roaster tailings, from sites contaminated with mercury (Hg) mining wastes. Leaching profiles of waste rock over a range of different pH and oxidation-reduction (...

Functional characterisation of a tropine-forming reductase gene from Brugmansia arborea, a woody plant species producing tropane alkaloids.

PubMed

Qiang, Wei; Xia, Ke; Zhang, Qiaozhuo; Zeng, Junlan; Huang, Yuanshe; Yang, Chunxian; Chen, Min; Liu, Xiaoqiang; Lan, Xiaozhong; Liao, Zhihua

2016-07-01

Brugmansia arborea is a woody plant species that produces tropane alkaloids (TAs). The gene encoding tropine-forming reductase or tropinone reductase I (BaTRI) in this plant species was functionally characterised. The full-length cDNA of BaTRI encoded a 272-amino-acid polypeptide that was highly similar to tropinone reductase I from TAs-producing herbal plant species. The purified 29kDa recombinant BaTRI exhibited maximum reduction activity at pH 6.8-8.0 when tropinone was used as substrate; it also exhibited maximum oxidation activity at pH 9.6 when tropine was used as substrate. The Km, Vmax and Kcat values of BaTRI for tropinone were 2.65mM, 88.3nkatmg(-1) and 2.93S(-1), respectively, at pH 6.4; the Km, Vmax and Kcat values of TRI from Datura stramonium (DsTRI) for tropinone were respectively 4.18mM, 81.20nkatmg(-1) and 2.40S(-1) at pH 6.4. At pH 6.4, 6.8 and 7.0, BaTRI had a significantly higher activity than DsTRI. Analogues of tropinone, 4-methylcyclohexanone and 3-quinuclidinone hydrochloride, were also used to investigate the enzymatic kinetics of BaTRI. The Km, Vmax and Kcat values of BaTRI for tropine were 0.56mM, 171.62nkat.mg(-1) and 5.69S(-1), respectively, at pH 9.6; the Km, Vmax and Kcat values of DsTRI for tropine were 0.34mM, 111.90nkatmg(-1) and 3.30S(-1), respectively, at pH 9.6. The tissue profiles of BaTRI differed from those in TAs-producing herbal plant species. BaTRI was expressed in all examined organs but was most abundant in secondary roots. Finally, tropane alkaloids, including hyoscyamine, anisodamine and scopolamine, were detected in various organs of B. arborea by HPLC. Interestingly, scopolamine constituted most of the tropane alkaloids content in B. arborea, which suggests that B. arborea is a scopolamine-rich plant species. The scopolamine content was much higher in the leaves and stems than in other organs. The gene expression and TAs accumulation suggest that the biosynthesis of hyoscyamine, especially scopolamine, occurred not only in the roots but also in the aerial parts of B. arborea. Copyright © 2016 Elsevier Ltd. All rights reserved.

Glycolytic potential and activity of adenosine monophosphate kinase (AMPK), glycogen phosphorylase (GP) and glycogen debranching enzyme (GDE) in steer carcasses with normal (<5.8) or high (>5.9) 24h pH determined in M. longissimus dorsi.

PubMed

Apaoblaza, A; Galaz, A; Strobel, P; Ramírez-Reveco, A; Jeréz-Timaure, N; Gallo, C

2015-03-01

Muscle glycogen concentration (MGC) and lactate (LA), activity of glycogen debranching enzyme (GDE), glycogen phosphorylase (GP) and adenosine monophosphate kinase (AMPK) were determined at 0.5h (T0) and 24h (T24) post-mortem in Longissimus dorsi samples from 38 steers that produced high pH (>5.9) and normal pH (<5.8) carcasses at 24h postmortem. MGC, LA and glycolytic potential were higher (P<0.05) in normal pH carcasses. GDE activity was similar (P>0.05) in both pH categories. GP activity increased between T0 and T24 only in normal pH carcasses. AMPK activity was four times higher in normal pH v/s high pH carcasses, without changing its activity over time. Results reinforce the idea that differences in postmortem glycogenolytic/glycolytic flow in L. dorsi of steers showing normal v/s high muscle pH at 24h, could be explained not only by the higher initial MGC in normal pH carcasses, but also by a high and sustained activity of AMPK and an increased GP activity at 24h postmortem. Copyright © 2014 Elsevier Ltd. All rights reserved.

Tight Coupling of Astrocyte pH Dynamics to Epileptiform Activity Revealed by Genetically Encoded pH Sensors.

PubMed

Raimondo, Joseph V; Tomes, Hayley; Irkle, Agnese; Kay, Louise; Kellaway, Lauriston; Markram, Henry; Millar, Robert P; Akerman, Colin J

2016-06-29

Astrocytes can both sense and shape the evolution of neuronal network activity and are known to possess unique ion regulatory mechanisms. Here we explore the relationship between astrocytic intracellular pH dynamics and the synchronous network activity that occurs during seizure-like activity. By combining confocal and two-photon imaging of genetically encoded pH reporters with simultaneous electrophysiological recordings, we perform pH measurements in defined cell populations and relate these to ongoing network activity. This approach reveals marked differences in the intracellular pH dynamics between hippocampal astrocytes and neighboring pyramidal neurons in rodent in vitro models of epilepsy. With three different genetically encoded pH reporters, astrocytes are observed to alkalinize during epileptiform activity, whereas neurons are observed to acidify. In addition to the direction of pH change, the kinetics of epileptiform-associated intracellular pH transients are found to differ between the two cell types, with astrocytes displaying significantly more rapid changes in pH. The astrocytic alkalinization is shown to be highly correlated with astrocytic membrane potential changes during seizure-like events and mediated by an electrogenic Na(+)/HCO3 (-) cotransporter. Finally, comparisons across different cell-pair combinations reveal that astrocytic pH dynamics are more closely related to network activity than are neuronal pH dynamics. This work demonstrates that astrocytes exhibit distinct pH dynamics during periods of epileptiform activity, which has relevance to multiple processes including neurometabolic coupling and the control of network excitability. Dynamic changes in intracellular ion concentrations are central to the initiation and progression of epileptic seizures. However, it is not known how changes in intracellular H(+) concentration (ie, pH) differ between different cell types during seizures. Using recently developed pH-sensitive proteins, we demonstrate that astrocytes undergo rapid alkalinization during periods of seizure-like activity, which is in stark contrast to the acidification that occurs in neighboring neurons. Rapid astrocytic pH changes are highly temporally correlated with seizure activity, are mediated by an electrogenic Na(+)/HCO3- cotransporter, and are more tightly coupled to network activity than are neuronal pH changes. As pH has profound effects on signaling in the nervous system, this work has implications for our understanding of seizure dynamics. Copyright © 2016 the authors 0270-6474/16/367002-12$15.00/0.

A novel assay for monoacylglycerol hydrolysis suitable for high-throughput screening.

PubMed

Brengdahl, Johan; Fowler, Christopher J

2006-12-01

A simple assay for monoacylglycerol hydrolysis suitable for high-throughput screening is described. The assay uses [(3)H]2-oleoylglycerol as substrate, with the tritium label in the glycerol part of the molecule and the use of phenyl sepharose gel to separate the hydrolyzed product ([(3)H]glycerol) from substrate. Using cytosolic fractions derived from rat cerebella as a source of hydrolytic activity, the assay gives the appropriate pH profile and sensitivity to inhibition with compounds known to inhibit hydrolysis of this substrate. The assay could also be adapted to a 96-well plate format, using C6 cells as the source of hydrolytic activity. Thus the assay is simple and appropriate for high-throughput screening of inhibitors of monoacylglycerol hydrolysis.

Enterocin HZ produced by a wild Enterococcus faecium strain isolated from a traditional, starter-free pickled cheese.

PubMed

Yildirim, Zeliha; Bilgin, Harun; Isleroglu, Hilal; Tokatli, Kader; Sahingil, Didem; Yildirim, Metin

2014-05-01

Bacteriogenic Enterococcus faecium HZ was identified by using biochemical (Strep-API 20, API-50 CHL, fatty acid profile) and 16S rRNA analysis (99·99 %). Ent. faecium HZ was sensitive to clinically important antibiotics such as vancomycin, and did not have gelatinase and haemolysis activities. Enterocin HZ, a bacteriocin from Ent. faecium HZ, was sensitive to papain and tyripsin, but resistant to pepsin, lipase, catalase, α-amylase, organic solvents, detergents, ß-mercaptoethanol, and heat treatment (90 °C/30 min). It was biologically active at pH 2·0-9·0 and synthesised at the highest level in MRS or M17 broth at 32 or 37 °C with an inoculum amount of 0·1-0·5 % and an initial pH of 6·0-7·0. Enterocin HZ production reached maximum level at middle and late logarithmic phase and its molecular weight was ∼4·5 kDa. It was active against some Gram-positive foodborne bacteria. Ent. faecium HZ or its bacteriocin enterocin HZ is a good candidate to be studied as a food biopreservative since enterocin HZ showed strong bactericidal activity against Listeria monocytogenes in UHT milk and also Ent. faecium HZ grew very well in milk and produced enterocin HZ at maximum level.

Improvement of strain Penicillium sp. EZ-ZH190 for tannase production by induced mutation.

PubMed

Zakipour-Molkabadi, E; Hamidi-Esfahani, Z; Sahari, M A; Azizi, M H

2013-11-01

In the search for an efficient producer of tannase, Penicillium sp. EZ-ZH190 was subjected to mutagenesis using heat treatment and strain EZ-ZH290 was isolated. The maximum tannase in this mutant strain was 4.32 U/mL with an incubation period of 84 h as compared to wild strain EZ-ZH190 where the incubation period was 96 h with a maximum enzyme activity of 4.33 U/mL. Also, the Penicillium sp. EZ-ZH290 tannase had a maximum activity at 40 °C and pH 5.5. Then, the spores of strain EZ-ZH290 were subjected to γ irradiation mutagenesis and strain EZ-ZH390 was isolated. Strain EZ-ZH390 exhibited higher tannase activity (7.66 U/mL) than the parent strain EZ-ZH290. It was also found that Penicillium sp. EZ-ZH390 tannase had an optimum activity at 35 °C and a broad pH profile with an optimum at pH 5.5. The tannase pH stability of Penicillium sp. EZ-ZH390 and its maximum production of tannase followed the same trend for five generations confirming the occurrence of stable mutant. This paper is shown that γ irradiation can mutate the Penicillium sp. leading to increase the tannase production.

From Amorphous to Defined: Balancing the Risks of Spiral Development

DTIC Science & Technology

2007-04-30

630 675 720 765 810 855 900 Time (Week) Work started and active PhIt [Requirements,Iter1] : JavelinCalibration work packages1 1 1 Work started and...active PhIt [Technology,Iter1] : JavelinCalibration work packages2 2 2 Work started and active PhIt [Design,Iter1] : JavelinCalibration work packages3 3 3 3...Work started and active PhIt [Manufacturing,Iter1] : JavelinCalibration work packages4 4 Work started and active PhIt [Use,Iter1] : JavelinCalibration

Molecular Determinants for Functional Differences between Alanine-Serine-Cysteine Transporter 1 and Other Glutamate Transporter Family Members*

PubMed Central

Scopelliti, Amanda J.; Ryan, Renae M.; Vandenberg, Robert J.

2013-01-01

The ASCTs (alanine, serine, and cysteine transporters) belong to the solute carrier family 1 (SLC1), which also includes the human glutamate transporters (excitatory amino acid transporters, EAATs) and the prokaryotic aspartate transporter GltPh. Despite the high degree of amino acid sequence identity between family members, ASCTs function quite differently from the EAATs and GltPh. The aim of this study was to mutate ASCT1 to generate a transporter with functional properties of the EAATs and GltPh, to further our understanding of the structural basis for the different transport mechanisms of the SLC1 family. We have identified three key residues involved in determining differences between ASCT1, the EAATs and GltPh. ASCT1 transporters containing the mutations A382T, T459R, and Q386E were expressed in Xenopus laevis oocytes, and their transport and anion channel functions were investigated. A382T and T459R altered the substrate selectivity of ASCT1 to allow the transport of acidic amino acids, particularly l-aspartate. The combination of A382T and T459R within ASCT1 generates a transporter with a similar profile to that of GltPh, with preference for l-aspartate over l-glutamate. Interestingly, the amplitude of the anion conductance activated by the acidic amino acids does not correlate with rates of transport, highlighting the distinction between these two processes. Q386E impaired the ability of ASCT1 to bind acidic amino acids at pH 5.5; however, this was reversed by the additional mutation A382T. We propose that these residues differences in TM7 and TM8 combine to determine differences in substrate selectivity between members of the SLC1 family. PMID:23393130

Effects of dietary forage and calf starter on ruminal pH and transcriptomic adaptation of the rumen epithelium in Holstein calves during the weaning transition.

PubMed

Kim, Yo-Han; Toji, Noriyuki; Kizaki, Keiichiro; Kushibiki, Shiro; Ichijo, Toshihiro; Sato, Shigeru

2016-11-01

We investigated the relationship between ruminal pH and transcriptomic adaptation of the rumen epithelium (RE) of calves fed calf starter with and without forage during the weaning transition. Holstein calves were assigned to groups fed calf starter either with forage (HAY group, n = 3) or without forage (CON group, n = 4). Ruminal pH was measured continuously, and rumen fluid and epithelium were collected 3 wk after weaning. mRNA expression profiles of the RE were examined by one-color microarray. Differentially expressed genes (DEGs) were investigated using the Ingenuity Pathway Analysis (IPA). Mean and maximum ruminal pH were significantly (P < 0.05) higher, and the duration of pH < 5.8 during 1 day was significantly (P < 0.05) shorter, in the HAY group. The proportion of ruminal acetate and the acetate-to-propionate ratio were significantly (P < 0.05) lower in the CON group. DEGs encoding transcription regulators (SREBP1), insulin-like growth factor binding proteins (IGFBP7 and CTGF), ketogenic enzymes (HMGCL, BDH1, and BDH2), and a transporter (SLC16A3) were identified (P < 0.05) between the two groups. A growth factor (TGFB1) and signaling pathway (EGF and EGFR) were activated as upstream regulators. These results suggest that dietary forage alleviates ruminal acidosis, and the decrease in ruminal pH may damage the RE, leading to changes in gene expression to repair the damage. Furthermore, rumen development may be regulated by growth factor (TGFB1) and signaling pathways (EGF and IGFBP) for adaptation to feeding on calf starter with and without forage during the weaning transition. Copyright © 2016 the American Physiological Society.

A pH dependent Raman and surface enhanced Raman spectroscopic studies of citrazinic acid aided by theoretical calculations

NASA Astrophysics Data System (ADS)

Sarkar, Sougata; Chowdhury, Joydeep; Dutta, Soumen; Pal, Tarasankar

2016-12-01

A pH dependent normal Raman scattering (NRS) and surface enhanced Raman scattering (SERS) spectral patterns of citrazinic acid (CZA), a biologically important molecule, have been investigated. The acid, with different pKa values ( 4 and 11) for the two different functional groups (-COOH and -OH groups), shows interesting range of color changes (yellow at pH 14 and brown at pH 2) with the variation in solution pH. Thus, depending upon the pH of the medium, CZA molecule can exist in various protonated and/or deprotonated forms. Here we have prescribed the existence different possible forms of CZA at different pH (Forms ;C;, ;H; and ;Dprot; at pH 14 and Forms ;A;, ;D;, and ;P; at pH 2 respectively). The NRS spectra of these solutions and their respective SERS spectra over gold nanoparticles were recorded. The spectra clearly differ in their spectral profiles. For example the SERS spectra recorded with the CZA solution at pH 2 shows blue shift for different bands compared to its NRS window e.g. 406 to 450 cm- 1, 616 to 632 cm- 1, 1332 to 1343 cm- 1 etc. Again, the most enhanced peak at 1548 cm- 1 in NRS while in the SERS window this appears at 1580 cm- 1. Similar observation was also made for CZA at pH 14. For example, the 423 cm- 1 band in the NRS profile experience a blue shift and appears at 447 cm- 1 in the SERS spectrum as well as other bands at 850, 1067 and 1214 cm- 1 in the SERS window are markedly enhanced. It is also worth noting that the SERS spectra at the different pH also differ from each other. These spectral differences indicate the existence of various adsorptive forms of the CZA molecule depending upon the pH of the solution. Therefore based on the experimental findings we propose different possible molecular forms of CZA at different pH (acidic and alkaline) conditions. For example forms 'A', 'D' and 'P' existing in acidic pH (pH 2) and three other deprotonated forms 'C', 'H' and 'Dprot' in alkaline pH (pH 14). The DFT calculations for these prescribed model systems were also carried out to have a plausible understanding of their equilibrium geometries and the vibrational wavenumbers. An idea about the molecular orientation of the adsorbate over nanocolloidal gold substrate is also documented.

A pH dependent Raman and surface enhanced Raman spectroscopic studies of citrazinic acid aided by theoretical calculations.

PubMed

Sarkar, Sougata; Chowdhury, Joydeep; Dutta, Soumen; Pal, Tarasankar

2016-12-05

A pH dependent normal Raman scattering (NRS) and surface enhanced Raman scattering (SERS) spectral patterns of citrazinic acid (CZA), a biologically important molecule, have been investigated. The acid, with different pKa values (~4 and ~11) for the two different functional groups (-COOH and -OH groups), shows interesting range of color changes (yellow at pH~14 and brown at pH~2) with the variation in solution pH. Thus, depending upon the pH of the medium, CZA molecule can exist in various protonated and/or deprotonated forms. Here we have prescribed the existence different possible forms of CZA at different pH (Forms "C", "H" and "Dprot" at pH~14 and Forms "A", "D", and "P" at pH~2 respectively). The NRS spectra of these solutions and their respective SERS spectra over gold nanoparticles were recorded. The spectra clearly differ in their spectral profiles. For example the SERS spectra recorded with the CZA solution at pH~2 shows blue shift for different bands compared to its NRS window e.g. 406 to 450cm(-1), 616 to 632cm(-1), 1332 to 1343cm(-1) etc. Again, the most enhanced peak at ~1548cm(-1) in NRS while in the SERS window this appears at ~1580cm(-1). Similar observation was also made for CZA at pH~14. For example, the 423cm(-1) band in the NRS profile experience a blue shift and appears at ~447cm(-1) in the SERS spectrum as well as other bands at ~850, ~1067 and ~1214cm(-1) in the SERS window are markedly enhanced. It is also worth noting that the SERS spectra at the different pH also differ from each other. These spectral differences indicate the existence of various adsorptive forms of the CZA molecule depending upon the pH of the solution. Therefore based on the experimental findings we propose different possible molecular forms of CZA at different pH (acidic and alkaline) conditions. For example forms 'A', 'D' and 'P' existing in acidic pH (pH~2) and three other deprotonated forms 'C', 'H' and 'Dprot' in alkaline pH (pH~14). The DFT calculations for these prescribed model systems were also carried out to have a plausible understanding of their equilibrium geometries and the vibrational wavenumbers. An idea about the molecular orientation of the adsorbate over nanocolloidal gold substrate is also documented. Copyright © 2016 Elsevier B.V. All rights reserved.

Development of Aspherical Active Gratings at NSRRC

NASA Astrophysics Data System (ADS)

Tseng, Tse-Chuan; Wang, Duan Jen; Perng, Shen-Yaw; Chen, Chien-Te; Lin, Chia-Jui; Kuan, Chien-Kuang; Ho, His-Chou; Wang, Jeremy; Fung, H. S.; Chang, Shuo-Hung

2007-01-01

An active grating based on a novel optical concept with bendable polynomial surface profile to reduce the coma and defocus aberrations had been designed and proved by the prototype testing. Due to the low glass transition temperature of the glue and the difference of thermal expansion coefficient between the 17-4 steel bender and silicon, the prototype distorted from flat polished condition when thermally de-blocked the polishing pitch. To improve the thermal deformation of the active grating in the polishing process, a new invar bender and high curing temperature glue were adapted to glue a silicon substrate on the bender. After some tests and manufacturer polishing, it showed acceptable conditions. In this paper we will present the design and preliminary tests of the invar active grating. Meanwhile, the design and analysis of a new 17-4 PH steel bender to be electro-less nickel plating and mechanical ruling for a new beamline will also be discussed.

Different Statistical Approaches to Investigate Porcine Muscle Metabolome Profiles to Highlight New Biomarkers for Pork Quality Assessment

PubMed Central

Welzenbach, Julia; Neuhoff, Christiane; Looft, Christian; Schellander, Karl; Tholen, Ernst; Große-Brinkhaus, Christine

2016-01-01

The aim of this study was to elucidate the underlying biochemical processes to identify potential key molecules of meat quality traits drip loss, pH of meat 1 h post-mortem (pH1), pH in meat 24 h post-mortem (pH24) and meat color. An untargeted metabolomics approach detected the profiles of 393 annotated and 1,600 unknown metabolites in 97 Duroc × Pietrain pigs. Despite obvious differences regarding the statistical approaches, the four applied methods, namely correlation analysis, principal component analysis, weighted network analysis (WNA) and random forest regression (RFR), revealed mainly concordant results. Our findings lead to the conclusion that meat quality traits pH1, pH24 and color are strongly influenced by processes of post-mortem energy metabolism like glycolysis and pentose phosphate pathway, whereas drip loss is significantly associated with metabolites of lipid metabolism. In case of drip loss, RFR was the most suitable method to identify reliable biomarkers and to predict the phenotype based on metabolites. On the other hand, WNA provides the best parameters to investigate the metabolite interactions and to clarify the complex molecular background of meat quality traits. In summary, it was possible to attain findings on the interaction of meat quality traits and their underlying biochemical processes. The detected key metabolites might be better indicators of meat quality especially of drip loss than the measured phenotype itself and potentially might be used as bio indicators. PMID:26919205

Nanotechnology Enabled Enhancement of Enzyme Activity and Thermostability: Study on Impaired Pectate Lyase from Attenuated Macrophomina phaseolina in Presence of Hydroxyapatite Nanoparticle

PubMed Central

Dutta, Nalok; Mukhopadhyay, Arka; Dasgupta, Anjan Kr.; Chakrabarti, Krishanu

2013-01-01

In this paper we show that hydroxyapatite nanoparticles (NP) can not only act as a chaperon (by imparting thermostability) but can serve as a synthetic enhancer of activity of an isolated extracellular pectate lyase (APL) with low native state activity. The purified enzyme (an attenuated strain of Macrophomina phaseolina) showed feeble activity at 50°C and pH 5.6. However, on addition of 10.5 µg/ml of hydroxyapatite nanoparticles (NP), APL activity increased 27.7 fold with a 51 fold increase in half-life at a temperature of 90°C as compared to untreated APL. The chaperon like activity of NP was evident from entropy–enthalpy compensation profile of APL. The upper critical temperature for such compensation was elevated from 50°C to 90°C in presence of NP. This dual role of NP in enhancing activity and conferring thermostability to a functionally impaired enzyme is reported for the first time. PMID:23691068

Relationship among Phosphorus Circulation Activity, Bacterial Biomass, pH, and Mineral Concentration in Agricultural Soil.

PubMed

Adhikari, Dinesh; Jiang, Tianyi; Kawagoe, Taiki; Kai, Takamitsu; Kubota, Kenzo; Araki, Kiwako S; Kubo, Motoki

2017-12-04

Improvement of phosphorus circulation in the soil is necessary to enhance phosphorus availability to plants. Phosphorus circulation activity is an index of soil's ability to supply soluble phosphorus from organic phosphorus in the soil solution. To understand the relationship among phosphorus circulation activity; bacterial biomass; pH; and Fe, Al, and Ca concentrations (described as mineral concentration in this paper) in agricultural soil, 232 soil samples from various agricultural fields were collected and analyzed. A weak relationship between phosphorus circulation activity and bacterial biomass was observed in all soil samples ( R ² = 0.25), and this relationship became significantly stronger at near-neutral pH (6.0-7.3; R ² = 0.67). No relationship between phosphorus circulation activity and bacterial biomass was observed at acidic (pH < 6.0) or alkaline (pH > 7.3) pH. A negative correlation between Fe and Al concentrations and phosphorus circulation activity was observed at acidic pH ( R ² = 0.72 and 0.73, respectively), as well as for Ca at alkaline pH ( R ² = 0.64). Therefore, bacterial biomass, pH, and mineral concentration should be considered together for activation of phosphorus circulation activity in the soil. A relationship model was proposed based on the effects of bacterial biomass and mineral concentration on phosphorus circulation activity. The suitable conditions of bacterial biomass, pH, and mineral concentration for phosphorus circulation activity could be estimated from the relationship model.

Regulating NETosis: Increasing pH Promotes NADPH Oxidase-Dependent NETosis.

PubMed

Khan, Meraj A; Philip, Lijy M; Cheung, Guillaume; Vadakepeedika, Shawn; Grasemann, Hartmut; Sweezey, Neil; Palaniyar, Nades

2018-01-01

Neutrophils migrating from the blood (pH 7.35-7.45) into the surrounding tissues encounter changes in extracellular pH (pH e ) conditions. Upon activation of NADPH oxidase 2 (Nox), neutrophils generate large amounts of H + ions reducing the intracellular pH (pH i ). Nevertheless, how extracellular pH regulates neutrophil extracellular trap (NET) formation (NETosis) is not clearly established. We hypothesized that increasing pH increases Nox-mediated production of reactive oxygen species (ROS) and neutrophil protease activity, stimulating NETosis. Here, we found that raising pH e (ranging from 6.6 to 7.8; every 0.2 units) increased pH i of both activated and resting neutrophils within 10-20 min (Seminaphtharhodafluor dual fluorescence measurements). Since Nox activity generates H + ions, pH i is lower in neutrophils that are activated compared to resting. We also found that higher pH stimulated Nox-dependent ROS production (R123 generation; flow cytometry, plate reader assay, and imaging) during spontaneous and phorbol myristate acetate-induced NETosis (Sytox Green assays, immunoconfocal microscopy, and quantifying NETs). In neutrophils that are activated and not resting, higher pH stimulated histone H4 cleavage (Western blots) and NETosis. Raising pH increased Escherichia coli lipopolysaccharide-, Pseudomonas aeruginosa (Gram-negative)-, and Staphylococcus aureus (Gram-positive)-induced NETosis. Thus, higher pH e promoted Nox-dependent ROS production, protease activity, and NETosis; lower pH has the opposite effect. These studies provided mechanistic steps of pH e -mediated regulation of Nox-dependent NETosis. Raising pH either by sodium bicarbonate or Tris base (clinically known as Tris hydroxymethyl aminomethane, tromethamine, or THAM) increases NETosis. Each Tris molecule can bind 3H + ions, whereas each bicarbonate HCO3 - ion binds 1H + ion. Therefore, the amount of Tris solution required to cause the same increase in pH level is less than that of equimolar bicarbonate solution. For that reason, regulating NETosis by pH with specific buffers such as THAM could be more effective than bicarbonate in managing NET-related diseases.

Soil characteristics and fallout and environmental radionuclides on different geomorphological features in Elephant Island for assessing environmental changes in maritime Antarctica

NASA Astrophysics Data System (ADS)

Navas, Ana; Serrano, Enrique; López-Martínez, Jerónimo; Gaspar, Leticia; Mink, Sandra

2013-04-01

Soils in ice-free areas of Elephant Island (South Shetland Islands, Antarctic Peninsula region) have been forming since the last deglaciation in an Antarctic maritime climate that is warmer and more humid than in interior Antarctica. The studied ice-free areas correspond mostly to coastal promontories and the underlain materials are composed of metamorphic rocks. A soil survey was carried out in the largest ice-free areas at the western coast of the island at Stinker Point and Lindsey Cape, as a part of a broader study on soils and geomorphology in maritime Antarctica. A soil sampling campaign was undertaken with the aim of characterizing soils developed on different geomorphic features and to investigate the processes involved in their development following the glacial retreat, that started in the area probably later than 4000 yr BP. Study sites have glacial deposits and raised marine surfaces and they include areas with different mosses and lichens coverage. Profiles were located at altitudes ranging from 30 to 90 m a.s.l. at Stinker Point and at 140 m a.s.l. at Lindsey Cape. A total of 8 soil profiles of variable depths were sampled at depth increment intervals of 5 cm until rock outcrop (15-30 cm). Distinctive geomorphic features have been described at the study sites to assess the extent of the relationship between soil characteristics and geomorphology. The main soil properties analysed were: pH, electrical conductivity, carbonate content, bulk density, soil texture and soil fertility indicators (organic matter and soil organic carbon content, nitrogen, available phosphorous and potassium). Analyses of stable elements and activities of fallout (FRN's) and environmental radionuclides (ERN's) were also performed in the interval samples. The studied Cryosols are stony with no clear horizon differentiation and the soil texture is mostly silty loam. The soils have in general low contents of organic matter (0.3-2.7 %), carbon (0.16 - 1.6 %) and nitrogen (< 0.33 %). Available K and P contents and N vary largely among the profiles in relation to ornithogenic activity. Carbonate contents are very low (< 1.0 %) and average electrical conductivity is 0.14 dS m-1. The pH ranges between 3.9 and 8.6 and variation from acid to alkaline profiles is related to the profile position. The major elements Al, Fe, Ca and Na, were the most abundant in that order, followed by Mg, K, Mn and then Pb, Ba and Sr whereas Cr, Zn, Li, Co, Ni and Cd are present as trace elements. In two profiles on intermediate marine platforms, the FRN's concentrate at the topsoil, where 137Cs and 210Pbex activities are 11 and 20 Bq/kg, respectively. The depth distribution of ERN's is quite homogeneous, especially for 226Ra and 232Th activities, whereas larger variations are observed for 40K and to less extent for 238U. The absence of 137Cs and depleted levels of 210Pbexin soils on till materials of moraines is likely related to the age of ice retreat but soil disturbance can not be disregarded. Cryogenic processes triggering the mechanical disintegration of bedrock by freezing-thaw cycles within the soil active layer and wetting-drying are main processes involved in soil development in Elephant Island. This research provides information on past environmental changes of interest to understand the soil response to actual changes.

Effects of starch content of calf starter on growth and rumen pH in Holstein calves during the weaning transition.

PubMed

Laarman, A H; Sugino, T; Oba, M

2012-08-01

The objective of this study was to evaluate the effects of substituting high fiber byproducts for dry ground corn in calf starter on growth and rumen pH during the weaning transition. Holstein bull calves were raised on an intensified nursing program using milk replacer containing 26% CP and 18% fat. Calves were fed a texturized calf starter containing either dry ground corn at 18.8% of dry matter (DM; CRN), beet pulp replacing dry ground corn at 10.2% dietary DM (BP), or triticale dried distillers grains with solubles replacing dry ground corn and high-protein feedstuffs at 18.6% of dietary DM (DDGS) in the pellet; treatment calf starters differed only in the pellet portion. Starch concentrations of CRN, BP, and DDGS were 35.3, 33.4, and 31.4%, respectively. After a calf consumed 2.50 kg of starter for 3 consecutive days, a small ruminant rumen pH data logger was inserted orally and rumen pH was measured continuously for 4d. Calves were then killed and rumen fluid was sampled to determine volatile fatty acid profile. No difference was found in overall average daily gain or growth rates of hip height, withers height, and heart girth. During the weaning transition, rate of increase in calf starter intake was greater for calves fed DDGS compared with those fed CRN (87.7 vs. 77.5 g/d), but lower for calves fed BP compared with CRN (68.1 vs. 77.5 g/d). The area under pH 5.8 (470 vs. 295 min × pH/d) or pH 5.2 (72.7 vs. 16.4 min × pH/d) was greater for calves fed DDGS than those fed CRN. Rumen pH profile was not affected by BP treatment compared with CRN, but calves fed BP tended to have greater water intake than those fed CRN (6.6 vs. 5.8 L/d). Volatile fatty acid profile was not affected by treatment with the exception of molar proportion of butyrate, which tended to be lower for calves fed BP compared with those fed CRN (15.0 vs. 16.6%). Hay intake was positively correlated to mean rumen pH for calves used in this study (r=0.48). Decreasing dietary starch concentration did not mitigate rumen acidosis in calves during weaning transition, and low rumen pH did not adversely affect growth during the weaning transition. Copyright © 2012 American Dairy Science Association. Published by Elsevier Inc. All rights reserved.

Cloning and identification of Group 1 mrp operon encoding a novel monovalent cation/proton antiporter system from the moderate halophile Halomonas zhaodongensis.

PubMed

Meng, Lin; Hong, Shan; Liu, Henan; Huang, Haipeng; Sun, Hao; Xu, Tong; Jiang, Juquan

2014-11-01

The novel species Halomonas zhaodongensis NEAU-ST10-25(T) recently identified by our group is a moderate halophile which can grow at the range of 0-2.5 M NaCl (optimum 0.5 M) and pH 6-12 (optimum pH 9). To explore its halo-alkaline tolerant mechanism, genomic DNA was screened from NEAU-ST10-25(T) in this study for Na(+)(Li(+))/H(+) antiporter genes by selection in Escherichia coli KNabc lacking three major Na(+)(Li(+))/H(+) antiporters. One mrp operon could confer tolerance of E. coli KNabc to 0.8 M NaCl and 100 mM LiCl, and an alkaline pH. This operon was previously mainly designated mrp (also mnh, pha or sha) due to its multiple resistance and pH-related activity. Here, we will also use mrp to designate the homolog from H. zhaodongensis (Hz_mrp). Sequence analysis and protein alignment showed that Hz_mrp should belong to Group 1 mrp operons. Further phylogenetic analysis reveals that Hz_Mrp system should represent a novel sub-class of Group 1 Mrp systems. This was confirmed by a significant difference in pH-dependent activity profile or the specificity and affinity for the transported monovalent cations between Hz_Mrp system and all the known Mrp systems. Therefore, we propose that Hz_Mrp should be categorized as a novel Group 1 Mrp system.

Purification and biochemical characterization of a thermostable and acid-stable alpha-amylase from Bacillus licheniformis B4-423.

PubMed

Wu, Xiangrong; Wang, Yuxia; Tong, Bending; Chen, Xianghua; Chen, Jianhua

2018-04-01

Novel thermostable amylase need to be continuously explored with the improvement of industrial requirements. A new acidophilic and thermostable amylase producing bacterium isolated from spring was identified as Bacillus strain on the basis of 16S rDNA. The amylase was purified by ammonium sulphate precipitation, gel chromatography and anion exchange chromatography. SDS-PAGE revealed that the enzyme was monomeric with a molecular weight of 58 kDa. The amylase exhibited optimal activity at pH 5.0 and temperature 100 °C. Then the enzyme showed high stability in pH ranges 4.0-10.0 and more than 90% of maximal activity was found from 20 °C to 80 °C. Apart from good stability toward SDS and non-ionic detergent, the purified enzyme exhibited high compatibility with some inhibitors such as urea and EDTA. The results demonstrated the stability of the enzyme in different organic solvents. Moreover, we determined the amylase gene, compared the structure with α-amylase BAA and BLA and found some thermostability determinants in our enzyme. Overall, presenting various properties were including high thermostability, Ca 2+ -independency, broad temperature and pH profiles, organic-solvent tolerance as well as excellent stability with detergents. Such characteristics have not been reported for this type of enzyme, and the α-amylase will be a suitable candidate in industrial fields. Copyright © 2017 Elsevier B.V. All rights reserved.

Hydrogel coated monoliths for enzymatic hydrolysis of penicillin G

PubMed Central

Smeltink, M. W.; Straathof, A. J. J.; Paasman, M. A.; van de Sandt, E. J. A. X.; Kapteijn, F.; Moulijn, J. A.

2008-01-01

The objective of this work was to develop a hydrogel-coated monolith for the entrapment of penicillin G acylase (E. coli, PGA). After screening of different hydrogels, chitosan was chosen as the carrier material for the preparation of monolithic biocatalysts. This protocol leads to active immobilized biocatalysts for the enzymatic hydrolysis of penicillin G (PenG). The monolithic biocatalyst was tested in a monolith loop reactor (MLR) and compared with conventional reactor systems using free PGA, and a commercially available immobilized PGA. The optimal immobilization protocol was found to be 5 g l−1 PGA, 1% chitosan, 1.1% glutaraldehyde and pH 7. Final PGA loading on glass plates was 29 mg ml−1 gel. For 400 cpsi monoliths, the final PGA loading on functionalized monoliths was 36 mg ml−1 gel. The observed volumetric reaction rate in the MLR was 0.79 mol s−1 m−3monolith. Apart from an initial drop in activity due to wash out of PGA at higher ionic strength, no decrease in activity was observed after five subsequent activity test runs. The storage stability of the biocatalysts is at least a month without loss of activity. Although the monolithic biocatalyst as used in the MLR is still outperformed by the current industrial catalyst (immobilized preparation of PGA, 4.5 mol s−1 m−3catalyst), the rate per gel volume is slightly higher for monolithic catalysts. Good activity and improved mechanical strength make the monolithic bioreactor an interesting alternative that deserves further investigation for this application. Although moderate internal diffusion limitations have been observed inside the gel beads and in the gel layer on the monolith channel, this is not the main reason for the large differences in reactor performance that were observed. The pH drop over the reactor as a result of the chosen method for pH control results in a decreased performance of both the MLR and the packed bed reactor compared to the batch system. A different reactor configuration including an optimal pH profile is required to increase the reactor performance. The monolithic stirrer reactor would be an interesting alternative to improve the performance of the monolith-PGA combination. PMID:18427849

Distant electric coupling between nitrate reduction and sulphide oxidation investigated by an improved nitrate microscale biosensor

NASA Astrophysics Data System (ADS)

Marzocchi, U.; Revsbech, N. P.; Nielsen, L. P.; Risgaard-Petersen, N.

2012-04-01

Bacteria are apparently able to transmit electrons to other bacteria (Summers et al. 2010) or to electrodes (Malvankar et al. 2011) by some kind of nanowires (Reguera et al. 2005, Gorbi et al. 2006). Lately it has been shown that such transfer may occur over distances of centimetres in sediments, thereby coupling sulphide oxidation in deeper layers with oxygen reduction near the surface (Nielsen 2011). The finding of these long-distance electrical connections originated from analysis of O2, H2S, and pH profiles measured with microsensors. Nitrate is thermodynamically almost as good an electron acceptor as O2, and we therefore set up an experiment to investigate whether long-distance electron transfer also happens with NO3-. Aquaria were filled with sulphidic marine sediment from Aarhus Bay that was previously used to show long-distance electron transfer to O2. The aquaria were equipped with a lid so that they could be completely filled without a gas phase. Anoxic seawater with 300 μM NO3- was supplied at a constant rate resulting in a steady state concentration in the aquatic phase of 250 μM NO3-. The reservoir with the nitrate-containing water was kept anoxic by bubbling it with a N2/CO2 mixture and was kept at an elevated temperature. The water was cooled on the way to the aquaria to keep the water in the aquaria undersaturated with gasses, so that bubble formation by denitrification in the sediment could be minimised. Profiles of NO3-, H2S, and pH were measured as a function of time (2 months) applying commercial sensors for H2S and pH and an improved microscale NO3- biosensor developed in our laboratory. The penetration of NO3- in the sediment was 4-5 mm after 2 months, whereas sulphide only could be detected below 8-9 mm depth. The electron acceptor and electron donor were thus separated by 4-5 mm, indicating long distance electron transfer. A pH maximum of about 8.6 pH units at the NO3- reduction zone similar to a pH maximum observed in the O2 reduction zone of electro-active sediments could be observed. This pH maximum was the strongest evidence for long-distance electron transfer in oxic sediments, but cannot be taken as proof in denitrifying sediments as conventional denitrification may also produce elevated pH. We are now searching for the NO3- reducing bacteria that may be active in long-distance electron transfer in our sediment. Gorby, Y. A., S. Yanina, et al. (2006). Electrically conductive bacterial nanowires produced by Shewanella oneidensis strain MR-1 and other microorganisms. Proceedings of the National Academy of Sciences of the United States of America 103(30): 11358-11363. Malvankar, N. S., M. Vargas, et al. (2011). Tunable metallic-like conductivity in microbial nanowire networks. Nature Nanotechnology 6(9): 573-579. Nielsen, L. P., N. Risgaard-Petersen, et al. (2010). Electric currents couple spatially separated biogeochemical processes in marine sediment. Nature 463(7284): 1071-1074. Reguera, G., K. D. McCarthy, et al. (2005). Extracellular electron transfer via microbial nanowires. Nature 435(7045): 1098-1101. Summers, Z. M., H. E. Fogarty, et al. (2010). Direct Exchange of Electrons Within Aggregates of an Evolved Syntrophic Coculture of Anaerobic Bacteria. Science 330(6009): 1413-1415.

Nanocomposites coated with xyloglucan for drug delivery: In vitro studies.

PubMed

Ribeiro, C; Arizaga, G G C; Wypych, F; Sierakowski, M-R

2009-02-09

Enalaprilate (Enal), an active pharmaceutical component, was intercalated into a layered double hydroxide (Mg/Al-LDH) by an ion exchange reaction. The use of a layered double hydroxide (LDH) to release active drugs is limited by the low pH of the stomach (pH approximately 1.2), in whose condition it is readily dissolved. To overcome this limitation, xyloglucan (XG) extracted from Hymenaea courbaril (jatobá) seeds, Brazilian species, was used to protect the LDH and allow the drug to pass through the gastrointestinal tract. All the materials were characterized by X-ray diffraction, Fourier transform infrared spectroscopy, elemental analyses, transmission electronic microscopy, thermal analyses, and a kinetic study of the in vitro release was monitored by ultraviolet spectroscopy. The resulting hybrid system containing HDL-Enal-XG(3) slowly released the Enal. In an 8-h of test, the system protected 40% (w/v) of the drug. The kinetic profile showed that the drug release was a co-effect behavior, involving dissolution of inorganic material and ion exchange between the intercalated anions in the lamella and those of phosphate in the buffer solution. The nanocomposite coated protection with XG was therefore efficient in obtaining a slow release of Enal.

Glycation of human erythrocyte glutathione peroxidase: effect on the physical and kinetic properties.

PubMed

Suravajjala, Sreekanth; Cohenford, Menashi; Frost, Leslie R; Pampati, Praveen K; Dain, Joel A

2013-06-05

Glutathione peroxidase (GPx) is a significant antioxidant enzyme that plays a key role in protecting the body from reactive oxygen species (ROS) and their toxicity. As a biocatalyst, the enzyme has been shown to reduce hydrogen peroxide to water and lipid hydroperoxides to their respective alcohols. The increased levels of ROS in patients with diabetes have been speculated to arise, in part, from alterations in the activity of glutathione antioxidant enzymes, perhaps, by mechanisms such as the glycation of the protein, in vivo. Under physiological conditions of temperature and pH, we investigated the susceptibility of human glutathione peroxidase to glycation, determined the effects of glycation on the physical and kinetic properties of the enzyme, and identified the protein's vulnerable amino acid sites of glycation. Circular dichroism, UV and mass spectrometry studies revealed that methylglyoxal and DL-glyceraldehyde are potent glycators of glutathione peroxidase; destabilizing its structure, altering its pH activity and stability profiles and increasing its Km value. In comparison to DL-glyceraldehyde, methylglyxol was a more potent glycator of the enzyme and was found to nonenzymatically condense with Arg-177, located near the glutathione binding site of GPx. Copyright © 2013 Elsevier B.V. All rights reserved.

Amelioration of an Ultisol profile acidity using crop straws combined with alkaline slag.

PubMed

Li, Jiu-yu; Masud, M M; Li, Zhong-yi; Xu, Ren-kou

2015-07-01

The acidity of Ultisols (pH <5) is detrimental to crop production. Technologies should be explored to promote base saturation and liming effect for amelioration of Ultisol pH. Column leaching experiments were conducted to investigate the amelioration effects of canola straw (CS) and peanut straw (PS) in single treatment and in combination whether with alkaline slag (AS) or with lime on Ultisol profile acidity. The treatment without liming materials was set as control, and the AS and lime in single treatment are set for comparison. Results indicated that all the liming materials increase soil profile pH and soil exchangeable base cations at the 0-40-cm depth, except that the lime had amelioration effect just on 0 to 15-cm profile. The amelioration effect of the liming materials on surface soil acidity was mainly dependent on the ash alkalinity in organic materials or acid neutralization capacity of inorganic materials. Specific adsorption of sulfate (SO4(2-)) or organic anions, decarboxylation of organic acids/anions, and the association of H(+) with organic anions induced a "liming effect" of crop residues and AS on subsoil acidity. Moreover, SO4(2-) and chloride (Cl(-)) in PS, CS, and AS primarily induced base cations to move downward to subsoil and exchange with exchangeable aluminum (Al(3+)) and protons (H(+)). These anions also promoted the exchangeable Al to leach out of the soil profile. The CS was more effective than PS in decreasing soil acidity in the subsoil, which mainly resulted from higher sulfur (S) and Cl content in CS compared to PS. The CS combined with AS was the better amendment choice in practical agricultural systems.

Too Little Too Soon? Modeling the Risks of Spiral Development

DTIC Science & Technology

2007-04-30

270 315 360 405 450 495 540 585 630 675 720 765 810 855 900 Time (Week) Work started and active PhIt [Requirements,Iter1] : JavelinCalibration work...packages1 1 1 Work started and active PhIt [Technology,Iter1] : JavelinCalibration work packages2 2 2 Work started and active PhIt [Design,Iter1...JavelinCalibration work packages3 3 3 3 Work started and active PhIt [Manufacturing,Iter1] : JavelinCalibration work packages4 4 Work started and active PhIt

Effect of pH and nitrite concentration on nitrite oxidation rate.

PubMed

Jiménez, E; Giménez, J B; Ruano, M V; Ferrer, J; Serralta, J

2011-10-01

The effect of pH and nitrite concentration on the activity of the nitrite oxidizing bacteria (NOB) in an activated sludge reactor has been determined by means of laboratory batch experiments based on respirometric techniques. The bacterial activity was measured at different pH and at different total nitrite concentrations (TNO₂). The experimental results showed that the nitrite oxidation rate (NOR) depends on the TNO₂ concentration independently of the free nitrous acid (FNA) concentration, so FNA cannot be considered as the real substrate for NOB. NOB were strongly affected by low pH values (no activity was detected at pH 6.5) but no inhibition was observed at high pH values (activity was nearly the same for the pH range 7.5-9.95). A kinetic expression for nitrite oxidation process including switch functions to model the effect of TNO₂ concentration and pH inhibition is proposed. Substrate half saturation constant and pH inhibition constants have been obtained. Copyright © 2011 Elsevier Ltd. All rights reserved.

The biochemical characterization of three imine-reducing enzymes from Streptosporangium roseum DSM43021, Streptomyces turgidiscabies and Paenibacillus elgii.

PubMed

Scheller, Philipp N; Nestl, Bettina M

2016-12-01

Recently imine reductases (IREDs) have emerged as promising biocatalysts for the synthesis of a wide variety of chiral amines. To promote their application, many novel enzymes were reported, but only a few of them were biochemically characterized. To expand the available knowledge about IREDs, we report the characterization of two recently identified (R)-selective IREDs from Streptosporangium roseum DSM43021 and Streptomyces turgidiscabies and one (S)-selective IRED from Paenibacillus elgii. The biochemical properties including pH profiles, temperature stabilities, and activities of the enzymes in the presence of organic solvents were investigated. All three enzymes showed relatively broad pH spectra with maximum activities in the neutral range. While the (R)-selective IREDs displayed only limited thermostabilities, the (S)-selective enzyme was found to be the most thermostable IRED known to date. The activity of this IRED proved also to be most tolerant towards the investigated co-solvents DMSO and methanol. We further studied activities and selectivities towards a panel of cyclic imine model substrates to compare these enzymes with other IREDs. In biotransformations, IREDs showed high conversions and the amine products were obtained with up to 99 % ee. By recording the kinetic constants for these compounds, substrate preferences of the IREDs were investigated and it was shown that the (S)-IRED favors the transformation of bulky imines contrary to the (R)-selective IREDs. Finally, novel exocyclic imine substrates were tested and also high activities and selectivities detected.

DOE Office of Scientific and Technical Information (OSTI.GOV)

Grissom, C.B.; Cleland, W.W.

The catalytic mechanism of porcine heart NADP isocitrate dehydrogenase has been investigated by use of the variation of deuterium and /sup 13/C kinetic isotope effects with pH. The observed /sup 13/C isotope effect on VK for isocitrate increases from 1.0038 at neutral pH to a limiting value of 1.040 at low pH. The limiting /sup 13/C isotope effect with deuteriated isocitrate at low pH is 1.016. This decrease in /sup 13/(VK/sub Ic/) upon deuteriation indicates a stepwise mechanism for the oxidation and decarboxylation of isocitrate. This predicts a deuterium isotope effect on VK of 2.9, but /sup D/(VK) at lowmore » pH only increases to a maximum of 1.08. The pK seen in the /sup 13/(VK/sub Ic/) pH profile for isocitrate if 4.5. This pK is displaced 1.2 pH units from the true pK of the acidbase functionality of 5.7 seen in the pK/sub i/ profile for oxalylglycine. From this displacement, catalysis is estimated to be 16 times faster than substrate dissociation. By use of the pH-dependent partitioning ratio of the reaction intermediate oxalosuccinate between decarboxylation to 2-ketoglutarate and reduction to isocitrate, the forward commitment to catalysis for decarboxylation was determined to be 7.3 at pH 5.4 and 3.2 at pH 5.0. This gives in intrinsic /sup 13/C isotope effect for decarboxylation of 1.050. The product of oxidative decarboxylation of 3-hydroxyisocitrate by NADP isocitrate dehydrogenase is 2-hydroxy-3-ketoglutarate. This results from enzymatic protonation of the cis-enediol intermediate at C/sub 2/ rather than C/sub 3/ (as seen with isocitrate and 3-fluoroisocitrate). 2-Hydroxy-3-ketoglutarate further decarboxylates in solution to 2-hydroxy-3-ketobutyrate, which further decarboxylates to acetol. This makes 3-hydroxyisocitrate unsuitable for /sup 13/C isotope effect studies.« less

Different rate-limiting activities of intracellular pH regulators for HCO3- secretion stimulated by forskolin and carbachol in rat parotid intralobular ducts.

PubMed

Ueno, Kaori; Hirono, Chikara; Kitagawa, Michinori; Shiba, Yoshiki; Sugita, Makoto

2016-11-01

Intracellular pH (pH i ) regulation fundamentally participates in maintaining HCO 3 - release from HCO 3 - -secreting epithelia. We used parotid intralobular ducts loaded with BCECF to investigate the contributions of a carbonic anhydrase (CA), anion channels and a Na + -H + exchanger (NHE) to pH i regulation for HCO 3 - secretion by cAMP and Ca 2+ signals. Resting pH i was dispersed between 7.4 and 7.9. Forskolin consistently decreased pH i showing the dominance of pH i -lowering activities, but carbachol gathered pH i around 7.6. CA inhibition suppressed the forskolin-induced decrease in pH i , while it allowed carbachol to consistently increase pH i by revealing that carbachol prominently activated NHE via Ca 2+ -calmodulin. Under NHE inhibition, forskolin and carbachol induced the remarkable decreases in pH i , which were slowed predominantly by CA inhibition and by CA or anion channel inhibition, respectively. Our results suggest that forskolin and carbachol primarily activate the pH i -lowering CA and pH i -raising NHE, respectively, to regulate pH i for HCO 3 - secretion.

Identification of novel candidate genes involved in mineralization of dental enamel by genome-wide transcript profiling.

PubMed

Lacruz, Rodrigo S; Smith, Charles E; Bringas, Pablo; Chen, Yi-Bu; Smith, Susan M; Snead, Malcolm L; Kurtz, Ira; Hacia, Joseph G; Hubbard, Michael J; Paine, Michael L

2012-05-01

The gene repertoire regulating vertebrate biomineralization is poorly understood. Dental enamel, the most highly mineralized tissue in mammals, differs from other calcifying systems in that the formative cells (ameloblasts) lack remodeling activity and largely degrade and resorb the initial extracellular matrix. Enamel mineralization requires that ameloblasts undergo a profound functional switch from matrix-secreting to maturational (calcium transport, protein resorption) roles as mineralization progresses. During the maturation stage, extracellular pH decreases markedly, placing high demands on ameloblasts to regulate acidic environments present around the growing hydroxyapatite crystals. To identify the genetic events driving enamel mineralization, we conducted genome-wide transcript profiling of the developing enamel organ from rat incisors and highlight over 300 genes differentially expressed during maturation. Using multiple bioinformatics analyses, we identified groups of maturation-associated genes whose functions are linked to key mineralization processes including pH regulation, calcium handling, and matrix turnover. Subsequent qPCR and Western blot analyses revealed that a number of solute carrier (SLC) gene family members were up-regulated during maturation, including the novel protein Slc24a4 involved in calcium handling as well as other proteins of similar function (Stim1). By providing the first global overview of the cellular machinery required for enamel maturation, this study provide a strong foundation for improving basic understanding of biomineralization and its practical applications in healthcare. Copyright © 2011 Wiley Periodicals, Inc.

Bacterial diversity in a nonsaline alkaline environment: heterotrophic aerobic populations.

PubMed

Tiago, Igor; Chung, Ana Paula; Veríssimo, António

2004-12-01

Heterotrophic populations were isolated and characterized from an alkaline groundwater environment generated by active serpentinization, which results in a Ca(OH)2-enriched, extremely diluted groundwater with pH 11.4. One hundred eighty-five strains were isolated in different media at different pH values during two sampling periods. To assess the degree of diversity present in the environment and to select representative strains for further characterization of the populations, we screened the isolates by using random amplified polymorphic DNA-PCR profiles and grouped them based on similarities determined by fatty acid methyl ester analysis. Phenotypic characterization, determinations of G+C content, phylogenetic analyses by direct sequencing of 16S rRNA genes, and determinations of pH tolerance were performed with the selected isolates. Although 38 different populations were identified and characterized, the vast majority of the isolates were gram positive with high G+C contents and were affiliated with three distinct groups, namely, strains closely related to the species Dietzia natrolimnae (32% of the isolates), to Frigoribacterium/Clavibacter lineages (29% of the isolates), and to the type strain of Microbacterium kitamiense (20% of the isolates). Other isolates were phylogenetically related to strains of the genera Agrococcus, Leifsonia, Kytococcus, Janibacter, Kocuria, Rothia, Nesterenkonia, Citrococcus, Micrococcus, Actinomyces, Rhodococcus, Bacillus, and Staphylococcus. Only five isolates were gram negative: one was related to the Sphingobacteria lineage and the other four were related to the alpha-Proteobacteria lineage. Despite the pH of the environment, the vast majority of the populations were alkali tolerant, and only two strains were able to grow at pH 11.

Constant pH Accelerated Molecular Dynamics Investigation of the pH Regulation Mechanism of Dinoflagellate Luciferase.

PubMed

Donnan, Patrick H; Ngo, Phong D; Mansoorabadi, Steven O

2018-01-23

The bioluminescence reaction in dinoflagellates involves the oxidation of an open-chain tetrapyrrole by the enzyme dinoflagellate luciferase (LCF). The activity of LCF is tightly regulated by pH, where the enzyme is essentially inactive at pH ∼8 and optimally active at pH ∼6. Little is known about the mechanism of LCF or the structure of the active form of the enzyme, although it has been proposed that several intramolecularly conserved histidine residues in the N-terminal region are important for the pH regulation mechanism. Here, constant pH accelerated molecular dynamics was employed to gain insight into the conformational activation of LCF induced by acidification.

H2-broadening, shifting and mixing coefficients of the doublets in the ν2 and ν4 bands of PH3 at room temperature

NASA Astrophysics Data System (ADS)

Salem, Jamel; Blanquet, Ghislain; Lepère, Muriel; Younes, Rached ben

2018-05-01

The broadening, shifting and mixing coefficients of the doublet spectral lines in the ν2 and ν4 bands of PH3 perturbed by H2 have been determined at room temperature. Indeed, the collisional spectroscopic parameters: intensities, line widths, line shifts and line mixing parameters, are all grouped together in the collisional relaxation matrix. To analyse the collisional process and physical effects on spectra of phosphine (PH3), we have used the measurements carried out using a tunable diode-laser spectrometer in the ν2 and ν4 bands of PH3 perturbed by hydrogen (H2) at room temperature. The recorded spectra are fitted by the Voigt profile and the speed-dependent uncorrelated hard collision model of Rautian and Sobelman. These profiles are developed in the studies of isolated lines and are modified to account for the line mixing effects in the overlapping lines. The line widths, line shifts and line mixing parameters are given for six A1 and A2 doublet lines with quantum numbers K = 3n, (n = 1, 2, …) and overlapped by collisional broadening at pressures of less than 50 mbar.

A pH Switch Regulates the Inverse Relationship between Membranolytic and Chaperone-like Activities of HSP-1/2, a Major Protein of Horse Seminal Plasma.

PubMed

Kumar, C Sudheer; Swamy, Musti J

2016-07-05

HSP-1/2, a major protein of horse seminal plasma binds to choline phospholipids present on the sperm plasma membrane and perturbs its structure by intercalating into the hydrophobic core, which results in an efflux of choline phospholipids and cholesterol, an important event in sperm capacitation. HSP-1/2 also exhibits chaperone-like activity (CLA) in vitro and protects target proteins against various kinds of stress. In the present study we show that HSP-1/2 exhibits destabilizing activity toward model supported and cell membranes. The membranolytic activity of HSP-1/2 is found to be pH dependent, with lytic activity being high at mildly acidic pH (6.0-6.5) and low at mildly basic pH (8.0-8.5). Interestingly, the CLA is also found to be pH dependent, with high activity at mildly basic pH and low activity at mildly acidic pH. Taken together the present studies demonstrate that the membranolytic and chaperone-like activities of HSP-1/2 have an inverse relationship and are regulated via a pH switch, which is reversible. The higher CLA observed at mildly basic pH could be correlated to an increase in surface hydrophobicity of the protein. To the best of our knowledge, this is the first study reporting regulation of two different activities of a chaperone protein by a pH switch.

Acidic pH sensing in the bacterial cytoplasm is required for Salmonella virulence.

PubMed

Choi, Jeongjoon; Groisman, Eduardo A

2016-09-01

pH regulates gene expression, biochemical activities and cellular behaviors. A mildly acidic pH activates the master virulence regulatory system PhoP/PhoQ in the facultative intracellular pathogen Salmonella enterica serovar Typhimurium. The sensor PhoQ harbors an extracytoplasmic domain implicated in signal sensing, and a cytoplasmic domain controlling activation of the regulator PhoP. We now report that, surprisingly, a decrease in Salmonella's own cytoplasmic pH induces transcription of PhoP-activated genes even when the extracytoplasmic pH remains neutral. Amino acid substitutions in PhoQ's cytoplasmic domain hindered activation by acidic pH and attenuated virulence in mice, but did not abolish activation by low Mg(2+) or the antimicrobial peptide C18G. Conversely, removal of PhoQ's extracytoplasmic domains prevented the response to the latter PhoQ-activating signals but not to acidic pH. PhoP-dependent genes were minimally induced by acidic pH in the non-pathogenic species Salmonella bongori but were activated by low Mg(2+) and C18G as in pathogenic S. enterica. Our findings indicate that the sensor PhoQ enables S. enterica to respond to both host- and bacterial-derived signals that alter its cytoplasmic pH. © 2016 John Wiley & Sons Ltd.

Biomedical Engineering | Classification | College of Engineering & Applied

Science.gov Websites

Engineering, Biomedical Engineering(414) 229-6614wjchang@uwm.eduEng & Math Sciences 1113 profile photo Malkoc, Ph.D.Visiting Assistant ProfessorBiomedical Engineering414-229-6919malkoc@uwm.eduEng & Math Engineering / Electrical Engineering(414) 229-3327misra@uwm.eduEng & Math Sciences E-314 profile photo

A multivariate approach to the study of the composting process by means of analytical electrofocusing.

PubMed

Grigatti, Marco; Cavani, Luciano; Ciavatta, Claudio

2007-01-01

Three blends formed by: agro-industrial waste, wastewater sewage sludge, and their mixture, blended with tree pruning as bulking agent, were composted over a 3-month period. During the composting process the blends were monitored for the main physical and chemical characteristics. Electrofocusing (EF) was carried out on the extracted organic matter. The EF profiles were analyzed by principal component analysis (PCA) in order to assess the suitability of EF to evaluate the stabilisation level during the composting process. Throughout the process, the blends showed a general shifting of focused bands, from low to high pH, even though the compost origin affected the EF profiles. If the EF profile is analyzed by dividing it into pH regions, the interpretation of the results can be affected by the origin of compost. A good clustering of compost samples depending on the process time was obtained by analyzing the whole profile by PCA. Analysis of EF results with PCA represents a useful analytical technique to study the evolution and the stabilisation of composted organic matter.

Segment density profiles of polyelectrolyte brushes determined by Fourier transform ellipsometry

NASA Astrophysics Data System (ADS)

Biesalski, Markus; Rühe, Jürgen; Johannsmann, Diethelm

1999-10-01

We describe a method for the explicit determination of the segment density profile φ(z) of surface-attached polymer brushes with multiple angle of incidence null-ellipsometry. Because the refractive index contrast between the brush layer and the solvent is weak, multiple reflections are of minor influence and the ellipsometric spectrum is closely related to the Fourier transform of the refractive index profile, thereby allowing for explicit inversion of the ellipsometric data. We chose surface-attached monolayers of polymethacrylic acid (PMAA), a weak polyelectrolyte, as a model system and determined the segment density profile of this system as a function of the pH value of the surrounding medium by the Fourier method. Complementary to the Fourier analysis, fits with error functions are given as well. The brushes were prepared on the bases of high refractive index prisms with the "grafting-from" technique. In water, the brushes swell by more than a factor of 30. The swelling increases with increasing pH because of a growing fraction of dissociated acidic groups leading to a larger electrostatic repulsion.

Development of a Native Fractionation Antigen Microarray for Autoantibody Profiling in Breast Cancer

DTIC Science & Technology

2011-10-01

Antigen Microarray for Autoantibody Profiling in Breast Cancer PRINCIPAL INVESTIGATOR: Brian C.-S. Liu, Ph.D...Profiling in Breast Cancer 5b. GRANT NUMBER W81XWH-09-1-0684 5c. PROGRAM ELEMENT NUMBER 6. AUTHOR(S) 5d. PROJECT NUMBER Brian C.-S. Liu...NOTES 14. ABSTRACT The humoral response of a cancer patient may allow earlier detection of cancer than current methods allow. If so, the serum

Process-driven bacterial community dynamics are key to cured meat colour formation by coagulase-negative staphylococci via nitrate reductase or nitric oxide synthase activities.

PubMed

Sánchez Mainar, María; Leroy, Frédéric

2015-11-06

The cured colour of European raw fermented meats is usually achieved by nitrate-into-nitrite reduction by coagulase-negative staphylococci (CNS), subsequently generating nitric oxide to form the relatively stable nitrosomyoglobin pigment. The present study aimed at comparing this classical curing procedure, based on nitrate reductase activity, with a potential alternative colour formation mechanism, based on nitric oxide synthase (NOS) activity, under different acidification profiles. To this end, meat models with and without added nitrate were fermented with cultures of an acidifying strain (Lactobacillus sakei CTC 494) and either a nitrate-reducing Staphylococcus carnosus strain or a rare NOS-positive CNS strain (Staphylococcus haemolyticus G110), or by relying on the background microbiota. Satisfactory colour was obtained in the models prepared with added nitrate and S. carnosus. In the presence of nitrate but absence of added CNS, however, cured colour was only obtained when L. sakei CTC 494 was also omitted. This was ascribed to the pH dependency of the emerging CNS background microbiota, selecting for nitrate-reducing Staphylococcus equorum strains at mild acidification conditions but for Staphylococcus saprophyticus strains with poor colour formation capability when the pH decrease was more rapid. This reliance of colour formation on the composition of the background microbiota was further explored by a side experiment, demonstrating the heterogeneity in nitrate reduction of a set of 88 CNS strains from different species. Finally, in all batches prepared with S. haemolyticus G110, colour generation failed as the strain was systematically outcompeted by the background microbiota, even when imposing milder acidification profiles. Thus, when aiming at colour formation through CNS metabolism, technological processing can severely interfere with the composition and functionality of the meat-associated CNS communities, for both nitrate reductase and NOS activities. Several major bottlenecks, among which the rareness of phenotypic NOS activity in meat-compatible CNS, need to be considered, which is seriously questioning the relevance of this pathway in fermented meats. Copyright © 2015 Elsevier B.V. All rights reserved.

Smart pH- and reduction-dual-responsive folate-PEG-coated polymeric lipid vesicles for tumor-triggered targeted drug delivery

NASA Astrophysics Data System (ADS)

Wang, Sheng; Wang, Hanjie; Liu, Zhongyun; Wang, Liangliang; Wang, Xiaomin; Su, Lin; Chang, Jin

2014-06-01

To improve their therapeutic index, designed nanocarriers should preferentially accumulate in tumor tissues and then rapidly enter tumor cells to release the encapsulated drugs in a triggered manner. In this article, a new kind of a smart pH- and reduction-dual-responsive drug delivery system based on folate-PEG-coated polymeric lipid vesicles (FPPLVs) formed from amphiphilic dextran derivatives was designed and prepared successfully. PEG chains with pH-sensitive hydrazone bonds, stearyl alcohol (SA) chains with reduction-sensitive disulfide bonds and folate were connected to a dextran main chain. The newly developed FPPLVs had a nano-sized structure (~50 nm) with a PEG coating. The in vitro DOX release profiles showed that the FPPLVs achieved a triggered drug release in response to acidic pH and reducing environments due to the cleavage of hydrazone bonds and disulfide bonds. It has also been demonstrated by an in vitro cellular uptake study that the FPPLVs lose their PEG coating as well as expose the folate in acidic conditions, which allows them to efficiently enter tumor cells through ligand-receptor interactions. In vitro cytotoxicity measurements also confirmed that FPPLVs exhibited pronounced antitumor activity against HeLa cells. These results suggest that FPPLVs are promising carriers for smart antitumor drug delivery applications.To improve their therapeutic index, designed nanocarriers should preferentially accumulate in tumor tissues and then rapidly enter tumor cells to release the encapsulated drugs in a triggered manner. In this article, a new kind of a smart pH- and reduction-dual-responsive drug delivery system based on folate-PEG-coated polymeric lipid vesicles (FPPLVs) formed from amphiphilic dextran derivatives was designed and prepared successfully. PEG chains with pH-sensitive hydrazone bonds, stearyl alcohol (SA) chains with reduction-sensitive disulfide bonds and folate were connected to a dextran main chain. The newly developed FPPLVs had a nano-sized structure (~50 nm) with a PEG coating. The in vitro DOX release profiles showed that the FPPLVs achieved a triggered drug release in response to acidic pH and reducing environments due to the cleavage of hydrazone bonds and disulfide bonds. It has also been demonstrated by an in vitro cellular uptake study that the FPPLVs lose their PEG coating as well as expose the folate in acidic conditions, which allows them to efficiently enter tumor cells through ligand-receptor interactions. In vitro cytotoxicity measurements also confirmed that FPPLVs exhibited pronounced antitumor activity against HeLa cells. These results suggest that FPPLVs are promising carriers for smart antitumor drug delivery applications. Electronic supplementary information (ESI) available. See DOI: 10.1039/c4nr00843j

Mechanistic studies of the agmatine deiminase from Listeria monocytogenes.

PubMed

Soares, Charles A; Knuckley, Bryan

2016-06-01

Listeria monocytogenes is a Gram-positive food-borne pathogen that is capable of living within extreme environments (i.e. low temperatures and pH). This ability to survive in such conditions may arise, at least in part, from agmatine catabolism via the agmatine deiminase system (AgDS). This catabolic pathway utilizes an agmatine deiminase (AgD) to hydrolyse agmatine into N-carbamoylputrescine (NCP), with concomitant release of ammonia, which increases the pH, thus mitigating the ill effects of the acidic environment. Given the potential significance of this pathway for cell survival, we set out to study the catalytic mechanism of the AgD encoded by L. monocytogenes In the present paper, we describe the catalytic mechanism employed by this enzyme based on pH profiles, pKa measurements of the active site cysteine and solvent isotope effects (SIE). In addition, we report inhibition of this enzyme by two novel AgD inhibitors, i.e. N-(4-aminobutyl)-2-fluoro-ethanimidamide (ABFA) and N-(4-aminobutyl)-2-chloro-ethanimidamide (ABCA). In contrast with other orthologues, L. monocytogenes AgD does not use the reverse protonation or substrate-assisted mechanism, which requires an active site cysteine with a high pKa and has been commonly seen in other members of the guanidinium-modifying enzyme (GME) superfamily. Instead, the L. monocytogenes AgD has a low pKa cysteine in the active site leading to an alternative mechanism of catalysis. This is the first time that this mechanism has been observed in the GME superfamily and is significant because it explains why previously developed mechanism-based inactivators of AgDs are ineffective against this orthologue. © 2016 The Author(s). published by Portland Press Limited on behalf of the Biochemical Society.

Production and partial characterization of serine and metallo peptidases secreted by Aspergillus fumigatus Fresenius in submerged and solid state fermentation.

PubMed

da Silva, Ronivaldo Rodrigues; de Freitas Cabral, Tatiana Pereira; Rodrigues, André; Cabral, Hamilton

2013-01-01

Enzyme production varies in different fermentation systems. Enzyme expression in different fermentation systems yields important information for improving our understanding of enzymatic production induction. Comparative studies between solid-state fermentation (SSF) using agro-industrial waste wheat bran and submerged fermentation (SmF) using synthetic media were carried out to determinate the best parameters for peptidase production by the fungus Aspergillus fumigatus Fresen. Variables tested include: the concentration of carbon and protein nitrogen sources, the size of the inoculum, the pH of the media, temperature, and the length of the fermentation process. The best peptidase production during SSF was obtained after 96 hours using wheat bran at 30 °C with an inoculum of 1 × 10(6) spores and yielded 1500 active units (U/mL). The best peptidase production using SmF was obtained after periods of 72 and 96 hours of fermentation in media containing 0.5% and 0.25% of casein, respectively, at a pH of 6.0 and at 30 °C and yielded 40 U/mL. We also found examples of catabolite repression of peptidase production under SmF conditions. Biochemical characterization of the peptidases produced by both fermentative processes showed optimum activity at pH 8.0 and 50 °C, and also showed that their proteolytic activity is modulated by surfactants. The enzymatic inhibition profile using phenylmethylsulfonyl fluoride (PMSF) in SmF and SSF indicated that both fermentative processes produced a serine peptidase. Additionally, the inhibitory effect of the ethylene-diaminetetraacetic acid (EDTA) chelating agent on the peptidase produced by SmF indicated that this fermentative process also produced a metallopeptidase.

Pharmacokinetics and pharmacodynamics of the proton pump inhibitors.

PubMed

Shin, Jai Moo; Kim, Nayoung

2013-01-01

Proton pump inhibitor (PPI) is a prodrug which is activated by acid. Activated PPI binds covalently to the gastric H(+), K(+)-ATPase via disulfide bond. Cys813 is the primary site responsible for the inhibition of acid pump enzyme, where PPIs bind. Omeprazole was the first PPI introduced in market, followed by pantoprazole, lansoprazole and rabeprazole. Though these PPIs share the core structures benzimidazole and pyridine, their pharmacokinetics and pharmacodynamics are a little different. Several factors must be considered in understanding the pharmacodynamics of PPIs, including: accumulation of PPI in the parietal cell, the proportion of the pump enzyme located at the canaliculus, de novo synthesis of new pump enzyme, metabolism of PPI, amounts of covalent binding of PPI in the parietal cell, and the stability of PPI binding. PPIs have about 1hour of elimination half-life. Area under the plasmic concentration curve and the intragastric pH profile are very good indicators for evaluating PPI efficacy. Though CYP2C19 and CYP3A4 polymorphism are major components of PPI metabolism, the pharmacokinetics and pharmacodynamics of racemic mixture of PPIs depend on the CYP2C19 genotype status. S-omeprazole is relatively insensitive to CYP2C19, so better control of the intragastric pH is achieved. Similarly, R-lansoprazole was developed in order to increase the drug activity. Delayed-release formulation resulted in a longer duration of effective concentration of R-lansoprazole in blood, in addition to metabolic advantage. Thus, dexlansoprazole showed best control of the intragastric pH among the present PPIs. Overall, PPIs made significant progress in the management of acid-related diseases and improved health-related quality of life.

Classification of andisol soil on robusta coffee plantation in Silima Pungga - Pungga District

NASA Astrophysics Data System (ADS)

Marbun, P.; Nasution, Z.; Hanum, H.; Karim, A.

2018-02-01

The survey study aims to classify the Inceptisol soil on Robusta coffee plantation in Silima Pugga-Pungga District, from Order level to Sub Group level. The study was conducted on location of sample soil profiles which were determined based on Soil Map Unit (SMU) with the main Andisol Order, i.e. SMU 12, SMU 15 and SMU 17 of 18 existing SMU. The soil profiles were described to determine the morphological characteristics of the soil, while the physical and chemical properties were done by laboratory analysis. The soil samples were taken from each horizon in each profile and analyzed in the laboratory in the form of soil texture, bulk density, pH H2O, pH KCl, pH NaF, C-organic, exchangeable bases (Ca2+, Mg2+, K+, Na+), ZPC (zero point charge), base saturation, cation exchange capasity (CEC), P-retention, Al-Oxalate (Al-O) and Si-Oxalate (Si-O). The results showed that the classification of Andisol soil based on Soil Taxonomy only has one Sub Group namely Typic Hapludand. It is expected that the results of this study can provide information for more appropriate land management in order to increase the production of Robusta coffee plant in Silima Pungga-Pungga Sub district.

Combined effects of high pressure homogenization treatment and citral on microbiological quality of apricot juice.

PubMed

Patrignani, Francesca; Tabanelli, Giulia; Siroli, Lorenzo; Gardini, Fausto; Lanciotti, Rosalba

2013-01-01

High pressure homogenization (HPH) technique is able to significantly reduce spoilage microbiota in fruit juice. On the other hand, aroma compounds and essential oils can have a key role in the microbial stability of these products. For this reason, the aim of this work was to evaluate the combined effects of an aroma compound (citral, used at a concentration of 50 mg/l) and HPH treatments (performed at 100 MPa for 1-8 successive passes) on the inactivation dynamics of Saccharomyces cerevisiae SPA strain inoculated in apricot juices at level of about 4.5 log CFU/ml. Moreover, growth of surviving yeast cells was measured during the storage of the treated juice at 10°C and pH, water activity, viscosity and volatile molecule profile of apricot juice were studied. Since citral had been diluted in ethanol before the addition to juice, also samples with only ethanol added at the same volume used to dissolve citral were considered. The results showed that yeast cell viability decreased with the increases of passes at 100 MPa and the relationship between yeast cell loads and number of passes at 100 MPa followed a linear trend. In addition, the effect of HPH treatment can be notably potentiated throughout the presence of citral and ethanol, increasing the time necessary to reach a spoilage threshold during storage. The volatile profiles of the juices added with citral showed a substitution by yeast metabolism of this aldehyde with molecule characterized by a lower antimicrobial activity such as alcohols. The HPH treatments had also a significant effect on pH and viscosity of apricot juices while did not affect a(w). Copyright © 2012 Elsevier B.V. All rights reserved.

Regulators of Slc4 bicarbonate transporter activity

PubMed Central

Thornell, Ian M.; Bevensee, Mark O.

2015-01-01

The Slc4 family of transporters is comprised of anion exchangers (AE1-4), Na+-coupled bicarbonate transporters (NCBTs) including electrogenic Na/bicarbonate cotransporters (NBCe1 and NBCe2), electroneutral Na/bicarbonate cotransporters (NBCn1 and NBCn2), and the electroneutral Na-driven Cl-bicarbonate exchanger (NDCBE), as well as a borate transporter (BTR1). These transporters regulate intracellular pH (pHi) and contribute to steady-state pHi, but are also involved in other physiological processes including CO2 carriage by red blood cells and solute secretion/reabsorption across epithelia. Acid-base transporters function as either acid extruders or acid loaders, with the Slc4 proteins moving HCO−3 either into or out of cells. According to results from both molecular and functional studies, multiple Slc4 proteins and/or associated splice variants with similar expected effects on pHi are often found in the same tissue or cell. Such apparent redundancy is likely to be physiologically important. In addition to regulating pHi, a HCO−3 transporter contributes to a cell's ability to fine tune the intracellular regulation of the cotransported/exchanged ion(s) (e.g., Na+ or Cl−). In addition, functionally similar transporters or splice variants with different regulatory profiles will optimize pH physiology and solute transport under various conditions or within subcellular domains. Such optimization will depend on activated signaling pathways and transporter expression profiles. In this review, we will summarize and discuss both well-known and more recently identified regulators of the Slc4 proteins. Some of these regulators include traditional second messengers, lipids, binding proteins, autoregulatory domains, and less conventional regulators. The material presented will provide insight into the diversity and physiological significance of multiple members within the Slc4 gene family. PMID:26124722

Regulators of Slc4 bicarbonate transporter activity.

PubMed

Thornell, Ian M; Bevensee, Mark O

2015-01-01

The Slc4 family of transporters is comprised of anion exchangers (AE1-4), Na(+)-coupled bicarbonate transporters (NCBTs) including electrogenic Na/bicarbonate cotransporters (NBCe1 and NBCe2), electroneutral Na/bicarbonate cotransporters (NBCn1 and NBCn2), and the electroneutral Na-driven Cl-bicarbonate exchanger (NDCBE), as well as a borate transporter (BTR1). These transporters regulate intracellular pH (pHi) and contribute to steady-state pHi, but are also involved in other physiological processes including CO2 carriage by red blood cells and solute secretion/reabsorption across epithelia. Acid-base transporters function as either acid extruders or acid loaders, with the Slc4 proteins moving HCO(-) 3 either into or out of cells. According to results from both molecular and functional studies, multiple Slc4 proteins and/or associated splice variants with similar expected effects on pHi are often found in the same tissue or cell. Such apparent redundancy is likely to be physiologically important. In addition to regulating pHi, a HCO(-) 3 transporter contributes to a cell's ability to fine tune the intracellular regulation of the cotransported/exchanged ion(s) (e.g., Na(+) or Cl(-)). In addition, functionally similar transporters or splice variants with different regulatory profiles will optimize pH physiology and solute transport under various conditions or within subcellular domains. Such optimization will depend on activated signaling pathways and transporter expression profiles. In this review, we will summarize and discuss both well-known and more recently identified regulators of the Slc4 proteins. Some of these regulators include traditional second messengers, lipids, binding proteins, autoregulatory domains, and less conventional regulators. The material presented will provide insight into the diversity and physiological significance of multiple members within the Slc4 gene family.

Contribution of starter cultures to the proteolytic process of a fermented non-dried whole muscle ham product.

PubMed

Scannell, Amalia G M; Kenneally, Paul M; Arendt, Elke K

2004-06-01

Porcine longissimus dorsi muscles were cured by brine injection. Curing brine containing 15% (w/v) NaCl, 1.33% (w/v) glucose, 750 ppm sodium nitrite, and appropriate levels of either Lactobacillus sakei LAD, L. sakei LAD plus Kocuria varians FT4 (formally Micrococcus varians), L. sakei LAD plus papain and GDL (glucono-delta-lactone) plus K. varians FT4, was injected to the muscle at a pumping rate 15% w/v. The effect of these treatments on the proteolysis in the ham system was compared to a control ham, produced without starter culture and containing GDL acidulant to control pH and antibiotics to reduce the contribution of background microflora. Hydrolysis of sarcoplasmic and myofibrillar protein fractions was evaluated by SDS-PAGE and reverse phase-HPLC. Hams with different treatments were also investigated for differences in amino acid profile, protein and non-protein nitrogen level, colour, pH, water activity and moisture and microbiological evolution. There was no significant difference in the gross compositional analysis of any of the treatments compared to the control. There was no significant difference (p>0.05) in the protein content, non-protein nitrogen level, SDS-PAGE and free amino acid analysis between the control ham and ham inoculated with proteolytic starter culture. However, it was observed that hams containing starter cultures exhibited decreases in certain peptide fractions and corresponding increases in some free amino acids compared to the uninoculated control. It can be concluded that, while the principle mechanisms resulting in the proteolysis of this non-dried ham product involve the activity of endogeneous cathepsins, the addition of proteolytic starter cultures influence the amino acid profile thereby potentially enhancing the sensorial attributes of the ham. Copyright 2004 Elsevier B.V.

Effect of mechanically deboned poultry meat content on technological properties and sensory characteristics of lamb and mutton sausages

PubMed Central

Perez, Juan Ramon Olalquiaga

2018-01-01

Objective This study aimed to develop a value-added product concerning technological and sensory characteristics changes of the use of mechanically deboned poultry meat (MDPM) as meat replacer in lamb and mutton emulsion-type sausages (mortadella). Methods Sausages were produced with lamb and mutton and with different contents of MDPM. Six treatments, using lamb or mutton and 0%, 30%, and 60% of MDPM in relation to the meat batter, were produced and analyzed for pH, proximal composition, calcium and residual nitrite content, water activity, 2-thiobarbituric acid reactive substances (TBARS), instrumental color and texture profile. The sensory profile of the mortadella’s was also evaluated by acceptance test and check-all-that-applies (CATA) analysis. Results The MDPM addition increased (p<0.05) fat, residual nitrite and calcium content in the all sausage formulations, but mutton sausage had (p<0.05) higher fat and lower moisture content than lamb sausage. The pH, water activity, TBARS index and color was not affected by MDPM additions, while the mutton sausages were significantly redder (higher a*, C*, and lower h°) and darker (lower L*) than lamb sausages. Adding up to 60% of MDPM reduced (p<0.05) sausages hardness and chewiness. Overall, the meat replacement by MDPM increased the sausages acceptance, but the mutton sausage with 30% of MDPM replacer were the most preferred. Consumers related that pink color, glossy appearance, poultry meat-like taste, soft texture, juicy and greasy mouth feel to all sausages contain MDPM according to CATA analysis. Conclusion Mutton from culled ewes can be utilized for mortadella production with 30% replacement of lean mutton and fat by MDPM. PMID:28920407

Effect of mechanically deboned poultry meat content on technological properties and sensory characteristics of lamb and mutton sausages.

PubMed

Massingue, Armando Abel; de Almeida Torres Filho, Robledo; Fontes, Paulo Rogério; de Lemos Souza Ramos, Alcinéia; Fontes, Edimar Aparecida Filomeno; Perez, Juan Ramon Olalquiaga; Ramos, Eduardo Mendes

2018-04-01

This study aimed to develop a value-added product concerning technological and sensory characteristics changes of the use of mechanically deboned poultry meat (MDPM) as meat replacer in lamb and mutton emulsion-type sausages (mortadella). Sausages were produced with lamb and mutton and with different contents of MDPM. Six treatments, using lamb or mutton and 0%, 30%, and 60% of MDPM in relation to the meat batter, were produced and analyzed for pH, proximal composition, calcium and residual nitrite content, water activity, 2-thiobarbituric acid reactive substances (TBARS), instrumental color and texture profile. The sensory profile of the mortadella's was also evaluated by acceptance test and check-all-that-applies (CATA) analysis. The MDPM addition increased (p<0.05) fat, residual nitrite and calcium content in the all sausage formulations, but mutton sausage had (p<0.05) higher fat and lower moisture content than lamb sausage. The pH, water activity, TBARS index and color was not affected by MDPM additions, while the mutton sausages were significantly redder (higher a *, C *, and lower h °) and darker (lower L *) than lamb sausages. Adding up to 60% of MDPM reduced (p<0.05) sausages hardness and chewiness. Overall, the meat replacement by MDPM increased the sausages acceptance, but the mutton sausage with 30% of MDPM replacer were the most preferred. Consumers related that pink color, glossy appearance, poultry meat-like taste, soft texture, juicy and greasy mouth feel to all sausages contain MDPM according to CATA analysis. Mutton from culled ewes can be utilized for mortadella production with 30% replacement of lean mutton and fat by MDPM.

Interactions between soy protein from water-soluble soy extract and polysaccharides in solutions with polydextrose.

PubMed

Spada, Jordana C; Marczak, Ligia D F; Tessaro, Isabel C; Cardozo, Nilo S M

2015-12-10

This study focuses on the investigation of the interactions between polysaccharides (carrageenan and carboxymethylcellulose--CMC) and soy proteins from the water-soluble soy extract. The influence of pH (2-7) and protein-polysaccharide ratio (5:1-40:1) on the interaction between these polyelectrolytes was investigated in aqueous solutions with 10% of polydextrose and without polydextrose. The studied systems were analyzed in terms of pH-solubility profile of protein, ζ-potential, methylene blue-polysaccharide interactions, differential scanning calorimetry (DSC), Fourier transform infrared spectroscopy (FTIR), and confocal laser scanning microscopy. Although the mixtures of soy extract with both carrageenan and CMC showed dependency on the pH and protein-polysaccharide ratio, they did not present the same behavior. Both polysaccharides modified the pH-solubility profile of the soy protein, shifting the pH range in which the coacervate is formed to a lower pH region with the decrease of the soy extract-polysaccharide ratio. The samples also presented detectable differences regarding to ζ-potential, DSC, FTIR and microscopy analyses. The complex formation was also detected even in a pH range where both biopolymers were net-negatively charged. The changes promoted by the presence of polydextrose were mainly detected by blue-polysaccharide interactions measures and confocal microscopy. Copyright © 2015 Elsevier Ltd. All rights reserved.

[24-h intraesophageal pH determination in children allergic to cow's milk protein at a tertiary care hospital].

PubMed

Ramírez-Mayans, J A; Toro-Monjaraz, E M; Romero-Trujillo, J; Cervantes-Bustamante, R; Zárate-Mondragón, F; Montijo-Barrios, E; Cadena-León, J; Cazares-Méndez, M

2014-01-01

Cow's milk protein allergy (CMPA) is being seen more frequently on a daily basis in pediatric consultations. It shares symptoms with gastroesophageal reflux (GER), which can complicate the differential diagnosis. To attempt to corroborate the presence of acid GER in children with CMPA, as well as to find a characteristic profile through the 24-hour pH monitoring study in children with GER and CMPA METHODS: The intraesophageal pH monitoring studies performed on 47 children with CMPA were reviewed. The measurements in all the studies were carried out within a 24-hour period using Digitrapper® equipment with a multi-use GeroFlex® catheter, after calibration with pH 7 and pH 1 buffer solutions. Of the 47 children, 23 were boys (32.4%) and 24 were girls (33.8%) and the mean age was 5±3.7 years. Fourteen of the 47 children (29%) presented with GER, according to the result of the 24-hour intraesophageal measurement. Only 2 of the 47 patients studied fit the phasic profile. The findings show the existing relation between the two pathologies. Nevertheless, it is important to determine the presence of non-acid or weak acid reflux, because their existence can increase this association. Copyright © 2013 Asociación Mexicana de Gastroenterología. Published by Masson Doyma México S.A. All rights reserved.

Development and evaluation of a novel polymeric hydrogel of sucrose acrylate-co-polymethylacrylic acid for oral curcumin delivery.

PubMed

Huang, Sijin; Wang, Jialei; Shang, Qing

2017-02-01

A monomer of sucrose acrylate (AC-sucrose) was synthesized by conjugating starting compound sucrose with methyl acrylate (MA). The obtained AC-sucrose was characterized by mass spectrometry (MS) and Fourier transform infrared (FTIR) spectroscopy. AC-sucrose was selected as a monomer to fabricate a novel pH sensitive hydrogel via free radical polymerization. The inner morphology of the final hydrogel was observed with an S-4800 scanning electron microscope (SEM). The swelling and de-swelling behaviors of the hydrogel chips were also studied. Curcumin (CUR) was selected as a model drug and loaded into the final hydrogel. The release profiles of CUR were performed via dialysis method in pH 1.2, 6.8 and 7.4 buffers, respectively. Mass and FTIR spectra confirmed the synthesis of AC-sucrose. SEM photographs showed that poly(AC-sucrose-co-MAA) hydrogels had many 3D meshes. In pH 1.2 buffer, the hydrogel chips showed the biggest swelling ratio (SR) of 34.4 ± 1.9%. However, in pH 7.4 buffer, the SRs of the hydrogel chips reached to 368.7 ± 28.0%, which suggested that the hydrogel had an excellent pH sensibility. The releasing profiles showed that only 4.6 ± 0.4% of CUR was released in pH 1.2 buffer but 93.7 ± 4.7% of CUR was diffused into pH 7.4 buffer. These data suggested that the CUR-loaded poly (AC-sucrose-co-MAA) hydrogel could direct CUR to release in basic environments.

Andrographis paniculata: Dissolution investigation and pharmacokinetic studies of four major active diterpenoids after multiple oral dose administration in healthy Thai volunteers.

PubMed

Pholphana, Nanthanit; Panomvana, Duangchit; Rangkadilok, Nuchanart; Suriyo, Tawit; Puranajoti, Porranee; Ungtrakul, Teerapat; Pongpun, Wanwisa; Thaeopattha, Saichit; Songvut, Phanit; Satayavivad, Jutamaad

2016-12-24

Andrographis paniculata is included in 'The National List of Essential Herbal Drugs A.D. 1999' of Thailand as an herbal drug for the treatment of common cold symptoms and non-infectious diarrhea. The therapeutic activities of A. paniculata are attributed to four major active diterpenoids: andrographolide (1), 14-deoxy-11, 12-didehydroandrographolide (2), neoandrographolide (3), and 14-deoxyandrographolide (4). However, the pharmacokinetic studies in humans of this plant were performed after a single oral dose administration and reported the parameters related to be of only 1. This study aims to determine the pharmacokinetic parameters of four major active diterpenoids after multiple oral dose administration of A. paniculata capsules in healthy volunteers. The dissolution testing of these four diterpenoids was also performed. The dissolution testing of four major active diterpenoids was conducted in pH 1.2, pH 4.5, and pH 6.8 for 10-100min. The pharmacokinetic study of these active diterpenoids was designed as an open-label, multiple oral dose administration of A. paniculata capsules in 20 healthy Thai volunteers at 1:1 ratio of female and male. Each volunteer was given four A. paniculata capsules each time which contained 1, 2, 3, and 4 in the quantities of 32.64, 5.40, 3.60, and 3.84mg, respectively, three times a day for three consecutive days. On the fourth day, after the first dose of the day was administered, blood samples were collected at the predefined time points. The validated LC-MS/MS method was used to simultaneously determine the concentrations of these diterpenoids in the human plasma samples. The pharmacokinetic parameters of each active diterpenoid were determined. All four major active diterpenoids have been completely dissolved in the simulated pH of gastrointestinal tract within 60min of dissolution. The dissolution profiles were found to be highest in pH 6.8 and lowest in pH 1.2, especially for 3. In the pharmacokinetic study, although 1 was administered at the highest dose among these four diterpenoids, 2 exhibited the highest maximum concentrations (C max ) of 44.89ng/mL and area under the plasma concentration-time curve (AUC) of 128.17h×ng/mL. Compound 1 had the second highest C max and AUC as 32.41ng/mL and 55.23h×ng/mL, respectively. The relative systemic exposure, represented by the dose normalized AUC [(h×ng/mL)/(mg/kg)], of 2 was approximately 14 times higher than that of 1, while those of 3 and 4 were approximately 1.5 and 1.6 times higher, respectively. C max , AUC, apparent volume of distribution, and apparent clearance of 2 were found to be significant difference between female and male. However, when these parameters were calculated as dose normalized basis, no statistically significant difference was found. The four major active diterpenoids in the A. paniculata capsules were soluble in all studied dissolution media. The pharmacokinetic parameters of these active diterpenoids in the present study could be applied for dose optimization of A. paniculata product in order to obtain good therapeutic efficacy and reduce the possible side effects that may occur from different active diterpenoids in this medicinal plant. Copyright © 2016 Elsevier Ireland Ltd. All rights reserved.

Synthesis and activity of Helicobacter pylori urease and catalase at low pH.

PubMed Central

Bauerfeind, P; Garner, R; Dunn, B E; Mobley, H L

1997-01-01

BACKGROUND: Helicobacter pylori produces large amounts of urease presumably to be prepared for the rare event of a sudden acid exposure. The hypothesis that H pylori is acid sensitive and protein production is inhibited by low pH was examined. METHODS: H pylori or its soluble enzymes were incubated buffered or unbuffered at a pH ranging from 2-7 in the presence of 5 mM urea for 30 minutes. After exposure, urease and catalase activities of whole cells, supernatants, and soluble enzyme preparations were measured at pH 6.8. Newly synthesised enzyme was quantified by immunoprecipitation of [35S]-methionine labelled protein. RESULTS: Exposure to buffer below pH 4 resulted in loss of intracellular urease activity. In soluble enzyme preparations and supernatant, no urease activity was measurable after incubation at pH < 5. In contrast, catalase in whole cells, supernatant, and soluble enzyme preparations remained active after exposure to pH > or = 3. Exposure below pH 5 inhibited synthesis of total protein including nascent urease and catalase. At pH 6 or 7, urease represented 10% of total protein, catalase 1.5%. Exposure of H pylori to unbuffered HCl (pH > 2) resulted in an immediate neutralisation; urease and catalase activities and synthesis were unchanged. CONCLUSION: Low surrounding pH reduces activity of urease and synthesis of nascent urease, catalase, and presumably of most other proteins. This suggests that H pylori is not acidophilic although it tolerates short-term exposure to low pH. PMID:9155571

Alkaline pH activates the transport activity of GLUT1 in L929 fibroblast cells.

PubMed

Gunnink, Stephen M; Kerk, Samuel A; Kuiper, Benjamin D; Alabi, Ola D; Kuipers, David P; Praamsma, Riemer C; Wrobel, Kathryn E; Louters, Larry L

2014-04-01

The widely expressed mammalian glucose transporter, GLUT1, can be acutely activated in L929 fibroblast cells by a variety of conditions, including glucose deprivation, or treatment with various respiration inhibitors. Known thiol reactive compounds including phenylarsine oxide and nitroxyl are the fastest acting stimulators of glucose uptake, implicating cysteine biochemistry as critical to the acute activation of GLUT1. In this study, we report that in L929 cells glucose uptake increases 6-fold as the pH of the uptake solution is increased from 6 to 9 with the half-maximal activation at pH 7.5; consistent with the pKa of cysteine residues. This pH effect is essentially blocked by the pretreatment of the cells with either iodoacetamide or cinnamaldehyde, compounds that form covalent adducts with reduced cysteine residues. In addition, the activation by alkaline pH is not additive at pH 8 with known thiol reactive activators such as phenylarsine oxide or hydroxylamine. Kinetic analysis in L929 cells at pH 7 and 8 indicate that alkaline conditions both increases the Vmax and decreases the Km of transport. This is consistent with the observation that pH activation is additive to methylene blue, which activates uptake by increasing the Vmax, as well as to berberine, which activates uptake by decreasing the Km. This suggests that cysteine biochemistry is utilized in both methylene blue and berberine activation of glucose uptake. In contrast a pH increase from 7 to 8 in HCLE cells does not further activate glucose uptake. HCLE cells have a 25-fold higher basal glucose uptake rate than L929 cells and the lack of a pH effect suggests that the cysteine biochemistry has already occurred in HCLE cells. The data are consistent with pH having a complex mechanism of action, but one likely mediated by cysteine biochemistry. Copyright © 2013 Elsevier Masson SAS. All rights reserved.

Characterization of callase (β-1,3-D-glucanase) activity during microsporogenesis in the sterile anthers of Allium sativum L. and the fertile anthers of A. atropurpureum.

PubMed

Winiarczyk, Krystyna; Jaroszuk-Ściseł, Jolanta; Kupisz, Kamila

2012-06-01

We examined callase activity in anthers of sterile Allium sativum (garlic) and fertile Allium atropurpureum. In A. sativum, a species that produces sterile pollen and propagates only vegetatively, callase was extracted from the thick walls of A. sativum microspore tetrads exhibited maximum activity at pH 4.8, and the corresponding in vivo values ranged from 4.5 to 5.0. Once microspores were released, in vitro callase activity peaked at three distinct pH values, reflecting the presence of three callase isoforms. One isoform, which was previously identified in the tetrad stage, displayed maximum activity at pH 4.8, and the remaining two isoforms, which were novel, were most active at pH 6.0 and 7.3. The corresponding in vivo values ranged from pH 4.75 to 6.0. In contrast, in A. atropurpureum, a sexually propagating species, three callase isoforms, active at pH 4.8-5.2, 6.1, and 7.3, were identified in samples of microsporangia that had released their microspores. The corresponding in vivo value for this plant was 5.9. The callose wall persists around A. sativum meiotic cells, whereas only one callase isoform, with an optimum activity of pH 4.8, is active in the acidic environment of the microsporangium. However, this isoform is degraded when the pH rises to 6.0 and two other callase isoforms, maximally active at pH 6.0 and 7.3, appear. Thus, factors that alter the pH of the microsporangium may indirectly affect the male gametophyte development by modulating the activity of callase and thereby regulating the degradation of the callose wall.

High-intensity ultrasound production of Maillard reaction flavor compounds in a cysteine-xylose model system.

PubMed

Ong, Olivia X H; Seow, Yi-Xin; Ong, Peter K C; Zhou, Weibiao

2015-09-01

Application of high intensity ultrasound has shown potential in the production of Maillard reaction odor-active flavor compounds in model systems. The impact of initial pH, sonication duration, and ultrasound intensity on the production of Maillard reaction products (MRPs) by ultrasound processing in a cysteine-xylose model system were evaluated using Response Surface Methodology (RSM) with a modified mathematical model. Generation of selected MRPs, 2-methylthiophene and tetramethyl pyrazine, was optimal at an initial pH of 6.00, accompanied with 78.1 min of processing at an ultrasound intensity of 19.8 W cm(-2). However, identification of volatiles using gas chromatography-mass spectrometry (GC/MS) revealed that ultrasound-assisted Maillard reactions generated fewer sulfur-containing volatile flavor compounds as compared to conventional heat treatment of the model system. Likely reasons for this difference in flavor profile include the expulsion of H2S due to ultrasonic degassing and inefficient transmission of ultrasonic energy. Copyright © 2015 Elsevier B.V. All rights reserved.

Partial purification and kinetic characterization of acid phosphatase from garlic seedling.

PubMed

Yenigün, Begüm; Güvenilir, Yüksel

2003-01-01

The objective of this study was to obtain purer acid phosphatases than produced by prior art by operating under conditions that improve the final product. The study features are the use of a mild nonionic detergent, 40-80% saturation with (NH4)2SOm4, maintained at low temperature to remove impurity, and the use of chromatografic columns to concentrate the acid phosphatase and remove non-acid phosphatase proteins with lower or higher molecular weights. Acid phosphatase was isolated and purified from garlic seedlings by a streamline method without the use of proteolytic and lipolytic enzymes, butanol, or other organic solvents. Grown garlic seedlings of 10- 15 cm height were homogenized with 0.1 M acetate buffer containing 0.1 M NaCl and 0.1% Triton X-100. After homogenization, the supernatant was filtered with paper filters. Filtrated supernatant was cooled to 4 degrees C, followed by a threestep fractionation of the proteins with ammonium sulfate. The crude enzyme was isolated as a green precipitate that was dissolved in a small amount of 0.1 M acetate buffer containing 0.1 M NaCl and 0.1% Triton X-100. Garlic seedling acid phosphatase was purified with ion-exchange chromatography (DEAE cellulose). The column was equilibrated with 0.1 M acetate buffer. Acid phosphatase was purified 40-fold from the starting material. The specific activity of the pure enzyme was 168 U/mg. A variety of stability and activity profiles were determined for the purified garlic seedling acid phosphatase: optimum pH, optimum temperature, pH stability, temperature stability, thermal inactivation, substrate specificity, effect of enzyme concentration, effect of substrate concentration, activation energy, and effect of inhibitor and activator. The molecular mass of acid phosphatase was estimated to be 58 kDa by sodium dodecyl sulfate polyacrylamide gel electrophoresis. The optimum pH was 5.7 and the optimum temperature was 50 degrees C. The enzyme was stable at pH 4.0-10.0 and 40-60 degrees C. Activation energy was between 10 and 20 kcal, and as Michaelis Menten coefficients, Vm values were 100 and 20 mM/s and Km values were 21.27 and 8.33 mM for paranitrophenylphosphate and paranitrophenyl, respectively. Studies of the effect of metal ions on enzyme activity showed both an activating and a deactivating effect. While Cu, Mo, and Mn showed strong inhibitory effects, Na, Ca, and K were the significant activators of acid phosphatase.

Change of pH during excess sludge fermentation under alkaline, acidic and neutral conditions.

PubMed

Yuan, Yue; Peng, Yongzhen; Liu, Ye; Jin, Baodan; Wang, Bo; Wang, Shuying

2014-12-01

The change in pH during excess sludge (ES) fermentation of varying sludge concentrations was investigated in a series of reactors at alkaline, acidic, and neutral pHs. The results showed that the changes were significantly affected by fermentative conditions. Under different conditions, pH exhibited changing profiles. When ES was fermented under alkaline conditions, pH decreased in a range of (10±1). At the beginning of alkaline fermentation, pH dropped significantly, at intervals of 4h, 4h, and 5h with sludge concentrations of 8665.6mg/L, 6498.8mg/L, and 4332.5mg/L, then it would become moderate. However, under acidic conditions, pH increased from 4 to 5. Finally, under neutral conditions pH exhibited a decrease then an increase throughout entire fermentation process. Further study showed short-chain fatty acids (SCFAs), ammonia nitrogen and cations contributed to pH change under various fermentation conditions. This study presents a novel strategy based on pH change to predict whether SCFAs reach their stable stage. Copyright © 2014 Elsevier Ltd. All rights reserved.

Kinetic and CD/MCD spectroscopic studies of the atypical, three-His-ligated, non-heme Fe2+ center in diketone dioxygenase: the role of hydrophilic outer shell residues in catalysis.

PubMed

Straganz, Grit D; Diebold, Adrienne R; Egger, Sigrid; Nidetzky, Bernd; Solomon, Edward I

2010-02-09

Diketone cleaving enzyme (Dke1) is a dioxygenase with an atypical, three-histidine-ligated, mononuclear non-heme Fe(2+) center. To assess the role in enzyme catalysis of the hydrophilic residues in the active site pocket, residues Glu98, Arg80, Tyr70, and Thr107 were subjected to mutational analysis. Steady state and pre-steady state kinetics indicated a role for Glu98 in promoting both substrate binding and O(2) reduction. Additionally, the Glu98 substitution eliminated the pH dependence of substrate binding (k(cat)(app)/K(M)(app)-pH profile) present in wild-type Dke1 (pK(a) = 6.3 +/- 0.4 and 8.4 +/- 0.4). MCD spectroscopy revealed that the Glu98 --> Gln mutation leads to the conversion of the six-coordinate (6C) resting Fe(2+) center present in the wild-type enzyme at pH 7.0 to a mixture of five-coordinate (5C) and 6C sites. The 6C geometry was restored with a pH shift to 9.5 which also resulted in ligand field (LF) energy splittings identical to that found for wild-type (WT) Dke1 at pH 9.5. In WT Dke1, these LF transitions are shifted up in energy by approximately 300 cm(-1) at pH 9.5 relative to pH 7.0. These data, combined with CD pH titrations which reveal a pK(a) of approximately 8.2 for resting WT Dke1 and the Glu98 --> Gln variant, indicate the deprotonation of a metal-ligated water. Together, the kinetic and spectroscopic data reveal a stabilizing effect of Glu98 on the 6C geometry of the metal center, priming it for substrate ligation. Arg80 and Tyr70 are shown to promote O(2) reduction, while Thr107 stabilizes the Fe(II) cofactor.

Organohalide Respiration with Chlorinated Ethenes under Low pH Conditions

DOE PAGES

Yang, Yi; Cápiro, Natalie L.; Marcet, Tyler F.; ...

2017-06-30

Bioremediation at chlorinated solvent sites often leads to groundwater acidification due to electron donor fermentation and enhanced dechlorination activity. The microbial reductive dechlorination process is robust at circumneutral pH, but activity declines at groundwater pH values below 6.0. Consistent with this observation, the activity of tetrachloroethene (PCE) dechlorinating cultures declined at pH 6.0 and was not sustained in pH 5.5 medium, with one notable exception. Sulf urospirillum multivorans dechlorinated PCE to cis-1,2-dichloroethene (cDCE) in pH 5.5 medium and maintained this activity upon repeated transfers. Microcosms established with soil and aquifer materials from five distinct locations dechlorinated PCE-to-ethene at pH 5.5more » and pH 7.2. Dechlorination to ethene was maintained following repeated transfers at pH 7.2, but no ethene was produced at pH 5.5, and only the transfer cultures derived from the Axton Cross Superfund (ACS) microcosms sustained PCE dechlorination to cDCE as a final product. 16S rRNA gene amplicon sequencing of pH 7.2 and pH 5.5 ACS enrichments revealed distinct microbial communities, with the dominant dechlorinator being Dehalococcoides in pH 7.2 and Sulf urospirillum in pH 5.5 cultures. PCE-to-trichloroethene- (TCE-) and PCE-to-cDCEdechlorinating isolates obtained from the ACS pH 5.5 enrichment shared 98.6%, and 98.5% 16S rRNA gene sequence similarities to Sulf urospirillum multivorans. Lastly, these findings imply that sustained Dehalococcoides activity cannot be expected in low pH (i.e., ≤ 5.5) groundwater, and organohalide-respiring Sulf urospirillum spp. are key contributors to in situ PCE reductive dechlorination under low pH conditions.« less

Organohalide Respiration with Chlorinated Ethenes under Low pH Conditions

DOE Office of Scientific and Technical Information (OSTI.GOV)

Yang, Yi; Cápiro, Natalie L.; Marcet, Tyler F.

Bioremediation at chlorinated solvent sites often leads to groundwater acidification due to electron donor fermentation and enhanced dechlorination activity. The microbial reductive dechlorination process is robust at circumneutral pH, but activity declines at groundwater pH values below 6.0. Consistent with this observation, the activity of tetrachloroethene (PCE) dechlorinating cultures declined at pH 6.0 and was not sustained in pH 5.5 medium, with one notable exception. Sulf urospirillum multivorans dechlorinated PCE to cis-1,2-dichloroethene (cDCE) in pH 5.5 medium and maintained this activity upon repeated transfers. Microcosms established with soil and aquifer materials from five distinct locations dechlorinated PCE-to-ethene at pH 5.5more » and pH 7.2. Dechlorination to ethene was maintained following repeated transfers at pH 7.2, but no ethene was produced at pH 5.5, and only the transfer cultures derived from the Axton Cross Superfund (ACS) microcosms sustained PCE dechlorination to cDCE as a final product. 16S rRNA gene amplicon sequencing of pH 7.2 and pH 5.5 ACS enrichments revealed distinct microbial communities, with the dominant dechlorinator being Dehalococcoides in pH 7.2 and Sulf urospirillum in pH 5.5 cultures. PCE-to-trichloroethene- (TCE-) and PCE-to-cDCEdechlorinating isolates obtained from the ACS pH 5.5 enrichment shared 98.6%, and 98.5% 16S rRNA gene sequence similarities to Sulf urospirillum multivorans. Lastly, these findings imply that sustained Dehalococcoides activity cannot be expected in low pH (i.e., ≤ 5.5) groundwater, and organohalide-respiring Sulf urospirillum spp. are key contributors to in situ PCE reductive dechlorination under low pH conditions.« less

The microbial mats of Pavilion Lake microbialites: examining the relationship between photosynthesis and carbonate precipitation

NASA Astrophysics Data System (ADS)

Lim, D. S. S.; Hawes, I.; Mackey, T. J.; Brady, A. L.; Biddle, J.; Andersen, D. T.; Belan, M.; Slater, G.; Abercromby, A.; Squyres, S. W.; Delaney, M.; Haberle, C. W.; Cardman, Z.

2014-12-01

Pavilion Lake in British Columbia, Canada is an ultra-oligotrophic lake that has abundant microbialite growth. Recent research has shown that photoautotrophic microbial communities are important to modern microbialite development in Pavilion Lake. However, questions remain as to the relationship between changing light levels within the lake, variation in microbialite macro-structure, microbial consortia, and the preservation of associated biosignatures within the microbialite fabrics. The 2014 Pavilion Lake Research Project (PLRP) field program was focused on data gathering to understand these complex relationships by determining if a) light is the immediate limit to photosynthetic activity and, if so, if light is distributed around microbialites in ways that are consistent with emergent microbialite structure; and b) if at more local scales, the filamentous pink and green cyanobacterial nodular colonies identified in previous PLRP studies are centers of photosynthetic activity that create pH conditions suitable for carbonate precipitation. A diver-deployed pulse-amplitude modulated (PAM) fluorometer was used to collect synoptic in situ measurements of fluorescence yield and irradiance and across microbialites, focusing on comparing flat and vertical structural elements at a range of sites and depths. As well, we collected time series measurements of photosynthetic activity and irradiance at a set depth of 18 m across three different regions in Pavilion Lake. Our initial findings suggest that all microbialite surfaces are primarily light-limited regardless of depth or location within the lake. Shore based PAM fluorometry and microelectrode profiling of diver-collected samples suggest that pink and green nodules have different photosynthetic properties and pH profiles, and that nodular growth is likely to be the primary route of calcification due to the gelatinous covering the nodule creates. On-going tests for molecular signatures and isotopic shifts will allow for further examination of surface microvariation and the associated influence on microbialite development.

Synthesis of Cross-Linked Polymeric Micelle pH Nanosensors: An Investigation of Design Flexibility.

PubMed

Kumar, E K Pramod; Jølck, Rasmus I; Andresen, Thomas L

2015-09-01

The design flexibility that polymeric micelles offer in the fabrication of optical nanosensors for ratiometric pH measurements is investigated. pH nanosensors based on polymeric micelles are synthesized either by a mixed-micellization approach or by a postmicelle modification strategy. In the mixed-micellization approach, self-assembly of functionalized unimers followed by shell cross-linking by copper-catalyzed azide-alkyne cycloaddition (CuAAC) results in stabilized cRGD-functionalized micelle pH nanosensors. In the postmicelle modification strategy, simultaneous cross-linking and fluorophore conjugation at the micelle shell using CuAAC results in a stabilized micelle pH nanosensor. Compared to the postmicelle modification strategy, the mixed-micellization approach increases the control of the overall composition of the nanosensors. Both approaches provide stable nanosensors with similar pKa profiles and thereby nanosensors with similar pH sensitivity. © 2015 WILEY-VCH Verlag GmbH & Co. KGaA, Weinheim.

Size control of Au NPs supported by pH operation

NASA Astrophysics Data System (ADS)

Ichiji, Masumi; Akiba, Hiroko; Hirasawa, Izumi

2017-07-01

Au NPs are expected to become useful functional particles, as particle gun used for plant gene transfer and also catalysts. We have studied PSD (particle size distribution) control of Au NPs by reduction crystallization. Previous study found out importance of seeds policy and also feeding profile. In this paper, effect of pH in the reduction crystallization was investigated to clarify the possibility of Au NPs PSD control by pH operation and also their growth process. Au NPs of size range 10-600 nm were obtained in single-jet system using ascorbic acid (AsA) as a reducing agent with adjusting pH of AsA. Au NPs are found to grow in the process of nucleation, agglomeration, agglomeration growth and surface growth. Au NPs tend to grow by agglomeration and become larger size in lower pH regions, and to grow only by surface growth and become smaller size in higher pH regions.

Characterization of glutamate decarboxylase from Lactobacillus plantarum and its C-terminal function for the pH dependence of activity.

PubMed

Shin, Sun-Mi; Kim, Hana; Joo, Yunhye; Lee, Sang-Jae; Lee, Yong-Jik; Lee, Sang Jun; Lee, Dong-Woo

2014-12-17

The gadB gene encoding glutamate decarboxylase (GAD) from Lactobacillus plantarum was cloned and expressed in Escherichia coli. The recombinant enzyme exhibited maximal activity at 40 °C and pH 5.0. The 3D model structure of L. plantarum GAD proposed that its C-terminal region (Ile454-Thr468) may play an important role in the pH dependence of catalysis. Accordingly, C-terminally truncated (Δ3 and Δ11 residues) mutants were generated and their enzyme activities compared with that of the wild-type enzyme at different pH values. Unlike the wild-type GAD, the mutants showed pronounced catalytic activity in a broad pH range of 4.0-8.0, suggesting that the C-terminal region is involved in the pH dependence of GAD activity. Therefore, this study may provide effective target regions for engineering pH dependence of GAD activity, thereby meeting industrial demands for the production of γ-aminobutyrate in a broad range of pH values.

Genetically encoded proton sensors reveal activity-dependent pH changes in neurons.

PubMed

Raimondo, Joseph V; Irkle, Agnese; Wefelmeyer, Winnie; Newey, Sarah E; Akerman, Colin J

2012-01-01

The regulation of hydrogen ion concentration (pH) is fundamental to cell viability, metabolism, and enzymatic function. Within the nervous system, the control of pH is also involved in diverse and dynamic processes including development, synaptic transmission, and the control of network excitability. As pH affects neuronal activity, and can also itself be altered by neuronal activity, the existence of tools to accurately measure hydrogen ion fluctuations is important for understanding the role pH plays under physiological and pathological conditions. Outside of their use as a marker of synaptic release, genetically encoded pH sensors have not been utilized to study hydrogen ion fluxes associated with network activity. By combining whole-cell patch clamp with simultaneous two-photon or confocal imaging, we quantified the amplitude and time course of neuronal, intracellular, acidic transients evoked by epileptiform activity in two separate in vitro models of temporal lobe epilepsy. In doing so, we demonstrate the suitability of three genetically encoded pH sensors: deGFP4, E(2)GFP, and Cl-sensor for investigating activity-dependent pH changes at the level of single neurons.

Over Time, How Do Post-Ph.D. Scientists Locate Teaching and Supervision within Their Academic Practice?

ERIC Educational Resources Information Center

McAlpine, Lynn

2014-01-01

While building a strong research profile is usually seen as key for those seeking a traditional academic position, teaching is also understood as central to academic practice. Still, we know little of how post-Ph.D. researchers seeking academic posts locate teaching and supervision in their academic practice, nor how their views may shift as they…

Simulation of stimuli-triggered release of molecular species from halloysite nanotubes

NASA Astrophysics Data System (ADS)

Elumalai, Divya Narayan; Tully, Joshua; Lvov, Yuri; Derosa, Pedro A.

2016-10-01

A Monte Carlo model is used to study the effect of environmental variables (pH and temperature) on the transport and release of dexamethasone molecules from Halloysite Nanotubes (HNTs) in a dielectric fluid medium. The model used for this study was introduced elsewhere and it is based on basic physics interactions without experimental parameters for these interactions. An intermediate phase between the burst and saturation phase is found and explained. Molecules experience a 1-D diffusion process that is different from the diffusion in the burst phase or the surface diffusion experienced by molecules attached to the wall. It is predicted that this phase exists when the molecule-wall interaction is attractive but not always noticeable in the release profile. In this work, it is shown that an agreement with the experiment better than previously reported is obtained when simulated delivery curves are produced by the weighted average of the release profiles from a collection of HNTs with diameters and lengths distributed according to the experimental sample, highlighting the relevance of HNTs' morphology in the release. HNTs are suitable for environment-triggered release and thus the effect of temperature, molecule zeta potential, and pH is studied. It is observed that for temperatures that significantly differ from room temperature (by 100's of degrees), the release profile changes significantly, increasing the delivery speed at high temperature and reducing that speed at low temperature. Finally, it is observed that as the pH becomes more acidic, both the molecule and inner wall surface become more positive (or less negative) with both eventually becoming positive leading to a repulsive interaction; thus, molecules are pushed out by electrostatic repulsion. On the contrary, as the pH becomes more basic, positive molecules become more positive while the wall becomes less negative, but even at pH 12, the wall remains negative and the interaction is attractive. Changes in pH between different regions may act as a trigger for delivery or as a control in the delivery rate.

Different Recovery Profiles of Coagulation Factors, Thrombin Generation, and Coagulation Function After Hemorrhagic Shock in Pigs

DTIC Science & Technology

2012-06-06

Different recovery profiles of coagulation factors, thrombin generation, and coagulation function after hemorrhagic shock in pigs Wenjun Z. Martini ...Defense. Address for reprints: Wenjun Z. Martini , PhD, The US Army Institute of Surgical Research, 3698 Chambers Pass, Ft. Sam Houston, San Antonio, TX...control number 1. REPORT DATE 01 SEP 2015 2. REPORT TYPE N/A 3. DATES COVERED - 4. TITLE AND SUBTITLE Different recovery profiles of

Apiaceous vegetable consumption decreases PhIP-induced DNA adducts and increases methylated PhIP metabolites in the urine metabolome in rats.

PubMed

Kim, Jae Kyeom; Gallaher, Daniel D; Chen, Chi; Yao, Dan; Trudo, Sabrina P

2015-03-01

Heterocyclic aromatic amines, such as 2-amino-1-methyl-6-phenylimidazo[4,5-b]pyridine (PhIP), are carcinogenic compounds produced during heating of protein-containing foods. Apiaceous vegetables inhibit PhIP-activating enzymes, whereas cruciferous vegetables induce both PhIP-activating and -detoxifying enzymes. We investigated the effects of these vegetables, either alone or combined, on PhIP metabolism and colonic DNA adduct formation in rats. Male Wistar rats were fed cruciferous vegetables (21%, wt:wt), apiaceous vegetables (21%, wt:wt), or a combination of both vegetables (10.5% wt:wt of each). Negative and positive control groups were fed an AIN-93G diet. After 6 d, all groups received an intraperitoneal injection of PhIP (10 mg · kg body weight(-1)) except for the negative control group, which received only vehicle. Urine was collected for 24 h after the injection for LC-tandem mass spectrometry metabolomic analyses. On day 7, rats were killed and tissues processed. Compared with the positive control, cruciferous vegetables increased the activity of hepatic PhIP-activating enzymes [39.5% and 45.1% for cytochrome P450 (CYP) 1A1 (P = 0.0006) and CYP1A2 (P < 0.0001), respectively] and of uridine 5'-diphospho-glucuronosyltransferase 1A (PhIP-detoxifying) by 24.5% (P = 0.0267). Apiaceous vegetables did not inhibit PhIP-activating enzymes, yet reduced colonic PhIP-DNA adducts by 20.4% (P = 0.0496). Metabolomic analyses indicated that apiaceous vegetables increased the relative abundance of urinary methylated PhIP metabolites. The sum of these methylated metabolites inversely correlated with colonic PhIP-DNA adducts (r = -0.43, P = 0.01). We detected a novel methylated urinary PhIP metabolite and demonstrated that methylated metabolites are produced in the human liver S9 fraction. Apiaceous vegetables did not inhibit the activity of PhIP-activating enzymes in rats, suggesting that the reduction in PhIP-DNA adducts may involve other pathways. Further investigation of the importance of PhIP methylation in carcinogen metabolism is warranted, given the inverse correlation of methylated PhIP metabolites with a biomarker of carcinogenesis and the detection of a novel methylated PhIP metabolite. © 2015 American Society for Nutrition.

Apiaceous Vegetable Consumption Decreases PhIP-Induced DNA Adducts and Increases Methylated PhIP Metabolites in the Urine Metabolome in Rats123

PubMed Central

Kim, Jae Kyeom; Gallaher, Daniel D; Chen, Chi; Yao, Dan; Trudo, Sabrina P

2015-01-01

Background: Heterocyclic aromatic amines, such as 2-amino-1-methyl-6-phenylimidazo[4,5-b]pyridine (PhIP), are carcinogenic compounds produced during heating of protein-containing foods. Apiaceous vegetables inhibit PhIP-activating enzymes, whereas cruciferous vegetables induce both PhIP-activating and -detoxifying enzymes. Objective: We investigated the effects of these vegetables, either alone or combined, on PhIP metabolism and colonic DNA adduct formation in rats. Methods: Male Wistar rats were fed cruciferous vegetables (21%, wt:wt), apiaceous vegetables (21%, wt:wt), or a combination of both vegetables (10.5% wt:wt of each). Negative and positive control groups were fed an AIN-93G diet. After 6 d, all groups received an intraperitoneal injection of PhIP (10 mg · kg body weight−1) except for the negative control group, which received only vehicle. Urine was collected for 24 h after the injection for LC–tandem mass spectrometry metabolomic analyses. On day 7, rats were killed and tissues processed. Results: Compared with the positive control, cruciferous vegetables increased the activity of hepatic PhIP-activating enzymes [39.5% and 45.1% for cytochrome P450 (CYP) 1A1 (P = 0.0006) and CYP1A2 (P < 0.0001), respectively] and of uridine 5′-diphospho-glucuronosyltransferase 1A (PhIP-detoxifying) by 24.5% (P = 0.0267). Apiaceous vegetables did not inhibit PhIP-activating enzymes, yet reduced colonic PhIP-DNA adducts by 20.4% (P = 0.0496). Metabolomic analyses indicated that apiaceous vegetables increased the relative abundance of urinary methylated PhIP metabolites. The sum of these methylated metabolites inversely correlated with colonic PhIP-DNA adducts (r = −0.43, P = 0.01). We detected a novel methylated urinary PhIP metabolite and demonstrated that methylated metabolites are produced in the human liver S9 fraction. Conclusions: Apiaceous vegetables did not inhibit the activity of PhIP-activating enzymes in rats, suggesting that the reduction in PhIP-DNA adducts may involve other pathways. Further investigation of the importance of PhIP methylation in carcinogen metabolism is warranted, given the inverse correlation of methylated PhIP metabolites with a biomarker of carcinogenesis and the detection of a novel methylated PhIP metabolite. PMID:25733458

3,4-Dihydroxyphenylacetate 2,3-dioxygenase from Pseudomonas aeruginosa: An Fe(II)-containing enzyme with fast turnover

PubMed Central

Kamutira, Philaiwarong; Watthaisong, Pratchaya; Thotsaporn, Kittisak; Tongsook, Chanakan; Juttulapa, Maneerat; Nijvipakul, Sarayut; Chaiyen, Pimchai

2017-01-01

3,4-dihydroxyphenylacetate (DHPA) dioxygenase (DHPAO) from Pseudomonas aeruginosa (PaDHPAO) was overexpressed in Escherichia coli and purified to homogeneity. As the enzyme lost activity over time, a protocol to reactivate and conserve PaDHPAO activity has been developed. Addition of Fe(II), DTT and ascorbic acid or ROS scavenging enzymes (catalase or superoxide dismutase) was required to preserve enzyme stability. Metal content and activity analyses indicated that PaDHPAO uses Fe(II) as a metal cofactor. NMR analysis of the reaction product indicated that PaDHPAO catalyzes the 2,3-extradiol ring-cleavage of DHPA to form 5-carboxymethyl-2-hydroxymuconate semialdehyde (CHMS) which has a molar absorptivity of 32.23 mM-1cm-1 at 380 nm and pH 7.5. Steady-state kinetics under air-saturated conditions at 25°C and pH 7.5 showed a Km for DHPA of 58 ± 8 μM and a kcat of 64 s-1, indicating that the turnover of PaDHPAO is relatively fast compared to other DHPAOs. The pH-rate profile of the PaDHPAO reaction shows a bell-shaped plot that exhibits a maximum activity at pH 7.5 with two pKa values of 6.5 ± 0.1 and 8.9 ± 0.1. Study of the effect of temperature on PaDHPAO activity indicated that the enzyme activity increases as temperature increases up to 55°C. The Arrhenius plot of ln(k’cat) versus the reciprocal of the absolute temperature shows two correlations with a transition temperature at 35°C. Two activation energy values (Ea) above and below the transition temperature were calculated as 42 and 14 kJ/mol, respectively. The data imply that the rate determining steps of the PaDHPAO reaction at temperatures above and below 35°C may be different. Sequence similarity network analysis indicated that PaDHPAO belongs to the enzyme clusters that are largely unexplored. As PaDHPAO has a high turnover number compared to most of the enzymes previously reported, understanding its biochemical and biophysical properties should be useful for future applications in biotechnology. PMID:28158217

Advances in protease engineering for laundry detergents.

PubMed

Vojcic, Ljubica; Pitzler, Christian; Körfer, Georgette; Jakob, Felix; Ronny Martinez; Maurer, Karl-Heinz; Schwaneberg, Ulrich

2015-12-25

Proteases are essential ingredients in modern laundry detergents. Over the past 30 years, subtilisin proteases employed in the laundry detergent industry have been engineered by directed evolution and rational design to tailor their properties towards industrial demands. This comprehensive review discusses recent success stories in subtilisin protease engineering. Advances in protease engineering for laundry detergents comprise simultaneous improvement of thermal resistance and activity at low temperatures, a rational strategy to modulate pH profiles, and a general hypothesis for how to increase promiscuous activity towards the production of peroxycarboxylic acids as mild bleaching agents. The three protease engineering campaigns presented provide in-depth analysis of protease properties and have identified principles that can be applied to improve or generate enzyme variants for industrial applications beyond laundry detergents. Copyright © 2015 Elsevier B.V. All rights reserved.

pH profile of the adsorption of nucleotides onto montmorillonite. I - Selected homoionic clays

NASA Technical Reports Server (NTRS)

Lawless, J. G.; Church, F. M.; Mazzurco, J.; Banin, A.; Huff, R.; Kao, J.; Cook, A.; Lowe, T.; Orenberg, J. B.; Edelson, E.

1985-01-01

The effect of pH and adsorbed ions on the adsorption of purine and pyrimidine nucleotides on montmorillonite clay was studied experimentally. The specific nucleotides examined were: 5 prime-AMP; 3-prime AMP; and 5 prime-CMP. The pH of the clay samples was adjusted to various levels in the 2-12 pH range using microliter volumes of concentrated acid (1N HCl) and base (1NHNaOH). It was found that preferential adsorption among nulceotides was dependent on the pH level and on the characteristics of the substituted metal cation and anion exchange mechanisms. Below pH 4, adsorption was attributed to cation and anion exchange mechanisms. Above pH 4, however, adsorption was attributed to the complexation mechanisms occurring between the metal cations in the clay exchange site and in the biomolecule. The possible role of homoionic clays in the concentration mechanisms of biomonomers in the prebiotic environment is discussed.

Effects of pH changes in water-based solvents to isolate antibacterial activated extracts of natural products

NASA Astrophysics Data System (ADS)

Buang, Yohanes; Suwari, Ola, Antonius R. B.

2017-12-01

Effects of pH changes in solvents on isolation of antibacterial activities of natural product extracts were conducted in the present study. Sarang semut (M. pendens) tubers as the model material for the study was considered to be the strategic resource of natural products based on its biochemical and therapeutical effects. The water with pH 5, 7, 9, and 13 was used as the solvents. The antibacterial activities of the resulted extracts indicated that higher the working pH, higher activities of the resulted extracts. The extent activities of the resulted extracts followed the increasing pH of the maceration system. The study also found that higher pH of the working solvent, higher the amounts of the antibacterial extracts isolated from the sample matrix of the natural product. The higher pH of the water solvents plays essential roles to promote the antibacterial activities of the natural product extracts from M. pendens tubers.

Influence of the metal ion on the enzyme activity and kinetics of PepA from Lactobacillus delbrueckii.

PubMed

Ewert, Jacob; Glück, Claudia; Strasdeit, Henry; Fischer, Lutz; Stressler, Timo

2018-03-01

The aminopeptidase A (PepA; EC 3.4.11.7) belongs to the group of metallopeptidases with two bound metal ions per subunit (M1M2(PepA)) and is specific for the cleavage of N-terminal glutamic (Glu) and aspartic acid (Asp) and, in low amounts, serine (Ser) residues. Our group recently characterized the first PepA from a Lactobacillus strain. However, the characterization was performed using synthetic para-nitroaniline substrates and not original peptide substrates, as was done in the current study. Prior to the characterization using original peptide substrates, the PepA purified was converted to its inactive apo-form and eight different metal ions were tested to restore its activity. It was found that five of the metal ions were able to reactivate apo-PepA: Co 2+ , Cu 2+ , Mn 2+ , Ni 2+ and Zn 2+ . Interestingly, depending on the metal ion used for reactivation, the activity and the pH and temperature profile differed. Exemplarily, MnMn(PepA), NiNi(PepA) and ZnZn(PepA) had an activity optimum using MES buffer (50mM, pH 6.0) and 60°C, whereas the activity optimum changed to Na/K-phosphate-buffer (50mM, pH 7.0) and 55°C for CuCu(PepA). However, more important than the changes in optimum pH and temperature, the kinetic properties of PepA were affected by the metal ion used. While all PepA variants could release N-terminal Glu or Asp, only CoCo(PepA), NiNi(PepA) and CuCu(PepA) could release Ser from the particular peptide substrate. In addition, it was found that the enzyme efficiency (V max /K M ) and catalytic mechanism (positive cooperative binding (Hill coefficent; n), substrate inhibition (K IS )) were influenced by the metal ion. Exemplarily, a high cooperativity (n>2),K IS value >20mM and preference for N-terminal Glu were detected for CuCu(PepA). In summary, the results suggested that an exchange of the metal ion can be used for tailoring the properties of PepA for specific hydrolysis requirements. Copyright © 2017 Elsevier Inc. All rights reserved.

Immobilization of Chloroperoxidase on Aminopropyl-Glass

PubMed Central

Kadima, Tenshuk A.; Pickard, Michael A.

1990-01-01

Chloroperoxidase (CPO) purified from Caldariomyces fumago CMI 89362 was covalently bound to aminopropyl-glass by using a modification of an established method. Acid-washed glass was derivatized by using aminopropyltriethoxysilane, and the enzyme was ionically bound at low ionic strength. Further treatment with glutaraldehyde covalently linked the enzyme to the glass beads in an active form. No elution of bound activity from glass beads could be detected with a variety of washings. The loading of enzyme protein to the glass beads was highest, 100 mg of CPO per g of glass, at high reaction ratios of CPO to glass, but the specific activity of the immobilized enzyme was highest, 36% of theoretical, at low enzyme-to-carrier ratios. No differences in the properties of the soluble and immobilized enzymes could be detected by a number of criteria: their pH-activity and pH-stability profiles were similar, as were their thermal stabilities. After five uses, the immobilized enzyme retained full activity between pH 6.0 and 6.7. PMID:16348352

Electrokinetic enhancement on phytoremediation in Zn, Pb, Cu and Cd contaminated soil using potato plants.

PubMed

Aboughalma, Hanssan; Bi, Ran; Schlaak, Michael

2008-07-01

The use of a combination of electrokinetic remediation and phytoremediation to decontaminate soil polluted with heavy metals has been demonstrated in a laboratory-scale experiment. Potato tubers were planted in plastic vessels filled with Zn, Pb, Cu and Cd contaminated soil and grown in a greenhouse. Three of these vessels were treated with direct current electric field (DC), three with alternative current (AC) and three remained untreated as control vessels. The soil pH varied from anode to cathode with a minimum of pH 3 near the anode and a maximum of pH 8 near the cathode in the DC treated soil profile. There was an accumulation of Zn, Cu and Cd at about 12 cm distance from anode when soil pH was 5 in the DC treated soil profile. There was no significant metal redistribution and pH variation between anode and cathode in the AC soil profile. The biomass production of the plants was 72% higher under AC treatment and 27% lower under DC treatment compared to the control. Metal accumulation was generally higher in the plant roots treated with electrical fields than the control. The overall metal uptake in plant shoots was lower under DC treatment compared to AC treatment and control, although there was a higher accumulation of Zn and Cu in the plant roots treated with electrical fields. The Zn uptake in plant shoots under AC treatment was higher compared to the control and DC treatment. Zn and Cu accumulation in the plant roots under AC and DC treatment was similar, and both were higher comparing to control. Cd content in plant roots under all three treatments was found to be higher than that in the soil. The Pb accumulation in the roots and the uptake into the shoots was lower compared to its content in the soil.

Improvement of activity, thermo-stability and fruit juice clarification characteristics of fungal exo-polygalacturonase.

PubMed

Amin, Faiza; Bhatti, Haq Nawaz; Bilal, Muhammad; Asgher, Muhammad

2017-02-01

An extracellular exo-polygalacturonase (exo-PG) from Penicillium notatum was immobilized in sodium-alginate matrix through two different protocols, viz. covalent bonding and adsorption to enhance its catalytic activity, thermal stability and life-time properties for industrial applications. Covalent immobilization was more efficient in terms of high relative activity (45.89%) and immobilization yield (71.6%) as compared to adsorption. Immobilized exo-PG derivatives displayed maximum activities at pH 5.5 and 55°C as compared to free enzyme which showed its optimum activity at pH 6.0 and 50°C. The affinity of enzyme towards its substrate (K m(app) ) was reduced after immobilization and V max of covalently immobilized exo-PG decreased to 66.7% while the V max value of adsorbed enzyme increased up to 150% as compared to free counterpart. Both immobilization techniques greatly enhanced the thermal stability profile of the enzyme. At 60°C, immobilized exo-PGs retained more than 90% of their residual activities after 60min of heating, while free enzyme did not show any activity at the same temperature. Thermodynamic properties (i.e., Ea, ΔH*, ΔS*and ΔG*) of the free and immobilized enzymes were also investigated. Sodium-alginate covalently immobilized and adsorbed enzymes showed excellent recycling efficiencies and retained 50.0% and 41.0% of original activities, respectively after seven consecutive batch reactions. Moreover, the immobilized enzymes treatment achieved promising results in turbidity and viscosity reduction as well as clarity amelioration in various fruit juices. Altogether catalytic, thermo-stability and fruit juices clarification characteristics of the immobilized ex-PGs suggest a high potential for biotechnological exploitability. Copyright © 2016 Elsevier B.V. All rights reserved.

pH-Induced interfacial properties of Chaplin E from Streptomyces coelicolor.

PubMed

Dokouhaki, Mina; Hung, Andrew; Prime, Emma L; Qiao, Greg G; Day, Li; Gras, Sally L

2017-12-01

Chaplin E, or Chp E, is a surface active peptide secreted by Streptomyces coelicolor that adopts different structures depending on solution pH but the effect of these structures on the interfacial properties of Chp E is not known. In experiments paired with simulations, Chp E was found to display pH-dependent interfacial assembly and surface activity. At pH 3.0, Chp E formed an ordered non-amyloidal interfacial film with high surface activity; while at pH 10.0, Chp E self-assembled into a heterogeneous film containing randomly arranged fibrils at the interface that was less surface active compared to the film formed at pH 3.0. In simulations at pH 10.0, Chp E molecules showed a higher propensity for dimerization within the solution phase, lower rate of adsorption to the interface and tighter inter-molecular associations at the interface, consistent with the lower surface activity and smaller interfacial area coverage per molecule measured at this pH compared to at pH 3.0. A model is presented for the role of Chp E in the developmental differentiation of Streptomyces coelicolor, where Chp E contributes to changes in surface tension at low pH and the formation of fibrils on the surface of aerial hyphae at high pH. Our data also suggest Chp E could be a promising surface active agent with functional activity that can be controlled by pH. Copyright © 2017 Elsevier B.V. All rights reserved.

Chemometrics-assisted spectrophotometric green method for correcting interferences in biowaiver studies: Application to assay and dissolution profiling study of donepezil hydrochloride tablets

NASA Astrophysics Data System (ADS)

Korany, Mohamed A.; Mahgoub, Hoda; Haggag, Rim S.; Ragab, Marwa A. A.; Elmallah, Osama A.

2018-06-01

A green, simple and cost effective chemometric UV-Vis spectrophotometric method has been developed and validated for correcting interferences that arise during conducting biowaiver studies. Chemometric manipulation has been done for enhancing the results of direct absorbance, resulting from very low concentrations (high incidence of background noise interference) of earlier points in the dissolution timing in case of dissolution profile using first and second derivative (D1 & D2) methods and their corresponding Fourier function convoluted methods (D1/FF& D2/FF). The method applied for biowaiver study of Donepezil Hydrochloride (DH) as a representative model was done by comparing two different dosage forms containing 5 mg DH per tablet as an application of a developed chemometric method for correcting interferences as well as for the assay and dissolution testing in its tablet dosage form. The results showed that first derivative technique can be used for enhancement of the data in case of low concentration range of DH (1-8 μg mL-1) in the three different pH dissolution media which were used to estimate the low drug concentrations dissolved at the early points in the biowaiver study. Furthermore, the results showed similarity in phosphate buffer pH 6.8 and dissimilarity in the other 2 pH media. The method was validated according to ICH guidelines and USP monograph for both assays (HCl of pH 1.2) and dissolution study in 3 pH media (HCl of pH 1.2, acetate buffer of pH 4.5 and phosphate buffer of pH 6.8). Finally, the assessment of the method greenness was done using two different assessment techniques: National Environmental Method Index label and Eco scale methods. Both techniques ascertained the greenness of the proposed method.

Chemometrics-assisted spectrophotometric green method for correcting interferences in biowaiver studies: Application to assay and dissolution profiling study of donepezil hydrochloride tablets.

PubMed

Korany, Mohamed A; Mahgoub, Hoda; Haggag, Rim S; Ragab, Marwa A A; Elmallah, Osama A

2018-06-15

A green, simple and cost effective chemometric UV-Vis spectrophotometric method has been developed and validated for correcting interferences that arise during conducting biowaiver studies. Chemometric manipulation has been done for enhancing the results of direct absorbance, resulting from very low concentrations (high incidence of background noise interference) of earlier points in the dissolution timing in case of dissolution profile using first and second derivative (D1 & D2) methods and their corresponding Fourier function convoluted methods (D1/FF& D2/FF). The method applied for biowaiver study of Donepezil Hydrochloride (DH) as a representative model was done by comparing two different dosage forms containing 5mg DH per tablet as an application of a developed chemometric method for correcting interferences as well as for the assay and dissolution testing in its tablet dosage form. The results showed that first derivative technique can be used for enhancement of the data in case of low concentration range of DH (1-8μgmL -1 ) in the three different pH dissolution media which were used to estimate the low drug concentrations dissolved at the early points in the biowaiver study. Furthermore, the results showed similarity in phosphate buffer pH6.8 and dissimilarity in the other 2pH media. The method was validated according to ICH guidelines and USP monograph for both assays (HCl of pH1.2) and dissolution study in 3pH media (HCl of pH1.2, acetate buffer of pH4.5 and phosphate buffer of pH6.8). Finally, the assessment of the method greenness was done using two different assessment techniques: National Environmental Method Index label and Eco scale methods. Both techniques ascertained the greenness of the proposed method. Copyright © 2018 Elsevier B.V. All rights reserved.

Biochemistry and Cell Wall Changes Associated with Noni (Morinda citrifolia L.) Fruit Ripening.

PubMed

Cárdenas-Coronel, Wendy G; Carrillo-López, Armando; Vélez de la Rocha, Rosabel; Labavitch, John M; Báez-Sañudo, Manuel A; Heredia, José B; Zazueta-Morales, José J; Vega-García, Misael O; Sañudo-Barajas, J Adriana

2016-01-13

Quality and compositional changes were determined in noni fruit harvested at five ripening stages, from dark-green to thaslucent-grayish. Fruit ripening was accompanied by acidity and soluble solids accumulation but pH diminution, whereas the softening profile presented three differential steps named early (no significant softening), intermediate (significant softening), and final (dramatic softening). At early step the extensive depolymerization of hydrosoluble pectins and the significantly increment of pectinase activities did not correlate with the slight reduction in firmness. The intermediate step showed an increment of pectinases and hemicellulases activities. The final step was accompanied by the most significant reduction in the yield of alcohol-insoluble solids as well as in the composition of uronic acids and neutral sugars; pectinases increased their activity and depolymerization of hemicellulosic fractions occurred. Noni ripening is a process conducted by the coordinated action of pectinases and hemicellulases that promote the differential dissasembly of cell wall polymers.

Heat Priming Induces Trans-generational Tolerance to High Temperature Stress in Wheat

PubMed Central

Wang, Xiao; Xin, Caiyun; Cai, Jian; Zhou, Qin; Dai, Tingbo; Cao, Weixing; Jiang, Dong

2016-01-01

Wheat plants are very sensitive to high temperature stress during grain filling. Effects of heat priming applied to the first generation on tolerance of the successive generation to post-anthesis high temperature stress were investigated. Compared with the progeny of non-heat primed plants (NH), the progeny of heat-primed plants (PH) possessed higher grain yield, leaf photosynthesis and activities of antioxidant enzymes and lower cell membrane damage under high temperature stress. In the transcriptome profile, 1430 probes showed obvious difference in expression between PH and NH. These genes were related to signal transduction, transcription, energy, defense, and protein destination and storage, respectively. The gene encoding the lysine-specific histone demethylase 1 (LSD1) which was involved in histone demethylation related to epigenetic modification was up-regulated in the PH compared with NH. The proteome analysis indicated that the proteins involved in photosynthesis, energy production and protein destination and storage were up-regulated in the PH compared with NH. In short, thermos-tolerance was induced through heritable epigenetic alternation and signaling transduction, both processes further triggered prompt modifications of defense related responses in anti-oxidation, transcription, energy production, and protein destination and storage in the progeny of the primed plants under high temperature stress. It was concluded that trans-generation thermo-tolerance was induced by heat priming in the first generation, and this might be an effective measure to cope with severe high-temperature stresses during key growth stages in wheat production. PMID:27148324

Probiotic Properties of Lactobacillus plantarum RYPR1 from an Indigenous Fermented Beverage Raabadi

PubMed Central

Yadav, Ruby; Puniya, Anil K.; Shukla, Pratyoosh

2016-01-01

Present study documents the potential probiotic Lactobacillus isolated from indigenous fermented beverage Raabadi, consumed during summers in Haryana and Rajasthan regions of India. A total of five Raabadi samples were collected aseptically and 54 isolates were purified using MRS medium. All the isolates were assessed for tolerance to low pH and bile salts. It was observed that out of 54 only 24 isolates could survive the simulated gastric conditions. These isolates were further evaluated in vitro for cell surface hydrophobicity, cell surface hydrophobicity, hypocholesteramic activity, anti-oxidative potential, BSH activity, antagonistic activity, and antibiotic resistance profile. In addition, the confirmation of phenol resistance was also done. On the basis of results obtained, the survival rate of isolates was noted and six isolates were finally selected for further studies. Among them Lactobacillus plantarum RYPR1 and RYPC7 showed good survival at pH 2 which shows good acid tolerance. Moreover, L. plantarum RYPR1 showed the highest hydrophobicity (79.13%) and represented the deconjugation of bile salts, which help in their adhesion to epithelial cells and colonization. Furthermore, RYPR1 also exhibited highest cholesterol reduction (59%) and subsequent analysis of results revealed that the above mentioned isolates further exhibit a good hypocholesterolemic effect and could be possibly used to prevent hypercholesterolemia. The present study divulges that L. plantarum RYPR1 has an excellent probiotic potential. PMID:27818658

Phospholipid bilayer-perturbing properties underlying lysis induced by pH-sensitive cationic lysine-based surfactants in biomembranes.

PubMed

Nogueira, Daniele Rubert; Mitjans, Montserrat; Busquets, M Antonia; Pérez, Lourdes; Vinardell, M Pilar

2012-08-14

Amino acid-based surfactants constitute an important class of natural surface-active biomolecules with an unpredictable number of industrial applications. To gain a better mechanistic understanding of surfactant-induced membrane destabilization, we assessed the phospholipid bilayer-perturbing properties of new cationic lysine-based surfactants. We used erythrocytes as biomembrane models to study the hemolytic activity of surfactants and their effects on cells' osmotic resistance and morphology, as well as on membrane fluidity and membrane protein profile with varying pH. The antihemolytic capacity of amphiphiles correlated negatively with the length of the alkyl chain. Anisotropy measurements showed that the pH-sensitive surfactants, with the positive charge on the α-amino group of lysine, significantly increased membrane fluidity at acidic conditions. SDS-PAGE analysis revealed that surfactants induced significant degradation of membrane proteins in hypo-osmotic medium and at pH 5.4. By scanning electron microscopy examinations, we corroborated the interaction of surfactants with lipid bilayer. We found that varying the surfactant chemical structure is a way to modulate the positioning of the molecule inside bilayer and, thus, the overall effect on the membrane. Our work showed that pH-sensitive lysine-based surfactants significantly disturb the lipid bilayer of biomembranes especially at acidic conditions, which suggests that these compounds are promising as a new class of multifunctional bioactive excipients for active intracellular drug delivery.

Improvement of the Insecticidal Capacity of Two Purpureocillium Lilacinum Strains against Tribolium Confusum

PubMed Central

Barra, Paula; Etcheverry, Miriam; Nesci, Andrea

2015-01-01

Entomopathogenic fungi can regulate insect populations. They have extracellular enzymes that degrade cuticle components, mainly hydrocarbons, used as an energy source. The increase in insecticidal activity of fungi in a medium supplemented with cuticular hydrocarbons was assayed and the hydrolytic enzyme profiles of two strains of Purpureocillium lilacinum were evaluated. A spore suspension of P. lilacinum was inoculated in Petri plates with different values (0.99–0.97–0.95) of water activity (Aw) using the substrates gelatin, starch and tween-20. Growth rate on the different substrates and the enzymatic activity index for proteases, amylases and lipases at different incubation times, pH and Aw, was evaluated. Moreover, the insecticidal efficiency of strains grown in media supplemented with n-hexadecane and n-octacosane was analyzed. LT50 was calculated against adults of Tribolium confusum and showed that mortality increased about 15% when the strains grew in amended culture medium. High amylolytic activity was detected, but proteases were the main enzymes produced. Optimal protease production was observed in a range of acid and alkaline pH and lower Aw. The greatest growth rate was obtained in presence of gelatin. Lipase and amylase production was detected in small amounts. Fungal growth in media with hydrocarbon mixtures increased the pathogenicity of the two strains of P. lilacinum, with the strain JQ926223 being more virulent. The information obtained is important for achieving both an increase in insecticidal capacity and an understanding of physiological adaptation of the fungus. PMID:26463076

Improvement of the Insecticidal Capacity of Two Purpureocillium Lilacinum Strains against Tribolium Confusum.

PubMed

Barra, Paula; Etcheverry, Miriam; Nesci, Andrea

2015-03-18

Entomopathogenic fungi can regulate insect populations. They have extracellular enzymes that degrade cuticle components, mainly hydrocarbons, used as an energy source. The increase in insecticidal activity of fungi in a medium supplemented with cuticular hydrocarbons was assayed and the hydrolytic enzyme profiles of two strains of Purpureocillium lilacinum were evaluated. A spore suspension of P. lilacinum was inoculated in Petri plates with different values (0.99-0.97-0.95) of water activity (Aw) using the substrates gelatin, starch and tween-20. Growth rate on the different substrates and the enzymatic activity index for proteases, amylases and lipases at different incubation times, pH and Aw, was evaluated. Moreover, the insecticidal efficiency of strains grown in media supplemented with n-hexadecane and n-octacosane was analyzed. LT50 was calculated against adults of Tribolium confusum and showed that mortality increased about 15% when the strains grew in amended culture medium. High amylolytic activity was detected, but proteases were the main enzymes produced. Optimal protease production was observed in a range of acid and alkaline pH and lower Aw. The greatest growth rate was obtained in presence of gelatin. Lipase and amylase production was detected in small amounts. Fungal growth in media with hydrocarbon mixtures increased the pathogenicity of the two strains of P. lilacinum, with the strain JQ926223 being more virulent. The information obtained is important for achieving both an increase in insecticidal capacity and an understanding of physiological adaptation of the fungus.

Influence of Lactobacillus plantarum on yogurt fermentation properties and subsequent changes during postfermentation storage.

PubMed

Li, Changkun; Song, Jihong; Kwok, Lai-Yu; Wang, Jicheng; Dong, Yan; Yu, Haijing; Hou, Qiangchuan; Zhang, Heping; Chen, Yongfu

2017-04-01

This study aimed to evaluate the influence of 9 Lactobacillusplantarum with broad-spectrum antibacterial activity on fermented milk, including changes to the fermentation characteristics (pH, titration acidity, and viable counts), texture profile, relative content of volatile compounds, and sensory evaluation during 28-d storage at 4°C. First, L. plantarum IMAU80106, IMAU10216, and IMAU70095 were selected as candidates for further study because of their excellent coagulation and proteolytic activities. Subsequently, these L. plantarum strainswere supplemented to fermented milk produced by commercial yogurt starters (Streptococcus thermophilus and Lactobacillus delbrueckii ssp. bulgaricus) and a panel of parameters reflecting product quality was subsequently monitored during 28 d of postfermentation storage. The pH value and titration acidity of the fermented milk mildly fluctuated, whereas the L. plantarum viable counts remained stable along the storage period. Fourteen key volatile compounds were detected in the fermented milk by gas chromatography-mass spectrometry, and some flavor compounds were uniquely present in the L. plantarum-supplemented fermented milk (including 2,3-pentanedione, acetaldehyde, and acetate). No significant difference was shown in the sensory evaluation scores between samples with or without L. plantarum supplementation, but a gradual decrease was observed over storage in all samples. However, when L. plantarum was added, apparent shifts were observed in the overall quality of the fermented milk based on principal component analysis and multivariate ANOVA, particularly in the texture (adhesiveness) and volatile flavor compound profiles (acetaldehyde). Compared with L. plantarum IMAU80106 and IMAU10216, both the texture and volatile flavor profiles of IMAU70095 were closest to those of the control without adding the adjunct bacteria, suggesting that IMAU70095 might be the most suitable strain for further application in functional dairy product development. The current work has explored the potential of applying L. plantarum in fermented milk by performing thorough physical and chemical characterization. Our work is of intense interest to the dairy industry. Copyright © 2017 American Dairy Science Association. Published by Elsevier Inc. All rights reserved.

Ancient thioredoxins evolved to modern-day stability-function requirement by altering native state ensemble.

PubMed

Modi, Tushar; Huihui, Jonathan; Ghosh, Kingshuk; Ozkan, S Banu

2018-06-19

Thioredoxins (THRXs)-small globular proteins that reduce other proteins-are ubiquitous in all forms of life, from Archaea to mammals. Although ancestral thioredoxins share sequential and structural similarity with the modern-day (extant) homologues, they exhibit significantly different functional activity and stability. We investigate this puzzle by comparative studies of their (ancient and modern-day THRXs') native state ensemble, as quantified by the dynamic flexibility index (DFI), a metric for the relative resilience of an amino acid to perturbations in the rest of the protein. Clustering proteins using DFI profiles strongly resemble an alternative classification scheme based on their activity and stability. The DFI profiles of the extant proteins are substantially different around the α3, α4 helices and catalytic regions. Likewise, allosteric coupling of the active site with the rest of the protein is different between ancient and extant THRXs, possibly explaining the decreased catalytic activity at low pH with evolution. At a global level, we note that the population of low-flexibility (called hinges) and high-flexibility sites increases with evolution. The heterogeneity (quantified by the variance) in DFI distribution increases with the decrease in the melting temperature typically associated with the evolution of ancient proteins to their modern-day counterparts.This article is part of a discussion meeting issue 'Allostery and molecular machines'. © 2018 The Author(s).

Growth and consumption of L-malic acid in wine-like medium by acclimated and non-acclimated cultures of Patagonian Oenococcus oeni strains.

PubMed

Bravo-Ferrada, Bárbara Mercedes; Hollmann, Axel; Brizuela, Natalia; La Hens, Danay Valdés; Tymczyszyn, Elizabeth; Semorile, Liliana

2016-09-01

Five Oenococcus oeni strains, selected from spontaneous malolactic fermentation (MLF) of Patagonic Pinot noir wine, were assessed for their use as MLF starter cultures. After the individual evaluation of tolerance to some stress conditions, usually found in wine (pH, ethanol, SO2, and lysozyme), the behavior of the strains was analyzed in MLO broth with 14 % ethanol and pH 3.5 in order to test for the synergistic effect of high ethanol level and low pH and, finally, in a wine-like medium. Although the five strains were able to grow in MLO broth under low pH and/or high ethanol, they must be acclimated to grow in a wine-like medium. Additionally, glycosidase and tannase activities were evaluated, showing differences among the strains. The potential of the strains to ferment citrate was tested and two of the five strains showed the ability to metabolize this substrate. We did not detect the presence of genes encoding histidine, tyrosine descarboxylase, and putrescine carbamoyltransferase. All the strains tested exhibited good growth capacity and ability to consume L-malic acid in a wine-like medium after cell acclimation, and each of them showed a particular enzyme profile, which might confer different organoleptic properties to the wine.

Cellulase and cell differentiation in Acer pseudoplatanus.

PubMed

Sheldrake, A R

1970-06-01

Homogenates of differentiating xylem and phloem tissue have higher cellulase activities than cambial samples; the highest activity is always found in phloem. Callus tissue, in which no vascular differentiation occurs, contains only low cellulase activity. The results suggest that cellulase is involved in vascular differentiation. Different pH optima of cellulase activity were found: in cambium, xylem and phloem tissue, cellulase activity with an optimum at about pH 5.9 is predominantly membrane-bound; it is sedimentable at 100,000 g and releasable by Triton X-100. The same may be true of activity with an optimum at pH 5.3. Phloem tissue also contains a soluble, cytoplasmic cellulase of high activity at pH 7.1, and xylem tissue contains cytoplasmic cellulase with an optimum at pH 6.5. Low cellulase activity with a pH optimum similar to that of xylem homogenates was found in xylem sap. Cellulase activity in abscission zones increases greatly just before leaf abscission. Abscission zone cellulase has two pH optima, et 5.3 and 5.9; both activities are increased by Triton treatment of homogenates. The possible existence of several different cellulases forming part of a cellulase complex, and the rôle of the enzymes in hydrolysing wall material during cell differentiation are discussed.

Effect of mixing during fermentation in yogurt manufacturing.

PubMed

Aguirre-Ezkauriatza, E J; Galarza-González, M G; Uribe-Bujanda, A I; Ríos-Licea, M; López-Pacheco, F; Hernández-Brenes, C M; Alvarez, M M

2008-12-01

In traditional yogurt manufacturing, the yogurt is not agitated during fermentation. However, stirring could be beneficial, particularly for improving heat and mass transport across the fermentation tank. In this contribution, we studied the effect of low-speed agitation during fermentation on process time, acidity profile, and microbial dynamics during yogurt fermentation in 2 laboratory-scale fermenters (3 and 5 L) with different heat-transfer characteristics. Lactobacillus bulgaricus and Streptococcus thermophilus were used as fermenting bacteria. Curves of pH, lactic acid concentration, lactose concentration, and bacterial population profiles during fermentation are presented for static and low-agitation conditions during fermentation. At low-inoculum conditions, agitation reduced the processing time by shortening the lag phase. However, mixing did not modify the duration or the shape of the pH profiles during the exponential phase. In fermentors with poor heat-transfer characteristics, important differences in microbial dynamics were observed between the agitated and nonagitated fermentation experiments; that is, agitation significantly increased the observable specific growth rate and the final microbial count of L. bulgaricus.

Effects of coating layer and release medium on release profile from coated capsules with Eudragit FS 30D: an in vitro and in vivo study.

PubMed

Moghimipour, Eskandar; Rezaei, Mohsen; Kouchak, Maryam; Fatahiasl, Jafar; Angali, Kambiz Ahmadi; Ramezani, Zahra; Amini, Mohsen; Dorkoosh, Farid Abedin; Handali, Somayeh

2018-05-01

The aim of the present research was to evaluate the impact of coating layers on release profile from enteric coated dosage forms. Capsules were coated with Eudragit FS 30D using dipping method. The drug profile was evaluated in both phosphate buffer and Hank's solutions. Utilization X-ray imaging, gastrointestinal transmission of enteric coated capsules was traced in rats. According to the results, no release of the drug was found at pH 1.2, and the extent of release drug in pH 6.8 medium was decreased by adding the coating layers. The results indicated single-layer coated capsules in phosphate buffer were significantly higher than that in Hank's solution. However, no significant difference was observed from capsules with three coating layers in two different dissolution media. X-ray imaging showed that enteric coated capsules were intact in the stomach and in the small intestine, while disintegrated in the colon.

Clinical utility of pH paper versus pH meter in the measurement of critical gastric pH in stress ulcer prophylaxis.

PubMed

Bradley, J S; Phillips, J O; Cavanaugh, J E; Metzler, M H

1998-11-01

To evaluate the clinical utility of measuring gastric pH with a pH meter vs. pH paper in critical care patients. Prospective comparison of gastric pH measurements, using both pH meter and pH paper. Surgical intensive care unit (ICU) at a rural Midwestern university medical center. Fifty-one patients who received therapy for prophylaxis of stress ulcers in the surgical ICU. Therapy for stress ulcer prophylaxis was monitored. The pH of 985 gastric samples, taken from 51 patients, was measured with both pH meter and pH paper. The pH meter and pH paper measures demonstrated a concordance correlation coefficient of .896. The mean difference between the two measures (pH paper - pH meter) was estimated to be between -0.4 and 1.4, suggesting a positive bias for the paper. The prevalence of events representing clinically relevant differences between the pH meter and pH paper in the measurement of the same gastric sample was calculated. The frequency with which each of the events occurred consecutively (or, in one case, two nearly consecutive events on the same day) was also calculated. Bias in a clinically relevant range was estimated. A set of "probability profiles" was constructed. A hand-held pH meter and pH paper are not interchangeable measures of gastric pH. The pH paper exhibits an appreciable positive bias compared with a hand-held pH meter in the clinically relevant range of 2 to 6. More research is needed to determine if that bias affects treatment outcomes. We recommend the use of a pH meter for patients who demonstrate pH readings of < or = 4, consecutive with readings of < or = 5.

Effect of pH on growth and lipid accumulation kinetics of the microalga Chlorella vulgaris grown heterotrophically under sulfur limitation.

PubMed

Sakarika, Myrsini; Kornaros, Michael

2016-11-01

The goal of the present study was to investigate the pH range that can support the growth of C. vulgaris, and, more specifically, to identify the optimal pH for the microalga's growth, under heterotrophic conditions. Furthermore, the effect of pH on the accumulation of intracellular lipids was studied. A wide range of pH values was tested using the respective buffer solutions. The optimal pH for biomass growth and lipid accumulation under sulfur limitation was found to be 7.5, resulting in maximum specific growth rate of 0.541days(-1) and maximum total lipid content of 53.43%ggDW(-1). The fatty acid composition of C. vulgaris was found to be unrelated to pH, as the lipid content did not present significant variations in the pH values tested. The fatty acid profile was mainly composed of monounsaturated fatty acids (MUFAs) with the dominant one being oleic acid (C18:1). Copyright © 2016 Elsevier Ltd. All rights reserved.

Characterization of a Bacteriophage-Induced, Host-Specific Lytic Enzyme from Lysates of Bacillus stearothermophilus Infected with Bacteriophage TP-8

PubMed Central

Brehm, Sylvia P.; Welker, N. E.

1974-01-01

Phage TP-8 lysates of Bacillus stearothermophilus 4S or 4S(8) contain lytic activity exhibiting two pH optima, one at pH 6.5 and the other at pH 7.5. Using a variety of fractionation procedures, the two lytic activities could not be separated. At pH 7.5 the lytic enzyme is an endopeptidase which hydrolyzes the l-alanyl-d-glutamyl linkage in the peptide subunits of the cell wall peptidoglycan and at pH 6.5 it exhibits N-acetylmuramidase activity. Endopeptidase activity is inhibited by NaCl and neither lytic activity was significantly affected by divalent cations or ethylenediaminetetraacetic acid. Crude lysates contain 2.5 to 3.0 times more endopeptidase activity than N-acetylmuramidase activity. The ratio of the two lytic activities (endopeptidase/N-acetylmuramidase) changes to 1.3 to 1.7 during the course of purification, to 1.0 after isoelectric focusing, and 3.9 and 6.00 after exposure for 2 h at 60 and 65 C, respectively. We conclude that the two lytic activities may be associated with a single protein or a lytic enzyme complex composed of two enzymes. Lytic activity at pH 7.5 is more effective in solubilizing cells or cell walls than the lytic activity at pH 6.5. LiCl extracts of 4S and 4S(8) cells contain lytic activity exhibiting endopeptidase activity at pH 7.5 and N-acetylmuramidase activity at pH 6.5. Lytic activity in these LiCl extracts also has a number of other properties in common with those in lysates of phage TP-8. We proposed that the lytic enzyme(s) are not coded for by the phage genome but are part of the host autolytic system. PMID:4218232

Adsorption of aqueous Cd(II) and Pb(II) on activated carbon nanopores prepared by chemical activation of doum palm shell.

PubMed

Gaya, Umar Ibrahim; Otene, Emmanuel; Abdullah, Abdul Halim

2015-01-01

Non-uniformly sized activated carbons were derived from doum palm shell, a new precursor, by carbonization in air and activation using KOH, NaOH and ZnCl2. The activated carbon fibres were characterised by X-ray diffraction, N2 adsorption-desorption, scanning electron microscopy, particle size analysis and evaluated for Cd(II) and Pb(II) removal. The 40-50 nm size, less graphitic, mesoporous NaOH activated carbon yielded high adsorption efficiency, pointing largely to the influence surface area. The performance of the KOH based activated carbon was arguably explained for the first time in terms of crystallinity. The efficiencies of the mesoporous ZnCl2-formulated activated carbon diminished due to the presence of larger particles. Batch adsorption of divalent metals revealed dependence on adsorbent dose, agitation time, pH and adsorbate concentrations with high adsorption efficiencies at optimum operating parameters. The equilibrium profiles fitted Langmuir and Freundlich isotherms, and kinetics favoured pseudo-second order model. The study demonstrated the practicability of the removal of alarming levels of cadmium and lead ions from industrial effluents.

Effect of pH on structure, function, and stability of mitochondrial carbonic anhydrase VA.

PubMed

Idrees, Danish; Shahbaaz, Mohd; Bisetty, Krishna; Islam, Asimul; Ahmad, Faizan; Hassan, Md Imtaiyaz

2017-02-01

Mitochondrial carbonic anhydrase VA (CAVA) catalyzes the hydration of carbon dioxide to produce proton and bicarbonate which is primarily expressed in the mitochondrial matrix of liver, and involved in numerous physiological processes including lipogenesis, insulin secretion from pancreatic cells, ureagenesis, gluconeogenesis, and neuronal transmission. To understand the effect of pH on the structure, function, and stability of CAVA, we employed spectroscopic techniques such as circular dichroism, fluorescence, and absorbance measurements in wide range of pH (from pH 2.0 to pH 11.5). CAVA showed an aggregation at acidic pH range from pH 2.0 to pH 5.0. However, it remains stable and maintains its secondary structure in the pH range, pH 7.0-pH 11.5. Furthermore, this enzyme has an appreciable activity at more than pH 7.0 (7.0 < pH ≤ 11.5) with maximum activity at pH 9.0. The maximal values of k cat and k cat /K m at pH 9.0 are 3.7 × 10 6  s -1 and 5.5 × 10 7  M -1  s -1 , respectively. However, this enzyme loses its activity in the acidic pH range. We further performed 20-ns molecular dynamics simulation of CAVA to see the dynamics at different pH values. An excellent agreement was observed between in silico and in vitro studies. This study provides an insight into the activity of CAVA in the pH range of subcellular environment.

Lime retention in anthracite coal-breaker refuse

Treesearch

Miroslaw M. Czapowskyj; Edward A. Sowa

1973-01-01

Hydrated lime was applied to extremely acid anthracite coal-breaker refuse at rates of 2.5 and 5.0 tons per acre. The lime raised the pH to neutral range, and this range was still in evidence 7 years after treatment. The pH readings decreased with the depth of the refuse profile, and below 9 inches they approximated those of the control plots. The 2.5-tons-of-lime-per-...

Understanding the spatial formation and accumulation of fats, oils and grease deposits in the sewer collection system.

PubMed

Dominic, Christopher Cyril Sandeep; Szakasits, Megan; Dean, Lisa O; Ducoste, Joel J

2013-01-01

Sanitary sewer overflows are caused by the accumulation of insoluble calcium salts of fatty acids, which are formed by the reaction between fats, oils and grease (FOG) and calcium found in wastewaters. Different sewer structural configurations (i.e., manholes, pipes, wet wells), which vary spatially, along with other obstructions (roots intrusion) and pipe deformations (pipe sags), may influence the detrimental buildup of FOG deposits. The purpose of this study was to quantify the spatial variation in FOG deposit formation and accumulation in a pilot-scale sewer collection system. The pilot system contained straight pipes, manholes, roots intrusion, and a pipe sag. Calcium and oil were injected into the system and operated at alkaline (pH = 10) and neutral (pH = 7) pH conditions. Results showed that solid accumulations were slightly higher at neutral pH. Fourier transform infrared (FTIR) analysis on the solids samples confirmed that the solids were indeed calcium-based fatty acid salts. However, the fatty acid profiles of the solids deviated from the profile found from FOG deposits in sewer systems, which were primarily saturated fatty acids. These results confirm the work done previously by researchers and suggest an alternative fate of unsaturated fatty acids that does not lead to their incorporation in FOG deposits in full-scale sewer systems.

Production of a bacteriocin-like inhibitory substance by Leuconostoc mesenteroides subsp. dextranicum 213M0 isolated from Mongolian fermented mare milk, airag.

PubMed

Arakawa, Kensuke; Yoshida, Saki; Aikawa, Hiroki; Hano, Chihiro; Bolormaa, Tsognemekh; Burenjargal, Sedkhuu; Miyamoto, Taku

2016-03-01

Strain 213M0 was selected with productivity of a bacteriocin-like inhibitory substance (BLIS) among 235 strains of lactic acid bacteria (LAB) isolated from Mongolian fermented milk 'airag'. Strain 213M0 was species-identified as Leuconostoc mesenteroides subsp. dextranicum by morphological observation, carbohydrate fermentation profiling and sequencing the 16S rRNA gene. Incubation temperature proper to produce the BLIS was 25°C rather than 30 and 37°C, and the production actively proceeded during the exponential growth phase of the producer cells. Antibacterial effect of BLIS 213M0 was limited to all nine strains of Listeria sp. bacteria and seven strains of LAB cocci among 53 tested strains, which corresponds to a typical feature of the class IIa pediocin-like bacteriocins. BLIS 213M0 was not inactivated in every broad pH range solution (pH 2.0-11.0), and was stable against storage at 25°C for 1 week and heating at 121°C for 15 min under pH 4.5. Peptide frame of BLIS 213M0 was confirmed by inactivation with some peptidases, and then its molecular weight was estimated to be 2.6-3.0 kDa using an in situ activity assay following sodium dodecyl sulfate polyacrylamide gel electrophoresis. The estimated size was different from the other Leuconostoc bacteriocins already reported. These results suggest that BLIS 213M0 would be a novel listericidal bacteriocin. © 2015 Japanese Society of Animal Science.

Surface acidity and solid-state compatibility of excipients with an acid-sensitive API: case study of atorvastatin calcium.

PubMed

Govindarajan, Ramprakash; Landis, Margaret; Hancock, Bruno; Gatlin, Larry A; Suryanarayanan, Raj; Shalaev, Evgenyi Y

2015-04-01

The objectives of this study were to measure the apparent surface acidity of common excipients and to correlate the acidity with the chemical stability of an acid-sensitive active pharmaceutical ingredient (API) in binary API-excipient powder mixtures. The acidity of 26 solid excipients was determined by two methods, (i) by measuring the pH of their suspensions or solutions and (ii) the pH equivalent (pHeq) measured via ionization of probe molecules deposited on the surface of the excipients. The chemical stability of an API, atorvastatin calcium (AC), in mixtures with the excipients was evaluated by monitoring the appearance of an acid-induced degradant, atorvastatin lactone, under accelerated storage conditions. The extent of lactone formation in AC-excipient mixtures was presented as a function of either solution/suspension pH or pHeq. No lactone formation was observed in mixtures with excipients having pHeq > 6, while the lactone levels were pronounced (> 0.6% after 6 weeks at 50°C/20% RH) with excipients exhibiting pHeq < 3. The three pHeq regions (> 6, 3-6, and < 3) were consistent with the reported solution pH-stability profile of AC. In contrast to the pHeq scale, lactone formation did not show any clear trend when plotted as a function of the suspension/solution pH. Two mechanisms to explain the discrepancy between the suspension/solution pH and the chemical stability data were discussed. Acidic excipients, which are expected to be incompatible with an acid-sensitive API, were identified based on pHeq measurements. The incompatibility prediction was confirmed in the chemical stability tests using AC as an example of an acid-sensitive API.

Proof of concept of a "greener" protein purification/enrichment method based on carboxylate-terminated carbosilane dendrimer-protein interactions.

PubMed

González-García, Estefanía; Maly, Marek; de la Mata, Francisco Javier; Gómez, Rafael; Marina, María Luisa; García, María Concepción

2016-11-01

Protein sample preparation is a critical and an unsustainable step since it involves the use of tedious methods that usually require high amount of solvents. The development of new materials offers additional opportunities in protein sample preparation. This work explores, for the first time, the potential application of carboxylate-terminated carbosilane dendrimers to the purification/enrichment of proteins. Studies on dendrimer binding to proteins, based on protein fluorescence intensity and emission wavelengths measurements, demonstrated the interaction between carboxylate-terminated carbosilane dendrimers and proteins at all tested pH levels. Interactions were greatly affected by the protein itself, pH, and dendrimer concentration and generation. Especially interesting was the interaction at acidic pH since it resulted in a significant protein precipitation. Dendrimer-protein interactions were modeled observing stable complexes for all proteins. Carboxylate-terminated carbosilane dendrimers at acidic pH were successfully used in the purification/enrichment of proteins extracted from a complex sample. Graphical Abstract Images showing the growing turbidity of solutions containing a mixture of proteins (lysozyme, myoglobin, and BSA) at different protein:dendrimer ratios (1:0, 1:1, 1:8, and 1:20) at acidic pH and SDS-PAGE profiles of the corresponsing supernatants. Comparison of SDS-PAGE profiles for the pellets obtained during the purification of proteins present in a complex sample using a conventional "no-clean" method based on acetone precipitation and the proposed "greener" method using carboxylate-terminated carbosilane dendrimer at a 1:20 protein:dendrimer ratio.

Computational studies of the binding profile of phosphoinositide PtdIns (3,4,5) P3 with the pleckstrin homology domain of an oomycete cellulose synthase

PubMed Central

Kuang, Guanglin; Bulone, Vincent; Tu, Yaoquan

2016-01-01

Saprolegnia monoica is a model organism to investigate Saprolegnia parasitica, an important oomycete which causes considerable loss in aquaculture every year. S. monoica contains cellulose synthases vital for oomycete growth. However, the molecular mechanism of the cellulose biosynthesis process in the oomycete growth is still poorly understood. Some cellulose synthases of S. monoica, such as SmCesA2, are found to contain a plecsktrin homology (PH) domain, which is a protein module widely found in nature and known to bind to phosphoinositides, a class of signaling compounds involved in many biological processes. Understanding the molecular interactions between the PH domain and phosphoinositides would help to unravel the cellulose biosynthesis process of oomycetes. In this work, the binding profile of PtdIns (3,4,5) P3, a typical phosphoinositide, with SmCesA2-PH was studied by molecular docking, molecular dynamics and metadynamics simulations. PtdIns (3,4,5) P3 is found to bind at a specific site located at β1, β2 and β1-β2 loop of SmCesA2-PH. The high affinity of PtdIns (3,4,5) P3 to SmCesA2-PH is contributed by the free phosphate groups, which have electrostatic and hydrogen-bond interactions with Lys88, Lys100 and Arg102 in the binding site. PMID:26857031

Dynamic changes in water and salinity in saline-alkali soils after simulated irrigation and leaching.

PubMed

Wang, Shutao; Feng, Qian; Zhou, Yapeng; Mao, Xiaoxi; Chen, Yaheng; Xu, Hao

2017-01-01

Soil salinization is a global problem that limits agricultural development and impacts human life. This study aimed to understand the dynamic changes in water and salinity in saline-alkali soil based on an indoor soil column simulation. We studied the changes in the water and salt contents of soils with different degrees of salinization under various irrigation conditions. The results showed that after seven irrigations, the pH, conductivity and total soluble salt content of the percolation samples after irrigation generally increased initially then decreased with repeated irrigation. The soil moisture did not change significantly after irrigation. The pH, conductivity, and total soluble salt content of each layer of the soil profile exhibited general declining trends. In the soil profile from Changguo Township (CG), the pH decreased from 8.21-8.35 to 7.71-7.88, the conductivity decreased from 0.95-1.14 ms/cm to 0.45-0.68 ms/cm, and the total soluble salt content decreased from 2.63-2.81 g/kg to 2.28-2.51 g/kg. In the soil profile from Zhongjie Industrial Park (ZJ), the pH decreased from 8.36-8.54 to 7.73-7.96, the conductivity decreased from 1.58-1.68 ms/cm to 1.45-1.54 ms/cm, and the total soluble salt decreased from 2.81-4.03 g/kg to 2.56-3.28 g/kg. The transported salt ions were primarily K+, Na+ and Cl-. After several irrigations, a representative desalination effect was achieved. The results of this study can provide technical guidance for the comprehensive management of saline-alkali soils.

Dynamic changes in water and salinity in saline-alkali soils after simulated irrigation and leaching

PubMed Central

Feng, Qian; Mao, Xiaoxi

2017-01-01

Soil salinization is a global problem that limits agricultural development and impacts human life. This study aimed to understand the dynamic changes in water and salinity in saline-alkali soil based on an indoor soil column simulation. We studied the changes in the water and salt contents of soils with different degrees of salinization under various irrigation conditions. The results showed that after seven irrigations, the pH, conductivity and total soluble salt content of the percolation samples after irrigation generally increased initially then decreased with repeated irrigation. The soil moisture did not change significantly after irrigation. The pH, conductivity, and total soluble salt content of each layer of the soil profile exhibited general declining trends. In the soil profile from Changguo Township (CG), the pH decreased from 8.21–8.35 to 7.71–7.88, the conductivity decreased from 0.95–1.14 ms/cm to 0.45–0.68 ms/cm, and the total soluble salt content decreased from 2.63–2.81 g/kg to 2.28–2.51 g/kg. In the soil profile from Zhongjie Industrial Park (ZJ), the pH decreased from 8.36–8.54 to 7.73–7.96, the conductivity decreased from 1.58–1.68 ms/cm to 1.45–1.54 ms/cm, and the total soluble salt decreased from 2.81–4.03 g/kg to 2.56–3.28 g/kg. The transported salt ions were primarily K+, Na+ and Cl-. After several irrigations, a representative desalination effect was achieved. The results of this study can provide technical guidance for the comprehensive management of saline-alkali soils. PMID:29091963

The effect of pH, buffer capacity and ionic strength on quetiapine fumarate release from matrix tablets prepared using two different polymeric blends.

PubMed

Hamed, Rania; AlJanabi, Reem; Sunoqrot, Suhair; Abbas, Aiman

2017-08-01

The objective of this study was to investigate the effect of the different physiological parameters of the gastrointestinal (GI) fluid (pH, buffer capacity, and ionic strength) on the in vitro release of the weakly basic BCS class II drug quetiapine fumarate (QF) from two once-a-day matrix tablet formulations (F1 and F2) developed as potential generic equivalents to Seroquel ® XR. F1 tablets were prepared using blends of high and low viscosity grades of hydroxypropyl methylcellulose (HPMC K4M and K100LV, respectively), while F2 tablets were prepared from HPMC K4M and PEGylated glyceryl behenate (Compritol ® HD5 ATO). The two formulations attained release profiles of QF over 24 h similar to that of Seroquel ® XR using the dissolution medium published by the Food and Drug Administration (FDA). A series of solubility and in vitro dissolution studies was then carried out using media that simulate the gastric and intestinal fluids and cover the physiological pH, buffer capacity and ionic strength range of the GIT. Solubility studies revealed that QF exhibits a typical weak base pH-dependent solubility profile and that the solubility of QF increases with increasing the buffer capacity and ionic strength of the media. The release profiles of QF from F1, F2 and Seroquel ® XR tablets were found to be influenced by the pH, buffer capacity and ionic strength of the dissolution media to varying degrees. Results highlight the importance of studying the physiological variables along the GIT in designing controlled release formulations for more predictive in vitro-in vivo correlations.

The hydrolysis kinetics of monobasic and dibasic aminoalkyl esters of ketorolac.

PubMed

Qandil, Amjad M; Jamhawi, Noor M; Tashtoush, Bassam M; Al-Ajlouni, Ahmad M; Idkaidek, Nasir M; Obaidat, Aiman A

2013-09-01

Six aminoethyl and aminobutyl esters of ketorolac containing 1-methylpiperazine (MPE and MPB), N-acetylpiperazine (APE and APB) or morpholine (ME and MB), were synthesized and their hydrolysis kinetics were studied. The hydrolysis was studied at pH 1 to 9 (for MPE, APE and ME) and pH 1 to 8 (for MPB, APB and MB) in aqueous phosphate buffer (0.16 M) with ionic strength (0.5 M) at 37°C. Calculation of k(obs), construction of the pH-rate profiles and determination of the rate equations were performed using KaleidaGraph® 4.1. The hydrolysis displays pseudo-first order kinetics and the pH-rate profiles shows that the aminobutyl esters, MPE, APB and MB, are the most stable. The hydrolysis of the ethyl esters MPE, APE and ME, depending on the pH, is either fast and catalyzed by the hydroxide anion or slow and uncatalyzed for the diprotonated, monoprotonated and nonprotonated forms. The hydrolysis of the butyl esters showed a similar profile, albeit it was also catalyzed by hydronium cation. In addition, the hydroxide anion is 105 more effective in catalyzing the hydrolysis than the hydronium cation. The hydrolysis pattern of the aminoethyl esters is affected by the number and pKa of its basic nitrogen atoms. The monobasic APE and ME, show a similar hydrolysis pattern that is different than the dibasic MPE. The length of the side chain and the pKa of the basic nitrogen atoms in the aminoethyl moiety affect the mechanism of hydrolysis as the extent of protonation at a given pH is directly related to the pKa.

Fermentation pH influences the physiological-state dynamics of Lactobacillus bulgaricus CFL1 during pH-controlled culture.

PubMed

Rault, Aline; Bouix, Marielle; Béal, Catherine

2009-07-01

This study aims at better understanding the effects of fermentation pH and harvesting time on Lactobacillus bulgaricus CFL1 cellular state in order to improve knowledge of the dynamics of the physiological state and to better manage starter production. The Cinac system and multiparametric flow cytometry were used to characterize and compare the progress of the physiological events that occurred during pH 6 and pH 5 controlled cultures. Acidification activity, membrane damage, enzymatic activity, cellular depolarization, intracellular pH, and pH gradient were determined and compared during growing conditions. Strong differences in the time course of viability, membrane integrity, and acidification activity were displayed between pH 6 and pH 5 cultures. As a main result, the pH 5 control during fermentation allowed the cells to maintain a more robust physiological state, with high viability and stable acidification activity throughout growth, in opposition to a viability decrease and fluctuation of activity at pH 6. This result was mainly explained by differences in lactate concentration in the culture medium and in pH gradient value. The elevated content of the ionic lactate form at high pH values damaged membrane integrity that led to a viability decrease. In contrast, the high pH gradient observed throughout pH 5 cultures was associated with an increased energetic level that helped the cells maintain their physiological state. Such results may benefit industrial starter producers and fermented-product manufacturers by allowing them to better control the quality of their starters, before freezing or before using them for food fermentation.

Hybrid hard- and soft-modeling of spectrophotometric data for monitoring of ciprofloxacin and its main photodegradation products at different pH values

NASA Astrophysics Data System (ADS)

Razuc, Mariela; Garrido, Mariano; Caro, Yamile S.; Teglia, Carla M.; Goicoechea, Héctor C.; Fernández Band, Beatriz S.

2013-04-01

A simple and fast on line spectrophotometric method combined with a hybrid hard-soft modeling multivariate curve resolution (HS-MCR) was proposed for the monitoring of photodegradation reaction of ciprofloxacin under UV radiation. The studied conditions attempt to emulate the effect of sunlight on these antibiotics that could be eventually present in the environment. The continuous flow system made it possible to study the ciprofloxacin degradation at different pH values almost at real time, avoiding errors that could arise from typical batch monitoring of the reaction. On the base of a concentration profiles obtained by previous pure soft-modeling approach, reaction pathways have been proposed for the parent compound and its photoproducts at different pH values. These kinetic models were used as a constraint in the HS-MCR analysis. The kinetic profiles and the corresponding pure response profile (UV-Vis spectra) of ciprofloxacin and its main degradation products were recovered after the application of HS-MCR analysis to the spectra recorded throughout the reaction. The observed behavior showed a good agreement with the photodegradation studies reported in the bibliography. Accordingly, the photodegradation reaction was studied by high performance liquid chromatography coupled with UV-Vis diode array detector (HPLC-DAD). The spectra recorded during the chromatographic analysis present a good correlation with the ones recovered by UV-Vis/HS-MCR method.

Short-column anion-exchange chromatography for soil and peat humic substances profiling by step-wise gradient of high pH aqueous sodium ethylenediaminetetraacetate.

PubMed

Hutta, Milan; Ráczová, Janka; Góra, Róbert; Pessl, Juraj

2015-08-21

Novel anion-exchange liquid chromatographic method with step gradient of aqueous EDTA(4-) based mobile phase elution has been developed to profile available Slovak soil humic substances and alkaline extracts of various soils. The method utilize short glass column (30mm×3mm) filled in with hydrolytically stable particles (60μm diameter) Separon HEMA-BIO 1000 having (diethylamino)ethyl functional groups. Step gradient was programmed by mixing mobile phase composed of aqueous solution of sodium EDTA (pH 12.0; 5mmolL(-1)) and mobile phase constituted of aqueous solution of sodium EDTA (pH 12.0, 500mmolL(-1)). The FLD of HSs was set to excitation wavelength 480nm and emission wavelength 530nm (λem). Separation mechanism was studied by use of selected aromatic acids related to humic acids with the aid of UV spectrophotometric detection at 280nm. The proposed method benefits from high ionic strength (I=5molL(-1)) of the end mobile phase buffer and provides high recovery of humic acids (98%). Accurate and reproducible profiling of studied humic substances, alkaline extracts of various types of soils enables straightforward characterization and differentiation of HSs in arable and forest soils. Selected model aromatic acids were used for separation mechanism elucidation. Copyright © 2015 Elsevier B.V. All rights reserved.

[Effects of root-knot nematodes on cucumber leaf N and P contents, soil pH, and soil enzyme activities].

PubMed

Xu, Hua; Ruan, Wei-Bin; Gao, Yu-Bao; Song, Xiao-Yan; Wei, Yu-Kun

2010-08-01

A pot experiment was conducted to study the effects of inoculation with root-knot nematodes on the cucumber leaf N and P contents, and the rhizospheric and non-rhizospheric soil pH and enzyme activities. The rhizospheric soil pH didn't have a significant decrease until the inoculation rate reached 6000 eggs per plant. With the increase of inoculation rate, the leaf N and P contents, rhizospheric soil peroxidase activity, and rhizospheric and non-rhizospheric soil polyphenol oxidase activity all decreased gradually, rhizospheric soil catalase activity was in adverse, non-rhizospheric soil pH decreased after an initial increase, and non-rhizospheric soil catalase activity had no regular change. After inoculation, rhizospheric soil urease activity decreased significantly, but rhizospheric and non-rhizospheric soil phosphatase activity and non-rhizospheric soil peroxidase activity only had a significant decrease under high inoculation rate. In most cases, there existed significant correlations between rhizospheric soil pH, enzyme activities, and leaf N and P contents; and in some cases, there existed significant correlations between non-rhizospheric soil pH, enzyme activities, and leaf N and P contents.

Development of a Physiologically Relevant Population Pharmacokinetic in Vitro-in Vivo Correlation Approach for Designing Extended-Release Oral Dosage Formulation.

PubMed

Kim, Tae Hwan; Shin, Soyoung; Bulitta, Jürgen B; Youn, Yu Seok; Yoo, Sun Dong; Shin, Beom Soo

2017-01-03

Establishing a level A in vitro-in vivo correlation (IVIVC) for a drug with complex absorption kinetics is challenging. The objective of the present study was to develop an IVIVC approach based on population pharmacokinetic (POP-PK) modeling that incorporated physiologically relevant absorption kinetics. To prepare three extended release (ER) tablets of loxoprofen, three types of hydroxypropyl methylcellulose (HPMC 100, 4000, and 15000 cps) were used as drug release modifiers, while lactose and magnesium stearate were used as the diluent and lubricant, respectively. An in vitro dissolution test in various pH conditions showed that loxoprofen dissolution was faster at higher pH. The in vivo pharmacokinetics of loxoprofen was assessed following oral administration of the different loxoprofen formulations to Beagle dogs (n = 22 in total). Secondary peaks or shoulders were observed in many of the individual plasma concentration vs time profiles after ER tablet administration, which may result from secondary absorption in the intestine due to a dissolution rate increase under intestinal pH compared to that observed at stomach pH. In addition, in vivo oral bioavailability was found to decrease with prolonged drug dissolution, indicating site-specific absorption. Based on the in vitro dissolution and in vivo absorption data, a POP-PK IVIVC model was developed using S-ADAPT software. pH-dependent biphasic dissolution kinetics, described using modified Michaelis-Menten kinetics with varying V max , and site-specific absorption, modeled using a changeable absorbed fraction parameter, were applied to the POP-PK IVIVC model. To experimentally determine the biphasic dissolution profiles of the ER tablets, another in vitro dissolution test was conducted by switching dissolution medium pH based on an in vivo estimate of gastric emptying time. The model estimated, using linear regression, that in vivo initial maximum dissolution rate (V max (0) in vivo ) was highly correlated (r 2 > 0.998) with in vitro (V max (0) in vitro ), indicating that in vivo dissolution profiles obtained from POP-PK modeling could be converted to in vitro dissolution profiles and vice versa. Monte Carlo simulations were performed for model validation, and prediction errors for C max and AUC were all within the acceptable range (90 to 110%) according to the FDA guidelines. The developed model was successfully applied for the prediction of in vivo pharmacokinetics of a loxoprofen double-layered tablet using the in vitro dissolution profile. In conclusion, a level A IVIVC approach was developed and validated using population modeling that accounted for pH-dependent dissolution and site-specific absorption. Excellent correlations were observed between in vitro and in vivo dissolution profiles. This new approach holds great promise for the establishment of IVIVCs for drug and formulation development where absorption kinetics strongly depend on complex physiologically absorption processes.

Phenolic contents, antioxidant and anticholinesterase potentials of crude extract, subsequent fractions and crude saponins from Polygonum hydropiper L

PubMed Central

2014-01-01

Background We investigated Polygonum hydropiper L. (P. hydropiper) for phenolic contents, antioxidant, anticholinesterase activities, in an attempt to rationalize its use in neurological disorders. Methods Plant crude extract (Ph.Cr), its subsequent fractions: n-hexane (Ph.Hex), chloroform (Ph.Chf), ethyl acetate (Ph.EtAc), n-Butanol (Ph.Bt), aqueous (Ph.Aq) and saponins (Ph.Sp) were evaluated for 1,1-diphenyl,2-picrylhydrazyl (DPPH), 2,2-azinobis[3-ethylbenzthiazoline]-6-sulfonic acid (ABTS) free radical scavenging potential. Further, acetylcholinesterase (AChE) & butyrylcholinesterase (BChE) inhibitory activities were performed using Ellman's assay. Moreover, total phenolic contents of plant extracts were determined and expressed in mg of gallic acid equivalent per gram of dry sample (mg GAE/g dry weight). Results Among different fractions, Ph.Cr (90.82), Ph.Chf (178.16), Ph.EtAc (203.44) and Ph.Bt (153.61) exhibited high phenolic contents. All fractions showed concentration dependent DPPH scavenging activity, with Ph.EtAc 71.33% (IC50 15 μg/ml), Ph.Bt 71.40% (IC50 3 μg/ml) and Ph.Sp 71.40% (IC50 35 μg/ml) were most potent. The plant extracts exhibited high ABTS scavenging ability i.e. Ph.Bt (91.03%), Ph.EtAc (90.56%), Ph.Sp (90.84%), Ph.Aq (90.56%) with IC50 < 0.01 μg/ml. All fractions showed moderate to high AChE inhibitory activity as; Ph.Cr, 86.87% (IC50 330 μg/ml), Ph.Hex, 87.49% (IC50 35 μg/ml), Ph.Chf, 84.76% (IC50 55 μg/ml), Ph.Sp, 87.58% (IC50 108 μg/ml) and Ph.EtAc 79.95% (IC50 310 μg/ml) at 1 mg/ml). Furthermore the BChE inhibitory activity was most prominent in Ph.Hex 90.30% (IC50 40 μg/ml), Ph.Chf 85.94% (IC50 215 μg/ml), Ph.Aq 87.62% (IC50 3 μg/ml) and Ph.EtAc 81.01% (IC50 395 μg/ml) fractions. Conclusions In this study, for the first time, we determined phenolic contents, isolated crude saponins, investigated antioxidant and anticholinestrase potential of P. hydropiper extracts. The results indicate that P. hydropiper is enriched with potent bioactive compounds and warrant further investigation by isolation and structural elucidation to find novel and affordable compounds for the treatment of various neurological disorders. PMID:24884823

Phenolic contents, antioxidant and anticholinesterase potentials of crude extract, subsequent fractions and crude saponins from Polygonum hydropiper L.

PubMed

Ayaz, Muhammad; Junaid, Muhammad; Ahmed, Jawad; Ullah, Farhat; Sadiq, Abdul; Ahmad, Sajjad; Imran, Muhammad

2014-05-03

We investigated Polygonum hydropiper L. (P. hydropiper) for phenolic contents, antioxidant, anticholinesterase activities, in an attempt to rationalize its use in neurological disorders. Plant crude extract (Ph.Cr), its subsequent fractions: n-hexane (Ph.Hex), chloroform (Ph.Chf), ethyl acetate (Ph.EtAc), n-Butanol (Ph.Bt), aqueous (Ph.Aq) and saponins (Ph.Sp) were evaluated for 1,1-diphenyl,2-picrylhydrazyl (DPPH), 2,2-azinobis[3-ethylbenzthiazoline]-6-sulfonic acid (ABTS) free radical scavenging potential. Further, acetylcholinesterase (AChE) & butyrylcholinesterase (BChE) inhibitory activities were performed using Ellman's assay. Moreover, total phenolic contents of plant extracts were determined and expressed in mg of gallic acid equivalent per gram of dry sample (mg GAE/g dry weight). Among different fractions, Ph.Cr (90.82), Ph.Chf (178.16), Ph.EtAc (203.44) and Ph.Bt (153.61) exhibited high phenolic contents. All fractions showed concentration dependent DPPH scavenging activity, with Ph.EtAc 71.33% (IC50 15 μg/ml), Ph.Bt 71.40% (IC50 3 μg/ml) and Ph.Sp 71.40% (IC50 35 μg/ml) were most potent. The plant extracts exhibited high ABTS scavenging ability i.e. Ph.Bt (91.03%), Ph.EtAc (90.56%), Ph.Sp (90.84%), Ph.Aq (90.56%) with IC50<0.01 μg/ml. All fractions showed moderate to high AChE inhibitory activity as; Ph.Cr, 86.87% (IC50 330 μg/ml), Ph.Hex, 87.49% (IC50 35 μg/ml), Ph.Chf, 84.76% (IC50 55 μg/ml), Ph.Sp, 87.58% (IC50 108 μg/ml) and Ph.EtAc 79.95% (IC50 310 μg/ml) at 1 mg/ml). Furthermore the BChE inhibitory activity was most prominent in Ph.Hex 90.30% (IC50 40 μg/ml), Ph.Chf 85.94% (IC50 215 μg/ml), Ph.Aq 87.62% (IC50 3 μg/ml) and Ph.EtAc 81.01% (IC50 395 μg/ml) fractions. In this study, for the first time, we determined phenolic contents, isolated crude saponins, investigated antioxidant and anticholinestrase potential of P. hydropiper extracts. The results indicate that P. hydropiper is enriched with potent bioactive compounds and warrant further investigation by isolation and structural elucidation to find novel and affordable compounds for the treatment of various neurological disorders.

Antitumour, antimicrobial and catalytic activity of gold nanoparticles synthesized by different pH propolis extracts

NASA Astrophysics Data System (ADS)

Gatea, Florentina; Teodor, Eugenia Dumitra; Seciu, Ana-Maria; Covaci, Ovidiu Ilie; Mănoiu, Sorin; Lazăr, Veronica; Radu, Gabriel Lucian

2015-07-01

The Romanian propolis was extracted in five different media, respectively, in water (pH 6.8), glycine buffer (pH 2.5), acetate buffer (pH 5), phosphate buffer (pH 7.4) and carbonate buffer (pH 9.2). The extracts presented different amounts of flavonoids and phenolic acids, increasing pH leading to higher concentrations of active compounds. Five variants of gold nanoparticles suspensions based on different pH Romanian propolis aqueous extracts were successfully synthesized. The obtained nanoparticles presented dimensions between 20 and 60 nm in dispersion form and around 18 nm in dried form, and different morphologies (spherical, hexagonal, triangular). Fourier transform infrared spectroscopy proved the attachment of organic compounds from propolis extracts to the colloidal gold suspensions and X-ray diffraction certified that the suspensions contain metallic gold. The obtained propolis gold nanoparticles do not exhibit any antibacterial or antifungal activity, but presented different catalytic activities and toxicity on tumour cells.

Comparative reactivity of TpRu(L)(NCMe)Ph (L = CO or PMe3): impact of ancillary ligand l on activation of carbon-hydrogen bonds including catalytic hydroarylation and hydrovinylation/oligomerization of ethylene.

PubMed

Foley, Nicholas A; Lail, Marty; Lee, John P; Gunnoe, T Brent; Cundari, Thomas R; Petersen, Jeffrey L

2007-05-30

Complexes of the type TpRu(L)(NCMe)R [L = CO or PMe3; R = Ph or Me; Tp = hydridotris(pyrazolyl)borate] initiate C-H activation of benzene. Kinetic studies, isotopic labeling, and other experimental evidence suggest that the mechanism of benzene C-H activation involves reversible dissociation of acetonitrile, reversible benzene coordination, and rate-determining C-H activation of coordinated benzene. TpRu(PMe3)(NCMe)Ph initiates C-D activation of C6D6 at rates that are approximately 2-3 times more rapid than that for TpRu(CO)(NCMe)Ph (depending on substrate concentration); however, the catalytic hydrophenylation of ethylene using TpRu(PMe3)(NCMe)Ph is substantially less efficient than catalysis with TpRu(CO)(NCMe)Ph. For TpRu(PMe3)(NCMe)Ph, C-H activation of ethylene, to ultimately produce TpRu(PMe3)(eta3-C4H7), is found to kinetically compete with catalytic ethylene hydrophenylation. In THF solutions containing ethylene, TpRu(PMe3)(NCMe)Ph and TpRu(CO)(NCMe)Ph separately convert to TpRu(L)(eta3-C4H7) (L = PMe3 or CO, respectively) via initial Ru-mediated ethylene C-H activation. Heating mesitylene solutions of TpRu(L)(eta3-C4H7) under ethylene pressure results in the catalytic production of butenes (i.e., ethylene hydrovinylation) and hexenes.

Fermentation of sarshir (kaymak) by lactic acid bacteria: antibacterial activity, antioxidant properties, lipid and protein oxidation and fatty acid profile.

PubMed

Hashemi, Seyed Mohammad Bagher; Mousavi Khaneghah, Amin; Kontominas, Michael G; Eş, Ismail; Sant'Ana, Anderson S; Martinez, Rafael R; Drider, Djamel

2017-10-01

The antioxidant and antibacterial activities of fermented sarshir (traditional dairy food), with three probiotic Lactobacillus plantarum strains (LP3, AF1, and LU5), were investigated. The oxidative stability and the lipid profile of non-fermented and fermented sarshir were compared, in addition to radical scavenging activity, as well as peroxide, anisidine and carbonyl values (PV, AnV and CV, respectively). The strong antibacterial activity of fermented sarshir against common pathogenic bacteria, including Gram-negative Escherichia coli O157: H7 ATCC 35150 and Pseudomonas aeruginosa ATCC 27853, as well as Gram-positive Bacillus cereus ATCC 10876 and Staphylococcus aureus ATCC 25923, was established. Among the strains examined, L. plantarum LP3 exhibited the highest radical scavenging activity (53.1 ± 1.8%) and lowest PV (3.0 meq kg -1 ), AnV (1.31 ± 0.06) and CV (1.4 ± 0.08). The pH of sarshir decreased from 6.2 ± 0 to 3.5 ± 0.1 during 14 h of fermentation. Incorporated bacterial cells exhibited notable viability during 10 days of cold storage (4 °C). The fermentation of sarshir by L. plantarum strains, especially LP3, resulted in beneficial changes in radical scavenging activity, as well as PV, AnV and carbonyl values, in addition to a broad spectrum of inhibitory activity against strains of P. aeruginosa, E. coli O157:H7, B. cereus and S. aureus. © 2017 Society of Chemical Industry. © 2017 Society of Chemical Industry.

Evidence of a major locus for lipoprotein lipase (LPL) activity in addition to a pleiotropic locus for both LPL and fasting insulin: results from the HERITAGE Family Study.

PubMed

Hong, Y; Rice, T; Després, J P; Gagnon, J; Nadeau, A; Bergeron, J; Pérusse, L; Bouchard, C; Leon, A S; Skinner, J S; Wilmore, J H; Rao, D C

1999-06-01

A major gene hypothesis for heparin releasable plasma lipoprotein lipase (PH-LPL) activity was assessed using segregation analyses of data on 495 members in 98 normolipidemic sedentary families of Caucasian descent who participated in the HERITAGE Family Study. Segregation analyses were performed on PH-LPL adjusted for age, and on PH-LPL activity adjusted for age and fasting insulin. Prior to adjustment for insulin, neither a major gene effect nor a multifactorial component could be rejected, and support for a major gene was equivocal i.e. neither the Mendelian transmission nor the no transmission (equal tau s) models were rejected. However, after adjusting for the effects of insulin, a major gene effect on PH-LPL activity was unambiguous. The putative locus accounted for 60% of the total phenotypic variance, and the homozygous recessive form affected 10% (q2) of the sample (i.e. gene frequency (q) = 0.31), and led to a low PH-LPL value. The lack of a significant multifactorial effect suggested that the familial etiology of PH-LPL activity adjusted for insulin was likely to be primarily a function of the major locus. In conclusion, the present study is the first to report segregation analyses on PH-LPL activity prior to and after adjusting for insulin, and suggests that there is an indication of a pleiotropic genetic effect on PH-LPL activity and insulin, in addition to a major gene effect on PH-LPL activity alone.

Acidic Extracellular pH Promotes Activation of Integrin αvβ3

PubMed Central

Paradise, Ranjani K.; Lauffenburger, Douglas A.; Van Vliet, Krystyn J.

2011-01-01

Acidic extracellular pH is characteristic of the cell microenvironment in several important physiological and pathological contexts. Although it is well established that acidic extracellular pH can have profound effects on processes such as cell adhesion and migration, the underlying molecular mechanisms are largely unknown. Integrin receptors physically connect cells to the extracellular matrix, and are thus likely to modulate cell responses to extracellular conditions. Here, we examine the role of acidic extracellular pH in regulating activation of integrin αvβ3. Through computational molecular dynamics simulations, we find that acidic extracellular pH promotes opening of the αvβ3 headpiece, indicating that acidic pH can thereby facilitate integrin activation. This prediction is consistent with our flow cytometry and atomic force microscope-mediated force spectroscopy assays of integrin αvβ3 on live cells, which both demonstrate that acidic pH promotes activation at the intact cell surface. Finally, quantification of cell morphology and migration measurements shows that acidic extracellular pH affects cell behavior in a manner that is consistent with increased integrin activation. Taken together, these computational and experimental results suggest a new and complementary mechanism of integrin activation regulation, with associated implications for cell adhesion and migration in regions of altered pH that are relevant to wound healing and cancer. PMID:21283814

Product development studies of amino acid conjugate of Aceclofenac.

PubMed

Singh, Ajay Pal; Ramadan, Wafa Mossa; Dahiya, Rajiv; Sarpal, A S; Pathak, Kamla

2009-04-01

The prodrugs designed by classical approach increase lipophilicity of the drug, which decreases the water solubility thus decreasing the concentration gradient, which controls drug absorption. To overcome the limitations of traditional prodrug approach, water soluble prodrugs can be designed by adding selected amino acid to the drug moiety that are the substrates for the enzyme located at the intestinal brush border thus overcoming pharmaceutical problem without compromising bioavailability. ACaa (Amino acid conjugate of Aceclofenac) was synthesized by conjugation with l-phenylalanine by conventional coupling method using N, N-dicyclohexylcarbodiimide and ACaa was characterized by melting point, TLC, photomicrograph, UV, FT-IR, FT-NMR, MS-FAB, XRD and DSC. As a part of product development study ACaa was subjected to studies like In-vivo in albino rats and in-vitro like ACaa reversion to AC (Aceclofenac) in aqueous buffers of pH 1.21, 2.38. 3.10, 6.22 and 7.41, at a constant concentration (0.05M), ionic strength (micro = 0.5) and at a temperature of 37 degrees C +/- 0.5 degrees C, ACaa showed negligible reversion (2.15 %) up to 24 hrs study at acidic pH thus suggesting stability in acidic environment of stomach, the rate of reversion increased as pH of medium increased. pH- partition profile, pH- solubility profile and micromeritic studies were also carried out in comparison to pure drug. The solubility and lipophilicity of ACaa exhibited higher values at all pH range when compared to AC. The micromeritic properties also evaluated in terms of particle shape and size, IQCS and kurtosis. Resulting IQCS value approached zero thus suggesting reducing in the degree of skewness.

The preparation and evaluation of salt forms of linogliride with reduced solubilities as candidates for extended release.

PubMed

Chrzanowski, Frank A; Ahmad, Kaleem

2017-03-01

Salts of linogliride with reduced solubilities were prepared and evaluated as potential candidates for extended-release oral dosage forms. A once-daily dose of 300-800 mg was intended. Seven acids were selected: p-acetamidobenzoic, benzoic, p-hydroxybenzoic, 3-hydroxy-2-naphthoic, 1-napsylic, pamoic, and p-toluenesulfonic acids but only four salts were able to be prepared in suitable quantities for evaluation: linogliride pamoate, p-hydroxybenzoate, 3-hydroxy-2-naphthoate, and 1-napsylate. The pH-solubility profiles of the four new salts, free base, and fumarate salt were compared over the pH 1.43-8.3 range and the intrinsic dissolution rates of the four new salts and the free base were determined at pH 1.43, 4.4, and 7.5. The range of the pH-solubility profile and intrinsic dissolution rates of the p-hydroxybenzoate salt were less than the free base and fumarate and higher than the other three new salts. The pH-solubilities and intrinsic dissolution rates of the 1-napsylate salt were pH-independent. The solubilities and intrinsic dissolution rates of the pamoate and 3-hydroxy-2-naphthoate were higher at pH 1.4-3.4 than at higher pH. At pH 4.4 and higher, the solubilities were essentially the same, in the 1-2 mg/mL range. The intrinsic dissolution rates were also very low and not very different. Dissolution studies with capsules containing 800 mg doses of the pamoate, 1-napsylate, free base, and fumarate performed in a dissolution medium of pH beginning at 2.2 and ending at 6.8 demonstrated that the pamoate and 1-napsylate salt forms dissolved slower and could be useful as extended-release forms.

Correlated patterns in hydrothermal plume distribution and apparent magmatic budget along 2500 km of the Southeast Indian Ridge

NASA Astrophysics Data System (ADS)

Baker, Edward T.; Hémond, Christophe; Briais, Anne; Maia, Marcia; Scheirer, Daniel S.; Walker, Sharon L.; Wang, Tingting; Chen, Yongshun John

2014-08-01

Multiple geological processes affect the distribution of hydrothermal venting along a mid-ocean ridge. Deciphering the role of a specific process is often frustrated by simultaneous changes in other influences. Here we take advantage of the almost constant spreading rate (65-71 mm/yr) along 2500 km of the Southeast Indian Ridge (SEIR) between 77°E and 99°E to examine the spatial density of hydrothermal venting relative to regional and segment-scale changes in the apparent magmatic budget. We use 227 vertical profiles of light backscatter and (on 41 profiles) oxidation-reduction potential along 27 first and second-order ridge segments on and adjacent to the Amsterdam-St. Paul (ASP) Plateau to map ph, the fraction of casts detecting a plume. At the regional scale, venting on the five segments crossing the magma-thickened hot spot plateau is almost entirely suppressed (ph = 0.02). Conversely, the combined ph (0.34) from all other segments follows the global trend of ph versus spreading rate. Off the ASP Plateau, multisegment trends in ph track trends in the regional axial depth, high where regional depth increases and low where it decreases. At the individual segment scale, a robust correlation between ph and cross-axis inflation for first-order segments shows that different magmatic budgets among first-order segments are expressed as different levels of hydrothermal spatial density. This correlation is absent among second-order segments. Eighty-five percent of the plumes occur in eight clusters totaling ˜350 km. We hypothesize that these clusters are a minimum estimate of the length of axial melt lenses underlying this section of the SEIR.

Solubility behavior and biopharmaceutical classification of novel high-solubility ciprofloxacin and norfloxacin pharmaceutical derivatives.

PubMed

Breda, Susana A; Jimenez-Kairuz, Alvaro F; Manzo, Ruben H; Olivera, María E

2009-04-17

The hydrochlorides of the 1:3 aluminum:norfloxacin and aluminum:ciprofloxacin complexes were characterized according to the Biopharmaceutics Classification System (BCS) premises in comparison with their parent compounds. The pH-solubility profiles of the complexes were experimentally determined at 25 and 37 degrees C in the range of pH 1-8 and compared to that of uncomplexed norfloxacin and ciprofloxacin. Both complexes are clearly more soluble than the antibiotics themselves, even at the lowest solubility pHs. The increase in solubility was ascribed to the species controlling solubility, which were analyzed in the solid phases at equilibrium at selected pHs. Additionally, permeability was set as low, based on data reported in the scientific literature regarding oral bioavailability, intestinal and cell cultures permeabilities and also considering the influence of stoichiometric amounts of aluminum. The complexes fulfill the BCS criterion to be classified as class 3 compounds (high solubility/low permeability). Instead, the active pharmaceutical ingredients (APIs) currently used in solid dosage forms, norfloxacin and ciprofloxacin hydrochloride, proved to be BCS class 4 (low solubility/low permeability). The solubility improvement turns the complexes as potential biowaiver candidates from the scientific point of view and may be a good way for developing more dose-efficient formulations. An immediate release tablet showing very rapid dissolution was obtained. Its dissolution profile was compared to that of the commercial ciprofloxacin hydrochloride tablets allowing to dissolution of the complete dose at a critical pH such as 6.8.

USGS Field Activities 11CEV01 and 11CEV02 on the West Florida Shelf, Gulf of Mexico, in January and February 2011

USGS Publications Warehouse

Robbins, Lisa L.; Knorr, Paul O.; Daly, Kendra L.; Taylor, Carl A.

2014-01-01

During January and February 2011 the U.S. Geological Survey (USGS), in cooperation with the University of South Florida (USF), conducted geochemical surveys on the west Florida Shelf. Data collected will allow USGS and USF scientists to investigate the effects of climate change on ocean acidification within the northern Gulf of Mexico, specifically, the effect of ocean acidification on marine organisms and habitats. This work is part of a larger USGS study on Climate and Environmental Variability (CEV). The first cruise was conducted from January 3 – 7 (11CEV01) and the second from February 17 - 27 (11CEV02). To view each cruise's survey lines, please see the Trackline page. Both cruises took place aboard the R/V Weatherbird II, a ship of opportunity led by Dr. Kendra Daly (USF), which departed and returned from Saint Petersburg, Florida. Data collection included sampling of the surface and water column (referred to as station samples) with lab analysis of pH, dissolved inorganic carbon (DIC), and total alkalinity. Augmenting the lab analysis was a continuous flow-through system with a Conductivity-Temperature-Depth (CTD) sensor, which also recorded salinity, and pH. Corroborating the USGS data are the vertical CTD profiles collected by USF. The CTD casts measured continuous vertical profiles of oxygen, chlorophyll fluorescence, optical backscatter, and transmissometer. Discrete samples for nutrients, chlorophyll, and particulate organic carbon/nitrogen were also collected during the CTD casts.

An enzyme kinetics study of the pH dependence of chloride activation of oxygen evolution in photosystem II.

PubMed

Baranov, Sergei; Haddy, Alice

2017-03-01

Oxygen evolution by photosystem II (PSII) involves activation by Cl - ion, which is regulated by extrinsic subunits PsbQ and PsbP. In this study, the kinetics of chloride activation of oxygen evolution was studied in preparations of PSII depleted of the PsbQ and PsbP subunits (NaCl-washed and Na 2 SO 4 /pH 7.5-treated) over a pH range from 5.3 to 8.0. At low pH, activation by chloride was followed by inhibition at chloride concentrations >100 mM, whereas at high pH activation continued as the chloride concentration increased above 100 mM. Both activation and inhibition were more pronounced at lower pH, indicating that Cl - binding depended on protonation events in each case. The simplest kinetic model that could account for the complete data set included binding of Cl - at two sites, one for activation and one for inhibition, and four protonation steps. The intrinsic (pH-independent) dissociation constant for Cl - activation, K S , was found to be 0.9 ± 0.2 mM for both preparations, and three of the four pK a s were determined, with the fourth falling below the pH range studied. The intrinsic inhibition constant, K I , was found to be 64 ± 2 and 103 ± 7 mM for the NaCl-washed and Na 2 SO 4 /pH7.5-treated preparations, respectively, and is considered in terms of the conditions likely to be present in the thylakoid lumen. This enzyme kinetics analysis provides a more complete characterization of chloride and pH dependence of O 2 evolution activity than has been previously presented.

Assessment of the pharmacodynamics of intranasal, intravenous and oral scopolamine

NASA Technical Reports Server (NTRS)

Tietze, Karen J.

1990-01-01

Space motion sickness is an important issue in the space medical sciences program. One of the objectives of the ongoing clinical experimental protocol Pharmacokinetics of Intranasal Scopolamine in Normal Subjects is to evaluate the pharmacodynamics of scopolamine using salivary flow rate and pH profiles and cognitive performance tests as pharmacodynamic parameters. Normal volunteers collected saliva and performed the NTI Multiresource Performance Battery tests at designed time intervals to establish control saliva flow rates, salivary pH profiles, and the characteristics of the learning curve for the performance program under normal conditions. In the clinical part of the study, saliva samples and performance test scores are collected from healthy nonsmoking subjects after receiving a single 0.4 mg dose of either intranasal, intravenous, or oral scopolamine.

Spoilage of sous vide cooked salmon (Salmo salar) stored under refrigeration.

PubMed

Díaz, P; Garrido, M D; Bañón, S

2011-02-01

The spoilage of Sous Vide 'SV' cooked salmon stored under refrigeration was studied. Samples were packaged under vacuum in polyamide-polypropylene pouches, cooked at an oven temperature/time of 80 (°)C/45 min, quickly chilled at 3 (°)C and stored at 2 (°)C for 0, 5 or 10 weeks for catering use. Microbial (aerobic and anaerobic psychrotrophs, lactic acid bacteria, molds and yeasts and Enterobacteriaceae), physical-chemical (pH, water activity, TBARS, acidity, L*a*b* color, texture profile analysis and shear force) and sensory (appearance, odor, flavor, texture and overall quality) parameters were determined. SV processing prevented the growth of aerobic and anaerobic psychrotrophs, lactic acid bacteria, molds and yeasts and Enterobacteriaceae. There were no relevant changes in pH, water activity, TBARS, CIELab color associated with cooked salmon spoilage. Instrumental texture data were contradictory. Slight decrease in lactic acid levels was found. In contrast, the SV cooked salmon suffered considerable sensory deterioration during its refrigerated storage, consisting of severe losses of cooked salmon odor and flavor, slight rancidity, discoloration associated with white precipitation, and moderates softness, and loss of chewiness and juiciness. No acidification, putrefaction or relevant rancidity was detected. The sensory spoilage preceded microbiological and physical-chemical spoilage, suggesting that microbiological quality alone may overestimate the shelf life of SV cooked salmon.

Screening, identification and characterization of bacteriocins produced by wine-isolated LAB strains.

PubMed

Ndlovu, B; Schoeman, H; Franz, C M A P; du Toit, M

2015-04-01

To screen and identify wine-isolated LAB strains for bacteriocin production, and to identify and characterize bacteriocins. One hundred and fifty-five LAB strains isolated from South African red wines undergoing spontaneous malolactic fermentation were screened for bacteriocin production. Eight isolates were identified to be bacteriocin producers and were identified as Enterococcus faecium. All eight isolates had the same phenotypic and genotypic profiles. The peptides were preliminarily identified as enterocin P using mass spectrometry and further confirmed by PCR-amplifying enterocin P gene. The enterocin activity was inhibited by α-Chymotrypsin, papain and proteinase K treatments. It was heat stable at 37, 60, 80 and 100°C and showed activity over a broad pH range of 2-10. The production of the enterocin followed that of primary metabolite kinetics and, it showed bactericidal effect to some wine spoilage LAB strains. Our study identified the presence of the enterocin-producing Enterococcus in wine. The enterocin was heat stable; with broad pH range and bactericidal effects to sensitive strains. This is one of very few studies that isolated Enterococcus species from wine. It is, however, the first to report presence of bacteriocin-producing Enterococcus in wine fermentation. © 2015 The Society for Applied Microbiology.

Biomimetics with a self-assembled monolayer of catalytically active tethered isoalloxazine on Au.

PubMed

Calvo, Ernesto J; Rothacher, M Silvina; Bonazzola, Cecilia; Wheeldon, Ian R; Salvarezza, Roberto C; Vela, Maria Elena; Benitez, Guillermo

2005-08-16

A new biomimetic nanostructured electrocatalyst comprised of a self-assembled monolayer (SAM) of flavin covalently attached to Au by reaction of methylformylisoalloxazine with chemisorbed cysteamine is introduced. Examinations by Fourier transform infrared spectroscopy and scanning tunneling microscopy (STM) show that the flavin molecules are oriented perpendicular to the surface with a 2 nm separation between flavin molecules. As a result of the contrast observed in the STM profiles between areas only covered by unreacted cysteamine and those covered by flavin-cysteamine moieties, it can be seen that the flavin molecules rise 0.7 nm above the chemisorbed cysteamines. The SAM flavin electrocatalyst undergoes fast electron transfer with the underlying Au and shows activity toward the oxidation of enzymatically active beta-NADH at pH 7 and very low potential (-0.2 V vs Ag/AgCl), a requirement for use in an enzymatic biofuel cell, and a 100-fold increase in activity with respect to the collisional reaction in solution.

A Comparative Study of New Aspergillus Strains for Proteolytic Enzymes Production by Solid State Fermentation

PubMed Central

Ortiz, Gastón Ezequiel; Noseda, Diego Gabriel; Ponce Mora, María Clara; Recupero, Matías Nicolás; Blasco, Martín; Albertó, Edgardo

2016-01-01

A comparative study of the proteolytic enzymes production using twelve Aspergillus strains previously unused for this purpose was performed by solid state fermentation. A semiquantitative and quantitative evaluation of proteolytic activity were carried out using crude enzymatic extracts obtained from the fermentation cultures, finding seven strains with high and intermediate level of protease activity. Biochemical, thermodynamics, and kinetics features such as optimum pH and temperature values, thermal stability, activation energy (E a), quotient energy (Q 10), K m, and V max were studied in four enzymatic extracts from the selected strains that showed the highest productivity. Additionally, these strains were evaluated by zymogram analysis obtaining protease profiles with a wide range of molecular weight for each sample. From these four strains with the highest productivity, the proteolytic extract of A. sojae ATCC 20235 was shown to be an appropriate biocatalyst for hydrolysis of casein and gelatin substrates, increasing its antioxidant activities in 35% and 125%, respectively. PMID:26989505

Hydrolytic Enzyme Activities and Protein Pattern of Avocado Fruit Ripened in Air and in Low Oxygen, with and without Ethylene 1

PubMed Central

Kanellis, Angelos K.; Solomos, Theophanes; Mattoo, Autar K.

1989-01-01

The effect of 2.5% O2 atmosphere with and without ethylene on the activities of hydrolytic enzymes associated with cell walls, and total protein profile during ripening of avocado fruits (Persea americana Mill., cv Hass) were investigated. The low 2.5% O2 atmosphere prevented the rise in the activities of cellulase, polygalacturonase, and acid phosphatase in avocado fruits whose ripening was initiated with ethylene. Addition of 100 microliters per liter ethylene to low O2 atmosphere did not alter these suppressive effects of 2.5% O2. Furthermore, 2.5% O2 atmosphere delayed the development of a number of polypeptides that appear during ripening of avocado fruits while at the same time new polypeptides accumulated. The composition of the extraction buffer and its pH greatly affected the recovery of cellulase activity and its total immunoreactive protein. Images Figure 1 Figure 2 Figure 5 PMID:16666746

A comparative study of the antihyaluronidase, antiurease, antioxidant, antimicrobial and physicochemical properties of different unifloral degrees of chestnut (Castanea sativa Mill.) honeys.

PubMed

Kolayli, Sevgi; Can, Zehra; Yildiz, Oktay; Sahin, Huseyin; Karaoglu, Sengul Alpay

2016-01-01

This study was planned to investigate some physicochemical and anti-inflammatory, antioxidant, antimicrobial properties of three different degrees of unifloral characters of chestnut honeys. Antihyaluronidase, antiurease and antimicrobial activities were evaluated as anti-inflammatory characteristics. Total phenolic contents, flavonoids, tannins, phenolic profiles, ferric-reducing antioxidant power (FRAP), scavenging activities of 2,2'-azinobis-(3-ethylbenzothiazoline-6-sulfonic acid (ABTS + ) and 2,2-diphenyl-1-picrylhydrazyl (DPPH) radicals were evaluated as antioxidant properties. Color, optical rotation, conductivity, moisture, pH and ash content were evaluated as physicochemical parameters, and some sugars content, prolin, diastase, HMF and minerals (Na, K, Ca, P, Fe, Cu and Zn) were evaluated as chemical and biochemical parameters. All studied physicochemical and biological active properties were changed in line with the unifloral character of the chestnut honeys. A higher unifloral character was found associated with greater apitherapeutic capacity of the honey, as well as biological active compounds.

Molecular characterization of southern bluefin tuna myoglobin (Thunnus maccoyii).

PubMed

Nurilmala, Mala; Ochiai, Yoshihiro

2016-10-01

The primary structure of southern bluefin tuna Thunnus maccoyii Mb has been elucidated by molecular cloning techniques. The cDNA of this tuna encoding Mb contained 776 nucleotides, with an open reading frame of 444 nucleotides encoding 147 amino acids. The nucleotide sequence of the coding region was identical to those of other bluefin tunas (T. thynnus and T. orientalis), thus giving the same amino acid sequences. Based on the deduced amino acid sequence, bioinformatic analysis was performed including phylogenic tree, hydropathy plot and homology modeling. In order to investigate the autoxidation profiles, the isolation of Mb was performed from the dark muscle. The water soluble fraction was subjected to ammonium sulfate fractionation (60-90 % saturation) followed by preparative gel electrophoresis. Autoxidation profiles of Mb were delineated at pH 5.6, 6.5 and 7.4 at temperature 37 °C. The autoxidation rate of tuna Mb was slightly higher than that of horse Mb at all pH examined. These results revealed that tuna myoglobin was unstable than that of horse Mb mainly at acidic pH.

A study of the effect of a boundary layer profile on the dynamic response and acoustic radiation of flat panels. Ph.D. Thesis - Virginia Univ.

NASA Technical Reports Server (NTRS)

Mixson, J. S.

1973-01-01

The response of a thin, elastic plate to a harmonic force which drives the plate from below and a compressible air stream with a viscous boundary layer flowing parallel to the upper surface along the length was investigated. Equations governing the forced response of the coupled plate-aerodynamic system are derived along with appropriate boundary conditions. Calculations of basic solution parameters for a linear velocity profile and for a Blasius profile showed that the same system response could be obtained from each profile if appropriate values of boundary layer thickness were chosen for each profile.

In Vitro Effects of Bromoalkyl Phenytoin Derivatives on Regulated Death, Cell Cycle and Ultrastructure of Leukemia Cells.

PubMed

Śladowska, Katarzyna; Opydo-Chanek, Małgorzata; Król, Teodora; Trybus, Wojciech; Trybus, Ewa; Kopacz-Bednarska, Anna; Handzlik, Jadwiga; Kieć-Kononowicz, Katarzyna; Mazur, Lidia

2017-11-01

To search for new antileukemic agents, the chemical structure of phenytoin was modified. A possible cytotoxic activity of three bromoalkyl phenytoin analogs, methyl 2-(1-(3-bromopropyl)-2,4-dioxo-5,5-diphenylimidazolidin-3-yl) propanoate (PH2), 1-(3-bromopropyl)-3-methyl-5,5-diphenylimidazolidine-2,4-dione (PH3) and 1-(4-bromobutyl)-3-methyl-5,5-diphenylimidazolidine-2,4-dione (PH4) on regulated cell death, the cell cycle and cell ultrastructure was assessed. The experiments were performed in vitro on HL-60 and U937 cells, using flow cytometry and electron microscopy methods. Application of PH2, PH3, and PH4 resulted in cell surface exposure of phosphatidylserine and plasma membrane impairment, caspase-8, -9, and -3/7 activation, dissipation of mitochondrial membrane potential, DNA breakage, cell-cycle disturbance and cell ultrastructural changes. In general, PH3 appeared to be the most active against the leukemia cells, and all bromoalkyl hydantoins, PH2-PH4, were more active in HL-60 cells than in U937 cells. The antileukemic activity of the bromoalkyl phenytoin analogs depended on the combination of N-hydantoin substituents and the human cell line used. Copyright© 2017, International Institute of Anticancer Research (Dr. George J. Delinasios), All rights reserved.

A high-throughput assay for the comprehensive profiling of DNA ligase fidelity

PubMed Central

Lohman, Gregory J. S.; Bauer, Robert J.; Nichols, Nicole M.; Mazzola, Laurie; Bybee, Joanna; Rivizzigno, Danielle; Cantin, Elizabeth; Evans, Thomas C.

2016-01-01

DNA ligases have broad application in molecular biology, from traditional cloning methods to modern synthetic biology and molecular diagnostics protocols. Ligation-based detection of polynucleotide sequences can be achieved by the ligation of probe oligonucleotides when annealed to a complementary target sequence. In order to achieve a high sensitivity and low background, the ligase must efficiently join correctly base-paired substrates, while discriminating against the ligation of substrates containing even one mismatched base pair. In the current study, we report the use of capillary electrophoresis to rapidly generate mismatch fidelity profiles that interrogate all 256 possible base-pair combinations at a ligation junction in a single experiment. Rapid screening of ligase fidelity in a 96-well plate format has allowed the study of ligase fidelity in unprecedented depth. As an example of this new method, herein we report the ligation fidelity of Thermus thermophilus DNA ligase at a range of temperatures, buffer pH and monovalent cation strength. This screen allows the selection of reaction conditions that maximize fidelity without sacrificing activity, while generating a profile of specific mismatches that ligate detectably under each set of conditions. PMID:26365241

A high-throughput assay for the comprehensive profiling of DNA ligase fidelity.

PubMed

Lohman, Gregory J S; Bauer, Robert J; Nichols, Nicole M; Mazzola, Laurie; Bybee, Joanna; Rivizzigno, Danielle; Cantin, Elizabeth; Evans, Thomas C

2016-01-29

DNA ligases have broad application in molecular biology, from traditional cloning methods to modern synthetic biology and molecular diagnostics protocols. Ligation-based detection of polynucleotide sequences can be achieved by the ligation of probe oligonucleotides when annealed to a complementary target sequence. In order to achieve a high sensitivity and low background, the ligase must efficiently join correctly base-paired substrates, while discriminating against the ligation of substrates containing even one mismatched base pair. In the current study, we report the use of capillary electrophoresis to rapidly generate mismatch fidelity profiles that interrogate all 256 possible base-pair combinations at a ligation junction in a single experiment. Rapid screening of ligase fidelity in a 96-well plate format has allowed the study of ligase fidelity in unprecedented depth. As an example of this new method, herein we report the ligation fidelity of Thermus thermophilus DNA ligase at a range of temperatures, buffer pH and monovalent cation strength. This screen allows the selection of reaction conditions that maximize fidelity without sacrificing activity, while generating a profile of specific mismatches that ligate detectably under each set of conditions. © The Author(s) 2015. Published by Oxford University Press on behalf of Nucleic Acids Research.

Detection of the 267 GHz J = 1-0 rotational transition of PH3 in Saturn with a new Fourier transform spectrometer

NASA Technical Reports Server (NTRS)

Weisstein, Eric W.; Serabyn, E.

1994-01-01

In recent observations at the Caltech Submillimeter Observatory, the highly pressure-broadened (FWHM = 11.2 GHz) J = 1-0 rotational transition of PH3 (phosphine) was detected on Saturn. By modeling the Saturnian atmosphere with a radiative transfer code, the observed line profile was consistent with a constant PH3 mole fraction of 3.0 plus or minus 1.0 ppm in the upper troposphere. A best-fit to the depth of the line implies a cutoff at high altitudes, with no PH3 present at pressures approximately less than 100 mbar. The observed line depth, combined with the lack of a detectable emission core, implies that a cutoff in the PH3 abundance occurs at a pressure between 13 and 140 mbar. PH3 in Jupiter was not detected, nor any other molecular lines between 195 and 295 GHz (1.54 mm and 1.02 mm, respectively) in either Jupiter or Saturn.

Application of quartz crystal microbalance to study the impact of pH and ionic strength on protein-silicone oil interactions.

PubMed

Dixit, Nitin; Maloney, Kevin M; Kalonia, Devendra S

2011-06-30

In this study, we have used quartz crystal microbalance (QCM) to quantitate the adsorption of a protein on silicone oil coated surfaces as a function of protein concentration, pH and ionic strength using a 5 MHz quartz crystal. Protein adsorption isotherms were generated at different solution pH and ionic strengths. Surface saturation concentrations were selected from adsorption isotherms and used to generate adsorption profiles from pH 3.0 to 9.0, and at ionic strengths of 10 mM and 150 mM. At low ionic strength (10mM) and pH 5.0 (close to the isoelectric point of the protein), maximum adsorption of protein to the silicone oil surface was observed. At higher ionic strength (150 mM), no significant pH influence on adsorption was observed. QCM could be used as a reliable technique to study the binding of proteins to silicone oil coated surfaces. Copyright © 2011 Elsevier B.V. All rights reserved.

Development of an autonomous, wireless pH and temperature sensing system for monitoring pig meat quality.

PubMed

Frisby, June; Raftery, Declan; Kerry, Joe P; Diamond, Dermot

2005-06-01

This paper focuses on the development of a unique wireless pH and temperature monitoring system to assess pig meat quality. Pale, soft and exudative (PSE) pig meat continues to be a major problem in the pig meat industry today. The PSE condition in pork is related to a number of factors including genetics, pre-slaughter stress and insufficient chilling of pig carcasses, which cause a rapid rate of glycolysis post-mortem (<1h). As a result the pH drops to low levels while the muscle temperature is still high. A wireless dual channel system that monitors pH and temperature simultaneously has been developed to provide pH and temperature data of the carcass during the first 24h after slaughter. We have demonstrated that this approach can distinguish in real time, pH and temperature profiles that are 'non-normal', and identify carcasses that are PSE positive quickly and easily.

Glyphosate inhibition of 5-enolpyruvylshikimate 3-phosphate synthease from suspension-cultured cells of Nicotiana silvestris

DOE Office of Scientific and Technical Information (OSTI.GOV)

Rubin, J.L.; Gaines, C.G.; Jensen, R.A.

1984-07-01

Treatment of isogenic suspension-cultured cells of Nicotiana silvestris Speg, et Comes with glyphosate (N-(phosphonomethyl)glycine) led to elevated levels of intracellular shikimate (364-fold increase by 1.0 millimolar glyphosate). In the presence of glyphosate, it is likely that most molecules of shikimate originate from the action of 3-deoxy-d-arabino-heptulosonate 7-phosphate (DAHP) synthase-Mn since this isozyme, in contrast to the DAHP synthase-Co isozyme, is insensitive to inhibition by glyphosate. 5-Enolpyruvylshikimate 3-phosphate (EPSP) synthase (EC 2.5.1.19) from N. silvestris was sensitive to micromolar concentrations of glyphosate and possessed a single inhibitor binding site. Rigorous kinetic studies of EPSP synthase required resolution from the multiple phosphatasemore » activities present in crude extracts, a result achieved by ion-exchange column chromatography. Although EPSP synthase exhibited a broad pH profile (50% of maximal activity between pH 6.2 and 8.5), sensitivity to glyphosate increased dramatically with increasing pH within this range. In accordance with these data and the pK/sub a/ values of glyphosate, it is likely that the ionic form of glyphosate inhibiting EPSP synthase is COO/sup -/CH/sub 2/NH/sub 2//sup +/CH/sub 2/PO/sub 3//sup 2 -/, and that a completely ionized phosphono group is essential for inhibition. At pH 7.0, inhibition was competitive with respect to phosphoenolpyruvate (K/sub i/ = 1.25 micromolar) and uncompetitive with respect to shikimate-3-P (K/sub i/ = 18.3 micromolar). All data were consistent with a mechanism of inhibition in which glyphosate competes with PEP for binding to an (enzyme:shikimate-3-P) complex and ultimately forms the dead-end complex of (enzyme:shikimate-3-P:glyphosate). 36 references, 8 figures, 1 table.« less

Yogurt made from milk heated at different pH values.

PubMed

Ozcan, Tulay; Horne, David S; Lucey, John A

2015-10-01

Milk for yogurt manufacture is subjected to high heat treatment to denature whey proteins. Low milk pH values (≤ 6.5) at heating result in most denatured whey proteins becoming associated with casein micelles, whereas high milk pH values (≥ 7.0) at heating result in the formation of mostly soluble (nonmicellar) denatured whey protein complexes. There are conflicting reports on the relative importance of soluble and casein-bound whey protein aggregates on the properties of acid gels. Prior studies investigating the effect of pH of milk at heating used model gels in which milk was acidified by glucono-δ-lactone; in this study, we prepared yogurt gels using commercial starter cultures. Model acid gels can have very different texture and physical properties from those made by fermentation with starter cultures. In this study, we investigated the effects of different pH values of milk at heating on the rheological, light backscatter, and microstructural properties of yogurt gels. Reconstituted skim milk was adjusted to pH values 6.2, 6.7, and 7.2 and heated at 85°C for 30 min. A portion of the heated milk samples was readjusted back to pH 6.7 after heating. Milks were inoculated with 3% (wt/wt) yogurt starter culture and incubated at 40°C until pH 4.6. Gel formation was monitored using dynamic oscillatory rheology, and parameters measured included the storage modulus (G') and loss tangent (LT) values. Light-backscattering properties, such as the backscatter ratio (R) and the first derivative of light backscatter ratio (R'), were also monitored during fermentation. Fluorescence microscopy was used to observe gel microstructure. The G' values at pH 4.6 were highest in gels made from milk heated at pH 6.7 and lowest in milk heated at pH 6.2, with or without pH adjustment after heating. The G' values at pH 4.6 were lower in samples after adjustment back to pH 6.7 after heating. No maximum in the LT parameter was observed during gelation for yogurts made from milk heated at pH 6.2; a maximum in LT was observed at pH ~4.8 for samples heated at pH 6.7 or 7.2, with or without pH adjustment after heating. Higher R-values were observed with an increase in pH of heating, with or without pH adjustment after heating. The sample heated at pH 6.2 had only one major peak in its R' profile during acidification, whereas samples heated at pH 6.7 and 7.2 had 2 large peaks. The lack of a maximum in LT parameter and the presence of a single peak in the R' profile for the samples heated at pH 6.2 were likely due to the partial solubilization of insoluble calcium phosphate when milk was acidified to this lower pH value. No clear differences were observed in the microstructures of gels between the different treatments. This study indicates that heating milk at the natural pH (~6.7) created an optimum balance of casein-bound and soluble denatured whey proteins, which resulted in yogurt with the highest gel stiffness. Copyright © 2015 American Dairy Science Association. Published by Elsevier Inc. All rights reserved.

Effect of HPMC - E15 LV premium polymer on release profile and compression characteristics of chitosan/ pectin colon targeted mesalamine matrix tablets and in vitro study on effect of pH impact on the drug release profile.

PubMed

Newton, A M J; Lakshmanan, Prabakaran

2014-04-01

The study was designed to investigate the in vitro dissolution profile and compression characteristics of colon targeted matrix tablets prepared with HPMC E15 LV in combination with pectin and Chitosan. The matrix tablets were subjected to two dissolution models in various simulated fluids such as pH 1.2, 6, 6.8, 7.2, 5.5. The fluctuations in colonic pH conditions during IBD (inflammatory bowel disease) and the nature of less fluid content in the colon may limit the expected drug release in the polysaccharide-based matrices when used alone. The Hydrophilic hydroxyl propyl methylcellulose ether premium polymer (HPMC E15 LV) of low viscosity grade was used in the formulation design, which made an excellent modification in physical and compression characteristics of the granules. The release studies indicated that the prepared matrices could control the drug release until the dosage form reaches the colon and the addition HPMC E15 LV showed the desirable changes in the dissolution profile by its hydrophilic nature since the colon is known for its less fluid content. The hydrophilic HPMC E15 LV allowed the colonic fluids to enter into the matrix and confirmed the drug release at the target site from a poorly water soluble polymer such as Chitosan and also from water soluble Pectin. The dramatic changes occurred in the drug release profile and physicochemical characteristics of the Pectin, Chitosan matrix tablets when a premium polymer HPMC E15 LV added in the formulation design in the optimized concentration. Various drug release mechanisms used for the examination of drug release characteristics. Drug release followed the combined mechanism of diffusion, erosion, swelling and polymer entanglement. In recent decade, IBD attracts many patents in novel treatment methods by using novel drug delivery systems.

Real-time pH monitoring of industrially relevant enzymatic reactions in a microfluidic side-entry reactor (μSER) shows potential for pH control.

PubMed

Gruber, Pia; Marques, Marco P C; Sulzer, Philipp; Wohlgemuth, Roland; Mayr, Torsten; Baganz, Frank; Szita, Nicolas

2017-06-01

Monitoring and control of pH is essential for the control of reaction conditions and reaction progress for any biocatalytic or biotechnological process. Microfluidic enzymatic reactors are increasingly proposed for process development, however typically lack instrumentation, such as pH monitoring. We present a microfluidic side-entry reactor (μSER) and demonstrate for the first time real-time pH monitoring of the progression of an enzymatic reaction in a microfluidic reactor as a first step towards achieving pH control. Two different types of optical pH sensors were integrated at several positions in the reactor channel which enabled pH monitoring between pH 3.5 and pH 8.5, thus a broader range than typically reported. The sensors withstood the thermal bonding temperatures typical of microfluidic device fabrication. Additionally, fluidic inputs along the reaction channel were implemented to adjust the pH of the reaction. Time-course profiles of pH were recorded for a transketolase and a penicillin G acylase catalyzed reaction. Without pH adjustment, the former showed a pH increase of 1 pH unit and the latter a pH decrease of about 2.5 pH units. With pH adjustment, the pH drop of the penicillin G acylase catalyzed reaction was significantly attenuated, the reaction condition kept at a pH suitable for the operation of the enzyme, and the product yield increased. This contribution represents a further step towards fully instrumented and controlled microfluidic reactors for biocatalytic process development. © 2017 The Authors. Biotechnology Journal published by WILEY-VCH Verlag GmbH & Co. KGaA, Weinheim.

Resilience and recovery of Dehalococcoides mccartyi following low pH exposure.

PubMed

Yang, Yi; Cápiro, Natalie L; Yan, Jun; Marcet, Tyler F; Pennell, Kurt D; Löffler, Frank E

2017-12-01

Bioremediation treatment (e.g. biostimulation) can decrease groundwater pH with consequences for Dehalococcoides mccartyi (Dhc) reductive dechlorination activity. To explore the pH resilience of Dhc, the Dhc-containing consortium BDI was exposed to pH 5.5 for up to 40 days. Following 8- and 16-day exposure periods to pH 5.5, dechlorination activity and growth recovered when returned to pH 7.2; however, the ability of the culture to dechlorinate vinyl chloride (VC) to ethene was impaired (i.e. decreased rate of VC transformation). Dhc cells exposed to pH 5.5 for 40 days did not recover the ethene-producing phenotype upon transfer to pH 7.2 even after 200 days of incubation. When returned to pH 7.2 conditions after an 8-, a 16- and a 40-day low pH exposure, tceA and vcrA genes showed distinct fold increases, suggesting Dhc strain-specific responses to low pH exposure. Furthermore, a survey of Dhc biomarker genes in groundwater samples revealed the average abundances of Dhc 16S rRNA, tceA and vcrA genes in pH 4.5-6 groundwater were significantly lower (P-value < 0.05) than in pH 6-8.3 groundwater. Overall, the results of the laboratory study and the assessment of field data demonstrate that sustained Dhc activity should not be expected in low pH groundwater, and the duration of low pH exposure affects the ability of Dhc to recover activity at circumneutral pH. © FEMS 2017. All rights reserved. For permissions, please e-mail: journals.permissions@oup.com.

The Spinal Cord Has an Intrinsic System for the Control of pH.

PubMed

Jalalvand, Elham; Robertson, Brita; Tostivint, Hervé; Wallén, Peter; Grillner, Sten

2016-05-23

For survival of the organism, acid-base homeostasis is vital [1, 2]. The respiratory and renal systems are central to this control. Here we describe a novel mechanism, intrinsic to the spinal cord, with sensors that detect pH changes and act to restore pH to physiological levels by reducing motor activity. This pH sensor consists of somatostatin-expressing cerebrospinal fluid-contacting (CSF-c) neurons, which target the locomotor network. They have a low level of activity at pH 7.4. However, at both alkaline and acidic pH, the activity of the individual CSF-c neuron is markedly enhanced through the action of two separate channel subtypes. The alkaline response depends on PKD2L1 channels that have a large conductance and an equilibrium potential around 0 mV, both characteristics of mouse PKD2L1 channels [3-5]. The acidic response is due to an activation of ASIC3 [6]. The discharge pattern of the CSF-c neurons is U-shaped with a minimum frequency around pH 7.4 and a marked increase already at slightly lower and higher pH. During ongoing locomotor activity in the isolated spinal cord, both an increase and as a decrease of pH will reduce the locomotor burst rate. A somatostatin antagonist blocks these effects, suggesting that CSF-c neurons are responsible for the suppression of locomotor activity. CSF-c neurons thus represent a novel innate homeostatic mechanism, designed to sense any deviation from physiological pH and to respond by causing a depression of the motor activity. Because CSF-c neurons are found in all vertebrates, their pH-sensing function is most likely conserved. Copyright © 2016 Elsevier Ltd. All rights reserved.

The pH and pCO2 dependence of sulfate reduction in shallow-sea hydrothermal CO2 – venting sediments (Milos Island, Greece)

PubMed Central

Bayraktarov, Elisa; Price, Roy E.; Ferdelman, Timothy G.; Finster, Kai

2013-01-01

Microbial sulfate reduction (SR) is a dominant process of organic matter mineralization in sulfate-rich anoxic environments at neutral pH. Recent studies have demonstrated SR in low pH environments, but investigations on the microbial activity at variable pH and CO2 partial pressure are still lacking. In this study, the effect of pH and pCO2 on microbial activity was investigated by incubation experiments with radioactive 35S targeting SR in sediments from the shallow-sea hydrothermal vent system of Milos, Greece, where pH is naturally decreased by CO2 release. Sediments differed in their physicochemical characteristics with distance from the main site of fluid discharge. Adjacent to the vent site (T ~40–75°C, pH ~5), maximal sulfate reduction rates (SRR) were observed between pH 5 and 6. SR in hydrothermally influenced sediments decreased at neutral pH. Sediments unaffected by hydrothermal venting (T ~26°C, pH ~8) expressed the highest SRR between pH 6 and 7. Further experiments investigating the effect of pCO2 on SR revealed a steep decrease in activity when the partial pressure increased from 2 to 3 bar. Findings suggest that sulfate reducing microbial communities associated with hydrothermal vent system are adapted to low pH and high CO2, while communities at control sites required a higher pH for optimal activity. PMID:23658555

The pH and pCO2 dependence of sulfate reduction in shallow-sea hydrothermal CO2 - venting sediments (Milos Island, Greece).

PubMed

Bayraktarov, Elisa; Price, Roy E; Ferdelman, Timothy G; Finster, Kai

2013-01-01

Microbial sulfate reduction (SR) is a dominant process of organic matter mineralization in sulfate-rich anoxic environments at neutral pH. Recent studies have demonstrated SR in low pH environments, but investigations on the microbial activity at variable pH and CO2 partial pressure are still lacking. In this study, the effect of pH and pCO2 on microbial activity was investigated by incubation experiments with radioactive (35)S targeting SR in sediments from the shallow-sea hydrothermal vent system of Milos, Greece, where pH is naturally decreased by CO2 release. Sediments differed in their physicochemical characteristics with distance from the main site of fluid discharge. Adjacent to the vent site (T ~40-75°C, pH ~5), maximal sulfate reduction rates (SRR) were observed between pH 5 and 6. SR in hydrothermally influenced sediments decreased at neutral pH. Sediments unaffected by hydrothermal venting (T ~26°C, pH ~8) expressed the highest SRR between pH 6 and 7. Further experiments investigating the effect of pCO2 on SR revealed a steep decrease in activity when the partial pressure increased from 2 to 3 bar. Findings suggest that sulfate reducing microbial communities associated with hydrothermal vent system are adapted to low pH and high CO2, while communities at control sites required a higher pH for optimal activity.

Enhanced membrane disruption and antibiotic action against pathogenic bacteria by designed histidine-rich peptides at acidic pH.

PubMed

Mason, A James; Gasnier, Claire; Kichler, Antoine; Prévost, Gilles; Aunis, Dominique; Metz-Boutigue, Marie-Hélène; Bechinger, Burkhard

2006-10-01

The histidine-rich amphipathic cationic peptide LAH4 has antibiotic and DNA delivery capabilities. Here, we explore the interaction of peptides from this family with model membranes as monitored by solid-state (2)H nuclear magnetic resonance and their antibiotic activities against a range of bacteria. At neutral pH, the membrane disruption is weak, but at acidic pH, the peptides strongly disturb the anionic lipid component of bacterial membranes and cause bacterial lysis. The peptides are effective antibiotics at both pH 7.2 and pH 5.5, although the antibacterial activity is strongly affected by the change in pH. At neutral pH, the LAH peptides were active against both methicillin-resistant and -sensitive Staphylococcus aureus strains but ineffective against Pseudomonas aeruginosa. In contrast, the LAH peptides were highly active against P. aeruginosa in an acidic environment, as is found in the epithelial-lining fluid of cystic fibrosis patients. Our results show that modest antibiotic activity of histidine-rich peptides can be dramatically enhanced by inducing membrane disruption, in this case by lowering the pH, and that histidine-rich peptides have potential as future antibiotic agents.

Effects of induced subacute ruminal acidosis on milk fat content and milk fatty acid profile.

PubMed

Enjalbert, F; Videau, Y; Nicot, M C; Troegeler-Meynadier, A

2008-06-01

Two lactating dairy cows fitted with a rumen cannula received successively diets containing 0%, 20%, 34% and again 0% of wheat on a dry matter basis. After 5, 10 and 11 days, ruminal pH was measured between 8:00 and 16:00 hours, and milk was analysed for fat content and fatty acid profile. Diets with 20% and 34% wheat induced a marginal and a severe subacute ruminal acidosis respectively. After 11 days, diets with wheat strongly reduced the milk yield and milk fat content, increased the proportions of C8:0 to C13:0 even- or odd-chain fatty acids, C18:2 n-6 and C18:3 n-3 fatty acids but decreased the proportions of C18:0 and cis-9 C18:1 fatty acids. Wheat also increased the proportions of trans-5 to trans-10 C18:1, the latter exhibiting a 10-fold increase with 34% of wheat compared with value during the initial 0% wheat period. There was also an increase of trans-10, cis-12 C18:2 fatty acid and a decrease of trans-11 to trans-16 C18:1 fatty acids. The evolution during adaptation or after return to a 0% wheat diet was rapid for pH but much slower for the fatty acid profile. The mean ruminal pH was closely related to milk fat content, the proportion of odd-chain fatty acids (linear relationship) and the ratio of trans-10 C18:1/trans-11 C18:1 (nonlinear relationship). Such changes in fatty acid profile suggested a possible use for non-invasive diagnosis of subacute ruminal acidosis.

Phenol removal performance and microbial community shift during pH shock in a moving bed biofilm reactor (MBBR).

PubMed

Zhou, Hao; Wang, Guochen; Wu, Minghuo; Xu, Weiping; Zhang, Xuwang; Liu, Lifen

2018-06-05

A moving bed biofilm reactor (MBBR) effectively removes pollutants and even runs under extreme conditions. However, the pH shock resistance of a biofilm in MBBRs has been rarely reported. In this study, simulated phenol wastewater with acidic shock (pH 7.5-3.0) was used. In the pH shock phase, the phenol and COD removal efficiencies initially decreased and gradually increased to more than 90%. Microscopic studies showed that the superficial biofilm was mainly composed of fungi (yeasts) in the acidic pH shock phase. The microbial community composition in the acidic pH shock phase was significantly different from those in other phases. Firmicutes and Ascomycota were the dominant bacterial and fungal phyla in this stage, respectively. 16S rRNA gene-based functional annotation indicated that functional profiles related to aromatic compound degradation existed in all of the stages. Therefore, MBBRs show potential for the treatment of phenolic wastewater exposed to pH shock. Copyright © 2018 Elsevier B.V. All rights reserved.

Extracellular pH Regulates Zinc Signaling via an Asp Residue of the Zinc-sensing Receptor (ZnR/GPR39)*

PubMed Central

Cohen, Limor; Asraf, Hila; Sekler, Israel; Hershfinkel, Michal

2012-01-01

Zinc activates a specific Zn2+-sensing receptor, ZnR/GPR39, and thereby triggers cellular signaling leading to epithelial cell proliferation and survival. Epithelial cells that express ZnR, particularly colonocytes, face frequent changes in extracellular pH that are of physiological and pathological implication. Here we show that the ZnR/GPR39-dependent Ca2+ responses in HT29 colonocytes were maximal at pH 7.4 but were reduced by about 50% at pH 7.7 and by about 62% at pH 7.1 and were completely abolished at pH 6.5. Intracellular acidification did not attenuate ZnR/GPR39 activity, indicating that the pH sensor of this protein is located on an extracellular domain. ZnR/GPR39-dependent activation of extracellular-regulated kinase (ERK)1/2 or AKT pathways was abolished at acidic extracellular pH of 6.5. A similar inhibitory effect was monitored for the ZnR/GPR39-dependent up-regulation of Na+/H+ exchange activity at pH 6.5. Focusing on residues putatively facing the extracellular domain, we sought to identify the pH sensor of ZnR/GPR39. Replacing the histidine residues forming the Zn2+ binding site, His17 or His19, or other extracellular-facing histidines to alanine residues did not abolish the pH dependence of ZnR/GPR39. In contrast, replacing Asp313 with alanine resulted in similar Ca2+ responses triggered by ZnR/GPR39 at pH 7.4 or 6.5. This mutant also showed similar activation of ERK1/2 and AKT pathways, and ZnR-dependent up-regulation of Na+/H+ exchange at pH 7.4 and pH 6.5. Substitution of Asp313 to His or Glu residues restored pH sensitivity of the receptor. This indicates that Asp313, which was shown to modulate Zn2+ binding, is an essential residue of the pH sensor of GPR39. In conclusion, ZnR/GPR39 is tuned to sense physiologically relevant changes in extracellular pH that thus regulate ZnR-dependent signaling and ion transport activity. PMID:22879599

Extracellular pH regulates zinc signaling via an Asp residue of the zinc-sensing receptor (ZnR/GPR39).

PubMed

Cohen, Limor; Asraf, Hila; Sekler, Israel; Hershfinkel, Michal

2012-09-28

Zinc activates a specific Zn(2+)-sensing receptor, ZnR/GPR39, and thereby triggers cellular signaling leading to epithelial cell proliferation and survival. Epithelial cells that express ZnR, particularly colonocytes, face frequent changes in extracellular pH that are of physiological and pathological implication. Here we show that the ZnR/GPR39-dependent Ca(2+) responses in HT29 colonocytes were maximal at pH 7.4 but were reduced by about 50% at pH 7.7 and by about 62% at pH 7.1 and were completely abolished at pH 6.5. Intracellular acidification did not attenuate ZnR/GPR39 activity, indicating that the pH sensor of this protein is located on an extracellular domain. ZnR/GPR39-dependent activation of extracellular-regulated kinase (ERK)1/2 or AKT pathways was abolished at acidic extracellular pH of 6.5. A similar inhibitory effect was monitored for the ZnR/GPR39-dependent up-regulation of Na(+)/H(+) exchange activity at pH 6.5. Focusing on residues putatively facing the extracellular domain, we sought to identify the pH sensor of ZnR/GPR39. Replacing the histidine residues forming the Zn(2+) binding site, His(17) or His(19), or other extracellular-facing histidines to alanine residues did not abolish the pH dependence of ZnR/GPR39. In contrast, replacing Asp(313) with alanine resulted in similar Ca(2+) responses triggered by ZnR/GPR39 at pH 7.4 or 6.5. This mutant also showed similar activation of ERK1/2 and AKT pathways, and ZnR-dependent up-regulation of Na(+)/H(+) exchange at pH 7.4 and pH 6.5. Substitution of Asp(313) to His or Glu residues restored pH sensitivity of the receptor. This indicates that Asp(313), which was shown to modulate Zn(2+) binding, is an essential residue of the pH sensor of GPR39. In conclusion, ZnR/GPR39 is tuned to sense physiologically relevant changes in extracellular pH that thus regulate ZnR-dependent signaling and ion transport activity.

[Cd Runoff Load and Soil Profile Movement After Implementation of Some Typical Contaminated Agricultural Soil Remediation Strategies].

PubMed

Liu, Xiao-li; Zeng, Zhao-xia; Tie, Bai-qing; Chen, Qiu-wen; Wei, Xiang-dong

2016-02-15

Owing to the strong ability to immobilize and hyperaccumulate some toxic heavy metals in contaminated soils, the biochar, lime and such as hyperaccumulator ramie received increasing interests from crops and environment safety in recent years. Outdoor pot experiment was conducted to compare the impacts of lime and biochar addition in paddy rice treatment, hyperaccumulator ramie and ramie combined with EDTA of plant Phytoremediation methods on soil available Cd dynamics in rainfall runoff and the mobility along soil profile, under both natural acid precipitation and acid soil conditions. The results showed that, biochar addition at a 2% mass ratio application amount significantly increased soil pH, while ramie with EDTA application obviously decreased soil pH compared to ramie monoculture. Within the same rainfall events, water soluble Cd concentration in surface runoff of ramie treatments was significantly higher than those of waterlogged rice treatments, and Cd concentration in runoff was obviously increased after EDTA addition, whereas lime at a 0.3% mass ratio application amount as additive had no obvious impact on soil pH and Cd speciation change, which may be due to the low application amount. During the whole experimental period , water soluble Cd concentration of rainfall runoff in spring was higher than that in summer, showing the same seasonal characteristics in all treatments. Biochar addition could significantly decrease available Cd content in 0-20 cm soil layer and with certain preferable persistency effects, whereas EDTA addition treatment obviously increased available Cd of 0-20 cm soil layer compared to other treatments, and obvious Cd element activation phenomenon in 20-40 cm soil layer was observed after EDTA addition. In conclusion, lime and biochar as environmental and friendly alkaline Cd immobilization materials showed lower environment risk to surface and ground receiving water, but attention should be paid to phytoremediation enhanced with EDTA or other organic acid before promotion and field application for heavy metals removal from contaminated soils.

Regulating NETosis: Increasing pH Promotes NADPH Oxidase-Dependent NETosis

PubMed Central

Khan, Meraj A.; Philip, Lijy M.; Cheung, Guillaume; Vadakepeedika, Shawn; Grasemann, Hartmut; Sweezey, Neil; Palaniyar, Nades

2018-01-01

Neutrophils migrating from the blood (pH 7.35–7.45) into the surrounding tissues encounter changes in extracellular pH (pHe) conditions. Upon activation of NADPH oxidase 2 (Nox), neutrophils generate large amounts of H+ ions reducing the intracellular pH (pHi). Nevertheless, how extracellular pH regulates neutrophil extracellular trap (NET) formation (NETosis) is not clearly established. We hypothesized that increasing pH increases Nox-mediated production of reactive oxygen species (ROS) and neutrophil protease activity, stimulating NETosis. Here, we found that raising pHe (ranging from 6.6 to 7.8; every 0.2 units) increased pHi of both activated and resting neutrophils within 10–20 min (Seminaphtharhodafluor dual fluorescence measurements). Since Nox activity generates H+ ions, pHi is lower in neutrophils that are activated compared to resting. We also found that higher pH stimulated Nox-dependent ROS production (R123 generation; flow cytometry, plate reader assay, and imaging) during spontaneous and phorbol myristate acetate-induced NETosis (Sytox Green assays, immunoconfocal microscopy, and quantifying NETs). In neutrophils that are activated and not resting, higher pH stimulated histone H4 cleavage (Western blots) and NETosis. Raising pH increased Escherichia coli lipopolysaccharide-, Pseudomonas aeruginosa (Gram-negative)-, and Staphylococcus aureus (Gram-positive)-induced NETosis. Thus, higher pHe promoted Nox-dependent ROS production, protease activity, and NETosis; lower pH has the opposite effect. These studies provided mechanistic steps of pHe-mediated regulation of Nox-dependent NETosis. Raising pH either by sodium bicarbonate or Tris base (clinically known as Tris hydroxymethyl aminomethane, tromethamine, or THAM) increases NETosis. Each Tris molecule can bind 3H+ ions, whereas each bicarbonate HCO3− ion binds 1H+ ion. Therefore, the amount of Tris solution required to cause the same increase in pH level is less than that of equimolar bicarbonate solution. For that reason, regulating NETosis by pH with specific buffers such as THAM could be more effective than bicarbonate in managing NET-related diseases. PMID:29487850

Ordered cubic nanoporous silica support MCM-48 for delivery of poorly soluble drug indomethacin

NASA Astrophysics Data System (ADS)

Zeleňák, Vladimír; Halamová, Dáša; Almáši, Miroslav; Žid, Lukáš; Zeleňáková, Adriána; Kapusta, Ondrej

2018-06-01

Ordered MCM-48 nanoporous silica (SBET = 923(3) m2·g-1, VP = 0.63(2) cm3·g-1) with cubic Ia3d symmetry was used as a support for drug delivery of anti-inflammatory poorly soluble drug indomethacin. The delivery from parent, unmodified MCM-48, and 3-aminopropyl modified silica carrier was studied into the simulated body fluids with the pH = 2 and pH = 7.4. The studied samples were characterized by thermal analysis (TG/DTG-DTA), N2 adsorption/desorption, infrared spectroscopy (FT-IR), powder XRD, SEM, HRTEM methods, measurements of zeta potential (ζ) and dynamic light scattering (DLS). The determined content of indomethacin in pure MCM-48 was 21 wt.% and in the amine-modified silica MCM-48A-I the content was 45 wt.%. The release profile of the drug, in the time period up to 72 h, was monitored by TLC chromatographic method. It as shown, that by the modification of the surface, the drug release can be controlled. The slower release of indomethacin was observed from amino modified sample MCM-48A-I in the both types of studied simulated body fluids (slightly alkaline intravenous solution with pH = 7.4 and acidic gastric fluid with pH = 2), which was supported and explained by zeta potential and DLS measurements. The amount of the released indomethacin into the fluids with various pH was different. The maximum released amount of the drug was 97% for sample containing unmodified silica, MCM-48-I at pH = 7.4 and lowest released amount, 57%, for amine modified sample MCM-48A-I at pH = 2. To compare the indomethacin release profile four kinetic models were tested. Results showed, that that the drug release based on diffusion Higuchi model, mainly governs the release.

Biodegradable polyurethane micelles with pH and reduction responsive properties for intracellular drug delivery.

PubMed

Guan, Yayuan; Su, Yuling; Zhao, Lili; Meng, Fancui; Wang, Quanxin; Yao, Yongchao; Luo, Jianbin

2017-06-01

Polyurethane micelles with disulfide linkage located at the interface of hydrophilic shell and hydrophobic core (PU-SS-I) have been shown enhanced drug release profiles. However, the payloads could not be released completely. The occurrence of aggregation of hydrophobic cores upon shedding hydrophilic PEG coronas was considered as the reason for the incomplete release. To verify the above hypothesis and to develop a new polyurethane based micelles with dual stimuli respond properties and controllable location of pH and reduction responsive groups in the PU main chains, a tertiary amine was incorporated into the hydrophobic core PU-SS-I, which resulted polyurethane with both reduction and pH sensitive properties (PU-SS-N). Biodegradable polyurethane with only disulfide linkages located between the hydrophilic PEG segment and the hydrophobic PCL segments (PU-SS-I) and polyurethane with only pH sensitive tertiary amine at the hydrophobic core (PU-N-C) were used as comparisons. Paclitaxel (PTX) was chosen as mode hydrophobic drug to evaluate the loading and redox triggered release profiles of the PU micelles. It was demonstrated that PU-SS-N micelles disassembled instantly at the presence of 10mM GSH and at an acidic environment (pH=5.5), which resulted the nearly complete release (~90%) of the payloads within 48h, while about ~70% PTX was released from PU-SS-I and PU-SS-N micelles at neutral environment (pH=7.4) with the presence of 10mM GSH. The rapid and complete redox and pH stimuli release properties of the PU-SS-N nanocarrier will be a promising anticancer drug delivery system to ensure sufficient drug concentration to kill the cancer cells and to prevent the emergency of MDR. The in vitro cytotoxicity and cell uptake of the PTX-loaded micelles was also assessed in H460 and HepG2 cells. Copyright © 2017 Elsevier B.V. All rights reserved.

pH-Dependent stability of azithromycin in aqueous solution and structure identification of two new degradation products.

PubMed

Saita, Maria Grazia; Aleo, Danilo; Melilli, Barbara; Mangiafico, Sergio; Cro, Melina; Sanfilippo, Claudia; Patti, Angela

2018-05-28

The degradation profile of azithromycin in buffered solutions was investigated using HPLC and found to be pH dependent in the range of 6.0-7.2. Desosaminylazitromycin, derived from hydrolytic loss of cladinose of the parent molecule, was the major degradation product at pH 6.0 but its amount progressively decreased moving toward pH 7.2. Two additional unreported degradation products were also observed and their structures were fully elucidated by MS- and NMR-spectroscopy to be associated with opening of the macrocyclic lactone ring. Copyright © 2018. Published by Elsevier B.V.

Effects of sources of protein and enzyme supplementation on protein digestibility and chyme characteristics in broilers.

PubMed

Yu, B; Lee, T T T; Chiou, P W S

2002-07-01

1. The purpose of this study was to evaluate the effects of protein source and enzyme supplementation on protein digestibility and chyme characteristics in broilers. 2. One hundred and twenty growing (13 d old) and 60 finishing (34 d old) Arbor Acre strain commercial male broilers were selected and placed into individual metabolic cages. 3. The experiment was a 5 x 2 factorial arrangement with 5 different sources of protein: casein, fish meal, soybean meal (SBM), soy protein concentrate (SPC), maize gluten meal (MGM) and two levels of protease (bromelain), 0 and 65 CDU/kg diets. 4. The diets were iso-nitrogenous and semi-purified, with Cr2O3 as an indicator for determination of ileal digestibility and chyme characteristics. 5. Apparent ileal protein digestibility (AIPD) in both growing and finishing chickens was highest on the casein diet, followed by fish meal, SBM, SPC and MGM. 6. Enzyme inclusion did not improve protein digestibility, but significantly decreased the digesta pH value in the gizzard and increased pH in the ileum in the 3-week-old broilers. 7. The digesta pH values in the gizzard and duodenum were significantly lower in the SBM and fish meal groups compared with the other protein groups. The molecular weight distribution pattern of the soluble protein in the chyme of the gastrointestinal (GI) segments showed a similar trend, regardless of the enzyme inclusion or the stage of growth. 8. The molecular weight profile of soluble protein changed dynamically in the casein fed broilers from the gizzard to ileum and the low molecular weight proteins, < 7 kDa, reached maximum levels at the ileum. The molecular weight profile of the soluble protein in the SBM and SPC changed between the jejunum and the ileum and in the intermediate molecular soluble protein weight (7 to 10 kDa) was significantly decreased. This indicated that the hydrolysis process began from the middle to the posterior end of the small intestine. 9. Similar profiles were also shown with fish meal protein. The pattern of distribution, however, did not show any prominent change in the GI segments of the MGM group. 10. The pepsin, trypsin and chymotrypsin protease activity in the gizzard and duodenum were highest in the casein group and lowest in the MGM group as compared with the other protein groups. 11. The rate change in the patterns of molecular weight distribution in soluble protein and the digestive enzyme activity provide indications of the partial digestibility of different protein sources. The exogenous enzyme, bromelain, did not show any beneficial effect on protein digestion.

How deep does disturbance go? The long-term effects of canopy disturbance on tropical forest soil biogeochemistry

NASA Astrophysics Data System (ADS)

Gutiérrez del Arroyo, O.; Silver, W. L.

2015-12-01

We used the Canopy Trimming Experiment (CTE), an ongoing ecosystem manipulation study in the Luquillo Experimental Forest (LEF), Puerto Rico to determine the decadal-scale effects of canopy disturbance and debris deposition on biogeochemistry throughout the soil profile of a wet tropical forest. These manipulations represent the most significant effects of hurricanes, which may increase in frequency or intensity with warming, strengthening their ecosystem-level effects on carbon (C) and nutrient cycling. Four replicated treatments were applied in 2005 using a complete randomized block design: canopy trimming + debris deposition, canopy trimming only, debris deposition only, and untreated control. In 2015, we sampled soils at 10 cm intervals to 1 m depth in each of 12 plots (3 per treatment). We measured gravimetric moisture content, pH, HCl and citrate-ascorbate (CA) extractable iron (Fe) species, organic (Po) and inorganic fractions of NaHCO3 and NaOH phosphorus (P), as well as total C and nitrogen (N). Soil moisture decreased markedly with depth up to ~60-70 cm, and then stabilized at ~33% down to 1 m. Across all treatments, pH increased significantly with depth, ranging from 4.6 in surface soils (0-10 cm) of trimmed plots to 5.2 in deep soils (80-90 cm) of control plots. Canopy trimming decreased pH significantly, possibly due to increased root activity in surface soils as vegetation recovered. Both HCl and CA extractable Fe showed strong depth dependance, decreasing linearly to 50 cm, and stabilizing at very low concentrations (<0.2 mg/g) down to 1 m. Inorganic P concentrations were low and did not vary significantly with depth. The majority of P was associated with organic matter, with significantly higher values in the upper soil profile (<50 cm). Debris deposition significantly increased Po, revealing the role of hurricanes in subsidizing the available soil P pool in these highly productive, low-P wet tropical forests. Debris deposition also increased soil C and N concentrations in surface soils (<20 cm). Our results suggest that the dominant effects of disturbance are limited to the upper soil profile in this wet tropical forest. However, effects were persistent and detectable after ten years of the CTE, suggesting that hurricanes result in long-term changes in tropical forest biogeochemistry.

Phospholipase B activity of a purified phospholipase A from Vipera palestinae venom.

PubMed

Shiloah, J; Klibansky, C; de Vries, A; Berger, A

1973-05-01

Phospholipase was isolated (in two fractions) from Vipera palestinae venom and it was shown to possess phospholipase A activity (hydrolyzing diacyl-sn-glycerophosphorylcholines, e.g., lecithin, in the 2-position) as well as lysophospholipase (phospholipase B) activity (hydrolyzing 1-monoacyl-sn-glycerophosphorylcholines, e.g., lysolecithin, yielding free fatty acid and glycerophosphorylcholine). Each of the two purified enzyme fractions was homogeneous as judged by electrophoresis on acrylamide gel and by immunodiffusion and immunoelectrophoresis, and both had essentially equal activities. The ratio of the specific activity, at various purification stages, to the specific activity of the whole venom was the same for A activity (substrate lecithin) as for B activity (substrate lysolecithin). The enzyme has a molecular weight of 16,000, six S-S bridges, and no free thiol groups. At pH 7, dimerization was observed in the ultracentrifuge. A dissociation constant of about 10(-5) m was estimated. The amino acid composition for both fractions (140 amino acid residues) was found to be essentially the same. The A activity had a pH optimum at 9; B activity was low at this pH but increased steadily beyond pH 10.5. For the hydrolysis of lysolecithin the Lineweaver-Burk plot was found to be linear, giving K(m) = 1.1 mm and k(cat) = 0.55 sec(-1) at 37 degrees C and pH 10. 2-Deoxylysolecithin was also hydrolyzed by the enzyme at pH 10, with k(cat) = 0.01 sec(-1) (zero-order kinetics in the range 0.5-2.5 mm). For lecithin these constants could not be determined, but at 0.25 mm substrate the hydrolysis rate (at pH 9) of lecithin was about 1000 times the hydrolysis rate of lysolecithin (at pH 10).

Effect of pH and glucose on cultured human peritoneal mesothelial cells.

PubMed

Shao, J C; Yorioka, N; Nishida, Y; Yamakido, M

1999-08-01

We investigated the effects of various pH and glucose concentrations on the growth of human peritoneal mesothelial cells and on coagulation and fibrinolytic factors. Cells were cultured at various pH values in Ham's F-12 medium containing 1.0% foetal calf serum and supplemented with D-glucose or D-mannitol at various concentrations. After 4-48 h, cell proliferation and 3H-thymidine incorporation were determined. Coagulation and fibrinolytic factors were measured after 48 h. Glucose caused concentration-dependent inhibition of cell growth at all pH values, but the deleterious effect of low pH on cell proliferation was faster and stronger than that of high glucose. At a similar osmolality, mannitol caused less inhibition of cell proliferation than glucose. There was a glucose concentration-dependent increase of thrombin-antithrombin III complex production at all pH values. At pH 5.2, tissue-type plasminogen activator production was far lower than at higher pH values, and production of the plasminogen activator inhibitor showed a glucose concentration-dependent increase. At pH 6.5 or 7.3, however, the plasminogen activator inhibitor production decreased and tissue-type plasminogen activator production increased in a glucose concentration-dependent manner. Low pH and/or high glucose culture medium had an inhibitory effect on peritoneal mesothelial cells, with the effect of high glucose being partially related to hyperosmolality. These cells may modulate peritoneal coagulant and fibrinolytic activity, with the balance between coagulation and fibrinolysis being disturbed by low pH and/or high glucose.

Observations on the influence of water and soil pH on the persistence of insecticides.

PubMed

Chapman, R A; Cole, C M

1982-01-01

The pH-disappearance rate profiles were determined at ca. 25 degrees C for 24 insecticides at 4 or 5 pH values over the range 4.5 to 8.0 in sterile phosphate buffers prepared in water-ethanol (99:1 v/v). Half-lives measured at pH 8 were generally smaller than at lower pH values. Changes in half lives between pH 8.0 and 4.5 were largest (greater than 1000x) for the aryl carbamates, carbofuran and carbaryl, the oxime carbamate, oxamyl, and the organophosphorus insecticide, trichlorfon. In contrast, half lives of phorate, terbufos, heptachlor, fensulfothion and aldicarb were affected only slightly by pH changes. Under the experimental conditions described half lives at pH8 varied from 1-2 days for trichlorfon and oxamyl to greater than 1 year for fensulfothion and cypermethrin. Insecticide persistence on alumina (acid, neutral and basic), mineral soils amended with aluminum sulfate or calcium hydroxide to different pH values and four natural soils of different pH was examined. No correlation was observed between the measured pH of these solids and the rate of disappearance of selected insecticides applied to them. These observations demonstrate the difficulty of extrapolating the pH dependent disappearance behaviour observed in homogeneous solution to partially solid heterogeneous systems such as soil.

Proteolytic and Trypsin Inhibitor Activity in Germinating Jojoba Seeds (Simmondsia chinensis).

PubMed

Samac, D; Storey, R

1981-12-01

Changes in proteolytic activity (aminopeptidase, carboxypeptidase, endopeptidase) were followed during germination (imbibition through seedling development) in extracts from cotyledons of jojoba seeds (Simmondsia chinensis). After imbibition, the cotyledons contained high levels of sulfhydryl aminopeptidase activity (APA) but low levels of serine carboxypeptidase activity (CPA). CPA increased with germination through the apparent loss of a CPA inhibitor substance in the seed. Curves showing changes in endopeptidase activity (EPA) assayed at pH 4, 5, 6, 7, and 8 during germination were distinctly different. EPA at pH 4, 5, 6, and 7 showed characteristics of sulfhydryl enzymes while activity at pH 8 was probably due to a serine type enzyme. EPA at pH 6 was inhibited early in germination by one or more substances in the seed. Activities at pH 5 and later at pH 6 were the highest of all EPA throughout germination and increases in these activities were associated with a rapid loss of protein from the cotyledons of the developing seedling.Jojoba cotyledonary extracts were found to inhibit the enzymic activity of trypsin, chymotrypsin, and pepsin but not the protease from Aspergillus saotoi. The heat-labile trypsin inhibitor substance(s) was found in commercially processed jojoba seed meal and the albumin fraction of seed proteins. Trypsin inhibitor activity decreased with germination.

Proteolytic and Trypsin Inhibitor Activity in Germinating Jojoba Seeds (Simmondsia chinensis) 1

PubMed Central

Samac, Deborah; Storey, Richard

1981-01-01

Changes in proteolytic activity (aminopeptidase, carboxypeptidase, endopeptidase) were followed during germination (imbibition through seedling development) in extracts from cotyledons of jojoba seeds (Simmondsia chinensis). After imbibition, the cotyledons contained high levels of sulfhydryl aminopeptidase activity (APA) but low levels of serine carboxypeptidase activity (CPA). CPA increased with germination through the apparent loss of a CPA inhibitor substance in the seed. Curves showing changes in endopeptidase activity (EPA) assayed at pH 4, 5, 6, 7, and 8 during germination were distinctly different. EPA at pH 4, 5, 6, and 7 showed characteristics of sulfhydryl enzymes while activity at pH 8 was probably due to a serine type enzyme. EPA at pH 6 was inhibited early in germination by one or more substances in the seed. Activities at pH 5 and later at pH 6 were the highest of all EPA throughout germination and increases in these activities were associated with a rapid loss of protein from the cotyledons of the developing seedling. Jojoba cotyledonary extracts were found to inhibit the enzymic activity of trypsin, chymotrypsin, and pepsin but not the protease from Aspergillus saotoi. The heat-labile trypsin inhibitor substance(s) was found in commercially processed jojoba seed meal and the albumin fraction of seed proteins. Trypsin inhibitor activity decreased with germination. PMID:16662104

[Phospholipase and proteinase production by Malassezia pachydermatis isolated in dogs with and without otitis].

PubMed

Ortiz, Gustavo; Martín, M Carmen; Carrillo-Muñoz, Alfonso J; Payá, M Jesús

2013-01-01

Malassezia pachydermatis is part of the skin microbiota of dogs and cats. M. pachydermatis has been associated with external otitis and seborrhoeic dermatitis, reported more often in dogs than in cats. When the physical, chemical or immunological mechanisms of the skin are altered, M. pachydermatis could act as a pathogen. Thus, several virulence factors, such as the ability to produce esterase, lipase, lipoxygenase, protease, chondroitin sulphatase, and hyaluronidase, have been studied. In the present study, we aim to identify the phospholipase activity measured at pH 6.3, and the proteinase activity measured at pH 6.3 and pH 6.8 (pH from ears of dogs with external otitis) of M. pachydermatis strains isolated from dogs with and without external otitis. The phospholipase activity was measured using a semi-quantitative method with egg yolk, and the proteinase activity with a semi-quantitative method using bovine serum albumin agar. The study was performed on 96 isolates of M. pachydermatis, 43 isolated from dogs without clinical symptoms of otitis, and 52 isolated from dogs with otitis. In our study, 75.8% of the isolates showed phospholipase activity at pH 6.3, and 81 and 97.9% of them showed proteinase activity measured at pH 6.3 and 6.8, respectively. A higher phospholipase activity was detected in strains isolated from dogs with otitis. The proteinase activity was increased at a pH of 6.8 (97.9%) in comparison to a pH of 6.3 (81%). Our results suggest that the phospholipase activity may play an important role in the invasion of host tissues in chronic canine otitis cases. The proteinase activity results obtained in this study suggest that a reduction in the pH of the treatment may improve its efficacy in the resolution of M. pachydermatis otitis. Copyright © 2012 Revista Iberoamericana de Micología. Published by Elsevier Espana. All rights reserved.

Activation of immunity, immune response, antioxidant ability, and resistance against Vibrio alginolyticus in white shrimp Litopenaeus vannamei decrease under long-term culture at low pH.

PubMed

Chen, Yu-Yuan; Chen, Jiann-Chu; Tseng, Kuei-Chi; Lin, Yong-Chin; Huang, Chien-Lun

2015-10-01

The growth, activation of immunity, immune parameters, and transcript levels of cytMnSOD, mtMnSOD, ecCuZnSOD, glutathione peroxidase (GPx), catalase, lysozyme, and penaeidin 3a were examined in white shrimp Litopenaeus vannamei reared at pH 6.8 and 8.1 after 24 weeks. No significant difference in growth was observed between the two groups. An in vitro study indicated that phenoloxidase activity and respiratory bursts (RB, release of the superoxide anion) were significantly higher in the haemocytes of pH 8.1 shrimp (shrimp reared at pH 8.1) than in pH 6.8 shrimp (shrimp reared at pH 6.8). An in vivo study indicated that the levels of immune parameters of pH 8.1 shrimp were significantly higher than in pH 6.8 shrimp, and the transcript levels of cytMnSOD, ecCuZnSOD, glutathione peroxidase, lysozyme, and penaeidin 3a were down-regulated in pH 6.8 shrimp. In another experiment, shrimp reared at pH 6.8 and 8.1 for 24 weeks were challenged with Vibrio alginolyticus. The mortality rate of pH 6.8 shrimp was significantly higher than in pH 8.1 shrimp over 12-168 h. Phagocytic activity, phagocytic index, and clearance efficiency to V. alginolyticus were significantly lower in pH 6.8 shrimp. We concluded that shrimp under long-term culture at pH 6.8 exhibited decreased resistance against V. alginolyticus as evidenced by reductions in the activation of immunity and immune parameters together with decreased transcript levels of cytMnSOD, ecCuZnSOD, GPx, lysozyme, and penaeidin 3a. Copyright © 2015 Elsevier Ltd. All rights reserved.

Monitoring microbial metabolites using an inductively coupled resonance circuit

NASA Astrophysics Data System (ADS)

Karnaushenko, Daniil; Baraban, Larysa; Ye, Dan; Uguz, Ilke; Mendes, Rafael G.; Rümmeli, Mark H.; de Visser, J. Arjan G. M.; Schmidt, Oliver G.; Cuniberti, Gianaurelio; Makarov, Denys

2015-08-01

We present a new approach to monitor microbial population dynamics in emulsion droplets via changes in metabolite composition, using an inductively coupled LC resonance circuit. The signal measured by such resonance detector provides information on the magnetic field interaction with the bacterial culture, which is complementary to the information accessible by other detection means, based on electric field interaction, i.e. capacitive or resistive, as well as optical techniques. Several charge-related factors, including pH and ammonia concentrations, were identified as possible contributors to the characteristic of resonance detector profile. The setup enables probing the ionic byproducts of microbial metabolic activity at later stages of cell growth, where conventional optical detection methods have no discriminating power.

Copper removal by algae Gelidium, agar extraction algal waste and granulated algal waste: kinetics and equilibrium.

PubMed

Vilar, Vítor J P; Botelho, Cidália M S; Boaventura, Rui A R

2008-03-01

Biosorption of copper ions by an industrial algal waste, from agar extraction industry has been studied in a batch system. This biosorbent was compared with the algae Gelidium itself, which is the raw material for agar extraction, and the industrial waste immobilized with polyacrylonitrile (composite material). The effects of contact time, pH, ionic strength (IS) and temperature on the biosorption process have been studied. Equilibrium data follow both Langmuir and Langmuir-Freundlich models. The parameters of Langmuir equilibrium model were: q(max)=33.0mgg(-1), K(L)=0.015mgl(-1); q(max)=16.7mgg(-1), K(L)=0.028mgl(-1) and q(max)=10.3mgg(-1), K(L)=0.160mgl(-1) respectively for Gelidium, algal waste and composite material at pH=5.3, T=20 degrees C and IS=0.001M. Increasing the pH, the number of deprotonated active sites increases and so the uptake capacity of copper ions. In the case of high ionic strengths, the contribution of the electrostatic component to the overall binding decreases, and so the uptake capacity. The temperature has little influence on the uptake capacity principally for low equilibrium copper concentrations. Changes in standard enthalpy, Gibbs energy and entropy during biosorption were determined. Kinetic data at different solution pH (3, 4 and 5.3) were fitted to pseudo-first-order and pseudo-second-order models. The adsorptive behaviour of biosorbent particles was modelled using a batch reactor mass transfer kinetic model, which successfully predicts Cu(II) concentration profiles.

Decolorization of Anthraquinonic Dyes from Textile Effluent Using Horseradish Peroxidase: Optimization and Kinetic Study

PubMed Central

Šekuljica, Nataša Ž.; Prlainović, Nevena Ž.; Stefanović, Andrea B.; Žuža, Milena G.; Čičkarić, Dragana Z.; Mijin, Dušan Ž.; Knežević-Jugović, Zorica D.

2015-01-01

Two anthraquinonic dyes, C.I. Acid Blue 225 and C.I. Acid Violet 109, were used as models to explore the feasibility of using the horseradish peroxidase enzyme (HRP) in the practical decolorization of anthraquinonic dyes in wastewater. The influence of process parameters such as enzyme concentration, hydrogen peroxide concentration, temperature, dye concentration, and pH was examined. The pH and temperature activity profiles were similar for decolorization of both dyes. Under the optimal conditions, 94.7% of C.I. Acid Violet 109 from aqueous solution was decolorized (treatment time 15 min, enzyme concentration 0.15 IU/mL, hydrogen peroxide concentration 0.4 mM, dye concentration 30 mg/L, pH 4, and temperature 24°C) and 89.36% of C.I. Acid Blue 225 (32 min, enzyme concentration 0.15 IU/mL, hydrogen peroxide concentration 0.04 mM, dye concentration 30 mg/L, pH 5, and temperature 24°C). The mechanism of both reactions has been proven to follow the two substrate ping-pong mechanism with substrate inhibition, revealing the formation of a nonproductive or dead-end complex between dye and HRP or between H2O2 and the oxidized form of the enzyme. Both chemical oxygen demand and total organic carbon values showed that there was a reduction in toxicity after the enzymatic treatment. This study verifies the viability of use of horseradish peroxidase for the wastewaters treatment of similar anthraquinonic dyes. PMID:25685837

Comparative Genomic and Transcriptomic Analysis of Wangiella dermatitidis, A Major Cause of Phaeohyphomycosis and a Model Black Yeast Human Pathogen

PubMed Central

Chen, Zehua; Martinez, Diego A.; Gujja, Sharvari; Sykes, Sean M.; Zeng, Qiandong; Szaniszlo, Paul J.; Wang, Zheng; Cuomo, Christina A.

2014-01-01

Black or dark brown (phaeoid) fungi cause cutaneous, subcutaneous, and systemic infections in humans. Black fungi thrive in stressful conditions such as intense light, high radiation, and very low pH. Wangiella (Exophiala) dermatitidis is arguably the most studied phaeoid fungal pathogen of humans. Here, we report our comparative analysis of the genome of W. dermatitidis and the transcriptional response to low pH stress. This revealed that W. dermatitidis has lost the ability to synthesize alpha-glucan, a cell wall compound many pathogenic fungi use to evade the host immune system. In contrast, W. dermatitidis contains a similar profile of chitin synthase genes as related fungi and strongly induces genes involved in cell wall synthesis in response to pH stress. The large portfolio of transporters may provide W. dermatitidis with an enhanced ability to remove harmful products as well as to survive on diverse nutrient sources. The genome encodes three independent pathways for producing melanin, an ability linked to pathogenesis; these are active during pH stress, potentially to produce a barrier to accumulated oxidative damage that might occur under stress conditions. In addition, a full set of fungal light-sensing genes is present, including as part of a carotenoid biosynthesis gene cluster. Finally, we identify a two-gene cluster involved in nucleotide sugar metabolism conserved with a subset of fungi and characterize a horizontal transfer event of this cluster between fungi and algal viruses. This work reveals how W. dermatitidis has adapted to stress and survives in diverse environments, including during human infections. PMID:24496724

Variation in pH optima of hydrolytic enzyme activities in tropical rain forest soils.

PubMed

Turner, Benjamin L

2010-10-01

Extracellular enzymes synthesized by soil microbes play a central role in the biogeochemical cycling of nutrients in the environment. The pH optima of eight hydrolytic enzymes involved in the cycles of carbon, nitrogen, phosphorus, and sulfur, were assessed in a series of tropical forest soils of contrasting pH values from the Republic of Panama. Assays were conducted using 4-methylumbelliferone-linked fluorogenic substrates in modified universal buffer. Optimum pH values differed markedly among enzymes and soils. Enzymes were grouped into three classes based on their pH optima: (i) enzymes with acidic pH optima that were consistent among soils (cellobiohydrolase, β-xylanase, and arylsulfatase), (ii) enzymes with acidic pH optima that varied systematically with soil pH, with the most acidic pH optima in the most acidic soils (α-glucosidase, β-glucosidase, and N-acetyl-β-glucosaminidase), and (iii) enzymes with an optimum pH in either the acid range or the alkaline range depending on soil pH (phosphomonoesterase and phosphodiesterase). The optimum pH values of phosphomonoesterase were consistent among soils, being 4 to 5 for acid phosphomonoesterase and 10 to 11 for alkaline phosphomonoesterase. In contrast, the optimum pH for phosphodiesterase activity varied systematically with soil pH, with the most acidic pH optima (3.0) in the most acidic soils and the most alkaline pH optima (pH 10) in near-neutral soils. Arylsulfatase activity had a very acidic optimum pH in all soils (pH ≤3.0) irrespective of soil pH. The differences in pH optima may be linked to the origins of the enzymes and/or the degree of stabilization on solid surfaces. The results have important implications for the interpretation of hydrolytic enzyme assays using fluorogenic substrates.

Physiological profiling of soil microbial communities in a Florida scrub-oak ecosystem: spatial distribution and nutrient limitations.

PubMed

Brown, Alisha L P; Garland, Jay L; Day, Frank P

2009-01-01

Rapid physiological profiling of heterotrophic microbial communities enables intensive analysis of the factors affecting activity in aerobic habitats, such as soil. Previous methods for performing such profiling were severely limited due to enrichment bias and inflexibility in incubation conditions. We tested a new physiological profiling approach based on a microtiter plate oxygen sensor system (Becton Dickinson Oxygen Biosensor System (BDOBS)), which allows for testing of lower substrate addition (i.e., lower enrichment potential) and manipulation of physiochemical assay conditions, such as pH and nutrients. Soil microbial communities associated with a scrub-oak forest ecosystem on Merritt Island Wildlife Refuge in central Florida, USA, were studied in order to evaluate microbial activity in a nutrient poor soil and to provide baseline data on the site for subsequent evaluation of the effects of elevated CO(2) on ecosystem function. The spatial variation in physiological activity amongst different habitats (litter, bulk soil, and rhizosphere) was examined as a function of adaptation to local resources (i.e., water soluble extracts of roots and leaf litter) and the degree of N and P limitation. All the communities were primarily N-limited, with a secondary P limitation, which was greater in the rhizosphere and bulk soil. The litter community showed greater overall oxygen consumption when exposed to litter extracts relative to the rhizosphere or soil, suggesting acclimation toward greater use of the mixed substrates in the extract. Root extracts were readily used by communities from all the habitats with no habitat specific acclimation observed. A priming effect was detected in all habitats; addition of glucose caused a significant increase in the use of soil organic carbon. Response to added glucose was only observed with N and P addition, suggesting that C may be lost to the groundwater from these porous soils because nutrient limitation prevents C immobilization.

Ultrasound promoted synthesis of 2-imidazolines in water: a greener approach toward monoamine oxidase inhibitors.

PubMed

Sant' Anna, Gabriela da S; Machado, Pablo; Sauzem, Patricia D; Rosa, Fernanda A; Rubin, Maribel A; Ferreira, Juliano; Bonacorso, Helio G; Zanatta, Nilo; Martins, Marcos A P

2009-01-15

A series of sixteen 2-substituted-2-imidazolines (where the substituent R=Ph, Me-4-Ph; MeO-4-Ph; (MeO)(2)-3,4-Ph; (MeO)(3)-3,4,5-Ph; Ph-4-O-C(O)-Ph; Cl-4-Ph; Cl-2-Ph; Cl(2)-2,4-Ph; NO(2)-4-Ph; NO(2)-3-Ph; Naphth-2-yl; Fur-2-yl; Benzofur-2-yl; Pyridin-2-yl; Quinolin-2-yl) has been synthesized from the reaction of the substituted-aldehydes and ethylenediamine by ultrasound irradiation with NBS in an aqueous medium in high yields (80-99%). The 2-imidazoline ability to inhibit the activity of the A and B isoforms of monoamine oxidase (MAO) was investigated and some of them showed potent and selective MAO inhibitory activity especially for the MAO-B isoform and could become promising candidates for future development.

Physical stability comparisons of IgG1-Fc variants: effects of N-glycosylation site occupancy and Asp/Gln residues at site Asn 297

PubMed Central

KIM, JAE HYUN; JOSHI, SANGEETA B.; MIDDAUGH, C. RUSSELL; TOLBERT, THOMAS J.; VOLKIN, DAVID B.

2014-01-01

The structural integrity and conformational stability of various IgG1-Fc proteins produced from the yeast Pichia pastoris with different glycosylation site occupancy (di-, mono-, and non- glycosylated) was determined. In addition, the physical stability profiles of three different forms of non-glycosylated Fc molecules (varying amino acid residues at site 297 in the CH2 domain due to point mutations and enzymatic digestion of the Fc glycoforms) were also examined. The physical stability of these IgG1-Fc glycoproteins was examined as a function of pH and temperature by high throughput biophysical analysis using multiple techniques combined with data visualization tools (three index empirical phase diagrams and radar charts). Across the pH range of 4.0 to 6.0, the di- and mono- glycosylated forms of the IgG1-Fc showed the highest and lowest levels of physical stability respectively, with the non-glycosylated forms showing intermediate stability depending on solution pH. In the aglycosylated Fc proteins, the introduction of Asp (D) residues at site 297 (QQ vs. DN vs. DD forms) resulted in more subtle changes in structural integrity and physical stability depending on solution pH. The utility of evaluating the conformational stability profile differences between the various IgG1-Fc glycoproteins is discussed in the context of analytical comparability studies. PMID:24740840

Production and characterisation of whey protein hydrolysate having antioxidant activity from cheese whey.

PubMed

Athira, Syamala; Mann, Bimlesh; Saini, Prerna; Sharma, Rajan; Kumar, Rajesh; Singh, Ashish Kumar

2015-11-01

Cheese whey is a rich by-product in nutritional terms, possessing components with high biological value, excellent functional properties, and an inert flavour profile. In the present study, mozzarella cheese whey was ultra-filtrated to remove lactose and mineral. The retentate was hydrolysed with food-grade enzyme alcalase and the hydrolysis conditions (pH, temperature and time) were optimised for maximum antioxidant activity using response surface methodology. Whey protein hydrolysed for 8 h at pH 9 and 55 °C showed a maximum antioxidant activity of 1.18 ± 0.015 µmol Trolox mg(-1) protein. The antioxidant peptides were further enriched by ultra-filtration through a 3 kDa membrane. Seven peptides - β-Lg f(123-131), β-Lg f(122-131), β-Lg f(124-131), β-Lg f(123-134), β-Lg f(122-131), β-Lg f(96-100) and β-Lg f(94-100) - were identified by LC-MS/MS in the 3 kDa permeate of the hydrolysate. The incorporation of whey protein hydrolysate (WPH) in lemon whey drink (5-10 g L(-1)) increased the antioxidant activity from 76% to 90% as compared to control. Hydrolysis of ultra-filtrated retentate of whey can be an energy- and cost-effective method for the direct production of WPH from whey compared to the industrial production of WPH from whey protein concentrate. This study suggests that WPH with good nutritional and biological properties can be effectively used in health-promoting foods as a biofunctional ingredient. © 2014 Society of Chemical Industry.

Effect of low extracellular pH on NF-κB activation in macrophages.

PubMed

Gerry, A B; Leake, D S

2014-04-01

Many diseases, including atherosclerosis, involve chronic inflammation. The master transcription factor for inflammation is NF-κB. Inflammatory sites have a low extracellular pH. Our objective was to demonstrate the effect of pH on NF-κB activation and cytokine secretion. Mouse J774 macrophages or human THP-1 or monocyte-derived macrophages were incubated at pH 7.0-7.4 and inflammatory cytokine secretion and NF-κB activity were measured. A pH of 7.0 greatly decreased pro-inflammatory cytokine secretion (TNF or IL-6) by J774 macrophages, but not THP-1 or human monocyte-derived macrophages. Upon stimulation of mouse macrophages, the levels of IκBα, which inhibits NF-κB, fell but low pH prevented its later increase, which normally restores the baseline activity of NF-κB, even though the levels of mRNA for IκBα were increased. pH 7.0 greatly increased and prolonged NF-κB binding to its consensus promoter sequence, especially the anti-inflammatory p50:p50 homodimers. Human p50 was overexpressed using adenovirus in THP-1 macrophages and monocyte-derived macrophages to see if it would confer pH sensitivity to NF-κB activity in human cells. Overexpression of p50 increased p50:p50 DNA-binding and in THP-1 macrophages inhibited considerably TNF and IL-6 secretion, but there was still no effect of pH on p50:p50 DNA binding or cytokine secretion. A modest decrease in pH can sometimes have marked effects on NF-κB activation and cytokine secretion and might be one reason to explain why mice normally develop less atherosclerosis than do humans. Copyright © 2014 The Authors. Published by Elsevier Ireland Ltd.. All rights reserved.

Carbonic Anhydrase Activity Monitored In Vivo by Hyperpolarized 13C-Magnetic Resonance Spectroscopy Demonstrates Its Importance for pH Regulation in Tumors.

PubMed

Gallagher, Ferdia A; Sladen, Helen; Kettunen, Mikko I; Serrao, Eva M; Rodrigues, Tiago B; Wright, Alan; Gill, Andrew B; McGuire, Sarah; Booth, Thomas C; Boren, Joan; McIntyre, Alan; Miller, Jodi L; Lee, Shen-Han; Honess, Davina; Day, Sam E; Hu, De-En; Howat, William J; Harris, Adrian L; Brindle, Kevin M

2015-10-01

Carbonic anhydrase buffers tissue pH by catalyzing the rapid interconversion of carbon dioxide (CO2) and bicarbonate (HCO3 (-)). We assessed the functional activity of CAIX in two colorectal tumor models, expressing different levels of the enzyme, by measuring the rate of exchange of hyperpolarized (13)C label between bicarbonate (H(13)CO3(-)) and carbon dioxide ((13)CO2), following injection of hyperpolarized H(13)CO3(-), using (13)C-magnetic resonance spectroscopy ((13)C-MRS) magnetization transfer measurements. (31)P-MRS measurements of the chemical shift of the pH probe, 3-aminopropylphosphonate, and (13)C-MRS measurements of the H(13)CO3(-)/(13)CO2 peak intensity ratio showed that CAIX overexpression lowered extracellular pH in these tumors. However, the (13)C measurements overestimated pH due to incomplete equilibration of the hyperpolarized (13)C label between the H(13)CO3(-) and (13)CO2 pools. Paradoxically, tumors overexpressing CAIX showed lower enzyme activity using magnetization transfer measurements, which can be explained by the more acidic extracellular pH in these tumors and the decreased activity of the enzyme at low pH. This explanation was confirmed by administration of bicarbonate in the drinking water, which elevated tumor extracellular pH and restored enzyme activity to control levels. These results suggest that CAIX expression is increased in hypoxia to compensate for the decrease in its activity produced by a low extracellular pH and supports the hypothesis that a major function of CAIX is to lower the extracellular pH. ©2015 American Association for Cancer Research.

The complex allosteric and redox regulation of the fumarate hydratase and malate dehydratase reactions of Arabidopsis thaliana Fumarase 1 and 2 gives clues for understanding the massive accumulation of fumarate.

PubMed

Zubimendi, Juan P; Martinatto, Andrea; Valacco, Maria P; Moreno, Silvia; Andreo, Carlos S; Drincovich, María F; Tronconi, Marcos A

2018-06-01

Arabidopsis thaliana possesses two fumarase genes (FUM), AtFUM1 (At2g47510) encoding for the mitochondrial Krebs cycle-associated enzyme and AtFUM2 (At5g50950) for the cytosolic isoform required for fumarate massive accumulation. Here, the comprehensive biochemical studies of AtFUM1 and AtFUM2 shows that they are active enzymes with similar kinetic parameters but differential regulation. For both enzymes, fumarate hydratase (FH) activity is favored over the malate dehydratase (MD) activity; however, MD is the most regulated activity with several allosteric activators. Oxalacetate, glutamine, and/or asparagine are modulators causing the MD reaction to become preferred over the FH reaction. Activity profiles as a function of pH suggest a suboptimal FUM activity in Arabidopsis cells; moreover, the direction of the FUM reaction is sensitive to pH changes. Under mild oxidation conditions, AtFUMs form high mass molecular aggregates, which present both FUM activities decreased to a different extent. The biochemical properties of oxidized AtFUMs (oxAtFUMs) were completely reversed by NADPH-supplied Arabidopsis leaf extracts, suggesting that the AtFUMs redox regulation can be accomplished in vivo. Mass spectrometry analyses indicate the presence of an active site-associated intermolecular disulfide bridge in oxAtFUMs. Finally, a phylogenetic approach points out that other plant species may also possess cytosolic FUM2 enzymes mainly encoded by paralogous genes, indicating that the evolutionary history of this trait has been drawn through a process of parallel evolution. Overall, according to our results, a multilevel regulatory pattern of FUM activities emerges, supporting the role of this enzyme as a carbon flow monitoring point through the organic acid metabolism in plants. © 2018 Federation of European Biochemical Societies.

pH Effects on solubility, zeta potential, and correlation between antibacterial activity and molecular weight of chitosan.

PubMed

Chang, Shun-Hsien; Lin, Hong-Ting Victor; Wu, Guan-James; Tsai, Guo Jane

2015-12-10

Six chitosans with molecular weights (MWs) of 300, 156, 72.1, 29.2, 7.1, and 3.3 kDa were prepared by cellulase degradation of chitosan (300 kDa) and ultrafiltration techniques. We examined the correlation between activity against Escherichia coli and Staphylococcus aureus and chitosan MW, and provided the underlying explanation. In acidic pH conditions, the chitosan activity increased with increasing MW, irrespective of the temperature and bacteria tested. However, at neutral pH, chitosan activity increased as the MW decreased, and little activity was observed for chitosans with MW >29.2 kDa. At pH 5.0 and 6.0, chitosans exhibited good water solubility and zeta potential (ZP) decreased with the MW, whereas the solubility and ZP of the chitosans decreased with increasing MW at pH 7.0. Particularly, low solubility and negative ZP values were determined for chitosans with MW >29.2 kDa, which may explain the loss of their antibacterial activity at pH 7.0. Copyright © 2015 Elsevier Ltd. All rights reserved.

Surface plasmon resonance measurement of pH-induced responses of immobilized biomolecules: conformational change or electrostatic interaction effects?

PubMed

Paynter, Sally; Russell, David A

2002-10-01

Recently, the observation of pH-induced conformational changes of biomolecules supported on carboxymethyldextran (CMD)-coated surfaces measured using surface plasmon resonance (SPR) has been reported. However, it is apparent that the evidence reported in the literature is ambiguous. The research presented in this paper describes investigations to study the changing SPR signal of immobilized biomolecules as a function of varying pH, to provide a detailed understanding of the origin of the pH-induced changes in the SPR profile. SPR measurements were performed with cytochrome c, concanavalin A, and poly-L-lysine, biomolecules that exhibit diverse conformational responses to changing pH, covalently immobilized onto CMD-coated supports. These SPR measurements were supported by circular dichroism (CD) solution studies. The SPR profiles recorded were not consistent with the conformational transitions of the biomolecules as observed using CD. An alternative explanation for the observed shifts in SPR is proposed, which explains the SPR profiles in terms of electrostatic interaction effects between the immobilized biomolecules and the carboxymethyldextran matrix.

Double-Track Electrochemical Green Approach for Simultaneous Dissolution Profiling of Naproxen Sodium and Diphenhydramine Hydrochloride.

PubMed

Shehata, Mostafa A; Fawaz, Esraa M; El-Rahman, Mohamed K Abd; Abdel-Moety, Ezzat M

2017-11-30

Acquisition of the dissolution profiles of more than single active ingredient in a multi-analyte pharmaceutical formulation is a mandatory manufacturing practice that is dominated by utilization of the off-line separation-based chromatographic methods. This contribution adopts a new "Double-Track" approach with the ultimate goal of advancing the in-line potentiometric sensors to their most effective applicability for simultaneous acquisition of the dissolution profiles of two active ingredients in a binary pharmaceutical formulation. The unique abilities of these sensors for real-time measurements is the key driver for adoption of "green analytical chemistry" (GAC) principles aiming to expand the application of eco-friendly analytical methods With the aim of performing a side-by-side comparison, this work investigates the degree of adherence of ISEs to the 12 principles of GAC in multicomponent dissolution profiling with respect to the HPLC. For the proof of concept, a binary mixture of naproxen sodium (NAPR) and diphenhydramine hydrochloride (DIPH) marketed as Aleve pm ® tablets was selected as a model for which dissolution profiles were attained by two techniques. The first "Double-Track" in-line strategy depends on dipping two highly integrated membrane sensors for continuous monitoring of the dissolution of each active pharmaceutical ingredient (API) by tracing the e.m.f change over the time scale. For the determination of NAPR, sensor I was developed using tridodecyl methyl ammonium chloride as an anion exchanger, while sensor II was developed for the determination of DIPH using potassium tetrakis (4-chlorophenyl) borate as a cation exchanger. The second off-line strategy utilizes a separation-based HPLC method via off-line tracking the increase of peak area by UV detection at 220nm over time using a mobile phase of acetonitrile: water (90:10) pH 3. The advantages of the newly introduced "Double-Track" approach regarding GAC principles are highlighted, and the merits of these benign real-time analyzers (ISEs) that can deliver equivalent analytical results as HPLC while significantly reducing solvent consumption/waste generation are described. Copyright © 2017 Elsevier B.V. All rights reserved.

Food preservative potential of essential oils and fractions from Cymbopogon citratus, Ocimum gratissimum and Thymus vulgaris against mycotoxigenic fungi.

PubMed

Nguefack, J; Dongmo, J B Lekagne; Dakole, C D; Leth, V; Vismer, H F; Torp, J; Guemdjom, E F N; Mbeffo, M; Tamgue, O; Fotio, D; Zollo, P H Amvam; Nkengfack, A E

2009-05-31

The food preservative potential of essential oils from three aromatic plants Cymbopogon citratus, Ocimum gratissimum and Thymus vulgaris and their fractions was investigated against two mycotoxigenic strains each of Aspergillus ochraceus, Penicillium expansum and P. verrucosum. The fungicidal activity was determined and expressed as a Number of Decimal Reduction of the colony forming units per ml (NDR cfu). The influence of pH variation on this activity was studied. The NDR cfu varied with the essential oils and its concentration, the pH of the medium and the strain tested. The essential oils from O. gratissimum exhibited the highest activity against the six fungal strains under the three pH tested. T. vulgaris and C. citratus essential oils were less active against the Penicillium species tested and A. ochraceus, respectively. Potassium sorbate did not present any activity at pH 6 and 9. At pH 3, its NDR cfu was the lowest against the six fungal strains. At the same pH and at 4000 ppm, the three essential oils presented a NRD cfu > or = 6 against strains of A. ochraceus and P. expansum. The same result was obtained with T. vulgaris and C. citratus at 8000 ppm against both strains of P. verrucosum. The highest activity of the three essential oils was recorded at pH 3 against A. ochraceus strains and at pH 9 against both species of Penicillium. From the fractionation, three active fractions were obtained each from C. citratus and O. gratissimum, and two active fractions from T. vulgaris. These active fractions exhibited a NDR cfu, two to seven folds higher than that of the complete essential oils.

Associations among body composition, inflammatory profile and disease extent in ulcerative colitis patients.

PubMed

Urbano, Ana Paula Signori; Sassaki, Ligia Yukie; Dorna, Mariana de Souza; Presti, Paula Torres; Carvalhaes, Maria Antonieta de Barros Leite; Martini, Ligia Araújo; Ferreira, Ana Lucia Anjos

2018-02-01

The aim of our study was to assess body composition status and its association with inflammatory profile and extent of intestinal damage in ulcerative colitis patients during clinical remission. This is a cross-sectional study in which body composition data (phase angle [PhA], fat mass [FM], triceps skin fold thickness [TSFt], mid-arm circumference [MAC], mid-arm muscle circumference [MAMC], adductor pollicis muscle thickness [APMt]), inflammatory profile (C-reactive protein [CRP], a1-acid glycoprotein, erythrocyte sedimentation rate [ESR]) and disease extent were recorded. The mean age of the 59 patients was 48.1 years; 53.3% were women. Most patients were in clinical remission (94.9%) and 3.4% was malnourished according to body mass index. PhA was inversely correlated with inflammatory markers such as CRP (R=-0.59; p<0.001) and ESR (R=-0.46; p<0.001) and directly correlated with lean mass: MAMC (R=0.31; p=0.01) and APMt (R=0.47; p<0.001). Lean mass was inversely correlated with non-specific inflammation marker (APMt vs. ESR) and directly correlated with hemoglobin values (MAMC vs. hemoglobin). Logistic regression analysis revealed that body cell mass was associated with disease extent (OR 0.92; 95CI 0.87-0.97; p<0.01). PhA was inversely correlated with inflammatory markers and directly correlated with lean mass. Acute inflammatory markers were correlated with disease extent. Body cell mass was associated with disease extent.

Activity of earthworm in Latosol under simulated acid rain stress.

PubMed

Zhang, Jia-En; Yu, Jiayu; Ouyang, Ying

2015-01-01

Acid rain is still an issue of environmental concerns. This study investigated the impacts of simulated acid rain (SAR) upon earthworm activity from the Latosol (acidic red soil). Laboratory experiment was performed by leaching the soil columns grown with earthworms (Eisenia fetida) at the SAR pH levels ranged from 2.0 to 6.5 over a 34-day period. Results showed that earthworms tended to escape from the soil and eventually died for the SAR at pH = 2.0 as a result of acid toxicity. The catalase activity in the earthworms decreased with the SAR pH levels, whereas the superoxide dismutases activity in the earthworms showed a fluctuate pattern: decreasing from pH 6.5 to 5.0 and increasing from pH 5.0 to 4.0. Results implied that the growth of earthworms was retarded at the SAR pH ≤ 3.0.

Daily and annually variation of unstimulated whole saliva flow rate and pH and their relation with body profile in healthy young adults.

PubMed

Foglio-Bonda, P L; Migliario, M; Rocchetti, V; Pattarino, F; Foglio-Bonda, A

2013-09-01

To analyse pH and flow rate (FR) of unstimulated whole saliva (UWS), detecting their possible correlations both among themselves and with body profile; in addition to identify daily, annually and gender differences. Eighty-one (47 ♀; 34 ♂) healthy young adults (mean age 22.7±4.09 years old) were enrolled. Saliva was sampled using spitting method. The data were statistically analysed using Pearson's coefficient, ANOVA or Kruskal-Wallis test, Student's t test or the Wilcoxon-Mann-Whitney test. The mean UWS/FR was 0.643 ml/min (range 0.164-1.656 ml/min; percentile 25 = 0.400 ml/min; percentile 50 = 0.643 ml/min, percentile 75 = 0.832 ml/min; median = 0.590 ml/min) and no significant differences were found in gender. The mean UWS/pH was 6.95 (range 6.06-7.91, S.D. 0.28, RSD % 4.08): pH was higher in males (7.02) than females (6.92; p = 0.009). The UWS/FR increased almost steadily during the day: from 0.593 ml/min at 9:00 to 0.669 ml/min at 17:00 (p = 0.04), the greatest increase was found between 9:00 and 11:00. Through the seasons the UWS/FR decreased from summer to spring with a difference of 0.048 ml/min (p < 0.05). The UWS/pH showed a slight increase between 9:00 and 17:00 (p < 0.05). There were little differences in UWS/pH among the seasons (max. 0.09; p < 0.05). Only a significant correlation between UWS/FR and pH was found (R = 0.20; p = 0.008). We did not find correlations between body profile vs UWS/FR or pH. UWS/FR varies more widely than UWS/pH: maintaining a proper acid/base balance is an essential factor for the homeostasis of the oral cavity and probably this would explain the reason for the lack of the variables evaluated influencing UWS/pH.

Probing surface charge potentials of clay basal planes and edges by direct force measurements.

PubMed

Zhao, Hongying; Bhattacharjee, Subir; Chow, Ross; Wallace, Dean; Masliyah, Jacob H; Xu, Zhenghe

2008-11-18

The dispersion and gelation of clay suspensions have major impact on a number of industries, such as ceramic and composite materials processing, paper making, cement production, and consumer product formulation. To fundamentally understand controlling mechanisms of clay dispersion and gelation, it is necessary to study anisotropic surface charge properties and colloidal interactions of clay particles. In this study, a colloidal probe technique was employed to study the interaction forces between a silica probe and clay basal plane/edge surfaces. A muscovite mica was used as a representative of 2:1 phyllosilicate clay minerals. The muscovite basal plane was prepared by cleavage, while the edge surface was obtained by a microtome cutting technique. Direct force measurements demonstrated the anisotropic surface charge properties of the basal plane and edge surface. For the basal plane, the long-range forces were monotonically repulsive within pH 6-10 and the measured forces were pH-independent, thereby confirming that clay basal planes have permanent surface charge from isomorphic substitution of lattice elements. The measured interaction forces were fitted well with the classical DLVO theory. The surface potentials of muscovite basal plane derived from the measured force profiles were in good agreement with those reported in the literature. In the case of edge surfaces, the measured forces were monotonically repulsive at pH 10, decreasing with pH, and changed to be attractive at pH 5.6, strongly suggesting that the charge on the clay edge surfaces is pH-dependent. The measured force profiles could not be reasonably fitted with the classical DLVO theory, even with very small surface potential values, unless the surface roughness was considered. The surface element integration (SEI) method was used to calculate the DLVO forces to account for the surface roughness. The surface potentials of the muscovite edges were derived by fitting the measured force profiles with the surface element integrated DLVO model. The point of zero charge of the muscovite edge surface was estimated to be pH 7-8.

A high resolution spectroscopic study of the oxygen molecule. Ph.D. Thesis Final Report

NASA Technical Reports Server (NTRS)

Ritter, K. J.

1984-01-01

A high resolution spectrometer which incorporates a narrow line width tunable dye laser was used to make absorption profiles of 57 spectral lines in the Oxygen A-Band at pressures up to one atmosphere in pure O2. The observed line profiles are compared to the Voigt, and a collisionally narrowed, profile using a least squares fitting procedure. The collisionally narrowed profile compares more favorable to the observed profiles. Values of the line strengths and self broadening coeffiencients, determined from the least square fitting process, are presented in tabular form. It is found that the experssion by Watson are in closest agreement with the experimentally determined strengths. The self broadening coefficients are compared with the measurements of several other investigators.

Study on Power Ultrasound Optimization and Its Comparison with Conventional Thermal Processing for Treatment of Raw Honey

PubMed Central

2017-01-01

Summary The present study was done to optimize the power ultrasound processing for maximizing diastase activity of and minimizing hydroxymethylfurfural (HMF) content in honey using response surface methodology. Experimental design with treatment time (1-15 min), amplitude (20-100%) and volume (40-80 mL) as independent variables under controlled temperature conditions was studied and it was concluded that treatment time of 8 min, amplitude of 60% and volume of 60 mL give optimal diastase activity and HMF content, i.e. 32.07 Schade units and 30.14 mg/kg, respectively. Further thermal profile analyses were done with initial heating temperatures of 65, 75, 85 and 95 ºC until temperature of honey reached up to 65 ºC followed by holding time of 25 min at 65 ºC, and the results were compared with thermal profile of honey treated with optimized power ultrasound. The quality characteristics like moisture, pH, diastase activity, HMF content, colour parameters and total colour difference were least affected by optimized power ultrasound treatment. Microbiological analysis also showed lower counts of aerobic mesophilic bacteria and in ultrasonically treated honey than in thermally processed honey samples complete destruction of coliforms, yeasts and moulds. Thus, it was concluded that power ultrasound under suggested operating conditions is an alternative nonthermal processing technique for honey. PMID:29540991