Enzymatic hydrolysis of oleuropein from Olea europea (olive) leaf extract and antioxidant activities.

PubMed

Yuan, Jiao-Jiao; Wang, Cheng-Zhang; Ye, Jian-Zhong; Tao, Ran; Zhang, Yu-Si

2015-02-11

Oleuropein (OE), the main polyphenol in olive leaf extract, is likely to decompose into hydroxytyrosol (HT) and elenolic acid under the action of light, acid, base, high temperature. In the enzymatic process, the content of OE in olive leaf extract and enzyme are key factors that affect the yield of HT. A selective enzyme was screened from among 10 enzymes with a high OE degradation rate. A single factor (pH, temperature, time, enzyme quantity) optimization process and a Box-Behnken design were studied for the enzymatic hydrolysis of 81.04% OE olive leaf extract. Additionally, enzymatic hydrolysis results with different substrates (38.6% and 81.04% OE) were compared and the DPPH antioxidant properties were also evaluated. The result showed that the performance of hydrolysis treatments was best using hemicellulase as a bio-catalyst, and the high purity of OE in olive extract was beneficial to biotransform OE into HT. The optimal enzymatic conditions for achieving a maximal yield of HT content obtained by the regression were as follows: pH 5, temperature 55 °C and enzyme quantity 55 mg. The experimental result was 11.31% ± 0.15%, and the degradation rate of OE was 98.54%. From the present investigation of the antioxidant activity determined by the DPPH method, the phenol content and radical scavenging effect were both decreased after enzymatic hydrolysis by hemicellulase. However, a high antioxidant activity of the ethyl acetate extract enzymatic hydrolysate (IC50 = 41.82 μg/mL) was demonstated. The results presented in this work suggested that hemicellulase has promising and attractive properties for industrial production of HT, and indicated that HT might be a valuable biological component for use in pharmaceutical products and functional foods.

13CO2 breath test to measure the hydrolysis of various starch formulations in healthy subjects.

PubMed Central

Hiele, M; Ghoos, Y; Rutgeerts, P; Vantrappen, G; de Buyser, K

1990-01-01

13CO2 starch breath test was used to study the effect of physicochemical characteristics of starch digestion. As starch is hydrolysed to glucose, which is subsequently oxidised to CO2, differences in 13CO2 excretion after ingestion of different starch products must be caused by differences in hydrolysis rate. To study the effect of the degree of chain branching, waxy starch, containing 98% amylopectin, was compared with high amylose starch, containing 30% amylopectin, and normal crystalline starch, containing 74% amylopectin. The effect of the extent of gelatinisation was studied by comparing extruded starch and crystalline starch. Finally, the possible inhibitory effect of adding wheat fibre to extruded starch on the hydrolysis rate was studied. The 13CO2 excretion from two to four hours after intake of crystalline starch was significantly lower than that of extruded starch. Waxy starch was hydrolysed much faster than high amylose starch, but there was no significant difference between waxy starch and normal crystalline starch. Addition of wheat fibre did not influence the hydrolysis rate. The 13CO2 starch breath test is an attractive test for the study of factors affecting carbohydrate assimilation. PMID:2107133

Hydrolysis kinetics in anaerobic degradation of particulate organic material: an overview.

PubMed

Vavilin, V A; Fernandez, B; Palatsi, J; Flotats, X

2008-01-01

The applicability of different kinetics to the hydrolysis of particulate organic material in anaerobic digestion is discussed. Hydrolysis has traditionally been modelled according to the first-order kinetics. For complex substrate, the first-order kinetics should be modified in order to take into account hardly degradable material. It has been shown that models in which hydrolysis is coupled to the growth of hydrolytic bacteria work well at high or at fluctuant organic loading. In particular, the surface-related two-phase and the Contois models showed good fits to experimental data from a wide range of organic waste. Both models tend to the first-order kinetics at a high biomass-to-waste ratio and, for this reason, they can be considered as more general models. Examples on different inhibition processes that might affect the degradation of solid waste are reported. Acetogenesis or methanogenesis might be the rate-limiting stages in complex waste. In such cases, stimulation of hydrolysis (mechanically, chemically or biologically) may lead to a further inhibition of these stages, which ultimately affects hydrolysis as well. Since the hydrolysis process is characterized by surface and transport phenomena, new developments in spatially distributed models are considered fundamental to provide new insights in this complex process.

Modeling residence-time distribution in horizontal screw hydrolysis reactors

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

Sievers, David A.; Stickel, Jonathan J.

The dilute-acid thermochemical hydrolysis step used in the production of liquid fuels from lignocellulosic biomass requires precise residence-time control to achieve high monomeric sugar yields. Difficulty has been encountered reproducing residence times and yields when small batch reaction conditions are scaled up to larger pilot-scale horizontal auger-tube type continuous reactors. A commonly used naive model estimated residence times of 6.2-16.7 min, but measured mean times were actually 1.4-2.2 the estimates. Here, this study investigated how reactor residence-time distribution (RTD) is affected by reactor characteristics and operational conditions, and developed a method to accurately predict the RTD based on key parameters.more » Screw speed, reactor physical dimensions, throughput rate, and process material density were identified as major factors affecting both the mean and standard deviation of RTDs. The general shape of RTDs was consistent with a constant value determined for skewness. The Peclet number quantified reactor plug-flow performance, which ranged between 20 and 357.« less

Modeling residence-time distribution in horizontal screw hydrolysis reactors

DOE PAGES

Sievers, David A.; Stickel, Jonathan J.

2017-10-12

The dilute-acid thermochemical hydrolysis step used in the production of liquid fuels from lignocellulosic biomass requires precise residence-time control to achieve high monomeric sugar yields. Difficulty has been encountered reproducing residence times and yields when small batch reaction conditions are scaled up to larger pilot-scale horizontal auger-tube type continuous reactors. A commonly used naive model estimated residence times of 6.2-16.7 min, but measured mean times were actually 1.4-2.2 the estimates. Here, this study investigated how reactor residence-time distribution (RTD) is affected by reactor characteristics and operational conditions, and developed a method to accurately predict the RTD based on key parameters.more » Screw speed, reactor physical dimensions, throughput rate, and process material density were identified as major factors affecting both the mean and standard deviation of RTDs. The general shape of RTDs was consistent with a constant value determined for skewness. The Peclet number quantified reactor plug-flow performance, which ranged between 20 and 357.« less

Quantitative predictions of bioconversion of aspen by dilute acid and SPORL pretreatments using a unified combined hydrolysis factor (CHF)

Treesearch

W. Zhu; Carl J. Houtman; J.Y. Zhu; Roland Gleisner; K.F. Chen

2012-01-01

A combined hydrolysis factor (CHF) was developed to predict xylan hydrolysis during pretreatments of native aspen (Populus tremuloides) wood chips. A natural extension of previously developed kinetic models allowed us to account for the effect of catalysts by dilute acid and two sulfite pretreatments at different pH values....

Efficiencies of acid catalysts in the hydrolysis of lignocellulosic biomass over a range of combined severity factors

Treesearch

Jae-Won Lee; Thomas W. Jeffries

2011-01-01

Dicarboxylic organic acids have properties that differ from those of sulfuric acid during hydrolysis of lignocellulose. To investigate the effects of different acid catalysts on the hydrolysis and degradation of biomass compounds over a range of thermochemical pretreatments, maleic, oxalic and sulfuric acids were each used at the same combined severity factor (CSF)...

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.

Phospholipase C-mediated hydrolysis of phosphatidylcholine is a target of transforming growth factor beta 1 inhibitory signals.

PubMed Central

Diaz-Meco, M T; Dominguez, I; Sanz, L; Municio, M M; Berra, E; Cornet, M E; Garcia de Herreros, A; Johansen, T; Moscat, J

1992-01-01

Cell growth and tumor transformation can be restrained in certain cell systems by the action of transforming growth factor beta (TGF-beta). It has been established that the mechanism whereby TGF-beta 1 inhibits cell growth does not interfere with the triggering of early mitogenic signal transduction mechanisms. Phospholipase C-catalyzed hydrolysis of phosphatidylcholine (PC) is a relatively late step in the cascade activated by growth factors. Therefore, conceivably activation of phospholipase C-catalyzed hydrolysis of PC could be the target of TGF-beta 1 action. In the study reported here, we demonstrate that TGF-beta 1 inhibits the coupling of ras p21 to the activation of PC hydrolysis, which appears to be critical for the antiproliferative effects of TGF-beta 1. Images PMID:1309592

Aspirin Hydrolysis in Plasma Is a Variable Function of Butyrylcholinesterase and Platelet-activating Factor Acetylhydrolase 1b2 (PAFAH1b2)*

PubMed Central

Zhou, Gang; Marathe, Gopal K.; Hartiala, Jaana; Hazen, Stanley L.; Allayee, Hooman; Tang, W. H. Wilson; McIntyre, Thomas M.

2013-01-01

Aspirin is rapidly hydrolyzed within erythrocytes by a heterodimer of PAFAH1b2/PAFAH1b3 but also in plasma by an unidentified activity. Hydrolysis in both compartments was variable, with a 12-fold variation in plasma among 2226 Cleveland Clinic GeneBank patients. Platelet inhibition by aspirin was suppressed in plasma that rapidly hydrolyzed aspirin. Plasma aspirin hydrolysis was significantly higher in patients with coronary artery disease compared with control subjects (16.5 ± 4.4 versus 15.1 ± 3.7 nmol/ml/min; p = 3.4 × 10−8). A genome-wide association study of 2054 GeneBank subjects identified a single locus immediately adjacent to the BCHE (butyrylcholinesterase) gene associated with plasma aspirin hydrolytic activity (lead SNP, rs6445035; p = 9.1 × 10−17). However, its penetrance was low, and plasma from an individual with an inactivating mutation in BCHE still effectively hydrolyzed aspirin. A second aspirin hydrolase was identified in plasma, the purification of which showed it to be homomeric PAFAH1b2. This is distinct from the erythrocyte PAFAH1b2/PAFAH1b3 heterodimer. Inhibitors showed that both butyrylcholinesterase (BChE) and PAFAH1b2 contribute to aspirin hydrolysis in plasma, with variation primarily reflecting non-genetic variation of BChE activity. Therefore, aspirin is hydrolyzed in plasma by two enzymes, BChE and a new extracellular form of platelet-activating factor acetylhydrolase, PAFAH1b2. Hydrolytic effectiveness varies widely primarily from non-genetic variation of BChE activity that affects aspirin bioavailability in blood and the ability of aspirin to inhibit platelet aggregation. PMID:23508960

Modeling and simulation of an enzymatic reactor for hydrolysis of palm oil.

PubMed

Bhatia, S; Naidu, A D; Kamaruddin, A H

1999-01-01

Hydrolysis of palm oil has become an important process in Oleochemical industries. Therefore, an investigation was carried out for hydrolysis of palm oil to fatty acid and glycerol using immobilized lipase in packed bed reactor. The conversion vs. residence time data were used in Michaelis-Menten rate equation to evaluate the kinetic parameters. A mathematical model for the rate of palm oil hydrolysis was proposed incorporating role of external mass transfer and pore diffusion. The model was simulated for steady-state isothermal operation of immobilized lipase packed bed reactor. The experimental data were compared with the simulated results. External mass transfer was found to affect the rate of palm oil hydrolysis at higher residence time.

Evaluation of chlorine dioxide as a supplementary pretreatment reagent for lignocellulosic biomass.

PubMed

Acharjee, Tapas C; Jiang, Zhihua; Haynes, Robert Daniel; Lee, Yoon Y

2017-11-01

Chlorine dioxide (ClO 2 ) is a bleaching reagent used in paper industry. Two different types of pretreatment methods were investigated incorporating ClO 2 as a secondary reagent: (a) alkaline followed by ClO 2 treatment; (b) dilute-sulfuric acid followed ClO 2 treatment. In these methods, ClO 2 treatment has shown little effect on delignification. Scheme-a has shown a significant improvement in enzymatic digestibility of glucan far above that treated by ammonia alone. On the contrary, dilute-acid followed by ClO 2 treatment has shown negative effect on the enzymatic hydrolysis. The main factors affecting the enzymatic hydrolysis are the changes of the chemical structure of lignin and its distribution on the biomass surface. ClO 2 treatment significantly increases the carboxylic acid content and reduces phenolic groups of lignin, affecting hydrophobicity of lignin and the H-bond induced association between the enzyme and lignin. This collectively led to reduction of unproductive binding of enzyme with lignin, consequently increasing the digestibility. Copyright © 2017 Elsevier Ltd. All rights reserved.

[State of Fungal Lipases of Rhizopus microsporus, Penicillium sp. and Oospora lactis in Border Layers Water-Solid Phase and Factors Affecting Catalytic Properties of Enzymes].

PubMed

Khasanov, Kh T; Davranov, K; Rakhimov, M M

2015-01-01

We demonstrated that a change in the catalytic activity of fungal lipases synthesized by Rhizopus microsporus, Penicillium sp. and Oospora lactis and their ability to absorb on different sorbents depended on the nature of groups on the solid phase surface in the model systems water: lipid and water: solid phase. Thus, the stability of Penicillium sp. lipases increased 85% in the presence ofsorsilen or DEAE-cellulose, and 55% of their initial activity respectively was preserved. In the presence of silica gel and CM-cellulose, a decreased rate of lipid hydrolysis by Pseudomonas sp. enzymes was observed in water medium, and the hydrolysis rate increased by 2.4 and 1.5 times respectively in the presence of aminoaerosil and polykefamid. In an aqueous-alcohol medium, aminoaerosil and polykefamid decreased the rate of substrate hydrolysis by more than 30 times. The addition of aerosil to aqueous and aqueous-alcohol media resulted in an increase in the hydrolysis rate by 1.2-1.3 times. Sorsilen stabilized Penicillium sp. lipase activity at 40, 45, 50 and 55 degrees C. Either stabilization or inactivation of lipases was observed depending on the pH of the medium and the nature of chemical groups localized on the surface of solid phase. The synthetizing activity of lipases also changed depending on the conditions.

Thermal properties of partially hydrolyzed starch-glycerophosphatidylcholine complexes with various acyl chains.

PubMed

Siswoyo, Tri Agus; Morita, Naofumi

2003-05-07

Complexes of starch and monoacyl-sn-glycerophosphatidylcholine (GPC) containing various acyl (myristoyl, palmitoyl, and stearoyl) chains were subjected to hydrolysis with glucoamylase (EC 3.2.1.3). The enzyme hydrolyzed approximately 40% of starch control and 20-28% of starch-GPC complexes. Among the GPCs examined, 1- and 2-monomyristoyl-sn-GPC showed the highest resistance to enzyme hydrolysis, and the hydrolysis rate of starch-GPCs was greater with longer chains. Enzymatic hydrolysis strongly affected the thermal properties of the starch. After enzymatic hydrolysis of starch-GPC complexes for 24 h, their thermograms had broader peaks with lower enthalpies than the corresponding starch without enzyme; however, the starch-GPC complexes showed little change. The surface of starch-GPC granules was less eroded. These results showed that the increasing amount of starch-GPC complexes could be more resistant to hydrolysis.

Study on the technology of compound enzymatic hydrolysis of whole passion fruit

NASA Astrophysics Data System (ADS)

Yang, Yu-xia; Duan, Zhen-hua; Kang, Chao; Zhu, Xiang-hao; Li, Ding-jin

2017-12-01

Fresh Whole Passion Fruit was used as raw material, The enzymatic hydrolysis technology of Passion Fruit by Complex enzyme were studied, The effects of enzyme dosage, Enzyme ratio(cellulose: pectinase), pH, temperature and time on the hydrolysis were investigated by single-tests and orthogonal tests, the hydrolysis indicators of single-factor tests and orthogonal tests were juice yield. The optimal hydrolysis conditions of Passion Fruit by Complex enzyme were enzyme dosage 0.12%, Enzyme ratio 5:1, hydrolysis temperature 50°C, pH4.0 and time 3.5 h. Under such conditions, juice yield of Passion Fruit was 92.91%.

The effect of chemical agents on the turnover of the bound phosphate associated with the sodium-and-potassium ion-stimulated adenosine triphosphatase in ox brain microsomes

PubMed Central

Rodnight, R.

1970-01-01

1. The effect of chemical agents on the turnover of the Na+-dependent bound phosphate and the simultaneous Na+-dependent hydrolysis of ATP by a membrane preparation from ox brain was studied at an ATP/protein ratio of 12.5pmol/μg. 2. The agents were added immediately after phosphorylation of the preparation in a medium containing 50mm-sodium chloride and 2.5μm-[γ-32P]ATP. 3. Concentrations of sodium chloride above 150mm, calcium chloride to 20mm and suramin to 1.4mm inhibited both phosphorylation and dephosphorylation and concomitantly slowed ATP hydrolysis. At 125mm-sodium chloride dephosphorylation and hydrolysis were slightly slowed without affecting phosphorylation. 4. Ethanol to 1.6m concentration inhibited dephosphorylation without affecting phosphorylation; the bound phosphate was increased and ATP hydrolysis slowed. 5. Ouabain to 4mm concentration partially inhibited ATP hydrolysis and caused a transient (1–2s) rise in bound phosphate followed by a rapid fall to a lower plateau value, which eventually declined to zero by the time ATP hydrolysis was complete. 6. Of the detergents examined Lubrol W, Triton X-100 and sodium deoxycholate had no significant effect on turnover. Sodium dodecyl sulphate and sodium decyl sulphate to 3.5mm and 20mm respectively completely inhibited turnover and ATP hydrolysis and stabilized the bound phosphate. PMID:4250238

Hydrolysis products generated by lipoprotein lipase and endothelial lipase differentially impact THP-1 macrophage cell signalling pathways.

PubMed

Essaji, Yasmin; Yang, Yanbo; Albert, Carolyn J; Ford, David A; Brown, Robert J

2013-08-01

Macrophages express lipoprotein lipase (LPL) and endothelial lipase (EL) within atherosclerotic plaques; however, little is known about how lipoprotein hydrolysis products generated by these lipases might affect macrophage cell signalling pathways. We hypothesized that hydrolysis products affect macrophage cell signalling pathways associated with atherosclerosis. To test our hypothesis, we incubated differentiated THP-1 macrophages with products from total lipoprotein hydrolysis by recombinant LPL or EL. Using antibody arrays, we found that the phosphorylation of six receptor tyrosine kinases and three signalling nodes--most associated with atherosclerotic processes--was increased by LPL derived hydrolysis products. EL derived hydrolysis products only increased the phosphorylation of tropomyosin-related kinase A, which is also implicated in playing a role in atherosclerosis. Using electrospray ionization-mass spectrometry, we identified the species of triacylglycerols and phosphatidylcholines that were hydrolyzed by LPL and EL, and we identified the fatty acids liberated by gas chromatography-mass spectrometry. To determine if the total liberated fatty acids influenced signalling pathways, we incubated differentiated THP-1 macrophages with a mixture of the fatty acids that matched the concentrations of liberated fatty acids from total lipoproteins by LPL, and we subjected cell lysates to antibody array analyses. The analyses showed that only the phosphorylation of Akt was significantly increased in response to fatty acid treatment. Overall, our study shows that macrophages display potentially pro-atherogenic signalling responses following acute treatments with LPL and EL lipoprotein hydrolysis products.

Role of asparagine 152 in catalysis of beta-lactam hydrolysis by Escherichia coli AmpC beta-lactamase studied by site-directed mutagenesis.

PubMed

Dubus, A; Normark, S; Kania, M; Page, M G

1995-06-13

The role of asparagine 152 in the catalytic mechanism of Escherichia coli AmpC beta-lactamase has been investigated by site-directed mutagenesis. The residue has been replaced by aspartic acid, glutamic acid, histidine, and leucine. All the substitutions had similar effects on the activity toward substrates and inhibitors. The rate of substrate hydrolysis decreased by factors of 500-5000. The rates of both acylation (2-50-fold decrease) and deacylation (50-500-fold decrease) were affected, indicating a role for Asn152 in both processes. The wild-type AmpC beta-lactamase appears to exist as an equilibrium mixture of two forms, identified by their different kinetic properties. The Asn152 mutations affected the activity of the slow-reacting form much more than that of the fast-reacting form, but they did not appear to affect the interconversion of these two kinetic forms. Comparison of these observations with results obtained with mutation of the equivalent residues in other classes of penicillin-sensitive enzyme indicates that there are quite profound differences between the catalytic mechanisms of these enzymes despite a high degree of conservation of amino acids in the active center, and of the overall three-dimensional structure.

Xylan hydrolysis in Populus trichocarpa × P. deltoides and model substrates during hydrothermal pretreatment.

PubMed

Trajano, Heather L; Pattathil, Sivakumar; Tomkins, Bruce A; Tschaplinski, Timothy J; Hahn, Michael G; Van Berkel, Gary J; Wyman, Charles E

2015-03-01

Previous studies defined easy and difficult to hydrolyze fractions of hemicellulose that may result from bonds among cellulose, hemicellulose, and lignin. To understand how such bonds affect hydrolysis, Populus trichocarpa × Populus deltoides, holocellulose isolated from P. trichocarpa × P. deltoides and birchwood xylan were subjected to hydrothermal flow-through pretreatment. Samples were characterized by glycome profiling, HPLC, and UPLC-MS. Glycome profiling revealed steady fragmentation and removal of glycans from solids during hydrolysis. The extent of polysaccharide fragmentation, hydrolysis rate, and total xylose yield were lowest for P. trichocarpa × P. deltoides and greatest for birchwood xylan. Comparison of results from P. trichocarpa × P. deltoides and holocellulose suggested that lignin-carbohydrate complexes reduce hydrolysis rates and limit release of large xylooligomers. Smaller differences between results with holocellulose and birchwood xylan suggest xylan-cellulose hydrogen bonds limited hydrolysis, but to a lesser extent. These findings imply cell wall structure strongly influences hydrolysis. Copyright © 2014 Elsevier Ltd. All rights reserved.

Subcritical carbon dioxide-water hydrolysis of sugarcane bagasse pith for reducing sugars production.

PubMed

Liang, Jiezhen; Chen, Xiaopeng; Wang, Linlin; Wei, Xiaojie; Wang, Huasheng; Lu, Songzhou; Li, Yunhua

2017-03-01

The aim of present study was to obtain total reducing sugars (TRS) by hydrolysis in subcritical CO 2 -water from sugarcane bagasse pith (SCBP), the fibrous residue remaining after papermaking from sugarcane bagasse. The optimum hydrolysis conditions were evaluated by L 16 (4 5 ) orthogonal experiments. The TRS yield achieved 45.8% at the optimal conditions: 200°C, 40min, 500rmin -1 , CO 2 initial pressure of 1MPa and liquid-to-solid ratio of 50:1. Fourier transform infrared spectrometry and two-dimensional heteronuclear single quantum coherence nuclear magnetic resonance were used to characterize hydrolysis liquor, treated and untreated SCBP, resulting in the removal of hemicelluloses to mainly produce xylose, glucose and arabinose during hydrolysis. The severity factors had no correlation to TRS yield, indicating that the simple kinetic processes of biomass solubilisation cannot perfectly describe the SCBP hydrolysis. The first-order kinetic model based on consecutive reaction was used to obtain rate constants, activation energies and pre-exponential factors. Copyright © 2016 Elsevier Ltd. All rights reserved.

Preparation of the oligosaccharides derived from Flammulina velutipes and their antioxidant activities.

PubMed

Xia, Zhenqiang

2015-03-15

The oligosaccharides were prepared from Flammulina velutipes by hydrolysis of F. velutipes polysaccharides with hydrogen peroxide (H2O2). The yields of F. velutipes derived oligosaccharides (FVOs) were monitored during the hydrolysis process. FVOs yields were affected by three factors, i.e. reaction temperature, H2O2 concentration, and time, which were optimized by using an orthogonal design experiments as follows: reaction temperature 70°C, H2O2 concentration 3%, and reaction time 6h. Under these optimum conditions, the maximal yield of the oligosaccharides reached 17.10%, which was higher than that of hot water extraction method. The oligosaccharides were partially characterized by Fourier transform infrared spectrum, monosaccharide composition, and antioxidant activity. The results indicate that the oligosaccharides derived from F. velutipes showed strong hydroxyl radical activity and reducing capacity at the concentration of 100 μg/mL. Copyright © 2014 Elsevier Ltd. All rights reserved.

Stimulation of phosphatidylcholine breakdown and diacylglycerol production by growth factors in Swiss-3T3 cells.

PubMed Central

Price, B D; Morris, J D; Hall, A

1989-01-01

The effect of a number of growth factors on phosphatidylcholine (PtdCho) turnover in Swiss-3T3 cells was studied. Phorbol 12-myristate 13-acetate (PMA), bombesin, platelet-derived growth factor (PDGF) and vasopressin rapidly stimulated PtdCho hydrolysis, diacylglycerol (DAG) production, and PtdCho synthesis. Insulin and prostaglandin F2 alpha (PGF2 alpha) stimulated PtdCho synthesis, but not its breakdown, whereas epidermal growth factor (EGF) and bradykinin were without effect. Stimulation of PtdCho hydrolysis by the above ligands resulted in increased production of phosphocholine and DAG (due to phospholipase C activity) and significant amounts of choline, suggesting activation of a phospholipase D as well. CDP-choline and glycerophosphocholine levels were unchanged. Down-regulation of protein kinase C with PMA (400 nM, 40 h) abolished the stimulation of PtdCho hydrolysis and PtdCho synthesis by PMA, bombesin, PDGF and vasopressin, but not the stimulation of PtdCho synthesis by insulin and PGF2 alpha. PtdCho hydrolysis therefore occurs predominantly by activation of protein kinase C (either by PMA or PtdIns hydrolysis) leading to elevation of DAG levels derived from non-PtdIns(4,5)P2 sources. PtdCho synthesis occurs by both a protein kinase C-dependent pathway (stimulated by PMA, PDGF, bombesin and vasopressin) and a protein kinase C-independent pathway (stimulated by insulin and PGF2 alpha). DAG production from PtdCho hydrolysis is not the primary signal to activate protein kinase C, but may contribute to long-term activation of this kinase. PMID:2690829

Using a combined hydrolysis factor to optimize high titer ethanol production from sulfite-pretreated poplar without detoxification

Treesearch

Jingzhi Zhang; Feng Gu; J.Y. Zhu; Ronald S. Zalesny Jr.

2015-01-01

Sulfite pretreatment to overcome the recalcitrance of lignocelluloses (SPORL) was applied to poplar NE222 chips in a range of chemical loadings, temperatures, and times. The combined hydrolysis factor (CHF) as a pretreatment severity accurately predicted xylan dissolution by SPORL. Good correlations between CHF and pretreated...

Non-persistent pesticides removal in constructed wetlands

NASA Astrophysics Data System (ADS)

Tu, Yue; Jiang, Lei; Li, Haixiang

2018-03-01

The heavy use of non-persistent pesticides, resulting in the accumulation of environment and destroy the aquatic environment. This paper presents the research status of using CWs to treat non-persistent pesticides in water. The removal mechanisms are mainly physical deposition, chemical hydrolysis and plant absorption. Analysis of the factors that affect the removal effect are mainly the nature of pesticides, HRT, plants. Some scholars have proposed that secondary products of non-persistent pesticides may be more harmful to the environment, However, the relevant reports are scarce.

High-throughput microplate technique for enzymatic hydrolysis of lignocellulosic biomass.

PubMed

Chundawat, Shishir P S; Balan, Venkatesh; Dale, Bruce E

2008-04-15

Several factors will influence the viability of a biochemical platform for manufacturing lignocellulosic based fuels and chemicals, for example, genetically engineering energy crops, reducing pre-treatment severity, and minimizing enzyme loading. Past research on biomass conversion has focused largely on acid based pre-treatment technologies that fractionate lignin and hemicellulose from cellulose. However, for alkaline based (e.g., AFEX) and other lower severity pre-treatments it becomes critical to co-hydrolyze cellulose and hemicellulose using an optimized enzyme cocktail. Lignocellulosics are appropriate substrates to assess hydrolytic activity of enzyme mixtures compared to conventional unrealistic substrates (e.g., filter paper, chromogenic, and fluorigenic compounds) for studying synergistic hydrolysis. However, there are few, if any, high-throughput lignocellulosic digestibility analytical platforms for optimizing biomass conversion. The 96-well Biomass Conversion Research Lab (BCRL) microplate method is a high-throughput assay to study digestibility of lignocellulosic biomass as a function of biomass composition, pre-treatment severity, and enzyme composition. The most suitable method for delivering milled biomass to the microplate was through multi-pipetting slurry suspensions. A rapid bio-enzymatic, spectrophotometric assay was used to determine fermentable sugars. The entire procedure was automated using a robotic pipetting workstation. Several parameters that affect hydrolysis in the microplate were studied and optimized (i.e., particle size reduction, slurry solids concentration, glucan loading, mass transfer issues, and time period for hydrolysis). The microplate method was optimized for crystalline cellulose (Avicel) and ammonia fiber expansion (AFEX) pre-treated corn stover. Copyright 2008 Wiley Periodicals, Inc.

Enhancing enzymatic hydrolysis of sugarcane bagasse by ferric chloride catalyzed organosolv pretreatment and Tween 80.

PubMed

Zhang, Hongdan; Fan, Meishan; Li, Xin; Zhang, Aiping; Xie, Jun

2018-06-01

In this work, a FeCl 3 -catalyzed organosolv pretreatment was employed at 160 °C to remove hemicellulose and lignin in sugarcane bagasse leaving the cellulose-enriched residue for enzymatic hydrolysis to sugars. The solubilized hemicellulose fractions consisted more monomer xylose than oligomer xylose. The FeCl 3 -catalyzed organosolv pretreatment significantly improved the enzymatic hydrolysis, nearly 100% of cellulose components were converted to glucose after pretreatment with 0.05 M FeCl 3 . Structural analysis was employed to reveal how pretreatment affected the enzymatic hydrolysis. With the addition of Tween 80, the same level of glucose was obtained with 50% reduction of enzyme dosage after 24 h. Furthermore, the influence of Tween 80 on different pretreatment systems was investigated, indicating that the improvement was increased as the lignin content increased, decreased with high enzyme loading and extending hydrolysis time. This work suggested that the addition of Tween 80 could improve the enzymatic hydrolysis, reduce the hydrolysis time and enzyme dosage. Copyright © 2018 Elsevier Ltd. All rights reserved.

Technological aspects of lactose-hydrolyzed milk powder.

PubMed

Torres, Jansen Kelis Ferreira; Stephani, Rodrigo; Tavares, Guilherme M; de Carvalho, Antônio Fernandes; Costa, Renata Golin Bueno; de Almeida, Carlos Eduardo Rocha; Almeida, Mariana Ramos; de Oliveira, Luiz Fernando Cappa; Schuck, Pierre; Perrone, Ítalo Tuler

2017-11-01

Few reports describe the effect of lactose hydrolysis on the properties of milk powder during production and storage. Hence, the aim of this study was to evaluate the effects of five different levels of enzymatic lactose hydrolysis during the production and storage of milk powder. As the lactose hydrolysis rate increased, adhesion to the drying chamber also increased, due to higher levels of particle agglomeration. Additionally, more brown powder was obtained when the lactose hydrolysis rate was increased, which in turn negatively affected rehydration ability. Using Raman spectroscopy, crystallization of the lactose residues in various samples was assessed over 6weeks of accelerated aging at a room temperature environment with 75.5% of air moisture. Products with 25% or greater lactose hydrolysis showed no signs of crystallization, in contrast to the non-hydrolyzed sample. Copyright © 2017. Published by Elsevier Ltd.

Thermal conductivity characteristics of dewatered sewage sludge by thermal hydrolysis reaction.

PubMed

Song, Hyoung Woon; Park, Keum Joo; Han, Seong Kuk; Jung, Hee Suk

2014-12-01

The purpose of this study is to quantify the thermal conductivity of sewage sludge related to reaction temperature for the optimal design of a thermal hydrolysis reactor. We continuously quantified the thermal conductivity of dewatered sludge related to the reaction temperature. As the reaction temperature increased, the dewatered sludge is thermally liquefied under high temperature and pressure by the thermal hydrolysis reaction. Therefore, the bound water in the sludge cells comes out as free water, which changes the dewatered sludge from a solid phase to slurry in a liquid phase. As a result, the thermal conductivity of the sludge was more than 2.64 times lower than that of the water at 20. However, above 200, it became 0.704 W/m* degrees C, which is about 4% higher than that of water. As a result, the change in physical properties due to thermal hydrolysis appears to be an important factor for heat transfer efficiency. Implications: The thermal conductivity of dewatered sludge is an important factor the optimal design of a thermal hydrolysis reactor. The dewatered sludge is thermally liquefied under high temperature and pressure by the thermal hydrolysis reaction. The liquid phase slurry has a higher thermal conductivity than pure water.

Hydrolysis of platelet-activating factor by human serum paraoxonase.

PubMed Central

Rodrigo, L; Mackness, B; Durrington, P N; Hernandez, A; Mackness, M I

2001-01-01

Human serum paraoxonase (human PON1) has been shown to be important in the metabolism of phospholipid and cholesteryl ester hydroperoxides, thereby preventing the oxidation of low-density lipoprotein (LDL) and retarding atherogenesis. However, the exact substrate specificity of PON1 has not been established. In the present study we show that purified PON1 hydrolyses platelet-activating factor (PAF). We could find no evidence for contamination of our preparation with authentic platelet-activating-factor acetylhydrolase (PAFAH) by immunoblotting with a PAFAH monoclonal antibody or by sequencing the purified protein. In addition the specific PAFAH inhibitor SB-222657 did not affect the ability of PON1 to hydrolyse PAF (30.1+/-2.8 micromol/min per mg of protein with no inhibitor; 31.4+/-2.2 micromol/min per mg of protein with 100 nM inhibitor) or phenyl acetate (242.6+/-30.8 versus 240.8+/-31.5 micromol/min per mg of protein with and without inhibitor respectively). SB-222657 was also unable to inhibit PAF hydrolysis by isolated human high-density lipoprotein (HDL), but completely abolished the activity of human LDL. Ostrich (Struthio camelus) HDL, which does not contain PON1, was unable to hydrolyse PAF. These data provide evidence that PON1 may limit the action of this bioactive pro-inflammatory phospholipid. PMID:11171072

Interactions between supplemented mineral phosphorus and phytase on phytate hydrolysis and inositol phosphates in the small intestine of broilers1,2.

PubMed

Zeller, Ellen; Schollenberger, Margit; Witzig, Maren; Shastak, Yauheni; Kühn, Imke; Hoelzle, Ludwig E; Rodehutscord, Markus

2015-05-01

Phytate breakdown in the digestive tract of broilers is affected by supplements of mineral phosphorus (P) and phytase with unknown interactions between the 2 factors. It was the objective to study phytate hydrolysis and the presence of inositol phosphate isomers (InsPs) as affected by supplements of mineral P and phytase in the small intestine of broilers. Fifteen-day old broilers were assigned to 48 pens of 20 broilers each (n = 8 pens/treatment). Two low-P corn-soybean meal-based diets without (BD-; 4.4 g P/kg dry matter) or with monocalcium phosphate (MCP; BD+; 5.2 g P/kg dry matter) were supplied without or with added phytase at 500 or 12,500 FTU/kg. On d 24, digesta from the duodenum/jejunum and lower ileum was pooled per segment on a by-pen basis, freeze-dried, and analyzed for P, InsPs, and the marker TiO2. Another 180 broilers (n = 6 pens/treatment, 10 birds each) were fed the 3 BD+ diets from d 1 to 21 to assess the influence of supplemented phytase on tibia mineralization and strength. Significant interactions between MCP and phytase supplements on myo-inositol 1,2,3,4,5,6-hexakis (dihydrogen phosphate) (InsP6) hydrolysis (duodenum/jejunum: P ≤ 0.001; ileum: P = 0.004) and level of specific lower InsPs were detected. Supplementation with 12,500 FTU/kg phytase resulted in 92% InsP6 hydrolysis and strong degradation of InsP5. This treatment resulted in higher P net absorption, affirmed by higher BW gain, tibia strength, and mineralization compared to treatments without or with 500 FTU/kg phytase (P ≤ 0.05). MCP supplementation reduced the degradation of InsP6 and specific lower InsPs in birds fed diets without or with 500 FTU/kg of phytase (P ≤ 0.05), but did not reduce InsP6 hydrolysis or degradation of InsP5 at the high phytase dose. Effects of added MCP on phytase efficacy depend on the dose of supplemented phytase. Differences in the concentrations of lower InsPs indicated that the initial step of InsP6 hydrolysis is not the only catabolic step that is influenced by MCP or phytase levels. © 2015 Poultry Science Association Inc.

Is enzymatic hydrolysis a reliable analytical strategy to quantify glucuronidated and sulfated polyphenol metabolites in human fluids?

PubMed

Quifer-Rada, Paola; Martínez-Huélamo, Miriam; Lamuela-Raventos, Rosa M

2017-07-19

Phenolic compounds are present in human fluids (plasma and urine) mainly as glucuronidated and sulfated metabolites. Up to now, due to the unavailability of standards, enzymatic hydrolysis has been the method of choice in analytical chemistry to quantify these phase II phenolic metabolites. Enzymatic hydrolysis procedures vary in enzyme concentration, pH and temperature; however, there is a lack of knowledge about the stability of polyphenols in their free form during the process. In this study, we evaluated the stability of 7 phenolic acids, 2 flavonoids and 3 prenylflavanoids in urine during enzymatic hydrolysis to assess the suitability of this analytical procedure, using three different concentrations of β-glucuronidase/sulfatase enzymes from Helix pomatia. The results indicate that enzymatic hydrolysis negatively affected the recovery of the precursor and free-form polyphenols present in the sample. Thus, enzymatic hydrolysis does not seem an ideal analytical strategy to quantify glucuronidated and sulfated polyphenol metabolites.

Pretreatment of wheat straw by nonionic surfactant-assisted dilute acid for enhancing enzymatic hydrolysis and ethanol production.

PubMed

Qi, Benkun; Chen, Xiangrong; Wan, Yinhua

2010-07-01

Pretreating wheat straw (WS) with combined use of varied sulfuric acid concentration (0-3%, w/v) and Tween 20 concentration (0-1%) was investigated in an attempt to enhance the hydrolysis and fermentability of pretreated WS. Enzymatic hydrolysis yield of glucan and xylan and ethanol production by simultaneous saccharification and fermentation (SSF) of water-insoluble solids (WIS) were significantly affected by the amount of Tween 20 added during acid pretreatment. Any further addition of Tween 20 in either hydrolysis stage or fermentation stage only led to small increase in glucan conversion and ethanol production. Determination of adsorption of cellulases during hydrolysis showed that Tween 20-assisted acid treated straw solution contained more free cellulases than individual acid treated straw solution, indicating that modification of lignin surface by Tween 20 added during pretreatment likely occurred. In addition, the effects of pretreatment conditions on overall recovery of glucose and xylose after pretreatment and enzymatic hydrolysis were also investigated. Copyright 2010 Elsevier Ltd. All rights reserved.

Effects of copper source and concentration on in vitro phytate phosphorus hydrolysis by phytase.

PubMed

Pang, Yanfang; Applegate, Todd J

2006-03-08

Five copper (Cu) sources were studied at pH 2.5, 5.5, and 6.5 to determine how Cu affects phytate phosphorus (PP) hydrolysis by phytase at concentrations up to 500 mg/kg diet (60 min, 40-41 degrees C). Subsequently, Cu solubility with and without sodium phytate was measured. Adding Cu inhibited PP hydrolysis at pH 5.5 and pH 6.5 (P < 0.05). This inhibition was greater with higher concentrations of Cu. Tri-basic copper chloride and copper lysinate inhibited PP hydrolysis much less than copper sulfate pentahydrate, copper chloride, and copper citrate (P < 0.05). A strong negative relationship was observed between PP hydrolysis and soluble Cu at pH 5.5 (r = -0.76, P < 0.0001) and 6.5 (r = -0.54, P < 0.0001). In conclusion, pH, Cu concentration, and source influenced PP hydrolysis by phytase in vitro and were related to the amount of soluble Cu and the formation of insoluble copper-phytin complexes.

A coarse-grained model for synergistic action of multiple enzymes on cellulose

DOE PAGES

Asztalos, Andrea; Daniels, Marcus; Sethi, Anurag; ...

2012-08-01

In this study, degradation of cellulose to glucose requires the cooperative action of three classes of enzymes, collectively known as cellulases. Endoglucanases randomly bind to cellulose surfaces and generate new chain ends by hydrolyzing -1,4-D-glycosidic bonds. Exoglucanases bind to free chain ends and hydrolyze glycosidic bonds in a processive manner releasing cellobiose units. Then, -glucosidases hydrolyze soluble cellobiose to glucose. Optimal synergistic action of these enzymes is essential for efficient digestion of cellulose. Experiments show that as hydrolysis proceeds and the cellulose substrate becomes more heterogeneous, the overall degradation slows down. As catalysis occurs on the surface of crystalline cellulose,more » several factors affect the overall hydrolysis. Therefore, spatial models of cellulose degradation must capture effects such as enzyme crowding and surface heterogeneity, which have been shown to lead to a reduction in hydrolysis rates. As a result, we present a coarse-grained stochastic model for capturing the key events associated with the enzymatic degradation of cellulose at the mesoscopic level. This functional model accounts for the mobility and action of a single cellulase enzyme as well as the synergy of multiple endo- and exo-cellulases on a cellulose surface. The quantitative description of cellulose degradation is calculated on a spatial model by including free and bound states of both endo- and exo-cellulases with explicit reactive surface terms (e.g., hydrogen bond breaking, covalent bond cleavages) and corresponding reaction rates. The dynamical evolution of the system is simulated by including physical interactions between cellulases and cellulose. In conclusion, our coarse-grained model reproduces the qualitative behavior of endoglucanases and exoglucanases by accounting for the spatial heterogeneity of the cellulose surface as well as other spatial factors such as enzyme crowding. Importantly, it captures the endo-exo synergism of cellulase enzyme cocktails. This model constitutes a critical step towards testing hypotheses and understanding approaches for maximizing synergy and substrate properties with a goal of cost effective enzymatic hydrolysis.« less

Particle size distribution of rice flour affecting the starch enzymatic hydrolysis and hydration properties.

PubMed

de la Hera, Esther; Gomez, Manuel; Rosell, Cristina M

2013-10-15

Rice flour is becoming very attractive as raw material, but there is lack of information about the influence of particle size on its functional properties and starch digestibility. This study evaluates the degree of dependence of the rice flour functional properties, mainly derived from starch behavior, with the particle size distribution. Hydration properties of flours and gels and starch enzymatic hydrolysis of individual fractions were assessed. Particle size heterogeneity on rice flour significantly affected functional properties and starch features, at room temperature and also after gelatinization; and the extent of that effect was grain type dependent. Particle size heterogeneity on rice flour induces different pattern in starch enzymatic hydrolysis, with the long grain having slower hydrolysis as indicated the rate constant (k). No correlation between starch digestibility and hydration properties or the protein content was observed. It seems that in intact granules interactions with other grain components must be taken into account. Overall, particle size fractionation of rice flour might be advisable for selecting specific physico-chemical properties. Copyright © 2013. Published by Elsevier Ltd.

Effects of hydrolysis on solid-state relaxation and stickiness behavior of sodium caseinate-lactose powders.

PubMed

Mounsey, J S; Hogan, S A; Murray, B A; O'Callaghan, D J

2012-05-01

Hydrolyzed or nonhydrolyzed sodium caseinate-lactose dispersions were spray dried, at a protein: lactose ratio of 0.5, to examine the effects of protein hydrolysis on relaxation behavior and stickiness of model powders. Sodium caseinate (NC) used included a nonhydrolyzed control (DH 0) and 2 hydrolyzed variants (DH 8.3 and DH 15), where DH = degree of hydrolysis (%). Prior to spray drying, apparent viscosities of liquid feeds (at 70°C) at a shear rate of 20/s were 37.6, 3.14, and 3.19 mPa·s, respectively, for DH 0, DH 8, and DH 15 dispersions. Powders containing hydrolyzed casein were more susceptible to sticking than those containing intact NC. The former had also lower bulk densities and powder particle sizes. Scanning electron microscopy showed that hydrolyzed powders had thinner particle walls and were more friable than powders containing intact NC. Secondary structure of caseinates, determined by Fourier transform infrared spectroscopy, was affected by the relative humidity of storage and the presence of lactose as co-solvent rather than its physical state. Glass transition temperatures and lactose crystallization temperatures, determined by differential scanning calorimetry were not affected by caseinate hydrolysis, although the effects of protein hydrolysis on glass-rubber transitions (T(gr)) could be determined by thermo-mechanical analysis. Powders containing hydrolyzed NC had lower T(gr) values (~30°C) following storage at a higher subcrystallization relative humidity (33%) compared with powder with nonhydrolyzed NC (T(gr) value of ~40°C), an effect that reflects more extensive plasticization of powder matrices by moisture. Results support that sodium caseinate-lactose interactions were weak but that relaxation behavior, as determined by the susceptibility of powder to sticking, was affected by hydrolysis of sodium caseinate. Copyright © 2012 American Dairy Science Association. Published by Elsevier Inc. All rights reserved.

Enhanced hydrolysis of cellulose hydrogels by morphological modification.

PubMed

Alfassi, Gilad; Rein, Dmitry M; Cohen, Yachin

2017-11-01

Cellulose is one of the most abundant bio-renewable materials on earth, yet the potential of cellulosic bio-fuels is not fully exploited, primarily due to the high costs of conversion. Hydrogel particles of regenerated cellulose constitute a useful substrate for enzymatic hydrolysis, due to their porous and amorphous structure. This article describes the influence of several structural aspects of the cellulose hydrogel on its hydrolysis. The hydrogel density was shown to be directly proportional to the cellulose concentration in the initial solution, thus affecting its hydrolysis rate. Using high-resolution scanning electron microscopy, we show that the hydrogel particles in aqueous suspension exhibit a dense external surface layer and a more porous internal network. Elimination of the external surface layer accelerated the hydrolysis rate by up to sixfold and rendered the process nearly independent of cellulose concentration. These findings may be of practical relevance to saccharification processing costs, by reducing required solvent quantities and enzyme load.

Hydrolysis rate constants at 10-25 °C can be more than doubled by a short anaerobic pre-hydrolysis at 35 °C.

PubMed

Zhang, L; Gao, R; Naka, A; Hendrickx, T L G; Rijnaarts, H H M; Zeeman, G

2016-11-01

Hydrolysis is the first step of the anaerobic digestion of complex wastewater and considered as the rate limiting step especially at low temperature. Low temperature (10-25 °C) hydrolysis was investigated with and without application of a short pre-hydrolysis at 35 °C. Batch experiments were executed using cellulose and tributyrin as model substrates for carbohydrates and lipids. The results showed that the low temperature anaerobic hydrolysis rate constants increased by a factor of 1.5-10, when the short anaerobic pre-hydrolysis at 35 °C was applied. After the pre-hydrolysis phase at 35 °C and decreasing the temperature, no lag phase was observed in any case. Without the pre-hydrolysis, the lag phase for cellulose hydrolysis at 35-10 °C was 4-30 days. Tributyrin hydrolysis showed no lag phase at any temperature. The hydrolysis efficiency of cellulose increased from 40 to 62%, and from 9.6 to 40% after 9.1 days at 15 and 10 °C, respectively, when the pre-hydrolysis at 35 °C was applied. The hydrolysis efficiency of tributyrin at low temperatures with the pre-hydrolysis at 35 °C was similar to those without the pre-hydrolysis. The hydrolytic activity of the supernatant collected from the digestate after batch digestion of cellulose and tributyrin at 35 °C was higher than that of the supernatants collected from the low temperature (≤25 °C) digestates. Copyright © 2016 Elsevier Ltd. All rights reserved.

Effect of acid hydrolysis on morphology, structure and digestion property of starch from Cynanchum auriculatum Royle ex Wight.

PubMed

Wang, Xingchi; Wen, Fanting; Zhang, Shurong; Shen, Ruru; Jiang, Wei; Liu, Jun

2017-03-01

Effect of acid hydrolysis on the morphology, structure and digestion property of starch from Cynanchum auriculatum Royle ex Wight was investigated in this study. The hydrolysis degree of C. auriculatum starch rapidly increased to 63.69% after 4days and reached 78.67% at the end of 9days. Morphology observation showed that the starch granules remained intact during the first 4days of hydrolysis. However, serious erosion phenomenon was observed after 5days and starch granules completely fell into pieces after 7days. During acid hydrolysis process, the crystal type of hydrolyzed starch changed from original C B -type to final A-type. Small-angle X-ray scattering patterns showed the semi-crystalline growth rings started to be hydrolyzed after 4days. The proportions of single helix and amorphous components as well as amylose content in starch gradually decreased, whereas the proportion of double helix components continuously increased during acid hydrolysis. However, the contents of rapidly digestible starch, slowly digestible starch and resistant starch were almost constant during acid hydrolysis process, indicating the in vitro digestion property of C. auriculatum starch was not affected by acid hydrolysis. Our results provided novel information on the inner structure of C. auriculatum starch granules. Copyright © 2017 Elsevier B.V. All rights reserved.

Cooperative mechanism of RNA packaging motor.

PubMed

Lísal, Jirí; Tuma, Roman

2005-06-17

P4 is a hexameric ATPase that serves as the RNA packaging motor in double-stranded RNA bacteriophages from the Cystoviridae family. P4 shares sequence and structural similarities with hexameric helicases. A structure-based mechanism for mechano-chemical coupling has recently been proposed for P4 from bacteriophage phi12. However, coordination of ATP hydrolysis among the subunits and coupling with RNA translocation remains elusive. Here we present detailed kinetic study of nucleotide binding, hydrolysis, and product release by phi12 P4 in the presence of different RNA and DNA substrates. Whereas binding affinities for ATP and ADP are not affected by RNA binding, the hydrolysis step is accelerated and the apparent cooperativity is increased. No nucleotide binding cooperativity is observed. We propose a stochastic-sequential cooperativity model to describe the coordination of ATP hydrolysis within the hexamer. In this model the apparent cooperativity is a result of hydrolysis stimulation by ATP and RNA binding to neighboring subunits rather than cooperative nucleotide binding. The translocation step appears coupled to hydrolysis, which is coordinated among three neighboring subunits. Simultaneous interaction of neighboring subunits with RNA makes the otherwise random hydrolysis sequential and processive.

Extraterrestrial material analysis: loss of amino acids during liquid-phase acid hydrolysis

NASA Astrophysics Data System (ADS)

Buch, Arnaud; Brault, Amaury; Szopa, Cyril; Freissinet, Caroline

2015-04-01

Searching for building blocks of life in extraterrestrial material is a way to learn more about how life could have appeared on Earth. With this aim, liquid-phase acid hydrolysis has been used, since at least 1970 , in order to extract amino acids and other organic molecules from extraterrestrial materials (e.g. meteorites, lunar fines) or Earth analogues (e.g. Atacama desert soil). This procedure involves drastic conditions such as heating samples in 6N HCl for 24 h, either under inert atmosphere/vacuum, or air. Analysis of the hydrolyzed part of the sample should give its total (free plus bound) amino acid content. The present work deals with the influence of the 6N HCl hydrolysis on amino acid degradation. Our experiments have been performed on a standard solution of 17 amino acids. After liquid-phase acid hydrolysis (6N HCl) under argon atmosphere (24 h at 100°C), the liquid phase was evaporated and the dry residue was derivatized with N-Methyl-N-(t-butyldimethylsilyl)trifluoroacetamide (MTBSTFA) and dimethylformamide (DMF), followed by gas chromatography-mass spectrometry analysis. After comparison with derivatized amino acids from the standard solution, a significant reduction of the chromatographic peak areas was observed for most of the amino acids after liquid-phase acid hydrolysis. Furthermore, the same loss pattern was observed when the amino acids were exposed to cold 6N HCl for a short amount of time. The least affected amino acid, i.e. glycine, was found to be 73,93% percent less abundant compared to the non-hydrolyzed standard, while the most affected, i.e. histidine, was not found in the chromatograms after hydrolysis. Our experiments thereby indicate that liquid-phase acid hydrolysis, even under inert atmosphere, leads to a partial or total loss of all of the 17 amino acids present in the standard solution, and that a quick cold contact with 6N HCl is sufficient to lead to a loss of amino acids. Therefore, in the literature, the reported increase of the total quantity of amino acids after acid hydrolysis, due to the formation/release of amino acids during the whole water extraction / liquid-phase acid hydrolysis, could have hidden a loss of amino acids. Thus, in extraterrestrial material studies involving liquid-phase acid hydrolysis, the quantities of total amino acids may have been underestimated.

Hydrothermal pretreatment of palm oil empty fruit bunch

NASA Astrophysics Data System (ADS)

Simanungkalit, Sabar Pangihutan; Mansur, Dieni; Nurhakim, Boby; Agustin, Astrid; Rinaldi, Nino; Muryanto, Fitriady, Muhammad Ariffudin

2017-01-01

Hydrothermal pretreatment methods in 2nd generation bioethanol production more profitable to be developed, since the conventional pretreatment, by using acids or alkalis, is associated with the serious economic and environmental constraints. The current studies investigate hydrothermal pretreatment of palm oil empty fruit bunch (EFB) in a batch tube reactor system with temperature and time range from 160 to 240 C and 15 to 30 min, respectively. The EFB were grinded and separated into 3 different particles sizes i.e. 10 mesh, 18 mesh and 40 mesh, prior to hydrothermal pretreatment. Solid yield and pH of the treated EFB slurries changed over treatment severities. The chemical composition of EFB was greatly affected by the hydrothermal pretreatment especially hemicellulose which decreased at higher severity factor as determined by HPLC. Both partial removal of hemicellulose and migration of lignin during hydrothermal pretreatment caused negatively affect for enzymatic hydrolysis. This studies provided important factors for maximizing hydrothermal pretreatment of EFB.

The polycyclic aromatic hydrocarbons benzo[a]pyrene and phenanthrene inhibit intestinal lipase activity in rainbow trout (Oncorhynchus mykiss).

PubMed

de Gelder, Stefan; Sæle, Øystein; de Veen, Bas T H; Vos, Joëlle; Flik, Gert; Berntssen, Marc H G; Klaren, Peter H M

2017-08-01

Elevated levels of polycyclic aromatic hydrocarbons (PAHs) are detected in aquafeeds where fish oils are (partially) replaced by vegetable oils. The highly lipophilic PAHs solubilize readily in oil droplets and micelles in the intestinal lumen that can affect enzymatic lipid digestion by altering lipase activity. We therefore investigated the effect of two PAHs, benzo[a]pyrene (BaP) and phenanthrene (PHE), on bile salt-activated lipase (BAL) activity in desalted luminal extracts of the proximal intestine of rainbow trout (Oncorhynchus mykiss) using the triacylglycerides rapeseed oil and fish oil as substrates. The hydrolysis of rapeseed oil and fish oil measured at a calculated substrate concentration of 2.2mM, increased linearly up to 30min at 15°C. Substrate dependency under initial velocity conditions was described by simple Michaelis-Menten kinetics with a K m value of 1.2mM for rapeseed and fish oil. Rapeseed oil hydrolysis was inhibited by 1nM BaP and 10nM PHE. The hydrolysis of fish oil was only inhibited by 10μM BaP. The in vitro lipase activity data were corroborated by TLC/HPLC analysis of the reaction products, showing that in the presence of BaP and PHE, 46-80% less free fatty acids (FFA) were hydrolysed from rapeseed and fish oil triacylglycerides. The presence of low concentrations of BaP and PHE decreased rapeseed oil hydrolysis by BAL whereas fish oil hydrolysis was not affected. The replacement of fish oil by rapeseed oil in aquafeeds introduces PAHs that could affect lipid digestion. Copyright © 2017 Elsevier Inc. All rights reserved.

Effect of enzymatic depolymerization on physicochemical and rheological properties of guar gum.

PubMed

Mudgil, Deepak; Barak, Sheweta; Khatkar, B S

2012-09-01

Depolymerization of guar gum using enzymatic hydrolysis was performed to obtain depolymerized guar gum having functional application as soluble dietary fiber. Enzymatic hydrolysis of guar gum significantly affected the physicochemical and rheological characteristics of guar gum. The depolymerized guar gum showed a significant increase in crystallinity index from 3.86% to 13.2% and flow behavior index from 0.31 to 1.7 as compared to native guar gum. Remarkable decrease in intrinsic viscosity and consistency index was also observed from 9 to 0.28 and 4.04 to 0.07, respectively. Results revealed that enzymatic hydrolysis of guar gum resulted in a polysaccharide with low degree of polymerization, viscosity and consistency which could make it useful for incorporation in food products as dietary fiber without affecting the rheology, consistency and texture of the products. Copyright © 2012 Elsevier Ltd. All rights reserved.

Effects of grain species and cultivar, thermal processing, and enzymatic hydrolysis on gluten quantitation.

PubMed

Pahlavan, Autusa; Sharma, Girdhari M; Pereira, Marion; Williams, Kristina M

2016-10-01

Gluten from wheat, rye, and barley can trigger IgE-mediated allergy or Celiac disease in sensitive individuals. Gluten-free labeled foods are available as a safe alternative. Immunoassays such as the enzyme-linked immunosorbent assay (ELISA) are commonly used to quantify gluten in foods. However, various non-assay related factors can affect gluten quantitation. The effect of gluten-containing grain cultivars, thermal processing, and enzymatic hydrolysis on gluten quantitation by various ELISA kits was evaluated. The ELISA kits exhibited variations in gluten quantitation depending on the gluten-containing grain and their cultivars. Acceptable gluten recoveries were obtained in 200mg/kg wheat, rye, and barley-spiked corn flour thermally processed at various conditions. However, depending on the enzyme, gluten grain source, and ELISA kit used, measured gluten content was significantly reduced in corn flour spiked with 200mg/kg hydrolyzed wheat, rye, and barley flour. Thus, the gluten grain source and processing conditions should be considered for accurate gluten analysis. Published by Elsevier Ltd.

FeCl3-catalyzed ethanol pretreatment of sugarcane bagasse boosts sugar yields with low enzyme loadings and short hydrolysis time.

PubMed

Zhang, Hongdan; Zhang, Shuaishuai; Yuan, Hongyou; Lyu, Gaojin; Xie, Jun

2018-02-01

An organosolv pretreatment system consisting of 60% ethanol and 0.025 mol·L -1 FeCl 3 under various temperatures was developed in this study. During the pretreatment, the highest xylose yield was 11.4 g/100 g raw material, representing 49.8% of xylose in sugarcane bagasse. Structural features of raw material and pretreated substrates were characterized to better understand how hemicellulose removal and delignification affected subsequent enzymatic hydrolysis. The 160 °C pretreated solid presented a remarkable glucose yield of 93.8% for 72 h. Furthermore, the influence of different additives on the enzymatic hydrolysis of pretreated solid was investigated. The results indicated that the addition of Tween 80 shortened hydrolysis time to 6 h and allowed a 50% reduction of enzyme loading to achieve the same level of glucose yield. This work suggested that FeCl 3 -catalyzed organosolv pretreatment could improve the enzymatic hydrolysis significantly and reduce the hydrolysis time and enzyme dosage with the addition of Tween 80. Copyright © 2017 Elsevier Ltd. All rights reserved.

Impact of recycled effluent on the hydrolysis during anaerobic digestion of vegetable and flower waste.

PubMed

Lü, F; He, P J; Hao, L P; Shao, L M

2008-01-01

Two trials were established to investigate the effect of recycled effluent on hydrolysis during anaerobic co-digestion of vegetable and flower waste. Trial I evaluated the effect by regulating the flow rate of recycled effluent, while Trial II regulated the ratio of hydrolytic effluent to methanogenic effluent, which were recycled to hydrolysis reactor. Results showed that the recirculation of methanogenic effluent could enhance the buffer capability and operation stability of hydrolysis reactor. Higher recycled flow rate was favourable for microbial anabolism and further promoted hydrolysis. After 9 days of hydrolysis, the cumulative SCOD in the hydrolytic effluent reached 334, 407, 413, 581 mg/g at recycled flow rates of 0.1, 0.5, 1.0, 2.0 m3/(m3 x d), respectively. It was feasible to recycling a mixture of hydrolytic and methanogenic effluent to the hydrolysis reactor. This research showed that partially introducing hydrolytic effluent into the recycled liquid could enhance hydrolysis, while excessive recirculation of hydrolytic effluent will inhibit the hydrolysis. The flow ratio 1:3 of hydrolytic to methanogenic effluent was found to provide the highest hydrolysis efficiency and degradation rate of lignocelluloses-type biomass, among four ratios of 0:1, 1:3, 1:1 and 3:1. Under this regime, after 9 days of hydrolysis, the cumulative TOC and TN in the hydrolytic effluent reached 162 mg/g and 15 mg/g, the removal efficiency of TS, VS, C and cellulose in the solid phase were 60.66%, 62.88%, 58.35% and 49.12%, respectively. The flow ratio affected fermentation pathways, i.e. lower ratio favoured propionic acid fermentation and the generation of lactic acid while higher ratio promoted butyric acid fermentation. IWA Publishing 2008.

Ruminal bacteria and protozoa composition, digestibility, and amino acid profile determined by multiple hydrolysis times.

PubMed

Fessenden, S W; Hackmann, T J; Ross, D A; Foskolos, A; Van Amburgh, M E

2017-09-01

Microbial samples from 4 independent experiments in lactating dairy cattle were obtained and analyzed for nutrient composition, AA digestibility, and AA profile after multiple hydrolysis times ranging from 2 to 168 h. Similar bacterial and protozoal isolation techniques were used for all isolations. Omasal bacteria and protozoa samples were analyzed for AA digestibility using a new in vitro technique. Multiple time point hydrolysis and least squares nonlinear regression were used to determine the AA content of omasal bacteria and protozoa, and equivalency comparisons were made against single time point hydrolysis. Formalin was used in 1 experiment, which negatively affected AA digestibility and likely limited the complete release of AA during acid hydrolysis. The mean AA digestibility was 87.8 and 81.6% for non-formalin-treated bacteria and protozoa, respectively. Preservation of microbe samples in formalin likely decreased recovery of several individual AA. Results from the multiple time point hydrolysis indicated that Ile, Val, and Met hydrolyzed at a slower rate compared with other essential AA. Singe time point hydrolysis was found to be nonequivalent to multiple time point hydrolysis when considering biologically important changes in estimated microbial AA profiles. Several AA, including Met, Ile, and Val, were underpredicted using AA determination after a single 24-h hydrolysis. Models for predicting postruminal supply of AA might need to consider potential bias present in postruminal AA flow literature when AA determinations are performed after single time point hydrolysis and when using formalin as a preservative for microbial samples. Copyright © 2017 American Dairy Science Association. Published by Elsevier Inc. All rights reserved.

ESTIMATION OF CARBOXYLIC ACID ESTER HYDROLYSIS RATE CONSTANTS

EPA Science Inventory

SPARC chemical reactivity models were extended to calculate hydrolysis rate constants for carboxylic acid esters from molecular structure. The energy differences between the initial state and the transition state for a molecule of interest are factored into internal and external...

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

Jope, R.S.; Casebolt, T.L.; Johnson, G.V.

Cortical slices from rat brain were used to study carbachol-stimulated inositol phospholipid hydrolysis. Omission of calcium during incubation of slices with (/sup 3/H)inositol increased its incorporation into receptor-coupled phospholipids. Carbachol-stimulated hydrolysis of (/sup 3/H)inositol phospholipids in slices was dose-dependent, was affected by the concentrations of calcium and lithium present and resulted in the accumulation of mostly (/sup 3/H)inositol-1-phosphate. Incubation of slices with N-ethylmaleimide or a phorbol ester reduced the response to carbachol. Membranes prepared from cortical slices labeled with (/sup 3/H)inositol retained the receptor-stimulated inositol phospholipid hydrolysis reaction. The basal rate of inositol phospholipid hydrolysis was higher than in slicesmore » and addition of carbachol further stimulated the process. Addition of GTP stimulated inositol phospholipid hydrolysis, suggesting the presence of a guanine nucleotide-binding protein coupled to phospholipase C. Carbachol and GTP-stimulated inositol phospholipid hydrolysis in membranes was detectable following a 3 min assay period. In contrast to slices, increased levels of inositol bisphosphate and inositol trisphosphate were detected following incubation of membranes with carbachol. These results demonstrate that agonist-responsive receptors are present in cortical membranes, that the receptors may be coupled to phosphatidylinositol 4, 5-bisphosphate, rather than phosphatidylinositol, hydrolysis and that a guanine nucleotide-binding protein may mediate the coupling of receptor activation to inositol phospholipid hydrolysis in brain.« less

Differential reinforcement of enzymatic hydrolysis by adding chemicals and accessory proteins to high solid loading substrates with different pretreatments.

PubMed

Du, Jian; Song, Wenxia; Zhang, Xiu; Zhao, Jian; Liu, Guodong; Qu, Yinbo

2018-04-23

High dosage of enzyme is required to achieve effective lignocellulose hydrolysis, especially at high-solid loadings, which is a significant barrier to large-scale bioconversion of lignocellulose. Here, we screened four chemical additives and three accessory proteins for their effects on the enzymatic hydrolysis of various lignocellulosic materials. The effects were found to be highly dependent on the composition and solid loadings of substrates. For xylan-extracted lignin-rich corncob residue, the enhancing effect of PEG 6000 was most pronounced and negligibly affected by solid content, which reduced more than half of enzyme demand at 20% dry matter (DM). Lytic polysaccharide monooxygenase enhanced the hydrolysis of ammonium sulfite wheat straw pulp, and its addition reduced about half of protein demand at the solid loading of 20% DM. Supplementation of the additives in the hydrolysis of pure cellulose and complex lignocellulosic materials revealed that their effects are tightly linked to pretreatment strategies.

Effects of Kraft lignin on hydrolysis/dehydration of sugars, cellulosic and lignocellulosic biomass under hot compressed water.

PubMed

Daorattanachai, Pornlada; Viriya-empikul, Nawin; Laosiripojana, Navadol; Faungnawakij, Kajornsak

2013-09-01

The effect of Kraft lignin presenting on the hydrolysis and dehydration of C5 and C6 sugars, cellulose, hemicelluloses and biomass under hot compressed water (HCW) in the presence of H3PO4 catalyst was intensively studied. The lignin strongly inhibited the acid hydrolysis of cellulose and hemicellulose to glucose and xylose, respectively. Interestingly, the admixed lignin markedly promoted the isomerization of glucose to fructose, and dehydration of fructose (except at the low catalyst loading), resulting in high 5-hydroxymethylfurfural yields. Nonetheless, lignin inhibited the hydrolysis of xylan to xylose and dehydration of xylose to furfural. Moreover, the acidity of the system significantly affects the hydrolysis/dehydration of biomass. It was revealed that the presence of lignin strongly interfered the yields of sugars and furans produced from raw corncob, while the delignified corncob provided significant improvement of product yields, confirming the observed role of lignin in the biomass conversion system via sugar platforms. Copyright © 2013 Elsevier Ltd. All rights reserved.

Optimization of cellulose nanocrystal length and surface charge density through phosphoric acid hydrolysis

NASA Astrophysics Data System (ADS)

Vanderfleet, Oriana M.; Osorio, Daniel A.; Cranston, Emily D.

2017-12-01

Cellulose nanocrystals (CNCs) are emerging nanomaterials with a large range of potential applications. CNCs are typically produced through acid hydrolysis with sulfuric acid; however, phosphoric acid has the advantage of generating CNCs with higher thermal stability. This paper presents a design of experiments approach to optimize the hydrolysis of CNCs from cotton with phosphoric acid. Hydrolysis time, temperature and acid concentration were varied across nine experiments and a linear least-squares regression analysis was applied to understand the effects of these parameters on CNC properties. In all but one case, rod-shaped nanoparticles with a high degree of crystallinity and thermal stability were produced. A statistical model was generated to predict CNC length, and trends in phosphate content and zeta potential were elucidated. The CNC length could be tuned over a relatively large range (238-475 nm) and the polydispersity could be narrowed most effectively by increasing the hydrolysis temperature and acid concentration. The CNC phosphate content was most affected by hydrolysis temperature and time; however, the charge density and colloidal stability were considered low compared with sulfuric acid hydrolysed CNCs. This study provides insight into weak acid hydrolysis and proposes `design rules' for CNCs with improved size uniformity and charge density. This article is part of a discussion meeting issue `New horizons for cellulose nanotechnology'.

Phase composition of lipoprotein SM/cholesterol/PtdCho affects FA specificity of sPLA2s.

PubMed

Kuksis, Arnis; Pruzanski, Waldemar

2008-10-01

We have previously reported preferential release of polyunsaturated FAs during hydrolysis of lipoprotein phosphatidylcholine (PtdCho) by group X secretory phospholipase A2 (sPLA2) and preferential release of oligounsaturated FAs during hydrolysis of lipoprotein PtdCho by group V sPLA2, but the mechanism of this selectivity has remained unknown. We now show that the rate and specificity of hydrolysis are affected by relative increases in endogenous SM and free cholesterol (FC) during the lipase digestion. The highest preference for arachidonate release from LDL and HDL by group X sPLA2 was observed for residual SM/PtdCho molar ratio of 1.2 and 0.4, compared with the respective starting ratios of 0.4 and 0.2, as measured by liquid chromatography/electrospray ionization-mass spectrometry. Group V sPLA2 showed preferential release of linoleate from LDL and HDL at SM/PtdCho ratio 1.5 and 0.6, respectively. We have attributed the change in FA specificity to segregation of molecular species of PtdCho and of sPLA2s between disordered and ordered SM/FC/PtdCho lipid phases. The increases in SM and FC during digestion with group IIA sPLA2 were more limited, and a preferential hydrolysis of any FAs was not observed. The significance of SM and FC SM and FC accumulation during sPLA2 hydrolysis of lipoprotein PtdCho has been previously overlooked.

Optimization of parameters for enhanced oil recovery from enzyme treated wild apricot kernels.

PubMed

Rajaram, Mahatre R; Kumbhar, Baburao K; Singh, Anupama; Lohani, Umesh Chandra; Shahi, Navin C

2012-08-01

Present investigation was undertaken with the overall objective of optimizing the enzymatic parameters i.e. moisture content during hydrolysis, enzyme concentration, enzyme ratio and incubation period on wild apricot kernel processing for better oil extractability and increased oil recovery. Response surface methodology was adopted in the experimental design. A central composite rotatable design of four variables at five levels was chosen. The parameters and their range for the experiments were moisture content during hydrolysis (20-32%, w.b.), enzyme concentration (12-16% v/w of sample), combination of pectolytic and cellulolytic enzyme i.e. enzyme ratio (30:70-70:30) and incubation period (12-16 h). Aspergillus foetidus and Trichoderma viride was used for production of crude enzyme i.e. pectolytic and cellulolytic enzyme respectively. A complete second order model for increased oil recovery as the function of enzymatic parameters fitted the data well. The best fit model for oil recovery was also developed. The effect of various parameters on increased oil recovery was determined at linear, quadric and interaction level. The increased oil recovery ranged from 0.14 to 2.53%. The corresponding conditions for maximum oil recovery were 23% (w.b.), 15 v/w of the sample, 60:40 (pectolytic:cellulolytic), 13 h. Results of the study indicated that incubation period during enzymatic hydrolysis is the most important factor affecting oil yield followed by enzyme ratio, moisture content and enzyme concentration in the decreasing order. Enzyme ratio, incubation period and moisture content had insignificant effect on oil recovery. Second order model for increased oil recovery as a function of enzymatic hydrolysis parameters predicted the data adequately.

Optimization of enzymatic hydrolysis of guar gum using response surface methodology.

PubMed

Mudgil, Deepak; Barak, Sheweta; Khatkar, B S

2014-08-01

Guar gum is a polysaccharide obtained from guar seed endosperm portion. Enzymatically hydrolyzed guar gum is low in viscosity and has several health benefits as dietary fiber. In this study, response surface methodology was used to determine the optimum conditions for hydrolysis that give minimum viscosity of guar gum. Central composite was employed to investigate the effects of pH (3-7), temperature (20-60 °C), reaction time (1-5 h) and cellulase concentration (0.25-1.25 mg/g) on viscosity during enzymatic hydrolysis of guar (Cyamopsis tetragonolobus) gum. A second order polynomial model was developed for viscosity using regression analysis. Results revealed statistical significance of model as evidenced from high value of coefficient of determination (R(2) = 0.9472) and P < 0.05. Viscosity was primarily affected by cellulase concentration, pH and hydrolysis time. Maximum viscosity reduction was obtained when pH, temperature, hydrolysis time and cellulase concentration were 6, 50 °C, 4 h and 1.00 mg/g, respectively. The study is important in optimizing the enzymatic process for hydrolysis of guar gum as potential source of soluble dietary fiber for human health benefits.

Improvement of Functional Properties of Wheat Gluten Using Acid Protease from Aspergillus usamii

PubMed Central

Deng, Lingli; Wang, Zhaoxia; Yang, Sheng; Song, Junmei; Que, Fei; Zhang, Hui; Feng, Fengqin

2016-01-01

Hydrolysis parameters (temperature, E/S ratio, pH, and time) for acid protease (from Aspergillus usamii) hydrolysis of wheat gluten were optimized by response surface methodology (RSM) using emulsifying activity index (EAI) as the response factor. A temperature of 48.9°C, E/S ratio of 1.60%, pH 3.0, hydrolysis time of 2.5 h was found to be the optimum condition to obtain wheat gluten hydrolysate with higher EAI. The solubility of wheat gluten was greatly improved by hydrolysis and became independent of pH over the studied range. Enzymatic hydrolysis resulted in dramatically increase in EAI, water and oil holding capacity. Molecular weight distribution results showed that most of the peptides above 10 kDa have been hydrolyzed into smaller peptides. The results of FTIR spectra and disulfide bond (SS) and sulfhydryl (SH) content suggested that a more extensional conformation was formed after hydrolysis, which could account for the improved functional properties. PMID:27467884

Improvement of Functional Properties of Wheat Gluten Using Acid Protease from Aspergillus usamii.

PubMed

Deng, Lingli; Wang, Zhaoxia; Yang, Sheng; Song, Junmei; Que, Fei; Zhang, Hui; Feng, Fengqin

2016-01-01

Hydrolysis parameters (temperature, E/S ratio, pH, and time) for acid protease (from Aspergillus usamii) hydrolysis of wheat gluten were optimized by response surface methodology (RSM) using emulsifying activity index (EAI) as the response factor. A temperature of 48.9°C, E/S ratio of 1.60%, pH 3.0, hydrolysis time of 2.5 h was found to be the optimum condition to obtain wheat gluten hydrolysate with higher EAI. The solubility of wheat gluten was greatly improved by hydrolysis and became independent of pH over the studied range. Enzymatic hydrolysis resulted in dramatically increase in EAI, water and oil holding capacity. Molecular weight distribution results showed that most of the peptides above 10 kDa have been hydrolyzed into smaller peptides. The results of FTIR spectra and disulfide bond (SS) and sulfhydryl (SH) content suggested that a more extensional conformation was formed after hydrolysis, which could account for the improved functional properties.

Investigating Mass Transport Limitations on Xylan Hydrolysis During Dilute Acid Pretreatment of Poplar

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

Mittal, Ashutosh; Pilath, Heid M.; Parent, Yves

2014-04-28

Mass transport limitations could be an impediment to achieving high sugar yields during biomass pretreatment and thus be a critical factor in the economics of biofuels production. The objective of this work was to study the mass transfer restrictions imposed by the structure of biomass on the hydrolysis of xylan during dilute acid pretreatment of biomass. Mass transfer effects were studied by pretreating poplar wood at particle sizes ranging from 10 micrometers to 10 mm. This work showed a significant reduction in the rate of xylan hydrolysis in poplar when compared to the intrinsic rate of hydrolysis for isolated xylanmore » that is possible in the absence of mass transfer. In poplar samples we observed no significant difference in the rates of xylan hydrolysis over more than two orders of magnitude in particle size. It appears that no additional mass transport restrictions are introduced by increasing particle size from 10 micrometers to 10 mm. This work suggests that the rates of xylan hydrolysis in biomass particles are limited primarily by the diffusion of hydrolysis products out of plant cell walls. A mathematical description is presented to describe the kinetics of xylan hydrolysis that includes transport of the hydrolysis products through biomass into the bulk solution. The modeling results show that the effective diffusion coefficient of the hydrolysis products in the cell wall is several orders of magnitude smaller than typical values in other applications signifying the role of plant cell walls in offering resistance to diffusion of the hydrolysis products.« less

Fundamental factors affecting biomass enzymatic reactivity.

PubMed

Chang, V S; Holtzapple, M T

2000-01-01

Poplar wood was treated with peracetic acid, KOH, and ball milling to produce 147 model lignocelluloses with a broad spectrum of lignin contents, acetyl contents, and crystallinity indices (CrIs), respectively. An empirical model was identified that describes the roles of these three properties in enzymatic hydrolysis. Lignin content and CrI have the greatest impact on biomass digestibility, whereas acetyl content has a minor impact. The digestibility of several lime-treated biomass samples agreed with the empirical model. Lime treatment removes all acetyl groups and a moderate amount of lignin and increases CrI slightly; lignin removal is the dominant benefit from lime treatment.

Effect of particle size on enzymatic hydrolysis of pretreated Miscanthus

USDA-ARS?s Scientific Manuscript database

Particle size reduction is a crucial factor in transportation logistics as well as cellulosic conversion. The effect of particle size on enzymatic hydrolysis of pretreated Miscanthus x giganteus was determined. Miscanthus was ground using a hammer mill equipped with screens having 0.08, 2.0 or 6.0...

Characterization of partially hydrolyzed OCP crystals deposited in a gelatin matrix as a scaffold for bone tissue engineering

NASA Astrophysics Data System (ADS)

Ezoe, Yushi; Anada, Takahisa; Yamazaki, Hajime; Handa, Takuto; Kobayashi, Kazuhito; Takahashi, Tetsu; Suzuki, Osamu

2015-03-01

The present study was designed to investigate how hydrolysis of octacalcium phosphate (OCP) into hydroxyapatite is affected by the presence of gelatin (Gel) molecules and how osteoblastic cells respond to the resultant OCP hydrolyzate/Gel composites as the hydrolysis advances. OCP was prepared from a solution containing calcium and phosphate ions and Gel molecules, having a composition to produce a 40 wt% OCP as a final co-precipitate as the OCP/Gel. The precipitate was further incubated up to 40 h to advance the hydrolysis of OCP. These precipitates were processed to mold OCP/Gel sponges through lyophilization and dehydrothermal treatment. Chemical analysis, X-ray diffraction, Fourier transform infrared spectroscopy, transmission electron microscopy, and selected area electron diffraction revealed that the hydrolysis of OCP/Gel composite in hot water advanced in a time-dependent manner and faster than hydrolysis of OCP alone. The effect of Gel on the OCP hydrolysis was further examined in the presence of distinct concentrations of Gel molecules in hot water, showing that the Gel enhanced the hydrolysis as the concentration increased. Proliferation and differentiation of mouse bone marrow stromal ST-2 cells on the hydrolyzed OCP/Gel composites were compatible with Gel sponge alone after 21 days of culture, suggesting that these composites could be a candidate as a scaffold in bone tissue engineering.

The mechanism of hydrothermal hydrolysis for glycyrrhizic acid into glycyrrhetinic acid and glycyrrhetinic acid 3-O-mono-β-D-glucuronide in subcritical water.

PubMed

Fan, Rui; Li, Nan; Xu, Honggao; Xiang, Jun; Wang, Lei; Gao, Yanxiang

2016-01-01

To improve the bioactivity and sweetness properties of glycyrrhizic acid (GL), the hydrothermal hydrolysis of GL into glycyrrhetinic acid (GA) and glycyrrhetinic acid 3-O-mono-β-D-glucuronide (GAMG) in subcritical water was investigated. The effects of temperature, time and their interaction on the conversion ratios were analyzed and the reactions were elaborated with kinetics and thermodynamics. The results showed that GL hydrothermal hydrolysis was significantly (P < 0.05) affected by reaction time and temperature, as well as their interaction, and could be fitted into first-order kinetics. The thermodynamic analysis indicated that the hydrolysis of GL was endergonic and non-spontaneous. The hydrolytic pathways were composed of complex consecutive and parallel reactions. It was concluded that subcritical water may be a potential medium for producing GAMG and GA. Copyright © 2015 Elsevier Ltd. All rights reserved.

Biological strategies for enhanced hydrolysis of lignocellulosic biomass during anaerobic digestion: Current status and future perspectives.

PubMed

Shrestha, Shilva; Fonoll, Xavier; Khanal, Samir Kumar; Raskin, Lutgarde

2017-12-01

Lignocellulosic biomass is the most abundant renewable bioresource on earth. In lignocellulosic biomass, the cellulose and hemicellulose are bound with lignin and other molecules to form a complex structure not easily accessible to microbial degradation. Anaerobic digestion (AD) of lignocellulosic biomass with a focus on improving hydrolysis, the rate limiting step in AD of lignocellulosic feedstocks, has received considerable attention. This review highlights challenges with AD of lignocellulosic biomass, factors contributing to its recalcitrance, and natural microbial ecosystems, such as the gastrointestinal tracts of herbivorous animals, capable of performing hydrolysis efficiently. Biological strategies that have been evaluated to enhance hydrolysis of lignocellulosic biomass include biological pretreatment, co-digestion, and inoculum selection. Strategies to further improve these approaches along with future research directions are outlined with a focus on linking studies of microbial communities involved in hydrolysis of lignocellulosics to process engineering. Copyright © 2017 Elsevier Ltd. All rights reserved.

Enhanced mannan-derived fermentable sugars of palm kernel cake by mannanase-catalyzed hydrolysis for production of biobutanol.

PubMed

Shukor, Hafiza; Abdeshahian, Peyman; Al-Shorgani, Najeeb Kaid Nasser; Hamid, Aidil Abdul; Rahman, Norliza A; Kalil, Mohd Sahaid

2016-10-01

Catalytic depolymerization of mannan composition of palm kernel cake (PKC) by mannanase was optimized to enhance the release of mannan-derived monomeric sugars for further application in acetone-butanol-ethanol (ABE) fermentation. Efficiency of enzymatic hydrolysis of PKC was studied by evaluating effects of PKC concentration, mannanase loading, hydrolysis pH value, reaction temperature and hydrolysis time on production of fermentable sugars using one-way analysis of variance (ANOVA). The ANOVA results revealed that all factors studied had highly significant effects on total sugar liberated (P<0.01). The optimum conditions for PKC hydrolysis were 20% (w/v) PKC concentration, 5% (w/w) mannanase loading, hydrolysis pH 4.5, 45°C temperature and 72h hydrolysis time. Enzymatic experiments in optimum conditions revealed total fermentable sugars of 71.54±2.54g/L were produced including 67.47±2.51g/L mannose and 2.94±0.03g/L glucose. ABE fermentation of sugar hydrolysate by Clostridium saccharoperbutylacetonicum N1-4 resulted in 3.27±1.003g/L biobutanol. Copyright © 2016 Elsevier Ltd. All rights reserved.

Disintegration impact on sludge digestion process.

PubMed

Dauknys, Regimantas; Rimeika, Mindaugas; Jankeliūnaitė, Eglė; Mažeikienė, Aušra

2016-11-01

The anaerobic sludge digestion is a widely used method for sludge stabilization in wastewater treatment plant. This process can be improved by applying the sludge disintegration methods. As the sludge disintegration is not investigated enough, an analysis of how the application of thermal hydrolysis affects the sludge digestion process based on full-scale data was conducted. The results showed that the maximum volatile suspended solids (VSS) destruction reached the value of 65% independently on the application of thermal hydrolysis. The average VSS destruction increased by 14% when thermal hydrolysis was applied. In order to have the maximum VSS reduction and biogas production, it is recommended to keep the maximum defined VSS loading of 5.7 kg VSS/m(3)/d when the thermal hydrolysis is applied and to keep the VSS loading between 2.1-2.4 kg VSS/m(3)/d when the disintegration of sludge is not applied. The application of thermal hydrolysis leads to an approximately 2.5 times higher VSS loading maintenance comparing VSS loading without the disintegration; therefore, digesters with 1.8 times smaller volume is required.

Hypo-and hyperthyroidism affect the ATP, ADP and AMP hydrolysis in rat hippocampal and cortical slices.

PubMed

Bruno, Alessandra Nejar; Diniz, Gabriela Placoná; Ricachenevsky, Felipe Klein; Pochmann, Daniela; Bonan, Carla Denise; Barreto-Chaves, Maria Luiza M; Sarkis, João José Freitas

2005-05-01

The presence of severe neurological symptoms in thyroid diseases has highlighted the importance of thyroid hormones in the normal functioning of the mature brain. Since, ATP is an important excitatory neurotransmitter and adenosine acts as a neuromodulatory structure inhibiting neurotransmitters release in the central nervous system (CNS), the ectonucleotidase cascade that hydrolyzes ATP to adenosine, is also involved in the control of brain functions. Thus, we investigated the influence of hyper-and hypothyroidism on the ATP, ADP and AMP hydrolysis in hippocampal and cortical slices from adult rats. Hyperthyroidism was induced by daily injections of l-thyroxine (T4) 25 microg/100 g body weight, for 14 days. Hypothyroidism was induced by thyroidectomy and methimazole (0.05%) added to their drinking water for 14 days. Hypothyroid rats were hormonally replaced by daily injections of T4 (5 microg/100 g body weight, i.p.) for 5 days. Hyperthyroidism significantly inhibited the ATP, ADP and AMP hydrolysis in hippocampal slices. In brain cortical slices, hyperthyroidism inhibited the AMP hydrolysis. In contrast, hypothyroidism increased the ATP, ADP and AMP hydrolysis in both hippocampal and cortical slices and these effects were reverted by T4 replacement. Furthermore, hypothyroidism increased the expression of NTPDase1 and 5'-nucleotidase, whereas hyperthyroidism decreased the expression of 5'-nucleotidase in hippocampus of adult rats. These findings demonstrate that thyroid disorders may influence the enzymes involved in the complete degradation of ATP to adenosine and possibly affects the responses mediated by adenine nucleotides in the CNS of adult rats.

Pectinase from Trichoderma reesei QM 9414

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

Haltmeier, T.; Leisola, M.; Ulmer, D.

1983-06-01

The present study was undertaken to determine whether T. reesei produces pectinolytic enzymes and, if so, to determine their action pattern and specificity. The aim was also to find out the practical importance of pectinolytic activity in the hydrolysis of cellulosic materials. It was found that the presence of pectinase does not appear to facilitate the hydrolysis of cellulose in plant material through any cell-separating mechanism. It is concluded that cellulase and xylanase activities are the important activities in the practical hydrolysis of cellulosic materials and that lignin is the real limiting factor. (Refs. 12).

The effect of pH, temperature and plaque thickness on the hydrolysis of monofluorophosphate in experimental dental plaque.

PubMed

Pearce, E I F; Dibdin, G H

2003-01-01

Monofluorophosphate (MFP), an anti-caries agent commonly used in toothpaste, is known to be degraded to fluoride and orthophosphate by bacterial phosphatases in dental plaque. We have examined the effect of pH, temperature, plaque thickness and some ions on this process. Both natural plaque and artificial microcosm plaque incubated with purified MFP at pH 4-10 showed an optimum pH of approximately 8 for hydrolysis. Diffusion and concomitant hydrolysis were examined in an apparatus in which artificial plaque was held between rigid membranes separating two chambers. When MFP diffused through a plaque of 0.51-mm thickness over 4 h it was almost completely hydrolysed at pH 8, but hydrolysis on diffusion decreased as the pH deviated from 8. MFP in toothpaste extract showed a similar pH susceptibility to hydrolysis, according to the inherent pH of the toothpaste. Hydrolysis of MFP in the toothpaste was reduced by no more than 10% when compared with a matched-pH control, suggesting that other toothpaste ingredients had no major influence on hydrolysis. Transport was slower and hydrolysis at pH 6 more complete the thicker the plaque, but hydrolysis was not significantly slower at 23 degrees C than at 37 degrees C. The addition of various potential activating or inhibiting ions at 0.1 and 1.0 mmol/l had small and non-significant effects on hydrolysis. The results suggest that MFP toothpaste should be formulated and used to maximise enzymic hydrolysis of this complex anion, and that plaque pH control is probably the most important factor. Copyright 2003 S. Karger AG, Basel

Analysis of a Modern Hybrid and an Ancient Sugarcane Implicates a Complex Interplay of Factors in Affecting Recalcitrance to Cellulosic Ethanol Production

PubMed Central

Ricci-Silva, Maria Esther; Rhys Williams, Thomas Christopher; Alves Peçanha, Diego; Contin Ventrella, Marília; Rencoret, Jorge; Ralph, John; Pereira Barbosa, Márcio Henrique; Loureiro, Marcelo

2015-01-01

Abundant evidence exists to support a role for lignin as an important element in biomass recalcitrance. However, several independent studies have also shown that factors apart from lignin are also relevant and overall, the relative importance of different recalcitrance traits remains in dispute. In this study we used two genetically distant sugarcane genotypes, and performed a correlational study with the variation in anatomical parameters, cell wall composition, and recalcitrance factors between these genotypes. In addition we also tracked alterations in these characteristics in internodes at different stages of development. Significant differences in the development of the culm between the genotypes were associated with clear differential distributions of lignin content and composition that were not correlated with saccharification and fermentation yield. Given the strong influence of the environment on lignin content and composition, we hypothesized that sampling within a single plant could allow us to more easily interpret recalcitrance and changes in lignin biosynthesis than analysing variations between different genotypes with extensive changes in plant morphology and culm anatomy. The syringyl/guaiacyl (S/G) ratio was higher in the oldest internode of the modern genotype, but S/G ratio was not correlated with enzymatic hydrolysis yield nor fermentation efficiency. Curiously we observed a strong positive correlation between ferulate ester level and cellulose conversion efficiency. Together, these data support the hypothesis that biomass enzymatic hydrolysis recalcitrance is governed by a quantitative heritage rather than a single trait. PMID:26252208

Antioxidant, antihypertensive and antimicrobial properties of ovine milk caseinate hydrolyzed with a microbial protease.

PubMed

Corrêa, Ana Paula F; Daroit, Daniel J; Coelho, Julise; Meira, Stela M M; Lopes, Fernanda C; Segalin, Jéferson; Risso, Patrícia H; Brandelli, Adriano

2011-09-01

Bioactive peptides might be released from precursor proteins through enzymatic hydrolysis. These molecules could be potentially employed in health and food products. In this investigation, ovine milk caseinate hydrolysates obtained with a novel microbial protease derived from Bacillus sp. P7 were evaluated for antioxidant, antimicrobial, and angiotensin I-converting enzyme (ACE)-inhibitory activities. Antioxidant activity measured by the 2,2'-azino-bis-(3-ethylbenzothiazoline)-6-sulfonic acid method increased with hydrolysis time up to 2 h, remaining stable for up to 4 h. Hydrolysates showed low 2,2-diphenyl-1-picrylhydrazyl radical-scavenging abilities, with higher activity (31%) reached after 1 h of hydrolysis. Fe(2+) -chelating ability was maximum for 0.5 h hydrolysates (83.3%), decreasing thereafter; and the higher reducing power was observed after 1 h of hydrolysis. ACE-inhibitory activity was observed to increase up to 2 h of hydrolysis (94% of inhibition), declining afterwards. 3 h hydrolysates were shown to inhibit the growth of Bacillus cereus, Corynebacterium fimi, Aspergillus fumigatus, and Penicillium expansum. Ovine caseinate hydrolyzed with Bacillus sp. P7 protease presented antioxidant, antihypertensive, and antimicrobial activities. Hydrolysis time was observed to affect the evaluated bioactivities. Such hydrolysates might have potential applications in the food industry. Copyright © 2011 Society of Chemical Industry.

Evaluation of hyper thermal acid hydrolysis of Kappaphycus alvarezii for enhanced bioethanol production.

PubMed

Ra, Chae Hun; Nguyen, Trung Hau; Jeong, Gwi-Taek; Kim, Sung-Koo

2016-06-01

Hyper thermal (HT) acid hydrolysis of Kappaphycus alvarezii, a red seaweed, was optimized to 12% (w/v) seaweed slurry content, 180mM H2SO4 at 140°C for 5min. The maximum monosaccharide concentration of 38.3g/L and 66.7% conversion from total fermentable monosaccharides of 57.6g/L with 120gdw/L K. alvarezii slurry were obtained from HT acid hydrolysis and enzymatic saccharification. HT acid hydrolysis at a severity factor of 0.78 efficiently converted the carbohydrates of seaweed to monosaccharides and produced a low concentration of inhibitory compounds. The levels of ethanol production by separate hydrolysis and fermentation with non-adapted and adapted Kluyveromyces marxianus to high concentration of galactose were 6.1g/L with ethanol yield (YEtOH) of 0.19 at 84h and 16.0g/L with YEtOH of 0.42 at 72h, respectively. Development of the HT acid hydrolysis process and adapted yeast could enhance the overall ethanol fermentation yields of K. alvarezii seaweed. Copyright © 2016 Elsevier Ltd. All rights reserved.

Effects of protonation on the hydrolysis of triphosphate in vacuum and the implications for catalysis by nucleotide hydrolyzing enzymes.

PubMed

Kiani, Farooq Ahmad; Fischer, Stefan

2016-06-29

Nucleoside triphosphate (NTP) hydrolysis is a key reaction in biology. It involves breaking two very stable bonds (one P-O bond and one O-H bond of water), in either a concurrent or a sequential way. Here, we systematically examine how protonation of the triphosphate affects the mechanism of hydrolysis. The hydrolysis reaction of methyl triphosphate in vacuum is computed with protons in various numbers and position on the three phosphate groups. Protonation is seen to have a strong catalytic effect, with the reaction mechanism depending highly on the protonation pattern. This dependence is apparently complicated, but is shown to obey a well-defined set of rules: Protonation of the α- and β-phosphate groups favors a sequential hydrolysis mechanism, whereas γ-protonation favors a concurrent mechanism, the two effects competing with each other in cases of simultaneous protonation. The rate-limiting step is always the breakup of the water molecule while it attacks the γ-phosphorus, and its barrier is lowered by γ-protonation. This step has significantly lower barriers in the sequential reactions, because the dissociated γ-metaphosphate intermediate (P γ O 3 - ) is a much better target for water attack than the un-dissociated γ-phosphate (-P γ O 4 2- ). The simple chemical logic behind these rules helps to better understand the catalytic strategy used by NTPase enzymes, as illustrated here for the catalytic pocket of myosin. A set of rules was determined that describes how protonating the phosphate groups affects the hydrolysis mechanism of methyl triphosphate: Protonation of the α- and/or β- phosphate groups promotes a sequential mechanism in which P-O bond breaking precedes the breakup of the attacking water, whereas protonation of the γ-phosphate promotes a concurrent mechanism and lowers the rate-limiting barrier of water breakup. The role played by individual protein residues in the catalytic pocket of triphosphate hydrolysing enzymes can be assigned accordingly.

DFT investigations of phosphotriesters hydrolysis in aqueous solution: a model for DNA single strand scission induced by N-nitrosoureas.

PubMed

Liu, Tingting; Zhao, Lijiao; Zhong, Rugang

2013-02-01

DNA phosphotriester adducts are common alkylation products of DNA phosphodiester moiety induced by N-nitrosoureas. The 2-hydroxyethyl phosphotriester was reported to hydrolyze more rapidly than other alkyl phosphotriesters both in neutral and in alkaline conditions, which can cause DNA single strand scission. In this work, DFT calculations have been employed to map out the four lowest activation free-energy profiles for neutral and alkaline hydrolysis of triethyl phosphate (TEP) and diethyl 2-hydroxyethyl phosphate (DEHEP). All the hydrolysis pathways were illuminated to be stepwise involving an acyclic or cyclic phosphorane intermediate for TEP or DEHEP, respectively. The rate-limiting step for all the hydrolysis reactions was found to be the formation of phosphorane intermediate, with the exception of DEHEP hydrolysis in alkaline conditions that the decomposition process turned out to be the rate-limiting step, owing to the extraordinary low formation barrier of cyclic phosphorane intermediate catalyzed by hydroxide. The rate-limiting barriers obtained for the four reactions are all consistent with the available experimental information concerning the corresponding hydrolysis reactions of phosphotriesters. Our calculations performed on the phosphate triesters hydrolysis predict that the lower formation barriers of cyclic phosphorane intermediates compared to its acyclic counter-part should be the dominant factor governing the hydrolysis rate enhancement of DEHEP relative to TEP both in neutral and in alkaline conditions.

The effect of medium viscosity on kinetics of ATP hydrolysis by the chloroplast coupling factor CF1.

PubMed

Malyan, Alexander N

2016-05-01

The coupling factor CF1 is a catalytic part of chloroplast ATP synthase which is exposed to stroma whose viscosity is many-fold higher than that of reaction mixtures commonly used to measure kinetics of CF1-catalyzed ATP hydrolysis. This study is focused on the effect of medium viscosity modulated by sucrose or bovine serum albumin (BSA) on kinetics of Ca(2+)- and Mg(2+)-dependent ATP hydrolysis by CF1. These agents were shown to reduce the maximal rate of Ca(2+)-dependent ATPase without changing the apparent Michaelis constant (К m), thus supporting the hypothesis on viscosity dependence of CF1 activity. For the sulfite- and ethanol-stimulated Mg(2+)-dependent reaction, the presence of sucrose increased К m without changing the maximal rate that is many-fold as high as that of Ca(2+)-dependent hydrolysis. The hydrolysis reaction was shown to be stimulated by low concentrations of BSA and inhibited by its higher concentrations, with the increasing maximal reaction rate estimated by extrapolation. Sucrose- or BSA-induced inhibition of the Mg(2+)-dependent ATPase reaction is believed to result from diffusion-caused deceleration, while its BSA-induced stimulation is probably caused by optimization of the enzyme structure. Molecular mechanisms of the inhibitory effect of viscosity are discussed. Taking into account high protein concentrations in the chloroplast stroma, it was suggested that kinetic parameters of ATP hydrolysis, and probably those of ATP synthesis in vivo as well, must be quite different from measurements taken at a viscosity level close to that of water.

Assessing mechanical deconstruction of softwood cell wall for cellulosic biofuels production

NASA Astrophysics Data System (ADS)

Jiang, Jinxue

Mechanical deconstruction offers a promising strategy to overcome biomass recalcitrance for facilitating enzymatic hydrolysis of pretreated substrates with zero chemicals input and presence of inhibitors. The goal of this dissertation research is to gain a more fundamental understanding on the impact of mechanical pretreatment on generating digestible micronized-wood and how the physicochemical characteristics influence the subsequent enzymatic hydrolysis of micronized wood. The initial moisture content of feedstock was found to be the key factor affecting the development of physical features and enzymatic hydrolysis of micronized wood. Lower moisture content resulted in much rounder particles with lower crystallinity, while higher moisture content resulted in the milled particles with larger aspect ratio and crystallinity. The enzymatic hydrolysis of micronized wood was improved as collectively increasing surface area (i.e., reducing particle size and aspect ratio) and decreasing crystallinity during mechanical milling pretreatment. Energy efficiency analysis demonstrated that low-moisture content feedstock with multi-step milling process would contribute to cost-effectiveness of mechanical pretreatment for achieving more than 70% of total sugars conversion. In the early stage of mechanical pretreatment, the types of cell fractures were distinguished by the initial moisture contents of wood, leading to interwall fracture at the middle lamella region for low moisture content samples and intrawall fracture at the inner cell wall for high moisture content samples. The changes in cell wall fractures also resulted in difference in the distribution of surface chemical composition and energy required for milling process. In an effort to exploit the underlying mechanism associated with the reduced recalcitrance in micronized wood, we reported the increased enzymatic sugar yield and correspondingly structural and accessible properties of micronized feedstock. Electronic microscopy analysis detailed the structural alternation of cell wall during mechanical process, including cell fracture and delamination, ultrastructure disintegration, and cell wall fragments amorphization, as coincident with the particle size reduction. It was confirmed with Simons' staining that longer milling time resulted in increased substrate accessibility and porosity. The changes in cellulose molecular structure with respect to degree of polymerization (DP) and crystallinity index (CrI) also benefited to decreasing recalcitrance and facilitating enzymatic hydrolysis of micronized wood.

The influence of cosolvent and heat on the solubility and reactivity of organophosphorous pesticide DNAPL alkaline hydrolysis.

PubMed

Muff, Jens; MacKinnon, Leah; Durant, Neal D; Bennedsen, Lars Frausing; Rügge, Kirsten; Bondgaard, Morten; Pennell, Kurt

2016-11-01

The presented research concerned the compatibility of cosolvents with in situ alkaline hydrolysis (ISAH) for treatment of organophosphorous (OPP) pesticide contaminated sites. In addition, the influence of moderate temperature heat increments was studied as a possible enhancement method. A complex dense non-aqueous phase liquid (DNAPL) of primarily parathion (~50 %) and methyl parathion (~15 %) obtained from the Danish Groyne 42 site was used as a contaminant source, and ethanol and propan-2-ol (0, 25, and 50 v/v%) was used as cosolvents in tap water and 0.34 M NaOH. Both cosolvents showed OPP solubility enhancement at 50 v/v% cosolvent content, with slightly higher OPP concentrations reached with propan-2-ol. Data on hydrolysis products did not show a clear trend with respect to alkaline hydrolysis reactivity in the presence of cosolvents. Results indicated that the hydrolysis rate of methyl-parathion (MP3) decreased with addition of cosolvent, whereas the hydrolysis rate of ethyl-parathion (EP3) remained constant, and overall indications were that the hydrolysis reactions were limited by the rate of hydrolysis rather than NAPL dissolution. In addition to cosolvents, the influence of low-temperature heating on ISAH was studied. Increasing reaction temperature from 10 to 30 °C provided an average rate of hydrolysis enhancement by a factor of 1.4-4.8 dependent on the base of calculation. When combining 50 v/v% cosolvent addition and heating to 30 °C, EP3 solubility was significantly enhanced and results for O,O-diethyl-thiophosphoric acid (EP2 acid) showed a significant enhancement of hydrolysis as well. However, this could not be supported by para-nitrophenol (PNP) data indicating the instability of this product in the presence of cosolvent.

Application of the Initial Rate Method in Anaerobic Digestion of Kitchen Waste

PubMed Central

Lang, Xianming; Liu, Yiwei; Li, Rundong; Yu, Meiling; Shao, Lijie; Wang, Xiaoming

2017-01-01

This article proposes a methane production approach through sequenced anaerobic digestion of kitchen waste, determines the hydrolysis constants and reaction orders at both low total solid (TS) concentrations and high TS concentrations using the initial rate method, and examines the population growth model and first-order hydrolysis model. The findings indicate that the first-order hydrolysis model better reflects the kinetic process of gas production. During the experiment, all the influential factors of anaerobic fermentation retained their optimal values. The hydrolysis constants and reaction orders at low TS concentrations are then employed to demonstrate that the first-order gas production model can describe the kinetics of the gas production process. At low TS concentrations, the hydrolysis constants and reaction orders demonstrated opposite trends, with both stabilizing after 24 days at 0.99 and 1.1252, respectively. At high TS concentrations, the hydrolysis constants and the reaction orders stabilized at 0.98 (after 18 days) and 0.3507 (after 14 days), respectively. Given sufficient reaction time, the hydrolysis involved in anaerobic fermentation of kitchen waste can be regarded as a first-order reaction in terms of reaction kinetics. This study serves as a good reference for future studies regarding the kinetics of anaerobic digestion of kitchen waste. PMID:28546964

Physicochemical structural changes of poplar and switchgrass during biomass pretreatment and enzymatic hydrolysis

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

Meng, Xianzhi; Sun, Qining; Kosa, Matyas

Converting lignocellulosics to simple sugars for second generation bioethanol is complicated due to biomass recalcitrance, and it requires a pretreatment stage prior to enzymatic hydrolysis. In this study, native, pretreated (acid and alkaline) and partially hydrolyzed poplar and switchgrass were characterized by using Simons’ staining for cellulose accessibility, GPC for degree of polymerization (DP), and FTIR for chemical structure of plant cell wall. The susceptibility of the pretreated biomass to enzymatic hydrolysis could not be easily predicted from differences in cellulose DP and accessibility. During hydrolysis, the most significant DP reduction occurred at the very beginning of hydrolysis, and themore » DP began to decrease at a significantly slower rate after this initial period, suggesting an existence of a synergistic action of endo- and exoglucanases that contribute to the occurrence of a “peeling off” mechanism. Cellulose accessibility was found to be increased at the beginning of hydrolysis, after reaching a maximum value then started to decrease. In conclusion, the fresh enzyme restart hydrolysis experiment along with the accessibility data indicated that the factors associated with the nature of enzyme such as irreversible nonspecific binding of cellulases by lignin and steric hindrance of enzymes should be responsible for the gradual slowing down of the reaction rate.« less

Physicochemical structural changes of poplar and switchgrass during biomass pretreatment and enzymatic hydrolysis

DOE PAGES

Meng, Xianzhi; Sun, Qining; Kosa, Matyas; ...

2016-07-27

Converting lignocellulosics to simple sugars for second generation bioethanol is complicated due to biomass recalcitrance, and it requires a pretreatment stage prior to enzymatic hydrolysis. In this study, native, pretreated (acid and alkaline) and partially hydrolyzed poplar and switchgrass were characterized by using Simons’ staining for cellulose accessibility, GPC for degree of polymerization (DP), and FTIR for chemical structure of plant cell wall. The susceptibility of the pretreated biomass to enzymatic hydrolysis could not be easily predicted from differences in cellulose DP and accessibility. During hydrolysis, the most significant DP reduction occurred at the very beginning of hydrolysis, and themore » DP began to decrease at a significantly slower rate after this initial period, suggesting an existence of a synergistic action of endo- and exoglucanases that contribute to the occurrence of a “peeling off” mechanism. Cellulose accessibility was found to be increased at the beginning of hydrolysis, after reaching a maximum value then started to decrease. In conclusion, the fresh enzyme restart hydrolysis experiment along with the accessibility data indicated that the factors associated with the nature of enzyme such as irreversible nonspecific binding of cellulases by lignin and steric hindrance of enzymes should be responsible for the gradual slowing down of the reaction rate.« less

Kinetic study of alkaline protease 894 for the hydrolysis of the pearl oyster Pinctada martensii

NASA Astrophysics Data System (ADS)

Chen, Xin; Chen, Hua; Cai, Bingna; Liu, Qingqin; Sun, Huili

2013-05-01

A new enzyme (alkaline protease 894) obtained from the marine extremophile Flavobacterium yellowsea (YS-80-122) has exhibited strong substrate-binding and catalytic activity, even at low temperature, but the characteristics of the hydrolysis with this enzyme are still unclear. The pearl oyster Pinctada martensii was used in this study as the raw material to illustrate the kinetic properties of protease 894. After investigating the intrinsic relationship between the degree of hydrolysis and several factors, including initial reaction pH, temperature, substrate concentration, enzyme concentration, and hydrolysis time, the kinetics model was established. This study showed that the optimal conditions for the enzymatic hydrolysis were an initial reaction pH of 5.0, temperature of 30°C, substrate concentration of 10% (w/v), enzyme concentration of 2 500 U/g, and hydrolysis time of 160 min. The kinetic characteristics of the protease for the hydrolysis of P. martensii were obtained. The inactivation constant was found to be 15.16/min, and the average relative error between the derived kinetics model and the actual measurement was only 3.04%, which indicated a high degree of fitness. Therefore, this study provides a basis for the investigation of the concrete kinetic characteristics of the new protease, which has potential applications in the food industry.

Application of the Initial Rate Method in Anaerobic Digestion of Kitchen Waste.

PubMed

Feng, Lei; Gao, Yuan; Kou, Wei; Lang, Xianming; Liu, Yiwei; Li, Rundong; Yu, Meiling; Shao, Lijie; Wang, Xiaoming

2017-01-01

This article proposes a methane production approach through sequenced anaerobic digestion of kitchen waste, determines the hydrolysis constants and reaction orders at both low total solid (TS) concentrations and high TS concentrations using the initial rate method, and examines the population growth model and first-order hydrolysis model. The findings indicate that the first-order hydrolysis model better reflects the kinetic process of gas production. During the experiment, all the influential factors of anaerobic fermentation retained their optimal values. The hydrolysis constants and reaction orders at low TS concentrations are then employed to demonstrate that the first-order gas production model can describe the kinetics of the gas production process. At low TS concentrations, the hydrolysis constants and reaction orders demonstrated opposite trends, with both stabilizing after 24 days at 0.99 and 1.1252, respectively. At high TS concentrations, the hydrolysis constants and the reaction orders stabilized at 0.98 (after 18 days) and 0.3507 (after 14 days), respectively. Given sufficient reaction time, the hydrolysis involved in anaerobic fermentation of kitchen waste can be regarded as a first-order reaction in terms of reaction kinetics. This study serves as a good reference for future studies regarding the kinetics of anaerobic digestion of kitchen waste.

Sulfated Pentagalloylglucoside is a Potent, Allosteric, and Selective Inhibitor of Factor XIa

PubMed Central

Al-Horani, Rami A.; Ponnusamy, Pooja; Mehta, Akul Y.; Gailani, David; Desai, Umesh R.

2013-01-01

Inhibition of factor XIa (FXIa) is a novel paradigm for developing anticoagulants without major bleeding consequences. We present the discovery of sulfated pentagalloylglucoside (6) as a highly selective inhibitor of human FXIa. Biochemical screening of a focused library led to the identification of 6, a sulfated aromatic mimetic of heparin. Inhibitor 6 displayed a potency of 551 nM against FXIa, which was at least 200-fold more selective than other relevant enzymes. It also prevented activation of factor IX and prolonged human plasma and whole blood clotting. Inhibitor 6 reduced VMAX of FXIa hydrolysis of chromogenic substrate without affecting the KM suggesting an allosteric mechanism. Competitive studies showed that 6 bound in the heparin-binding site of FXIa. No allosteric small molecule has been discovered to date that exhibits equivalent potency against FXIa. Inhibitor 6 is expected to open up a major route to allosteric FXIa anticoagulants with clinical relevance. PMID:23316863

Ezetimibe selectively inhibits intestinal cholesterol absorption in rodents in the presence and absence of exocrine pancreatic function

PubMed Central

van Heek, Margaret; Farley, Constance; Compton, Douglas S; Hoos, Lizbeth; Davis, Harry R

2001-01-01

Ezetimibe potently inhibits the transport of cholesterol across the intestinal wall, thereby reducing plasma cholesterol in preclinical animal models of hypercholesterolemia. The effect of ezetimibe on known absorptive processes was determined in the present studies.Experiments were conducted in the hamster and/or rat to determine whether ezetimibe would affect the absorption of molecules other than free cholesterol, namely cholesteryl ester, triglyceride, ethinylestradiol, progesterone, vitamins A and D, and taurocholic acid. In addition, to determine whether exocrine pancreatic function is involved in the mechanism of action of ezetimibe, a biliary anastomosis model, which eliminates exocrine pancreatic function from the intestine while maintaining bile flow, was established in the rat.Ezetimibe reduced plasma cholesterol and hepatic cholesterol accumulation in cholesterol-fed hamsters with an ED50 of 0.04 mg kg−1. Utilizing cholesteryl esters labelled on either the cholesterol or the fatty acid moiety, we demonstrated that ezetimibe did not affect cholesteryl ester hydrolysis and the absorption of fatty acid thus generated in both hamsters and rats. The free cholesterol from this hydrolysis, however, was not absorbed (92 – 96% inhibition) in the presence of ezetimibe. Eliminating pancreatic function in rats abolished hydrolysis of cholesteryl esters, but did not affect the ability of ezetimibe to block absorption of free cholesterol (−94%). Ezetimibe did not affect the absorption of triglyceride, ethinylestradiol, progesterone, vitamins A and D, and taurocholic acid in rats.Ezetimibe is a potent inhibitor of intestinal free cholesterol absorption that does not require exocrine pancreatic function for activity. Ezetimibe does not affect the absorption of triglyceride as a pancreatic lipase inhibitor (Orlistat) would, nor does it affect the absorption of vitamin A, D or taurocholate, as a bile acid sequestrant (cholestyramine) would. PMID:11564660

Enzymatic treatment of whey proteins in cow's milk results in differential inhibition of IgE-mediated mast cell activation compared to T-cell activation.

PubMed

Knipping, Karen; van Esch, Betty C A M; van Ieperen-van Dijk, Adrie G; van Hoffen, Els; van Baalen, Ton; Knippels, Léon M J; van der Heide, Sicco; Dubois, Anthony E J; Garssen, Johan; Knol, Edward F

2012-01-01

Cow's milk (CM) hydrolysates are frequently used as milk substitutes for children with CM allergy. In hydrolysates, allergenic epitopes within CM proteins are diminished by enzymatic treatment. The aim of this study was to examine the allergenic and immunogenic properties of whey proteins during hydrolysis. During hydrolysis, samples were obtained at 0, 10, 15, 30, 45, 60, 75 and 90 min. Degradation was checked by HPLC and SDS-PAGE. Allergenic potential was analyzed by IgE crosslinking capacity of human Fcε receptor type 1-transduced rat basophilic leukemia cells sensitized with serum of CM-allergic patients. Whey-sensitized C3H/HeOuJ mice were ear challenged intracutaneously with the hydrolysates. Immunogenicity was tested using whey-specific human T-cell clones and T-cell lines at the level of proliferation and release of IL-4, IL-10, IL-13 and IFN-γ. After 15 min of hydrolysis, the majority of the proteins were degraded. Hydrolysis for 15 min resulted in 92% inhibition of mast cell degranulation and in 82% reduction of ear swelling in the mouse model. In contrast, T-cell-stimulatory capacity was less affected by hydrolysis: reduction of human T-cell proliferation was only 9%. This was further reduced to 57 and 74% after 30 and 45 min of hydrolysis, respectively. Cytokine production followed the pattern of T-cell proliferation. Via differential analysis of allergenic versus immunogenic properties of the time kinetics of hydrolysis of whey proteins, we have demonstrated specific hydrolysis conditions with reduced IgE-crosslinking responses but retained T-cell activating properties. This approach might be useful in better defining CM hydrolysates. Copyright © 2012 S. Karger AG, Basel.

The Role of Factor XIa (FXIa) Catalytic Domain Exosite Residues in Substrate Catalysis and Inhibition by the Kunitz Protease Inhibitor Domain of Protease Nexin 2*

PubMed Central

Su, Ya-Chi; Miller, Tara N.; Navaneetham, Duraiswamy; Schoonmaker, Robert T.; Sinha, Dipali; Walsh, Peter N.

2011-01-01

To select residues in coagulation factor XIa (FXIa) potentially important for substrate and inhibitor interactions, we examined the crystal structure of the complex between the catalytic domain of FXIa and the Kunitz protease inhibitor (KPI) domain of a physiologically relevant FXIa inhibitor, protease nexin 2 (PN2). Six FXIa catalytic domain residues (Glu98, Tyr143, Ile151, Arg3704, Lys192, and Tyr5901) were subjected to mutational analysis to investigate the molecular interactions between FXIa and the small synthetic substrate (S-2366), the macromolecular substrate (factor IX (FIX)) and inhibitor PN2KPI. Analysis of all six Ala mutants demonstrated normal Km values for S-2366 hydrolysis, indicating normal substrate binding compared with plasma FXIa; however, all except E98A and K192A had impaired values of kcat for S-2366 hydrolysis. All six Ala mutants displayed deficient kcat values for FIX hydrolysis, and all were inhibited by PN2KPI with normal values of Ki except for K192A, and Y5901A, which displayed increased values of Ki. The integrity of the S1 binding site residue, Asp189, utilizing p-aminobenzamidine, was intact for all FXIa mutants. Thus, whereas all six residues are essential for catalysis of the macromolecular substrate (FIX), only four (Tyr143, Ile151, Arg3704, and Tyr5901) are important for S-2366 hydrolysis; Glu98 and Lys192 are essential for FIX but not S-2366 hydrolysis; and Lys192 and Tyr5901 are required for both inhibitor and macromolecular substrate interactions. PMID:21778227

Studies on the metabolism of benoxinate by human pseudocholinesterase.

PubMed

Dubbels, R; Schloot, W

1983-01-01

The local anesthetic drug benoxinate (oxybuprocaine, Novesine) is hydrolyzed to 3-butoxy-4-aminobenzoic acid. A rapid and simple spectrophotometric method for benoxinate hydrolysis by human plasma was developed. Benoxinate is hydrolyzed enzymatically by an esterase present in the serum. Heat stability characteristics and apparent affinity values of the benoxinate metabolizing enzyme were in the same range compared to benzoylcholine chloride hydrolysis. Apparent Vmax-values differ by a mean factor of about 18 between the hydrolysis of both substrates. Considerable interindividual variability of benoxinate hydrolysis and inhibition of the enzymatic reaction by dibucaine and sodium fluoride has been observed. Furthermore, enzyme activity with benoxinate as substrate is positively correlated (P less than 0.001) with benzoylcholine chloride hydrolysis. Therefore, we assume that benoxinate is metabolized by human pseudocholinesterase (PCHE, E.C. 3.1.1.8) and that ocular side effects after benoxinate application may be caused by altered metabolism of this drug, depending on genetically determined variants of pseudocholinesterase.

Rapid saccharification for production of cellulosic biofuels.

PubMed

Lee, Dae-Seok; Wi, Seung Gon; Lee, Soo Jung; Lee, Yoon-Gyo; Kim, Yeong-Suk; Bae, Hyeun-Jong

2014-04-01

The economical production of biofuels is hindered by the recalcitrance of lignocellulose to processing, causing high consumption of processing enzymes and impeding hydrolysis of pretreated lignocellulosic biomass. We determined the major rate-limiting factor in the hydrolysis of popping pre-treated rice straw (PPRS) by examining cellulase adsorption to lignin and cellulose, amorphogenesis of PPRS, and re-hydrolysis. Based on the results, equivalence between enzyme loading and the open structural area of cellulose was required to significantly increase productive adsorption of cellulase and to accelerate enzymatic saccharification of PPRS. Amorphogenesis of PPRS by phosphoric acid treatment to expand open structural area of the cellulose fibers resulted in twofold higher cellulase adsorption and increased the yield of the first re-hydrolysis step from 13% to 46%. The total yield from PPRS was increased to 84% after 3h. These results provide evidence that cellulose structure is one of major effects on the enzymatic hydrolysis. Copyright © 2014 Elsevier Ltd. All rights reserved.

Origin Licensing Requires ATP Binding and Hydrolysis by the MCM Replicative Helicase

PubMed Central

Coster, Gideon; Frigola, Jordi; Beuron, Fabienne; Morris, Edward P.; Diffley, John F.X.

2014-01-01

Summary Loading of the six related Minichromosome Maintenance (MCM) proteins as head-to-head double hexamers during DNA replication origin licensing is crucial for ensuring once-per-cell-cycle DNA replication in eukaryotic cells. Assembly of these prereplicative complexes (pre-RCs) requires the Origin Recognition Complex (ORC), Cdc6, and Cdt1. ORC, Cdc6, and MCM are members of the AAA+ family of ATPases, and pre-RC assembly requires ATP hydrolysis. Here we show that ORC and Cdc6 mutants defective in ATP hydrolysis are competent for origin licensing. However, ATP hydrolysis by Cdc6 is required to release nonproductive licensing intermediates. We show that ATP binding stabilizes the wild-type MCM hexamer. Moreover, by analyzing MCM containing mutant subunits, we show that ATP binding and hydrolysis by MCM are required for Cdt1 release and double hexamer formation. This work alters our view of how ATP is used by licensing factors to assemble pre-RCs. PMID:25087873

Kinetic characterization of factor Xa binding using a quenched fluorescent substrate based on the reactive site of factor Xa inhibitor from Bauhinia ungulata seeds.

PubMed

Oliva, M L V; Andrade, S A; Juliano, M A; Sallai, R C; Torquato, R J; Sampaio, M U; Pott, V J; Sampaio, C A M

2003-07-01

The specific Kunitz Bauhinia ungulata factor Xa inhibitor (BuXI) and the Bauhinia variegata trypsin inhibitor (BvTI) blocked the activity of trypsin, chymotrypsin, plasmin, plasma kallikrein and factor XIIa, and factor Xa inhibition was achieved only by BuXI (K(i) 14 nM). BuXI and BvTI are highly homologous (70%). The major differences are the methionine residues at BuXI reactive site, which are involved in the inhibition, since the oxidized protein no longer inhibits factor Xa but maintains the trypsin inhibition. Quenched fluorescent substrates based on the reactive site sequence of the inhibitors were synthesized and the kinetic parameters of the hydrolysis were determined using factor Xa and trypsin. The catalytic efficiency k(cat)/K(m) 4.3 x 10(7) M(-1)sec(>-1) for Abz-VMIAALPRTMFIQ-EDDnp (lead peptide) hydrolysis by factor Xa was 10(4)-fold higher than that of Boc-Ile-Glu-Gly-Arg-AMC, widely used as factor Xa substrate. Lengthening of the substrate changed its susceptibility to factor Xa hydrolysis. Both methionine residues in the substrate influence the binding to factor Xa. Serine replacement of threonine (P(1)') decreases the catalytic efficiency by four orders of magnitude. Factor Xa did not hydrolyze the substrate containing the reactive site sequence of BvTI, that inhibits trypsin inhibitor but not factor Xa. Abz-VMIAALPRTMFIQ-EDDnp prolonged both the prothrombin time and the activated partial thromboplastin time, and the other modified substrates used in this experiment altered blood-clotting assays.

Unraveling the effects of laccase treatment on enzymatic hydrolysis of steam-exploded wheat straw.

PubMed

Oliva-Taravilla, Alfredo; Moreno, Antonio D; Demuez, Marie; Ibarra, David; Tomás-Pejó, Elia; González-Fernández, Cristina; Ballesteros, Mercedes

2015-01-01

Laccase enzymes are promising detoxifying agents during lignocellulosic bioethanol production from wheat straw. However, they affect the enzymatic hydrolysis of this material by lowering the glucose recovery yields. This work aimed at explaining the negative effects of laccase on enzymatic hydrolysis. Relative glucose recovery in presence of laccase (10IU/g substrate) with model cellulosic substrate (Sigmacell) at 10% (w/v) was almost 10% points lower (P<0.01) than in the absence of laccase. This fact could be due to an increase in the competition of cellulose binding sites between the enzymes and a slight inhibition of β-glucosidase activity. However, enzymatic hydrolysis and infrared spectra of laccase-treated and untreated wheat straw filtered pretreated residue (WS-FPR), revealed that a grafting process of phenoxy radicals onto the lignin fiber could be the cause of diminished accessibility of cellulases to cellulose in pretreated wheat straw. Copyright © 2014 Elsevier Ltd. All rights reserved.

Statistical optimization of recycled-paper enzymatic hydrolysis for simultaneous saccharification and fermentation via central composite design.

PubMed

Liu, Qing; Cheng, Ke-ke; Zhang, Jian-an; Li, Jin-ping; Wang, Ge-hua

2010-01-01

A central composite design of the response surface methodology (RSM) was employed to study the effects of temperature, enzyme concentration, and stirring rate on recycled-paper enzymatic hydrolysis. Among the three variables, temperature and enzyme concentration significantly affected the conversion efficiency of substrate, whereas stirring rate was not effective. A quadratic polynomial equation was obtained for enzymatic hydrolysis by multiple regression analysis using RSM. The results of validation experiments were coincident with the predicted model. The optimum conditions for enzymatic hydrolysis were temperature, enzyme concentration, and stirring rate of 43.1 degrees C, 20 FPU g(-1) substrate, and 145 rpm, respectively. In the subsequent simultaneous saccharification and fermentation (SSF) experiment under the optimum conditions, the highest 28.7 g ethanol l(-1) was reached in the fed-batch SSF when 5% (w/v) substrate concentration was used initially, and another 5% added after 12 h fermentation. This ethanol output corresponded to 77.7% of the theoretical yield based on the glucose content in the raw material.

Structure and optical properties of ZnO produced from microwave hydrothermal hydrolysis of tris(ethylenediamine)zinc nitrate complex

NASA Astrophysics Data System (ADS)

Mostafa, Nasser Y.; Heiba, Zein K.; Ibrahim, Mohamed M.

2015-01-01

ZnO powders were synthesized using a solution microwave hydrothermal hydrolysis process and tris(ethylenediamine)zinc nitrate {[Zn(en)3](NO3)2} (en = ethylenediamine) as a precursor. Hydrolysis of the precursor complex at different pH produced zinc oxide with a diversity of well-defined morphologies. The effect of hydrolysis pH values on the structural and optical properties has been explored using XRD, SEM, and UV-visible diffuse reflectance spectroscopy (DRS). At pH = 7.0, randomly dispersed rods were formed. Whereas flower-like morphologies were obtained by treating the complex precursor in water at pH = 10.0 and 12.0. The ZnO4 tetrahedrons are greatly affected by the pH value. The band gap decreased sharply with increasing the pH value from 7.0 to 10.0, then slightly decreased with further increasing the pH to 12.0. The relationship between band gap and both structure and surface defects of the samples is also discussed.

Bitterness in sodium caseinate hydrolysates: role of enzyme preparation and degree of hydrolysis.

PubMed

O'Sullivan, Dara; Nongonierma, Alice B; FitzGerald, Richard J

2017-10-01

Enzymatic hydrolysis of sodium caseinate (NaCas) may lead to the development of bitterness. Careful selection of hydrolysis conditions (i.e. enzyme preparation and duration) yielding different degrees of hydrolysis (DH) may aid in the development of low bitterness. Eighteen NaCas hydrolysates were generated with four enzyme preparations (Alcalase 2.4L, Prolyve 1000, FlavorPro Whey and pepsin) to different DH values. Hydrolysate bitterness score, assessed using a trained panel (ten assessors), generally increased at higher DH values for Alcalase, Prolyve and pepsin hydrolysates. However, all FlavorPro Whey hydrolysates (DH 0.38-10.62%) displayed low bitterness score values (<26.0%) comparable to that of intact NaCas (13.8 ± 2.0%, P > 0.05). Enzyme preparation and DH affect the bitterness of NaCas hydrolysates. The results are relevant for the generation of NaCas hydrolysates with reduced bitterness. © 2017 Society of Chemical Industry. © 2017 Society of Chemical Industry.

Simultaneous Improvement of Oxidative and Hydrolytic Resistance of Polycarbonate Urethanes Based on Polydimethylsiloxane/Poly(hexamethylene carbonate) Mixed Macrodiols.

PubMed

Li, Zhen; Yang, Jian; Ye, Heng; Ding, Mingming; Luo, Feng; Li, Jianshu; Li, Jiehua; Tan, Hong; Fu, Qiang

2018-06-11

The degradation behaviors including oxidation and hydrolysis of silicone modified polycarbonate urethanes were thoroughly investigated. These polyurethanes were based on polyhexamethylene carbonate (PHMC)/polydimethylsiloxane (PDMS) mixed macrodiols with molar ratio of PDMS ranging from 5% to 30%. It was proved that PDMS tended to migrate toward surface and even a small amount of PDMS could form a silicone-like surface. Macrophages-mediated oxidation process indicated that the PDMS surface layer was desirable to protect the fragile soft PHMC from the attack of degradative species. Hydrolysis process was probed in detail after immersing in boiling buffered water using combined analytical tools. Hydrolytically stable PDMS could act as protective shields for the bulk to hinder the chain scission of polycarbonate carbonyls whereas the hydrolysis of urethane linkages was less affected. Although the promoted phase separation at higher PDMS fractions lead to possible physical defects and mechanical compromise after degradation, simultaneously enhanced oxidation and hydrolysis resistance could be achieved for the polyurethanes with proper PDMS incorporation.

Modeling of Anaerobic Digestion with a Focus on Estimation of Hydrolysis Constants at 35, 55, and 60 °C.

PubMed

Haghighatafshar, Salar; Ossiansson, Elin; Koch, Konrad; Kjerstadius, Hamse; Jansen, Jes la Cour; Davidsson, Åsa

2015-07-01

Hydrolysis constants of mixed sludge at 35, 55, and 60 °C were found to be 0.32, 0.44, and 0.50 1/d, respectively, in pilot-scale, semicontinuously operated anaerobic digesters. The hydrolysis constants and estimated chemical oxygen demand fractions in the feed were introduced to a mathematical model for anaerobic digestion published by Siegrist et al. (2002), which is similar to Anaerobic Digestion Model No. 1. First-order and Monod-type kinetics were tested for estimation of hydrolysis constants. The applied kinetics were found to affect the outcome of the regression study. Moreover, the free ammonia inhibition model was excluded for both propionate oxidation and acetate conversion, thanks to the apparent acclimatized biomass. No substantial accumulation of volatile fatty acids was observed in the reactors at 35, 55, and 60 °C, corresponding to free ammonia nitrogen concentrations of about 20, 110, and 130 g N/m³, respectively.

Production and characterization of cowpea protein hydrolysate with optimum nitrogen solubility by enzymatic hydrolysis using pepsin.

PubMed

Mune Mune, Martin Alain; Minka, Samuel René

2017-06-01

Cowpea is a source of low-cost and good nutritional quality protein for utilization in food formulations in replacement of animal proteins. Therefore it is necessary that cowpea protein exhibits good functionality, particularly protein solubility which affects the other functional properties. The objective of this study was to produce cowpea protein hydrolysate exhibiting optimum solubility by the adequate combination of hydrolysis parameters, namely time, solid/liquid ratio (SLR) and enzyme/substrate ratio (ESR), and to determine its functional properties and molecular characteristics. A Box-Behnken experimental design was used for the experiments, and a second-order polynomial to model the effects of hydrolysis time, SLR and ESR on the degree of hydrolysis and nitrogen solubility index. The optimum hydrolysis conditions of time 208.61 min, SLR 1/15 (w/w) and ESR 2.25% (w/w) yielded a nitrogen solubility of 75.71%. Protein breakdown and the peptide profile following enzymatic hydrolysis were evaluated by sodium dodecyl sulfate polyacrylamide gel electrophoresis and size exclusion chromatography. Cowpea protein hydrolysate showed higher oil absorption capacity, emulsifying activity and foaming ability compared with the concentrate. The solubility of cowpea protein hydrolysate was adequately optimized by response surface methodology, and the hydrolysate showed adequate functionality for use in food. © 2016 Society of Chemical Industry. © 2016 Society of Chemical Industry.

Enhanced enzymatic hydrolysis of waste paper for ethanol production using separate saccharification and fermentation.

PubMed

Guerfali, Mohamed; Saidi, Adel; Gargouri, Ali; Belghith, Hafedh

2015-01-01

Ethanol produced from lignocellulosic biomass is a renewable alternative to diminishing petroleum-based liquid fuels. In this study, the feasibility of ethanol production from waste paper using the separate hydrolysis and fermentation (SHF) was investigated. Two types of waste paper materials, newspaper and office paper, were evaluated for their potential to be used as a renewable feedstock for the production of fermentable sugars via enzymatic hydrolysis of their cellulose fractions. Hydrolysis step was conducted with a mixture of cellulolytic enzymes produced locally by Trichoderma reesei Rut-C30 (cellulase-overproducing mutant) and Aspergillus niger F38 cultures. Surfactant pretreatment effect on waste paper enzymatic digestibility was studied and Triton X-100 at 0.5 % (w w(-1)) has improved the digestibility of newspaper about 45 %. The effects of three factors (dry matter quantity, phosphoric acid pretreatment and hydrolysis time) on the extent of saccharification were also assessed and quantified by using a methodical approach based on response surface methodology. Under optimal hydrolysis conditions, maximum degrees of saccharification of newspaper and office paper were 67 and 92 %, respectively. Sugars released from waste paper were subsequently converted into ethanol (0.38 g ethanol g(-1) sugar) with Saccharomyces cerevisiae CTM-30101.

Determination of Flavonol Aglycones in Ginkgo biloba Dietary Supplement Crude Materials and Finished Products by High-Performance Liquid Chromatography: Single Laboratory Validation

PubMed Central

Gray, Dean; LeVanseler, Kerri; Pan, Meide

2008-01-01

A single laboratory validation (SLV) was completed for a method to determine the flavonol aglycones quercetin, kaempferol, and isorhamnetin in Ginkgo biloba products. The method calculates total glycosides based on these aglycones formed following acid hydrolysis. Nine matrixes were chosen for the study, including crude leaf material, standardized dry powder extract, single and multiple entity finished products, and ethanol and glycerol tinctures. For the 9 matrixes evaluated as part of this SLV, the method appeared to be selective and specific, with no observed interferences. The simplified 60 min oven heating hydrolysis procedure was effective for each of the matrixes studied, with no apparent or consistent differences between 60, 75, and 90 min at 90°C. A Youden ruggedness trial testing 7 factors with the potential to affect quantitative results showed that 2 factors (volume hydrolyzed and test sample extraction/hydrolysis weight) were the most important parameters for control during sample preparation. The method performed well in terms of precision, with 4 matrixes tested in triplicate over a 3-day period showing an overall repeatability (relative standard deviation, RSD) of 2.3%. Analysis of variance testing at α = 0.05 showed no significant differences among the within- or between-group sources of variation, although comparisons of within-day (Sw), between-day (Sb), and total (St) precision showed that a majority of the standard deviation came from within-day determinations for all matrixes. Accuracy testing at 2 levels (approximately 30 and 90% of the determined concentrations in standardized dry powder extract) from 2 complex negative control matrixes showed an overall 96% recovery and RSD of 1.0% for the high spike, and 94% recovery and RSD of 2.5% for the low spike. HorRat scores were within the limits for performance acceptability, ranging from 0.4 to 1.3. Based on the performance results presented herein, it is recommended that this method progress to the collaborative laboratory trial. PMID:16001841

Engineering secondary cell wall deposition in plants

PubMed Central

Yang, Fan; Mitra, Prajakta; Zhang, Ling; Prak, Lina; Verhertbruggen, Yves; Kim, Jin-Sun; Sun, Lan; Zheng, Kejian; Tang, Kexuan; Auer, Manfred; Scheller, Henrik V; Loqué, Dominique

2013-01-01

Lignocellulosic biomass was used for thousands of years as animal feed and is now considered a great sugar source for biofuels production. It is composed mostly of secondary cell walls built with polysaccharide polymers that are embedded in lignin to reinforce the cell wall structure and maintain its integrity. Lignin is the primary material responsible for biomass recalcitrance to enzymatic hydrolysis. During plant development, deep reductions of lignin cause growth defects and often correlate with the loss of vessel integrity that adversely affects water and nutrient transport in plants. The work presented here describes a new approach to decrease lignin content while preventing vessel collapse and introduces a new strategy to boost transcription factor expression in native tissues. We used synthetic biology tools in Arabidopsis to rewire the secondary cell network by changing promoter-coding sequence associations. The result was a reduction in lignin and an increase in polysaccharide depositions in fibre cells. The promoter of a key lignin gene, C4H, was replaced by the vessel-specific promoter of transcription factor VND6. This rewired lignin biosynthesis specifically for vessel formation while disconnecting C4H expression from the fibre regulatory network. Secondly, the promoter of the IRX8 gene, secondary cell wall glycosyltransferase, was used to express a new copy of the fibre transcription factor NST1, and as the IRX8 promoter is induced by NST1, this also created an artificial positive feedback loop (APFL). The combination of strategies—lignin rewiring with APFL insertion—enhances polysaccharide deposition in stems without over-lignifying them, resulting in higher sugar yields after enzymatic hydrolysis. PMID:23140549

Bauhinia proteinase inhibitor-based synthetic fluorogenic substrates for enzymes isolated from insect midgut and caterpillar bristles.

PubMed

Andrade, Sonia A; Santomauro-Vaz, Eugênio M; Lopes, Adriana R; Chudzinski-Tavassi, Ana M; Juliano, Maria A; Terra, Walter R; Sampaio, Misako U; Sampaio, Claudio A M; Oliva, Maria Luiza V

2003-03-01

Bauhinia ungulata factor Xa inhibitor (BuXI) inactivates factor Xa and LOPAP, a prothrombin activator proteinase isolated from the venom of Lonomia obliqua caterpillar bristles. The reactive site of the enzyme-inhibitor interaction was explored to design specific substrates for both enzymes. Methionine is crucial for LOPAP and factor Xa substrate interaction, since the change of both Met residues in the substrates abolished the hydrolysis. Synthetic substrates containing the sequence around the reactive site of BbKI, a plasma kallikrein inhibitor, were shown to be specific for trypsin hydrolysis. Therefore, these substrates may be an alternative in studies aiming at a characterization of trypsin-like enzyme activities, especially non-mammalian enzymes.

Hydrolysis of phosphatidylcholine during LDL oxidation is mediated by platelet-activating factor acetylhydrolase.

PubMed

Steinbrecher, U P; Pritchard, P H

1989-03-01

Degradation of phosphatidylcholine to lysophosphatidylcholine occurs during oxidative modification of low density lipoproteins (LDL). In this study, we have shown that this phospholipid hydrolysis is brought about by an LDL-associated phospholipase A2 that can hydrolyze oxidized but not intact LDL phosphatidylcholine. The chemical nature of the oxidized phospholipids that can act as substrates for this enzyme was not fully characterized, but we hypothesized that the specificity of the enzyme for oxidized LDL phosphatidylcholine might be explained by fragmentation of polyunsaturated sn-2 fatty acyl groups in LDL phosphatidylcholine during oxidation. To facilitate characterization of this enzyme, we therefore selected a fluorescent phosphatidylcholine substrate that had a short-chain, polar residue in the sn-2 position: 1-palmitoyl 2-(6-[7-nitrobenzoxadiazolyl]amino) caproyl phosphatidylcholine, (C6NBD PC). This substrate was efficiently hydrolyzed by LDL, but the dodecanoyl analogue of C6NBD PC, which differed only in that a 12-carbon rather than a 6-carbon acyl derivative was present in the sn-2 position, was not hydrolyzed. The phospholipase activity was heat-stable, calcium-independent, and was inhibited by the serine esterase inhibitors phenylmethylsulfonyl-fluoride and diisopropylfluorophosphate, but was resistant to p-bromophenacylbromide and dithiobisnitrobenzoic acid. The phospholipid hydrolysis could not be attributed to the action of lecithin:cholesterol acyltransferase or lipoprotein lipase. Nearly all of the activity in EDTA-anticoagulated normal plasma was physically associated with apoB-containing lipoproteins, but this apoprotein was not essential as enzyme activity was present in plasma from abetalipoproteinemic patients. These properties are very similar to those recently reported for human plasma platelet-activating factor (PAF) acetylhydrolase. In the present study, we found that acylhydrolase activity against C6NBD PC, PAF, and oxidized phosphatidylcholine copurfied through gel filtration and ion-exchange chromatography. Substrate competition was demonstrated between C6NBD PC, PAF, and oxidized 2-arachidonyl phosphatidylcholine, suggesting that a single enzyme was active against all three substrates. The enzyme had an apparent molecular weight of 40,000-45,000 by high pressure gel exclusion chromatography. Inhibition of this activity with disopropyfluorophosphate prior to oxidative modification of LDL prevented phospholipid hydrolysis but did not affect the production of thiobarbituric acid reactive compounds or the change in electrophoretic mobility. In addition, this inhibition of phospholipase did not prevent the rapid degradati

Cold active β-galactosidase from Thalassospira sp. 3SC-21 to use in milk lactose hydrolysis: a novel source from deep waters of Bay-of-Bengal.

PubMed

Ghosh, Mrinmoy; Pulicherla, K K; Rekha, V P B; Raja, P Kumar; Sambasiva Rao, K R S

2012-09-01

The cold active β-galactosidase from psychrophilic bacteria accelerate the possibility of outperforming the current commercial β-galactosidase production from mesophilic sources. The present study is carried out to screen and isolate a cold active β-galactosidase producing bacterium from profound marine waters of Bay-of-Bengal and to optimize the factors for lactose hydrolysis in milk. Isolated bacterium 3SC-21 was characterized as marine psychrotolerant, halophile, gram negative, rod shaped strain producing an intracellular cold active β-galactosidase enzyme. Further, based upon the 16S rRNA gene sequence, bacterium 3SC-21 was identified as Thalassospira sp. The isolated strain Thalassospira sp. 3SC-21 had shown the enzyme activity between 4 and 20 °C at pH of 6.5 and the enzyme was completely inactivated at 45 °C. The statistical method, central composite rotatable design of response surface methodology was employed to optimize the hydrolysis of lactose and to reveal the interactions between various factors behind this hydrolysis. It was found that maximum of 80.18 % of lactose in 8 ml of raw milk was hydrolysed at pH of 6.5 at 20 °C in comparison to 40 % of lactose hydrolysis at 40 °C, suggesting that the cold active β-galactosidase from Thalassospira sp. 3SC-21 would be best suited for manufacturing the lactose free dairy products at low temperature.

BSA treatment to enhance enzymatic hydrolysis of cellulose in lignin containing substrates.

PubMed

Yang, Bin; Wyman, Charles E

2006-07-05

Cellulase and bovine serum albumin (BSA) were added to Avicel cellulose and solids containing 56% cellulose and 28% lignin from dilute sulfuric acid pretreatment of corn stover. Little BSA was adsorbed on Avicel cellulose, while pretreated corn stover solids adsorbed considerable amounts of this protein. On the other hand, cellulase was highly adsorbed on both substrates. Adding a 1% concentration of BSA to dilute acid pretreated corn stover prior to enzyme addition at 15 FPU/g cellulose enhanced filter paper activity in solution by about a factor of 2 and beta-glucosidase activity in solution by about a factor of 14. Overall, these results suggested that BSA treatment reduced adsorption of cellulase and particularly beta-glucosidase on lignin. Of particular note, BSA treatment of pretreated corn stover solids prior to enzymatic hydrolysis increased 72 h glucose yields from about 82% to about 92% at a cellulase loading of 15 FPU/g cellulose or achieved about the same yield at a loading of 7.5 FPU/g cellulose. Similar improvements were also observed for enzymatic hydrolysis of ammonia fiber explosion (AFEX) pretreated corn stover and Douglas fir treated by SO(2) steam explosion and for simultaneous saccharification and fermentation (SSF) of BSA pretreated corn stover. In addition, BSA treatment prior to hydrolysis reduced the need for beta-glucosidase supplementation of SSF. The results are consistent with non-specific competitive, irreversible adsorption of BSA on lignin and identify promising strategies to reduce enzyme requirements for cellulose hydrolysis. (c) 2006 Wiley Periodicals, Inc.

Effect of high hydrostatic pressure on the enzymatic hydrolysis of bovine serum albumin.

PubMed

De Maria, Serena; Ferrari, Giovanna; Maresca, Paola

2017-08-01

The extent of enzymatic proteolysis mainly depends on accessibility of the peptide bonds, which stabilize the protein structure. The high hydrostatic pressure (HHP) process is able to induce, at certain operating conditions, protein displacement, thus suggesting that this technology can be used to modify protein resistance to the enzymatic attack. This work aims at investigating the mechanism of enzymatic hydrolysis assisted by HHP performed under different processing conditions (pressure level, treatment time). Bovine serum albumin was selected for the experiments, solubilized in sodium phosphate buffer (25 mg mL -1 , pH 7.5) with α-chymotrypsin or trypsin (E/S ratio = 1/10) and HPP treatment (100-500 MPa, 15-25 min). HHP treatment enhanced the extent of the hydrolysis reaction of globular proteins, being more effective than conventional hydrolysis. At HHP treatment conditions maximizing the protein unfolding, the hydrolysis degree of proteins was increased as a consequence of the increased exposure of peptide bonds to the attack of proteolytic enzymes. The maximum hydrolysis degree (10% and 7% respectively for the samples hydrolyzed with α-chymotrypsin and trypsin) was observed for the samples processed at 400 MPa for 25 min. At pressure levels higher than 400 MPa the formation of aggregates was likely to occur; thus the degree of hydrolysis decreased. Protein unfolding represents the key factor controlling the efficiency of HHP-assisted hydrolysis treatments. The peptide produced under high pressure showed lower dimensions and a different structure with respect to those of the hydrolysates obtained when the hydrolysis was carried out at atmospheric pressure, thus opening new frontiers of application in food science and nutrition. © 2016 Society of Chemical Industry. © 2016 Society of Chemical Industry.

Immobilization of lipases in hydrophobic chitosan for selective hydrolysis of fish oil: The impact of support functionalization on lipase activity, selectivity and stability.

PubMed

Urrutia, P; Arrieta, R; Alvarez, L; Cardenas, C; Mesa, M; Wilson, L

2018-03-01

The objective of this paper was to carry out an integral study of the use of hydrophobic chitosan as a low-cost support for immobilizing lipases and their further application in the selective hydrolysis of fish oil. Chitosan functionalized with different alkyl chains (C4, C8, C12) were characterized by FTIR, TGA, SEM, and Rose Bengal adsorption. Lipase B from Candida antarctica (CalB) and lipase from Rhizomucor miehei (RML) were immobilized obtaining a higher expressed activity at a longer alkyl chain length of support. Biocatalyst thermal stability showed that the impact of the alkyl chain length on enzyme stabilization varied according to the lipase source. The biocatalysts were applied in menhaden oil hydrolysis. Total polyunsaturated fatty acids released after 30 h of reaction with lipases immobilized in butyl, octyl and dodecyl-chitosan was 60, 107, and 90 mM for CalB biocatalysts, and 560, 392, and 50 mM for RML biocatalysts, respectively. Selectivity of CalB was not affected by the alkyl chain, while in the case of RML, a higher selectivity to cis-4,7,10,13,16,19-docohexaenoic acid release was obtained with dodecyl-chitosan. In conclusion, the adequate functionalization of chitosan varied according to lipase source, affecting their activity, stability and performance in the hydrolysis of fish oil. Copyright © 2017 Elsevier B.V. All rights reserved.

Regulation of glucose transport by insulin, bombesin, and bradykinin in Swiss 3T3 fibroblasts: Involvement of protein kinase C-dependent and -independent mechanisms

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

Dettori, C.; Meldolesi, J.

1989-05-01

Glucose transport stimulation by insulin, bombesin, and bradykinin in Swiss 3T3 fibroblasts was compared with the phosphoinositide hydrolysis effects of the same stimulants in a variety of experimental paradigms known to affect generation and/or functioning of intracellular second messengers: short- and long-term treatments with phorbol dibutyrate, that cause activation and down-regulation of protein kinase C, respectively; cell loading with high (quin2), that causes clamping of (Ca{sup 2+}){sub i} near the resting level; poisoning with pertussis toxin, that affects the GTP binding proteins of the Go/Gi class; treatment with Ca{sup 2+} ionophores. ({sup 14}C) glucose transport stimulation by maximal (insulin) wasmore » affected by neither pertussis toxin nor protein kinase C down-regulation. This result correlates with the lack of effect of insulin on phosphoinositide hydrolysis. In contrast, part of the glucose transport responses induced by bombesin and bradykinin appeared to be mediated by protein kinase C in proportion with the stimulation induced by these peptides on the phosphoinositide hydrolysis. The protein kinase C-independent portion of the response to bradykinin was found to be inhibitable by pertussis toxin. This latter result might suggest an interaction between the bradykinin receptor and a glucose transporter, mediated by a protein of the Go/Gi class.« less

Digestibility index and factors affecting rate of starch digestion in vitro in conventional food preparation.

PubMed

Urooj, A; Puttraj, S

1999-02-01

The rate of starch hydrolysis in ten cereal-based food preparations was studied using an in vitro dialysis system. The foods were incubated with human saliva and porcine pancreatin. The sugars released after 3 h digestion were expressed as digestibility index (DI), the percentage starch digested was determined and correlated with the degree of gelatinization (DG). Granule morphology was also investigated and related with starch availability for hydrolysis. Significant differences were observed in the in vitro starch digestibility of the 10 foods (P < 0.05). The DI ranged from 53 for chapathi to 78 for rice flakes. DI was inversely related to the protein (r = -0.79, P < 0.01), fat (r = -0.63, P < 0.05) and energy (r = -0.61, P < 0.01). Percent starch digested was inversely related to the insoluble (r = -0.49, P < 0.05) and total dietary fiber (r = -0.63, P < 0.01) content of the foods. The SEM results provided a better understanding of granular morphology on cooking and the effect of protein on limiting DG. The results suggest that carbohydrate foods of potential use in the therapeutic diets may be identified by their in vitro digestion characteristics.

Cloning and Biochemical Characterization of TAF-172, a Human Homolog of Yeast Mot1

PubMed Central

Chicca, John J.; Auble, David T.; Pugh, B. Franklin

1998-01-01

The TATA binding protein (TBP) is a central component of the eukaryotic transcriptional machinery and is the target of positive and negative transcriptional regulators. Here we describe the cloning and biochemical characterization of an abundant human TBP-associated factor (TAF-172) which is homologous to the yeast Mot1 protein and a member of the larger Snf2/Swi2 family of DNA-targeted ATPases. Like Mot1, TAF-172 binds to the conserved core of TBP and uses the energy of ATP hydrolysis to dissociate TBP from DNA (ADI activity). Interestingly, ATP also causes TAF-172 to dissociate from TBP, which has not been previously observed with Mot1. Unlike Mot1, TAF-172 requires both TBP and DNA for maximal (∼100-fold) ATPase activation. TAF-172 inhibits TBP-driven RNA polymerase II and III transcription but does not appear to affect transcription driven by TBP-TAF complexes. As it does with Mot1, TFIIA reverses TAF-172-mediated repression of TBP. Together, these findings suggest that human TAF-172 is the functional homolog of yeast Mot1 and uses the energy of ATP hydrolysis to remove TBP (but apparently not TBP-TAF complexes) from DNA. PMID:9488487

Evaluation of Bacterial Expansin EXLX1 as a Cellulase Synergist for the Saccharification of Lignocellulosic Agro-Industrial Wastes

PubMed Central

Lin, Hui; Shen, Qi; Zhan, Ju-Mei; Wang, Qun; Zhao, Yu-Hua

2013-01-01

Various types of lignocellulosic wastes extensively used in biofuel production were provided to assess the potential of EXLX1 as a cellulase synergist. Enzymatic hydrolysis of natural wheat straw showed that all the treatments using mixtures of cellulase and an optimized amount of EXLX1, released greater quantities of sugars than those using cellulase alone, regardless of cellulase dosage and incubation time. EXLX1 exhibited different synergism and binding characteristics for different wastes, but this can be related to their lignocellulosic components. The cellulose proportion could be one of the important factors. However, when the cellulose proportion of different biomass samples exhibited no remarkable differences, a higher synergism of EXLX1 is prone to occur on these materials, with a high proportion of hemicellulose and a low proportion of lignin. The information could be favorable to assess whether EXLX1 is effective as a cellulase synergist for the hydrolysis of the used materials. Binding assay experiments further suggested that EXLX1 bound preferentially to alkali pretreated materials, as opposed to acid pretreated materials under the assay condition and the binding preference would be affected by incubation temperature. PMID:24086425

Lignin content in natural Populus variants affects sugar release

PubMed Central

Studer, Michael H.; DeMartini, Jaclyn D.; Davis, Mark F.; Sykes, Robert W.; Davison, Brian; Keller, Martin; Tuskan, Gerald A.; Wyman, Charles E.

2011-01-01

The primary obstacle to producing renewable fuels from lignocellulosic biomass is a plant's recalcitrance to releasing sugars bound in the cell wall. From a sample set of wood cores representing 1,100 individual undomesticated Populus trichocarpa trees, 47 extreme phenotypes were selected across measured lignin content and ratio of syringyl and guaiacyl units (S/G ratio). This subset was tested for total sugar release through enzymatic hydrolysis alone as well as through combined hot-water pretreatment and enzymatic hydrolysis using a high-throughput screening method. The total amount of glucan and xylan released varied widely among samples, with total sugar yields of up to 92% of the theoretical maximum. A strong negative correlation between sugar release and lignin content was only found for pretreated samples with an S/G ratio < 2.0. For higher S/G ratios, sugar release was generally higher, and the negative influence of lignin was less pronounced. When examined separately, only glucose release was correlated with lignin content and S/G ratio in this manner, whereas xylose release depended on the S/G ratio alone. For enzymatic hydrolysis without pretreatment, sugar release increased significantly with decreasing lignin content below 20%, irrespective of the S/G ratio. Furthermore, certain samples featuring average lignin content and S/G ratios exhibited exceptional sugar release. These facts suggest that factors beyond lignin and S/G ratio influence recalcitrance to sugar release and point to a critical need for deeper understanding of cell-wall structure before plants can be rationally engineered for reduced recalcitrance and efficient biofuels production. PMID:21444820

Optimization of hydrolysis conditions for the production of glucomanno-oligosaccharides from konjac using β-mannanase by response surface methodology.

PubMed

Chen, Junfan; Liu, Desheng; Shi, Bo; Wang, Hai; Cheng, Yongqiang; Zhang, Wenjing

2013-03-01

Glucomanno-oligosaccharides (GMO), usually produced from hydrolysis of konjac tubers with a high content of glucomannan, have a positive effect on Bifidobacterium as well as a variety of other physiological activities. Response surface methodology (RSM) was employed to optimize the hydrolysis time, hydrolysis temperature, pH and enzyme to substrate ratio (E/S) to obtain a high GMO yield from konjac tubers. From the signal-factor experiments, it was concluded that the change in the direct reducing sugar (DRS) is consistent with total reducing sugar (TRS) but contrary to the degree of polymerization (DP). DRS was used as an indicator of the content of GMO in the RSM study. The optimum RSM operating conditions were: reaction time of 3.4 h, reaction temperature of 41.0°C, pH of 7.1 and E/S of 0.49. The results suggested that the enzymatic hydrolysis was enhanced by temperature, pH and incubation time. Model validation showed good agreement between experimental results and the predicted responses. Copyright © 2012 Elsevier Ltd. All rights reserved.

Improvement of the enzymatic hydrolysis of furfural residues by pretreatment with combined green liquor and ethanol organosolv.

PubMed

Yu, Hailong; Xing, Yang; Lei, Fuhou; Liu, Zhiping; Liu, Zuguang; Jiang, Jianxin

2014-09-01

Furfural residues (FRs) were pretreated with ethanol and a green liquor (GL) catalyst to produce fermentable sugar. Anthraquinone (AQ) was used as an auxiliary reagent to improve delignification and reduce cellulose decomposition. The results showed that 42.7% of lignin was removed and 96.5% of cellulose was recovered from substrates pretreated with 1.0 mL GL/g of dry substrate and 0.4% (w/w) AQ at 140°C for 1h. Compared with raw material, ethanol-GL pretreatment of FRs increased the glucose yield from 69.0% to 85.9% after 96 h hydrolysis with 18 FPU/g-cellulose for cellulase, 27 CBU/g-cellulose for β-glucosidase. The Brauner-Emmett-Teller surface area was reduced during pretreatment, which did not inhibit the enzymatic hydrolysis. Owing to the reduced surface area, the unproductive binding of cellulase to lignin was decreased, thus improving the enzymatic hydrolysis. The degree of polymerization of cellulose from FRs was too low to be a key factor for improving enzymatic hydrolysis. Copyright © 2014 Elsevier Ltd. All rights reserved.

Improving enzymatic hydrolysis efficiency of wheat straw through sequential autohydrolysis and alkaline post-extraction.

PubMed

Wu, Xinxing; Huang, Chen; Zhai, Shengcheng; Liang, Chen; Huang, Caoxing; Lai, Chenhuan; Yong, Qiang

2018-03-01

In this work, a two-step pretreatment process of wheat straw was established by combining autohydrolysis pretreatment and alkaline post-extraction. The results showed that employing alkaline post-extraction to autohydrolyzed wheat straw could significantly improve its enzymatic hydrolysis efficiency from 36.0% to 83.7%. Alkaline post-extraction lead to the changes of the structure characteristics of autohydrolyzed wheat straw. Associations between enzymatic hydrolysis efficiency and structure characteristics were also studied. The results showed that the factors of structure characteristics such as delignification, xylan removal yield, crystallinity, accessibility and hydrophobicity are positively related to enzymatic hydrolysis efficiency within a certain range for alkaline post-extracted wheat straw. The results demonstrated that autohydrolysis coupled with alkaline post-extraction is an effective and promising method to gain fermentable sugars from biomass. Copyright © 2017 Elsevier Ltd. All rights reserved.

Pretreatment of corn stover using wet oxidation to enhance enzymatic digestibility.

PubMed

Varga, Eniko; Schmidt, Anette S; Réczey, Kati; Thomsen, Anne Belinda

2003-01-01

Corn stover is an abundant, promising raw material for fuel ethanol production. Although it has a high cellulose content, without pretreatment it resists enzymatic hydrolysis, like most lignocellulosic materials. Wet oxidation (water, oxygen, mild alkali or acid, elevated temperature and pressure) was investigated to enhance the enzymatic digestibility of corn stover. Six different combinations of reaction temperature, time, and pH were applied. The best conditions (60 g/L of corn stover, 195 degrees C, 15 min, 12 bar O2, 2 g/L of Na2CO3) increased the enzymatic conversion of corn stover four times, compared to untreated material. Under these conditions 60% of hemicellulose and 30% of lignin were solubilized, whereas 90% of cellulose remained in the solid fraction. After 24-h hydrolysis at 50 degrees C using 25 filter paper units (FPU)/g of drymatter (DM) biomass, the achieved conversion of cellulose to glucose was about 85%. Decreasing the hydrolysis temperature to 40 degrees C increased hydrolysis time from 24 to 72 h. Decreasing the enzyme loading to 5 FPU/g of DM biomass slightly decreased the enzymatic conversion from 83.4 to 71%. Thus, enzyme loading can be reduced without significantly affecting the efficiency of hydrolysis, an important economical aspect.

Kinetic study of the thermal hydrolysis of Agave salmiana for mezcal production.

PubMed

Garcia-Soto, M J; Jimenez-Islas, H; Navarrete-Bolanos, J L; Rico-Martinez, R; Miranda-Lopez, R; Botello-Alvarez, J E

2011-07-13

The kinetics of the thermal hydrolysis of the fructans of Agave salmiana were determined during the cooking step of mezcal production in a pilot autoclave. Thermal hydrolysis was achieved at different temperatures and cooking times, ranging from 96 to 116 °C and from 20 to 80 h. A simple kinetic model of the depolymerization of fructans to monomers and other reducing sugars and of the degradation of reducing sugars to furans [principally 5-(hydroxymethyl)furfural, HMF] was developed. From this model, the rate constants of the reactions were calculated, as well as the pre-exponential factors and activation energies of the Arrhenius equation. The model was found to fit the experimental data well. The tradeoff between a maximum fructan hydrolysis and a critical furan concentration in allowing for the best ethanol yield during fermentation was investigated. The results indicated that the thermal hydrolysis of agave was optimal, from the point of view of ethanol yield in the ensuing fermentation, in the temperature range of 106-116 °C and the cooking range time of 6-14 h. The optimal conditions corresponded to a fructan hydrolysis of 80%, producing syrups with furan and reducing sugar concentrations of 1 ± 0.1 and 110 ± 10 g/L, respectively.

Modeling evolution of hydrogen bonding and stabilization of transition states in the process of cocaine hydrolysis catalyzed by human butyrylcholinesterase.

PubMed

Gao, Daquan; Zhan, Chang-Guo

2006-01-01

Molecular dynamics (MD) simulations and quantum mechanical/molecular mechanical (QM/MM) calculations were performed on the prereactive enzyme-substrate complex, transition states, intermediates, and product involved in the process of human butyrylcholinesterase (BChE)-catalyzed hydrolysis of (-)-cocaine. The computational results consistently reveal a unique role of the oxyanion hole (consisting of G116, G117, and A199) in BChE-catalyzed hydrolysis of cocaine, compared to acetylcholinesterase (AChE)-catalyzed hydrolysis of acetylcholine. During BChE-catalyzed hydrolysis of cocaine, only G117 has a hydrogen bond with the carbonyl oxygen (O31) of the cocaine benzoyl ester in the prereactive BChE-cocaine complex, and the NH groups of G117 and A199 are hydrogen-bonded with O31 of cocaine in all of the transition states and intermediates. Surprisingly, the NH hydrogen of G116 forms an unexpected hydrogen bond with the carboxyl group of E197 side chain and, therefore, is not available to form a hydrogen bond with O31 of cocaine in the acylation. The NH hydrogen of G116 is only partially available to form a weak hydrogen bond with O31 of cocaine in some structures involved in the deacylation. The change of the estimated hydrogen-bonding energy between the oxyanion hole and O31 of cocaine during the reaction process demonstrates how the protein environment can affect the energy barrier for each step of the BChE-catalyzed hydrolysis of cocaine. These insights concerning the effects of the oxyanion hole on the energy barriers provide valuable clues on how to rationally design BChE mutants with a higher catalytic activity for the hydrolysis of (-)-cocaine. 2005 Wiley-Liss, Inc.

Modeling Evolution of Hydrogen Bonding and Stabilization of Transition States in the Process of Cocaine Hydrolysis Catalyzed by Human Butyrylcholinesterase

PubMed Central

Gao, Daquan; Zhan, Chang-Guo

2010-01-01

Molecular dynamics (MD) simulations and quantum mechanical/molecular mechanical (QM/MM) calculations were performed on the prereactive enzyme-substrate complex, transition states, intermediates, and product involved in the process of human butyrylcholinesterase (BChE)-catalyzed hydrolysis of (−)-cocaine. The computational results consistently reveal a unique role of the oxyanion hole (consisting of G116, G117, and A199) in BChE-catalyzed hydrolysis of cocaine, as compared to acetylcholinesterase (AChE)-catalyzed hydrolysis of acetylcholine. During BChE-catalyzed hydrolysis of cocaine, only G117 has a hydrogen bond with the carbonyl oxygen (O31) of the cocaine benzoyl ester in the prereactive BChE-cocaine complex, and the NH groups of G117 and A199 are hydrogen-bonded with O31 of cocaine in all of the transition states and intermediates. Surprisingly, the NH hydrogen of G116 forms an unexpected hydrogen bond with the carboxyl group of E197 side chain and, therefore, is not available to form a hydrogen bond with O31 of cocaine in the acylation. The NH hydrogen of G116 is only partially available to form a weak hydrogen bond with O31 of cocaine in some structures involved in the deacylation. The change of the estimated hydrogen bonding energy between the oxyanion hole and O31 of cocaine during the reaction process demonstrates how the protein environment can affect the energy barrier for each step of the BChE-catalyzed hydrolysis of cocaine. These insights concerning the effects of the oxyanion hole on the energy barriers provide valuable clues on how to rationally design BChE mutants with a higher catalytic activity for the hydrolysis of (−)-cocaine. PMID:16288482

Natural genetic variability reduces recalcitrance in poplar

DOE PAGES

Bhagia, Samarthya; Muchero, Wellington; Kumar, Rajeev; ...

2016-05-20

Here, lignin content and structure are known to affect recalcitrance of lignocellulosic biomass to chemical/biochemical conversion. Previously, we identified rare Populus trichocarpa natural variants with significantly reduced lignin content. Because reduced lignin content may lower recalcitrance, 18 rare variants along with 4 comparators, and BESC standard Populus was analyzed for composition of structural carbohydrates and lignin. Sugar yields from these plants were measured at 5 process conditions: one for just enzymatic hydrolysis without pretreatment and four via our combined high-throughput hot water pretreatment and co-hydrolysis (HTPH) technique.

Volatility of ten priority pollutants from fortified avian toxicity test diets

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

McCrady, J.K.; Johnson, D.E.; Turner, L.W.

1985-01-01

With a few important exceptions, most pesticides have low volatility. However, a significant number of the industrial chemicals under the jurisdiction of TSCA have sufficient volatility to affect availability in fortified test diets and consequently the test results. Although extremely volatile chemicals might evaportate from test diets, there is insufficient data to indicate what levels of volatility are of concern. Volatility may be only one of a variety of factors influencing the fate of organic chemicals in test diets. Other mechanisms such as hydrolysis, adsorption, and photolysis should also be considered, but for many chemicals having significant vapor pressures, volatilitymore » is likely to be the major source of loss.« less

Deproteinated palm kernel cake-derived oligosaccharides: A preliminary study

NASA Astrophysics Data System (ADS)

Fan, Suet Pin; Chia, Chin Hua; Fang, Zhen; Zakaria, Sarani; Chee, Kah Leong

2014-09-01

Preliminary study on microwave-assisted hydrolysis of deproteinated palm kernel cake (DPKC) to produce oligosaccharides using succinic acid was performed. Three important factors, i.e., temperature, acid concentration and reaction time, were selected to carry out the hydrolysis processes. Results showed that the highest yield of DPKC-derived oligosaccharides can be obtained at a parameter 170 °C, 0.2 N SA and 20 min of reaction time.

Bioethanol Production By Utilizing Cassava Peels Waste Through Enzymatic And Microbiological Hydrolysis

NASA Astrophysics Data System (ADS)

Witantri, R. G.; Purwoko, T.; Sunarto; Mahajoeno, E.

2017-07-01

Cassava peels waste contains, cellulose which is quite high at 43.626%, this is a potential candidate as a raw for bioethanol production. The purpose of this study was to determine the performance of the enzymatic hydrolysis, microbiological (Effective microbe) and fermentation in cassava peel waste is known from the results of quantitative measurement of multiple ethanol parameters (DNS Test, pH, ethanol concentration). This research was carried out in stages, the first stage is hydrolysis with completely randomized design with single factor variation of the catalyst, consisting of three levels ie cellulase enzymes, multienzyme and effective microbial EM4. The second stage is fermentation with factorial randomized block design, consisting of three groups of variations of catalyst, and has two factors: variations of fermipan levels 1, 2, 3% and the duration of fermentation, 2,4,6 days. The parameters in the test is a reducing sugar, pH and concentration of ethanol. The results showed that variation of hydrolysis treatment, fermentation time, and fermipan levels has real effect on the fermentation process. On average the highest ethanol content obtained from the treatment with multienzyme addition, with the addition of 2% fermipan levels and on the 2nd day of fermentation that is equal to 3.76%.

Hyperthyroidism modifies ecto-nucleotidase activities in synaptosomes from hippocampus and cerebral cortex of rats in different phases of development.

PubMed

Bruno, Alessandra Nejar; Da Silva, Rosane Souza; Bonan, Carla Denise; Battastini, Ana Maria Oliveira; Barreto-chaves, Maria Luiza M; Sarkis, João José Freitas

2003-11-01

Here we investigate the possible effects of the hyperthyroidism on the hydrolysis of the ATP to adenosine in the synaptosomes of hippocampus, cerebral cortex and blood serum of rats in different developmental phases. Manifestations of hyperthyroidism include anxiety, nervousness, tachycardia, physical hyperactivity and weight loss amongst others. The thyroid hormones modulate a number of physiological functions in central nervous system, including development, function, expression of adenosine A(1) receptors and transport of neuromodulator adenosine. Thus, hyperthyroidism was induced in male Wistar rats (5-, 60-, 150- and 330-day old) by daily injections of L-thyroxine (T4) for 14 days. Nucleotide hydrolysis was decreased by about 14-52% in both hippocampus and cerebral cortex in 5 to 60-day-old rats. These changes were also observed in rat blood serum. In addition, in 11-month-old rats, inhibition of ADP and AMP hydrolysis persisted in the hippocampus, whereas, in cerebral cortex, an increase in AMP hydrolysis was detected. Thus, hyperthyroidism affects the extracellular nucleotides balance and adenosine production, interfering in neurotransmitter release, development and others physiological processes in different systems.

Joint action of ultrasonic and Fe³⁺ to improve selectivity of acid hydrolysis for microcrystalline cellulose.

PubMed

Li, Jinbao; Qiang, Dandan; Zhang, Meiyun; Xiu, Huijuan; Zhang, Xiangrong

2015-09-20

In this study, the combination of Fe(3+)/HCl and ultrasonic treatment was applied to selectively hydrolyze cellulose for the preparation of microcrystalline cellulose (MCC). It was found that the crystallinity and specific surface area of hydrocellulose samples were higher (78.92% and 2.23581 m(2)g(-1), respectively), compared with the method that only used Fe(3+)/HCl catalyst without ultrasonic treatment. Meanwhile, the hydrolysate can be extracted and reused for cellulose hydrolysis for three runs, which was effective in saving the dosage of chemicals and reducing the pollution of the environment without affecting the properties of hydrocellulose. Moreover, the increased concentration of total reducing sugar (TRS) after three runs may be used as a valuable source in biofuels production. The technology of cellulose hydrolysis, by retaining the crystalline region for MCC products while promoting hydrolysis of amorphous region for further utilization is of great novelty, which may prove valuable in converting biomass into chemicals and biofuels, environmentally and economically. Copyright © 2015 Elsevier Ltd. All rights reserved.

Deactivation of Cellulase at the Air-Liquid Interface Is the Main Cause of Incomplete Cellulose Conversion at Low Enzyme Loadings.

PubMed

Bhagia, Samarthya; Dhir, Rachna; Kumar, Rajeev; Wyman, Charles E

2018-01-22

Amphiphilic additives such as bovine serum albumin (BSA) and Tween have been used to improve cellulose hydrolysis by cellulases. However, there has been a lack of clarity to explain their mechanism of action in enzymatic hydrolysis of pure or low-lignin cellulosic substrates. In this work, a commercial Trichoderma reesei enzyme preparation and the amphiphilic additives BSA and Tween 20 were applied for hydrolysis of pure Avicel cellulose. The results showed that these additives only had large effects on cellulose conversion at low enzyme to substrate ratios when the reaction flasks were shaken. Furthermore, changes in the air-liquid interfacial area profoundly affected cellulose conversion, but surfactants reduced or prevented cellulase deactivation at the air-liquid interface. Not shaking the flasks or adding low amounts of surfactant resulted in near theoretical cellulose conversion at low enzyme loadings given enough reaction time. At low enzyme loadings, hydrolysis of cellulose in lignocellulosic biomass with low lignin content suffered from enhanced enzyme deactivation at the air-liquid interface.

Measurement of the rates of acetyl-CoA hydrolysis and synthesis from acetate in rat hepatocytes and the role of these fluxes in substrate cycling.

PubMed Central

Crabtree, B; Gordon, M J; Christie, S L

1990-01-01

1. Acetyl-CoA hydrolysis, acetyl-CoA synthesis from acetate and several related fluxes were measured in rat hepatocytes. 2. In contrast with acetyl-CoA hydrolysis, most of the acetyl-CoA synthesis from acetate occurred in the mitochondria. 3. Acetyl-CoA hydrolysis was not significantly affected by 24 h starvation or (-)-hydroxycitrate. 4. In the cytoplasm there was a net flux of acetyl-CoA to acetate, and substrate cycling between acetate and acetyl-CoA in this compartment was very low, accounting for less than 0.1% of the total heat production by the animal. 5. A larger cycle, involving mitochondrial and cytoplasmic acetate and acetyl-CoA, may operate in fed animals, but would account for only approx 1% of total heat production. 6. It is proposed that the opposing fluxes of mitochondrial acetate utilization and cytoplasmic net acetate production may provide sensitivity, feedback and buffering, even when these fluxes are not linked to form a conventional substrate cycle. PMID:2396982

Using mechanisms of hydrolysis and sorption to reduce siloxanes occurrence in biogas of anaerobic sludge digesters.

PubMed

Gatidou, Georgia; Arvaniti, Olga S; Stasinakis, Athanasios S; Thomaidis, Nikolaos S; Andersen, Henrik R

2016-12-01

Hydrolysis of hexamethylcyclotrisiloxane (D3), octamethylcyclotetrasiloxane (D4), decamethylcyclopentasiloxane (D5), dodecamethylcyclohexasiloxane (D6) and dodecamethylcyclohexasilane (D6_silane) and their sorption to digested sludge was studied in batch experiments. Hydrolysis was affected by the type of the compound and the applied temperature, while the relevant half-life values ranged between 0.07±0.01d (D3, 55°C) and 48.4±17.1d (D6_silane, 4°C). D5 showed the greatest affinity for sorption to digested sludge (logK d : 3.84±3.42), the lowest LogK d value was found for D3 (1.46±0.95). Prediction of investigated compounds' fate in a single-stage anaerobic digestion system indicated that volatilization seems to be the major fate in both mesophilic and thermophilic conditions. The addition of a pre-digester with 3d retention time would significantly decrease the expected concentrations of all siloxanes in biogas, enhancing their removal through hydrolysis and sorption to sludge. Copyright © 2016 Elsevier Ltd. All rights reserved.

Mechanistic kinetic models of enzymatic cellulose hydrolysis-A review.

PubMed

Jeoh, Tina; Cardona, Maria J; Karuna, Nardrapee; Mudinoor, Akshata R; Nill, Jennifer

2017-07-01

Bioconversion of lignocellulose forms the basis for renewable, advanced biofuels, and bioproducts. Mechanisms of hydrolysis of cellulose by cellulases have been actively studied for nearly 70 years with significant gains in understanding of the cellulolytic enzymes. Yet, a full mechanistic understanding of the hydrolysis reaction has been elusive. We present a review to highlight new insights gained since the most recent comprehensive review of cellulose hydrolysis kinetic models by Bansal et al. (2009) Biotechnol Adv 27:833-848. Recent models have taken a two-pronged approach to tackle the challenge of modeling the complex heterogeneous reaction-an enzyme-centric modeling approach centered on the molecularity of the cellulase-cellulose interactions to examine rate limiting elementary steps and a substrate-centric modeling approach aimed at capturing the limiting property of the insoluble cellulose substrate. Collectively, modeling results suggest that at the molecular-scale, how rapidly cellulases can bind productively (complexation) and release from cellulose (decomplexation) is limiting, while the overall hydrolysis rate is largely insensitive to the catalytic rate constant. The surface area of the insoluble substrate and the degrees of polymerization of the cellulose molecules in the reaction both limit initial hydrolysis rates only. Neither enzyme-centric models nor substrate-centric models can consistently capture hydrolysis time course at extended reaction times. Thus, questions of the true reaction limiting factors at extended reaction times and the role of complexation and decomplexation in rate limitation remain unresolved. Biotechnol. Bioeng. 2017;114: 1369-1385. © 2017 Wiley Periodicals, Inc. © 2017 Wiley Periodicals, Inc.

Identifying and overcoming the effect of mass transfer limitation on decreased yield in enzymatic hydrolysis of lignocellulose at high solid concentrations.

PubMed

Du, Jian; Cao, Yuan; Liu, Guodong; Zhao, Jian; Li, Xuezhi; Qu, Yinbo

2017-04-01

Cellulose conversion decreases significantly with increasing solid concentrations during enzymatic hydrolysis of insoluble lignocellulosic materials. Here, mass transfer limitation was identified as a significant determining factor of this decrease by studying the hydrolysis of delignified corncob residue in shake flask, the most used reaction vessel in bench scale. Two mass transfer efficiency-related factors, mixing speed and flask filling, were shown to correlate closely with cellulose conversion at solid loadings higher than 15% DM. The role of substrate characteristics in mass transfer performance was also significant, which was revealed by the saccharification of two corn stover substrates with different pretreatment methods at the same solid loading. Several approaches including premix, fed-batch operation, and particularly the use of horizontal rotating reactor were shown to be valid in facilitating cellulose conversion via improving mass transfer efficiency at solid concentrations higher than 15% DM. Copyright © 2017 Elsevier Ltd. All rights reserved.

Mutations in Elongation Factor Ef-1α Affect the Frequency of Frameshifting and Amino Acid Misincorporation in Saccharomyces Cerevisiae

PubMed Central

Sandbaken, M. G.; Culbertson, M. R.

1988-01-01

A mutational analysis of the eukaryotic elongation factor EF-1α indicates that this protein functions to limit the frequency of errors during genetic code translation. We found that both amino acid misincorporation and reading frame errors are controlled by EF-1α. In order to examine the function of this protein, the TEF2 gene, which encodes EF-1α in Saccharomyces cerevisiae, was mutagenized in vitro with hydroxylamine. Sixteen independent TEF2 alleles were isolated by their ability to suppress frameshift mutations. DNA sequence analysis identified eight different sites in the EF-1α protein that elevate the frequency of mistranslation when mutated. These sites are located in two different regions of the protein. Amino acid substitutions located in or near the GTP-binding and hydrolysis domain of the protein cause suppression of frameshift and nonsense mutations. These mutations may effect mistranslation by altering the binding or hydrolysis of GTP. Amino acid substitutions located adjacent to a putative aminoacyl-tRNA binding region also suppress frameshift and nonsense mutations. These mutations may alter the binding of aminoacyl-tRNA by EF-1α. The identification of frameshift and nonsense suppressor mutations in EF-1α indicates a role for this protein in limiting amino acid misincorporation and reading frame errors. We suggest that these types of errors are controlled by a common mechanism or closely related mechanisms. PMID:3066688

Substitution of Asp-309 by Asn in the Arg-Asp-Pro (RDP) motif of Acetobacter diazotrophicus levansucrase affects sucrose hydrolysis, but not enzyme specificity.

PubMed Central

Batista, F R; Hernández, L; Fernández, J R; Arrieta, J; Menéndez, C; Gómez, R; Támbara, Y; Pons, T

1999-01-01

beta-Fructofuranosidases share a conserved aspartic acid-containing motif (Arg-Asp-Pro; RDP) which is absent from alpha-glucopyranosidases. The role of Asp-309 located in the RDP motif of levansucrase (EC 2.4.1.10) from Acetobacter diazotrophicus SRT4 was studied by site-directed mutagenesis. Substitution of Asp-309 by Asn did not affect enzyme secretion. The kcat of the mutant levansucrase was reduced 75-fold, but its Km was similar to that of the wild-type enzyme, indicating that Asp-309 plays a major role in catalysis. The two levansucrases showed optimal activity at pH 5.0 and yielded similar product profiles. Thus the mutation D309N affected the efficiency of sucrose hydrolysis, but not the enzyme specificity. Since the RDP motif is present in a conserved position in fructosyltransferases, invertases, levanases, inulinases and sucrose-6-phosphate hydrolases, it is likely to have a common functional role in beta-fructofuranosidases. PMID:9895294

Biomass enzymatic saccharification is determined by the non-KOH-extractable wall polymer features that predominately affect cellulose crystallinity in corn.

PubMed

Jia, Jun; Yu, Bin; Wu, Leiming; Wang, Hongwu; Wu, Zhiliang; Li, Ming; Huang, Pengyan; Feng, Shengqiu; Chen, Peng; Zheng, Yonglian; Peng, Liangcai

2014-01-01

Corn is a major food crop with enormous biomass residues for biofuel production. Due to cell wall recalcitrance, it becomes essential to identify the key factors of lignocellulose on biomass saccharification. In this study, we examined total 40 corn accessions that displayed a diverse cell wall composition. Correlation analysis showed that cellulose and lignin levels negatively affected biomass digestibility after NaOH pretreatments at p<0.05 & 0.01, but hemicelluloses did not show any significant impact on hexoses yields. Comparative analysis of five standard pairs of corn samples indicated that cellulose and lignin should not be the major factors on biomass saccharification after pretreatments with NaOH and H2SO4 at three concentrations. Notably, despite that the non-KOH-extractable residues covered 12%-23% hemicelluloses and lignin of total biomass, their wall polymer features exhibited the predominant effects on biomass enzymatic hydrolysis including Ara substitution degree of xylan (reverse Xyl/Ara) and S/G ratio of lignin. Furthermore, the non-KOH-extractable polymer features could significantly affect lignocellulose crystallinity at p<0.05, leading to a high biomass digestibility. Hence, this study could suggest an optimal approach for genetic modification of plant cell walls in bioenergy corn.

Biomass Enzymatic Saccharification Is Determined by the Non-KOH-Extractable Wall Polymer Features That Predominately Affect Cellulose Crystallinity in Corn

PubMed Central

Wu, Leiming; Wang, Hongwu; Wu, Zhiliang; Li, Ming; Huang, Pengyan; Feng, Shengqiu; Chen, Peng; Zheng, Yonglian; Peng, Liangcai

2014-01-01

Corn is a major food crop with enormous biomass residues for biofuel production. Due to cell wall recalcitrance, it becomes essential to identify the key factors of lignocellulose on biomass saccharification. In this study, we examined total 40 corn accessions that displayed a diverse cell wall composition. Correlation analysis showed that cellulose and lignin levels negatively affected biomass digestibility after NaOH pretreatments at p<0.05 & 0.01, but hemicelluloses did not show any significant impact on hexoses yields. Comparative analysis of five standard pairs of corn samples indicated that cellulose and lignin should not be the major factors on biomass saccharification after pretreatments with NaOH and H2SO4 at three concentrations. Notably, despite that the non-KOH-extractable residues covered 12%–23% hemicelluloses and lignin of total biomass, their wall polymer features exhibited the predominant effects on biomass enzymatic hydrolysis including Ara substitution degree of xylan (reverse Xyl/Ara) and S/G ratio of lignin. Furthermore, the non-KOH-extractable polymer features could significantly affect lignocellulose crystallinity at p<0.05, leading to a high biomass digestibility. Hence, this study could suggest an optimal approach for genetic modification of plant cell walls in bioenergy corn. PMID:25251456

Influence of lactose hydrolysis on galacto-oligosaccharides, lactose, volatile profile and physicochemical parameters of different yogurt varieties.

PubMed

Vénica, Claudia I; Wolf, Irma V; Bergamini, Carina V; Perotti, María C

2016-12-01

Different types of reduced-lactose yogurt, obtained by lactose hydrolysis using β-galactosidase enzyme, are commercially available. The breakdown of lactose modifies the carbohydrate profile, including the production of prebiotic galacto-oligosaccharides (GOS), which could affect the survival and activity of starter and probiotic cultures and the parameters of yogurt quality. The extension of these changes is dependent on the yogurt matrix composition. This study aimed to evaluate the influence of lactose hydrolysis on GOS, lactose, volatile profile and physicochemical parameters of different yogurt varieties during storage. The presence of β-galactosidase enzyme did not affect either the global composition or the survival of cultures. Overall, the hydrolyzed products had lower acidity than traditional ones. GOS were found at similar levels in fresh hydrolyzed yogurts, whereas in traditional yogurts they were not detected. The proportion of ketones, acids and aldehydes seems to be more dependent on yogurt variety than on addition of the enzyme. Likewise, the storage period affected the volatile fraction to different degree; the increase in acid compounds was more pronounced in hydrolyzed than in traditional yogurts. This work shows that it is possible to obtain different varieties of reduced-lactose yogurt, some of them with additional benefits to health such as reduced fat, reduced calories, added with probiotic/inulin and enriched in GOS, with similar characteristics to traditional products. © 2016 Society of Chemical Industry. © 2016 Society of Chemical Industry.

Rate of hydrolysis and degradation of the cyanogenic glycoside - dhurrin - in soil.

PubMed

Johansen, Henrik; Rasmussen, Lars Holm; Olsen, Carl Erik; Bruun Hansen, Hans Christian

2007-02-01

Cyanogenic glycosides are common plant toxins. Toxic hydrogen cyanide originating from cyanogenic glycosides may affect soil processes and water quality. In this study, hydrolysis, degradation and sorption of dhurrin (4-hydroxymandelonitrile-beta-d-glucoside) produced by sorghum has been studied in order to assess its fate in soil. The log K(ow) of dhurrin was -1.18+/-0.08 (22 degrees C). Hydrolysis was a first-order reaction with respect to dhurrin and hydroxyl ion concentrations. Half lives ranged from 1.2h (pH 8.6; 25 degrees C) to 530d (pH 4; 25 degrees C). The activation energy of hydrolysis was 112+9kJ. At pH 5.8 and room temperature, addition of humic acids (50gl(-1)) increased the rate of hydrolysis tenfold, while addition of kaolinite or goethite (100-250gl(-1)) both decreased the rate considerably. No significant sorption to soil components could be observed. The degradation rates of dhurrin in top and subsoils of Oxisols, Ultisols, Alfisols and Mollisols were studied at 22 degrees C (25mgl(-1), soil:liquid 1:1 (w:V), pH 3.8-8.1). Half-lives were 0.25-2h for topsoils, and 5-288h in subsoils. Hydrolysis in solution explained up to 45% of the degradation in subsoils whereas the contribution in topsoils was less than 14%, indicating the importance of enzymatic degradation processes. The highest risk of dhurrin leaching will take place when the soil is a low activity acid shallow soil with low content of clay minerals, iron oxides and humic acids.

Effect of endogenous and exogenous enzymatic treatment of green vanilla beans on extraction of vanillin and main aromatic compounds.

PubMed

Pardío, Violeta T; Flores, Argel; López, Karla M; Martínez, David I; Márquez, Ofelia; Waliszewski, Krzysztof N

2018-06-01

Endogenous and exogenous enzymatic hydrolysis carried out to obtain vanilla extracts with higher concentrations of vanillin using green vanilla beans. Sequences initiated with freezing of green vanilla beans at - 1 °C for 24 h, followed by endogenous hydrolysis under optimal β-glucosidase activity at 4.2 and 35 °C for 96 h, exogenous hydrolysis with Crystalzyme PML-MX at pH 5.0 and 40 °C for 72 h, and ethanol extraction at 40% (v v -1 ) for 30 days. In the proposed method, 200 g of fresh green vanilla beans with 84% moisture (32 g dry base) were used to obtain a liter of single fold vanilla extract. This method allowed the release of 82.57% of the theoretically available vanillin from its precursor glucovanillin with 5.78 g 100 g -1 green vanilla beans (dry base). Vanillic acid, p -hydroxybenzaldehyde and vanillyl alcohol were also released and found in commercial and enzymatic extracts. Glucovanillin was detected in commercial and traditional extracts but was absent in enzymatic extracts, indicating incomplete hydrolysis during the curing process. An in vitro assay was conducted to determine if the presence of peroxidase during hydrolysis might affect overall vanillin concentration. Results showed that POD can use vanillin as a substrate under conditions similar to those in which hydrolysis was conducted (pH 5.0 and 50 °C), possibly explaining why vanillin concentration was not complete at the end of the process.

Enzymatic hydrolysis of Grass Carp fish skin hydrolysates able to promote the proliferation of Streptococcus thermophilus.

PubMed

Wang, Xiao-Nan; Qin, Mei; Feng, Yu-Ying; Chen, Jian-Kang; Song, Yi-Shan

2017-09-01

The promotion effect on proliferation of Streptococcus thermophilus by enzymatic hydrolysates of aquatic products was firstly studied. The effect of influencing factors of the hydrolysis on the growth of S. thermophilus was investigated. Grass Carp fish skin was hydrolysed to peptides by enzymatic hydrolysis using protease ProteAX, and for the S. thermophilus growth, the optimal enzymatic hydrolysis conditions were temperature of 60 °C, initial pH of 9.0, enzyme concentration of 10 g kg -1 , hydrolysis time of 80 min, and ratio of material to liquid of 1:2. The Grass Carp fish skin hydrolysate (GCFSH) prepared under the optimum conditions was fractionated to five fragments (GCFSH 1, GCFSH 2, GCFSH 3, GCFSH 4, GCFSH 5) according to molecular weight sizes, in which the fragments GCFSH 4 and GCFSH 5, with molecular weights of less than 1000 Da, significantly promoted the growth of S. thermophilus. The hydrolysis process of Grass Carp fish skin can be simplified, and the peptides with molecular weights below 1000 Da in the hydrolysates are the best nitrogen source for proliferation of S. thermophilus. This work can provide a fundamental theoretical basis for the production of multi-component functional foods, especially in milk drinks or yogurt. © 2017 Society of Chemical Industry. © 2017 Society of Chemical Industry.

Exploring crystalline-structural variations of cellulose during alkaline pretreatment for enhanced enzymatic hydrolysis.

PubMed

Ling, Zhe; Chen, Sheng; Zhang, Xun; Xu, Feng

2017-01-01

The study aimed to explore the crystallinity and crystalline structure of alkaline pretreated cellulose. The enzymatic hydrolysis followed by pretreatment was conducted for measuring the efficiency of sugar conversion. For cellulose Iβ dominated samples, alkaline pretreatment (<8wt%) caused increased cellulose crystallinity and depolymerized hemicelluloses, that were superimposed to affect the enzymatic conversion to glucose. Varying crystallite sizes and lattice spacings indicated the separation of cellulose crystals during mercerization (8-12wt% NaOH). Completion of mercerization was proved under higher alkaline concentration (14-18wt% NaOH), leading to distortion of crystalline cellulose to some extent. Cellulose II crystallinity showed a stimulative impact on enzymatic hydrolysis due to the weakened hydrophobic interactions within cellulose chains. The current study may provide innovative explanations for enhanced enzymatic digestibility of alkaline pretreated lignocellulosic materials. Copyright © 2016 Elsevier Ltd. All rights reserved.

Constant Enthalpy Change Value during Pyrophosphate Hydrolysis within the Physiological Limits of NaCl*

PubMed Central

Wakai, Satoshi; Kidokoro, Shun-ichi; Masaki, Kazuo; Nakasone, Kaoru; Sambongi, Yoshihiro

2013-01-01

A decrease in water activity was thought to result in smaller enthalpy change values during PPi hydrolysis, indicating the importance of solvation for the reaction. However, the physiological significance of this phenomenon is unknown. Here, we combined biochemistry and calorimetry to solve this problem using NaCl, a physiologically occurring water activity-reducing reagent. The pyrophosphatase activities of extremely halophilic Haloarcula japonica, which can grow at ∼4 m NaCl, and non-halophilic Escherichia coli and Saccharomyces cerevisiae were maximal at 2.0 and 0.1 m NaCl, respectively. Thus, halophilic and non-halophilic pyrophosphatases exhibit distinct maximal activities at different NaCl concentration ranges. Upon calorimetry, the same exothermic enthalpy change of −35 kJ/mol was obtained for the halophile and non-halophiles at 1.5–4.0 and 0.1–2.0 m NaCl, respectively. These results show that solvation changes caused by up to 4.0 m NaCl (water activity of ∼0.84) do not affect the enthalpy change in PPi hydrolysis. It has been postulated that PPi is an ATP analog, having a so-called high energy phosphate bond, and that the hydrolysis of both compounds is enthalpically driven. Therefore, our results indicate that the hydrolysis of high energy phosphate compounds, which are responsible for biological energy conversion, is enthalpically driven within the physiological limits of NaCl. PMID:23965994

Hydrogen generation from Al-NiCl2/NaBH4 mixture affected by lanthanum metal.

PubMed

Sun, Wen Qiang; Fan, Mei-Qiang; Fei, Yong; Pan, Hua; Wang, Liang Liang; Yao, Jun

2012-01-01

The effect of La on Al/NaBH(4) hydrolysis was elaborated in the present paper. Hydrogen generation amount increases but hydrogen generation rate decreases with La content increasing. There is an optimized composition that Al-15 wt% La-5 wt% NiCl(2)/NaBH(4) mixture (Al-15 wt% La-5 wt% NiCl(2)/NaBH(4) weight ratio, 1 : 3) has 126 mL g(-1 )min(-1) maximum hydrogen generation rate and 1764 mL g(-1) hydrogen generation amount within 60 min. The efficiency is 88%. Combined with NiCl(2), La has great effect on NaBH(4) hydrolysis but has little effect on Al hydrolysis. Increasing La content is helpful to decrease the particle size of Al-La-NiCl(2) in the milling process, which induces that the hydrolysis byproduct Ni(2)B is highly distributed into Al(OH)(3) and the catalytic reactivity of Ni(2)B/Al(OH)(3) is increased therefore. But hydrolysis byproduct La(OH)(3) deposits on Al surface and leads to some side effect. The Al-La-NiCl(2)/NaBH(4) mixture has good stability in low temperature and its hydrolytic performance can be improved with increasing global temperature. Therefore, the mixture has good safety and can be applied as on board hydrogen generation material.

PDE7B is involved in nandrolone decanoate hydrolysis in liver cytosol and its transcription is up-regulated by androgens in HepG2.

PubMed

Strahm, Emmanuel; Rane, Anders; Ekström, Lena

2014-01-01

Most androgenic drugs are available as esters for a prolonged depot action. However, the enzymes involved in the hydrolysis of the esters have not been identified. There is one study indicating that PDE7B may be involved in the activation of testosterone enanthate. The aims are to identify the cellular compartments where the hydrolysis of testosterone enanthate and nandrolone decanoate occurs, and to investigate the involvement of PDE7B in the activation. We also determined if testosterone and nandrolone affect the expression of the PDE7B gene. The hydrolysis studies were performed in isolated human liver cytosolic and microsomal preparations with and without specific PDE7B inhibitor. The gene expression was studied in human hepatoma cells (HepG2) exposed to testosterone and nandrolone. We show that PDE7B serves as a catalyst of the hydrolysis of testosterone enanthate and nandrolone decanoate in liver cytosol. The gene expression of PDE7B was significantly induced 3- and 5- fold after 2 h exposure to 1 μM testosterone enanthate and nandrolone decanoate, respectively. These results show that PDE7B is involved in the activation of esterified nandrolone and testosterone and that the gene expression of PDE7B is induced by supra-physiological concentrations of androgenic drugs.

Improving the performance of enzymes in hydrolysis of high solids paper pulp derived from MSW.

PubMed

Puri, Dhivya J; Heaven, Sonia; Banks, Charles J

2013-01-01

The research aimed to improve the overall conversion efficiency of the CTec® family of enzymes by identifying factors that lead to inhibition and seeking methods to overcome these through process modification and manipulation. The starting material was pulp derived from municipal solid waste and processed in an industrial-scale washing plant. Analysis of the pulp by acid hydrolysis showed a ratio of 55 : 12 : 6 : 24 : 3 of glucan : xylan : araban/galactan/mannan : lignin : ash. At high total solids content (>18.5% TS) single-stage enzyme hydrolysis gave a maximum glucan conversion of 68%. It was found that two-stage hydrolysis could give higher conversion if sugar inhibition was removed by an intermediate fermentation step between hydrolysis stages. This, however, was not as effective as direct removal of the sugar products, including xylose, by washing of the residual pulp at pH 5. This improved the water availability and allowed reactivation of the pulp-bound enzymes. Inhibition of enzyme activity could further be alleviated by replenishment of β-glucosidase which was shown to be removed during the wash step. The two-stage hydrolysis process developed could give an overall glucan conversion of 88%, with an average glucose concentration close to 8% in 4 days, thus providing an ideal starting point for ethanol fermentation with a likely yield of 4 wt%. This is a significant improvement over a single-step process. This hydrolysis configuration also provides the potential to recover the sugars associated with residual solids which are diluted when washing hydrolysed pulp.

Determination of alcohol sulfates in wastewater treatment plant influents and effluents by gas chromatography-mass spectrometry.

PubMed

Fernández-Ramos, C; Ballesteros, O; Blanc, R; Zafra-Gómez, A; Jiménez-Díaz, I; Navalón, A; Vílchez, J L

2012-08-30

In the present paper, we developed an accurate method for the analysis of alcohol sulfates (AS) in wastewater samples from wastewater treatment plant (WWTP) influents and effluents. Although many methodologies have been published in the literature concerning the study of anionic surfactants in environmental samples, at present, the number of analytical methodologies that focus in the determination of AS by gas chromatography in the different environmental compartments is limited. The reason for this is that gas chromatography-mass spectrometry (GC-MS) technique requires a previous hydrolysis reaction followed by derivatization reactions. In the present work, we proposed a new procedure in which the hydrolysis and derivatization reactions take place in one single step and AS are directly converted to trimethylsilyl derivatives. The main factors affecting solid-phase extraction (SPE), hydrolysis/derivatization and GC-MS procedures were accurately optimised. Quantification of the target compounds was performed by using GC-MS in selected ion monitoring (SIM) mode. The limits of detection (LOD) obtained ranged from 0.2 to 0.3 μg L(-1), and limits of quantification (LOQ) from 0.5 to 1.0 μg L(-1), while inter- and intra-day variability was under 5%. A recovery assay was also carried out. Recovery rates for homologues in spiked samples ranged from 96 to 103%. The proposed method was successfully applied for the determination of anionic surfactants in wastewater samples from one WWTP located in Granada (Spain). Concentration levels for the homologues up to 39.4 μg L(-1) in influent and up to 8.1 μg L(-1) in effluent wastewater samples. Copyright © 2012 Elsevier B.V. All rights reserved.

Role of a ribosomal RNA phosphate oxygen during the EF-G–triggered GTP hydrolysis

PubMed Central

Koch, Miriam; Flür, Sara; Kreutz, Christoph; Ennifar, Eric; Micura, Ronald; Polacek, Norbert

2015-01-01

Elongation factor-catalyzed GTP hydrolysis is a key reaction during the ribosomal elongation cycle. Recent crystal structures of G proteins, such as elongation factor G (EF-G) bound to the ribosome, as well as many biochemical studies, provide evidence that the direct interaction of translational GTPases (trGTPases) with the sarcin-ricin loop (SRL) of ribosomal RNA (rRNA) is pivotal for hydrolysis. However, the precise mechanism remains elusive and is intensively debated. Based on the close proximity of the phosphate oxygen of A2662 of the SRL to the supposedly catalytic histidine of EF-G (His87), we probed this interaction by an atomic mutagenesis approach. We individually replaced either of the two nonbridging phosphate oxygens at A2662 with a methyl group by the introduction of a methylphosphonate instead of the natural phosphate in fully functional, reconstituted bacterial ribosomes. Our major finding was that only one of the two resulting diastereomers, the SP methylphosphonate, was compatible with efficient GTPase activation on EF-G. The same trend was observed for a second trGTPase, namely EF4 (LepA). In addition, we provide evidence that the negative charge of the A2662 phosphate group must be retained for uncompromised activity in GTP hydrolysis. In summary, our data strongly corroborate that the nonbridging proSP phosphate oxygen at the A2662 of the SRL is critically involved in the activation of GTP hydrolysis. A mechanistic scenario is supported in which positioning of the catalytically active, protonated His87 through electrostatic interactions with the A2662 phosphate group and H-bond networks are key features of ribosome-triggered activation of trGTPases. PMID:25941362

Hydrolysis of short-chain phosphatidylcholines by bee venom phospholipase A2.

PubMed

Raykova, D; Blagoev, B

1986-01-01

In order to find out the aggregation state of the substrate, preferred by bee venom phospholipase A2 (EC 3.1.1.4), its action on short-chain phosphatidylcholines with two identical (C6-C10) fatty acids has been tested. The rate of hydrolysis as a function of acyl chain length showed a maximum at dioctanoylphosphatidylcholine. The effects of alcohols, NaCl and Triton X-100, which affect the aggregation state of phospholipids in water, were also studied. The addition of n-alcohol led to a significant inhibition of the hydrolysis of the substrates present in micellar form and activated the hydrolysis of substrates which form liposomes. The inhibitory effect increased with increasing length of the aliphatic carbon chain of the alcohol. Triton X-100 at low Triton/phospholipid molar ratios enhanced enzyme activity. These results do not agree with the accepted idea that bee venom phospholipase A2 hydrolyzes short-chain lecithins in their molecularly dispersed form and that micelles cannot act as substrates. The data indicate that short-chain lecithins in the aggregated state are hydrolyzed and that the requirements of bee venom phospholipase A2 for the aggregation state of the substrate are not strict.

Interactions of chlorphenesin and divalent metal ions with phosphodiesterase.

PubMed

Edelson, J; McMullen, J P

1976-09-01

Chlorphenesin inhibition of the hydrolysis of cyclic AMP by guinea-pig lung phosphodiesterase was reversed by the addition of exogenous magnesium ions. Chlorphenesin and theophylline inhibition of this enzyme was shown to be noncompetitive when the substrate concentration was low. Kinetic studies of the inhibition of beef heart phosphodiesterase by chlorphenesin and theophylline indicated that the substrate concentration was a factor in determining whether inhibition was competitive or noncompetitive. Calcium, cobalt and copper ions were inhibitory to guinea-pig lung phosphodiesterase. The inhibition due to chlorphenesin was partially reversed by low (40 mM or less) concentrations of barium ions; high concentrations of barium ions, or manganese ions, were inhibitory. The concentration of the divalent cation did not affect the type of inhibition that was observed.

Formation of ethyl ferulate from feruloylated oligosaccharide by transesterification of rice koji enzyme under sake mash conditions.

PubMed

Suzuki, Nobukazu; Ito, Toshihiko; Hiroshima, Kai; Tokiwano, Tetsuo; Hashizume, Katsumi

2016-03-01

Formation of ethyl ferulate (EF) and ferulic acid (FA) under sake mash conditions was studied using feruloylated oligosaccharide (FO), prepared from rice grains, as the substrate for rice koji enzyme. EF and FA were produced from FO over six times faster than from alkyl ferulates however, under the same ethanol concentration, only small differences were observed between the EF/FA ratios when either FO or methyl ferulate were used as substrates. Esterification and hydrolysis of FO or methyl ferulate showed similar pH dependencies and similar EF/FA ratios for each substrate in all of the pH ranges tested. Ethanol concentration clearly affected the EF/FA ratio; the ratio increased as ethanol concentration increased. Formation of EF and FA in the sake mash simulated rice digest was accelerated by addition of exogenous FO. These results indicated that supply of FO to sake mash is a crucial step for EF and FA formation, and ethanol is an influencing factor in the EF/FA ratio. The rice koji enzyme reaction suggested that EF and FA are formed through a common feruloylated enzyme intermediate complex by transesterification or hydrolysis, and these reactions occur competitively. Copyright © 2015 The Society for Biotechnology, Japan. Published by Elsevier B.V. All rights reserved.

Long chain fatty acids (LCFA) evolution for inhibition forecasting during anaerobic treatment of lipid-rich wastes: Case of milk-fed veal slaughterhouse waste.

PubMed

Rodríguez-Méndez, R; Le Bihan, Y; Béline, F; Lessard, P

2017-09-01

A detailed study of a solid slaughterhouse waste (SHW) anaerobic treatment is presented. The waste used in this study is rich in lipids and proteins residue. Long chain fatty acids (LCFA), coming from the hydrolysis of lipids were inhibitory to anaerobic processes at different degrees. Acetogenesis and methanogenesis processes were mainly affected by inhibition whereas disintegration and hydrolysis processes did not seem to be affected by high LCFA concentrations. Nevertheless, because of the high energy content, this kind of waste is very suitable for anaerobic digestion but strict control of operating conditions is required to prevent inhibition. For that, two inhibition indicators were identified in this study. Those two indicators, LCFA dynamics and LCFA/VS biomass ratio proved to be useful to predict and to estimate the process inhibition degree. Copyright © 2017 Elsevier Ltd. All rights reserved.

Dilute phosphoric acid-catalysed hydrolysis of municipal bio-waste wood shavings using autoclave parr reactor system.

PubMed

Orozco, Angela M; Al-Muhtaseb, Ala'a H; Albadarin, Ahmad B; Rooney, David; Walker, Gavin M; Ahmad, Mohammad N M

2011-10-01

The visibility of using municipal bio-waste, wood shavings, as a potential feedstock for ethanol production was investigated. Dilute acid hydrolysis of wood shavings with H₃PO₄ was undertaken in autoclave parr reactor. A combined severity factor (CSF) was used to integrate the effects of hydrolysis times, temperature and acid concentration into a single variable. Xylose concentration reached a maximum value of 17 g/100 g dry mass corresponding to a yield of 100% at the best identified conditions of 2.5 wt.% H₃PO₄, 175 °C and 10 min reaction time corresponding to a CSF of 1.9. However, for glucose, an average yield of 30% was obtained at 5 wt.% H₃PO₄, 200 °C and 10 min. Xylose production increased with increasing temperature and acid concentration, but its transformation to the degradation product furfural was also catalysed by those factors. The maximum furfural formed was 3 g/100 g dry mass, corresponding to the 24% yield. Copyright © 2011 Elsevier Ltd. All rights reserved.

Producing standard damaged DNA samples by heating: pitfalls and suggestions.

PubMed

Fattorini, Paolo; Marrubini, Giorgio; Bonin, Serena; Bertoglio, Barbara; Grignani, Pierangela; Recchia, Elisa; Pitacco, Paola; Procopio, Francesca; Cantoni, Carolina; Pajnič, Irena Zupanič; Sorçaburu-Cigliero, Solange; Previderè, Carlo

2018-05-15

Heat-mediated hydrolysis of DNA is a simple and inexpensive method for producing damaged samples in vitro. Despite heat-mediated DNA hydrolysis is being widely used in forensic and clinical validation procedures, the lack of standardized procedures makes it impossible to compare the intra and inter-laboratory outcomes of the damaging treatments. In this work, a systematic approach to heat induced DNA hydrolysis was performed at 70 °C for 0-18 h to test the role both of the hydrolysis buffer and of the experimental conditions. Specifically, a trial DNA sample, resuspended in three different media (ultrapure water, 0.1% DEPC-water and, respectively, TE) was treated both in Eppendorf tubes ("Protocol P") and in Eppendorf tubes provided with screwcaps ("Protocol S"). The results of these comparative tests were assessed by normalization of the qPCR results. DEPC-water increased the degradation of the samples up to about 100 times when compared to the ultrapure water. Conversely, the TE protected the DNA from degradation whose level was about 1700 times lower than in samples treated in ultrapure water. Even the employment of the "Protocol S" affected the level of degradation, by consistently increasing it (up to about 180 times in DEPC-water). Thus, this comparative approach showed that even seemingly apparently trivial and often underestimated parameters modify the degradation level up to 2-3 orders of magnitude. The chemical-physical reasons of these findings are discussed together with the role of potential factors such as enhanced reactivity of CO 2 , ROS, NO x and pressure, which are likely to be involved. Since the intra and inter-laboratory comparison of the outcomes of the hydrolytic procedure is the first step toward its standardization, the normalization of the qPCR data by the UV/qPCR ratio seems to be the simplest and most reliable way to allow this. Finally, the supplying (provided with the commercial qPCR kits) of a DNA sample whose degree of degradation is well documented could be helpful in ISO/IEC 17025 validation procedures and in proficiency testing. Copyright © 2018 Elsevier Inc. All rights reserved.

Deletion of a unique loop in the mycobacterial F-ATP synthase γ subunit sheds light on its inhibitory role in ATP hydrolysis-driven H(+) pumping.

PubMed

Hotra, Adam; Suter, Manuel; Biuković, Goran; Ragunathan, Priya; Kundu, Subhashri; Dick, Thomas; Grüber, Gerhard

2016-05-01

The F1 FO -ATP synthase is one of the enzymes that is essential to meet the energy requirement of both the proliferating aerobic and hypoxic dormant stages of the life cycle of mycobacteria. Most F-ATP synthases consume ATP in the α3 :β3 headpiece to drive the γ subunit, which couples ATP cleavage with proton pumping in the c ring of FO via the bottom of the γ subunit. ATPase-driven H(+) pumping is latent in mycobacteria. The presence of a unique 14 amino acid residue loop of the mycobacterial γ subunit has been described and aligned in close vicinity to the c-ring loop Priya R et al. (2013) J Bioenerg Biomembr 45, 121-129 Here, we used inverted membrane vesicles (IMVs) of fast-growing Mycobacterium smegmatis and a variety of covalent and non-covalent inhibitors to characterize the ATP hydrolysis activity of the F-ATP synthase inside IMVs. These vesicles formed a platform to investigate the function of the unique mycobaterial γ loop by deleting the respective loop-encoding sequence (γ166-179 ) in the genome of M. smegmatis. ATP hydrolysis-driven H(+) pumping was observed in IMVs containing the Δγ166-179 mutant protein but not for IMVs containing the wild-type F-ATP synthase. In addition, when compared to the wild-type enzyme, IMVs containing the Δγ166-179 mutant protein showed increased ATP cleavage and lower levels of ATP synthesis, demonstrating that the loop affects ATPase activity, ATPase-driven H(+) pumping and ATP synthesis. These results further indicate that the loop may affect coupling of ATP hydrolysis and synthesis in a different mode. © 2016 Federation of European Biochemical Societies.

Potential of hydrolysis of particulate COD in extended anaerobic conditions to enhance biological phosphorous removal.

PubMed

Jabari, P; Yuan, Q; Oleszkiewicz, J A

2016-11-01

The effect of anaerobic hydrolysis of particulate COD (pCOD) on biological phosphorous removal in extended anaerobic condition was investigated through (i) sequencing batch reactors (SBR)s with anaerobic hydraulic retention time (HRT) of 0.8, 2, and 4 h; (ii) batch tests using biomass from a full scale biological nutrient removal (BNR) plant; and (iii) activated sludge modeling (BioWin 4.1 simulation). The results from long-term SBRs operation showed that phosphorus removal was correlated to the ratio of filtered COD (FCOD) to total phosphorus (TP) in the influent. Under conditions with low FCOD/TP ratio (average of 20) in the influent, extending anaerobic HRT to 4 h in the presence of pCOD did not significantly improve overall phosphorous removal. During the period with high FCOD/TP ratio (average of 37) in the influent, all SBRs removed phosphorous completely, and the long anaerobic HRT did not have negative effect on overall phosphorous removal. The batch tests also showed that pCOD at different concentration during 4 h test did not affect the rate of anaerobic phosphorus release. The rate of anaerobic hydrolysis of pCOD was significantly low and extending the anaerobic HRT was ineffective. The simulation (BioWin 4.1) of SBRs with low influent FCOD/TP ratio showed that the default kinetics of anaerobic hydrolysis in ASM2d overestimated phosphorous removal in the SBRs (high anaerobic hydrolysis of pCOD). The default anaerobic hydrolysis rate in BioWin 4.1 (ten times lower) could produce similar phosphorous removal to that in the experiment. Results showed that the current kinetics of anaerobic hydrolysis in ASM2d could lead to considerable error in predicting phosphorus removal in processes with extended anaerobic HRT. Biotechnol. Bioeng. 2016;113: 2377-2385. © 2016 Wiley Periodicals, Inc. © 2016 Wiley Periodicals, Inc.

Effects of organosolv and ammonia pretreatments on lignin properties and its inhibition for enzymatic hydrolysis

DOE PAGES

Yoo, Chang Geun; Li, Mi; Meng, Xianzhi; ...

2017-04-05

Lignin offers structural support and protection for plant cell walls; but, it also contributes to biomass recalcitrance and the costs of biofuel production via the biological pathway. Organosolv and ammonia pretreatments have been developed to reduce biomass recalcitrance and improve sugar release performance during enzymatic hydrolysis. It is believed that lignin properties are related to its inhibition on enzymatic hydrolysis; therefore, understanding the characteristics of lignin is a key for effective biomass conversion to biofuels. In this study, an organosolv pretreatment using 60% ethanol with 1.25% H 2SO 4 significantly deconstructed poplar lignin and reduced its molecular weights due tomore » the cleavage of lignin inter-unit linkages. The organosolv pretreatment increased the contents of phenolic OH units and the lignin residue showed a high cellulase maximum adsorption capacity. Ammonia pretreatment with 5% ammonium hydroxide was not as effective as organosolv pretreatment on lignin deconstruction. Organosolv lignin residue had lower lignin S/G ratio than the untreated one. Furthermore, when compared to the organosolv lignin residue and untreated lignin, ammonia lignin residue had a higher cellulase adsorption affinity. In addition, the effects of lignin on cellulose hydrolysis was investigated and the results suggested that the presence of lignin with cellulose substrates reduced cellulose hydrolysis, and its inhibitory effect was primarily determined by the lignin properties after each pretreatment. The organosolv pretreatment resulted in a slightly lower cellulase binding strength (249.7 mL g -1) on poplar lignin than that on untreated samples (261.1 mL g -1), while ammonia lignin residue showed a higher cellulase binding strength (402.8 mL g -1) and had more significant inhibition effect on cellulose hydrolysis. Our results demonstrated that the binding strength significantly affected the lignin-derived inhibition on enzymatic hydrolysis of cellulose in the cellulose-lignin mixtures.« less

Effects of organosolv and ammonia pretreatments on lignin properties and its inhibition for enzymatic hydrolysis

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

Yoo, Chang Geun; Li, Mi; Meng, Xianzhi

Lignin offers structural support and protection for plant cell walls; but, it also contributes to biomass recalcitrance and the costs of biofuel production via the biological pathway. Organosolv and ammonia pretreatments have been developed to reduce biomass recalcitrance and improve sugar release performance during enzymatic hydrolysis. It is believed that lignin properties are related to its inhibition on enzymatic hydrolysis; therefore, understanding the characteristics of lignin is a key for effective biomass conversion to biofuels. In this study, an organosolv pretreatment using 60% ethanol with 1.25% H 2SO 4 significantly deconstructed poplar lignin and reduced its molecular weights due tomore » the cleavage of lignin inter-unit linkages. The organosolv pretreatment increased the contents of phenolic OH units and the lignin residue showed a high cellulase maximum adsorption capacity. Ammonia pretreatment with 5% ammonium hydroxide was not as effective as organosolv pretreatment on lignin deconstruction. Organosolv lignin residue had lower lignin S/G ratio than the untreated one. Furthermore, when compared to the organosolv lignin residue and untreated lignin, ammonia lignin residue had a higher cellulase adsorption affinity. In addition, the effects of lignin on cellulose hydrolysis was investigated and the results suggested that the presence of lignin with cellulose substrates reduced cellulose hydrolysis, and its inhibitory effect was primarily determined by the lignin properties after each pretreatment. The organosolv pretreatment resulted in a slightly lower cellulase binding strength (249.7 mL g -1) on poplar lignin than that on untreated samples (261.1 mL g -1), while ammonia lignin residue showed a higher cellulase binding strength (402.8 mL g -1) and had more significant inhibition effect on cellulose hydrolysis. Our results demonstrated that the binding strength significantly affected the lignin-derived inhibition on enzymatic hydrolysis of cellulose in the cellulose-lignin mixtures.« less

Batch bioethanol production via the biological and chemical saccharification of some Egyptian marine macroalgae.

PubMed

Soliman, Ramadan M; Younis, Sherif A; El-Gendy, Nour Sh; Mostafa, Soha S M; El-Temtamy, Seham A; Hashim, Ahmed I

2018-04-19

Marine seaweeds (macroalgae) cause eutrophication problem and affects the touristic activities. The success of the production of the third generation bioethanol from marine macroalgae depends mainly on the development of an ecofriendly and eco-feasible pretreatment (i.e. hydrolysis) technique, a highly effective saccharification step and finally an efficient bioethanol fermentation step. Therefore, this study aimed to investigate the potentiality of different marine macroalgal strains, collected from Egyptian coasts, for bioethanol production via different saccharification processes. Different marine macroalgal strains; red Jania rubens, green Ulva lactuca. and brown Sargassum latifolium, have been collected from Egyptian Mediterranean and Red Sea shores. Different hydrolysis processes were evaluated to maximize the extraction of fermentable sugars; thermo-chemical hydrolysis with diluted acids (HCl and H 2 SO 4 ) and base (NaOH), hydrothermal hydrolysis followed by saccharification with different fungal strains and finally, thermo-chemical hydrolysis with diluted HCl, followed by fungal saccharification. The hydrothermal hydrolysis of Sargassum latifolium followed by biological saccharification using Trichoderma asperellum RM1 produced maximum total sugars of 510 mg g -1 macroalgal biomass. The integration of the hydrothermal and fungal hydrolyses of the macroalgal biomass with a separate batch fermentation of the produced sugars using two Saccharomyces cerevisiae strains, produced approximately 0.29 g bioethanol g -1 total reducing sugars. A simulated regression modeling for the batch bioethanol fermentation was also performed. This study, supported the possibility of using seaweeds as a renewable source of bioethanol, throughout a suggested integration of macroalgal biomass hydrothermal- and fungal- hydrolysis with a separate batch bioethanol fermentation process of the produced sugars. The usage of marine macroalgae (i.e. seaweeds) as feedstock for bioethanol; an alternative and/or complimentary to petro-fuel, would act as triple fact solution; bioremediation process for ecosystem, renewable energy source and economy savings. This article is protected by copyright. All rights reserved. This article is protected by copyright. All rights reserved.

The Multi Domain Caldicellulosiruptor bescii CelA Cellulase Excels at the Hydrolysis of Crystalline Cellulose

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

Brunecky, Roman; Donohoe, Bryon S.; Yarbrough, John M.

The crystalline nature of cellulose microfibrils is one of the key factors influencing biomass recalcitrance which is a key technical and economic barrier to overcome to make cellulosic biofuels a commercial reality. To date, all known fungal enzymes tested have great difficulty degrading highly crystalline cellulosic substrates. We have demonstrated that the CelA cellulase from Caldicellulosiruptor bescii degrades highly crystalline cellulose as well as low crystallinity substrates making it the only known cellulase to function well on highly crystalline cellulose. Unlike the secretomes of cellulolytic fungi, which typically comprise multiple, single catalytic domain enzymes for biomass degradation, some bacterial systemsmore » employ an alternative strategy that utilizes multi-catalytic domain cellulases. Additionally, CelA is extremely thermostable and highly active at elevated temperatures, unlike commercial fungal cellulases. Furthermore we have determined that the factors negatively affecting digestion of lignocellulosic materials by C. bescii enzyme cocktails containing CelA appear to be significantly different from the performance barriers affecting fungal cellulases. Furthermore, we explore the activity and degradation mechanism of CelA on a variety of pretreated substrates to better understand how the different bulk components of biomass, such as xylan and lignin, impact its performance.« less

The Multi Domain Caldicellulosiruptor bescii CelA Cellulase Excels at the Hydrolysis of Crystalline Cellulose

DOE PAGES

Brunecky, Roman; Donohoe, Bryon S.; Yarbrough, John M.; ...

2017-08-29

The crystalline nature of cellulose microfibrils is one of the key factors influencing biomass recalcitrance which is a key technical and economic barrier to overcome to make cellulosic biofuels a commercial reality. To date, all known fungal enzymes tested have great difficulty degrading highly crystalline cellulosic substrates. We have demonstrated that the CelA cellulase from Caldicellulosiruptor bescii degrades highly crystalline cellulose as well as low crystallinity substrates making it the only known cellulase to function well on highly crystalline cellulose. Unlike the secretomes of cellulolytic fungi, which typically comprise multiple, single catalytic domain enzymes for biomass degradation, some bacterial systemsmore » employ an alternative strategy that utilizes multi-catalytic domain cellulases. Additionally, CelA is extremely thermostable and highly active at elevated temperatures, unlike commercial fungal cellulases. Furthermore we have determined that the factors negatively affecting digestion of lignocellulosic materials by C. bescii enzyme cocktails containing CelA appear to be significantly different from the performance barriers affecting fungal cellulases. Furthermore, we explore the activity and degradation mechanism of CelA on a variety of pretreated substrates to better understand how the different bulk components of biomass, such as xylan and lignin, impact its performance.« less

Isothiocyanates, Nitriles, and Epithionitriles from Glucosinolates Are Affected by Genotype and Developmental Stage in Brassica oleracea Varieties.

PubMed

Hanschen, Franziska S; Schreiner, Monika

2017-01-01

Vegetables of the Brassica oleracea group, such as broccoli, cauliflower, and cabbage, play an important role for glucosinolate consumption in the human diet. Upon maceration of the vegetable tissue, glucosinolates are degraded enzymatically to form volatile isothiocyanates, nitriles, and epithionitriles. However, only the uptake of isothiocyanates is linked to the cancer-preventive effects. Thus, it is of great interest to evaluate especially the isothiocyanate formation. Here, we studied the formation of glucosinolates and their respective hydrolysis products in sprouts and fully developed vegetable heads of different genotypes of the five B. oleracea varieties: broccoli, cauliflower as well as white, red, and savoy cabbages. Further, the effect of ontogeny (developmental stages) during the head development on the formation of glucosinolates and their respective hydrolysis products was evaluated at three different developmental stages (mini, fully developed, and over-mature head). Broccoli and red cabbage were mainly rich in 4-(methylsulfinyl)butyl glucosinolate (glucoraphanin), whereas cauliflower, savoy cabbage and white cabbage contained mainly 2-propenyl (sinigrin) and 3-(methylsulfinyl)propyl glucosinolate (glucoiberin). Upon hydrolysis, epithionitriles or nitriles were often observed to be the main hydrolysis products, with 1-cyano-2,3-epithiopropane being most abundant with up to 5.7 μmol/g fresh weight in white cabbage sprouts. Notably, sprouts often contained more than 10 times more glucosinolates or their hydrolysis products compared to fully developed vegetables. Moreover, during head development, both glucosinolate concentrations as well as hydrolysis product concentrations changed and mini heads contained the highest isothiocyanate concentrations. Thus, from a cancer-preventive point of view, consumption of mini heads of the B. oleracea varieties is recommended.

Temperature induced decoupling of enzymatic hydrolysis and carbon remineralization in long-term incubations of Arctic and temperate sediments

NASA Astrophysics Data System (ADS)

Robador, Alberto; Brüchert, Volker; Steen, Andrew D.; Arnosti, Carol

2010-04-01

Extracellular enzymatic hydrolysis of high-molecular weight organic matter is the initial step in sedimentary organic carbon degradation and is often regarded as the rate-limiting step. Temperature effects on enzyme activities may therefore exert an indirect control on carbon mineralization. We explored the temperature sensitivity of enzymatic hydrolysis and its connection to subsequent steps in anoxic organic carbon degradation in long-term incubations of sediments from the Arctic and the North Sea. These sediments were incubated under anaerobic conditions for 24 months at temperatures of 0, 10, and 20 °C. The short-term temperature response of the active microbial community was tested in temperature gradient block incubations. The temperature optimum of extracellular enzymatic hydrolysis, as measured with a polysaccharide (chondroitin sulfate), differed between Arctic and temperate habitats by about 8-13 °C in fresh sediments and in sediments incubated for 24 months. In both Arctic and temperate sediments, the temperature response of chondroitin sulfate hydrolysis was initially similar to that of sulfate reduction. After 24 months, however, hydrolysis outpaced sulfate reduction rates, as demonstrated by increased concentrations of dissolved organic carbon (DOC) and total dissolved carbohydrates. This effect was stronger at higher incubation temperatures, particularly in the Arctic sediments. In all experiments, concentrations of volatile fatty acids (VFA) were low, indicating tight coupling between VFA production and consumption. Together, these data indicate that long-term incubation at elevated temperatures led to increased decoupling of hydrolytic DOC production relative to fermentation. Temperature increases in marine sedimentary environments may thus significantly affect the downstream carbon mineralization and lead to the increased formation of refractory DOC.

Isothiocyanates, Nitriles, and Epithionitriles from Glucosinolates Are Affected by Genotype and Developmental Stage in Brassica oleracea Varieties

PubMed Central

Hanschen, Franziska S.; Schreiner, Monika

2017-01-01

Vegetables of the Brassica oleracea group, such as broccoli, cauliflower, and cabbage, play an important role for glucosinolate consumption in the human diet. Upon maceration of the vegetable tissue, glucosinolates are degraded enzymatically to form volatile isothiocyanates, nitriles, and epithionitriles. However, only the uptake of isothiocyanates is linked to the cancer-preventive effects. Thus, it is of great interest to evaluate especially the isothiocyanate formation. Here, we studied the formation of glucosinolates and their respective hydrolysis products in sprouts and fully developed vegetable heads of different genotypes of the five B. oleracea varieties: broccoli, cauliflower as well as white, red, and savoy cabbages. Further, the effect of ontogeny (developmental stages) during the head development on the formation of glucosinolates and their respective hydrolysis products was evaluated at three different developmental stages (mini, fully developed, and over-mature head). Broccoli and red cabbage were mainly rich in 4-(methylsulfinyl)butyl glucosinolate (glucoraphanin), whereas cauliflower, savoy cabbage and white cabbage contained mainly 2-propenyl (sinigrin) and 3-(methylsulfinyl)propyl glucosinolate (glucoiberin). Upon hydrolysis, epithionitriles or nitriles were often observed to be the main hydrolysis products, with 1-cyano-2,3-epithiopropane being most abundant with up to 5.7 μmol/g fresh weight in white cabbage sprouts. Notably, sprouts often contained more than 10 times more glucosinolates or their hydrolysis products compared to fully developed vegetables. Moreover, during head development, both glucosinolate concentrations as well as hydrolysis product concentrations changed and mini heads contained the highest isothiocyanate concentrations. Thus, from a cancer-preventive point of view, consumption of mini heads of the B. oleracea varieties is recommended. PMID:28690627

Adenosine receptor activation potentiates phosphoinositide hydrolysis and arachidonic acid release in DDT1-MF2 cells: putative interrelations.

PubMed

Schachter, J B; Yasuda, R P; Wolfe, B B

1995-09-01

Studies were undertaken in an effort to discern possible mechanisms by which the A1 adenosine receptor agonist cyclopentyladenosine (CPA) enhances the norepinephrine-stimulated (NE-stimulated) hydrolysis of phosphoinositides in DDT1-MF2 cells. Measurements of arachidonic acid release revealed similar behaviours to those observed in measurements of phosphoinositide hydrolysis. In the presence of NE, both second messenger responses were potentiated by the addition of CPA, whereas in the absence of NE, CPA had little or no effect on either second messenger. The stimulation and potentiation of both second messenger responses were enhanced in the presence of extracellular calcium, and in each case these effects were persistent over time. For either second messenger system the stimulation by NE and the potentiation by CPA appeared to utilize separate mechanisms as evidenced by the fact that the potentiations by CPA were selectively antagonized by a cAMP analogue or by pertussis toxin, whereas the stimulations by NE were essentially unaffected by these agents. Inhibition of phospholipase A2 (PLA2) also blocked the potentiation of PLC by CPA, without affecting NE-stimulated phosphoinositide hydrolysis. Furthermore, in the presence of CPA, the exogenous administration of PLA2 was found to stimulate phosphoinositide hydrolysis in these cells. These data are consistent with a hypothesis whereby the apparent potentiation of NE-stimulated phosphoinositide hydrolysis by CPA is actually due to the stimulation by CPA of a second pathway of phospholipase C activity which is additive to that of NE. The activation of PLC and PLA2 by NE produces phospholipid products which may play a permissive role in the pathway coupling adenosine A1 receptors to these phospholipases. The formation of lysophosphatidic acid is suggested as one possible mediator of this permissive effect.

An integrated green process: Subcritical water, enzymatic hydrolysis, and fermentation, for biohydrogen production from coconut husk.

PubMed

Muharja, Maktum; Junianti, Fitri; Ranggina, Dian; Nurtono, Tantular; Widjaja, Arief

2018-02-01

The objective of this work is to develop an integrated green process of subcritical water (SCW), enzymatic hydrolysis and fermentation of coconut husk (CCH) to biohydrogen. The maximum sugar yield was obtained at mild severity factor. This was confirmed by the degradation of hemicellulose, cellulose and lignin. The tendency of the changing of sugar yield as a result of increasing severity factor was opposite to the tendency of pH change. It was found that CO 2 gave a different tendency of severity factor compared to N 2 as the pressurizing gas. The result of SEM analysis confirmed the structural changes during SCW pretreatment. This study integrated three steps all of which are green processes which ensured an environmentally friendly process to produce a clean biohydrogen. Copyright © 2017 Elsevier Ltd. All rights reserved.

In vitro succinylcholine hydrolysis in plasma of the African elephant (Loxodonta africana) and impala (Aepyceros melampus).

PubMed

Pitts, Neville I; Mitchell, Graham

2003-01-01

In elephants the time lapsed from i.m. injection of an overdose of the muscle relaxant succinylcholine (SuCh) until death, is significantly longer than in impala. To determine a difference in the rate of SuCh hydrolysis, once the drug enters the circulation, contributes to this phenomenon we have measured the rate of hydrolysis of SuCh in elephant and impala plasma, and by elephant erythrocytes. Rate of hydrolysis was determined by incubating SuCh in plasma or erythrocyte lysate at 37 degrees C and quantifying the choline produced. Plasma SuCh hydrolytic activity in elephant plasma (12.1+/-1.7 Ul(-1) mean+/-S.D.; n=9) was significantly higher than it was in impala plasma (6.6+/-0.6 Ul(-1); n=5), but were approximately 12 and 21 times lower, respectively, than in human plasma. Elephant erythrocyte lysate had no SuCh hydrolytic activity. Applying this data to previous studies, we can show that the ratio of SuCh absorption to SuCh hydrolysis is expected to be 1.25:1 and 1.41:1 for elephants and impala respectively. It will thus take at least 1.7 times longer for elephant to achieve a plasma SuCh concentration similar to that in impala. We conclude that a more rapid hydrolysis of SuCh in elephant plasma is one factor that contributes to the longer time to death compared to impala.

Kinetics and mechanism of imazosulfuron hydrolysis.

PubMed

Morrica, P; Barbato, F; Della Iacovo, R; Seccia, S; Ungaro, F

2001-08-01

Knowledge of the kinetics and pathways of hydrolytic degradation is crucial to the prediction of the fate and transport mechanism of chemicals. This work first describes the kinetics of the chemical hydrolysis of imazosulfuron, a new sulfonylurea herbicide, and evaluates the results to propose a degradation pathway. The hydrolysis of imazosulfuron has been studied in aqueous buffers both within the pH range 1.9-12.3 at ambient temperature (thermostated at 25 +/- 2 degrees C) and at pH 3.6 within the temperature range of 15-55 degrees C. The hydrolysis rate of imazosulfuron was characterized by a first-order kinetics, pH- and temperature-dependent, and accelerated by acidic conditions and higher temperatures. The calculated half-lives at pH 4.5 and 5.9 were 36.5 and 578 days, respectively. At pH 6.6, 7.4, 9.2, and 12.3 no significant change in imazosulfuron concentration was observed after 150 days. Half-lives were much lower at pH <4 (= imazosulfuron pK(a)), at which they ranged from 3.3 to 6.3 days. Moreover, a change in temperature from 15 to 25 degrees C in acidic conditions (pH 3.6) decreased the half-life of imazosulfuron by a factor of approximately 4.0; in any case, a 3-5-fold increase in the rate of hydrolysis was found for each 10 degrees C increase in temperature. In acidic conditions the only hydrolysis products were the two molecules resulting from the cleavage of the sulfonylurea bridge.

Hydrolysis of oligosaccharides over solid acid catalysts: a review.

PubMed

Vilcocq, Léa; Castilho, Paula C; Carvalheiro, Florbela; Duarte, Luís C

2014-04-01

Mild fractionation/pretreatment processes are becoming the most preferred choices for biomass processing within the biorefinery framework. To further explore their advantages, new developments are needed, especially to increase the extent of the hydrolysis of poly- and oligosaccharides. A possible way forward is the use of solid acid catalysts that may overcome many current drawbacks of other common methods. In this Review, the advantages and limitations of the use of heterogeneous catalysis for the main groups of solid acid catalysts (zeolites, resins, carbon materials, clays, silicas, and other oxides) and their relation to the hydrolysis of model soluble disaccharides and soluble poly- and oligosaccharides are presented and discussed. Special attention is given to the hydrolysis of hemicelluloses and hemicellulose-derived saccharides into monosaccharides, the impact on process performance of potential catalyst poisons originating from biomass and biomass hydrolysates (e.g., proteins, mineral ions, etc.). The data clearly point out the need for studying hemicelluloses in natura rather than in model compound solutions that do not retain the relevant factors influencing process performance. Furthermore, the desirable traits that solid acid catalysts must possess for the efficient hemicellulose hydrolysis are also presented and discussed with regard to the design of new catalysts. © 2014 WILEY-VCH Verlag GmbH & Co. KGaA, Weinheim.

Effect of protonation on the mechanism of phosphate monoester hydrolysis and comparison with the hydrolysis of nucleoside triphosphate in biomolecular motors.

PubMed

Hassan, Hammad Ali; Rani, Sadaf; Fatima, Tabeer; Kiani, Farooq Ahmad; Fischer, Stefan

2017-11-01

Hydrolysis of phosphate groups is a crucial reaction in living cells. It involves the breaking of two strong bonds, i.e. the O a H bond of the attacking water molecule, and the PO l bond of the substrate (O a and O l stand for attacking and leaving oxygen atoms). Mechanism of the hydrolysis reaction can proceed either by a concurrent or a sequential mechanism. In the concurrent mechanism, the breaking of O a H and PO l bonds occurs simultaneously, whereas in the sequential mechanism, the O a H and PO l bonds break at different stages of the reaction. To understand how protonation affects the mechanism of hydrolysis of phosphate monoester, we have studied the mechanism of hydrolysis of protonated and deprotonated phosphate monoester at M06-2X/6-311+G**//M06-2X/6-31+G*+ZPE level of theory (where ZPE stands for zero point energy). Our calculations show that in both protonated and deprotonated cases, the breaking of the water O a H bond occurs before the breaking of the PO l bond. Because the two events are not separated by a stable intermediate, the mechanism can be categorized as semi-concurrent. The overall energy barrier is 41kcalmol -1 in the unprotonated case. Most (5/6th) of this is due to the initial breaking of the water O a H bond. This component is lowered from 34 to 25kcalmol -1 by adding one proton to the phosphate. The rest of the overall energy barrier comes from the subsequent breaking of the PO l bond and is not sensitive to protonation. This is consistent with previous findings about the effect of triphosphate protonation on the hydrolysis, where the equivalent protonation (on the γ-phosphate) was seen to lower the barrier of breaking the water O a H bond and to have little effect on the PO l bond breaking. Hydrolysis pathways of phosphate monoester with initial breaking of the PO l bond could not be found here. This is because the leaving group in phosphate monoester cannot be protonated, unlike in triphosphate hydrolysis, where protonation of the β- and γ-phosphates had been shown to promote a mechanism where the PO l bond breaks before the O a H bond does. We also point out that the charge shift due to PO l bond breaking during sequential ATP hydrolysis in bio-molecular motors onsets the week unbinding of hydrolysis product that finally leads to the product release during power stroke. Copyright © 2017 Elsevier B.V. All rights reserved.

Hydrogen Generation from Al-NiCl2/NaBH4 Mixture Affected by Lanthanum Metal

PubMed Central

Qiang Sun, Wen; Fan, Mei-Qiang; Fei, Yong; Pan, Hua; Wang, Liang Liang; Yao, Jun

2012-01-01

The effect of La on Al/NaBH4 hydrolysis was elaborated in the present paper. Hydrogen generation amount increases but hydrogen generation rate decreases with La content increasing. There is an optimized composition that Al-15 wt% La-5 wt% NiCl2/NaBH4 mixture (Al-15 wt% La-5 wt% NiCl2/NaBH4 weight ratio, 1 : 3) has 126 mL g−1 min−1 maximum hydrogen generation rate and 1764 mL g−1 hydrogen generation amount within 60 min. The efficiency is 88%. Combined with NiCl2, La has great effect on NaBH4 hydrolysis but has little effect on Al hydrolysis. Increasing La content is helpful to decrease the particle size of Al-La-NiCl2 in the milling process, which induces that the hydrolysis byproduct Ni2B is highly distributed into Al(OH)3 and the catalytic reactivity of Ni2B/Al(OH)3 is increased therefore. But hydrolysis byproduct La(OH)3 deposits on Al surface and leads to some side effect. The Al-La-NiCl2/NaBH4 mixture has good stability in low temperature and its hydrolytic performance can be improved with increasing global temperature. Therefore, the mixture has good safety and can be applied as on board hydrogen generation material. PMID:22619596

Optimization of papain hydrolysis conditions for release of glycosaminoglycans from the chicken keel cartilage

NASA Astrophysics Data System (ADS)

Le Vien, Nguyen Thi; Nguyen, Pham Bao; Cuong, Lam Duc; An, Trinh Thi Thua; Dao, Dong Thi Anh

2017-09-01

Glycosaminoglycans (GAGs) are natural biocompounds which join to construct cartilage tissuses, it can be extracted from cartilage of sharks, pigs, cows, chickens, etc. GAGs contain a Chondroitin sulfate (CS) content which is a supplement of functional food used for preventing and supporting treatment of arthritis and eye diseases. Therefore, the GAGs extraction from byproducts of the industry of cattle and poultry slaughter to identify the CS content by papain enzyme is necessary. In this study, the optimal hydrolysis conditions were obtained by response surface methodology (RSM). The independent variables were coded as: pH (x1), enzyme concentration (x2), incubation temperature (x3) and hydrolysis time (x4). The results of the analysis of variance (ANOVA) shown that the variables actively affected GAGs content. The optimal conditions of hydrolysis were derived at pH of 7.1, ratio of enzyme per substances of 0.62% w/wpo, temperature of 65°C and hydrolysis time of 230 minutes, GAGs content reached 14.3% of the dry matter of raw material. Analyzes by HPLC revealed that 56.17% of the dry preparations of GAGs were CS compound, were equivalent to 8.11% of the dry matter of chicken keel cartilage. Molecular weight of the dry preparations GAGs was 259.6 kDa. The dry preparations included the contents of moisture 12.2%, protein 8.42%, lipid 0%, ash 10.03% and extracted GAGs 69.35%.

An evaluation of the role of a pyroglutamyl peptidase, a post-proline cleaving enzyme and a post-proline dipeptidyl amino peptidase, each purified from the soluble fraction of guinea-pig brain, in the degradation of thyroliberin in vitro.

PubMed

Browne, P; O'Cuinn, G

1983-12-01

The degradation of thyroliberin (less than Glu-His-Pro-NH2) to its component amino acids by the soluble fraction of guinea pig brain is catalysed by four enzymes namely a pyroglutamate aminopeptidase, a post-proline cleaving enzyme, a post-proline dipeptidyl aminopeptidase and a proline dipeptidase. 1. The pyroglutamate aminopeptidase was purified to over 90% homogeneity with a purification factor of 2868-fold and a yield of 5.7%. In addition to catalysing the hydrolysis of thyroliberin, acid thyroliberin and pyroglutamate-7-amido-4-methylcoumarin the pyroglutamate aminopeptidase catalysed the hydrolysis of the peptide bond adjacent to the pyroglutamic acid residue in luliberin, neurotensin bombesin, bradykinin-potentiating peptide B, the anorexogenic peptide and the dipeptides pyroglutamyl alanine and pyroglutamyl valine. Pyroglutamyl proline and eledoisin were not hydrolysed. 2. The post-proline cleaving enzyme was purified to apparent electrophoretic homogeneity with a purification factor of 2298-fold and a yield of 10.6%. The post-proline cleaving enzyme catalysed the hydrolysis of thyroliberin and N-benzyloxycarbonyl-glycylproline-7-amido-4-methylcoumarin. It did not catalyse the hydrolysis of glycylproline-7-amido-4-methylcoumarin or His-Pro-NH2. 3. The post-proline dipeptidyl aminopeptidase was partially purified with a purification factor of 301-fold and a yield of 8.9%. The post-proline dipeptidyl aminopeptidase catalysed the hydrolysis of His-Pro-NH2 and glycylproline-7-amido-4-methylcoumarin but did not exhibit any post-proline cleaving endopeptidase activity against thyroliberin or N-benzyloxycarbonyl-glycylproline-7-amido-4-methylcoumarin. 4. Studies with various functional reagents indicated that the pyroglutamate aminopeptidase could be specifically inhibited by 2-iodoacetamide (100% inhibition at an inhibitor concentration of 5 microM), the post-proline cleaving enzyme by bacitracin (IC50 = 42 microM) and the post-proline dipeptidyl aminopeptidase by puromycin (IC50 = 46 microM). Because of their specific inhibitory effects these three reagents were key elements in the elucidation of the overall pathway for the metabolism of thyroliberin by guinea pig brain tissue enzymes.

Effects of Dilute Acid Pretreatment on Cellulose DP and the Relationship Between DP Reduction and Cellulose Digestibility

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

Wang, W.; Chen, X.; Tucker, M.

2012-01-01

The degree of polymerization(DP) of cellulose is considered to be one of the most important properties affecting the enzymatic hydrolysis of cellulose. Various pure cellulosic and biomass materials have been used in a study of the effect of dilute acid treatment on cellulose DP. A substantial reduction in DP was found for all pure cellulosic materials studied even at conditions that would be considered relatively mild for pretreatment. The effect of dilute acid pretreatment on cellulose DP in biomass samples was also investigated. Corn stover pretreated with dilute acid under the most optimal conditions contained cellulose with a DPw inmore » the range of 1600{approx}3500, which is much higher than the level-off DP(DPw 150{approx}300) obtained with pure celluloses. The effect of DP reduction on the saccharification of celluloses was also studied. From this study it does not appear that cellulose DP is a main factor affecting cellulose saccharification.« less

Short communication: Urea hydrolysis in dairy cattle manure under different temperature, urea, and pH conditions.

PubMed

Moraes, L E; Burgos, S A; DePeters, E J; Zhang, R; Fadel, J G

2017-03-01

The objective of the study was to quantify the rate of urea hydrolysis in dairy cattle manure under different initial urea concentration, temperature, and pH conditions. In particular, by varying all 3 factors simultaneously, the interactions between them could also be determined. Fresh feces and artificial urine solutions were combined into a slurry to characterize the rate of urea hydrolysis under 2 temperatures (15°C and 35°C), 3 urea concentrations in urine solutions (500, 1,000, and 1,500 mg of urea-N/dL), and 3 pH levels (6, 7, and 8). Urea N concentration in slurry was analyzed at 0.0167, 1, 2, 4, 6, 8, 12, 16, 20, and 24 h after initial mixing. A nonlinear mixed effects model was used to determine the effects of urea concentration, pH, and temperature treatments on the exponential rate of urea hydrolysis and to predict the hydrolysis rate for each treatment combination. We detected a significant interaction between pH and initial urea level. Increasing urea concentration from 1,000 to 1,500 mg of urea-N/dL decreased the rate of urea hydrolysis across all pH levels. Across all pH and initial urea levels, the rate of urea hydrolysis increased with temperature, but the effect of pH was only observed for pH 6 versus pH 8 at the intermediate initial urea concentration. The fast rates of urea hydrolysis indicate that urea was almost completely hydrolyzed within a few hours of urine mixing with feces. The estimated urea hydrolysis rates from this study are likely maximum rates because of the thorough mixing before each sampling. Although considerable mixing of feces and urine occurs on the barn floor of commercial dairy operations from cattle walking through the manure, such mixing may be not as quick and thorough as in this study. Consequently, the urea hydrolysis rates from this study indicate the maximum loss of urea and should be accounted for in management aimed at mitigating ammonia emissions from dairy cattle manure under similar urea concentration, pH, and temperature conditions reported in this experiment. Copyright © 2017 American Dairy Science Association. Published by Elsevier Inc. All rights reserved.

Human plasma kallikrein and tissue kallikrein binding to a substrate based on the reactive site of a factor Xa inhibitor isolated from Bauhinia ungulata seeds.

PubMed

Oliva, M L; Andrade, S A; Batista, I F; Sampaio, M U; Juliano, M; Fritz, H; Auerswald, E A; Sampaio, C A

1999-12-01

Kunitz type Bauhinia ungulata factor Xa inhibitor (BuXI) was purified from B. ungulata seeds. BuXI inactivates factor Xa and human plasma kallikrein (HuPK) with Ki values of 18.4 and 6.9 nM, respectively. However, Bauhinia variegata trypsin inhibitor (BvTI) which is 70% homologous to BuXI does not inhibit factor Xa and is less efficient on HuPK (Ki = 80 nM). The comparison between BuXI and BvTI reactive site structure indicates differences at Met59, Thr66 and Met67 residues. The hydrolysis rate of quenched fluorescence peptide substrates based on BuXI reactive site sequence, Abz-VMIAALPRTMFIQ-EDDnp (leading peptide), by HuPK and porcine pancreatic kallikrein (PoPK) is low, but hydrolysis is enhanced with Abz-VMIAALPRTMQ-EDDnp, derived from the leading peptide shortened by removing the dipeptide Phe-Ileu from the C-terminal portion, for HuPK (Km = 0.68 microM, k(cat)/Km = 1.3 x 10(6) M(-1) s(-1)), and the shorter substrate Abz-LPRTMQ-EDDnp is better for PoPK (Km = 0.66 microM, k(cat)/Km = 2.2 x 10(3) M(-1) s(-1)). The contribution of substrate methionine residues to HuPK and PoPK hydrolysis differs from that observed with factor Xa. The determined Km and k(cat) values suggest that the substrates interact with kallikreins the same as an enzyme and inhibitor interacts to form complexes.

Lignocellulosic hydrolysate inhibitors selectively inhibit/deactivate cellulase performance.

PubMed

Mhlongo, Sizwe I; den Haan, Riaan; Viljoen-Bloom, Marinda; van Zyl, Willem H

2015-12-01

In this study, we monitored the inhibition and deactivation effects of various compounds associated with lignocellulosic hydrolysates on individual and combinations of cellulases. Tannic acid representing polymeric lignin residues strongly inhibited cellobiohydrolase 1 (CBH1) and β-glucosidase 1 (BGL1), but had a moderate inhibitory effect on endoglucanase 2 (EG2). Individual monomeric lignin residues had little or no inhibitory effect on hydrolytic enzymes. However, coniferyl aldehyde and syringaldehyde substantially decreased the activity of CBH1 and deactivated BGL1. Acetic and formic acids also showed strong inhibition of BGL1 but not CBH1 and EG2, whereas tannic, acetic and formic acid strongly inhibited a combination of CBH1 and EG2 during Avicel hydrolysis. Diminishing enzymatic hydrolysis is largely a function of inhibitor concentration and the enzyme-inhibitor relationship, rather than contact time during the hydrolysis process (i.e. deactivation). This suggests that decreased rates of hydrolysis during the enzymatic depolymerisation of lignocellulosic hydrolysates may be imparted by other factors related to substrate crystallinity and accessibility. Copyright © 2015 Elsevier Inc. All rights reserved.

Activation of Elongation Factor G by Phosphate Analogues

PubMed Central

Salsi, Enea; Farah, Elie

2016-01-01

EF-G is a universally conserved translational GTPase that promotes the translocation of tRNA and mRNA through the ribosome. EF-G binds to the ribosome in a GTP-bound form and subsequently catalyzes GTP hydrolysis. The contribution of the ribosome-stimulated GTP hydrolysis by EF-G to tRNA/mRNA translocation remains debated. Here, we show that while EF-G•GDP does not stably bind to the ribosome and induce translocation, EFG• GDP in complex with phosphate group analogues BeF3− and AlF4− promotes the translocation of tRNA and mRNA. Furthermore, the rates of mRNA translocation induced by EF-G in the presence of GTP and a non-hydrolysable analogue of GTP, GDP•BeF3−are similar. Our results are consistent with the model suggesting that GTP hydrolysis is not directly coupled to mRNA/tRNA translocation. Hence, GTP binding is required to induce the activated, translocation-competent conformation of EF-G while GTP hydrolysis triggers EF-G release from the ribosome. PMID:27063503

Tassel Removal Positively Affects Biomass Production Coupled with Significantly Increasing Stem Digestibility in Switchgrass

PubMed Central

Zhao, Chunqiao; Fan, Xifeng; Hou, Xincun; Zhu, Yi; Yue, Yuesen; Zhang, Shuang; Wu, Juying

2015-01-01

In this study, tassels of Cave-in-Rock (upland) and Alamo (lowland) were removed at or near tassel emergence to explore its effects on biomass production and quality. Tassel-removed (TR) Cave-in-Rock and Alamo both exhibited a significant (P<0.05) increase in plant heights (not including tassel length), tiller number, and aboveground biomass dry weight (10% and 12%, 30% and 13%, 13% and 18%, respectively by variety) compared to a control (CK) treatment. Notably, total sugar yields of TR Cave-in-Rock and Alamo stems increased significantly (P<0.05 or 0.01) by 19% and 19%, 21% and 14%, 52% and 18%, respectively by variety, compared to those of control switchgrass under 3 treatments by direct enzymatic hydrolysis (DEH), enzymatic hydrolysis after 1% NaOH pretreatment (EHAL) and enzymatic hydrolysis after 1% H2SO4 pretreatment (EHAC). These differences were mainly due to significantly (P<0.05 or 0.01) higher cellulose content, lower cellulose crystallinity indexes (CrI) caused by higher arabinose (Ara) substitution in xylans, and lower S/G ratio in lignin. However, the increases of nitrogen (N) and sulphur (S) concentration negatively affects the combustion quality of switchgrass aboveground biomass. This work provides information for increasing biomass production and quality in switchgrass and also facilitates the inhibition of gene dispersal of switchgrass in China. PMID:25849123

Phosphoric acid pretreatment of Achyranthes aspera and Sida acuta weed biomass to improve enzymatic hydrolysis.

PubMed

Siripong, Premjet; Duangporn, Premjet; Takata, Eri; Tsutsumi, Yuji

2016-03-01

Achyranthes aspera and Sida acuta, two types of weed biomass are abundant and waste in Thailand. We focus on them as novel feedstock for bio-ethanol production because they contain high-cellulose content (45.9% and 46.9%, respectively) and unutilized material. Phosphoric acid (70%, 75%, and 80%) was employed for the pretreatment to improve by enzymatic hydrolysis. The pretreatment process removed most of the xylan and a part of the lignin from the weeds, while most of the glucan remained. The cellulose conversion to glucose was greater for pretreated A. aspera (86.2 ± 0.3%) than that of the pretreated S. acuta (82.2 ± 1.1%). Thus, the removal of hemicellulose significantly affected the efficiency of the enzymatic hydrolysis. The scanning electron microscopy images showed the exposed fibrous cellulose on the cell wall surface, and this substantial change of the surface structure contributed to improving the enzyme accessibility. Copyright © 2015 Elsevier Ltd. All rights reserved.

[Study of surface enhanced Raman scattering of trace trinitrotoluene based on silver colloid nanoparticles].

PubMed

Zhang, Chun-ling; Li, Zhe; Wu, Zheng-long; Han, De-jun

2012-03-01

Trinitrotoluene (TNT), a representative nitroexplosive, attracts more and more attentions because of the urgent demand for trace analysis of explosives in recent years. The present study investigated the experiment condition of the surface enhanced Raman scattering (SERS) of 10(-6) mol x L(-1) TNT solution, especially the influence of NaCl and basic hydrolysis. The results indicate that SERS spectra of TNT can not be obtained when preparing the SERS samples without NaCl, and it was also shown that the intensity of Raman peaks has a relationship with the concentration of NaCl. With the increase in the concentration of NaCl, the intensity of Raman peak at 1 392 cm(-1) has a maximum value. This report explained the reason why NaCl can affect the intensity of SERS theoretically. It was also shown that the SERS spectrum of TNT treated with basic hydrolysis is more intense than that without basic hydrolysis.

Feasibility of reusing the black liquor for enzymatic hydrolysis and ethanol fermentation.

PubMed

Wang, Wen; Chen, Xiaoyan; Tan, Xuesong; Wang, Qiong; Liu, Yunyun; He, Minchao; Yu, Qiang; Qi, Wei; Luo, Yu; Zhuang, Xinshu; Yuan, Zhenhong

2017-03-01

The black liquor (BL) generated in the alkaline pretreatment process is usually thought as the environmental pollutant. This study found that the pure alkaline lignin hardly inhibited the enzymatic hydrolysis of cellulose (EHC), which led to the investigation on the feasibility of reusing BL as the buffer via pH adjustment for the subsequent enzymatic hydrolysis and fermentation. The pH value of BL was adjusted from 13.23 to 4.80 with acetic acid, and the alkaline lignin was partially precipitated. It deposited on the surface of cellulose and negatively influenced the EHC via blocking the access of cellulase to cellulose and adsorbing cellulase. The supernatant separated from the acidified BL scarcely affected the EHC, but inhibited the ethanol fermentation. The 4-times diluted supernatant and the last-time waste wash water of the alkali-treated sugarcane bagasse didn't inhibit the EHC and ethanol production. This work gives a clue of saving water for alkaline pretreatment. Copyright © 2016 Elsevier Ltd. All rights reserved.

Thermochemical hydrolysis of macroalgae Ulva for biorefinery: Taguchi robust design method

PubMed Central

Jiang, Rui; Linzon, Yoav; Vitkin, Edward; Yakhini, Zohar; Chudnovsky, Alexandra; Golberg, Alexander

2016-01-01

Understanding the impact of all process parameters on the efficiency of biomass hydrolysis and on the final yield of products is critical to biorefinery design. Using Taguchi orthogonal arrays experimental design and Partial Least Square Regression, we investigated the impact of change and the comparative significance of thermochemical process temperature, treatment time, %Acid and %Solid load on carbohydrates release from green macroalgae from Ulva genus, a promising biorefinery feedstock. The average density of hydrolysate was determined using a new microelectromechanical optical resonator mass sensor. In addition, using Flux Balance Analysis techniques, we compared the potential fermentation yields of these hydrolysate products using metabolic models of Escherichia coli, Saccharomyces cerevisiae wild type, Saccharomyces cerevisiae RN1016 with xylose isomerase and Clostridium acetobutylicum. We found that %Acid plays the most significant role and treatment time the least significant role in affecting the monosaccharaides released from Ulva biomass. We also found that within the tested range of parameters, hydrolysis with 121 °C, 30 min 2% Acid, 15% Solids could lead to the highest yields of conversion: 54.134–57.500 gr ethanol kg−1 Ulva dry weight by S. cerevisiae RN1016 with xylose isomerase. Our results support optimized marine algae utilization process design and will enable smart energy harvesting by thermochemical hydrolysis. PMID:27291594

Isolation, Solubility, and Characterization of D-Mannitol Esters of 4-Methoxybenzeneboronic Acid.

PubMed

Lopalco, Antonio; Marinaro, William A; Day, Victor W; Stella, Valentino J

2017-02-01

The purpose of this study was to determine the aqueous solubility of a model phenyl boronic acid, 4-methoxybenzeneboronic acid, as a function of pH both in the absence and in the presence of varying D-mannitol concentration. Solid isolated D-mannitol esters were characterized by differential scanning calorimetry, thermogravimetric analysis, powder X-ray diffraction, and single-crystal X-ray studies, and the boronic acid-to-D-mannitol ratio was quantified by HPLC. Hydrolysis of the monoester was studied using UV spectral differences between the monoester and the parent boronic acid. Two D-mannitol esters of 4-methoxybenzeneboronic acid were isolated. The triboronate ester was very insoluble whereas a symmetrical monoboronate monohydrate was also less soluble than the parent. Both esters were crystalline. The monoboronate monohydrate was, however, more soluble than the parent at alkaline pH values due to its lower pKa value (6.53) compared to the parent acid (9.41). Hydrolysis of the monoboronate was extremely fast when even small amount of water was added to dry acetonitrile solutions of the ester. The hydrolysis was buffer concentration dependent and apparent pH sensitive with hydrolysis accelerated by acid. Implications affecting the formulation of future boronic acid drugs are discussed. Copyright © 2017 American Pharmacists Association®. Published by Elsevier Inc. All rights reserved.

Thermochemical hydrolysis of macroalgae Ulva for biorefinery: Taguchi robust design method

NASA Astrophysics Data System (ADS)

Jiang, Rui; Linzon, Yoav; Vitkin, Edward; Yakhini, Zohar; Chudnovsky, Alexandra; Golberg, Alexander

2016-06-01

Understanding the impact of all process parameters on the efficiency of biomass hydrolysis and on the final yield of products is critical to biorefinery design. Using Taguchi orthogonal arrays experimental design and Partial Least Square Regression, we investigated the impact of change and the comparative significance of thermochemical process temperature, treatment time, %Acid and %Solid load on carbohydrates release from green macroalgae from Ulva genus, a promising biorefinery feedstock. The average density of hydrolysate was determined using a new microelectromechanical optical resonator mass sensor. In addition, using Flux Balance Analysis techniques, we compared the potential fermentation yields of these hydrolysate products using metabolic models of Escherichia coli, Saccharomyces cerevisiae wild type, Saccharomyces cerevisiae RN1016 with xylose isomerase and Clostridium acetobutylicum. We found that %Acid plays the most significant role and treatment time the least significant role in affecting the monosaccharaides released from Ulva biomass. We also found that within the tested range of parameters, hydrolysis with 121 °C, 30 min 2% Acid, 15% Solids could lead to the highest yields of conversion: 54.134-57.500 gr ethanol kg-1 Ulva dry weight by S. cerevisiae RN1016 with xylose isomerase. Our results support optimized marine algae utilization process design and will enable smart energy harvesting by thermochemical hydrolysis.

An in-depth understanding of biomass recalcitrance using natural poplar variants as the feedstock

DOE PAGES

Meng, Xianzhi; Pu, Yunqiao; Yoo, Chang Geun; ...

2016-12-12

Here, in an effort to better understand the biomass recalcitrance, six natural poplar variants were selected as feedstocks based on previous sugar release analysis. Compositional analysis and physicochemical characterizations of these poplars were performed and the correlations between these physicochemical properties and enzymatic hydrolysis yield were investigated. Gel permeation chromatography (GPC) and 13C solid state NMR were used to determine the degree of polymerization (DP) and crystallinity index (CrI) of cellulose, and the results along with the sugar release study indicated that cellulose DP likely played a more important role in enzymatic hydrolysis. Simons’ stain revealed that the accessible surface area of substrate significantly varied among these variants from 17.3 to 33.2 mg gmore » $$–1\\atop{biomass}$$ as reflected by dye adsorption, and cellulose accessibility was shown as one of the major factors governing substrates digestibility. HSQC and 31P NMR analysis detailed the structural features of poplar lignin variants. Overall, cellulose relevant factors appeared to have a stronger correlation with glucose release, if any, than lignin structural features. Lignin structural features, such as a phenolic hydroxyl group and the ratio of syringyl and guaiacyl (S/G), were found to have a more convincing impact on xylose release. Low lignin content, low cellulose DP, and high cellulose accessibility generally favor enzymatic hydrolysis; however, recalcitrance cannot be simply judged on any single substrate factor.« less

An in-depth understanding of biomass recalcitrance using natural poplar variants as the feedstock

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

Meng, Xianzhi; Pu, Yunqiao; Yoo, Chang Geun

Here, in an effort to better understand the biomass recalcitrance, six natural poplar variants were selected as feedstocks based on previous sugar release analysis. Compositional analysis and physicochemical characterizations of these poplars were performed and the correlations between these physicochemical properties and enzymatic hydrolysis yield were investigated. Gel permeation chromatography (GPC) and 13C solid state NMR were used to determine the degree of polymerization (DP) and crystallinity index (CrI) of cellulose, and the results along with the sugar release study indicated that cellulose DP likely played a more important role in enzymatic hydrolysis. Simons’ stain revealed that the accessible surface area of substrate significantly varied among these variants from 17.3 to 33.2 mg gmore » $$–1\\atop{biomass}$$ as reflected by dye adsorption, and cellulose accessibility was shown as one of the major factors governing substrates digestibility. HSQC and 31P NMR analysis detailed the structural features of poplar lignin variants. Overall, cellulose relevant factors appeared to have a stronger correlation with glucose release, if any, than lignin structural features. Lignin structural features, such as a phenolic hydroxyl group and the ratio of syringyl and guaiacyl (S/G), were found to have a more convincing impact on xylose release. Low lignin content, low cellulose DP, and high cellulose accessibility generally favor enzymatic hydrolysis; however, recalcitrance cannot be simply judged on any single substrate factor.« less

Utilization of fish meal and fish oil for production of Cryptococcus sp. MTCC 5455 lipase and hydrolysis of polyurethane thereof.

PubMed

Thirunavukarasu, K; Purushothaman, S; Gowthaman, M K; Nakajima-Kambe, T; Rose, C; Kamini, N R

2015-09-01

Fish meal has been used as an additional nitrogen source and fish oil as inducer for the growth and production of lipase from Cryptococcus sp. MTCC 5455. A response surface design illustrated that the optimum factors influencing lipase production were fish meal, 1.5 %, w/v, Na2HPO4, 0.2 %, w/v, yeast extract, 0.25 %, w/v and sardine oil, 2.0 %, w/v with an activity of 71.23 U/mL at 96 h and 25 °C, which was 48.39 % higher than the conventional one-factor-at-a-time method. The crude concentrated enzyme hydrolyzed polyurethane (PUR) efficiently and hydrolysis was 94 % at 30 °C and 96 h. The products, diethylene glycol and adipic acid were quantified by HPLC and scanning electron microscopic studies of the degraded polymer showed significant increase in size of the holes from 24 to 72 h of incubation. Hydrolysis of PUR within 96 h makes the lipase novel for disposal of PUR and provides an innovative solution to the problems created by plastic wastes.

Hydrolysis and acidification of agricultural waste in a non-airtight system: Effect of solid content, temperature, and mixing mode.

PubMed

Yu, Jiadong; Zhao, Yubin; Zhang, Huan; Hua, Binbin; Yuan, Xufeng; Zhu, Wanbin; Wang, Xiaofen; Cui, Zongjun

2017-01-01

A two-phase digestion system for treating agricultural waste is beneficial for methane production. This study explored the effect of solid content, temperature, and mixing mode on the process of hydrolysis and acidification using rice straw and cow dung launched in non-airtight acidogenic system. The results showed that the substrate could be hydrolyzed efficiently in the initial stage, the hydrolysis coefficient (k) of maximum cellulose and hemicellulose can be increased by 217.9% and 290.5%, respectively, compared with those of middle and last stages. High solid content played a leading role in promoting hydrolysis, resulted in hydrolysate content (sCOD) that was significantly higher than in treatments with low solid content (P<0.01), and led to organic acids accumulation up to 5.8 and 6.7g/L at mesophilic and thermophilic temperatures. Thermophilic temperature stimulated the hydrolysis and acidification of low solid content (P<0.05), and improved organic acid accumulation of high solid content only during the middle stage (P<0.01). Mixing mode was not a major factor, but increasing the mixing time was necessary for organic acid accumulation during the last stage (P<0.05). In addition, the study comprehensively analyzed a series of corresponding relationships among each operating parameter during the whole treatment process using canonical correspondence analysis. Copyright © 2016 Elsevier Ltd. All rights reserved.

In vitro and in vivo evaluation of the metabolism and pharmacokinetics of sebacoyl dinalbuphine.

PubMed

Pao, Li-Heng; Hsiong, Cheng-Huei; Hu, Oliver Yoa-Pu; Wang, Jhi-Jung; Ho, Shung-Tai

2005-03-01

A diester prodrug of nalbuphine, sebacoyl dinalbuphine (SDN), and its long-acting formulation are currently being developed to prolong the duration of nalbuphine. A comparative in vitro hydrolysis study was conducted for SDN in rat, rabbit, dog, and human blood. Both SDN and nalbuphine in blood or plasma were measured by high-performance liquid chromatography. The hydrolysis rates of SDN in blood were ranked as follows: rat > rabbit > human > dog. The rapid formation of nalbuphine in the blood accounted for almost 100% of the prodrug, which supported the contention that nalbuphine is the major metabolite after SDN hydrolysis. The hydrolysis profiles of SDN were similar both in plasma and in red blood cells when compared in the blood. In vitro release results of SDN long-acting formulation showed that the rate-limited step of SDN hydrolysis to nalbuphine in blood is the penetration of SDN from oil into the blood. After intravenous administration of SDN in sesame oil into rats, nalbuphine quickly appeared in plasma and, thereafter, exhibited monoexponential decay. Pharmaceutical dosage forms affecting the drug disposition kinetics were demonstrated after intravenous administration. The AUC of nalbuphine was significantly higher and clearance was significantly lower, without changes in the t(1/2) of nalbuphine after intravenous dosing of SDN in sesame oil when compared with that of intravenous dosing with nalbuphine HCl in rats. Overall, these results suggest that SDN fulfilled the original pro-soft drug design in which the prodrug can rapidly metabolize to nalbuphine, and no other unexpected compounds were apparent in the blood.

Toxicovigilance: new biochemical tool used in sulfonylurea herbicides toxicology studies.

PubMed

Belhadj-Tahar, Hafid; Adamczewski, Nicolas; Nassar, Bertrand; Coulais, Yvon

2003-06-01

In vitro toxic effects of sulfonylurea herbicides (thifensulfuron-methyl and metsulfuron-methyl) were evaluated according to a new protocol. Physiological conditions were reproduced in order to boost toxicovigilance. Sulfonylureas and their hydrolysis products were added to biological substrates such as urea, alanine, aspartic acid, alpha-ketoglutarate, oxaloacetate, pyruvate and then incubated with some specific enzymes. Addition of these sulfonylureas and their degradation products did not significantly change the enzymatic activity of the urease, aspartate-aminotransferase, glutamate dehydrogenase, malate dehydrogenase and lactate dehydrogenase. However, the acid hydrolysis products inhibited up to 95% of the activity of the alanine-aminotransferase at low concentrations (0.27 micromol L(-1)). Inhibition did not affect the mitochondrial aspartate-aminotransferase.

Secretome analysis of Trichoderma reesei and Aspergillus niger cultivated by submerged and sequential fermentation processes: Enzyme production for sugarcane bagasse hydrolysis.

PubMed

Florencio, Camila; Cunha, Fernanda M; Badino, Alberto C; Farinas, Cristiane S; Ximenes, Eduardo; Ladisch, Michael R

2016-08-01

Cellulases and hemicellulases from Trichoderma reesei and Aspergillus niger have been shown to be powerful enzymes for biomass conversion to sugars, but the production costs are still relatively high for commercial application. The choice of an effective microbial cultivation process employed for enzyme production is important, since it may affect titers and the profile of protein secretion. We used proteomic analysis to characterize the secretome of T. reesei and A. niger cultivated in submerged and sequential fermentation processes. The information gained was key to understand differences in hydrolysis of steam exploded sugarcane bagasse for enzyme cocktails obtained from two different cultivation processes. The sequential process for cultivating A. niger gave xylanase and β-glucosidase activities 3- and 8-fold higher, respectively, than corresponding activities from the submerged process. A greater protein diversity of critical cellulolytic and hemicellulolytic enzymes were also observed through secretome analyses. These results helped to explain the 3-fold higher yield for hydrolysis of non-washed pretreated bagasse when combined T. reesei and A. niger enzyme extracts from sequential fermentation were used in place of enzymes obtained from submerged fermentation. An enzyme loading of 0.7 FPU cellulase activity/g glucan was surprisingly effective when compared to the 5-15 times more enzyme loadings commonly reported for other cellulose hydrolysis studies. Analyses showed that more than 80% consisted of proteins other than cellulases whose role is important to the hydrolysis of a lignocellulose substrate. Our work combined proteomic analyses and enzymology studies to show that sequential and submerged cultivation methods differently influence both titers and secretion profile of key enzymes required for the hydrolysis of sugarcane bagasse. The higher diversity of feruloyl esterases, xylanases and other auxiliary hemicellulolytic enzymes observed in the enzyme mixtures from the sequential fermentation could be one major reason for the more efficient enzyme hydrolysis that results when using the combined secretomes from A. niger and T. reesei. Copyright © 2016 Elsevier Inc. All rights reserved.

Nucleotide Catabolism on the Surface of Aortic Valve Xenografts; Effects of Different Decellularization Strategies.

PubMed

Kutryb-Zajac, Barbara; Yuen, Ada H Y; Khalpey, Zain; Zukowska, Paulina; Slominska, Ewa M; Taylor, Patricia M; Goldstein, Steven; Heacox, Albert E; Lavitrano, Marialuisa; Chester, Adrian H; Yacoub, Magdi H; Smolenski, Ryszard T

2016-04-01

Extracellular nucleotide metabolism controls thrombosis and inflammation and may affect degeneration and calcification of aortic valve prostheses. We evaluated the effect of different decellularization strategies on enzyme activities involved in extracellular nucleotide metabolism. Porcine valves were tested intact or decellularized either by detergent treatment or hypotonic lysis and nuclease digestion. The rates of ATP hydrolysis, AMP hydrolysis, and adenosine deamination were estimated by incubation of aorta or valve leaflet sections with substrates followed by HPLC analysis. We demonstrated relatively high activities of ecto-enzymes on porcine valve as compared to the aortic wall. Hypotonic lysis/nuclease digestion preserved >80 % of ATP and AMP hydrolytic activity but reduced adenosine deamination to <10 %. Detergent decellularization completely removed (<5 %) all these activities. These results demonstrate high intensity of extracellular nucleotide metabolism on valve surface and indicate that various valve decellularization techniques differently affect ecto-enzyme activities that could be important in the development of improved valve prostheses.

Effects of ethylene oxide sterilization on 82: 18 PLLA/PGA copolymer craniofacial fixation plates.

PubMed

Pietrzak, William S

2010-01-01

Bioabsorbable devices are generally susceptible to some form of degradation or alteration of material properties in response to exposure to the terminal sterilization cycle. In addition to affecting the material strength, sterilization can also increase the rate of hydrolysis, both of which can impact clinical performance. The impact of sterilization on the material/device is unpredictable and must be empirically determined. This study examined the effects of ethylene oxide treatment on the material properties of LactoSorb 82:18 poly(L-lactic acid)-poly(glycolic acid) craniofacial plates. Compared with untreated control plates, there was no effect on the initial inherent viscosity (1.3 dL/g), the glass transition temperature (58 degrees C), or on the flexural mechanical properties. Furthermore, there was no effect on the in vitro rate of hydrolysis and mechanical strength loss profile. This provides evidence that the ethylene oxide sterilization cycle is compatible with these copolymer plates and that such treatment should not affect the clinical performance.

Use of enzyme inhibitors to evaluate the conversion pathways of ester and amide prodrugs: a case study example with the prodrug ceftobiprole medocaril.

PubMed

Eichenbaum, Gary; Skibbe, Jennifer; Parkinson, Andrew; Johnson, Mark D; Baumgardner, Dawn; Ogilvie, Brian; Usuki, Etsuko; Tonelli, Fred; Holsapple, Jeff; Schmitt-Hoffmann, Anne

2012-03-01

An approach was developed that uses enzyme inhibitors to support the assessment of the pathways that are responsible for the conversion of intravenously administered ester and amide prodrugs in different biological matrices. The methodology was applied to ceftobiprole medocaril (BAL5788), the prodrug of the cephalosporin antibiotic, ceftobiprole. The prodrug was incubated in plasma, postmitochondrial supernatant fractions from human liver (impaired and nonimpaired), kidney, and intestine as well as erythrocytes, in the presence and absence of different enzyme inhibitors (acetylcholinesterase, pseudocholinesterase, retinyl palmitoyl hydrolase, serine esterases, amidases, and cholinesterase). Hydrolysis was rapid, extensive, and not dependent on the presence of β-nicotinamide-adenine dinucleotide phosphate (reduced form) in all matrices tested, suggesting the involvement of carboxylesterases but not P450 enzymes. Hydrolysis in healthy human plasma was rapid and complete and only partially inhibited in the presence of paraoxonase inhibitors or in liver from hepatic impaired patients, suggesting involvement of nonparaoxonase pathways. The results demonstrate the utility of this approach in confirming the presence of multiple conversion pathways of intravenously administered prodrugs and in the case of BAL5788 demonstrated that this prodrug is unlikely to be affected by genetic polymorphisms, drug interactions, or other environmental factors that might inhibit or induce the enzymes involved in its conversion. Copyright © 2011 Wiley Periodicals, Inc.

Improved sludge dewaterability and hydrolysis performance after pretreatment with Fenton's reagent.

PubMed

Yuan, Hongying; Yang, Yuping; Yuan, Jian; Wang, Yanning; Song, Yameng; Lu, Jingfang; Song, Jianyang

2018-01-01

The dewaterability of excess sludge significantly improved upon pretreatment with Fenton's reagent in this study. After 0.9 g/L of Fe 2+ and 5.0 g/L of H 2 O 2 were added to the sludge, and reacted for 2 h at pH = 4, the specific resistance to filtration (SRF) of the excess sludge decreased from an initial value of 29.74 × 10 12 m/kg to 6.49 × 10 12 m/kg. The factors that affected this improvement in sludge dewaterability as evaluated by SRF reduction showed the following order: H 2 O 2 > pH > Fe 2+ > reaction time. Furthermore, the hydrolysis performance of the sludge under the optimal reaction conditions was investigated. The results indicated that the concentration of soluble chemical oxygen demand in the supernatant increased almost 14 times compared to raw sludge, and the contents of soluble protein and soluble polysaccharide were more than 8 and 17 times higher, respectively, than for the untreated situation. However, the amounts of ammonia nitrogen (NH 4 + -N) and phosphate (PO 4 3- -P) released from the sludge showed different trends: NH 4 + -N increased by 200%, while PO 4 3- -P decreased by 82%. The production of volatile fatty acids (VFAs) from the treated sludge showed that total VFAs increased by 66%, and iso-butylacetic acid was the dominant product among the total VFAs.

Cadmium effects on embryo growth of pea seeds during germination: Investigation of the mechanisms of interference of the heavy metal with protein mobilization-related factors.

PubMed

Jaouani, Khadija; Karmous, Inès; Ostrowski, Maciej; Ferjani, Ezzedine El; Jakubowska, Anna; Chaoui, Abdelilah

2018-04-16

This work aims to give more insight into mechanisms of action of cadmium (Cd) on germinating pea seeds (Pisum sativum L. var. douce province), specifically the different ways by which Cd cations may interfere with the principal factors involved during germination process, notably storage proteins mobilization, amino acids freeing and proteolytic activities. Obtained results revealed that the process of hydrolysis of main storage proteins showed a significant disruption, which resulted in the decrease of the release of free amino acids, thus imposing a lack in nitrogen supply of essential nutrients to growing embryo under Cd stress. This hypothesis was evidenced by Cd-induced changes occurring in main purified protein fractions; Albumins, Legumins and Vicilins, during their breakdown. Besides, at enzymatic level, the activities of main proteases responsible for this hydrolysis were altered. Indeed, assays using synthetic substrates and specific protease inhibitors followed by protease activity measurements demonstrated that Cd inhibited drastically the total azocaseinolytic activity (ACA) and activities of different proteolytic classes: cysteine-, aspartic-, serine- and metallo-endopeptidases (EP), leucine- and proline-aminopeptidases (LAP and PAP, respectively), and glycine-carboxypeptidases (Gly-CP). The data here presented may suggest that the vulnerability of the embryonic axes towards Cd toxicity could be explained as a result of eventual disruption of metabolic pathways that affect mobilization of reserves and availability of nutrients. In vitro studies suggest that Cd cations may act either directly on the catalytic sites of the proteolytic enzymes, which may cause their deactivation, or indirectly via the generation of oxidative stress and overproduction of free radicals that can interact with enzymes, by altering their activity and structure. Copyright © 2018 Elsevier GmbH. All rights reserved.

Determination of genotoxic effects of methidathion alkaline hydrolysis in human lymphocytes using the micronucleus assay and square-wave voltammetry.

PubMed

Stivaktakis, Polychronis D; Giannakopoulos, Evangelos; Vlastos, Dimitris; Matthopoulos, Demetrios P

2017-02-01

The interaction of pesticides with environmental factors, such as pH, may result in alterations of their physicochemical properties and should be taken into consideration in regard to their classification. This study investigates the genotoxicity of methidathion and its alkaline hydrolysis by-products in cultured human lymphocytes, using the square-wave voltammetry (square wave-adsorptive cathodic stripping voltammetry (SW-AdCSV) technique) and the cytokinesis block micronucleus assay (CBMN assay). According to the SW-AdCSV data the alkaline hydrolysis of methidathion results in two new molecules, one non-electro-active and a second electro-active which is more genotoxic than methidathion itself in cultured human lymphocytes, inducing higher micronuclei frequencies. The present study confirms the SW-AdCSV technique as a voltammetric method which can successfully simulates the electrodynamics of the cellular membrane. Copyright © 2016 Elsevier B.V. All rights reserved.

Surface characterization studies of walnut-shell biochar catalysts for simultaneously removing of organic sulfur from yellow phosphorus tail gas

NASA Astrophysics Data System (ADS)

Song, Xin; Li, Kai; Ning, Ping; Wang, Chi; Sun, Xin; Tang, Lihong; Ruan, Haotian; Han, Shuang

2017-12-01

The influences of different preparation conditions for surface characteristics on removing organic sulfur were studied. From BET, XRD, FTIR, DRIFTS, TG/DTA, CO2-TPD results, it can be seen that these preparation conditions had great influences on the pore structure, specific surface area, crystal structure and surface functional groups. The micropore volume, amorphous structure and alkalinity site strength played major roles in desulfurization process. H2S was oxidized by oxygen containing functional groups, such as sbnd COO, sbnd Cdbnd O. H2O molecule could be converted into some groups, such as sbnd CH and Csbnd OH groups, and promoted the hydrolysis reaction. The strong alkalinity site was the key factor for chemical adsorption and hydrolysis. H2O molecule, sbnd CH, Csbnd OH groups promoted the hydrolysis reaction and sbnd COO, sbnd Cdbnd O groups promoted the oxidation of H2S on the surface of WSB. Meanwhile, the main desulfurization process over WSB after carbonization was adsorption and it changed to hydrolysis reaction after activation on the surface of WSB. Furthermore, the reaction mechanism was investigated by DRIFTS measurement according to the change of surface functional groups.

Arrhenius equation for modeling feedyard ammonia emissions using temperature and diet crude protein

USDA-ARS?s Scientific Manuscript database

Temperature controls many processes of ammonia volatilization. For example, urea hydrolysis is an enzymatically catalyzed reaction described by the Arrhenius equation. Diet crude protein (CP) controls ammonia emission by affecting N excretion. Objectives were to use the Arrhenius equation to model a...

AGE-DEPENDENT EFFECTS OF AROCLOR 1254 ON CALCIUM UPTAKE BY SUBCELLULAR ORGANELLES IN SELECTED BRAIN REGIONS OF RATS.

EPA Science Inventory

Earlier reports from our laboratory have indicated that polychlorinated biphenyls (PCBs) affect signal transduction mechanisms in brain, including Ca2+ homeostasis, phosphoinositol hydrolysis, and protein kinase C (PKC) translocation in mature neurons and adult brain homogenate p...

Effects of ionic strength, temperature, and pH on degradation of selected antibiotics

USGS Publications Warehouse

Loftin, K.A.; Adams, C.D.; Meyer, M.T.; Surampalli, R.

2008-01-01

Aqueous degradation rates, which include hydrolysis and epimerization, for chlorretracycline (CTC), oxytetracycline (OTC), tetracycline (TET), lincomycin (LNC), sulfachlorpyridazine (SCP), sulfadimethoxine (SDM), sulfathiazole (STZ), trimethoprim (TRM), and tylosin A (TYL) were studied as a function of ionic strength (0.0015, 0.050, or 0.084 mg/L as Na2HPO4), temperature (7, 22, and 35??C), and pH (2, 5, 7, 9, and 11). Multiple linear regression revealed that ionic strength did not significantly affect (?? = 0.05) degradation rates for all compounds, but temperature and pH affected rates for CTC, OTC, and TET significandy (?? = 0.05). Degradation also was observed for TYL at pH 2 and 11. No significant degradation was observed for LNC, SCR SDM, STZ, TRM, and TYL (pH 5, 7, and 9) under study conditions. Pseudo first-order rate constants, half-lives, and Arrhenius coefficients were calculated where appropriate. In general, hydrolysis rates for CTC, OTC, and TET increased as pH and temperature increased following Arrhenius relationships. Known degradation products were used to confirm that degradation had occurred, but these products were not quantified. Half-lives ranged from less than 6 h up to 9.7 wk for the tetracyclines and for TYL (pH 2 and 11), but no degradation of LIN, the sulfonamides, or TRM was observed during the study period. These results indicate that tetracyclines and TYL at pH 2 and 11 are prone to pH-mediated transformation and hydrolysis in some cases, but not the sulfonamides, LIN nor TRM are inclined to degrade under study conditions. This indicates that with the exception of CTC OTC, and TET, pH-mediated reactions such as hydrolysis and epimerization are not likely removal mechanisms in surface water, anaerobic swine lagoons, wastewater, and ground water. Copyright ?? 2008 by the American Society of Agronomy, Crop Science Society of America, and Soil Science Society of America. All rights reserved.

Collagen extraction from mussel byssus: a new marine collagen source with physicochemical properties of industrial interest.

PubMed

Rodríguez, F; Morán, L; González, G; Troncoso, E; Zúñiga, R N

2017-04-01

Mussel byssus is a by-product of mussel production and is a potential source of collagen. The goal of this study was to extract collagen from the byssus of Chilean mussel using an enzymatic method and characterize it. A pepsin-aided extraction method was employed where first an enzymatic hydrolysis at two pepsin/substrate ratios (1:50 or 4:50) and times (4 or 24 h) was done. Extraction was conducted at 80 °C for 24 h, in a 0.5 N acetic acid solution. All samples were analyzed for collagen content, amino acid profile, turbidity, viscosity, solubility, denaturation temperature and surface tension. Hydrolysis time had significant effect on collagen content, hydroxyproline content and extraction yield. Hydrolysis with a pepsin/byssus ratio of 4:50 for 24 h gave the better extraction performance with values of 69 mg/g protein, 1.8 mg/g protein and 30%, for collagen content, hydroxyproline content and extraction yield, respectively. No differences were found for the viscosity and surface tension of collagen dispersions, suggesting that the enzymatic hydrolysis did not affect the integrity of the collagen molecule. Denaturation temperature of freeze-dried byssus collagen presented a high value (83-91 °C), making this kind of collagen a very interesting material for encapsulation of bioactive molecules and for biomedical applications.

Optimization of a thermal hydrolysis process for sludge pre-treatment.

PubMed

Sapkaite, I; Barrado, E; Fdz-Polanco, F; Pérez-Elvira, S I

2017-05-01

At industrial scale, thermal hydrolysis is the most used process to enhance biodegradability of the sludge produced in wastewater treatment plants. Through statistically guided Box-Behnken experimental design, the present study analyses the effect of TH as pre-treatment applied to activated sludge. The selected process variables were temperature (130-180 °C), time (5-50 min) and decompression mode (slow or steam-explosion effect), and the parameters evaluated were sludge solubilisation and methane production by anaerobic digestion. A quadratic polynomial model was generated to compare the process performance for the 15 different combinations of operation conditions by modifying the process variables evaluated. The statistical analysis performed exhibited that methane production and solubility were significantly affected by pre-treatment time and temperature. During high intensity pre-treatment (high temperature and long times), the solubility increased sharply while the methane production exhibited the opposite behaviour, indicating the formation of some soluble but non-biodegradable materials. Therefore, solubilisation is not a reliable parameter to quantify the efficiency of a thermal hydrolysis pre-treatment, since it is not directly related to methane production. Based on the operational parameters optimization, the estimated optimal thermal hydrolysis conditions to enhance of sewage sludge digestion were: 140-170 °C heating temperature, 5-35min residence time, and one sudden decompression. Copyright © 2017 Elsevier Ltd. All rights reserved.

Modeling the Effect of pH and Temperature for Cellulases Immobilized on Enzymogel Nanoparticles.

PubMed

Samaratunga, Ashani; Kudina, Olena; Nahar, Nurun; Zakharchenko, Andrey; Minko, Sergiy; Voronov, Andriy; Pryor, Scott W

2015-06-01

Production costs of cellulosic biofuels can be lowered if cellulases are recovered and reused using particulate carriers that can be extracted after biomass hydrolysis. Such enzyme recovery was recently demonstrated using enzymogel nanoparticles with grafted polymer brushes loaded with cellulases. In this work, cellulase (NS50013) and β-glucosidase (Novozyme 188) were immobilized on enzymogels made of poly(acrylic acid) polymer brushes grafted to the surface of silica nanoparticles. Response surface methodology was used to model effects of pH and temperature on hydrolysis and recovery of free and attached enzymes. Hydrolysis yields using both enzymogels and free cellulase and β-glucosidase were highest at the maximum temperature tested, 50 °C. The optimal pH for cellulase enzymogels and free enzyme was 5.0 and 4.4, respectively, while both free β-glucosidase and enzymogels had an optimal pH near 4.4. Highest hydrolysis sugar concentrations with cellulase and β-glucosidase enzymogels were 69 and 53 % of those with free enzymes, respectively. Enzyme recovery using enzymogels decreased with increasing pH, but cellulase recovery remained greater than 88 % throughout the operating range of pH values less than 5.0 and was greater than 95 % at pH values below 4.3. Recovery of β-glucosidase enzymogels was not affected by temperature and had little impact on cellulase recovery.

Physicochemical properties of cross-linked and acetylated starches and products of their hydrolysis in continuous recycle membrane reactor.

PubMed

Prochaska, Krystyna; Konował, Emilia; Sulej-Chojnacka, Joanna; Lewandowicz, Grazyna

2009-11-01

The aim of the present work was to study the physicochemical properties of doubly modified, by cross-linking and acetylating, starches as well as the products of their enzymatic hydrolysis. A two step procedure of hydrolysis, including the batch and membrane reactors, were investigated. The second step of enzymatic processes were carried out in a continuous recycle membrane reactor (CRMR). Three kinds of commercial starches--two preparations of acetylated distarch adipate E1422 of different degrees of cross-linking, as well as one preparation of acetylated distarch phosphate E1414 were examined. It was found that the degree of substitution of acetyl groups in the macromolecules of starch did not influence the effectiveness of hydrolysis. However, the degree of cross-linking with adipate groups slightly decreased the efficiency of processing in the CRMR. Additionally, the relationship between the type of hydrocolloid and its adsorption activity in the air/water and oil/water systems was considered. All obtained derivatives revealed adsorption properties and reduced the surface/interface tension in the air/water and oil/water systems. The efficiency and effectiveness of adsorption of the investigated hydrocolloids were affected by the type of modification as well as the degree of substitution of acetyl groups in the macromolecules of starch. Particle size distributions formed in aqueous solutions for all investigated hydrolyses were determined and compared with results obtained for commercial products.

Particulate Size of Microalgal Biomass Affects Hydrolysate Properties and Bioethanol Concentration

PubMed Central

Harun, Razif; Danquah, Michael K.; Thiruvenkadam, Selvakumar

2014-01-01

Effective optimization of microalgae-to-bioethanol process systems hinges on an in-depth characterization of key process parameters relevant to the overall bioprocess engineering. One of the such important variables is the biomass particle size distribution and the effects on saccharification levels and bioethanol titres. This study examined the effects of three different microalgal biomass particle size ranges, 35 μm ≤ x ≤ 90 μm, 125 μm ≤ x ≤ 180 μm, and 295 μm ≤ x ≤ 425 μm, on the degree of enzymatic hydrolysis and bioethanol production. Two scenarios were investigated: single enzyme hydrolysis (cellulase) and double enzyme hydrolysis (cellulase and cellobiase). The glucose yield from biomass in the smallest particle size range (35 μm ≤ x ≤ 90 μm) was the highest, 134.73 mg glucose/g algae, while the yield from biomass in the larger particle size range (295 μm ≤ x ≤ 425 μm) was 75.45 mg glucose/g algae. A similar trend was observed for bioethanol yield, with the highest yield of 0.47 g EtOH/g glucose obtained from biomass in the smallest particle size range. The results have shown that the microalgal biomass particle size has a significant effect on enzymatic hydrolysis and bioethanol yield. PMID:24971327

Hydrolysis of Glycosidic Flavonoids during the Preparation of Danggui Buxue Tang: An Outcome of Moderate Boiling of Chinese Herbal Mixture

PubMed Central

Zhang, Wendy Li; Chen, Jian-Ping; Lam, Kelly Yin-Ching; Zhan, Janis Ya-Xian; Yao, Ping; Dong, Tina Ting-Xia; Tsim, Karl Wah-Keung

2014-01-01

Chemical change during boiling of herbal mixture is a puzzle. By using Danggui Buxue Tang (DBT), a herbal decoction that contains Astragali Radix (AR) and Angelicae Sinensis Radix (ASR), we developed a model in analyzing the hydrolysis of flavonoid glycosides during the boiling of herbal mixture in water. A proper preparation of DBT is of great benefit to the complete extraction of bioactive ingredients. Boiling of DBT in water increased the solubility of AR-derived astragaloside IV, calycosin, formononetin, calycosin-7-O-β-D-glucoside, and ononin in a time- and temperature-dependent manner: the amounts of these chemicals reached a peak at 2 h. The glycosidic resides of AR, calycosin-7-O-β-D-glucoside, and ononin could be hydrolyzed during the moderate boiling process to form calycosin and formononetin, respectively. The hydrolysis efficiency was strongly affected by pH, temperature, and amount of herbs. Interestingly, the preheated herbs were not able to show this hydrolytic activity. The current results supported the rationality of ancient preparation of DBT in boiling water by moderate heat. PMID:24744813

Graphene oxide/MnO2 nanocomposite as destructive adsorbent of nerve-agent simulants in aqueous media

NASA Astrophysics Data System (ADS)

Šťastný, Martin; Tolasz, Jakub; Štengl, Václav; Henych, Jiří; Žižka, David

2017-08-01

Graphene oxide/MnO2 nanocomposite was prepared by thermal hydrolysis of potassium permanganate (KMnO4) and 2-chloroacetamide aqueous solutions with graphene oxide (GO) suspension. The synthesized samples were characterized by specific surface area (BET) and porosity determination (BJH), X-ray Diffraction (XRD) and high-resolution electron microscopes (HRSEM, HRTEM). These nanocomposites were used in an experimental evaluation of their adsorption activity with nerve agent simulants dimethyl methyl phosphonate (DMMP) and triethyl phosphate (TEP) in aqueous media. The nanocomposites exhibited enhanced adsorptive degradation ability compared to pure manganese oxide (MnO2) and GO. The GO amount in the nanocomposites affected their degradation activity substantially. The best adsorption efficiency was observed for samples with moderate GO amount. Three methods were used to observe the mechanism of the nerve-agent simulants deactivation: Gas chromatography with mass spectrometry (GC-MS), High-Performance Liquid Chromatography (HPLC) and in situ Infrared spectroscopy (FTIR). It was shown that the hydrolysis on the surface of prepared nanocomposites yields volatile primary alcohols (methanol and ethanol) as the main hydrolysis products.

Pretreating wheat straw by phosphoric acid plus hydrogen peroxide for enzymatic saccharification and ethanol production at high solid loading.

PubMed

Qiu, Jingwen; Ma, Lunjie; Shen, Fei; Yang, Gang; Zhang, Yanzong; Deng, Shihuai; Zhang, Jing; Zeng, Yongmei; Hu, Yaodong

2017-08-01

Wheat straw was pretreated by phosphoric acid plus hydrogen peroxide (PHP) for enzymatic hydrolysis and ethanol fermentation at high solid loadings. Results indicated solid loading could reach 20% with 77.4% cellulose-glucose conversion and glucose concentration of 164.9g/L in hydrolysate, it even was promoted to 25% with only 3.4% decrease on cellulose-glucose conversion as the pretreated-wheat straw was dewatered by air-drying. 72.9% cellulose-glucose conversion still was achieved as the minimized enzyme input of 20mg protein/g cellulose was employed for hydrolysis at 20% solid loading. In the corresponding conditions, 100g wheat straw can yield 11.2g ethanol with concentration of 71.2g/L by simultaneous saccharification and fermentation. Thus, PHP-pretreatment benefitted the glucose or ethanol yield at high solid loadings with lower enzyme input. Additionally, decreases on the maximal cellulase adsorption and the direct-orange/direct-blue indicated drying the PHP-pretreated substrates negatively affected the hydrolysis due to the shrinkage of cellulase-size-accommodable pores. Copyright © 2017 Elsevier Ltd. All rights reserved.

Further analysis of the structure and immunological activity of an RG-I type pectin from Panax ginseng.

PubMed

Zhang, Xu; Li, Shanshan; Sun, Lin; Ji, Li; Zhu, Jingjing; Fan, Yuying; Tai, Guihua; Zhou, Yifa

2012-06-20

In this paper, we further analysed the structure of a type I rhamnogalacturonan (RG-I) pectin (WGPA-2-RG) fractionated from ginseng polysaccharides. Methylation and periodate oxidation analyses showed that WGPA-2-RG has a backbone consisting of alternating rhamnose (Rha) and galacturonic acid (GalA) residues and side chains consisting of type II arabinogalactan (AG-II). Partial acidic hydrolysis for 6h completely removed arabinose (Ara), partial galactose (Gal), but little GalA and Rha. During partial hydrolysis, the molecular weight of WGPA-2-RG decreased smoothly, suggesting that the Ara and cleavable Gal residues exist on the surface of the molecule, while GalA and Rha residues exist in the core of the molecule. The bioactivity assay showed that the arabinogalactan side chains of WGPA-2-RG are essential structures for stimulating NO secretion and lymphocyte proliferation. However, removal of the Ara and Gal residues through hydrolysis did not appreciably affect the ability of WGPA-2-RG to enhance macrophage phagocytosis. Copyright © 2012 Elsevier Ltd. All rights reserved.

Structure of the protein core of translation initiation factor 2 in apo, GTP-bound and GDP-bound forms

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

Simonetti, Angelita; Marzi, Stefano; Fabbretti, Attilio

2013-06-01

The crystal structures of the eubacterial translation initiation factor 2 in apo form and with bound GDP and GTP reveal conformational changes upon nucleotide binding and hydrolysis, notably of the catalytically important histidine in the switch II region. Translation initiation factor 2 (IF2) is involved in the early steps of bacterial protein synthesis. It promotes the stabilization of the initiator tRNA on the 30S initiation complex (IC) and triggers GTP hydrolysis upon ribosomal subunit joining. While the structure of an archaeal homologue (a/eIF5B) is known, there are significant sequence and functional differences in eubacterial IF2, while the trimeric eukaryotic IF2more » is completely unrelated. Here, the crystal structure of the apo IF2 protein core from Thermus thermophilus has been determined by MAD phasing and the structures of GTP and GDP complexes were also obtained. The IF2–GTP complex was trapped by soaking with GTP in the cryoprotectant. The structures revealed conformational changes of the protein upon nucleotide binding, in particular in the P-loop region, which extend to the functionally relevant switch II region. The latter carries a catalytically important and conserved histidine residue which is observed in different conformations in the GTP and GDP complexes. Overall, this work provides the first crystal structure of a eubacterial IF2 and suggests that activation of GTP hydrolysis may occur by a conformational repositioning of the histidine residue.« less

Microbial Cellulose Utilization: Fundamentals and Biotechnology

PubMed Central

Lynd, Lee R.; Weimer, Paul J.; van Zyl, Willem H.; Pretorius, Isak S.

2002-01-01

Fundamental features of microbial cellulose utilization are examined at successively higher levels of aggregation encompassing the structure and composition of cellulosic biomass, taxonomic diversity, cellulase enzyme systems, molecular biology of cellulase enzymes, physiology of cellulolytic microorganisms, ecological aspects of cellulase-degrading communities, and rate-limiting factors in nature. The methodological basis for studying microbial cellulose utilization is considered relative to quantification of cells and enzymes in the presence of solid substrates as well as apparatus and analysis for cellulose-grown continuous cultures. Quantitative description of cellulose hydrolysis is addressed with respect to adsorption of cellulase enzymes, rates of enzymatic hydrolysis, bioenergetics of microbial cellulose utilization, kinetics of microbial cellulose utilization, and contrasting features compared to soluble substrate kinetics. A biological perspective on processing cellulosic biomass is presented, including features of pretreated substrates and alternative process configurations. Organism development is considered for “consolidated bioprocessing” (CBP), in which the production of cellulolytic enzymes, hydrolysis of biomass, and fermentation of resulting sugars to desired products occur in one step. Two organism development strategies for CBP are examined: (i) improve product yield and tolerance in microorganisms able to utilize cellulose, or (ii) express a heterologous system for cellulose hydrolysis and utilization in microorganisms that exhibit high product yield and tolerance. A concluding discussion identifies unresolved issues pertaining to microbial cellulose utilization, suggests approaches by which such issues might be resolved, and contrasts a microbially oriented cellulose hydrolysis paradigm to the more conventional enzymatically oriented paradigm in both fundamental and applied contexts. PMID:12209002

Xylanase supplementation on enzymatic saccharification of dilute acid pretreated poplars at different severities

Treesearch

Chao Zhang; Xinshu Zhuang; Zhao Jiang Wang; Fred Matt; Franz St. John; J.Y. Zhu

2013-01-01

Three pairs of solid substrates from dilute acid pretreatment of two poplar wood samples were enzymatically hydrolyzed by cellulase preparations supplemented with xylanase. Supplementation of xylanase improved cellulose saccharification perhaps due to improved cellulose accessibility by xylan hydrolysis. Total xylan removal directly affected enzymatic cellulose...

Development of non-forage based incubation system for culturing ruminal lipase-producing bacteria in vitro

USDA-ARS?s Scientific Manuscript database

Metabolism of dietary lipids within the ruminant gastrointestinal tract can differentially affect populations of gut microbes residing in the rumen or intestine. For instance, hydrolysis of dietary fats in the rumen can liberate different mixtures of free fatty acids, depending on the dietary sourc...

CAN FLUORIDATION AFFECT LEAD (II) IN POTABLE WATER? HEXAFLUOROSILICATE AND FLUORIDE EQUILIBRIA IN AQUEOUS SOLUTION

EPA Science Inventory

Recent reports have attempted to show that fluoridating potable water is linked to increased levels of lead(II) in the blood. We examine these claims in light of the established science and critically evaluate their significance. The completeness of hexafluorosilicate hydrolysi...

Rapid estimation of sugar release from winter wheat straw during bioethanol production using FTIR-photoacoustic spectroscopy

DOE PAGES

Bekiaris, Georgios; Lindedam, Jane; Peltre, Clément; ...

2015-06-18

Complexity and high cost are the main limitations for high-throughput screening methods for the estimation of the sugar release from plant materials during bioethanol production. In addition, it is important that we improve our understanding of the mechanisms by which different chemical components are affecting the degradability of plant material. In this study, Fourier transform infrared photoacoustic spectroscopy (FTIR-PAS) was combined with advanced chemometrics to develop calibration models predicting the amount of sugars released after pretreatment and enzymatic hydrolysis of wheat straw during bioethanol production, and the spectra were analysed to identify components associated with recalcitrance. A total of 1122more » wheat straw samples from nine different locations in Denmark and one location in the United Kingdom, spanning a large variation in genetic material and environmental conditions during growth, were analysed. The FTIR-PAS spectra of non-pretreated wheat straw were correlated with the measured sugar release, determined by a high-throughput pretreatment and enzymatic hydrolysis (HTPH) assay. A partial least square regression (PLSR) calibration model predicting the glucose and xylose release was developed. The interpretation of the regression coefficients revealed a positive correlation between the released glucose and xylose with easily hydrolysable compounds, such as amorphous cellulose and hemicellulose. Additionally, we observed a negative correlation with crystalline cellulose and lignin, which inhibits cellulose and hemicellulose hydrolysis. FTIR-PAS was used as a reliable method for the rapid estimation of sugar release during bioethanol production. The spectra revealed that lignin inhibited the hydrolysis of polysaccharides into monomers, while the crystallinity of cellulose retarded its hydrolysis into glucose. Amorphous cellulose and xylans were found to contribute significantly to the released amounts of glucose and xylose, respectively.« less

Hydrolysis of clavulanate by Mycobacterium tuberculosis β-lactamase BlaC harboring a canonical SDN motif.

PubMed

Soroka, Daria; Li de la Sierra-Gallay, Inès; Dubée, Vincent; Triboulet, Sébastien; van Tilbeurgh, Herman; Compain, Fabrice; Ballell, Lluis; Barros, David; Mainardi, Jean-Luc; Hugonnet, Jean-Emmanuel; Arthur, Michel

2015-09-01

Combinations of β-lactams with clavulanate are currently being investigated for tuberculosis treatment. Since Mycobacterium tuberculosis produces a broad spectrum β-lactamase, BlaC, the success of this approach could be compromised by the emergence of clavulanate-resistant variants, as observed for inhibitor-resistant TEM variants in enterobacteria. Previous analyses based on site-directed mutagenesis of BlaC have led to the conclusion that this risk was limited. Here, we used a different approach based on determination of the crystal structure of β-lactamase BlaMAb of Mycobacterium abscessus, which efficiently hydrolyzes clavulanate. Comparison of BlaMAb and BlaC allowed for structure-assisted site-directed mutagenesis of BlaC and identification of the G(132)N substitution that was sufficient to switch the interaction of BlaC with clavulanate from irreversible inactivation to efficient hydrolysis. The substitution, which restored the canonical SDN motif (SDG→SDN), allowed for efficient hydrolysis of clavulanate, with a more than 10(4)-fold increase in k cat (0.41 s(-1)), without affecting the hydrolysis of other β-lactams. Mass spectrometry revealed that acylation of BlaC and of its G(132)N variant by clavulanate follows similar paths, involving sequential formation of two acylenzymes. Decarboxylation of the first acylenzyme results in a stable secondary acylenzyme in BlaC, whereas hydrolysis occurs in the G(132)N variant. The SDN/SDG polymorphism defines two mycobacterial lineages comprising rapidly and slowly growing species, respectively. Together, these results suggest that the efficacy of β-lactam-clavulanate combinations may be limited by the emergence of resistance. β-Lactams active without clavulanate, such as faropenem, should be prioritized for the development of new therapies. Copyright © 2015, American Society for Microbiology. All Rights Reserved.

Hydrolysis of Clavulanate by Mycobacterium tuberculosis β-Lactamase BlaC Harboring a Canonical SDN Motif

PubMed Central

Soroka, Daria; Li de la Sierra-Gallay, Inès; Dubée, Vincent; Triboulet, Sébastien; van Tilbeurgh, Herman; Compain, Fabrice; Ballell, Lluis; Barros, David; Mainardi, Jean-Luc

2015-01-01

Combinations of β-lactams with clavulanate are currently being investigated for tuberculosis treatment. Since Mycobacterium tuberculosis produces a broad spectrum β-lactamase, BlaC, the success of this approach could be compromised by the emergence of clavulanate-resistant variants, as observed for inhibitor-resistant TEM variants in enterobacteria. Previous analyses based on site-directed mutagenesis of BlaC have led to the conclusion that this risk was limited. Here, we used a different approach based on determination of the crystal structure of β-lactamase BlaMAb of Mycobacterium abscessus, which efficiently hydrolyzes clavulanate. Comparison of BlaMAb and BlaC allowed for structure-assisted site-directed mutagenesis of BlaC and identification of the G132N substitution that was sufficient to switch the interaction of BlaC with clavulanate from irreversible inactivation to efficient hydrolysis. The substitution, which restored the canonical SDN motif (SDG→SDN), allowed for efficient hydrolysis of clavulanate, with a more than 104-fold increase in kcat (0.41 s−1), without affecting the hydrolysis of other β-lactams. Mass spectrometry revealed that acylation of BlaC and of its G132N variant by clavulanate follows similar paths, involving sequential formation of two acylenzymes. Decarboxylation of the first acylenzyme results in a stable secondary acylenzyme in BlaC, whereas hydrolysis occurs in the G132N variant. The SDN/SDG polymorphism defines two mycobacterial lineages comprising rapidly and slowly growing species, respectively. Together, these results suggest that the efficacy of β-lactam–clavulanate combinations may be limited by the emergence of resistance. β-Lactams active without clavulanate, such as faropenem, should be prioritized for the development of new therapies. PMID:26149997

Rapid estimation of sugar release from winter wheat straw during bioethanol production using FTIR-photoacoustic spectroscopy

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

Bekiaris, Georgios; Lindedam, Jane; Peltre, Clément

Complexity and high cost are the main limitations for high-throughput screening methods for the estimation of the sugar release from plant materials during bioethanol production. In addition, it is important that we improve our understanding of the mechanisms by which different chemical components are affecting the degradability of plant material. In this study, Fourier transform infrared photoacoustic spectroscopy (FTIR-PAS) was combined with advanced chemometrics to develop calibration models predicting the amount of sugars released after pretreatment and enzymatic hydrolysis of wheat straw during bioethanol production, and the spectra were analysed to identify components associated with recalcitrance. A total of 1122more » wheat straw samples from nine different locations in Denmark and one location in the United Kingdom, spanning a large variation in genetic material and environmental conditions during growth, were analysed. The FTIR-PAS spectra of non-pretreated wheat straw were correlated with the measured sugar release, determined by a high-throughput pretreatment and enzymatic hydrolysis (HTPH) assay. A partial least square regression (PLSR) calibration model predicting the glucose and xylose release was developed. The interpretation of the regression coefficients revealed a positive correlation between the released glucose and xylose with easily hydrolysable compounds, such as amorphous cellulose and hemicellulose. Additionally, we observed a negative correlation with crystalline cellulose and lignin, which inhibits cellulose and hemicellulose hydrolysis. FTIR-PAS was used as a reliable method for the rapid estimation of sugar release during bioethanol production. The spectra revealed that lignin inhibited the hydrolysis of polysaccharides into monomers, while the crystallinity of cellulose retarded its hydrolysis into glucose. Amorphous cellulose and xylans were found to contribute significantly to the released amounts of glucose and xylose, respectively.« less

Site-directed mutagenesis of α-L-rhamnosidase from Alternaria sp. L1 to enhance synthesis yield of reverse hydrolysis based on rational design.

PubMed

Xu, Li; Liu, Xiaohong; Yin, Zhenhao; Liu, Qian; Lu, Lili; Xiao, Min

2016-12-01

The α-L-rhamnosidase catalyzes the hydrolytic release of rhamnose from polysaccharides and glycosides and is widely used due to its applications in a variety of industrial processes. Our previous work reported that a wild-type α-L-rhamnosidase (RhaL1) from Alternaria sp. L1 could synthesize rhamnose-containing chemicals (RCCs) though reverse hydrolysis reaction with inexpensive rhamnose as glycosyl donor. To enhance the yield of reverse hydrolysis reaction and to determine the amino acid residues essential for the catalytic activity of RhaL1, site-directed mutagenesis of 11 residues was performed in this study. Through rationally designed mutations, the critical amino acid residues which may form direct or solvent-mediated hydrogen bonds with donor rhamnose (Asp 252 , Asp 257 , Asp 264 , Glu 530 , Arg 548 , His 553 , and Trp 555 ) and may form the hydrophobic pocket in stabilizing donor (Trp 261 , Tyr 302 , Tyr 316 , and Trp 369 ) in active-site of RhaL1 were analyzed, and three positive mutants (W261Y, Y302F, and Y316F) with improved product yield stood out. From the three positive variants, mutant W261Y accelerated the reverse hydrolysis with a prominent increase (43.7 %) in relative yield compared to the wild-type enzyme. Based on the 3D structural modeling, we supposed that the improved yield of mutant W261Y is due to the adjustment of the spatial position of the putative catalytic acid residue Asp 257 . Mutant W261Y also exhibited a shift in the pH-activity profile in hydrolysis reaction, indicating that introducing of a polar residue in the active site cavity may affect the catalysis behavior of the enzyme.

The catalytic effect of H2O on the hydrolysis of CO32- in hydrated clusters and its implication in the humidity driven CO2 air capture.

PubMed

Xiao, Hang; Shi, Xiaoyang; Zhang, Yayun; Liao, Xiangbiao; Hao, Feng; Lackner, Klaus S; Chen, Xi

2017-10-18

The hydration of ions in nanoscale hydrated clusters is ubiquitous and essential in many physical and chemical processes. Here we show that the hydrolysis reaction is strongly affected by relative humidity. The hydrolysis of CO 3 2- with n = 1-8 water molecules is investigated using an ab initio method. For n = 1-5 water molecules, all the reactants follow a stepwise pathway to the transition state. For n = 6-8 water molecules, all the reactants undergo a direct proton transfer to the transition state with overall lower activation free energy. The activation free energy of the reaction is dramatically reduced from 10.4 to 2.4 kcal mol -1 as the number of water molecules increases from 1 to 6. Meanwhile, the degree of hydrolysis of CO 3 2- is significantly increased compared to the bulk water solution scenario. Incomplete hydration shells facilitate the hydrolysis of CO 3 2- with few water molecules to be not only thermodynamically favorable but also kinetically favorable. We showed that the chemical kinetics is not likely to constrain the speed of CO 2 air capture driven by the humidity-swing. Instead, the pore-diffusion of ions is expected to be the time-limiting step in the humidity driven CO 2 air capture. The effect of humidity on the speed of CO 2 air capture was studied by conducting a CO 2 absorption experiment using IER with a high ratio of CO 3 2- to H 2 O molecules. Our result is able to provide valuable insights into designing efficient CO 2 air-capture sorbents.

Effects of Chemical Structure on Hydrolysis Pathways of Small Peptides in Coastal Seawater

NASA Astrophysics Data System (ADS)

Liu, S.; Reyna, N. E.; Hamdan, L. J.; Liu, Z.

2016-02-01

Deciphering peptide hydrolysis pathways is key to understanding the mechanism of peptide hydrolysis, in particular the types of extracellular enzymes that are active in seawater. From the hydrolyzed fragments of small peptides, one can estimate the role of amino-, carboxy-, and endopeptidases in a quantitative way. In this study, we incubated several small peptides with different amino acid compositions, alanine-valine-phenylalanine-alanine (AVFA), phenylalanine-alanine-serine-tryptophan-glycine-alanine (FASWGA), VFA, SWGA, VVFA, arginine-valine-phenylalanine-alanine (RVFA), SVFA, aspartic acid-valine-phenylalanine-alanine (DVFA), trialanine (AAA), and AVF in two coastal seawaters (ship channel seawater in the western Gulf of Mexico and Sta. C6 seawater in the northern Gulf of Mexico). In both seawaters, aminopeptidases played a more dominant role (22-67%) in hydrolyzing peptides with hydrophobic amino acid at the N-terminus, such as AVFA, VVFA, VFA, and AAA, or with basic amino acid at the N-terminus (RVFA), as compared to those with N-terminal polar amino acid (SVFA, SWGA) or acidic amino acid (DVFA) (0-24%). This result indicates that amino acid composition in a peptide structure affects how the peptide is hydrolyzed. We also found that peptides in the C6 seawater were hydrolyzed dominantly by aminopeptidases (10-59%), while those in the ship channel seawater also by endo- or carboxypeptidases (9-69%). This pattern suggests that peptide hydrolysis pathways depend on specific environment conditions, such as bacterial community structure, that can lead to variations in abundances or activities among amino-, carboxy- and endopeptidases. Overall, the results provide insights into the effects of chemical structure and seawater environment on peptide hydrolysis pathways.

Characterization of Clostridium difficile Spores Lacking Either SpoVAC or Dipicolinic Acid Synthetase

PubMed Central

Donnelly, M. Lauren; Fimlaid, Kelly A.

2016-01-01

ABSTRACT The spore-forming obligate anaerobe Clostridium difficile is a leading cause of antibiotic-associated diarrhea around the world. In order for C. difficile to cause infection, its metabolically dormant spores must germinate in the gastrointestinal tract. During germination, spores degrade their protective cortex peptidoglycan layers, release dipicolinic acid (DPA), and hydrate their cores. In C. difficile, cortex hydrolysis is necessary for DPA release, whereas in Bacillus subtilis, DPA release is necessary for cortex hydrolysis. Given this difference, we tested whether DPA synthesis and/or release was required for C. difficile spore germination by constructing mutations in either spoVAC or dpaAB, which encode an ion channel predicted to transport DPA into the forespore and the enzyme complex predicted to synthesize DPA, respectively. C. difficile spoVAC and dpaAB mutant spores lacked DPA but could be stably purified and were more hydrated than wild-type spores; in contrast, B. subtilis spoVAC and dpaAB mutant spores were unstable. Although C. difficile spoVAC and dpaAB mutant spores exhibited wild-type germination responses, they were more readily killed by wet heat. Cortex hydrolysis was not affected by this treatment, indicating that wet heat inhibits a stage downstream of this event. Interestingly, C. difficile spoVAC mutant spores were significantly more sensitive to heat treatment than dpaAB mutant spores, indicating that SpoVAC plays additional roles in conferring heat resistance. Taken together, our results demonstrate that SpoVAC and DPA synthetase control C. difficile spore resistance and reveal differential requirements for these proteins among the Firmicutes. IMPORTANCE Clostridium difficile is a spore-forming obligate anaerobe that causes ∼500,000 infections per year in the United States. Although spore germination is essential for C. difficile to cause disease, the factors required for this process have been only partially characterized. This study describes the roles of two factors, DpaAB and SpoVAC, which control the synthesis and release of dipicolinic acid (DPA), respectively, from bacterial spores. Previous studies of these proteins in other spore-forming organisms indicated that they are differentially required for spore formation, germination, and resistance. We now show that the proteins are dispensable for C. difficile spore formation and germination but are necessary for heat resistance. Thus, our study further highlights the diverse functions of DpaAB and SpoVAC in spore-forming organisms. PMID:27044622

Steam-exploded biomass saccharification is predominately affected by lignocellulose porosity and largely enhanced by Tween-80 in Miscanthus.

PubMed

Sun, Dan; Alam, Aftab; Tu, Yuanyuan; Zhou, Shiguang; Wang, Yanting; Xia, Tao; Huang, Jiangfeng; Li, Ying; Zahoor; Wei, Xiaoyang; Hao, Bo; Peng, Liangcai

2017-09-01

In this study, total ten Miscanthus accessions exhibited diverse cell wall compositions, leading to largely varied hexoses yields at 17%-40% (% cellulose) released from direct enzymatic hydrolysis of steam-exploded (SE) residues. Further supplied with 2% Tween-80 into the enzymatic digestion, the Mis7 accession showed the higher hexose yield by 14.8-fold than that of raw material, whereas the Mis10 had the highest hexoses yield at 77% among ten Miscanthus accessions. Significantly, this study identified four wall polymer features that negatively affect biomass saccharification as p<0.05 or 0.01 in the SE residues, including cellulose DP, Xyl and Ara of hemicellulose, and S-monomer of lignin. Based on Simons' stain, the SE porosity (defined by DY/DB) was examined to be the unique positive factor on biomass enzymatic digestion. Hence, this study provides the potential strategy to enhance biomass saccharification using optimal biomass process technology and related genetic breeding in Miscanthus and beyond. Copyright © 2017 Elsevier Ltd. All rights reserved.

Watch Out for the "Living Dead": Cell-Free Enzymes and Their Fate.

PubMed

Baltar, Federico

2017-01-01

Microbes are the engines driving biogeochemical cycles. Microbial extracellular enzymatic activities (EEAs) are the "gatekeepers" of the carbon cycle. The total EEA is the sum of cell-bound (i.e., cell-attached), and dissolved (i.e., cell-free) enzyme activities. Cell-free enzymes make up a substantial proportion (up to 100%) of the total marine EEA. Although we are learning more about how microbial diversity and function (including total EEA) will be affected by environmental changes, little is known about what factors control the importance of the abundant cell-free enzymes. Since cell-attached EEAs are linked to the cell, their fate will likely be linked to the factors controlling the cell's fate. In contrast, cell-free enzymes belong to a kind of "living dead" realm because they are not attached to a living cell but still are able to perform their function away from the cell; and as such, the factors controlling their activity and fate might differ from those affecting cell-attached enzymes. This article aims to place cell-free EEA into the wider context of hydrolysis of organic matter, deal with recent studies assessing what controls the production, activity and lifetime of cell-free EEA, and what their fate might be in response to environmental stressors. This perspective article advocates the need to go "beyond the living things," studying the response of cells/organisms to different stressors, but also to study cell-free enzymes, in order to fully constrain the future and evolution of marine biogeochemical cycles.

Multimodal analysis of pretreated biomass species highlights generic markers of lignocellulose recalcitrance.

PubMed

Herbaut, Mickaël; Zoghlami, Aya; Habrant, Anouck; Falourd, Xavier; Foucat, Loïc; Chabbert, Brigitte; Paës, Gabriel

2018-01-01

Biomass recalcitrance to enzymatic hydrolysis has been assigned to several structural and chemical factors. However, their relative importance remains challenging to evaluate. Three representative biomass species (wheat straw, poplar and miscanthus) were submitted to four standard pretreatments (dilute acid, hot water, ionic liquid and sodium chlorite) in order to generate a set of contrasted samples. A large array of techniques, including wet chemistry analysis, porosity measurements using NMR spectroscopy, electron and fluorescence microscopy, were used in order to determine possible generic factors of biomass recalcitrance. The pretreatment conditions selected allowed obtaining samples displaying different susceptibility to enzymatic hydrolysis (from 3 up to 98% of the initial glucose content released after 96 h of saccharification). Generic correlation coefficients were calculated between the measured chemical and structural features and the final saccharification rates. Increases in porosity displayed overall strong positive correlations with saccharification efficiency, but different porosity ranges were concerned depending on the considered biomass. Lignin-related factors displayed highly negative coefficients for all biomasses. Lignin content, which is likely involved in the correlations observed for porosity, was less detrimental to enzymatic hydrolysis than lignin composition. Lignin influence was highlighted by the strong negative correlation with fluorescence intensity which mainly originates from monolignols in mature tissues. Our results provide a better understanding of the factors responsible for biomass recalcitrance that can reasonably be considered as generic. The correlations with specific porosity ranges are biomass species-dependent, meaning that enzymes cocktails with fitted enzyme size are likely to be needed to optimise saccharification depending on the biomass origin. Lignin composition, which probably influences its structure, is the most important parameter to overcome to enhance enzymes access to the polysaccharides. Accordingly, fluorescence intensity was found to be a rapid and simple method to assess recalcitrance after pretreatment.

Understanding longitudinal wood fiber ultra-structure for producing cellulose nanofibrils using disk milling with diluted acid prehydrolysis

Treesearch

Yanlin Qin; Xueqing Qiu; Junyong Zhu

2016-01-01

Here we used dilute oxalic acid to pretreat a kraft bleached Eucalyptus pulp (BEP) fibers to facilitate mechanical fibrillation in producing cellulose nanofibrils using disk milling with substantial mechanical energy savings. We successfully applied a reaction kinetics based combined hydrolysis factor (CHFx) as a severity factor to quantitatively...

Hydrolytic stability of pneumococcal group 6 (type 6A and 6B) capsular polysaccharides.

PubMed Central

Zon, G; Szu, S C; Egan, W; Robbins, J D; Robbins, J B

1982-01-01

The hydrolyses of the immunologically cross-reactive and constitutionally isomeric group 6 pneumococcal polysaccharides, types 6A and 6B, were investigated by 31P nuclear magnetic resonance spectroscopy, gel filtration through Sepharose 4B, reducing-sugar analysis, and rocket immunoelectrophoresis. Phosphorus nuclear magnetic resonance spectroscopy showed that cleavage of the repeating-unit phosphodiester linkages at pH 10, 60 degrees C was considerably faster (greater than 10(3) ) for the type 6A than the type 6B polysaccharide. Under these reaction conditions, 31P nuclear magnetic resonance kinetic measurements showed that the Na+ form of the type 6A polysaccharide underwent phosphodiester-linkage hydrolysis two times slower than the corresponding Ca+2 form; a stoichiometrically excess amount of Ca+2 caused a 30-fold enhancement of the latter hydrolysis rate. The spectroscopic characterization of phosphorus-containing end groups resulting from hydrolysis of the type 6A polymer provided additional mechanistic information. Heating the type 6A and 6B polysaccharides at 56 degrees C for various times led to gel filtration coefficients of distribution (Kd values) which indicated that the type 6A material underwent size reductions considerably faster than did the type 6B antigen; these increased Kd values qualitatively correlated with the loss of immunochemical reactivity measured by rocket immunoelectrophoresis. The application of a statistical theory to the depolymerization of the type 6A and 6B polysaccharides was consistent with random bond cleavage, as evidenced by the calculated versus measured gel filtration patterns. Although the molecular changes causing the size reductions were not fully elaborated, it was established that the acetal linkages of the type 6A and 6B polysaccharides were comparatively resistant to hydrolysis and that depolymerization by hydrolysis of the phosphodiester linkage was a major factor only in the type 6A structure. It was concluded that the hydrolytic stability of the type 6B antigen would favor its use in the polyvalent pneumococcal vaccine rather than the cross-reactive, but comparatively unstable, type 6A polysaccharide, if all other factors are equal. Images PMID:7107011

A structured approach to target starch solubilization and hydrolysis for the sugarcane industry

USDA-ARS?s Scientific Manuscript database

In sugarcane processing, starch is considered an impurity that negatively affects processing and reduces the quality of the sugar end-product. In the last decade, there has been a general world-wide increase in starch concentrations in sugarcane. Industrial a-amylases have been used for many years ...

The regulatory properties of rubisco activase differ among species and affect photosynthetic induction during light transitions

USDA-ARS?s Scientific Manuscript database

Rubisco’s catalytic chaperone, Rubisco activase (Rca), uses the energy from ATP hydrolysis to restore catalytic competence to Rubisco. In Arabidopsis, inhibition of Rca activity by ADP is fine-tuned by redox regulation of the a-isoform. To elucidate the mechanism for Rca regulation in species contai...

Hydrolysis of soybean protein improves iron bioavailability

USDA-ARS?s Scientific Manuscript database

Iron is an important trace metal element in human body. Iron deficiency affects human health, especially pregnant women and children. Soybean protein is a popular food in Asia and can contain a high amount of iron (145.70±0.74 ug/g); however, it is usually reported as an inhibitor of iron absorption...

Enzymatic production of trans fatty acid free fat from partially hydrogenated soybean oil (PHSO)--theory, strategy and practicability.

PubMed

Jala, Ram Chandra Reddy; Xu, Xuebing; Guo, Zheng

2013-12-01

Development of an advanced process/production technology for healthful fats constitutes a major interest of plant oil refinery industry. In this work, a strategy to produce trans fatty acid (TFA) free (or low TFA) products from partially hydrogenated soybean oil by lipase-catalysed selective hydrolysis was proposed, where a physically founded mathematic model to delineate the multi-responses of the reaction as a function of selectivity factor was defined for the first time. The practicability of this strategy was assessed with commercial trans-selective Candida antarctica lipase A (CAL-A) as a model biocatalyst based on a parameter study and fitting to the model. CAL-A was found to have a selectivity factor 4.26 and to maximally remove 73.3% of total TFAs at 46.5% hydrolysis degree. Copyright © 2013 Elsevier Ltd. All rights reserved.

Enhancement of β-xylosidase productivity in cellulase producing fungus Acremonium cellulolyticus

PubMed Central

2011-01-01

Enzymatic hydrolysis is one of the most important processes in bioethanol production from lignocellulosic biomass. Acremonium cellulolyticus is a filamentous fungus with high cellulase production but productivity of hemicellulase, especially β-xylosidase, is lower than other filamentous fungi. We identified 2.4 Kb β-xylosidase gene in the A. cellulolyticus genome sequence information and it encoded 798 amino acids without introns. To enhance hemicellulase productivity in A. cellulolyticus, we transformed this fungus with the identified β-xylosidase gene driven by the cellobiohydrolase Ι (cbh1) promoter, using the protoplast-polyethyleneglycol (PEG) method, and obtained a transformant, YKX1. Hydrolysis rate of xylooligosaccharides was more than 50-fold higher using culture supernatant from YKX1 than that from the parental strain, Y-94. Total cellulase activity (measured by filter paper assay) in YKX1 was not affected by the cbh1 promoter used for expression of β-xylosidase, and induced by cellulose. Since YKX1 can produce larger amount of β-xylosidase without affecting cellulase productivity, it is considered to be beneficial for practical monosaccharide recoveries from lignocellulosic biomass. PMID:21906369

Enhancement of β-xylosidase productivity in cellulase producing fungus Acremonium cellulolyticus.

PubMed

Kanna, Machi; Yano, Shinichi; Inoue, Hiroyuki; Fujii, Tatsuya; Sawayama, Shigeki

2011-06-30

Enzymatic hydrolysis is one of the most important processes in bioethanol production from lignocellulosic biomass. Acremonium cellulolyticus is a filamentous fungus with high cellulase production but productivity of hemicellulase, especially β-xylosidase, is lower than other filamentous fungi. We identified 2.4 Kb β-xylosidase gene in the A. cellulolyticus genome sequence information and it encoded 798 amino acids without introns. To enhance hemicellulase productivity in A. cellulolyticus, we transformed this fungus with the identified β-xylosidase gene driven by the cellobiohydrolase Ι (cbh1) promoter, using the protoplast-polyethyleneglycol (PEG) method, and obtained a transformant, YKX1. Hydrolysis rate of xylooligosaccharides was more than 50-fold higher using culture supernatant from YKX1 than that from the parental strain, Y-94. Total cellulase activity (measured by filter paper assay) in YKX1 was not affected by the cbh1 promoter used for expression of β-xylosidase, and induced by cellulose. Since YKX1 can produce larger amount of β-xylosidase without affecting cellulase productivity, it is considered to be beneficial for practical monosaccharide recoveries from lignocellulosic biomass.

The correlation between cellulose allomorphs (I and II) and conversion after removal of hemicellulose and lignin of lignocellulose.

PubMed

Song, Yanliang; Zhang, Jingzhi; Zhang, Xu; Tan, Tianwei

2015-10-01

H2SO4, NaOH and H3PO4 were applied to decompose lignocellulose samples (giant reeds, pennisetum and cotton stalks) to investigate the correlation between cellulose allomorphs (cellulose I and II) and conversion of cellulose. The effect of removal of hemicellulose and lignin on the surface morphology, crystallinity index (CrI), cellulose allomorphs (cellulose I and II), and enzymatic hydrolysis under different pretreatments was also studied. CrI caused by H3PO4 pretreatment reached 11.19%, 24.93% and 8.15% for the three samples, respectively. Corn stalk showed highest conversion of cellulose among three samples, irrespective of the pretreatment used. This accounted for the widely use of corn stalk as the renewable crop substrate to synthesize biofuels like ethanol. CrI of cellulose I (CrI-I) negatively affects cellulose conversion but CrI of cellulose II (CrI-II) positively affects cellulose conversion. It contributes to make the strategy to transform cellulose I to cellulose II and enhancing enzymatic hydrolysis of lignocellulose. Copyright © 2015 Elsevier Ltd. All rights reserved.

Cell growth and proteolytic activity of Lactobacillus acidophilus, Lactobacillus helveticus, Lactobacillus delbrueckii ssp. bulgaricus, and Streptococcus thermophilus in milk as affected by supplementation with peptide fractions.

PubMed

Gandhi, Akanksha; Shah, Nagendra P

2014-12-01

The present investigation examined the effects of supplementation of milk peptide fractions produced by enzymatic hydrolysis on the fermentation of reconstituted skim milk (RSM). Changes in pH, cell growth, proteolytic activity, and angiotensin-converting enzyme (ACE)-inhibitory activity were monitored during fermentation of RSM by pure cultures of Lactobacillus acidophilus, Lactobacillus helveticus, Lactobacillus delbrueckii ssp. bulgaricus, and Streptococcus thermophilus. The study showed that supplementation with peptide fractions of different molecular weights did not significantly affect the bacterial growth in RSM. All bacteria showed an increased proteolytic activity in RSM supplemented with large peptides (>10 kDa), and L. helveticus in general exhibited the highest proteolytic activity among the bacteria studied. The ACE-inhibitory activity was observed to be the maximum in RSM supplemented with larger peptides (>10 kDa) for all bacteria. The results suggest that proteolysis by bacteria leads to increased production of ACE-inhibitory peptides compared to the supplemented peptides produced by enzymatic hydrolysis.

Antibacterial Activity of Free Fatty Acids from Hydrolyzed Virgin Coconut Oil Using Lipase from Candida rugosa.

PubMed

Nguyen, Van Thi Ai; Le, Truong Dang; Phan, Hoa Ngoc; Tran, Lam Bich

2017-01-01

Free fatty acids (FFAs) were obtained from hydrolyzed virgin coconut oil (VCO) by Candida rugosa lipase (CRL). Four factors' influence on hydrolysis degree (HD) was examined. The best hydrolysis conditions in order to get the highest HD value were determined at VCO to buffer ratio 1 : 5 (w/w), CRL concentration 1.5% (w/w oil), pH 7, and temperature 40°C. After 16 hours' reaction, the HD value achieved 79.64%. FFAs and residual hydrolyzed virgin coconut oil (HVCO) were isolated from the hydrolysis products. They were tested for their antibacterial activity against Gram-negative and Gram-positive bacteria, which can be found in contaminated food and cause food poisoning. FFAs showed their inhibition against Bacillus subtilis (ATCC 11774), Escherichia coli (ATCC 25922), Salmonella enteritidis (ATCC 13076), and Staphylococcus aureus (ATCC 25923) at minimum inhibitory concentration (MIC) of 50%, 60%, 20%, and 40%, respectively. However, VCO and HVCO did not show their antibacterial activity against these tested bacteria.

Characterization of Proteins in Filtrate from Biodegradation of Crop Residue

NASA Technical Reports Server (NTRS)

Horton, Wileatha; Trotman, A. A.

1997-01-01

Biodegradation of plant biomass is a feasible path for transformation of crop residue and recycling of nutrients for crop growth. The need to model the effects of factors associated with recycling of plant biomass resulting from hydroponic sweet potato production has led to investigation of natural soil isolates with the capacity for starch hydrolysis. This study sought to use nondenaturing gel electrophoresis to characterize the proteins present in filtered effluent from bioreactors seeded with starch hydrolyzing bacterial culture used in the biodegradation of senesced sweet potato biomass. The study determined the relative molecular weight of proteins in sampled effluent and the protein banding pattern was characterized. The protein profiles of effluent were similar for samples taken from independent runs under similar conditions of starch hydrolysis. The method can be used as a quality control tool for confirmation of starch hydrolysis of crop biomass. In addition, this method will allow monitoring for presence of contaminants within the system-protein profiles indicative of new enzymes in the bioreactors.

Combined Enzymatic and Mechanical Cell Disruption and Lipid Extraction of Green Alga Neochloris oleoabundans

PubMed Central

Wang, Dongqin; Li, Yanqun; Hu, Xueqiong; Su, Weimin; Zhong, Min

2015-01-01

Microalgal biodiesel is one of the most promising renewable fuels. The wet technique for lipids extraction has advantages over the dry method, such as energy-saving and shorter procedure. The cell disruption is a key factor in wet oil extraction to facilitate the intracellular oil release. Ultrasonication, high-pressure homogenization, enzymatic hydrolysis and the combination of enzymatic hydrolysis with high-pressure homogenization and ultrasonication were employed in this study to disrupt the cells of the microalga Neochloris oleoabundans. The cell disruption degree was investigated. The cell morphology before and after disruption was assessed with scanning and transmission electron microscopy. The energy requirements and the operation cost for wet cell disruption were also estimated. The highest disruption degree, up to 95.41%, assessed by accounting method was achieved by the combination of enzymatic hydrolysis and high-pressure homogenization. A lipid recovery of 92.6% was also obtained by the combined process. The combined process was found to be more efficient and economical compared with the individual process. PMID:25853267

Antibacterial Activity of Free Fatty Acids from Hydrolyzed Virgin Coconut Oil Using Lipase from Candida rugosa

PubMed Central

Phan, Hoa Ngoc; Tran, Lam Bich

2017-01-01

Free fatty acids (FFAs) were obtained from hydrolyzed virgin coconut oil (VCO) by Candida rugosa lipase (CRL). Four factors' influence on hydrolysis degree (HD) was examined. The best hydrolysis conditions in order to get the highest HD value were determined at VCO to buffer ratio 1 : 5 (w/w), CRL concentration 1.5% (w/w oil), pH 7, and temperature 40°C. After 16 hours' reaction, the HD value achieved 79.64%. FFAs and residual hydrolyzed virgin coconut oil (HVCO) were isolated from the hydrolysis products. They were tested for their antibacterial activity against Gram-negative and Gram-positive bacteria, which can be found in contaminated food and cause food poisoning. FFAs showed their inhibition against Bacillus subtilis (ATCC 11774), Escherichia coli (ATCC 25922), Salmonella enteritidis (ATCC 13076), and Staphylococcus aureus (ATCC 25923) at minimum inhibitory concentration (MIC) of 50%, 60%, 20%, and 40%, respectively. However, VCO and HVCO did not show their antibacterial activity against these tested bacteria. PMID:29259829

Parameter and Process Significance in Mechanistic Modeling of Cellulose Hydrolysis

NASA Astrophysics Data System (ADS)

Rotter, B.; Barry, A.; Gerhard, J.; Small, J.; Tahar, B.

2005-12-01

The rate of cellulose hydrolysis, and of associated microbial processes, is important in determining the stability of landfills and their potential impact on the environment, as well as associated time scales. To permit further exploration in this field, a process-based model of cellulose hydrolysis was developed. The model, which is relevant to both landfill and anaerobic digesters, includes a novel approach to biomass transfer between a cellulose-bound biofilm and biomass in the surrounding liquid. Model results highlight the significance of the bacterial colonization of cellulose particles by attachment through contact in solution. Simulations revealed that enhanced colonization, and therefore cellulose degradation, was associated with reduced cellulose particle size, higher biomass populations in solution, and increased cellulose-binding ability of the biomass. A sensitivity analysis of the system parameters revealed different sensitivities to model parameters for a typical landfill scenario versus that for an anaerobic digester. The results indicate that relative surface area of cellulose and proximity of hydrolyzing bacteria are key factors determining the cellulose degradation rate.

Determination of effect factor for effective parameter on saccharification of lignocellulosic material by concentrated acid

NASA Astrophysics Data System (ADS)

Aghili, Sina; Nodeh, Ali Arasteh

2015-12-01

Tamarisk usage as a new group of lignocelluloses material to produce fermentable sugars in bio ethanol process was studied. The overall aim of this work was to establish the optimum condition for acid hydrolysis of this new material and a mathematical model predicting glucose release as a function of operation variable. Sulfuric acid concentration in the range of 20 to 60%(w/w), process temperature between 60 to 95oC, hydrolysis time from 120 to 240 min and solid content 5,10,15%(w/w) were used as hydrolysis conditions. HPLC was used to analysis of the product. This analysis indicated that glucose was the main fermentable sugar and was increase with time, temperature and solid content and acid concentration was a parabola influence in glucose production. The process was modeled by a quadratic equation. Curve study and model were found that 42% acid concentration, 15 % solid content and 90oC were optimum condition.

Immunological characterization of the gluten fractions and their hydrolysates from wheat, rye and barley.

PubMed

Rallabhandi, Prasad; Sharma, Girdhari M; Pereira, Marion; Williams, Kristina M

2015-02-18

Gluten proteins in wheat, rye and barley cause celiac disease, an autoimmune disorder of the small intestine, which affects approximately 1% of the world population. Gluten is comprised of prolamin and glutelin. Since avoidance of dietary gluten is the only option for celiac patients, a sensitive gluten detection and quantitation method is warranted. Most regulatory agencies have set a threshold of 20 ppm gluten in foods labeled gluten-free, based on the currently available ELISA methods. However, these methods may exhibit differences in gluten quantitation from different gluten-containing grains. In this study, prolamin and glutelin fractions were isolated from wheat, rye, barley, oats and corn. Intact and pepsin-trypsin (PT)-digested prolamin and glutelin fractions were used to assess their immunoreactivity and gluten recovery by three sandwich and two competitive ELISA kits. The Western blots revealed varied affinity of ELISA antibodies to gluten-containing grain proteins and no reactivity to oat and corn proteins. ELISA results showed considerable variation in gluten recoveries from both intact and PT-digested gluten fractions among different kits. Prolamin fractions showed higher gluten recovery compared to their respective glutelin fractions. Among prolamins, barley exhibited higher recovery compared to wheat and rye with most of the ELISA kits used. Hydrolysis resulted in reduced gluten recovery of most gluten fractions. These results suggest that the suitability of ELISA for accurate gluten quantitation is dependent upon various factors, such as grain source, antibody specificity, gluten proteins and the level of their hydrolysis in foods.

Impact of magnesium:calcium ratio on calcification of the aortic wall.

PubMed

Villa-Bellosta, Ricardo

2017-01-01

An inverse relationship between serum magnesium concentration and vascular calcification has been reported following observational clinical studies. Moreover, several studies have been suggesting a protective effect of magnesium on the vascular calcification. However, the exact mechanism remains elusive, and investigators have speculated among a myriad of potential actions. The effect of magnesium on calcification of the aortic wall is yet to be investigated. In the present study, the effects of magnesium and calcium on the metabolism of extracellular PPi, the main endogenous inhibitor of vascular calcification, were investigated in the rat aorta. Calcium and magnesium have antagonist effects on PPi hydrolysis in the aortic wall. Km and Ki values for PPi hydrolysis in rat aortic rings were 1.1 mmol/L magnesium and 32 μmol/L calcium, respectively, but ATP hydrolysis was not affected with calcium. Calcium deposition in the rat aortic wall dramatically increased when the magnesium concentration was increased (ratio of Mg:Ca = 1:1; 1.5 mmol/L calcium and 1.5 mmol/L magnesium) respect to low magnesium concentration (ratio Mg:Ca = 1:3, 1.5 mmol/L calcium and 0.75 mmol/L magnesium). Data from observational clinical studies showing that the serum magnesium concentration is inversely correlated with vascular calcification could be reinterpreted as a compensatory regulatory mechanism that reduces both PPi hydrolysis and vascular calcification. The impact of magnesium in vascular calcification in humans could be studied in association with calcium levels, for example, as the magnesium:calcium ratio.

Detoxification of a lignocellulosic biomass slurry by soluble polyelectrolyte adsorption for improved fermentation efficiency.

PubMed

Carter, Brian; Squillace, Phillip; Gilcrease, Patrick C; Menkhaus, Todd J

2011-09-01

This study investigated the detoxification of a dilute acid pretreated Ponderosa pine slurry using the polyelectrolyte polyethyleneimine (PEI). The addition of polyelectrolyte to remove enzymatic and/or fermentation inhibitory compounds, that is, acetic acid, furfural, and 5-hydroxymethylfurfural (HMF), was performed either before or after enzymatic hydrolysis to determine the optimal process sequence. Negligible acetic acid, glucose, and xylose were removed regardless of where in the process the polymer addition was made. Maximum furfural and HMF separation was achieved with the addition of PEI to a clarified pre-enzymatic hydrolysis liquor, which showed that 88.3% of furfural and 66.4% of HMF could be removed. On the other hand, only 23.1% and 13.4% of furfural and HMF, respectively, were removed from a post-enzymatic hydrolysis sample; thus, the effects of enzymes, glucose, and wood solids on inhibitor removal were also investigated. The presence of solid particles >0.2 µm and unknown soluble components <10 kDa reduced inhibitory compound removal, but the presence of elevated glucose levels and enzymes (cellulases) did not affect the separation. The fermentability of detoxified versus undetoxified hydrolysate was also investigated. An ethanol yield of 92.6% of theoretical was achieved with Saccharomyces cerevisiae fermenting the detoxified hydrolyzate, while no significant ethanol was produced in the undetoxified hydrolyzate. These results indicate that PEI may provide a practical alternative for furan removal and detoxification of lignocellolosic hydrolysates, and that application before enzymatic hydrolysis minimizes separation interferences. Copyright © 2011 Wiley Periodicals, Inc.

Production of wheat gluten hydrolysates with reduced antigenicity employing enzymatic hydrolysis combined with downstream unit operations.

PubMed

Merz, Michael; Kettner, Lucas; Langolf, Emma; Appel, Daniel; Blank, Imre; Stressler, Timo; Fischer, Lutz

2016-08-01

Due to allergies or other health disorders a certain segment of the population is not able to safely consume some plant proteins, which are the main protein support in human nutrition. Coeliac disease is a prominent autoimmune disorder and requires a strict adherence to a gluten-free diet. The aim of this study was to identify suitable combinations of enzymatic hydrolysis and common unit operations in food processing (centrifugation, ultra-filtration) to produce gluten-free wheat gluten hydrolysates for food application. To analyse the hydrolysates, a simple and cheap competitive ELISA protocol was designed and validated in this study as well. The competitive ELISA was validated using gliadin spiked skim milk protein hydrolysates, due to the latter application of the assay. The limit of quantification was 4.19 mg kg(-1) , which allowed the identification of gluten-free (<20 mg kg(-1) ) hydrolysates. Enzymatic hydrolysis, including the type of peptidase, and the downstream processing greatly affected the antigenicity of the hydrolysates. Enzymatic hydrolysis and downstream processing operations, such as centrifugation and ultra-filtration, reduced the antigenicity of wheat gluten hydrolysates. Gluten-free hydrolysates were obtained with Flavourzyme after centrifugation (25 g L(-1) substrate) and after 1 kDa ultra-filtration (100 g L(-1) substrate). A multiple peptidase complex, such as Flavourzyme, seems to be required for the production of gluten-free hydrolysates. © 2015 Society of Chemical Industry. © 2015 Society of Chemical Industry.

A review on the kinetics of microbially induced calcium carbonate precipitation by urea hydrolysis

NASA Astrophysics Data System (ADS)

van Paassen, L. A.

2017-12-01

In this study the kinetics of calcium carbonate precipitation induced by the ureolytic bacteria are reviewed based on experiments and mathematical modelling. The study shows how urea hydrolysis rate depends on the amount of bacteria and the conditions during growth, storage, hydrolysis and precipitation. The dynamics of Microbially Induced Carbonate Precipitation has been monitored in non-seeded liquid batch experiments. Results show that particulary for a fast hydrolysis of urea (>1 M-urea day-1) in a highly concentrated equimolar solution with calcium chloride (>0.25 M) the solubility product of CaCO3 is exceeded within a short period (less than 30 minutes), the supersaturation remains high for an exended period, resulting in prolonged periods of nucleation and crystal growth and extended growth of metastable precursor mineral phases. The pH, being a result of the speciation, quickly rises until critical supersaturation is reached and precipitation is initiated. Then pH drops (sometimes showing oscillating behaviour) to about neutral where it stays until all substrates are depleted. Higher hydrolysis rates lead to higher supersaturation and pH and relatively many small crystals, whereas higher concentrations of urea and calcium chloride mainly lead to lower pH values. The conversion can be reasonably monitored by electrical conductivity and reasonably predicted, using a simplified model based on a single reaction as long as the urea hydrolysis rate is known. Complex geochemical models, which include chemical speciciation through acid-base equilibria and kinetic equations to describe mineral precipitation, do not show significant difference from the simplified model regarding the bulk chemistry and the total amount of precipitates. However, experiments show that ureolytic MICP can result in a highly variable crystal morphologies with large variation in the affected hydraulic properties when applied in a porous medium. In order to calculate the number, size and type of crystals, use of these more complex models is essential. Quantitative prediction to a level at which the pH and conductivity are simulated accurately is not yet possible as experimental data regarding the interaction between existing mineral surfaces are the surface interaction between ions and micro-organisms is still lacking.

A parameterization of the heterogeneous hydrolysis of N2O5 for mass-based aerosol models: improvement of particulate nitrate prediction

NASA Astrophysics Data System (ADS)

Chen, Ying; Wolke, Ralf; Ran, Liang; Birmili, Wolfram; Spindler, Gerald; Schröder, Wolfram; Su, Hang; Cheng, Yafang; Tegen, Ina; Wiedensohler, Alfred

2018-01-01

The heterogeneous hydrolysis of N2O5 on the surface of deliquescent aerosol leads to HNO3 formation and acts as a major sink of NOx in the atmosphere during night-time. The reaction constant of this heterogeneous hydrolysis is determined by temperature (T), relative humidity (RH), aerosol particle composition, and the surface area concentration (S). However, these parameters were not comprehensively considered in the parameterization of the heterogeneous hydrolysis of N2O5 in previous mass-based 3-D aerosol modelling studies. In this investigation, we propose a sophisticated parameterization (NewN2O5) of N2O5 heterogeneous hydrolysis with respect to T, RH, aerosol particle compositions, and S based on laboratory experiments. We evaluated closure between NewN2O5 and a state-of-the-art parameterization based on a sectional aerosol treatment. The comparison showed a good linear relationship (R = 0.91) between these two parameterizations. NewN2O5 was incorporated into a 3-D fully online coupled model, COSMO-MUSCAT, with the mass-based aerosol treatment. As a case study, we used the data from the HOPE Melpitz campaign (10-25 September 2013) to validate model performance. Here, we investigated the improvement of nitrate prediction over western and central Europe. The modelled particulate nitrate mass concentrations ([NO3-]) were validated by filter measurements over Germany (Neuglobsow, Schmücke, Zingst, and Melpitz). The modelled [NO3-] was significantly overestimated for this period by a factor of 5-19, with the corrected NH3 emissions (reduced by 50 %) and the original parameterization of N2O5 heterogeneous hydrolysis. The NewN2O5 significantly reduces the overestimation of [NO3-] by ˜ 35 %. Particularly, the overestimation factor was reduced to approximately 1.4 in our case study (12, 17-18 and 25 September 2013) when [NO3-] was dominated by local chemical formations. In our case, the suppression of organic coating was negligible over western and central Europe, with an influence on [NO3-] of less than 2 % on average and 20 % at the most significant moment. To obtain a significant impact of the organic coating effect, N2O5, SOA, and NH3 need to be present when RH is high and T is low. However, those conditions were rarely fulfilled simultaneously over western and central Europe. Hence, the organic coating effect on the reaction probability of N2O5 may not be as significant as expected over western and central Europe.

A supramolecular structure insight for conversion property of cellulose in hot compressed water: Polymorphs and hydrogen bonds changes.

PubMed

Wang, Yan; Lian, Jie; Wan, Jinquan; Ma, Yongwen; Zhang, Yingshi

2015-11-20

Waste paper samples with different cellulose supramolecular structure were treated in hot compressed water (HCW) at 375°C and 22.5MPa within 200s to evaluate the specific effect mechanism of cellulose supramolecular structure on the conversion of waste paper to reusable resource. Although the distribution of liquid products and the oligosaccharides were related to reaction time, depolymerization and decrystallization of the cellulose, the characteristics absorption peak of cellulose from FTIR analysis and crystal structure of the cellulose detected in the residues with hydrolysis rate up 96.5% indicated crystal structure was the dominant factor that affect conversion behavior of waste paper. The conversion of cellulose Iβ to cellulose Iα or cellulose I(α+β) in HCW demonstrated that the recrystallization occurred during the decrystallization of cellulose through the rearrangement of hydrogen bonds. Copyright © 2015 Elsevier Ltd. All rights reserved.

Preparation and antioxidant activities of oligosaccharides from Crassostrea gigas.

PubMed

Wu, Shengjun; Huang, Xiaolian

2017-02-01

Oligosaccharides were prepared from Crassostrea gigas by hydrolysis of polysaccharide in C. gigas with peroxide oxygen (H2O2). The hydrolysates were cleared of protein, filtered, ultrafiltered and precipitated with absolute ethanol to give C. gigas oligosaccharides (CGOs). Factors affecting CGO yields, i.e., reaction time, temperature, and H2O2 concentration, were optimised as follows: 2.96h reaction time, 84.71°C reaction temperature, and 2.46% H2O2 concentration. Under these conditions, the maximum yield of CGOs reached 10.61%. The CGOs were then partially characterised by Fourier transform infrared spectroscopy, UV spectroscopy, monosaccharide composition, and antioxidant activities. Results indicate that CGOs possessed strong hydroxyl radical activity, 2,2-diphenyl-β-picrylhydrazyl-radical-scavenging activity and reducing capacity at a concentration of 100μg/mL. Copyright © 2016 Elsevier Ltd. All rights reserved.

Angiotensin-I-Converting Enzyme (ACE)-Inhibitory Peptides from Plants

PubMed Central

Daskaya-Dikmen, Ceren; Yucetepe, Aysun; Karbancioglu-Guler, Funda; Daskaya, Hayrettin; Ozcelik, Beraat

2017-01-01

Hypertension is an important factor in cardiovascular diseases. Angiotensin-I-converting enzyme (ACE) inhibitors like synthetic drugs are widely used to control hypertension. ACE-inhibitory peptides from food origins could be a good alternative to synthetic drugs. A number of plant-based peptides have been investigated for their potential ACE inhibitor activities by using in vitro and in vivo assays. These plant-based peptides can be obtained by solvent extraction, enzymatic hydrolysis with or without novel food processing methods, and fermentation. ACE-inhibitory activities of peptides can be affected by their structural characteristics such as chain length, composition and sequence. ACE-inhibitory peptides should have gastrointestinal stability and reach the cardiovascular system to show their bioactivity. This paper reviews the current literature on plant-derived ACE-inhibitory peptides including their sources, production and structure, as well as their activity by in vitro and in vivo studies and their bioavailability. PMID:28333109

Investigating the Affinities and Persistence of VX Nerve Agent in Environmental Matrices

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

Love, A H; Vance, A L; Reynolds, J G

2004-03-09

Laboratory experiments were conducted to determine environmental variables that affect the affinities and persistence of the nerve agent O-ethyl S-(2-diisopropylaminoethyl) methylphosphonothiolate (VX) at dilute concentrations in environmental matrices. Quantitative analyses of VX and its degradation products were performed using LC-MS. Batch hydrolysis experiments demonstrated an increasing hydrolysis rate as pH increased, as shown in previous studies, but also indicated that dissolved aqueous constituents can cause significant differences in the absolute hydrolysis rate. Adsorption isotherms from batch aqueous experiments revealed that VX has a high affinity for hydrophobic organics, a moderate affinity for montmorillonite clay, and a very low affinity formore » an iron-oxyhydroxide soil mineral, goethite. The adsorption on goethite was increased with the presence of dissolved organic matter in solution. VX degraded rapidly when dried onto goethite, when an inner-sphere complex was forced. No enhanced degradation occurred with goethite in small amounts water. These results suggest that aqueous conditions have important controls on VX adsorption and degradation in the environment and a more mechanistic understanding of these controls is needed in order to enable accurate predictions of its long-term fate and persistence.« less

Enzymatic properties of Staphylococcus aureus adenosine synthase (AdsA)

PubMed Central

2011-01-01

Background Staphylococcus aureus is a human pathogen that produces extracellular adenosine to evade clearance by the host immune system, an activity attributed to the 5'-nucleotidase activity of adenosine synthase (AdsA). In mammals, conversion of adenosine triphosphate to adenosine is catalyzed in a two-step process: ecto-nucleoside triphosphate diphosphohydrolases (ecto-NTDPases) hydrolyze ATP and ADP to AMP, whereas 5'-nucleotidases hydrolyze AMP to adenosine. NTPDases harbor apyrase conserved regions (ACRs) that are critical for activity. Results NTPDase ACR motifs are absent in AdsA, yet we report here that recombinant AdsA hydrolyzes ADP and ATP in addition to AMP. Competition assays suggest that hydrolysis occurs following binding of all three substrates at a unique site. Alanine substitution of two amino acids, aspartic acid 127 and histidine 196 within the 5'-nucleotidase signature sequence, leads to reduced AMP or ADP hydrolysis but does not affect the binding of these substrates. Conclusion Collectively, these results provide insight into the unique ability of AdsA to produce adenosine through the consecutive hydrolysis of ATP, ADP and AMP, thereby endowing S. aureus with the ability to modulate host immune responses. PMID:22035583

Degradation mechanisms of bioresorbable polyesters. Part 2. Effects of initial molecular weight and residual monomer.

PubMed

Gleadall, Andrew; Pan, Jingzhe; Kruft, Marc-Anton; Kellomäki, Minna

2014-05-01

This paper presents an understanding of how initial molecular weight and initial monomer fraction affect the degradation of bioresorbable polymers in terms of the underlying hydrolysis mechanisms. A mathematical model was used to analyse the effects of initial molecular weight for various hydrolysis mechanisms including noncatalytic random scission, autocatalytic random scission, noncatalytic end scission or autocatalytic end scission. Different behaviours were identified to relate initial molecular weight to the molecular weight half-life and to the time until the onset of mass loss. The behaviours were validated by fitting the model to experimental data for molecular weight reduction and mass loss of samples with different initial molecular weights. Several publications that consider initial molecular weight were reviewed. The effect of residual monomer on degradation was also analysed, and shown to accelerate the reduction of molecular weight and mass loss. An inverse square root law relationship was found between molecular weight half-life and initial monomer fraction for autocatalytic hydrolysis. The relationship was tested by fitting the model to experimental data with various residual monomer contents. Copyright © 2014 Acta Materialia Inc. Published by Elsevier Ltd. All rights reserved.

Lactobacillus reuteri 100-23 modulates urea hydrolysis in the murine stomach.

PubMed

Wilson, Charlotte M; Loach, Diane; Lawley, Blair; Bell, Tracey; Sims, Ian M; O'Toole, Paul W; Zomer, Aldert; Tannock, Gerald W

2014-10-01

Comparisons of in vivo (mouse stomach) and in vitro (laboratory culture) transcriptomes of Lactobacillus reuteri strain 100-23 were made by microarray analysis. These comparisons revealed the upregulation of genes associated with acid tolerance, including urease production, in the mouse stomach. Inactivation of the ureC gene reduced the acid tolerance of strain 100-23 in vitro, and the mutant was outcompeted by the wild type in the gut of ex-Lactobacillus-free mice. Urine analysis showed that stable isotope-labeled urea, administered by gavage, was metabolized to a greater extent in Lactobacillus-free mice than animals colonized by strain 100-23. This surprising observation was associated with higher levels of urease activity and fecal-type bacteria in the stomach digesta of Lactobacillus-free mice. Despite the modulation of urea hydrolysis in the stomach, recycling of urea nitrogen in the murine host was not affected since the essential amino acid isoleucine, labeled with a stable isotope, was detected in the livers of both Lactobacillus-free and 100-23-colonized animals. Therefore, our experiments reveal a new and unexpected impact of Lactobacillus colonization on urea hydrolysis in the murine gut. Copyright © 2014, American Society for Microbiology. All Rights Reserved.

Lactobacillus reuteri 100-23 Modulates Urea Hydrolysis in the Murine Stomach

PubMed Central

Wilson, Charlotte M.; Loach, Diane; Lawley, Blair; Bell, Tracey; Sims, Ian M.; O'Toole, Paul W.; Zomer, Aldert

2014-01-01

Comparisons of in vivo (mouse stomach) and in vitro (laboratory culture) transcriptomes of Lactobacillus reuteri strain 100-23 were made by microarray analysis. These comparisons revealed the upregulation of genes associated with acid tolerance, including urease production, in the mouse stomach. Inactivation of the ureC gene reduced the acid tolerance of strain 100-23 in vitro, and the mutant was outcompeted by the wild type in the gut of ex-Lactobacillus-free mice. Urine analysis showed that stable isotope-labeled urea, administered by gavage, was metabolized to a greater extent in Lactobacillus-free mice than animals colonized by strain 100-23. This surprising observation was associated with higher levels of urease activity and fecal-type bacteria in the stomach digesta of Lactobacillus-free mice. Despite the modulation of urea hydrolysis in the stomach, recycling of urea nitrogen in the murine host was not affected since the essential amino acid isoleucine, labeled with a stable isotope, was detected in the livers of both Lactobacillus-free and 100-23-colonized animals. Therefore, our experiments reveal a new and unexpected impact of Lactobacillus colonization on urea hydrolysis in the murine gut. PMID:25063664

Consumption Safety of Pastries, Confectioneries, and Potato Products as Related to Fat Content.

PubMed

Żbikowska, Anna; Rutkowska, Jarosława; Kowalska, Małgorzata

2015-01-01

To determine the content of the main groups of fatty acids in pastries, confectionery, and potato products, paying special attention to trans fatty acids and the products of fat oxidation and hydrolysis, as factors affecting the safety of consumption. A total of 157 products were collected in Poland in 2009-2010. In fats extracted from samples, products of oxidation and hydrolysis were assayed using peroxide (PV), anisidine (AnV), and acid (AV) values. The fatty acid (FA) composition, especially the trans FAs (TFAs) content, was determined by gas chromatography. When assessing the TFA intake, Household Budget Surveys were considered. Highest content of fat was found for wafers with filling and crisps (32.3% and 29.3%, respectively). In 4 out of 9 groups of pastry and confectionery products studied, the quality of fat was decreased due to an excessive oxidation, as evidenced by a substantial content of secondary products of fat oxidation. The extracted fat was rich in SFA (on average, 50 g SFA/100 g FA) except fries and mixes. A great diversity of TFA content in fat of the products was found (0.1-24.8 g TFA/100 g FA). Wafers were characterized by the highest average content of TFA in the group of pastries (1.94 g TFA/100 g of product). Products of natural origin supplied 0.496 g of TFA per day, and those of industrial origin supplied about 1.5 g. The average TFA consumption, about 2 g/day, does not seem to affect health. However, an excessive consumption of pastry and confectionery products may present a risk, due not only to TFA but also to a high consumption of toxic secondary oxidation lipid products. Moreover, because 75% of TFAs in the diet were isomers of industrial origin, their further limitation and monitoring of their level in food seems highly recommendable.

The Conformational Change in Elongation Factor Tu Involves Separation of Its Domains

DOE PAGES

Lai, Jonathan; Ghaemi, Zhaleh; Luthey-Schulten, Zaida

2017-10-18

Elongation factor Tu (EF-Tu) is a highly conserved GTPase that is responsible for supplying the aminoacylated tRNA to the ribosome. Upon binding to the ribosome, EF-Tu undergoes GTP hydrolysis, which drives a major conformational change, triggering the release of aminoacylated tRNA to the ribosome. Using a combination of molecular simulation techniques, we studied the transition between the pre- and post-hydrolysis structures through two distinct pathways. Here, we show that the transition free energy is minimal along a non-intuitive pathway that involves “separation” of the GTP binding domain (domain 1) from the OB folds (domains 2 and 3), followed by domainmore » 1 rotation, and, eventually, locking the EF-Tu conformation in the post-hydrolysis state. The domain separation also leads to a slight extension of the linker connecting domain 1 to domain 2. Using docking tools and correlation-based analysis, we identified and characterized the EF-Tu conformations that release the tRNA. These calculations suggest that EF-Tu can release the tRNA before the domains separate and after domain 1 rotates by 25°. Lastly, we also examined the EF-Tu conformations in the context of the ribosome. Given the high degrees of sequence similarity with other translational GTPases, we predict a similar separation mechanism is followed.« less

Effect of pretreatment severity on accumulation of major degradation products from dilute acid pretreated corn stover and subsequent inhibition of enzymatic hydrolysis of cellulose.

PubMed

Um, Byung-Hwan; van Walsum, G Peter

2012-09-01

The concept of reaction severity, which combines residence time and temperature, is often used in the pulp and paper and biorefining industries. The influence of corn stover pretreatment severity on yield of sugar and major degradation products and subsequent effects on enzymatic cellulose hydrolysis was investigated. The pretreatment residence time and temperature, combined into the severity factor (Log R(o)), were varied with constant acid concentration. With increasing severity, increasing concentrations of furfural and 5-hydroxymethylfurfural (5-HMF) coincided with decreasing yields of oligosaccharides. With further increase in severity factor, the concentrations of furans decreased, while the formation of formic acid and lactic acid increased. For example, from severity 3.87 to 4.32, xylose decreased from 6.39 to 5.26 mg/mL, while furfural increased from 1.04 to 1.33 mg/mL; as the severity was further increased to 4.42, furfural diminished to 1.23 mg/mL as formate rose from 0.62 to 1.83 mg/mL. The effects of dilute acid hydrolyzate, acetic acid, and lignin, in particular, on enzymatic hydrolysis were investigated with a rapid microassay method. The microplate method gave considerable time and cost savings compared to the traditional assay protocol, and it is applicable to a broad range of lignocellulosic substrates.

The Conformational Change in Elongation Factor Tu Involves Separation of Its Domains

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

Lai, Jonathan; Ghaemi, Zhaleh; Luthey-Schulten, Zaida

Elongation factor Tu (EF-Tu) is a highly conserved GTPase that is responsible for supplying the aminoacylated tRNA to the ribosome. Upon binding to the ribosome, EF-Tu undergoes GTP hydrolysis, which drives a major conformational change, triggering the release of aminoacylated tRNA to the ribosome. Using a combination of molecular simulation techniques, we studied the transition between the pre- and post-hydrolysis structures through two distinct pathways. Here, we show that the transition free energy is minimal along a non-intuitive pathway that involves “separation” of the GTP binding domain (domain 1) from the OB folds (domains 2 and 3), followed by domainmore » 1 rotation, and, eventually, locking the EF-Tu conformation in the post-hydrolysis state. The domain separation also leads to a slight extension of the linker connecting domain 1 to domain 2. Using docking tools and correlation-based analysis, we identified and characterized the EF-Tu conformations that release the tRNA. These calculations suggest that EF-Tu can release the tRNA before the domains separate and after domain 1 rotates by 25°. Lastly, we also examined the EF-Tu conformations in the context of the ribosome. Given the high degrees of sequence similarity with other translational GTPases, we predict a similar separation mechanism is followed.« less

Kinetic hydrogen isotope effects in the concerted mechanism for the hydrolysis of acetals, ketals, and ortho esters

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

Eliason, R.; Kreevoy, M.M.

1978-10-25

The hydrolysis of many ortho esters, and some acetals and ketals, is general acid catalyzed, and in some examples these generate linear Bronsted plots over substantial ranges of catalyst acidity. This suggests that the reaction coordinate is primarily a reorganization of heavy atoms since proton transfer from one oxygen to another has been shown to generate strongly curved Bronsted plots. However, the isotopic fractionation factor for the catalytically active proton in these transition states is substantially less than 1.0; in several examples it is less than 0.5. Such values have been thought to require that the reaction coordinate be largelymore » a motion of the hydrogen giving the low fractionation factor. This dilemma has been resolved by the observation and rationalization of fractionation factors as low as 0.28 for stable, hydrogen bridge-bonded complexes, AHA/sup -/. A similar, bounded coordinate is now suggested for the catalytically active protons in question. This permits the reaction coordinate to be pictured. 3 figures, 2 tables.« less

Hexeneuronic acid content of chemical pulp

Treesearch

Junyong Zhu; X.S. Chai

2007-01-01

This method describes a procedure to determine hexeneuronic acid groups (HexA) in chemical pulps. HexA affects the kappa number determination by reaction with permanganate, and can react with certain bleaching chemicals, e.g. chlorine dioxide and ozone, but not with some others such as oxygen and peroxide. The method is based on the highly selective hydrolysis of HexA...

[Research of imidazo[1,2-a]benzimidazole derivatives. XXX. Synthesis and properties of (imidazo[1,2-a]benzimidazolyl-2)acetic acid derivatives].

PubMed

Anisimova, V A; Tolpygin, I E; Spasov, A A; Serdiuk, T S; Sukhov, A G

2011-01-01

Ethyl esters of (9-subtituted-imidazo[1,2-a]benzimidazolyl-2)acetic acids were synthesized. The chemical properties of these esters (hydrolysis, decarboxylation, hydrazinolysis) and biological activity (fungicidal, antimicrobial, antiarrhythmic activity, and also affects on the brain rhythmogenesis) of the prepared compounds were studied.

Molecular switch-like regulation in motor proteins.

PubMed

Tafoya, Sara; Bustamante, Carlos

2018-06-19

Motor proteins are powered by nucleotide hydrolysis and exert mechanical work to carry out many fundamental biological tasks. To ensure their correct and efficient performance, the motors' activities are allosterically regulated by additional factors that enhance or suppress their NTPase activity. Here, we review two highly conserved mechanisms of ATP hydrolysis activation and repression operating in motor proteins-the glutamate switch and the arginine finger-and their associated regulatory factors. We examine the implications of these regulatory mechanisms in proteins that are formed by multiple ATPase subunits. We argue that the regulatory mechanisms employed by motor proteins display features similar to those described in small GTPases, which require external regulatory elements, such as dissociation inhibitors, exchange factors and activating proteins, to switch the protein's function 'on' and 'off'. Likewise, similar regulatory roles are taken on by the motor's substrate, additional binding factors, and even adjacent subunits in multimeric complexes. However, in motor proteins, more than one regulatory factor and the two mechanisms described here often underlie the machine's operation. Furthermore, ATPase regulation takes place throughout the motor's cycle, which enables a more complex function than the binary 'active' and 'inactive' states.This article is part of a discussion meeting issue 'Allostery and molecular machines'. © 2018 The Author(s).

Muscarinic-agonist and guanine nucleotide stimulation of myo-inositol trisphosphate formation in membranes isolated from bovine iris sphincter smooth muscle: effects of short-term cholinergic desensitization.

PubMed

Honkanen, R E; Abdel-Latif, A A

1989-01-01

The effect of short-term cholinergic desensitization on muscarinic acetylcholine receptor (mAChR)-mediated activation of phospholipase C was investigated in membranes isolated from the bovine iris sphincter smooth muscle. Membranes prepared from normal or desensitized muscles, prelabeled with either [3H]myo-inositol or 32P from [gamma-32P]ATP, were incubated with a hydrolysis-resistant analogue of GTP, GTP gamma S, or GTP gamma S plus carbachol (CCh), and the production of [3H]myo-inositol 1,4,5-trisphosphate (IP3) and the breakdown of polyphosphoinositides were assessed. In normal membranes, GTP (greater than or equal to 1 mM), GTP gamma S (greater than 10 microM) and GTP gamma S (1 microM) plus CCh (10 microM), but not GDP or GDP beta S, increased phosphatidylinositol 4,5-bisphosphate (PIP2) hydrolysis and IP3 production. GTP gamma S increased IP3 accumulation in a time- and dose-dependent manner, and CCh, which had no effect on phospholipase C activity in the absence of GTP gamma S, potentiated the effects of GTP gamma S. The effect of CCh plus GTP gamma S on IP3 production was inhibited by atropine, had an absolute requirement for nM amounts of Ca2+ and was not affected by pertussis toxin. At higher concentrations (greater than 1 microM), Ca2+ alone induced PIP2 hydrolysis. Short-term exposure (less than 60 min) of the muscle to CCh (100 microM) did not affect the total number (Bmax) of mAChRs nor their affinity (KD) for [3H]-N-methylscopolamine. Desensitization did, however, result in: (1) a loss of the CCh-high affinity binding state of the sphincter mAChRs in a manner analogous to that produced by GTP gamma S; (2) a loss of the ability of GTP gamma S to affect CCh binding to the receptors; and (3) an attenuation of the GTP gamma S plus CCh-stimulated PIP2 hydrolysis. In conclusion, the data presented suggest that, in the iris smooth muscle, G-proteins are involved in the coupling of mAChRs to phospholipase C and that short-term cholinergic desensitization results in (1) the uncoupling of the receptor-G-protein complex and (2) the attenuation of mAChR-activation of phospholipase C.

Reaction pathways and free energy profiles for spontaneous hydrolysis of urea and tetramethylurea: Unexpected substituent effects

PubMed Central

Yao, Min; Tu, Wenlong; Chen, Xi; Zhan, Chang-Guo

2013-01-01

It has been difficult to directly measure the spontaneous hydrolysis rate of urea and, thus, 1,1,3,3-tetramethylurea (Me4U) was used as a model to determine the “experimental” rate constant for urea hydrolysis. The use of Me4U was based on an assumption that the rate of urea hydrolysis should be 2.8 times that of Me4U hydrolysis because the rate of acetamide hydrolysis is 2.8 times that of N,N-dimethyl-acetamide hydrolysis. The present first-principles electronic-structure calculations on the competing non-enzymatic hydrolysis pathways have demonstrated that the dominant pathway is the neutral hydrolysis via the CN addition for both urea (when pH<~11.6) and Me4U (regardless of pH), unlike the non-enzymatic hydrolysis of amides where alkaline hydrolysis is dominant. Based on the computational data, the substituent shift of free energy barrier calculated for the neutral hydrolysis is remarkably different from that for the alkaline hydrolysis, and the rate constant for the urea hydrolysis should be ~1.3×109-fold lower than that (4.2×10−12 s−1) measured for the Me4U hydrolysis. As a result, the rate enhancement and catalytic proficiency of urease should be 1.2×1025 and 3×1027 M−1, respectively, suggesting that urease surpasses proteases and all other enzymes in its power to enhance the rate of reaction. All of the computational results are consistent with available experimental data for Me4U, suggesting that the computational prediction for urea is reliable. PMID:24097048

Dissecting the role of conformational change and membrane binding by the bacterial cell division regulator MinE in the stimulation of MinD ATPase activity.

PubMed

Ayed, Saud H; Cloutier, Adam D; McLeod, Laura J; Foo, Alexander C Y; Damry, Adam M; Goto, Natalie K

2017-12-15

The bacterial cell division regulators MinD and MinE together with the division inhibitor MinC localize to the membrane in concentrated zones undergoing coordinated pole-to-pole oscillation to help ensure that the cytokinetic division septum forms only at the mid-cell position. This dynamic localization is driven by MinD-catalyzed ATP hydrolysis, stimulated by interactions with MinE's anti-MinCD domain. This domain is buried in the 6-β-stranded MinE "closed" structure, but is liberated for interactions with MinD, giving rise to a 4-β-stranded "open" structure through an unknown mechanism. Here we show that MinE-membrane interactions induce a structural change into a state resembling the open conformation. However, MinE mutants lacking the MinE membrane-targeting sequence stimulated higher ATP hydrolysis rates than the full-length protein, indicating that binding to MinD is sufficient to trigger this conformational transition in MinE. In contrast, conformational change between the open and closed states did not affect stimulation of ATP hydrolysis rates in the absence of membrane binding, although the MinD-binding residue Ile-25 is critical for this conformational transition. We therefore propose an updated model where MinE is brought to the membrane through interactions with MinD. After stimulation of ATP hydrolysis, MinE remains bound to the membrane in a state that does not catalyze additional rounds of ATP hydrolysis. Although the molecular basis for this inhibited state is unknown, previous observations of higher-order MinE self-association may explain this inhibition. Overall, our findings have general implications for Min protein oscillation cycles, including those that regulate cell division in bacterial pathogens. © 2017 by The American Society for Biochemistry and Molecular Biology, Inc.

Effects of production and market factors on ethanol profitability for an integrated first and second generation ethanol plant using the whole sugarcane as feedstock

PubMed Central

2014-01-01

Background Sugarcane is an attractive feedstock for ethanol production, especially if the lignocellulosic fraction can also be treated in second generation (2G) ethanol plants. However, the profitability of 2G ethanol is affected by the processing conditions, operating costs and market prices. This study focuses on the minimum ethanol selling price (MESP) and maximum profitability of ethanol production in an integrated first and second generation (1G + 2G) sugarcane-to-ethanol plant. The feedstock used was sugarcane juice, bagasse and leaves. The lignocellulosic fraction was hydrolysed with enzymes. Yields were assumed to be 95% of the theoretical for each of the critical steps in the process (steam pretreatment, enzymatic hydrolysis (EH), fermentation, solid/liquid separation, anaerobic digestion) in order to obtain the best conditions possible for ethanol production, to assess the lowest production costs. Techno-economic analysis was performed for various combinations of process options (for example use of pentoses, addition of leaves), EH conditions (water-insoluble solids (WIS) and residence time), operating cost (enzymes) and market factors (wholesale prices of electricity and ethanol, cost of the feedstock). Results The greatest reduction in 2G MESP was achieved when using the pentoses for the production of ethanol rather than biogas. This was followed, in decreasing order, by higher enzymatic hydrolysis efficiency (EHE), by increasing the WIS to 30% and by a short residence time (48 hours) in the EH. The addition of leaves was found to have a slightly negative impact on 1G + 2G MESP, but the effect on 2G MESP was negligible. Sugarcane price significantly affected 1G + 2G MESP, while the price of leaves had a much lower impact. Net present value (NPV) analysis of the most interesting case showed that integrated 1G + 2G ethanol production including leaves could be more profitable than 1G ethanol, despite the fact that the MESP was higher than in 1G ethanol production. Conclusions A combined 1G + 2G ethanol plant could potentially outperform a 1G plant in terms of NPV, depending on market wholesale prices of ethanol and electricity. Therefore, although it is more expensive than 1G ethanol production, 2G ethanol production can make the integrated 1G + 2G process more profitable. PMID:24559312

Effects of production and market factors on ethanol profitability for an integrated first and second generation ethanol plant using the whole sugarcane as feedstock.

PubMed

Macrelli, Stefano; Galbe, Mats; Wallberg, Ola

2014-02-21

Sugarcane is an attractive feedstock for ethanol production, especially if the lignocellulosic fraction can also be treated in second generation (2G) ethanol plants. However, the profitability of 2G ethanol is affected by the processing conditions, operating costs and market prices. This study focuses on the minimum ethanol selling price (MESP) and maximum profitability of ethanol production in an integrated first and second generation (1G + 2G) sugarcane-to-ethanol plant. The feedstock used was sugarcane juice, bagasse and leaves. The lignocellulosic fraction was hydrolysed with enzymes. Yields were assumed to be 95% of the theoretical for each of the critical steps in the process (steam pretreatment, enzymatic hydrolysis (EH), fermentation, solid/liquid separation, anaerobic digestion) in order to obtain the best conditions possible for ethanol production, to assess the lowest production costs. Techno-economic analysis was performed for various combinations of process options (for example use of pentoses, addition of leaves), EH conditions (water-insoluble solids (WIS) and residence time), operating cost (enzymes) and market factors (wholesale prices of electricity and ethanol, cost of the feedstock). The greatest reduction in 2G MESP was achieved when using the pentoses for the production of ethanol rather than biogas. This was followed, in decreasing order, by higher enzymatic hydrolysis efficiency (EHE), by increasing the WIS to 30% and by a short residence time (48 hours) in the EH. The addition of leaves was found to have a slightly negative impact on 1G + 2G MESP, but the effect on 2G MESP was negligible. Sugarcane price significantly affected 1G + 2G MESP, while the price of leaves had a much lower impact. Net present value (NPV) analysis of the most interesting case showed that integrated 1G + 2G ethanol production including leaves could be more profitable than 1G ethanol, despite the fact that the MESP was higher than in 1G ethanol production. A combined 1G + 2G ethanol plant could potentially outperform a 1G plant in terms of NPV, depending on market wholesale prices of ethanol and electricity. Therefore, although it is more expensive than 1G ethanol production, 2G ethanol production can make the integrated 1G + 2G process more profitable.

Optimization of hydrolysis conditions for bovine plasma protein using response surface methodology.

PubMed

Seo, Hyun-Woo; Jung, Eun-Young; Go, Gwang-Woong; Kim, Gap-Don; Joo, Seon-Tea; Yang, Han-Sul

2015-10-15

The purpose of this study was to establish optimal conditions for the hydrolysis of bovine plasma protein. Response surface methodology was used to model and optimize responses [degree of hydrolysis (DH), 2,2-diphenyl-1-picrydrazyl (DPPH) radical-scavenging activity and Fe(2+)-chelating activity]. Hydrolysis conditions, such as hydrolysis temperature (46.6-63.4 °C), hydrolysis time (98-502 min), and hydrolysis pH (6.32-9.68) were selected as the main processing conditions in the hydrolysis of bovine plasma protein. Optimal conditions for maximum DH (%), DPPH radical-scavenging activity (%) and Fe(2+)-chelating activity (%) of the hydrolyzed bovine plasma protein, were respectively established. We discovered the following three conditions for optimal hydrolysis of bovine plasma: pH of 7.82-8.32, temperature of 54.1 °C, and time of 338.4-398.4 min. We consequently succeeded in hydrolyzing bovine plasma protein under these conditions and confirmed the various desirable properties of optimal hydrolysis. Copyright © 2015 Elsevier Ltd. All rights reserved.

Watch Out for the “Living Dead”: Cell-Free Enzymes and Their Fate

PubMed Central

Baltar, Federico

2018-01-01

Microbes are the engines driving biogeochemical cycles. Microbial extracellular enzymatic activities (EEAs) are the “gatekeepers” of the carbon cycle. The total EEA is the sum of cell-bound (i.e., cell-attached), and dissolved (i.e., cell-free) enzyme activities. Cell-free enzymes make up a substantial proportion (up to 100%) of the total marine EEA. Although we are learning more about how microbial diversity and function (including total EEA) will be affected by environmental changes, little is known about what factors control the importance of the abundant cell-free enzymes. Since cell-attached EEAs are linked to the cell, their fate will likely be linked to the factors controlling the cell’s fate. In contrast, cell-free enzymes belong to a kind of “living dead” realm because they are not attached to a living cell but still are able to perform their function away from the cell; and as such, the factors controlling their activity and fate might differ from those affecting cell-attached enzymes. This article aims to place cell-free EEA into the wider context of hydrolysis of organic matter, deal with recent studies assessing what controls the production, activity and lifetime of cell-free EEA, and what their fate might be in response to environmental stressors. This perspective article advocates the need to go “beyond the living things,” studying the response of cells/organisms to different stressors, but also to study cell-free enzymes, in order to fully constrain the future and evolution of marine biogeochemical cycles. PMID:29354095

Cellobionic acid inhibition of cellobiohydrolase I and cellobiose dehydrogenase

USDA-ARS?s Scientific Manuscript database

End-product inhibition by cellobiose and glucose is a rate-limiting factor in cellulose hydrolysis by cellulases. While cellobiose and glucose inhibition have been extensively investigated, cellobionate inhibition has been minimally studied despite the discovery that accessory proteins such as cello...

How Glucosinolates Affect Generalist Lepidopteran Larvae: Growth, Development and Glucosinolate Metabolism

PubMed Central

Jeschke, Verena; Kearney, Emily E.; Schramm, Katharina; Kunert, Grit; Shekhov, Anton; Gershenzon, Jonathan; Vassão, Daniel G.

2017-01-01

Multiple lepidopteran larvae feed successfully on plants containing glucosinolates despite the diverse array of toxic and deterrent breakdown products, such as isothiocyanates (ITCs), formed upon plant damage. While much is known about how specialist lepidopterans metabolize and tolerate glucosinolates, there is little information about the metabolic fate of these plant defense compounds in specialized herbivores. Employing 13C- and 14C-labeled 4-methylsulfinylbutyl glucosinolate (glucoraphanin), we identified and quantified the major detoxification products of glucosinolates and ITCs in selected specialized and generalist larvae. While specialists prevented glucosinolate hydrolysis or diverted hydrolysis to form nitriles, hydrolysis in generalists proceeded to toxic ITCs, of which a portion were conjugated to glutathione. However, a large amount of ITCs remained unmodified, which may have led to the observed negative effects on growth and development. The performance of two generalist-feeding caterpillars, Spodoptera littoralis (African cotton leafworm) and Mamestra brassicae (cabbage moth) on Arabidopsis thaliana Col-0 and various glucosinolate-deficient mutants was investigated from hatching until pupation. We found that glucosinolates negatively affected larval growth and development, but not survival, with aliphatic glucosinolates having stronger effects than indolic glucosinolates, and the combination of the two glucosinolate types being even more detrimental to growth and development. Curiously, last instar larvae grew better on wild type than on non-glucosinolate-containing plant lines, but this could not be attributed to a change in detoxification rate or feeding behavior. Glucosinolates thus appear to be effective defenses against generalist lepidopteran herbivores at least during most stages of larval development. Nevertheless, the reversal of negative effects in the oldest instar is intriguing, and further investigation of this phenomenon may shed light on how generalists adjust their physiology to feed on diets with many different types of plant defense compounds. PMID:29209354

Pressure Modulation of the Enzymatic Activity of Phospholipase A2, A Putative Membrane-Associated Pressure Sensor.

PubMed

Suladze, Saba; Cinar, Suleyman; Sperlich, Benjamin; Winter, Roland

2015-10-07

Phospholipases A2 (PLA2) catalyze the hydrolysis reaction of sn-2 fatty acids of membrane phospholipids and are also involved in receptor signaling and transcriptional pathways. Here, we used pressure modulation of the PLA2 activity and of the membrane's physical-chemical properties to reveal new mechanistic information about the membrane association and subsequent enzymatic reaction of PLA2. Although the effect of high hydrostatic pressure (HHP) on aqueous soluble and integral membrane proteins has been investigated to some extent, its effect on enzymatic reactions operating at the water/lipid interface has not been explored, yet. This study focuses on the effect of HHP on the structure, membrane binding and enzymatic activity of membrane-associated bee venom PLA2, covering a pressure range up to 2 kbar. To this end, high-pressure Fourier-transform infrared and high-pressure stopped-flow fluorescence spectroscopies were applied. The results show that PLA2 binding to model biomembranes is not significantly affected by pressure and occurs in at least two kinetically distinct steps. Followed by fast initial membrane association, structural reorganization of α-helical segments of PLA2 takes place at the lipid water interface. FRET-based activity measurements reveal that pressure has a marked inhibitory effect on the lipid hydrolysis rate, which decreases by 75% upon compression up to 2 kbar. Lipid hydrolysis under extreme environmental conditions, such as those encountered in the deep sea where pressures up to the kbar-level are encountered, is hence markedly affected by HHP, rendering PLA2, next to being a primary osmosensor, a good candidate for a sensitive pressure sensor in vivo.

DMR (deacetylation and mechanical refining) processing of corn stover achieves high monomeric sugar concentrations (230 g L -1) during enzymatic hydrolysis and high ethanol concentrations (>10% v/v) during fermentation without hydrolysate purification or concentration

DOE PAGES

Chen, Xiaowen; Kuhn, Erik; Jennings, Edward W.; ...

2016-04-01

Distilling and purifying ethanol and other products from second generation lignocellulosic biorefineries adds significant capital and operating costs to biofuel production. The energy usage associated with distillation negatively affects plant gate costs and causes environmental and life-cycle impacts, and the lower titers in fermentation caused by lower sugar concentrations from pretreatment and enzymatic hydrolysis increase energy and water usage and ethanol production costs. In addition, lower ethanol titers increase the volumes required for enzymatic hydrolysis and fermentation vessels increase capital expenditure (CAPEX). Therefore, increasing biofuel titers has been a research focus in renewable biofuel production for several decades. In thismore » work, we achieved approximately 230 g L -1 of monomeric sugars after high solid enzymatic hydrolysis using deacetylation and mechanical refining (DMR) processed corn stover substrates produced at the 100 kg per day scale. The high sugar concentrations and low chemical inhibitor concentrations achieved by the DMR process allowed fermentation to ethanol with titers as high as 86 g L -1, which translates into approximately 10.9% v/v ethanol. To our knowledge, this is the first time that titers greater than 10% v/v ethanol in fermentations derived from corn stover without any sugar concentration or purification steps have been reported. As a result, the potential cost savings from high sugar and ethanol titers achieved by the DMR process are also reported using TEA analysis.« less

DMR (deacetylation and mechanical refining) processing of corn stover achieves high monomeric sugar concentrations (230 g L -1) during enzymatic hydrolysis and high ethanol concentrations (>10% v/v) during fermentation without hydrolysate purification or concentration

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

Chen, Xiaowen; Kuhn, Erik; Jennings, Edward W.

Distilling and purifying ethanol and other products from second generation lignocellulosic biorefineries adds significant capital and operating costs to biofuel production. The energy usage associated with distillation negatively affects plant gate costs and causes environmental and life-cycle impacts, and the lower titers in fermentation caused by lower sugar concentrations from pretreatment and enzymatic hydrolysis increase energy and water usage and ethanol production costs. In addition, lower ethanol titers increase the volumes required for enzymatic hydrolysis and fermentation vessels increase capital expenditure (CAPEX). Therefore, increasing biofuel titers has been a research focus in renewable biofuel production for several decades. In thismore » work, we achieved approximately 230 g L -1 of monomeric sugars after high solid enzymatic hydrolysis using deacetylation and mechanical refining (DMR) processed corn stover substrates produced at the 100 kg per day scale. The high sugar concentrations and low chemical inhibitor concentrations achieved by the DMR process allowed fermentation to ethanol with titers as high as 86 g L -1, which translates into approximately 10.9% v/v ethanol. To our knowledge, this is the first time that titers greater than 10% v/v ethanol in fermentations derived from corn stover without any sugar concentration or purification steps have been reported. As a result, the potential cost savings from high sugar and ethanol titers achieved by the DMR process are also reported using TEA analysis.« less

Impact of magnesium:calcium ratio on calcification of the aortic wall

PubMed Central

2017-01-01

Objective An inverse relationship between serum magnesium concentration and vascular calcification has been reported following observational clinical studies. Moreover, several studies have been suggesting a protective effect of magnesium on the vascular calcification. However, the exact mechanism remains elusive, and investigators have speculated among a myriad of potential actions. The effect of magnesium on calcification of the aortic wall is yet to be investigated. In the present study, the effects of magnesium and calcium on the metabolism of extracellular PPi, the main endogenous inhibitor of vascular calcification, were investigated in the rat aorta. Approach and results Calcium and magnesium have antagonist effects on PPi hydrolysis in the aortic wall. Km and Ki values for PPi hydrolysis in rat aortic rings were 1.1 mmol/L magnesium and 32 μmol/L calcium, respectively, but ATP hydrolysis was not affected with calcium. Calcium deposition in the rat aortic wall dramatically increased when the magnesium concentration was increased (ratio of Mg:Ca = 1:1; 1.5 mmol/L calcium and 1.5 mmol/L magnesium) respect to low magnesium concentration (ratio Mg:Ca = 1:3, 1.5 mmol/L calcium and 0.75 mmol/L magnesium). Conclusion Data from observational clinical studies showing that the serum magnesium concentration is inversely correlated with vascular calcification could be reinterpreted as a compensatory regulatory mechanism that reduces both PPi hydrolysis and vascular calcification. The impact of magnesium in vascular calcification in humans could be studied in association with calcium levels, for example, as the magnesium:calcium ratio. PMID:28570619

Direct Biotransformation of Dioscin into Diosgenin in Rhizome of Dioscorea zingiberensis by Penicillium dioscin.

PubMed

Dong, Jingzhou; Lei, Can; Lu, Dayan; Wang, Ying

2015-06-01

Diosgenin is an important precursor for synthesis of more than 200 steroidal hormone medicines. Rhizome of Dioscorea zingiberensis C. H. Wright (RDZ) contained the highest content of diosgenin in Dioscorea plant species. Diosgenin is traditionally extracted by acid hydrolysis from RDZ. However, the acid hydrolysis process produces massive wastewater which caused serious environment pollution. In this study, diosgenin extraction by direct biotransformation with Penicillium dioscin was investigated. The spawn cultivation conditions were optimized as: Czapeks liquid culture medium without sugar and agar (1,000 ml) + 6.0 g dioscin/6.0 g DL, 30 °C, 36 h; solid fermentation of RDZ: mycelia/RDZ of 0.05 g/kg, 30 °C, 50 h; the yield of diosgenin was over 90 %. Spawn cultivation was crucial for the direct biotransformation. In the spawn cultivation, amount and ratio of dioscin/DL were the key factors to promote biotransformation activity of P. dioscin. This biotransformation method was environment-friendly, simple and energy saving, and might be a potential substitute for acid hydrolysis in diosgenin extraction industry.

Direct Spectroscopic Detection of ATP Turnover Reveals Mechanistic Divergence of ABC Exporters.

PubMed

Collauto, Alberto; Mishra, Smriti; Litvinov, Aleksei; Mchaourab, Hassane S; Goldfarb, Daniella

2017-08-01

We have applied high-field (W-band) pulse electron-nuclear double resonance (ENDOR) and electron-electron double resonance (ELDOR)-detected nuclear magnetic resonance (EDNMR) to characterize the coordination sphere of the Mn 2+ co-factor in the nucleotide binding sites (NBSs) of ABC transporters. MsbA and BmrCD are two efflux transporters hypothesized to represent divergent catalytic mechanisms. Our results reveal distinct coordination of Mn 2+ to ATP and transporter residues in the consensus and degenerate NBSs of BmrCD. In contrast, the coordination of Mn 2+ at the two NBSs of MsbA is similar, which provides a mechanistic rationale for its higher rate constant of ATP hydrolysis relative to BmrCD. Direct detection of vanadate ion, trapped in a high-energy post-hydrolysis intermediate, further supports the notion of asymmetric hydrolysis by the two NBSs of BmrCD. The integrated spectroscopic approach presented here, which link energy input to conformational dynamics, can be applied to a variety of systems powered by ATP turnover. Copyright © 2017 Elsevier Ltd. All rights reserved.

Effects of Impurities in Alkali-Extracted Xylan on Its Enzymatic Hydrolysis to Produce Xylo-Oligosaccharides.

PubMed

Shen, Rui; Li, Hong-Qiang; Zhang, Jie; Xu, Jian

2016-07-01

As the second abundant natural carbohydrate, xylan is normally prepared through alkaline extraction and then used for xylo-oligosaccharides (XOS) production. However, the extracted xylan inevitably contains salt, ethanol, and pigment. In order to investigate the effects of these impurities on XOS production, the alkaline-extracted xylan with different kinds and concentrations of impurities was made and then hydrolyzed using alkaline xylanase (EC 3.2.1.8) to produce XOS. The results showed that a certain concentration of salt (NaCl) promoted the XOS production, while ethanol and pigment inhibited the enzymatic hydrolysis process significantly. The color value mainly ascribed to the phenolic compounds binding to xylan was a key restriction factor in the enzymatic hydrolysis later stage. Using optimal xylan sample (with 10 mg/mL NaCl, color value of 4.6 × 10(5), without ethanol) as substrate, the highest XOS yield of 58.58 % was obtained. As the substrate of XOS production, prepared xylan should contain colored materials and ethanol as less as possible, however, retains appropriate salt.

Insight into the substrate specificity change caused by the Y227H mutation of α-glucosidase III from the European honeybee (Apis mellifera) through molecular dynamics simulations.

PubMed

Na Ayutthaya, Pratchaya Pramoj; Chanchao, Chanpen; Chunsrivirot, Surasak

2018-01-01

Honey from the European honeybee, Apis mellifera, is produced by α-glucosidases (HBGases) and is widely used in food, pharmaceutical, and cosmetic industries. Categorized by their substrate specificities, HBGases have three isoforms: HBGase I, II and III. Previous experimental investigations showed that wild-type HBGase III from Apis mellifera (WT) preferred sucrose to maltose as a substrate, while the Y227H mutant (MT) preferred maltose to sucrose. This mutant can potentially be used for malt hydrolysis because it can efficiently hydrolyze maltose. In this work, to elucidate important factors contributing to substrate specificity of this enzyme and gain insight into how the Y227H mutation causes substrate specificity change, WT and MT homology models were constructed, and sucrose/maltose was docked into active sites of the WT and MT. AMBER14 was employed to perform three independent molecular dynamics runs for these four complexes. Based on the relative binding free energies calculated by the MM-GBSA method, sucrose is better than maltose for WT binding, while maltose is better than sucrose for MT binding. These rankings support the experimentally observed substrate specificity that WT preferred sucrose to maltose as a substrate, while MT preferred maltose to sucrose, suggesting the importance of binding affinity for substrate specificity. We also found that the Y227H mutation caused changes in the proximities between the atoms necessary for sucrose/maltose hydrolysis that may affect enzyme efficiency in the hydrolysis of sucrose/maltose. Moreover, the per-residue binding free energy decomposition results show that Y227/H227 may be a key residue for preference binding of sucrose/maltose in the WT/MT active site. Our study provides important and novel insight into the binding of sucrose/maltose in the active site of Apis mellifera HBGase III and into how the Y227H mutation leads to the substrate specificity change at the molecular level. This knowledge could be beneficial in the design of this enzyme for increased production of desired products.

Substrate-Induced Formation of Ribosomal Decoding Center for Accurate and Rapid Genetic Code Translation.

PubMed

Pavlov, Michael Y; Ehrenberg, Måns

2018-05-20

Accurate translation of genetic information is crucial for synthesis of functional proteins in all organisms. We use recent experimental data to discuss how induced fit affects accuracy of initial codon selection on the ribosome by aminoacyl transfer RNA in ternary complex ( T 3 ) with elongation factor Tu (EF-Tu) and guanosine-5'-triphosphate (GTP). We define actual accuracy ([Formula: see text]) of a particular protein synthesis system as its current accuracy and the effective selectivity ([Formula: see text]) as [Formula: see text] in the limit of zero ribosomal binding affinity for T 3 . Intrinsic selectivity ([Formula: see text]), defined as the upper thermodynamic limit of [Formula: see text], is determined by the free energy difference between near-cognate and cognate T 3 in the pre-GTP hydrolysis state on the ribosome. [Formula: see text] is much larger than [Formula: see text], suggesting the possibility of a considerable increase in [Formula: see text] and [Formula: see text] at negligible kinetic cost. Induced fit increases [Formula: see text] and [Formula: see text] without affecting [Formula: see text], and aminoglycoside antibiotics reduce [Formula: see text] and [Formula: see text] at unaltered [Formula: see text].

Optimization of enzyme complexes for efficient hydrolysis of corn stover to produce glucose.

PubMed

Yu, Xiaoxiao; Liu, Yan; Meng, Jiatong; Cheng, Qiyue; Zhang, Zaixiao; Cui, Yuxiao; Liu, Jiajing; Teng, Lirong; Lu, Jiahui; Meng, Qingfan; Ren, Xiaodong

2015-05-01

Hydrolysis of cellulose to glucose is the critical step for transferring the lignocellulose to the industrial chemicals. For improving the conversion rate of cellulose of corn stover to glucose, the cocktail of celllulase with other auxiliary enzymes and chemicals was studied in this work. Single factor tests and Response Surface Methodology (RSM) were applied to optimize the enzyme mixture, targeting maximum glucose release from corn stover. The increasing rate of glucan-to-glucose conversion got the higher levels while the cellulase was added 1.7μl tween-80/g cellulose, 300μg β-glucosidase/g cellulose, 400μg pectinase/g cellulose and 0.75mg/ml sodium thiosulphate separately in single factor tests. To improve the glucan conversion, the β-glucosidase, pectinase and sodium thiosulphate were selected for next step optimization with RSM. It is showed that the maximum increasing yield was 45.8% at 377μg/g cellulose Novozyme 188, 171μg/g cellulose pectinase and 1mg/ml sodium thiosulphate.

Efficient and low-cost alternative of lipase concentration aiming at the application in the treatment of waste cooking oils.

PubMed

Preczeski, Karina P; Kamanski, Angela B; Scapini, Thamarys; Camargo, Aline F; Modkoski, Tatiani A; Rossetto, Vanusa; Venturin, Bruno; Mulinari, Jéssica; Golunski, Simone M; Mossi, Altemir J; Treichel, Helen

2018-06-01

In this study, we evaluated the concentration of lipases from Aspergillus niger using efficient and low-cost methods aiming at application in the treatment of waste cooking oils. The change in ionic strength of the medium by the addition of salt and precipitation with ethanol increased the specific activity from 2.90 to 28.50 U/mg, resulting in a purification factor of 9.82-fold. The use of acetone resulted in a specific activity of 33.63 U/mg, resulting in a purification factor of 11.60-fold. After that, the concentrated lipase was used in the hydrolysis of waste cooking oil and 753.07 and 421.60 µmol/mL of free fatty acids were obtained for the enzyme precipitated with ethanol and acetone, respectively. The hydrolysis of waste cooking oil catalyzed by homemade purified lipase in ultrasonic media can be considered a pretreatment of oil by converting a significant amount of triglycerides into free fatty acids.

Alkaline-assisted screw press pretreatment affecting enzymatic hydrolysis of wheat straw.

PubMed

Yan, Qingqi; Wang, Yumei; Rodiahwati, Wawat; Spiess, Antje; Modigell, Michael

2017-02-01

Screw press processing of biomass can be considered as a suitable mechanically based pretreatment for biofuel production since it disrupts the structure of lignocellulosic biomass with high shear and pressure forces. The combination with chemical treatment has been suggested to increase the conversion of lignocellulosic biomass to fermentable sugars. Within the study, the synergetic effect of alkaline (sodium hydroxide, NaOH) soaking and screw press pretreatment on wheat straw was evaluated based on, e.g., sugar recovery and energy efficiency. After alkaline soaking (at 0.1 M for 30 min) and sequential screw press pretreatment with various screw press configurations and modified screw barrel, the lignin content of pretreated wheat straw was quantified. In addition, the structure of pretreated wheat straw was investigated by scanning electron microscopy and measurement of specific surface area. It could be shown that removal of lignin is more important than increase of surface area of the biomass to reach a high sugar recovery. The rate constant of the enzymatic hydrolysis increased from 1.1 × 10 -3  1/h for the non-treated material over 2.3 × 10 -3  1/h for the alkaline-soaked material to 26.9 × 10 -3  1/h for alkaline-assisted screw press pretreated material, indicating a nearly 25-fold improvement of the digestibility by the combined chemo-mechanical pretreatment. Finally, the screw configuration was found to be an important factor for improving the sugar recovery and for reducing the specific energy consumption of the screw press pretreatment.

Accelerated hydrolysis of substituted cellulose for potential biofuel production: kinetic study and modeling.

PubMed

Mu, Bingnan; Xu, Helan; Yang, Yiqi

2015-11-01

In this work, kinetics of substitution accelerated cellulose hydrolysis with multiple reaction stages was investigated to lay foundation for mechanism study and molecular design of substituting compounds. High-efficiency hydrolysis of cellulose is critical for cellulose-based bioethanol production. It is known that, substitution could substantially decrease activation energy and increase reaction rate of acidic hydrolysis of glycosidic bonds in cellulose. However, reaction kinetics and mechanism of the accelerated hydrolysis were not fully revealed. In this research, it was proved that substitution therefore accelerated hydrolysis only occurred in amorphous regions of cellulose fibers, and was a process with multiple reaction stages. With molar ratio of substitution less than 1%, the overall hydrolysis rate could be increased for around 10 times. We also quantified the relationship between the hydrolysis rate of individual reaction stage and its major influences, including molar ratio of substitution, activation energy of acidic hydrolysis, pH and temperature. Copyright © 2015 Elsevier Ltd. All rights reserved.

Physicochemical structural changes of cellulosic substrates during enzymatic saccharification

DOE PAGES

Meng, Xianzhi; Yoo, Chang Geun; Li, Mi; ...

2016-12-30

Enzymatic hydrolysis represents one of the major steps and barriers in the commercialization process of converting cellulosic substrates into biofuels and other value added products. It is usually achieved by a synergistic action of enzyme mixture typically consisting of multiple enzymes such as glucanase, cellobiohydrolase and β-glucosidase with different mode of actions. Due to the innate biomass recalcitrance, enzymatic hydrolysis normally starts with an initial fast rate of hydrolysis followed by a rapid decrease of rate toward the end of hydrolysis. With majority of literature studies focusing on the effect of key substrate characteristics on the initial rate or finalmore » yield of enzymatic hydrolysis, information about physicochemical structural changes of cellulosic substrates during enzymatic hydrolysis is still quite limited. Consequently, what slows down the reaction rate toward the end of hydrolysis is not well understood. Lastly, this review highlights recent advances in understanding the structural changes of cellulosic substrates during the hydrolysis process, to better understand the fundamental mechanisms of enzymatic hydrolysis.« less

A process for producing lignin and volatile compounds from hydrolysis liquor.

PubMed

Khazraie, Tooran; Zhang, Yiqian; Tarasov, Dmitry; Gao, Weijue; Price, Jacquelyn; DeMartini, Nikolai; Hupa, Leena; Fatehi, Pedram

2017-01-01

Hot water hydrolysis process is commercially applied for treating wood chips prior to pulping or wood pellet production, while it produces hydrolysis liquor as a by-product. Since the hydrolysis liquor is dilute, the production of value-added materials from it would be challenging. In this study, acidification was proposed as a viable method to extract (1) furfural and acetic acid from hot water hydrolysis liquor and (2) lignin compounds from the liquor. The thermal properties of the precipitates made from the acidification of hydrolysis liquor confirmed the volatile characteristics of precipitates. Membrane dialysis was effective in removing inorganic salts associated with lignin compounds. The purified lignin compounds had a glass transition temperature (Tg) of 180-190 °C, and were thermally stable. The results confirmed that lignin compounds present in hot water hydrolysis liquor had different characteristics. The acidification of hydrolysis liquor primarily removed the volatile compounds from hydrolysis liquor. Based on these results, a process for producing purified lignin and precipitates of volatile compounds was proposed.

Physicochemical structural changes of cellulosic substrates during enzymatic saccharification

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

Meng, Xianzhi; Yoo, Chang Geun; Li, Mi

Enzymatic hydrolysis represents one of the major steps and barriers in the commercialization process of converting cellulosic substrates into biofuels and other value added products. It is usually achieved by a synergistic action of enzyme mixture typically consisting of multiple enzymes such as glucanase, cellobiohydrolase and β-glucosidase with different mode of actions. Due to the innate biomass recalcitrance, enzymatic hydrolysis normally starts with an initial fast rate of hydrolysis followed by a rapid decrease of rate toward the end of hydrolysis. With majority of literature studies focusing on the effect of key substrate characteristics on the initial rate or finalmore » yield of enzymatic hydrolysis, information about physicochemical structural changes of cellulosic substrates during enzymatic hydrolysis is still quite limited. Consequently, what slows down the reaction rate toward the end of hydrolysis is not well understood. Lastly, this review highlights recent advances in understanding the structural changes of cellulosic substrates during the hydrolysis process, to better understand the fundamental mechanisms of enzymatic hydrolysis.« less

Polyhydroxyalkanoates in waste activated sludge enhances anaerobic methane production through improving biochemical methane potential instead of hydrolysis rate.

PubMed

Wang, Qilin; Sun, Jing; Zhang, Chang; Xie, Guo-Jun; Zhou, Xu; Qian, Jin; Yang, Guojing; Zeng, Guangming; Liu, Yiqi; Wang, Dongbo

2016-01-21

Anaerobic sludge digestion is the main technology for sludge reduction and stabilization prior to sludge disposal. Nevertheless, methane production from anaerobic digestion of waste activated sludge (WAS) is often restricted by the poor biochemical methane potential and slow hydrolysis rate of WAS. This work systematically investigated the effect of PHA levels of WAS on anaerobic methane production, using both experimental and mathematical modeling approaches. Biochemical methane potential tests showed that methane production increased with increased PHA levels in WAS. Model-based analysis suggested that the PHA-based method enhanced methane production by improving biochemical methane potential of WAS, with the highest enhancement being around 40% (from 192 to 274 L CH4/kg VS added; VS: volatile solid) when the PHA levels increased from 21 to 143 mg/g VS. In contrast, the hydrolysis rate (approximately 0.10 d(-1)) was not significantly affected by the PHA levels. Economic analysis suggested that the PHA-based method could save $1.2/PE/y (PE: population equivalent) in a typical wastewater treatment plant (WWTP). The PHA-based method can be easily integrated into the current WWTP to enhance methane production, thereby providing a strong support to the on-going paradigm shift in wastewater management from pollutant removal to resource recovery.

Comparative studies on the alcohol types presence in Gracilaria sp. and rice fermentation using Sasad

NASA Astrophysics Data System (ADS)

Mansa, R.; Mansuit, H.; Sipaut, C. S.; Yee, C. F.; Yasir, S. M.

2016-06-01

Alternative fuel sources such as biofuels are needed in order to overcome environmental problem caused by fossil fuel consumption. Currently, most biofuel are produced from land based crops and there is a possibility that marine biomass such as macroalgae can be an alternative source for biofuel production. The carbohydrate in macroalgae can be broken down into simple sugar through thermo-chemical hydrolysis and enzymatic hydrolysis. Dilute-acid hydrolysis was believed to be the most available and affordable method. However, the process may release inhibitors which would affect alcohol yield from fermentation. Thus, this work was aimed at investigating if it is possible to avoid this critical pre-treatment step in macroalgae fermentation process by using Sasad, a local Sabahan fermentation agent and to compare the yield with rice wine fermentation. This work hoped to determine and compare the alcohol content from Gracilaria sp. and rice fermentation with Sasad. Rice fermentation was found containing ethanol and 2 - methyl - 1 - propanol. Fermentation of Gracilaria sp. had shown the positive presence of 3 - methyl - 1 - butanol. It was found that Sasad can be used as a fermentation agent for bioalcohol production from Gracilaria sp. without the need for a pretreatment step. However further investigations are needed to determine if pre-treatment would increase the yield of alcohol.

GM2-ganglioside metabolism in hexosaminidase A deficiency states: determination in situ using labeled GM2 added to fibroblast cultures

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

Raghavan, S.S.; Krusell, A.; Krusell, J.

1985-11-01

To clarify the relationship between hexosaminidase A (HEX A) activity and GM2-ganglioside hydrolysis in atypical clinical situations of HEX A deficiency, we have developed a simple method to assess GM2-ganglioside metabolism in cultured fibroblasts utilizing GM2 labeled with tritium in the sphingosine portion of the molecule. The radioactive lipid is added to the media of cultured skin fibroblasts, and after 10 days the cells are thoroughly washed, then harvested, and their lipid composition analyzed by HPLC. The degree of hydrolysis of the ingested GM2 is determined by comparing the amount of radioactive counts recovered in undegraded substrate with total cellularmore » radioactivity. A deficiency in GM2-ganglioside hydrolysis was demonstrated in seven HEX A-deficient adults with neurological signs and in two healthy-appearing adolescents with older affected siblings. In each case, an analysis of endogenous monosialoganglioside composition revealed an increase in GM2-ganglioside, confirming the presence of a block in the metabolism of GM2. No defect in GM2-catabolism was found in four other healthy individuals with HEX A deficiency. This method of assay is especially helpful in the evaluation of atypical cases of HEX A deficiency for the definitive diagnosis of GM2-gangliosidosis.« less

Exploring the A22-Bacterial Actin MreB Interaction through Molecular Dynamics Simulations.

PubMed

Awuni, Yaw; Jiang, Shimin; Robinson, Robert C; Mu, Yuguang

2016-09-22

MreB is an actin-like cytoskeleton protein that plays a vital role in the maintenance of the rod-shaped morphology of many bacteria. S-(3,4-Dichlorobenzyl) isothiourea (A22) is an antibiotic-like small molecule that perturbs the rod cell shape and has been suggested to inhibit MreB by targeting ATP hydrolysis. However, without the elucidation of the structure of the ATP-bound state of MreB in the presence of A22, the mechanism of A22 inhibition is still not clear. Here we apply conventional molecular dynamics simulations to explore the dynamics of the active site of MreB in complex with A22 and different nucleotides. We observe that hydrogen bonding between A22 and the catalytic Glu140 residue is not favored in the ATP-A22-bound state of MreB. Water dynamics analysis in the MreB active site reveals that in the presence of A22 water molecules are able to occupy positions suitable for ATP hydrolysis. Overall, our results are consistent with a mechanism in which A22 affects MreB polymerization/depolymerization dynamics in part through slowing phosphate release rather than by inhibiting ATP hydrolysis. These data can be incorporated in the design/development of the next generation of MreB inhibitors.

Polyhydroxyalkanoates in waste activated sludge enhances anaerobic methane production through improving biochemical methane potential instead of hydrolysis rate

PubMed Central

Wang, Qilin; Sun, Jing; Zhang, Chang; Xie, Guo-Jun; Zhou, Xu; Qian, Jin; Yang, Guojing; Zeng, Guangming; Liu, Yiqi; Wang, Dongbo

2016-01-01

Anaerobic sludge digestion is the main technology for sludge reduction and stabilization prior to sludge disposal. Nevertheless, methane production from anaerobic digestion of waste activated sludge (WAS) is often restricted by the poor biochemical methane potential and slow hydrolysis rate of WAS. This work systematically investigated the effect of PHA levels of WAS on anaerobic methane production, using both experimental and mathematical modeling approaches. Biochemical methane potential tests showed that methane production increased with increased PHA levels in WAS. Model-based analysis suggested that the PHA-based method enhanced methane production by improving biochemical methane potential of WAS, with the highest enhancement being around 40% (from 192 to 274 L CH4/kg VS added; VS: volatile solid) when the PHA levels increased from 21 to 143 mg/g VS. In contrast, the hydrolysis rate (approximately 0.10 d−1) was not significantly affected by the PHA levels. Economic analysis suggested that the PHA-based method could save $1.2/PE/y (PE: population equivalent) in a typical wastewater treatment plant (WWTP). The PHA-based method can be easily integrated into the current WWTP to enhance methane production, thereby providing a strong support to the on-going paradigm shift in wastewater management from pollutant removal to resource recovery. PMID:26791952

Novel DDR Processing of Corn Stover Achieves High Monomeric Sugar Concentrations from Enzymatic Hydrolysis (230 g/L) and High Ethanol Concentration (10% v/v) During Fermentation

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

Chen, Xiaowen; Jennings, Ed; Shekiro, Joe

Distilling and purifying ethanol, butanol, and other products from second and later generation lignocellulosic biorefineries adds significant capital and operating cost for biofuels production. The energy costs associated with distillation affects plant gate and life cycle analysis costs. Lower titers in fermentation due to lower sugar concentrations from pretreatment increase both energy and production costs. In addition, higher titers decrease the volumes required for enzymatic hydrolysis and fermentation vessels. Therefore, increasing biofuels titers has been a research focus in renewable biofuels production for several decades. In this work, we achieved over 200 g/L of monomeric sugars after high solids enzymaticmore » hydrolysis using the novel deacetylation and disc refining (DDR) process on corn stover. The high sugar concentrations and low chemical inhibitor concentrations from the DDR process allowed ethanol titers as high as 82 g/L in 22 hours, which translates into approximately 10 vol% ethanol. To our knowledge, this is the first time that 10 vol% ethanol in fermentation derived from corn stover without any sugar concentration or purification steps has been reported. Techno-economic analysis shows the higher titer ethanol achieved from the DDR process could significantly reduce the minimum ethanol selling price from cellulosic biomass.« less

Force Generation in Single Conventional Actomyosin Complexes under High Dynamic Load

PubMed Central

Takagi, Yasuharu; Homsher, Earl E.; Goldman, Yale E.; Shuman, Henry

2006-01-01

The mechanical load borne by a molecular motor affects its force, sliding distance, and its rate of energy transduction. The control of ATPase activity by the mechanical load on a muscle tunes its efficiency to the immediate task, increasing ATP hydrolysis as the power output increases at forces less than isometric (the Fenn effect) and suppressing ATP hydrolysis when the force is greater than isometric. In this work, we used a novel ‘isometric’ optical clamp to study the mechanics of myosin II molecules to detect the reaction steps that depend on the dynamic properties of the load. An actin filament suspended between two beads and held in separate optical traps is brought close to a surface that is sparsely coated with motor proteins on pedestals of silica beads. A feedback system increases the effective stiffness of the actin by clamping the force on one of the beads and moving the other bead electrooptically. Forces measured during actomyosin interactions are increased at higher effective stiffness. The results indicate that single myosin molecules transduce energy nearly as efficiently as whole muscle and that the mechanical control of the ATP hydrolysis rate is in part exerted by reversal of the force-generating actomyosin transition under high load without net utilization of ATP. PMID:16326899

Hydrolysis of Mg(BH4)2 and its coordination compounds as a way to obtain hydrogen

NASA Astrophysics Data System (ADS)

Solovev, Mikhail V.; Chashchikhin, Oleg V.; Dorovatovskii, Pavel V.; Khrustalev, Victor N.; Zyubin, A. S.; Zyubina, T. S.; Kravchenko, O. V.; Zaytsev, Alexey A.; Dobrovolsky, Yu. A.

2018-02-01

Three ligand-stabilized Mg(BH4)2-based complexes have been synthesized and evaluated as potential hydrogen storage media for portable fuel cell applications. The new borohydrides: Mg(BH4)2 × 0.5Et2O and Mg(BH4)2 × diglyme (diglyme - CH3O(CH2)2O(CH2)2OCH3) have been synthesized and examined by X-ray single crystal diffraction method. Hydrolysis reactions of the compounds liberate hydrogen in quantities ranging from 46 to 96% of the theoretical yield. The hydrolysis of Mg(BH4)2 and other borohydrides is also accompanied by the diborane formation. The amount of liberated diborane depends on the Mg-coordination environment. To explain this fact quantum-chemical calculations have been performed. It is shown that formation of Mg-O-Mg-bridges enables the side process of diborane generation. It means that the size and denticity of the ligand directly affects the amount of released diborane. In general, the larger the ligand and the higher its denticity, the smaller is amount of diborane produced. The new compound Mg(BH4)2 × diglyme decomposes without diborane formation that allows one to be considered as a new promising chemical hydrogen storage compound for the practical usage.

Polyhydroxyalkanoates in waste activated sludge enhances anaerobic methane production through improving biochemical methane potential instead of hydrolysis rate

NASA Astrophysics Data System (ADS)

Wang, Qilin; Sun, Jing; Zhang, Chang; Xie, Guo-Jun; Zhou, Xu; Qian, Jin; Yang, Guojing; Zeng, Guangming; Liu, Yiqi; Wang, Dongbo

2016-01-01

Anaerobic sludge digestion is the main technology for sludge reduction and stabilization prior to sludge disposal. Nevertheless, methane production from anaerobic digestion of waste activated sludge (WAS) is often restricted by the poor biochemical methane potential and slow hydrolysis rate of WAS. This work systematically investigated the effect of PHA levels of WAS on anaerobic methane production, using both experimental and mathematical modeling approaches. Biochemical methane potential tests showed that methane production increased with increased PHA levels in WAS. Model-based analysis suggested that the PHA-based method enhanced methane production by improving biochemical methane potential of WAS, with the highest enhancement being around 40% (from 192 to 274 L CH4/kg VS added; VS: volatile solid) when the PHA levels increased from 21 to 143 mg/g VS. In contrast, the hydrolysis rate (approximately 0.10 d-1) was not significantly affected by the PHA levels. Economic analysis suggested that the PHA-based method could save $1.2/PE/y (PE: population equivalent) in a typical wastewater treatment plant (WWTP). The PHA-based method can be easily integrated into the current WWTP to enhance methane production, thereby providing a strong support to the on-going paradigm shift in wastewater management from pollutant removal to resource recovery.

Bioethanol production: an integrated process of low substrate loading hydrolysis-high sugars liquid fermentation and solid state fermentation of enzymatic hydrolysis residue.

PubMed

Chu, Qiulu; Li, Xin; Ma, Bin; Xu, Yong; Ouyang, Jia; Zhu, Junjun; Yu, Shiyuan; Yong, Qiang

2012-11-01

An integrated process of enzymatic hydrolysis and fermentation was investigated for high ethanol production. The combination of enzymatic hydrolysis at low substrate loading, liquid fermentation of high sugars concentration and solid state fermentation of enzymatic hydrolysis residue was beneficial for conversion of steam explosion pretreated corn stover to ethanol. The results suggested that low substrate loading hydrolysis caused a high enzymatic hydrolysis yield; the liquid fermentation of about 200g/L glucose by Saccharomyces cerevisiae provided a high ethanol concentration which could significantly decrease cost of the subsequent ethanol distillation. A solid state fermentation of enzymatic hydrolysis residue was combined, which was available to enhance ethanol production and cellulose-to-ethanol conversion. The results of solid state fermentation demonstrated that the solid state fermentation process accompanied by simultaneous saccharification and fermentation. Copyright © 2012 Elsevier Ltd. All rights reserved.

Enzymatic hydrolysis of lignocellulosic biomass by Kitasatospora sp. to produce xylo-oligosaccharides (XOS)

NASA Astrophysics Data System (ADS)

Rahmani, Nanik; Jannah, Alifah Mafatikhul; Lisdiyanti, Puspita; Prasetya, Bambang; Yopi

2017-11-01

The optimizations of enzymatic hydrolysis to produce of xylo-oligosaccharides (XOs) from three different lignocellulosic biomasses were investigated. Sugarcane bagasse, oil palm empty fruit bunch, and rice straw contain rich hemicelluloses especially hetero-xylan which can be hydrolyzes by endo-xylanase enzyme. Enzymatic hydrolysis of sugarcane bagasse by endo-xylanase from Kitasatospora sp. was optimum at temperature hydrolysis 30 °C using 16 U of enzyme concentrations and 4 % substrate concentrations, while oil palm empty fruit bunchwas optimum at temperature hydrolysis 30 °C using 16 U of enzyme concentrations and 5 % substrate concentrations, and rice straw was optimum at 40 °C temperature hydrolysis using 16 U of enzyme concentrations and 4 % substrate concentrations. The hydrolysis products were analyzed by TLC and HPLC. The main product hydrolysis for sugarcane bagasse, oil palm empty fruit bunch and rice straw are xylobiose.

[Impact of liquid volume of recycled methanogenic effluent on anaerobic hydrolysis].

PubMed

Hao, Li-ping; Lü, Fan; He, Pin-jing; Shao, Li-ming

2008-09-01

Methanogenic effluent was recycled to regulate hydrolysis during two-phase anaerobic digestion of organic solid wastes. In order to study the impact of recycled effluent's volume on hydrolysis, four hydrolysis reactors filled with vegetable and flower wastes were constructed, with different liquid volumes of recycled methanogenic effluent, i.e., 0.1, 0.5, 1.0, 2.0 m3/(m3 x d), respectively. The parameters related to hydrolytic environment (pH, alkalinity, ORP, concentrations of ammonia and reducing sugar), microbial biomass and hydrolysis efficiency (accumulated SCOD, accumulated reducing sugar, and hydrolysis rate constants) were monitored. This research shows that recycling methanogenic effluent into the hydrolysis reactor can enhance its buffer capability and operation stability; higher recycled volume is favorable for microbial anabolism and further promotes hydrolysis. After 9 days of reaction, the accumulated SCOD in the hydrolytic effluent reach 334, 407, 413, 581 mg/g at recycled volumes of 0.1, 0.5, 1.0, 2.0 m3/(m3 x d) and their first-order hydrolysis rate kinetic constants are 0.065, 0.083, 0.089, 0.105 d(-1), respectively.

Effects of pH on the Production of Phosphate and Pyrophosphate by Matrix Vesicles' Biomimetics

PubMed Central

Simão, Ana Maria S.; Bolean, Maytê; Hoylaerts, Marc F.; Millán, José Luis; Ciancaglini, Pietro

2013-01-01

During endochondral bone formation, chondrocytes and osteoblasts synthesize and mineralize the extracellular matrix through a process that initiates within matrix vesicles (MVs) and ends with bone mineral propagation onto the collagenous scaffold. pH gradients have been identified in the growth plate of long bones, but how pH changes affect the initiation of skeletal mineralization is not known. Tissue-nonspecific alkaline phosphatase (TNAP) degrades extracellular inorganic pyrophosphate (ePPi), a mineralization inhibitor produced by ectonucleotide pyrophosphatase/ phosphodiesterase-1 (NPP1), while contributing Pi from ATP to initiate mineralization. TNAP and NPP1, alone or combined, were reconstituted in dipalmitoylphosphatidylcholine (DPPC) liposomes to mimic the microenvironment of MVs. The hydrolysis of ATP, ADP, AMP and PPi was studied at pH 8 and 9 and compared to the data determined at pH 7.4. While catalytic efficiencies in general were higher at alkaline pH, PPi hydrolysis was maximal at pH 8 and indicated a preferential utilization of PPi over ATP, at pH 8 versus 9. In addition, all proteoliposomes induced mineral formation when incubated in a synthetic cartilage lymph (SCL) containing 1 mM ATP as substrate and amorphous calcium phosphate (ACP) or calciumphosphate- phosphatidylserine complexes (PS-CPLX) as nucleators. Propagation of mineralization was significantly more efficient at pHs 7.5 and 8 than at pH 9. Since a slight pH elevation from 7.4 to 8 promotes considerably more hydrolysis of ATP, ADP and AMP primarily by TNAP, this small pH change facilitates mineralization, especially via upregulated PPi hydrolysis by both NPP1 and TNAP, further elevating the Pi/PPi ratio, thus enhancing bone mineralization. PMID:23942722

Effects of pH on the production of phosphate and pyrophosphate by matrix vesicles' biomimetics.

PubMed

Simão, Ana Maria S; Bolean, Maytê; Hoylaerts, Marc F; Millán, José Luis; Ciancaglini, Pietro

2013-09-01

During endochondral bone formation, chondrocytes and osteoblasts synthesize and mineralize the extracellular matrix through a process that initiates within matrix vesicles (MVs) and ends with bone mineral propagation onto the collagenous scaffold. pH gradients have been identified in the growth plate of long bones, but how pH changes affect the initiation of skeletal mineralization is not known. Tissue-nonspecific alkaline phosphatase (TNAP) degrades extracellular inorganic pyrophosphate (PPi), a mineralization inhibitor produced by ectonucleotide pyrophosphatase/phosphodiesterase-1 (NPP1), while contributing Pi from ATP to initiate mineralization. TNAP and NPP1, alone or combined, were reconstituted in dipalmitoylphosphatidylcholine liposomes to mimic the microenvironment of MVs. The hydrolysis of ATP, ADP, AMP, and PPi was studied at pH 8 and 9 and compared to the data determined at pH 7.4. While catalytic efficiencies in general were higher at alkaline pH, PPi hydrolysis was maximal at pH 8 and indicated a preferential utilization of PPi over ATP at pH 8 versus 9. In addition, all proteoliposomes induced mineral formation when incubated in a synthetic cartilage lymph containing 1 mM ATP as substrate and amorphous calcium phosphate or calcium-phosphate-phosphatidylserine complexes as nucleators. Propagation of mineralization was significantly more efficient at pH 7.5 and 8 than at pH 9. Since a slight pH elevation from 7.4 to 8 promotes considerably more hydrolysis of ATP, ADP, and AMP primarily by TNAP, this small pH change facilitates mineralization, especially via upregulated PPi hydrolysis by both NPP1 and TNAP, further elevating the Pi/PPi ratio, thus enhancing bone mineralization.

Photocatalytic degradation of 4-amino-6-chlorobenzene-1,3-disulfonamide stable hydrolysis product of hydrochlorothiazide: Detection of intermediates and their toxicity.

PubMed

Armaković, Sanja J; Armaković, Stevan; Četojević-Simin, Dragana D; Šibul, Filip; Abramović, Biljana F

2018-02-01

In this work we have investigated in details the process of degradation of the 4-amino-6-chlorobenzene-1,3-disulfonamide (ABSA), stable hydrolysis product of frequently used pharmaceutical hydrochlorothiazide (HCTZ), as one of the most ubiquitous contaminants in the sewage water. The study encompassed investigation of degradation by hydrolysis, photolysis, and photocatalysis employing commercially available TiO 2 Degussa P25 catalyst. The process of direct photolysis and photocatalytic degradation were investigated under different type of lights. Detailed insights into the reactive properties of HCTZ and ABSA have been obtained by density functional theory calculations and molecular dynamics simulations. Specifically, preference of HCTZ towards hydrolysis was confirmed experimentally and explained using computational study. Results obtained in this study indicate very limited efficiency of hydrolytic and photolytic degradation in the case of ABSA, while photocatalytic degradation demonstrated great potential. Namely, after 240 min of photocatalytic degradation, 65% of ABSA was mineralizated in water/TiO 2 suspension under SSI, while the nitrogen was predominantly present as NH 4 + . Reaction intermediates were studied and a number of them were detected using LC-ESI-MS/MS. This study also involves toxicity assessment of HCTZ, ABSA, and their mixtures formed during the degradation processes towards mammalian cell lines (rat hepatoma, H-4-II-E, human colon adenocarcinoma, HT-29, and human fetal lung, MRC-5). Toxicity assessments showed that intermediates formed during the process of photocatalysis exerted only mild cell growth effects in selected cell lines, while direct photolysis did not affect cell growth. Copyright © 2017 Elsevier Ltd. All rights reserved.

Structure and catalytic mechanism of LigI: insight into the amidohydrolase enzymes of cog3618 and lignin degradation.

PubMed

Hobbs, Merlin Eric; Malashkevich, Vladimir; Williams, Howard J; Xu, Chengfu; Sauder, J Michael; Burley, Stephen K; Almo, Steven C; Raushel, Frank M

2012-04-24

LigI from Sphingomonas paucimobilis catalyzes the reversible hydrolysis of 2-pyrone-4,6-dicarboxylate (PDC) to 4-oxalomesaconate and 4-carboxy-2-hydroxymuconate in the degradation of lignin. This protein is a member of the amidohydrolase superfamily of enzymes. The protein was expressed in Escherichia coli and then purified to homogeneity. The purified recombinant enzyme does not contain bound metal ions, and the addition of metal chelators or divalent metal ions to the assay mixtures does not affect the rate of product formation. This is the first enzyme from the amidohydrolase superfamily that does not require a divalent metal ion for catalytic activity. The kinetic constants for the hydrolysis of PDC are 340 s(-1) and 9.8 × 10(6) M(-1) s(-1) (k(cat) and k(cat)/K(m), respectively). The pH dependence on the kinetic constants suggests that a single active site residue must be deprotonated for the hydrolysis of PDC. The site of nucleophilic attack was determined by conducting the hydrolysis of PDC in (18)O-labeled water and subsequent (13)C nuclear magnetic resonance analysis. The crystal structures of wild-type LigI and the D248A mutant in the presence of the reaction product were determined to a resolution of 1.9 Å. The C-8 and C-11 carboxylate groups of PDC are coordinated within the active site via ion pair interactions with Arg-130 and Arg-124, respectively. The hydrolytic water molecule is activated by the transfer of a proton to Asp-248. The carbonyl group of the lactone substrate is activated by electrostatic interactions with His-180, His-31, and His-33.

Degradation of Biofumigant Isothiocyanates and Allyl Glucosinolate in Soil and Their Effects on the Microbial Community Composition

PubMed Central

Hanschen, Franziska S.; Yim, Bunlong; Winkelmann, Traud; Smalla, Kornelia; Schreiner, Monika

2015-01-01

Brassicales species rich in glucosinolates are used for biofumigation, a process based on releasing enzymatically toxic isothiocyanates into the soil. These hydrolysis products are volatile and often reactive compounds. Moreover, glucosinolates can be degraded also without the presence of the hydrolytic enzyme myrosinase which might contribute to bioactive effects. Thus, in the present study the stability of Brassicaceae plant-derived and pure glucosinolates hydrolysis products was studied using three different soils (model biofumigation). In addition, the degradation of pure 2-propenyl glucosinolate was investigated with special regard to the formation of volatile breakdown products. Finally, the influence of pure glucosinolate degradation on the bacterial community composition was evaluated using denaturing gradient gel electrophoresis of 16S rRNA gene amplified from total community DNA. The model biofumigation study revealed that the structure of the hydrolysis products had a significant impact on their stability in the soil but not the soil type. Following the degradation of pure 2-propenyl glucosinolate in the soils, the nitrile as well as the isothiocyanate can be the main degradation products, depending on the soil type. Furthermore, the degradation was shown to be both chemically as well as biologically mediated as autoclaving reduced degradation. The nitrile was the major product of the chemical degradation and its formation increased with iron content of the soil. Additionally, the bacterial community composition was significantly affected by adding pure 2-propenyl glucosinolate, the effect being more pronounced than in treatments with myrosinase added to the glucosinolate. Therefore, glucosinolates can have a greater effect on soil bacterial community composition than their hydrolysis products. PMID:26186695

Kinetics of PTEN-mediated PI(3,4,5)P3 hydrolysis on solid supported membranes

PubMed Central

Liu, Chun; Deb, Sanghamitra; Ferreira, Vinicius S.; Xu, Eric; Baumgart, Tobias

2018-01-01

Phosphatidylinositides play important roles in cellular signaling and migration. Phosphatidylinositol-3,4,5-trisphosphate (PI(3,4,5)P3) is an important phosphatidylinositide because it acts as a secondary messenger to trigger cell movement and proliferation. A high level of PI(3,4,5)P3 at the plasma membrane is known to contribute to tumorigenesis. One key enzyme that regulates PI(3,4,5)P3 levels at the plasma membrane is phosphatase and tensin homologue deleted on chromosome 10 (PTEN), which dephosphorylates PI(3,4,5)P3 through hydrolysis to form phosphatidylinositol-4,5-bisphosphate (PI(4,5)P2). It has been reported that PI(4,5)P2 is involved in positive feedback in the PI(3,4,5)P3 hydrolysis by PTEN. However, how PI(3,4,5)P3 dephosphorylation by PTEN is regulated, is still under debate. How other PI(3,4,5)P3-binding proteins affect the dephosphorylation kinetics catalyzed by PTEN also remains unclear. Here, we develop a fluorescent-protein biosensor approach to study how PI(3,4,5)P3 dephosphorylation is regulated by PTEN as well as its membrane-mediated feedback mechanisms. Our observation of sigmoidal kinetics of the PI(3,4,5)P3 hydrolysis reaction supports the notion of autocatalysis in PTEN function. We developed a kinetic model to describe the observed reaction kinetics, which allowed us to i) distinguish between membrane-recruitment and allosteric activation of PTEN by PI(4,5)P2, ii) account for the influence of the biosensor on the observed reaction kinetics, and iii) demonstrate that all of these mechanisms contribute to the kinetics of PTEN-mediated catalysis. PMID:29447222

Kinetics of PTEN-mediated PI(3,4,5)P3 hydrolysis on solid supported membranes.

PubMed

Liu, Chun; Deb, Sanghamitra; Ferreira, Vinicius S; Xu, Eric; Baumgart, Tobias

2018-01-01

Phosphatidylinositides play important roles in cellular signaling and migration. Phosphatidylinositol-3,4,5-trisphosphate (PI(3,4,5)P3) is an important phosphatidylinositide because it acts as a secondary messenger to trigger cell movement and proliferation. A high level of PI(3,4,5)P3 at the plasma membrane is known to contribute to tumorigenesis. One key enzyme that regulates PI(3,4,5)P3 levels at the plasma membrane is phosphatase and tensin homologue deleted on chromosome 10 (PTEN), which dephosphorylates PI(3,4,5)P3 through hydrolysis to form phosphatidylinositol-4,5-bisphosphate (PI(4,5)P2). It has been reported that PI(4,5)P2 is involved in positive feedback in the PI(3,4,5)P3 hydrolysis by PTEN. However, how PI(3,4,5)P3 dephosphorylation by PTEN is regulated, is still under debate. How other PI(3,4,5)P3-binding proteins affect the dephosphorylation kinetics catalyzed by PTEN also remains unclear. Here, we develop a fluorescent-protein biosensor approach to study how PI(3,4,5)P3 dephosphorylation is regulated by PTEN as well as its membrane-mediated feedback mechanisms. Our observation of sigmoidal kinetics of the PI(3,4,5)P3 hydrolysis reaction supports the notion of autocatalysis in PTEN function. We developed a kinetic model to describe the observed reaction kinetics, which allowed us to i) distinguish between membrane-recruitment and allosteric activation of PTEN by PI(4,5)P2, ii) account for the influence of the biosensor on the observed reaction kinetics, and iii) demonstrate that all of these mechanisms contribute to the kinetics of PTEN-mediated catalysis.

Hydrolysis Batteries: Generating Electrical Energy during Hydrogen Absorption.

PubMed

Xiao, Rui; Chen, Jun; Fu, Kai; Zheng, Xinyao; Wang, Teng; Zheng, Jie; Li, Xingguo

2018-02-19

The hydrolysis reaction of aluminum can be decoupled into a battery by pairing an Al foil with a Pd-capped yttrium dihydride (YH 2 -Pd) electrode. This hydrolysis battery generates a voltage around 0.45 V and leads to hydrogen absorption into the YH 2 layer. This represents a new hydrogen absorption mechanism featuring electrical energy generation during hydrogen absorption. The hydrolysis battery converts 8-15 % of the thermal energy of the hydrolysis reaction into usable electrical energy, leading to much higher energy efficiency compared to that of direct hydrolysis. © 2018 Wiley-VCH Verlag GmbH & Co. KGaA, Weinheim.

Nucleotide and bivalent cation specificity of the insulin-granule proton translocase.

PubMed Central

Hutton, J C; Peshavaria, M

1983-01-01

1. The nucleotide and bivalent cation specificity of the proton translocase activity of insulin secretory granules was investigated by assessing the inhibitor-sensitive rates of nucleotide hydrolysis by these organelles in relation to their chemiosmotic properties. 2. The relative rates of nucleotide hydrolysis by freeze/thawed granule preparations were: Mg2+ATP (100%) greater than Mg2+GTP (55%) greater than Mg2+UTP (48%) greater than Mg2+ITP (44%) greater than Mg2+CTP (23%) greater than Mg2+TTP (20%), and by intact granules were: Mg2+ATP (100%) greater than Mg2+ITP (74%) greater than Mg2+GTP (60%) greater than Mg2+CTP (35%). Mg2+ATP, Mg2+GTP and Mg2+ITP hydrolyses were inhibited by tributyltin and stimulated, in intact granules, by the protonophore carbonyl cyanide p-trifluoromethoxyphenylhydrazone; Mg2+CTP hydrolysis was not markedly affected by these compounds. Correspondingly, only Mg2+ATP, Mg2+GTP and Mg2+ITP produced large changes in the delta psi and delta mu H+ across the granule membrane. 3. The relative rates of maximal ATPase activity stimulated by bivalent cations in freeze/thawed granule preparations were: Mg2+ (100%) greater than Mn2+ (82%) greater than Ca2+ (40%) greater than Co2+ (36%) greater than Zn2+ (0%), and in intact granules were: Mg2+ (100%) greater than Mn2+ (85%) greater than Co2+ (61%) greater than Ca2+ (42%). Tributyltin and carbonyl cyanide p-trifluoromethoxyphenylhydrazone affected Mg2+-, Mn2+- and Co2+-activated, but not Ca2+-activated, ATP hydrolysis. Correspondingly, only Mg2+, Mn2+ and Co2+ supported the generation of a delta psi and delta mu H+ across granule membranes in the presence of ATP. 4. The results were consistent with a single proton translocase that had its catalytic site exposed on the external face of the granule membrane. The indicated specificity (Mg2+ATP = Mn2+ATP greater than Co2+ATP greater than Mg2+GTP greater than Mg2+ITP) was similar to that of enzymes described in membrane fractions prepared from adenohypophyseal tissue, adrenal chromaffin granules and yeast vacuoles. The insulin-granule activity thus appears to be a type of proton translocase, which is characteristic of intracellular storage vesicles in eukaryotic cells. PMID:6303313

The hydrolysis of proteins by microwave energy

PubMed Central

Margolis, Sam A.; Jassie, Lois; Kingston, H. M.

1991-01-01

Microwave energy, at manually-adjusted, partial power settings has been used to hydrolyse bovine serum albumin at 125 °C. Hydrolysis was complete within 2 h, except for valine and isoleucine which were completely liberated within 4 h. The aminoacid destruction was less than that observed at similar hydrolysis conditions with other methods and complete hydrolysis was achieved more rapidly. These results provide a basis for automating the process of amino-acid hydrolysis. PMID:18924889

Depolymerization dynamics of individual filaments of bacterial cytoskeletal protein FtsZ

PubMed Central

Mateos-Gil, Pablo; Paez, Alfonso; Hörger, Ines; Rivas, Germán; Vicente, Miguel; Tarazona, Pedro; Vélez, Marisela

2012-01-01

We report observation and analysis of the depolymerization filaments of the bacterial cytoskeletal protein FtsZ (filament temperature-sensitive Z) formed on a mica surface. At low concentration, proteins adsorbed on the surface polymerize forming curved filaments that close into rings that remain stable for some time before opening irreversibly and fully depolymerizing. The distribution of ring lifetimes (T) as a function of length (N), shows that the rate of ring aperture correlates with filament length. If this ring lifetime is expressed as a bond survival time, (Tb ≡ NT), this correlation is abolished, indicating that these rupture events occur randomly and independently at each monomer interface. After rings open irreversibly, depolymerization of the remaining filaments is fast, but can be slowed down and followed using a nonhydrolyzing GTP analogue. The histogram of depolymerization velocities of individual filaments has an asymmetric distribution that can be fit with a computer model that assumes two rupture rates, a slow one similar to the one observed for ring aperture, affecting monomers in the central part of the filaments, and a faster one affecting monomers closer to the open ends. From the quantitative analysis, we conclude that the depolymerization rate is affected both by nucleotide hydrolysis rate and by its exchange along the filament, that all monomer interfaces are equally competent for hydrolysis, although depolymerization is faster at the open ends than in central filament regions, and that all monomer–monomer interactions, regardless of the nucleotide present, can adopt a curved configuration. PMID:22566654

Gastric protein hydrolysis of raw and roasted almonds in the growing pig.

PubMed

Bornhorst, Gail M; Drechsler, Krista C; Montoya, Carlos A; Rutherfurd, Shane M; Moughan, Paul J; Singh, R Paul

2016-11-15

Gastric protein hydrolysis may influence gastric emptying rate and subsequent protein digestibility in the small intestine. This study examined the gastric hydrolysis of dietary protein from raw and roasted almonds in the growing pig as a model for the adult human. The gastric hydrolysis of almond proteins was quantified by performing tricine-sodium dodecyl sulfate-polyacrylamide gel electrophoresis and subsequent image analysis. There was an interaction between digestion time, stomach region, and almond type for gastric protein hydrolysis (p<0.05). Gastric emptying rate of protein was a significant (p<0.05) covariate in the gastric protein hydrolysis. In general, greater gastric protein hydrolysis was observed in raw almonds (compared to roasted almonds), hypothesized to be related to structural changes in almond proteins during roasting. Greater gastric protein hydrolysis was observed in the distal stomach (compared to the proximal stomach), likely related to the lower pH in the distal stomach. Copyright © 2016 Elsevier Ltd. All rights reserved.

Enhanced efficiency of biological excess sludge hydrolysis under anaerobic digestion by additional enzymes.

PubMed

Yang, Qi; Luo, Kun; Li, Xiao-ming; Wang, Dong-bo; Zheng, Wei; Zeng, Guang-ming; Liu, Jing-jin

2010-05-01

In this investigation, the effects of commercial enzyme preparation containing alpha amylase and neutral protease on hydrolysis of excess sludge and the kinetic analysis of hydrolysis process were evaluated. The results indicated that amylase treatment displayed higher hydrolysis efficiency than that of protease. VSS reduction greatly increased to 39.70% for protease and 54.24% for amylase at the enzyme dosage of 6% (w/w), respectively. The hydrolysis rate of sludge improved with temperature increasing from 40 to 50 degrees Celsius, which could be well described by the amended Arrhenius equation. Mixed-enzyme had great impact on sludge solubilisation than single enzyme. The mixture of two enzymes (protease:amylase=1:3) resulted in optimum hydrolysis efficiency, the efficiency of solids hydrolysis increased from 10% (control test) to 68.43% at the temperature of 50 degrees Celsius. Correspondingly, the concentration of reducing sugar and NH(4)(+)-N improved about 377% and 201%, respectively. According to the kinetic analysis of enzymatic hydrolysis process, VSS solubilisation process within prior 4 h followed first-order kinetics. Compared with control test, the hydrolysis rate improved significantly at 50 degrees Celsius when either single enzyme or mixed-enzyme was added. Copyright 2009. Published by Elsevier Ltd.

Formation of ethyl ferulate by rice koji enzyme in sake and mirin mash conditions.

PubMed

Hashizume, Katsumi; Ito, Toshihiko; Ishizuka, Takahiro; Takeda, Naoki

2013-08-01

Formation mechanism of ethyl ferulate (EF) in sake and mirin mash conditions was investigated to understand EF level control in the manufacturing process. Rice koji formed EF from ferulic acid (FA) and ethanol and decomposed EF to FA. This did not occur in sake yeast and chemical esterification was rare. Esterification of FA and hydrolysis of EF by rice koji might be due to feruloyl esterase(s). The rice koji enzyme showed normal Michaelis-Menten kinetics for FA in ethyl esterification and for EF in hydrolysis, but not for ethanol in the esterification reaction. Substrate specificity of the rice koji enzyme for hydroxycinnamic acids suggested that the main enzyme involved might be similar to type A feruloyl esterase. We studied the rice koji enzyme properties, short-term digestion of steamed rice grains with exogenous ethanol and small scale mirin making with pH adjustment. Our results suggested differences in the esterification and hydrolysis properties of the enzyme, in particular, different pH dependencies and different behaviors under high ethanol conditions; these factors might cause the differing EF levels in sake and mirin mashes. Copyright © 2013 The Society for Biotechnology, Japan. Published by Elsevier B.V. All rights reserved.

Phospholipids and products of their hydrolysis as dietary preventive factors for civilization diseases.

PubMed

Parchem, Karol; Bartoszek, Agnieszka

2016-12-31

The results of numerous epidemiological studies indicate that phospholipids play an important role in the prevention of chronic diseases faced by contemporary society. Firstly, these compounds are responsible for the proper functioning of cell membranes, by ensuring liquidity and permeability, which is pivotal for normal activity of membrane proteins, including receptors. These mechanisms are at the core of prevention of cancer, autoimmune or neurological disorders. Secondly, structure and properties of phospholipids cause that they are highly available source of biologically active fatty acids. Thirdly, also products of endogenous hydrolysis of phospholipids exhibit biological activity. These include lysophospholipids formed as a result of disconnecting free fatty acid from glycerophospholipids in the reaction catalyzed by phospholipase A, phosphatidic acid and hydrophilic subunits released by the activity of phospholipase D. The bioactive products of hydrolysis also include ceramides liberated from phosphosphingolipids after removal of a hydrophilic unit catalyzed by sphingomyelinase. Phospholipids are supplied to the human body with food. A high content of phospholipids is characteristic for egg yolk, liver, pork and poultry, as well as some soy products. Particularly beneficial are phospholipids derived from seafood because they are a rich source of essential fatty acids of the n-3 family.

Microwave pretreatment of paramylon enhances the enzymatic production of soluble β-1,3-glucans with immunostimulatory activity.

PubMed

Gissibl, Alexander; Care, Andrew; Parker, Lindsay M; Iqbal, Sameera; Hobba, Graham; Nevalainen, Helena; Sunna, Anwar

2018-09-15

A hydrothermal microwave pretreatment was established to facilitate the enzymatic production of soluble bioactive β-1,3-glucans from the recalcitrant substrate paramylon. The efficacy of this pretreatment was monitored with a newly developed direct Congo Red dye-based assay over a range of temperatures. Microwave pretreatment at 170 °C for 2 min resulted in a significantly enhanced enzymatic hydrolysis of paramylon. The action of endo-β-1,3- and exo- β-1,3-glucanases on the microwave-pretreated paramylon produced soluble β-1,3-glucans with degrees of polymerisation (DP) ranging from 2-59 and 2-7, respectively. In comparison, acid-mediated hydrolysis of untreated paramylon resulted in β-1,3-glucans with a DP range of 2-38. The hydrolysates were assayed on their immunostimulatory effect on murine macrophages by measuring the production of the inflammation-linked marker tumour necrosis factor alpha (TNFα) using immunofluorescence. All of the tested hydrolysis products were shown to induce TNFα production, with the most significant immunostimulatory effect observed with the hydrolysate from the exo-β-1,3-glucanase treatment. Copyright © 2018 Elsevier Ltd. All rights reserved.

Symmetry broken and rebroken during the ATP hydrolysis cycle of the mitochondrial Hsp90 TRAP1

PubMed Central

Elnatan, Daniel; Betegon, Miguel; Liu, Yanxin; Ramelot, Theresa; Kennedy, Michael A; Agard, David A

2017-01-01

Hsp90 is a homodimeric ATP-dependent molecular chaperone that remodels its substrate ‘client’ proteins, facilitating their folding and activating them for biological function. Despite decades of research, the mechanism connecting ATP hydrolysis and chaperone function remains elusive. Particularly puzzling has been the apparent lack of cooperativity in hydrolysis of the ATP in each protomer. A crystal structure of the mitochondrial Hsp90, TRAP1, revealed that the catalytically active state is closed in a highly strained asymmetric conformation. This asymmetry, unobserved in other Hsp90 homologs, is due to buckling of one of the protomers and is most pronounced at the broadly conserved client-binding region. Here, we show that rather than being cooperative or independent, ATP hydrolysis on the two protomers is sequential and deterministic. Moreover, dimer asymmetry sets up differential hydrolysis rates for each protomer, such that the buckled conformation favors ATP hydrolysis. Remarkably, after the first hydrolysis, the dimer undergoes a flip in the asymmetry while remaining in a closed state for the second hydrolysis. From these results, we propose a model where direct coupling of ATP hydrolysis and conformational flipping rearranges client-binding sites, providing a paradigm of how energy from ATP hydrolysis can be used for client remodeling. DOI: http://dx.doi.org/10.7554/eLife.25235.001 PMID:28742020

Catalytic hydrolysis of carbonyl sulphide and carbon disulphide over Fe2O3 cluster: Competitive adsorption and reaction mechanism.

PubMed

Ning, Ping; Song, Xin; Li, Kai; Wang, Chi; Tang, Lihong; Sun, Xin

2017-10-31

The competitive adsorption and reaction mechanism for the catalytic hydrolysis of carbonyl sulphide (COS) and carbon disulphide (CS 2 ) over Fe 2 O 3 cluster was investigated. Compared with experimental results, the theoretical study was used to further investigate the competitive adsorption and effect of H 2 S in the hydrolysis reaction of COS and CS 2 . Experimental results showed that Fe 2 O 3 cluster enhanced the catalytic hydrolysis effect. Meanwhile, H 2 S was not conducive to the hydrolysis of COS and CS 2 . Theoretical calculations indicated that the order of competitive adsorption on Fe 2 O 3 is as follows: H 2 O (strong) >CS 2 (medium) >COS (weak). In the hydrolysis process, the C=S bond cleavage occurs easier than C=O bond cleavage. The hydrolysis reaction is initiated via the migration of an H-atom, which triggers C=S bond cleavage and S-H bond formation. Additionally, we find the first step of CS 2 hydrolysis to be rate limiting. The presence of H 2 S increases the reaction energy barrier, which is not favourable for COS hydrolysis. Fe 2 O 3 can greatly decrease the maximum energy barrier, which decreases the minimum energy required for hydrolysis, making it relatively facile to occur. In general, the theoretical results were consistent with experimental results, which proved that the theoretical study was reliable.

Comparing the performance of Miscanthus x giganteus and wheat straw biomass in sulfuric acid based pretreatment.

PubMed

Kärcher, M A; Iqbal, Y; Lewandowski, I; Senn, T

2015-03-01

The objective of this study was to assess and compare the suitability of Miscanthus x giganteus and wheat straw biomass in dilute acid catalyzed pretreatment. Miscanthus and wheat straw were treated in a dilute sulfuric acid/steam explosion pretreatment. As a result of combining dilute sulfuric acid- and steam explosion pretreatment the hemicellulose hydrolysis yields (96% in wheat straw and 90% in miscanthus) in both substrates were higher than reported in literature. The combined severity factor (=CSF) for optimal hemicellulose hydrolysis was 1.9 and 1.5 in for miscanthus and wheat straw respectively. Because of the higher CSF value more furfural, furfuryl alcohol, 5-hydroxymethylfurfural and acetic acid was formed in miscanthus than in wheat straw pretreatment. Copyright © 2015 Elsevier Ltd. All rights reserved.

Evaluation of preservation methods for improving biogas production and enzymatic conversion yields of annual crops

PubMed Central

2011-01-01

Background The use of energy crops and agricultural residues is expected to increase to fulfil the legislative demands of bio-based components in transport fuels. Ensiling methods, adapted from the feed sector, are suitable storage methods to preserve fresh crops throughout the year for, for example, biogas production. Various preservation methods, namely ensiling with and without acid addition for whole crop maize, fibre hemp and faba bean were investigated. For the drier fibre hemp, alkaline urea treatment was studied as well. These treatments were also explored as mild pretreatment methods to improve the disassembly and hydrolysis of these lignocellulosic substrates. Results The investigated storage treatments increased the availability of the substrates for biogas production from hemp and in most cases from whole maize but not from faba bean. Ensiling of hemp, without or with addition of formic acid, increased methane production by more than 50% compared to fresh hemp. Ensiling resulted in substantially increased methane yields also from maize, and the use of formic acid in ensiling of maize further enhanced methane yields by 16%, as compared with fresh maize. Ensiled faba bean, in contrast, yielded somewhat less methane than the fresh material. Acidic additives preserved and even increased the amount of the valuable water-soluble carbohydrates during storage, which affected most significantly the enzymatic hydrolysis yield of maize. However, preservation without additives decreased the enzymatic hydrolysis yield especially in maize, due to its high content of soluble sugars that were already converted to acids during storage. Urea-based preservation significantly increased the enzymatic hydrolysability of hemp. Hemp, preserved with urea, produced the highest carbohydrate increase of 46% in enzymatic hydrolysis as compared to the fresh material. Alkaline pretreatment conditions of hemp improved also the methane yields. Conclusions The results of the present work show that ensiling and alkaline preservation of fresh crop materials are useful pretreatment methods for methane production. Improvements in enzymatic hydrolysis were also promising. While all three crops still require a more powerful pretreatment to release the maximum amount of carbohydrates, anaerobic preservation is clearly a suitable storage and pretreatment method prior to production of platform sugars from fresh crops. PMID:21771298

Comparison of rates of ureolysis between Sporosarcina pasteurii and an indigenous groundwater community under conditions required to precipitate large volumes of calcite

NASA Astrophysics Data System (ADS)

Tobler, Dominique J.; Cuthbert, Mark O.; Greswell, Richard B.; Riley, Michael S.; Renshaw, Joanna C.; Handley-Sidhu, Stephanie; Phoenix, Vernon R.

2011-06-01

Ureolysis-driven calcite precipitation has potential to seal porosity and fracture networks in rocks thus preventing groundwater flow and contaminant transport. In this study urea hydrolysis and calcite precipitation rates for the model bacterium Sporosarcina pasteurii were compared with those of indigenous groundwater communities under conditions required to precipitate large volumes of calcite (up to 50 g L -1). We conducted microcosm experiments in oxic artificial and anoxic natural groundwaters (collected from the Permo-Triassic sandstone aquifer at Birmingham, UK) that were inoculated with aerobically grown S. pasteurii. The rate constants for urea hydrolysis, kurea, ranged between 0.06 and 3.29 d -1 and were only affected by inoculum density. Higher Ca 2+ concentration (50-500 mM Ca 2+) as well as differences in fO 2 did not inhibit the ureolytic activity of S. pasteurii and did not significantly impact kurea. These results demonstrate that S. pasteurii has potential to improve calcite precipitation in both oxic and anoxic groundwaters, especially if indigenous communities lack ureolytic activity. Urea hydrolysis by indigenous groundwater communities was investigated in anoxic, natural groundwaters amended with urea and CaCl 2. A notable increase in ureolysis rates was measured only when these communities were stimulated with dilute nutrients (with best results from blackstrap molasses). Furthermore, there was a considerable lag time (12-20 days) before ureolysis and calcite precipitation began. Calculated ureolysis rate constants, kurea, ranged between 0.03 and 0.05 d -1 and were similar to kurea values produced by S. pasteurii at low inoculum densities. Overall, this comparative study revealed that the growth of ureolytic microorganisms present within groundwaters can easily be stimulated to enhance rates of urea hydrolysis in the subsurface, and thus can be used to induce calcite precipitation in these environments. The time required for urea hydrolysis to begin is almost instantaneous if an inoculum of S. pasteurii is included, while it may take several weeks for ureolytic groundwater communities to grow and become ureolytically active.

Process simulation and economic assessment of hydrothermal pretreatment and enzymatic hydrolysis of multi-feedstock lignocellulose - Separate vs combined processing.

PubMed

Ashraf, Muhammad Tahir; Schmidt, Jens Ejbye

2018-02-01

Biorefinery based on multi-feedstock lignocellulose can be viable where a sustainable supply of a single substrate is limited, for example in arid regions. Processing of mixed feedstocks has been studied in lab scale, however, its economics are less studied. In this study, an economic comparison was made between separate and combined (mixed) processing approaches for multi-feedstock lignocellulose for the production of monomeric sugars. This modular approach of focusing on sugar platform makes the results applicable for many applications using the sugars as feedstock. Feedstock considered in this study were the green and woody lignocellulose residues: Bermuda grass, Jasmine hedges, and date palm fronds. Results showed that, at an identical total feed rate, combined processing was more advantageous as compared to separate processing. A further sensitivity analysis on mixed combined processing showed that the cellulase enzyme price and feed price are the two major factors affecting the production cost. Copyright © 2017 Elsevier Ltd. All rights reserved.

A Novel High-Affinity Sucrose Transporter Is Required for Virulence of the Plant Pathogen Ustilago maydis

PubMed Central

Goos, Sarah; Kämper, Jörg; Sauer, Norbert

2010-01-01

Plant pathogenic fungi cause massive yield losses and affect both quality and safety of food and feed produced from infected plants. The main objective of plant pathogenic fungi is to get access to the organic carbon sources of their carbon-autotrophic hosts. However, the chemical nature of the carbon source(s) and the mode of uptake are largely unknown. Here, we present a novel, plasma membrane-localized sucrose transporter (Srt1) from the corn smut fungus Ustilago maydis and its characterization as a fungal virulence factor. Srt1 has an unusually high substrate affinity, is absolutely sucrose specific, and allows the direct utilization of sucrose at the plant/fungal interface without extracellular hydrolysis and, thus, without the production of extracellular monosaccharides known to elicit plant immune responses. srt1 is expressed exclusively during infection, and its deletion strongly reduces fungal virulence. This emphasizes the central role of this protein both for efficient carbon supply and for avoidance of apoplastic signals potentially recognized by the host. PMID:20161717

Characterization of amylose nanoparticles prepared via nanoprecipitation: Influence of chain length distribution.

PubMed

Chang, Yanjiao; Yang, Jingde; Ren, Lili; Zhou, Jiang

2018-08-15

The influence of chain length distribution of amylose on size and structure of the amylose nanoparticles (ANPs) prepared through nanoprecipitation was investigated. Amylose with different chain length distributions was obtained by β-amylase treating amylose paste for different times and measured by size exclusion chromatography (SEC) and fluorophore-assisted carbohydrate electrophoresis (FACE). ANPs prepared via precipitation were characterized by using dynamic light scattering (DLS), scanning electron microscopy (SEM) and X-ray diffraction (XRD). Results showed that the β-amylase treatments led to decrease in chain length of amylose, and it was the most important factor affecting size of ANPs. When hydrolysis degree of amylose was 52.8%, mean size of ANPs decreased from 206.4 nm to 102.7 nm. All the ANPs displayed a V-type crystalline structure and the effect of amylose chain length on crystallinity of the precipitated ANPs was negligible in the investigated range. Copyright © 2018 Elsevier Ltd. All rights reserved.

Chemopreventive potential of the tannase-mediated biotransformation of green tea.

PubMed

Macedo, J A; Ferreira, L R; Camara, L E; Santos, J C; Gambero, A; Macedo, G A; Ribeiro, M L

2012-07-15

Green tea (Camellia sinensis) is one of the most widely consumed beverages in the world. The cancer chemopreventive qualities of green tea have been well documented. Epigallocatechin gallate (EGCG) is often described as the most potently chemopreventive green tea catechin; however, the low bioavailability of EGCG is a limiting factor for its biological effect. Thus, the aim of this work was to test the chemopreventive potential of green tea extract and EGCG after tannase-mediated hydrolysis. The results showed that the biotransformed compounds retained most of the beneficial properties of the original compounds, and some beneficial properties were improved in the biotransformed compounds. Biotransformation of EGCG decreased its toxicity without affecting its antiproliferative effects. Furthermore, human cells gene expression profiling showed that the biotransformed compounds modulated the expression of several genes related to carcinogenesis. These results demonstrate the benefits of the biotechnological modification of natural food molecules, allowing the improvement of the nutraceutical potential of a beverage as green tea. Copyright © 2012 Elsevier Ltd. All rights reserved.

Optimization of antioxidant activity by response surface methodology in hydrolysates of jellyfish (Rhopilema esculentum) umbrella collagen.

PubMed

Zhuang, Yong-liang; Zhao, Xue; Li, Ba-fang

2009-08-01

To optimize the hydrolysis conditions to prepare hydrolysates of jellyfish umbrella collagen with the highest hydroxyl radical scavenging activity, collagen extracted from jellyfish umbrella was hydrolyzed with trypsin, and response surface methodology (RSM) was applied. The optimum conditions obtained from experiments were pH 7.75, temperature (T) 48.77 degrees C, and enzyme-to-substrate ratio ([E]/[S]) 3.50%. The analysis of variance in RSM showed that pH and [E]/[S] were important factors that significantly affected the process (P<0.05 and P<0.01, respectively). The hydrolysates of jellyfish umbrella collagen were fractionated by high performance liquid chromatography (HPLC), and three fractions (HF-1>3000 Da, 1000 Da

Optimization of antioxidant activity by response surface methodology in hydrolysates of jellyfish (Rhopilema esculentum) umbrella collagen*

PubMed Central

Zhuang, Yong-liang; Zhao, Xue; Li, Ba-fang

2009-01-01

To optimize the hydrolysis conditions to prepare hydrolysates of jellyfish umbrella collagen with the highest hydroxyl radical scavenging activity, collagen extracted from jellyfish umbrella was hydrolyzed with trypsin, and response surface methodology (RSM) was applied. The optimum conditions obtained from experiments were pH 7.75, temperature (T) 48.77 °C, and enzyme-to-substrate ratio ([E]/[S]) 3.50%. The analysis of variance in RSM showed that pH and [E]/[S] were important factors that significantly affected the process (P<0.05 and P<0.01, respectively). The hydrolysates of jellyfish umbrella collagen were fractionated by high performance liquid chromatography (HPLC), and three fractions (HF-1>3000 Da, 1000 Da

Effects of operational conditions on sludge degradation and organic acids formation in low-critical wet air oxidation.

PubMed

Chung, Jinwook; Lee, Mikyung; Ahn, Jaehwan; Bae, Wookeun; Lee, Yong-Woo; Shim, Hojae

2009-02-15

Wet air oxidation processes are to treat highly concentrated organic compounds including refractory materials, sludge, and night soil, and usually operated at supercritical water conditions of high temperature and pressure. In this study, the effects of operational conditions including temperature, pressure, and oxidant dose on sludge degradation and conversion into subsequent intermediates such as organic acids were investigated at low critical wet oxidation conditions. The reaction time and temperature in the wet air oxidation process was shown an important factor affecting the liquefaction of volatile solids, with more significant effect on the thermal hydrolysis reaction rather than the oxidation reaction. The degradation efficiency of sludge and the formation of organic acids were improved with longer reaction time and higher reaction temperature. For the sludge reduction and the organic acids formation under the wet air oxidation, the optimal conditions for reaction temperature, time, pressure, and oxidant dose were shown approximately 240 degrees C, 30min, 60atm, and 2.0L/min, respectively.

Bermuda grass as feedstock for biofuel production: a review.

PubMed

Xu, Jiele; Wang, Ziyu; Cheng, Jay J

2011-09-01

Bermuda grass is a promising feedstock for the production of fuel ethanol in the Southern United States. This paper presents a review of the significant amount of research on the conversion of Bermuda grass to ethanol and a brief discussion on the factors affecting the biomass production in the field. The biggest challenge of biomass conversion comes from the recalcitrance of lignocellulose. A variety of chemical, physico-chemical, and biological pretreatment methods have been investigated to improve the digestibility of Bermuda grass with encouraging results reported. The subsequent enzymatic hydrolysis and fermentation steps have also been extensively studied and effectively optimized. It is expected that the development of genetic engineering technologies for the grass and fermenting organisms has the potential to greatly improve the economic viability of Bermuda grass-based fuel ethanol production systems. Other energy applications of Bermuda grass include anaerobic digestion for biogas generation and pyrolysis for syngas production. Copyright © 2011 Elsevier Ltd. All rights reserved.

Influence of sugarcane bagasse variability on sugar recovery for cellulosic ethanol production.

PubMed

Andrade, Liliane Pires; Crespim, Elaine; de Oliveira, Nilton; de Campos, Rafael Carinha; Teodoro, Juliana Conceição; Galvão, Célia Maria Araújo; Maciel Filho, Rubens

2017-10-01

In the context of cellulosic ethanol production, special attention must be given to the raw material, as it affects final product yield. As observed for sugarcane, bagasse variations may derive from several elements, for instance edaphoclimatic factors, seasonality, maturation stage and harvesting techniques. Therefore, in the present work, to investigate the impact of raw material characteristics on process performance, sugarcane bagasse from four harvests from October/2010 to October/2011 was pretreated by steam explosion and had its soluble and insoluble solids contents measured, following enzymatic hydrolysis to assess glucan conversion. As confirmed by ANOVA, glucose concentration was related to the solids content in the reactor, whereas glucan conversion was related to the enzymatic load. Variations in raw material composition were indeed observed to significantly interfere in the final sugar recovery, probably due to the increase in the impurities observed as a result of the type of harvest performed in 2011. Copyright © 2017 Elsevier Ltd. All rights reserved.

Lignin degradation in corn stalk by combined method of H2O2 hydrolysis and Aspergillus oryzae CGMCC5992 liquid-state fermentation.

PubMed

Zhang, Zhicai; Xia, Lili; Wang, Feng; Lv, Peng; Zhu, Maxiaoqi; Li, Jinhua; Chen, Keping

2015-01-01

Lignin peroxidase (LiP) is the primary enzyme responsible for lignin degradation. In our previous work, in order to shorten the pretreatment time and increase the lignin degradation, we have pretreated the corn stalk (CS) using a combination of Aspergillus oryzae CGMCC 5992 solid-state fermentation and H2O2 treatment. In the present study, one-factor-at-a-time design and response surface design were applied to optimize the nutritional constituents for LiP production in liquid-state fermentation by A. oryzae CGMCC 5992 and the conditions for CS degradation by A. oryzae CGMCC 5992. The optimal medium included CS of 30 g/L, glucose of 4.6 g/L, sodium nitrate of 1.2 g/L, corn steep liquor of 1 g/L, yeast extract of 1.2 g/L, and vitamin B1 of 0.15 g/L. Under these optimal conditions, the LiP production reached its maximum of 652.34 U/L. The optimal condition for CS degradation included CS of 20 g, A. oryzae CGMCC 5992 broth of 50 mL, 1.5 % H2O2 solution of 80 mL, H2O2 flow rate of 0.4 mL/min, water volume of 240 mL (water/material ratio of 12:1), hydrolysis temperature of 39 °C, and hydrolysis time of 8 h. Before hydrolysis, CS and water were pretreated at 113 °C for 11 min. Under these optimal conditions, the sugar yield reached its maximum of 46.28 %. Our newly developed method had great advantages in pretreatment of CS due to its quickness, convenience, safety, no special equipment and high sugar yield.Graphical abstractThe schematic diagram of corn straw hydrolysis.

Stimulation and inhibition of enzymatic hydrolysis by organosolv lignins as determined by zeta potential and hydrophobicity

Treesearch

Yang Huang; Shaolong Sun; Chen Huang; Qiang Yong; Thomas Elder; Maobing Tu

2017-01-01

Background: Lignin typically inhibits enzymatic hydrolysis of cellulosic biomass, but certain organosolv lignins or lignosulfonates enhance enzymatic hydrolysis. The hydrophobic and electrostatic interactions between lignin and cellulases play critical roles in the enzymatic hydrolysis process. However, how to incorporate these two...

Laboratory-Scale Demonstration Using Dilute Ammonia Gas-Induced Alkaline Hydrolysis of Soil Contaminants (Chlorinated Propanes and Explosives)

DTIC Science & Technology

2016-06-01

Hydrolysis of Soil Contaminants (Chlorinated Propanes and Explosives) En vi ro nm en ta l L ab or at or y Victor F. Medina, Scott A. Waisner, Charles...Using Dilute Ammonia Gas-Induced Alkaline Hydrolysis of Soil Contaminants (Chlorinated Propanes and Explosives) Victor F. Medina, Scott A. Waisner...hydrolysis. This project explored the use of ammonia gas to raise soil pH in order to stimulate alkaline hydrolysis. When ammonia gas dissolves in water

Visualizing cellulase activity.

PubMed

Bubner, Patricia; Plank, Harald; Nidetzky, Bernd

2013-06-01

Commercial exploitation of lignocellulose for biotechnological production of fuels and commodity chemicals requires efficient-usually enzymatic-saccharification of the highly recalcitrant insoluble substrate. A key characteristic of cellulose conversion is that the actual hydrolysis of the polysaccharide chains is intrinsically entangled with physical disruption of substrate morphology and structure. This "substrate deconstruction" by cellulase activity is a slow, yet markedly dynamic process that occurs at different length scales from and above the nanometer range. Little is currently known about the role of progressive substrate deconstruction on hydrolysis efficiency. Application of advanced visualization techniques to the characterization of enzymatic degradation of different celluloses has provided important new insights, at the requisite nano-scale resolution and down to the level of single enzyme molecules, into cellulase activity on the cellulose surface. Using true in situ imaging, dynamic features of enzyme action and substrate deconstruction were portrayed at different morphological levels of the cellulose, thus providing new suggestions and interpretations of rate-determining factors. Here, we review the milestones achieved through visualization, the methods which significantly promoted the field, compare suitable (model) substrates, and identify limiting factors, challenges and future tasks. Copyright © 2013 Wiley Periodicals, Inc.

Fuzzy logic feedback control for fed-batch enzymatic hydrolysis of lignocellulosic biomass.

PubMed

Tai, Chao; Voltan, Diego S; Keshwani, Deepak R; Meyer, George E; Kuhar, Pankaj S

2016-06-01

A fuzzy logic feedback control system was developed for process monitoring and feeding control in fed-batch enzymatic hydrolysis of a lignocellulosic biomass, dilute acid-pretreated corn stover. Digested glucose from hydrolysis reaction was assigned as input while doser feeding time and speed of pretreated biomass were responses from fuzzy logic control system. Membership functions for these three variables and rule-base were created based on batch hydrolysis data. The system response was first tested in LabVIEW environment then the performance was evaluated through real-time hydrolysis reaction. The feeding operations were determined timely by fuzzy logic control system and efficient responses were shown to plateau phases during hydrolysis. Feeding of proper amount of cellulose and maintaining solids content was well balanced. Fuzzy logic proved to be a robust and effective online feeding control tool for fed-batch enzymatic hydrolysis.

The Hydrolysis of Carbonyl Sulfide at Low Temperature: A Review

PubMed Central

Zhao, Shunzheng; Yi, Honghong; Tang, Xiaolong; Jiang, Shanxue; Gao, Fengyu; Zhang, Bowen; Zuo, Yanran; Wang, Zhixiang

2013-01-01

Catalytic hydrolysis technology of carbonyl sulfide (COS) at low temperature was reviewed, including the development of catalysts, reaction kinetics, and reaction mechanism of COS hydrolysis. It was indicated that the catalysts are mainly involved metal oxide and activated carbon. The active ingredients which can load on COS hydrolysis catalyst include alkali metal, alkaline earth metal, transition metal oxides, rare earth metal oxides, mixed metal oxides, and nanometal oxides. The catalytic hydrolysis of COS is a first-order reaction with respect to carbonyl sulfide, while the reaction order of water changes as the reaction conditions change. The controlling steps are also different because the reaction conditions such as concentration of carbonyl sulfide, reaction temperature, water-air ratio, and reaction atmosphere are different. The hydrolysis of carbonyl sulfide is base-catalyzed reaction, and the force of the base site has an important effect on the hydrolysis of carbonyl sulfide. PMID:23956697

Quantitative characterization of nonstructural carbohydrates of mezcal Agave (Agave salmiana Otto ex Salm-Dick).

PubMed

Michel-Cuello, Christian; Juárez-Flores, Bertha Irene; Aguirre-Rivera, Juan Rogelio; Pinos-Rodríguez, Juan Manuel

2008-07-23

Fructans are the reserve carbohydrates in Agave spp. plants. In mezcal factories, fructans undergoes thermal hydrolysis to release fructose and glucose, which are the basis to produce this spirit. Carbohydrate content determines the yield of the final product, which depends on plant organ, ripeness stage, and thermal hydrolysis. Thus, a qualitative and quantitative characterization of nonstructural carbohydrates was conducted in raw and hydrolyzed juices extracted from Agave salmiana stems and leaves under three ripeness stages. By high-performance liquid chromatography (HPLC), fructose, glucose, sucrose, xylose, and maltose were identified in agave juice. Only the plant fraction with hydrolysis interaction was found to be significant in the glucose concentration plant. Interactions of the fraction with hydrolysis and ripeness with hydrolysis were statistically significant in fructose concentration. Fructose concentration rose considerably with hydrolysis, but only in juice extracted from ripe agave stems (early mature and castrated). This increase was statistically significant only with acid hydrolysis.

Study of Enzymatic Hydrolysis of Fructans from Agave salmiana Characterization and Kinetic Assessment

PubMed Central

Michel-Cuello, Christian; Ortiz-Cerda, Imelda; Moreno-Vilet, Lorena; Grajales-Lagunes, Alicia; Moscosa-Santillán, Mario; Bonnin, Johanne; González-Chávez, Marco Martín; Ruiz-Cabrera, Miguel

2012-01-01

Fructans were extracted from Agave salmiana juice, characterized and subjected to hydrolysis process using a commercial inulinase preparation acting freely. To compare the performance of the enzymatic preparation, a batch of experiments were also conducted with chicory inulin (reference). Hydrolysis was performed for 6 h at two temperatures (50, 60°C) and two substrate concentrations (40, 60 mg/ml). Hydrolysis process was monitored by measuring the sugars released and residual substrate by HPLC. A mathematical model which describes the kinetics of substrate degradation as well as fructose production was proposed to analyze the hydrolysis assessment. It was found that kinetics were significantly influenced by temperature, substrate concentration, and type of substrate (P < 0.01). The extent of substrate hydrolysis varied from 82 to 99%. Hydrolysis product was mainly constituted of fructose, obtaining from 77 to 96.4% of total reducing sugars. PMID:22629216

Enzymatic hydrolysis of biomimetic bacterial cellulose-hemicellulose composites.

PubMed

Penttilä, Paavo A; Imai, Tomoya; Hemming, Jarl; Willför, Stefan; Sugiyama, Junji

2018-06-15

The production of biofuels and other chemicals from lignocellulosic biomass is limited by the inefficiency of enzymatic hydrolysis. Here a biomimetic composite material consisting of bacterial cellulose and wood-based hemicelluloses was used to study the effects of hemicelluloses on the enzymatic hydrolysis with a commercial cellulase mixture. Bacterial cellulose synthesized in the presence of hemicelluloses, especially xylan, was found to be more susceptible to enzymatic hydrolysis than hemicellulose-free bacterial cellulose. The reason for the easier hydrolysis could be related to the nanoscale structure of the substrate, particularly the packing of cellulose microfibrils into ribbons or bundles. In addition, small-angle X-ray scattering was used to show that the average nanoscale morphology of bacterial cellulose remained unchanged during the enzymatic hydrolysis. The reported easier enzymatic hydrolysis of bacterial cellulose produced in the presence of wood-based xylan offers new insights to overcome biomass recalcitrance through genetic engineering. Copyright © 2018 Elsevier Ltd. All rights reserved.

Evaluation of physical structural features on influencing enzymatic hydrolysis efficiency of micronized wood

Treesearch

Jinxue Jiang; Jinwu Wang; Xiao Zhang; Michael Wolcott

2016-01-01

Enzymatic hydrolysis of lignocellulosic biomass is highly dependent on the changes in structural features after pretreatment. Mechanical milling pretreatment is an effective approach to alter the physical structure of biomass and thus improve enzymatic hydrolysis. This study examined the influence of structural characteristics on the enzymatic hydrolysis of micronized...

Short-time ultrasonication treatment in enzymatic hydrolysis of biomass

Treesearch

Zengqian Shi; Zhiyong Cai; Siqun Wang; Qixin Zhong; Joseph J. Bozell

2013-01-01

To improve the conversion of enzymatic hydrolysis of biomass in an energy-efficient manner, two shorttime ultrasonication strategies were applied on six types of biomass with different structures and components. The strategies include pre-sonication before the hydrolysis and intermittent sonication during the ongoing hydrolysis. The microstructures of each type of...

Effect of hydrolysis on identifying prenatal cannabis exposure

PubMed Central

Gray, Teresa R.; Barnes, Allan J.

2011-01-01

Identification of prenatal cannabis exposure is important due to potential cognitive and behavioral consequences. A two-dimensional gas chromatography–mass spectrometry method for cannabinol, Δ9-tetrahydrocannabinol (THC), 11-hydroxy-THC (11-OH-THC), 8β,11-dihydroxy-THC, and 11-nor-9-carboxy-THC (THCCOOH) quantification in human meconium was developed and validated. Alkaline, enzymatic, and enzyme–alkaline tandem hydrolysis conditions were optimized with THC- and THCCOOH-glucuronide reference standards. Limits of quantification ranged from 10 to 15 ng/g, and calibration curves were linear to 500 ng/g. Bias and intra-day and inter-day imprecision were <12.3%. Hydrolysis efficiencies were analyte-dependent; THC-glucuronide was effectively cleaved by enzyme, but not base. Conversely, THCCOOH-glucuronide was most sensitive to alkaline hydrolysis. Enzyme–alkaline tandem hydrolysis maximized efficiency for both glucuronides. Identification of cannabinoid-positive meconium specimens nearly doubled following alkaline and enzyme–alkaline hydrolysis. Although no 11-OH-THC glucuronide standard is available, enzymatic hydrolysis improved 11-OH-THC detection in authentic specimens. Maximal identification of cannabis-exposed neonates and the widest range of cannabis biomarkers are achieved with enzyme–alkaline tandem hydrolysis. PMID:20517601

Hydrolysis of Extracellular ATP by Ectonucleoside Triphosphate Diphosphohydrolase (ENTPD) Establishes the Set Point for Fibrotic Activity of Cardiac Fibroblasts*

PubMed Central

Lu, David; Insel, Paul A.

2013-01-01

The establishment of set points for cellular activities is essential in regulating homeostasis. Here, we demonstrate key determinants of the fibrogenic set point of cardiac fibroblasts (CFs) by focusing on the pro-fibrotic activity of ATP, which is released by CFs. We tested the hypothesis that the hydrolysis of extracellular ATP by ectonucleoside triphosphate diphosphohydrolases (ENTPDs) regulates pro-fibrotic nucleotide signaling. We detected two ENTPD isoforms, ENTPD-1 and -2, in adult rat ventricular CFs. Partial knockdown of ENTPD-1 and -2 with siRNA increased basal extracellular ATP concentration and enhanced the pro-fibrotic effect of ATP stimulation. Sodium polyoxotungstate-1, an ENTPD inhibitor, not only enhanced the pro-fibrotic effects of exogenously added ATP but also increased basal expression of α-smooth muscle actin, plasminogen activator inhibitor-1 and transforming growth factor (TGF)-β, collagen synthesis, and gel contraction. Furthermore, we found that adenosine, a product of ATP hydrolysis by ENTPD, acts via A2B receptors to counterbalance the pro-fibrotic response to ATP. Removal of extracellular adenosine or inhibition of A2B receptors enhanced pro-fibrotic ATP signaling. Together, these results demonstrate the contribution of basally released ATP in establishing the set point for fibrotic activity in adult rat CFs and identify a key role for the modulation of this activity by hydrolysis of released ATP by ENTPDs. These findings also imply that cellular homeostasis and fibrotic response involve the integration of signaling that is pro-fibrotic by ATP and anti-fibrotic by adenosine and that is regulated by ENTPDs. PMID:23677997

Human Paraoxonase1 Hydrolysis of Nanomolar Chlorpyrifos-oxon Concentrations is Unaffected by Phenotype or Q192R Genotype

PubMed Central

Coombes, R. Hunter; Meek, Edward C.; Dail, Mary Beth; Chambers, Howard W.; Chambers, Janice E.

2016-01-01

The organophosphorus insecticide chlorpyrifos has been widely used. Its active metabolite chlorpyrifos-oxon (CPO) is a potent anticholinesterase and is detoxified by paraoxonase-1 (PON1). PON1 activity is influenced by numerous factors including a Q192R polymorphism. Using forty human blood samples bearing homozygous genotypes and either high or low activity phenotypes (as determined by high concentration assays of paraoxon and diazoxon hydrolysis) the serum PON1 hydrolysis of high (320 μM) and low (178 nM) CPO concentrations was assessed using direct or indirect spectrophotometric methods, respectively. PON1 activity at high CPO concentration reflected the phenotype and genotype differences; subjects with the high activity phenotype and homozygous for the PON1R192 alloform hydrolyzed significantly more CPO than subjects with the low activity phenotype and/or PON1Q192 alloform (High RR=11023±722, Low RR=9467±798, High QQ=8809±672, Low QQ=6030±1015 μmoles CPO hydrolyzed/min/L serum). However, PON1 hydrolysis of CPO at the lower, more environmentally relevant concentration showed no significant differences between the PON1192 genotypes and/or between high and low activity phenotypes (High RR=231±27, Low RR=219±52, High QQ=193±59, Low QQ=185±43 nmoles CPO/min/L serum). Low CPO concentrations were probably not saturating, so PON1 did not display maximal velocity and the PON1 genotype/phenotype might not influence the extent of metabolism at environmental exposures. PMID:25093614

Hydrolysis of membrane phospholipids by phospholipases of rat liver lysosomes

PubMed Central

Richards, Donald E.; Irvine, Robin F.; Dawson, Rex M. C.

1979-01-01

(1) The hydrolysis of 32P- or myo-[2-3H]inositol-labelled rat liver microsomal phospholipids by rat liver lysosomal enzymes has been studied. (2) The relative rates of hydrolysis of phospholipids at pH4.5 are: sphingomyelin>phosphatidylethanolamine>phosphatidylcholine> phosphatidylinositol. (3) The predominant products of phosphatidylcholine and phosphatidylethanolamine hydrolysis are their corresponding lyso-compounds, indicating a slow rate of total deacylation. (4) Ca2+ inhibits the hydrolysis of all phospholipids, though only appreciably at high (>5mm) concentration. The hydrolysis of sphingomyelin is considerably less sensitive to Ca2+ than that of glycerophospholipids. (5) Analysis of the water-soluble products of phosphatidylinositol hydrolysis (by using myo-[3H]inositol-labelled microsomal fraction as a substrate) produced evidence that more than 95% of the product is phosphoinositol, which was derived by direct cleavage from phosphatidylinositol, rather than by hydrolysis of glycerophosphoinositol. (6) This production of phosphoinositol, allied with negligible lysophosphatidylinositol formation and a detectable accumulation of diacylglycerol, indicates that lysosomes hydrolyse membrane phosphatidylinositol almost exclusively in a phospholipase C-like manner. (7) Comparisons are drawn between the hydrolysis by lysosomal enzymes of membrane substrates and that of pure phospholipid substrates, and also the possible role of phosphatidylinositol-specific lysosomal phospholipase C in cellular phosphatidylinositol catabolism is discussed. PMID:508301

Compounds inhibiting the bioconversion of hydrothermally pretreated lignocellulose.

PubMed

Ko, Ja Kyong; Um, Youngsoon; Park, Yong-Cheol; Seo, Jin-Ho; Kim, Kyoung Heon

2015-05-01

Hydrothermal pretreatment using liquid hot water, steam explosion, or dilute acids enhances the enzymatic digestibility of cellulose by altering the chemical and/or physical structures of lignocellulosic biomass. However, compounds that inhibit both enzymes and microbial activity, including lignin-derived phenolics, soluble sugars, furan aldehydes, and weak acids, are also generated during pretreatment. Insoluble lignin, which predominantly remains within the pretreated solids, also acts as a significant inhibitor of cellulases during hydrolysis of cellulose. Exposed lignin, which is modified to be more recalcitrant to enzymes during pretreatment, adsorbs cellulase nonproductively and reduces the availability of active cellulase for hydrolysis of cellulose. Similarly, lignin-derived phenolics inhibit or deactivate cellulase and β-glucosidase via irreversible binding or precipitation. Meanwhile, the performance of fermenting microorganisms is negatively affected by phenolics, sugar degradation products, and weak acids. This review describes the current knowledge regarding the contributions of inhibitors present in whole pretreatment slurries to the enzymatic hydrolysis of cellulose and fermentation. Furthermore, we discuss various biological strategies to mitigate the effects of these inhibitors on enzymatic and microbial activity to improve the lignocellulose-to-biofuel process robustness. While the inhibitory effect of lignin on enzymes can be relieved through the use of lignin blockers and by genetically engineering the structure of lignin or of cellulase itself, soluble inhibitors, including phenolics, furan aldehydes, and weak acids, can be detoxified by microorganisms or laccase.

Microwave irradiation decreases ATP, increases free [Mg2+], and alters in vivo intracellular reactions in rat brain

PubMed Central

Srivastava, Shireesh; Kashiwaya, Yoshihiro; Chen, Xuesong; Geiger, Jonathan D.; Pawlosky, Robert; Veech, Richard L.

2012-01-01

Rapid inactivation of metabolism is essential for accurately determining the concentrations of metabolic intermediates in the in vivo state. We compared a broad spectrum of energetic intermediate metabolites and neurotransmitters in brains obtained by microwave irradiation to those obtained by freeze blowing, the most rapid method of extracting and freezing rat brain. The concentrations of many intermediates, cytosolic free NAD(P)+/NAD(P)H ratios, as well as neurotransmitters were not affected by the microwave procedure. However, the brain concentrations of ATP were about 30% lower, whereas those of ADP, AMP, and GDP were higher in the microwave-irradiated compared with the freeze-blown brains. In addition, the hydrolysis of approximately 1 μmol/g of ATP, a major in vivo Mg2+-binding site, was related to approximately five-fold increase in free [Mg2+] (0.53 ± 0.07 mM in freeze blown vs. 2.91 mM ± 0.48 mM in microwaved brains), as determined from the ratio [citrate]/[isocitrate]. Consequently, many intracellular properties, such as the phosphorylation potential and the ΔG’ of ATP hydrolysis were significantly altered in microwaved tissue. The determinations of some glycolytic and TCA cycle metabolites, the phosphorylation potential, and the ΔG’ of ATP hydrolysis do not represent the in vivo state when using microwave-fixed brain tissue. PMID:23013291

Rapid enzymatic hydrolysis using a novel recombinant β-glucuronidase in benzodiazepine urinalysis.

PubMed

Morris, Ayodele A; Chester, Scot A; Strickland, Erin C; McIntire, Gregory L

2014-10-01

Only trace amounts of parent benzodiazepines are present in urine following extensive metabolism and conjugation. Thus, hydrolysis of glucuronides is necessary for improved detection. Enzyme hydrolysis is preferred to retain identification specificity, but can be costly and time-consuming. The assessment of a novel recombinant β-glucuronidase for rapid hydrolysis in benzodiazepine urinalysis is presented. Glucuronide controls for oxazepam, lorazepam and temazepam were treated with IMCSzyme™ recombinant β-glucuronidase. Hydrolysis efficiency was assessed at 55°C and at room temperature (RT) using the recommended optimum pH. Hydrolysis efficiency for four other benzodiazepines was evaluated solely with positive patient samples. Maximum hydrolysis of glucuronide controls at 5 min at RT (mean analyte recovery ≥ 94% for oxazepam and lorazepam and ≥ 80% for temazepam) was observed. This was considerably faster than the optimized 30 min incubation time for the abalone β-glucuronidase at 65°C. Mean analyte recovery increased at longer incubation times at 55°C for temazepam only. Total analyte in patient samples compared well to targets from abalone hydrolysis after recombinant β-glucuronidase hydrolysis at RT with no incubation. Some matrix effect, differential reactivity, conjugation variability and transformation impacting total analyte recovery were indicated. The unique potential of the IMCSzyme™ recombinant β-glucuronidase was demonstrated with fast benzodiazepine hydrolysis at RT leading to decreased processing time without the need for heat activation. © The Author 2014. Published by Oxford University Press. All rights reserved. For Permissions, please email: journals.permissions@oup.com.

Rooster semen cryopreservation: effect of pedigree line and male age on postthaw sperm function.

PubMed

Long, J A; Bongalhardo, D C; Pelaéz, J; Saxena, S; Settar, P; O'Sullivan, N P; Fulton, J E

2010-05-01

The fertility rates of cryopreserved poultry semen are highly variable and not reliable for use in preservation of commercial genetic stocks. Our objective was to evaluate the cryosurvival of semen from 8 pedigreed layer lines at 2 different ages: the onset and end of commercial production. Semen from 160 roosters (20/line) was frozen individually with 11% glycerol at 6 and 12 mo of age. Glycerol was removed from thawed semen by Accudenz gradient centrifugation. The viability of thawed sperm from each male was determined using fluorescent live-dead staining and flow cytometry; sperm velocity parameters were measured using computerized motion analysis. The fertilizing ability of thawed sperm was evaluated in vitro by assessing hydrolysis of the inner perivitelline membrane. The postthaw function of sperm from the elite lines varied widely, despite the fact that fresh semen from all of these lines typically yielded high fertility rates. The percentage of thawed sperm with intact plasma membranes ranged from 27.8 + or - 2.1 to 49.6 + or - 1.9 and varied among lines and between age groups. Thawed sperm from 2 lines consistently demonstrated the highest and lowest motility parameters, whereas the velocity parameters of the remaining 6 lines varied widely. The mean number of hydrolysis points per square millimeter of inner perivitelline membrane ranged from 12.5 + or - 4.1 (line 2) to 103.3 + or - 30.2 (line 6). Age effects were observed for 4 out of 8 lines; however, improved postthaw sperm function at 12 mo of age was not consistent for all 3 assays. These results demonstrate variability among pedigreed lines in withstanding glycerol-based semen cryopreservation and provide a model for delineating genotypic and phenotypic factors affecting sperm cryosurvival.

Alkaline twin-screw extrusion pretreatment for fermentable sugar production.

PubMed

Liu, Chao; van der Heide, Evert; Wang, Haisong; Li, Bin; Yu, Guang; Mu, Xindong

2013-01-01

The inevitable depletion of fossil fuels has resulted in an increasing worldwide interest in exploring alternative and sustainable energy sources. Lignocellulose, which is the most abundant biomass on earth, is widely regarded as a promising raw material to produce fuel ethanol. Pretreatment is an essential step to disrupt the recalcitrance of lignocellulosic matrix for enzymatic saccharification and bioethanol production. This paper established an ATSE (alkaline twin-screw extrusion pretreatment) process using a specially designed twin-screw extruder in the presence of alkaline solution to improve the enzymatic hydrolysis efficiency of corn stover for the production of fermentable sugars. The ATSE pretreatment was conducted with a biomass/liquid ratio of 1/2 (w/w) at a temperature of 99°C without heating equipment. The results indicated that ATSE pretreatment is effective in improving the enzymatic digestibility of corn stover. Sodium hydroxide loading is more influential factor affecting both sugar yield and lignin degradation than heat preservation time. After ATSE pretreatment under the proper conditions (NaOH loading of 0.06 g/g biomass during ATSE and 1 hour heat preservation after extrusion), 71% lignin removal was achieved and the conversions of glucan and xylan in the pretreated biomass can reach to 83% and 89% respectively via subsequent enzymatic hydrolysis (cellulase loading of 20 FPU/g-biomass and substrate consistency of 2%). About 78% of the original polysaccharides were converted into fermentable sugars. With the physicochemical functions in extrusion, the ATSE method can effectively overcome the recalcitrance of lignocellulose for the production of fermentable sugars from corn stover. This process can be considered as a promising pretreatment method due to its relatively low temperature (99°C), high biomass/liquid ratio (1/2) and satisfied total sugar yield (78%), despite further study is needed for process optimization and cost reduction.

Effect of salting on back fat hydrolysis and oxidation

NASA Astrophysics Data System (ADS)

Tunieva, E. К; Nasonova, V. V.; Stanovova, I. A.; Spiridonov, К I.; Kurzova, A. A.

2017-09-01

Technological factors significantly affect the rate of hydrolytic and oxidative changes in fat. The aim of the research was to study the effect of sodium chloride on hydrolysis and oxidation of fat raw material, including the impact of thermal treatment. Back fat was minced, sodium chloride was added (in amounts of 0.0, 2.0, 3.5 or 5.0%), then it was thermally treated or not. Determination of the acid value (AV) was carried out by titration with aqueous potassium hydroxide of free fatty acids in the ether-alcohol solution of back fat; the peroxide value (PV) was based on oxidation of iodhydric acid with peroxides contained in fat followed by titration of released iodine with sodium thiosulphate. The thiobarbituric acid value (TBAV) was determined by the development of stained substances due to interaction of fat oxidation products with 2-thiobarbituric acid and measurement of color intensity using a spectrophotometer. Adding 5.0% sodium chloride to back fat led to a 30.1% decrease in AV. Addition of 2.0% sodium chloride inhibited the development of the oxidation products and led to a 17% decrease in the PV and to a 25% decrease in TBAV (p<0.05). In the presence of 5.0% sodium chloride, the secondary oxidation products significantly increased by 34.1% (p<0.05) and 24.3% (p<0.05) on days 1 and 3 of storage, respectively. Thermal treatment mitigated the effect of sodium chloride on the indicators of hydrolytic and oxidative spoilage (p>0.05). The results obtained showed an ambiguous effect of sodium chloride on the processes of fat oxidation, depending on dosage and the use of thermal treatment, justifying the necessity to develop approaches that allow reduction of the sodium chloride content in meat products that are not subjected to thermal treatment.

Techno-economic analysis of the industrial production of a low-cost enzyme using E. coli: the case of recombinant β-glucosidase.

PubMed

Ferreira, Rafael da Gama; Azzoni, Adriano Rodrigues; Freitas, Sindelia

2018-01-01

The enzymatic conversion of lignocellulosic biomass into fermentable sugars is a promising approach for producing renewable fuels and chemicals. However, the cost and efficiency of the fungal enzyme cocktails that are normally employed in these processes remain a significant bottleneck. A potential route to increase hydrolysis yields and thereby reduce the hydrolysis costs would be to supplement the fungal enzymes with their lacking enzymatic activities, such as β-glucosidase. In this context, it is not clear from the literature whether recombinant E. coli could be a cost-effective platform for the production of some of these low-value enzymes, especially in the case of on-site production. Here, we present a conceptual design and techno-economic evaluation of the production of a low-cost industrial enzyme using recombinant E. coli . In a simulated baseline scenario for β-glucosidase demand in a hypothetical second-generation ethanol (2G) plant in Brazil, we found that the production cost (316 US$/kg) was higher than what is commonly assumed in the literature for fungal enzymes, owing especially to the facility-dependent costs (45%) and to consumables (23%) and raw materials (25%). Sensitivity analyses of process scale, inoculation volume, and volumetric productivity indicated that optimized conditions may promote a dramatic reduction in enzyme cost and also revealed the most relevant factors affecting production costs. Despite the considerable technical and economic uncertainties that surround 2G ethanol and the large-scale production of low-cost recombinant enzymes, this work sheds light on some relevant questions and supports future studies in this field. In particular, we conclude that process optimization, on many fronts, may strongly reduce the costs of E. coli recombinant enzymes, in the context of tailor-made enzymatic cocktails for 2G ethanol production.

Trihalomethane hydrolysis in drinking water at elevated temperatures.

PubMed

Zhang, Xiao-Lu; Yang, Hong-Wei; Wang, Xiao-Mao; Karanfil, Tanju; Xie, Yuefeng F

2015-07-01

Hydrolysis could contribute to the loss of trihalomethanes (THMs) in the drinking water at elevated temperatures. This study was aimed at investigating THM hydrolysis pertaining to the storage of hot boiled water in enclosed containers. The water pH value was in the range of 6.1-8.2 and the water temperature was varied from 65 to 95 °C. The effects of halide ions, natural organic matter, and drinking water matrix were investigated. Results showed that the hydrolysis rates declined in the order following CHBrCl2 > CHBr2Cl > CHBr3 > CHCl3. THM hydrolysis was primarily through the alkaline pathway, except for CHCl3 in water at relatively low pH value. The activation energies for the alkaline hydrolysis of CHCl3, CHBrCl2, CHBr2Cl and CHBr3 were 109, 113, 115 and 116 kJ/mol, respectively. No hydrolysis intermediates could accumulate in the water. The natural organic matter, and probably other constituents, in drinking water could substantially decrease THM hydrolysis rates by more than 50%. When a drinking water was at 90 °C or above, the first order rate constants for THM hydrolysis were in the magnitude of 10(-2)‒10(-1) 1/h. When the boiled real tap water was stored in an enclosed container, THMs continued increasing during the first few hours and then kept decreasing later on due to the competition between hydrolysis and further formation. The removal of THMs, especially brominated THMs, by hydrolysis would greatly reduce one's exposure to disinfection by-products by consuming the boiled water stored in enclosed containers. Copyright © 2015 Elsevier Ltd. All rights reserved.

Optimizing isothiocyanate formation during enzymatic glucosinolate breakdown by adjusting pH value, temperature and dilution in Brassica vegetables and Arabidopsis thaliana

NASA Astrophysics Data System (ADS)

Hanschen, Franziska S.; Klopsch, Rebecca; Oliviero, Teresa; Schreiner, Monika; Verkerk, Ruud; Dekker, Matthijs

2017-01-01

Consumption of glucosinolate-rich Brassicales vegetables is associated with a decreased risk of cancer with enzymatic hydrolysis of glucosinolates playing a key role. However, formation of health-promoting isothiocyanates is inhibited by the epithiospecifier protein in favour of nitriles and epithionitriles. Domestic processing conditions, such as changes in pH value, temperature or dilution, might also affect isothiocyanate formation. Therefore, the influences of these three factors were evaluated in accessions of Brassica rapa, Brassica oleracea, and Arabidopsis thaliana. Mathematical modelling was performed to determine optimal isothiocyanate formation conditions and to obtain knowledge on the kinetics of the reactions. At 22 °C and endogenous plant pH, nearly all investigated plants formed nitriles and epithionitriles instead of health-promoting isothiocyanates. Response surface models, however, clearly demonstrated that upon change in pH to domestic acidic (pH 4) or basic pH values (pH 8), isothiocyanate formation considerably increases. While temperature also affects this process, the pH value has the greatest impact. Further, a kinetic model showed that isothiocyanate formation strongly increases due to dilution. Finally, the results show that isothiocyanate intake can be strongly increased by optimizing the conditions of preparation of Brassicales vegetables.

Optimization of Polygalacturonase Production from a Newly Isolated Thalassospira frigidphilosprofundus to Use in Pectin Hydrolysis: Statistical Approach

PubMed Central

Rekha, V. P. B.; Ghosh, Mrinmoy; Adapa, Vijayanand; Oh, Sung-Jong; Pulicherla, K. K.; Sambasiva Rao, K. R. S.

2013-01-01

The present study deals with the production of cold active polygalacturonase (PGase) by submerged fermentation using Thalassospira frigidphilosprofundus, a novel species isolated from deep waters of Bay of Bengal. Nonlinear models were applied to optimize the medium components for enhanced production of PGase. Taguchi orthogonal array design was adopted to evaluate the factors influencing the yield of PGase, followed by the central composite design (CCD) of response surface methodology (RSM) to identify the optimum concentrations of the key factors responsible for PGase production. Data obtained from the above mentioned statistical experimental design was used for final optimization study by linking the artificial neural network and genetic algorithm (ANN-GA). Using ANN-GA hybrid model, the maximum PGase activity (32.54 U/mL) was achieved at the optimized concentrations of medium components. In a comparison between the optimal output of RSM and ANN-GA hybrid, the latter favored the production of PGase. In addition, the study also focused on the determination of factors responsible for pectin hydrolysis by crude pectinase extracted from T. frigidphilosprofundus through the central composite design. Results indicated 80% degradation of pectin in banana fiber at 20°C in 120 min, suggesting the scope of cold active PGase usage in the treatment of raw banana fibers. PMID:24455722

Optimization of polygalacturonase production from a newly isolated Thalassospira frigidphilosprofundus to use in pectin hydrolysis: statistical approach.

PubMed

Rekha, V P B; Ghosh, Mrinmoy; Adapa, Vijayanand; Oh, Sung-Jong; Pulicherla, K K; Sambasiva Rao, K R S

2013-01-01

The present study deals with the production of cold active polygalacturonase (PGase) by submerged fermentation using Thalassospira frigidphilosprofundus, a novel species isolated from deep waters of Bay of Bengal. Nonlinear models were applied to optimize the medium components for enhanced production of PGase. Taguchi orthogonal array design was adopted to evaluate the factors influencing the yield of PGase, followed by the central composite design (CCD) of response surface methodology (RSM) to identify the optimum concentrations of the key factors responsible for PGase production. Data obtained from the above mentioned statistical experimental design was used for final optimization study by linking the artificial neural network and genetic algorithm (ANN-GA). Using ANN-GA hybrid model, the maximum PGase activity (32.54 U/mL) was achieved at the optimized concentrations of medium components. In a comparison between the optimal output of RSM and ANN-GA hybrid, the latter favored the production of PGase. In addition, the study also focused on the determination of factors responsible for pectin hydrolysis by crude pectinase extracted from T. frigidphilosprofundus through the central composite design. Results indicated 80% degradation of pectin in banana fiber at 20 °C in 120 min, suggesting the scope of cold active PGase usage in the treatment of raw banana fibers.

Central or Peripheral Delivery of an Adenosine A1 Receptor Agonist Improves Mechanical Allodynia in a Mouse Model of Painful Diabetic Neuropathy

PubMed Central

Katz, N. K.; Ryals, J. M.; Wright, D. E.

2014-01-01

Diabetic peripheral neuropathy is a common complication of diabetes mellitus, and a significant proportion of individuals suffer debilitating pain that significantly affects their quality of life. Unfortunately, symptomatic treatment options have limited efficacy, and often carry significant risk of systemic adverse effects. Activation of the adenosine A1 receptor (A1R) by the analgesic small molecule adenosine has been shown to have antinociceptive benefits in models of inflammatory and neuropathic pain. The current study used a mouse model of painful diabetic neuropathy to determine the effect of diabetes on endogenous adenosine production, and if central or peripheral delivery of adenosine receptor agonists could alleviate signs of mechanical allodynia in diabetic mice. Diabetes was induced using streptozocin in male A/J mice. Mechanical withdrawal thresholds were measured weekly to characterize neuropathy phenotype. Hydrolysis of AMP into adenosine by ectonucleotidases was determined in the dorsal root ganglia (DRG) and spinal cord at 8-weeks post-induction of diabetes. AMP, adenosine and the specific A1R agonist, N6-cyclopentyladenosine (CPA), were administered both centrally (intrathecal) and peripherally (intraplantar) to determine the effect of activation of adenosine receptors on mechanical allodynia in diabetic mice. Eight weeks post-induction, diabetic mice displayed significantly decreased hydrolysis of extracellular AMP in the DRG; at this same time, diabetic mice displayed significantly decreased mechanical withdrawal thresholds compared to nondiabetic controls. Central delivery AMP, adenosine and CPA significantly improved mechanical withdrawal thresholds in diabetic mice. Surprisingly, peripheral delivery of CPA also improved mechanical allodynia in diabetic mice. This study provides new evidence that diabetes significantly affects endogenous AMP hydrolysis, suggesting that altered adenosine production could contribute to the development of painful diabetic neuropathy. Moreover, central and peripheral activation of A1R significantly improved mechanical sensitivity, warranting further investigation into this important antinociceptive pathway as a novel therapeutic option for the treatment of painful diabetic neuropathy. PMID:25451280

Phenotype overlap in Xylella fastidiosa is controlled by the cyclic di-GMP phosphodiesterase Eal in response to antibiotic exposure and diffusible signal factor-mediated cell-cell signaling.

PubMed

de Souza, Alessandra A; Ionescu, Michael; Baccari, Clelia; da Silva, Aline M; Lindow, Steven E

2013-06-01

Eal is an EAL domain protein in Xylella fastidiosa homologous to one involved in resistance to tobramycin in Pseudomonas aeruginosa. EAL and HD-GYP domain proteins are implicated in the hydrolysis of the secondary messenger bis-(3'-5')-cyclic dimeric GMP (cyclic di-GMP). Cell density-dependent communication mediated by a Diffusible Signal Factor (DSF) also modulates cyclic di-GMP levels in X. fastidiosa, thereby controlling the expression of virulence genes and genes involved in insect transmission. The possible linkage of Eal to both extrinsic factors such as antibiotics and intrinsic factors such as quorum sensing, and whether both affect virulence, was thus addressed. Expression of eal was induced by subinhibitory concentrations of tobramycin, and an eal deletion mutant was more susceptible to this antibiotic than the wild-type strain and exhibited phenotypes similar to those of an rpfF deletion mutant blocked in DSF production, such as hypermotility, reduced biofilm formation, and hypervirulence to grape. Consistent with that, the rpfF mutant was more susceptible than the wild-type strain to tobramycin. Therefore, we propose that cell-cell communication and antibiotic stress can apparently lead to similar modulations of cyclic di-GMP in X. fastidiosa, resulting in similar phenotypes. However, the effect of cell density is dominant compared to that of antibiotic stress, since eal is suppressed by RpfF, which may prevent inappropriate behavioral changes in response to antibiotic stress when DSF accumulates.

Kinetics, mechanisms, and influencing factors on the treatment of haloacetonitriles (HANs) in water by two household heating devices.

PubMed

Shi, Wendong; Wang, Lei; Chen, Baiyang

2017-04-01

Haloacetonitriles (HANs) are a group of nitrogenous disinfection by-products (DBPs) commonly found in treated water with potential carcinogenic, cytotoxic, and genotoxic risks. In order to control HANs and understand their real intake levels by people via drinking water, this study evaluated a list of structural, operational, and environmental factors affecting the treatment of HANs by two domestic heating devices, i.e., an electric boiler and a microwave oven. Results show that the concentrations of HANs decreased exponentially over time with increasing temperature, water turbulence, and bubbles, and the phenomena were most likely due to a combined effect of volatilization and hydrolysis. Among HANs, the lability increased with increasing halogenation degrees (i.e., tri- > di- > mono- HANs) yet decreasing halogen molecular weights (i.e., Cl- > Br- > I- HANs); such trends were well captured by quantitative structure activity relationship models (R 2  = 0.99). Operational factors played critical roles in controlling HANs too, including the rate of heating, water volume, water temperature at time of pouring, cooling method, and capping condition, suggesting that people could benefit from proper handling methods and procedures. In addition, HANs added to tap water exhibited higher removals than those added to ultrapure water, probably because of the presence of free chlorine in tap water. Copyright © 2017 Elsevier Ltd. All rights reserved.

Hydrolysis mechanism of methyl parathion evidenced by Q-Exactive mass spectrometry.

PubMed

Liu, Yuan; Zhang, Caixiang; Liao, Xiaoping; Luo, Yinwen; Wu, Sisi; Wang, Jianwei

2015-12-01

Organophosphorus pesticides (OPPs), a kind of widely used pesticides, are currently attracting great attention due to their adverse effects on human central nervous systems, particularly in children. Although the hydrolysis behavior of OPPs has been studied well, its hydrolysis mechanism remained controversial, especially at various pH conditions, partly due to their relatively complex structures and abundant moieties that were prone to be attacked by nucleophiles. The Q-Exactive mass spectrometer, part of those hybrid high-resolution mass spectrometers (HRMS), was used to determine hydrolysis products of methyl parathion (MP), a kind of OPPs in situ buffer aqueous solution with pH ranging from 1 to 13 in this study. Most of the complex hydrolysis products of MP were identified due to the high sensitivity and accuracy of HRMS. The results demonstrated that the hydrolysis rate and pathway of MP were strong pH dependent. With the increase of pH, the hydrolysis rate of MP increased, and two different reaction mechanisms were identified: SN (2)@P pathway dominated the hydrolysis process at high pH (e.g., pH ≥ 11) while SN (2)@C was the main behavior at low pH (e.g., pH ≤ 9). This study helps understand the hydrolysis mechanism of OPPs at various pH and extends the use of Q-Exactive mass spectrometry in identifying organic pollutants and their degradation products in environmental matrices.

Optimization of enzymatic hydrolysis and fermentation conditions for improved bioethanol production from potato peel residues.

PubMed

Ben Taher, Imen; Fickers, Patrick; Chniti, Sofien; Hassouna, Mnasser

2017-03-01

The aim of this work was the optimization of the enzyme hydrolysis of potato peel residues (PPR) for bioethanol production. The process included a pretreatment step followed by an enzyme hydrolysis using crude enzyme system composed of cellulase, amylase and hemicellulase, produced by a mixed culture of Aspergillus niger and Trichoderma reesei. Hydrothermal, alkali and acid pretreatments were considered with regards to the enhancement of enzyme hydrolysis of potato peel residues. The obtained results showed that hydrothermal pretreatment lead to a higher enzyme hydrolysis yield compared to both acid and alkali pretreatments. Enzyme hydrolysis was also optimized for parameters such as temperature, pH, substrate loading and surfactant loading using a response surface methodology. Under optimized conditions, 77 g L -1 of reducing sugars were obtained. Yeast fermentation of the released reducing sugars led to an ethanol titer of 30 g L -1 after supplementation of the culture medium with ammonium sulfate. Moreover, a comparative study between acid and enzyme hydrolysis of potato peel residues was investigated. Results showed that enzyme hydrolysis offers higher yield of bioethanol production than acid hydrolysis. These results highlight the potential of second generation bioethanol production from potato peel residues treated with onsite produced hydrolytic enzymes. © 2017 American Institute of Chemical Engineers Biotechnol. Prog., 33:397-406, 2017. © 2017 American Institute of Chemical Engineers.

Sequential Desorption of Nitroaromatic Compounds (NAC) from Soils

DTIC Science & Technology

2005-03-01

the soil solution . Weissmahr et al. [20] suggest an electron donor acceptor (EDA) complex between oxygens of the siloxane surface of the clays and the...release of NACs into the soil solution . At high pH values desorption is superimposed by NACs hydrolysis. Therefore, in- creasing pH values impedes the...presented demonstrate that both the cation present in the soil solution and its concentration may affect the desorption behavior of NACs in contaminated soils

Hydrolysis kinetics of secoisolariciresinol diglucoside oligomers from flaxseed.

PubMed

Yuan, Jian-Ping; Li, Xin; Xu, Shi-Ping; Wang, Jiang-Hai; Liu, Xin

2008-11-12

Flaxseed is the richest dietary source of the lignan secoisolariciresinol diglucoside (SDG) and contains the largest amount of SDG oligomers, which are often hydrolyzed to break the ester linkages for the release of SDG and the glycosidic bonds for the release of secoisolariciresinol (SECO). The alkaline hydrolysis reaction kinetics of SDG oligomers from flaxseed and the acid hydrolysis process of SDG and other glucosides were investigated. For the kinetic modeling, a pseudo-first-order reaction was assumed. The results showed that the alkaline hydrolysis of SDG oligomers followed first-order reaction kinetics under mild alkaline hydrolytic conditions and that the concentration of sodium hydroxide had a strong influence on the activation energy of the alkaline hydrolysis of SDG oligomers. The results also indicated that the main acid hydrolysates of SDG included secoisolariciresinol monoglucoside (SMG), SECO, and anhydrosecoisolariciresinol (anhydro-SECO) and that the extent and the main hydrolysates of the acid hydrolysis reaction depended on the acid concentration, hydrolysis temperature, and time. In addition, the production and change of p-coumaric acid glucoside, ferulic acid glucoside and their methyl esters and p-coumaric acid, ferulic acid, and their methyl esters during the process of hydrolysis was also investigated.

Impacts of microalgae pre-treatments for improved anaerobic digestion: thermal treatment, thermal hydrolysis, ultrasound and enzymatic hydrolysis.

PubMed

Ometto, Francesco; Quiroga, Gerardo; Pšenička, Pavel; Whitton, Rachel; Jefferson, Bruce; Villa, Raffaella

2014-11-15

Anaerobic digestion (AD) of microalgae is primarily inhibited by the chemical composition of their cell walls containing biopolymers able to resist bacterial degradation. Adoption of pre-treatments such as thermal, thermal hydrolysis, ultrasound and enzymatic hydrolysis have the potential to remove these inhibitory compounds and enhance biogas yields by degrading the cell wall, and releasing the intracellular algogenic organic matter (AOM). This work investigated the effect of four pre-treatments on three microalgae species, and their impact on the quantity of soluble biomass released in the media and thus on the digestion process yields. The analysis of the composition of the soluble COD released and of the TEM images of the cells showed two main degradation actions associated with the processes: (1) cell wall damage with the release of intracellular AOM (thermal, thermal hydrolysis and ultrasound) and (2) degradation of the cell wall constituents with the release of intracellular AOM and the solubilisation of the cell wall biopolymers (enzymatic hydrolysis). As a result of this, enzymatic hydrolysis showed the greatest biogas yield increments (>270%) followed by thermal hydrolysis (60-100%) and ultrasounds (30-60%). Copyright © 2014 Elsevier Ltd. All rights reserved.

Application of solid-acid catalyst and marine macro-algae Gracilaria verrucosa to production of fermentable sugars.

PubMed

Jeong, Gwi-Taek; Kim, Sung-Koo; Park, Don-Hee

2015-04-01

In this study, the hydrolysis of marine macro-algae Gracilaria verrucosa with a solid-acid catalyst was investigated. To optimize the hydrolysis, four reaction factors, including liquid-to-solid ratio, catalyst loading, reaction temperature, and reaction time, were investigated. In the results, the highest total reducing sugar (TRS) yield, 61 g/L (51.9%), was obtained under the following conditions: 1:7.5 solid-to-liquid ratio, 15% (w/v) catalyst loading, 140 °C reaction temperature, and 150 min reaction time. Under these conditions, 10.7 g/L of 5-HMF and 2.5 g/L of levulinic acid (LA) were generated. The application of solid-acid catalyst and marine macro-algae resources shows a very high potential for production of fermentable sugars. Copyright © 2015 Elsevier Ltd. All rights reserved.

Site- and species-specific hydrolysis rates of heroin.

PubMed

Szöcs, Levente; Orgován, Gábor; Tóth, Gergő; Kraszni, Márta; Gergó, Lajos; Hosztafi, Sándor; Noszál, Béla

2016-06-30

The hydroxide-catalyzed non-enzymatic, simultaneous and consecutive hydrolyses of diacetylmorphine (DAM, heroin) are quantified in terms of 10 site- and species-specific rate constants in connection with also 10 site- and species-specific acid-base equilibrium constants, comprising all the 12 coexisting species in solution. This characterization involves the major and minor decomposition pathways via 6-acetylmorphine and 3-acetylmorphine, respectively, and morphine, the final product. Hydrolysis has been found to be 18-120 times faster at site 3 than at site 6, depending on the status of the amino group and the rest of the molecule. Nitrogen protonation accelerates the hydrolysis 5-6 times at site 3 and slightly less at site 6. Hydrolysis rate constants are interpreted in terms of intramolecular inductive effects and the concomitant local electron densities. Hydrolysis fraction, a new physico-chemical parameter is introduced and determined to quantify the contribution of the individual microspecies to the overall hydrolysis. Hydrolysis fractions are depicted as a function of pH. Copyright © 2016 Elsevier B.V. All rights reserved.

The effect of biological pretreatment with the selective white-rot fungus Echinodontium taxodii on enzymatic hydrolysis of softwoods and hardwoods.

PubMed

Yu, Hongbo; Guo, Guoning; Zhang, Xiaoyu; Yan, Keliang; Xu, Chunyan

2009-11-01

Selective white-rot fungi have shown potential for lignocellulose pretreatment. In the study, a new fungal isolate, Echinodontium taxodii 2538, was used in biological pretreatment to enhance the enzymatic hydrolysis of two native woods: Chinese willow (hardwood) and China-fir (softwood). E. taxodii preferentially degraded the lignin during the pretreatment, and the pretreated woods showed significant increases in enzymatic hydrolysis ratios (4.7-fold for hardwood and 6.3-fold for softwood). To better understand effects of biological pretreatment on enzymatic hydrolysis, enzyme-substrate interactions were investigated. It was observed that E. taxodii enhanced initial adsorption of cellulase but which did not always translate to high initial hydrolysis rate. However, the rate of change in hydrolysis rate declined dramatically with decreasing irreversible adsorption of cellulase. Thus, the enhancement of enzymatic hydrolysis was attributed to the decline of irreversible adsorption which may result from partial lignin degradation and alteration in lignin structure after biological pretreatment.

Enzymatic saccharification of pretreated wheat straw: comparison of solids-recycling, sequential hydrolysis and batch hydrolysis.

PubMed

Pihlajaniemi, Ville; Sipponen, Satu; Sipponen, Mika H; Pastinen, Ossi; Laakso, Simo

2014-02-01

In the enzymatic hydrolysis of lignocellulose materials, the recycling of the solid residue has previously been considered within the context of enzyme recycling. In this study, a steady state investigation of a solids-recycling process was made with pretreated wheat straw and compared to sequential and batch hydrolysis at constant reaction times, substrate feed and liquid and enzyme consumption. Compared to batch hydrolysis, the recycling and sequential processes showed roughly equal hydrolysis yields, while the volumetric productivity was significantly increased. In the 72h process the improvement was 90% due to an increased reaction consistency, while the solids feed was 16% of the total process constituents. The improvement resulted primarily from product removal, which was equally efficient in solids-recycling and sequential hydrolysis processes. No evidence of accumulation of enzymes beyond the accumulation of the substrate was found in recycling. A mathematical model of solids-recycling was constructed, based on a geometrical series. Copyright © 2013 Elsevier Ltd. All rights reserved.

Mechanistic investigation in ultrasound induced enhancement of enzymatic hydrolysis of invasive biomass species.

PubMed

Borah, Arup Jyoti; Agarwal, Mayank; Poudyal, Manisha; Goyal, Arun; Moholkar, Vijayanand S

2016-08-01

This study has assessed four invasive weeds, viz. Saccharum spontaneum (SS), Mikania micrantha (MM), Lantana camara (LC) and Eichhornia crassipes (EC) for enzymatic hydrolysis prior to bioalcohol fermentation. Enzymatic hydrolysis of pretreated biomasses of weeds has been conducted with mechanical agitation and sonication under constant (non-optimum) conditions. Profiles of total reducible sugar release have been fitted to HCH-1 model of enzymatic hydrolysis using Genetic Algorithm. Trends in parameters of this model reveal physical mechanism of ultrasound-induced enhancement of enzymatic hydrolysis. Sonication accelerates hydrolysis kinetics by ∼10-fold. This effect is contributed by several causes, attributed to intense micro-convection generated during sonication: (1) increase in reaction velocity, (2) increase in enzyme-substrate affinity, (3) reduction in product inhibition, and (4) enhancement of enzyme activity due to conformational changes in its secondary structure. Enhancement effect of sonication is revealed to be independent of conditions of enzymatic hydrolysis - whether optimum or non-optimum. Copyright © 2016 Elsevier Ltd. All rights reserved.

Mechanism of Elongation Factor-G-mediated Fusidic Acid Resistance and Fitness Compensation in Staphylococcus aureus *

PubMed Central

Koripella, Ravi Kiran; Chen, Yang; Peisker, Kristin; Koh, Cha San; Selmer, Maria; Sanyal, Suparna

2012-01-01

Antibiotic resistance in bacteria is often associated with fitness loss, which is compensated by secondary mutations. Fusidic acid (FA), an antibiotic used against pathogenic bacteria Staphylococcus aureus, locks elongation factor-G (EF-G) to the ribosome after GTP hydrolysis. To clarify the mechanism of fitness loss and compensation in relation to FA resistance, we have characterized three S. aureus EF-G mutants with fast kinetics and crystal structures. Our results show that a significantly slower tRNA translocation and ribosome recycling, plus increased peptidyl-tRNA drop-off, are the causes for fitness defects of the primary FA-resistant mutant F88L. The double mutant F88L/M16I is three to four times faster than F88L in both reactions and showed no tRNA drop-off, explaining its fitness compensatory phenotype. The M16I mutation alone showed hypersensitivity to FA, higher activity, and somewhat increased affinity to GTP. The crystal structures demonstrate that Phe-88 in switch II is a key residue for FA locking and also for triggering interdomain movements in EF-G essential for its function, explaining functional deficiencies in F88L. The mutation M16I loosens the hydrophobic core in the G domain and affects domain I to domain II contact, resulting in improved activity both in the wild-type and F88L background. Thus, FA-resistant EF-G mutations causing fitness loss and compensation operate by affecting the conformational dynamics of EF-G on the ribosome. PMID:22767604

Effect of environmental factors on the complexation of iron and humic acid.

PubMed

Fang, Kai; Yuan, Dongxing; Zhang, Lei; Feng, Lifeng; Chen, Yaojin; Wang, Yuzhou

2015-01-01

A method of size exclusion chromatography coupled with ultraviolet spectrophotometry and off-line graphite furnace atomic absorption spectrometry was developed to assess the complexation properties of iron (Fe) and humic acid (HA) in a water environment. The factors affecting the complexation of Fe and HA, such as ionic strength, pH, temperature and UV radiation, were investigated. The Fe-HA complex residence time was also studied. Experimental results showed that pH could influence the deprotonation of HA and hydrolysis of Fe, and thus affected the complexation of Fe and HA. The complexation was greatly disrupted by the presence of NaCl. Temperature had some influence on the complexation. The yield of Fe-HA complexes showed a small decrease at high levels of UV radiation, but the effect of UV radiation on Fe-HA complex formation at natural levels could be neglected. It took about 10 hr for the complexation to reach equilibrium, and the Fe-HA complex residence time was about 20 hr. Complexation of Fe and HA reached a maximum level under the conditions of pH 6, very low ionic strength, in the dark and at a water temperature of about 25°C, for 10 hr. It was suggested that the Fe-HA complex could form mainly in freshwater bodies and reach high levels in the warm season with mild sunlight radiation. With changing environmental parameters, such as at lower temperature in winter or higher pH and ionic strength in an estuary, the concentration of the Fe-HA complex would decrease. Copyright © 2014. Published by Elsevier B.V.

Low frequency ultrasonic-assisted hydrolysis of starch in the presence of α-amylase.

PubMed

Gaquere-Parker, Anne; Taylor, Tamera; Hutson, Raihannah; Rizzo, Ashley; Folds, Aubrey; Crittenden, Shastina; Zahoor, Neelam; Hussein, Bilal; Arruda, Aaron

2018-03-01

Hydrolysis of starch is an important process in the food industry and in the production of bioethanol or smaller carbohydrate molecules that can be used as starting blocks for chemical synthesis. Such hydrolysis can be enhanced by lowering the pH, heating the reaction mixture or catalyzing the reaction with enzymes. This study reports the effect of sonication on the reaction rate of starch hydrolysis at different temperatures, in the presence or absence of alpha-amylase. Starch Azure, a commercially available potato starch covalently linked with Remazol Brilliant Blue, has been chosen since its hydrolysis releases a blue dye, which concentration can be monitored by UV Vis spectroscopy. Ultrasounds, regardless of experimental conditions, provide the highest reaction rate for such hydrolysis. Copyright © 2017 Elsevier B.V. All rights reserved.

The hydrolysis of polyimides

NASA Technical Reports Server (NTRS)

Hoagland, P. D.; Fox, S. W.

1973-01-01

Thermal polymerization of aspartic acid produces a polysuccinimide (I), a chain of aspartoyl residues. An investigation was made of the alkaline hydrolysis of the imide rings of (I) which converts the polyimide to a polypeptide. The alkaline hydrolysis of polyimides can be expected to be kinetically complex due to increasing negative charge generated by carboxylate groups. For this reason, a diimide, phthaloyl-DL-aspartoyl-beta-alanine (IIA) was synthesized for a progressive study of the hydrolysis of polyimides. In addition, this diimide (IIA) can be related to thalidomide and might be expected to exhibit similar reactivity during hydrolysis of the phthalimide ring.

The Effect of Sulphate Anions on the Ultrafine Titania Nucleation

NASA Astrophysics Data System (ADS)

Kotsyubynsky, Volodymyr O.; Myronyuk, Ivan F.; Chelyadyn, Volodymyr L.; Hrubiak, Andriy B.; Moklyak, Volodymyr V.; Fedorchenko, Sofia V.

2017-05-01

The phenomenological model of sulphate anions effect on the nanodispersed titania synthesis during hydrolysis of titanium tetrachloride was studied. It was proposed that both chelating and bridging bidentate complexes formation between sulphate anions and octahedrally coordinated [Ti(OH)h(OH2)6-h](4-h)+ mononers is the determinative factor for anatase phase nucleation.

Hydrolysis of ammonia borane as a hydrogen source: fundamental issues and potential solutions towards implementation.

PubMed

Sanyal, Udishnu; Demirci, Umit B; Jagirdar, Balaji R; Miele, Philippe

2011-12-16

In today's era of energy crisis and global warming, hydrogen has been projected as a sustainable alternative to depleting CO(2)-emitting fossil fuels. However, its deployment as an energy source is impeded by many issues, one of the most important being storage. Chemical hydrogen storage materials, in particular B-N compounds such as ammonia borane, with a potential storage capacity of 19.6 wt % H(2) and 0.145 kg(H2)L(-1), have been intensively studied from the standpoint of addressing the storage issues. Ammonia borane undergoes dehydrogenation through hydrolysis at room temperature in the presence of a catalyst, but its practical implementation is hindered by several problems affecting all of the chemical compounds in the reaction scheme, including ammonia borane, water, borate byproducts, and hydrogen. In this Minireview, we exhaustively survey the state of the art, discuss the fundamental problems, and, where applicable, propose solutions with the prospect of technological applications. Copyright © 2011 WILEY-VCH Verlag GmbH & Co. KGaA, Weinheim.

Synthesis, Aqueous Reactivity, and Biological Evaluation of Carboxylic Acid Ester-Functionalized Platinum–Acridine Hybrid Anticancer Agents

PubMed Central

Graham, Leigh A.; Suryadi, Jimmy; West, Tiffany K.; Kucera, Gregory L.; Bierbach, Ulrich

2012-01-01

The synthesis of platinum–acridine hybrid agents containing carboxylic acid ester groups is described. The most active derivatives and the unmodified parent compounds showed up to 6-fold higher activity in ovarian cancer (OVCAR-3) and breast cancer (MCF-7, MDA-MB-23) cell lines than cisplatin. Inhibition of cell proliferation at nanomolar concentrations was observed in pancreatic (PANC-1) and non-small cell lung cancer cells (NSCLC, NCI-H460) of 80- and 150-fold, respectively. Introduction of the ester groups did not affect the cytotoxic properties of the hybrids, which form the same monofunctional–intercalative DNA adducts as the parent compounds, as demonstrated in a plasmid unwinding assay. In-line high-performance liquid chromatography and electrospray mass spectrometry (LC-ESMS) shows that the ester moieties undergo platinum-mediated hydrolysis in a chloride concentration-dependent manner to form carboxylate chelates. Potential applications of the chloride-sensitive ester hydrolysis as a self-immolative release mechanism for tumor-selective delivery of platinum–acridines are discussed. PMID:22871158

Effect of banana pulp and peel flour on physicochemical properties and in vitro starch digestibility of yellow alkaline noodles.

PubMed

Ramli, Saifullah; Alkarkhi, Abbas F M; Shin Yong, Yeoh; Min-Tze, Liong; Easa, Azhar Mat

2009-01-01

The present study describes the utilization of banana--Cavendish (Musa acuminata L., cv cavendshii) and Dream (Musa acuminata colla. AAA, cv 'Berangan')--pulp and peel flours as functional ingredients in yellow alkaline noodles. Noodles were prepared by partial substitution of wheat flour with ripe banana pulp or peel flours. In most cases, the starch hydrolysis index, predicted glycaemic index (pGI) and physicochemical properties of cooked noodles were affected by banana flour addition. In general, the pGI values of cooked noodles were in the order; banana peel noodles < banana pulp noodles < control noodles. Since the peel flour was higher in total dietary fibre but lower in resistant starch contents than the pulp flour, the low pGI of banana peel noodles was mainly due to its high dietary fibre content. In conclusion, banana pulp and peel flour could be useful for controlling starch hydrolysis of yellow noodles, even though some physicochemical properties of the noodles were altered.

Importance of cellulase cocktails favoring hydrolysis of cellulose.

PubMed

Victoria, Juliet; Odaneth, Annamma; Lali, Arvind

2017-07-03

Depolymerization of lignocellulosic biomass is catalyzed by groups of enzymes whose action is influenced by substrate features and the composition of cellulase preparation. Cellulases contain a mixture of variety of enzymes, whose proportions dictate the saccharification of biomass. In the current study, four cellulase preparation varying in their composition were used to hydrolyze two types of alkali-treated biomass (aqueous ammonia-treated rice straw and sodium hydroxide-treated rice straw) to study the effect on catalytic rate, saccharification yields, and sugar release profile. We found that substrate features affected the extent of saccharification but had minimal effect on the sugar release pattern. In addition, complete hydrolysis to glucose was observed with enzyme preparation having at least a cellobiase units (CBU)/carboxymethyl cellulose (CMC) ratio (>0.15), while a modified enzyme ratio can be used for oligosaccharide synthesis. Thus, cellulase preparation with defined ratios of the three main enzymes can improve the saccharification which is of utmost importance in defining the success of lignocellulose-based economies.

Optimization and evaluation of alkaline potassium permanganate pretreatment of corncob.

PubMed

Ma, Lijuan; Cui, Youzhi; Cai, Rui; Liu, Xueqiang; Zhang, Cuiying; Xiao, Dongguang

2015-03-01

Alkaline potassium permanganate solution (APP) was applied to the pretreatment of corncob with a simple and effective optimization of APP concentration, reaction time, temperature and solid to liquid ratio (SLR). The optimized pretreatment conditions were at 2% (w/v) potassium permanganate with SLR of 1:10 treating for 6h at 50°C. This simple one-step treatment resulted in significant 94.56% of the cellulose and 81.47% of the hemicellulose recoveries and 46.79% of the lignin removal of corncob. The reducing sugar in the hydrolysate from APP-pretreated corncob was 8.39g/L after 12h enzymatic hydrolysis, which was 1.44 and 1.29 folds higher than those from raw and acid pretreated corncobs. Physical characteristics, crystallinity and structure of the pretreated corncob were analyzed and assessed by SEM, XRD and FTIR. The APP pretreatment process was novel and enhanced enzymatic hydrolysis of lignocellulose by affecting composition and structural features. Copyright © 2014 Elsevier Ltd. All rights reserved.

Recycling the liquid fraction of alkaline hydrogen peroxide in the pretreatment of corn stover.

PubMed

Alencar, Bárbara Ribeiro Alves; Reis, Alexandre Libanio Silva; de Souza, Raquel de Fatima Rodrigues; Morais, Marcos Antônio; Menezes, Rômulo Simões Cezar; Dutra, Emmanuel Damilano

2017-10-01

The aim of this study was to evaluate the influence of recycling the liquid fraction of pretreatment with alkaline hydrogen peroxide (AHP) on the hydrolysis of corn stover. Corn stover was pretreated in the traditional condition with 7.5% v/v H 2 O 2 . After pretreatment, the solids were separated from the liquid fraction and five successive reuse cycles of the liquid fraction were tested. The solid fraction from pretreatment in each recycle was submitted to enzymatic hydrolysis. The number of recycles had a linear negative effect (R 2 =0.98) on biomass delignification efficiency and also affected negatively the enzymatic conversion efficiency. Despite the decrease in efficiency after each recycling step, reuse of the liquid fraction leads to reduction in water, H 2 O 2 and NaOH consumption of up to 57.6%, 59.6% and 57.6%, respectively. These findings point to an efficient recycling technology, which may reduce costs and save water. Copyright © 2017 Elsevier Ltd. All rights reserved.

Hydrolytic activities of extracellular enzymes in thermophilic and mesophilic anaerobic sequencing-batch reactors treating organic fractions of municipal solid wastes.

PubMed

Kim, Hyun-Woo; Nam, Joo-Youn; Kang, Seok-Tae; Kim, Dong-Hoon; Jung, Kyung-Won; Shin, Hang-Sik

2012-04-01

Extracellular enzymes offer active catalysis for hydrolysis of organic solid wastes in anaerobic digestion. To evidence the quantitative significance of hydrolytic enzyme activities for major waste components, track studies of thermophilic and mesophilic anaerobic sequencing-batch reactors (TASBR and MASBR) were conducted using a co-substrate of real organic wastes. During 1day batch cycle, TASBR showed higher amylase activity for carbohydrate (46%), protease activity for proteins (270%), and lipase activity for lipids (19%) than MASBR. In particular, the track study of protease identified that thermophilic anaerobes degraded protein polymers much more rapidly. Results revealed that differences in enzyme activities eventually affected acidogenic and methanogenic performances. It was demonstrated that the superior nature of enzymatic capability at thermophilic condition led to successive high-rate acidogenesis and 32% higher CH(4) recovery. Consequently, these results evidence that the coupling thermophilic digestion with sequencing-batch operation is a viable option to promote enzymatic hydrolysis of organic particulates. Copyright © 2012 Elsevier Ltd. All rights reserved.

Characterization of a novel debranching enzyme from Nostoc punctiforme possessing a high specificity for long branched chains

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

Choi, Ji-Hye; Lee, Heeseob; Kim, Young-Wan

2009-01-09

A novel debranching enzyme from Nostoc punctiforme PCC73102 (NPDE) exhibits hydrolysis activity toward both {alpha}-(1,6)- and {alpha}-(1,4)-glucosidic linkages. The action patterns of NPDE revealed that branched chains are released first, and the resulting maltooligosaccharides are then hydrolyzed. Analysis of the reaction with maltooligosaccharide substrates labeled with {sup 14}C-glucose at the reducing end shows that NPDE specifically liberates glucose from the reducing end. Kinetic analyses showed that the hydrolytic activity of NPDE is greatly affected by the length of the substrate. The catalytic efficiency of NPDE increased considerably upon using substrates that can occupy at least eight glycone subsites such asmore » maltononaose and maltooctaosyl-{alpha}-(1,6)-{beta}-cyclodextrin. These results imply that NPDE has a unique subsite structure consisting of -8 to +1 subsites. Given its unique subsite structure, side chains shorter than maltooctaose in amylopectin were resistant to hydrolysis by NPDE, and the population of longer side chains was reduced.« less

Evaluation of Brewer's spent yeast to produce flavor enhancer nucleotides: influence of serial repitching.

PubMed

Vieira, Elsa; Brandão, Tiago; Ferreira, Isabel M P L V O

2013-09-18

The present work evaluates the influence of serial yeast repitching on nucleotide composition of brewer's spent yeast extracts produced without addition of exogenous enzymes. Two procedures for disrupting cell walls were compared, and the conditions for low-cost and efficient RNA hydrolysis were selected. A HILIC methodology was validated for the quantification of nucleotides and nucleosides in yeast extracts. Thirty-seven samples of brewer's spent yeast ( Saccharomyces pastorianus ) organized according to the number of serial repitchings were analyzed. Nucleotides accounted for 71.1-88.2% of the RNA products; 2'AMP was the most abundant (ranging between 0.08 and 2.89 g/100 g dry yeast). 5'GMP content ranged between 0.082 and 0.907 g/100 g dry yeast. The sum of 5'GMP, 5'IMP, and 5'AMP represented between 25 and 32% of total nucleotides. This works highlights for the first time that although serial repitching influences the content of monophosphate nucleotides and nucleosides, the profiles of these RNA hydrolysis products are not affected.

Role of lecithin-cholesterol acyltransferase in the metabolism of oxidized phospholipids in plasma: studies with platelet-activating factor-acetyl hydrolase-deficient plasma.

PubMed

Subramanian, V S; Goyal, J; Miwa, M; Sugatami, J; Akiyama, M; Liu, M; Subbaiah, P V

1999-07-09

To determine the relative importance of platelet-activating factor-acetylhydrolase (PAF-AH) and lecithin-cholesterol acyltransferase (LCAT) in the hydrolysis of oxidized phosphatidylcholines (OXPCs) to lyso-phosphatidylcholine (lyso-PC), we studied the formation and metabolism of OXPCs in the plasma of normal and PAF-AH-deficient subjects. Whereas the loss of PC following oxidation was similar in the deficient and normal plasmas, the formation of lyso-PC was significantly lower, and the accumulation of OXPC was higher in the deficient plasma. Isolated LDL from the PAF-AH-deficient subjects was more susceptible to oxidation, and stimulated adhesion molecule synthesis in endothelial cells, more than the normal LDL. Oxidation of 16:0-[1-14C]-18:2 PC, equilibrated with plasma PC, resulted in the accumulation of labeled short- and long-chain OXPCs, in addition to the labeled aqueous products. The formation of the aqueous products decreased by 80%, and the accumulation of short-chain OXPC increased by 110% in the deficient plasma, compared to the normal plasma, showing that PAF-AH is predominantly involved in the hydrolysis of the truncated OXPCs. Labeled sn-2-acyl group from the long-chain OXPC was not only hydrolyzed to free fatty acid, but was preferentially transferred to diacylglycerol, in both the normal and deficient plasmas. In contrast, the acyl group from unoxidized PC was transferred only to cholesterol, showing that the specificity of LCAT is altered by OXPC. It is concluded that, while PAF-AH carries out the hydrolysis of mainly truncated OXPCs, LCAT hydrolyzes and transesterifies the long-chain OXPCs.

Kunitz trypsin inhibitor in addition to Bowman-Birk inhibitor influence stability of lunasin against pepsin-pancreatin hydrolysis.

PubMed

Price, Samuel J; Pangloli, Philipus; Krishnan, Hari B; Dia, Vermont P

2016-12-01

Soybean contains several biologically active components and one of this belongs to the bioactive peptide group. The objectives of this study were to produce different lunasin-enriched preparations (LEP) and determine the effect of Bowman-Birk inhibitor (BBI) and Kunitz trypsin inhibitor (KTI) concentrations on the stability of lunasin against pepsin-pancreatin hydrolysis (PPH). In addition, the effect of KTI mutation on lunasin stability against PPH was determined. LEP were produced by calcium and pH precipitation methods of 30% aqueous ethanol extract from defatted soybean flour. LEP, lunasin-enriched commercially available products and KTI control and mutant flours underwent PPH and samples were taken after pepsin and pepsin-pancreatin hydrolysis. The concentrations of BBI, KTI, and lunasin all decreased after hydrolysis, but they had varying results. BBI concentration ranged from 167.5 to 655.8μg/g pre-hydrolysis and 171.5 to 250.1μg/g after hydrolysis. KTI concentrations ranged from 0.3 to 122.3μg/g pre-hydrolysis and 9.0 to 18.7μg/g after hydrolysis. Lunasin concentrations ranged from 8.5 to 71.0μg/g pre-hydrolysis and 4.0 to 13.2μg/g after hydrolysis. In all products tested, lunasin concentration after PPH significantly correlated with BBI and KTI concentrations. Mutation in two KTI isoforms led to a lower concentration of lunasin after PPH. This is the first report on the potential role of KTI in lunasin stability against PPH and must be considered in designing lunasin-enriched products that could potentially survive digestion after oral ingestion. Copyright © 2016 Elsevier Ltd. All rights reserved.

Unexpected Hydrolytic Instability of N-Acylated Amino Acid Amides and Peptides

PubMed Central

2015-01-01

Remote amide bonds in simple N-acyl amino acid amide or peptide derivatives 1 can be surprisingly unstable hydrolytically, affording, in solution, variable amounts of 3 under mild acidic conditions, such as trifluoroacetic acid/water mixtures at room temperature. This observation has important implications for the synthesis of this class of compounds, which includes N-terminal-acylated peptides. We describe the factors contributing to this instability and how to predict and control it. The instability is a function of the remote acyl group, R2CO, four bonds away from the site of hydrolysis. Electron-rich acyl R2 groups accelerate this reaction. In the case of acyl groups derived from substituted aromatic carboxylic acids, the acceleration is predictable from the substituent’s Hammett σ value. N-Acyl dipeptides are also hydrolyzed under typical cleavage conditions. This suggests that unwanted peptide truncation may occur during synthesis or prolonged standing in solution when dipeptides or longer peptides are acylated on the N-terminus with electron-rich aromatic groups. When amide hydrolysis is an undesired secondary reaction, as can be the case in the trifluoroacetic acid-catalyzed cleavage of amino acid amide or peptide derivatives 1 from solid-phase resins, conditions are provided to minimize that hydrolysis. PMID:24617596

A SENSITIVE FLUORESCENCE-BASED ASSAY FOR MONITORING GM2 GANGLIOSIDE HYDROLYSIS IN LIVE PATIENT CELLS AND THEIR LYSATES

PubMed Central

Tropak, Michael B.; Bukovac, Scott W.; Rigat, Brigitte A.; Yonekawa, Sayuri; Wakarchuk, Warren; Mahuran, Don J.

2010-01-01

Enzyme enhancement therapy, utilizing small molecules as pharmacological chaperones, is anattractive approach for the treatment of lysosomal storage diseases that are associated with protein misfolding. However, pharmacological chaperones are alsoinhibitors of their target enzyme. Thus, a major concern with this approach is that, despite enhancing protein folding within, and intracellular transport of the functional mutant enzyme out of the endoplasmic reticulum, the chaperone will continue to inhibit the enzyme in the lysosome, preventing substrate clearance. Herewe demonstrate that the in vitro hydrolysis of a fluorescent derivative of lyso-GM2 ganglioside, like natural GM2 ganglioside, is specifically carried out by the β-hexosaminidase A isozyme, requires the GM2 activator protein as a co-factor, increases when the derivative is incorporated into anionic liposomes and follows similar Michaelis-Menten kinetics. This substrate can also be used to differentiate between lysates from normal and GM2 activator-deficient cells. When added to the growth medium of cells, the substrate is internalized and primarily incorporated into lysosomes. Utilizing adult Tay-Sachs fibroblasts that have been pre-treated with the pharmacological chaperone Pyrimethamine and subsequently loaded with this substrate, we demonstrate an increase in both the levels of mutant β-hexosaminidase A and substrate-hydrolysis as compared to mock treated cells. PMID:19917668

A sensitive fluorescence-based assay for monitoring GM2 ganglioside hydrolysis in live patient cells and their lysates.

PubMed

Tropak, Michael B; Bukovac, Scott W; Rigat, Brigitte A; Yonekawa, Sayuri; Wakarchuk, Warren; Mahuran, Don J

2010-03-01

Enzyme enhancement therapy, utilizing small molecules as pharmacological chaperones, is an attractive approach for the treatment of lysosomal storage diseases that are associated with protein misfolding. However, pharmacological chaperones are also inhibitors of their target enzyme. Thus, a major concern with this approach is that, despite enhancing protein folding within, and intracellular transport of the functional mutant enzyme out of the endoplasmic reticulum, the chaperone will continue to inhibit the enzyme in the lysosome, preventing substrate clearance. Here we demonstrate that the in vitro hydrolysis of a fluorescent derivative of lyso-GM2 ganglioside, like natural GM2 ganglioside, is specifically carried out by the beta-hexosaminidase A isozyme, requires the GM2 activator protein as a co-factor, increases when the derivative is incorporated into anionic liposomes and follows similar Michaelis-Menten kinetics. This substrate can also be used to differentiate between lysates from normal and GM2 activator-deficient cells. When added to the growth medium of cells, the substrate is internalized and primarily incorporated into lysosomes. Utilizing adult Tay-Sachs fibroblasts that have been pre-treated with the pharmacological chaperone Pyrimethamine and subsequently loaded with this substrate, we demonstrate an increase in both the levels of mutant beta-hexosaminidase A and substrate-hydrolysis as compared to mock-treated cells.

A Factorial Analysis Study on Enzymatic Hydrolysis of Fiber Pressed Oil Palm Frond for Bioethanol Production

NASA Astrophysics Data System (ADS)

Hashim, F. S.; Yussof, H. W.; Zahari, M. A. K. M.; Illias, R. M.; Rahman, R. A.

2016-03-01

Different technologies have been developed to for the conversion of lignocellulosic biomass to suitable fermentation substrates for bioethanol production. The enzymatic conversion of cellulose seems to be the most promising technology as it is highly specific and does not produce substantial amounts of unwanted byproducts. The effects of agitation speed, enzyme loading, temperature, pH and reaction time on the conversion of glucose from fiber pressed oil palm frond (FPOPF) for bioethanol production were screened by statistical analysis using response surface methodology (RSM). A half fraction two-level factorial analysis with five factors was selected for the experimental design to determine the best enzymatic conditions that produce maximum amount of glucose. FPOPF was pre-treated with alkaline prior to enzymatic hydrolysis. The enzymatic hydrolysis was performed using a commercial enzyme Cellic CTec2. From this study, the highest yield of glucose concentration was 9.736 g/L at 72 hours reaction time at 35 °C, pH 5.6, and 1.5% (w/v) of enzyme loading. The model obtained was significant with p-value <0.0001. It is suggested that this model had a maximum point which is likely to be the optimum point and possible for the optimization process.

Histopathological and Reproductive Evaluation in Male Rats Fed Jatropha curcas Seed Cake with or without Alkaline Hydrolysis and Subjected to Heat Treatment.

PubMed

Teixeira Sousa Moura, Laiane; Palomaris Mariano Souza, Domenica; Mendonça, Simone; de Aquino Ribeiro, José Antônio; Fernandes Sousa, Luciano; Tony Ramos, Adriano; Maiorka, Paulo César; de Araújo, Vera Lúcia; Mayumi Maruo, Viviane

2017-01-01

Jatropha curcas cake, a by-product of biodiesel production, is rich in protein and has potential to be used in livestock feed; however, the presence of antinutritional factors and phorbol esters limits its use. Thus, this study investigated toxicological and reproductive effects in male Wistar rats after subchronic exposure to J. curcas cake subjected to detoxification procedures. Rats were divided into seven groups ( n = 10) and treated for 60 days. The control group received commercial feed, while experimental groups received a diet containing 5% J . curcas cake nonhydrolyzed or hydrolyzed with 5 M NaOH. The cakes were unwashed or washed with ethanol or water and were autoclaved at 121°C for 30 minutes. Alkaline hydrolysis combined with ethanol washing decreased the phorbol ester concentration in the cake by 98%. Histopathological findings included diffuse degeneration of the liver and edema around the pulmonary vessels in the nonhydrolyzed groups. In addition, nontreated females mated with males of nonhydrolyzed unwashed group showed a decreased number of live fetuses and an increased placental weight. There were no signs of toxicity in rats given hydrolyzed cakes washed and unwashed, indicating that alkaline hydrolysis associated with heat treatment is an efficient method for detoxification of the J. curcas cake.

Histopathological and Reproductive Evaluation in Male Rats Fed Jatropha curcas Seed Cake with or without Alkaline Hydrolysis and Subjected to Heat Treatment

PubMed Central

Palomaris Mariano Souza, Domenica; Mendonça, Simone; de Aquino Ribeiro, José Antônio; Fernandes Sousa, Luciano; Maiorka, Paulo César; Mayumi Maruo, Viviane

2017-01-01

Jatropha curcas cake, a by-product of biodiesel production, is rich in protein and has potential to be used in livestock feed; however, the presence of antinutritional factors and phorbol esters limits its use. Thus, this study investigated toxicological and reproductive effects in male Wistar rats after subchronic exposure to J. curcas cake subjected to detoxification procedures. Rats were divided into seven groups (n = 10) and treated for 60 days. The control group received commercial feed, while experimental groups received a diet containing 5% J. curcas cake nonhydrolyzed or hydrolyzed with 5 M NaOH. The cakes were unwashed or washed with ethanol or water and were autoclaved at 121°C for 30 minutes. Alkaline hydrolysis combined with ethanol washing decreased the phorbol ester concentration in the cake by 98%. Histopathological findings included diffuse degeneration of the liver and edema around the pulmonary vessels in the nonhydrolyzed groups. In addition, nontreated females mated with males of nonhydrolyzed unwashed group showed a decreased number of live fetuses and an increased placental weight. There were no signs of toxicity in rats given hydrolyzed cakes washed and unwashed, indicating that alkaline hydrolysis associated with heat treatment is an efficient method for detoxification of the J. curcas cake. PMID:28620618

Cytidine 5′-Diphosphocholine (CDP-Choline) in Stroke and Other CNS Disorders

PubMed Central

Adibhatla, Rao Muralikrishna; Hatcher, J. F.

2007-01-01

Brain phosphatidylcholine (PC) levels are regulated by a balance between synthesis and hydrolysis. Pro-inflammatory cytokines such as tumor necrosis factor-α (TNF-α) and interleukin-1 (IL-1α/β) activate phospholipase A2 (PLA2) and PC-phospholipase C (PC-PLC) to hydrolyze PC. PC hydrolysis by PLA2 releases free fatty acids including arachidonic acid, and lyso-PC, an inhibitor of CTP-phosphocholine cytidylyltransferase (CCT). Arachidonic acid metabolism by cyclooxygenases/lipoxygenases is a significant source of reactive oxygen species. CDP-choline might increase the PC levels by attenuating PLA2 stimulation and loss of CCT activity. TNF-α also stimulates proteolysis of CCT. TNF-α and IL-1β are induced in brain ischemia and may disrupt PC homeostasis by increasing its hydrolysis (increase PLA2 and PC-PLC activities) and inhibiting its synthesis (decrease CCT activity). The beneficial effects of CDP-choline may result by counteracting TNF-α and/or IL-1 mediated events, integrating cytokine biology and lipid metabolism. Re-evaluation of CDP-choline phase III stroke clinical trial data is encouraging and future trails are warranted. CDP-choline is non-xenobiotic, safe, well tolerated, and can be considered as one of the agents in multi-drug treatment of stroke. PMID:15756928

Aminoacyl-nucleotide reactions - Studies related to the origin of the genetic code and protein synthesis

NASA Technical Reports Server (NTRS)

Mullins, D. W., Jr.; Senaratne, N.; Lacey, J. C., Jr.

1984-01-01

In the present paper, a report is presented on the effect of pH and carbonate on the hydrolysis rate constants of N-blocked and free aminoacyl adenylate anhydrides. Whereas the hydrolysis of free aminoacyl adenylates seems principally catalyzed by OH(-), the hydrolysis of the N-blocked species is also catalyzed by H(+), giving this compound a U-shaped hydrolysis vs. pH curve. Furthermore, at pH's less than 8, carbonate has an extreme catalytic effect on the hydrolysis of free aminoacyl-AMP anhydride, but essentially no effect on the hydrolysis of N-blocked aminoacyl-AMP anhydride. Furthermore, the N-blocked aminoacyl-AMP anhydride is a very efficient generator of peptides using free glycine as acceptor. The possible significance of the observations to prebiological peptide synthesis is discussed.

Where does hydrolysis of nandrolone decanoate occur in the human body after release from an oil depot?

PubMed

Kalicharan, R W; Bout, M R; Oussoren, C; Vromans, H

2016-12-30

Long-term therapy of nandrolone (N) is recommended to increase mineral density and muscle strength. Using a parenteral sustained release drug formulation with nandrolone decanoate (ND), therapeutic N levels can be achieved and maintained. Until now, it is unknown if hydrolysis of ND into N occurs in tissue at the injection site or after systemic absorption. Therefore, hydrolysis studies were conducted to investigate the location and rate of ND hydrolysis after its release from the oil depot. ND hydrolysis was studied in porcine tissues, to mimic the human muscular and subcutaneous tissues. Additionally, the ND hydrolysis was studied in human whole blood, plasma and serum at a concentration range of 23.3-233.3μM. ND hydrolysis only occurred in human whole blood. The hydrolysis did not start immediately, but after a lag time. The mean lag time for all studied concentrations was 34.9±2.5min. Because of a slow penetration into tissue, hydrolysis of ND is found to be very low in surrounding tissue. Therefore the local generation of the active compound is clinically irrelevant. It is argued that after injection of the oil depot, ND molecules will be transported via the lymphatic system towards lymph nodes. From here, it will enter the central circulation and within half an hour it will hydrolyse to the active N compound. Copyright © 2016 Elsevier B.V. All rights reserved.

Hydrolysis of amphenicol and macrolide antibiotics: Chloramphenicol, florfenicol, spiramycin, and tylosin.

PubMed

Mitchell, Shannon M; Ullman, Jeffrey L; Teel, Amy L; Watts, Richard J

2015-09-01

Antibiotics that enter the environment can present human and ecological health risks. An understanding of antibiotic hydrolysis rates is important for predicting their environmental persistence as biologically active contaminants. In this study, hydrolysis rates and Arrhenius constants were determined as a function of pH and temperature for two amphenicol (chloramphenicol and florfenicol) and two macrolide (spiramycin and tylosin) antibiotics. Antibiotic hydrolysis rates in pH 4-9 buffer solutions at 25°C, 50°C, and 60°C were quantified, and degradation products were characterized. All of the antibiotics tested remained stable and exhibited no observable hydrolysis under ambient conditions typical of aquatic ecosystems. Acid- and base-catalyzed hydrolysis occurred at elevated temperatures (50-60°C), and hydrolysis rates increased considerably below pH 5 and above pH 8. Hydrolysis rates also increased approximately 1.5- to 2.9-fold for each 10°C increase in temperature. Based on the degradation product masses found, the functional groups that underwent hydrolysis were alkyl fluoride, amide, and cyclic ester (lactone) moieties; some of the resultant degradation products may remain bioactive, but to a lesser extent than the parent compounds. The results of this research demonstrate that amphenicol and macrolide antibiotics persist in aquatic systems under ambient temperature and pH conditions typical of natural waters. Thus, these antibiotics may present a risk in aquatic ecosystems depending on the concentration present. Copyright © 2015. Published by Elsevier Ltd.

[Structural characterization of Astragalus polysaccharides using partial acid hydrolysis-hydrophilic interaction liquid chromatography-mass spectrometry].

PubMed

Liang, Tu; Fu, Qing; Xin, Huaxia; Li, Fangbing; Jin, Yu; Liang, Xinmiao

2014-12-01

Water-soluble polysaccharides from traditional Chinese medicine (TCM) have properties of broad-spectrum treatment and low toxicity, making them as important components in natural medicines and health products. In order to solve the problem of polysaccharides characterization caused by their complex structures, a "bottom-up" approach was developed to complete the characterization of polysaccharides from Astragalus. Firstly, Astragalus pieces were extracted with hot water and then were precipitated by ethanol to obtain Astragalus polysaccharides. Secondly, a partial acid hydrolysis method was carried out and the effects of time, acid concentration and temperature on hydrolysis were investigated. The degree of hydrolysis increased along with the increase of hydrolysis time and acid concentration. The temperature played a great role in the hydrolysis process. No hydrolysis of the polysaccharides occurred at low temperature, while the polysaccharides were almost hydrolyzed to monosaccharide at high temperature. Under the optimum hydrolysis conditions (4 h, 1.5 mol/L trifluoroacetic acid, and 80 °C), Astragalus polysaccharides were hydrolyzed to characteristic oligosaccharide fragments. At last, a hydrophilic liquid chromatography-mass spectrometry method was used for the separation and structural characterization of the polysaccharide hydrolysates. The results showed that the resulting polysaccharides were mainly 1--> 4 linear glucan, and gluco-oligosaccharides with the degrees of polymerization (DP) of 4 - 11 were obtained after partial acid hydrolysis. The significance of this study is that it is the guidance for the characterization of other TCM polysaccharides.

Electrospray ionization mass spectrometry for the hydrolysis complexes of cisplatin: implications for the hydrolysis process of platinum complexes.

PubMed

Feifan, Xie; Pieter, Colin; Jan, Van Bocxlaer

2017-07-01

Non-enzyme-dependent hydrolysis of the drug cisplatin is important for its mode of action and toxicity. However, up until today, the hydrolysis process of cisplatin is still not completely understood. In the present study, the hydrolysis of cisplatin in an aqueous solution was systematically investigated by using electrospray ionization mass spectrometry coupled to liquid chromatography. A variety of previously unreported hydrolysis complexes corresponding to monomeric, dimeric and trimeric species were detected and identified. The characteristics of the Pt-containing complexes were investigated by using collision-induced dissociation (CID). The hydrolysis complexes demonstrate distinctive and correlative CID characteristics, which provides tools for an informative identification. The most frequently observed dissociation mechanism was sequential loss of NH 3 , H 2 O and HCl. Loss of the Pt atom was observed as the final step during the CID process. The formation mechanisms of the observed complexes were explored and experimentally examined. The strongly bound dimeric species, which existed in solution, are assumed to be formed from the clustering of the parent compound and its monohydrated or dihydrated complexes. The role of the electrospray process in the formation of some of the observed ions was also evaluated, and the electrospray ionization-related cold clusters were identified. The previously reported hydrolysis equilibria were tested and subsequently refined via a hydrolysis study resulting in a renewed mechanistic equilibrium system of cisplatin as proposed from our results. Copyright © 2017 John Wiley & Sons, Ltd. Copyright © 2017 John Wiley & Sons, Ltd.

Acid hydrolysis of cellulose to yield glucose

DOEpatents

Tsao, George T.; Ladisch, Michael R.; Bose, Arindam

1979-01-01

A process to yield glucose from cellulose through acid hydrolysis. Cellulose is recovered from cellulosic materials, preferably by pretreating the cellulosic materials by dissolving the cellulosic materials in Cadoxen or a chelating metal caustic swelling solvent and then precipitating the cellulose therefrom. Hydrolysis is accomplished using an acid, preferably dilute sulfuric acid, and the glucose is yielded substantially without side products. Lignin may be removed either before or after hydrolysis.

Quantification of quercetin glycosides in 6 onion cultivars and comparisons of hydrolysis-HPLC and spectrophotometric methods in measuring total quercetin concentrations.

PubMed

Yoo, Kil Sun; Lee, Eun Jin; Patil, Bhimanagouda S

2010-03-01

This study was performed to purify and quantify quercetin glycosides (QG) and aglycone (free) quercetin (Q) in 6 selected onion cultivars and to compare analytical approaches based on high-performance liquid chromatography (HPLC) and spectrophotometry for the quantification of total quercetin (TQ) concentrations. Individual mono- and di-glycoside Q compounds were purified using a semipreparative HPLC and identified by comparing spectral data and by confirming corresponding peaks of QG and Q after incomplete enzyme-hydrolysis. Purified QG were quantified as Q by enzyme-hydrolysis/HPLC. TQ concentrations obtained from 20 onion bulbs with enzyme-hydrolysis/HPLC, no-hydrolysis/HPLC, and a spectrophotometric method without prior hydrolysis were significantly correlated (r(2)= 0.99) and were about 15% higher, identical, or 10% less than those concentrations by a standard acid-hydrolysis/HPLC method, respectively. During enzyme-hydrolysis of onion extracts, progressive reduction of the QG and formation of the corresponding mono-glycosides and Q were monitored using an analytical HPLC. TQ ranged from 83 to 330 microg/g F.W. in 6 selected cultivars of long-day or short-day onions. Q3,4'G and Q4'G were the 2 major compounds and comprised approximately between 94% and 97% of TQ in onions.

Ethanol and anaerobic conditions reversibly inhibit commercial cellulase activity in thermophilic simultaneous saccharification and fermentation (tSSF)

PubMed Central

2012-01-01

Background A previously developed mathematical model of low solids thermophilic simultaneous saccharification and fermentation (tSSF) with Avicel was unable to predict performance at high solids using a commercial cellulase preparation (Spezyme CP) and the high ethanol yield Thermoanaerobacterium saccharolyticum strain ALK2. The observed hydrolysis proceeded more slowly than predicted at solids concentrations greater than 50 g/L Avicel. Factors responsible for this inaccuracy were investigated in this study. Results Ethanol dramatically reduced cellulase activity in tSSF. At an Avicel concentration of 20 g/L, the addition of ethanol decreased conversion at 96 hours, from 75% in the absence of added ethanol down to 32% with the addition of 34 g/L initial ethanol. This decrease is much greater than expected based on hydrolysis inhibition results in the absence of a fermenting organism. The enhanced effects of ethanol were attributed to the reduced, anaerobic conditions of tSSF, which were shown to inhibit cellulase activity relative to hydrolysis under aerobic conditions. Cellulose hydrolysis in anaerobic conditions was roughly 30% slower than in the presence of air. However, this anaerobic inhibition was reversed by exposing the cellulase enzymes to air. Conclusion This work demonstrates a previously unrecognized incompatibility of enzymes secreted by an aerobic fungus with the fermentation conditions of an anaerobic bacterium and suggests that enzymes better suited to industrially relevant fermentation conditions would be valuable. The effects observed may be due to inactivation or starvation of oxygen dependent GH61 activity, and manipulation or replacement of this activity may provide an opportunity to improve biomass to fuel process efficiency. PMID:22703989

Elucidation of Factors Effecting Enzymatic Saccharification using Transgenic Hardwoods

NASA Astrophysics Data System (ADS)

Min, Douyong

Three groups of transgenic wood samples were used as starting materials to elucidate the recalcitrance of enzymatic saccharification with/without pretreatments. The first group of transgenic wood samples is low lignin P. trichocarpa. The second group is low xylan P. trichocarpa. The third one is 12 hybrid poplars which have different levels of S/V ratio and lignin content. Four pretreatments were carried out in this research including dilute sulfuric acid, green liquor, auto hydrolysis and ozone delignification. The behavior among pretreatments as a function of removal of lignin appears to be different. Lignin is the major factor of recalcitrance of the lignocellulosic material to ethanol conversion process. Xylan also plays key role in this process. In addition, the crude milled wood lignin was isolated from these three groups of transgenic samples. Lignin carbohydrate complexes was characterized by 1H-13C HMQC and 13C NMR. Thus the effect of LCCs on enzymatic saccharification was elucidated. High S/V ratio propels the lignin removal during pretreatments however; high S/V ratio retards the enzymatic saccharification on the lignocellulosic material without pretreatments. The level of LCCs linkages accounts for additional recalcitrance of the lignocellulosic material to ethanol conversion process. The amount of LCCs linkages is affected by xylan content, lignin content and S/V ratio.

Adsorption of monocomponent enzymes in enzyme mixture analyzed quantitatively during hydrolysis of lignocellulose substrates.

PubMed

Várnai, Anikó; Viikari, Liisa; Marjamaa, Kaisa; Siika-aho, Matti

2011-01-01

The adsorption of purified Trichoderma reesei cellulases (TrCel7A, TrCel6A and TrCel5A) and xylanase TrXyn11 and Aspergillus niger β-glucosidase AnCel3A was studied in enzyme mixture during hydrolysis of two pretreated lignocellulosic materials, steam pretreated and catalytically delignified spruce, along with microcrystalline cellulose (Avicel). The enzyme mixture was compiled to resemble the composition of commercial cellulase preparations. The hydrolysis was carried out at 35 °C to mimic the temperature of the simultaneous saccharification and fermentation (SSF). Enzyme adsorption was followed by analyzing the activity and the protein amount of the individual free enzymes in the hydrolysis supernatant. Most enzymes adsorbed quickly at early stages of the hydrolysis and remained bound throughout the hydrolysis, although the conversion reached was fairly high. Only with the catalytically oxidized spruce samples, the bound enzymes started to be released as the hydrolysis degree reached 80%. The results based on enzyme activities and protein assay were in good accordance. Copyright © 2010 Elsevier Ltd. All rights reserved.

Effect of various pH values, ionic strength, and temperature on papain hydrolysis of salivary film.

PubMed

Yao, Jiang-Wu; Xiao, Yin; Lin, Feng

2012-04-01

Stimulated human whole saliva (WS) was used to study the dynamics of papain hydrolysis at defined pH, ionic strength, and temperature with the view of reducing an acquired pellicle. A quartz crystal microbalance with dissipation (QCM-D) was used to monitor the changes in frequency caused by enzyme hydrolysis of WS films, and the hydrolytic parameters were calculated using an empirical model. The morphological and conformational changes of the salivary films before and after enzymatic hydrolysis were characterized by atomic force microscopy (AFM) imaging and grazing-angle Fourier transform infrared (GA-FTIR ) spectra, respectively. The characteristics of papain hydrolysis of WS films were pH-, ionic strength-, and temperature-dependent. The WS films were partially removed by the action of papain, resulting in thinner and smoother surfaces. The infrared data suggested that hydrolysis-induced deformation did not occur on the remnants of salivary films. The processes of papain hydrolysis of WS films can be controlled by properly regulating pH, ionic strength, and temperature. © 2012 Eur J Oral Sci.

Continuous-flow electro-assisted acid hydrolysis of granular potato starch via inductive methodology.

PubMed

Li, Dandan; Yang, Na; Jin, Yamei; Guo, Lunan; Zhou, Yuyi; Xie, Zhengjun; Jin, Zhengyu; Xu, Xueming

2017-08-15

The induced electric field assisted hydrochloric acid (IEF-HCl) hydrolysis of potato starch was investigated in a fluidic system. The impact of various reaction parameters on the hydrolysis rate, including reactor number (1-4), salt type (KCl, MgCl 2 , FeCl 3 ), salt concentration (3-12%), temperature (40-55°C), and hydrolysis time (0-60h), were comprehensively assessed. Under optimal conditions, the maximum reducing sugar content in the hydrolysates was 10.59g/L. X-ray diffraction suggested that the crystallinity of IEF-HCl-modified starches increased with the intensification of hydrolysis but was lower than that of native starch. Scanning electron microscopy indicated that the surface and interior regions of starch granules were disrupted by the hydrolysis. The solubility of IEF-HCl-modified starches increased compared to native starch while their swelling power decreased, contributing to a decline in paste viscosity. These results suggest that IEF is a notable potential electrotechnology to conventional hydrolysis under mild conditions without any electrode touching the subject. Copyright © 2017 Elsevier Ltd. All rights reserved.

Extraction of cellulose nano-crystals from old corrugated container fiber using phosphoric acid and enzymatic hydrolysis followed by sonication.

PubMed

Tang, Yanjun; Shen, Xiaochuang; Zhang, Junhua; Guo, Daliang; Kong, Fangong; Zhang, Nan

2015-07-10

Due to its amazing physicochemical properties and high environmental compatibility, cellulose nano-crystals (CNC) hold great promise for serving as a strategic platform for sustainable development. Now, there has been growing interest in the development of processes using waste or residual biomass as CNC source for addressing economic and environmental concerns. In the present work, a combined process involving phosphoric acid hydrolysis, enzymatic hydrolysis and sonication was proposed aiming to efficiently exact CNC from low-cost old corrugated container (OCC) pulp fiber. The effect of enzymatic hydrolysis on the yield and microstructure of resulting CNC was highlighted. Results showed that the enzymatic hydrolysis was effective in enhancing CNC yield after phosphoric acid hydrolysis. CNC was obtained with a yield of 23.98 wt% via the combined process with phosphoric acid concentration of 60 wt%, cellulase dosage of 2 mL (84 EGU) per 2g fiber and sonication intensity of 200 W. Moreover, the presence of enzymatic hydrolysis imparted the obtained CNC with improved dispersion, increased crystallinity and thermal stability. Copyright © 2015 Elsevier Ltd. All rights reserved.

In silico kinetics of alkaline hydrolysis of 1,3,5-trinitro-1,3,5-triazinane (RDX): M06-2X investigation.

PubMed

Sviatenko, L K; Gorb, L; Leszczynska, D; Okovytyy, S I; Shukla, M K; Leszczynski, J

2017-03-22

Alkaline hydrolysis of RDX (hexahydro-1,3,5-trinitro-1,3,5-triazine), as one of the most promising methods for nitrocompound remediation, was investigated computationally at the PCM(Pauling)/M06-2X/6-311++G(d,p) level of theory. Computational simulation shows that RDX hydrolysis is a highly exothermic multistep process involving initial deprotonation and nitrite elimination, cycle cleavage, further transformation of cycle-opened intermediates to end products caused by a series of C-N bond ruptures, hydroxide attachments, and proton transfers. Computationally predicted products of RDX hydrolysis such as nitrite, nitrous oxide, formaldehyde, formate, and ammonia correspond to experimentally observed ones. Accounting of specific hydration of hydroxide is critical to create an accurate kinetic model for alkaline hydrolysis. Simulated kinetics of the hydrolysis are in good agreement with available experimental data. A period of one month is necessary for 99% RDX decomposition at pH 10. Computations predict significant increases of the reaction rate of hydrolysis at pH 11, pH 12, and pH 13.

Extracellular ATP is a mitogen for 3T3, 3T6, and A431 cells and acts synergistically with other growth factors.

PubMed Central

Huang, N; Wang, D J; Heppel, L A

1989-01-01

Extracellular ATP in concentrations of 5-50 microM displayed very little mitogenic activity by itself but it caused synergistic stimulation of [3H]thymidine incorporation in the presence of phorbol 12-tetradecanoate 13-acetate, epidermal growth factor, platelet-derived growth factor, insulin, adenosine, or 5'-(N-ethyl)carboxamidoadenosine. Cultures of Swiss 3T3, Swiss 3T6, A431, DDT1-MF2, and HFF cells were used. The percent of cell nuclei labeled with [3H]thymidine and cell number were also increased. ADP was equally mitogenic, while UTP and ITP were much less active. The effect of ATP was not due to hydrolysis by ectoenzymes to form adenosine, a known growth factor. Thus, the nonhydrolyzable analogue adenosine 5'-[beta, gamma-imido]triphosphate was mitogenic. In addition, it was found that ATP showed synergism in 3T6 and 3T3 cells when present for only the first hour of an incorporation assay, during which time no significant hydrolysis occurred. Furthermore, prolonged preincubation of cells with ATP reduced the mitogenic response to ATP but not to adenosine; preincubation with adenosine or N6-(R-phenylisopropyl)adenosine had the reverse effect. Finally, the effect of adenosine, but not of ATP, was inhibited by aminophylline. We conclude that extracellular ATP is a mitogen that interacts with P2 purinoceptors on the plasma membrane. PMID:2813367

Type-7 metabotropic glutamate receptors negatively regulate α1-adrenergic receptor signalling.

PubMed

Iacovelli, Luisa; Di Menna, Luisa; Peterlik, Daniel; Stangl, Christina; Orlando, Rosamaria; Molinaro, Gemma; De Blasi, Antonio; Bruno, Valeria; Battaglia, Giuseppe; Flor, Peter J; Uschold-Schmidt, Nicole; Nicoletti, Ferdinando

2017-02-01

We studied the interaction between mGlu7 and α 1 -adrenergic receptors in heterologous expression systems, brain slices, and living animals. L-2-Amino-4-phosphonobutanoate (L-AP4), and l-serine-O-phosphate (L-SOP), which activate group III mGlu receptors, restrained the stimulation of polyphosphoinositide (PI) hydrolysis induced by the α 1 -adrenergic receptor agonist, phenylephrine, in HEK 293 cells co-expressing α 1 -adrenergic and mGlu7 receptors. The inibitory action of L-AP4 was abrogated by (i) the mGlu7 receptor antagonist, XAP044; (ii) the C-terminal portion of type-2 G protein coupled receptor kinase; and (iii) the MAP kinase inhibitors, UO126 and PD98059. This suggests that the functional interaction between mGlu7 and α 1 -adrenergic receptors was mediated by the βγ-subunits of the G i protein and required the activation of the MAP kinase pathway. Remarkably, activation of neither mGlu2 nor mGlu4 receptors reduced α 1 -adrenergic receptor-mediated PI hydrolysis. In mouse cortical slices, both L-AP4 and L-SOP were able to attenuate norepinephrine- and phenylephrine-stimulated PI hydrolysis at concentrations consistent with the activation of mGlu7 receptors. L-AP4 failed to affect norepinephrine-stimulated PI hydrolysis in cortical slices from mGlu7 -/- mice, but retained its inhibitory activity in slices from mGlu4 -/- mice. At behavioural level, i.c.v. injection of phenylephrine produced antidepressant-like effects in the forced swim test. The action of phenylephrine was attenuated by L-SOP, which was inactive per se. Finally, both phenylephrine and L-SOP increased corticosterone levels in mice, but the increase was halved when the two drugs were administered in combination. Our data demonstrate that α 1 -adrenergic and mGlu7 receptors functionally interact and suggest that this interaction might be targeted in the treatment of stress-related disorders. Copyright © 2016 Elsevier Ltd. All rights reserved.

Impact of cultivar selection and process optimization on ethanol yield from different varieties of sugarcane

PubMed Central

2014-01-01

Background The development of ‘energycane’ varieties of sugarcane is underway, targeting the use of both sugar juice and bagasse for ethanol production. The current study evaluated a selection of such ‘energycane’ cultivars for the combined ethanol yields from juice and bagasse, by optimization of dilute acid pretreatment optimization of bagasse for sugar yields. Method A central composite design under response surface methodology was used to investigate the effects of dilute acid pretreatment parameters followed by enzymatic hydrolysis on the combined sugar yield of bagasse samples. The pressed slurry generated from optimum pretreatment conditions (maximum combined sugar yield) was used as the substrate during batch and fed-batch simultaneous saccharification and fermentation (SSF) processes at different solid loadings and enzyme dosages, aiming to reach an ethanol concentration of at least 40 g/L. Results Significant variations were observed in sugar yields (xylose, glucose and combined sugar yield) from pretreatment-hydrolysis of bagasse from different cultivars of sugarcane. Up to 33% difference in combined sugar yield between best performing varieties and industrial bagasse was observed at optimal pretreatment-hydrolysis conditions. Significant improvement in overall ethanol yield after SSF of the pretreated bagasse was also observed from the best performing varieties (84.5 to 85.6%) compared to industrial bagasse (74.5%). The ethanol concentration showed inverse correlation with lignin content and the ratio of xylose to arabinose, but it showed positive correlation with glucose yield from pretreatment-hydrolysis. The overall assessment of the cultivars showed greater improvement in the final ethanol concentration (26.9 to 33.9%) and combined ethanol yields per hectare (83 to 94%) for the best performing varieties with respect to industrial sugarcane. Conclusions These results suggest that the selection of sugarcane variety to optimize ethanol production from bagasse can be achieved without adversely affecting juice ethanol and cane yield, thus maintaining first generation ethanol production levels while maximizing second generation ethanol production. PMID:24725458

Continuous measurement of galactolipid hydrolysis by pancreatic lipolytic enzymes using the pH-stat technique and a medium chain monogalactosyl diglyceride as substrate.

PubMed

Amara, Sawsan; Lafont, Dominique; Fiorentino, Brice; Boullanger, Paul; Carrière, Frédéric; De Caro, Alain

2009-10-01

Galactolipids are the main lipids from plants and galactolipases play a major role in their metabolism. These enzymes were however poorly studied so far and only few assays have been developed. A specific and continuous galactolipase assay using synthetic medium chain monogalactosyl diacylglycerol (MGDG) as substrate was developed using the pH-stat technique and recombinant human (rHPLRP2) and guinea pig (rGPLRP2) pancreatic lipase-related protein 2 as model enzymes. PLRP2s are the main enzymes involved in the digestion of galactolipids in the gastrointestinal tract. Monogalactosyl di-octanoylglycerol was mixed with bile salt solutions by sonication to form a micellar substrate before launching the assay. The nature of the bile salt and the bile salt to MGDG ratio were found to significantly affect the rate of MGDG hydrolysis by rHPLRP2 and rGPLRP2. The maximum galactolipase activity of both enzymes was recorded with sodium deoxycholate (NaDC) and at a NaDC to MGDG ratio of 1.33 and at basic pH values (8.0-9.0). The maximum rates of hydrolysis were obtained using a MGDG concentration of 10(-2) M and calcium chloride was found to be not necessary to obtain the maximum of activity. Under these conditions, the maximum turnovers of rGPLRP2 and rHPLRP2 on mixed NaDC/MGDG micelles were found to be 8000+/-500 and 2800+/-60 micromol/min/mg (U/mg), respectively. These activities are in the same order of magnitude as the activities on triglycerides of lipases and they are the highest specific activities ever reported for galactolipases. For the sake of comparison, the hydrolysis of mixed bile salt/MGDG micelles was also tested using other pancreatic lipolytic enzymes and only native and recombinant human carboxyl ester hydrolase were found to display significant but lower activities (240+/-17 and 432+/-62 U/mg, respectively) on MGDG.

Modeling of autocatalytic hydrolysis of adefovir dipivoxil in solid formulations.

PubMed

Dong, Ying; Zhang, Yan; Xiang, Bingren; Deng, Haishan; Wu, Jingfang

2011-04-01

The stability and hydrolysis kinetics of a phosphate prodrug, adefovir dipivoxil, in solid formulations were studied. The stability relationship between five solid formulations was explored. An autocatalytic mechanism for hydrolysis could be proposed according to the kinetic behavior which fits the Prout-Tompkins model well. For the classical kinetic models could hardly describe and predict the hydrolysis kinetics of adefovir dipivoxil in solid formulations accurately when the temperature is high, a feedforward multilayer perceptron (MLP) neural network was constructed to model the hydrolysis kinetics. The build-in approaches in Weka, such as lazy classifiers and rule-based learners (IBk, KStar, DecisionTable and M5Rules), were used to verify the performance of MLP. The predictability of the models was evaluated by 10-fold cross-validation and an external test set. It reveals that MLP should be of general applicability proposing an alternative efficient way to model and predict autocatalytic hydrolysis kinetics for phosphate prodrugs.

ATP hydrolysis assists phosphate release and promotes reaction ordering in F1-ATPase

PubMed Central

Li, Chun-Biu; Ueno, Hiroshi; Watanabe, Rikiya; Noji, Hiroyuki; Komatsuzaki, Tamiki

2015-01-01

F1-ATPase (F1) is a rotary motor protein that can efficiently convert chemical energy to mechanical work of rotation via fine coordination of its conformational motions and reaction sequences. Compared with reactant binding and product release, the ATP hydrolysis has relatively little contributions to the torque and chemical energy generation. To scrutinize possible roles of ATP hydrolysis, we investigate the detailed statistics of the catalytic dwells from high-speed single wild-type F1 observations. Here we report a small rotation during the catalytic dwell triggered by the ATP hydrolysis that is indiscernible in previous studies. Moreover, we find in freely rotating F1 that ATP hydrolysis is followed by the release of inorganic phosphate with low synthesis rates. Finally, we propose functional roles of the ATP hydrolysis as a key to kinetically unlock the subsequent phosphate release and promote the correct reaction ordering. PMID:26678797

Hydrolysis of dilute acid-pretreated cellulose under mild hydrothermal conditions.

PubMed

Chimentão, R J; Lorente, E; Gispert-Guirado, F; Medina, F; López, F

2014-10-13

The hydrolysis of dilute acid-pretreated cellulose was investigated in a conventional oven and under microwave heating. Two acids--sulfuric and oxalic--were studied. For both hydrothermal conditions (oven and microwave) the resultant total organic carbon (TOC) values obtained by the hydrolysis of the cellulose pretreated with sulfuric acid were higher than those obtained by the hydrolysis of the cellulose pretreated with oxalic acid. However, the dicarboxylic acid exhibited higher hydrolytic efficiency towards glucose. The hydrolysis of cellulose was greatly promoted by microwave heating. The Rietveld method was applied to fit the X-ray patterns of the resultant cellulose after hydrolysis. Oxalic acid preferentially removed the amorphous region of the cellulose and left the crystalline region untouched. On the other hand, sulfuric acid treatment decreased the ordering of the cellulose by partially disrupting its crystalline structure. Copyright © 2014 Elsevier Ltd. All rights reserved.

Process development of starch hydrolysis using mixing characteristics of Taylor vortices.

PubMed

Masuda, Hayato; Horie, Takafumi; Hubacz, Robert; Ohmura, Naoto; Shimoyamada, Makoto

2017-04-01

In food industries, enzymatic starch hydrolysis is an important process that consists of two steps: gelatinization and saccharification. One of the major difficulties in designing the starch hydrolysis process is the sharp change in its rheological properties. In this study, Taylor-Couette flow reactor was applied to continuous starch hydrolysis process. The concentration of reducing sugar produced via enzymatic hydrolysis was evaluated by varying operational variables: rotational speed of the inner cylinder, axial velocity (reaction time), amount of enzyme, and initial starch content in the slurry. When Taylor vortices were formed in the annular space, efficient hydrolysis occurred because Taylor vortices improved the mixing of gelatinized starch with enzyme. Furthermore, a modified inner cylinder was proposed, and its mixing performance was numerically investigated. The modified inner cylinder showed higher potential for enhanced mixing of gelatinized starch and the enzyme than the conventional cylinder.

Optimization of the Hydrolysis of Safflower Oil for the Production of Linoleic Acid, Used as Flavor Precursor.

PubMed

Aziz, Marya; Husson, Florence; Kermasha, Selim

2015-01-01

Commercial lipases, from porcine pancreas (PPL), Candida rugosa (CRL), and Thermomyces lanuginosus (Lipozyme TL IM), were investigated in terms of their efficiency for the hydrolysis of safflower oil (SO) for the liberation of free linoleic acid (LA), used as a flavor precursor. Although PPL, under the optimized conditions, showed a high degree of hydrolysis (91.6%), its low tolerance towards higher substrate concentrations could limit its use for SO hydrolysis. In comparison to the other investigated lipases, Lipozyme TL IM required higher amount of enzyme and an additional 3 h of reaction time to achieve its maximum degree of SO hydrolysis (90.2%). On the basis of the experimental findings, CRL was selected as the most appropriate biocatalyst, with 84.1% degree of hydrolysis. The chromatographic analyses showed that the CRL-hydrolyzed SO is composed mainly of free LA.

DNA-Catalyzed Amide Hydrolysis.

PubMed

Zhou, Cong; Avins, Joshua L; Klauser, Paul C; Brandsen, Benjamin M; Lee, Yujeong; Silverman, Scott K

2016-02-24

DNA catalysts (deoxyribozymes) for a variety of reactions have been identified by in vitro selection. However, for certain reactions this identification has not been achieved. One important example is DNA-catalyzed amide hydrolysis, for which a previous selection experiment instead led to DNA-catalyzed DNA phosphodiester hydrolysis. Subsequent efforts in which the selection strategy deliberately avoided phosphodiester hydrolysis led to DNA-catalyzed ester and aromatic amide hydrolysis, but aliphatic amide hydrolysis has been elusive. In the present study, we show that including modified nucleotides that bear protein-like functional groups (any one of primary amino, carboxyl, or primary hydroxyl) enables identification of amide-hydrolyzing deoxyribozymes. In one case, the same deoxyribozyme sequence without the modifications still retains substantial catalytic activity. Overall, these findings establish the utility of introducing protein-like functional groups into deoxyribozymes for identifying new catalytic function. The results also suggest the longer-term feasibility of deoxyribozymes as artificial proteases.

Organosolv pretreatment by crude glycerol from oleochemicals industry for enzymatic hydrolysis of wheat straw.

PubMed

Sun, Fubao; Chen, Hongzhang

2008-09-01

In order to defray the cost of biodiesel production, the ensuing work was to further investigate utilization of the crude glycerol (CG) from oleochemicals industry in the atmospheric autocatalytic organosolv pretreatment (AAOP) to enhance enzymatic hydrolysis. The AAOP-CG enabled wheat straw to achieve with reasonable enzymatic hydrolysis yields, reaching 75% for the wet substrate and 63% for the dried. Lipophilic compounds from the CG formed pitch deposition on the fiber, which was responsible for low delignification (30%) and also troublesome in practical operation. Pitch deposits itself had no significant role on enzymatic hydrolysis. A striking finding of the lignin recondensation and/or lignin-carbohydrate complex helped explain why dried pretreated wheat straw had a low enzymatic hydrolysis yield. The CG was suitable for the AAOP to enhance enzymatic hydrolysis of lignocellulosic biomass. But it was advisable to remove lipophilic compounds from crude glycerol before utilization.

Deinhibition of cardiac Na/sup +/-K/sup +/-ATPase after exposure to exogenous phospholipase A/sub 2/

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

Colvin, R.A.

1987-01-01

After 2 h of exogenous phospholipase A/sub 2/ (PLA/sub 2/) exposure, membrane phospholipid decreased from 3.22 +/- 0.31 to 1.06 +/- 0.13 ..mu..mol/mg (33% of control). All classes of phospholipid, except sphingomyelin, were hydrolyzed, whereas total cholesterol content was unaffected. Increases in nonesterified fatty acids (NEFA) were reflected primarily in oleic (18:1), linoleic (18:2), and arachidonic (20:4). Na/sup +/-K/sup +/-adenosinetriphosphatase (ATPase) activity was inhibited to 29% of control by 2 h of PLA/sub 2/ treatment, and this inhibition was reversed (albeit, not completely after 5 min of PLA/sub 2/ treatment) by removal of the hydrolysis products with 0.1% bovine serummore » albumin (BSA). In contrast, the apparent binding capacity for (/sup 3/H)ouabain was not affected by PLA/sub 2/ treatment. Unmasking of latent (/sup 3/H)ouabain binding by alamethicin was utilized to estimate changes in the proportion of sealed vesicles present before and after PLA/sub 2/ treatment. PLA/sub 2/ treatment resulted in a time-dependent loss of sealed vesicles that paralleled the time course of phospholipid hydrolysis and was not reversed by washing with BSA. These studies demonstrate that cardiac Na/sup +/-K/sup +/-ATPase activity is inhibited by accumulation of endogenously produced lysophospholipids and NEFA. In contrast, loss of vesicle integrity may result from both accumulation of endogenously produced hydrolysis products and membrane phospholipid depletion.« less

Multi-spectral and thermodynamic analysis of the interaction mechanism between Cu2+ and α-amylase and impact on sludge hydrolysis.

PubMed

Zhou, Ruiqi; Liu, Hong; Hou, Guangying; Ju, Lei; Liu, Chunguang

2017-04-01

An increasing amount of heavy metals (e.g., Cu 2+ ) is being discharged into sewage treatment plants and is accumulating in sludge, which is toxic to the enzyme in sludge or soil when the sludge is used as fertilizer, resulting in unfavorable effect on the biological treatment of sludge and the circulation and conversion of materials in soil. In this research, effect of Cu 2+ on sludge hydrolysis by α-amylase is studied from the respect of concentration and components of soluble organic matter in sludge, using three-dimensional fluorescence spectra. Results show that Cu 2+ exposure not only inhibits the hydrolysis of sludge due to the denaturation of α-amylase but also affects the components of soluble organic matter in sludge. In order to illuminate the interaction mechanism between Cu 2+ and α-amylase (a model of hydrolase in sludge), multi-spectra and isothermal titration microcalorimetry techniques are applied. Results show that the secondary structure of α-amylase is changed as that the α-helical content increases and the structure loosens. The microenvironment of amino acid residue in α-amylase is changed that the hydrophobicity decreases and the polarity increases with Cu 2+ exposure. Isothermal titration calorimetry results show that Van der Waals force and hydrogen bond exist in the interaction between Cu 2+ and α-amylase. Results from this research would favor the development of advanced process for the biological treatment of sludge containing heavy metals.

In Silico Screening for Inhibitors of P-Glycoprotein That Target the Nucleotide Binding Domains

PubMed Central

Brewer, Frances K.; Follit, Courtney A.; Vogel, Pia D.

2014-01-01

Multidrug resistances and the failure of chemotherapies are often caused by the expression or overexpression of ATP-binding cassette transporter proteins such as the multidrug resistance protein, P-glycoprotein (P-gp). P-gp is expressed in the plasma membrane of many cell types and protects cells from accumulation of toxins. P-gp uses ATP hydrolysis to catalyze the transport of a broad range of mostly hydrophobic compounds across the plasma membrane and out of the cell. During cancer chemotherapy, the administration of therapeutics often selects for cells which overexpress P-gp, thereby creating populations of cancer cells resistant to a variety of chemically unrelated chemotherapeutics. The present study describes extremely high-throughput, massively parallel in silico ligand docking studies aimed at identifying reversible inhibitors of ATP hydrolysis that target the nucleotide-binding domains of P-gp. We used a structural model of human P-gp that we obtained from molecular dynamics experiments as the protein target for ligand docking. We employed a novel approach of subtractive docking experiments that identified ligands that bound predominantly to the nucleotide-binding domains but not the drug-binding domains of P-gp. Four compounds were found that inhibit ATP hydrolysis by P-gp. Using electron spin resonance spectroscopy, we showed that at least three of these compounds affected nucleotide binding to the transporter. These studies represent a successful proof of principle demonstrating the potential of targeted approaches for identifying specific inhibitors of P-gp. PMID:25270578

Effect of enzymatic treatment of extracted sunflower proteins on solubility, amino acid composition, and surface activity.

PubMed

Conde, José Miñones; Escobar, María del Mar Yust; Pedroche Jiménez, Justo J; Rodríguez, Francisco Millán; Rodríguez Patino, Juan M

2005-10-05

Industrial proteins from agriculture of either animal or vegetable origin, including their peptide derivatives, are of great importance, from the qualitative and quantitative point of view, in food formulations (emulsions and foams). A fundamental understanding of the physical, chemical, and functional properties of these proteins is essential if the performance of proteins in foods is to be improved and if underutilized proteins, such as plant proteins (and their hydrolysates and peptides derivatives), are to be increasingly used in traditional and new processed food products (safe, high-quality, health foods with good nutritional value). In this contribution we have determined the main physicochemical characteristics (solubility, composition, and analysis of amino acids) of a sunflower protein isolate (SPI) and its hydrolysates with low (5.62%), medium (23.5%), and high (46.3%) degrees of hydrolysis. The hydrolysates were obtained by enzymatic treatment with Alcalase 2.4 L for DH 5.62 and 23.5% and with Alcalase 2.4 L and Flavorzyme 1000 MG sequentially for DH 46.3%. The protein concentration dependence on surface pressure (surface pressure isotherm), a measure of the surface activity of the products (SPI and its hydrolysates), was obtained by tensiometry. We have observed that the degree of hydrolysis has an effect on solubility, composition, and content of the amino acids of the SPI and its hydrolysates. The superficial activity and the adsorption efficiency were also affected by the degree of hydrolysis.

Cancer-Associated Mutants of RNA Helicase DDX3X Are Defective in RNA-Stimulated ATP Hydrolysis

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

Epling, Leslie B.; Grace, Christy R.; Lowe, Brandon R.

The DEAD-box RNA helicase DDX3X is frequently mutated in pediatric medulloblastoma. We dissect how these mutants affect DDX3X function with structural, biochemical, and genetic experiments. We identify an N-terminal extension (“ATP-binding loop”, ABL) that is critical for the stimulation of ATP hydrolysis by RNA. We present crystal structures suggesting that the ABL interacts dynamically with ATP and confirming that the interaction occurs in solution by NMR chemical shift perturbation and isothermal titration calorimetry. DEAD-box helicases require interaction between two conserved RecA-like helicase domains, D1 and D2 for function. We use NMR chemical shift perturbation to show that DDX3X interacts specificallymore » with double-stranded RNA through its D1 domain, with contact mediated by residues G302 and G325. Mutants of these residues, G302V and G325E, are associated with pediatric medulloblastoma. These mutants are defective in RNA-stimulated ATP hydrolysis. We show that DDX3X complements the growth defect in a ded1 temperature-sensitive strain of Schizosaccharomyces pombe, but the cancer-associated mutants G302V and G325E do not complement and exhibit protein expression defects. In conclusion, taken together, our results suggest that impaired translation of important mRNA targets by mutant DDX3X represents a key step in the development of medulloblastoma.« less

Polymeric amylase nanoparticles as a new semi-synthetic enzyme system for hydrolysis of starch.

PubMed

Say, R; Şenay, R Hilal; Biçen, Özlem; Ersöz, Arzu; Şişman Yılmaz, Filiz; Akgöl, Sinan; Denizli, Adil

2013-05-01

α-Amylase (EC 3.2.1.1; α-D-1,4,glucan glucanohydrolase) catalyzes the hydrolysis of α-D-(1,4)-glucosidic linkages in starch, glycogen, and various malto-oligosaccharides, by releasing α-anomeric products. In this study, a novel method has been developed to prepare nanoprotein particles that carry α-amylase as a monomer by using a photosensitive microemulsion polymerization process. The nanostructured α-amylase with photosensitive features have been characterized by fluorescence spectroscopy, transmission electron microscopy (TEM) and Zeta Sizer. The fluorescence intensity of amylase nanoparticles was determined to be 658 a.u. at 610 nm and the average particle size of nanoamylase was found to be about 71.8 nm. Both free α-amylase and nanoparticles were used in the hydrolysis of starch under varying reaction conditions such as pH and temperature that affect enzyme activity and the results were compared to each other. Km values were 0.26 and 0.87 mM and Vmax values were 0.36 IU mg(-1) and 22.32 IU mg(-1) for nanoenzyme and free enzyme, respectively. Then, thermal stability, storage stability and reusability were investigated and according to the results, activity was preserved 60% at 60 °C; 20% at 70-80 °C temperature values and 80% after 105 days storage. Finally after 10 cycles, the activity was preserved 90% and this novel enzymatic polymeric amylase nanoparticle has showed considerable potential as reusable catalyst. Copyright © 2012 Elsevier B.V. All rights reserved.

Cancer-Associated Mutants of RNA Helicase DDX3X Are Defective in RNA-Stimulated ATP Hydrolysis

DOE PAGES

Epling, Leslie B.; Grace, Christy R.; Lowe, Brandon R.; ...

2015-02-25

The DEAD-box RNA helicase DDX3X is frequently mutated in pediatric medulloblastoma. We dissect how these mutants affect DDX3X function with structural, biochemical, and genetic experiments. We identify an N-terminal extension (“ATP-binding loop”, ABL) that is critical for the stimulation of ATP hydrolysis by RNA. We present crystal structures suggesting that the ABL interacts dynamically with ATP and confirming that the interaction occurs in solution by NMR chemical shift perturbation and isothermal titration calorimetry. DEAD-box helicases require interaction between two conserved RecA-like helicase domains, D1 and D2 for function. We use NMR chemical shift perturbation to show that DDX3X interacts specificallymore » with double-stranded RNA through its D1 domain, with contact mediated by residues G302 and G325. Mutants of these residues, G302V and G325E, are associated with pediatric medulloblastoma. These mutants are defective in RNA-stimulated ATP hydrolysis. We show that DDX3X complements the growth defect in a ded1 temperature-sensitive strain of Schizosaccharomyces pombe, but the cancer-associated mutants G302V and G325E do not complement and exhibit protein expression defects. In conclusion, taken together, our results suggest that impaired translation of important mRNA targets by mutant DDX3X represents a key step in the development of medulloblastoma.« less

An alternative analytical method based on ultrasound micro bath hydrolysis and GC-MS analysis for the characterization of organic biomarkers in archaeological ceramics.

PubMed

Blanco-Zubiaguirre, Laura; Olivares, Maitane; Castro, Kepa; Iñañez, Javier G; Madariaga, Juan Manuel

2016-11-01

The analysis of organic biomarkers in ancient and valuable archaeological remains provides a worthwhile source of information regarding their management. This work was focused on the development of an analytical procedure to characterize organic residues that have remained in archaeological ceramic samples. A novel analytical approach based on an alkaline hydrolysis by means of an ultrasound micro bath followed by liquid extraction was proposed to isolate saturated and unsaturated fatty acids, degradation products such as dihydroxy acids or dienoic fatty acids, isoprenoid fatty acids, and many other biomarkers from archaeological remains. This main goal has been achieved after the optimization of the main parameters affecting the hydrolysis step, the extraction procedure, and the derivatization step prior to the gas chromatography-mass spectrometry analysis. In this work, archaeological ceramic remains suspected to have been used by Basque Whalers to store whale oil in the period from the sixteenth to the seventeenth century were studied. Nevertheless, the proposed method is useful to determine the organic remains preserved in many other archaeological ceramic remains. Moreover, this methodology can be used to determine organic remains in any porous ceramic, archaeological or not. The preliminary results of the analysis of ceramic vessels led to the determination of some interesting unsaturated compounds such as 11-eicosenoic acid, an important biomarker of marine commodities, and several saturated fatty acids, which could be indicative of having used the vessels to store whale oil. Graphical abstract ᅟ.

Response surface methodology applied to the generation of casein hydrolysates with antioxidant and dipeptidyl peptidase IV inhibitory properties.

PubMed

Nongonierma, Alice B; Maux, Solène Le; Esteveny, Claire; FitzGerald, Richard J

2017-03-01

Hydrolysis parameters affecting the release of dipeptidyl peptidase IV (DPP-IV) inhibitory and antioxidant peptides from milk proteins have not been extensively studied. Therefore, a multifactorial (i.e. pH, temperature and hydrolysis time) composite design was used to optimise the release of bioactive peptides (BAPs) with DPP-IV inhibitory and antioxidant [oxygen radical absorbance capacity (ORAC)] properties from sodium caseinate. Fifteen sodium caseinate hydrolysates (H1-H15) were generated with Protamex TM , a bacillus proteinase activity. Hydrolysis time (1 to 5 h) had the highest influence on both DPP-IV inhibitory properties and ORAC activity (P < 0.05). Alteration of incubation temperature (40 to 60 °C) and pH (6.5 to 8.0) had an effect on the DPP-IV inhibitory properties but not the ORAC activity of the Protamex sodium caseinate hydrolysates. A multi-functional hydrolysate, H12, was identified having DPP-IV inhibitory (actual: 0.82 ± 0.24 vs. predicted optimum: 0.68 mg mL -1 ) and ORAC (actual: 639 ± 66 vs. predicted optimum: 639 µmol TE g -1 ) activity of the same order (P > 0.05) as the response surface methodology (RSM) predicted optimum bioactivities. Generation of milk protein hydrolysates through multifactorial design approaches may aid in the optimal enzymatic release of BAPs with serum glucose lowering and antioxidant properties. © 2016 Society of Chemical Industry. © 2016 Society of Chemical Industry.

Woody biomass: Niche position as a source of sustainable renewable chemicals and energy and kinetics of hot-water extraction/hydrolysis.

PubMed

Liu, Shijie

2010-01-01

The conversion of biomass to chemicals and energy is imperative to sustaining our way of life as known to us today. Fossil chemical and energy sources are traditionally regarded as wastes from a distant past. Petroleum, natural gas, and coal are not being regenerated in a sustainable manner. However, biomass sources such as algae, grasses, bushes and forests are continuously being replenished. Woody biomass represents the most abundant and available biomass source. Woody biomass is a reliably sustainable source of chemicals and energy that could be replenished at a rate consistent with our needs. The biorefinery is a concept describing the collection of processes used to convert biomass to chemicals and energy. Woody biomass presents more challenges than cereal grains for conversion to platform chemicals due to its stereochemical structures. Woody biomass can be thought of as comprised of at least four components: extractives, hemicellulose, lignin and cellulose. Each of these four components has a different degree of resistance to chemical, thermal and biological degradation. The biorefinery concept proposed at ESF (State University of New York - College of Environmental Science and Forestry) aims at incremental sequential deconstruction, fractionation/conversion of woody biomass to achieve efficient separation of major components. The emphasis of this work is on the kinetics of hot-water extraction, filling the gap in the fundamental understanding, linking engineering developments, and completing the first step in the biorefinery processes. This first step removes extractives and hemicellulose fractions from woody biomass. While extractives and hemicellulose are largely removed in the extraction liquor, cellulose and lignin largely remain in the residual woody structure. Xylo-oligomers and acetic acid in the extract are the major components having the greatest potential value for development. Extraction/hydrolysis involves at least 16 general reactions that could be divided into four categories: adsorption of proton onto woody biomass, hydrolysis reactions on the woody biomass surface, dissolution of soluble substances into the extraction liquor, and hydrolysis and dehydration decomposition in the extraction liquor. The extraction/hydrolysis rates are significantly simplified when the reactivity of all the intermonomer bonds are regarded as identical within each macromolecule, and the overall reactivity are identical for all the extractable macromolecules on the surface. A pseudo-first order extraction rate expression has been derived based on concentrations in monomer units. The reaction rate constant is however lower at the beginning of the extraction than that towards the end of the extraction. Furthermore, the H-factor and/or severity factor can be applied to lump the effects of temperature and residence time on the extraction process, at least for short times. This provides a means to control and optimize the performance of the extraction process effectively. Copyright 2010 Elsevier Inc. All rights reserved.

Internal Hydrolysis Indicator for Sample Specific Monitoring of β-Glucuronidase Activity.

PubMed

Taylor, Lacy L; Flint, Noah A; Ma, Vinh; Hill, Brandy M; Clark, Chantry J; Strathmann, Frederick G

2017-06-01

Metabolized forms of benzodiazepines (benzos) can cause issues with mass spectrometry identification. Benzodiazepines undergo a process called glucuronidation during metabolism that attaches a glucuronic acid for increased solubility. Often in clinical testing an enzymatic hydrolysis step is implemented to increase the sensitivity of benzodiazepines by hydrolyzing β-D-glucuronic acid from benzodiazepine-glucuronide conjugates in urine samples using the β-Glucuronidase enzyme. In this study resorufin β-D-glucuronide, a substrate of the β-Glucuronidase enzyme, was added to patient samples to determine if proper hydrolysis had occurred. The presence of resorufin as an Internal Hydrolysis Indicator (IHI) shows the activity and efficiency of the enzyme in each patient sample. Synthetic/patient urine samples were obtained and mixed with hydrolysis buffer containing resorufin β-D-glucuronide. The β-Glucuronidase enzyme was used to hydrolyze the benzodiazepine analytes as well as resorufin β-D-glucuronide. The enzymatic hydrolysis addition increased the positivity rate of benzodiazepines by 42.5%. The β-Glucuronidase substrate resorufin (IHI) displayed variability in area counts between patient samples. Comparative studies with internal standards and resorufin (IHI) showed no correlation between recovery and analyte variability. Hydrolysis reactions greatly improved the sensitivity of benzodiazepines by liquid chromatography time-of-flight mass spectrometry analysis. The large variation in resorufin (IHI) area counts amongst patient samples indicates possible variability in enzymatic hydrolysis activity. The enzymatic hydrolysis step is a part of the extraction procedure and should be controlled for in each patient sample. © The Author 2017. Published by Oxford University Press. All rights reserved. For Permissions, please email: journals.permissions@oup.com.

Hydrolysis of substance P in the presence of the osteosarcoma cell line SaOS-2: release of free amino acids.

PubMed

Cavazza, Antonella; Marini, Mario; Roda, L Giorgio; Tarantino, Umberto; Valenti, Angela

2011-12-01

The possible hydrolysis of substance P (Arg-Pro-Lys-Pro-Gln-Gln-Phe-Phe-Gly-Leu-Met) in presence of the osteoblastic cell line SaOS-2 was measured by capillary electrophoresis coupled to mass detection. The results obtained indicate that a very rapid disappearance of the intact undecapeptide was associated to a slower appearance of seven of its eight component amino acids. These results can be interpreted as indicating that an extremely fast hydrolysis of substance P by endopeptidases, which released peptidic by-products, was followed by a noticeably slower secondary degradation which released free amino acids. In decreasing quantitative importance, these phenomena appear to originate by the hydrolysis of the Pro(4)-Gln(5) bond, followed by C-terminal sequential degradation of the Arg(1)-Pro(4) tetrapeptide; by the hydrolysis of or Phe(7)-Phe(8) bond (or, possibly, of Gln(6)-Phe(7)) leading to release of free Phe and Gln; by hydrolysis of the Gly(9)-Leu(10) bond with subsequent release of Met and Leu. Results obtained appear to be compatible with the expression by SaOS-2 cells of enzymes already known to catalyze substance P hydrolysis, together with an apparent low efficiency of aminopeptidases. Because of the activity of C-terminal fragments on NK1 receptors, the delay between primary hydrolysis of substance P and secondary hydrolysis of its peptidic fragments indicated by the data shown implies a possible persistence of substance P physiological effects even after degradation of the intact peptide.

PSA Converts Parathyroid Hormone-Related Protein (THrP) from an Osteolytic to an Osteoblastic Factor: Role in Bone Metastasis

DTIC Science & Technology

2004-12-01

bone density associated with DLX3 mutation in the tricho-dento-osseous syndrome . Bone. 35:988-997. Hassan MQ, Javed A, Morasso MI, Karlin J, Montecino M...putative role in hydrolysis of osteogenic peptides. J Bone Miner Res. 15:1266-1274. Ryan CW, Vogelzang NJ, Vokes EE, Kindler HL, Undevia SD, Humerickhouse

ABC transporters and immunity: mechanism of self-defense.

PubMed

Hinz, Andreas; Tampé, Robert

2012-06-26

The transporter associated with antigen processing (TAP) is a prototype of an asymmetric ATP-binding cassette (ABC) transporter, which uses ATP binding and hydrolysis to translocate peptides from the cytosol to the lumen of the endoplasmic reticulum (ER). Here, we review molecular details of peptide binding and ATP binding and hydrolysis as well as the resulting allosteric cross-talk between the nucleotide-binding domains and the transmembrane domains that drive translocation of the solute across the ER membrane. We also discuss the general molecular architecture of ABC transporters and demonstrate the importance of structural and functional studies for a better understanding of the role of the noncanonical site of asymmetric ABC transporters. Several aspects of peptide binding and specificity illustrate details of peptide translocation by TAP. Furthermore, this ABC transporter forms the central part of the major histocompatibility complex class I (MHC I) peptide-loading machinery. Hence, TAP is confronted with a number of viral factors, which prevent antigen translocation and MHC I loading in virally infected cells. We review how these viral factors have been used as molecular tools to decipher mechanistic aspects of solute translocation and discuss how they can help in the structural analysis of TAP.

Evidence against involvement of the acid lysosomal sphingomyelinase in the tumor-necrosis-factor- and interleukin-1-induced sphingomyelin cycle and cell proliferation in human fibroblasts.

PubMed Central

Andrieu, N; Salvayre, R; Levade, T

1994-01-01

The hydrolysis of sphingomyelin (SPM) has been reported to mediate a number of responses to extracellular agents, including cytokines. The so-called SPM cycle may result from the activation of different types of sphingomyelinases (SPMases). We investigated the hypothetical contribution of acid lysosomal SPMase in the SPM signal-transduction pathway. We examined the ability of human skin fibroblasts with a genetic deficiency of acid lysosomal SPMase activity to respond to tumour necrosis factor alpha (TNF-alpha) or interleukin-1 beta (IL-1 beta). We report that both cytokines promoted SPM hydrolysis in fibroblasts derived from patients with Niemann-Pick disease or I-cell disease, similar to that observed in normal cells. Treatment of normal fibroblasts with cationic amphiphilic drugs resulted in inhibition of acid SPMase activity, but had no effect on cytokine-induced SPM turnover. In addition, TNF-alpha and IL-1 beta stimulated [3H]thymidine incorporation in Niemann-Pick fibroblasts, as in normal cells. Thus our results argue against a role for acid endolysosomal SPMase in mediating the cytokine-induced SPM signalling cascade. Images Figure 2 PMID:7980390

Understanding the synergistic effect and the main factors influencing the enzymatic hydrolyzability of corn stover at low enzyme loading by hydrothermal and/or ultrafine grinding pretreatment.

PubMed

Zhang, Haiyan; Li, Junbao; Huang, Guangqun; Yang, Zengling; Han, Lujia

2018-05-26

A thorough assessment of the microstructural changes and synergistic effects of hydrothermal and/or ultrafine grinding pretreatment on the subsequent enzymatic hydrolysis of corn stover was performed in this study. The mechanism of pretreatment was elucidated by characterizing the particle size, specific surface area (SSA), pore volume (PV), average pore size, cellulose crystallinity (CrI) and surface morphology of the pretreated samples. In addition, the underlying relationships between the structural parameters and final glucose yields were elucidated, and the relative significance of the factors influencing enzymatic hydrolyzability were assessed by principal component analysis (PCA). Hydrothermal pretreatment at a lower temperature (170 °C) combined with ultrafine grinding achieved a high glucose yield (80.36%) at a low enzyme loading (5 filter paper unit (FPU)/g substrate) which is favorable. The relative significance of structural parameters in enzymatic hydrolyzability was SSA > PV > average pore size > CrI/cellulose > particle size. PV and SSA exhibited logarithmic correlations with the final enzymatic hydrolysis yield. Copyright © 2018 Elsevier Ltd. All rights reserved.

Mammalian translation elongation factor eEF1A2: X-ray structure and new features of GDP/GTP exchange mechanism in higher eukaryotes

PubMed Central

Crepin, Thibaut; Shalak, Vyacheslav F.; Yaremchuk, Anna D.; Vlasenko, Dmytro O.; McCarthy, Andrew; Negrutskii, Boris S.; Tukalo, Michail A.; El'skaya, Anna V.

2014-01-01

Eukaryotic elongation factor eEF1A transits between the GTP- and GDP-bound conformations during the ribosomal polypeptide chain elongation. eEF1A*GTP establishes a complex with the aminoacyl-tRNA in the A site of the 80S ribosome. Correct codon–anticodon recognition triggers GTP hydrolysis, with subsequent dissociation of eEF1A*GDP from the ribosome. The structures of both the ‘GTP’- and ‘GDP’-bound conformations of eEF1A are unknown. Thus, the eEF1A-related ribosomal mechanisms were anticipated only by analogy with the bacterial homolog EF-Tu. Here, we report the first crystal structure of the mammalian eEF1A2*GDP complex which indicates major differences in the organization of the nucleotide-binding domain and intramolecular movements of eEF1A compared to EF-Tu. Our results explain the nucleotide exchange mechanism in the mammalian eEF1A and suggest that the first step of eEF1A*GDP dissociation from the 80S ribosome is the rotation of the nucleotide-binding domain observed after GTP hydrolysis. PMID:25326326

Platinum-catalyzed hydrolysis etching of SiC in water: A density functional theory study

NASA Astrophysics Data System (ADS)

Van Bui, Pho; Toh, Daisetsu; Isohashi, Ai; Matsuyama, Satoshi; Inagaki, Kouji; Sano, Yasuhisa; Yamauchi, Kazuto; Morikawa, Yoshitada

2018-05-01

A comprehensive study of the physicochemical interactions and the reaction mechanism of SiC etching with water by Pt catalysts can reveal key details about the surface treatment and catalytic phenomena at interfaces. Therefore, density functional theory simulations were performed to study the kinetics of Pt-assisted water dissociation and breaking of a Si–C bond compared to the HF-assisted mechanism. These calculations carefully considered the elastic and chemical interaction energies at the Pt–SiC interface, activation barriers of Si–C bond dissociation, and the catalytic role of Pt. It was found that the Pt-catalyzed etching of SiC in water is initiated via hydrolysis reactions that break the topmost Si–C bonds. The activation barrier strongly depends on the elastic and chemical interactions. However, chemical interactions are a dominant factor and mainly contribute to the lowering of the activation barrier, resulting in an increased rate of reaction.

Continuous thermal hydrolysis and energy integration in sludge anaerobic digestion plants.

PubMed

Fdz-Polanco, F; Velazquez, R; Perez-Elvira, S I; Casas, C; del Barrio, D; Cantero, F J; Fdz-Polanco, M; Rodriguez, P; Panizo, L; Serrat, J; Rouge, P

2008-01-01

A thermal hydrolysis pilot plant with direct steam injection heating was designed and constructed. In a first period the equipment was operated in batch to verify the effect of sludge type, pressure and temperature, residence time and solids concentration. Optimal operation conditions were reached for secondary sludge at 170 degrees C, 7 bar and 30 minutes residence time, obtaining a disintegration factor higher than 10, methane production increase by 50% and easy centrifugation In a second period the pilot plant was operated working with continuous feed, testing the efficiency by using two continuous anaerobic digester operating in the mesophilic and thermophilic range. Working at 12 days residence time, biogas production increases by 40-50%. Integrating the energy transfer it is possible to design a self-sufficient system that takes advantage of this methane increase to produce 40% more electric energy. (c) IWA Publishing 2008.

Study on treatment technology of wastewater from hydrolysis of acid oil

NASA Astrophysics Data System (ADS)

Li, Yuejin; Lin, Zhiyong; Han, Yali

2017-06-01

In this paper, the degumming of ferric chloride, calcium hydroxide after the removal of acid acidification hydrolysis of waste oil as raw material, through the treatment process to purify the wastewater. Choose different chemical additives, investigation of different temperature, pH value and other factors, find the best extraction condition. Through the orthogonal test of sodium carbonate, sodium oxalate, barium carbonate, compared with three kinds of chemical additives. The best chemical assistant is sodium carbonate, the best treatment temperature is 80 degrees Celsius, pH value is 8.0. After the reaction, the content of calcium and iron ions were determined by suitable methods. The removal rate of calcium ion is 98%, the removal rate of iron ion is 99%, and the effect of calcium and iron ion precipitation on the subsequent evaporation operation is reduced. Finally, the comparison is made to clarify the Dilute Glycerol water solution.

Identification and characterization of core cellulolytic enzymes from Talaromyces cellulolyticus (formerly Acremonium cellulolyticus) critical for hydrolysis of lignocellulosic biomass

DOE PAGES

Inoue, Hiroyuki; Decker, Stephen R.; Taylor, Larry E.; ...

2014-10-09

Background: Enzymatic hydrolysis of pretreated lignocellulosic biomass is an essential process for the production of fermentable sugars for industrial use. A better understanding of fungal cellulase systems will provide clues for maximizing the hydrolysis of target biomass. Talaromyces cellulolyticus is a promising fungus for cellulase production and efficient biomass hydrolysis. Several cellulolytic enzymes purified from T. cellulolyticus were characterized in earlier studies, but the core enzymes critical for hydrolysis of lignocellulosic biomass remain unknown. Results: Six cellulolytic enzymes critical for the hydrolysis of crystalline cellulose were purified from T. cellulolyticus culture supernatant using an enzyme assay based on synergistic hydrolysismore » of Avicel. The purified enzymes were identified by their substrate specificities and analyses of trypsin-digested peptide fragments and were classified into the following glycosyl hydrolase (GH) families: GH3 (β-glucosidase, Bgl3A), GH5 (endoglucanase, Cel5A), GH6 (cellobiohydrolase II, Cel6A), GH7 (cellobiohydrolase I and endoglucanase, Cel7A and Cel7B, respectively), and GH10 (xylanase, Xyl10A). Hydrolysis of dilute acid-pretreated corn stover (PCS) with mixtures of the purified enzymes showed that Cel5A, Cel7B, and Xyl10A each had synergistic effects with a mixture of Cel6A and Cel7A. Cel5A seemed to be more effective in the synergistic hydrolysis of the PCS than Cel7B. The ratio of Cel5A, Cel6A, Cel7A, and Xyl10A was statistically optimized for the hydrolysis of PCS glucan in the presence of Bgl3A. The resultant mixture achieved higher PCS glucan hydrolysis at lower enzyme loading than a culture filtrate from T. cellulolyticus or a commercial enzyme preparation, demonstrating that the five enzymes play a role as core enzymes in the hydrolysis of PCS glucan. In Conclusion: Core cellulolytic enzymes in the T. cellulolyticus cellulase system were identified to Cel5A, Cel6A, Cel7A, Xyl10A, and Bgl3A and characterized. The optimized mixture of these five enzymes was highly effective for the hydrolysis of PCS glucan, providing a foundation for future improvement of the T. cellulolyticus cellulase system.« less

Identification and characterization of core cellulolytic enzymes from Talaromyces cellulolyticus (formerly Acremonium cellulolyticus) critical for hydrolysis of lignocellulosic biomass

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

Inoue, Hiroyuki; Decker, Stephen R.; Taylor, Larry E.

Background: Enzymatic hydrolysis of pretreated lignocellulosic biomass is an essential process for the production of fermentable sugars for industrial use. A better understanding of fungal cellulase systems will provide clues for maximizing the hydrolysis of target biomass. Talaromyces cellulolyticus is a promising fungus for cellulase production and efficient biomass hydrolysis. Several cellulolytic enzymes purified from T. cellulolyticus were characterized in earlier studies, but the core enzymes critical for hydrolysis of lignocellulosic biomass remain unknown. Results: Six cellulolytic enzymes critical for the hydrolysis of crystalline cellulose were purified from T. cellulolyticus culture supernatant using an enzyme assay based on synergistic hydrolysismore » of Avicel. The purified enzymes were identified by their substrate specificities and analyses of trypsin-digested peptide fragments and were classified into the following glycosyl hydrolase (GH) families: GH3 (β-glucosidase, Bgl3A), GH5 (endoglucanase, Cel5A), GH6 (cellobiohydrolase II, Cel6A), GH7 (cellobiohydrolase I and endoglucanase, Cel7A and Cel7B, respectively), and GH10 (xylanase, Xyl10A). Hydrolysis of dilute acid-pretreated corn stover (PCS) with mixtures of the purified enzymes showed that Cel5A, Cel7B, and Xyl10A each had synergistic effects with a mixture of Cel6A and Cel7A. Cel5A seemed to be more effective in the synergistic hydrolysis of the PCS than Cel7B. The ratio of Cel5A, Cel6A, Cel7A, and Xyl10A was statistically optimized for the hydrolysis of PCS glucan in the presence of Bgl3A. The resultant mixture achieved higher PCS glucan hydrolysis at lower enzyme loading than a culture filtrate from T. cellulolyticus or a commercial enzyme preparation, demonstrating that the five enzymes play a role as core enzymes in the hydrolysis of PCS glucan. In Conclusion: Core cellulolytic enzymes in the T. cellulolyticus cellulase system were identified to Cel5A, Cel6A, Cel7A, Xyl10A, and Bgl3A and characterized. The optimized mixture of these five enzymes was highly effective for the hydrolysis of PCS glucan, providing a foundation for future improvement of the T. cellulolyticus cellulase system.« less

Structure-activity correlations for organophosphorus ester anticholinesterases. Part 2: CNDO/2 calculations applied to ester hydrolysis rates

NASA Technical Reports Server (NTRS)

Johnson, H.; Kenley, R. A.; Rynard, C.; Golub, M. A.

1984-01-01

Quantitative structure-activity relationships are presented for the hydrolysis of organophosphorus esters, RR'P(O)X, where R and R' are alkyl and/or alkoxy groups and X is fluorine, chlorine or a phenoxy group. CNDO/2 calculations provide values for molecular parameters that correlate with alkaline hydrolysis rates. For each subset of esters with the same leaving group, X, the CNDO-derived net atomic charge at the central phosphorus atom correlates well with the alkaline hydrolysis rate constants. For the whole set of esters with different leaving groups, equations are derived that relate charge, orbital energy and bond order to the hydrolysis rate constants.

Progressing batch hydrolysis process

DOEpatents

Wright, J.D.

1985-01-10

A progressive batch hydrolysis process is disclosed for producing sugar from a lignocellulosic feedstock. It comprises passing a stream of dilute acid serially through a plurality of percolation hydrolysis reactors charged with feed stock, at a flow rate, temperature and pressure sufficient to substantially convert all the cellulose component of the feed stock to glucose. The cooled dilute acid stream containing glucose, after exiting the last percolation hydrolysis reactor, serially fed through a plurality of pre-hydrolysis percolation reactors, charged with said feedstock, at a flow rate, temperature and pressure sufficient to substantially convert all the hemicellulose component of said feedstock to glucose. The dilute acid stream containing glucose is cooled after it exits the last prehydrolysis reactor.

Stepwise hydrolysis to improve carbon releasing efficiency from sludge.

PubMed

Liu, Hongbo; Wang, Yuanyuan; Wang, Ling; Yu, Tiantian; Fu, Bo; Liu, He

2017-08-01

Based on thermal alkaline hydrolysis (TAH), a novel strategy of stepwise hydrolysis was developed to improve carbon releasing efficiency from waste activated sludge (WAS). By stepwise increasing hydrolysis intensity, conventional sludge hydrolysis (the control) was divided into four stages for separately recovering sludge carbon sources with different bonding strengths, namely stage 1 (60 °C, pH 6.0-8.0), stage 2 (80 °C, pH 6.0-8.0), stage 3 (80 °C, pH 10.0) and stage 4 (90 °C, pH 12.0). Results indicate stepwise hydrolysis could enhance the amount of released soluble chemical oxygen demand (SCOD) for almost 2 times, from 7200 to 14,693 mg/L, and the released carbon presented better biodegradability, with BOD/COD of 0.47 and volatile fatty acids (VFAs) yield of 0.37 g VFAs/g SCOD via anaerobic fermentation. Moreover, stepwise hydrolysis also improved the dewaterability of hydrolyzed sludge, capillary suction time (CST) reducing from 2500 to 1600 s. Economic assessment indicates stepwise hydrolysis shows less alkali demand and lower thermal energy consumption than those of the control. Furthermore, results of this study help support the concepts of improving carbon recovery in wastewater by manipulating WAS composition and the idea of classifiably recovering the nutrients in WAS. Copyright © 2017 Elsevier Ltd. All rights reserved.

Transcriptome and Metabolome Analyses in Exogenous FABP4- and FABP5-Treated Adipose-Derived Stem Cells

PubMed Central

Sugaya, Takeshi; Oikawa, Tsuyoshi; Matsumoto, Megumi; Funahashi, Yasuhito; Matsukawa, Yoshihisa; Gotoh, Momokazu; Miura, Tetsuji

2016-01-01

Adipose-derived stem cells (ADSC), which exist near adipocytes in adipose tissue, have been used as a potential tool of regenerative medicine. Lipid chaperones, fatty acid-binding protein 4 (FABP4) and 5 (FABP5), are abundantly expressed in adipocytes. FABP4 has recently been shown to be secreted from adipocytes during lipolysis in a non-classical pathway and may act as an adipokine. Here, we investigated the role of exogenous FABP4 and FABP5 in transcriptional and metabolic regulation in ADSC. FABP4 and FABP5 were little expressed in ADSC. However, both FABP4 and FABP5 were significantly induced after adipocyte differentiation of ADSC and were secreted from the differentiated adipocytes. Analysis of microarray data, including gene ontology enrichment analysis and cascade analysis of the protein-protein interaction network using a transcription factor binding site search, demonstrated that treatment of ADSC with FABP4 or FABP5 affected several kinds of genes related to inflammatory and metabolic responses and the process of cell differentiation. Notably, myogenic factors, including myocyte enhancer factors, myogenic differentiation 1 and myogenin, were modulated by treatment of ADSC with FABP4, indicating that exogenous FABP4 treatment is partially associated with myogenesis in ADSC. Metabolome analysis showed that treatment of ADSC with FABP4 and with FABP5 similarly, but differently in extent, promoted hydrolysis and/or uptake of lipids, consequentially together with enhancement of β oxidation, inhibition of downstream of the glycolysis pathway, accumulation of amino acids, reduction of nucleic acid components and increase in the ratio of reduced and oxidized nicotinamide adenine dinucleotide phosphates (NADPH/NADP+), an indicator of reducing power, and the ratio of adenosine triphosphate and adenosine monophosphate (ATP/AMP), an indicator of the energy state, in ADSC. In conclusion, secreted FABP4 and FABP5 from adipocytes as adipokines differentially affect transcriptional and metabolic regulation in ADSC near adipocytes. The adiposity condition in the host of regenerative medicine may affect characteristics of ADSC by exposure of the balance of FABP4 and FABP5. PMID:27936164

Transcriptome and Metabolome Analyses in Exogenous FABP4- and FABP5-Treated Adipose-Derived Stem Cells.

PubMed

Yamamoto, Tokunori; Furuhashi, Masato; Sugaya, Takeshi; Oikawa, Tsuyoshi; Matsumoto, Megumi; Funahashi, Yasuhito; Matsukawa, Yoshihisa; Gotoh, Momokazu; Miura, Tetsuji

2016-01-01

Adipose-derived stem cells (ADSC), which exist near adipocytes in adipose tissue, have been used as a potential tool of regenerative medicine. Lipid chaperones, fatty acid-binding protein 4 (FABP4) and 5 (FABP5), are abundantly expressed in adipocytes. FABP4 has recently been shown to be secreted from adipocytes during lipolysis in a non-classical pathway and may act as an adipokine. Here, we investigated the role of exogenous FABP4 and FABP5 in transcriptional and metabolic regulation in ADSC. FABP4 and FABP5 were little expressed in ADSC. However, both FABP4 and FABP5 were significantly induced after adipocyte differentiation of ADSC and were secreted from the differentiated adipocytes. Analysis of microarray data, including gene ontology enrichment analysis and cascade analysis of the protein-protein interaction network using a transcription factor binding site search, demonstrated that treatment of ADSC with FABP4 or FABP5 affected several kinds of genes related to inflammatory and metabolic responses and the process of cell differentiation. Notably, myogenic factors, including myocyte enhancer factors, myogenic differentiation 1 and myogenin, were modulated by treatment of ADSC with FABP4, indicating that exogenous FABP4 treatment is partially associated with myogenesis in ADSC. Metabolome analysis showed that treatment of ADSC with FABP4 and with FABP5 similarly, but differently in extent, promoted hydrolysis and/or uptake of lipids, consequentially together with enhancement of β oxidation, inhibition of downstream of the glycolysis pathway, accumulation of amino acids, reduction of nucleic acid components and increase in the ratio of reduced and oxidized nicotinamide adenine dinucleotide phosphates (NADPH/NADP+), an indicator of reducing power, and the ratio of adenosine triphosphate and adenosine monophosphate (ATP/AMP), an indicator of the energy state, in ADSC. In conclusion, secreted FABP4 and FABP5 from adipocytes as adipokines differentially affect transcriptional and metabolic regulation in ADSC near adipocytes. The adiposity condition in the host of regenerative medicine may affect characteristics of ADSC by exposure of the balance of FABP4 and FABP5.

Microwave Pretreatment For Hydrolysis Of Cellulose

NASA Technical Reports Server (NTRS)

Cullingford, Hatice S.; George, Clifford E.; Lightsey, George R.

1993-01-01

Microwave pretreatment enhances enzymatic hydrolysis of cellulosic wastes into soluble saccharides used as feedstocks for foods, fuels, and other products. Low consumption of energy, high yield, and low risk of proposed hydrolysis process incorporating microwave pretreatment makes process viable alternative to composting.

A kinetic study of hydrolysis of polyester elastomer in magnetic tape

NASA Technical Reports Server (NTRS)

Yamamoto, K.; Watanabe, H.

1994-01-01

A useful method for kinetic study of the hydrolysis of polyester elastomer is established which uses the number-average molecular weight. The reasonableness of this method is confirmed and the effect of magnetic particles on hydrolysis is considered.

ESTIMATION OF PHOSPHATE ESTER HYDROLYSIS RATE CONSTANTS - ALKALINE HYDROLYSIS

EPA Science Inventory

SPARC (SPARC Performs Automated Reasoning in Chemistry) chemical reactivity models were extended to allow the calculation of alkaline hydrolysis rate constants of phosphate esters in water. The rate is calculated from the energy difference between the initial and transition state...

Environmental Fate of Organophosphorus Compounds Related to Chemical Weapons

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

Davisson, M L; Love, A H; Vance, A

2005-02-08

Man-made organophosphorus compounds have been widely distributed throughout our environment as pesticides since their development during and after WWII. Many important studies have documented their relative persistence and toxicity. Development and use of some organophosphorus compounds as nerve agents gave rise to a separate but parallel effort to understand environmental persistence. In this latter case, the experiments have focused mainly on evaporation rates and first-order reaction kinetics. However, because organophosphorus compounds are easily polarized, the ionic content of a surrounding media directly factors into these reaction rates, but limited work in this regard has been done under environmentally relevant conditions.more » Furthermore, limited experiments investigating persistence of these agents on soil has resulted in widely varying degradation rates. Not surprisingly, no studies have investigated affinities of organophosphorus nerve agents to mineral or organic matter typically found in soil. As a result, we initiated laboratory experiments on dilute concentrations of nerve agent O-ethyl S-(2-diisopropylaminoethyl) methylphosphonothiolate (VX) to quantify persistence in simulated environmental aqueous conditions. A quantitative analytical method was developed for VX and its degradation products using High Performance Liquid Chromatography-Electrospray Ionization-Mass Spectrometry (HPLC-ESI-MS). VX hydrolysis rate is known to have a pH-dependency, however, the type of buffer and the relative proportion of different nucleophiles in solution significantly affect the overall rate and mechanism of degradation. For example, dissolved carbonate, a weak nucleophile dominating natural water, yielded pseudo-first order rate constants of {approx} 8 x 10{sup -3}/hr at pH 5 and 2 x 10{sup -2}/hr at pH 11. This small pH-dependent variation departs significantly from widely accepted rates at this pH range (4 x 10{sup -4}/hr to 8 x 10{sup -2}/hr) that were based on chloride and hydroxyl (strong nucleophile) dominated experimental solutions. Because of its overwhelming abundance in solution relative to hydroxyl ion, bicarbonate likely effectively competes in nucleophilic attack on phosphorus. The addition of natural dissolved organic matter at 100 mg/L in pH 7 bicarbonate buffered solution slowed VX hydrolysis rates {approx}2 times relative to controls, suggesting hydrophobic interaction. Adsorption experiments derived isotherms from batch aqueous experiments on montmorillonite clay, iron-oxyhydroxide goethite, and on amorphous silica. VX had moderate affinity for montmorillonite and amorphous silica, and very low affinity toward goethite. The addition of dissolved organic matter into solution enhanced VX adsorption to goethite, consistent with its high affinity for hydrophobic organic matter (log K{sub oc} = 2.52). Diisopropylaminoethylthiol (DESH), a hydrolysis product of VX showed equivalent adsorption to montmorillonite, and poor affinity to goethite and silica. However, hydrolysis products O-Ethylmethylphosphonic acid (EMPA) and methylphosphonic acid (MPA) strongly adsorbed on goethite, but not on montmorillonite or silica, suggesting a ligand-exchange mechanism. VX degraded rapidly when completely dried onto goethite followed by rehydration, consistent with an irreversible chemical adsorption mechanism.« less

Standard Gibbs energy of metabolic reactions: II. Glucose-6-phosphatase reaction and ATP hydrolysis.

PubMed

Meurer, Florian; Do, Hoang Tam; Sadowski, Gabriele; Held, Christoph

2017-04-01

ATP (adenosine triphosphate) is a key reaction for metabolism. Tools from systems biology require standard reaction data in order to predict metabolic pathways accurately. However, literature values for standard Gibbs energy of ATP hydrolysis are highly uncertain and differ strongly from each other. Further, such data usually neglect the activity coefficients of reacting agents, and published data like this is apparent (condition-dependent) data instead of activity-based standard data. In this work a consistent value for the standard Gibbs energy of ATP hydrolysis was determined. The activity coefficients of reacting agents were modeled with electrolyte Perturbed-Chain Statistical Associating Fluid Theory (ePC-SAFT). The Gibbs energy of ATP hydrolysis was calculated by combining the standard Gibbs energies of hexokinase reaction and of glucose-6-phosphate hydrolysis. While the standard Gibbs energy of hexokinase reaction was taken from previous work, standard Gibbs energy of glucose-6-phosphate hydrolysis reaction was determined in this work. For this purpose, reaction equilibrium molalities of reacting agents were measured at pH7 and pH8 at 298.15K at varying initial reacting agent molalities. The corresponding activity coefficients at experimental equilibrium molalities were predicted with ePC-SAFT yielding the Gibbs energy of glucose-6-phosphate hydrolysis of -13.72±0.75kJ·mol -1 . Combined with the value for hexokinase, the standard Gibbs energy of ATP hydrolysis was finally found to be -31.55±1.27kJ·mol -1 . For both, ATP hydrolysis and glucose-6-phosphate hydrolysis, a good agreement with own and literature values were obtained when influences of pH, temperature, and activity coefficients were explicitly taken into account in order to calculate standard Gibbs energy at pH7, 298.15K and standard state. Copyright © 2017 Elsevier B.V. All rights reserved.

Interdependence of the kinetics of NTP hydrolysis and the stability of the RecA-ssDNA complex.

PubMed

Katz, F S; Bryant, F R

2001-09-18

The ssDNA-dependent NTP hydrolysis activity of the RecA protein was examined using a series of dTn oligomers ranging in size from dT10 to dT2000 as the ssDNA effector. There were three distinct manifestations of the dTn-dependent NTP hydrolysis reaction, depending on the length of the dTn effector that was used. With longer dTn oligomers, NTP hydrolysis occurred with a turnover number of 20-25 min(-1) and the observed S0.5 value for the NTP was independent of the concentration of the dTn oligomer (DNA concentration-independent hydrolysis). With dTn oligomers of intermediate length, NTP hydrolysis still occurred with a turnover number of 20-25 min(-1), but the observed S0.5 for the NTP decreased with increasing dTn concentration until reaching a value similar to that obtained with the longer dTn oligomers (DNA concentration-dependent hydrolysis). With shorter dTn oligomers, the NTP hydrolysis activity was effectively eliminated. Although this general progression of kinetic behavior was observed for the three structurally related NTPs (dATP, ATP, and GTP), the dTn oligomer length at which DNA concentration-independent, DNA concentration-dependent, and no NTP hydrolysis was observed depended on the NTP being considered. For example, dATP (S0.5 = 35 microM) was hydrolyzed in the presence of dT20, whereas ATP (S0.5 = 70 microM) and GTP (S0.5 = 1200 microM) required at least dT50 and dT200 for hydrolysis, respectively. These results are discussed in terms of a kinetic model in which the stability of the RecA-ssDNA-NTP complex is dependent on the intrinsic S0.5 value of the NTP being hydrolyzed.

Prediction of Hydrolysis Products of Organic Chemicals under Environmental pH Conditions.

PubMed

Tebes-Stevens, Caroline; Patel, Jay M; Jones, W Jack; Weber, Eric J

2017-05-02

Cheminformatics-based software tools can predict the molecular structure of transformation products using a library of transformation reaction schemes. This paper presents the development of such a library for abiotic hydrolysis of organic chemicals under environmentally relevant conditions. The hydrolysis reaction schemes in the library encode the process science gathered from peer-reviewed literature and regulatory reports. Each scheme has been ranked on a scale of one to six based on the median half-life in a data set compiled from literature-reported hydrolysis rates. These ranks are used to predict the most likely transformation route when more than one structural fragment susceptible to hydrolysis is present in a molecule of interest. Separate rank assignments are established for pH 5, 7, and 9 to represent standard conditions in hydrolysis studies required for registration of pesticides in Organisation for Economic Co-operation and Development (OECD) member countries. The library is applied to predict the likely hydrolytic transformation products for two lists of chemicals, one representative of chemicals used in commerce and the other specific to pesticides, to evaluate which hydrolysis reaction pathways are most likely to be relevant for organic chemicals found in the natural environment.

Aspartic acid 405 contributes to the substrate specificity of aminopeptidase B.

PubMed

Fukasawa, Kayoko M; Hirose, Junzo; Hata, Toshiyuki; Ono, Yukio

2006-09-26

Aminopeptidase B (EC 3.4.11.6, ApB) specifically cleaves in vitro the N-terminal Arg or Lys residue from peptides and synthetic derivatives. Ap B was shown to have a consensus sequence found in the metallopeptidase family. We determined the putative zinc binding residues (His324, His328, and Glu347) and the essential Glu325 residue for the enzyme using site-directed mutagenesis (Fukasawa, K. M., et al. (1999) Biochem. J. 339, 497-502). To identify the residues binding to the amino-terminal basic amino acid of the substrate, rat cDNA encoding ApB was cloned into pGEX-4T-3 so that recombinant protein was expressed as a GST fusion protein. Twelve acidic amino acid residues (Glu or Asp) in ApB were replaced with a Gln or Asn using site-directed mutagenesis. These mutants were isolated to characterize the kinetic parameters of enzyme activity toward Arg-NA and compare them to those of the wild-type ApB. The catalytic efficiency (kcat/Km) of the mutant D405N was 1.7 x 10(4) M(-1) s(-1), markedly decreased compared with that of the wild-type ApB (6.2 x 10(5) M(-1) s(-1)). The replacement of Asp405 with an Asn residue resulted in the change of substrate specificity such that the specific activity of the mutant D405N toward Lys-NA was twice that toward Arg-NA (in the case of wild-type ApB; 0.4). Moreover, when Asp405 was replaced with an Ala residue, the kcat/Km ratio was 1000-fold lower than that of the wild-type ApB for hydrolysis of Arg-NA; in contrast, in the hydrolysis of Tyr-NA, the kcat/Km ratios of the wild-type (1.1 x 10(4) M(-1) s(-1)) and the mutated (8.2 x 10(3) M(-1) s(-1)) enzymes were similar. Furthermore, the replacement of Asp-405 with a Glu residue led to the reduction of the kcat/Km ratio for the hydrolysis of Arg-NA by a factor of 6 and an increase of that for the hydrolysis of Lys-NA. Then the kcat/Km ratio of the D405E mutant for the hydrolysis of Lys-NA was higher than that for the hydrolysis of Arg-NA as opposed to that of wild-type ApB. These data strongly suggest that the Asp 405 residue is involved in substrate binding via an interaction with the P1 amino group of the substrate's side chain.

ESTIMATION OF PHOSPHATE ESTER HYDROLYSIS RATE CONSTANTS. I. ALKALINE HYDROLYSIS

EPA Science Inventory

SPARC (SPARC Performs Automated Reasoning in Chemistry) chemical reactivity models were extended to allow the calculation of alkaline hydrolysis rate constants of phosphate esters in water. The rate is calculated from the energy difference between the initial and transition state...

Bioabatement with hemicellulase supplementation to reduce enzymatic hydrolysis inhibitors

USDA-ARS?s Scientific Manuscript database

Removal of inhibitory compounds by bioabatement, combined with xylan hydrolysis, enables effective cellulose hydrolysis of pretreated corn stover, for fermentation of the sugars to fuel ethanol or other products. The fungus Coniochaeta ligniaria NRRL30616 eliminates most enzyme and fermentation inhi...

Species differences in the hydrolysis of 2-cyanoethylene oxide, the epoxide metabolite of acrylonitrile.

PubMed

Kedderis, G L; Batra, R

1993-04-01

The carcinogenic effects of acrylonitrile in rats are believed to be mediated by its DNA-reactive epoxide metabolite, 2-cyanoethylene oxide (CEO). Previous studies have shown that conjugation with glutathione is the major detoxication pathway for both acrylonitrile and CEO. This study investigated the role of epoxide hydrolase in the hydrolysis of CEO by HPLC analysis of the products from [2,3-14C]CEO. CEO is a relatively stable epoxide with a half-life of 99 min at 37 degrees C in sodium phosphate buffer (0.1 M), pH 7.3. Incubation with hepatic microsomes or cytosols from male F-344 rats or B6C3F1 mice did not enhance the rate of hydrolysis of CEO (0.69 nmol/min). Human hepatic microsomes significantly increased the rate of hydrolysis of CEO, whereas human hepatic cytosols did not. Human hepatic microsomal hydrolysis activity was heat-sensitive and potently inhibited by 1,1,1-trichloropropene oxide (IC50 of 23 microM), indicating that epoxide hydrolase was the catalyst. The hydrolysis of CEO catalyzed by hepatic microsomes from six individuals exhibited normal saturation kinetics with KM ranging from 0.6 to 3.2 mM and Vmax from 8.3 to 18.8 nmol hydrolysis products/min/mg protein. Pretreatment of rodents with phenobarbital or acetone induced hepatic microsomal hydrolysis activity toward CEO, whereas treatment with beta-naphthoflavone, dexamethasone or acrylonitrile itself was without effect. These data show that humans possess an additional detoxication pathway for CEO that is not active in rodents (but is inducible). The presence of an active epoxide hydrolase hydrolysis activity toward CEO in humans should be considered in assessments of cancer risk from acrylonitrile exposure.

Phenotype Overlap in Xylella fastidiosa Is Controlled by the Cyclic Di-GMP Phosphodiesterase Eal in Response to Antibiotic Exposure and Diffusible Signal Factor-Mediated Cell-Cell Signaling

PubMed Central

de Souza, Alessandra A.; Ionescu, Michael; Baccari, Clelia; da Silva, Aline M.

2013-01-01

Eal is an EAL domain protein in Xylella fastidiosa homologous to one involved in resistance to tobramycin in Pseudomonas aeruginosa. EAL and HD-GYP domain proteins are implicated in the hydrolysis of the secondary messenger bis-(3′-5′)-cyclic dimeric GMP (cyclic di-GMP). Cell density-dependent communication mediated by a Diffusible Signal Factor (DSF) also modulates cyclic di-GMP levels in X. fastidiosa, thereby controlling the expression of virulence genes and genes involved in insect transmission. The possible linkage of Eal to both extrinsic factors such as antibiotics and intrinsic factors such as quorum sensing, and whether both affect virulence, was thus addressed. Expression of eal was induced by subinhibitory concentrations of tobramycin, and an eal deletion mutant was more susceptible to this antibiotic than the wild-type strain and exhibited phenotypes similar to those of an rpfF deletion mutant blocked in DSF production, such as hypermotility, reduced biofilm formation, and hypervirulence to grape. Consistent with that, the rpfF mutant was more susceptible than the wild-type strain to tobramycin. Therefore, we propose that cell-cell communication and antibiotic stress can apparently lead to similar modulations of cyclic di-GMP in X. fastidiosa, resulting in similar phenotypes. However, the effect of cell density is dominant compared to that of antibiotic stress, since eal is suppressed by RpfF, which may prevent inappropriate behavioral changes in response to antibiotic stress when DSF accumulates. PMID:23542613

The origin of mouth-exhaled ammonia.

PubMed

Chen, W; Metsälä, M; Vaittinen, O; Halonen, L

2014-09-01

It is known that the oral cavity is a production site for mouth-exhaled NH3. However, the mechanism of NH3 production in the oral cavity has been unclear. Since bacterial urease in the oral cavity has been found to produce ammonia from oral fluid urea, we hypothesize that oral fluid urea is the origin of mouth-exhaled NH3. Our results show that under certain conditions a strong correlation exists between oral fluid urea and oral fluid ammonia (NH4(+)+NH3) (rs = 0.77, p < 0.001). We also observe a strong correlation between oral fluid NH3 and mouth-exhaled NH3 (rs = 0.81, p < 0.001). We conclude that three main factors affect the mouth-exhaled NH3 concentration: urea concentration, urease activity and oral fluid pH. Bacterial urease catalyses the hydrolysis of oral fluid urea to ammonia (NH4(+)+NH3). Oral fluid ammonia (NH4(+)+NH3) and pH determine the concentration of oral fluid NH3, which evaporates from oral fluid into gas phase and turns to mouth-exhaled NH3.

A mathematical model for the generation and control of a pH gradient in an immobilized enzyme system involving acid generation.

PubMed

Chen, G; Fournier, R L; Varanasi, S

1998-02-20

An optimal pH control technique has been developed for multistep enzymatic synthesis reactions where the optimal pH differs by several units for each step. This technique separates an acidic environment from a basic environment by the hydrolysis of urea within a thin layer of immobilized urease. With this technique, a two-step enzymatic reaction can take place simultaneously, in proximity to each other, and at their respective optimal pH. Because a reaction system involving an acid generation represents a more challenging test of this pH control technique, a number of factors that affect the generation of such a pH gradient are considered in this study. The mathematical model proposed is based on several simplifying assumptions and represents a first attempt to provide an analysis of this complex problem. The results show that, by choosing appropriate parameters, the pH control technique still can generate the desired pH gradient even if there is an acid-generating reaction in the system. Copyright 1998 John Wiley & Sons, Inc.

Biomass saccharification is largely enhanced by altering wall polymer features and reducing silicon accumulation in rice cultivars harvested from nitrogen fertilizer supply.

PubMed

Zahoor; Sun, Dan; Li, Ying; Wang, Jing; Tu, Yuanyuan; Wang, Yanting; Hu, Zhen; Zhou, Shiguang; Wang, Lingqiang; Xie, Guosheng; Huang, Jianliang; Alam, Aftab; Peng, Liangcai

2017-11-01

In this study, two rice cultivars were collected from experimental fields with seven nitrogen fertilizer treatments. All biomass samples contained significantly increased cellulose contents and reduced silica levels, with variable amounts of hemicellulose and lignin from different nitrogen treatments. Under chemical (NaOH, CaO, H 2 SO 4 ) and physical (hot water) pretreatments, biomass samples exhibited much enhanced hexoses yields from enzymatic hydrolysis, with high bioethanol production from yeast fermentation. Notably, both degree of polymerization (DP) of cellulose and xylose/arabinose (Xyl/Ara) ratio of hemicellulose were reduced in biomass residues, whereas other wall polymer features (cellulose crystallinity and monolignol proportion) were variable. Integrative analysis indicated that cellulose DP, hemicellulosic Xyl/Ara and silica are the major factors that significantly affect cellulose crystallinity and biomass saccharification. Hence, this study has demonstrated that nitrogen fertilizer supply could largely enhance biomass saccharification in rice cultivars, mainly by reducing cellulose DP, hemicellulosic Xyl/Ara and silica in cell walls. Copyright © 2017 Elsevier Ltd. All rights reserved.

Molecular Dynamics Simulations of Family 7 Cellobiohydrolase Mutants Aimed at Reducing Product Inhibition.

PubMed

Silveira, Rodrigo L; Skaf, Munir S

2015-07-23

Enzymatic conversion of lignocellulosic biomass into biofuels and chemicals constitutes a potential route for sustainable development. Cellobiohydrolases are key enzymes used in industrial cocktails for depolymerization of crystalline cellulose, and their mechanism of action has been intensely studied in the past several years. Provided with a tunnel-like substrate-binding cavity, cellobiohydrolases possess the ability to processively hydrolyze glycosidic bonds of crystalline cellulose, yielding one molecule of cellobiose per catalytic cycle. As such, cellobiose expulsion from the product binding site is a necessary step in order to allow for the processive hydrolysis mechanism. However, the high-affinity binding of cellobiose to the enzyme impairs the process and causes activity inhibition due to reaction products. Here, we use molecular dynamics simulations to study the binding of cellobiose to the Trichoderma reesei Cel7A (TrCel7A) cellobiohydrolase and the effects of mutations that reduce cellobiose binding, without affecting the structural and dynamical integrities of the enzyme. We observe that the product binding site exhibits an intrinsic flexibility that can sterically hinder cellobiose release. Several point mutations in the product binding site reduce cellobiose-enzyme interactions, but not all modifications are able to maintain the structural integrity of the enzyme. In particular, mutation of charged residues in the TrCel7A product binding site causes perturbations that affect the structure of the loops that form the substrate-binding tunnel of the enzyme and, hence, may affect TrCel7A function in other steps of the hydrolysis mechanism. Our results suggest there is a trade-off between product inhibition and catalytic efficiency, and they provide directions for cellulases engineering.

Effect of pretreatment severity in continuous steam explosion on enzymatic conversion of wheat straw: Evidence from kinetic analysis of hydrolysis time courses.

PubMed

Monschein, Mareike; Nidetzky, Bernd

2016-01-01

Focusing on continuous steam explosion, the influence of pretreatment severity due to varied acid loading on hydrolysis of wheat straw by Trichoderma reesei cellulases was investigated based on kinetic evaluation of the saccharification of each pretreated substrate. Using semi-empirical descriptors of the hydrolysis time course, key characteristics of saccharification efficiency were captured in a quantifiable fashion. Not only hydrolysis rates per se, but also the transition point of their bi-phasic decline was crucial for high saccharification degree. After 48h the highest saccharification was achieved for substrate pretreated at relatively low severity (1.2% acid). Higher severity increased enzyme binding to wheat straw, but reduced the specific hydrolysis rates. Higher affinity of the lignocellulosic material for cellulases does not necessarily result in increased saccharification, probably because of lignin modifications occurring at high pretreatment severities. At comparable severity, continuous pretreatment produced a substrate more susceptible to enzymatic hydrolysis than the batch process. Copyright © 2015 Elsevier Ltd. All rights reserved.

Two stage hydrolysis of corn stover at high solids content for mixing power saving and scale-up applications.

PubMed

Liu, Ke; Zhang, Jian; Bao, Jie

2015-11-01

A two stage hydrolysis of corn stover was designed to solve the difficulties between sufficient mixing at high solids content and high power input encountered in large scale bioreactors. The process starts with the quick liquefaction to convert solid cellulose to liquid slurry with strong mixing in small reactors, then followed the comprehensive hydrolysis to complete saccharification into fermentable sugars in large reactors without agitation apparatus. 60% of the mixing energy consumption was saved by removing the mixing apparatus in large scale vessels. Scale-up ratio was small for the first step hydrolysis reactors because of the reduced reactor volume. For large saccharification reactors in the second step, the scale-up was easy because of no mixing mechanism was involved. This two stage hydrolysis is applicable for either simple hydrolysis or combined fermentation processes. The method provided a practical process option for industrial scale biorefinery processing of lignocellulose biomass. Copyright © 2015 Elsevier Ltd. All rights reserved.

Hydrolysis of cellulose catalyzed by quaternary ammonium perrhenates in 1-allyl-3-methylimidazolium chloride.

PubMed

Wang, Jingyun; Zhou, Mingdong; Yuan, Yuguo; Zhang, Quan; Fang, Xiangchen; Zang, Shuliang

2015-12-01

Quaternary ammonium perrhenates were applied as catalyst to promote the hydrolysis of cellulose in 1-allyl-3-methylimidazolium chloride ([Amim]Cl). The quaternary ammonium perrhenates displayed good catalytic performance for cellulose hydrolysis. Water was also proven to be effective to promote cellulose hydrolysis. Accordingly, 97% of total reduced sugar (TRS) and 42% of glucose yields could be obtained under the condition of using 5mol% of tetramethyl ammonium perrhenate as catalyst, 70μL of water, ca. 0.6mmol of microcrystalline cellulose (MCC) and 2.0g of [Amim]Cl as solvent under microwave irradiation for 30min at 150°C (optimal conditions). The influence of quaternary ammonium cation on the efficiency of cellulose hydrolysis was examined based on different cation structures of perrhenates. The mechanism on perrhenate catalyzed cellulose hydrolysis is also discussed, whereas hydrogen bonding between ReO4 anion and hydroxyl groups of cellulose is assumed to be the key step for depolymerization of cellulose. Copyright © 2015. Published by Elsevier Ltd.

Effect of alkali lignins with different molecular weights from alkali pretreated rice straw hydrolyzate on enzymatic hydrolysis.

PubMed

Li, Yun; Qi, Benkun; Luo, Jianquan; Wan, Yinhua

2016-01-01

This study investigated the effect of alkali lignins with different molecular weights on enzymatic hydrolysis of lignocellulose. Different alkali lignins fractions, which were obtained from cascade ultrafiltration, were added into the dilute acid pretreated (DAP) and alkali pretreated (AP) rice straws respectively during enzymatic hydrolysis. The results showed that the addition of alkali lignins enhanced the hydrolysis and the enhancement for hydrolysis increased with increasing molecular weights of alkali lignins, with maximum enhancement being 28.69% for DAP and 20.05% for AP, respectively. The enhancement was partly attributed to the improved cellulase activity, and filter paper activity increased by 18.03% when adding lignin with highest molecular weight. It was found that the enhancement of enzymatic hydrolysis was correlated with the adsorption affinity of cellulase on alkali lignins, and the difference in surface charge and hydrophobicity of alkali lignins were responsible for the difference in affinity between cellulase and lignins. Copyright © 2015 Elsevier Ltd. All rights reserved.

Impact of electrical conductivity on acid hydrolysis of guar gum under induced electric field.

PubMed

Li, Dandan; Zhang, Yao; Yang, Na; Jin, Zhengyu; Xu, Xueming

2018-09-01

This study aimed to improve induced electric field (IEF)-assisted hydrolysis of polysaccharide by controlling electrical conductivity. As the conductivity of reaction medium was increased, the energy efficiency of IEF was increased because of deceased impedance, as well as enhanced output voltage and temperature, thus the hydrolysis of guar gum (GG) was accelerated under IEF. Changes in weight-average molecular weight (Mw) suggested that IEF-assisted hydrolysis of GG could be described by the first-order kinetics 1/Mw ∝ kt, with the rate constant (k), varying directly with the medium conductivity. Although IEF-assisted hydrolysis largely disrupted the morphological structure of GG, it had no impact on the chemical structure. In comparison to native GG, the steady shear viscosity of hydrolyzed GG dramatically declined while the thermal stability slightly decreased. This study extended the knowledge of electrical conductivity upon IEF-assisted acid hydrolysis of GG and might contribute to a better utilization of IEF for polysaccharide modification. Copyright © 2018 Elsevier Ltd. All rights reserved.

Enzymatic hydrolysis of pretreated waste paper--source of raw material for production of liquid biofuels.

PubMed

Brummer, Vladimir; Jurena, Tomas; Hlavacek, Viliam; Omelkova, Jirina; Bebar, Ladislav; Gabriel, Petr; Stehlik, Petr

2014-01-01

Enzymatic hydrolysis of waste paper is becoming a perspective way to obtain raw material for production of liquid biofuels. Reducing sugars solutions that arise from the process of saccharification are a precursors for following or simultaneous fermentation to ethanol. Different types of waste paper were evaluated, in terms of composition and usability, in order to select the appropriate type of the waste paper for the enzymatic hydrolysis process. Novozymes® enzymes NS50013 and NS50010 were used in a laboratory scale trials. Technological conditions, which seem to be the most suitable for hydrolysis after testing on cellulose pulp and filter paper, were applied to hydrolysis of widely available waste papers - offset paper, cardboard, recycled paper in two qualities, matte MYsol offset paper and for comparison again on model materials. The highest yields were achieved for the cardboard, which was further tested using various pretreatment combinations in purpose of increasing the hydrolysis yields. Copyright © 2013 Elsevier Ltd. All rights reserved.

Comparison of sulfuric and hydrochloric acids as catalysts in hydrolysis of Kappaphycus alvarezii (cottonii).

PubMed

Meinita, Maria Dyah Nur; Hong, Yong-Ki; Jeong, Gwi-Taek

2012-01-01

In this study, hydrolysis of marine algal biomass Kappaphhycus alvarezii using two different acid catalysts was examined with the goal of identifying optimal reaction conditions for the formation of sugars and by-products. K. alvarezii were hydrolyzed by autoclave using sulfuric acid or hydrochloric acid as catalyst with different acid concentrations (0.1-1.0 M), substrate concentrations (1.0-13.5%), hydrolysis time (10-90 min) and hydrolysis temperatures (100-130 (°)C). A difference in galactose, glucose, reducing sugar and total sugar content was observed under the different hydrolysis conditions. Different by-product compounds such as 5-hydroxymethylfurfural and levulinic acid were also observed under the different reaction conditions. The optimal conditions for hydrolysis were achieved at a sulfuric acid concentration, temperature and reaction time of 0.2 M, 130 °C and 15 min, respectively. These results may provide useful information for the development of more efficient systems for biofuel production from marine biomass.

Study of a specific lignin model: γ-oxidation and how it influences the hydrolysis efficiency of alcohol-aldehyde dehydrogenation copolymers.

PubMed

Bouxin, Florent; Baumberger, Stéphanie; Renault, Jean-Hugues; Dole, Patrice

2011-05-01

Six coniferyl alcohol-coniferaldehyde dehydrogenation copolymers (DHcoPs) were synthesized in order to determine the influence of an increased number of aldehyde functions on hydrolysis. After heterogeneous hydrolysis using acidic Montmorillonite K10 clay, the DHcoPs were thioacidolyzed and analyzed by gel permeation chromatography (GPC). Comparison of the thioacidolyzed products, with and without the hydrolysis step, showed that there was a greater proportion of condensation reaction in the absence of aldehyde. When the coniferaldehyde content in the initial synthetic mixture was more than 30% (w/w), only a low fraction of condensed products was generated during the K10 clay hydrolysis step. This suggests that condensation pathways are mainly due to the alcohol present in the γ-position in the DHcoPs. Investigation of the reactivity and the potential condensation of aldehyde and alcohol monomers under hydrolysis conditions showed the important conversion of coniferyl alcohol and conversely the stability of coniferaldehyde. Copyright © 2011 Elsevier Ltd. All rights reserved.

Factors that influence properties of FOG deposits and their formation in sewer collection systems.

PubMed

Iasmin, Mahbuba; Dean, Lisa O; Lappi, Simon E; Ducoste, Joel J

2014-02-01

Understanding the formation of Fat, Oil, and Grease (FOG) deposits in sewer systems is critical to the sustainability of sewer collection systems since they have been implicated in causing sewerage blockages that leads to sanitary sewer overflows (SSOs). Recently, FOG deposits in sewer systems displayed strong similarities with calcium-based fatty acid salts as a result of a saponification reaction. The objective of this study was to quantify the factors that may affect the formation of FOG deposits and their chemical and rheological properties. These factors included the types of fats used in FSEs, environmental conditions (i.e. pH and temperature), and the source of calcium in sewer systems. The results of this study showed that calcium content in the calcium based salts seemed to depend on the solubility limit of the calcium source and influenced by pH and temperature conditions. The fatty acid profile of the calcium-based fatty acid salts produced under alkali driven hydrolysis were identical to the profile of the fat source and did not match the profile of field FOG deposits, which displayed a high fraction of palmitic, a long chain saturated fatty acid. It is hypothesized that selective microbial metabolism of fats and/or biologically induced hydrogenation may contribute to the FOG deposit makeup in sewer system. Therefore, selective removal of palmitic in pretreatment processes may be necessary prior to the discharge of FSE wastes into the sewer collection system. Copyright © 2013 Elsevier Ltd. All rights reserved.

Pseudomonas aeruginosa arylsulfatase: a purified enzyme for the mild hydrolysis of steroid sulfates.

PubMed

Stevenson, Bradley J; Waller, Christopher C; Ma, Paul; Li, Kunkun; Cawley, Adam T; Ollis, David L; McLeod, Malcolm D

2015-10-01

The hydrolysis of sulfate ester conjugates is frequently required prior to analysis for a range of analytical techniques including gas chromatography-mass spectrometry (GC-MS). Sulfate hydrolysis may be achieved with commercial crude arylsulfatase enzyme preparations such as that derived from Helix pomatia but these contain additional enzyme activities such as glucuronidase, oxidase, and reductase that make them unsuitable for many analytical applications. Strong acid can also be used to hydrolyze sulfate esters but this can lead to analyte degradation or increased matrix interference. In this work, the heterologously expressed and purified arylsulfatase from Pseudomonas aeruginosa is shown to promote the mild enzyme-catalyzed hydrolysis of a range of steroid sulfates. The substrate scope of this P. aeruginosa arylsulfatase hydrolysis is compared with commercial crude enzyme preparations such as that derived from H. pomatia. A detailed kinetic comparison is reported for selected examples. Hydrolysis in a urine matrix is demonstrated for dehydroepiandrosterone 3-sulfate and epiandrosterone 3-sulfate. The purified P. aeruginosa arylsulfatase contains only sulfatase activity allowing for the selective hydrolysis of sulfate esters in the presence of glucuronide conjugates as demonstrated in the short three-step chemoenzymatic synthesis of 5α-androstane-3β,17β-diol 17-glucuronide (ADG, 1) from epiandrosterone 3-sulfate. The P. aeruginosa arylsulfatase is readily expressed and purified (0.9 g per L of culture) and thus provides a new and selective method for the hydrolysis of steroid sulfate esters in analytical sample preparation. Copyright © 2015 John Wiley & Sons, Ltd.

Differential effects of pertussis toxin on insulin-stimulated phosphatidylcholine hydrolysis and glycerolipid synthesis de novo. Studies in BC3H-1 myocytes and rat adipocytes

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

Hoffman, J.M.; Standaert, M.L.; Nair, G.P.

1991-04-02

Insulin-induced increases in diacylglycerol (DAG) have been suggested to result from stimulation of de novo phosphatidic acid (PA) synthesis and phosphatidylcholine (PC) hydrolysis. Presently, the authors found that insulin decreased PC levels of BC3H-1 myocytes and rat adipocytes by approximately 10-25% within 30 s. These decreases were rapidly reversed in both cell types, apparently because of increased PC synthesis de novo. In BC3H-1 myocytes, pertussis toxin inhibited PC resynthesis and insulin effects on the pathway of de novo PA-DAG-PC synthesis, as evidenced by changes in ({sup 3}H)glycerol incorporation, but did not inhibit insulin-stimulated PC hydrolysis. Pertussis toxin also blocked themore » later, but not the initial, increase in DAG production in the myocytes. Phorbol esters activated PC hydrolysis in both myocytes and adipocytes, but insulin-induced stimulation of PC hydrolysis was not dependent upon activation of PKC, since this hydrolysis was not inhibited by 500 {mu}M sangivamycin, an effective PKC inhibitor. The results indicate that insulin increases DAG by pertussis toxin sensitive and insensitive (PC hydrolysis) mechanisms, which are mechanistically separate, but functionally interdependent and integrated. PC hydrolysis may contribute importantly to initial increases in DAG, but later sustained increases are apparently largely dependent on insulin-induced stimulation of the pathway of de novo phospholipid synthesis.« less

Combined heat treatment and acid hydrolysis of cassava grate waste (CGW) biomass for ethanol production

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

Agu, R.C.; Amadife, A.E.; Ude, C.M.

1997-12-31

The effect of combined heat treatment and acid hydrolysis (various concentrations) on cassava grate waste (CGW) biomass for ethanol production was investigated. At high concentrations of H{sub 2}SO{sub 4} (1--5 M), hydrolysis of the CGW biomass was achieved but with excessive charring or dehydration reaction. At lower acid concentrations, hydrolysis of CGW biomass was also achieved with 0.3--0.5 M H{sub 2}SO{sub 4}, while partial hydrolysis was obtained below 0.3 M H{sub 2}SO{sub 4} (the lowest acid concentration that hydrolyzed CGW biomass) at 120 C and 1 atm pressure for 30 min. A 60% process efficiency was achieved with 0.3 Mmore » H{sub 2}SO{sub 4} in hydrolyzing the cellulose and lignin materials present in the CGW biomass. High acid concentration is therefore not required for CGW biomass hydrolysis. The low acid concentration required for CGW biomass hydrolysis, as well as the minimal cost required for detoxification of CGW biomass because of low hydrogen cyanide content of CGW biomass would seem to make this process very economical. From three liters of the CGW biomass hydrolysate obtained from hydrolysis with 0.3M H{sub 2}SO{sub 4}, ethanol yield was 3.5 (v/v%) after yeast fermentation. However, although the process resulted in gainful utilization of CGW biomass, additional costs would be required to effectively dispose new by-products generated from CGW biomass processing.« less

Towards the design of organocatalysts for nerve agents remediation: The case of the active hydrolysis of DCNP (a Tabun mimic) catalyzed by simple amine-containing derivatives.

PubMed

Barba-Bon, Andrea; Martínez-Máñez, Ramón; Sancenón, Félix; Costero, Ana M; Gil, Salvador; Pérez-Pla, Francisco; Llopis, Elisa

2015-11-15

We report herein a study of the hydrolysis of Tabun mimic DCNP in the presence of different amines, aminoalcohols and glycols as potential suitable organocatalysts for DCNP degradation. Experiments were performed in CD3CN in the presence of 5% D2O, which is a suitable solvent mixture to follow the DCNP hydrolysis. These studies allowed the definition of different DCNP depletion paths, resulting in the formation of diethylphosphoric acid, tetraethylpyrophosphate and phosphoramide species as final products. Without organocatalysts, DCNP hydrolysis occurred mainly via an autocatalysis path. Addition of tertiary amines in sub-stoichiometric amounts largely enhanced DCNP depletion whereas non-tertiary polyamines reacted even faster. Glycols induced very slight increment in the DCNP hydrolysis, whereas DCNP hydrolysis increased sharply in the presence of certain aminoalcohols especially, 2-(2-aminoethylamino)ethanol. For the latter compound, DCNP depletion occurred ca. 80-fold faster than in the absence of organocatalysts. The kinetic studies revealed that DCNP hydrolysis in the presence of 2-(2-aminoethylamino)ethanol occurred via a catalytic process, in which the aminoalcohol was involved. DCNP hydrolysis generally depended strongly on the structure of the amine, and it was found that the presence of the OHCH2CH2N moiety in the organocatalyst structure seems important to induce a fast degradation of DCNP. Copyright © 2015 Elsevier B.V. All rights reserved.

High yield hydrolysis of seaweed-waste biomass using peracetic acid and ionic liquid treatments

NASA Astrophysics Data System (ADS)

Uju, Wijayanta, Agung Tri; Goto, Masahiro; Kamiya, Noriho

2018-02-01

Seaweed is one of the most promising bioethanol feedstocks. This water plant has high carbohydrate content but low lignin content, as a result it will be easier to be hydrolysed. This paper described hydrolysis of seaweed-waste biomass from the carrageenan (SWBC) industry using enzymatic saccharification or ionic liquids-HCl hydrolysis. In the first work, SWBC pretreated by peracetic acid (PAA) followed by ionic liquid (IL) caused enhance the cellulose conversion of enzymatic saccharification. At 48h saccharification, the value conversion almost reached 100%. In addition, the untreated SWBC also produced the cellulose conversion 77%. In the second work, SWBC or Bagasse with or without pretreated by PAA was hydrolyzed using ILs-HCl hydrolysis. The ILs used were 1-buthyl-3-methylpyridium chloride, [Bmpy][Cl] and 1-butyl-3-metyl imidazolium chloride ([Bmim][Cl]). [Bmpy][Cl]-HCl hydrolysis produced higher cellulose conversion than [Bmim][Cl]-HCl hydrolysis. The phenomenon was clearly observed on the Bagasse, which without pretreated by PAA. Furthermore, SWBC hydrolyzed by both ILs in the presence low concentration of HCl produced cellulose conversion 70-98% at 60-90 min of hydrolysis time. High cellulose conversion of SWBC on the both hydrolysis was caused by SWBC had the low lignin (4%). Moreover, IL treatments caused lowering of cellulose hydrogen bonds or even changed the cellulose characteristics from cellulose I to cellulose II which easily to be hydrolyzed. In the case of [Bmpy][Cl], this IL may reduce the degree polymerization of celluloses.

The role of pH on the thermodynamics and kinetics of muscle biochemistry: an in vivo study by (31)P-MRS in patients with myo-phosphorylase deficiency.

PubMed

Malucelli, E; Iotti, S; Manners, D N; Testa, C; Martinuzzi, A; Barbiroli, B; Lodi, R

2011-09-01

In this study we assessed ΔG'(ATP) hydrolysis, cytosolic [ADP], and the rate of phosphocreatine recovery using Phosphorus Magnetic Resonance Spectroscopy in the calf muscle of a group of patients affected by glycogen myo-phosphorylase deficiency (McArdle disease). The goal was to ascertain whether and to what extent the deficit of the glycogenolytic pathway would affect the muscle energy balance. A typical feature of this pathology is the lack of intracellular acidosis. Therefore we posed the question of whether, in the absence of pH decrease, the rate of phosphocreatine recovery depends on the amount of phosphocreatine consumed during exercise. Results showed that at the end of exercise both [ADP] and ΔG'(ATP) of patients were significantly higher than those of matched control groups reaching comparable levels of phosphocreatine concentration. Furthermore, in these patients we found that the rate of phosphocreatine recovery is not influenced by the amount of phosphocreatine consumed during exercise. These outcomes provide experimental evidence that: i) the intracellular acidification occurring in exercising skeletal muscle is a protective factor for the energy consumption; and ii) the influence of pH on the phosphocreatine recovery rate is at least in part related to the kinetic mechanisms of mitochondrial creatine kinase enzyme. 2011 Elsevier B.V. All rights reserved.

The Effect of Ionic Liquid Pretreatment on the Bioconversion of Tomato Processing Waste to Fermentable Sugars and Biogas.

PubMed

Allison, Brittany J; Cádiz, Juan Canales; Karuna, Nardrapee; Jeoh, Tina; Simmons, Christopher W

2016-08-01

Tomato pomace is an abundant lignocellulosic waste stream from industrial tomato processing and therefore a potential feedstock for production of renewable biofuels. However, little research has been conducted to determine if pretreatment can enhance release of fermentable sugars from tomato pomace. Ionic liquids (ILs) are an emerging pretreatment technology for lignocellulosic biomass to increase enzymatic digestibility and biofuel yield while utilizing recyclable chemicals with low toxicity. In this study, pretreatment of tomato pomace with the ionic liquid 1-ethyl-3-methylimidazolium acetate ([C2mim][OAc]) was investigated. Changes in pomace enzymatic digestibility were affected by pretreatment time and temperature. Certain pretreatment conditions significantly improved reducing sugar yield and hydrolysis time compared to untreated pomace. Compositional analyses suggested that pretreatment primarily removed water-soluble compounds and enriched for lignocellulose in pomace, with only subtle changes to the composition of the lignocellulose. While tomato pomace was effectively pretreated with [C2mim][OAc] to improve enzymatic digestibility, as of yet, unknown factors in the pomace caused ionic liquid pretreatment to negatively affect anaerobic digestion of pretreated material. This result, which is unique compared to similar studies on IL pretreatment of grasses and woody biomass, highlights the need for additional research to determine how the unique chemical composition of tomato pomace and other lignocellulosic fruit residues may interact with ionic liquids to generate inhibitors for downstream fermentation to biofuels.

Use of an algal hydrolysate to improve enzymatic hydrolysis of anaerobically digested fiber

USDA-ARS?s Scientific Manuscript database

This study investigated the use of acid hydrolyzed algae to enhance the enzymatic hydrolysis of cellulosic biomass. We first characterized wastewater-grown algal samples and determined the optimal conditions (acid concentration, reaction temperature, and reaction time) for algal hydrolysis using di...

ESTIMATION OF PHOSPHATE ESTER HYDROLYSIS RATE CONSTANTS. II. ACID AND GENERAL BASE CATALYZED HYDROLYSIS

EPA Science Inventory

SPARC (SPARC Performs Automated Reasoning in Chemistry) chemical reactivity models were extended to calculate acid and neutral hydrolysis rate constants of phosphate esters in water. The rate is calculated from the energy difference between the initial and transition states of a ...

Bioconversion of beetle-killed lodgepole pine using SPORL: Process scale-up design, lignin co-product, and high solids fermentation without detoxification

USDA-ARS?s Scientific Manuscript database

Mountain pine beetle killed Lodgepole pine (Pinus contorta Douglas ex Loudon) wood chips were pretreated using an acidic sulfite solution of approximately pH = 2.0 at a liquor to wood ratio of 3 and sodium bisulfite loading of 8 wt % on wood. The combined hydrolysis factor (CHF), formulated from rea...

Observations on the action of limit dextrinases on amylopectin-like polysaccharides (Short Communication)

PubMed Central

Dunn, Graham; Hardie, D. Grahame; Manners, David J.

1973-01-01

The rate of hydrolysis of amylopectin by three different limit dextrinase preparations is only about 15–23% of that of amylopectin β-limit dextrin under similar conditions. On dilution of the enzymes there was no change in specificity. The factors controlling the specificity of the enzyme and the possible significance in vivo of the results are discussed. PMID:4734116

Molecular Level Understanding of the Factors Affecting the Stability of Dimethoxy Benzene Catholyte Candidates from First-Principles Investigations

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

Assary, Rajeev S.; Zhang, Lu; Huang, Jinhua

First-principles simulations are performed to gain molecular level insights into the factors affecting the stability of seven 1,4-dimethoxybenzene (DMB) derivatives. These molecules are potential catholyte candidates for nonaqueous redox flow battery systems. Computations are performed to predict oxidation potentials in various dielectric mediums, intrinsic-reorganization energies, and structural changes of these representative catholyte molecules during the redox process. In order to understand the stability of the DMB-based radical cations, the thermodynamic feasibility of the following reactions is computed using density functional theory: (a) deprotonation, (b) dimerization, (c) hydrolysis, and (d) demethylation. The computations indicate that radical cations of the 2,3-dimethyl andmore » 2,5-dimethyl derivatives are the most stable among the DMB derivatives considered in this study. In the presence of solvents with high-proton solvating ability (water, DMSO, acetonitrile), degradation of cation radical occurring via deprotonation is the most likely mechanism. In the presence of solvents such as propylene carbonate (PC), demethylation was found to be the most likely reaction that causes degradation of radical cations. From the computed enthalpy of activation (Delta H-double dagger) for a demethylation reaction in PC, the 2,5-dimethyl DMB cation radical would exhibit better kinetic stability in comparison to the other candidates. Finally, this investigation suggests that computational studies of structural properties such as redox potentials, reorganization energies, and the computed reaction energetics (deprotonation and demethylation) of charged species can be used to predict the relative stability of a large set of molecules required for the discovery of novel redox active materials for flow battery applications« less

Molecular Level Understanding of the Factors Affecting the Stability of Dimethoxy Benzene Catholyte Candidates from First-Principles Investigations

DOE PAGES

Assary, Rajeev S.; Zhang, Lu; Huang, Jinhua; ...

2016-06-14

First-principles simulations are performed to gain molecular level insights into the factors affecting the stability of seven 1,4-dimethoxybenzene (DMB) derivatives. These molecules are potential catholyte candidates for nonaqueous redox flow battery systems. Computations are performed to predict oxidation potentials in various dielectric mediums, intrinsic-reorganization energies, and structural changes of these representative catholyte molecules during the redox process. In order to understand the stability of the DMB-based radical cations, the thermodynamic feasibility of the following reactions is computed using density functional theory: (a) deprotonation, (b) dimerization, (c) hydrolysis, and (d) demethylation. The computations indicate that radical cations of the 2,3-dimethyl andmore » 2,5-dimethyl derivatives are the most stable among the DMB derivatives considered in this study. In the presence of solvents with high-proton solvating ability (water, DMSO, acetonitrile), degradation of cation radical occurring via deprotonation is the most likely mechanism. In the presence of solvents such as propylene carbonate (PC), demethylation was found to be the most likely reaction that causes degradation of radical cations. From the computed enthalpy of activation (Delta H-double dagger) for a demethylation reaction in PC, the 2,5-dimethyl DMB cation radical would exhibit better kinetic stability in comparison to the other candidates. Finally, this investigation suggests that computational studies of structural properties such as redox potentials, reorganization energies, and the computed reaction energetics (deprotonation and demethylation) of charged species can be used to predict the relative stability of a large set of molecules required for the discovery of novel redox active materials for flow battery applications« less