Reversible inactivation of CO dehydrogenase with thiol compounds

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

Kreß, Oliver; Gnida, Manuel; Pelzmann, Astrid M.

2014-05-09

Highlights: • Rather large thiols (e.g. coenzyme A) can reach the active site of CO dehydrogenase. • CO- and H{sub 2}-oxidizing activity of CO dehydrogenase is inhibited by thiols. • Inhibition by thiols was reversed by CO or upon lowering the thiol concentration. • Thiols coordinate the Cu ion in the [CuSMo(=O)OH] active site as a third ligand. - Abstract: Carbon monoxide dehydrogenase (CO dehydrogenase) from Oligotropha carboxidovorans is a structurally characterized member of the molybdenum hydroxylase enzyme family. It catalyzes the oxidation of CO (CO + H{sub 2}O → CO{sub 2} + 2e{sup −} + 2H{sup +}) which proceedsmore » at a unique [CuSMo(=O)OH] metal cluster. Because of changing activities of CO dehydrogenase, particularly in subcellular fractions, we speculated whether the enzyme would be subject to regulation by thiols (RSH). Here we establish inhibition of CO dehydrogenase by thiols and report the corresponding K{sub i}-values (mM): L-cysteine (5.2), D-cysteine (9.7), N-acetyl-L-cysteine (8.2), D,L-homocysteine (25.8), L-cysteine–glycine (2.0), dithiothreitol (4.1), coenzyme A (8.3), and 2-mercaptoethanol (9.3). Inhibition of the enzyme was reversed by CO or upon lowering the thiol concentration. Electron paramagnetic resonance spectroscopy (EPR) and X-ray absorption spectroscopy (XAS) of thiol-inhibited CO dehydrogenase revealed a bimetallic site in which the RSH coordinates to the Cu-ion as a third ligand ([Mo{sup VI}(=O)OH{sub (2)}SCu{sup I}(SR)S-Cys]) leaving the redox state of the Cu(I) and the Mo(VI) unchanged. Collectively, our findings establish a regulation of CO dehydrogenase activity by thiols in vitro. They also corroborate the hypothesis that CO interacts with the Cu-ion first. The result that thiol compounds much larger than CO can freely travel through the substrate channel leading to the bimetallic cluster challenges previous concepts involving chaperone function and is of importance for an understanding how the sulfuration

Oral-Fluid Thiol-Detection Test Identifies Underlying Active Periodontal Disease Not Detected by the Visual Awake Examination.

PubMed

Queck, Katherine E; Chapman, Angela; Herzog, Leslie J; Shell-Martin, Tamara; Burgess-Cassler, Anthony; McClure, George David

Periodontal disease in dogs is highly prevalent but can only be accurately diagnosed by performing an anesthetized oral examination with periodontal probing and dental radiography. In this study, 114 dogs had a visual awake examination of the oral cavity and were administered an oral-fluid thiol-detection test prior to undergoing a a full-mouth anesthetized oral examination and digital dental radiographs. The results show the visual awake examination underestimated the presence and severity of active periodontal disease. The thiol-detection test was superior to the visual awake examination at detecting the presence and severity of active periodontal disease and was an indicator of progression toward alveolar bone loss. The thiol-detection test detected active periodontal disease at early stages of development, before any visual cues were present, indicating the need for intervention to prevent periodontal bone loss. Early detection is important because without intervention, dogs with gingivitis (active periodontal disease) progress to irreversible periodontal bone loss (stage 2+). As suggested in the current AAHA guidelines, a thiol-detection test administered in conjunction with the visual awake examination during routine wellness examinations facilitates veterinarian-client communication and mitigates under-diagnosis of periodontal disease and underutilization of dental services. The thiol-detection test can be used to monitor the periodontal health status of the conscious patient during follow-up examinations based on disease severity.

Determination of thiol metabolites in human urine by stable isotope labeling in combination with pseudo-targeted mass spectrometry analysis

NASA Astrophysics Data System (ADS)

Liu, Ping; Qi, Chu-Bo; Zhu, Quan-Fei; Yuan, Bi-Feng; Feng, Yu-Qi

2016-02-01

Precursor ion scan and multiple reaction monitoring scan (MRM) are two typical scan modes in mass spectrometry analysis. Here, we developed a strategy by combining stable isotope labeling (IL) with liquid chromatography-mass spectrometry (LC-MS) under double precursor ion scan (DPI) and MRM for analysis of thiols in 5 types of human cancer urine. Firstly, the IL-LC-DPI-MS method was applied for non-targeted profiling of thiols from cancer samples. Compared to traditional full scan mode, the DPI method significantly improved identification selectivity and accuracy. 103 thiol candidates were discovered in all cancers and 6 thiols were identified by their standards. It is worth noting that pantetheine, for the first time, was identified in human urine. Secondly, the IL-LC-MRM-MS method was developed for relative quantification of thiols in cancers compared to healthy controls. All the MRM transitions of light and heavy labeled thiols were acquired from urines by using DPI method. Compared to DPI method, the sensitivity of MRM improved by 2.1-11.3 folds. In addition, the concentration of homocysteine, γ-glutamylcysteine and pantetheine enhanced more than two folds in cancer patients compared to healthy controls. Taken together, the method demonstrated to be a promising strategy for identification and comprehensive quantification of thiols in human urines.

Determination of thiol metabolites in human urine by stable isotope labeling in combination with pseudo-targeted mass spectrometry analysis

PubMed Central

Liu, Ping; Qi, Chu-Bo; Zhu, Quan-Fei; Yuan, Bi-Feng; Feng, Yu-Qi

2016-01-01

Precursor ion scan and multiple reaction monitoring scan (MRM) are two typical scan modes in mass spectrometry analysis. Here, we developed a strategy by combining stable isotope labeling (IL) with liquid chromatography-mass spectrometry (LC-MS) under double precursor ion scan (DPI) and MRM for analysis of thiols in 5 types of human cancer urine. Firstly, the IL-LC-DPI-MS method was applied for non-targeted profiling of thiols from cancer samples. Compared to traditional full scan mode, the DPI method significantly improved identification selectivity and accuracy. 103 thiol candidates were discovered in all cancers and 6 thiols were identified by their standards. It is worth noting that pantetheine, for the first time, was identified in human urine. Secondly, the IL-LC-MRM-MS method was developed for relative quantification of thiols in cancers compared to healthy controls. All the MRM transitions of light and heavy labeled thiols were acquired from urines by using DPI method. Compared to DPI method, the sensitivity of MRM improved by 2.1–11.3 folds. In addition, the concentration of homocysteine, γ-glutamylcysteine and pantetheine enhanced more than two folds in cancer patients compared to healthy controls. Taken together, the method demonstrated to be a promising strategy for identification and comprehensive quantification of thiols in human urines. PMID:26888486

The yeast Hsp70 Ssa1 is a sensor for activation of the heat shock response by thiol-reactive compounds

PubMed Central

Wang, Yanyu; Gibney, Patrick A.; West, James D.; Morano, Kevin A.

2012-01-01

The heat shock transcription factor HSF1 governs the response to heat shock, oxidative stresses, and xenobiotics through unknown mechanisms. We demonstrate that diverse thiol-reactive molecules potently activate budding yeast Hsf1. Hsf1 activation by thiol-reactive compounds is not consistent with the stresses of misfolding of cytoplasmic proteins or cytotoxicity. Instead, we demonstrate that the Hsp70 chaperone Ssa1, which represses Hsf1 in the absence of stress, is hypersensitive to modification by a thiol-reactive probe. Strikingly, mutation of two conserved cysteine residues to serine in Ssa1 rendered cells insensitive to Hsf1 activation and subsequently induced thermotolerance by thiol-reactive compounds, but not by heat shock. Conversely, substitution with the sulfinic acid mimic aspartic acid resulted in constitutive Hsf1 activation. Cysteine 303, located within the nucleotide-binding domain, was found to be modified in vivo by a model organic electrophile, demonstrating that Ssa1 is a direct target for thiol-reactive molecules through adduct formation. These findings demonstrate that Hsp70 is a proximal sensor for Hsf1-mediated cytoprotection and can discriminate between two distinct environmental stressors. PMID:22809627

1,2,4-triazole derivative with Schiff base; thiol-thione tautomerism, DFT study and antileishmanial activity

NASA Astrophysics Data System (ADS)

Süleymanoğlu, Nevin; Ustabaş, Reşat; Direkel, Şahin; Alpaslan, Yelda Bingöl; Ünver, Yasemin

2017-12-01

Thiol-thione tautomerism of 1,2,4-triazole derivative with Schiff base was investigated by spectroscopic methods and quantum mechanical calculations. Theoretical study of thiol-thione tautomeric forms of 1,2,4-triazole derivative with Schiff base; 1,2,4-triazole-thiol form, 1-((5-mercapto-4-(thiophene-2-ylmethyleneamino)-4H-1,2,4-triazole-3-yl)methyl)-3-(thiophene-2-ylmethyl)-4-(thiophene-2-ylmethyleneamino)-1H-1,2,4-triazole-5(4H)-one (I) and 1,2,4-triazole-thione form, 3-(thiophene-2-ylmethyl)-4-(thiophene-2-ylmethyleneamino)-1-((4-(thiophene-2-ylmethyleneamino)-5-thioxo-4,5-dihydro-1H-1,2,4-triazole-3-yl)methyl)-1H-1,2,4-triazole-5(4H)-one (II) was performed by the density functional theory (DFT) method with 6-311++G(d,p) basis set. Structural parameters were obtained and spectral parameters of NMR, FTIR and UV-vis were compared with experimental ones to determine structural details. In vitro antileishmanial activity was studied against Leishmania infantum promastigots by microdilution broth assay with Alamar Blue Dye. The results indicate that 1,2,4-triazole derivative exists in both thiol and thione form and, can be evaluated as antiparasitic in term of antileishmanial activity.

Jack bean urease: the effect of active-site binding inhibitors on the reactivity of enzyme thiol groups.

PubMed

Krajewska, Barbara; Zaborska, Wiesława

2007-10-01

In view of the complexity of the role of the active site flap cysteine in the urease catalysis, in this work we studied how the presence of typical active-site binding inhibitors of urease, phenylphosphorodiamidate (PPD), acetohydroxamic acid (AHA), boric acid and fluoride, affects the reactivity of enzyme thiol groups, the active site flap thiol in particular. For that the inhibitor-urease complexes were prepared with excess inhibitors and had their thiol groups titrated with DTNB. The effects observed were analyzed in terms of the structures of the inhibitor-urease complexes reported in the literature. We found that the effectiveness in preventing the active site cysteine from the modification by disulfides, varied among the inhibitors studied, even though they all bind to the active site. The variations were accounted for by different extents of geometrical distortion in the active site that the inhibitors introduced upon binding, leaving the flap either open in AHA-, boric acid- and fluoride-inhibited urease, like in the native enzyme or closed in PPD-inhibited urease. Among the inhibitors, only PPD was found to be able to thoroughly protect the flap cysteines from the further reaction with disulfides, this apparently resulting from the closed conformation of the flap. Accordingly, in practical terms PPD may be regarded as the most suitable inhibitor for active-site protection experiments in inhibition studies of urease.

Improvement of aromatic thiol release through the selection of yeasts with increased β-lyase activity.

PubMed

Belda, Ignacio; Ruiz, Javier; Navascués, Eva; Marquina, Domingo; Santos, Antonio

2016-05-16

The development of a selective medium for the rapid differentiation of yeast species with increased aromatic thiol release activity has been achieved. The selective medium was based on the addition of S-methyl-l-cysteine (SMC) as β-lyase substrate. In this study, a panel of 245 strains of Saccharomyces cerevisiae strains was tested for their ability to grow on YCB-SMC medium. Yeast strains with an increased β-lyase activity grew rapidly because of their ability to release ammonium from SMC in comparison to others, and allowed for the easy isolation and differentiation of yeasts with promising properties in oenology, or another field, for aromatic thiol release. The selective medium was also helpful for the discrimination between those S. cerevisiae strains, which present a common 38-bp deletion in the IRC7 sequence (present in around 88% of the wild strains tested and are likely to be less functional for 4-mercapto-4-methylpentan-2-one (4MMP) production), and those S. cerevisiae strains homozygous for the full-length IRC7 allele. The medium was also helpful for the selection of non-Saccharomyces yeasts with increased β-lyase activity. Based on the same medium, a highly sensitive, reproducible and non-expensive GC-MS method for the evaluation of the potential volatile thiol release by different yeast isolates was developed. Copyright © 2016 Elsevier B.V. All rights reserved.

Thiol/disulphide homeostasis in celiac disease

PubMed Central

Kaplan, Mustafa; Ates, Ihsan; Yuksel, Mahmut; Ozderin Ozin, Yasemin; Alisik, Murat; Erel, Ozcan; Kayacetin, Ertugrul

2017-01-01

AIM To determine dynamic thiol/disulphide homeostasis in celiac disease and to examine the associate with celiac autoantibodies and gluten-free diet. METHODS Seventy three patients with celiac disease and 73 healthy volunteers were enrolled in the study. In both groups, thiol/disulphide homeostasis was examined with a new colorimetric method recently developed by Erel and Neselioglu. RESULTS In patients with celiac disease, native thiol (P = 0.027) and total thiol (P = 0.031) levels were lower, while disulphide (P < 0.001) level, disulphide/native thiol (P < 0.001) and disulphide/total thiol (P < 0.001) ratios were higher compared to the control group. In patients who do not comply with a gluten-free diet, disulphide/native thiol ratio was found higher compared to the patients who comply with the diet (P < 0.001). In patients with any autoantibody-positive, disulphide/native thiol ratio was observed higher compared to the patients with autoantibody-negative (P < 0.05). It is found that there is a negative correlation between celiac autoantibodies, and native thiol, total thiol levels and native thiol/total thiol ratio, while a positive correlation is observed between disulphide, disulphide/native thiol and disulphide/total thiol levels. CONCLUSION This study is first in the literature which found that the patients with celiac disease the dynamic thiol/disulphide balance shifts through disulphide form compared to the control group. PMID:28533921

Soybean P34 Probable Thiol Protease Probably Has Proteolytic Activity on Oleosins.

PubMed

Zhao, Luping; Kong, Xiangzhen; Zhang, Caimeng; Hua, Yufei; Chen, Yeming

2017-07-19

P34 probable thiol protease (P34) and Gly m Bd 30K (30K) show high relationship with the protease of 24 kDa oleosin of soybean oil bodies. In this study, 9 day germinated soybean was used to separate bioprocessed P34 (P32) from bioprocessed 30K (28K). Interestingly, P32 existed as dimer, whereas 28K existed as monomer; a P32-rich sample had proteolytic activity and high cleavage site specificity (Lys-Thr of 24 kDa oleosin), whereas a 28K-rich sample showed low proteolytic activity; the P32-rich sample contained one thiol protease. After mixing with purified oil bodies, all P32 dimers were dissociated and bound to 24 kDa oleosins to form P32-24 kDa oleosin complexes. By incubation, 24 kDa oleosin was preferentially hydrolyzed, and two hydrolyzed products (HPs; 17 and 7 kDa) were confirmed. After most of 24 kDa oleosin was hydrolyzed, some P32 existed as dimer, and the other as P32-17 kDa HP. It was suggested that P32 was the protease.

Thiols in the alphaIIbbeta3 integrin are necessary for platelet aggregation.

PubMed

Manickam, Nagaraj; Sun, Xiuhua; Hakala, Kevin W; Weintraub, Susan T; Essex, David W

2008-07-01

Sulfhydryl groups of platelet surface proteins are important in platelet aggregation. While p-chloromercuribenzene sulphonate (pCMBS) has been used in most studies on platelet surface thiols, the specific thiol-proteins that pCMBS reacts with to inhibit aggregation have not been well defined. Since the thiol-containing P2Y(12) ADP receptor is involved in most types of platelet aggregation, we used the ADP scavenger apyrase and the P2Y(12) receptor antagonist 2-MeSAMP to examine thiol-dependent reactions in the absence of contributions from this receptor. We provide evidence for a non-P2Y(12) thiol-dependent reaction near the final alphaIIbbeta3-dependent events of aggregation. We then used 3-(N-maleimidylpropionyl)biocytin (MPB) and pCMBS to study thiols in alphaIIbbeta3. As previously reported, disruption of the receptor was required to obtain labelling of thiols with MPB. Specificity of labelling for thiols in the alphaIIb and beta3 subunits was confirmed by identification of the purified proteins by mass spectrometry and by inhibition of labelling with 5,5'-dithiobis-(2-nitrobenzoic acid). In contrast to MPB, pCMBS preferentially reacted with thiols in alphaIIbbeta3 and blocked aggregation under physiological conditions. Similarly, pCMBS preferentially inhibited signalling-independent activation of alphaIIbbeta3 by Mn(2+). Our results suggest that the thiols in alphaIIbbeta3 that are blocked by pCMBS are important in the activation of this integrin.

Thiol Specific and Mitochondria Selective Fluorogenic Benzofurazan Sulfide for Live Cell Nonprotein Thiol Imaging and Quantification in Mitochondria.

PubMed

Wang, Shenggang; Yin, Huihui; Huang, Yue; Guan, Xiangming

2018-06-11

Cellular thiols are divided into two major categories: nonprotein thiols (NPSH) and protein thiols (PSH). Thiols are unevenly distributed inside the cell and compartmentalized in subcellular structures. Most of our knowledge on functions/dysfunctions of cellular/subcellular thiols is based on the quantification of cellular/subcellular thiols through homogenization of cellular/subcellular structures followed by a thiol quantification method. We would like to report a thiol-specific mitochondria-selective fluorogenic benzofurazan sulfide {7,7'-thiobis( N-rhodamine-benzo[c][1,2,5]oxadiazole-4-sulfonamide) (TBROS)} that can effectively image and quantify live cell NPSH in mitochondria through fluorescence intensity. Limited methods are available for imaging thiols in mitochondria in live cells especially in a quantitative manner. The thiol specificity of TBROS was demonstrated by its ability to react with thiols and inability to react with biologically relevant nucleophilic functional groups other than thiols. TBROS, with minimal fluorescence, formed strong fluorescent thiol adducts (λ ex = 550 nm, λ em = 580 nm) when reacting with NPSH confirming its fluorogenicity. TBROS failed to react with PSH from bovine serum albumin and cell homogenate proteins. The high mitochondrial thiol selectivity of TBROS was achieved by its mitochondria targeting structure and its higher reaction rate with NPSH at mitochondrial pH. Imaging of mitochondrial NPSH in live cells was confirmed by two colocalization methods and use of a thiol-depleting reagent. TBROS effectively imaged NPSH changes in a quantitative manner in mitochondria in live cells. The reagent will be a useful tool in exploring physiological and pathological roles of mitochondrial thiols.

Thiol-redox signaling, dopaminergic cell death, and Parkinson's disease.

PubMed

Garcia-Garcia, Aracely; Zavala-Flores, Laura; Rodriguez-Rocha, Humberto; Franco, Rodrigo

2012-12-15

Parkinson's disease (PD) is characterized by the selective loss of dopaminergic neurons of the substantia nigra pars compacta, which has been widely associated with oxidative stress. However, the mechanisms by which redox signaling regulates cell death progression remain elusive. Early studies demonstrated that depletion of glutathione (GSH), the most abundant low-molecular-weight thiol and major antioxidant defense in cells, is one of the earliest biochemical events associated with PD, prompting researchers to determine the role of oxidative stress in dopaminergic cell death. Since then, the concept of oxidative stress has evolved into redox signaling, and its complexity is highlighted by the discovery of a variety of thiol-based redox-dependent processes regulating not only oxidative damage, but also the activation of a myriad of signaling/enzymatic mechanisms. GSH and GSH-based antioxidant systems are important regulators of neurodegeneration associated with PD. In addition, thiol-based redox systems, such as peroxiredoxins, thioredoxins, metallothioneins, methionine sulfoxide reductases, transcription factors, as well as oxidative modifications in protein thiols (cysteines), including cysteine hydroxylation, glutathionylation, and nitrosylation, have been demonstrated to regulate dopaminergic cell loss. In this review, we summarize major advances in the understanding of the role of thiol-redox signaling in dopaminergic cell death in experimental PD. Future research is still required to clearly understand how integrated thiol-redox signaling regulates the activation of the cell death machinery, and the knowledge generated should open new avenues for the design of novel therapeutic approaches against PD.

Hydrogen sulfide deactivates common nitrobenzofurazan-based fluorescent thiol labeling reagents.

PubMed

Montoya, Leticia A; Pluth, Michael D

2014-06-17

Sulfhydryl-containing compounds, including thiols and hydrogen sulfide (H2S), play important but differential roles in biological structure and function. One major challenge in separating the biological roles of thiols and H2S is developing tools to effectively separate the reactivity of these sulfhydryl-containing compounds. To address this challenge, we report the differential responses of common electrophilic fluorescent thiol labeling reagents, including nitrobenzofurazan-based scaffolds, maleimides, alkylating agents, and electrophilic aldehydes, toward cysteine and H2S. Although H2S reacted with all of the investigated scaffolds, the photophysical response to each scaffold was significantly different. Maleimide-based, alkylating, and aldehydic thiol labeling reagents provided a diminished fluorescence response when treated with H2S. By contrast, nitrobenzofurazan-based labeling reagents were deactivated by H2S addition. Furthermore, the addition of H2S to thiol-activated nitrobenzofurazan-based reagents reduced the fluorescence signal, thus establishing the incompatibility of nitrobenzofurazan-based thiol labeling reagents in the presence of H2S. Taken together, these studies highlight the differential reactivity of thiols and H2S toward common thiol-labeling reagents and suggest that sufficient care must be taken when labeling or measuring thiols in cellular environments that produce H2S due to the potential for both false-positive and eroded responses.

Synthesis of soybean oil-based thiol oligomers.

PubMed

Wu, Jennifer F; Fernando, Shashi; Weerasinghe, Dimuthu; Chen, Zhigang; Webster, Dean C

2011-08-22

Industrial grade soybean oil (SBO) and thiols were reacted to generate thiol-functionalized oligomers via a thermal, free radical initiated thiol-ene reaction between the SBO double bond moieties and the thiol functional groups. The effect of the reaction conditions, including thiol concentration, catalyst loading level, reaction time, and atmosphere, on the molecular weight and the conversion to the resultant soy-thiols were examined in a combinatorial high-throughput fashion using parallel synthesis, combinatorial FTIR, and rapid gel permeation chromatography (GPC). High thiol functionality and concentration, high thermal free radical catalyst concentration, long reaction time, and the use of a nitrogen reaction atmosphere were found to favor fast consumption of the SBO, and produced high molecular weight products. The thiol conversion during the reaction was inversely affected by a high thiol concentration, but was favored by a long reaction time and an air reaction atmosphere. These experimental observations were explained by the initial low affinity of the SBO and thiol, and the improved affinity between the generated soy-thiol oligomers and unreacted SBO during the reaction. The synthesized soy-thiol oligomers can be used for renewable thiol-ene UV curable materials and high molecular solids and thiourethane thermal cure materials. Copyright © 2011 WILEY-VCH Verlag GmbH & Co. KGaA, Weinheim.

Evaluation of dynamic thiol/disulphide homeostasis as a novel indicator of oxidative stress in maple syrup urine disease patients under treatment.

PubMed

Zubarioglu, Tanyel; Kiykim, Ertugrul; Cansever, Mehmet Serif; Neselioglu, Salim; Aktuglu-Zeybek, Cigdem; Erel, Ozcan

2017-02-01

Maple syrup urine disease (MSUD) is a metabolic disorder that is caused by deficiency of branched-chain α-keto acid dehydrogenase complex. Although accumulation of toxic metabolites is associated with neurotoxicity, mechanisms underlying brain damage remain unclear. Aim of this study is to evaluate thiol/disulphide homeostasis as a novel indicator of oxidative stress in MSUD patients under treatment. Twenty patients with MSUD and 20 healthy individuals were included in study. All patients were under regular follow-up and had a good metabolic control. Serum native thiol (-SH), total thiol (-SH + -S-S-), disulphide (-S-S) levels were measured in all subjects. Disulphide/native thiol, disulphide/total thiol and native thiol/total thiol ratios were calculated from these values. Simultaneous blood sampling for plasma quantitative amino acid analysis was performed in both groups. Any significant difference was not observed in -SH, -SH + -S-S-, -S-S levels between two groups. In addition no increase of disulphide/native thiol and disulphide/total thiol ratios was detected in patient group. This study is the first study that evaluates dynamic thiol/disulphide homeostasis as an indicator of oxidative stress in MSUD patients. Among previous studies that were made to determine oxidative stress in treated MSUD patients, this study had the largest sample size also. In recent studies, it was claimed that oxidative stress could be responsible from neurotoxicity even in treated patients. Here, dynamic thiol/disulfide homeostasis status showed that providing good metabolic control in MSUD patients prevent oxidative stress. Under regular follow-up and good compliance with diet, additional antioxidant therapies would possibly not be necessary.

Thiol/disulfide homeostasis in postmenopausal osteoporosis.

PubMed

Korkmaz, V; Kurdoglu, Z; Alisik, M; Turgut, E; Sezgın, O O; Korkmaz, H; Ergun, Y; Erel, O

2017-04-01

To evaluate the impact of postmenopausal osteoporosis on thiol/disulfide homeostasis. A total of 75 participants were divided into two groups: Group 1 (n = 40) was composed of healthy postmenopausal women, and group 2 (n = 35) was composed of women with postmenopausal osteoporosis. Clinical findings and thiol/disulfide homeostasis were compared between the two groups. The disulfide/native thiol ratio was 8.6% ± 3.6 in group 1 and 12.7% ± 8.4 in group 2 (p = 0.04). The disulfide/native thiol percent ratio was significantly higher in group 2 after adjustment for the years since menopause and age (p < 0.05). The native thiol/total thiol percent ratio was 85.6% ± 4.8 in group 1 and 73.8% ± 24.9 in group 2 (p = 0.01). The native thiol/total thiol percent ratio was significantly lower in group 2 after adjustment for the years since menopause and age (p < 0.05). Thiol/disulfide homeostasis shifted to the disulfide side independent of age and years since menopause in postmenopausal osteoporosis.

Pharmaceutically active secondary metabolites of marine actinobacteria.

PubMed

Manivasagan, Panchanathan; Venkatesan, Jayachandran; Sivakumar, Kannan; Kim, Se-Kwon

2014-04-01

Marine actinobacteria are one of the most efficient groups of secondary metabolite producers and are very important from an industrial point of view. Many representatives of the order Actinomycetales are prolific producers of thousands of biologically active secondary metabolites. Actinobacteria from terrestrial sources have been studied and screened since the 1950s, for many important antibiotics, anticancer, antitumor and immunosuppressive agents. However, frequent rediscovery of the same compounds from the terrestrial actinobacteria has made them less attractive for screening programs in the recent years. At the same time, actinobacteria isolated from the marine environment have currently received considerable attention due to the structural diversity and unique biological activities of their secondary metabolites. They are efficient producers of new secondary metabolites that show a range of biological activities including antibacterial, antifungal, anticancer, antitumor, cytotoxic, cytostatic, anti-inflammatory, anti-parasitic, anti-malaria, antiviral, antioxidant, anti-angiogenesis, etc. In this review, an evaluation is made on the current status of research on marine actinobacteria yielding pharmaceutically active secondary metabolites. Bioactive compounds from marine actinobacteria possess distinct chemical structures that may form the basis for synthesis of new drugs that could be used to combat resistant pathogens. With the increasing advancement in science and technology, there would be a greater demand for new bioactive compounds synthesized by actinobacteria from various marine sources in future. Copyright © 2013 Elsevier GmbH. All rights reserved.

Synthesis of cyclic, multivalent Arg-Gly-Asp using sequential thiol-ene/thiol-yne photoreactions

PubMed Central

Aimetti, Alex A.; Feaver, Kristen R.

2014-01-01

A unique method has been developed for the formation of multivalent cyclic peptides. This procedure exploits on-resin peptide cyclization using a photoinitiated thiol-ene click reaction and subsequent clustering using thiol-yne photochemistry. Both reactions utilize the sulfhydryl group on natural cysteine amino acids to participate in the thiol-mediated reactions. PMID:20552127

Impaired Thiol-Disulfide Balance in Acute Brucellosis.

PubMed

Kolgelier, Servet; Ergin, Merve; Demir, Lutfi Saltuk; Inkaya, Ahmet Cagkan; Aktug Demir, Nazlim; Alisik, Murat; Erel, Ozcan

2017-05-24

The objective of this study was to examine a novel profile: thiol-disulfide homeostasis in acute brucellosis. The study included 90 patients with acute brucellosis, and 27 healthy controls. Thiol-disulfide profile tests were analyzed by a recently developed method, and ceruloplasmin levels were determined. Native thiol levels were 256.72 ± 48.20 μmol/L in the acute brucellosis group and 461.13 ± 45.37 μmol/L in the healthy group, and total thiol levels were 298.58 ± 51.78 μmol/L in the acute brucellosis group and 504.83 ± 51.05 μmol/L in the healthy group (p < 0.001, for both). The disulfide/native thiol ratios and disulfide/total thiol ratios were significantly higher, and native thiol/total thiol ratios were significantly lower in patients with acute brucellosis than in the healthy controls (p < 0.001, for all ratios). There were either positive or negative relationships between ceruloplasmin levels and thiol-disulfide parameters. The thiol-disulfide homeostasis was impaired in acute brucellosis. The strong associations between thiol-disulfide parameters and a positive acute-phase reactant reflected the disruption of the balance between the antioxidant and oxidant systems. Since thiol groups act as anti-inflammatory mediators, the alteration in the thiol-disulfide homeostasis may be involved in brucellosis.

Mechanism of covalent modification of glyceraldehyde-3-phosphate dehydrogenase at its active site thiol by nitric oxide, peroxynitrite and related nitrosating agents.

PubMed

Mohr, S; Stamler, J S; Brüne, B

1994-07-18

Previous studies have suggested that glyceraldehyde-3-phosphate dehydrogenase (GAPDH) undergoes covalent modification of an active site thiol by a NO.-induced [32P]NAD(+)-dependent mechanism. However, the efficacy of GAPDH modification induced by various NO donors was found to be independent of spontaneous rates of NO. release. To further test the validity of this mechanism, we studied the effects of nitrosonium tertrafluoroborate (BF4NO), a strong NO+ donor. BF4NO potently induces GAPDH labeling by the radioactive nucleotide. In this case, the addition of thiol significantly attenuates enzyme modification by competing for the NO moiety in the formation of RS-NO. Peroxynitrite (ONOO-) also induces GAPDH modification in the presence of thiol, consistent with the notion that this species can transfer NO+ (or NO2+) through the intermediacy of RS-NO. However, the efficiency of this reaction is limited by ONOO- -induced oxidation of protein SH groups at the active site. ONOO- generation appears to account for the modification of GAPDH by SIN-1. Thus, S-nitrosylation of the active site thiol is a prequisite for subsequent post-translational modification with NAD+, and emphasizes the role of NO+ transfer in the initial step of this pathway. Our findings thus provide a uniform mechanism by which nitric oxide and related NO donors initiate non-enzymatic ADP-ribosylation (like) reactions. In biological systems, endogenous RS-NO are likely to support the NO group transfer to thiol-containing proteins.

Thiol-Based Redox Switches and Gene Regulation

PubMed Central

2011-01-01

Abstract Cysteine is notable among the universal, proteinogenic amino acids for its facile redox chemistry. Cysteine thiolates are readily modified by reactive oxygen species (ROS), reactive electrophilic species (RES), and reactive nitrogen species (RNS). Although thiol switches are commonly triggered by disulfide bond formation, they can also be controlled by S-thiolation, S-alkylation, or modification by RNS. Thiol-based switches are common in both prokaryotic and eukaryotic organisms and activate functions that detoxify reactive species and restore thiol homeostasis while repressing functions that would be deleterious if expressed under oxidizing conditions. Here, we provide an overview of the best-understood examples of thiol-based redox switches that affect gene expression. Intra- or intermolecular disulfide bond formation serves as a direct regulatory switch for several bacterial transcription factors (OxyR, OhrR/2-Cys, Spx, YodB, CrtJ, and CprK) and indirectly regulates others (the RsrA anti-σ factor and RegB sensory histidine kinase). In eukaryotes, thiol-based switches control the yeast Yap1p transcription factor, the Nrf2/Keap1 electrophile and oxidative stress response, and the Chlamydomonas NAB1 translational repressor. Collectively, these regulators reveal a remarkable range of chemical modifications exploited by Cys residues to effect changes in gene expression. Antioxid. Redox Signal. 14, 1049—1063. PMID:20626317

Mitochondrial thiol modification by a targeted electrophile inhibits metabolism in breast adenocarcinoma cells by inhibiting enzyme activity and protein levels.

PubMed

Smith, M Ryan; Vayalil, Praveen K; Zhou, Fen; Benavides, Gloria A; Beggs, Reena R; Golzarian, Hafez; Nijampatnam, Bhavitavya; Oliver, Patsy G; Smith, Robin A J; Murphy, Michael P; Velu, Sadanandan E; Landar, Aimee

2016-08-01

Many cancer cells follow an aberrant metabolic program to maintain energy for rapid cell proliferation. Metabolic reprogramming often involves the upregulation of glutaminolysis to generate reducing equivalents for the electron transport chain and amino acids for protein synthesis. Critical enzymes involved in metabolism possess a reactive thiolate group, which can be modified by certain oxidants. In the current study, we show that modification of mitochondrial protein thiols by a model compound, iodobutyl triphenylphosphonium (IBTP), decreased mitochondrial metabolism and ATP in MDA-MB 231 (MB231) breast adenocarcinoma cells up to 6 days after an initial 24h treatment. Mitochondrial thiol modification also depressed oxygen consumption rates (OCR) in a dose-dependent manner to a greater extent than a non-thiol modifying analog, suggesting that thiol reactivity is an important factor in the inhibition of cancer cell metabolism. In non-tumorigenic MCF-10A cells, IBTP also decreased OCR; however the extracellular acidification rate was significantly increased at all but the highest concentration (10µM) of IBTP indicating that thiol modification can have significantly different effects on bioenergetics in tumorigenic versus non-tumorigenic cells. ATP and other adenonucleotide levels were also decreased by thiol modification up to 6 days post-treatment, indicating a decreased overall energetic state in MB231 cells. Cellular proliferation of MB231 cells was also inhibited up to 6 days post-treatment with little change to cell viability. Targeted metabolomic analyses revealed that thiol modification caused depletion of both Krebs cycle and glutaminolysis intermediates. Further experiments revealed that the activity of the Krebs cycle enzyme, aconitase, was attenuated in response to thiol modification. Additionally, the inhibition of glutaminolysis corresponded to decreased glutaminase C (GAC) protein levels, although other protein levels were unaffected. This study

Location of the redox-active thiols of ribonucleotide reductase: sequences similarity between the Escherichia coli and Lactobacillus leichmannii enzymes

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

Lin, A.N.I.; Ashley, G.W.; Stubbe, J.

1987-11-03

The redox-active thiols of Escherichia coli ribonucleoside diphosphate reductase and of Lactobacillus leichmannii ribonucleoside triphosphate reductase have been located by a procedure involving (1) prereduction of enzyme with dithiothreitol, (2) specific oxidation of the redox-active thiols by treatment with substrate in the absence of exogenous reductant, (3) alkylation of other thiols with iodoacetamide, and (4) reduction of the disulfides with dithiothreitol and alkylation with (1-/sup 14/C)iodoacetamide. The dithiothreitol-reduce E. coli B1 subunit is able to convert 3 equiv of CDP to dCDP and is labeled with 5.4 equiv of /sup 14/C. Sequencing of tryptic peptides shows that 2.8 equiv ofmore » /sup 14/C is on cysteines-752 and -757 at the C-terminus of B1, while 1.0-1.5 equiv of /sup 14/C is on cysteines-222 and -227. It thus appears that two sets of redox-active dithiols are involved in substrate reduction. The L. leichmannii reductase is able to convert 1.1 equiv of CTP to dCTP and is labeled with 2.1 equiv of /sup 14/C. Sequencing of tryptic peptides shows that 1.4 equiv of /sup 14/C is located on the two cysteines of C-E-G-G-A-C-P-I-K. This peptide shows remarkable and unexpected similarity to the thiol-containing region of the C-terminal peptide of E. coli B1, C-E-S-G-A-C-K-I.« less

Preparation of thiol-functionalized activated carbon from sewage sludge with coal blending for heavy metal removal from contaminated water.

PubMed

Li, Juan; Xing, Xing; Li, Jiao; Shi, Mei; Lin, Aijun; Xu, Congbin; Zheng, Jianzhong; Li, Ronghua

2018-03-01

Sewage sludge produced from wastewater treatment is a pressing environmental issue. Mismanagement of the massive amount of sewage sludge would threat our valuble surface and shallow ground water resources. Use of activated carbon prepared from carbonization of these sludges for heavy metal removal can not only minimize and stabilize these hazardous materials but also realize resources reuse. In this study, thiol-functionalized activated carbon was synthesized from coal-blended sewage sludge, and its capacity was examined for removing Cu(II), Pb(II), Cd(II) and Ni(II) from water. Pyrolysis conditions to prepare activated carbons from the sludge and coal mixture were examined, and the synthesized material was found to achieve the highest BET surface area of 1094 m 2 /g under 500 °C and 30 min. Batch equilibrium tests indicated that the thiol-functionalized activated carbon had a maximum sorption capacity of 238.1, 96.2, 87.7 and 52.4 mg/g for Pb(II), Cd(II), Cu(II) and Ni(II) removal from water, respectively. Findings of this study suggest that thiol-functionalized activated carbon prepared from coal-blended sewage sludge would be a promising sorbent material for heavy metal removal from waters contaminated with Cu(II), Pb(II), Cd(II) and Ni(II). Copyright © 2017 Elsevier Ltd. All rights reserved.

Conferring specificity in redox pathways by enzymatic thiol/disulfide exchange reactions.

PubMed

Netto, Luis Eduardo S; de Oliveira, Marcos Antonio; Tairum, Carlos A; da Silva Neto, José Freire

2016-01-01

Thiol-disulfide exchange reactions are highly reversible, displaying nucleophilic substitutions mechanism (S(N)2 type). For aliphatic, low molecular thiols, these reactions are slow, but can attain million times faster rates in enzymatic processes. Thioredoxin (Trx) proteins were the first enzymes described to accelerate thiol-disulfide exchange reactions and their high reactivity is related to the high nucleophilicity of the attacking thiol. Substrate specificity in Trx is achieved by several factors, including polar, hydrophobic, and topological interactions through a groove in the active site. Glutaredoxin (Grx) enzymes also contain the Trx fold, but they do not share amino acid sequence similarity with Trx. A conserved glutathione binding site is a typical feature of Grx that can reduce substrates by two mechanisms (mono and dithiol). The high reactivity of Grx enzymes is related to the very acid pK(a) values of reactive Cys that plays roles as good leaving groups. Therefore, although distinct oxidoreductases catalyze similar thiol–disulfide exchange reactions, their enzymatic mechanisms vary. PDI and DsbA are two other oxidoreductases, but they are involved in disulfide bond formation, instead of disulfide reduction, which is related to the oxidative environment where they are found. PDI enzymes and DsbC are endowed with disulfide isomerase activity, which is related with their tetra-domain architecture. As illustrative description of specificity in thiol-disulfide exchange, redox aspects of transcription activation in bacteria, yeast, and mammals are presented in an evolutionary perspective. Therefore, thiol-disulfide exchange reactions play important roles in conferring specificity to pathways, a required feature for signaling.

A sensitive and rapid ultra HPLC-MS/MS method for the simultaneous detection of clopidogrel and its derivatized active thiol metabolite in human plasma.

PubMed

Peer, Cody J; Spencer, Shawn D; VanDenBerg, Dustin A H; Pacanowski, Michael A; Horenstein, Richard B; Figg, William D

2012-01-01

A sensitive, selective, and rapid ultra-high performance liquid chromatography-tandem mass spectrometry (uHPLC-MS/MS) was developed for the simultaneous quantification of clopidogrel (Plavix(®)) and its derivatized active metabolite (CAMD) in human plasma. Derivatization of the active metabolite in blood with 2-bromo-3'-methoxy acetophenone (MPB) immediately after collection ensured metabolite stability during sample handling and storage. Following addition of ticlopidine as an internal standard and simple protein precipitation, the analytes were separated on a Waters Acquity UPLC™ sub-2 μm-C(18) column via gradient elution before detection on a triple-quadrupole MS with multiple-reaction-monitoring via electrospray ionization. The method was validated across the clinically relevant concentration range of 0.01-50 ng/mL for parent clopidogrel and 0.1-150 ng/mL (r(2)=0.99) for CAMD, with a fast run time of 1.5 min to support pharmacokinetic studies using 75, 150, or 300 mg oral doses of clopidogrel. The analytical method measured concentrations of clopidogrel and CAMD with accuracy (%DEV) <±12% and precision (%CV) of <±6%. The method was successfully applied to measure the plasma concentrations of clopidogrel and CAMD in three subjects administered single oral doses of 75, 150, and 300 mg clopidogrel. It was further demonstrated that the derivatizing agent (MPB) does not affect clopidogrel levels, thus from one aliquot of blood drawn clinically, this method can simultaneously quantify both clopidogrel and CAMD with sensitivity in the picogram per mL range. Published by Elsevier B.V.

A Sensitive and Rapid Ultra HPLC-MS/MS Method for the Simultaneous Detection of Clopidogrel and its Derivatized Active Thiol Metabolite in Human Plasma

PubMed Central

Peer, Cody J.; Spencer, Shawn D.; VanDenBerg, Dustin A. H.; Pacanowski, Michael A.; Horenstein, Richard B.; Figg, William D.

2011-01-01

A sensitive, selective, and rapid ultra-high performance liquid chromatography-tandem mass spectrometry (uHPLC-MS/MS) was developed for the simultaneous quantification of clopidogrel (Plavix®) and its derivatized active metabolite (CAMD) in human plasma. Derivatization of the active metabolite in blood with 2-bromo-3’-methoxy acetophenone (MPB) immediately after collection ensured metabolite stability during sample handling and storage. Following addition of ticlopidine as an internal standard and simple protein precipitation, the analytes were separated on a Waters Acquity UPLC™ sub-2µm-C18 column via gradient elution before detection on a triple-quadrupole MS with multiple-reaction-monitoring via electrospray ionization. The method was validated across the clinically-relevant concentration range of 0.01–50 ng/mL for parent clopidogrel and 0.1–150 ng/mL (r2= 0.99) for CAMD, with a fast run time of 1.5 min to support pharmacokinetic studies using 75, 150, or 300 mg oral doses of clopidogrel. The analytical method measured concentrations of clopidogrel and CAMD with accuracy (%DEV) < ±12% and precision (%CV) of < ±6%. The method was successfully applied to measure the plasma concentrations of clopidogrel and CAMD in three subjects administered single oral doses of 75, 150, and 300 mg clopidogrel. It was further demonstrated that the derivatizing agent (MPB) does not affect clopidogrel levels, thus from one aliquot of blood drawn clinically, this method can simultaneously quantify both clopidogrel and CAMD with sensitivity in the picogram per mL range. PMID:22169056

Thiol Reactivity of Curcumin and Its Oxidation Products.

PubMed

Luis, Paula B; Boeglin, William E; Schneider, Claus

2018-04-16

The polypharmacological effects of the turmeric compound curcumin may be partly mediated by covalent adduction to cellular protein. Covalent binding to small molecule and protein thiols is thought to occur through a Michael-type addition at the enone moiety of the heptadienedione chain connecting the two methoxyphenol rings of curcumin. Here we show that curcumin forms the predicted thiol-Michael adducts with three model thiols, glutathione, N-acetylcysteine, and β-mercaptoethanol. More abundant, however, are respective thiol adducts of the dioxygenated spiroepoxide intermediate of curcumin autoxidation. Two electrophilic sites at the quinone-like ring of the spiroepoxide are identified. Addition of β-mercaptoethanol at the 5'-position of the ring gives a 1,7-dihydroxycyclopentadione-5' thioether, and addition at the 1'-position results in cleavage of the aromatic ring from the molecule, forming methoxyphenol-thioether and a tentatively identified cyclopentadione aldehyde. The curcuminoids demethoxy- and bisdemethoxycurcumin do not form all of the possible thioether adducts, corresponding with their increased stability toward autoxidation. RAW264.7 macrophage-like cells activated with phorbol ester form curcumin-glutathionyl and the 1,7-dihydroxycyclopentadione-5'-glutathionyl adducts. These studies indicate that the enone of the parent compound is not the only functional electrophile in curcumin, and that its oxidation products provide additional electrophilic sites. This suggests that protein binding by curcumin may involve oxidative activation into reactive quinone methide and spiroepoxide electrophiles.

Submicrodeterminations of thiols, disulphides and thiol esters in serum by using o-hydroxymercuribenzoic acid and dithiofluorescein

PubMed Central

Wroński, Mieczysław

1967-01-01

1. Methods are described for selective estimation of thiols, disulphides and thiol esters in standard solutions and in serum. The methods are based on the reaction with the excess of o-hydroxymercuribenzoic acid (HMB) in alkaline solution with subsequent addition of dithiofluorescein in excess and determination of the extinction at 588mμ. The sensitivity of the methods amounts to 1·5×10−9g.equiv. in 5ml. of final solution. Of results obtained on standard solutions 80% have the errors within the range ±4%. 2. It has been found that serum contains an unidentified substance (substance X) producing green complexes with dithiofluorescein which undergo decomposition on addition of formaldehyde. The correction for substance X must be estimated in a separate sample and taken into account. The concentration of substance X can be calculated from extinctions measured at 588mμ and 635mμ in the presence of dithiofluorescein in excess. 3. The selective determination of thiols and disulphides is based on different reaction rates with formaldehyde. The complexes between HMB and cysteine can be selectively decomposed by formaldehyde, and free glutathione can be selectively removed by formaldehyde in the presence of protein thiols. 4. Thiols are determined in the presence of triethylamine, thiols plus disulphides in the presence of triethylamine and sulphite, and thiols plus thiol esters in the presence of dimethylamine, with subsequent addition of ammonium sulphate. PMID:6049936

Transcriptional Activation by Heat and Cold of a Thiol Protease Gene in Tomato

PubMed Central

Schaffer, Mark A.; Fischer, Robert L.

1990-01-01

We previously determined that low temperature induces the accumulation in tomato (Lycopersicon esculentum) fruit of a cloned mRNA, designated C14, encoding a polypeptide related to thiol proteases (MA Schaffer, RL Fischer [1988] Plant Physiol 87: 431-436). We now demonstrate that C14 mRNA accumulation is a response common to both high (40°C) and low (4°C) temperature stresses. Exposure of tomato fruit to 40°C results in the accumulation of C14 mRNA, by 8 hours. This response is more rapid than that to 4°C, but slower than the induction of many heat shock messages by 40°C, and therefore unique. We have also studied the mechanism by which heat and cold exposure activate C14 gene expression. Both high and low temperature regulate protease gene expression through transcriptional induction of a single C14 gene. A hypothesis for the function of C14 thiol protease gene expression in response to heat and cold is discussed. Images Figure 1 Figure 2 Figure 3 Figure 4 PMID:16667644

Biologically Active Secondary Metabolites from the Fungi.

PubMed

Bills, Gerald F; Gloer, James B

2016-11-01

Many Fungi have a well-developed secondary metabolism. The diversity of fungal species and the diversification of biosynthetic gene clusters underscores a nearly limitless potential for metabolic variation and an untapped resource for drug discovery and synthetic biology. Much of the ecological success of the filamentous fungi in colonizing the planet is owed to their ability to deploy their secondary metabolites in concert with their penetrative and absorptive mode of life. Fungal secondary metabolites exhibit biological activities that have been developed into life-saving medicines and agrochemicals. Toxic metabolites, known as mycotoxins, contaminate human and livestock food and indoor environments. Secondary metabolites are determinants of fungal diseases of humans, animals, and plants. Secondary metabolites exhibit a staggering variation in chemical structures and biological activities, yet their biosynthetic pathways share a number of key characteristics. The genes encoding cooperative steps of a biosynthetic pathway tend to be located contiguously on the chromosome in coregulated gene clusters. Advances in genome sequencing, computational tools, and analytical chemistry are enabling the rapid connection of gene clusters with their metabolic products. At least three fungal drug precursors, penicillin K and V, mycophenolic acid, and pleuromutilin, have been produced by synthetic reconstruction and expression of respective gene clusters in heterologous hosts. This review summarizes general aspects of fungal secondary metabolism and recent developments in our understanding of how and why fungi make secondary metabolites, how these molecules are produced, and how their biosynthetic genes are distributed across the Fungi. The breadth of fungal secondary metabolite diversity is highlighted by recent information on the biosynthesis of important fungus-derived metabolites that have contributed to human health and agriculture and that have negatively impacted crops

Thiol biochemistry of prokaryotes

NASA Technical Reports Server (NTRS)

Fahey, Robert C.

1986-01-01

The present studies have shown that GSH metabolism arose in the purple bacteria and cyanobacteria where it functions to protect against oxygen toxicity. Evidence was obtained indicating that GSH metabolism was incorporated into eucaryotes via the endosymbiosis giving rise to mitochrondria and chloroplasts. Aerobic bacteria lacking GSH utilize other thiols for apparently similar functions, the thiol being coenzyme A in Gram positive bacteria and chi-glutamylcysteine in the halobacteria. The thiol biochemistry of prokaryotes is thus seen to be much more highly diversified than that of eucaryotes and much remains to be learned about this subject.

Thiol/disulfide redox states in signaling and sensing

PubMed Central

Go, Young-Mi; Jones, Dean P.

2015-01-01

Rapid advances in redox systems biology are creating new opportunities to understand complexities of human disease and contributions of environmental exposures. New understanding of thiol-disulfide systems have occurred during the past decade as a consequence of the discoveries that thiol and disulfide systems are maintained in kinetically controlled steady-states displaced from thermodynamic equilibrium, that a widely distributed family of NADPH oxidases produces oxidants that function in cell signaling, and that a family of peroxiredoxins utilize thioredoxin as a reductant to complement the well-studied glutathione antioxidant system for peroxide elimination and redox regulation. This review focuses on thiol/disulfide redox state in biologic systems and the knowledge base available to support development of integrated redox systems biology models to better understand the function and dysfunction of thiol-disulfide redox systems. In particular, central principles have emerged concerning redox compartmentalization and utility of thiol/disulfide redox measures as indicators of physiologic function. Advances in redox proteomics show that, in addition to functioning in protein active sites and cell signaling, cysteine residues also serve as redox sensors to integrate biologic functions. These advances provide a framework for translation of redox systems biology concepts to practical use in understanding and treating human disease. Biological responses to cadmium, a widespread environmental agent, are used to illustrate the utility of these advances to the understanding of complex pleiotropic toxicities. PMID:23356510

Secondary metabolites and insecticidal activity of Anemone pavonina.

PubMed

Varitimidis, Christos; Petrakis, Panos V; Vagias, Constantinos; Roussis, Vassilios

2006-01-01

The insecticidal properties of the crude extracts of the leaves and flowers of Anemone pavonina were evaluated on Pheidole pallidula ants and showed significant levels of activity. Bioassay-guided fractionations led to the isolation of the butenolide ranunculin (1) as the active principle. Chemical investigations of the extracts showed them to contain as major components the sitosterol glycopyranoside lipids 2-5 and the glycerides 6-8. The structures of the metabolites were elucidated, following acetylation and hydrolysis of the natural products, by interpretation of their NMR and mass spectral data. The uncommon lipid metabolites 2-8 were isolated for the first time from the genus Anemone and this is the first report of insecticidal activity of the Anemone metabolite ranunculin against ants.

Thiol/disulfide homeostasis in asphalt workers.

PubMed

Yilmaz, Ömer Hınç; Bal, Ceylan; Neşelioglu, Salim; Büyükşekerci, Murat; Gündüzöz, Meşide; Eren, Funda; Tutkun, Lutfiye; Yilmaz, Fatma Meric

2016-09-02

The aim of this study was to investigate thiol/disulfide homeostasis in asphalt workers who are exposed to polycyclic aromatic hydrocarbons occupationally. The study was carried out in 34 nonsmoker asphalt workers. Additionally, 35 healthy nonsmoker volunteers were recruited as control group. Thiol and disulfide concentrations were determined using the novel automated measurement method. Levels of urinary 1-OH-pyrene were analyzed by liquid chromatography. Disulfide/thiol ratio was significantly higher in exposed group (p = .034). Also, a positive correlation was detected between disulfide/thiol ratio and 1-OH-pyrene values (r = .249, p = .036). Thiol/disulfide homeostasis was found to be disturbed in asphalt workers. The novel test used in this study may be useful for evaluating the oxidative status in polycyclic aromatic hydrocarbon (PAH) exposure.

Effects of thiol antioxidants on the atropselective oxidation of 2,2',3,3',6,6'-hexachlorobiphenyl (PCB 136) by rat liver microsomes.

PubMed

Wu, Xianai; Lehmler, Hans-Joachim

2016-02-01

Chiral polychlorinated biphenyl (PCB) congeners, such as PCB 136, are atropselectively metabolized to various hydroxylated PCB metabolites (HO-PCBs). The present study investigates the effect of two thiol antioxidants, glutathione and N-acetyl-cysteine (NAC), on profiles and chiral signatures of PCB 136 and its HO-PCB metabolites in rat liver microsomal incubations. Liver microsomes prepared from rats pretreated with phenobarbital were incubated with PCB 136 (5 μM) in the presence of the respective antioxidant (0-10 mM), and levels and chiral signatures of PCB 136 and its HO-PCB metabolites were determined. Three metabolites, 5-136 (2,2',3,3',6,6'-hexachlorobiphenyl-5-ol), 4-136 (2,2',3,3',6,6'-hexachlorobiphenyl-4-ol), and 4,5-136 (2,2',3,3',6,6'-hexachlorobiphenyl-4,5-diol), were detected in all incubations, with 5-136 being the major metabolite. Compared to microsomal incubations without antioxidant, levels of 4,5-136 increased with increasing antioxidant concentration, whereas levels of PCB 136 and both mono-HO-PCBs were not affected by the presence of either antioxidant. PCB 136, 4-136, and 5-136 displayed significant atropisomeric enrichment; however, the direction and extent of the atropisomeric enrichment was not altered in the presence of an antioxidant. Because 4,5-136 can either be conjugated to a sulfate or glucuronide metabolite that is readily excreted or further oxidized a potentially toxic PCB 136 quinone, the effect of both thiol antioxidants on 4,5-136 formation suggests that disruptions of glutathione homeostasis may alter the balance between both metabolic pathways and, thus, PCB 136 toxicity in vivo.

Synthesis of a thiol-β-cyclodextrin, a potential agent for controlling enzymatic browning in fruits and vegetables.

PubMed

Manta, Carmen; Peralta-Altier, Gabriela; Gioia, Larissa; Méndez, María F; Seoane, Gustavo; Ovsejevi, Karen

2013-11-27

A thiol-β-cyclodextrin was synthesized by a simple and environmentally friendly three-step method comprising epoxy activation of β-cyclodextrin, thiosulfate-mediated oxirane opening, and further reduction of the S-alkyl thiosulfate to a thiol group. The final step was optimized by using thiopropyl-agarose, a solid phase reducing agent with many advantages over soluble ones. β-Cyclodextrin thiolation was confirmed by titration with a thiol-reactive reagent, NMR studies, and MALDI-TOF/TOF. Thiolated cyclodextrin had an average value of one thiol group per molecule. Thiol-β-cyclodextrin proved to be an excellent agent for controlling polyphenol oxidase activity. This copper-containing enzyme is responsible for browning in fruits and vegetables. Under the same conditions, thiol-β-cyclodextrin generated a reductive microenvironment that increased the antibrowning effect on Red Delicious apples compared to unmodified β-cyclodextrin.

Physical activity, sedentary behavior, and vitamin D metabolites.

PubMed

Hibler, Elizabeth A; Sardo Molmenti, Christine L; Dai, Qi; Kohler, Lindsay N; Warren Anderson, Shaneda; Jurutka, Peter W; Jacobs, Elizabeth T

2016-02-01

Physical activity is associated with circulating 25-hydroxyvitamin D (25(OH)D). However, the influence of activity and/or sedentary behavior on the biologically active, seco-steroid hormone 1α,25-dihydroxyvitamin D (1,25(OH)2D) is unknown. We conducted a cross-sectional analysis among ursodeoxycholic acid (UDCA) randomized trial participants (n=876) to evaluate associations between physical activity, sedentary behavior, and circulating vitamin D metabolite concentrations. Continuous vitamin D metabolite measurements and clinical thresholds were evaluated using multiple linear and logistic regression models, mutually adjusted for either 1,25(OH)2D or 25(OH)D and additional confounding factors. A statistically significant linear association between 1,25(OH)2D and moderate-vigorous physical activity per week was strongest among women (β (95% CI): 3.10 (1.51-6.35)) versus men (β (95% CI): 1.35 (0.79-2.29)) in the highest tertile of activity compared to the lowest (p-interaction=0.003). Furthermore, 25(OH)D was 1.54ng/ml (95% CI 1.09-1.98) higher per hour increase in moderate-vigorous activity (p=0.001) and odds of sufficient 25(OH)D status was higher among physically active participants (p=0.001). Sedentary behavior was not significantly associated with either metabolite in linear regression models, nor was a statistically significant interaction by sex identified. The current study identified novel associations between physical activity and serum 1,25(OH)2D levels, adjusted for 25(OH)D concentrations. These results identify the biologically active form of vitamin D as a potential physiologic mechanism related to observed population-level associations between moderate-vigorous physical activity with bone health and chronic disease risk. However, future longitudinal studies are needed to further evaluate the role of physical activity and vitamin D metabolites in chronic disease prevention. Copyright © 2015 Elsevier Inc. All rights reserved.

Thiol synthetases of legumes: immunogold localization and differential gene regulation by phytohormones.

PubMed

Clemente, Maria R; Bustos-Sanmamed, Pilar; Loscos, Jorge; James, Euan K; Pérez-Rontomé, Carmen; Navascués, Joaquín; Gay, Marina; Becana, Manuel

2012-06-01

In plants and other organisms, glutathione (GSH) biosynthesis is catalysed sequentially by γ-glutamylcysteine synthetase (γECS) and glutathione synthetase (GSHS). In legumes, homoglutathione (hGSH) can replace GSH and is synthesized by γECS and a specific homoglutathione synthetase (hGSHS). The subcellular localization of the enzymes was examined by electron microscopy in several legumes and gene expression was analysed in Lotus japonicus plants treated for 1-48 h with 50 μM of hormones. Immunogold localization studies revealed that γECS is confined to chloroplasts and plastids, whereas hGSHS is also in the cytosol. Addition of hormones caused differential expression of thiol synthetases in roots. After 24-48 h, abscisic and salicylic acids downregulated GSHS whereas jasmonic acid upregulated it. Cytokinins and polyamines activated GSHS but not γECS or hGSHS. Jasmonic acid elicited a coordinated response of the three genes and auxin induced both hGSHS expression and activity. Results show that the thiol biosynthetic pathway is compartmentalized in legumes. Moreover, the similar response profiles of the GSH and hGSH contents in roots of non-nodulated and nodulated plants to the various hormonal treatments indicate that thiol homeostasis is independent of the nitrogen source of the plants. The differential regulation of the three mRNA levels, hGSHS activity, and thiol contents by hormones indicates a fine control of thiol biosynthesis at multiple levels and strongly suggests that GSH and hGSH play distinct roles in plant development and stress responses.

Adsorption of Dissolved Metals in the Berkeley Pit using Thiol-Functionalized Self-Assembled Monolayers on Mesoporous Supports (Thiol-SAMMS)

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

Betancourt, Amaury P.; Mattigod, Shas V.; Wellman, Dawn M.

2010-03-07

The Berkeley Pit in Butte, Montana, is heavily contaminated with dissolved metals. Adsorption and extraction of these metals can be accomplished through the use of a selective adsorbent. For this research, the adsorbent used was thiol-functionalized Self-Assembled Monolayers on Mesoporous Supports (thiol-SAMMS), which was developed at Pacific Northwest National Laboratory (PNNL). Thiol-SAMMS selectively binds to numerous types of dissolved metals. The objective of this research was to evaluate the loading and kinetics of aluminum, beryllium, copper, and zinc on thiol-SAMMS. For the loading tests, a series of Berkeley Pit water to thiol-SAMMS ratios (mL:g) were tested. These ratios were 1000:1,more » 500:1, 100:1, and 50:1. Berkeley Pit water is acidic (pH {approx} 2.5). This can affect the performance of SAMMS materials. Therefore, the effect of pH was evaluated by conducting parallel series of loading tests wherein the Berkeley Pit water was neutralized before or after addition of thiol-SAMMS, and a series of kinetics tests wherein the Berkeley Pit water was neutralized before addition of thiol-SAMMS for the first test and was not neutralized for the second test. For the kinetics tests, one Berkeley Pit water to thiol-SAMMS ratio was tested, which was 2000:1. The results of the loading and kinetics tests suggest that a significant decrease in dissolved metal concentration at Berkeley Pit could be realized through neutralization of Berkeley Pit water. Thiol-SAMMS technology has a limited application under the highly acidic conditions posed by the Berkeley Pit. However, thiol-SAMMS could provide a secondary remedial technique which would complete the remedial system and remove dissolved metals from the Berkeley Pit to below drinking water standards.« less

A Novel Oxidative Stress Mediator in Acute Appendicitis: Thiol/Disulphide Homeostasis

PubMed Central

Turan, Umit; Kuvvetli, Adnan; Kilavuz, Huseyin; Karakaya, Burak; Ozaltun, Pınar; Alısık, Murat; Erel, Ozcan

2016-01-01

Aim. To investigate the role of a novel oxidative stress marker, thiol/disulphide homeostasis, in patients diagnosed with acute appendicitis (AA). Methods. In this study, seventy-one (43 male and 28 female) patients diagnosed with AA and 71 (30 male and 41 female) healthy volunteers were included. Age, gender, body mass index (BMI), haemoglobin (Hb), white blood cell (WBC), c-reactive protein (CRP), and thiol/disulphide homeostasis parameters (native thiol, total thiol, disulphide, disulphide/native thiol, native thiol/total thiol, and disulphide/total thiol ratios) were compared between the groups. Thiol/disulphide homeostasis was determined by a newly developed method by Erel and Neselioglu. Results. The native thiol, total thiol, and the native thiol/total thiol ratio levels were statistically significantly decreased in the AA compared with the control group (p < 0.001). Disulphide level and the ratios of disulphide/native thiol and disulphide/total thiol were higher in the AA group than in the control group (p < 0.001). There was a negative correlation of CRP with native thiol, total thiol, and native thiol/total thiol ratio while there was a positive correlation of CRP with disulphide/native thiol and disulphide/total thiol in the AA group. In the stepwise regression model, risk factors as disulphide/native thiol (OR = 1.368; p = 0.018) and CRP (OR = 1.635; p = 0.003) were determined as predictors of perforated appendicitis compared to the nonperforated group. Conclusion. This is the first study examining the thiol/disulphide homeostasis as a diagnostic aid in AA and establishing thiol/disulphide homeostatis balance shifted towards the disulphide formation due to thiol oxidation. Further studies are needed to optimize the use of this novel oxidative stress marker in AA. PMID:27642237

Biologically Active Metabolites Synthesized by Microalgae

PubMed Central

Costa, Jorge Alberto Vieira

2015-01-01

Microalgae are microorganisms that have different morphological, physiological, and genetic traits that confer the ability to produce different biologically active metabolites. Microalgal biotechnology has become a subject of study for various fields, due to the varied bioproducts that can be obtained from these microorganisms. When microalgal cultivation processes are better understood, microalgae can become an environmentally friendly and economically viable source of compounds of interest, because production can be optimized in a controlled culture. The bioactive compounds derived from microalgae have anti-inflammatory, antimicrobial, and antioxidant activities, among others. Furthermore, these microorganisms have the ability to promote health and reduce the risk of the development of degenerative diseases. In this context, the aim of this review is to discuss bioactive metabolites produced by microalgae for possible applications in the life sciences. PMID:26339647

Localization of ATP Sulfurylase and O-Acetylserine(thiol)lyase in Spinach Leaves.

PubMed

Lunn, J E; Droux, M; Martin, J; Douce, R

1990-11-01

The intracellular compartmentation of ATP sulfurylase and O-acetylserine(thiol)lyase in spinach (Spinacia oleracea L.) leaves has been investigated by isolation of organelles and fractionation of protoplasts. ATP sulfurylase is located predominantly in the chloroplasts, but is also present in the cytosol. No evidence was found for ATP sulfurylase activity in the mitochondria. Two forms of ATP sulfurylase were separated by anion-exchange chromatography. The more abundant form is present in the chloroplasts, the second is cytosolic. O-Acetylserine(thiol)lyase activity is located primarily in the chloroplasts and cytosol, but is also present in the mitochondria. Three forms of O-acetylserine(thiol)lyase were separated by anion-exchange chromatography, and each was found to be specific to one intracellular compartment. The cytosolic ATP sulfurylase may not be active in vivo due to the unfavorable equilibrium constant of the reaction, and the presence of micromolar concentrations of inorganic pyrophosphate in the cytosol, therefore its role remains unknown. It is suggested that the plant cell may be unable to transport cysteine between the different compartments, so that the cysteine required for protein synthesis must be synthesized in situ, hence the presence of O-acetylserine(thiol)lyase in the three compartments where proteins are synthesized.

Localization of ATP Sulfurylase and O-Acetylserine(thiol)lyase in Spinach Leaves

PubMed Central

Lunn, John E.; Droux, Michel; Martin, Jacqueline; Douce, Roland

1990-01-01

The intracellular compartmentation of ATP sulfurylase and O-acetylserine(thiol)lyase in spinach (Spinacia oleracea L.) leaves has been investigated by isolation of organelles and fractionation of protoplasts. ATP sulfurylase is located predominantly in the chloroplasts, but is also present in the cytosol. No evidence was found for ATP sulfurylase activity in the mitochondria. Two forms of ATP sulfurylase were separated by anion-exchange chromatography. The more abundant form is present in the chloroplasts, the second is cytosolic. O-Acetylserine(thiol)lyase activity is located primarily in the chloroplasts and cytosol, but is also present in the mitochondria. Three forms of O-acetylserine(thiol)lyase were separated by anion-exchange chromatography, and each was found to be specific to one intracellular compartment. The cytosolic ATP sulfurylase may not be active in vivo due to the unfavorable equilibrium constant of the reaction, and the presence of micromolar concentrations of inorganic pyrophosphate in the cytosol, therefore its role remains unknown. It is suggested that the plant cell may be unable to transport cysteine between the different compartments, so that the cysteine required for protein synthesis must be synthesized in situ, hence the presence of O-acetylserine(thiol)lyase in the three compartments where proteins are synthesized. PMID:16667839

Thiol synthetases of legumes: immunogold localization and differential gene regulation by phytohormones

PubMed Central

Clemente, Maria R.; Bustos-Sanmamed, Pilar; Loscos, Jorge; James, Euan K.; Pérez-Rontomé, Carmen; Navascués, Joaquín; Gay, Marina; Becana, Manuel

2012-01-01

In plants and other organisms, glutathione (GSH) biosynthesis is catalysed sequentially by γ-glutamylcysteine synthetase (γECS) and glutathione synthetase (GSHS). In legumes, homoglutathione (hGSH) can replace GSH and is synthesized by γECS and a specific homoglutathione synthetase (hGSHS). The subcellular localization of the enzymes was examined by electron microscopy in several legumes and gene expression was analysed in Lotus japonicus plants treated for 1–48 h with 50 μM of hormones. Immunogold localization studies revealed that γECS is confined to chloroplasts and plastids, whereas hGSHS is also in the cytosol. Addition of hormones caused differential expression of thiol synthetases in roots. After 24–48 h, abscisic and salicylic acids downregulated GSHS whereas jasmonic acid upregulated it. Cytokinins and polyamines activated GSHS but not γECS or hGSHS. Jasmonic acid elicited a coordinated response of the three genes and auxin induced both hGSHS expression and activity. Results show that the thiol biosynthetic pathway is compartmentalized in legumes. Moreover, the similar response profiles of the GSH and hGSH contents in roots of non-nodulated and nodulated plants to the various hormonal treatments indicate that thiol homeostasis is independent of the nitrogen source of the plants. The differential regulation of the three mRNA levels, hGSHS activity, and thiol contents by hormones indicates a fine control of thiol biosynthesis at multiple levels and strongly suggests that GSH and hGSH play distinct roles in plant development and stress responses. PMID:22442424

A Central Role for Thiols in Plant Tolerance to Abiotic Stress

PubMed Central

Zagorchev, Lyuben; Seal, Charlotte E.; Kranner, Ilse; Odjakova, Mariela

2013-01-01

Abiotic stress poses major problems to agriculture and increasing efforts are being made to understand plant stress response and tolerance mechanisms and to develop new tools that underpin successful agriculture. However, the molecular mechanisms of plant stress tolerance are not fully understood, and the data available is incomplete and sometimes contradictory. Here, we review the significance of protein and non-protein thiol compounds in relation to plant tolerance of abiotic stress. First, the roles of the amino acids cysteine and methionine, are discussed, followed by an extensive discussion of the low-molecular-weight tripeptide, thiol glutathione, which plays a central part in plant stress response and oxidative signalling and of glutathione-related enzymes, including those involved in the biosynthesis of non-protein thiol compounds. Special attention is given to the glutathione redox state, to phytochelatins and to the role of glutathione in the regulation of the cell cycle. The protein thiol section focuses on glutaredoxins and thioredoxins, proteins with oxidoreductase activity, which are involved in protein glutathionylation. The review concludes with a brief overview of and future perspectives for the involvement of plant thiols in abiotic stress tolerance. PMID:23549272

Facile quantitation of free thiols in a recombinant monoclonal antibody by reversed-phase high performance liquid chromatography with hydrophobicity-tailored thiol derivatization.

PubMed

Welch, Leslie; Dong, Xiao; Hewitt, Daniel; Irwin, Michelle; McCarty, Luke; Tsai, Christina; Baginski, Tomasz

2018-06-02

Free thiol content, and its consistency, is one of the product quality attributes of interest during technical development of manufactured recombinant monoclonal antibodies (mAbs). We describe a new, mid/high-throughput reversed-phase-high performance liquid chromatography (RP-HPLC) method coupled with derivatization of free thiols, for the determination of total free thiol content in an E. coli-expressed therapeutic monovalent monoclonal antibody mAb1. Initial selection of the derivatization reagent used an hydrophobicity-tailored approach. Maleimide-based thiol-reactive reagents with varying degrees of hydrophobicity were assessed to identify and select one that provided adequate chromatographic resolution and robust quantitation of free thiol-containing mAb1 forms. The method relies on covalent derivatization of free thiols in denatured mAb1 with N-tert-butylmaleimide (NtBM) label, followed by RP-HPLC separation with UV-based quantitation of native (disulfide containing) and labeled (free thiol containing) forms. The method demonstrated good specificity, precision, linearity, accuracy and robustness. Accuracy of the method, for samples with a wide range of free thiol content, was demonstrated using admixtures as well as by comparison to an orthogonal LC-MS peptide mapping method with isotope tagging of free thiols. The developed method has a facile workflow which fits well into both R&D characterization and quality control (QC) testing environments. The hydrophobicity-tailored approach to the selection of free thiol derivatization reagent is easily applied to the rapid development of free thiol quantitation methods for full-length recombinant antibodies. Copyright © 2018 Elsevier B.V. All rights reserved.

Dynamic asymmetry and the role of the conserved active-site thiol in rabbit muscle creatine kinase.

PubMed

Londergan, Casey H; Baskin, Rachel; Bischak, Connor G; Hoffman, Kevin W; Snead, David M; Reynoso, Christopher

2015-01-13

Symmetric and asymmetric crystal structures of the apo and transition state analogue forms, respectively, of the dimeric rabbit muscle creatine kinase have invoked an "induced fit" explanation for asymmetry between the two subunits and their active sites. However, previously reported thiol reactivity studies at the dual active-site cysteine 283 residues suggest a more latent asymmetry between the two subunits. The role of that highly conserved active-site cysteine has also not been clearly determined. In this work, the S-H vibrations of Cys283 were observed in the unmodified MM isoform enzyme via Raman scattering, and then one and both Cys283 residues in the same dimeric enzyme were modified to covalently attach a cyano group that reports on the active-site environment via its infrared CN stretching absorption band while maintaining the catalytic activity of the enzyme. Unmodified and Cys283-modified enzymes were investigated in the apo and transition state analogue forms of the enzyme. The narrow and invariant S-H vibrational bands report a homogeneous environment for the unmodified active-site cysteines, indicating that their thiols are hydrogen bonded to the same H-bond acceptor in the presence and absence of the substrate. The S-H peak persists at all physiologically relevant pH's, indicating that Cys283 is protonated at all pH's relevant to enzymatic activity. Molecular dynamics simulations identify the S-H hydrogen bond acceptor as a single, long-resident water molecule and suggest that the role of the conserved yet catalytically unnecessary thiol may be to dynamically rigidify that part of the active site through specific H-bonding to water. The asymmetric and broad CN stretching bands from the CN-modified Cys283 suggest an asymmetric structure in the apo form of the enzyme in which there is a dynamic exchange between spectral subpopulations associated with water-exposed and water-excluded probe environments. Molecular dynamics simulations indicate a

Quantitative interference by cysteine and N-acetylcysteine metabolites during the LC-MS/MS bioanalysis of a small molecule.

PubMed

Barricklow, Jason; Ryder, Tim F; Furlong, Michael T

2009-08-01

During LC-MS/MS quantification of a small molecule in human urine samples from a clinical study, an unexpected peak was observed to nearly co-elute with the analyte of interest in many study samples. Improved chromatographic resolution revealed the presence of at least 3 non-analyte peaks, which were identified as cysteine metabolites and N-acetyl (mercapturic acid) derivatives thereof. These metabolites produced artifact responses in the parent compound MRM channel due to decomposition in the ionization source of the mass spectrometer. Quantitative comparison of the analyte concentrations in study samples using the original chromatographic method and the improved chromatographic separation method demonstrated that the original method substantially over-estimated the analyte concentration in many cases. The substitution of electrospray ionization (ESI) for atmospheric pressure chemical ionization (APCI) nearly eliminated the source instability of these metabolites, which would have mitigated their interference in the quantification of the analyte, even without chromatographic separation. These results 1) demonstrate the potential for thiol metabolite interferences during the quantification of small molecules in pharmacokinetic samples, and 2) underscore the need to carefully evaluate LC-MS/MS methods for molecules that can undergo metabolism to thiol adducts to ensure that they are not susceptible to such interferences during quantification.

Thiol peroxidases mediate specific genome-wide regulation of gene expression in response to hydrogen peroxide

PubMed Central

Fomenko, Dmitri E.; Koc, Ahmet; Agisheva, Natalia; Jacobsen, Michael; Kaya, Alaattin; Malinouski, Mikalai; Rutherford, Julian C.; Siu, Kam-Leung; Jin, Dong-Yan; Winge, Dennis R.; Gladyshev, Vadim N.

2011-01-01

Hydrogen peroxide is thought to regulate cellular processes by direct oxidation of numerous cellular proteins, whereas antioxidants, most notably thiol peroxidases, are thought to reduce peroxides and inhibit H2O2 response. However, thiol peroxidases have also been implicated in activation of transcription factors and signaling. It remains unclear if these enzymes stimulate or inhibit redox regulation and whether this regulation is widespread or limited to a few cellular components. Herein, we found that Saccharomyces cerevisiae cells lacking all eight thiol peroxidases were viable and withstood redox stresses. They transcriptionally responded to various redox treatments, but were unable to activate and repress gene expression in response to H2O2. Further studies involving redox transcription factors suggested that thiol peroxidases are major regulators of global gene expression in response to H2O2. The data suggest that thiol peroxidases sense and transfer oxidative signals to the signaling proteins and regulate transcription, whereas a direct interaction between H2O2 and other cellular proteins plays a secondary role. PMID:21282621

Secondary Metabolites from Three Florida Sponges with Antidepressant Activity

PubMed Central

Kochanowska, Anna J.; Rao, Karumanchi V.; Childress, Suzanne; El-Alfy, Abir; Matsumoto, Rae R.; Kelly, Michelle; Stewart, Gina S.; Sufka, Kenneth J.; Hamann, Mark T.

2016-01-01

Brominated indole alkaloids are a common class of metabolites reported from sponges of the order Verongida. Herein we report the isolation, structure determination, and activity of metabolites from three Florida sponges, namely, Verongula rigida (order Verongida, family Aplysinidae), Smenospongia aurea, and S. cerebriformis (order Dictyoceratida, family Thorectidae). All three species were investigated chemically, revealing similarities in secondary metabolites. Brominated compounds, as well as sesquiterpene quinones and hydroquinones, were identified from both V. rigida and S. aurea despite their apparent taxonomic differences at the ordinal level. Similar metabolites found in these distinct sponge species of two different genera provide evidence for a microbial origin of the metabolites. Isolated compounds were evaluated in the Porsolt forced swim test (FST) and the chick anxiety–depression continuum model. Among the isolated compounds, 5,6-dibromo-N,N-dimethyltryptamine (1) exhibited significant antidepressant-like action in the rodent FST model, while 5-bromo-N,N-dimethyltryptamine (2) caused significant reduction of locomotor activity indicative of a potential sedative action. The current study provides ample evidence that marine natural products with the diversity of brominated marine alkaloids will provide potential leads for antidepressant and anxiolytic drugs. PMID:18217716

Secondary metabolites from three Florida sponges with antidepressant activity.

PubMed

Kochanowska, Anna J; Rao, Karumanchi V; Childress, Suzanne; El-Alfy, Abir; Matsumoto, Rae R; Kelly, Michelle; Stewart, Gina S; Sufka, Kenneth J; Hamann, Mark T

2008-02-01

Brominated indole alkaloids are a common class of metabolites reported from sponges of the order Verongida. Herein we report the isolation, structure determination, and activity of metabolites from three Florida sponges, namely, Verongula rigida (order Verongida, family Aplysinidae), Smenospongia aurea, and S. cerebriformis (order Dictyoceratida, family Thorectidae). All three species were investigated chemically, revealing similarities in secondary metabolites. Brominated compounds, as well as sesquiterpene quinones and hydroquinones, were identified from both V. rigida and S. aurea despite their apparent taxonomic differences at the ordinal level. Similar metabolites found in these distinct sponge species of two different genera provide evidence for a microbial origin of the metabolites. Isolated compounds were evaluated in the Porsolt forced swim test (FST) and the chick anxiety-depression continuum model. Among the isolated compounds, 5,6-dibromo- N,N-dimethyltryptamine ( 1) exhibited significant antidepressant-like action in the rodent FST model, while 5-bromo- N,N-dimethyltryptamine ( 2) caused significant reduction of locomotor activity indicative of a potential sedative action. The current study provides ample evidence that marine natural products with the diversity of brominated marine alkaloids will provide potential leads for antidepressant and anxiolytic drugs.

Dynamic thiol/disulphide homeostasis in patients with basal cell carcinoma.

PubMed

Demirseren, Duriye Deniz; Cicek, Cagla; Alisik, Murat; Demirseren, Mustafa Erol; Aktaş, Akın; Erel, Ozcan

2017-09-01

The aim of this study is to measure and compare the dynamic thiol/disulphide homeostasis of patients with basal cell carcinoma and healthy subjects with a newly developed and original method. Thirty four patients attending our outpatient clinic and clinically and histopathologically diagnosed as nodular basal cell carcinoma, and age and gender matched 30 healthy individuals have been involved in the study. Thiol/disulphide homeostasis tests have been measured with a novel automatic spectrophotometric method developed and the results have been compared statistically. Serum native thiol and disulphide levels in the patient and control group show a considerable variance statistically (p = 0.028, 0.039, respectively). Total thiol levels do not reveal a considerable variation (p = 0.094). Disulphide/native thiol ratios and native thiol/total thiol ratios also show a considerable variance statistically (p = 0.012, 0.013, 0.010, respectively). Thiol disulphide homeostasis in patients with basal cell carcinoma alters in the way that disulphide gets lower and thiols get higher. Thiol/disulphide level is likely to have a role in basal cell carcinoma pathogenesis.

Pathway Activity Profiling (PAPi): from the metabolite profile to the metabolic pathway activity.

PubMed

Aggio, Raphael B M; Ruggiero, Katya; Villas-Bôas, Silas Granato

2010-12-01

Metabolomics is one of the most recent omics-technologies and uses robust analytical techniques to screen low molecular mass metabolites in biological samples. It has evolved very quickly during the last decade. However, metabolomics datasets are considered highly complex when used to relate metabolite levels to metabolic pathway activity. Despite recent developments in bioinformatics, which have improved the quality of metabolomics data, there is still no straightforward method capable of correlating metabolite level to the activity of different metabolic pathways operating within the cells. Thus, this kind of analysis still depends on extremely laborious and time-consuming processes. Here, we present a new algorithm Pathway Activity Profiling (PAPi) with which we are able to compare metabolic pathway activities from metabolite profiles. The applicability and potential of PAPi was demonstrated using a previously published data from the yeast Saccharomyces cerevisiae. PAPi was able to support the biological interpretations of the previously published observations and, in addition, generated new hypotheses in a straightforward manner. However, PAPi is time consuming to perform manually. Thus, we also present here a new R-software package (PAPi) which implements the PAPi algorithm and facilitates its usage to quickly compare metabolic pathways activities between different experimental conditions. Using the identified metabolites and their respective abundances as input, the PAPi package calculates pathways' Activity Scores, which represents the potential metabolic pathways activities and allows their comparison between conditions. PAPi also performs principal components analysis and analysis of variance or t-test to investigate differences in activity level between experimental conditions. In addition, PAPi generates comparative graphs highlighting up- and down-regulated pathway activity. These datasets are available in http://www.4shared

Are free radicals involved in thiol-based redox signaling?

PubMed

Winterbourn, Christine C

2015-03-01

Cells respond to many stimuli by transmitting signals through redox-regulated pathways. It is generally accepted that in many instances signal transduction is via reversible oxidation of thiol proteins, although there is uncertainty about the specific redox transformations involved. The prevailing view is that thiol oxidation occurs by a two electron mechanism, most commonly involving hydrogen peroxide. Free radicals, on the other hand, are considered as damaging species and not generally regarded as important in cell signaling. This paper examines whether it is justified to dismiss radicals or whether they could have a signaling role. Although there is no direct evidence that radicals are involved in transmitting thiol-based redox signals, evidence is presented that they are generated in cells when these signaling pathways are activated. Radicals produce the same thiol oxidation products as two electron oxidants, although by a different mechanism, and at this point radical-mediated pathways should not be dismissed. There are unresolved issues about how radical mechanisms could achieve sufficient selectivity, but this could be possible through colocalization of radical-generating and signal-transducing proteins. Colocalization is also likely to be important for nonradical signaling mechanisms and identification of such associations should be a priority for advancing the field. Copyright © 2014 Elsevier Inc. All rights reserved.

Depletion of Intracellular Thiols and Increased Production of 4-Hydroxynonenal that Occur During Cryopreservation of Stallion Spermatozoa Lead to Caspase Activation, Loss of Motility, and Cell Death.

PubMed

Martin Muñoz, Patricia; Ortega Ferrusola, Cristina; Vizuete, Guillermo; Plaza Dávila, Maria; Rodriguez Martinez, Heriberto; Peña, Fernando J

2015-12-01

Oxidative stress has been linked to sperm death and the accelerated senescence of cryopreserved spermatozoa. However, the molecular mechanisms behind this phenomenon remain poorly understood. Reactive oxygen species (ROS) are considered relevant signaling molecules for sperm function, only becoming detrimental when ROS homeostasis is lost. We hereby hypothesize that a major component of the alteration of ROS homeostasis in cryopreserved spermatozoa is the exhaustion of intrinsic antioxidant defense mechanisms. To test this hypothesis, semen from seven stallions was frozen using a standard technique. The parameters of sperm quality (motility, velocity, and membrane integrity) and markers of sperm senescence (caspase 3, 4-hydroxynonenal, and mitochondrial membrane potential) were assessed before and after cryopreservation. Changes in the intracellular thiol content were also monitored. Cryopreservation caused significant increases in senescence markers as well as dramatic depletion of intracellular thiols to less than half of the initial values (P < 0.001) postthaw. Interestingly, very high and positive correlations were observed among thiol levels with sperm functionality postthaw: total motility (r = 0.931, P < 0.001), progressive motility (r = 0.904, P < 0.001), and percentage of live spermatozoa without active caspase 3 (r = 0.996, P < 0.001). In contrast, negative correlations were detected between active caspase 3 and thiol content both in living (r = -0.896) and dead (r = -0.940) spermatozoa; additionally, 4-hydroxynonenal levels were negatively correlated with thiol levels (r = -0.856). In conclusion, sperm functionality postthaw correlates with the maintenance of adequate levels of intracellular thiols. The accelerated senescence of thawed spermatozoa is related to oxidative and electrophilic stress induced by increased production of 4-hydroxynoneal in thawed samples once intracellular thiols are depleted. © 2015 by the Society for the Study of Reproduction

Kinetic Resolution of sec-Thiols by Enantioselective Oxidation with Rationally Engineered 5-(Hydroxymethyl)furfural Oxidase.

PubMed

Pickl, Mathias; Swoboda, Alexander; Romero, Elvira; Winkler, Christoph K; Binda, Claudia; Mattevi, Andrea; Faber, Kurt; Fraaije, Marco W

2018-03-05

Various flavoprotein oxidases were recently shown to oxidize primary thiols. Herein, this reactivity is extended to sec-thiols by using structure-guided engineering of 5-(hydroxymethyl)furfural oxidase (HMFO). The variants obtained were employed for the oxidative kinetic resolution of racemic sec-thiols, thus yielding the corresponding thioketones and nonreacted R-configured thiols with excellent enantioselectivities (E≥200). The engineering strategy applied went beyond the classic approach of replacing bulky amino acid residues with smaller ones, as the active site was additionally enlarged by a newly introduced Thr residue. This residue established a hydrogen-bonding interaction with the substrates, as verified in the crystal structure of the variant. These strategies unlocked HMFO variants for the enantioselective oxidation of a range of sec-thiols. © 2018 Wiley-VCH Verlag GmbH & Co. KGaA, Weinheim.

The Expanding Landscape of the Thiol Redox Proteome*

PubMed Central

Yang, Jing; Carroll, Kate S.; Liebler, Daniel C.

2016-01-01

Cysteine occupies a unique place in protein chemistry. The nucleophilic thiol group allows cysteine to undergo a broad range of redox modifications beyond classical thiol-disulfide redox equilibria, including S-sulfenylation (-SOH), S-sulfinylation (-SO2H), S-sulfonylation (-SO3H), S-nitrosylation (-SNO), S-sulfhydration (-SSH), S-glutathionylation (-SSG), and others. Emerging evidence suggests that these post-translational modifications (PTM) are important in cellular redox regulation and protection against oxidative damage. Identification of protein targets of thiol redox modifications is crucial to understanding their roles in biology and disease. However, analysis of these highly labile and dynamic modifications poses challenges. Recent advances in the design of probes for thiol redox forms, together with innovative mass spectrometry based chemoproteomics methods make it possible to perform global, site-specific, and quantitative analyses of thiol redox modifications in complex proteomes. Here, we review chemical proteomic strategies used to expand the landscape of thiol redox modifications. PMID:26518762

Can thiol compounds be used as biomarkers of aquatic ecosystem contamination by cadmium?

PubMed Central

Kovářová, Jana; Svobodová, Zdeňka

2009-01-01

Due to anthropogenic activities, heavy metals still represent a threat for various trophic levels. If aquatic animals are exposed to heavy metals we can obviously observe considerable toxicity. It is well known that an organism affected by cadmium (Cd) synthesize low molecular mass thiol compounds rich in cysteine (Cys), such as metallothioneins (MT) and glutathione (GSH/GSSG). The aim of this study was to summarize the effect of Cd on level of thiol compounds in aquatic organisms, and evaluate that the concentrations of thiol compounds are effective indicators of Cd water pollution and explain their potential use in biomonitoring applications. PMID:21217850

Hydrophobic thiol-ene surfaces fabricated via plasma activation and photo polymerization

NASA Astrophysics Data System (ADS)

Champathet, P.; Ervithayasuporn, V.; Osotchan, T.; Dangtip, S.

2017-09-01

Alumina, such as glazed alumina for electrical insulator, operated in an open field subjects to a very harsh condition; resulting in lifetime shortening. Coating hydrophobic layer on alumina surface can help prolonging its lifetime. In this study, 25 ×25 mm alumina sheets were used as substrates. The hydrophobic composite polymers were prepared from (3-mercaptopropyl)trimethoxysilane(MPTMS), 2,4,6,8-tetramethyl-2,4,6,8tetravinylcyclotetra siloxane(TMTVSi), pentaerythritoltetra(3-mercaptopropionate)(PETMP), 2,2-dimethoxy-2-phe nylaceto phenone(photoinitiator) and heptadecafluorodecylmethacrylate(HEFDMA) via the thiol-ene reaction. The alumina sheets were first activated by dielectric-barrier discharge plasma to improve its adhesion. All the polymers were found to optimize at the ratio of (MPTMS:TMTVSi:PETMP:HDFDMA) to 4:2:1:2 for coating on the alumina substrate. To enhance polymerization, 2,2-dimethoxy-2-phenylaceto phenome was also used as a photoinitiator A proper mixing sequence in the thiol-ene reaction results in film with excellent surface retention after prolong soaking in solvent such as acetone. FTIR shows that S-H and C=C functional groups have significantly changed after photopolymerization and thermally cured. The static contact angle increase from mere 53.0°±1.5° of the uncoated substrate to 120.0°±1.2° after coating. SEM shows the film with clear appearance of a few-micron thick. Under AFM, the coated surface roughness was about 9.3 nm with evenly distributed spikes of a few nanometer in height. The cross-cut test also confirmed the film was very smooth and none of the square of the films detached.

Evolution of thiol protective systems in prokaryotes

NASA Technical Reports Server (NTRS)

Fahey, R. C.; Newton, G. L.

1986-01-01

Biological thiols are essential elements in most aspects of cell function but undergo rapid oxidation to disulfides in the presence of oxygen. The evolution of systems to protect against such oxygen toxicity was essential to the emergence of aerobic life. The protection system used by eukaryotes is based upon glutathione (GSH) and GSH-dependent enzymes but many bacteria lack GSH and apparently use other mechanisms. The objective of this research is to elaborate the thiol protective mechanisms employed by prokaryotes of widely divergent evolutionary origin and to understand why GSH became the central thiol employed in essentially all higher organisms. Thiol-selective fluorescent labeling and HPLC analysis has been used to determine key monothiol components.

EFFECTS OF THIOL ANTIOXIDANTS ON THE ATROPSELECTIVE OXIDATION OF 2,2′,3,3′,6,6′-HEXACHLOROBIPHENYL (PCB 136) BY RAT LIVER MICROSOMES

PubMed Central

Wu, Xianai; Lehmler, Hans-Joachim

2015-01-01

Chiral polychlorinated biphenyl (PCB) congeners, such as PCB 136, are atropselectively metabolized to various hydroxylated PCB metabolites (HO-PCBs). The present study investigates the effect of two thiol antioxidants, glutathione and N-acetyl-cysteine (NAC), on profiles and chiral signatures of PCB 136 and its HO-PCB metabolites in rat liver microsomal incubations. Liver microsomes prepared from rats pretreated with phenobarbital were incubated with PCB 136 (5 μM) in the presence of the respective antioxidant (0–10 mM), and levels and chiral signatures of PCB 136 and its HO-PCB metabolites were determined. Three metabolites, 5-136 (2,2′,3,3′,6,6′-hexachlorobiphenyl-5-ol), 4-136 (2,2′,3,3′,6,6′-hexachlorobiphenyl-4-ol) and 4,5-136 (2,2′,3,3′,6,6′-hexachlorobiphenyl-4,5-diol), were detected in all incubations, with 5-136 being the major metabolite. Compared to microsomal incubations without antioxidant, levels of 4,5-136 increased with increasing antioxidant concentration, whereas levels of PCB 136 and both mono-HO-PCBs were not affected by the presence of either antioxidant. PCB 136, 4-136 and 5-136 displayed significant atropisomeric enrichment; however, the direction and extent of the atropisomeric enrichment was not altered in the presence of an antioxidant. Because 4,5-136 can either be conjugated to a sulfate or glucuronide metabolite that is readily excreted or further oxidized a potentially toxic PCB 136 quinone, the effect of both thiol antioxidants on 4,5-136 formation suggests that disruptions of glutathione homeostasis may alter the balance between both metabolic pathways and, thus, PCB 136 toxicity in vivo. PMID:26155892

Purification and Kinetic Properties of Serine Acetyltransferase Free of O-Acetylserine(thiol)lyase from Spinach Chloroplasts.

PubMed

Ruffet, M. L.; Droux, M.; Douce, R.

1994-02-01

Serine acetyltransferase, a key enzyme in the L-cysteine biosynthetic pathway, was purified over 300,000-fold from the stroma of spinach (Spinacia oleracea) leaf chloroplasts. The purification procedure consisted of ammonium sulfate precipitation, anion-exchange chromatography (Trisacryl M DEAE and Mono Q HR10/10), hydroxylapatite chromatography, and gel filtration (Superdex 200). The purified enzyme exhibited a specific activity higher than 200 units mg-1 and a subunit molecular mass of about 33 kD upon polyacrylamide gel electrophoresis in the presence of sodium dodecyl sulfate. Moreover, the purified serine acetyltransferase appeared to be essentially free of O-acetyleserine(thiol)lyase, another enzyme component in the L-cysteine biosynthetic pathway. A steady-state kinetic analysis indicated that the mechanism of the enzyme-catalyzed reaction involves a double displacement. The apparent Km for the two substrates, L-serine and acetyl-coenzyme A, were 2.29 [plus or minus] 0.43 and 0.35 [plus or minus] 0.02 mM, respectively. The rate of L-cysteine synthesis in vitro was measured in a coupled enzyme assay using extensively purified O-acetylserine(thiol)lyase and serine acetyltransferase. This rate was maximum when the assay contained approximately a 400-fold excess of O-acetylserine(thiol)lyase over serine acetyltransferase. Measurements of the relative level of O-acetylserine(thiol)lyase and serine acetyltransferase activities in the stroma indicated that the former enzyme was present in much larger quantities than the latter. Thus, the activity ratio for these two enzymes [O-acetylserine(thiol)lyase activity/serine acetyltransferase activity] measured in the stromal protein extract was 345. This strongly suggested that all the O-acetylserine(thiol)lyase and serine acetyltransferase activities in the stroma are involved in bringing a full synthesis of L-cysteine in the chloroplast.

Purification and Kinetic Properties of Serine Acetyltransferase Free of O-Acetylserine(thiol)lyase from Spinach Chloroplasts.

PubMed Central

Ruffet, M. L.; Droux, M.; Douce, R.

1994-01-01

Serine acetyltransferase, a key enzyme in the L-cysteine biosynthetic pathway, was purified over 300,000-fold from the stroma of spinach (Spinacia oleracea) leaf chloroplasts. The purification procedure consisted of ammonium sulfate precipitation, anion-exchange chromatography (Trisacryl M DEAE and Mono Q HR10/10), hydroxylapatite chromatography, and gel filtration (Superdex 200). The purified enzyme exhibited a specific activity higher than 200 units mg-1 and a subunit molecular mass of about 33 kD upon polyacrylamide gel electrophoresis in the presence of sodium dodecyl sulfate. Moreover, the purified serine acetyltransferase appeared to be essentially free of O-acetyleserine(thiol)lyase, another enzyme component in the L-cysteine biosynthetic pathway. A steady-state kinetic analysis indicated that the mechanism of the enzyme-catalyzed reaction involves a double displacement. The apparent Km for the two substrates, L-serine and acetyl-coenzyme A, were 2.29 [plus or minus] 0.43 and 0.35 [plus or minus] 0.02 mM, respectively. The rate of L-cysteine synthesis in vitro was measured in a coupled enzyme assay using extensively purified O-acetylserine(thiol)lyase and serine acetyltransferase. This rate was maximum when the assay contained approximately a 400-fold excess of O-acetylserine(thiol)lyase over serine acetyltransferase. Measurements of the relative level of O-acetylserine(thiol)lyase and serine acetyltransferase activities in the stroma indicated that the former enzyme was present in much larger quantities than the latter. Thus, the activity ratio for these two enzymes [O-acetylserine(thiol)lyase activity/serine acetyltransferase activity] measured in the stromal protein extract was 345. This strongly suggested that all the O-acetylserine(thiol)lyase and serine acetyltransferase activities in the stroma are involved in bringing a full synthesis of L-cysteine in the chloroplast. PMID:12232109

Electrophilic Triterpenoid Enones: A Comparative Thiol-Trapping and Bioactivity Study.

PubMed

Del Prete, Danilo; Taglialatela-Scafati, Orazio; Minassi, Alberto; Sirignano, Carmina; Cruz, Cristina; Bellido, Maria L; Muñoz, Eduardo; Appendino, Giovanni

2017-08-25

Bardoxolone methyl (1) is the quintessential member of triterpenoid cyanoacrylates, an emerging class of bioactive compounds capable of transient covalent binding to thiols. The mechanistic basis for this unusual "pulsed reactivity" profile and the mode of its biological translation are unknown. To provide clues on these issues, a series of Δ 1 -dehydrooleanolates bearing an electron-withdrawing group at C-2 (7a-m) were prepared from oleanolic acid (3a) and comparatively investigated in terms of reactivity with thiols and bioactivity against a series of electrophile-sensitive transcription factors (Nrf2, NF-κB, STAT3). The emerging picture suggests that the triterpenoid scaffold sharply decreases the reactivity of the enone system by steric encumbrance and that only strongly electrophilic and sterically undemanding substituents such as a cyanide or a carboxylate group can re-establish Michael reactivity, albeit in a transient way for the cyanide group. In general, a substantial dissection between the thiol-trapping ability and the modulation of biological end-points sensitive to thiol alkylation was observed, highlighting the role of shape complementarity for the activity of triterpenoid thia-Michael acceptors.

Presence of closely spaced protein thiols on the surface of mammalian cells.

PubMed Central

Donoghue, N.; Yam, P. T.; Jiang, X. M.; Hogg, P. J.

2000-01-01

It has been proposed that certain cell-surface proteins undergo redox reactions, that is, transfer of hydrogens and electrons between closely spaced cysteine thiols that can lead to reduction, formation, or interchange of disulfide bonds. This concept was tested using a membrane-impermeable trivalent arsenical to identify closely spaced thiols in cell-surface proteins. We attached the trivalent arsenical, phenylarsenoxide, to the thiol of reduced glutathione to produce 4-(N-(S-glutathionylacetyl)amino)phenylarsenoxide (GSAO). GSAO bound tightly to synthetic, peptide, and protein dithiols like thioredoxin, but not to monothiols. To identify cell-surface proteins that contain closely spaced thiols, we attached a biotin moiety through a spacer arm to the primary amino group of the gamma-glutamyl residue of GSAO (GSAO-B). Incorporation of GSAO-B into proteins was assessed by measuring the biotin using streptavidin-peroxidase. Up to 12 distinct proteins were labeled with GSAO-B on the surface of endothelial and fibrosarcoma cells. The pattern of labeled proteins differed between the different cell types. Protein disulfide isomerase was one of the proteins on the endothelial and fibrosarcoma cell surface that incorporated GSAO-B. These findings demonstrate that the cell-surface environment can support the existence of closely spaced protein thiols and suggest that at least some of these thiols are redox active. PMID:11206065

Inactivation of Penicillins by Thiol Broth

PubMed Central

Murray, Patrick R.; Niles, Ann C.

1982-01-01

Thiol broth with sodium polyanetholesulfonate inactivated penicillin G, carbenicillin, nafcillin, oxacillin, and gentamicin, but had no effect on cephalothin, cefoxitin, clindamycin, chloramphenicol, erythromycin, and tetracycline. Only Thiol broth was capable of this inactivation, which was not influenced by the presence of blood. PMID:7153352

Purification and characterization of O-acetylserine (thiol) lyase from spinach chloroplasts.

PubMed

Droux, M; Martin, J; Sajus, P; Douce, R

1992-06-01

O-Acetylserine (thiol) lyase, the last enzyme in the cysteine biosynthetic pathway, was purified to homogeneity from spinach leaf chloroplasts. The enzyme has a molecular mass of 68,000 and consists of two identical subunits of Mr 35,000. The absorption spectrum obtained at pH 7.5 exhibited a peak at 407 nm due to pyridoxal phosphate, and addition of O-acetylserine induced a considerable modification of the spectrum. The pyridoxal phosphate content was found to be 1.1 per subunit of 35,000, and the chromophore was displaced from the enzyme by O-acetylserine, leading to a progressive inactivation of the holoenzyme. Upon gel filtration chromatography on Superdex 200, part of the chloroplastic O-acetylserine (thiol) lyase eluted in association with serine acetyltransferase at a position corresponding to a molecular mass of 310,000 (such a complex called cysteine synthase has been characterized in bacteria). The activity of O-acetylserine (thiol) lyase was optimum between pH 7.5 and 8.5. The apparent Km for O-acetylserine was 1.3 mM and for sulfide was 0.25 mM. The calculated activation energy was 12.6 kcal/mol at 10 mM O-acetylserine. The overall amino-acid composition of spinach chloroplast O-acetylserine (thiol) lyase was different than that determined for the same enzyme (cytosolic?) obtained from a crude extract of spinach leaves. A polyclonal antibody prepared against the chloroplastic O-acetylserine (thiol) lyase exhibited a very low cross-reactivity with a preparation of mitochondrial matrix and cytosolic proteins suggesting that the chloroplastic isoform was distinct from the mitochondrial and cytosolic counterparts.

Redox regulation of mitochondrial proteins and proteomes by cysteine thiol switches.

PubMed

Nietzel, Thomas; Mostertz, Jörg; Hochgräfe, Falko; Schwarzländer, Markus

2017-03-01

Mitochondria are hotspots of cellular redox biochemistry. Respiration as a defining mitochondrial function is made up of a series of electron transfers that are ultimately coupled to maintaining the proton motive force, ATP production and cellular energy supply. The individual reaction steps involved require tight control and flexible regulation to maintain energy and redox balance in the cell under fluctuating demands. Redox regulation by thiol switching has been a long-standing candidate mechanism to support rapid adjustment of mitochondrial protein function at the posttranslational level. Here we review recent advances in our understanding of cysteine thiol switches in the mitochondrial proteome with a focus on their operation in vivo. We assess the conceptual basis for thiol switching in mitochondria and discuss to what extent insights gained from in vitro studies may be valid in vivo, considering thermodynamic, kinetic and structural constraints. We compare functional proteomic approaches that have been used to assess mitochondrial protein thiol switches, including thioredoxin trapping, redox difference gel electrophoresis (redoxDIGE), isotope-coded affinity tag (OxICAT) and iodoacetyl tandem mass tag (iodoTMT) labelling strategies. We discuss conditions that may favour active thiol switching in mitochondrial proteomes in vivo, and appraise recent advances in dissecting their impact using combinations of in vivo redox sensing and quantitative redox proteomics. Finally we focus on four central facets of mitochondrial biology, aging, carbon metabolism, energy coupling and electron transport, exemplifying the current emergence of a mechanistic understanding of mitochondrial regulation by thiol switching in living plants and animals. Copyright © 2016 Elsevier B.V. and Mitochondria Research Society. All rights reserved.

Radicals are required for thiol etching of gold particles

PubMed Central

Dreier, Timothy A.

2016-01-01

Etching of gold with excess thiol ligand is used in both synthesis and analysis of gold particles. Mechanistically, the process of etching gold with excess thiol is opaque. Previous studies have obliquely considered the role of oxygen in thiolate etching of gold. Herein, we show that oxygen or a radical initator is a necessary component for efficient etching of gold by thiolates. Attenuation of the etching process by radical scavengers in the presence of oxygen, and the restoration of activity by radical initiators under inert atmosphere, strongly implicate the oxygen radical. These data led us to propose an atomistic mechanism in which the oxygen radical initiates the etching process. PMID:26089294

An interactive human carbonic anhydrase-II (hCA-II) receptor--pharmacophore molecular model & anti-convulsant activity of the designed and synthesized 5-amino-1,3,4-thiadiazole-2-thiol conjugated imine derivatives.

PubMed

Yusuf, Mohammad; Khan, Riaz A; Khan, Maria; Ahmed, Bahar

2013-05-01

New imines, derived from aromatic aldehyde, chalcones and 5-amino-1,3,4-thiadiazole-2-thiol exhibited promising anti-convulsant activity which is explained through chemo-biological interactions at receptor site producing the inhibition of human Carbonic Anhydrase-II enzyme (hCA-II) through the proposed pharmacophore model at molecular levels as basis for pharmacological activity. The compounds 5-{1-(4-Chlorophenyl)-3-[4-(methoxy-phenyl)-prop-2-en-1-ylidene]amino}-1,3,4-thiadiazole-2-thiol (2b), 5-{[1-(4-chloro-phenyl)]-3-[4-(dimethyl-amino-phenyl)-prop-2-en-1-ylidene]amino}-1,3,4-thiadiazole-2-thiol (2c) and 5-{[1-(4-chloro-phenyl)]-3-[(4-amino-phenyl)-prop-2-en-1-ylidene]amino}-1,3,4-thiadiazole-2-thiol (2f) showed 100% activity in comparison with standard Acetazolamide, a known anti-convulsant drug. The compounds 2c, 2f also passed the Rotarod and Ethanol Potentiation tests which further confirmed them to be safe in motor coordination activity and safe from generating neurological toxicity. © 2013 John Wiley & Sons A/S.

Antioxidant activity of selenenamide-based mimic as a function of the aromatic thiols nucleophilicity, a DFT-SAPE model.

PubMed

Kheirabadi, Ramesh; Izadyar, Mohammad

2018-05-18

The mechanism of action of the selenenamide 1 as a mimic of the glutathione peroxidase (GPx) was investigated by the density functional theory. The solvent-assisted proton exchange procedure was applied to model the catalytic behavior and antioxidant activity of this mimic. To have an insight into the charge transfer effect, different aromatic thiols, including electron donating substituents on the phenyl ring were considered. The catalytic behavior of the selenenamide was modeled in a four-step mechanism, described by the oxidation of the mimic, the reduction of the obtained product, selenoxide, the reduction of the selenenylsulfide and dehydration of selenenic acid. On the basis of the activation parameters, the final step of the proposed mechanism is the rate determining states of the catalytic cycle. Turnover frequency (TOF) analysis showed that the electron donating groups at the para-position of the phenyl ring of the PhSH do not affect the catalytic activity of the selenenamide in contrast to p-methyl thiophenol which indicates the highest nucleophilicity. The evaluation of the electronic contribution of the various donating groups on the phenyl ring of the aromatic thiols shows that the antioxidant activity of the selenenamide sufficiently increases in the presence of the electron-donating substitutions. Finally, the charge transfer process at the rate-determining state was investigated based on the natural bond orbital analysis. Copyright © 2018 Elsevier Ltd. All rights reserved.

Transdermal thiol-acrylate polyethylene glycol hydrogel synthesis using near infrared light

NASA Astrophysics Data System (ADS)

Chung, Solchan; Lee, Hwangjae; Kim, Hyung-Seok; Kim, Min-Gon; Lee, Luke P.; Lee, Jae Young

2016-07-01

Light-induced polymerization has been widely applied for hydrogel synthesis, which conventionally involves the use of ultraviolet or visible light to activate a photoinitiator for polymerization. However, with these light sources, transdermal gelation is not efficient and feasible due to their substantial interactions with biological systems, and thus a high power is required. In this study, we used biocompatible and tissue-penetrating near infrared (NIR) light to remotely trigger a thiol-acrylate reaction for efficient in vivo gelation with good controllability. Our gelation system includes gold nanorods as a photothermal agent, a thermal initiator, diacrylate polyethylene glycol (PEG), and thiolated PEG. Irradiation with a low-power NIR laser (0.3 W cm-2) could induce gelation via a mixed-mode reaction with a small increase in temperature (~5 °C) under the optimized conditions. We also achieved successful transdermal gelation via the NIR-assisted photothermal thiol-acryl reactions. This new type of NIR-assisted thiol-acrylate polymerization provides new opportunities for in situ hydrogel formation for injectable hydrogels and delivery of drugs/cells for various biomedical applications.Light-induced polymerization has been widely applied for hydrogel synthesis, which conventionally involves the use of ultraviolet or visible light to activate a photoinitiator for polymerization. However, with these light sources, transdermal gelation is not efficient and feasible due to their substantial interactions with biological systems, and thus a high power is required. In this study, we used biocompatible and tissue-penetrating near infrared (NIR) light to remotely trigger a thiol-acrylate reaction for efficient in vivo gelation with good controllability. Our gelation system includes gold nanorods as a photothermal agent, a thermal initiator, diacrylate polyethylene glycol (PEG), and thiolated PEG. Irradiation with a low-power NIR laser (0.3 W cm-2) could induce gelation

Distribution and abundance of organic thiols

NASA Technical Reports Server (NTRS)

Fahey, R.

1985-01-01

The role of glutathione (GSH) in protecting against the toxicity of oxygen and oxygen by products is well established for all eukaryotes studied except Entamoeba histolytica which lacks mitochrondria, chloroplasts, and microtubules. The GSH is not universal among prokaryotes. Entamoeba histolytica does not produce GSH or key enzymes of GSH metabolism. A general method of thiol analysis based upon fluorescent labeling with monobromobimane and HPLC separation of the resulting thiol derivatives was developed to determine the occurrence of GSH and other low molecular weight thiols in bacteria. Glutathione is the major thiol in cyanobacteria and in most bacteria closely related to the purple photosynthetic bacteria, but GSH was not found in archaebacteria, green bacteria, or GRAM positive bacteria. It suggested that glutathione metabolism was incorporated into eukaryotes at the time that mitochondria and chloroplasts were acquired by endosymbiosis. In Gram positive aerobes, coenzyme A occurs at millimolar levels and CoA disulfide reductases are identified. The CoA, rather than glutathione, may function in the oxygen detoxification processes of these organisms.

A Search for Interstellar Monohydric Thiols

NASA Astrophysics Data System (ADS)

Gorai, Prasanta; Das, Ankan; Das, Amaresh; Sivaraman, Bhalamurugan; Etim, Emmanuel E.; Chakrabarti, Sandip K.

2017-02-01

It has been pointed out by various astronomers that a very interesting relationship exists between interstellar alcohols and the corresponding thiols (sulfur analog of alcohols) as far as the spectroscopic properties and chemical abundances are concerned. Monohydric alcohols such as methanol and ethanol are widely observed and 1-propanol was recently claimed to have been seen in Orion KL. Among the monohydric thiols, methanethiol (chemical analog of methanol) has been firmly detected in Orion KL and Sgr B2(N2) and ethanethiol (chemical analog of ethanol) has been observed in Sgr B2(N2), though the confirmation of this detection is yet to come. It is very likely that higher order thiols could be observed in these regions. In this paper, we study the formation of monohydric alcohols and their thiol analogs. Based on our quantum chemical calculation and chemical modeling, we find that the Tg conformer of 1-propanethiol is a good candidate of astronomical interest. We present various spectroscopically relevant parameters of this molecule to assist in its future detection in the interstellar medium.

Comparative study of the anti-HIV activities of ascorbate and thiol-containing reducing agents in chronically HIV-infected cells.

PubMed

Harakeh, S; Jariwalla, R J

1991-12-01

To elucidate the action of vitamin C on pathogenic human retroviruses, we investigated and compared the effects of noncytoxic concentrations of ascorbic acid (AA), its calcium salt (Ca-ascorbate), and two thiol-based reducing agents [glutathione (GSH) and N-acetyl-L-cysteine (NAC)] against human immunodeficiency virus (HIV)-1 replication in chronically infected T lymphocytes. Ca-ascorbate reduced extracellular HIV reverse transcriptase (RT) activity by about the same magnitude as the equivalent dose of AA. Long-term experiments showed that continuous presence of ascorbate was necessary for HIV suppression. NAC (10 mmol/L) caused less than twofold inhibition of HIV RT and conferred a synergistic effect (approximately eightfold inhibition) when tested simultaneously with AA (0.426 mmol/L). In contrast, nonesterified GSH (less than or equal to 1.838 mmol/L) had no effect on RT concentrations and did not potentiate the anti-HIV effect of AA. These results further support the potent antiviral activity of ascorbate and suggest its therapeutic value in controlling HIV infection in combination with thiols.

Molecular cloning and characterization of Bacillus alvei thiol-dependent cytolytic toxin expressed in Escherichia coli.

PubMed

Geoffroy, C; Alouf, J E

1988-07-01

A chromosomal DNA fragment from Bacillus alvei, encoding a thiol-dependent haemolytic product known as alveolysin (Mr 60,000, pI 5.0) was cloned in Escherichia coli SK1592, using pBR322 as the vector plasmid. Only a single haemolysin-positive clone was identified, by testing for haemolysis on blood agar plates. The haemolytic material was associated with the host bacterial cell. It was released by ultrasonic disruption and purified 267-fold. A 64 kDa polypeptide of pI 8.2 cofractionated with haemolytic activity during gel filtration chromatography and isoelectric focusing. It behaved identically to alveolysin in its activation by thiols, inactivation by thiol group reagents, inhibition by cholesterol, and neutralization, immunoprecipitation and immunoblotting by immune sera raised against alveolysin and streptolysin O.

Vinyl functionalized silica hybrid monolith-based trypsin microreactor for on line digestion and separation via thiol-ene "click" strategy.

PubMed

Chen, Yingzhuang; Wu, Minghuo; Wang, Keyi; Chen, Bo; Yao, Shouzhuo; Zou, Hanfa; Nie, Lihua

2011-11-04

A novel thiol-ene "click" strategy for the preparation of monolithic trypsin microreactor was proposed. The hybrid organic-inorganic monolithic capillary column with ene-functionality was fabricated by sol-gel process using tetramethoxysilane (TMOS) and γ-methacryloxypropyltrimethoxysilane (γ-MAPS) as precursors. The disulfide bonds of trypsin were reduced to form free thiol groups. Then the trypsin containing free thiol groups was attached on the γ-MAPS hybrid monolithic column with ene-functionality via thiol-ene click chemistry to form a trypsin microreactor. The activity of the trypsin microreactor was characterized by detecting the substrate (Nα-p-tosyl-L-arginine methyl ester hydrochloride, TAME) and the product (Nα-p-tosyl-L-arginine, TA) with on-line capillary zone electrophoresis. After investigating various synthesizing conditions, it was found that the microreactor with poly(N,N'-methylenebisacrylamide) as spacer can deliver the highest activity, yielding a rapid reaction rate. After repeatedly sampling and analyzing for 100 times, the monolithic trypsin microreactor still remained 87.5% of its initial activity. It was demonstrated that thiol-ene "click" strategy for the construction of enzyme microreactor is a promising method for the highly selective immobilization of proteins under mild conditions, especially enzymes with free thiol radicals. Copyright © 2011 Elsevier B.V. All rights reserved.

ortho- and meta-substituted aromatic thiols are efficient redox buffers that increase the folding rate of a disulfide-containing protein.

PubMed

Gough, Jonathan D; Barrett, Elvis J; Silva, Yenia; Lees, Watson J

2006-08-20

Thiol based redox buffers are used to enhance the folding rates of disulfide-containing proteins in vitro. Traditionally, small molecule aliphatic thiols such as glutathione are employed. Recently, we have demonstrated that aromatic thiols can further enhance protein-folding rates. In the presence of para-substituted aromatic thiols the folding rate of a disulfide-containing protein was increased by 4-23 times over that measured for glutathione. However, several important practical issues remain to be addressed. Aromatic thiols have never been tested in the presence of denaturants such as guanidine hydrochloride. Only two of the para-substituted aromatic thiols previously examined are commercially available. To expand the number of aromatic thiols for protein folding, several commercially available meta- and ortho-substituted aromatic thiols were studied. Furthermore, an ortho-substituted aromatic thiol, easily obtained from inexpensive starting materials, was investigated. Folding rates of scrambled ribonuclease A at pH 6.0, 7.0 and 7.7, with ortho- and meta-substituted aromatic thiols, were up to 10 times greater than those with glutathione. In the presence of the common denaturant guanidine hydrochloride (0.5M) aromatic thiols provided 100% yield of active protein while maintaining equivalent folding rates.

Thioredoxin Selectivity for Thiol-based Redox Regulation of Target Proteins in Chloroplasts*

PubMed Central

Yoshida, Keisuke; Hara, Satoshi; Hisabori, Toru

2015-01-01

Redox regulation based on the thioredoxin (Trx) system is believed to ensure light-responsive control of various functions in chloroplasts. Five Trx subtypes have been reported to reside in chloroplasts, but their functional diversity in the redox regulation of Trx target proteins remains poorly clarified. To directly address this issue, we studied the Trx-dependent redox shifts of several chloroplast thiol-modulated enzymes in vitro and in vivo. In vitro assays using a series of Arabidopsis recombinant proteins provided new insights into Trx selectivity for the redox regulation as well as the underpinning for previous suggestions. Most notably, by combining the discrimination of thiol status with mass spectrometry and activity measurement, we identified an uncharacterized aspect of the reductive activation of NADP-malate dehydrogenase; two redox-active Cys pairs harbored in this enzyme were reduced via distinct utilization of Trxs even within a single polypeptide. In our in vitro assays, Trx-f was effective in reducing all thiol-modulated enzymes analyzed here. We then investigated the in vivo physiological relevance of these in vitro findings, using Arabidopsis wild-type and Trx-f-deficient plants. Photoreduction of fructose-1,6-bisphosphatase was partially impaired in Trx-f-deficient plants, but the global impact of Trx-f deficiency on the redox behaviors of thiol-modulated enzymes was not as striking as expected from the in vitro data. Our results provide support for the in vivo functionality of the Trx system and also highlight the complexity and plasticity of the chloroplast redox network. PMID:25878252

Benzofurazan Sulfides for Thiol Imaging and Quantification in Live Cells through Fluorescence Microscopy

PubMed Central

Li, Yinghong; Yang, Yang; Guan, Xiangming

2012-01-01

Thiol groups play a significant role in various cellular functions. Cellular thiol concentrations can be affected by various physiological or pathological factors. A fluorescence imaging agent that can effectively and specifically image thiols in live cells through fluorescence microscopy is desirable for live cell thiol monitoring. Benzofurazan sulfides 1a–e were synthesized and found to be thiol specific fluorogenic agents except 1d. They are not fluorescent but form strong fluorescent thiol adducts after reacting with thiols through a sulfide-thiol exchange reaction. On the other hand, they exhibit no reaction with other biologically relevant nucleophilic functional groups such as -NH2, -OH, or -COOH revealing the specificity for the detection of thiols. Sulfide 1a was selected to confirm its ability to image cellular thiols through fluorescence microscopy. The compound was demonstrated to effectively image and quantify thiol changes in live cells through fluorescence microscopy using 430 nm and 520 nm as the excitation and emission wavelengths respectively. The quantification results of total thiol in live cells obtained from fluorescence microscopy were validated by an HPLC/UV total thiol assay method. The reagents and method will be of a great value to thiol redox-related research. PMID:22794193

Radicals Are Required for Thiol Etching of Gold Particles.

PubMed

Dreier, Timothy A; Ackerson, Christopher J

2015-08-03

Etching of gold with an excess of thiol ligand is used in both synthesis and analysis of gold particles. Mechanistically, the process of etching gold with excess thiol is unclear. Previous studies have obliquely considered the role of oxygen in thiolate etching of gold. Herein, we show that oxygen or a radical initiator is a necessary component for efficient etching of gold by thiolates. Attenuation of the etching process by radical scavengers in the presence of oxygen, and the restoration of activity by radical initiators under inert atmosphere, strongly implicate the oxygen radical. These data led us to propose an atomistic mechanism in which the oxygen radical initiates the etching process. © 2015 WILEY-VCH Verlag GmbH & Co. KGaA, Weinheim.

Mitochondrial respiratory chain complexes as sources and targets of thiol-based redox-regulation.

PubMed

Dröse, Stefan; Brandt, Ulrich; Wittig, Ilka

2014-08-01

The respiratory chain of the inner mitochondrial membrane is a unique assembly of protein complexes that transfers the electrons of reducing equivalents extracted from foodstuff to molecular oxygen to generate a proton-motive force as the primary energy source for cellular ATP-synthesis. Recent evidence indicates that redox reactions are also involved in regulating mitochondrial function via redox-modification of specific cysteine-thiol groups in subunits of respiratory chain complexes. Vice versa the generation of reactive oxygen species (ROS) by respiratory chain complexes may have an impact on the mitochondrial redox balance through reversible and irreversible thiol-modification of specific target proteins involved in redox signaling, but also pathophysiological processes. Recent evidence indicates that thiol-based redox regulation of the respiratory chain activity and especially S-nitrosylation of complex I could be a strategy to prevent elevated ROS production, oxidative damage and tissue necrosis during ischemia-reperfusion injury. This review focuses on the thiol-based redox processes involving the respiratory chain as a source as well as a target, including a general overview on mitochondria as highly compartmentalized redox organelles and on methods to investigate the redox state of mitochondrial proteins. This article is part of a Special Issue entitled: Thiol-Based Redox Processes. Copyright © 2014 Elsevier B.V. All rights reserved.

Investigation of thiol-disulphide balance in patients with acute urticaria and chronic spontaneous urticaria.

PubMed

Akbas, Ayse; Kilinc, Fadime; Sener, Sertac; Aktaş, Akın; Baran, Pervin; Ergin, Merve

2017-09-01

Thiol-disulphide balance plays a major role in health and diseases. This balance may be disrupted by various diseases. We aimed to determine status of the effect of thiol-disulphide balance in urticaria. We aimed to investigate the thiol-disulphide balance in patients with acute urticaria (AUP) and chronic spontaneous urticaria (CSU). Study included 53 AUP and 47 healthy controls plus 57 patients with chronic spontaneous urticaria (CSUP) and 57 healthy controls. Levels of native thiols, disulphides and total thiols were evaluated in plasma using a new and automated spectrophotometric method. Ratios of disulphides/total thiols, disulphides/native thiols and native thiols/total thiols were calculated. For AU, there was no statistical difference compared to control group in levels of native thiols, disulphides and total thiols. For CSU, however, there was an increase in levels of native thiols, disulphides and total thiols and the ratio of thiol/disulphide in favour of disulphide. Thiol-disulphide balance was not affected by AU but shifted towards to disulphide in CSU indicating the presence of oxidative stress (OS).

Fast and Selective Modification of Thiol Proteins/Peptides by N-(Phenylseleno)phthalimide

NASA Astrophysics Data System (ADS)

Wang, Zhengfang; Zhang, Yun; Zhang, Hao; Harrington, Peter B.; Chen, Hao

2012-03-01

We previously reported that selenamide reagents such as ebselen and N-(phenylseleno)phthalimide (NPSP) can be used to selectively derivatize thiols for mass spectrometric analysis, and the introduced selenium tags are useful as they could survive or removed with collision-induced dissociation (CID). Described herein is the further study of the reactivity of various protein/peptide thiols toward NPSP and its application to derivatize thiol peptides in protein digests. With a modified protocol (i.e., dissolving NPSP in acetonitrile instead of aqueous solvent), we found that quantitative conversion of thiols can be obtained in seconds, using NPSP in a slight excess amount (NPSP:thiol of 1.1-2:1). Further investigation shows that the thiol reactivity toward NPSP reflects its chemical environment and accessibility in proteins/peptides. For instance, adjacent basic amino acid residues increase the thiol reactivity, probably because they could stabilize the thiolate form to facilitate the nucleophilic attack of thiol on NPSP. In the case of creatine phosphokinase, the native protein predominately has one thiol reacted with NPSP while all of four thiol groups of the denatured protein can be derivatized, in accordance with the corresponding protein conformation. In addition, thiol peptides in protein/peptide enzymatic digests can be quickly and effectively tagged by NPSP following tri- n-butylphosphine (TBP) reduction. Notably, all three thiols of the peptide QCCASVCSL in the insulin peptic digest can be modified simultaneously by NPSP. These results suggest a novel and selective method for protecting thiols in the bottom-up approach for protein structure analysis.

Thiol Redox and pKa Properties of Mycothiol, the Predominant Low-Molecular-Weight Thiol Cofactor in the Actinomycetes.

PubMed

Sharma, Sunil V; Van Laer, Koen; Messens, Joris; Hamilton, Chris J

2016-09-15

The thiol pKa and standard redox potential of mycothiol, the major low-molecular-weight thiol cofactor in the actinomycetes, are reported. The measured standard redox potential reveals substantial discrepancies in one or more of the other previously measured intracellular parameters that are relevant to mycothiol redox biochemistry. © 2016 WILEY-VCH Verlag GmbH & Co. KGaA, Weinheim.

Multiplexed Thiol Reactivity Profiling for Target Discovery of Electrophilic Natural Products.

PubMed

Tian, Caiping; Sun, Rui; Liu, Keke; Fu, Ling; Liu, Xiaoyu; Zhou, Wanqi; Yang, Yong; Yang, Jing

2017-11-16

Electrophilic groups, such as Michael acceptors, expoxides, are common motifs in natural products (NPs). Electrophilic NPs can act through covalent modification of cysteinyl thiols on functional proteins, and exhibit potent cytotoxicity and anti-inflammatory/cancer activities. Here we describe a new chemoproteomic strategy, termed multiplexed thiol reactivity profiling (MTRP), and its use in target discovery of electrophilic NPs. We demonstrate the utility of MTRP by identifying cellular targets of gambogic acid, an electrophilic NP that is currently under evaluation in clinical trials as anticancer agent. Moreover, MTRP enables simultaneous comparison of seven structurally diversified α,β-unsaturated γ-lactones, which provides insights into the relative proteomic reactivity and target preference of diverse structural scaffolds coupled to a common electrophilic motif and reveals various potential druggable targets with liganded cysteines. We anticipate that this new method for thiol reactivity profiling in a multiplexed manner will find broad application in redox biology and drug discovery. Copyright © 2017 Elsevier Ltd. All rights reserved.

An evaluation of thiol/disulphide homeostasis in patients with psoriasis

PubMed Central

Yorulmaz, Ahu; Erdogan, Serpil; Cakmak, Seray Kulcu; Guney, Elif; Sen, Orhan; Erel, Ozcan

2017-01-01

Introduction The role of oxidative stress in the pathogenesis of psoriasis has been investigated in previous studies with conflicting results. On the other hand, well-established treatments currently used in psoriasis exert their effects via a boost of oxidative stress. Recently, a strong positive association between psoriasis, metabolic syndrome and dyslipidemia has also been described showing the complex nature of the disease. Aim To examine thiol/disulphide homeostasis, a newly developed homeostasis assay in psoriasis and evaluate the possible association between thiol/disulphide homeostasis and dyslipidemia in psoriasis. Material and methods The study population included 92 psoriasis patients and 71 healthy subjects. Serum native thiol, total thiol and disulphide levels were investigated in patients with psoriasis and in healthy subjects. In addition, lipid profile (serum total cholesterol, triglyceride, high-density lipoprotein cholesterol and low-density lipoprotein cholesterol) levels were investigated in both groups. The association between thiol-disulphide parameters and dyslipidemia was also evaluated. Results Serum total cholesterol and triglyceride levels were found to be higher in patients with psoriasis than in the healthy group. Lower plasma disulphide and higher native thiol levels were found in patients with psoriasis indicating an antioxidant status. Conclusions To our knowledge, this is the first study showing the shift of dynamic thiol/disulphide homeostasis towards the thiol form in psoriasis which indicate higher antioxidant status. PMID:29507562

The Role of Follicular Fluid Thiol/Disulphide Homeostasis in Polycystic Ovary Syndrome.

PubMed

Tola, Esra Nur; Köroğlu, Nadiye; Ergin, Merve; Oral, Hilmi Baha; Turgut, Abdülkadir; Erel, Özcan

2018-04-04

Oxidative stress is suggested as a potential triggering factor in the etiopathogenesis of Polycystic ovary syndrome related infertility. Thiol/disulphide homeostasis, a recently oxidative stress marker, is one of the antioxidant mechanism in human which have critical roles in folliculogenesis and ovulation. The aim of our study is to investigate follicular fluid thiol/disulphide homeostasis in the etiopathogenesis of Polycystic ovary syndrome and to determine its' association with in vitro fertilization outcome. The study procedures were approved by local ethic committee. Cross sectional design Methods: Follicular fluid of twenty-two Polycystic ovary syndrome women and twenty ovulatory controls undergoing in vitro fertilization treatment were recruited. Thiol/disulphide homeostasis was analyzed via a novel spectrophotometric method. Follicular native thiol levels were found to be lower in Polycystic ovary syndrome group than non- Polycystic ovary syndrome group (p=0.041) as well as native thiol/total thiol ratio (p<0.0001). Disulphide level, disulphide/native thiol and disulphide/total thiol ratios were increased in Polycystic ovary syndrome group (p<0.0001). A positive correlation between fertilization rate and native thiol (p=0.01, r=0.53) and total thiol (p=0.01, r=0.052) among Polycystic ovary syndrome patients was found. A positive predictive effect of native thiol on fertilization rate among Polycystic ovary syndrome group was also found (p=0.03, β=0.45, 95% CI=0.031-0.643). Deterioration in thiol/disulphide homeostasis, especially elevated disulphide levels could be one of the etiopathogenetic mechanism in Polycystic ovary syndrome. Increased native thiol levels is related to fertilization rate among Polycystic ovary syndrome patients and also positive predictor marker of fertilization rate among Polycystic ovary syndrome patients. Improvement of thiol/disulphide homeostasis could be of importance in the treatment of Polycystic ovary syndrome to increase in

Plasma protein hydroperoxides during aging in humans: correlation with paraoxonase 1 (PON1) arylesterase activity and plasma total thiols.

PubMed

Mehdi, Mohammad Murtaza; Rizvi, Syed Ibrahim

2013-02-01

Oxidative stress is thought to play a major role in the development of several age-dependent diseases. Proteins are major targets for oxidative attack. Protein hydroperoxides are formed by hydroxyl and singlet oxygen attack on protein, forming relatively stable hydroperoxides on histidine, tyrosine and tryptophan residues. This study investigated the levels of plasma protein hydroperoxides and antioxidant potential of plasma during aging in humans. We correlated the protein hydroperoxide formation with plasma antioxidant potential, paraoxonase 1 (PON1) arylesterase activity and plasma total thiols. The protein hydroperoxides and antioxidant potential were measured in plasma of human subjects aged between 20 and 81 years of both genders. Increase in plasma protein hydroperoxides and decrease in plasma antioxidant potential were observed as function of human age. This study provides strong correlation between plasma protein hydroperoxides formation and decrease in plasma antioxidant potential during aging. PON1 arylesterase activity and plasma total thiols levels were also found to show significant correlation with increasing levels of plasma protein hydroperoxides during aging. The plasma protein hydroperoxides provide a reliable marker of long-term redox balance and degree of oxidative stress during aging process. Copyright © 2013 IMSS. Published by Elsevier Inc. All rights reserved.

Structural and Mechanistic Insights into Unusual Thiol Disulfide Oxidoreductase

PubMed Central

Garcin, Edwige B.; Bornet, Olivier; Elantak, Latifa; Vita, Nicolas; Pieulle, Laetitia; Guerlesquin, Françoise; Sebban-Kreuzer, Corinne

2012-01-01

Cytoplasmic desulfothioredoxin (Dtrx) from the anaerobe Desulfovibrio vulgaris Hildenborough has been identified as a new member of the thiol disulfide oxidoreductase family. The active site of Dtrx contains a particular consensus sequence, CPHC, never seen in the cytoplasmic thioredoxins and generally found in periplasmic oxidases. Unlike canonical thioredoxins (Trx), Dtrx does not present any disulfide reductase activity, but it presents instead an unusual disulfide isomerase activity. We have used NMR spectroscopy to gain insights into the structure and the catalytic mechanism of this unusual Dtrx. The redox potential of Dtrx (−181 mV) is significantly less reducing than that of canonical Trx. A pH dependence study allowed the determination of the pKa of all protonable residues, including the cysteine and histidine residues. Thus, the pKa values for the thiol group of Cys31 and Cys34 are 4.8 and 11.3, respectively. The His33 pKa value, experimentally determined for the first time, differs notably as a function of the redox states, 7.2 for the reduced state and 4.6 for the oxidized state. These data suggest an important role for His33 in the molecular mechanism of Dtrx catalysis that is confirmed by the properties of mutant DtrxH33G protein. The NMR structure of Dtrx shows a different charge repartition compared with canonical Trx. The results presented are likely indicative of the involvement of this protein in the catalysis of substrates specific of the anaerobe cytoplasm of DvH. The study of Dtrx is an important step toward revealing the molecular details of the thiol-disulfide oxidoreductase catalytic mechanism. PMID:22128175

Metabolites of ginger component [6]-shogaol remain bioactive in cancer cells and have low toxicity in normal cells: chemical synthesis and biological evaluation.

PubMed

Zhu, Yingdong; Warin, Renaud F; Soroka, Dominique N; Chen, Huadong; Sang, Shengmin

2013-01-01

Our previous study found that [6]-shogaol, a major bioactive component in ginger, is extensively metabolized in cancer cells and in mice. It is unclear whether these metabolites retain bioactivity. The aim of the current study is to synthesize the major metabolites of [6]-shogaol and evaluate their inhibition of growth and induction of apoptosis in human cancer cells. Twelve metabolites of [6]-shogaol (M1, M2, and M4-M13) were successfully synthesized using simple and easily accessible chemical methods. Growth inhibition assays showed that most metabolites of [6]-shogaol had measurable activities against human cancer cells HCT-116 and H-1299. In particular, metabolite M2 greatly retained the biological activities of [6]-shogaol, with an IC(50) of 24.43 µM in HCT-116 human colon cancer cells and an IC(50) of 25.82 µM in H-1299 human lung cancer cells. Also exhibiting a relatively high potency was thiol-conjugate M13, with IC(50) values of 45.47 and 47.77 µM toward HCT-116 and H-1299 cells, respectively. The toxicity evaluation of the synthetic metabolites (M1, M2, and M4-M13) against human normal fibroblast colon cells CCD-18Co and human normal lung cells IMR-90 demonstrated a detoxifying metabolic biotransformation of [6]-shogaol. The most active metabolite M2 had almost no toxicity to CCD-18Co and IMR-90 normal cells with IC(50)s of 99.18 and 98.30 µM, respectively. TUNEL (Terminal deoxynucleotidyl transferase dUTP nick end labeling) assay indicated that apoptosis was triggered by metabolites M2, M13, and its two diastereomers M13-1 and M13-2. There was no significant difference between the apoptotic effect of [6]-shogaol and the effect of M2 and M13 after 6 hour treatment.

Estrogenic activities of diuron metabolites in female Nile tilapia (Oreochromis niloticus).

PubMed

Pereira, Thiago Scremin Boscolo; Boscolo, Camila Nomura Pereira; Felício, Andreia Arantes; Batlouni, Sergio Ricardo; Schlenk, Daniel; de Almeida, Eduardo Alves

2016-03-01

Some endocrine disrupting chemicals (EDCs) can alter the estrogenic activities of the organism by directly interacting with estrogen receptors (ER) or indirectly through the hypothalamus-pituitary-gonadal axis. Recent studies in male Nile tilapia (Oreochromis niloticus) indicated that diuron may have anti-androgenic activity augmented by biotransformation. In this study, the effects of diuron and three of its metabolites were evaluated in female tilapia. Sexually mature female fish were exposed for 25 days to diuron, as well as to its metabolites 3,4-dichloroaniline (DCA), 3,4-dichlorophenylurea (DCPU) and 3,4-dichlorophenyl-N-methylurea (DCPMU), at concentrations of 100 ng/L. Diuron metabolites caused increases in E2 plasma levels, gonadosomatic indices and in the percentage of final vitellogenic oocytes. Moreover, diuron and its metabolites caused a decrease in germinative cells. Significant differences in plasma concentrations of the estrogen precursor and gonadal regulator17α-hydroxyprogesterone (17α-OHP) were not observed. These results show that diuron metabolites had estrogenic effects potentially mediated through enhanced estradiol biosynthesis and accelerated the ovarian development of O. niloticus females. Copyright © 2015 Elsevier Ltd. All rights reserved.

Copper toxicity in Chinese cabbage is not influenced by plant sulphur status, but affects sulphur metabolism-related gene expression and the suggested regulatory metabolites.

PubMed

Shahbaz, M; Stuiver, C E E; Posthumus, F S; Parmar, S; Hawkesford, M J; De Kok, L J

2014-01-01

The toxicity of high copper (Cu) concentrations in the root environment of Chinese cabbage (Brassica pekinensis) was little influenced by the sulphur nutritional status of the plant. However, Cu toxicity removed the correlation between sulphur metabolism-related gene expression and the suggested regulatory metabolites. At high tissue Cu levels, there was no relation between sulphur metabolite levels viz. total sulphur, sulphate and water-soluble non-protein thiols, and the expression and activity of sulphate transporters and expression of APS reductase under sulphate-sufficient or-deprived conditions, in the presence or absence of H2 S. This indicated that the regulatory signal transduction pathway of sulphate transporters was overruled or by-passed upon exposure to elevated Cu concentrations. © 2013 German Botanical Society and The Royal Botanical Society of the Netherlands.

The Role of Thiol/Disulphide Homeostasis in Anthracycline Associated Cardiac Toxicity.

PubMed

Topuz, Mustafa; Şen, Omer; Kaplan, Mehmet; Akkus, Oguz; Erel, Ozcan; Gur, Mustafa

2017-02-07

The aim of the present study was to evaluate whether the baseline thiol/disulfide state can predict the occurrence of anthracycline induced cardiac toxicity. A total of 186 cancer patients receiving anthracycline (doxorubicin)-based chemotherapy were enrolled. All patients underwent 2-dimensional (2D) speckle tracking echocardiography (STE) to determine their left ventricular ejection fraction (LVEF) and blood samples for measuring thiol forms were obtained before treatment and 4 weeks after completion of the chemotherapy. The mean dose of doxorubicin exposure was 255 ± 39.2 mg/m 2 . Baseline native thiol was found to be lower whereas baseline disulfide and the disulfide/total thiol ratio were found to be higher in patients who had a decrease in LVEF after anthracycline therapy. Also, the amount of decrease in LVEF was well correlated with the delta value of the thiol forms. Logistic regression analysis revealed that changes in BNP and global longitudinal strain (GLS), baseline level of native thiol, disulfide, and the disulfide/total thiol ratio were strong predictors for a decrease in LVEF.The thiol/disulfide pathway may be a factor for predicting chemotherapy-induced cardiac toxicity as one of the oxidative stress mechanisms.

Involvement of thiol-based mechanisms in plant development.

PubMed

Rouhier, Nicolas; Cerveau, Delphine; Couturier, Jérémy; Reichheld, Jean-Philippe; Rey, Pascal

2015-08-01

Increasing knowledge has been recently gained regarding the redox regulation of plant developmental stages. The current state of knowledge concerning the involvement of glutathione, glutaredoxins and thioredoxins in plant development is reviewed. The control of the thiol redox status is mainly ensured by glutathione (GSH), a cysteine-containing tripeptide and by reductases sharing redox-active cysteines, glutaredoxins (GRXs) and thioredoxins (TRXs). Indeed, thiol groups present in many regulatory proteins and metabolic enzymes are prone to oxidation, ultimately leading to post-translational modifications such as disulfide bond formation or glutathionylation. This review focuses on the involvement of GSH, GRXs and TRXs in plant development. Recent studies showed that the proper functioning of root and shoot apical meristems depends on glutathione content and redox status, which regulate, among others, cell cycle and hormone-related processes. A critical role of GRXs in the formation of floral organs has been uncovered, likely through the redox regulation of TGA transcription factor activity. TRXs fulfill many functions in plant development via the regulation of embryo formation, the control of cell-to-cell communication, the mobilization of seed reserves, the biogenesis of chloroplastic structures, the metabolism of carbon and the maintenance of cell redox homeostasis. This review also highlights the tight relationships between thiols, hormones and carbon metabolism, allowing a proper development of plants in relation with the varying environment and the energy availability. GSH, GRXs and TRXs play key roles during the whole plant developmental cycle via their antioxidant functions and the redox-regulation of signaling pathways. This article is part of a Special Issue entitled Redox regulation of differentiation and de-differentiation. Copyright © 2015 Elsevier B.V. All rights reserved.

Thiol-ene immobilisation of carbohydrates onto glass slides as a simple alternative to gold-thiol monolayers, amines or lipid binding.

PubMed

Biggs, Caroline I; Edmondson, Steve; Gibson, Matthew I

2015-01-01

Carbohydrate arrays are a vital tool in studying infection, probing the mechanisms of bacterial, viral and toxin adhesion and the development of new treatments, by mimicking the structure of the glycocalyx. Current methods rely on the formation of monolayers of carbohydrates that have been chemically modified with a linker to enable interaction with a functionalised surface. This includes amines, biotin, lipids or thiols. Thiol-addition to gold to form self-assembled monolayers is perhaps the simplest method for immobilisation as thiolated glycans are readily accessible from reducing carbohydrates in a single step, but are limited to gold surfaces. Here we have developed a quick and versatile methodology which enables the use of thiolated carbohydrates to be immobilised as monolayers directly onto acrylate-functional glass slides via a 'thiol-ene'/Michael-type reaction. By combining the ease of thiol chemistry with glass slides, which are compatible with microarray scanners this offers a cost effective, but also useful method to assemble arrays.

Cysteine-conjugated metabolite of ginger component [6]-shogaol serves as a carrier of [6]-shogaol in cancer cells and in mice

PubMed Central

Chen, Huadong; Soroka, Dominique N.; Zhu, Yingdong; Hu, Yuhui; Chen, Xiaoxin; Sang, Shengmin

2013-01-01

Shogaols, a series of major constituents in dried ginger (Zingiber officinale), show high anti-cancer potencies. Previously, we reported that a major metabolite resulting from the mercapturic acid pathway, 5-cysteinyl-[6]-shogaol (M2), showed comparable growth inhibitory effects towards cancer cells to [6]-shogaol (6S). Here we probe the mechanism by which M2 exerts its bioactivity. We utilized a series of chemical stability tests in conjunction with bioassays to show that thiol-conjugates display chemopreventative potency by acting as carriers of active ginger component 6S. M2 chemical degradation to 6S was observed in an environment most resembling physiological conditions, with a pH of 7.4 at 37°C. The metabolic profiles of M2 in cancer cells HCT-116 and H-1299 resembled those of 6S, indicating that its biotransformation route was initiated by deconjugation. Further, the presence of excess glutathione significantly delayed 6S and M2 metabolism and counteracted cell death induced by 6S and M2, suggesting that increasing available free thiols exogenously both promoted formation of 5-glutathionyl-[6]-shogaol (M13) and inhibited the production of free 6S from M2 deconjugation, resulting in delayed 6S cell entry and bioactivity. Given the chemopreventative properties of M2 and our observations in vitro, we investigated its metabolism in mice. M2 and 6S showed similar metabolic profiles in mouse urine and fecal samples. Six new thiol-conjugated metabolites (M16–M21), together with previously reported ones, were identified by LC/MS. In particular, the increase of 5-N-acetylcystenyl-[6]-shogaol (M5) and its 3′-demethylated product (M16) abundance in mouse feces after treatment with M2 indicate that in addition to acting as a carrier of 6S, M2 is also directly acetylated to M5, which is further demethylated to M16 in vivo. In conclusion, cysteine-conjugated metabolite of [6]-shogaol M2 exerts its bioactivity by acting as a carrier of 6S in both cancer cells and in

Cysteine-conjugated metabolite of ginger component [6]-shogaol serves as a carrier of [6]-shogaol in cancer cells and in mice.

PubMed

Chen, Huadong; Soroka, Dominique N; Zhu, Yingdong; Hu, Yuhui; Chen, Xiaoxin; Sang, Shengmin

2013-06-17

Shogaols, a series of major constituents in dried ginger (Zingiber officinale), show high anticancer potencies. Previously, we reported that a major metabolite resulting from the mercapturic acid pathway, 5-cysteinyl-[6]-shogaol (M2), showed comparable growth inhibitory effects toward cancer cells to [6]-shogaol (6S). Here, we probe the mechanism by which M2 exerts its bioactivity. We utilized a series of chemical stability tests in conjunction with bioassays to show that thiol-conjugates display chemopreventative potency by acting as carriers of active ginger component 6S. M2 chemical degradation to 6S was observed in an environment most resembling physiological conditions, with a pH of 7.4 at 37 °C. The metabolic profiles of M2 in cancer cells HCT-116 and H-1299 resembled those of 6S, indicating that its biotransformation route was initiated by deconjugation. Further, the presence of excess glutathione significantly delayed 6S and M2 metabolism and counteracted cell death induced by 6S and M2, suggesting that increasing available free thiols exogenously both promoted the formation of 5-glutathionyl-[6]-shogaol (M13) and inhibited the production of free 6S from M2 deconjugation, resulting in delayed 6S cell entry and bioactivity. Given the chemopreventative properties of M2 and our observations in vitro, we investigated its metabolism in mice. M2 and 6S showed similar metabolic profiles in mouse urine and fecal samples. Six new thiol-conjugated metabolites (M16-M21), together with previously reported ones, were identified by LC/MS. In particular, the increase of 5-N-acetylcystenyl-[6]-shogaol (M5) and its 3'-demethylated product (M16) abundance in mouse feces after treatment with M2 indicates that in addition to acting as a carrier of 6S, M2 is also directly acetylated to M5, which is further demethylated to M16 in vivo. In conclusion, the cysteine-conjugated metabolite of [6]-shogaol M2 exerts its bioactivity by acting as a carrier of 6S in both cancer cells and

Phytol metabolites are circulating dietary factors that activate the nuclear receptor RXR.

PubMed Central

Kitareewan, S; Burka, L T; Tomer, K B; Parker, C E; Deterding, L J; Stevens, R D; Forman, B M; Mais, D E; Heyman, R A; McMorris, T; Weinberger, C

1996-01-01

RXR is a nuclear receptor that plays a central role in cell signaling by pairing with a host of other receptors. Previously, 9-cis-retinoic acid (9cRA) was defined as a potent RXR activator. Here we describe a unique RXR effector identified from organic extracts of bovine serum by following RXR-dependent transcriptional activity. Structural analyses of material in active fractions pointed to the saturated diterpenoid phytanic acid, which induced RXR-dependent transcription at concentrations between 4 and 64 microM. Although 200 times more potent than phytanic acid, 9cRA was undetectable in equivalent amounts of extract and cannot be present at a concentration that could account for the activity. Phytanic acid, another phytol metabolite, was synthesized and stimulated RXR with a potency and efficacy similar to phytanic acid. These metabolites specifically displaced [3H]-9cRA from RXR with Ki values of 4 microM, indicating that their transcriptional effects are mediated by direct receptor interactions. Phytol metabolites are compelling candidates for physiological effectors, because their RXR binding affinities and activation potencies match their micromolar circulating concentrations. Given their exclusive dietary origin, these chlorophyll metabolites may represent essential nutrients that coordinate cellular metabolism through RXR-dependent signaling pathways. PMID:8856661

Proteomic detection of oxidized and reduced thiol proteins in cultured cells.

PubMed

Cuddihy, Sarah L; Baty, James W; Brown, Kristin K; Winterbourn, Christine C; Hampton, Mark B

2009-01-01

The oxidation and reduction of cysteine residues is emerging as an important post-translational control of protein function. We describe a method for fluorescent labelling of either reduced or oxidized thiols in combination with two-dimensional sodium dodecyl sulphate polyacrylamide gel electrophoresis (2DE) to detect changes in the redox proteome of cultured cells. Reduced thiols are labelled with the fluorescent compound 5-iodoacetamidofluorescein. To monitor oxidized thiols, the reduced thiols are first blocked with N-ethyl-maleimide, then the oxidized thiols reduced with dithiothreitol and labelled with 5-iodoacetamidofluorescein. The method is illustrated by treating Jurkat T-lymphoma cells with hydrogen peroxide and monitoring increased labelling of oxidized thiol proteins. A decrease in labelling can also be detected, and this is attributed to the formation of higher oxidation states of cysteine that are not reduced by dithiothreitol.

Modification of the mitochondrial sulfonylurea receptor by thiol reagents.

PubMed

Szewczyk, A; Wójcik, G; Lobanov, N A; Nalecz, M J

1999-08-19

The purpose of this study was to investigate the effects exerted by thiol-modifying reagents on themitochondrial sulfonylurea receptor. The thiol-oxidizing agents (timerosal and 5, 5'-dithio-bis(2-nitrobenzoic acid)) were found to produce a large inhibition (70% to 80%) of specific binding of [(3)H]glibenclamide to the beef heart mitochondrial membrane. Similar effects were observed with membrane permeable (N-ethylmaleimide) and non-permeable (mersalyl) thiol modifying agents. Glibenclamide binding was also decreased by oxidizing agents (hydrogen peroxide) but not by reducing agents (reduced gluthatione, dithiothreitol and the 2,3-dihydroxy-1,4-dithiolbutane). The results suggest that intact thiol groups, facing the mitochondrial matrix, are essential for glibenclamide binding to the mitochondrial sulfonylurea receptor. Copyright 1999 Academic Press.

Widespread occurrence of neuro-active pharmaceuticals and metabolites in 24 Minnesota rivers and wastewaters

USGS Publications Warehouse

Writer, Jeffrey; Ferrer, Imma; Barber, Larry B.; Thurman, E. Michael

2013-01-01

Concentrations of 17 neuro-active pharmaceuticals and their major metabolites (bupropion, hydroxy-bupropion, erythro-hydrobupropion, threo-hydrobupropion, carbamazepine, 10,11,-dihydro-10,11,-dihydroxycarbamazepine, 10-hydroxy-carbamazepine, citalopram, N-desmethyl-citalopram, fluoxetine, norfluoxetine, gabapentin, lamotrigine, 2-N-glucuronide-lamotrigine, oxcarbazepine, venlafaxine and O-desmethyl-venlafaxine), were measured in treated wastewater and receiving surface waters from 24 locations across Minnesota, USA. The analysis of upstream and downstream sampling sites indicated that the wastewater treatment plants were the major source of the neuro-active pharmaceuticals and associated metabolites in surface waters of Minnesota. Concentrations of parent compound and the associated metabolite varied substantially between treatment plants (concentrations ± standard deviation of the parent compound relative to its major metabolite) as illustrated by the following examples; bupropion and hydrobupropion 700 ± 1000 ng L−1, 2100 ± 1700 ng L−1, carbamazepine and 10-hydroxy-carbamazepine 480 ± 380 ng L−1, 360 ± 400 ng L−1, venlafaxine and O-desmethyl-venlafaxine 1400 ± 1300 ng L−1, 1800 ± 2300 ng L−1. Metabolites of the neuro-active compounds were commonly found at higher or comparable concentrations to the parent compounds in wastewater effluent and the receiving surface water. Neuro-active pharmaceuticals and associated metabolites were detected only sporadically in samples upstream from the effluent outfall. Metabolite to parent ratios were used to evaluate transformation, and we determined that ratios in wastewater were much lower than those reported in urine, indicating that the metabolites are relatively more labile than the parent compounds in the treatment plants and in receiving waters. The widespread occurrence of neuro-active pharmaceuticals and metabolites in Minnesota effluents and surface waters indicate that

Inhibition of Urease by Disulfiram, an FDA-Approved Thiol Reagent Used in Humans.

PubMed

Díaz-Sánchez, Ángel Gabriel; Alvarez-Parrilla, Emilio; Martínez-Martínez, Alejandro; Aguirre-Reyes, Luis; Orozpe-Olvera, Jesica Aline; Ramos-Soto, Miguel Armando; Núñez-Gastélum, José Alberto; Alvarado-Tenorio, Bonifacio; de la Rosa, Laura Alejandra

2016-11-26

Urease is a nickel-dependent amidohydrolase that catalyses the decomposition of urea into carbamate and ammonia, a reaction that constitutes an important source of nitrogen for bacteria, fungi and plants. It is recognized as a potential antimicrobial target with an impact on medicine, agriculture, and the environment. The list of possible urease inhibitors is continuously increasing, with a special interest in those that interact with and block the flexible active site flap. We show that disulfiram inhibits urease in Citrullus vulgaris (CVU), following a non-competitive mechanism, and may be one of this kind of inhibitors. Disulfiram is a well-known thiol reagent that has been approved by the FDA for treatment of chronic alcoholism. We also found that other thiol reactive compounds (l-captopril and Bithionol) and quercetin inhibits CVU. These inhibitors protect the enzyme against its full inactivation by the thiol-specific reagent Aldrithiol (2,2'-dipyridyl disulphide, DPS), suggesting that the three drugs bind to the same subsite. Enzyme kinetics, competing inhibition experiments, auto-fluorescence binding experiments, and docking suggest that the disulfiram reactive site is Cys592, which has been proposed as a "hinge" located in the flexible active site flap. This study presents the basis for the use of disulfiram as one potential inhibitor to control urease activity.

Short communication: characterization of soluble thiols in bovine milk.

PubMed

Niero, G; De Marchi, M; Masi, A; Penasa, M; Cassandro, M

2015-09-01

Antioxidants are molecules essential for the maintenance of cell homeostasis and their intake through the diet has positive effects on human health. Among antioxidants, low-molecular-weight (LMW) thiols represent an important class of compounds. The aim of this study was to identify LMW thiols in bovine milk. A total of 96 individual milk samples from Brown Swiss, Holstein-Friesian, Alpine Grey, and Simmental cattle breeds were collected in 8 herds. The LMW thiols were extracted from the soluble fraction of milk and, following a derivatization protocol, they were separated by reverse phase HPLC and detected fluorimetrically. Six thiol species were detected and 2, glutathione (GSH) and cysteine-glycine (Cys-Gly), were identified and quantified. Regardless of the breed, the average concentration of Cys-Gly in milk was greater than that of GSH. Overall, milk from dual-purpose breeds (Simmental and Alpine Grey) was richer in LMW thiols than milk from dairy cows (Holstein-Friesian and Brown Swiss). Glutathione and Cys-Gly, closely linked metabolically, were strongly correlated. Pearson correlations of Cys-Gly with protein and casein contents were moderately low, and no relationship was found between GSH and milk chemical composition. Future research should focus on the identification of all detected LMW thiol species. Copyright © 2015 American Dairy Science Association. Published by Elsevier Inc. All rights reserved.

Interfacial thiol-ene photoclick reactions for forming multilayer hydrogels.

PubMed

Shih, Han; Fraser, Andrew K; Lin, Chien-Chi

2013-03-13

Interfacial visible light-mediated thiol-ene photoclick reactions were developed for preparing step-growth hydrogels with multilayer structures. The effect of a noncleavage type photoinitiator eosin-Y on visible-light-mediated thiol-ene photopolymerization was first characterized using in situ photorheometry, gel fraction, and equilibrium swelling ratio. Next, spectrophotometric properties of eosin-Y in the presence of various relevant macromer species were evaluated using ultraviolet-visible light (UV-vis) spectrometry. It was determined that eosin-Y was able to reinitiate the thiol-ene photoclick reaction, even after light exposure. Because of its small molecular weight, most eosin-Y molecules readily leached out from the hydrogels. The diffusion of residual eosin-Y from preformed hydrogels was exploited for fabricating multilayer step-growth hydrogels. Interfacial hydrogel coating was formed via the same visible-light-mediated gelation mechanism without adding fresh initiator. The thickness of the thiol-ene gel coating could be easily controlled by adjusting visible light exposure time, eosin-Y concentration initially loaded in the core gel, or macromer concentration in the coating solution. The major benefits of this interfacial thiol-ene coating system include its simplicity and cytocompatibility. The formation of thiol-ene hydrogels and coatings neither requires nor generates any cytotoxic components. This new gelation chemistry may have great utilities in controlled release of multiple sensitive growth factors and encapsulation of multiple cell types for tissue regeneration.

Interactions between serine acetyltransferase and O-acetylserine (thiol) lyase in higher plants--structural and kinetic properties of the free and bound enzymes.

PubMed

Droux, M; Ruffet, M L; Douce, R; Job, D

1998-07-01

presence of a large excess of free O-acetylserine (thiol) lyase. These findings contradict the widely held belief that such a bienzyme complex is required to channel the metabolite intermediate O-acetylserine.

Resveratrol glucuronides as the metabolites of resveratrol in humans: characterization, synthesis, and anti-HIV activity.

PubMed

Wang, Lai-Xi; Heredia, Alonso; Song, Haijing; Zhang, Zhaojun; Yu, Biao; Davis, Charles; Redfield, Robert

2004-10-01

Resveratrol is a natural product with diverse biological activities. We have previously reported that resveratrol possesses potent synergistic inhibitory activity against human immunodeficiency virus (HIV)-1 infection in combination with nucleoside analogs (Heredia et al. 2000. J Acquir Immune Defic Syndr 25:246-255). As a part of our program in developing resveratrol as a component for anti-HIV chemotherapy, we describe in this article the characterization, chemical synthesis, and biological effects of the human metabolites of resveratrol. We found that resveratrol was metabolized in humans into two metabolites, which were characterized as resveratrol-3-O- and 4'-O-glucuronides. For further biological studies, we reported two simple, alternative methods for the synthesis of the metabolites. The cytotoxic and antiviral activities of resveratrol and its metabolites were compared in cell culture experiments using human peripheral blood mononuclear cells. Whereas resveratrol was cytotoxic at > or =30 microM, no cytotoxicity was observed for the metabolites at concentrations as high as 300 microM. However, resveratrol showed strong synergistic anti-HIV activity with didanosine at 10 microM, but no synergistic effects were observed for either of the metabolites at up to 300 microM. Nevertheless, the in vitro activity of the metabolites (resveratrol glucuronides) may not necessarily reflect their in vivo function, given the fact that the ubiquitously existing human beta-glucuronidase could convert the metabolites back to resveratrol locally or systematically in vivo. The present studies have implications for future development of resveratrol and/or its derivatives as a chemotherapeutic agent. Copyright 2004 Wiley-Liss, Inc. and the American Pharmacists Association

[Influence of corynebacteria metabolites on antagonistic activity of H2O2 producing lactobacilli].

PubMed

Bukharin, O V; Sgibnev, A V

2012-01-01

Study combined influence of Corynebacterium genus bacteria metabolites and H2O2 producing lactobacilli on survival rate of Staphylococcus aureus, Escherichia coli and Lactobacillus acidophilus. The ability to inhibit catalase of the test strains used and to reduce bactericidal effect of hydroxyl radical were determined in corynebacteria. H2O2 containing metabolites were obtained by cultivating lactobacilli in mineral medium, the amount of H2O2 was determined by oxidation of TMB by peroxidase. Bactericidal effect of lactobacilli metabolites for test strains treated by corynebacteria metabolites was evaluated by seeding results. Results. Inhibitio by corynebacteria metabolites of S. aureus catalase activity by 30-40% and E. coli catalase activ ity by 40-70% was shown. A reduction of bactericidal effect of hydroxyl radicals by corynebacteria metabolites by 30-35% for S. aureus, 38-42% for E. coli and 70-73% for L. acidophilus was noted. The enchantment of bactericidal effect of lactobacilli after treatment of the test strain by corynebacteria metabolites against S. aureus and E. coli manifested by reduction of the numbe of viable cells by 2-3 lg CFU. For L. acidophilus the bactericidal effect oflactobacilli metabolite in the same conditions reduced, and that led to the increase ofviability by 2-4 lg PFU. A conclusion on the possibility of regulation by associative bacteria the manifestations of antagonistic activity of H2O2 producing dominant microorganisms is made based on the data obtained.

Tumor Xenograft Response to Redox-Active Therapies Assessed by Magnetic Resonance Imaging Using a Thiol-Bearing DOTA Complex of Gadolinium1

PubMed Central

Guntle, Gerald P; Jagadish, Bhumasamudram; Mash, Eugene A; Powis, Garth; Dorr, Robert T; Raghunand, Natarajan

2012-01-01

Gd-LC6-SH is a thiol-bearing DOTA complex of gadolinium designed to bind plasma albumin at the conserved Cys34 site. The binding of Gd-LC6-SH shows sensitivity to the presence of competing thiols. We hypothesized that Gd-LC6-SH could provide magnetic resonance imaging (MRI) enhancement that is sensitive to tumor redox state and that the prolonged retention of albumin-bound Gd-LC6-SH in vivo can be exploited to identify a saturating dose above which the shortening of MRI longitudinal relaxation time (T1) of tissue is insensitive to the injected gadolinium dose. In the Mia-PaCa-2 pancreatic tumor xenograft model in SCID mice, both the small-molecule Gd-DTPA-BMA and the macromolecule Galbumin MRI contrast agents produced dose-dependent decreases in tumor T1. By contrast, the decreases in tumor T1 provided by Gd-LC6-SH at 0.05 and 0.1 mmol/kg were not significantly different at longer times after injection. SCID mice bearing Mia-PaCa-2 or NCI-N87 tumor xenografts were treated with either the glutathione synthesis inhibitor buthionine sulfoximine or the thiol-oxidizing anticancer drug Imexon, respectively. In both models, there was a significantly greater increase in tumor R1 (=1/T1) 60 minutes after injection of Gd-LC6-SH in drug-treated animals relative to saline-treated controls. In addition, Mercury Orange staining for nonprotein sulfhydryls was significantly decreased by drug treatment relative to controls in both tumor models. In summary, these studies show that thiol-bearing complexes of gadolinium such as Gd-LC6-SH can serve as redox-sensitive MRI contrast agents for detecting differences in tumor redox status and can be used to evaluate the effects of redox-active drugs. PMID:22741038

Secondary metabolites profiles and antioxidant activities of germinated brown and red rice

NASA Astrophysics Data System (ADS)

Nurnaistia, Y.; Aisyah, S.; Munawaroh, H. S. H.; Zackiyah

2018-05-01

The research aims to investigate the effect of germination on the secondary metabolite profiles and antioxidant activity of brown and red rice. The germination was performed by using a simple laboratory-scale machine that was designed and optimized to provide conditions that support the germination process. The germination was carried out for 2 days in dark conditions at 26°C and 99% humidity. Analysis of the secondary metabolite profile of ungerminated and germinated rice was performed using LC-MS. The antioxidant activities of ungerminated and germinated rice were done by using DPPH method. The results showed that the profiles of secondary metabolites of brown and red rice changed after germination. Some peaks were found to be induced in the germinated rice. However, some peaks were also loss during germination. The antioxidant activity of brown rice was slightly increased due to the germination, from 11.2% to 22.5%. Meanwhile the antioxidant activity of red rice was decreased after germination, from 73.8% to 60.0%.

Moderate physical exercise induces the oxidation of human blood protein thiols.

PubMed

Inayama, Takayo; Oka, Jun; Kashiba, Misato; Saito, Makoto; Higuchi, Mitsuru; Umegaki, Keizo; Yamamoto, Yorihiro; Matsuda, Mitsuo

2002-03-15

Exercise is known to induce the oxidation of blood low-molecular-weight (LMW) thiols such as reduced glutathione (GSH). We previously reported that full-marathon running induced a decrease in human plasma levels of protein-bound sulfhydryl groups (p-SHs). Moderate exercise, a 30-min running at the intensity of the individual ventilatory threshold, performed by untrained healthy females caused a significant decrease in erythrocyte levels of p-SHs (mostly hemoglobin cysteine residues) and LMW thiols, but their levels returned to each baseline by 2 h. No significant change in plasma LMW thiols was observed. However, plasma levels of p-SHs significantly decreased after running and remained unchanged after 24 h. These results suggest that moderate exercise causes the oxidation of blood thiols, especially protein-bound thiols.

Dynamic thiol/disulfide homeostasis and effects of smoking on homeostasis parameters in patients with psoriasis.

PubMed

Emre, Selma; Demirseren, Duriye Deniz; Alisik, Murat; Aktas, Akin; Neselioglu, Salim; Erel, Ozcan

2017-12-01

Recently, increased reactive oxygen species (ROS), reduced antioxidant capacity, and oxidative stress have been suggested in the pathogenesis of psoriasis. The aim of this study to evaluate the thiol/disulfide homeostasis in patients with psoriasis. Ninety patients with psoriasis who did not receive any systemic treatment in the last six  months were included in the study. Seventy-six age and gender-matched healthy volunteers served as control group. Thiol/disulfide homeostasis was measured in venous blood samples obtained from patient and control groups. Native thiol and total thiol levels were significantly higher in patients than in control group. When thiol/disulfide hemostasis parameters and clinical and demographic characteristics were compared, a negative correlation was detected between native thiol and total thiol with age. The levels of total thiols had also negative correlation with PASI and duration of the disease. When we divided the patients into smokers and non-smokers, native thiol and total thiol levels were significantly higher in smokers than in controls, whereas native thiol and total thiol levels were comparable in non-smoker patients and controls. Thiol/disulfide balance shifted towards thiol in psoriasis patients and this may be responsible for increased keratinocyte proliferation in the pathogenesis of psoriasis.

Mutational analysis of RsrA, a zinc-binding anti-sigma factor with a thiol-disulphide redox switch.

PubMed

Paget, M S; Bae, J B; Hahn, M Y; Li, W; Kleanthous, C; Roe, J H; Buttner, M J

2001-02-01

In the Gram-positive bacterium, Streptomyces coelicolor A3(2), expression of the thioredoxin system is modulated by a sigma factor called sigmaR in response to changes in the cytoplasmic thiol-disulphide status, and the activity of sigmaR is controlled post-translationally by an anti-sigma factor, RsrA. In vitro, the anti-sigma factor activity of RsrA, which contains seven cysteines, correlates with its thiol-disulphide redox status. Here, we investigate the function of RsrA in vivo. A constructed rsrA null mutant had very high constitutive levels of disulphide reductase activity and sigmaR-dependent transcription, confirming that RsrA is a negative regulator of sigmaR and a key sensor of thiol-disulphide status. Targeted mutagenesis revealed that three of the seven cysteines in RsrA (C11, C41 and C44) were essential for anti-sigma factor activity and that a mutant RsrA protein containing only these three cysteines was active and still redox sensitive in vivo. We also show that RsrA is a metalloprotein, containing near-stoichiometric amounts of zinc. On the basis of these data, we propose that a thiol-disulphide redox switch is formed between two of C11, C41 and C44, and that all three residues play an essential role in anti-sigma factor activity in their reduced state, perhaps by acting as ligands for zinc. Unexpectedly, rsrA null mutants were blocked in sporulation, probably as a consequence of an increase in the level of free sigmaR.

Oxidative stress, thiols, and redox profiles.

PubMed

Harris, Craig; Hansen, Jason M

2012-01-01

Oxidative stress has been recognized as a contributing factor in the toxicity of a large number of developmental toxicants. Traditional definitions of oxidative stress state that a shift in the balance between reduced and oxidized biomolecules within cells, in favor of the latter, result in changes that are deleterious to vital cell functions and can culminate in malformations and death. The glutathione (GSH)/glutathione disulfide (GSSG) redox couple has been the traditional marker of choice for characterization of oxidative stress because of its high concentrations and direct roles as antioxidant and cellular protectant. Steady state depletion of GSH through conjugation, oxidation, or export has often been reported as the sole criteria for invoking oxidative stress and a myriad of associated deleterious consequences. Numerous other, mostly qualitative, observations have also been reported to suggest oxidative stress has occurred but it is not always clear how well they reflect the state of a cell or its functions. Our emerging understanding of redox signaling and the roles of reactive oxygen species (ROS), thiols, oxidant molecules, and cellular antioxidants, all acting as second messengers, has prompted a redefinition of oxidative stress based on changes in the real posttranslational protein thiol modifications that are central to redox regulation and control. Thiol-based redox couples such as GSH/GSSG, cysteine/cystine (cys/cySS), thioredoxin-reduced/thioredoxin-oxidized (TRX(red)/TRX(ox)) form independent signaling nodes that selectively regulate developmental events and are closely linked to changes in intracellular redox potentials. Accurate assessment of the consequences of increased free radicals in developing conceptuses should best be made using a battery of measurements including the quantitative assessment of intracellular redox potential, ROS, redox status of biomolecules, and induced changes in specific redox signaling nodes. Methods are presented for

Maillard reaction products derived from thiol compounds as inhibitors of enzymatic browning of fruits and vegetables: the structure-activity relationship.

PubMed

Billaud, C; Maraschin, C; Peyrat-Maillard, M-N; Nicolas, J

2005-06-01

Some thiol-derived Maillard reaction products (MRPs) may exert antioxidant activity, depending on the reaction conditions as well as on the sugar and the sulphydryl compound. Recently, we reported that MRPs derived from glucose or fructose with cysteine (CSH) or glutathione (GSH) mixtures greatly inhibited polyphenoloxidases (PPOs), oxidoreductases responsible for discoloration of fresh or minimally processed fruits and vegetables. Glucose and GSH were shown to be the most active in producing inhibitory MRPs. Therefore, we examined the way in which the nature of the reactants affected their synthesis, in order to establish a structure-activity relationship for the inhibitory products. Various aqueous (0.083 M, 0.125 M, or 0.25 M) mixtures of a sugar (hexose, pentose, or diholoside) with either a CSH-related compound (CSH, GSH, N-acetyl-cysteine, cysteamine, cysteic acid, methyl-cysteine, cysteine methyl ester), an amino acid (gamma-glutamic acid, glycine, methionine), or other sulfur compound (thiourea, 1,4-dithiothreitol, 2-mercaptoethanol) were heated at 103 degrees C for 14 h. Soluble MRPs were compared for their ability to inhibit apple PPO activity. In the presence of CSH, the rated sugars (same molar concentration) ranked as to inhibitory effect were pentoses > sucrose > hexoses > or = maltose. In the presence of glucose, the simultaneous presence of an amino group, a carboxyl group, and a free thiol group on the same molecule seemed essential for the production of highly inhibitory compounds.

Antifungal activity of microbial secondary metabolites.

PubMed

Coleman, Jeffrey J; Ghosh, Suman; Okoli, Ikechukwu; Mylonakis, Eleftherios

2011-01-01

Secondary metabolites are well known for their ability to impede other microorganisms. Reanalysis of a screen of natural products using the Caenorhabditis elegans-Candida albicans infection model identified twelve microbial secondary metabolites capable of conferring an increase in survival to infected nematodes. In this screen, the two compound treatments conferring the highest survival rates were members of the epipolythiodioxopiperazine (ETP) family of fungal secondary metabolites, acetylgliotoxin and a derivative of hyalodendrin. The abundance of fungal secondary metabolites indentified in this screen prompted further studies investigating the interaction between opportunistic pathogenic fungi and Aspergillus fumigatus, because of the ability of the fungus to produce a plethora of secondary metabolites, including the well studied ETP gliotoxin. We found that cell-free supernatant of A. fumigatus was able to inhibit the growth of Candida albicans through the production of a secreted product. Comparative studies between a wild-type and an A. fumigatus ΔgliP strain unable to synthesize gliotoxin demonstrate that this secondary metabolite is the major factor responsible for the inhibition. Although toxic to organisms, gliotoxin conferred an increase in survival to C. albicans-infected C. elegans in a dose dependent manner. As A. fumigatus produces gliotoxin in vivo, we propose that in addition to being a virulence factor, gliotoxin may also provide an advantage to A. fumigatus when infecting a host that harbors other opportunistic fungi.

Antifungal Activity of Microbial Secondary Metabolites

PubMed Central

Okoli, Ikechukwu; Mylonakis, Eleftherios

2011-01-01

Secondary metabolites are well known for their ability to impede other microorganisms. Reanalysis of a screen of natural products using the Caenorhabditis elegans-Candida albicans infection model identified twelve microbial secondary metabolites capable of conferring an increase in survival to infected nematodes. In this screen, the two compound treatments conferring the highest survival rates were members of the epipolythiodioxopiperazine (ETP) family of fungal secondary metabolites, acetylgliotoxin and a derivative of hyalodendrin. The abundance of fungal secondary metabolites indentified in this screen prompted further studies investigating the interaction between opportunistic pathogenic fungi and Aspergillus fumigatus, because of the ability of the fungus to produce a plethora of secondary metabolites, including the well studied ETP gliotoxin. We found that cell-free supernatant of A. fumigatus was able to inhibit the growth of Candida albicans through the production of a secreted product. Comparative studies between a wild-type and an A. fumigatus ΔgliP strain unable to synthesize gliotoxin demonstrate that this secondary metabolite is the major factor responsible for the inhibition. Although toxic to organisms, gliotoxin conferred an increase in survival to C. albicans-infected C. elegans in a dose dependent manner. As A. fumigatus produces gliotoxin in vivo, we propose that in addition to being a virulence factor, gliotoxin may also provide an advantage to A. fumigatus when infecting a host that harbors other opportunistic fungi. PMID:21966496

The effects of plant nutritional strategy on soil microbial denitrification activity through rhizosphere primary metabolites.

PubMed

Guyonnet, Julien P; Vautrin, Florian; Meiffren, Guillaume; Labois, Clément; Cantarel, Amélie A M; Michalet, Serge; Comte, Gilles; Haichar, Feth El Zahar

2017-04-01

The aim of this study was to determine (i) whether plant nutritional strategy affects the composition of primary metabolites exuded into the rhizosphere and (ii) the impact of exuded metabolites on denitrification activity in soil. We answered this question by analysing primary metabolite content extracted from the root-adhering soil (RAS) and the roots of three grasses representing different nutrient management strategies: conservative (Festuca paniculata), intermediate (Bromus erectus) and exploitative (Dactylis glomerata). We also investigated the impact of primary metabolites on soil microbial denitrification enzyme activity without carbon addition, comparing for each plant RAS and bulk soils. Our data show that plant nutritional strategy impacts on primary metabolite composition of root extracts or RAS. Further we show, for the first time, that RAS-extracted primary metabolites are probably better indicators to explain plant nutrient strategy than root-extracted ones. In addition, our results show that some primary metabolites present in the RAS were well correlated with soil microbial denitrification activity with positive relationships found between denitrification and the presence of some organic acids and negative ones with the presence of xylose. We demonstrated that the analysis of primary metabolites extracted from the RAS is probably more pertinent to evaluate the impact of plant on soil microbial community functioning. © FEMS 2017. All rights reserved. For permissions, please e-mail: journals.permissions@oup.com.

Thiol Disulfide Homeostasis in Schizophrenic Patients Using Atypical Antipsychotic Drugs

PubMed Central

Ersan, Etem Erdal; Aydin, Hüseyin; Erdoğan, Serpil; Erşan, Serpil; Alişik, Murat; Bakir, Sevtap; Erel, Özcan; Koç, Derya

2018-01-01

Objective Schizophrenia is a severe, debilitating mental disorder characterized by behavioral abnormalities. Although several studies have investigated the role of oxidative stress and the effects of antipsychotic drugs on oxidative markers in schizophrenia, adequate information is not available on these issues. The aim of this study is to determine the changes in oxidative status and thiol disulfide homeostasis in schizophrenic patients using atypical antipsychotic drugs. Methods Thirteen schizophrenic patients using atypical antipsychotic drugs and 30 healthy controls were included this study. The concentrations of total oxidant status (TOS), total antioxidant status (TAS), native thiol, total thiol, and disulfide levels were determined in the study population. Results The TAS (p=0.001), total thiol, and native thiol levels (p<0.001) were higher in the patients compared to the controls, whereas the TOS and disulfide levels were lower in the patients than in the controls (p<0.001). Conclusion These results may suggest that atypical antipsychotic drugs have a useful therapeutic effect by reducing oxidative stress via the inhibition of the formation of disulfide bonds. The study population number was one of the limitations of this study. Therefore, further studies are needed to establish the association between thiol disulfide homeostasis in schizophrenic patients using atypical antipsychotic drugs. PMID:29397665

Sulindac metabolites inhibit epidermal growth factor receptor activation and expression.

PubMed

Pangburn, Heather A; Kraus, Hanna; Ahnen, Dennis J; Rice, Pamela L

2005-09-02

Regular use of nonsteroidal anti-inflammatory drugs (NSAIDs) is associated with a decreased mortality from colorectal cancer (CRC). NSAIDs induce apoptotic cell death in colon cancer cells in vitro and inhibit growth of neoplastic colonic mucosa in vivo however, the biochemical mechanisms required for these growth inhibitory effects are not well defined. We previously reported that metabolites of the NSAID sulindac downregulate extracellular-signal regulated kinase 1/2 (ERK1/2) signaling and that this effect is both necessary and sufficient for the apoptotic effects of these drugs. The goal of this project was to specifically test the hypothesis that sulindac metabolites block activation and/or expression of the epidermal growth factor (EGF) receptor (EGFR). HT29 human colon cancer cells were treated with EGF, alone, or in the presence of sulindac sulfide or sulindac sulfone. Cells lysates were assayed by immunoblotting for phosphorylated EGFR (pEGFR, pY1068), total EGFR, phosphorylated ERK1/2 (pERK1/2), total ERK1/2, activated caspase-3, and alpha-tubulin. EGF treatment rapidly induced phosphorylation of both EGFR and ERK1/2 in HT29 colon cancer cells. Pretreatment with sulindac metabolites for 24 h blocked EGF-induced phosphorylation of both EGFR and ERK1/2 and decreased total EGFR protein expression. Under basal conditions, downregulation of pEGFR and total EGFR was detected as early as 12 h following sulindac sulfide treatment and persisted through at least 48 h. Sulindac sulfone induced downregulation of pEGFR and total EGFR was detected as early as 1 h and 24 h, respectively, following drug treatment, and persisted through at least 72 h. EGFR downregulation by sulindac metabolites was observed in three different CRC cell lines, occurred prior to the observed downregulation of pERK1/2 and induction of apoptosis by these drugs, and was not dependent of caspase activation. These results suggest that downregulation of EGFR signaling by sulindac metabolites may

Phenolics from Garcinia mangostana Inhibit Advanced Glycation Endproducts Formation: Effect on Amadori Products, Cross-Linked Structures and Protein Thiols.

PubMed

Abdallah, Hossam M; El-Bassossy, Hany; Mohamed, Gamal A; El-Halawany, Ali M; Alshali, Khalid Z; Banjar, Zainy M

2016-02-22

Accumulation of Advanced Glycation Endproducts (AGEs) in body tissues plays a major role in the development of diabetic complications. Here, the inhibitory effect of bioactive metabolites isolated from fruit hulls of Garcinia mangostana on AGE formation was investigated through bio-guided approach using aminoguanidine (AG) as a positive control. Including G. mangostana total methanol extract (GMT) in the reaction mixture of bovine serum albumin (BSA) and glucose or ribose inhibited the fluorescent and non-fluorescent AGEs formation in a dose dependent manner. The bioassay guided fractionation of GMT revealed isolation of four bioactive constituents from the bioactive fraction; which were identified as: garcimangosone D (1), aromadendrin-8-C-glucopyranoside (2), epicatechin (3), and 2,3',4,5',6-pentahydroxybenzophenone (4). All the tested compounds significantly inhibited fluorescent and non-fluorescent AGEs formation in a dose dependent manner whereas compound 3 (epicatechin) was found to be the most potent. In search for the level of action, addition of GMT, and compounds 2-4 inhibited fructosamine (Amadori product) and protein aggregation formation in both glucose and ribose. To explore the mechanism of action, it was found that addition of GMT and only compound (3) to reaction mixture increased protein thiol in both glucose and ribose while compounds 1, 2 and 4 only increased thiol in case of ribose. In conclusion, phenolic compounds 1-4 inhibited AGEs formation at the levels of Amadori product and protein aggregation formation through saving protein thiol.

Clustering of 3D-Structure Similarity Based Network of Secondary Metabolites Reveals Their Relationships with Biological Activities.

PubMed

Ohtana, Yuki; Abdullah, Azian Azamimi; Altaf-Ul-Amin, Md; Huang, Ming; Ono, Naoaki; Sato, Tetsuo; Sugiura, Tadao; Horai, Hisayuki; Nakamura, Yukiko; Morita Hirai, Aki; Lange, Klaus W; Kibinge, Nelson K; Katsuragi, Tetsuo; Shirai, Tsuyoshi; Kanaya, Shigehiko

2014-12-01

Developing database systems connecting diverse species based on omics is the most important theme in big data biology. To attain this purpose, we have developed KNApSAcK Family Databases, which are utilized in a number of researches in metabolomics. In the present study, we have developed a network-based approach to analyze relationships between 3D structure and biological activity of metabolites consisting of four steps as follows: construction of a network of metabolites based on structural similarity (Step 1), classification of metabolites into structure groups (Step 2), assessment of statistically significant relations between structure groups and biological activities (Step 3), and 2-dimensional clustering of the constructed data matrix based on statistically significant relations between structure groups and biological activities (Step 4). Applying this method to a data set consisting of 2072 secondary metabolites and 140 biological activities reported in KNApSAcK Metabolite Activity DB, we obtained 983 statistically significant structure group-biological activity pairs. As a whole, we systematically analyzed the relationship between 3D-chemical structures of metabolites and biological activities. © 2014 WILEY-VCH Verlag GmbH & Co. KGaA, Weinheim.

Diversity of Secondary Metabolites from Marine Bacillus Species: Chemistry and Biological Activity

PubMed Central

Mondol, Muhammad Abdul Mojid; Shin, Hee Jae; Islam, Mohammad Tofazzal

2013-01-01

Marine Bacillus species produce versatile secondary metabolites including lipopeptides, polypeptides, macrolactones, fatty acids, polyketides, and isocoumarins. These structurally diverse compounds exhibit a wide range of biological activities, such as antimicrobial, anticancer, and antialgal activities. Some marine Bacillus strains can detoxify heavy metals through reduction processes and have the ability to produce carotenoids. The present article reviews the chemistry and biological activities of secondary metabolites from marine isolates. Side by side, the potential for application of these novel natural products from marine Bacillus strains as drugs, pesticides, carotenoids, and tools for the bioremediation of heavy metal toxicity are also discussed. PMID:23941823

Thiol versus hydroxamate as zinc binding group in HDAC inhibition: An ab initio QM/MM molecular dynamics study.

PubMed

Gong, Wenjing; Wu, Ruibo; Zhang, Yingkai

2015-11-15

Zinc-dependent histone deacetylases (HDACs) play a critical role in transcriptional repression and gene silencing, and are among the most attractive targets for the development of new therapeutics against cancer and various other diseases. Two HDAC inhibitors have been approved by FDA as anti-cancer drugs: one is SAHA whose hydroxamate is directly bound to zinc, the other is FK228 whose active form may use thiol as the zinc binding group. In spite of extensive studies, it remains to be ambiguous regarding how thiol and hydroxamate are bound to the zinc active site of HDACs. In this work, our computational approaches center on Born-Oppenheimer ab initio quantum mechanical/molecular mechanical (QM/MM) molecular dynamics with umbrella sampling, which allow for modeling of the zinc active site with reasonable accuracy while properly including dynamics and effects of protein environment. Meanwhile, an improved short-long effective function (SLEF2) to describe non-bonded interactions between zinc and other atoms has been employed in initial MM equilibrations. Our ab initio QM/MM MD simulations have confirmed that hydroxamate is neutral when it is bound to HDAC8, and found that thiol is deprotonated when directly bound to zinc in the HDAC active site. By comparing thiol and hydroxamate, our results elucidated the differences in their binding environment in the HDAC active sites, and emphasized the importance of the linker design to achieve more specific binding toward class IIa HDACs. © 2015 Wiley Periodicals, Inc.

Thiol Versus Hydroxamate as Zinc Binding Group In HDAC Inhibition: An Ab Initio QM/MM Molecular Dynamics Study

PubMed Central

Gong, Wenjing; Wu, Ruibo; Zhang, Yingkai

2015-01-01

Zinc-dependent histone deacetylases (HDACs) play a critical role in transcriptional repression and gene silencing, and are among the most attractive targets for the development of new therapeutics against cancer and various other diseases. Two HDAC inhibitors have been approved by FDA as anti-cancer drugs: one is SAHA whose hydroxamate is directly bound to zinc, the other is FK228 whose active form may use thiol as the zinc binding group. In spite of extensive studies, it remains to be ambiguous regarding how thiol and hydroxamate are bound to the zinc active site of HDACs. In this work, our computational approaches center on Born-Oppenheimer ab initio quantum mechanical/molecular mechanical (QM/MM) molecular dynamics with umbrella sampling, which allow for modeling of the zinc active site with reasonable accuracy while properly including dynamics and effects of protein environment. Meanwhile, an improved short-long effective function (SLEF2) to describe non-bonded interactions between zinc and other atoms has been employed in initial MM equilibrations. Our ab initio QM/MM MD simulations have confirmed that hydroxamate is neutral when it is bound to HDAC8, and found that thiol is deprotonated when directly bound to zinc in the HDAC active site. By comparing thiol and hydroxamate, our results elucidated the differences in their binding environment in the HDAC active sites, and emphasized the importance of the linker design to achieve more specific binding towards class IIa HDACs. PMID:26452222

Identification of Thiols in Yellow Onion (Allium cepa L.) Using Solvent Vented Large Volume Injection GC-MS.

PubMed

Wermes, Clint; Cannon, Robert; Haasnoot, Sytze; Colstee, Hans; Niedeveld, Cor; Koopmanschap, Gijs; Da Costa, Neil C

2017-11-01

Thiols are often highly odor active molecules and as such can significantly contribute to aroma while being present at extremely low concentrations. This paper details the identification of thiols in yellow onion juice by solvent extraction followed by thiol enrichment using a mercuric agarose gel column. Due to the inherent thermal instability and low concentrations of thiols in onion, chromatographic analysis utilized larger volume solvent elimination injections. New sulfur compounds in onion included 1,1-propanedithiol, bis-(1-sulfanylpropyl)-sulfide, 1-methylsulfanyl-1-propanethiol, 1-propylsulfanyl-1-propanethiol, and 1-allylsulfanyl-1-propanethiol. A discussion on the potential route of formation for each compound is included along with the orthonasal and retronasal evaluations of the synthesized molecules. This work investigated and identified 5 newly identified compounds present in onions that can impart onion character at low concentrations levels. © 2017 Institute of Food Technologists®.

Low-molecular-weight thiols in streptomycetes and their potential role as antioxidants.

PubMed Central

Newton, G L; Fahey, R C; Cohen, G; Aharonowitz, Y

1993-01-01

The intracellular low-molecular-weight thiols present in five gram-positive Streptomyces species and one Flavobacterium species were analyzed by high-performance liquid chromatography after fluorescence labeling with monobromobimane. Bacteria were chosen to include penicillin and cephalosporin beta-lactam producers and nonproducers. No significant amount of glutathione was found in any of the streptomycetes. Major intracellular thiols in all strains examined were cysteine, coenzyme A, sulfide, thiosulfate, and an unknown thiol designated U17. Those streptomycetes that make beta-lactam antibiotics also produce significant amounts of delta-(L-alpha-aminoadipyl)-L-cysteinyl-D-valine (ACV), a key intermediate in their biosynthesis. In Streptomyces clavuligerus, a potent producer of beta-lactams, the level of ACV was low during the early phase of growth and increased rapidly toward the end of exponential growth, paralleling that of antibiotic production. These and other observations indicate that ACV does not function as a protective thiol in streptomycetes. U17 may have this role since it was the major thiol in all streptomycetes and appeared to occur at levels about 10-fold higher than those of the other thiols measured, including ACV. Purification and amino acid analysis of U17 indicated that it contains cysteine and an unusual amine that is not one of the common amino acids. This thiol is identical to an unknown thiol found previously in Micrococcus roseus and Streptomyces griseus. A high level of ergothioneine was found in Streptomyces lactamdurans, and several unidentified thiols were detected in this and other streptomycetes. PMID:8478335

Metabolites of Ginger Component [6]-Shogaol Remain Bioactive in Cancer Cells and Have Low Toxicity in Normal Cells: Chemical Synthesis and Biological Evaluation

PubMed Central

Zhu, Yingdong; Chen, Huadong; Sang, Shengmin

2013-01-01

Our previous study found that [6]-shogaol, a major bioactive component in ginger, is extensively metabolized in cancer cells and in mice. It is unclear whether these metabolites retain bioactivity. The aim of the current study is to synthesize the major metabolites of [6]-shogaol and evaluate their inhibition of growth and induction of apoptosis in human cancer cells. Twelve metabolites of [6]-shogaol (M1, M2, and M4–M13) were successfully synthesized using simple and easily accessible chemical methods. Growth inhibition assays showed that most metabolites of [6]-shogaol had measurable activities against human cancer cells HCT-116 and H-1299. In particular, metabolite M2 greatly retained the biological activities of [6]-shogaol, with an IC50 of 24.43 µM in HCT-116 human colon cancer cells and an IC50 of 25.82 µM in H-1299 human lung cancer cells. Also exhibiting a relatively high potency was thiol-conjugate M13, with IC50 values of 45.47 and 47.77 µM toward HCT-116 and H-1299 cells, respectively. The toxicity evaluation of the synthetic metabolites (M1, M2, and M4–M13) against human normal fibroblast colon cells CCD-18Co and human normal lung cells IMR-90 demonstrated a detoxifying metabolic biotransformation of [6]-shogaol. The most active metabolite M2 had almost no toxicity to CCD-18Co and IMR-90 normal cells with IC50s of 99.18 and 98.30 µM, respectively. TUNEL (Terminal deoxynucleotidyl transferase dUTP nick end labeling) assay indicated that apoptosis was triggered by metabolites M2, M13, and its two diastereomers M13-1 and M13-2. There was no significant difference between the apoptotic effect of [6]-shogaol and the effect of M2 and M13 after 6 hour treatment. PMID:23382939

Soft-lithography fabrication of microfluidic features using thiol-ene formulations.

PubMed

Ashley, John F; Cramer, Neil B; Davis, Robert H; Bowman, Christopher N

2011-08-21

In this work, a novel thiol-ene based photopolymerizable resin formulation was shown to exhibit highly desirable characteristics, such as low cure time and the ability to overcome oxygen inhibition, for the photolithographic fabrication of microfluidic devices. The feature fidelity, as well as various aspects of the feature shape and quality, were assessed as functions of various resin attributes, particularly the exposure conditions, initiator concentration and inhibitor to initiator ratio. An optical technique was utilized to evaluate the feature fidelity as well as the feature shape and quality. These results were used to optimize the thiol-ene resin formulation to produce high fidelity, high aspect ratio features without significant reductions in feature quality. For structures with aspect ratios below 2, little difference (<3%) in feature quality was observed between thiol-ene and acrylate based formulations. However, at higher aspect ratios, the thiol-ene resin exhibited significantly improved feature quality. At an aspect ratio of 8, raised feature quality for the thiol-ene resin was dramatically better than that achieved by using the acrylate resin. The use of the thiol-ene based resin enabled fabrication of a pinched-flow microfluidic device that has complex channel geometry, small (50 μm) channel dimensions, and high aspect ratio (14) features. This journal is © The Royal Society of Chemistry 2011

Synthesis and Antioxidant Activity of 2-Amino-5-methylthiazol Derivatives Containing 1,3,4-Oxadiazole-2-thiol Moiety.

PubMed

Mohana, Kikkeri N; Kumar, Chikkur B Pradeep

2013-01-01

A series of new 5-(2-amino-5-methylthiazol-4-yl)-1,3,4-oxadiazole-2-thiol derivatives 6(a-j) were designed and synthesized with various substituted aldehydes. The chemical structures were confirmed by elemental analyses, FT-IR, (1)H NMR, and mass spectral studies. The antioxidant activity of the synthesized compounds was evaluated by 2,2-diphenyl-1-picrylhydrazyl (DPPH), hydroxyl, nitric oxide, and superoxide radical scavenging assay methods. Compounds 6a, 6e, and 6c showed significant radical scavenging potential due to the presence of electron donating substituent on substituted aldehydes.

Thiol oxidation by nitrosative stress: Cellular localization in human spermatozoa.

PubMed

Cabrillana, María E; Uribe, Pamela; Villegas, Juana V; Álvarez, Juan; Sánchez, Raúl; Fornés, Miguel W

2016-10-01

Peroxynitrite is a highly reactive nitrogen species and when it is generated at high levels it causes nitrosative stress, an important cause of impaired sperm function. High levels of peroxynitrite have been shown to correlate with decreased semen quality in infertile men. Thiol groups in sperm are mainly found in enzymes, antioxidant molecules, and structural proteins in the axoneme. Peroxynitrite primarily reacts with thiol groups of cysteine-containing proteins. Although it is well known that peroxynitrite oxidizes sulfhydryl groups in sperm, the subcellular localization of this oxidation remains unknown. The main objective of this study was to establish the subcellular localization of peroxynitrite-induced nitrosative stress in thiol groups and its relation to sperm motility in human spermatozoa. For this purpose, spermatozoa from healthy donors were exposed in vitro to 3-morpholinosydnonimine (SIN-1), a compound which generates peroxynitrite. In order to detect peroxynitrite and reduced thiol groups, the fluorescent probes, dihydrorhodamine 123 and monobromobimane (mBBr), were used respectively. Sperm viability was analyzed by propidium iodide staining. Peroxynitrite generation and thiol redox state were monitored by confocal microscopy whereas sperm viability was evaluated by flow cytometry. Sperm motility was analyzed by CASA using the ISAS(®) system. The results showed that exposure of human spermatozoa to peroxynitrite results in increased thiol oxidation which is mainly localized in the sperm head and principal piece regions. Thiol oxidation was associated with motility loss. The high susceptibility of thiol groups to peroxynitrite-induced oxidation could explain, at least in part, the negative effect of reactive nitrogen species on sperm motility. DHR: dihydrorhodamine 123; mBBr: monobromobimane ONOO(-): peroxynitrite RNS: reactive nitrogen species RFI: relative fluorescence intensity SIN-1: 3-morpholinosydnonimine CASA: Computer-Aided Sperm Analysis

Strategies to enhance biologically active-secondary metabolites in cell cultures of Artemisia - current trends.

PubMed

Ali, Mohammad; Abbasi, Bilal Haider; Ahmad, Nisar; Khan, Haji; Ali, Gul Shad

2017-11-01

The genus Artemisia has been utilized worldwide due to its immense potential for protection against various diseases, especially malaria. Artemisia absinthium, previously renowned for its utilization in the popular beverage absinthe, is gaining resurgence due to its extensive pharmacological activities. Like A. annua, this species exhibits strong biological activities like antimalarial, anticancer and antioxidant. Although artemisinin was found to be the major metabolite for its antimalarial effects, several flavonoids and terpenoids are considered to possess biological activities when used alone and also to synergistically boost the bioavailability of artemisinin. However, due to the limited quantities of these metabolites in wild plants, in vitro cultures were established and strategies have been adopted to enhance medicinally important secondary metabolites in these cultures. This review elaborates on the traditional medicinal uses of Artemisia species and explains current trends to establish cell cultures of A. annua and A. absinthium for enhanced production of medicinally important secondary metabolites.

Long-Chain Metabolites of Vitamin E: Metabolic Activation as a General Concept for Lipid-Soluble Vitamins?

PubMed

Schubert, Martin; Kluge, Stefan; Schmölz, Lisa; Wallert, Maria; Galli, Francesco; Birringer, Marc; Lorkowski, Stefan

2018-01-12

Vitamins E, A, D and K comprise the class of lipid-soluble vitamins. For vitamins A and D, a metabolic conversion of precursors to active metabolites has already been described. During the metabolism of vitamin E, the long-chain metabolites (LCMs) 13'-hydroxychromanol (13'-OH) and 13'-carboxychromanol (13'-COOH) are formed by oxidative modification of the side-chain. The occurrence of these metabolites in human serum indicates a physiological relevance. Indeed, effects of the LCMs on lipid metabolism, apoptosis, proliferation and inflammatory actions as well as tocopherol and xenobiotic metabolism have been shown. Interestingly, there are several parallels between the actions of the LCMs of vitamin E and the active metabolites of vitamin A and D. The recent findings that the LCMs exert effects different from that of their precursors support their putative role as regulatory metabolites. Hence, it could be proposed that the mode of action of the LCMs might be mediated by a mechanism similar to vitamin A and D metabolites. If the physiological relevance and this concept of action of the LCMs can be confirmed, a general concept of activation of lipid-soluble vitamins via their metabolites might be deduced.

Evaluation of low molecular mass thiols content in carotid atherosclerotic plaques.

PubMed

Zinellu, Angelo; Lepedda, Antonio; Sotgia, Salvatore; Zinellu, Elisabetta; Scanu, Bastianina; Turrini, Franco; Spirito, Rita; Deiana, Luca; Formato, Marilena; Carru, Ciriaco

2009-06-01

Despite the evidence that both homocysteine and cysteine are important risk factors for vascular disease and atherosclerosis no information are reported about their effective amount in plaque and on the relationship with the other low molecular weight thiols. We used capillary electrophoresis to measure thiols in both carotid plaque specimens and plasma samples from 37 patients undergoing carotid endarterectomy. Pearson's correlation shows that intraplaque homocysteine, cysteine and cysteinylglycine levels are related to their plasma concentrations. The distribution of intraplaque GSH and Glu-Cys was higher than that of the same thiols in plasma, whereas the other thiols were significantly less prevalent in plaque than in plasma. Intraplaque haemoglobin and GSH levels were correlated, thus suggesting their common origin from erythrocytes lysis. Data suggest that increased levels of intraplaque glutathione may induce important effects on plaque fate by perturbing the normal LMW thiol redox state.

Detection of intracellular glutathione using ThiolTracker violet stain and fluorescence microscopy.

PubMed

Mandavilli, Bhaskar S; Janes, Michael S

2010-07-01

Glutathione plays an important role in protecting mammalian cells from oxidative stress and cell death. Because reduced glutathione (GSH) represents the large majority of intracellular free thiols, cell-permeant, thiol-reactive fluorescent probes represent potentially useful indicators of intracellular GSH. The ThiolTracker Violet stain (a registered trademark of Invitrogen) is a bright fluorescent probe that is highly reactive to thiols and can be used as a convenient and effective indicator of intracellular GSH and general redox status by a variety of detection modalities. While this probe has been validated in flow cytometry and microplate fluorimetry assays, the following method will describe details on the use of the ThiolTracker Violet dye in traditional fluorescence microscopy, as well as high-content imaging and analysis.

Antifeedant Activity of Ginkgo biloba Secondary Metabolites against Hyphantria cunea Larvae: Mechanisms and Applications

PubMed Central

Ren, Lili; Chen, Fang; Feng, Yuqian

2016-01-01

Ginkgo biloba is a typical relic plant that rarely suffers from pest hazards. This study analyzed the pattern of G. biloba pest hazards in Beijing; tested the antifeedant activity of G. biloba extracts, including ginkgo flavonoids, ginkgolide, and bilobalide, against Hyphantria cunea larvae; determined the activities of glutathione transferase (GSTs), acetylcholinesterase (AChE), carboxylesterase (CarE) and mixed-functional oxidase (MFO), in larvae after feeding on these G. biloba secondary metabolites; and screened for effective botanical antifeedants in the field. In this study, no indicators of insect infestation were found for any of the examined leaves of G. biloba; all tested secondary metabolites showed significant antifeedant activity and affected the activity of the four larval detoxifying enzymes. Ginkgolide had the highest antifeedant activity and the most significant effect on the detoxifying enzymes (P<0.05). Spraying leaves with G. biloba extracts or ginkgolide both significantly repelled H. cunea larvae in the field (P<0.05), although the former is more economical and practical. This study investigated the antifeedant activity of G. biloba secondary metabolites against H. cunea larvae, and the results provide new insights into the mechanism of G. biloba pest resistance. This study also developed new applications of G. biloba secondary metabolites for effective pest control. PMID:27214257

Antifeedant Activity of Ginkgo biloba Secondary Metabolites against Hyphantria cunea Larvae: Mechanisms and Applications.

PubMed

Pan, Long; Ren, Lili; Chen, Fang; Feng, Yuqian; Luo, Youqing

2016-01-01

Ginkgo biloba is a typical relic plant that rarely suffers from pest hazards. This study analyzed the pattern of G. biloba pest hazards in Beijing; tested the antifeedant activity of G. biloba extracts, including ginkgo flavonoids, ginkgolide, and bilobalide, against Hyphantria cunea larvae; determined the activities of glutathione transferase (GSTs), acetylcholinesterase (AChE), carboxylesterase (CarE) and mixed-functional oxidase (MFO), in larvae after feeding on these G. biloba secondary metabolites; and screened for effective botanical antifeedants in the field. In this study, no indicators of insect infestation were found for any of the examined leaves of G. biloba; all tested secondary metabolites showed significant antifeedant activity and affected the activity of the four larval detoxifying enzymes. Ginkgolide had the highest antifeedant activity and the most significant effect on the detoxifying enzymes (P<0.05). Spraying leaves with G. biloba extracts or ginkgolide both significantly repelled H. cunea larvae in the field (P<0.05), although the former is more economical and practical. This study investigated the antifeedant activity of G. biloba secondary metabolites against H. cunea larvae, and the results provide new insights into the mechanism of G. biloba pest resistance. This study also developed new applications of G. biloba secondary metabolites for effective pest control.

New brominated flame retardants and their metabolites as activators of the pregnane X receptor.

PubMed

Gramec Skledar, Darja; Tomašič, Tihomir; Carino, Adriana; Distrutti, Eleonora; Fiorucci, Stefano; Peterlin Mašič, Lucija

2016-09-30

The present study investigated the activities on different nuclear receptors of the new brominated flame retardants 2-ethylhexyl 2,3,4,5-tetrabromobenzoate (TBB) and bis(2-ethylhexyl) 2,3,4,5-tetrabromophthalate (TBPH), and their main carboxylic acid metabolites 2,3,4,5-tetrabromobenzoic acid (TBBA) and mono(2-ethylhexyl) tetrabromophthalate (TBMEPH). None of selected chemicals exhibited marked activity towards PPARα and PPARγ by the use of transactivation assays in HepG2 cells transfected with peroxisome proliferator-activated receptors. In contrast, selected flame retardants all exhibited potent agonist activity on pregnane X receptor (PXR), with EC50 values of 5.5μM for TBPH and 2.0μM for its metabolite TBMEPH. Molecular docking of TBPH and TBMEPH to the PXR ligand binding site revealed similar interactions, with differences only for conformation and orientation of the alkyl chains. Additionally, TBPH showed antagonist activity on PXR (IC50, 13.9μM). Moreover, there was significant up-regulation of CYP3A4 expression via PXR activation for TBB and TBPH and their metabolites. Induction of CYP3A4 might cause undesired drug-drug interactions, lower bioavailability of pharmaceutical drugs, higher formation of reactive toxic metabolites, or enhanced elimination of endogenous hormones, such as T3/T4, to lead to endocrine disruption. These data provide new and important insights into the toxicity of these new polybrominated flame retardants, TBB and TBPH, and their metabolites. Copyright © 2016 Elsevier Ireland Ltd. All rights reserved.

Dinitrosyl iron complexes with thiol-containing ligands as a "working form" of endogenous nitric oxide.

PubMed

Vanin, Anatoly F

2016-04-01

The material presented herein is an overview of the results obtained by our research team during the many years' study of biological activities and occurrence of dinitrosyl iron complexes (DNIC) with thiol-containing ligands in human and animal organisms. With regard to their dose dependence and vast diversity of biological activities, DNIC are similar to the system of endogenous NO, one of the most universal regulators of biological processes. The role of biologically active components in DNIC is played by their iron-dinitrosyl fragments, [Fe(NO)2], endowed with the ability to generate neutral NO molecules and nitrosonium ions (NO(+)). Their release is effected by heme-and thiol-containing proteins, which fulfill the function of biological targets and acceptors of NO and NO(+). Beneficial regulatory effects of DNIC on physiological and metabolic processes are numerous and diverse and include, among other things, lowering of arterial pressure and accelerated healing of skin wounds. In the course of fast decomposition of their Fe(NO)2 fragments (e.g., in the presence of iron chelators), DNIC produce adverse (cytotoxic) effects, which can best be exemplified by their ability to suppress the development of experimental endometriosis in animals. In animal tissues, DNIC with thiol-containing ligands are predominantly represented by the binuclear form, which, contrary to mononuclear DNIC detectable by the 2.03 signal, is EPR-silent. The ample body of evidence on biological activities and occurrence of DNIC gained so far clearly demonstrates that in human and animal organisms DNIC with thiol-containing ligands represent a "working form" of the system of endogenous NO responsible for its accumulation and stabilization in animal tissues as well as its further transfer to its biological targets. Copyright © 2016 Elsevier Inc. All rights reserved.

Spectrophotometric Determination of Phenolic Antioxidants in the Presence of Thiols and Proteins

PubMed Central

Avan, Aslı Neslihan; Demirci Çekiç, Sema; Uzunboy, Seda; Apak, Reşat

2016-01-01

Development of easy, practical, and low-cost spectrophotometric methods is required for the selective determination of phenolic antioxidants in the presence of other similar substances. As electron transfer (ET)-based total antioxidant capacity (TAC) assays generally measure the reducing ability of antioxidant compounds, thiols and phenols cannot be differentiated since they are both responsive to the probe reagent. In this study, three of the most common TAC determination methods, namely cupric ion reducing antioxidant capacity (CUPRAC), 2,2′-azinobis(3-ethylbenzothiazoline-6-sulfonic acid) diammonium salt/trolox equivalent antioxidant capacity (ABTS/TEAC), and ferric reducing antioxidant power (FRAP), were tested for the assay of phenolics in the presence of selected thiol and protein compounds. Although the FRAP method is almost non-responsive to thiol compounds individually, surprising overoxidations with large positive deviations from additivity were observed when using this method for (phenols + thiols) mixtures. Among the tested TAC methods, CUPRAC gave the most additive results for all studied (phenol + thiol) and (phenol + protein) mixtures with minimal relative error. As ABTS/TEAC and FRAP methods gave small and large deviations, respectively, from additivity of absorbances arising from these components in mixtures, mercury(II) compounds were added to stabilize the thiol components in the form of Hg(II)-thiol complexes so as to enable selective spectrophotometric determination of phenolic components. This error compensation was most efficient for the FRAP method in testing (thiols + phenols) mixtures. PMID:27529232

Spectrophotometric Determination of Phenolic Antioxidants in the Presence of Thiols and Proteins.

PubMed

Avan, Aslı Neslihan; Demirci Çekiç, Sema; Uzunboy, Seda; Apak, Reşat

2016-08-12

Development of easy, practical, and low-cost spectrophotometric methods is required for the selective determination of phenolic antioxidants in the presence of other similar substances. As electron transfer (ET)-based total antioxidant capacity (TAC) assays generally measure the reducing ability of antioxidant compounds, thiols and phenols cannot be differentiated since they are both responsive to the probe reagent. In this study, three of the most common TAC determination methods, namely cupric ion reducing antioxidant capacity (CUPRAC), 2,2'-azinobis(3-ethylbenzothiazoline-6-sulfonic acid) diammonium salt/trolox equivalent antioxidant capacity (ABTS/TEAC), and ferric reducing antioxidant power (FRAP), were tested for the assay of phenolics in the presence of selected thiol and protein compounds. Although the FRAP method is almost non-responsive to thiol compounds individually, surprising overoxidations with large positive deviations from additivity were observed when using this method for (phenols + thiols) mixtures. Among the tested TAC methods, CUPRAC gave the most additive results for all studied (phenol + thiol) and (phenol + protein) mixtures with minimal relative error. As ABTS/TEAC and FRAP methods gave small and large deviations, respectively, from additivity of absorbances arising from these components in mixtures, mercury(II) compounds were added to stabilize the thiol components in the form of Hg(II)-thiol complexes so as to enable selective spectrophotometric determination of phenolic components. This error compensation was most efficient for the FRAP method in testing (thiols + phenols) mixtures.

A periodic mixed gaussians-plane waves DFT study on simple thiols on Au(111): adsorbate species, surface reconstruction, and thiols functionalization.

PubMed

Rajaraman, Gopalan; Caneschi, Andrea; Gatteschi, Dante; Totti, Federico

2011-03-07

Here we present DFT calculations based on a periodic mixed gaussians/plane waves approach to study the energetics, structure, bonding of SAMs of simple thiols on Au(111). Several open issues such as structure, bonding and the nature of adsorbate are taken into account. We started with methyl thiols (MeSH) on Au(111) to establish the nature of the adsorbate. We have considered several structural models embracing the reconstructed surface scenario along with the MeS˙-Au(ad)-MeS˙ type motif put forward in recent years. Our calculations suggest a clear preference for the homolytic cleavage of the S-H bond leading to a stable MeS˙ on a gold surface. In agreement with the recent literature studies, the reconstructed models of the MeS˙ species are found to be energetically preferred over unreconstructed models. Besides, our calculations reveal that the model with 1:2 Au(ad)/thiols ratio, i.e. MeS˙-Au(ad)-MeS˙, is energetically preferred compared to the clean and 1:1 ratio models, in agreement with the experimental and theoretical evidences. We have also performed Molecular Orbital/Natural Bond Orbital, MO/NBO, analysis to understand the electronic structure and bonding in different structural motifs and many useful insights have been gained. Finally, the studies have then been extended to alkyl thiols of the RSR' (R, R' = Me, Et and Ph) type and here our calculations again reveal a preference for the RS˙ type species adsorption for clean as well as for reconstructed 1:2 Au(ad)/thiols ratio models.

Iron and thiols as two major players in carcinogenesis: friends or foes?

PubMed

Toyokuni, Shinya

2014-01-01

Iron is the most abundant metal in the human body and mainly works as a cofactor for proteins such as hemoglobin and various enzymes. No independent life forms on earth can survive without iron. However, excess iron is intimately associated with carcinogenesis by increasing oxidative stress via its catalytic activity to generate hydroxyl radicals. Biomolecules with redox-active sulfhydryl function(s) (thiol compounds) are necessary for the maintenance of mildly reductive cellular environments to counteract oxidative stress, and for the execution of redox reactions for metabolism and detoxification. Involvement of glutathione S-transferase and thioredoxin has long attracted the attention of cancer researchers. Here, I update recent findings on the involvement of iron and thiol compounds during carcinogenesis and in cancer cells. It is now recognized that the cystine/glutamate transporter (antiporter) is intimately associated with ferroptosis, an iron-dependent, non-apoptotic form of cell death, observed in cancer cells, and also with cancer stem cells; the former with transporter blockage but the latter with its stabilization. Excess iron in the presence of oxygen appears the most common known mutagen. Ironically, the persistent activation of antioxidant systems via genetic alterations in Nrf2 and Keap1 also contributes to carcinogenesis. Therefore, it is difficult to conclude the role of iron and thiol compounds as friends or foes, which depends on the quantity/distribution and induction/flexibility, respectively. Avoiding further mutation would be the most helpful strategy for cancer prevention, and myriad of efforts are being made to sort out the weaknesses of cancer cells.

Heavy metal ion removal by thiol functionalized aluminum oxide hydroxide nanowhiskers

NASA Astrophysics Data System (ADS)

Xia, Zhiyong; Baird, Lance; Zimmerman, Natasha; Yeager, Matthew

2017-09-01

In this study, we developed a cost effective method of using thiol functionalized γ-aluminum oxide hydroxide (γ-AlOOH) filters for removing three key heavy metals from water: mercury, lead, and cadmium under non-concomitant conditions. Compared to non-thiol treated γ-AlOOH filters, the introduction of thiol functional groups greatly improved the heavy metal removal efficiency under both static and dynamic filtration conditions. The adsorption kinetics of thiol functionalized γ-AlOOH were investigated using the Lagergren first order and pseudo-second order kinetics models; whereas the isothermal adsorption behavior of these membranes was revealed through the Langmuir and Freundlich models. Heavy metal concentration was quantified by Inductively Coupled Plasma-Mass Spectroscopy, and the thiol level on γ-AlOOH surface was measured by a colorimetric assay using Ellman's reagent. X-ray photoelectron spectroscopy was used to further address the surface sulfur state on the membranes after heavy metal exposure. Mechanisms for heavy metal adsorption were also discussed.

The synthesis of novel hybrid thiol-functionalized nano-structured SBA-15

NASA Astrophysics Data System (ADS)

Hoang, Van Duc; Phuong Dang, Tuyet; Khieu Dinh, Quang; Phu Nguyen, Huu; Vu, Anh Tuan

2010-09-01

Mesoporous thiol-functionalized SBA-15 has been directly synthesized by co-condensation of tetraethyl orthosilicate (TEOS) and 3-mercaptopropyltrimethoxysilane (MPTMS) with triblock copolymer P123 as-structure-directing agent under hydrothermal conditions. Surfactant removal was performed by Soxhlet ethanol extraction. These materials have been characterized by powder x-ray diffraction (XRD), nitrogen adsorption/desorption (BET model), transmission electron microscopy (TEM), thermal analysis, infrared spectroscopy (IR) and energy-dispersive x-ray spectroscopy (EDX). The main parameters, such as the initial molar ratio of MPTMS to TEOS, the time of adding MPTMS to synthesized gel and the Soxhlet ethanol extraction on the thiol functionalized SBA-15 with high thiol content and highly ordered hexagonal mesostructure, were investigated and evaluated. The adsorption capacity of the thiol-functionalized and non-functionalized SBA-15 materials for Pb2+ ion from aqueous solution was tested. It was found that the Pb2+ adsorption capacity of the thiol functionalized SBA-15 is three times higher than that of non-functionalized SBA-15.

Thiol/disulfide homeostasis in pregnant women with obstructive sleep apnea syndrome.

PubMed

Üstündağ, Yasemin; Demirci, Hakan; Balık, Rifat; Erel, Ozcan; Özaydın, Fahri; Kücük, Bilgen; Ertaş, Dilber; Ustunyurt, Emin

2017-11-27

Repetitive episodes of hypoxia and reoxygenation during sleep in patients with obstructive sleep apnea syndrome (OSAS) resemble an ischemia-reperfusion injury. We aimed to test the hypothesis that oxidative stress occurs in pregnant women with OSAS. We also aimed to compare thiol/disulfide homeostasis with ischemia-modified albumin (IMA) and total antioxidant capacity (TAC) as markers of ischemia-reperfusion injury in pregnant women with and without OSAS and healthy control. This study included 29 pregnant women with OSAS, 30 women without OSAS in the third trimester applying for periodic examinations, and 30 healthy women. Serum IMA and TAC (using the ferric reducing power of plasma method) were measured. Serum thiol/disulfide homeostasis was determined by a novel automated method. The mean age of the pregnant women with OSAS was 31.0 ± 4.7 years with a mean gestational age of 36.5 ± 3.0 weeks. The mean age of pregnant women without OSAS was 29.8 ± 4.9 years with a mean gestational age of 36.9 ± 2.7 weeks. The mean age of the nonpregnant control group was 29.7 ± 6.4 years. Both native thiol (291 ± 29 μmol/L versus 314 ± 30 μmol/L; p = .018) and total thiol (325 ± 32 versus 350 ± 32, p = .025) levels were lower in pregnant women with OSAS compared to pregnant women without OSAS, respectively (p < .01). This is the first study demonstrating the thiol/disulfide homeostasis in pregnant women with OSAS. Native thiol and total thiol levels were lower in pregnant women with OSAS compared to those without OSAS. However, dynamic thiol/disulfide homeostasis parameters cannot provide valuable information to discriminate OSAS in pregnant women.

Redox mediators in visible light photocatalysis: photocatalytic radical thiol-ene additions.

PubMed

Tyson, Elizabeth L; Niemeyer, Zachary L; Yoon, Tehshik P

2014-02-07

Synthetically useful radical thiol-ene reactions can be initiated by visible light irradiation in the presence of transition metal polypyridyl photocatalysts. The success of this method relies upon the use of p-toluidine as an essential additive. Using these conditions, high-yielding thiol-ene reactions of cysteine-containing biomolecules can be accomplished using biocompatibile wavelengths of visible light, under aqueous conditions, and with the thiol component as the limiting reagent. We present evidence that p-toluidine serves as a redox mediator that is capable of catalyzing the otherwise inefficient photooxidation of thiols to the key thiyl radical intermediate. Thus, we show that co-catalytic oxidants can be important in the design of synthetic reactions involving visible light photoredox catalysis.

Gel Permeation Chromatography Characterization of the Chain Length Distributions in Thiol-Acrylate Photopolymer Networks

PubMed Central

Rydholm, Amber E.; Held, Nicole L.; Bowman, Christopher N.; Anseth, Kristi S.

2008-01-01

Crosslinked, degradable networks formed from the photopolymerization of thiol and acrylate monomers are explored as potential biomaterials. The degradation behavior and material properties of these networks are influenced by the molecular weight of the nondegradable thiol-polyacrylate backbone chains that form during photopolymerization. Here, gel permeation chromatography was used to characterize the thiol-polyacrylate backbone chain lengths in degraded thiol-acrylate networks. Increasing thiol functionality from 1 to 4 increased the backbone molecular weight (M̄w = 2.3 ± 0.07 × 104 Da for monothiol and 3.6 ± 0.1 × 104 Da for tetrathiol networks). Decreasing thiol functional group concentration from 30 to 10 mol% also increased the backbone lengths (M̄w = 7.3 ± 1.1 × 104 Da for the networks containing 10 mol% thiol groups as compared to 3.6 ± 0.1 × 104 Da for 30 mol% thiol). Finally, the backbone chain lengths were probed at various stages of degradation and an increase in backbone molecular weight was observed as mass loss progressed from 10 to 70%. PMID:19079733

[Secondary Metabolites from Marine Microorganisms. I. Secondary Metabolites from Marine Actinomycetes].

PubMed

Orlova, T I; Bulgakova, V G; Polin, A N

2015-01-01

Review represents data on new active metabolites isolated from marine actinomycetes published in 2007 to 2014. Marine actinomycetes are an unlimited source of novel secondary metabolites with various biological activities. Among them there are antibiotics, anticancer compounds, inhibitors of biochemical processes.

Long-Chain Metabolites of Vitamin E: Metabolic Activation as a General Concept for Lipid-Soluble Vitamins?

PubMed Central

Schubert, Martin; Kluge, Stefan; Schmölz, Lisa; Wallert, Maria

2018-01-01

Vitamins E, A, D and K comprise the class of lipid-soluble vitamins. For vitamins A and D, a metabolic conversion of precursors to active metabolites has already been described. During the metabolism of vitamin E, the long-chain metabolites (LCMs) 13′-hydroxychromanol (13′-OH) and 13′-carboxychromanol (13′-COOH) are formed by oxidative modification of the side-chain. The occurrence of these metabolites in human serum indicates a physiological relevance. Indeed, effects of the LCMs on lipid metabolism, apoptosis, proliferation and inflammatory actions as well as tocopherol and xenobiotic metabolism have been shown. Interestingly, there are several parallels between the actions of the LCMs of vitamin E and the active metabolites of vitamin A and D. The recent findings that the LCMs exert effects different from that of their precursors support their putative role as regulatory metabolites. Hence, it could be proposed that the mode of action of the LCMs might be mediated by a mechanism similar to vitamin A and D metabolites. If the physiological relevance and this concept of action of the LCMs can be confirmed, a general concept of activation of lipid-soluble vitamins via their metabolites might be deduced. PMID:29329238

Buprenorphine metabolites, buprenorphine-3-glucuronide and norbuprenorphine-3-glucuronide, are biologically active

PubMed Central

Brown, Sarah M.; Holtzman, Michael; Kim, Thomas; Kharasch, Evan D.

2012-01-01

Background The long-lasting high affinity opioid buprenorphine has complex pharmacology including ceiling effects with respect to analgesia and respiratory depression. Plasma concentrations of the major buprenorphine metabolites norbuprenorphine, buprenorphine-3-glucuronide, and norbuprenorphine-3-glucuronide approximate or exceed those of the parent drug. Buprenorphine glucuronide metabolites pharmacology is undefined. This investigation determined binding and pharmacological activity of the two glucuronide metabolites, and in comparison with buprenorphine and norbuprenorphine. Methods Competitive inhibition of radioligand binding to human mu, kappa, delta opioid and nociceptin receptors was used to determine glucuronide binding affinities for these receptors. Common opiate effects were assessed in vivo in Swiss Webster mice. Antinociception was assessed using a tail-flick assay, respiratory effects were measured using unrestrained whole-body plethysmography, and sedation was assessed by inhibition of locomotion measured by open-field testing. Results Buprenorphine-3-glucuronide had high affinity for human mu (Ki = 4.9±2.7 pM), delta (Ki = 270±0.4 nM), and nociceptin (Ki = 36±0.3 μM) but not kappa receptors. Norbuprenorphine-3-glucuronide had affinity for human kappa (Ki = 300±0.5 nM) and nociceptin (Ki= 18±0.2 μM) but not mu or delta receptors. At the dose tested, buprenorphine-3-glucuronide had a small antinociceptive effect. Neither glucuronide had significant effects on respiratory rate, but norbuprenorphine-3-glucuronide decreased tidal volume. Norbuprenorphine-3-glucuronide also caused sedation. Conclusions Both glucuronide metabolites of buprenorphine are biologically active at doses relevant to metabolite exposures which occur after buprenorphine. Activity of the glucuronides may contribute to the overall pharmacology of buprenorphine. PMID:22037640

Simulation studies on structural and thermal properties of alkane thiol capped gold nanoparticles.

PubMed

Devi, J Meena

2017-06-01

The structural and thermal properties of the passivated gold nanoparticles were explored employing molecular dynamics simulation for the different surface coverage densities of the self-assembled monolayer (SAM) of alkane thiol. The structural properties of the monolayer protected gold nanoparticles such us overall shape, organization and conformation of the capping alkane thiol chains were found to be influenced by the capping density. The structural order of the thiol capped gold nanoparticles enhances with the increase in the surface coverage density. The specific heat capacity of the alkane thiol capped gold nanoparticles was found to increase linearly with the thiol coverage density. This may be attributed to the enhancement in the lattice vibrational energy. The present simulation results suggest, that the structural and thermal properties of the alkane thiol capped gold nanoparticles may be modified by the suitable selection of the SAM coverage density. Copyright © 2017 Elsevier Inc. All rights reserved.

Bisphenol A and its analogs: Do their metabolites have endocrine activity?

PubMed

Gramec Skledar, Darja; Peterlin Mašič, Lucija

2016-10-01

Structural analogs of bisphenol A are commonly used as its alternatives in industrial and commercial applications. Nevertheless, the question arises whether the use of other bisphenols is justified as replacements for bisphenol A in mass production of plastic materials. To evaluate the influence of metabolic reactions on endocrine activities of bisphenols, we conducted a systematic review of the literature. Knowledge about the metabolic pathways and enzymes involved in metabolic biotransformations is essential for understanding and predicting mechanisms of toxicity. Bisphenols are metabolized predominantly by the glucuronidation reaction, which is considered their most important detoxification pathway, as based on current knowledge, glucuronides do not have activity on endocrine receptors. In contrast, several oxidative metabolites of bisphenols with enhanced endocrine activities are presented, and these findings indicate that oxidative metabolites of bisphenols can still have endocrine activities in humans. Copyright © 2016 Elsevier B.V. All rights reserved.

Synthesis and Antioxidant Activity of 2-Amino-5-methylthiazol Derivatives Containing 1,3,4-Oxadiazole-2-thiol Moiety

PubMed Central

Mohana, Kikkeri N.; Kumar, Chikkur B. Pradeep

2013-01-01

A series of new 5-(2-amino-5-methylthiazol-4-yl)-1,3,4-oxadiazole-2-thiol derivatives 6(a–j) were designed and synthesized with various substituted aldehydes. The chemical structures were confirmed by elemental analyses, FT-IR, 1H NMR, and mass spectral studies. The antioxidant activity of the synthesized compounds was evaluated by 2,2-diphenyl-1-picrylhydrazyl (DPPH), hydroxyl, nitric oxide, and superoxide radical scavenging assay methods. Compounds 6a, 6e, and 6c showed significant radical scavenging potential due to the presence of electron donating substituent on substituted aldehydes. PMID:24052865

Thiol-Disulfide Exchange in Peptides Derived from Human Growth Hormone

PubMed Central

Chandrasekhar, Saradha; Epling, Daniel E.; Sophocleous, Andreas M.; Topp, Elizabeth M.

2014-01-01

Disulfide bonds stabilize proteins by crosslinking distant regions into a compact three-dimensional structure. They can also participate in hydrolytic and oxidative pathways to form non-native disulfide bonds and other reactive species. Such covalent modifications can contribute to protein aggregation. Here we present experimental data for the mechanism of thiol-disulfide exchange in tryptic peptides derived from human growth hormone in aqueous solution. Reaction kinetics were monitored to investigate the effect of pH (6.0-10.0), temperature (4-50 °C), oxidation suppressants (EDTA and N2 sparging) and peptide secondary structure (amide cyclized vs. open form). The concentrations of free thiol containing peptides, scrambled disulfides and native disulfide-linked peptides generated via thiol-disulfide exchange and oxidation reactions were determined using RP-HPLC and LC-MS. Concentration vs. time data were fitted to a mathematical model using non-linear least squares regression analysis. At all pH values, the model was able to fit the data with R2≥0.95. Excluding oxidation suppressants (EDTA and N2 sparging) resulted in an increase in the formation of scrambled disulfides via oxidative pathways but did not influence the intrinsic rate of thiol-disulfide exchange. In addition, peptide secondary structure was found to influence the rate of thiol-disulfide exchange. PMID:24549831

High-throughput identification of promiscuous inhibitors from screening libraries with the use of a thiol-containing fluorescent probe.

PubMed

McCallum, Megan M; Nandhikonda, Premchendar; Temmer, Jonathan J; Eyermann, Charles; Simeonov, Anton; Jadhav, Ajit; Yasgar, Adam; Maloney, David; Arnold, Alexander Leggy

2013-07-01

Testing small molecules for their ability to modify cysteine residues of proteins in the early stages of drug discovery is expected to accelerate our ability to develop more selective drugs with lesser side effects. In addition, this approach also enables the rapid evaluation of the mode of binding of new drug candidates with respect to thiol reactivity and metabolism by glutathione. Herein, we describe the development of a fluorescence-based high-throughput assay that allows the identification of thiol-reactive compounds. A thiol-containing fluorescent probe, MSTI, was synthesized and used to evaluate small molecules from the Library of Pharmacologically Active Compounds (LOPAC) collection of bioactive molecules. LOPAC compounds that are known to react with sulfur nucleophiles were identified with this assay, for example, irreversible protease inhibitors, nitric oxide-releasing compounds, and proton-pump inhibitors. The results confirm that both electrophilic and redox reactive compounds can be quickly identified in a high-throughput manner, enabling the assessment of screening libraries with respect to thiol-reactive compounds.

Intercalation of gaseous thiols and sulfides into Ag+ ion-exchanged aluminum dihydrogen triphosphate.

PubMed

Hayashi, Aki; Saimen, Hiroki; Watanabe, Nobuaki; Kimura, Hitomi; Kobayashi, Ayumi; Nakayama, Hirokazu; Tsuhako, Mitsutomo

2005-08-02

Ag(+) ion-exchanged layered aluminum dihydrogen triphosphate (AlP) with the interlayer distance of 0.85 nm was synthesized by the ion-exchange of proton in triphosphate with Ag(+) ion. The amount of exchanged Ag(+) ion depended on the concentration of AgNO(3) aqueous solution. Ag(+) ion-exchanged AlP adsorbed gaseous thiols and sulfides into the interlayer region. The adsorption amounts of thiols were more than those of sulfides, thiols with one mercapto group > thiol with two mercapto groups > sulfides, and depended on the amount of exchanged Ag(+) ion in the interlayer region. The thiols with one mercapto group were intercalated to expand the interlayer distance of Ag(+) ion-exchanged AlP, whereas there was no expansion in the adsorption of sulfide. In the case of thiol with two mercapto groups, there was observed contraction of the interlayer distance through the bridging with Ag(+) ions of the upper and lower sides of the interlayer region.

Kinetics and mechanisms of thiol-disulfide exchange covering direct substitution and thiol oxidation-mediated pathways.

PubMed

Nagy, Péter

2013-05-01

Disulfides are important building blocks in the secondary and tertiary structures of proteins, serving as inter- and intra-subunit cross links. Disulfides are also the major products of thiol oxidation, a process that has primary roles in defense mechanisms against oxidative stress and in redox regulation of cell signaling. Although disulfides are relatively stable, their reduction, isomerisation, and interconversion as well as their production reactions are catalyzed by delicate enzyme machineries, providing a dynamic system in biology. Redox homeostasis, a thermodynamic parameter that determines which reactions can occur in cellular compartments, is also balanced by the thiol-disulfide pool. However, it is the kinetic properties of the reactions that best represent cell dynamics, because the partitioning of the possible reactions depends on kinetic parameters. This review is focused on the kinetics and mechanisms of thiol-disulfide substitution and redox reactions. It summarizes the challenges and advances that are associated with kinetic investigations in small molecular and enzymatic systems from a rigorous chemical perspective using biological examples. The most important parameters that influence reaction rates are discussed in detail. Kinetic studies of proteins are more challenging than small molecules, and quite often investigators are forced to sacrifice the rigor of the experimental approach to obtain the important kinetic and mechanistic information. However, recent technological advances allow a more comprehensive analysis of enzymatic systems via using the systematic kinetics apparatus that was developed for small molecule reactions, which is expected to provide further insight into the cell's machinery.

Antimalarial Activity of Plant Metabolites.

PubMed

Pan, Wen-Hui; Xu, Xin-Ya; Shi, Ni; Tsang, Siu Wai; Zhang, Hong-Jie

2018-05-06

Malaria, as a major global health problem, continues to affect a large number of people each year, especially those in developing countries. Effective drug discovery is still one of the main efforts to control malaria. As natural products are still considered as a key source for discovery and development of therapeutic agents, we have evaluated more than 2000 plant extracts against Plasmodium falciparum . As a result, we discovered dozens of plant leads that displayed antimalarial activity. Our phytochemical study of some of these plant extracts led to the identification of several potent antimalarial compounds. The prior comprehensive review article entitled “Antimalarial activity of plant metabolites” by Schwikkard and Van Heerden (2002) reported structures of plant-derived compounds with antiplasmodial activity and covered literature up to the year 2000. As a continuation of this effort, the present review covers the antimalarial compounds isolated from plants, including marine plants, reported in the literature from 2001 to the end of 2017. During the span of the last 17 years, 175 antiplasmodial compounds were discovered from plants. These active compounds are organized in our review article according to their plant families. In addition, we also include ethnobotanical information of the antimalarial plants discussed.

Study of Highly Selective and Efficient Thiol Derivatization using Selenium Reagents by Mass Spectrometry

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

Xu, Kehua; Zhang, Yun W.; Tang, Bo

2010-08-15

Biological thiols are critical physiological components and their detection often involves derivatization. This paper reports a systemic mass spectrometry (MS) investigation of the cleavage of Se-N bond by thiol to form a new Se-S bond, the new selenium chemistry for thiol labeling. Our data shows that the reaction is highly selective, rapid, reversible and efficient. For instance, among twenty amino acids, only cysteine was found to be reactive with Se-N containing reagents and the reaction takes place in seconds. By adding dithiothreitol (DTT), the newly formed Se-S bond of peptides/proteins can be reduced back to free thiol. The high selectivitymore » and excellent reversibility of the reaction provide potential of using this chemistry for selective identification of thiol compounds or enriching and purifying thiol peptides/proteins. In addition, the derivatized thiol peptides have interesting dissociation behavior, which is tunable using different selenium reagents. For example, by introducing an adjacent nucleophilic group into the selenium reagent in the case of using ebselen, the reaction product of ebselen with glutathione (GSH) is easy to lose the selenium tag upon collision-induced dissociation (CID), which is useful to "fish out" those peptides containing free cysteine residues by precursor ion scan. By contrast, the selenium tag of N-(phenylseleno) phthalimide reagent can be stable and survive in CID process, which would be of value in pinpointing thiol location using a top-down proteomic approach. Also, the high conversion yield of the reaction allows the counting of total number of thiol in proteins. We believe that ebselen or N-(phenylseleno) phthalimide as tagging thiol-protein reagents will have important applications in both qualitative and quantitative analysis of different thiol-proteins derived from living cells by MS method.« less

A Potential Biofilm Metabolite Signature for Caries Activity - A Pilot Clinical Study

PubMed Central

Zandona, F; Soini, HA; Novotny, MV; Santiago, E; Eckert, GJ; Preisser, JS; Benecha, HK; Arthur, RA; Zero, DT

2016-01-01

Background This study's aim was to compare the dental biofilm metabolite-profile of caries-active (N=11) or caries-free (N=4) children by gas chromatography-mass spectrometry (GC/MS) analyses. Methods Samples collected after overnight fasting, with or without a previous glucose rinse, were combined for each child based on the caries status of the site, re-suspended in ethanol and analyzed by GC/MS. Results Biofilm from caries-active sites exhibited a different chromatographic profile compared to caries-free sites. Qualitative and quantitative analysis suggested a special cluster of branched alcohols and esters present at substantially higher intensity in biofilms of caries-active sites. Conclusions This pilot study indicates that there are metabolites present in the biofilm which have the potential to provide a characteristic metabolomics signature for caries activity. PMID:27885354

Thiol-Ene functionalized siloxanes for use as elastomeric dental impression materials

PubMed Central

Cole, Megan A.; Jankousky, Katherine C.; Bowman, Christopher N.

2014-01-01

Objectives Thiol- and allyl-functionalized siloxane oligomers are synthesized and evaluated for use as a radical-mediated, rapid set elastomeric dental impression material. Thiol-ene siloxane formulations are crosslinked using a redox-initiated polymerization scheme, and the mechanical properties of the thiol-ene network are manipulated through the incorporation of varying degrees of plasticizer and kaolin filler. Formulations with medium and light body consistencies are further evaluated for their ability to accurately replicate features on both the gross and microscopic levels. We hypothesize that thiol-ene functionalized siloxane systems will exhibit faster setting times and greater detail reproduction than commercially available polyvinylsiloxane (PVS) materials of comparable consistencies. Methods Thiol-ene functionalized siloxane mixtures formulated with varying levels of redox initiators, plasticizer, and kaolin filler are made and evaluated for their polymerization speed (FTIR), consistency (ISO4823.9.2), and surface energy (goniometer). Feature replication is evaluated quantitatively by SEM. The Tg, storage modulus, and creep behavior are determined by DMA. Results Increasing redox initiation rate increases the polymerization rate but at high levels also limits working time. Combining 0.86 wt% oxidizing agent with up to 5 wt% plasticizer gave a working time of 3 min and a setting time of 2 min. The selected medium and light body thiol-ene formulations also achieved greater qualitative detail reproduction than the commercial material and reproduced micrometer patterns with 98% accuracy. Significance Improving detail reproduction and setting speed is a primary focus of dental impression material design and synthesis. Radical-mediated polymerizations, particularly thiol-ene reactions, are recognized for their speed, reduced shrinkage, and ‘click’ nature. PMID:24553250

Confirmation of 1-Phenylethane-1-thiol as the Character Impact Aroma Compound in Curry Leaves and Its Behavior during Tissue Disruption, Drying, and Frying.

PubMed

Steinhaus, Martin

2017-03-15

The most odor-active compounds previously identified by application of an aroma extract dilution analysis were quantitated in freshly picked curry leaves, either by stable isotope dilution assays in combination with GC-GC-MS or by GC-FID after simultaneous extraction/fractionation. Odor activity values (OAVs) were calculated as ratios of concentrations to odor threshold values. The topmost OAVs were obtained for (3Z)-hex-3-enal (grassy; OAV 180 000), (1S)-1-phenylethane-1-thiol (sulfury, burnt; OAV 150 000), (1R)-1-phenylethane-1-thiol (sulfury, burnt; OAV 120 000), (3R)-linalool (citrusy; OAV 58 000), and myrcene (geranium leaf-like; OAV 23 000). The high OAVs calculated for its enantiomers confirmed 1-phenylethane-1-thiol as character impact compound of the typical sulfury and burnt aroma of curry leaves. The 1-phenylethane-1-thiol concentration in curry leaves decreased upon tissue disruption and drying, as well as upon frying of fresh leaves. By contrast, frying of dried leaves led to an increase of 1-phenylethane-1-thiol, indicating a yet unknown thermolabile precursor.

A Study of Functional Polymer Colloids Prepared Using Thiol-Ene/Yne Click Chemistry

NASA Astrophysics Data System (ADS)

Durham, Olivia Z.

This project demonstrates the first instance of thiol-ene chemistry as the polymerization method for the production of polymer colloids in two-phase heterogeneous suspensions, miniemulsions, and emulsions. This work was also expanded to thiol-yne chemistry for the production of polymer particles containing increased crosslinking density. The utility of thiol-ene and thiol-yne chemistries for polymerization and polymer modification is well established in bulk systems. These reactions are considered 'click' reactions, which can be defined as processes that are both facile and simple, offering high yields with nearly 100% conversion, no side products, easy product separation, compatibility with a diverse variety of commercially available starting materials, and orthogonality with other chemistries. In addition, thiol-ene and thiol-yne chemistry follow a step-growth mechanism for the development of highly uniform polymer networks, where polymer growth is dependent on the coupling of functional groups. These step-growth polymerization systems are in stark contrast to the chain-growth mechanisms of acrylic and styrenic monomers that have dominated the field of conventional heterogeneous polymerizations. Preliminary studies evaluated the mechanism of particle production in suspension and miniemulsion systems. Monomer droplets were compared to the final polymer particles to confirm that particle growth occurred through the polymerization of monomer droplets. Additional parameters examined include homogenization energy (mechanical mixing), diluent species and concentration, and monomer content. These reactions were conducted using photoinitiation to yield particles in a matter of minutes with diameters in the size range of several microns to hundreds of microns in suspensions or submicron particles in miniemulsions. Improved control over the particle size and size distribution was examined through variation of reaction parameters. In addition, a method of seeded suspension

Occurrence of low molecular weight thiols in biological systems

NASA Technical Reports Server (NTRS)

Fahey, Robert C.; Newton, Gerald L.

1983-01-01

Bromobimane labeling and high performance chromatography analysis were applied to various species of bacteria, plant tissues, and animal tissues. The reaction between thiols and monobromobimane is studied. Chromatograms revealing peaks produced by nonthiols and thiols are analyzed and compared. It is observed that all the bacteria species contain hydrogen sulfide, and glutathione is contained in facultative and aerobic gram-negative bacteria. For the plant tissues, the data reveal that mung bean sprouts contain homoglutathione and no glutathione; alfalfa sprouts contain homoglutathione and glutathione; the pea seed, nonlegumes, and fungi contain glutathione and no homoglutathione. It is detected that the main thiol in the animal tissues is glutathione. Based on the data, it is suggested that glutathione has an essential function in higher organisms.

Non-protein thiol imaging and quantification in live cells with a novel benzofurazan sulfide triphenylphosphonium fluorogenic compound.

PubMed

Yang, Yang; Guan, Xiangming

2017-05-01

Thiols (-SH) play various roles in biological systems. They are divided into protein thiols (PSH) and non-protein thiols (NPSH). Due to the significant roles thiols play in various physiological/pathological functions, numerous analytical methods have been developed for thiol assays. Most of these methods are developed for glutathione, the major form of NPSH. Majority of these methods require tissue/cell homogenization before analysis. Due to a lack of effective thiol-specific fluorescent/fluorogenic reagents, methods for imaging and quantifying thiols in live cells are limited. Determination of an analyte in live cells can reveal information that cannot be revealed by analysis of cell homogenates. Previously, we reported a thiol-specific thiol-sulfide exchange reaction. Based on this reaction, a benzofurazan sulfide thiol-specific fluorogenic reagent was developed. The reagent was able to effectively image and quantify total thiols (PSH+NPSH) in live cells through fluorescence microscopy. The reagent was later named as GUALY's reagent. Here we would like to report an extension of the work by synthesizing a novel benzofurazan sulfide triphenylphosphonium derivative [(((7,7'-thiobis(benzo[c][1,2,5]oxadiazole-4,4'-sulfonyl))bis(methylazanediyl))bis(butane-4,1-diyl))bis(triphenylphosphonium) (TBOP)]. Like GUALY's reagent, TBOP is a thiol-specific fluorogenic agent that is non-fluorescent but forms fluorescent thiol adducts in a thiol-specific fashion. Different than GUALY's reagent, TBOP reacts only with NPSH but not with PSH. TBOP was effectively used to image and quantify NPSH in live cells using fluorescence microscopy. TBOP is a complementary reagent to GUALY's reagent in determining the roles of PSH, NPSH, and total thiols in thiol-related physiological/pathological functions in live cells through fluorescence microscopy. Graphical Abstract Live cell imaging and quantification of non-protein thiols by TBOP.

Transcriptional activation by heat and cold of a thiol protease gene in tomato. [Lycopersicon esculentum

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

Schaffer, M.A.; Fischer, R.L.

We previously determined that low temperature induces the accumulation in tomato (Lycopersicon esculentum) fruit of a cloned mRNA, designated C14, encoding a polypeptide related to thiol proteases. We now demonstrate that C14 mRNA accumulation is a response common to both high (40{degree}C) and low (4{degree}C) temperature stresses. Exposure of tomato fruit to 40{degree}C results in the accumulation of C14 mRNA, by 8 hours. This response is more rapid than that to 4{degree}C, but slower than the induction of many heat shock messages by 40{degree}C, and therefore unique. We have also studied the mechanism by which heat and cold exposure activatemore » C14 gene expression. Both high and low temperature regulate protease gene expression through transcriptional induction of a single C14 gene. A hypothesis for the function of C14 thiol protease gene expression in response to heat and cold is discussed.« less

Intracellular studies of the nucleoside reverse transcriptase inhibitor active metabolites: a review.

PubMed

Rodriguez Orengo, J F; Santana, J; Febo, I; Diaz, C; Rodriguez, J L; Garcia, R; Font, E; Rosario, O

2000-03-01

Nucleoside reverse transcriptase inhibitors (NRTIs) plasma concentrations do not correlate with clinical efficacy or toxicity. These agents need to be phosphorylated to become active against HIV-infection. Thus, the characterization of the NRTIs intracellular metabolite pharmacological parameters will provide a better understanding that could lead to the development of more rational dose regimens in the HIV-infected population. Furthermore, intracellular measurements of NRTIs may provide a better marker with respect to clinical efficacy and toxicity than plasma concentrations. Thus, in this article we review the latest information regarding the intracellular pharmacological parameters of zidovudine (ZDV) and lamivudine (3TC) active metabolites in HIV-infected patients including the results from our recent clinical studies. We will start the discussion with ZDV and 3TC clinical efficacy, followed by systemic pharmacokinetics studies. We will then discuss the in vitro and in vivo intracellular studies with particular emphasis in the method development to measure these metabolites and we will conclude with the most current data from our clinical trials.

Marine Invertebrate Metabolites with Anticancer Activities: Solutions to the “Supply Problem”

PubMed Central

Gomes, Nelson G. M.; Dasari, Ramesh; Chandra, Sunena; Kiss, Robert; Kornienko, Alexander

2016-01-01

Marine invertebrates provide a rich source of metabolites with anticancer activities and several marine-derived agents have been approved for the treatment of cancer. However, the limited supply of promising anticancer metabolites from their natural sources is a major hurdle to their preclinical and clinical development. Thus, the lack of a sustainable large-scale supply has been an important challenge facing chemists and biologists involved in marine-based drug discovery. In the current review we describe the main strategies aimed to overcome the supply problem. These include: marine invertebrate aquaculture, invertebrate and symbiont cell culture, culture-independent strategies, total chemical synthesis, semi-synthesis, and a number of hybrid strategies. We provide examples illustrating the application of these strategies for the supply of marine invertebrate-derived anticancer agents. Finally, we encourage the scientific community to develop scalable methods to obtain selected metabolites, which in the authors’ opinion should be pursued due to their most promising anticancer activities. PMID:27213412

The formation of estrogen-like tamoxifen metabolites and their influence on enzyme activity and gene expression of ADME genes.

PubMed

Johänning, Janina; Kröner, Patrick; Thomas, Maria; Zanger, Ulrich M; Nörenberg, Astrid; Eichelbaum, Michel; Schwab, Matthias; Brauch, Hiltrud; Schroth, Werner; Mürdter, Thomas E

2018-03-01

Tamoxifen, a standard therapy for breast cancer, is metabolized to compounds with anti-estrogenic as well as estrogen-like action at the estrogen receptor. Little is known about the formation of estrogen-like metabolites and their biological impact. Thus, we characterized the estrogen-like metabolites tamoxifen bisphenol and metabolite E for their metabolic pathway and their influence on cytochrome P450 activity and ADME gene expression. The formation of tamoxifen bisphenol and metabolite E was studied in human liver microsomes and Supersomes™. Cellular metabolism and impact on CYP enzymes was analyzed in upcyte® hepatocytes. The influence of 5 µM of tamoxifen, anti-estrogenic and estrogen-like metabolites on CYP activity was measured by HPLC MS/MS and on ADME gene expression using RT-PCR analyses. Metabolite E was formed from tamoxifen by CYP2C19, 3A and 1A2 and from desmethyltamoxifen by CYP2D6, 1A2 and 3A. Tamoxifen bisphenol was mainly formed from (E)- and (Z)-metabolite E by CYP2B6 and CYP2C19, respectively. Regarding phase II metabolism, UGT2B7, 1A8 and 1A3 showed highest activity in glucuronidation of tamoxifen bisphenol and metabolite E. Anti-estrogenic metabolites (Z)-4-hydroxytamoxifen, (Z)-endoxifen and (Z)-norendoxifen inhibited the activity of CYP2C enzymes while tamoxifen bisphenol consistently induced CYPs similar to rifampicin and phenobarbital. On the transcript level, highest induction up to 5.6-fold was observed for CYP3A4 by tamoxifen, (Z)-4-hydroxytamoxifen, tamoxifen bisphenol and (E)-metabolite E. Estrogen-like tamoxifen metabolites are formed in CYP-dependent reactions and are further metabolized by glucuronidation. The induction of CYP activity by tamoxifen bisphenol and the inhibition of CYP2C enzymes by anti-estrogenic metabolites may lead to drug-drug-interactions.

The change in serum Thiol/Disulphide homeostasis after transrectal ultrasound guided prostate biopsy

PubMed Central

Tokgöz, Hüsnü; Taş, Selim; Giray, Özlem; Yalçınkaya, Soner; Tokgöz, Özlem; Koca, Cemile; Savaş, Murat; Erel, Özcan

2017-01-01

ABSTRACT Objectives The aim of this prospective clinical study was to investigate variations in a novel oxidative stress marker (thiol/disulphide homeostasis) in men who underwent transrectal ultrasound guided prostate biopsy (TRUSB). Materials and Methods A total of 22 men undergoing TRUSB of the prostate were enrolled in the study. Patients with abnormal digital rectal examination and/or total prostate specific antigen (PSA) over 4ng/mL underwent TRUSB with 12 cores. Serum samples were obtained before and just after the procedure to evaluate the possible changes in thiol/disulphide homeostasis. Mean age, total PSA and free PSA, prostate volume and histopathological data were also recorded. Results Mean age of the study population was 65.05±8.89 years. Significant decreases in native and total thiol levels were documented after the biopsy procedure. However, serum disulphide levels and disulphide/native thiol, disulphide/total thiol and native/total thiol ratios did not significantly change after TRUSB. No correlation was observed between oxidative parameters and total PSA and free PSA levels, prostate volume and histopathology of the prostate. However, mean patient age was significantly correlated with mean native and total thiol levels. Conclusion Significant decreases in serum native and total thiol levels related to the prostate biopsy procedure suggest that TRUSB causes acute oxidative stress in the human body. Since our trial is the first in the current literature to investigate these oxidative stress markers in urology practice, additional studies are warranted. PMID:28128906

The change in serum Thiol/Disulphide homeostasis after transrectal ultrasound guided prostate biopsy.

PubMed

Tokgöz, Hüsnü; Taş, Selim; Giray, Özlem; Yalçınkaya, Soner; Tokgöz, Özlem; Koca, Cemile; Savaş, Murat; Erel, Özcan

2017-01-01

The aim of this prospective clinical study was to investigate variations in a novel oxidative stress marker (thiol/disulphide homeostasis) in men who underwent transrectal ultrasound guided prostate biopsy (TRUSB). A total of 22 men undergoing TRUSB of the prostate were enrolled in the study. Patients with abnormal digital rectal examination and/or total prostate specific antigen (PSA) over 4ng/mL underwent TRUSB with 12 cores. Serum samples were obtained before and just after the procedure to evaluate the possible changes in thiol/disulphide homeostasis. Mean age, total PSA and free PSA, prostate volume and histopathological data were also recorded. Mean age of the study population was 65.05±8.89 years. Significant decreases in native and total thiol levels were documented after the biopsy procedure. However, serum disulphide levels and disulphide/native thiol, disulphide/total thiol and native / total thiol ratios did not significantly change after TRUSB. No correlation was observed between oxidative parameters and total PSA and free PSA levels, prostate volume and histopathology of the prostate. However, mean patient age was significantly correlated with mean native and total thiol levels. Significant decreases in serum native and total thiol levels related to the prostate biopsy procedure suggest that TRUSB causes acute oxidative stress in the human body. Since our trial is the first in the current literature to investigate these oxidative stress markers in urology practice, additional studies are warranted. Copyright® by the International Brazilian Journal of Urology.

Lignin-Based Materials Through Thiol-Maleimide "Click" Polymerization.

PubMed

Buono, Pietro; Duval, Antoine; Averous, Luc; Habibi, Youssef

2017-03-09

In the present report an environmentally friendly approach to transforming renewable feedstocks into value-added materials is proposed. This transformation pathway was conducted under green conditions, without the use of solvents or catalyst. First, controlled modification of lignin, a major biopolymer present in wood and plants, was achieved by esterification with 11-maleimidoundecylenic acid (11-MUA), a derivative from castor oil that contains maleimide groups, following its transformation into 11-maleimidoundecanoyl chloride (11-MUC). Different degrees of substitution were achieved by using various amounts of the 11-MUC, leading to an efficient conversion of lignin hydroxy groups, as demonstrated by 1 H and 31 P NMR analyses. These fully biobased maleimide-lignin derivatives were subjected to an extremely fast (ca. 1 min) thiol-ene "click" polymerization with thiol-containing linkers. Aliphatic and aromatic thiol linkers bearing two to four thiol groups were used to tune the reactivity and crosslink density. The properties of the resulting materials were evaluated by swelling tests and thermal and mechanical analyses, which showed that varying the degree of functionality of the linker and the linker structure allowed accurate tailoring of the thermal and mechanical properties of the final materials, thus providing interesting perspectives for lignin in functional aromatic polymers. © 2017 Wiley-VCH Verlag GmbH & Co. KGaA, Weinheim.

Simulation of interindividual differences in inactivation of reactive para-benzoquinone imine metabolites of diclofenac by glutathione S-transferases in human liver cytosol.

PubMed

den Braver, Michiel W; Zhang, Yongjie; Venkataraman, Harini; Vermeulen, Nico P E; Commandeur, Jan N M

2016-07-25

Diclofenac is a widely prescribed NSAID that causes severe idiosyncratic drug induced liver injury (IDILI) in a small part of the patient population. Formation of protein-reactive metabolites is considered to play a role in the development of diclofenac-induced IDILI. Therefore, a high hepatic activity of enzymes involved in bioactivation of diclofenac is expected to increase the risk for liver injury. However, the extent of covalent protein binding may also be determined by activity of protective enzymes, such as glutathione S-transferases (GSTs). This is supported by an association study in which a correlation was found between NSAID-induced IDILI and the combined null genotypes of GSTM1 and GSTT1. In the present study, the activity of 10 different recombinant human GSTs in inactivation of protein-reactive quinoneimine (QI) metabolites of diclofenac was tested. Both at low and high GSH concentrations, high activities of GSTA1-1, A2-2, A3-3, M1-1, M3-3 and P1-1 in the inactivation of these QIs were found. By using the expression levels of GSTs in livers of 22 donors, a 6-fold variation in GST-dependent inactivation of reactive diclofenac metabolites was predicted. Moreover, it was shown in vitro that GSTs can strongly increase the efficiency of GSH to protect against the alkylation of the model thiol N-acetylcysteine by reactive diclofenac metabolites. The results of this study demonstrate that variability of GST expression may significantly contribute to the inter-individual differences in susceptibility to diclofenac-induced liver injury. In addition, expression levels of GSTs in in vitro models for hepatotoxicity may be important factors determining sensitivity to diclofenac cytotoxicity. Copyright © 2016 Elsevier Ireland Ltd. All rights reserved.

Functional thiols as repair and doping agents of defective MoS2 monolayers

NASA Astrophysics Data System (ADS)

Förster, Anja; Gemming, Sibylle; Seifert, Gotthard

2018-06-01

Recent experimental and theoretical studies indicate that thiols (R-SH) can be used to repair sulfur vacancy defects in MoS2 monolayers (MLs). This density functional theory study investigates how the thiol repair mechanism process can be used to dope MoS2 MLs. Fluorinated thiols as well as amine-containing ones are used to p- and n-dope the MoS2 ML, respectively. It is shown that functional groups are only physisorbed on the repaired MoS2 surface. This explains the reversible doping with fluorinated thiols.

Monohalogenated acetamide-induced cellular stress and genotoxicity are related to electrophilic softness and thiol/thiolate reactivity.

PubMed

Pals, Justin A; Wagner, Elizabeth D; Plewa, Michael J; Xia, Menghang; Attene-Ramos, Matias S

2017-08-01

Haloacetamides (HAMs) are cytotoxic, genotoxic, and mutagenic byproducts of drinking water disinfection. They are soft electrophilic compounds that form covalent bonds with the free thiol/thiolate in cysteine residues through an S N 2 reaction mechanism. Toxicity of the monohalogenated HAMs (iodoacetamide, IAM; bromoacetamide, BAM; or chloroacetamide, CAM) varied depending on the halogen substituent. The aim of this research was to investigate how the halogen atom affects the reactivity and toxicological properties of HAMs, measured as induction of oxidative/electrophilic stress response and genotoxicity. Additionally, we wanted to determine how well in silico estimates of electrophilic softness matched thiol/thiolate reactivity and in vitro toxicological endpoints. Each of the HAMs significantly induced nuclear Rad51 accumulation and ARE signaling activity compared to a negative control. The rank order of effect was IAM>BAM>CAM for Rad51, and BAM≈IAM>CAM for ARE. In general, electrophilic softness and in chemico thiol/thiolate reactivity provided a qualitative indicator of toxicity, as the softer electrophiles IAM and BAM were more thiol/thiolate reactive and were more toxic than CAM. Copyright © 2017. Published by Elsevier B.V.

Determination of acidity and nucleophilicity in thiols by reaction with monobromobimane and fluorescence detection.

PubMed

Sardi, Florencia; Manta, Bruno; Portillo-Ledesma, Stephanie; Knoops, Bernard; Comini, Marcelo A; Ferrer-Sueta, Gerardo

2013-04-01

A method based on the differential reactivity of thiol and thiolate with monobromobimane (mBBr) has been developed to measure nucleophilicity and acidity of protein and low-molecular-weight thiols. Nucleophilicity of the thiolate is measured as the pH-independent second-order rate constant of its reaction with mBBr. The ionization constants of the thiols are obtained through the pH dependence of either second-order rate constant or initial rate of reaction. For readily available thiols, the apparent second-order rate constant is measured at different pHs and then plotted and fitted to an appropriate pH function describing the observed number of ionization equilibria. For less available thiols, such as protein thiols, the initial rate of reaction is determined in a wide range of pHs and fitted to the appropriate pH function. The method presented here shows excellent sensitivity, allowing the use of nanomolar concentrations of reagents. The method is suitable for scaling and high-throughput screening. Example determinations of nucleophilicity and pK(a) are presented for captopril and cysteine as low-molecular-weight thiols and for human peroxiredoxin 5 and Trypanosoma brucei monothiol glutaredoxin 1 as protein thiols. Copyright © 2013 Elsevier Inc. All rights reserved.

The antibacterial activity of syringopicroside, its metabolites and natural analogues from Syringae Folium.

PubMed

Zhou, Zhengyuan; Han, Na; Liu, Zhihui; Song, Zehai; Wu, Peng; Shao, Jingxuan; Zhang, Jia Ming; Yin, Jun

2016-04-01

In the present study, the in vitro antibacterial activity of an effective fraction (ESF) from Syringae Folium (SF) on Methicillin-resistant Staphylococcus aureus (MRSA) was evaluated and then its in vivo activity was evaluated by using the MRSA-infected mouse peritonitis model. The ESF showed a significant in vitro and in vivo activity on decreasing the Minimum Inhibitory Concentrations (MICs) and increasing the survival rate of mouse from 42.8% to 100%. Six iridoid glucosides (IGs) of ESF were characterized by UPLC-TOF-MS method and also isolated by column chromatography. Most of them showed in vitro anti MRSA activity. Syringopicroside (Sy), the major compound of IGs, was found to increase the survival rate from 42.8% to 92.8% of the MRSA-infected mouse, which revealed Sy is also the main active components of ESF. In order to know why the effect of oral administration of SF is better than its injections in clinic and the metabolites of Sy, seven metabolites of Sy were isolated from rat urine and identified on the basis of NMR and MS spectra. Most of metabolites possessed stronger in vitro anti-MRSA activity than that of Sy, which furtherly proved the clinical result. Copyright © 2016 Elsevier B.V. All rights reserved.

Visible light-initiated interfacial thiol-norbornene photopolymerization for forming islet surface conformal coating

PubMed Central

Shih, Han; Mirmira, Raghavendra G.; Lin, Chien-Chi

2015-01-01

A cytocompatible visible light-mediated interfacial thiol-norbornene photopolymerization scheme was developed for creating hydrogel conformal coating on pancreatic islets. The step-growth thiol-norbornene reaction affords high consistency and tunability in gel coating thickness. Furthermore, isolated islets coated with thiol-norbornene gel maintained their viability and function in vitro. PMID:26509035

Synthesis of Single-walled Carbon Nanotubes Coated with Thiol-reactive Gel via Emulsion Polymerization.

PubMed

Nagai, Yukiko; Tsutsumi, Yusuke; Nakashima, Naotoshi; Fujigaya, Tsuyohiko

2018-06-15

Single-walled carbon nanotubes (SWNTs) have unique near-infrared absorption and photoemission properties that are attractive for in vivo biological applications such as photothermal cancer treatment and bioimaging. Therefore, a smart functionalization strategy for SWNTs to create biocompatible surfaces and introduce various ligands to target active cancer cells without losing the unique optical properties of the SWNTs is strongly desired. This paper reports the de-sign and synthesis of a SWNT/gel hybrid containing maleimide groups, which react with various thiol compounds through Michael addition reactions. In this hybrid, the method called carbon nanotube micelle polymerization was used to non-covalently modify the surface of SWNTs with a cross-linked polymer gel layer. This method can form an extremely stable gel layer on SWNTs; such stability is essential for in vivo biological applications. The monomer used to form the gel layer contained a maleimide group, which was protected with furan in endo-form. The resulting hybrid was treated in water to induce deprotection via retro Diels-Alder reaction and then functionalized with thiol com-pounds through Michael addition. The functionalization of the hybrid was explored using a thiol-containing fluores-cent dye as a model thiol and the formation of the SWNT-dye conjugate was confirmed by energy transfer from the dye to SWNTs. Our strategy offers a promising SWNT-based platform for biological functionalization for cancer targeting, imaging, and treatment.

Induction Curing of Thiol-acrylate and Thiolene Composite Systems

PubMed Central

Ye, Sheng; Cramer, Neil B.; Stevens, Blake E.; Sani, Robert L.; Bowman, Christopher N.

2011-01-01

Induction curing is demonstrated as a novel type of in situ radiation curing that maintains most of the advantages of photocuring while eliminating the restriction of light accessibility. Induction curing is utilized to polymerize opaque composites comprised of thiol-acrylate and thiol-ene resins, nanoscale magnetic particles, and carbon nanotubes. Nanoscale magnetic particles are dispersed in the resin and upon exposure to the magnetic field, these particles lead to induction heating that rapidly initiates the polymerization. Heat transfer profiles and reaction kinetics of the samples are modeled during the reactions with varying induction heater power, species concentration, species type and sample thickness, and the model is compared with the experimental results. Thiol-ene polymerizations achieved full conversion between 1.5 minutes and 1 hour, depending on the field intensity and the composition, with the maximum reaction temperature decreasing from 146 – 87 °C when the induction heater power was decreased from 8 – 3 kW. The polymerization reactions of the thiol-acrylate system were demonstrated to achieve full conversion between 0.6 and 30 minutes with maximum temperatures from 139 to 86 °C. The experimental behavior was characterized and the temperature profile modeled for the thiol-acrylate composite comprised of sub100nm nickel particles and induction heater power in the range of 32 to 20 kW. A 9°C average deviation was observed between the modeling and experimental results for the maximum temperature rise. The model also was utilized to predict reaction temperatures and kinetics for systems with varying thermal initiator concentration, initiator half-life, monomer molecular weight and temperature gradients in samples with varying thickness, thereby demonstrating that induction curing represents a designable and tunable polymerization method. Finally, induction curing was utilized to cure thiol-acrylate systems containing carbon nanotubes where 1 wt

“Turn-on” fluorescence probe integrated polymer nanoparticles for sensing biological thiol molecules

NASA Astrophysics Data System (ADS)

Ang, Chung Yen; Tan, Si Yu; Lu, Yunpeng; Bai, Linyi; Li, Menghuan; Li, Peizhou; Zhang, Quan; Selvan, Subramanian Tamil; Zhao, Yanli

2014-11-01

A ``turn-on'' thiol-responsive fluorescence probe was synthesized and integrated into polymeric nanoparticles for sensing intracellular thiols. There is a photo-induced electron transfer process in the off state of the probe, and this process is terminated upon the reaction with thiol compounds. Configuration interaction singles (CIS) calculation was performed to confirm the mechanism of this process. A series of sensing studies were carried out, showing that the probe-integrated nanoparticles were highly selective towards biological thiol compounds over non-thiolated amino acids. Kinetic studies were also performed to investigate the relative reaction rate between the probe and the thiolated amino acids. Subsequently, the Gibbs free energy of the reactions was explored by means of the electrochemical method. Finally, the detection system was employed for sensing intracellular thiols in cancer cells, and the sensing selectivity could be further enhanced with the use of a cancer cell-targeting ligand in the nanoparticles. This development paves a path for the sensing and detection of biological thiols, serving as a potential diagnostic tool in the future.

Pharmacologically active plant metabolites as survival strategy products.

PubMed

Attardo, C; Sartori, F

2003-01-01

The fact that plant organisms produce chemical substances that are able to positively or negatively interfere with the processes which regulate human life has been common knowledge since ancient times. One of the numerous possible examples in the infusion of Conium maculatum, better known as Hemlock, a plant belonging to the family umbelliferae, used by the ancient Egyptians to cure skin diseases. The current official pharmacopoeia includes various chemical substances produced by secondary plant metabolisms. For example, the immunosuppressive drugs used to prevent organ transplant rejection and the majority of antibiotics are metabolites produced by fungal organisms, pilocarpin, digitalis, strophantus, salicylic acid and curare are examples of plant organism metabolites. For this reason, there has been an increase in research into plants, based on information on their medicinal use in the areas where they grow. The study of plants in relation to local culture and traditions is known as "ethnobotany". Careful study of the behaviour of sick animals has also led to the discovery of medicinal plants. The study of this subject is known as "zoopharmacognosy". The aim of this article is to discuss the fact that "ad hoc" production of such chemical substances, defined as "secondary metabolites", is one of the modes in which plant organisms respond to unfavourable environmental stimuli, such as an attack by predatory phytophagous animals or an excessive number of plant individuals, even of the same species, in a terrain. In the latter case, the plant organisms produce toxic substances, called "allelopathic" which limit the growth of other individuals. "Secondary metabolites" are produced by metabolic systems that are shunts of the primary systems which, when required, may be activated from the beginning, or increased to the detriment of others. The study of the manner in which such substances are produced is the subject of a new branch of learning called "ecological

Accessibility of hepatocyte protein thiols to monobromobimane.

PubMed

Weis, M; Cotgreave, I C; Moore, G A; Norbeck, K; Moldéus, P

1993-03-10

The amino-acid residue specificity of monobromobimane (mBBr) and its accessibility to cellular protein cysteine residues were investigated. mBBr reacted selectively with the sulfhydryl group of both the free amino acid cysteine and bovine serum albumin. Incubation of isolated hepatocytes with mBBr resulted in a concentration-dependent formation of protein-bound mBBr fluorescence in the cytosolic, mitochondrial and microsomal fractions, which was not fully saturated with up to 16 mM mBBr. SDS-PAGE resolution of the proteins revealed that the major portion of increased protein-bound mBBr fluorescence that occurred at high mBBr concentrations was due to covalent binding to proteins. A minor portion (10-16% in the microsomal fraction) of protein-bound mBBr fluorescence was removed by SDS-PAGE and is therefore concluded to be due to physical entrapment of fluorescent mBBr reaction products. The accessibility of mBBr, assayed as the degree of depletion of total protein cysteine residues, was similar to N-ethylmaleimide (NEM) in isolated microsomes. By contrast, in the cytosol a markedly lower amount of protein cysteine residues were labelled by mBBr as compared to NEM. In both organelle fractions p-BQ was the most efficient thiol-depleting reagent. It is concluded that mBBr is a suitable reagent for the analysis of the cellular protein thiol status and of its xenobiotic-induced alterations when used at high concentrations; however, it should be considered that, (i) the relative accessibility of mBBr and a particular xenobiotic to cellular protein thiol residues may be different, and (ii) physically entrapped fluorescent reaction products of mBBr should be removed when quantitating protein thiol levels.

Thiol-Reactive Star Polymers Display Enhanced Association with Distinct Human Blood Components.

PubMed

Glass, Joshua J; Li, Yang; De Rose, Robert; Johnston, Angus P R; Czuba, Ewa I; Khor, Song Yang; Quinn, John F; Whittaker, Michael R; Davis, Thomas P; Kent, Stephen J

2017-04-12

Directing nanoparticles to specific cell types using nonantibody-based methods is of increasing interest. Thiol-reactive nanoparticles can enhance the efficiency of cargo delivery into specific cells through interactions with cell-surface proteins. However, studies to date using this technique have been largely limited to immortalized cell lines or rodents, and the utility of this technology on primary human cells is unknown. Herein, we used RAFT polymerization to prepare pyridyl disulfide (PDS)-functionalized star polymers with a methoxy-poly(ethylene glycol) brush corona and a fluorescently labeled cross-linked core using an arm-first method. PDS star polymers were examined for their interaction with primary human blood components: six separate white blood cell subsets, as well as red blood cells and platelets. Compared with control star polymers, thiol-reactive nanoparticles displayed enhanced association with white blood cells at 37 °C, particularly the phagocytic monocyte, granulocyte, and dendritic cell subsets. Platelets associated with more PDS than control nanoparticles at both 37 °C and on ice, but they were not activated in the duration examined. Association with red blood cells was minor but still enhanced with PDS nanoparticles. Thiol-reactive nanoparticles represent a useful strategy to target primary human immune cell subsets for improved nanoparticle delivery.

Urinary Estrogen Metabolites, Active and Sedentary Behaviors, and Breast Cancer Risk

Cancer.gov

A cross-sectional study of approximately 600 postmenopausal controls in the Breast Cancer Case-Control Study in Poland to assess urinary estrogen metabolites in relation to accelerometer-based measures of active and sedentary behaviors

Full inhibition of enzymatic browning in the presence of thiol-functionalised silica nanomaterial.

PubMed

Muñoz-Pina, Sara; Ros-Lis, José V; Argüelles, Ángel; Coll, Carmen; Martínez-Máñez, Ramón; Andrés, Ana

2018-02-15

Darkening processed fruits and vegetables is caused mainly by enzymatic browning through polyphenol oxidase (PPO) action. Accordingly, we explored the potential of four silica-based materials (MCM-41 nanometric size, MCM-41 micrometric size, UVM-7 and aerosil), non-functionalised and functionalised with thiol groups, to inhibit PPO activity in the model system and apple juice. All materials showed relevant performance when immobilising and inhibiting PPO in model systems, and support topology is a main factor for enzyme immobilisation and inhibition. Thiol-containing silica UVM7-SH showed the greatest inactivation, and similar browning values to those obtained by acidification. The enzyme's kinetic parameters in the presence of UVM-7-SH suggested non-competitive inhibition, which indicated that the material interacted with the enzyme, but beyond the active centre. In real systems, UVM-7-SH completely inhibited enzymatic browning in apple juice (cv. Granny Smith and cv. Golden Delicious) up to 9days after 5min of contact. Copyright © 2017 Elsevier Ltd. All rights reserved.

Changes in Thiol-Disulfide Homeostasis of the Body to Surgical Trauma in Laparoscopic Cholecystectomy Patients.

PubMed

Polat, Murat; Ozcan, Onder; Sahan, Leyla; Üstündag-Budak, Yasemin; Alisik, Murat; Yilmaz, Nigar; Erel, Özcan

2016-12-01

We aimed to investigate the short-term effect of laparoscopic surgery on serum thiol-disulfide homeostasis levels as a marker of oxidant stress of surgical trauma in elective laparoscopic cholecystectomy patients. Venous blood samples were collected, and levels of native thiols, total thiols, and disulfides were determined with a novel automated assay. Total antioxidant capacity (measured as the ferric-reducing ability of plasma) and serum ischemia modified albumin, expressed as absorbance units assayed by the albumin cobalt binding test, were determined. The major findings of the present study were that native thiol (283 ± 45 versus 241 ± 61 μmol/L), total thiol (313 ± 49 versus 263 ± 67 μmol/L), and disulfide (14.9 ± 4.6 versus 11.0 ± 6.1 μmol/L) levels were decreased significantly during operation and although they increased, they did not return to preoperation levels 24 hours after laparoscopic surgery compared to the levels at baseline. Disulfide/native thiol and disulfide/total thiol levels did not change during laparoscopic surgery. The decrease in plasma level of native and total thiol groups suggests impairment of the antioxidant capacity of plasma; however, the delicate balance between the different redox forms of thiols was maintained during surgery.

Green Tea Catechin Metabolites Exert Immunoregulatory Effects on CD4(+) T Cell and Natural Killer Cell Activities.

PubMed

Kim, Yoon Hee; Won, Yeong-Seon; Yang, Xue; Kumazoe, Motofumi; Yamashita, Shuya; Hara, Aya; Takagaki, Akiko; Goto, Keiichi; Nanjo, Fumio; Tachibana, Hirofumi

2016-05-11

Tea catechins, such as (-)-epigallocatechin-3-O-gallate (EGCG), have been shown to effectively enhance immune activity and prevent cancer, although the underlying mechanism is unclear. Green tea catechins are instead converted to catechin metabolites in the intestine. Here, we show that these green tea catechin metabolites enhance CD4(+) T cell activity as well as natural killer (NK) cell activity. Our data suggest that the absence of a 4'-hydroxyl on this phenyl group (B ring) is important for the effect on immune activity. In particular, 5-(3',5'-dihydroxyphenyl)-γ-valerolactone (EGC-M5), a major metabolite of EGCG, not only increased the activity of CD4(+) T cells but also enhanced the cytotoxic activity of NK cells in vivo. These data suggest that EGC-M5 might show immunostimulatory activity.

Chemopreventive Activities of Sulforaphane and Its Metabolites in Human Hepatoma HepG2 Cells.

PubMed

Liu, Peng; Wang, Wei; Zhou, Zhigang; Smith, Andrew J O; Bowater, Richard P; Wormstone, Ian Michael; Chen, Yuqiong; Bao, Yongping

2018-05-09

Sulforaphane (SFN) exhibits chemopreventive effects through various mechanisms. However, few studies have focused on the bioactivities of its metabolites. Here, three metabolites derived from SFN were studied, known as sulforaphane glutathione, sulforaphane cysteine and sulforaphane- N -acetylcysteine. Their effects on cell viability, DNA damage, tumorigenicity, cell migration and adhesion were measured in human hepatoma HepG2 cells, and their anti-angiogenetic effects were determined in a 3D co-culture model of human umbilical vein endothelial cells (HUVECs) and pericytes. Results indicated that these metabolites at high doses decreased cancer cell viability, induced DNA damage and inhibited motility, and impaired endothelial cell migration and tube formation. Additionally, pre-treatment with low doses of SFN metabolites protected against H₂O₂ challenge. The activation of the nuclear factor E2-related factor 2 (Nrf2)-antioxidant response element (ARE) pathway and the induction of intracellular glutathione (GSH) played an important role in the cytoprotective effects of SFN metabolites. In conclusion, SFN metabolites exhibited similar cytotoxic and cytoprotective effects to SFN, which proves the necessity to study the mechanisms of action of not only SFN but also of its metabolites. Based on the different tissue distribution profiles of these metabolites, the most relevant chemical forms can be selected for targeted chemoprevention.

Promoting Thiol Expression Increases The Durability of Antitumor T cell Functions

PubMed Central

Scurti, Gina; Thyagarajan, Krishnamurthy; Kaur, Navtej; Husain, Shahid; Fang, Quan; Naga, Osama S.; Simms, Patricia; Beeson, Gyda; Voelkel-Johnson, Christina; Garrett-Mayer, Elizabeth; Beeson, Craig C.; Nishimura, Michael I.; Mehrotra, Shikhar

2014-01-01

Ex vivo-expanded CD8+ T cells used for adoptive immunotherapy generally acquire an effector memory-like phenotype (TEM cells). With regard to therapeutic applications, two undesired features of this phenotype in vivo are limited persistence and reduced anti-tumor efficacy, relative to CD8+ T cells with a central memory-like phenotype (TCM cells). Further, there is incomplete knowledge about all the differences between TEM and TCM cells that may influence tumor treatment outcomes. Given that TCM cells survive relatively longer in oxidative tumor microenvironments, we investigated the hypothesis that TCM possess relatively greater anti-oxidative capacity than TEM cells. Here we report that TCM cells exhibit a relative increase compared to TEM cells in expression of cell surface thiols, a key target of cellular redox controls, along with other antioxidant molecules. Increased expression of redox regulators in TCM cells inversely correlated with the generation of reactive oxygen and nitrogen species, proliferative capacity and glycolytic enzyme levels. Notably, TCR-transduced T cells pretreated with thiol donors, such as N-acetyl cysteine or rapamycin, up-regulated thiol levels and antioxidant genes. A comparison of anti-tumor CD8+ T cell populations on the basis of surface thiol expression showed that thiol-high cells persisted longer in vivo and exerted superior tumor control. Our results suggest that higher levels of reduced cell surface thiols are a key characteristic of T cells that can control tumor growth, and that profiling this biomarker may have benefits to T cell adoptive immunotherapy protocols. PMID:25164014

Baicalin, a metabolite of baicalein with antiviral activity against dengue virus

PubMed Central

Moghaddam, Ehsan; Teoh, Boon-Teong; Sam, Sing-Sin; Lani, Rafidah; Hassandarvish, Pouya; Chik, Zamri; Yueh, Andrew; Abubakar, Sazaly; Zandi, Keivan

2014-01-01

Baicalin, a flavonoid derived from Scutellaria baicalensis, is the main metabolite of baicalein released following administration in different animal models and human. We previously reported the antiviral activity of baicalein against dengue virus (DENV). Here, we examined the anti-DENV properties of baicalin in vitro, and described the inhibitory potentials of baicalin at different steps of DENV-2 (NGC strain) replication. Our in vitro antiviral experiments showed that baicalin inhibited virus replication at IC50 = 13.5 ± 0.08 μg/ml with SI = 21.5 following virus internalization by Vero cells. Baicalin exhibited virucidal activity against DENV-2 extracellular particles at IC50 = 8.74 ± 0.08 μg/ml and showed anti-adsorption effect with IC50 = 18.07 ± 0.2 μg/ml. Our findings showed that baicalin as the main metabolite of baicalein exerting in vitro anti-DENV activity. Further investigations on baicalein and baicalin to deduce its antiviral therapeutic effects are warranted. PMID:24965553

Oxidative stress and decreased thiol level in patients with migraine: cross-sectional study.

PubMed

Eren, Yasemin; Dirik, Ebru; Neşelioğlu, Salim; Erel, Özcan

2015-12-01

Although migraine is a neurological disorder known since long, its physiopathology remains unclear. Recent studies suggest that migraine is associated with oxidative stress; however, they report divergent results. The aim of the present study was to evaluate total antioxidant status (TAS), total oxidant status (TOS), oxidative stress index (OSI), and serum thiol level in migraine patients with or without aura. The study group consisted of 141 migraine patients. The control group included 70 healthy subjects. TAS, TOS, OSI were evaluated using a method developed by Erel. Serum thiol level was measured using the Hu method. No difference was found in TAS, TOS, OSI between the patients and controls. The level of thiol was significantly lower in patients than in controls. Negative correlations were detected between thiol level and Migraine Disability Assessment score in patients. Although TAS, TOS, and OSI were similar to those of the control group, serum thiol level, an important marker of antioxidant capacity, was significantly lower in migraines compared with controls, and caused more serious disability. Novel treatment approaches may be developed based on these data, and compounds containing thiol, such as alpha lipoic acid and N-acetyl cysteine, may be used in prophylaxis.

Microbiome-Derived Tryptophan Metabolites and Their Aryl Hydrocarbon Receptor-Dependent Agonist and Antagonist Activities

PubMed Central

Jin, Un-Ho; Lee, Syng-Ook; Sridharan, Gautham; Lee, Kyongbum; Davidson, Laurie A.; Jayaraman, Arul; Chapkin, Robert S.; Alaniz, Robert

2014-01-01

The tryptophan metabolites indole, indole-3-acetate, and tryptamine were identified in mouse cecal extracts and fecal pellets by mass spectrometry. The aryl hydrocarbon receptor (AHR) agonist and antagonist activities of these microbiota-derived compounds were investigated in CaCo-2 intestinal cells as a model for understanding their interactions with colonic tissue, which is highly aryl hydrocarbon (Ah)–responsive. Activation of Ah-responsive genes demonstrated that tryptamine and indole 3-acetate were AHR agonists, whereas indole was an AHR antagonist that inhibited TCDD (2,3,7,8-tetrachlorodibenzo-p-dioxin)–induced CYP1A1 expression. In contrast, the tryptophan metabolites exhibited minimal anti-inflammatory activities, whereas TCDD decreased phorbol ester-induced CXCR4 [chemokine (C-X-C motif) receptor 4] gene expression, and this response was AHR dependent. These results demonstrate that the tryptophan metabolites indole, tryptamine, and indole-3-acetate modulate AHR-mediated responses in CaCo-2 cells, and concentrations of indole that exhibit AHR antagonist activity (100–250 μM) are detected in the intestinal microbiome. PMID:24563545

[Optimising azathioprine treatment: determination of thiopurine methyltransferase activity and thiopurine metabolites].

PubMed

Alvarez Beltran, M; Infante Pina, D; Tormo Carnicé, R; Segarra Cantón, O; Redecillas Ferreiro, S

2009-02-01

Individualised doses of azathioprine (AZA) may be prescribed by monitoring the levels of the enzyme thiopurine methyltransferase (TPMT). The measurements of thiopurine metabolites of AZA, 6-thioguanine (6-TGN) and 6-methylmercaptopurine (6-MMP), have also been reported as new markers of AZA activity. To describe TPMT phenotype in our population and to establish a relationship between thiopurine metabolites,and therapeutic activity and adverse effects. Data on TPMT were retrospectively collected from 107 patients, and 6-TGN and 6-MMP levels in 18 patients currently on treatment with AZA (Crohn's disease 5, ulcerative colitis 5, autoimmune hepatitis 5). Mean value of TPMT was 20.19U/ml. None of the patients had a TPMT activity<5U/ml. Of the 18 patients on treatment, 13 showed sub-therapeutic levels of 6-TGN (<235pmol/8x10(8) red blood cells). Clinical remission was maintained in 45% of patients. Mean levels of 6-TGN in patients with clinical remission were 259pmol/8x10(8) red blood cells versus 209pmol/8x10(8) red blood cells in non-responders (p=0.37). There was an inverse relationship (r=-0.28) between TPMT and 6-TGN levels. Toxic effects occurred in 6 of 18 patients, with leukopenia in 5 and hyperamylasemia in 1. Determination of TPMT and monitoring of thiopurine metabolites allows AZA treatment to be optimised, although further studies are necessary to establish therapeutic effectiveness and toxicity ranges.

A chromenoquinoline-based fluorescent off-on thiol probe for bioimaging.

PubMed

Kand, Dnyaneshwar; Kalle, Arunasree Marasanapalli; Varma, Sreejith Jayasree; Talukdar, Pinaki

2012-03-11

A new chromenoquinoline-based fluorescent off-on thiol probe 2 is reported. In aqueous buffer solutions at physiological pH, the probe exhibited 223-fold enhancement in fluorescence intensity by a Michael addition of cysteine to the maleimide appended to a chromenoquinoline. Cell permeability and live cell imaging of thiols are also demonstrated. This journal is © The Royal Society of Chemistry 2012

Selenocysteine in thiol/disulfide-like exchange reactions.

PubMed

Hondal, Robert J; Marino, Stefano M; Gladyshev, Vadim N

2013-05-01

Among trace elements used as cofactors in enzymes, selenium is unique in that it is incorporated into proteins co-translationally in the form of an amino acid, selenocysteine (Sec). Sec differs from cysteine (Cys) by only one atom (selenium versus sulfur), yet this switch dramatically influences important aspects of enzyme reactivity. The main focus of this review is an updated and critical discussion on how Sec might be used to accelerate thiol/disulfide-like exchange reactions in natural selenoenzymes, compared with their Cys-containing homologs. We discuss in detail three major aspects associated with thiol/disulfide exchange reactions: (i) nucleophilicity of the attacking thiolate (or selenolate); (ii) electrophilicity of the center sulfur (or selenium) atom; and (iii) stability of the leaving group (sulfur or selenium). In all these cases, we analyze the benefits that selenium might provide in these types of reactions. It is the biological thiol oxidoreductase-like function that benefits from the use of Sec, since Sec functions to chemically accelerate the rate of these reactions. We review various hypotheses that could help explain why Sec is used in enzymes, particularly with regard to competitive chemical advantages provided by the presence of the selenium atom in enzymes. Ultimately, these chemical advantages must be connected to biological functions of Sec.

Influence of reagents reacting with metal, thiol and amino sites of catalytic activity and l-phenylalanine inhibition of rat intestinal alkaline phosphatase

PubMed Central

Fishman, William H.; Ghosh, Nimai K.

1967-01-01

1. Studies on the inactivation of rat intestinal alkaline phosphatase by several metal-binding agents, namely EDTA, 8-hydroxyquinoline, pyridine-2,6-dicarboxylic acid, αα′-bipyridyl, o-phenanthroline and sodium cyanide, indicated the functional role of a metal, probably zinc, in the catalysis. The metal ligands lowered stereospecific uncompetitive inhibition of the enzyme by l-phenylalanine by an extent that paralleled the decline in enzyme activity. 2. The thiol reagents p-hydroxymercuribenzoate, iodoacetamide and iodine inactivated rat intestinal phosphatase. The enzyme could be protected from inactivation by either cysteine or substrate. The l-phenylalanine inhibition remained unchanged only in the presence of moderately inactivating concentrations of the thiol reagents. 3. Inactivation of the enzyme by the amino-group-blocking reagent, O-methylisourea, provided ample evidence for the participation in the catalysis of the ∈-amino group of lysine. At the same time, l-phenylalanine inhibition remained unaltered even when the enzyme was strongly inactivated. This ∈-amino-group-blocked enzyme exhibited no change in migration in starch gel, in contrast with enzyme treated with acetic anhydride, formaldehyde or succinic anhydride. The Michaelis constant of the enzyme was enhanced by such modifications, but the optimum pH remained the same. 4. d-Phenylalanine acted as a competitive or `co-operative' activator for intestinal alkaline phosphatase after it had been modified by acetylation. PMID:16742542

Identification and Characterization of CINPA1 Metabolites Facilitates Structure-Activity Studies of the Constitutive Androstane Receptor

PubMed Central

Cherian, Milu T.; Yang, Lei; Chai, Sergio C.; Lin, Wenwei

2016-01-01

The constitutive androstane receptor (CAR) regulates the expression of genes involved in drug metabolism and other processes. A specific inhibitor of CAR is critical for modulating constitutive CAR activity. We recently described a specific small-molecule inhibitor of CAR, CINPA1 (ethyl (5-(diethylglycyl)-10,11-dihydro-5H-dibenzo[b,f]azepin-3-yl)carbamate), which is capable of reducing CAR-mediated transcription by changing the coregulator recruitment pattern and reducing CAR occupancy at the promoter regions of its target genes. In this study, we showed that CINPA1 is converted to two main metabolites in human liver microsomes. By using cell-based reporter gene and biochemical coregulator recruitment assays, we showed that although metabolite 1 was very weak in inhibiting CAR function and disrupting CAR-coactivator interaction, metabolite 2 was inactive in this regard. Docking studies using the CAR ligand-binding domain structure showed that although CINPA1 and metabolite 1 can bind in the CAR ligand-binding pocket, metabolite 2 may be incapable of the molecular interactions required for binding. These results indicate that the metabolites of CINPA1 may not interfere with the action of CINPA1. We also used in vitro enzyme assays to identify the cytochrome P450 enzymes responsible for metabolizing CINPA1 in human liver microsomes and showed that CINPA1 was first converted to metabolite 1 by CYP3A4 and then further metabolized by CYP2D6 to metabolite 2. Identification and characterization of the metabolites of CINPA1 enabled structure-activity relationship studies of this family of small molecules and provided information to guide in vivo pharmacological studies. PMID:27519550

Thiol reduction of arsenite and selenite: DFT modeling of the pathways to an as-se bond.

PubMed

Harper, Lenora K; Antony, Sonia; Bayse, Craig A

2014-12-15

The reactivity of arsenite and selenite with biological thiols plays an important role in the toxicity of these elements. However, toxic effects are eliminated when the species are coadministered, due to the antagonistic relationship between selenium and arsenic. The reduction of arsenous acid and selenious acid by thiol and the formation of an As-Se species have been modeled using density functional theory (DFT) and solvent-assisted proton exchange (SAPE), a microsolvation technique that uses a network of water molecules to mimic the participation of bulk solvent in proton transfer processes. Activation barriers and relative energies were calculated for the stepwise thiol reduction of arsenite to form As(SR)3 and selenious acid to first form a selenotrisulfide (Se(SR)2) and then H2Se. Several pathways were explored for the formation of an As-Se bond: the nucleophilic attack of selenide or selenopersulfide on As(OH)3, (RS)As(OH)2, and (RS)2AsOH to form (RS)2AsSeH. On the basis of the lower activation barrier and bioavailability of (RS)2AsOH, the reaction of H2Se with (RS)2AsOH is deemed the most favorable, consistent with previous experimental studies.

Degraded protein adducts of cis-2-butene-1,4-dial are urinary and hepatocyte metabolites of furan.

PubMed

Lu, Ding; Sullivan, Mathilde M; Phillips, Martin B; Peterson, Lisa A

2009-06-01

Furan is a liver toxicant and carcinogen in rodents. On the basis of these observations and the large potential for human exposure, furan has been classified as a possible human carcinogen. The mechanism of tumor induction by furan is unknown. However, the toxicity requires cytochrome P450-catalyzed oxidation of furan. The product of this oxidation, cis-2-butene-1,4-dial (BDA), reacts readily with glutathione, amino acids, and DNA and is a bacterial mutagen in Ames assay strain TA104. Characterization of the urinary metabolites of furan is expected to provide information regarding the structure(s) of the reactive metabolite(s). Recently, several urinary metabolites have been identified. We reported the presence of a monoglutathione-BDA reaction product, N-[4-carboxy-4-(3-mercapto-1H-pyrrol-1-yl)-1-oxobutyl]-l-cysteinylglycine cyclic sulfide. Three additional urinary metabolites of furan were also characterized as follows: R-2-acetylamino-6-(2,5-dihydro-2-oxo-1H-pyrrol-1-yl)-1-hexanoic acid, N-acetyl-S-[1-(5-acetylamino-5-carboxypentyl)-1H-pyrrol-3-yl]-l-cysteine, and its sulfoxide. It was postulated that these three metabolites are derived from degraded protein adducts. However, the possibility that these metabolites result from the reaction of BDA with free lysine and/or cysteine was not ruled out. In this latter case, one might predict that the reaction of thiol-BDA with free lysine would not occur exclusively on the epsilon-amino group. Reaction of BDA with N-acetylcysteine or GSH in the presence of lysine indicated that both the alpha- and the epsilon-amino groups of lysine can be modified by thiol-BDA. The N-acetylcysteine-BDA-N-acetyllysine urinary metabolites were solely linked through the epsilon-amino group of lysine. A GSH-BDA-lysine cross-link was a significant hepatocyte metabolite of furan. In this case, the major product resulted from reaction with the epsilon-amino group of lysine; however, small amounts of the alpha-amino reaction product were also

Degraded protein adducts of cis-2-butene-1,4-dial are urinary and hepatocyte metabolites of furan

PubMed Central

Lu, Ding; Sullivan, Mathilde M.; Phillips, Martin B.; Peterson, Lisa A.

2009-01-01

Furan is a liver toxicant and carcinogen in rodents. Based on these observations and the large potential for human exposure, furan has been classified as a possible human carcinogen. The mechanism of tumor induction by furan is unknown. However, the toxicity requires cytochrome P450 catalyzed oxidation of furan. The product of this oxidation, cis-2-butene-1,4-dial (BDA), reacts readily with glutathione, amino acids and DNA and is a bacterial mutagen in Ames assay strain TA104. Characterization of the urinary metabolites of furan is expected to provide information regarding the structure(s) of the reactive metabolite(s). Recently, several urinary metabolites have been identified. We reported the presence of a mono-glutathione-BDA reaction product, N-[4-carboxy-4-(3-mercapto-1H-pyrrol-1-yl)-1-oxobutyl]-L-cysteinylglycine cyclic sulfide. Three additional urinary metabolites of furan were also characterized: R-2-acetylamino-6-(2,5-dihydro-2-oxo-1H-pyrrol-1-yl)-1-hexanoic acid, N-acetyl-S-[1-(5-acetylamino-5-carboxypentyl)-1H-pyrrol-3-yl]-L-cysteine and its sulfoxide. It was postulated that these three metabolites are derived from degraded protein adducts. However, the possibility that these metabolites result from reaction of BDA with free lysine and/or cysteine was not ruled out. In this latter case, one might predict that the reaction of thiol-BDA with free lysine would not occur exclusively on the ε-amino group. Reaction of BDA with N-acetylcysteine or GSH in the presence of lysine indicated that both the α- and ε-amino groups of lysine can be modified by thiol-BDA. The N-acetylcysteine-BDA-N-acetyllysine urinary metabolites were solely linked through the ε-amino group of lysine. A GSH-BDA-lysine crosslink was a significant hepatocyte metabolite of furan. In this case, the major product resulted from reaction with the ε-amino group of lysine, however, small amounts of the α-amino reaction product were also observed. Western analysis of liver and hepatocyte

Activation of the reticulothalamic cholinergic pathway by the major metabolites of aniracetam.

PubMed

Nakamura, K; Shirane, M

1999-09-10

The aim of the study was to further investigate the effects of aniracetam, a cognition enhancer, and its metabolites on the brain cholinergic system. We measured choline acetyltransferase activity and acetylcholine release using in vivo brain microdialysis in stroke-prone spontaneously hypertensive rats (SHRSP). The enzyme activity in the pons-midbrain and hippocampus, and basal acetylcholine release in the nucleus reticularis thalami were lower in SHRSP than in age-matched Wistar Kyoto rats, indicating central cholinergic deficits in SHRSP. Repeated treatment of aniracetam (50 mg/kg p.o. x 11 for 6 days) preferentially increased the enzyme activity in the thalamus, whereas decreased it in the striatum. Among the metabolites of aniracetam, local perfusion of N-anisoyl-gamma-aminobutyric acid (GABA, 0.1 and/or 1 microM) and p-anisic acid (1 microM) into the nucleus reticularis thalami, dorsal hippocampus and prefrontal cortex of SHRSP produced a significant but delayed increase of acetylcholine release. We failed, however, to find any effect of aniracetam itself. A direct injection of N-anisoyl-GABA (1 nmol) into the pedunculopontine tegmental nucleus of SHRSP enhanced the release in the nucleus reticularis thalami. Thus, these data prove that aniracetam can facilitate central cholinergic neurotransmission via both metabolites. Based on its pharmacokinetic profile, N-anisoyl-GABA may contribute to the clinical effects of aniracetam, mainly by acting on the reticulothalamic cholinergic pathway.

Amplified and in situ detection of redox-active metabolite using a biobased redox capacitor.

PubMed

Kim, Eunkyoung; Gordonov, Tanya; Bentley, William E; Payne, Gregory F

2013-02-19

Redox cycling provides a mechanism to amplify electrochemical signals for analyte detection. Previous studies have shown that diverse mediators/shuttles can engage in redox-cycling reactions with a biobased redox capacitor that is fabricated by grafting redox-active catechols onto a chitosan film. Here, we report that redox cycling with this catechol-chitosan redox capacitor can amplify electrochemical signals for detecting a redox-active bacterial metabolite. Specifically, we studied the redox-active bacterial metabolite pyocyanin that is reported to be a virulence factor and signaling molecule for the opportunistic pathogen P. aeruginosa. We demonstrate that redox cycling can amplify outputs from various electrochemical methods (cyclic voltammetry, chronocoulometry, and differential pulse voltammetry) and can lower the detection limit of pyocyanin to 50 nM. Further, the compatibility of this biobased redox capacitor allows the in situ monitoring of the production of redox-active metabolites (e.g., pyocyanin) during the course of P. aeruginosa cultivation. We anticipate that the amplified output of redox-active virulence factors should permit an earlier detection of life-threatening infections by the opportunistic pathogen P. aeruginosa while the "bio-compatibility" of this measurement approach should facilitate in situ study of the spatiotemporal dynamics of bacterial redox signaling.

Crystal structures of potent thiol-based inhibitors bound to carboxypeptidase B.

PubMed

Adler, Marc; Bryant, Judi; Buckman, Brad; Islam, Imadul; Larsen, Brent; Finster, Silke; Kent, Lorraine; May, Karen; Mohan, Raju; Yuan, Shendong; Whitlow, Marc

2005-07-05

This paper presents the crystal structure of porcine pancreatic carboxypeptidase B (pp-CpB) in complex with a variety of thiol-based inhibitors that were developed as antagonists of activated thrombin-activatable fibrinolysis inhibitor (TAFIa). Recent studies have indicated that a selective inhibitor of TAFIa could enhance the efficacy of existing thrombolytic agents for the treatment of acute myocardial infarction, one of the most prevalent forms of heart attacks. Unfortunately, activated TAFIa rapidly degrades in solution and cannot be used for crystallographic studies. In contrast, porcine pancreatic CpB is stable at room temperature and is available from commercial sources. Both pancreatic CpB and TAFIa are zinc-based exopeptidases, and the proteins share a 47% sequence identity. The homology improves considerably in the active site where nearly all of the residues are conserved. The inhibitors used in this study were designed to mimic a C-terminal arginine residue, one of the natural substrates of TAFIa. The X-ray structures show that the thiol group chelates the active site zinc, the carboxylic acid forms a salt bridge to Arg145, and the guanidine group forms two hydrogen bonds to Asp255. A meta-substituted phenyl was introduced into our inhibitors to reduce conformational freedom. This modification vastly improved the selectivity of compounds against other exopeptidases that cleave basic residues. Comparisons between structures indicate that selectivity derives from the interaction between the guanidine group in the inhibitors and an acidic active site residue. The location of this acidic residue is not conserved in the various carboxypeptidases.

Toposelective electrochemical desorption of thiol SAMs from neighboring polycrystalline gold surfaces.

PubMed

Tencer, Michal; Berini, Pierre

2008-11-04

We describe a method for the selective desorption of thiol self-assembled monolayers from gold surfaces having micrometer-scale separations on a substrate. In an electrolyte solution, the electrical resistance between the adjacent areas can be much lower than the resistance between a surface and the counter electrode. Also, both reductive and oxidative thiol desorption may occur. Therefore, the potentials of the surfaces must be independently controlled with a multichannel potentiostat and operating windows for a given thiol/electrolyte system must be established. In this study operating windows were established for 1-dodecanethiol-based SAMs in phosphate buffer, phosphate-buffered saline, and sodium hydroxide solution, and selective SAM removal was successfully performed in a four-electrode configuration.

Selenium-containing organic nanoparticles as silent precursors for ultra-sensitive thiol-responsive transmembrane anion transport.

PubMed

Lang, Chao; Zhang, Xin; Dong, Zeyuan; Luo, Quan; Qiao, Shanpeng; Huang, Zupeng; Fan, Xiaotong; Xu, Jiayun; Liu, Junqiu

2016-02-07

An anion transporter with a selenoxide group was able to form nanoparticles in water, whose activity was fully turned off due to the aggregation effect. The formed nanoparticles have a uniform size and can be readily dispersed in water at high concentrations. Turn-on of the nanoparticles by reducing molecules is proposed to be a combined process, including the reduction of selenoxide to selenide, disassembly of the nanoparticles and location of the transporter to the lipid membrane. Accordingly, a special acceleration phase can be observed in the turn-on kinetic curves. Since turn-on of the nanoparticles is quantitatively related to the amount of reductant, the nanoparticles can be activated in a step-by-step manner. Due to the sensibility of this system to thiols, cysteine can be detected at low nanomolar concentrations. This ultra-sensitive thiol-responsive transmembrane anion transport system is quite promising in biological applications.

Protein Thiol Redox Signaling in Monocytes and Macrophages.

PubMed

Short, John D; Downs, Kevin; Tavakoli, Sina; Asmis, Reto

2016-11-20

Monocyte and macrophage dysfunction plays a critical role in a wide range of inflammatory disease processes, including obesity, impaired wound healing diabetic complications, and atherosclerosis. Emerging evidence suggests that the earliest events in monocyte or macrophage dysregulation include elevated reactive oxygen species production, thiol modifications, and disruption of redox-sensitive signaling pathways. This review focuses on the current state of research in thiol redox signaling in monocytes and macrophages, including (i) the molecular mechanisms by which reversible protein-S-glutathionylation occurs, (ii) the identification of bona fide S-glutathionylated proteins that occur under physiological conditions, and (iii) how disruptions of thiol redox signaling affect monocyte and macrophage functions and contribute to atherosclerosis. Recent Advances: Recent advances in redox biochemistry and biology as well as redox proteomic techniques have led to the identification of many new thiol redox-regulated proteins and pathways. In addition, major advances have been made in expanding the list of S-glutathionylated proteins and assessing the role that protein-S-glutathionylation and S-glutathionylation-regulating enzymes play in monocyte and macrophage functions, including monocyte transmigration, macrophage polarization, foam cell formation, and macrophage cell death. Protein-S-glutathionylation/deglutathionylation in monocytes and macrophages has emerged as a new and important signaling paradigm, which provides a molecular basis for the well-established relationship between metabolic disorders, oxidative stress, and cardiovascular diseases. The identification of specific S-glutathionylated proteins as well as the mechanisms that control this post-translational protein modification in monocytes and macrophages will facilitate the development of new preventive and therapeutic strategies to combat atherosclerosis and other metabolic diseases. Antioxid. Redox Signal

Transporter-Mediated Disposition, Clinical Pharmacokinetics and Cholestatic Potential of Glyburide and Its Primary Active Metabolites.

PubMed

Li, Rui; Bi, Yi-An; Vildhede, Anna; Scialis, Renato J; Mathialagan, Sumathy; Yang, Xin; Marroquin, Lisa D; Lin, Jian; Varma, Manthena V S

2017-07-01

Glyburide is widely used for the treatment of type 2 diabetes. We studied the mechanisms involved in the disposition of glyburide and its pharmacologically active hydroxy metabolites M1 and M2b and evaluated their clinical pharmacokinetics and the potential role in glyburide-induced cholestasis employing physiologically based pharmacokinetic (PBPK) modeling. Transport studies of parent and metabolites in human hepatocytes and transfected cell systems imply hepatic uptake mediated by organic anion-transporting polypeptides. Metabolites are also subjected to basolateral and biliary efflux by P-glycoprotein, breast cancer resistance protein, and multidrug resistance-associated proteins, and are substrates to renal organic anion transporter 3. A PBPK model in combination with a Bayesian approach was developed considering the identified disposition mechanisms. The model reasonably described plasma concentration time profiles and urinary recoveries of glyburide and the metabolites, implying the role of multiple transport processes in their pharmacokinetics. Predicted free liver concentrations of the parent (∼30-fold) and metabolites (∼4-fold) were higher than their free plasma concentrations. Finally, all three compounds showed bile salt export pump inhibition in vitro; however, significant in vivo inhibition was not apparent for any compound on the basis of a predicted unbound liver exposure-response effect model using measured in vitro IC 50 values. In conclusion, this study demonstrates the important role of multiple drug transporters in the disposition of glyburide and its active metabolites, suggesting that variability in the function of these processes may lead to pharmacokinetic variability in the parent and the metabolites, potentially translating to pharmacodynamic variability. Copyright © 2017 by The American Society for Pharmacology and Experimental Therapeutics.

Human Metabolites of Cannabidiol: A Review on Their Formation, Biological Activity, and Relevance in Therapy

PubMed Central

Ujváry, István; Hanuš, Lumír

2016-01-01

Abstract Cannabidiol (CBD), the main nonpsychoactive constituent of Cannabis sativa, has shown a wide range of therapeutically promising pharmacological effects either as a sole drug or in combination with other drugs in adjunctive therapy. However, the targets involved in the therapeutic effects of CBD appear to be elusive. Furthermore, scarce information is available on the biological activity of its human metabolites which, when formed in pharmacologically relevant concentration, might contribute to or even account for the observed therapeutic effects. The present overview summarizes our current knowledge on the pharmacokinetics and metabolic fate of CBD in humans, reviews studies on the biological activity of CBD metabolites either in vitro or in vivo, and discusses relevant drug–drug interactions. To facilitate further research in the area, the reported syntheses of CBD metabolites are also catalogued. PMID:28861484

Human Metabolites of Cannabidiol: A Review on Their Formation, Biological Activity, and Relevance in Therapy.

PubMed

Ujváry, István; Hanuš, Lumír

2016-01-01

Cannabidiol (CBD), the main nonpsychoactive constituent of Cannabis sativa , has shown a wide range of therapeutically promising pharmacological effects either as a sole drug or in combination with other drugs in adjunctive therapy. However, the targets involved in the therapeutic effects of CBD appear to be elusive. Furthermore, scarce information is available on the biological activity of its human metabolites which, when formed in pharmacologically relevant concentration, might contribute to or even account for the observed therapeutic effects. The present overview summarizes our current knowledge on the pharmacokinetics and metabolic fate of CBD in humans, reviews studies on the biological activity of CBD metabolites either in vitro or in vivo , and discusses relevant drug-drug interactions. To facilitate further research in the area, the reported syntheses of CBD metabolites are also catalogued.

General and practical formation of thiocyanates from thiols.

PubMed

Frei, Reto; Courant, Thibaut; Wodrich, Matthew D; Waser, Jerome

2015-02-02

A new method for the cyanation of thiols and disulfides using cyanobenziodoxol(on)e hypervalent iodine reagents is described. Both aliphatic and aromatic thiocyanates can be accessed in good yields in a few minutes at room temperature starting from a broad range of thiols with high chemioselectivity. The complete conversion of disulfides to thiocyanates was also possible. Preliminary computational studies indicated a low energy concerted transition state for the cyanation of the thiolate anion or radical. The developed thiocyanate synthesis has broad potential for various applications in synthetic chemistry, chemical biology and materials science. © 2015 WILEY-VCH Verlag GmbH & Co. KGaA, Weinheim.

Selenocysteine in Thiol/Disulfide-Like Exchange Reactions

PubMed Central

Marino, Stefano M.

2013-01-01

Abstract Significance: Among trace elements used as cofactors in enzymes, selenium is unique in that it is incorporated into proteins co-translationally in the form of an amino acid, selenocysteine (Sec). Sec differs from cysteine (Cys) by only one atom (selenium versus sulfur), yet this switch dramatically influences important aspects of enzyme reactivity. Recent Advances: The main focus of this review is an updated and critical discussion on how Sec might be used to accelerate thiol/disulfide-like exchange reactions in natural selenoenzymes, compared with their Cys-containing homologs. Critical Issues: We discuss in detail three major aspects associated with thiol/disulfide exchange reactions: (i) nucleophilicity of the attacking thiolate (or selenolate); (ii) electrophilicity of the center sulfur (or selenium) atom; and (iii) stability of the leaving group (sulfur or selenium). In all these cases, we analyze the benefits that selenium might provide in these types of reactions. Future Directions: It is the biological thiol oxidoreductase-like function that benefits from the use of Sec, since Sec functions to chemically accelerate the rate of these reactions. We review various hypotheses that could help explain why Sec is used in enzymes, particularly with regard to competitive chemical advantages provided by the presence of the selenium atom in enzymes. Ultimately, these chemical advantages must be connected to biological functions of Sec. Antioxid. Redox Signal. 18, 1675–1689. PMID:23121622

Possible involvement of membrane lipids peroxidation and oxidation of catalytically essential thiols of the cerebral transmembrane sodium pump as component mechanisms of iron-mediated oxidative stress-linked dysfunction of the pump's activity

PubMed Central

Omotayo, T.I.; Akinyemi, G.S.; Omololu, P.A.; Ajayi, B.O.; Akindahunsi, A.A.; Rocha, J.B.T.; Kade, I.J.

2014-01-01

The precise molecular events defining the complex role of oxidative stress in the inactivation of the cerebral sodium pump in radical-induced neurodegenerative diseases is yet to be fully clarified and thus still open. Herein we investigated the modulation of the activity of the cerebral transmembrane electrogenic enzyme in Fe2+-mediated in vitro oxidative stress model. The results show that Fe2+ inhibited the transmembrane enzyme in a concentration dependent manner and this effect was accompanied by a biphasic generation of aldehydic product of lipid peroxidation. While dithiothreitol prevented both Fe2+ inhibitory effect on the pump and lipid peroxidation, vitamin E prevented only lipid peroxidation but not inhibition of the pump. Besides, malondialdehyde (MDA) inhibited the pump by a mechanism not related to oxidation of its critical thiols. Apparently, the low activity of the pump in degenerative diseases mediated by Fe2+ may involve complex multi-component mechanisms which may partly involve an initial oxidation of the critical thiols of the enzyme directly mediated by Fe2+ and during severe progression of such diseases; aldehydic products of lipid peroxidation such as MDA may further exacerbate this inhibitory effect by a mechanism that is likely not related to the oxidation of the catalytically essential thiols of the ouabain-sensitive cerebral electrogenic pump. PMID:25618580

An Antiestrogenic Activity Score for tamoxifen and its metabolites is associated with breast cancer outcome

PubMed Central

Alexi, X.; van Werkhoven, E.; Madlensky, L.; Natarajan, L.; Flatt, S. W.; Zwart, W.; Linn, S. C.; Parker, B. A.; Wu, A. H. B.; Pierce, J. P.; Huitema, A. D. R.; Beijnen, J. H.

2018-01-01

Purpose Endoxifen concentrations have been associated with breast cancer recurrence in tamoxifen-treated patients. However, tamoxifen itself and other metabolites also show antiestrogenic anti-tumor activity. Therefore, the aim of this study was to develop a comprehensive Antiestrogenic Activity Score (AAS), which accounts for concentration and antiestrogenic activity of tamoxifen and three metabolites. An association between the AAS and recurrence-free survival was investigated and compared to a previously published threshold for endoxifen concentrations of 5.97 ng/mL. Patients and methods The antiestrogenic activities of tamoxifen, (Z)-endoxifen, (Z)-4-hydroxytamoxifen, and N-desmethyltamoxifen were determined in a cell proliferation assay. The AAS was determined by calculating the sum of each metabolite concentration multiplied by an IC50 ratio, relative to tamoxifen. The AAS was calculated for 1370 patients with estrogen receptor alpha (ERα)-positive breast cancer. An association between AAS and recurrence was investigated using Cox regression and compared with the 5.97 ng/mL endoxifen threshold using concordance indices. Results An AAS threshold of 1798 was associated with recurrence-free survival, hazard ratio (HR) 0.67 (95% confidence interval (CI) 0.47–0.96), bias corrected after bootstrap HR 0.69 (95% CI 0.48–0.99). The concordance indices for AAS and endoxifen did not significantly differ; however, using the AAS threshold instead of endoxifen led to different dose recommendations for 5.2% of the patients. Conclusions Endoxifen concentrations can serve as a proxy for the antiestrogenic effect of tamoxifen and metabolites. However, for the aggregate effect of tamoxifen and three metabolites, defined by an integrative algorithm, a trend towards improving treatment is seen and moreover, is significantly associated with breast cancer recurrence. PMID:28005246

Advanced functionalization of polyhydroxyalkanoate via the UV-initiated thiol-ene click reaction.

PubMed

Tajima, Kenji; Iwamoto, Kosuke; Satoh, Yasuharu; Sakai, Ryosuke; Satoh, Toshifumi; Dairi, Tohru

2016-05-01

Polyhydroxyalkanoates (PHAs) incorporating vinyl-bearing 3-hydroxyalkanoates were prepared in 8.5-12.9 g L(-1) yield. The molar ratios (0-16 mol%) of the vinyl-bearing 3-hydroxyalkanoate derivatives were controlled by the continuous feeding of undecylenate at various concentrations. Subsequently, the PHAs were functionalized by UV-initiated thiol-ene click reaction and chemical modification. (1)H NMR spectra suggested that 3-mercaptopropionic acid and 2-aminoethanethiol were successfully introduced into the vinyl-bearing PHA. Subsequently, chemical modification using fluorescein or a fibronectin active fragment (GRGDS) was attempted. The former yielded a PHA derivative capable of emitting fluorescence under UV irradiation, which was useful for determining the miscibility of PHA in a composite film comprising poly-ʟ-lactic acid (PLLA) and PHA. In the latter case, PHA bearing GRGDS peptides exhibited cell adhesiveness, suggesting that its biocompatibility was improved upon peptide introduction. Taken together, the UV-initiated thiol-ene click reaction was demonstrated to be useful in PHA modification.

Highly Reactive Thiol-Norbornene Photo-Click Hydrogels: Toward Improved Processability.

PubMed

Van Hoorick, Jasper; Gruber, Peter; Markovic, Marica; Rollot, Mélanie; Graulus, Geert-Jan; Vagenende, Maxime; Tromayer, Maximilian; Van Erps, Jürgen; Thienpont, Hugo; Martins, José C; Baudis, Stefan; Ovsianikov, Aleksandr; Dubruel, Peter; Van Vlierberghe, Sandra

2018-06-10

In the present work, gelatin type B is modified with highly reactive norbornene functionalities (Gel-NB) following a one-pot synthesis approach to enable subsequent thiol-ene photo-click crosslinking. The modification strategy displays close control over the amount of introduced functionalities. Additionally, Gel-NB exhibits considerably improved processing capabilities in terms of two-photon polymerization when benchmarked to earlier-reported crosslinkable gelatin derivatives (e.g., gelatin-methacrylamide (Gel-MOD) and gelatin-methacrylamide-aminoethylmethacrylate (Gel-MOD-AEMA)). The improvement is especially apparent in terms of minimally required laser power (20 mW vs ≥60 mW (Gel-MOD) vs ≥40 mW (Gel-MOD-AEMA) at 100 mm s -1 scan speed) and processable concentration range (≥5 w/v% vs ≥10 w/v% (Gel-MOD/Gel-MOD-AEMA)). Furthermore, the proposed functionalization scheme maintains the excellent biocompatibility and cell interactivity of gelatin. Additionally, the norbornene functionalities have potential for straightforward postprocessing "thiol-ene" surface grafting of active molecules. As a consequence, a very promising material toward tissue engineering applications and more specifically, biofabrication, is presented. © 2018 WILEY-VCH Verlag GmbH & Co. KGaA, Weinheim.

Development of ionic gels using thiol-based monomers in ionic liquid

NASA Astrophysics Data System (ADS)

Ahmed, Kumkum; Naga, Naofumi; Kawakami, Masaru; Furukawa, Hidemitsu

2016-04-01

Ionic gels (IGs) using ionic liquids (ILs) can propose diverse applications in the field of optics, sensors and separation have opened wide prospects in materials science. ILs have attracted remarkable interest for gel polymer electrolytes and batteries based on their useful properties such as non-volatility, non-flammability, a wide electrochemical window, high thermal stability and a high ionic conductivity. The formation of gel in IL media makes it possible to immobilize ILs within organic or inorganic matrices and to take advantage of their unique properties in the solid state, thus eliminating some shortcomings related to shaping and risk of leakage. In this work for the first time we used multifunctional thiol monomers having uniform structure and good compatibility with the IL of our interest. Therefore we focused on developing thiol monomer-based IGs using multifunctional thiol monomers and acrylate crosslinkers utilizing thiol-ene reaction between monomer and crosslinking molecules in an IL medium and characterize their physico-chemical properties like thermal, conductive, mechanical properties etc.. This work has been focused mainly to improve the mechanical strength of IGs and make prospects of IGs in tribology and lubricants.

Linking diet, physical activity, cardiorespiratory fitness and obesity to serum metabolite networks: findings from a population-based study

PubMed Central

Floegel, A; Wientzek, A; Bachlechner, U; Jacobs, S; Drogan, D; Prehn, C; Adamski, J; Krumsiek, J; Schulze, M B; Pischon, T; Boeing, H

2014-01-01

Objective: It is not yet resolved how lifestyle factors and intermediate phenotypes interrelate with metabolic pathways. We aimed to investigate the associations between diet, physical activity, cardiorespiratory fitness and obesity with serum metabolite networks in a population-based study. Methods: The present study included 2380 participants of a randomly drawn subcohort of the European Prospective Investigation into Cancer and Nutrition-Potsdam. Targeted metabolomics was used to measure 127 serum metabolites. Additional data were available including anthropometric measurements, dietary assessment including intake of whole-grain bread, coffee and cake and cookies by food frequency questionnaire, and objectively measured physical activity energy expenditure and cardiorespiratory fitness in a subsample of 100 participants. In a data-driven approach, Gaussian graphical modeling was used to draw metabolite networks and depict relevant associations between exposures and serum metabolites. In addition, the relationship of different exposure metabolite networks was estimated. Results: In the serum metabolite network, the different metabolite classes could be separated. There was a big group of phospholipids and acylcarnitines, a group of amino acids and C6-sugar. Amino acids were particularly positively associated with cardiorespiratory fitness and physical activity. C6-sugar and acylcarnitines were positively associated with obesity and inversely with intake of whole-grain bread. Phospholipids showed opposite associations with obesity and coffee intake. Metabolite networks of coffee intake and obesity were strongly inversely correlated (body mass index (BMI): r=−0.57 and waist circumference: r=−0.59). A strong positive correlation was observed between metabolite networks of BMI and waist circumference (r=0.99), as well as the metabolite networks of cake and cookie intake with cardiorespiratory fitness and intake of whole-grain bread (r=0.52 and r=0

Linking diet, physical activity, cardiorespiratory fitness and obesity to serum metabolite networks: findings from a population-based study.

PubMed

Floegel, A; Wientzek, A; Bachlechner, U; Jacobs, S; Drogan, D; Prehn, C; Adamski, J; Krumsiek, J; Schulze, M B; Pischon, T; Boeing, H

2014-11-01

It is not yet resolved how lifestyle factors and intermediate phenotypes interrelate with metabolic pathways. We aimed to investigate the associations between diet, physical activity, cardiorespiratory fitness and obesity with serum metabolite networks in a population-based study. The present study included 2380 participants of a randomly drawn subcohort of the European Prospective Investigation into Cancer and Nutrition-Potsdam. Targeted metabolomics was used to measure 127 serum metabolites. Additional data were available including anthropometric measurements, dietary assessment including intake of whole-grain bread, coffee and cake and cookies by food frequency questionnaire, and objectively measured physical activity energy expenditure and cardiorespiratory fitness in a subsample of 100 participants. In a data-driven approach, Gaussian graphical modeling was used to draw metabolite networks and depict relevant associations between exposures and serum metabolites. In addition, the relationship of different exposure metabolite networks was estimated. In the serum metabolite network, the different metabolite classes could be separated. There was a big group of phospholipids and acylcarnitines, a group of amino acids and C6-sugar. Amino acids were particularly positively associated with cardiorespiratory fitness and physical activity. C6-sugar and acylcarnitines were positively associated with obesity and inversely with intake of whole-grain bread. Phospholipids showed opposite associations with obesity and coffee intake. Metabolite networks of coffee intake and obesity were strongly inversely correlated (body mass index (BMI): r = -0.57 and waist circumference: r = -0.59). A strong positive correlation was observed between metabolite networks of BMI and waist circumference (r = 0.99), as well as the metabolite networks of cake and cookie intake with cardiorespiratory fitness and intake of whole-grain bread (r = 0.52 and r = 0.50; respectively). Lifestyle factors

In-stream attenuation of neuro-active pharmaceuticals and their metabolites

USGS Publications Warehouse

Writer, Jeffrey; Antweiler, Ronald C.; Ferrar, Imma; Ryan, Joseph N.; Thurman, Michael

2013-01-01

In-stream attenuation was determined for 14 neuro-active pharmaceuticals and associated metabolites. Lagrangian sampling, which follows a parcel of water as it moves downstream, was used to link hydrological and chemical transformation processes. Wastewater loading of neuro-active compounds varied considerably over a span of several hours, and thus a sampling regime was used to verify that the Lagrangian parcel was being sampled and a mechanism was developed to correct measured concentrations if it was not. In-stream attenuation over the 5.4-km evaluated reach could be modeled as pseudo-first-order decay for 11 of the 14 evaluated neuro-active pharmaceutical compounds, illustrating the capacity of streams to reduce conveyance of neuro-active compounds downstream. Fluoxetine and N-desmethyl citalopram were the most rapidly attenuated compounds (t1/2 = 3.6 ± 0.3 h, 4.0 ± 0.2 h, respectively). Lamotrigine, 10,11,-dihydro-10,11,-dihydroxy-carbamazepine, and carbamazepine were the most persistent (t1/2 = 12 ± 2.0 h, 12 ± 2.6 h, 21 ± 4.5 h, respectively). Parent compounds (e.g., buproprion, carbamazepine, lamotrigine) generally were more persistent relative to their metabolites. Several compounds (citalopram, venlafaxine, O-desmethyl-venlafaxine) were not attenuated. It was postulated that the primary mechanism of removal for these compounds was interaction with bed sediments and stream biofilms, based on measured concentrations in stream biofilms and a column experiment using stream sediments.

The metabolism of primaquine to its active metabolite is dependent on CYP 2D6.

PubMed

Pybus, Brandon S; Marcsisin, Sean R; Jin, Xiannu; Deye, Gregory; Sousa, Jason C; Li, Qigui; Caridha, Diana; Zeng, Qiang; Reichard, Gregory A; Ockenhouse, Christian; Bennett, Jason; Walker, Larry A; Ohrt, Colin; Melendez, Victor

2013-06-20

The efficacy of the 8-aminoquinoline (8AQ) drug primaquine (PQ) has been historically linked to CYP-mediated metabolism. Although to date no clear evidence exists in the literature that unambiguously assigns the metabolic pathway or specific metabolites necessary for activity, recent literature suggests a role for CYP 2D6 in the generation of redox active metabolites. In the present study, the specific CYP 2D6 inhibitor paroxetine was used to assess its effects on the production of specific phenolic metabolites thought to be involved in PQ efficacy. Further, PQ causal prophylactic (developing liver stage) efficacy against Plasmodium berghei in CYP 2D knockout mice was assessed in comparison with a normal C57 background and with humanized CYP 2D6 mice to determine the direct effects of CYP 2D6 metabolism on PQ activity. PQ exhibited no activity at 20 or 40 mg/kg in CYP 2D knockout mice, compared to 5/5 cures in normal mice at 20 mg/kg. The activity against developing liver stages was partially restored in humanized CYP 2D6 mice. These results unambiguously demonstrate that metabolism of PQ by CYP 2D6 is essential for anti-malarial causal prophylaxis efficacy.

Activation and products of the cryptic secondary metabolite biosynthetic gene clusters by rifampin resistance (rpoB) mutations in actinomycetes.

PubMed

Tanaka, Yukinori; Kasahara, Ken; Hirose, Yutaka; Murakami, Kiriko; Kugimiya, Rie; Ochi, Kozo

2013-07-01

A subset of rifampin resistance (rpoB) mutations result in the overproduction of antibiotics in various actinomycetes, including Streptomyces, Saccharopolyspora, and Amycolatopsis, with H437Y and H437R rpoB mutations effective most frequently. Moreover, the rpoB mutations markedly activate (up to 70-fold at the transcriptional level) the cryptic/silent secondary metabolite biosynthetic gene clusters of these actinomycetes, which are not activated under general stressful conditions, with the exception of treatment with rare earth elements. Analysis of the metabolite profile demonstrated that the rpoB mutants produced many metabolites, which were not detected in the wild-type strains. This approach utilizing rifampin resistance mutations is characterized by its feasibility and potential scalability to high-throughput studies and would be useful to activate and to enhance the yields of metabolites for discovery and biochemical characterization.

The oxygen-centered radicals scavenging activity of sulfasalazine and its metabolites. A direct protection of the bowel.

PubMed

Prónai, L; Yukinobu, I; Láng, I; Fehér, J

1992-01-01

Oxygen-centered radicals, such as superoxide (O2-) and hydroxyl radicals (.OH) generated by phagocytes have been suggested to be involved in the pathogenesis of chronic inflammations of the bowel, such as Crohn's disease and colitis ulcerosa. Recently, sulfasalazine (SASP) and its metabolites have been reported to exert their effects as a direct scavenger of oxygen-centered radicals in the bowel. To scavenge oxygen-centered radicals in vivo, however, SASP and its metabolites have to react with O2- and/or .OH in vitro very rapidly, furthermore they have to reach an appropriate (possible millimolar) concentration range at the site of inflammation. To test this possibility, we investigated the direct O2- and .OH scavenging activity of SASP and its metabolites using the specific electron paramagnetic resonance/spin trapping method, and we compared the 50% inhibition rates of SASP and its metabolites with their known concentrations in the bowel and in the human plasma. It was found that SASP and its metabolites, such as 5-amino-salicylic acid (5-ASA), and acetyl-5-amino-salicylic acid (AC-5-ASA), but not sulfapyridine (SP) and acetyl-sulfapyridine (Ac-SP) have a direct O2- and .OH scavenging activity in vitro systems. Among the compounds, SASP and 5-ASA can reach a concentration which is appropriate to scavenge oxygen-centered radicals in the bowel but not in the human plasma. It was concluded that the in vivo antiinflammatory effects of SASP and its metabolites are, at least partly, due to the direct oxygen-centered scavenging activity of these drugs.

Interaction of benzene thiol and thiolate with small gold clusters.

PubMed

Letardi, Sara; Cleri, Fabrizio

2004-06-01

We studied the interaction between benzene thiol and thiolate molecules, and gold clusters made of 1 to 3 atoms, by means of ab initio density functional theory in the local density approximation. We find that the thiolate is energetically more stable than the thiol, however the process of detachment of H from the thiol appears to be possibly mediated by the intermediate step of H chemisorption on Au. Cleavage of the S-H bond is accompanied by a 90 degrees rotation of the molecule around the S-Au bond, showing a strong steric specificity. Such a rotation is induced by the relative energy shift of the S atom p orbitals with respect to the benzene pi ring and the Au d orbitals. By analyzing the correlation of the bond energy, bond lengths, and HOMO-LUMO gap with the number of S-Au bonds, we find that the thiolate S atom appears to prefer a low-coordination condition on Au clusters. (c) 2004 American Institute of Physics.

The First MS-Cleavable, Photo-Thiol-Reactive Cross-Linker for Protein Structural Studies

NASA Astrophysics Data System (ADS)

Iacobucci, Claudio; Piotrowski, Christine; Rehkamp, Anne; Ihling, Christian H.; Sinz, Andrea

2018-04-01

Cleavable cross-linkers are gaining increasing importance for chemical cross-linking/mass spectrometry (MS) as they permit a reliable and automated data analysis in structural studies of proteins and protein assemblies. Here, we introduce 1,3-diallylurea (DAU) as the first CID-MS/MS-cleavable, photo-thiol-reactive cross-linker. DAU is a commercially available, inexpensive reagent that efficiently undergoes an anti-Markovnikov hydrothiolation with cysteine residues in the presence of a radical initiator upon UV-A irradiation. Radical cysteine cross-linking proceeds via an orthogonal "click reaction" and yields stable alkyl sulfide products. DAU reacts at physiological pH and cross-linking reactions with peptides, and proteins can be performed at temperatures as low as 4 °C. The central urea bond is efficiently cleaved upon collisional activation during tandem MS experiments generating characteristic product ions. This improves the reliability of automated cross-link identification. Different radical initiators have been screened for the cross-linking reaction of DAU using the thiol-containing compounds cysteine and glutathione. Our concept has also been exemplified for the biologically relevant proteins bMunc13-2 and retinal guanylyl cyclase-activating protein-2. [Figure not available: see fulltext.

[Physico-chemical features of dinitrosyl iron complexes with natural thiol-containing ligands underlying biological activities of these complexes].

PubMed

Vanin, A F; Borodulin, R R; Kubrina, L N; Mikoian, V D; Burbaev, D Sh

2013-01-01

Current notions and new experimental data of the authors on physico-chemical features of dinitrosyl iron complexes with natural thiol-containing ligands (glutathione or cysteine), underlying the ability of the complexes to act as NO molecule and nitrosonium ion donors, are considered. This ability determines various biological activities of dinitrosyl iron complexes--inducing long-lasting vasodilation and thereby long-lasting hypotension in human and animals, inhibiting pellet aggregation, increasing red blood cell elasticity, thereby stimulating microcirculation, and reducing necrotic zone in animals with myocardial infarction. Moreover, dinitrosyl iron complexes are capable of accelerating skin wound healing, improving the function of penile cavernous tissue, blocking apoptosis development in cell cultures. When decomposed dinitrosyl iron complexes can exert cytotoxic effect that can be used for curing infectious and carcinogenic pathologies.

Thiols of flagellar proteins are essential for progressive motility in human spermatozoa.

PubMed

Cabrillana, María Eugenia; Monclus, María de Los Ángeles; Lancellotti, Tania Estefania Sáez; Boarelli, Paola Vanina; Vincenti, Amanda Edith; Fornés, Miguel Matias; Sanabria, Eduardo Alfredo; Fornés, Miguel Walter

2017-07-01

Male infertility is a disorder of the reproductive system defined by the failure to achieve a clinical pregnancy after 12 months or more of regular unprotected sexual intercourse. The presence of low-motile or immotile spermatozoa is one of many causes of infertility; however, this observation provides little or no information regarding the pathogenesis of the malfunction. Good sperm motility depends on correct assembly of the sperm tail in the testis and efficient maturation during epididymal transit. Thiols of flagellar proteins, such as outer dense fibre protein 1 (ODF1), are oxidised to form disulfides during epididymal transit and the spermatozoa become motile. This study was designed to determine how oxidative changes in protein thiol status affect progressive motility in human spermatozoa. Monobromobimane (mBBr) was used as a specific thiol marker and disruptor of sperm progressive motility. When mBBr was blocked by dithiothreitol it did not promote motility changes. The analysis of mBBr-treated spermatozoa revealed a reduction of progressive motility and an increased number of spermatozoa with non-progressive motility without affecting ATP production. Laser confocal microscopy and western blot analysis showed that one of the mBBr-positive proteins reacted with an antibody to ODF1. Monobromobimane fluorescence intensity of the sperm tail was lower in normozoospermic than asthenozoospermic men, suggesting that thiol oxidation in spermatozoa of asthenozoospermic men is incomplete. Our findings indicate that mBBr affects the thiol status of ODF1 in human spermatozoa and interferes with progressive motility.

Thiol surface functionalization via continuous phase plasma polymerization of allyl mercaptan, with subsequent maleimide-linked conjugation of collagen.

PubMed

Stynes, Gil D; Gengenbach, Thomas R; Kiroff, George K; Morrison, Wayne A; Kirkland, Mark A

2017-07-01

Thiol groups can undergo a large variety of chemical reactions and are used in solution phase to conjugate many bioactive molecules. Previous research on solid substrates with continuous phase glow discharge polymerization of thiol-containing monomers may have been compromised by oxidation. Thiol surface functionalization via glow discharge polymerization has been reported as requiring pulsing. Herein, continuous phase glow discharge polymerization of allyl mercaptan (2-propene-1-thiol) was used to generate significant densities of thiol groups on a mixed macrodiol polyurethane and tantalum. Three general classes of chemistry are used to conjugate proteins to thiol groups, with maleimide linkers being used most commonly. Here the pH specificity of maleimide reactions was used effectively to conjugate surface-bound thiol groups to amine groups in collagen. XPS demonstrated surface-bound thiol groups without evidence of oxidation, along with the subsequent presence of maleimide and collagen. Glow discharge reactor parameters were optimized by testing the resistance of bound collagen to degradation by 8 M urea. The nature of the chemical bonding of collagen to surface thiol groups was effectively assessed by colorimetric assay (ELISA) of residual collagen after incubation in 8 M urea over 8 days and after incubation with keratinocytes over 15 days. The facile creation of useable solid-supported thiol groups via continuous phase glow discharge polymerization of allyl mercaptan opens a route for attaching a vast array of bioactive molecules. © 2016 Wiley Periodicals, Inc. J Biomed Mater Res Part A: 105A: 1940-1948, 2017. © 2017 Wiley Periodicals, Inc.

Mass Spectrometry in Studies of Protein Thiol Chemistry and Signaling: Opportunities and Caveats

PubMed Central

Devarie Baez, Nelmi O.; Reisz, Julie A.; Furdui, Cristina M.

2014-01-01

Mass spectrometry (MS) has become a powerful and widely utilized tool in the investigation of protein thiol chemistry, biochemistry, and biology. Very early biochemical studies of metabolic enzymes have brought to light the broad spectrum of reactivity profiles that distinguish cysteine thiols with functions in catalysis and protein stability from other cysteine residues in proteins. The development of MS methods for the analysis of proteins using electrospray ionization (ESI) or matrix-assisted laser desorption/ionization (MALDI) coupled with the emergence of high-resolution mass analyzers have been instrumental in advancing studies of thiol modifications, both in single proteins and within the cellular context. This article reviews MS instrumentation and methods of analysis employed in investigations of thiols and their reactivity toward a range of small biomolecules. A selected number of studies are detailed to highlight the advantages brought about by the MS technologies along with the caveats associated with these analyses. PMID:25261734

Hydrangea-like magneto-fluorescent nanoparticles through thiol-inducing assembly

NASA Astrophysics Data System (ADS)

Chen, Shun; Zhang, Junjun; Song, Shaokun; Xiong, Chuanxi; Dong, Lijie

2017-01-01

Magneto-fluorescent nanoparticles (NPs), recognized as an emerging class of materials, have drawn much attention because of their potential applications. Due to surface functionalization and thiol-metal bonds, a simple method has been put forward for fabricating hydrangea-like magneto-fluorescent Fe3O4-SH@QD NPs, through assembling thiol-modified Fe3O4 NPs with sub-size multi-layer core/shell CdSe/CdS/ZnS QDs. After a refined but controllable silane hydrolysis process, thiol-modified Fe3O4 was fabricated, resulting in Fe3O4-SH@QD NPs with QDs, while preventing the quenching of the QDs. As a result, the core Fe3O4 NPs were 18 nm in diameter, while the scattered CdSe/CdS/ZnS QDs were 7 nm in diameter. The resultant magneto-fluorescent Fe3O4-SH@QD NPs exhibit efficient fluorescence, superparamagnetism at room temperature, and rapid response to the external field, which make them ideal candidates for difunctional probes in MRI and bio-labels, targeting and photodynamic therapy, and cell tracking and separation.

Contact-Engineered Electrical Properties of MoS2 Field-Effect Transistors via Selectively Deposited Thiol-Molecules.

PubMed

Cho, Kyungjune; Pak, Jinsu; Kim, Jae-Keun; Kang, Keehoon; Kim, Tae-Young; Shin, Jiwon; Choi, Barbara Yuri; Chung, Seungjun; Lee, Takhee

2018-05-01

Although 2D molybdenum disulfide (MoS 2 ) has gained much attention due to its unique electrical and optical properties, the limited electrical contact to 2D semiconductors still impedes the realization of high-performance 2D MoS 2 -based devices. In this regard, many studies have been conducted to improve the carrier-injection properties by inserting functional paths, such as graphene or hexagonal boron nitride, between the electrodes and 2D semiconductors. The reported strategies, however, require relatively time-consuming and low-yield transfer processes on sub-micrometer MoS 2 flakes. Here, a simple contact-engineering method is suggested, introducing chemically adsorbed thiol-molecules as thin tunneling barriers between the metal electrodes and MoS 2 channels. The selectively deposited thiol-molecules via the vapor-deposition process provide additional tunneling paths at the contact regions, improving the carrier-injection properties with lower activation energies in MoS 2 field-effect transistors. Additionally, by inserting thiol-molecules at the only one contact region, asymmetric carrier-injection is feasible depending on the temperature and gate bias. © 2018 WILEY-VCH Verlag GmbH & Co. KGaA, Weinheim.

Integrated circuit-based electrochemical sensor for spatially resolved detection of redox-active metabolites in biofilms.

PubMed

Bellin, Daniel L; Sakhtah, Hassan; Rosenstein, Jacob K; Levine, Peter M; Thimot, Jordan; Emmett, Kevin; Dietrich, Lars E P; Shepard, Kenneth L

2014-01-01

Despite advances in monitoring spatiotemporal expression patterns of genes and proteins with fluorescent probes, direct detection of metabolites and small molecules remains challenging. A technique for spatially resolved detection of small molecules would benefit the study of redox-active metabolites that are produced by microbial biofilms and can affect their development. Here we present an integrated circuit-based electrochemical sensing platform featuring an array of working electrodes and parallel potentiostat channels. 'Images' over a 3.25 × 0.9 mm(2) area can be captured with a diffusion-limited spatial resolution of 750 μm. We demonstrate that square wave voltammetry can be used to detect, identify and quantify (for concentrations as low as 2.6 μM) four distinct redox-active metabolites called phenazines. We characterize phenazine production in both wild-type and mutant Pseudomonas aeruginosa PA14 colony biofilms, and find correlations with fluorescent reporter imaging of phenazine biosynthetic gene expression.

Integrated circuit-based electrochemical sensor for spatially resolved detection of redox-active metabolites in biofilms

PubMed Central

Bellin, Daniel L.; Sakhtah, Hassan; Rosenstein, Jacob K.; Levine, Peter M.; Thimot, Jordan; Emmett, Kevin; Dietrich, Lars E. P.; Shepard, Kenneth L.

2014-01-01

Despite advances in monitoring spatiotemporal expression patterns of genes and proteins with fluorescent probes, direct detection of metabolites and small molecules remains challenging. A technique for spatially resolved detection of small molecules would benefit the study of redox-active metabolites produced by microbial biofilms, which can drastically affect colony development. Here we present an integrated circuit-based electrochemical sensing platform featuring an array of working electrodes and parallel potentiostat channels. “Images” over a 3.25 × 0.9 mm area can be captured with a diffusion-limited spatial resolution of 750 μm. We demonstrate that square wave voltammetry can be used to detect, identify, and quantify (for concentrations as low as 2.6 μM) four distinct redox-active metabolites called phenazines. We characterize phenazine production in both wild-type and mutant Pseudomonas aeruginosa PA14 colony biofilms, and find correlations with fluorescent reporter imaging of phenazine biosynthetic gene expression. PMID:24510163

UHPLC-MS/MS determination of varietal thiol precursors in Sauvignon Blanc grapes.

PubMed

Vanzo, Andreja; Janeš, Lucija; Požgan, Franc; Velikonja Bolta, Špela; Sivilotti, Paolo; Lisjak, Klemen

2017-10-13

Varietal thiol precursors in grapes are subject to metabolic changes during post-harvest treatments. Metabolic activity should therefore be limited after sampling to understand their biosynthesis in the berry and genetic regulation. In this study, berries were frozen in liquid nitrogen immediately after harvesting, transported in dry ice, stored briefly at -80 °C, cryo-milled and extracted without being thawed in cold methanol in a ratio of 1:4 (w/v). A UHPLC-MS/MS method for quantitative determination of the thiol precursors 3-S-glutathionylhexan-1-ol (G3MH), 3-S-cysteinylhexan-1-ol (Cys3MH), 4-S-glutathionyl-4-methylpentan-2-one (G4MMP) and 4-S-cysteinyl-4-methylpentan-2-one (Cys4MMP), glutathione, oxidized glutathione and L-methionine in grapes was developed. Reference material was provided through synthesis of precursors and their deuterium labelled analogues. The average thiol precursor content in grapes in 2013-15 was in the range 8-16 μg kg -1 for G3MH, 1-6 μg kg -1 for Cys3MH, 1-4 μg kg -1 for Cys4MMP and 0.3 μg kg -1 for G4MMP. In 2013 and 2014, the highest precursor content in mature Sauvignon Blanc grapes from vineyards located in Italy regarded G3MH, followed by Cys3MH, Cys4MMP and G4MMP. In 2015, G3MH was again the most abundant precursor, but followed by Cys4MMP, Cys3MH and G4MMP.

Detection of Free Thiols and Fluorescence Response of Phycoerythrin Chromophore after Ultraviolet-B Radiation Stress.

PubMed

Kannaujiya, Vinod K; Sinha, Rajeshwar P

2017-03-01

The chemistry of thiol-chromophore linkage plays a central role in the nature of fluorescence of phycoerythrin (PE). Interaction of thiol and chromophore is crucial for the energy transfer, redox signal and inhibition of oxidative damage. In the present investigation the effects of ultraviolet-B radiation on an emission fluorescence intensity and wavelength shift in PE due to interaction between thiol and chromophore by remarkable strategy of detection technique was studied. Purification of PE was done by using a gel permeation and ion exchange chromatography that yielded a quite high purity index (6.40) in a monomeric (αβ) form. UV-B radiation accelerated the quenching efficiency (24.9 ± 1.52%) by reducing fluorescence emission intensity of thiol linked chromophore after 240 min of UV-B exposure. However, after blocking of transiently released free thiol by N-ethylmaleimide, quenching efficiency was increased (36.8 ± 2.80%) with marked emission wavelength shift towards shorter wavelengths up to 562 nm as compared to 575 nm in control. Emission fluorescence of free thiol was at maximum after 240 min that was detected specifically by monobromobimane (mBrB) molecular probe. The association/dissociation of bilin chromophore was analyzed by SDS- and Native-PAGE that also indicated a complete reduction in emission fluorescence. Our work clearly shows an early detection of free thiols and relative interaction with chromophore after UV-B radiation which might play a significant role in structural and functional integrity of terminal PE.

Modified cupric reducing antioxidant capacity (CUPRAC) assay for measuring the antioxidant capacities of thiol-containing proteins in admixture with polyphenols.

PubMed

Cekiç, Sema Demirci; Başkan, Kevser Sözgen; Tütem, Esma; Apak, Reşat

2009-07-15

Proteins are not considered as true antioxidants but are known to protect antioxidants from oxidation in various antioxidant activity assays. This study aims to investigate the contribution of proteins, especially thiol-containing proteins, to the observed overall antioxidant capacity measured by known methods. To determine the antioxidant properties of thiol-containing proteins, the CUPRAC method of antioxidant assay using the oxidizing reagent Cu(II)-neocuproine previously used for simultaneous analysis of cystine and cysteine was adopted. While the CUPRAC method is capable of determining all antioxidant compounds including thiols in complex sample matrices, the Ellman method of thiol quantitation basically does not respond to other antioxidants. The antioxidant quantities in the selected samples were assayed with the ABTS and FRAP methods as well as with the CUPRAC method. In all applied methods, the dilutions were made with a standard pH 8 buffer used in the Ellman method by substituting the Na(2)EDTA component of the buffer with sodium citrate. On the other hand, the standard CUPRAC protocol was modified by substituting the pH 7 ammonium acetate buffer (at 1M concentration) with 8M urea buffer adjusted to pH 7 by neutralizing with 6M HCl. Urea helps to partly solubilize and denaturate proteins so that their buried thiols be oxidized more easily. All methods used in the estimation of antioxidant properties of proteins (i.e., CUPRAC, Ellman, ABTS, and FRAP) were first standardized with a simple thiol compound, cysteine, by constructing the calibration curves. The molar absorptivities of these methods for cysteine were: epsilon(CUPRAC)=7.71x10(3), epsilon(Ellman)=1.37x10(4), epsilon(ABTS)=2.06x10(4), and epsilon(FRAP)=2.98x10(3)L mol(-1)cm(-1). Then these methods were applied to various samples containing thiols, such as glutathione (reduced form:GSH), egg white, whey proteins, and gelatin. Additionally, known quantities of selected antioxidants were added to

Enzymatic Continuous Flow Synthesis of Thiol-Terminated Poly(δ-Valerolactone) and Block Copolymers.

PubMed

Zhu, Ning; Huang, Weijun; Hu, Xin; Liu, Yihuan; Fang, Zheng; Guo, Kai

2018-04-01

Thiol-terminated poly(δ-valerolactone) is directly synthesized via enzymatic 6-mercapto-1-hexanol initiated ring-opening polymerization in both batch and microreactor. By using Candida antartica Lipase B immobilized tubular reactor, narrowly dispersed poly(δ-valerolactone) with higher thiol fidelity is more efficiently prepared in contrast to the batch reactor. Moreover, the integrated enzyme packed tubular reactor system is established to perform the chain extension experiments. Thiol-terminated poly(δ-valerolactone)-block-poly(ε-caprolactone) and poly(ε-caprolactone)-block-poly(δ-valerolactone) are easily prepared by modulating the monomer introduction sequence. © 2018 WILEY-VCH Verlag GmbH & Co. KGaA, Weinheim.

Evaluation of the in vitro activity of levornidazole, its metabolites and comparators against clinical anaerobic bacteria.

PubMed

Hu, Jiali; Zhang, Jing; Wu, Shi; Zhu, Demei; Huang, Haihui; Chen, Yuancheng; Yang, Yang; Zhang, Yingyuan

2014-12-01

This study evaluated the in vitro anti-anaerobic activity and spectrum of levornidazole, its metabolites and comparators against 375 clinical isolates of anaerobic bacteria, including Gram-negative bacilli (181 strains), Gram-negative cocci (11 strains), Gram-positive bacilli (139 strains) and Gram-positive cocci (44 strains), covering 34 species. Minimum inhibitory concentrations (MICs) of levornidazole, its five metabolites and three comparators against these anaerobic isolates were determined by the agar dilution method. Minimum bactericidal concentrations (MBCs) of levornidazole and metronidazole were measured against 22 strains of Bacteroides fragilis. Levornidazole showed good activity against B. fragilis, other Bacteroides spp., Clostridium difficile, Clostridium perfringens and Peptostreptococcus magnus, evidenced by MIC90 values of 0.5, 1, 0.25, 2 and 1mg/L, respectively. The activity of levornidazole and the comparators was poor for Veillonella spp. Generally, levornidazole displayed activity similar to or slightly higher than that of metronidazole, ornidazole and dextrornidazole against anaerobic Gram-negative bacilli, Gram-positive bacilli and Gram-positive cocci, especially B. fragilis. Favourable anti-anaerobic activity was also seen with levornidazole metabolites M1 and M4 but not M2, M3 or M5. For the 22 clinical B. fragilis strains, MBC50 and MBC90 values of levornidazole were 2mg/L and 4mg/L, respectively. Both MBC50/MIC50 and MBC90/MIC90 ratios of levornidazole were 4, similar to those of metronidazole. Levornidazole is an important anti-anaerobic option in clinical settings in terms of its potent and broad-spectrum in vitro activity, bactericidal property, and the anti-anaerobic activity of its metabolites M1 and M4. Copyright © 2014 Elsevier B.V. and the International Society of Chemotherapy. All rights reserved.

Antioxidant potential of fungal metabolite nigerloxin during eye lens abnormalities in galactose-fed rats.

PubMed

Suresha, Bharathinagar S; Srinivasan, Krishnapura

2013-10-01

The role of osmotic and oxidative stress has been strongly implicated in the pathogenesis of cataract. Nigerloxin, a fungal metabolite, has been shown to possess aldose reductase inhibition and improved antioxidant defense system in lens of diabetic rats. In the present study, the beneficial influence of nigerloxin was investigated in galactose-induced cataract in experimental animals. Cataract was induced in Wistar rats by feeding 30% galactose in diet. Groups of galactose-fed rats were orally administered with nigerloxin (25 and 100 mg/kg body weight/day) for 24 days. Lens aldose reductase activity was increased significantly in galactose-fed animals. Lens lipid peroxides and advanced glycation end products were also significantly increased. Antioxidant molecule - reduced glutathione, total thiols and activities of antioxidant enzymes superoxide dismutase and glutathione peroxidase were decreased in the lens of galactose-fed animals. Oral administration of nigerloxin once a day for 24 days at a dose of 100 mg/kg body weight, significantly decreased lens lipid peroxides and advanced glycation end products in galactose-fed rats. Lens aldose reductase activity was reduced and lens antioxidant molecules and antioxidant enzyme activities were elevated significantly by nigerloxin administration. The results suggest that alteration in polyol pathway and antioxidant defense system were countered by nigerloxin in the lens of galactose-fed animals, suggesting the potential of nigerloxin in ameliorating the development of galactose-induced cataract in experimental animals.

In Vitro Antibacterial Activities of AF 3013, the Active Metabolite of Prulifloxacin, against Nosocomial and Community Italian Isolates

PubMed Central

Montanari, Maria Pia; Mingoia, Marina; Varaldo, Pietro Emanuele

2001-01-01

AF 3013, the active metabolite of prulifloxacin, was tested to determine its inhibitory and bactericidal activities against 396 nosocomial and 258 community Italian isolates. Compared with that of ciprofloxacin, its activity (assessed in MIC and minimal bactericidal concentration tests) was generally similar or greater against gram-positive bacteria and greater against gram-negative bacteria. In time-kill assays using selected isolates, its bactericidal activity was comparable to that of ciprofloxacin. PMID:11709353

A structurally driven analysis of thiol reactivity in mammalian albumins.

PubMed

Spiga, Ottavia; Summa, Domenico; Cirri, Simone; Bernini, Andrea; Venditti, Vincenzo; De Chiara, Matteo; Priora, Raffaella; Frosali, Simona; Margaritis, Antonios; Di Giuseppe, Danila; Di Simplicio, Paolo; Niccolai, Neri

2011-04-01

Understanding the structural basis of protein redox activity is still an open question. Hence, by using a structural genomics approach, different albumins have been chosen to correlate protein structural features with the corresponding reaction rates of thiol exchange between albumin and disulfide DTNB. Predicted structures of rat, porcine, and bovine albumins have been compared with the experimentally derived human albumin. High structural similarity among these four albumins can be observed, in spite of their markedly different reactivity with DTNB. Sequence alignments offered preliminary hints on the contributions of sequence-specific local environments modulating albumin reactivity. Molecular dynamics simulations performed on experimental and predicted albumin structures reveal that thiolation rates are influenced by hydrogen bonding pattern and stability of the acceptor C34 sulphur atom with donor groups of nearby residues. Atom depth evolution of albumin C34 thiol groups has been monitored during Molecular Dynamic trajectories. The most reactive albumins appeared also the ones presenting the C34 sulphur atom on the protein surface with the highest accessibility. High C34 sulphur atom reactivity in rat and porcine albumins seems to be determined by the presence of additional positively charged amino acid residues favoring both the C34 S⁻ form and the approach of DTNB. Copyright © 2011 Wiley Periodicals, Inc.

Reaction Mechanisms of Metals with Hydrogen Sulfide and Thiols in Model Wine. Part 1: Copper-Catalyzed Oxidation.

PubMed

Kreitman, Gal Y; Danilewicz, John C; Jeffery, David W; Elias, Ryan J

2016-05-25

Sulfidic off-odors as a result of hydrogen sulfide (H2S) and low-molecular-weight thiols are commonly encountered in wine production. These odors are usually removed by the process of Cu(II) fining, a process that remains poorly understood. The present study aims to elucidate the underlying mechanisms by which Cu(II) interacts with H2S and thiol compounds (RSH) under wine-like conditions. Copper complex formation was monitored along with H2S, thiol, oxygen, and acetaldehyde concentrations after the addition of Cu(II) (50 or 100 μM) to air-saturated model wine solutions containing H2S, cysteine, 6-sulfanylhexan-1-ol, or 3-sulfanylhexan-1-ol (300 μM each). The presence of H2S and thiols in excess to Cu(II) led to the rapid formation of ∼1.4:1 H2S/Cu and ∼2:1 thiol/Cu complexes, resulting in the oxidation of H2S and thiols and reduction of Cu(II) to Cu(I), which reacted with oxygen. H2S was observed to initially oxidize rather than form insoluble copper sulfide. The proposed reaction mechanisms provide insight into the extent to which H2S can be selectively removed in the presence of thiols in wine.

Effects of clopidogrel on the pharmacokinetics of sibutramine and its active metabolites.

PubMed

Bae, Jung-Woo; Jang, Choon-Gon; Lee, Seok-Young

2011-12-01

Sibutramine is metabolized by the enzymes CYP2B6 and CYP2C19 into 2 active metabolites, M1 (mono-desmethyl sibutramine) and M2 (di-desmethyl sibutramine). Clopidogrel is a mechanism-based inhibitor of CYP2B6 and CYP2C19. In this study, 13 extensive metabolizers of CYP2B6 and CYP2C19 were evaluated to clarify whether clopidogrel inhibits the formation of the active metabolites of sibutramine. In the control phase, each subject received a 15-mg oral dose of sibutramine. After a washout period of 2 weeks, in the clopidogrel phase, the subjects received 300 mg of clopidogrel on the first day and then 75-mg once daily for 6 days. One hour after the last dosing of clopidogrel, all subjects received 15-mg of sibutramine. Compared with the control phase, the mean sibutramine and M1 plasma concentrations were higher after clopidogrel treatment. Clopidogrel significantly increased the half-life (242% of control phase) and area under the plasma concentration-time curve from 0 to infinity (AUC(inf)) (227% of control phase) of sibutramine and decreased the apparent oral clearance (31% of control phase) of sibutramine. Pharmacokinetic analysis showed significant increases in the AUC(inf) (162% of control phase) of M1. The CYP2B6 and CYP2C19 inhibitor clopidogrel significantly inhibited the formations of M1 from sibutramine and M2 from sibutramine by 37% and 64%, respectively. Therefore, CYP2B6 and CYP2C19 are in vivo catalysts for the formation of the 2 active metabolites of sibutramine.

Bisphenol-A alters microbiota metabolites derived from aromatic amino acids and worsens disease activity during colitis.

PubMed

DeLuca, Jennifer Aa; Allred, Kimberly F; Menon, Rani; Riordan, Rebekah; Weeks, Brad R; Jayaraman, Arul; Allred, Clinton D

2018-06-01

Inflammatory bowel disease is a complex collection of disorders. Microbial dysbiosis as well as exposure to toxins including xenoestrogens are thought to be risk factors for inflammatory bowel disease development and relapse. Bisphenol-A has been shown to exert estrogenic activity in the colon and alter intestinal function, but the role that xenoestrogens, such as bisphenol-A , play in colonic inflammation has been previously described but with conflicting results. We investigated the ability of bisphenol-A to exacerbate colonic inflammation and alter microbiota metabolites derived from aromatic amino acids in an acute dextran sulfate sodium-induced colitis model. Female C57BL/6 mice were ovariectomized and exposed to bisphenol-A daily for 15 days. Disease activity measures include body weight, fecal consistency, and rectal bleeding. Colons were scored for inflammation, injury, and nodularity. Alterations in the levels of microbiota metabolites derived from aromatic amino acids known to reflect phenotypic changes in the gut microbiome were analyzed. Bisphenol-A exposure increased mortality and worsened disease activity as well as inflammation and nodularity scores in the middle colon region following dextran sulfate sodium exposure. Unique patterns of metabolites were associated with bisphenol-A consumption. Regardless of dextran sulfate sodium treatment, bisphenol-A reduced levels of tryptophan and several metabolites associated with decreased inflammation in the colon. This is the first study to show that bisphenol-A treatment alone can reduce microbiota metabolites derived from aromatic amino acids in the colon which may be associated with increased colonic inflammation and inflammatory bowel disease. Impact statement As rates of inflammatory bowel disease rise, discovery of the mechanisms related to the development of these conditions is important. Environmental exposure is hypothesized to play a role in etiology of the disease, as are alterations in the gut

Metabolites related to renal function, immune activation, and carbamylation are associated with muscle composition in older adults.

PubMed

Lustgarten, Michael S; Fielding, Roger A

2017-12-15

Reduced skeletal muscle density in older adults is associated with insulin resistance, decreased physical function, and an increased all-cause mortality risk. To elucidate mechanisms that may underlie the maintenance of skeletal muscle density, we conducted a secondary analysis of previously published muscle composition and serum metabolomic data in 73 older adults (average age, 78y). Multivariable-adjusted linear regression was used to examine associations between 321 metabolites with muscle composition, defined as the ratio between normal density (NDM) with low density (LDM) thigh muscle cross sectional area (NDM/LDM). Sixty metabolites were significantly (p≤0.05 and q<0.30) associated with NDM/LDM. Decreased renal function and the immune response have been previously linked with reduced muscle density, but the mechanisms underlying these connections are less clear. Metabolites that were significantly associated with muscle composition were then tested for their association with circulating markers of renal function (blood urea nitrogen, creatinine, uric acid), and with the immune response (neutrophils/lymphocytes) and activation (kynurenine/tryptophan). 43 significant NDM/LDM metabolites (including urea) were co-associated with at least 1 marker of renal function; 23 of these metabolites have been previously identified as uremic solutes. The neutrophil/lymphocyte ratio was significantly associated with NDM/LDM (β±SE: -0.3±0.1, p=0.01, q=0.04). 35 significant NDM/LDM metabolites were co-associated with immune activation. Carbamylation (defined as homocitrulline/lysine) was identified as a pathway that may link renal function and immune activation with muscle composition, as 29 significant NDM/LDM metabolites were co-associated with homocitrulline/lysine, with at least 2 markers of renal function, and with kynurenine/tryptophan. When considering that elevated urea and uremic metabolites have been linked with an increased systemic microbial burden, that

In vitro glutathione peroxidase mimicry of ebselen is linked to its oxidation of critical thiols on key cerebral suphydryl proteins - A novel component of its GPx-mimic antioxidant mechanism emerging from its thiol-modulated toxicology and pharmacology.

PubMed

Kade, I J; Balogun, B D; Rocha, J B T

2013-10-25

The antioxidant mechanism of ebselen in rats brain is largely linked with its glutathione peroxidase (GPx) rather than its peroxiredoxin mimicry ability. However, the precise molecular dynamics between the GPx-mimicry of ebselen and thiol utilization is yet to be fully clarified and thus still open. Herein, we investigated the influence of dithiothreitol (DTT) on the antioxidant action of ebselen against oxidant-induced cerebral lipid peroxidation and deoxyribose degradation. Furthermore, the critical inhibitory concentrations of ebselen on the activities of sulphydryl enzymes such as cerebral sodium pump, δ-aminolevulinic acid dehydratase (δ-ALAD) and lactate dehydrogenase (LDH) were also investigated. We observe that ebselen (at ≥42 μM) markedly inhibited lipid peroxidation in the presence and absence of DTT, whereas it inhibited deoxyribose degradation only in the presence of DTT. Furthermore, under in vitro conditions, ebselen inhibited the thiol containing enzymes; cerebral sodium pump (at ≥40 μM), δ-ALAD (≥10 μM) and LDH (≥1 μM) which were either prevented or reversed by DTT. However, the inhibition of the activities of these sulphydryl proteins in diabetic animals was prevented by ebselen. Summarily, it is apparent that the effective in vitro inhibitory doses of ebselen on the activity of the sulphydryl proteins are far less than its antioxidant doses. In addition, the presence of DTT is evidently a critical requirement for ebselen to effect its antioxidant action against deoxyribose degeradation and not lipid peroxidation. Consequently, we conclude that ebselen possibly utilizes available thiols on sulphydryl proteins to effect its GPx mimicry antioxidant action against lipid peroxidation in rat brain homogenate. Copyright © 2013 Elsevier Ireland Ltd. All rights reserved.

Secondary Metabolites Produced by an Endophytic Fungus Pestalotiopsis sydowiana and Their 20S Proteasome Inhibitory Activities.

PubMed

Xia, Xuekui; Kim, Soonok; Liu, Changheng; Shim, Sang Hee

2016-07-20

Fungal endophytes have attracted attention due to their functional diversity. Secondary metabolites produced by Pestalotiopsis sydowiana from a halophyte, Phragmites communis Trinus, were investigated. Eleven compounds, including four penicillide derivatives (1-4) and seven α-pyrone analogues (5-10) were isolated from cultures of P. sydowiana. The compounds were identified based on spectroscopic data. The inhibitory activities against the 20S proteasome were evaluated. Compounds 1-3, 5, and 9-10 showed modest proteasome inhibition activities, while compound 8 showed strong activity with an IC50 of 1.2 ± 0.3 μM. This is the first study on the secondary metabolites produced by P. sydowiana and their proteasome inhibitory activities. The endophytic fungus P. sydowiana might be a good resource for proteasome inhibitors.

Thiol redox transitions in cell signaling: a lesson from N-acetylcysteine.

PubMed

Parasassi, Tiziana; Brunelli, Roberto; Costa, Graziella; De Spirito, Marco; Krasnowska, Ewa; Lundeberg, Thomas; Pittaluga, Eugenia; Ursini, Fulvio

2010-06-29

The functional status of cells is under the control of external stimuli affecting the function of critical proteins and eventually gene expression. Signal sensing and transduction by messengers to specific effectors operate by post-translational modification of proteins, among which thiol redox switches play a fundamental role that is just beginning to be understood. The maintenance of the redox status is, indeed, crucial for cellular homeostasis and its dysregulation towards a more oxidized intracellular environment is associated with aberrant proliferation, ultimately related to diseases such as cancer, cardiovascular disease, and diabetes. Redox transitions occur in sensitive cysteine residues of regulatory proteins relevant to signaling, their evolution to metastable disulfides accounting for the functional redox switch. N-acetylcysteine (NAC) is a thiol-containing compound that is able to interfere with redox transitions of thiols and, thus, in principle, able to modulate redox signaling. We here review the redox chemistry of NAC, then screen possible mechanisms to explain the effects observed in NAC-treated normal and cancer cells; such effects involve a modification of global gene expression, thus of functions and morphology, with a leitmotif of a switch from proliferation to terminal differentiation. The regulation of thiol redox transitions in cell signaling is, therefore, proposed as a new tool, holding promise not only for a deeper explanation of mechanisms, but indeed for innovative pharmacological interventions.

Activation of the Silent Secondary Metabolite Production by Introducing Neomycin-Resistance in a Marine-Derived Penicillium purpurogenum G59

PubMed Central

Wu, Chang-Jing; Yi, Le; Cui, Cheng-Bin; Li, Chang-Wei; Wang, Nan; Han, Xiao

2015-01-01

Introduction of neomycin-resistance into a marine-derived, wild-type Penicillium purpurogenum G59 resulted in activation of silent biosynthetic pathways for the secondary metabolite production. Upon treatment of G59 spores with neomycin and dimethyl sulfoxide (DMSO), a total of 56 mutants were obtained by single colony isolation. The acquired resistance of mutants to neomycin was testified by the resistance test. In contrast to the G59 strain, the EtOAc extracts of 28 mutants inhibited the human cancer K562 cells, indicating that the 28 mutants have acquired the capability to produce bioactive metabolites. HPLC-photodiode array detector (PDAD)-UV and HPLC-electron spray ionization (ESI)-MS analyses further indicated that diverse secondary metabolites have been newly produced in the bioactive mutant extracts. Followed isolation and characterization demonstrated that five bioactive secondary metabolites, curvularin (1), citrinin (2), penicitrinone A (3), erythro-23-O-methylneocyclocitrinol (4) and 22E-7α-methoxy-5α,6α-epoxyergosta-8(14),22-dien-3β-ol (5), were newly produced by a mutant, 4-30, compared to the G59 strain. All 1–5 were also not yet found in the secondary metabolites of other wild type P. purpurogenum strains. Compounds 1–5 inhibited human cancer K562, HL-60, HeLa and BGC-823 cells to varying extents. Both present bioassays and chemical investigations demonstrated that the introduction of neomycin-resistance into the marine-derived fungal G59 strain could activate silent secondary metabolite production. The present work not only extended the previous DMSO-mediated method for introducing drug-resistance in fungi both in DMSO concentrations and antibiotics, but also additionally exemplified effectiveness of this method for activating silent fungal secondary metabolites. This method could be applied to other fungal isolates to elicit their metabolic potentials to investigate secondary metabolites from silent biosynthetic pathways. PMID:25913704

Activation of the silent secondary metabolite production by introducing neomycin-resistance in a marine-derived Penicillium purpurogenum G59.

PubMed

Wu, Chang-Jing; Yi, Le; Cui, Cheng-Bin; Li, Chang-Wei; Wang, Nan; Han, Xiao

2015-04-22

Introduction of neomycin-resistance into a marine-derived, wild-type Penicillium purpurogenum G59 resulted in activation of silent biosynthetic pathways for the secondary metabolite production. Upon treatment of G59 spores with neomycin and dimethyl sulfoxide (DMSO), a total of 56 mutants were obtained by single colony isolation. The acquired resistance of mutants to neomycin was testified by the resistance test. In contrast to the G59 strain, the EtOAc extracts of 28 mutants inhibited the human cancer K562 cells, indicating that the 28 mutants have acquired the capability to produce bioactive metabolites. HPLC-photodiode array detector (PDAD)-UV and HPLC-electron spray ionization (ESI)-MS analyses further indicated that diverse secondary metabolites have been newly produced in the bioactive mutant extracts. Followed isolation and characterization demonstrated that five bioactive secondary metabolites, curvularin (1), citrinin (2), penicitrinone A (3), erythro-23-O-methylneocyclocitrinol (4) and 22E-7α-methoxy-5α, 6α-epoxyergosta-8(14),22-dien-3β-ol (5), were newly produced by a mutant, 4-30, compared to the G59 strain. All 1-5 were also not yet found in the secondary metabolites of other wild type P. purpurogenum strains. Compounds 1-5 inhibited human cancer K562, HL-60, HeLa and BGC-823 cells to varying extents. Both present bioassays and chemical investigations demonstrated that the introduction of neomycin-resistance into the marine-derived fungal G59 strain could activate silent secondary metabolite production. The present work not only extended the previous DMSO-mediated method for introducing drug-resistance in fungi both in DMSO concentrations and antibiotics, but also additionally exemplified effectiveness of this method for activating silent fungal secondary metabolites. This method could be applied to other fungal isolates to elicit their metabolic potentials to investigate secondary metabolites from silent biosynthetic pathways.

Chromenoquinoline-based thiol probes: a study on the quencher position for controlling fluorescent Off-On characteristics.

PubMed

Kand, Dnyaneshwar; Kalle, Arunasree Marasanapalli; Talukdar, Pinaki

2013-02-13

The design, synthesis and thiol sensing ability of chromenoquinoline-based fluorescent probes 4, 5 and 6 and are reported here. The relative position of the maleimide moiety was varied along the chromenoquinoline fluorophore to decrease the background fluorescence. Lower background fluorescence in probes 4 and 6 was rationalized by the smaller k(r)/k(nr) values compared to that of probe 5. An intramolecular charge transfer (ICT) mechanism was proposed for quenching and the extent was dependent on the position of the maleimide quencher. Fluorescent Off-On characteristics were evaluated by theoretical calculations. All probes were selective only towards thiol containing amino acids. Thiol sensing by probes 4 and 6 were much better compared to 5. Probe 4 displayed a better fluorescence response for less hindered thiol (185-, 223- and 156-fold for Hcy, Cys and GSH, respectively), while for probe 6, a higher enhancement in fluorescence was observed with more hindered thiols (180-, 205- and 245-fold for Hcy, Cys and GSH, respectively). The better response to bulkier thiol, GSH by probe 6 was attributed to the steric crowding at the C-4 position and bulkiness of the GSH group which force the succinimide unit to be in a nearly orthogonal conformation. This spatial arrangement was important in reducing the fluorescence quenching ability of the succinimide moiety. The application of probes 4, 5 and 6 was demonstrated by naked eye detection thiols using a 96-well plate system as well as by live-cell imaging.

Selective chloroform sensor using thiol functionalized reduced graphene oxide at room temperature

NASA Astrophysics Data System (ADS)

Midya, Anupam; Mukherjee, Subhrajit; Roy, Shreyasee; Santra, Sumita; Manna, Nilotpal; Ray, Samit K.

2018-02-01

This paper presents a highly selective chloroform sensor using functionalised reduced graphene oxide (RGO) as a sensing layer. Thiol group is covalently attached on the basal plan of RGO film by a simple one-step aryl diazonium chemistry to improve its selectivity. Several spectroscopic techniques like X-ray photoelectron, Raman and Fourier transform infrared spectroscopy confirm successful thiol functionalization of RGO. Finally, the fabricated chemiresistor type sensor is exposed to chloroform in the concentration range 200-800 ppm (parts per million). The sensor shows a 4.3% of response towards 800 ppm chloroform. The selectivity of the sensor is analyzed using various volatile organic compounds as well. The devices show enhanced response and faster recovery attributed to the physiosorption of chloroform onto thiol functionalized graphene making them attractive for 2D materials based sensing applications.

Enhancing Electrophoretic Display Lifetime: Thiol-Polybutadiene Evaporation Barrier Property Response to Network Microstructure

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

Cook, Caitlyn Christian

An evaporation barrier is required to enhance the lifetime of electrophoretic deposition (EPD) displays. As EPD functions on the basis of reversible deposition and resuspension of colloids suspended in a solvent, evaporation of the solvent ultimately leads to device failure. Incorporation of a thiol-polybutadiene elastomer into EPD displays enabled display lifetime surpassing six months in counting and catalyzed rigid display transition into a flexible package. Final flexible display transition to mass production compels an electronic-ink approach to encapsulate display suspension within an elastomer shell. Final thiol-polybutadiene photosensitive resin network microstructure was idealized to be dense, homogeneous, and expose an elasticmore » response to deformation. Research at hand details an approach to understanding microstructural change within display elastomers. Polybutadiene-based resin properties are modified via polymer chain structure, with and without added aromatic urethane methacrylate difunctionality, and in measuring network response to variation in thiol and initiator concentration. Dynamic mechanical analysis results signify that cross-linked segments within a difunctionalized polybutadiene network were on average eight times more elastically active than that of linked segments within a non-functionalized polybutadiene network. Difunctionalized polybutadiene samples also showed a 2.5 times greater maximum elastic modulus than non-functionalized samples. Hybrid polymer composed of both polybutadiene chains encompassed TE-2000 stiffness and B-1000 elasticity for use in encapsulating display suspension. Later experiments measured kinetic and rheological response due to alteration in dithiol cross-linker chain length via real time Fourier transform infrared spectroscopy and real-time dynamic rheology. Distinct differences were discovered between dithiol resin systems, as maximum thiol conversion achieved in short and long chain length dithiols was

Dynamic Thiol/Disulphide Homeostasis in Children and Adolescents with Non-Autoimmune Subclinical Hypothyroidism

PubMed Central

Uçaktürk, Seyit Ahmet; Alışık, Murat; Uğur, Çağatay; Elmaoğulları, Selin; Mengen, Eda; Erel, Özcan

2018-01-01

Objective To evaluate the thiol/disulphide homeostasis in children with non-autoimmune subclinical hypothyroidism (SHT). Subjects and Methods Thiol/disulphide homeosta sis, involving native thiol (SH), disulphide (SS), and total thiol (SS + SH), was evaluated in 60 children and adolescents who were negative for thyroid auto-antibodies (anti-thyroid peroxidase, anti-thyroglobulin) and had a thyroid-stimulating hormone (TSH) value of > 5 mIU/L, and in 40 sex- and age-matched healthy control subjects who were negative for thyroid autoantibodies and had normal TSH levels. Lipid profiles and urine iodine levels were also determined. Results SH (466 ± 32.8 vs. 462 ± 32.1 μmol/L p = 0.59), SH + SS (508 ± 34.0 vs. 506 ± 32.7 μmol/L, p = 0.81), SS (21 ± 5.5 vs. 22 ± 5.8 μmol/L, p = 0.41), SS/SH (4.5 ± 1.2 vs. 4.8 ± 1.3%, p = 0.36), SS/SH + SS (4.1 ± 1.0 vs. 4.3 ± 1.1%, p = 0.36) and SH/SH + SS (91 ± 2.1 vs. 91 ± 2.1%, p = 0.31) levels were similar in children with SHT and control subjects (p > 0.05). There was no difference between total cholesterol, triglyceride, and low-density lipoprotein levels in SHT patients and controls. No difference was detected between the patients with or without iodine deficiency in the SHT group in terms of thiol/disulphide homeostasis parameters. Conclusion The status of dynamic thiol/disulphide homeostasis did not change in children and adolescents with non-autoimmune SHT. Future studies are needed for the evaluation of oxidative stress in patients with long-standing non-autoimmune SHT. PMID:29402856

Stable isotope labeling-solid phase extraction-mass spectrometry analysis for profiling of thiols and aldehydes in beer.

PubMed

Zheng, Shu-Jian; Wang, Ya-Lan; Liu, Ping; Zhang, Zheng; Yu, Lei; Yuan, Bi-Feng; Feng, Yu-Qi

2017-12-15

In this study, we developed a strategy for profiling of thiols and aldehydes in beer samples by stable isotope labeling-solid phase extraction-liquid chromatography-double precursor ion scan/double neutral loss scan-mass spectrometry analysis (SIL-SPE-LC-DPIS/DNLS-MS). A pair of isotope reagents (ω-bromoacetonylquinolinium bromide, BQB; ω-bromoacetonylquinolinium-d 7 bromide, BQB-d 7 ) were used to label thiols; while for the aldehydes, a pair of isotope reagents (4-(2-(trimethylammonio) ethoxy) benzenaminium halide, 4-APC; 4-(2-(trimethylammonio) ethoxy) benzenaminium halide-d 4 , 4-APC-d 4 ) were used. The labeled thiols and aldehydes were extracted and purified with solid-phase extraction, respectively, followed by LC-MS analysis. Using the proposed SIL-SPE-LC-DPIS/DNLS-MS methods, 76 thiol and 25 aldehyde candidates were found in beer. Furthermore, we established SIL-SPE-LC-MRM-MS methods for the relative quantitation of thiols and aldehydes in different beer samples. The results showed that the contents of thiols and aldehydes are closely related to the brands and origins of beers. Copyright © 2017 Elsevier Ltd. All rights reserved.

The chemical foundations of nitroalkene fatty acid signaling through addition reactions with thiols.

PubMed

Turell, Lucía; Steglich, Martina; Alvarez, Beatriz

2018-03-22

Nitroalkene fatty acids can be formed in vivo and administered exogenously. They exert pleiotropic signaling actions with cytoprotective and antiinflammatory effects. The presence of the potent electron withdrawing nitro group confers electrophilicity to the adjacent β-carbon. Thiols (precisely, thiolates) are strong nucleophiles and can react with nitroalkene fatty acids through reversible Michael addition reactions. In addition, nitroalkene fatty acids can undergo several other processes including metabolic oxidation, reduction, esterification, nitric oxide release and partition into hydrophobic compartments. The signaling actions of nitroalkenes are mainly mediated by reactions with critical thiols in regulatory proteins. Thus, the thio-Michael addition reaction provides a framework for understanding the molecular basis of the biological effects of nitroalkene fatty acids at the crossroads of thiol signaling and electrophilic lipid signaling. In this review, we describe the reactions of nitroalkene fatty acids in biological contexts. We focus on the Michael addition-elimination reaction with thiols and its mechanism, and extrapolate kinetic and thermodynamic considerations to in vivo settings. Copyright © 2018 Elsevier Inc. All rights reserved.

Selective chromogenic detection of thiol-containing biomolecules using carbonaceous nanospheres loaded with silver nanoparticles as carrier.

PubMed

Hu, Bo; Zhao, Yang; Zhu, Hai-Zhou; Yu, Shu-Hong

2011-04-26

Thiol-containing biomolecules show strong affinity with noble metal nanostructures and could not only stably protect them but also control the self-assembly process of these special nanostructures. A highly selective and sensitive chromogenic detection method has been designed for the low and high molecular weight thiol-containing biomolecules, including cysteine, glutathione, dithiothreitol, and bovine serum albumin, using a new type of carbonaceous nanospheres loaded with silver nanoparticles (Ag NPs) as carrier. This strategy relies upon the place-exchange process between the reporter dyes on the surface of Ag NPs and the thiol groups of thiol-containing biomolecules. The concentration of biomolecules can be determined by monitoring with the fluorescence intensity of reporter dyes dispersed in solution. This new chromogenic assay method could selectively detect these biomolecules in the presence of various other amino acids and monosaccharides and even sensitively detect the thiol-containing biomolecules with different molecular weight, even including proteins.

Impact of limited solvent capacity on metabolic rate, enzyme activities, and metabolite concentrations of S. cerevisiae glycolysis.

PubMed

Vazquez, Alexei; de Menezes, Marcio A; Barabási, Albert-László; Oltvai, Zoltan N

2008-10-01

The cell's cytoplasm is crowded by its various molecular components, resulting in a limited solvent capacity for the allocation of new proteins, thus constraining various cellular processes such as metabolism. Here we study the impact of the limited solvent capacity constraint on the metabolic rate, enzyme activities, and metabolite concentrations using a computational model of Saccharomyces cerevisiae glycolysis as a case study. We show that given the limited solvent capacity constraint, the optimal enzyme activities and the metabolite concentrations necessary to achieve a maximum rate of glycolysis are in agreement with their experimentally measured values. Furthermore, the predicted maximum glycolytic rate determined by the solvent capacity constraint is close to that measured in vivo. These results indicate that the limited solvent capacity is a relevant constraint acting on S. cerevisiae at physiological growth conditions, and that a full kinetic model together with the limited solvent capacity constraint can be used to predict both metabolite concentrations and enzyme activities in vivo.

Impact of Limited Solvent Capacity on Metabolic Rate, Enzyme Activities, and Metabolite Concentrations of S. cerevisiae Glycolysis

PubMed Central

Vazquez, Alexei; de Menezes, Marcio A.; Barabási, Albert-László; Oltvai, Zoltan N.

2008-01-01

The cell's cytoplasm is crowded by its various molecular components, resulting in a limited solvent capacity for the allocation of new proteins, thus constraining various cellular processes such as metabolism. Here we study the impact of the limited solvent capacity constraint on the metabolic rate, enzyme activities, and metabolite concentrations using a computational model of Saccharomyces cerevisiae glycolysis as a case study. We show that given the limited solvent capacity constraint, the optimal enzyme activities and the metabolite concentrations necessary to achieve a maximum rate of glycolysis are in agreement with their experimentally measured values. Furthermore, the predicted maximum glycolytic rate determined by the solvent capacity constraint is close to that measured in vivo. These results indicate that the limited solvent capacity is a relevant constraint acting on S. cerevisiae at physiological growth conditions, and that a full kinetic model together with the limited solvent capacity constraint can be used to predict both metabolite concentrations and enzyme activities in vivo. PMID:18846199

Major secondary metabolites produced by two commercial Trichoderma strains active against different phytopathogens.

PubMed

Vinale, F; Marra, R; Scala, F; Ghisalberti, E L; Lorito, M; Sivasithamparam, K

2006-08-01

Trichoderma harzianum strains T22 and T39 are two micro-organisms used as active agents in a variety of commercial biopesticides and biofertilizers and widely applied amongst field and greenhouse crops. The production, isolation, biological and chemical characterization of the main secondary metabolites produced by these strains are investigated. Of the three major compounds produced by strain T22, one is a new azaphilone that shows marked in vitro inhibition of Rhizoctonia solani, Pythium ultimum and Gaeumannomyces graminis var. tritici. In turn, filtrates from strain T39 were demonstrated to contain two compounds previously isolated from other T. harzianum strains and a new butenolide. The production of the isolated metabolites was also monitored by liquid chromatography/mass spectrometry during in vitro interaction with R. solani. This paper reports the isolation and characterization of the main secondary metabolites obtained from culture filtrates of two T. harzianum strains and their production during antagonistic interaction with the pathogen R. solani. This is the first work on secondary metabolites produced by the commercially applied strains T22 and T39. Our results provide a better understanding of the metabolism of these fungi, which are both widely used as biopesticides and/or biofertilizers in biocontrol.

The urinary metabolites of DINCH® have an impact on the activities of the human nuclear receptors ERα, ERβ, AR, PPARα and PPARγ.

PubMed

Engel, Anika; Buhrke, Thorsten; Kasper, Stefanie; Behr, Anne-Cathrin; Braeuning, Albert; Jessel, Sönke; Seidel, Albrecht; Völkel, Wolfgang; Lampen, Alfonso

2018-05-01

DINCH ® (di-isononyl cyclohexane-1,2-dicarboxylate) is a non-phthalate plasticizer that has been developed to replace phthalate plasticizers such as DEHP (di-2-ethylhexyl phthalate) or DINP (di-isononyl phthalate). DINCH ® is metabolized to its corresponding monoester and subsequently to oxidized monoester derivatives. These are conjugated to glucuronic acid and subject to urinary excretion. In contrast to DINCH ® , there are almost no toxicological data available regarding its primary and secondary metabolites. The present study aimed at the characterization of potential endocrine properties of DINCH ® and five DINCH ® metabolites by using reporter gene assays to monitor the activity of the human nuclear receptors ERα, ERβ, AR, PPARα and PPARγ in vitro. DINCH ® itself did not have any effect on the activity of these receptors whereas DINCH ® metabolites were shown to activate all these receptors. In the case of AR, DINCH ® metabolites predominantly enhanced dihydrotestosterone-stimulated AR activity. In the H295R steroidogenesis assay, neither DINCH ® nor any of its metabolites affected estradiol or testosterone synthesis. In conclusion, primary and secondary DINCH ® metabolites exert different effects at the molecular level compared to DINCH ® itself. All these in vitro effects of DINCH ® metabolites, however, were only observed at high concentrations such as 10 μM or above which is about three orders of magnitude above reported DINCH ® metabolite concentrations in human urine. Thus, the in vitro data do not support the notion that DINCH ® or any of the investigated metabolites may exert considerable endocrine effects in vivo at relevant human exposure levels. Copyright © 2018 Elsevier B.V. All rights reserved.

Silver Makes Better Electrical Contacts to Thiol-Terminated Silanes than Gold.

PubMed

Li, Haixing; Su, Timothy A; Camarasa-Gómez, María; Hernangómez-Pérez, Daniel; Henn, Simon E; Pokorný, Vladislav; Caniglia, Caravaggio D; Inkpen, Michael S; Korytár, Richard; Steigerwald, Michael L; Nuckolls, Colin; Evers, Ferdinand; Venkataraman, Latha

2017-11-06

We report that the single-molecule junction conductance of thiol-terminated silanes with Ag electrodes are higher than the conductance of those formed with Au electrodes. These results are in contrast to the trends in the metal work function Φ(Ag)<Φ(Au). As such, a better alignment of the Au Fermi level to the molecular orbital of silane that mediates charge transport would be expected. This conductance trend is reversed when we replace the thiols with amines, highlighting the impact of metal-S covalent and metal-NH 2 dative bonds in controlling the molecular conductance. Density functional theory calculations elucidate the crucial role of the chemical linkers in determining the level alignment when molecules are attached to different metal contacts. We also demonstrate that conductance of thiol-terminated silanes with Pt electrodes is lower than the ones formed with Au and Ag electrodes, again in contrast to the trends in the metal work-functions. © 2017 Wiley-VCH Verlag GmbH & Co. KGaA, Weinheim.

Mutagenic activity of austocystins - secondary metabolites of Aspergillus ustus

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

Kfir, R.; Johannsen, E.; Vleggaar, R.

1986-11-01

Mycotoxins constitute a group of toxic secondary fungal metabolites. Fungi that produce these toxins frequently contaminate food and feed, creating a potential threat to human and animal health. Biological activities of mycotoxins include, amongst others: toxicity, mutagenicity and carcinogenicity, which can be expressed with or without metabolic activation. Austocystins are similar in structure to aflatoxin B/sup 1/ and are probably synthesized in a similar manner. The Ames Salmonella test, a widely accepted method employed for the detection of mutagenic activity of various chemical compounds was used for testing the mutagenic activity of different mycotoxins. As aflatoxin B/sup 1/ was foundmore » by the Ames test to be highly mutagenic, the same test was applied for the study of possible mutagenicity of the austocystins. The mutagenic activity of these compounds was studied with and without metabolic activation using two tester strains of S. typhimurium, one capable of detecting frame shift mutation (strain TA98) and the other capable of detecting base pair substitution (strain TA100).« less

Simultaneous LC/MS/MS determination of thiols and disulfides in urine samples based on differential labeling with ferrocene-based maleimides.

PubMed

Seiwert, Bettina; Karst, Uwe

2007-09-15

A method for the simultaneous determination of a series of thiols and disulfides in urine samples has been developed based on the sequential labeling of free and bound thiol functionalities with two ferrocene-based maleimide reagents. The sample is first exposed to N-(2-ferroceneethyl)maleimide, thus leading to the derivatization of free thiol groups in the sample. After quantitative reaction and subsequent reduction of the disulfide-bound thiols by tris(2-carboxyethyl)phosphine, the newly formed thiol functionalities are reacted with ferrocenecarboxylic acid-(2-maleimidoyl)ethylamide. The reaction products are determined by LC/MS/MS in the multiple reaction mode, and precursor ion scan as well as neutral loss scan is applied to detect unknown further thiols. The method was successfully applied to the analysis of free and disulfide-bound thiols in urine samples. Limits of detection are 30 to 110 nM, and the linear range comprises two decades of concentration, thus covering the relevant concentration range of thiols in urine samples. The thiol and disulfide concentrations were referred to the creatinine content to compensate for different sample volumes. As some calibration standards for the disulfides are not commercially available, they were synthesized in an electrochemical flow-through cell. This allowed the synthesis of hetero- and homodimeric disulfides.

Eucalyptus tolerance mechanisms to lanthanum and cerium: subcellular distribution, antioxidant system and thiol pools.

PubMed

Shen, Yichang; Zhang, Shirong; Li, Sen; Xu, Xiaoxun; Jia, Yongxia; Gong, Guoshu

2014-12-01

Guanglin 9 (Eucalyptus grandis × Eucalyptus urophlla) and Eucalyptus grandis 5 are two eucalyptus species which have been found to grow normally in soils contaminated with lanthanum and cerium, but the tolerance mechanisms are not clear yet. In this study, a pot experiment was conducted to investigate the tolerance mechanisms of the eucalyptus to lanthanum and cerium. Cell walls stored 45.40-63.44% of the metals under lanthanum or cerium stress. Peroxidase and catalase activities enhanced with increasing soil La or Ce concentrations up to 200 mg kg(-1), while there were no obvious changes in glutathione and ascorbate concentrations. Non-protein thiols concentrations increased with increasing treatment levels up to 200 mg kg(-1), and then decreased. Phytochelatins concentrations continued to increase under La or Ce stress. Therefore, the two eucalyptus species are La and Ce tolerant plants, and the tolerance mechanisms include cell wall deposition, antioxidant system response, and thiol compound synthesis. Copyright © 2014 Elsevier Ltd. All rights reserved.

[The pharmacokinetics of the dipeptide analog of piracetam with nootropic activity GVS-111 and of its basic metabolites].

PubMed

Boĭko, S S; Zherdev, V P; Dvorianinov, A A; Gudasheva, T A; Ostrovskaia, R U; Voronina, T A; Rozantsev, G G; Seredenin, S B

1997-01-01

The pharmacokinetics of a new nootropic dipeptide analog of piracetam-N-phenylacetyl-L-prolylglycine (GWS-111) and its main metabolites were studied in rats by means of high performance liquid chromatography and gas-liquid chromatography. The compound under study showed a greater resistance to an enzymatic effect than natural neuropeptides. In addition to an unchanged compound three of its metabolites were found in the blood plasma of the rats. One of them, cyclo-Pro-Gly was an active metabolite of GWS-111.

Porritoxins, metabolites of Alternaria porri, as anti-tumor-promoting active compounds.

PubMed

Horiuchi, Masayuki; Tokuda, Harukuni; Ohnishi, Keiichiro; Yamashita, Masakazu; Nishino, Hoyoku; Maoka, Takashi

2006-02-01

To search for possible cancer chemopreventive agents from natural sources, we performed primary screening of metabolites of Alternaria porri by examining their possible inhibitory effects on Epstein-Barr virus early antigen (EBV-EA) activation induced by 12-O-tetradecanoylphorbol-13-acetate (TPA) in Raji cells. The ethyl acetate extract of A. porri showed the inhibitory effect on EBV-EA activation. Three porritoxins (1-3) were obtained as inhibitory active compounds for EBV-EA from ethyl acetate extract. 6-(3',3'-Dimethylallyloxy)-4-methoxy-5-methylphthalide (2) showed the strongest activity among them. Inhibitory effect of porritoxin (1) and (2) was superior to that of beta-carotene, a well-known anti-tumor promoter. Furthermore, the structure-activity correlation of porritoxins and their related compounds were discussed.

EPR Characterization of Dinitrosyl Iron Complexes with Thiol-Containing Ligands as an Approach to Their Identification in Biological Objects: An Overview.

PubMed

Vanin, Anatoly F

2018-06-01

The overview demonstrates how the use of only one physico-chemical approach, viz., the electron paramagnetic resonance method, allowed detection and identification of dinitrosyl iron complexes with thiol-containing ligands in various animal and bacterial cells. These complexes are formed in biological objects in the paramagnetic (electron paramagnetic resonance-active) mononuclear and diamagnetic (electron paramagnetic resonance-silent) binuclear forms and control the activity of nitrogen monoxide, one of the most universal regulators of metabolic processes in the organism. The analysis of electronic and spatial structures of dinitrosyl iron complex sheds additional light on the mechanism whereby dinitrosyl iron complex with thiol-containing ligands function in human and animal cells as donors of nitrogen monoxide and its ionized form, viz., nitrosonium ions (NO + ).

Microbial transformation of (+)-nootkatone and the antiproliferative activity of its metabolites.

PubMed

Gliszczyńska, Anna; Łysek, Agnieszka; Janeczko, Tomasz; Świtalska, Marta; Wietrzyk, Joanna; Wawrzeńczyk, Czesław

2011-04-01

Six metabolites were obtained as a result of microbial transformation of (+)-nootkatone (1) by the fungal strains: Botrytis, Didymosphaeria, Aspergillus, Chaetomium and Fusarium. Their structure were established as (+)-(4R,5S,7R,9R)-9α-hydroxynootkatone (2), (+)-(4R,5S,7R)-13-hydroxynootkatone (3) and (+)-(4R,5S,7R,9R,11S)-11,12-epoxy-9α-hydroxynootkatone (4), (+)-(4R,5S,7R,11S)-11,12-epoksynootkatone (5), (+)-(4R,5S,7R)-11,12-dihydroxynootkatone (6) and (+)-(4R,5S,7R)-7,11,12-trihydroxynootkatone (7) on the basis of their spectral data. Two products: (4) and (7) were not previously reported in the literature. The antiproliferative activity of (+)-nootkatone (1) and isolated metabolites (2-7) of its biotransformation has been evaluated. Copyright © 2011 Elsevier Ltd. All rights reserved.

Nitroolefin-based BODIPY as a novel water-soluble ratiometric fluorescent probe for detection of endogenous thiols

NASA Astrophysics Data System (ADS)

Kang, Jin; Huo, Fangjun; Chao, Jianbin; Yin, Caixia

2018-04-01

Small molecule biothiols, including cysteine (Cys), homocysteine (Hcy), and glutathione (GSH), play many crucial roles in physiological processes. In this work, we have prepared a nitroolefin-based BODIPY fluorescent probe with excellent water solubility for detection thiols, which displayed ratiometric fluorescent signal for thiols. Incorporation of a nitroolefin unit to the BODIPY dye would transform it into a strong Michael acceptor, which would be highly susceptible to sulfhydryl nucleophiles. This probe shows an obvious ratio change upon response with thiols, an increase of the emission at 517 nm along with a concomitant decrease of fluorescence peak at 573 nm. Moreover, these successes of intracellular imaging experiments in A549 cells indicated that this probe is suitable for imaging of ex-/endogenous thiols in living cells.

Passage of irinotecan and its active metabolite, SN-38, into human milk.

PubMed

Nakagawa, J; Terui, K; Hosoi, K; Ueno, K; Yokoyama, Y; Hayakari, M

2016-10-01

We measured the levels of irinotecan and its active metabolite, SN-38, in human milk after the administration of irinotecan to assess the potential risks when women treated with irinotecan nurse their infants. Human milk was collected for 6 days starting on the day after irinotecan was administered. The levels of irinotecan and SN-38 in human milk were measured using liquid chromatography-mass spectrometry. Irinotecan was detected on Days 2 and 3 but not after Day 4. A strong signal indicating the presence of SN-38 was detected on Day 2 and the signal was readily detected until Day 7, indicating that SN-38 remained in human milk. Intravenously administered CPT-11 continues to pass into human milk over a prolonged period in the form of its active metabolite, SN-38. The relationship between administration of CPT-11 and SN-38 exposure and toxicity is still not well defined, so patients should avoid nursing their infants while they are being treated with CPT-11. © 2016 John Wiley & Sons Ltd.

Regulatory requirements for genotoxicity assessment of plant protection product active ingredients, impurities, and metabolites.

PubMed

Booth, Ewan D; Rawlinson, Paul J; Maria Fagundes, Priscila; Leiner, Kevin A

2017-06-01

Active ingredients in plant protection products are subject to rigorous safety assessment during their development, including assessment of genotoxicity. Plant protection products are used for agriculture in multiple regions and for the registration of active ingredients it is necessary to satisfy the data requirements of these different regions. There are no overarching global agreements on which genotoxicity studies need to be conducted to satisfy the majority of regulatory authorities. The implementation of new OECD guidelines for the in vitro micronucleus, transgenic rodent somatic and germ cell gene mutation and in vivo comet assays, as well as the revision of a number of other OECD test guidelines has resulted in some changes to data requirements. This review describes the genotoxicity data requirements for chemical active ingredients as well as biologicals, microbials, ground water metabolites, metabolites, and impurities in a number of regions. Similarities and differences are highlighted. Environ. Mol. Mutagen. 58:325-344, 2017. © 2017 Wiley Periodicals, Inc. © 2017 Wiley Periodicals, Inc.

Quinone-induced protein modifications: Kinetic preference for reaction of 1,2-benzoquinones with thiol groups in proteins.

PubMed

Li, Yuting; Jongberg, Sisse; Andersen, Mogens L; Davies, Michael J; Lund, Marianne N

2016-08-01

Oxidation of polyphenols to quinones serves as an antioxidative mechanism, but the resulting quinones may induce damage to proteins as they react through a Michael addition with nucleophilic groups, such as thiols and amines to give protein adducts. In this study, rate constants for the reaction of 4-methylbenzoquinone (4MBQ) with proteins, thiol and amine compounds were determined under pseudo first-order conditions by UV-vis stopped-flow spectrophotometry. The chemical structures of the adducts were identified by LC-ESI-MS/MS. Proteins with free thiols were rapidly modified by 4MBQ with apparent second order rate constants, k2 of (3.1±0.2)×10(4)M(-1)s(-1) for bovine serum albumin (BSA) and (4.8±0.2)×10(3)M(-1)s(-1) for human serum albumin at pH 7.0. These values are at least 12-fold greater than that for α-lactalbumin (4.0±0.2)×10(2)M(-1)s(-1), which does not contain any free thiols. Reaction of Cys-34 of BSA with N-ethylmaleimide reduced the thiol concentration by ~59%, which resulted in a decrease in k2 by a similar percentage, consistent with rapid adduction at Cys-34. Reaction of 4MBQ with amines (Gly, Nα-acetyl-l-Lys, Nε-acetyl-l-Lys and l-Lys) and the guanidine group of Nα-acetyl-l-Arg was at least 5×10(5) slower than with low-molecular-mass thiols (l-Cys, Nα-acetyl-l-Cys, glutathione). The thiol-quinone interactions formed colorless thiol-phenol products via an intermediate adduct, while the amine-quinone interactions generated colored amine-quinone products that require oxygen involvement. These data provide strong evidence for rapid modification of protein thiols by quinone species which may be of considerable significance for biological and food systems. Copyright © 2016 Elsevier Inc. All rights reserved.

Fluorimetric determination of some thiol compounds in their dosage forms.

PubMed

Al-Ghannam, Sh M; El-Brashy, A M; Al-Farhan, B S

2002-08-01

A simple fluorimetric procedure was adopted for determination of three pharmaceutical compounds containing thiol groups namely, captopril, D-penicillamine and N-acetylcysteine. In this method, the drugs are treated with 1,2-naphthoquinone-4-sulfonic acid. The latter is reduced to 1,2-dihydroxynaphthalene-4-sulfonic acid which has a maximum fluorescence intensity at 480/318 nm (lambdaEm/Ex). The method is sensitive to 0.5-4.5 pg ml(- 1) with minimum detectability 0.05 microg ml(-1) (S/N = 2), and has been applied to determine these three thiols in their dosage forms. The results obtained are compared favourably with those obtained by their pharmacopeial methods.

Topography of Escherichia coli ribosomal proteins. The order of reactivity of thiol groups*

PubMed Central

Bakardjieva, Anastasia; Crichton, Robert R.

1974-01-01

1. 30S and 50S ribosomal subunits of Escherichia coli were treated with N-[2,3-14C]-ethylmaleimide and iodo[14C]acetamide. 2. The proteins in the native subunits which reacted with the reagents were S1,‡ S2, S12, S13, S18, S21, L2, L5, L6, L10, L11, L15, L17, L20, L26+28 and L27. 3. Several proteins, such as S1, S12, S14, S18, L2, L6, L10, L11 and either L26 or 28, had thiol groups in an oxidized form and reacted to a greater extent after reduction with β-mercaptoethanol or dithiothreitol. 4. The total number of thiol groups in 30S and 50S subunits was determined as 16–17 and 26–27 respectively. The total number of thiol groups in each ribosomal protein was also determined. 5. The reaction of 30S and 50S subunits with iodoacetamide under several different conditions established the order of reactivity of thiol groups. PMID:4618476

Molecular Characterization of Thiols in Fossil Fuels by Michael Addition Reaction Derivatization and Electrospray Ionization Fourier Transform Ion Cyclotron Resonance Mass Spectrometry.

PubMed

Wang, Meng; Zhao, Suoqi; Liu, Xuxia; Shi, Quan

2016-10-04

Thiols widely occur in sediments and fossil fuels. However, the molecular composition of these compounds is unclear due to the lack of appropriate analytical methods. In this work, a characterization method for thiols in fossil fuels was developed on the basis of Michael addition reaction derivatization followed by electrospray ionization Fourier transform ion cyclotron resonance mass spectrometry (ESI FT-ICR MS). Model thiol compound studies showed that thiols were selectively reacted with phenylvinylsulfone and transformed to sulfones with greater than 98% conversions. This method was applied to a coker naphtha, light and heavy gas oils, and crude oils from various geological sources. The results showed that long alkyl chain thiols are readily present in petroleum, which have up to 30 carbon atoms. Large DBE dispersity of thiols indicates that naphthenic and aromatic thiols are also present in the petroleum. This method is capable of detecting thiol compounds in the part per million range by weight. This method allows characterization of thiols in a complex hydrocarbon matrix, which is complementary to the comprehensive analysis of sulfur compounds in fossil fuels.

Differential effects of clinically used derivatives and metabolites of artemisinin in the activation of constitutive androstane receptor isoforms

PubMed Central

Burk, O; Piedade, R; Ghebreghiorghis, L; Fait, JT; Nussler, AK; Gil, JP; Windshügel, B; Schwab, M

2012-01-01

BACKGROUND AND PURPOSE Widespread resistance to antimalarial drugs requires combination therapies with increasing risk of pharmacokinetic drug–drug interactions. Here, we explore the capacity of antimalarial drugs to induce drug metabolism via activation of constitutive androstane receptors (CAR) by ligand binding. EXPERIMENTAL APPROACH A total of 21 selected antimalarials and 11 major metabolites were screened for binding to CAR isoforms using cellular and in vitro CAR-coactivator interaction assays, combined with in silico molecular docking. Identified ligands were further characterized by cell-based assays and primary human hepatocytes were used to elucidate induction of gene expression. KEY RESULTS Only two artemisinin derivatives arteether and artemether, the metabolite deoxyartemisinin and artemisinin itself demonstrated agonist binding to the major isoforms CAR1 and CAR3, while arteether and artemether were also inverse agonists of CAR2. Dihydroartemisinin and artesunate acted as weak inverse agonists of CAR1. While arteether showed the highest activities in vitro, it was less active than artemisinin in inducing hepatic CYP3A4 gene expression in hepatocytes. CONCLUSIONS AND IMPLICATIONS Artemisinin derivatives and metabolites differentially affect the activities of CAR isoforms and of the pregnane X receptor (PXR). This negates a common effect of these drugs on CAR/PXR-dependent induction of drug metabolism and further provides an explanation for artemisinin consistently inducing cytochrome P450 genes in vivo, whereas arteether and artemether do not. All these drugs are metabolized very rapidly, but only artemisinin is converted to an enzyme-inducing metabolite. For better understanding of pharmacokinetic drug–drug interaction possibilities, the inducing properties of artemisinin metabolites should be considered. PMID:22577882

Bromo- and thiomaleimides as a new class of thiol-mediated fluorescence 'turn-on' reagents.

PubMed

Youziel, Judith; Akhbar, Ahmed R; Aziz, Qadeer; Smith, Mark E B; Caddick, Stephen; Tinker, Andrew; Baker, James R

2014-01-28

Bromo- and thiomaleimides are shown to serve as highly effective quenchers of a covalently attached fluorophore. Reactions with thiols that lead to removal of the maleimide conjugation, or detachment of the fluorophore from the maleimide, result in 'turn-on' of the fluorescence. These reagents thus offer opportunities in thiol sensing and intracellular reporting.

Octadecyl Chains Immobilized onto Hyaluronic Acid Coatings by Thiol-ene "Click Chemistry" Increase the Surface Antimicrobial Properties and Prevent Platelet Adhesion and Activation to Polyurethane.

PubMed

Felgueiras, Helena P; Wang, L M; Ren, K F; Querido, M M; Jin, Q; Barbosa, M A; Ji, J; Martins, M C L

2017-03-08

Infection and thrombus formation are still the biggest challenges for the success of blood contact medical devices. This work aims the development of an antimicrobial and hemocompatible biomaterial coating through which selective binding of albumin (passivant protein) from the bloodstream is promoted and, thus, adsorption of other proteins responsible for bacterial adhesion and thrombus formation can be prevented. Polyurethane (PU) films were coated with hyaluronic acid, an antifouling agent, that was previously modified with thiol groups (HA-SH), using polydopamine as the binding agent. Octadecyl acrylate (C18) was used to attract albumin since it resembles the circulating free fatty acids and albumin is a fatty acid transporter. Thiol-ene "click chemistry" was explored for C18 immobilization on HA-SH through a covalent bond between the thiol groups from the HA and the alkene groups from the C18 chains. Surfaces were prepared with different C18 concentrations (0, 5, 10, and 20%) and successful immobilization was demonstrated by scanning electron microscopy (SEM), water contact angle determinations, X-ray photoelectron spectroscopy (XPS) and Fourier transform infrared spectroscopy (FTIR). The ability of surfaces to bind albumin selectively was determined by quartz crystal microbalance with dissipation (QCM-D). Albumin adsorption increased in response to the hydrophobic nature of the surfaces, which augmented with C18 saturation. HA-SH coating reduced albumin adsorption to PU. C18 immobilized onto HA-SH at 5% promoted selective binding of albumin, decreased Staphylococcus aureus adhesion and prevented platelet adhesion and activation to PU in the presence of human plasma. C18/HA-SH coating was established as an innovative and promising strategy to improve the antimicrobial properties and hemocompatibility of any blood contact medical device.

Comparison of pharmacokinetics of loxoprofen and its active metabolites after an intravenous, intramuscular, and oral administration of loxoprofen in rats: evidence for extrahepatic metabolism.

PubMed

Koo, Tae-Sung; Kim, Dae-Hyun; Ahn, Sung-Hoon; Kim, Kang-Pil; Kim, In-Wha; Seo, Seung-Yong; Suh, Young-Ger; Kim, Dae-Duk; Shim, Chang-Koo; Chung, Suk-Jae

2005-10-01

The objective of this study was to characterize the extent of the formation of the active (trans-alcohol form) and inactive (cis-alcohol) metabolites of loxoprofen and to compare the kinetics after its intragastric, intravenous, and intramuscular administrations in rats. After intravenous administration of the drug at doses of 5-20 mg/kg, the clearance and the volume of distribution for loxoprofen, and the ratios of the AUC for the metabolites to the parent drug were not statistically different with the dosage; the formation clearances were 1.08 and 0.87 mL/min/kg for the active and its isomeric metabolite, respectively. After the intragastric, intravenous, or intramuscular administration, AUC for loxoprofen and the metabolites at a dose of 10 mg/kg were not statistically different for the different routes of administration. The formation of the metabolites with the concomitant loss of loxoprofen was found in incubations with liver homogenates and blood but not with a muscle homogenate or plasma, indicating that the conversion of loxoprofen to the metabolites may occur both in the liver and extraheptic tissue(s). Thus, approximately 22% of the loxoprofen may have been converted to the active metabolite in the liver and the extraheptic tissue(s) and the pharmacokinetics of the active metabolite was independent of the route of administration. Copyright (c) 2005 Wiley-Liss, Inc. and the American Pharmacists Association

Protein S-Mycothiolation Functions as Redox-Switch and Thiol Protection Mechanism in Corynebacterium glutamicum Under Hypochlorite Stress

PubMed Central

Chi, Bui Khanh; Busche, Tobias; Van Laer, Koen; Bäsell, Katrin; Becher, Dörte; Clermont, Lina; Seibold, Gerd M.; Persicke, Marcus; Kalinowski, Jörn; Messens, Joris

2014-01-01

Abstract Aims: Protein S-bacillithiolation was recently discovered as important thiol protection and redox-switch mechanism in response to hypochlorite stress in Firmicutes bacteria. Here we used transcriptomics to analyze the NaOCl stress response in the mycothiol (MSH)-producing Corynebacterium glutamicum. We further applied thiol-redox proteomics and mass spectrometry (MS) to identify protein S-mycothiolation. Results: Transcriptomics revealed the strong upregulation of the disulfide stress σH regulon by NaOCl stress in C. glutamicum, including genes for the anti sigma factor (rshA), the thioredoxin and MSH pathways (trxB1, trxC, cg1375, trxB, mshC, mca, mtr) that maintain the redox balance. We identified 25 S-mycothiolated proteins in NaOCl-treated cells by liquid chromatography–tandem mass spectrometry (LC-MS/MS), including 16 proteins that are reversibly oxidized by NaOCl in the thiol-redox proteome. The S-mycothiolome includes the methionine synthase (MetE), the maltodextrin phosphorylase (MalP), the myoinositol-1-phosphate synthase (Ino1), enzymes for the biosynthesis of nucleotides (GuaB1, GuaB2, PurL, NadC), and thiamine (ThiD), translation proteins (TufA, PheT, RpsF, RplM, RpsM, RpsC), and antioxidant enzymes (Tpx, Gpx, MsrA). We further show that S-mycothiolation of the thiol peroxidase (Tpx) affects its peroxiredoxin activity in vitro that can be restored by mycoredoxin1. LC-MS/MS analysis further identified 8 proteins with S-cysteinylations in the mshC mutant suggesting that cysteine can be used for S-thiolations in the absence of MSH. Innovation and Conclusion: We identified widespread protein S-mycothiolations in the MSH-producing C. glutamicum and demonstrate that S-mycothiolation reversibly affects the peroxidase activity of Tpx. Interestingly, many targets are conserved S-thiolated across bacillithiol- and MSH-producing bacteria, which could become future drug targets in related pathogenic Gram-positives. Antioxid. Redox Signal. 20, 589–605

Acetate- and thiol-capped monodisperse ruthenium nanoparticles: XPS, XAS, and HRTEM studies.

PubMed

Chakroune, Nassira; Viau, Guillaume; Ammar, Souad; Poul, Laurence; Veautier, Delphine; Chehimi, Mohamed M; Mangeney, Claire; Villain, Françoise; Fiévet, Fernand

2005-07-19

Monodisperse ruthenium nanoparticles were prepared by reduction of RuCl3 in 1,2-propanediol. The mean particle size was controlled by appropriate choice of the reduction temperature and the acetate ion concentration. Colloidal solutions in toluene were obtained by coating the metal particles with dodecanethiol. High-resolution transmission electron microscopy (HRTEM), X-ray photoelectron spectroscopy (XPS), and X-ray absorption spectroscopy (XANES and EXAFS for the Ru K-absorption edge) were performed on particles of two different diameters, 2 and 4 nm, and in different environments, polyol/acetate or thiol. For particles stored in polyol/acetate XPS studies revealed superficial oxidation limited to one monolayer and a surface coating containing mostly acetate ions. Analysis of the EXAFS spectra showed both oxygen and ruthenium atoms around the ruthenium atoms with a Ru-Ru coordination number N smaller than the bulk value, as expected for fine particles. In the case of 2 nm acetate-capped particles N is consistent with particles made up of a metallic core and an oxidized monolayer. For 2 nm thiol-coated particles, a Ru-S bond was evidenced by XPS and XAS. For the 4 nm particles XANES and XPS studies showed that most of the ruthenium atoms are in the zerovalent state. Nevertheless, in both cases, when capped with thiol, the Ru-Ru coordination number inferred from EXAFS is much smaller than for particles of the same size stored in polyol. This is attributed to a structural disorganization of the particles by thiol chemisorption. HRTEM studies confirm the marked dependence of the structural properties of the ruthenium particles on their chemical environment; they show the acetate-coated particles to be single crystals, whereas the thiol-coated particles appear to be polycrystalline.

Modifying surface resistivity and liquid moisture management property of keratin fibers through thiol-ene click reactions.

PubMed

Yu, Dan; Cai, Jackie Y; Church, Jeffrey S; Wang, Lijing

2014-01-22

This paper reports on a new method for improving the antistatic and liquid moisture management properties of keratinous materials. The method involves the generation of thiols by controlled reduction of cystine disulfide bonds in keratin with tris(2-carboxyethyl) phosphine hydrochloride and subsequent grafting of hydrophilic groups onto the reduced keratin by reaction with an acrylate sulfonate or acrylamide sulfonate through thiol-ene click chemistry. The modified substrates were characterized with Raman spectroscopy and scanning electron microscopy and evaluated for their performance changes in liquid moisture management, surface resistivity, and wet burst strength. The results have revealed that the thiol-acrylate reaction is more efficient than the thiol-acrylamide reaction, and the keratinous substrate modified with an acrylate sulfonate salt exhibits significantly improved antistatic and liquid moisture management properties.

Enantioselective Synthesis of α-Mercapto-β-amino Esters via Rh(II)/Chiral Phosphoric Acid-Cocatalyzed Three-Component Reaction of Diazo Compounds, Thiols, and Imines.

PubMed

Xiao, Guolan; Ma, Chaoqun; Xing, Dong; Hu, Wenhao

2016-12-02

An enantioselective method for the synthesis of α-mercapto-β-amino esters has been developed via a rhodium(II)/chiral phosphoric acid-cocatalyzed three-component reaction of diazo compounds, thiols, and imines. This transformation is proposed to proceed through enantioselective trapping of the sulfonium ylide intermediate generated in situ from the diazo compound and thiol by the phosphoric acid-activated imine. With this method, a series of α-mercapto-β-amino esters were obtained in good yields with moderate to good stereoselectivities.

Photogenerated Lectin Sensors Produced by Thiol-Ene/Yne Photo-Click Chemistry in Aqueous Solution

PubMed Central

Norberg, Oscar; Lee, Irene H.; Aastrup, Teodor; Yan, Mingdi; Ramström, Olof

2012-01-01

The photoinitiated radical reactions between thiols and alkenes/alkynes (thiol-ene and thiol-yne chemistry) have been applied to a functionalization methodology to produce carbohydrate-presenting surfaces for analyses of biomolecular interactions. Polymer-coated quartz surfaces were functionalized with alkenes or alkynes in a straightforward photochemical procedure utilizing perfluorophenylazide (PFPA) chemistry. The alkene/alkyne surfaces were subsequently allowed to react with carbohydrate thiols in water under UV-irradiation. The reaction can be carried out in a drop of water directly on the surface without photoinitiator and any disulfide side products were easily washed away after the functionalization process. The resulting carbohydrate-presenting surfaces were evaluated in real-time studies of protein-carbohydrate interactions using a quartz crystal microbalance flow-through system with recurring injections of selected lectins with intermediate regeneration steps using low pH buffer. The resulting methodology proved fast, efficient and scalable to high-throughput analysis formats, and the produced surfaces showed significant protein binding with expected selectivities of the lectins used in the study. PMID:22341757

Relationship between Extracellular Low-Molecular-Weight Thiols and Mercury Species in Natural Lake Periphytic Biofilms.

PubMed

Leclerc, Maxime; Planas, Dolors; Amyot, Marc

2015-07-07

The uptake of mercury by microorganisms is a key step in the production of methylmercury, a biomagnifiable toxin. Mercury complexation by low-molecular-weight (LMW) thiols can affect its bioavailability and thus the production of methylmercury. Freshwater biofilms were sampled in the summer using artificial Teflon substrates submerged for over a year to allow natural community colonization in the littoral zone of a Boreal Shield lake. Inside biofilms, concentrations of different extracellular thiol species (thioglycolic acid, l-cysteine-l-glycine, cysteine, and glutathione) were up to 3 orders of magnitude greater than in the surrounding water column, potentially more readily controlling mercury speciation than in the water column. All biofilm thiols except thioglycolic acid were highly correlated to chlorophyll a, likely indicating an algal origin. Extracellular total mercury represented 3 ± 1% of all biofilm mercury and was preferentially found in the capsular fraction. Levels of LMW thiols of presumed algal origins were highly correlated with total mercury in the mobile colloidal fraction of biofilms. We propose that periphytic phototrophic microorganisms such as algae likely affect the bioavailability of mercury through the exudation of LMW thiols, and thus they may play a key role in the production of methylmercury in biofilms.

In vitro antimycoplasmal activities of rufloxacin and its metabolite MF 922.

PubMed Central

Furneri, P M; Bisignano, G; Cerniglia, G; Nicoletti, G; Cesana, M; Tempera, G

1994-01-01

The in vitro activities of rufloxacin and its metabolite, MF 922, were compared with those of ofloxacin, ciprofloxacin, erythromycin, and minocycline against Mycoplasma pneumoniae, Mycoplasma hominis, Mycoplasma fermentans, and Ureaplasma urealyticum. Rufloxacin, MF 922, and ciprofloxacin shared similar activities against all mycoplasmas tested. (MICs for 90% of isolates tested [MIC90s], 0.5 to 4 micrograms/ml. Ofloxacin had the lowest MIC90s for U. urealyticum, M. fermentans, and M. hominis (MIC90s, 0.25 to 1 micrograms/ml) and erythromycin had the lowest MIC90 for M. pneumoniae (MIC90, 0.004 micrograms/ml). PMID:7872762

Characterization and quantification of endogenous fatty acid nitroalkene metabolites in human urine[S

PubMed Central

Salvatore, Sonia R.; Vitturi, Dario A.; Baker, Paul R. S.; Bonacci, Gustavo; Koenitzer, Jeffrey R.; Woodcock, Steven R.; Freeman, Bruce A.; Schopfer, Francisco J.

2013-01-01

The oxidation and nitration of unsaturated fatty acids transforms cell membrane and lipoprotein constituents into mediators that regulate signal transduction. The formation of 9-NO2-octadeca-9,11-dienoic acid and 12-NO2-octadeca-9,11-dienoic acid stems from peroxynitrite- and myeloperoxidase-derived nitrogen dioxide reactions as well as secondary to nitrite disproportionation under the acidic conditions of digestion. Broad anti-inflammatory and tissue-protective responses are mediated by nitro-fatty acids. It is now shown that electrophilic fatty acid nitroalkenes are present in the urine of healthy human volunteers (9.9 ± 4.0 pmol/mg creatinine); along with electrophilic 16- and 14-carbon nitroalkenyl β-oxidation metabolites. High resolution mass determinations and coelution with isotopically-labeled metabolites support renal excretion of cysteine-nitroalkene conjugates. These products of Michael addition are in equilibrium with the free nitroalkene pool in urine and are displaced by thiol reaction with mercury chloride. This reaction increases the level of free nitroalkene fraction >10-fold and displays a KD of 7.5 × 10−6 M. In aggregate, the data indicates that formation of Michael adducts by electrophilic fatty acids is favored under biological conditions and that reversal of these addition reactions is critical for detecting both parent nitroalkenes and their metabolites. The measurement of this class of mediators can constitute a sensitive noninvasive index of metabolic and inflammatory status. PMID:23620137

Investigation of thiol derivatized gold nanoparticle sensors for gas analysis

NASA Astrophysics Data System (ADS)

Stephens, Jared S.

Analysis of volatile organic compounds (VOCs) in air and exhaled breath by sensor array is a very useful testing technique. It can provide non-invasive, fast, inexpensive testing for many diseases. Breath analysis has been very successful in identifying cancer and other diseases by using a chemiresistor sensor or array with gold nanoparticles to detect biomarkers. Acetone is a biomarker for diabetes and having a portable testing device could help to monitor diabetic and therapeutic progress. An advantage to this testing method is it is conducted at room temperature instead of 200 degrees Celsius. 3. The objective of this research is to determine the effect of thiol derivatized gold nanoparticles based on sensor(s) detection of VOCs. The VOCs to be tested are acetone, ethanol, and a mixture of acetone and ethanol. Each chip is tested under all three VOCs and three concentration levels (0.1, 1, and 5.0 ppm). VOC samples are used to test the sensors' ability to detect and differentiate VOCs. Sensors (also referred to as a chip) are prepared using several types of thiol derivatized gold nanoparticles. The factors are: thiol compound and molar volume loading of the thiol in synthesis. The average resistance results are used to determine the VOC selectivity of the sensors tested. The results show a trend of increasing resistance as VOC concentration is increased relative to dry air; which is used as baseline for VOCs. Several sensors show a high selectivity to one or more VOCs. Overall the 57 micromoles of 4-methoxy-toluenethiol sensor shows the strongest selectivity for VOCs tested. 3. Gerfen, Kurt. 2012. Detection of Acetone in Air Using Silver Ion Exchanged ZSM-5 and Zinc Oxide Sensing Films. Master of Science thesis, University of Louisville.

Stereoselective synthesis of an active metabolite of the potent PI3 kinase inhibitor PKI-179.

PubMed

Chen, Zecheng; Venkatesan, Aranapakam M; Dos Santos, Osvaldo; Delos Santos, Efren; Dehnhardt, Christoph M; Ayral-Kaloustian, Semiramis; Ashcroft, Joseph; McDonald, Leonard A; Mansour, Tarek S

2010-03-05

The synthesis and stereochemical determination of 1-(4-(4-((1R,5R,6R)-6-hydroxy-3-oxa-8-azabicyclo[3.2.1]octan-8-yl)-6-morpholino-1,3,5-triazin-2-yl)phenyl)-3-(pyridin-4-yl)urea (2), an active metabolite of the potent PI3 kinase inhibitor PKI-179 (1), is described. Stereospecific hydroboration of the double bond of 2,5-dihydro-1H-pyrrole 8 gave the 2,3-trans alcohol 9 exclusively. The configuration of the 3-hydroxyl group in 9 was inverted by an oxidation and stereoselective reduction sequence to give the corresponding 2,3-cis isomer 23. Both exo (21) and endo (27) isomers of the metabolite 2 were prepared via a practical synthetic route from 9 and 23, respectively, and the stereochemistry of 2 was determined to be endo. The endo isomer (27) was separated into two enantiomers 28 and 29 by chiral HPLC. Compound 2 was found to be enantiomerically pure and identical to the enantiomer 28. The absolute stereochemistry of the enantiomer 28 was determined by Mosher's method, thus establishing the stereochemistry of the active metabolite 2.

Spray-deposition and photopolymerization of organic-inorganic thiol-ene resins for fabrication of superamphiphobic surfaces.

PubMed

Xiong, Li; Kendrick, Laken L; Heusser, Hannele; Webb, Jamie C; Sparks, Bradley J; Goetz, James T; Guo, Wei; Stafford, Christopher M; Blanton, Michael D; Nazarenko, Sergei; Patton, Derek L

2014-07-09

Superamphiphobic surfaces, exhibiting high contact angles and low contact angle hysteresis to both water and low surface tension liquids, have attracted a great deal attention in recent years because of the potential of these materials in practical applications such as liquid-resistant textiles, self-cleaning surfaces, and antifouling/anticorrosion coatings. In this work, we present a simple strategy for fabricating of superamphiphobic coatings based on photopolymerization of hybrid thiol-ene resins. Spray-deposition and UV photopolymerization of thiol-ene resins containing hydrophobic silica nanoparticles and perfluorinated thiols provide a multiscale topography and low-energy surface that endows the surface with superamphiphobicity. The wettability and chemical composition of the surfaces were characterized by contact-angle goniometry and X-ray photoelectron spectroscopy, respectively. The hierarchical roughness features of the thiol-ene surfaces were investigated with field-emission scanning electron microscopy. Droplet impact and sandpaper abrasion tests indicate the coatings respectively possess a robust antiwetting behavior and good mechanical durability.

Graphene Oxide-Polymer Composite Langmuir Films Constructed by Interfacial Thiol-Ene Photopolymerization

NASA Astrophysics Data System (ADS)

Luo, Xiaona; Ma, Kai; Jiao, Tifeng; Xing, Ruirui; Zhang, Lexin; Zhou, Jingxin; Li, Bingbing

2017-02-01

The effective synthesis and self-assembly of graphene oxide (GO) nanocomposites are of key importance for a broad range of nanomaterial applications. In this work, a one-step chemical strategy is presented to synthesize stable GO-polymer Langmuir composite films by interfacial thiol-ene photopolymerization at room temperature, without use of any crosslinking agents and stabilizing agents. It is discovered that photopolymerization reaction between thiol groups modified GO sheets and ene in polymer molecules is critically responsible for the formation of the composite Langmuir films. The film formed by Langmuir assembly of such GO-polymer composite films shows potential to improve the mechanical and chemical properties and promotes the design of various GO-based nanocomposites. Thus, the GO-polymer composite Langmuir films synthesized by interfacial thiol-ene photopolymerization with such a straightforward and clean manner, provide new alternatives for developing chemically modified GO-based hybrid self-assembled films and nanomaterials towards a range of soft matter and graphene applications.

A novel strategy for global analysis of the dynamic thiol redox proteome.

PubMed

Martínez-Acedo, Pablo; Núñez, Estefanía; Gómez, Francisco J Sánchez; Moreno, Margoth; Ramos, Elena; Izquierdo-Álvarez, Alicia; Miró-Casas, Elisabet; Mesa, Raquel; Rodriguez, Patricia; Martínez-Ruiz, Antonio; Dorado, David Garcia; Lamas, Santiago; Vázquez, Jesús

2012-09-01

Nitroxidative stress in cells occurs mainly through the action of reactive nitrogen and oxygen species (RNOS) on protein thiol groups. Reactive nitrogen and oxygen species-mediated protein modifications are associated with pathophysiological states, but can also convey physiological signals. Identification of Cys residues that are modified by oxidative stimuli still poses technical challenges and these changes have never been statistically analyzed from a proteome-wide perspective. Here we show that GELSILOX, a method that combines a robust proteomics protocol with a new computational approach that analyzes variance at the peptide level, allows a simultaneous analysis of dynamic alterations in the redox state of Cys sites and of protein abundance. GELSILOX permits the characterization of the major endothelial redox targets of hydrogen peroxide in endothelial cells and reveals that hypoxia induces a significant increase in the status of oxidized thiols. GELSILOX also detected thiols that are redox-modified by ischemia-reperfusion in heart mitochondria and demonstrated that these alterations are abolished in ischemia-preconditioned animals.

Photoinduced Cross-Linking of Dynamic Poly(disulfide) Films via Thiol Oxidative Coupling.

PubMed

Feillée, Noémi; Chemtob, Abraham; Ley, Christian; Croutxé-Barghorn, Céline; Allonas, Xavier; Ponche, Arnaud; Le Nouen, Didier; Majjad, Hicham; Jacomine, Léandro

2016-01-01

Initially developed as an elastomer with an excellent record of barrier and chemical resistance properties, poly(disulfide) has experienced a revival linked to the dynamic nature of the S-S covalent bond. A novel photobase-catalyzed oxidative polymerization of multifunctional thiols to poly(disulfide) network is reported. Based solely on air oxidation, the single-step process is triggered by the photodecarboxylation of a xanthone acetic acid liberating a strong bicyclic guanidine base. Starting with a 1 μm thick film based on trithiol poly(ethylene oxide) oligomer, the UV-mediated oxidation of thiols to disulfides occurs in a matter of minutes both selectively, i.e., without overoxidation, and quantitatively as assessed by a range of spectroscopic techniques. Thiolate formation and film thickness determine the reaction rates and yield. Spatial control of the photopolymerization serves to generate robust micropatterns, while the reductive cleavage of S-S bridges allows the recycling of 40% of the initial thiol groups. © 2015 WILEY-VCH Verlag GmbH & Co. KGaA, Weinheim.

Metabolite profiling of bendamustine in urine of cancer patients after administration of [14C]bendamustine.

PubMed

Dubbelman, Anne-Charlotte; Jansen, Robert S; Rosing, Hilde; Darwish, Mona; Hellriegel, Edward; Robertson, Philmore; Schellens, Jan H M; Beijnen, Jos H

2012-07-01

Bendamustine is an alkylating agent consisting of a mechlorethamine derivative, a benzimidazole group, and a butyric acid substituent. A human mass balance study showed that bendamustine is extensively metabolized and subsequently excreted in urine. However, limited information is available on the metabolite profile of bendamustine in human urine. The objective of this study was to elucidate the metabolic pathways of bendamustine in humans by identification of its metabolites excreted in urine. Human urine samples were collected up to 168 h after an intravenous infusion of 120 mg/m(2) (80-95 μCi) [(14)C]bendamustine. Metabolites of [(14)C]bendamustine were identified using liquid chromatography (high-resolution)-tandem mass spectrometry with off-line radioactivity detection. Bendamustine and a total of 25 bendamustine-related compounds were detected. Observed metabolic conversions at the benzimidazole and butyric acid moiety were N-demethylation and γ-hydroxylation. In addition, various other combinations of these conversions with modifications at the mechlorethamine moiety were observed, including hydrolysis (the primary metabolic pathway), cysteine conjugation, and subsequent biotransformation to mercapturic acid and thiol derivatives, N-dealkylation, oxidation, and conjugation with phosphate, creatinine, and uric acid. Bendamustine-derived products containing phosphate, creatinine, and uric acid conjugates were also detected in control urine incubated with bendamustine. Metabolites that were excreted up to 168 h after the infusion included products of dihydrolysis and cysteine conjugation of bendamustine and γ-hydroxybendamustine. The range of metabolic reactions is generally consistent with those reported for rat urine and bile, suggesting that the overall processes involved in metabolic elimination are qualitatively the same in rats and humans.

Thiol-ene mediated neoglycosylation of collagen patches: a preliminary study.

PubMed

Russo, Laura; Battocchio, Chiara; Secchi, Valeria; Magnano, Elena; Nappini, Silvia; Taraballi, Francesca; Gabrielli, Luca; Comelli, Francesca; Papagni, Antonio; Costa, Barbara; Polzonetti, Giovanni; Nicotra, Francesco; Natalello, Antonino; Doglia, Silvia M; Cipolla, Laura

2014-02-11

Despite the relevance of carbohydrates as cues in eliciting specific biological responses, the covalent surface modification of collagen-based matrices with small carbohydrate epitopes has been scarcely investigated. We report thereby the development of an efficient procedure for the chemoselective neoglycosylation of collagen matrices (patches) via a thiol-ene approach, between alkene-derived monosaccharides and the thiol-functionalized material surface. Synchrotron radiation-induced X-ray photoelectron spectroscopy (SR-XPS), Fourier transform-infrared (FT-IR), and enzyme-linked lectin assay (ELLA) confirmed the effectiveness of the collagen neoglycosylation. Preliminary biological evaluation in osteoarthritic models is reported. The proposed methodology can be extended to any thiolated surface for the development of smart biomaterials for innovative approaches in regenerative medicine.

Chemical synthesis of oligonucleotides containing a free sulphydryl group and subsequent attachment of thiol specific probes.

PubMed Central

Connolly, B A; Rider, P

1985-01-01

Oligonucleotides containing a free sulphydryl group at their 5'-termini have been synthesised and further derivatised with thiol specific probes. The nucleotide sequence required is prepared using standard solid phase phosphoramidite techniques and an extra round of synthesis is then performed using the S-triphenylmethyl O-methoxymorpholinophosphite derivatives of 2-mercaptoethanol, 3-mercaptopropan (1) ol or 6-mercaptohexan (1) ol. After cleavage from the resin and removal of the phosphate and base protecting groups, this yields an oligonucleotide containing an S-triphenylmethyl group attached to the 5'-phosphate group via a two, three or six carbon chain. The triphenylmethyl group can be readily removed with silver nitrate to give the free thiol. With the three and six carbon chain oligonucleotides, this thiol can be used, at pH 8, for the attachment of thiol specific probes as illustrated by the reaction with fluorescent conjugates of iodoacetates and maleiimides. However, oligonucleotides containing a thiol attached to the 5'-phosphate group via a two carbon chain are unstable at pH 8 decomposing to the free 5'-phosphate and so are unsuitable for further derivatisation. PMID:4011448

Fungal secondary metabolites - strategies to activate silent gene clusters.

PubMed

Brakhage, Axel A; Schroeckh, Volker

2011-01-01

Filamentous fungi produce a multitude of low molecular weight bioactive compounds. The increasing number of fungal genome sequences impressively demonstrated that their biosynthetic potential is far from being exploited. In fungi, the genes required for the biosynthesis of a secondary metabolite are clustered. Many of these bioinformatically newly discovered secondary metabolism gene clusters are silent under standard laboratory conditions. Consequently, no product can be found. This review summarizes the current strategies that have been successfully applied during the last years to activate these silent gene clusters in filamentous fungi, especially in the genus Aspergillus. The techniques take advantage of genome mining, vary from the simple search for compounds with bioinformatically predicted physicochemical properties up to methods that exploit a probable interaction of microorganisms. Until now, the majority of successful approaches have been based on molecular biology like the generation of gene "knock outs", promoter exchange, overexpression of transcription factors or other pleiotropic regulators. Moreover, strategies based on epigenetics opened a new avenue for the elucidation of the regulation of secondary metabolite formation and will certainly continue to play a significant role for the elucidation of cryptic natural products. The conditions under which a given gene cluster is naturally expressed are largely unknown. One technique is to attempt to simulate the natural habitat by co-cultivation of microorganisms from the same ecosystem. This has already led to the activation of silent gene clusters and the identification of novel compounds in Aspergillus nidulans. These simulation strategies will help discover new natural products in the future, and may also provide fundamental new insights into microbial communication. Copyright © 2010 Elsevier Inc. All rights reserved.

The compromise of dynamic disulfide/thiol homeostasis as a biomarker of oxidative stress in trichloroethylene exposure.

PubMed

Bal, C; Büyükşekerci, M; Koca, C; Ağış, E R; Erdoğan, S; Baran, P; Gündüzöz, M; Yilmaz, Öh

2016-09-01

In this study, we aimed to investigate disulfide/thiol homeostasis in trichloroethylene (TCE) exposure. The study was carried out in 30 nonsmoker TCE-exposed workers with a variety of occupations. Additionally, 30 healthy nonsmoker volunteers were recruited as the control group. TCE exposure was determined by measuring urinary trichloroacetic acid (TCA) concentration. Median urinary TCA levels of exposed workers (20.5 mg/L) were significantly higher than control subjects (5 mg/L). Thiol and disulfide concentrations were determined using a novel automated method. Disulfide/thiol ratio was significantly higher in the exposed group (p < 0.001). Thiol/disulfide homeostasis was found to be disturbed in TCE-exposed workers. We predict that in TCE-exposed workers this disturbance can be a therapeutic target, and the efficiency of the treatment can easily be monitored by the novel method we used. © The Author(s) 2015.

Thimerosal Exposure and the Role of Sulfation Chemistry and Thiol Availability in Autism

PubMed Central

Kern, Janet K.; Haley, Boyd E.; Geier, David A.; Sykes, Lisa K.; King, Paul G.; Geier, Mark R.

2013-01-01

Autism spectrum disorder (ASD) is a neurological disorder in which a significant number of the children experience a developmental regression characterized by a loss of previously acquired skills and abilities. Typically reported are losses of verbal, nonverbal, and social abilities. Several recent studies suggest that children diagnosed with an ASD have abnormal sulfation chemistry, limited thiol availability, and decreased glutathione (GSH) reserve capacity, resulting in a compromised oxidation/reduction (redox) and detoxification capacity. Research indicates that the availability of thiols, particularly GSH, can influence the effects of thimerosal (TM) and other mercury (Hg) compounds. TM is an organomercurial compound (49.55% Hg by weight) that has been, and continues to be, used as a preservative in many childhood vaccines, particularly in developing countries. Thiol-modulating mechanisms affecting the cytotoxicity of TM have been identified. Importantly, the emergence of ASD symptoms post-6 months of age temporally follows the administration of many childhood vaccines. The purpose of the present critical review is provide mechanistic insight regarding how limited thiol availability, abnormal sulfation chemistry, and decreased GSH reserve capacity in children with an ASD could make them more susceptible to the toxic effects of TM routinely administered as part of mandated childhood immunization schedules. PMID:23965928

Au-thiol interaction chemistry to influence the structural transformation of semiconductor nanocrystals and formation of giant nanostructures.

PubMed

Bose, Riya; Manna, Goutam; Pradhan, Narayan

2014-04-09

Giant nanostructures which are difficult to design by the classical growth process can be fabricated in a facilitated and well programmed surface ligand removal protocol employing the thiol-gold strong interaction chemistry. When thiol capped small ZnSe seed nanocrystals are treated with amine capped gold particles, gold snatches the thiol ligands from ZnSe and forces them to agglomerate leading to the giant crystalline ZnSe nanostructures. © 2013 WILEY-VCH Verlag GmbH & Co. KGaA, Weinheim.

Salicylic acid metabolites and derivatives inhibit CDK activity: Novel insights into aspirin's chemopreventive effects against colorectal cancer

PubMed Central

Dachineni, Rakesh; Kumar, D. Ramesh; Callegari, Eduardo; Kesharwani, Siddharth S.; Sankaranarayanan, Ranjini; Seefeldt, Teresa; Tummala, Hemachand; Bhat, G. Jayarama

2017-01-01

Aspirin's potential as a drug continues to be evaluated for the prevention of colorectal cancer (CRC). Although multiple targets for aspirin and its metabolite, salicylic acid, have been identified, no unifying mechanism has been proposed to clearly explain its chemopreventive effects. Our goal here was to investigate the ability of salicylic acid metabolites, known to be generated through cytochrome P450 (CYP450) enzymes, and its derivatives as cyclin dependent kinase (CDK) inhibitors to gain new insights into aspirin's chemopreventive actions. Using in vitro kinase assays, for the first time, we demonstrate that salicylic acid metabolites, 2,3-dihydroxy-benzoic acid (2,3-DHBA) and 2,5-dihydroxybenzoic acid (2,5-DHBA), as well as derivatives 2,4-dihydroxybenzoic acid (2,4-DHBA), 2,6-dihydroxybenzoic acid (2,6-DHBA), inhibited CDK1 enzyme activity. 2,3-DHBA and 2,6-DHBA did not inhibit CDK2 and 4; however, both inhibited CDK-6 activity. Interestingly, another derivative, 2,4,6-trihydroxybenzoic acid (2,4,6-THBA) was highly effective in inhibiting CDK1, 2, 4 and 6 activity. Molecular docking studies showed that these compounds potentially interact with CDK1. Immunoblotting experiments showed that aspirin acetylated CDK1, and pre-incubation with salicylic acid and its derivatives prevented aspirin-mediated CDK1 acetylation, which supported the data obtained from molecular docking studies. We suggest that intracellularly generated salicylic acid metabolites through CYP450 enzymes within the colonic epithelial cells, or the salicylic acid metabolites generated by gut microflora may significantly contribute to the preferential chemopreventive effect of aspirin against CRC through inhibition of CDKs. This novel hypothesis and mechanism of action in aspirin's chemopreventive effects opens a new area for future research. In addition, structural modification to salicylic acid derivatives may prove useful in the development of novel CDK inhibitors in cancer prevention and

Proteome-wide Light/Dark Modulation of Thiol Oxidation in Cyanobacteria Revealed by Quantitative Site-specific Redox Proteomics*

PubMed Central

Guo, Jia; Nguyen, Amelia Y.; Dai, Ziyu; Su, Dian; Gaffrey, Matthew J.; Moore, Ronald J.; Jacobs, Jon M.; Monroe, Matthew E.; Smith, Richard D.; Koppenaal, David W.; Pakrasi, Himadri B.; Qian, Wei-Jun

2014-01-01

Reversible protein thiol oxidation is an essential regulatory mechanism of photosynthesis, metabolism, and gene expression in photosynthetic organisms. Herein, we present proteome-wide quantitative and site-specific profiling of in vivo thiol oxidation modulated by light/dark in the cyanobacterium Synechocystis sp. PCC 6803, an oxygenic photosynthetic prokaryote, using a resin-assisted thiol enrichment approach. Our proteomic approach integrates resin-assisted enrichment with isobaric tandem mass tag labeling to enable site-specific and quantitative measurements of reversibly oxidized thiols. The redox dynamics of ∼2,100 Cys-sites from 1,060 proteins under light, dark, and 3-(3,4-dichlorophenyl)-1,1-dimethylurea (a photosystem II inhibitor) conditions were quantified. In addition to relative quantification, the stoichiometry or percentage of oxidation (reversibly oxidized/total thiols) for ∼1,350 Cys-sites was also quantified. The overall results revealed broad changes in thiol oxidation in many key biological processes, including photosynthetic electron transport, carbon fixation, and glycolysis. Moreover, the redox sensitivity along with the stoichiometric data enabled prediction of potential functional Cys-sites for proteins of interest. The functional significance of redox-sensitive Cys-sites in NADP-dependent glyceraldehyde-3-phosphate dehydrogenase, peroxiredoxin (AhpC/TSA family protein Sll1621), and glucose 6-phosphate dehydrogenase was further confirmed with site-specific mutagenesis and biochemical studies. Together, our findings provide significant insights into the broad redox regulation of photosynthetic organisms. PMID:25118246

Modification of Ag nanoparticles with mixed thiols for improved SERS detection of poorly adsorbing target molecules: detection of MDMA.

PubMed

Stewart, Alan; Bell, Steven E J

2011-04-21

Here we report an example of a mixed thiol monolayer on the surface of Ag nanoparticles which promotes adsorption and quantitative SERS detection of 3,4-methylenedioxymethamphetamine (MDMA, "Ecstasy"); the thiols in the mixed monolayers act synergistically since MDMA does not adsorb onto colloids modified with either of the thiols separately. © The Royal Society of Chemistry 2011

Thiol-ene chemistry guided preparation of thiolated polymeric nanocomposite for anodic stripping voltammetric analysis of Cd2+ and Pb2+.

PubMed

Su, Zhaohong; Liu, Ying; Zhang, Yi; Xie, Qingji; Chen, Li; Huang, Yi; Fu, Yingchun; Meng, Yue; Li, Xuejiao; Ma, Ming; Yao, Shouzhuo

2013-02-21

We report on the thiol-ene chemistry guided preparation of a novel thiolated polymeric nanocomposite involving polyaniline (PANI), a functionalized thiol, e.g., sulfur-rich 2,5-dimercapto-1,3,4-thiadiazole (DMcT), and multiwalled carbon nanotubes (MWCNTs) for the sensitive differential pulse anodic stripping voltammetric determination of Cd(2+) and Pb(2+) on a glassy carbon electrode (GCE). Briefly, the thiol-ene reaction of a thiol with oxidized PANI that was chemically synthesized in the presence of solution-dispersed acidified MWCNTs yielded a thiolated polymeric nanocomposite of thiol-PANI/MWCNTs. The thiols examined include DMcT, 1,6-hexanedithiol and β-mercaptoethanol. Quartz crystal microbalance, cyclic voltammetry, scanning electron microscopy, Fourier transform infrared spectroscopy and ultraviolet-visible spectroscopy were used for film characterization and process monitoring. Under the optimized conditions, the obtained Bi/Nafion/DMcT-PANI/MWCNTs/GCE can sensitively sense Cd(2+) and Pb(2+) with limits of detection of 0.01 and 0.04 μg L(-1), respectively.

A Novel Tool for the Assessment Oxidative Stress in Age-Related Macular Degeneration: Thiol/Disulfide Homeostasis Revisited.

PubMed

Arıkan Yorgun, Mücella; Toklu, Yasin; Altınkaynak, Hasan; Tanrıverdi, Burak; Ergin, Merve; Biçer, Cemile

2016-12-01

To investigate thiol/disulfide status using a novel automated assay in patients with age-related macular degeneration (AMD) compared to age-matched healthy controls. A total of 64 AMD patients [51 (79%) non-exudative, 13 (21%) exudative AMD] and 21 age-matched healthy control subjects were enrolled in this study. Plasma total thiol, native thiol, disulfide levels were measured and native thiol/disulfide ratio (TDR) was calculated using a novel spectrophotometric assay. Patients with AMD had significantly lower levels of total thiol (434.8 ± 7.0 μmol/L vs. 472.2 ± 7.9 μmol/L, p < 0.001), native thiol (393.6 ± 6.5 μmol/L vs. 437.5 ± 7.1 μmol/L, p = 0.004) compared to healthy controls. However, plasma disulfide levels were higher in AMD patients (20.6 ± 0.9 μmol/L vs. 17.3 ± 1.3 μmol/L, p = 0.113) compared to healthy controls. The TDR was not statistically different between the early AMD group and healthy controls (24.2 ± 2.3 vs. 29.5 ± 3.1, p = 0.345). However, intermediate and advanced stage AMD groups had significantly lower levels of TDR compared to healthy controls (21.6 ± 2.6 vs. 29.5 ± 3.1, p = 0.023 and 20.3 ± 1.2 vs. 29.5 ± 3.1, p = 0.005, respectively). Native TDR was significantly lower in patients with exudative and non-exudative AMD (19.9 ± 2.3 vs. 29.5 ± 3.1, p = 0.024 and 21.8 ± 1.14 vs. 29.47 ± 3.1 respectively, p = 0.011). A greater extent of thiol consumption occurred in AMD patients compared to age-matched healthy controls. However, despite the similar levels of total thiol levels between several grades of AMD, the plasma native TDR value was decreased in accordance with the severity of the disease, which reflected the disease grade better.

Solid-Phase Extraction of Sulfur Mustard Metabolites Using an Activated Carbon Fiber Sorbent.

PubMed

Lee, Jin Young; Lee, Yong Han

2016-01-01

A novel solid-phase extraction method using activated carbon fiber (ACF) was developed and validated. ACF has a vast network of pores of varying sizes and microporous structures that result in rapid adsorption and selective extraction of sulfur mustard metabolites according to the pH of eluting solvents. ACF could not only selectively extract thiodiglycol and 1-methylsulfinyl-2-[2-(methylthio)-ethylsulfonyl]ethane eluting a 9:1 ratio of dichloromethane to acetone, and 1,1'-sulfonylbis[2-(methylsulfinyl)ethane] and 1,1'-sulfonylbis- [2-S-(N-acetylcysteinyl)ethane] eluting 3% hydrogen chloride in methanol, but could also eliminate most interference without loss of analytes during the loading and washing steps. A sample preparation method has been optimized for the extraction of sulfur mustard metabolites from human urine using an ACF sorbent. The newly developed extraction method was applied to the trace analysis of metabolites of sulfur mustard in human urine matrices in a confidence-building exercise for the analysis of biomedical samples provided by the Organisation for the Prohibition of Chemical Weapons. © The Author 2015. Published by Oxford University Press. All rights reserved. For Permissions, please email: journals.permissions@oup.com.

Activation of the dormant secondary metabolite production by introducing gentamicin-resistance in a marine-derived Penicillium purpurogenum G59.

PubMed

Chai, Yun-Jing; Cui, Cheng-Bin; Li, Chang-Wei; Wu, Chang-Jing; Tian, Cong-Kui; Hua, Wei

2012-03-01

A new approach to activate silent gene clusters for dormant secondary metabolite production has been developed by introducing gentamicin-resistance to an originally inactive, marine-derived fungal strain Penicillium purpurogenum G59. Upon treatment of the G59 spores with a high concentration of gentamicin in aqueous DMSO, a total of 181 mutants were obtained by single colony isolation. In contrast to the strain G59, the EtOAc extracts of nine mutant cultures showed inhibitory effects on K562 cells, indicating that the nine mutants had acquired capability to produce antitumor metabolites. This was evidenced by TLC and HPLC analysis of EtOAc extracts of G59 and the nine mutants. Further isolation and characterization demonstrated that four antitumor secondary metabolites, janthinone (1), fructigenine A (2), aspterric acid methyl ester (3) and citrinin (4), were newly produced by mutant 5-1-4 compared to the parent strain G59, and which were also not found in the secondary metabolites of other Penicillium purpurogenum strains. However, Compounds 1-4 inhibited the proliferation of K562 cells with inhibition rates of 34.6% (1), 60.8% (2), 31.7% (3) and 67.1% (4) at 100 μg/mL, respectively. The present study demonstrated the effectiveness of a simple, yet practical approach to activate the production of dormant fungal secondary metabolites by introducing acquired resistance to aminoglycoside antibiotics, which could be applied to the studies for eliciting dormant metabolic potential of fungi to obtain cryptic secondary metabolites.

Secondary metabolites from Ganoderma.

PubMed

Baby, Sabulal; Johnson, Anil John; Govindan, Balaji

2015-06-01

Ganoderma is a genus of medicinal mushrooms. This review deals with secondary metabolites isolated from Ganoderma and their biological significance. Phytochemical studies over the last 40years led to the isolation of 431 secondary metabolites from various Ganoderma species. The major secondary compounds isolated are (a) C30 lanostanes (ganoderic acids), (b) C30 lanostanes (aldehydes, alcohols, esters, glycosides, lactones, ketones), (c) C27 lanostanes (lucidenic acids), (d) C27 lanostanes (alcohols, lactones, esters), (e) C24, C25 lanostanes (f) C30 pentacyclic triterpenes, (g) meroterpenoids, (h) farnesyl hydroquinones (meroterpenoids), (i) C15 sesquiterpenoids, (j) steroids, (k) alkaloids, (l) prenyl hydroquinone (m) benzofurans, (n) benzopyran-4-one derivatives and (o) benzenoid derivatives. Ganoderma lucidum is the species extensively studied for its secondary metabolites and biological activities. Ganoderma applanatum, Ganoderma colossum, Ganoderma sinense, Ganoderma cochlear, Ganoderma tsugae, Ganoderma amboinense, Ganoderma orbiforme, Ganoderma resinaceum, Ganoderma hainanense, Ganoderma concinna, Ganoderma pfeifferi, Ganoderma neo-japonicum, Ganoderma tropicum, Ganoderma australe, Ganoderma carnosum, Ganoderma fornicatum, Ganoderma lipsiense (synonym G. applanatum), Ganoderma mastoporum, Ganoderma theaecolum, Ganoderma boninense, Ganoderma capense and Ganoderma annulare are the other Ganoderma species subjected to phytochemical studies. Further phytochemical studies on Ganoderma could lead to the discovery of hitherto unknown biologically active secondary metabolites. Copyright © 2015 Elsevier Ltd. All rights reserved.

Advanced Oxidation Protein Products-Modified Albumin Induces Differentiation of RAW264.7 Macrophages into Dendritic-Like Cells Which Is Modulated by Cell Surface Thiols.

PubMed

Garibaldi, Silvano; Barisione, Chiara; Marengo, Barbara; Ameri, Pietro; Brunelli, Claudio; Balbi, Manrico; Ghigliotti, Giorgio

2017-01-10

Local accumulation of Advanced Oxidation Protein Products (AOPP) induces pro-inflammatory and pro-fibrotic processes in kidneys and is an independent predictor of renal fibrosis and of rapid decline of eGFR in patients with chronic kidney disease (CKD). In addition to kidney damage, circulating AOPP may be regarded as mediators of systemic oxidative stress and, in this capacity, they might play a role in the progression of atherosclerotic damage of arterial walls. Atherosclerosis is a chronic inflammatory disease that involves activation of innate and adaptive immunity. Dendritic cells (DCs) are key cells in this process, due to their role in antigen presentation, inflammation resolution and T cell activation. AOPP consist in oxidative modifications of proteins (such as albumin and fibrinogen) that mainly occur through myeloperoxidase (MPO)-derived hypochlorite (HOCl). HOCl modified proteins have been found in atherosclerotic lesions. The oxidizing environment and the shifts in cellular redox equilibrium trigger inflammation, activate immune cells and induce immune responses. Thus, surface thiol groups contribute to the regulation of immune functions. The aims of this work are: (1) to evaluate whether AOPP-proteins induce activation and differentiation of mature macrophages into dendritic cells in vitro; and (2) to define the role of cell surface thiol groups and of free radicals in this process. AOPP-proteins were prepared by in vitro incubation of human serum albumin (HSA) with HOCl. Mouse macrophage-like RAW264.7 were treated with various concentrations of AOPP-HSA with or without the antioxidant N -acetyl cysteine (NAC). Following 48 h of HSA-AOPP treatment, RAW264.7 morphological changes were evaluated by microscopic observation, while markers of dendritic lineage and activation (CD40, CD86, and MHC class II) and allogeneic T cell proliferation were evaluated by flow cytometry. Cell surface thiols were measured by AlexaFluor-maleimide binding, and ROS

Dietary thiols in exercise: oxidative stress defence, exercise performance, and adaptation.

PubMed

McLeay, Yanita; Stannard, Stephen; Houltham, Stuart; Starck, Carlene

2017-01-01

Endurance athletes are susceptible to cellular damage initiated by excessive levels of aerobic exercise-produced reactive oxygen species (ROS). Whilst ROS can contribute to the onset of fatigue, there is increasing evidence that they play a crucial role in exercise adaptations. The use of antioxidant supplements such as vitamin C and E in athletes is common; however, their ability to enhance performance and facilitate recovery is controversial, with many studies suggesting a blunting of training adaptations with supplementation. The up-regulation of endogenous antioxidant systems brought about by exercise training allows for greater tolerance to subsequent ROS, thus, athletes may benefit from increasing these systems through dietary thiol donors. Recent work has shown supplementation with a cysteine donor (N-acetylcysteine; NAC) improves antioxidant capacity by augmenting glutathione levels and reducing markers of oxidative stress, as well as ergogenic potential through association with delayed fatigue in numerous experimental models. However, the use of this, and other thiol donors may have adverse physiological effects. A recent discovery for the use of a thiol donor food source, keratin, to potentially enhance endogenous antioxidants may have important implications for endurance athletes hoping to enhance performance and recovery without blunting training adaptations.

3D-QSAR Studies on a Series of Dihydroorotate Dehydrogenase Inhibitors: Analogues of the Active Metabolite of Leflunomide

PubMed Central

Li, Shun-Lai; He, Mao-Yu; Du, Hong-Guang

2011-01-01

The active metabolite of the novel immunosuppressive agent leflunomide has been shown to inhibit the enzyme dihydroorotate dehydrogenase (DHODH). This enzyme catalyzes the fourth step in de novo pyrimidine biosynthesis. Self-organizing molecular field analysis (SOMFA), a simple three-dimensional quantitative structure-activity relationship (3D-QSAR) method is used to study the correlation between the molecular properties and the biological activities of a series of analogues of the active metabolite. The statistical results, cross-validated rCV2 (0.664) and non cross-validated r2 (0.687), show a good predictive ability. The final SOMFA model provides a better understanding of DHODH inhibitor-enzyme interactions, and may be useful for further modification and improvement of inhibitors of this important enzyme. PMID:21686163

Modification of porous silicon rugate filters through thiol-yne photochemistry

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

Soeriyadi, Alexander H., E-mail: alexander.soeriyadi@unsw.edu.au; Zhu, Ying, E-mail: alexander.soeriyadi@unsw.edu.au; Gooding, J. Justin, E-mail: justin.gooding@unsw.edu.au

2014-02-24

Porous silicon (PSi) has a considerable potential as biosensor platform. In particular, the ability to modify the surface chemistry of porous silicon is of interest. Here we present a generic method to modify the surface of porous silicon through thiol-yne photochemistry initiated by a radical initiator. Firstly, a freshly etched porous silicon substrate is modified through thermal hydrosilylation with 1,8-nonadiyne to passivate the surface and introduce alkyne functionalities. The alkyne functional surface could then be further reacted with thiol species in the presence of a radical initiator and UV light. Functionalization of the PSi rugate filter is followed with opticalmore » reflectivity measurements as well as high resolution X-ray photoelectron spectroscopy (XPS)« less

Acid-Activatable Michael-Type Fluorescent Probes for Thiols and for Labeling Lysosomes in Live Cells.

PubMed

Dai, Chun-Guang; Du, Xiao-Jiao; Song, Qin-Hua

2015-12-18

A Michael addition is usually taken as a base-catalyzed reaction. Most fluorescent probes have been designed to detect thiols in slightly alkaline solutions (pH 7-9). The sensing reactions of almost all Michael-type fluorescent probes for thiols are faster in a high pH solution than in a low pH solution. In this work, we synthesized a series of 7-substituted 2-(quinolin-2-ylmethylene)malonic acids (QMAs, substituents: NEt2, OH, H, Cl, or NO2) and their ethyl esters (QMEs) as Michael-type fluorescent probes for thiols. The sensing reactions of QMAs and QMEs occur in distinct pH ranges, pH < 7 for QMAs and pH > 7 for QMEs. On the basis of experimental and theoretic studies, we have clarified the distinct pH effects on the sensing reactivity between QMAs and QMEs and demonstrated that two QMAs (NEt2, OH) are highly sensitive and selective fluorescent probes for thiols in acidic solutions (pH < 7) and promising dyes that can label lysosomes in live cells.

Sulforaphane inhibits damage-induced poly (ADP-ribosyl)ation via direct interaction of its cellular metabolites with PARP-1.

PubMed

Piberger, Ann Liza; Keil, Claudia; Platz, Stefanie; Rohn, Sascha; Hartwig, Andrea

2015-11-01

The isothiocyanate sulforaphane, a major breakdown product of the broccoli glucosinolate glucoraphanin, has frequently been proposed to exert anticarcinogenic properties. Potential underlying mechanisms include a zinc release from Kelch-like ECH-associated protein 1 followed by the induction of detoxifying enzymes. This suggests that sulforaphane may also interfere with other zinc-binding proteins, e.g. those essential for DNA repair. Therefore, we explored the impact of sulforaphane on poly (ADP-ribose)polymerase-1 (PARP-1), poly (ADP-ribosyl)ation (PARylation), and DNA single-strand break repair (SSBR) in cell culture. Immunofluorescence analyses showed that sulforaphane diminished H2 O2 -induced PARylation in HeLa S3 cells starting from 15 μM despite increased lesion induction under these conditions. Subcellular experiments quantifying the damage-induced incorporation of (32) P-ADP-ribose by PARP-1 displayed no direct impact of sulforaphane itself, but cellular metabolites, namely the glutathione conjugates of sulforaphane and its interconversion product erucin, reduced PARP-1 activity concentration dependently. Interestingly, this sulforaphane metabolite-induced PARP-1 inhibition was prevented by thiol compounds. PARP-1 is a stimulating factor for DNA SSBR-rate and we further demonstrated that 25 μM sulforaphane also delayed the rejoining of H2 O2 -induced DNA strand breaks, although this might be partly due to increased lesion frequencies. Sulforaphane interferes with damage-induced PARylation and SSBR, which implies a sulforaphane-dependent impairment of genomic stability. © 2015 WILEY-VCH Verlag GmbH & Co. KGaA, Weinheim.

Thiol functionalized polymethacrylic acid-based hydrogel microparticles for oral insulin delivery.

PubMed

Sajeesh, S; Vauthier, C; Gueutin, C; Ponchel, G; Sharma, Chandra P

2010-08-01

In the present study thiol functionalized polymethacrylic acid-polyethylene glycol-chitosan (PCP)-based hydrogel microparticles were utilized to develop an oral insulin delivery system. Thiol modification was achieved by grafting cysteine to the activated surface carboxyl groups of PCP hydrogels (Cys-PCP). Swelling and insulin loading/release experiments were conducted on these particles. The ability of these particles to inhibit protease enzymes was evaluated under in vitro experimental conditions. Insulin transport experiments were performed on Caco-2 cell monolayers and excised intestinal tissue with an Ussing chamber set-up. Finally, the efficacy of insulin-loaded particles in reducing the blood glucose level in streptozotocin-induced diabetic rats was investigated. Thiolated hydrogel microparticles showed less swelling and had a lower insulin encapsulation efficiency as compared with unmodified PCP particles. PCP and Cys-PCP microparticles were able to inhibit protease enzymes under in vitro conditions. Thiolation was an effective strategy to improve insulin absorption across Caco-2 cell monolayers, however, the effect was reduced in the experiments using excised rat intestinal tissue. Nevertheless, functionalized microparticles were more effective in eliciting a pharmacological response in diabetic animal, as compared with unmodified PCP microparticles. From these studies thiolation of hydrogel microparticles seems to be a promising approach to improve oral delivery of proteins/peptides. Copyright 2010 Acta Materialia Inc. Published by Elsevier Ltd. All rights reserved.

A crucial role for thiol antioxidants in estrogen-deficiency bone loss

PubMed Central

Lean, Jenny M.; Davies, Julie T.; Fuller, Karen; Jagger, Christopher J.; Kirstein, Barrie; Partington, Geoffrey A.; Urry, Zoë L.; Chambers, Timothy J.

2003-01-01

The mechanisms through which estrogen prevents bone loss are uncertain. Elsewhere, estrogen exerts beneficial actions by suppression of reactive oxygen species (ROS). ROS stimulate osteoclasts, the cells that resorb bone. Thus, estrogen might prevent bone loss by enhancing oxidant defenses in bone. We found that glutathione and thioredoxin, the major thiol antioxidants, and glutathione and thioredoxin reductases, the enzymes responsible for maintaining them in a reduced state, fell substantially in rodent bone marrow after ovariectomy and were rapidly normalized by exogenous 17-β estradiol. Moreover, administration of N-acetyl cysteine (NAC) or ascorbate, antioxidants that increase tissue glutathione levels, abolished ovariectomy-induced bone loss, while L-buthionine-(S,R)-sulphoximine (BSO), a specific inhibitor of glutathione synthesis, caused substantial bone loss. The 17-β estradiol increased glutathione and glutathione and thioredoxin reductases in osteoclast-like cells in vitro. Furthermore, in vitro NAC prevented osteoclast formation and NF-κB activation. BSO and hydrogen peroxide did the opposite. Expression of TNF-α, a target for NF-κB and a cytokine strongly implicated in estrogen-deficiency bone loss, was suppressed in osteoclasts by 17-β estradiol and NAC. These observations strongly suggest that estrogen deficiency causes bone loss by lowering thiol antioxidants in osteoclasts. This directly sensitizes osteoclasts to osteoclastogenic signals and entrains ROS-enhanced expression of cytokines that promote osteoclastic bone resorption. PMID:12975476

Linear glandular trichomes of Helianthus (Asteraceae): morphology, localization, metabolite activity and occurrence

PubMed Central

Aschenbrenner, Anna-Katharina; Horakh, Silke; Spring, Otmar

2013-01-01

Capitate glandular trichomes of sunflower are well investigated, but detailed studies are lacking for the linear glandular trichomes (LGT), a second type of physiologically active plant hair present on the surface of sunflowers. Light, fluorescence and scanning electron microscopy as well as histochemical staining were used to investigate the structure and metabolite deposition of LGT. Consisting of 6–11 linearly arranged cells, LGT were found on the surface of most plant organs of Helianthus annuus. They were associated with the leaf vascular system, and also occurred along petioles, stems and the abaxial surface of chaffy bracts, ray and disc florets. The highest density was found on the abaxial surface of phyllaries. Phenotypically similar LGT were common in all species of the genus, but also occurred in most other genera of the Helianthinae so far screened. Brownish and fluorescent metabolites of an as yet unknown chemical structure, together with terpenoids, were produced and stored in apical cells of LGT. The deposition of compounds gradually progressed from the tip cell to the basal cells of older trichomes. This process was accompanied by nucleus degradation in metabolite-accumulating cells. The localization of these trichomes on prominent plant parts of the apical bud and the capitulum combined with the accumulation of terpenoids and other as yet unknown compounds suggests a chemo-ecological function of the LGT in plant–insect or plant–herbivore interaction.

Thiol derivatization of Xanthan gum and its evaluation as a mucoadhesive polymer.

PubMed

Bhatia, Meenakshi; Ahuja, Munish; Mehta, Heena

2015-10-20

Thiol-derivatization of xanthan gum polysaccharide was carried out by esterification with mercaptopropionic acid and thioglycolic acid. Thiol-derivatization was confirmed by Fourier-transformed infra-red spectroscopy. Xanthan-mercaptopropionic acid conjugate and xanthan-thioglycolic acid conjugate were found to possess 432.68mM and 465.02mM of thiol groups as determined by Ellman's method respectively. Comparative evaluation of mucoadhesive property of metronidazole loaded buccal pellets of xanthan and thiolated xanthan gum using chicken buccal pouch membrane revealed higher ex vivo bioadhesion time of thiolated xanthan gum as compared to xanthan gum. Improved mucoadhesive property of thiolated xanthan gum over the xanthan gum can be attributed to the formation of disulfide bond between mucus and thiolated xanthan gum. In vitro release study conducted using phosphate buffer (pH 6.8) revealed a sustained release profile of metronidazole from thiolated xanthan pellets as compared to xanthan pellets. In conclusion, thiolation of xanthan improves its mucoadhesive property and sustained the release of metronidazole over a prolonged period. Copyright © 2015 Elsevier Ltd. All rights reserved.

Differentiation of Sclerotinia minor depends on thiol redox state and oxidative stress.

PubMed

Patsoukis, Nikolaos; Georgiou, Christos D

2008-01-01

Sclerotial differentiation in Sclerotinia minor is associated with oxidative stress and thiol redox state. The significance of oxidative stress to sclerotial differentiation was revealed by the higher oxidative stress of S. minor compared with a nonsclerotiogenic counterpart. The effect of thiol redox state on sclerotial differentiation was shown by the antioxidant action of the thiol (-SH) group of N-acetylcysteine and cysteine and by an unknown (not antioxidant) role of glutathione (GSH) on S. minor. The nonantioxidant role of GSH was indicated by the differentiation-inhibiting and differentiation-noninhibiting actions of the GSH biosynthesis inhibitor L-buthionine-S,R-sulfoximine and the GSH biosynthesis inducer L-2-oxo-thiazolidine-4-carboxylate, respectively, and by the increase of oxidative stress they caused during the transition from the undifferentiated to differentiated state of S. minor. Moreover, N-acetylcysteine can be used as a potent nontoxic fungicide against this phytopathogenic fungus by acting as a growth-inhibiting cytotoxic oxidant and by sustaining the fungus in the undifferentiated hyphal stage, which is vulnerable to degradation by soil microorganisms.

Effect of storage time on metabolite profile and alpha-glucosidase inhibitory activity of Cosmos caudatus leaves - GCMS based metabolomics approach.

PubMed

Javadi, Neda; Abas, Faridah; Mediani, Ahmed; Abd Hamid, Azizah; Khatib, Alfi; Simoh, Sanimah; Shaari, Khozirah

2015-09-01

Cosmos caudatus, which is a commonly consumed vegetable in Malaysia, is locally known as "Ulam Raja". It is a local Malaysian herb traditionally used as a food and medicinal herb to treat several maladies. Its bioactive or nutritional constituents consist of a wide range of metabolites, including glucosinolates, phenolics, amino acids, organic acids, and sugars. However, many of these metabolites are not stable and easily degraded or modified during storage. In order to investigate the metabolomics changes occurring during post-harvest storage, C. caudatus samples were subjected to seven different storage times (0 hours, 2 hours, 4 hours, 6 hours, 8 hours, 10 hours, and 12 hours) at room temperature. As the model experiment, the metabolites identified by gas chromatography-mass spectrometry (GC-MS) were correlated with α-glucosidase inhibitory activity analyzed with multivariate data analysis (MVDA) to find out the variation among samples and metabolites contributing to the activity. Orthogonal partial least squares (OPLS) analysis was applied to investigate the metabolomics changes. A profound chemical alteration, both in primary and secondary metabolites, was observed. The α-tocopherol, catechin, cyclohexen-1-carboxylic acid, benzoic acid, myo-inositol, stigmasterol, and lycopene compounds were found to be the discriminating metabolites at early storage; however, sugars such as sucrose, α-d-galactopyranose, and turanose were detected, which was attributed to the discriminating metabolites for late storage. The result shows that the MVDA method is a promising technique to identify biomarker compounds relative to storage at different times. Copyright © 2015. Published by Elsevier B.V.

O-acetylserine(thiol)lyase from spinach (Spinacia oleracea L.) leaf: cDNA cloning, characterization, and overexpression in Escherichia coli of the chloroplast isoform.

PubMed

Rolland, N; Droux, M; Lebrun, M; Douce, R

1993-01-01

The last enzymatic step for L-cysteine biosynthesis is catalyzed by O-acetylserine(thiol)lyase (OASTL, EC 4.2.99.8) which synthesizes L-cysteine from O-acetylserine and "sulfide." We have isolated and characterized a full-length cDNA (1432 bp) from a lambda gt11 library of spinach leaf encoding the complete precursor of the chloroplast isoform. The 1149-nucleotide open reading frame coding for O-acetylserine(thiol)lyase was in the direction opposite that of the lambda gt11 beta-galactosidase gene. The derived amino acid sequence indicates that the protein precursor consists of 383 amino acid residues including a N-terminal presequence peptide of 52 residues. The amino acid sequence of mature spinach chloroplast O-acetylserine(thiol)lyase shows 40 and 57% homology with its bacterial counterparts. Sequence comparison with several pyridoxal 5'-phosphate-containing proteins reveals the presence of a lysine residue assumed to be involved in cofactor binding. A synthetic cDNA was constructed, coding for the entire 331-amino-acid mature O-acetylserine(thiol)lyase and for an initiating methionine. A high level of expression of the active mature chloroplast isoform was achieved in an Escherichia coli strain carrying the T7 RNA polymerase system (F. W. Studier, A. H. Rosenberg, J. J. Dunn, and J. W. Dubendorff, 1990, in Methods in Enzymology, D. V. Goeddel, Ed., Vol. 185, pp. 60-89, Academic Press, San Diego, CA). Addition of pyridoxine to the bacterial growth medium enhanced the enzyme activity due to the recombinant protein. The extent of production is 25-fold higher than in chloroplast from spinach leaves and the recombinant protein presents the relative molecular mass and immunological properties of the natural enzyme from spinach leaf chloroplast. This work, together with our previous biochemical studies, are in accordance with a prokaryotic type enzyme for L-cysteine biosynthesis in higher plant chloroplasts. Southern blot analysis indicated that O-acetylserine(thiol

Analysis of volatile thiols in alcoholic beverages by simultaneous derivatization/extraction and liquid chromatography-high resolution mass spectrometry.

PubMed

Vichi, Stefania; Cortés-Francisco, Nuria; Caixach, Josep

2015-05-15

A simultaneous derivatization/extraction method followed by liquid chromatography-electrospray-high resolution mass spectrometry for the determination of volatile thiols in hydroalcoholic matrixes was optimized and used to identify and quantify volatile thiols in wine and beer samples. The method was evaluated in terms of sensitivity, precision, accuracy and selectivity. The experimental LOQs of eleven thiols tested ranged between 0.01 ng/L and 10 ng/L. Intra-day relative standard deviation (RSD) was in general lower than 10% and inter-day RSD ranged between 10% and 30%. Recovery in the model and real matrixes ranged from 45% to 129%. The method was then applied for the analysis of four white wines and six beers. Five out of the eleven reference thiols were identified and quantified in the samples analyzed. The non-target approach, carried out by monitoring the diagnostic ion at m/z 275.9922 [C13H10ONSe](+) in the fragmentation spectrum, allowed detecting, in the same samples, fourteen non-target thiols. Copyright © 2014 Elsevier Ltd. All rights reserved.

Structural Basis of a Thiol-Disulfide Oxidoreductase in the Hedgehog-Forming Actinobacterium Corynebacterium matruchotii.

PubMed

Luong, Truc Thanh; Tirgar, Reyhaneh; Reardon-Robinson, Melissa E; Joachimiak, Andrzej; Osipiuk, Jerzy; Ton-That, Hung

2018-05-01

The actinobacterium Corynebacterium matruchotii has been implicated in nucleation of oral microbial consortia leading to biofilm formation. Due to the lack of genetic tools, little is known about basic cellular processes, including protein secretion and folding, in this organism. We report here a survey of the C. matruchotii genome, which encodes a large number of exported proteins containing paired cysteine residues, and identified an oxidoreductase that is highly homologous to the Corynebacterium diphtheriae thiol-disulfide oxidoreductase MdbA (MdbA Cd ). Crystallization studies uncovered that the 1.2-Å resolution structure of C. matruchotii MdbA (MdbA Cm ) possesses two conserved features found in actinobacterial MdbA enzymes, a thioredoxin-like fold and an extended α-helical domain. By reconstituting the disulfide bond-forming machine in vitro , we demonstrated that MdbA Cm catalyzes disulfide bond formation within the actinobacterial pilin FimA. A new gene deletion method supported that mdbA is essential in C. matruchotii Remarkably, heterologous expression of MdbA Cm in the C. diphtheriae Δ mdbA mutant rescued its known defects in cell growth and morphology, toxin production, and pilus assembly, and this thiol-disulfide oxidoreductase activity required the catalytic motif CXXC. Altogether, the results suggest that MdbA Cm is a major thiol-disulfide oxidoreductase, which likely mediates posttranslocational protein folding in C. matruchotii by a mechanism that is conserved in Actinobacteria IMPORTANCE The actinobacterium Corynebacterium matruchotii has been implicated in the development of oral biofilms or dental plaque; however, little is known about the basic cellular processes in this organism. We report here a high-resolution structure of a C. matruchotii oxidoreductase that is highly homologous to the Corynebacterium diphtheriae thiol-disulfide oxidoreductase MdbA. By biochemical analysis, we demonstrated that C. matruchotii MdbA catalyzes disulfide

Activation of the Dormant Secondary Metabolite Production by Introducing Gentamicin-Resistance in a Marine-Derived Penicillium purpurogenum G59

PubMed Central

Chai, Yun-Jing; Cui, Cheng-Bin; Li, Chang-Wei; Wu, Chang-Jing; Tian, Cong-Kui; Hua, Wei

2012-01-01

A new approach to activate silent gene clusters for dormant secondary metabolite production has been developed by introducing gentamicin-resistance to an originally inactive, marine-derived fungal strain Penicillium purpurogenum G59. Upon treatment of the G59 spores with a high concentration of gentamicin in aqueous DMSO, a total of 181 mutants were obtained by single colony isolation. In contrast to the strain G59, the EtOAc extracts of nine mutant cultures showed inhibitory effects on K562 cells, indicating that the nine mutants had acquired capability to produce antitumor metabolites. This was evidenced by TLC and HPLC analysis of EtOAc extracts of G59 and the nine mutants. Further isolation and characterization demonstrated that four antitumor secondary metabolites, janthinone (1), fructigenine A (2), aspterric acid methyl ester (3) and citrinin (4), were newly produced by mutant 5-1-4 compared to the parent strain G59, and which were also not found in the secondary metabolites of other Penicillium purpurogenum strains. However, Compounds 1–4 inhibited the proliferation of K562 cells with inhibition rates of 34.6% (1), 60.8% (2), 31.7% (3) and 67.1% (4) at 100 μg/mL, respectively. The present study demonstrated the effectiveness of a simple, yet practical approach to activate the production of dormant fungal secondary metabolites by introducing acquired resistance to aminoglycoside antibiotics, which could be applied to the studies for eliciting dormant metabolic potential of fungi to obtain cryptic secondary metabolites. PMID:22611354

Actions of incretin metabolites on locomotor activity, cognitive function and in vivo hippocampal synaptic plasticity in high fat fed mice.

PubMed

Porter, David; Faivre, Emilie; Flatt, Peter R; Hölscher, Christian; Gault, Victor A

2012-05-01

The incretin hormones glucagon-like peptide-1 (GLP-1) and glucose-dependent insulinotropic polypeptide (GIP) improve markers of cognitive function in obesity-diabetes, however, both are rapidly degraded to their major metabolites, GLP-1(9-36)amide and GIP(3-42), respectively. Therefore, the present study investigated effects of GLP-1(9-36)amide and GIP(3-42) on locomotor activity, cognitive function and hippocampal synaptic plasticity in mice with diet-induced obesity and insulin resistance. High-fat fed Swiss TO mice treated with GLP-1(9-36)amide, GIP(3-42) or exendin(9-39)amide (twice-daily for 60 days) did not exhibit any changes in bodyweight, non-fasting plasma glucose and plasma insulin concentrations or glucose tolerance compared with high-fat saline controls. Similarly, locomotor and feeding activity, O(2) consumption, CO(2) production, respiratory exchange ratio and energy expenditure were not altered by chronic treatment with incretin metabolites. Administration of the truncated metabolites did not alter general behavior in an open field test or learning and memory ability as recorded during an object recognition test. High-fat mice exhibited a significant impairment in hippocampal long-term potentiation (LTP) which was not affected by treatment with incretin metabolites. These data indicate that incretin metabolites do not influence locomotor activity, cognitive function and hippocampal synaptic plasticity when administered at pharmacological doses to mice fed a high-fat diet. Copyright © 2012 Elsevier Inc. All rights reserved.

Screening of marine fungus from Nanji Island and activity of their metabolites against pathogenic Vibrio from Pseudosciaena crocea

NASA Astrophysics Data System (ADS)

Zhao, Shujiang; Li, Shuping; Liu, Huihui; Zhao, Qian; Wang, Jieyou; Yan, Maocang

2012-09-01

Seventy-eight marine fungal strains were isolated from sediment samples collected off the coast of Nanji Island, Wenzhou, Zhejiang Province, China. Antibacterial screening using the agar disc method showed that 19 of the isolated strains could inhibit at least one pathogenic V ibrio from P seudosciaena crocea. Subsequent screening confirmed that nine strains produced antibacterial metabolites that had activity against one or several types of pathogenic V ibrio. Strain NJ0104 had the widest antimicrobial spectrum and strong activity, particularly against Vibrio parahaemolyticus-MM0810072. A preliminary study of NJ0104 antibacterial metabolites demonstrated that they had thermal stability up to 80°C, ultraviolet stability up to 40 min and pH stability between 4.0-7.0. In addition, the antibacterial metabolites were readily soluble in butanol. To identify the specific strain, the ITS-5.8S rDNA regions of NJ0104 were PCR amplified and sequenced. Based on the combination of phenotypic and genotypic data, the strain was identified as Arthrinium sp.

Smelling Sulfur: Copper and Silver Regulate the Response of Human Odorant Receptor OR2T11 to Low-Molecular-Weight Thiols.

PubMed

Li, Shengju; Ahmed, Lucky; Zhang, Ruina; Pan, Yi; Matsunami, Hiroaki; Burger, Jessica L; Block, Eric; Batista, Victor S; Zhuang, Hanyi

2016-10-03

Mammalian survival depends on ultrasensitive olfactory detection of volatile sulfur compounds, since these compounds can signal the presence of rancid food, O 2 depleted atmospheres, and predators (through carnivore excretions). Skunks exploit this sensitivity with their noxious spray. In commerce, natural and liquefied gases are odorized with t-BuSH and EtSH, respectively, as warnings. The 100-million-fold difference in olfactory perception between structurally similar EtSH and EtOH has long puzzled those studying olfaction. Mammals detect thiols and other odorants using odorant receptors (ORs), members of the family of seven transmembrane G-protein-coupled receptors (GPCRs). Understanding the regulator cofactors and response of ORs is particularly challenging due to the lack of X-ray structural models. Here, we combine computational modeling and site-directed mutagenesis with saturation transfer difference (STD) NMR spectroscopy and measurements of the receptor response profiles. We find that human thiol receptor OR2T11 responds specifically to gas odorants t-BuSH and EtSH requiring ionic copper for its robust activation and that this role of copper is mimicked by ionic and nanoparticulate silver. While copper is both an essential nutrient for life and, in excess, a hallmark of various pathologies and neurodegenerative diseases, its involvement in human olfaction has not been previously demonstrated. When screened against a series of alcohols, thiols, sulfides, and metal-coordinating ligands, OR2T11 responds with enhancement by copper to the mouse semiochemical CH 3 SCH 2 SH and derivatives, to four-membered cyclic sulfide thietane and to one- to four-carbon straight- and branched-chain and five-carbon branched-chain thiols but not to longer chain thiols, suggesting compact receptor dimensions. Alcohols are unreactive.

Thiol X Click Foldamers for Polymer Affinity

DTIC Science & Technology

2016-06-24

polymers   e. Invention  of  a  novel,  robust,  and  ambient   polymerization ...efficiently   polymerized   to   moderate  sized   polymers  capable  of  forming  >>1012  sequence  distinct   polymers ... polymerization  of  nucleobase  appended   thiol-­‐ene  monomers.    Naturally,   the  average   composition  of  the  

In vitro antiprogestational/antiglucocorticoid activity and progestin and glucocorticoid receptor binding of the putative metabolites and synthetic derivatives of CDB-2914, CDB-4124, and mifepristone.

PubMed

Attardi, Barbara J; Burgenson, Janet; Hild, Sheri A; Reel, Jerry R

2004-03-01

In determining the biological profiles of various antiprogestins, it is important to assess the hormonal and antihormonal activity, selectivity, and potency of their proximal metabolites. The early metabolism of mifepristone is characterized by rapid demethylation and hydroxylation. Similar initial metabolic pathways have been proposed for CDB-2914 (CDB: Contraceptive Development Branch of NICHD) and CDB-4124, and their putative metabolites have been synthesized. We have examined the functional activities and potencies, in various cell-based assays, and relative binding affinities (RBAs) for progesterone receptors (PR) and glucocorticoid receptors (GR) of the putative mono- and didemethylated metabolites of CDB-2914, CDB-4124, and mifepristone and of the 17alpha-hydroxy and aromatic A-ring derivatives of CDB-2914 and CDB-4124. The binding affinities of the monodemethylated metabolites for rabbit uterine PR and human PR-A and PR-B were similar to those of the parent compounds. Monodemethylated mifepristone bound to rabbit thymic GR with higher affinity than monodemethylated CDB-2914 or CDB-4124. T47D-CO cells were used to assess inhibition of R5020-stimulated endogenous alkaline phosphatase activity and transactivation of the PRE(2)-thymidine kinase (tk)-luciferase (LUC) reporter plasmid in transient transfections. The antiprogestational potency was as follows: mifepristone/CDB-2914/CDB-4124/monodemethylated metabolites (IC(50)'s approximately 10(-9)M) > aromatic A-ring derivatives (IC(50)'s approximately 10(-8)M) > didemethylated/17alpha-hydroxy derivatives (IC(50)'s approximately 10(-7)M). Antiglucocorticoid activity was determined by inhibition of dexamethasone-stimulated transcriptional activity in HepG2 cells. The mono- and didemethylated metabolites of CDB-2914 and CDB-4124 had less antiglucocorticoid activity (IC(50)'s approximately 10(-6)M) than monodemethylated mifepristone (IC(50) approximately 10(-8)M) or the other test compounds. At 10(-6)M in

Allocation of secondary metabolites, photosynthetic capacity, and antioxidant activity of Kacip Fatimah (Labisia pumila Benth) in response to CO2 and light intensity.

PubMed

Ibrahim, Mohd Hafiz; Jaafar, Hawa Z E; Karimi, Ehsan; Ghasemzadeh, Ali

2014-01-01

A split plot 3 by 4 experiment was designed to investigate and distinguish the relationships among production of secondary metabolites, soluble sugar, phenylalanine ammonia lyase (PAL; EC 4.3.1.5) activity, leaf gas exchange, chlorophyll content, antioxidant activity (DPPH), and lipid peroxidation under three levels of CO2 (400, 800, and 1200 μ mol/mol) and four levels of light intensity (225, 500, 625, and 900 μ mol/m(2)/s) over 15 weeks in Labisia pumila. The production of plant secondary metabolites, sugar, chlorophyll content, antioxidant activity, and malondialdehyde content was influenced by the interactions between CO2 and irradiance. The highest accumulation of secondary metabolites, sugar, maliondialdehyde, and DPPH activity was observed under CO2 at 1200 μ mol/mol + light intensity at 225 μ mol/m(2)/s. Meanwhile, at 400 μ mol/mol CO2 + 900 μ mol/m(2)/s light intensity the production of chlorophyll and maliondialdehyde content was the highest. As CO2 levels increased from 400 to 1200 μ mol/mol the photosynthesis, stomatal conductance, f v /f m (maximum efficiency of photosystem II), and PAL activity were enhanced. The production of secondary metabolites displayed a significant negative relationship with maliondialdehyde indicating lowered oxidative stress under high CO2 and low irradiance improved the production of plant secondary metabolites that simultaneously enhanced the antioxidant activity (DPPH), thus improving the medicinal value of Labisia pumila under this condition.

Trichloroethylene and Its Oxidative Metabolites Enhance the Activated State and Th1 Cytokine Gene Expression in Jurkat Cells.

PubMed

Pan, Yao; Wei, Xuetao; Hao, Weidong

2015-08-28

Trichloroethylene (TCE) is an occupational and ubiquitous environmental contaminant, and TCE exposure will increase the risk of autoimmune diseases and allergic diseases. T cells play an important role in the pathogenesis of TCE-related immune disorders, but the effect of TCE and its oxidative metabolites, trichloroacetic acid (TCA) and dichloroacetic acid (DCA), on the activation of human T cells is still unknown. In this study, Jurkat cells were pre-treated with TCE, TCA and DCA overnight and then stimulated with phorbol 12-myristate 13-acetate and ionomycin for another 4, 8 and 24 hours. IL-2 secretion was detected by ELISA; the expressions of CD25 and CD69 were tested by flow cytometry; and IFN-γ and IL-2 mRNA expression levels were investigated by real-time PCR. The results showed that TCE and its oxidative metabolites, TCA and DCA, significantly enhanced IL-2 releasing and the expression of T cell activation markers, CD25 and CD69. Consistent with this result, these compounds markedly up-regulated the expression levels of IFN-γ and IL-2 mRNA. Collectively, these findings suggest that TCE and its metabolites, TCA and DCA, might enhance the activation of T cells and disrupt various activities of peripheral T cells.

Trichloroethylene and Its Oxidative Metabolites Enhance the Activated State and Th1 Cytokine Gene Expression inJurkat Cells

PubMed Central

Pan, Yao; Wei, Xuetao; Hao, Weidong

2015-01-01

Trichloroethylene (TCE) is an occupational and ubiquitous environmental contaminant, and TCE exposure will increase the risk of autoimmune diseases and allergic diseases. T cells play an important role in the pathogenesis of TCE-related immune disorders, but the effect of TCE and its oxidative metabolites, trichloroacetic acid (TCA) and dichloroacetic acid (DCA), on the activation of human T cells is still unknown. In this study, Jurkat cells were pre-treated with TCE, TCA and DCA overnight and then stimulated with phorbol 12-myristate 13-acetate and ionomycin for another 4, 8 and 24 hours. IL-2 secretion was detected by ELISA; the expressions of CD25 and CD69 were tested by flow cytometry; and IFN-γ and IL-2 mRNA expression levels were investigated by real-time PCR. The results showed that TCE and its oxidative metabolites, TCA and DCA, significantly enhanced IL-2 releasing and the expression of T cell activation markers, CD25 and CD69. Consistent with this result, these compounds markedly up-regulated the expression levels of IFN-γ and IL-2 mRNA. Collectively, these findings suggest that TCE and its metabolites, TCA and DCA, might enhance the activation of T cells and disrupt various activities of peripheral T cells. PMID:26343699

Thiol-vinyl systems as shape memory polymers and novel two-stage reactive polymer systems

NASA Astrophysics Data System (ADS)

Nair, Devatha P.

2011-12-01

The focus of this research was to formulate, characterize and tailor the reaction methodologies and material properties of thiol-vinyl systems to develop novel polymer platforms for a range of engineering applications. Thiol-ene photopolymers were demonstrated to exhibit several advantageous characteristics for shape memory polymer systems for a range of biomedical applications. The thiol-ene shape memory polymer systems were tough and flexible as compared to the acrylic control systems with glass transition temperatures between 30 and 40 °C; ideal for actuation at body temperature. The thiol-ene polymers also exhibited excellent shape fixity and a rapid and distinct shape memory actuation response along with free strain recoveries of greater than 96% and constrained stress recoveries of 100%. Additionally, two-stage reactive thiol-acrylate systems were engineered as a polymer platform technology enabling two independent sets of polymer processing and material properties. There are distinct advantages to designing polymer systems that afford two distinct sets of material properties -- an intermediate polymer that would enable optimum handling and processing of the material (stage 1), while maintaining the ability to tune in different, final properties that enable the optimal functioning of the polymeric material (stage 2). To demonstrate the range of applicability of the two-stage reactive systems, three specific applications were demonstrated; shape memory polymers, lithographic impression materials, and optical materials. The thiol-acrylate reactions exhibit a wide range of application versatility due to the range of available thiol and acrylate monomers as well as reaction mechanisms such as Michael Addition reactions and free radical polymerizations. By designing a series of non-stoichiometeric thiol-acrylate systems, a polymer network is initially formed via a base catalyzed 'click' Michael addition reaction. This self-limiting reaction results in a Stage 1

[Secondary metabolites, lethality and antimicrobial activity of extracts from three corals and three marine mollusks from Sucre, Venezuela].

PubMed

Ordaz, Gabriel; D'Armas, Haydelba; Yáñez, Dayanis; Hernández, Juan; Camacho, Angel

2010-06-01

The study of biochemical activity of extracts obtained from marine organisms is gaining interest as some have proved to have efficient health or industrial applications. To evaluate lethality and antimicrobial activities, some chemical tests were performed on crude extracts of the octocorals Eunicea sp., Muricea sp. and Pseudopterogorgia acerosa and the mollusks Pteria colymbus, Phyllonotus pomum and Chicoreus brevifrons, collected in Venezuelan waters. The presence of secondary metabolites like alkaloids, unsaturated sterols and pentacyclic triterpenes in all invertebrates, was evidenced. Additionally, sesquiterpenlactones, saponins, tannins, cyanogenic and cardiotonic glycosides were also detected in some octocoral extracts, suggesting that biosynthesis of these metabolites is typical in this group. From the lethality bioassays, all extracts resulted lethal to Artemia salina (LC50<1000 microg/ml) with an increased of lethal activity with exposition time. P. pomum extract showed the highest lethality rate (LC50=46.8 microg/ml). Compared to the octocorals, mollusks extracts displayed more activity and a greater action spectrum against different bacterial strains, whereas octocorals also inhibited some fungi strains growth. Staphylococcus aureus was the most susceptible to the antimicrobial power of the extracts (66.7%), whereas Pseudomonas aeruginosa, Candida albicans and Aspergillus niger were not affected. The antibiosis shown by marine organisms extracts indicates that some of their biosynthesized metabolites are physiologically active, and may have possible cytotoxic potential or as a source of antibiotic components.

Comparative evaluation of two Trichoderma harzianum strains for major secondary metabolite production and antifungal activity.

PubMed

Ahluwalia, Vivek; Kumar, Jitendra; Rana, Virendra S; Sati, Om P; Walia, S

2015-01-01

This investigation was undertaken to identify the major secondary metabolite, produced by two Trichoderma harzianum strains (T-4 and T-5) with their antifungal activity against phytopathogenic fungi using poison food technique. The ethyl acetate extract was subjected to column chromatography using n-hexane, ethyl acetate and methanol gradually. Chromatographic separation of ethyl acetate extract of T. harzianum (T-4) resulted in the isolation and identification of palmitic acid (1), 1,8-dihydroxy-3-methylanthraquinone (2), 6-pentyl-2H-pyran-2-one (3), 2(5H)-furanone (4), stigmasterol (5) and β-sitosterol (6), while T. harzianum (T-5) gave palmitic acid (1), 1-hydroxy-3-methylanthraquinone (7), δ-decanolactone (8), 6-pentyl-2H-pyran-2-one (3), ergosterol (9), harzianopyridone (10) and 6-methyl-1,3,8-trihydroxyanthraquinone (11) as major metabolites. Among compounds screened for antifungal activity, compound 10 was found to be most active (EC50 35.9-50.2 μg mL(-1)). In conclusion, the present investigation provided significant information about antifungal activity and compounds isolated from two different strains of T. harzianum obtained from two different Himalayan locations.

Biosynthesis of human diazepam and clonazepam metabolites.

PubMed

de Paula, Núbia C; Araujo Cordeiro, Kelly C F; de Melo Souza, Paula L; Nogueira, Diogo F; da Silva e Sousa, Diego B; Costa, Maísa B; Noël, François; de Oliveira, Valéria

2015-03-01

A screening of fungal and microbial strains allowed to select the best microorganisms to produce in high yields some of the human metabolites of two benzodiazepine drugs, diazepam and clonazepam, in order to study new pharmacological activities and for chemical standard proposes. Among the microorganisms tested, Cunninghamella echinulata ATCC 9244 and Rhizopus arrhizus ATCC 11145 strains, were the most active producers of the mains metabolites of diazepam which included demethylated, hydroxylated derivatives. Beauveria bassiana ATCC 7159 and Chaetomium indicum LCP 984200 produced the 7 amino-clonazepam metabolite and a product of acid hydrolysis of this benzodiazepine. Copyright © 2015 Elsevier Ltd. All rights reserved.

Modeling of S-Nitrosothiol-Thiol Reactions of Biological Significance: HNO Production by S-Thiolation Requires a Proton Shuttle and Stabilization of Polar Intermediates.

PubMed

Ivanova, Lena V; Cibich, Daniel; Deye, Gregory; Talipov, Marat R; Timerghazin, Qadir K

2017-04-18

Nitroxyl (HNO), a reduced form of the important gasotransmitter nitric oxide, exhibits its own unique biological activity. A possible biological pathway of HNO formation is the S-thiolation reaction between thiols and S-nitrosothiols (RSNOs). Our density functional theory (DFT) calculations suggested that S-thiolation proceeds through a proton transfer from the thiol to the RSNO nitrogen atom, which increases electrophilicity of the RSNO sulfur, followed by nucleophilic attack by thiol, yielding a charge-separated zwitterionic intermediate structure RSS + (R)N(H)O - (Zi), which decomposes to yield HNO and disulfide RSSR. In the gas phase, the proton transfer and the S-S bond formation are asynchronous, resulting in a high activation barrier (>40 kcal mol -1 ), making the reaction infeasible. However, the barrier can decrease below the S-N bond dissociation energy in RSNOs (≈30 kcal mol -1 ) upon transition into an aqueous environment that stabilizes Zi and provides a proton shuttle to synchronize the proton transfer and the S-S bond formation. These mechanistic features suggest that S-thiolation can easily lend itself to enzymatic catalysis and thus can be a possible route of endogenous HNO production. © 2017 Wiley-VCH Verlag GmbH & Co. KGaA, Weinheim.

Antitrypanosomal Activity of Fexinidazole Metabolites, Potential New Drug Candidates for Chagas Disease

PubMed Central

Nascimento, Alvaro F. S.; Mazzeti, Ana Lia; Marques, Luiz F.; Gonçalves, Karolina R.; Mota, Ludmilla W. R.; Diniz, Lívia de F.; Caldas, Ivo S.; Talvani, André; Shackleford, David M.; Koltun, Maria; Saunders, Jessica; White, Karen L.; Scandale, Ivan; Charman, Susan A.; Chatelain, Eric

2014-01-01

This study was designed to verify the in vivo efficacy of sulfoxide and sulfone fexinidazole metabolites following oral administration in a murine model of Chagas disease. Female Swiss mice infected with the Y strain of Trypanosoma cruzi were treated orally once per day with each metabolite at doses of 10 to 100 mg/kg of body weight for a period of 20 days. Parasitemia was monitored throughout, and cures were detected by parasitological and PCR assays. The results were compared with those achieved with benznidazole treatment at the same doses. Fexinidazole metabolites were effective in reducing the numbers of circulating parasites and protecting mice against death, compared with untreated mice, but without providing cures at daily doses of 10 and 25 mg/kg. Both metabolites were effective in curing mice at 50 mg/kg/day (30% to 40%) and 100 mg/kg/day (100%). In the benznidazole-treated group, parasitological cure was detected only in animals treated with the higher dose of 100 mg/kg/day (80%). Single-dose pharmacokinetic parameters for each metabolite were obtained from a parallel group of uninfected mice and were used to estimate the profiles following repeated doses. Pharmacokinetic data suggested that biological efficacy most likely resides with the sulfone metabolite (or subsequent reactive metabolites formed following reduction of the nitro group) following administration of either the sulfoxide or the sulfone and that prolonged plasma exposure over the 24-h dosing window is required to achieve high cure rates. Fexinidazole metabolites were effective in treating T. cruzi in a mouse model of acute infection, with cure rates superior to those achieved with either fexinidazole itself or benznidazole. PMID:24841257

Redox Proteomics Applied to the Thiol Secretome.

PubMed

Ghezzi, Pietro; Chan, Philippe

2017-03-01

Secreted proteins are important both as signaling molecules and potential biomarkers. Recent Advances: Protein can undergo different types of oxidation, both in physiological conditions or under oxidative stress. Several redox proteomics techniques have been successfully applied to the identification of glutathionylated proteins, an oxidative post-translational modification consisting in the formation of a mixed disulfide between a protein cysteine and glutathione. Redox proteomics has also been used to study other forms of protein oxidation. Because of the highest proportion of free cysteines in the cytosol, redox proteomics of protein thiols has focused, so far, on intracellular proteins. However, plasma proteins, such as transthyretin and albumin, have been described as glutathionylated or cysteinylated. The present review discusses the redox state of protein cysteines in relation to their cellular distribution. We describe the various approaches used to detect secreted glutathionylated proteins, the only thiol modification studied so far in secreted proteins, and the specific problems presented in the study of the secretome. This review focusses on glutathionylated proteins secreted under inflammatory conditions and that may act as soluble mediators (cytokines). Future studies on the redox secretome (including other forms of oxidation) might identify new soluble mediators and biomarkers of oxidative stress. Antioxid. Redox Signal. 26, 299-312.

Stoichiometry of mercury-thiol complexes on bacterial cell envelopes

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

Mishra, Bhoopesh; Shoenfelt, Elizabeth; Yu, Qiang

We have examined the speciation of Hg(II) complexed with intact cell suspensions (1013 cells L- 1) of Bacillus subtilis, a common gram-positive soil bacterium, Shewanella oneidensis MR-1, a facultative gram-negative aquatic organism, and Geobacter sulfurreducens, a gram-negative anaerobic bacterium capable of Hg-methylation at Hg(II) loadings spanning four orders of magnitude (120 nM to 350 μM) at pH 5.5 (± 0.2). The coordination environments of Hg on bacterial cells were analyzed using synchrotron based X-ray Absorption Near Edge Structure (XANES) and Extended X-ray Absorption Fine Structure (EXAFS) spectroscopy at the Hg LIII edge. The abundance of thiols on intact cells wasmore » determined by a fluorescence-spectroscopy based method using a soluble bromobimane, monobromo(trimethylammonio)bimane (qBBr) to block thiol sites, and potentiometric titrations of biomass with and without qBBr treatment. The chemical forms of S on intact bacterial cells were determined using S k-edge XANES spectroscopy.« less

Arsenolysis and Thiol-Dependent Arsenate Reduction

EPA Science Inventory

Conversion of arsenate to arsenite is a critical event in the pathway that leads from inorganic arsenic to a variety of methylated metabolites. The formation of methylated metabolites influences distribution and retention of arsenic and affects the reactivity and toxicity of thes...

Microsomal metabolism of trenbolone acetate metabolites ...

EPA Pesticide Factsheets

Trenbolone acetate (TBA) is a synthetic growth promoter widely used in animal agriculture, and its metabolites are suspected endocrine disrupting compounds in agriculturally impacted receiving waters. However, beyond the three widely recognized TBA metabolites (17-trenbolone, 17-trenbolone and trendione), little is known about other metabolites formed in vivo and subsequently discharged into the environment, with some evidence suggesting these unknown metabolites comprise a majority of the TBA mass dosed to the animal. Here, we explored the metabolism of the three known TBA metabolites using rat liver microsome studies. All TBA metabolites are transformed into a complex mixture of monohydroxylated products. Based on product characterization, the majority are more polar than the parent metabolites but maintain their characteristic trienone backbone. A minor degree of interconversion between known metabolites was also observed, as were higher order hydroxylated products with a greater extent of reaction. Notably, the distribution and yield of products were generally comparable across a series of variably induced rat liver microsomes, as well as during additional studies with human and bovine liver microsomes. Bioassays conducted with mixtures of these transformation products suggest that androgen receptor (AR) binding activity is diminished as a result of the microsomal treatment, suggesting that the transformation products are generally less potent than

Activation of cryptic metabolite production through gene disruption: Dimethyl furan-2,4-dicarboxylate produced by Streptomyces sahachiroi.

PubMed

Simkhada, Dinesh; Zhang, Huitu; Mori, Shogo; Williams, Howard; Watanabe, Coran M H

2013-01-01

At least 65% of all small molecule drugs on the market today are natural products, however, re-isolation of previously identified and characterized compounds has become a serious impediment to the discovery of new bioactive natural products. Here, genetic knockout of an unusual non-ribosomal peptide synthetase (NRPS) C-PCP-C module, aziA2, is performed resulting in the accumulation of the secondary metabolite, dimethyl furan-2,4-dicarboxylate. The cryptic metabolite represents the first non-azinomycin related compound to be isolated and characterized from the soil bacterium, S. sahachiroi. The results from this study suggest that abolishing production of otherwise predominant natural products through genetic knockout may constitute a means to "activate" the production of novel secondary metabolites that would otherwise lay dormant within microbial genome sequences.

Biotransformation of Bisphenol AF to Its Major Glucuronide Metabolite Reduces Estrogenic Activity

PubMed Central

Yin, Jie; Zhang, Jing; Feng, Yixing; Shao, Bing

2013-01-01

Bisphenol AF (BPAF), an endocrine disrupting chemical, can induce estrogenic activity through binding to estrogen receptor (ER). However, the metabolism of BPAF in vivo and the estrogenic activity of its metabolites remain unknown. In the present study, we identified four metabolites including BPAF diglucuronide, BPAF glucuronide (BPAF-G), BPAF glucuronide dehydrated and BPAF sulfate in the urine of Sprague-Dawley (SD) rats. BPAF-G was further characterized by nuclear magnetic resonance (NMR). After treatment with a single dose of BPAF, BPAF was metabolized rapidly to BPAF-G, as detected in the plasma of SD rats. Biotransformation of BPAF to BPAF-G was confirmed with human liver microsomes (HLM), and Vmax of glucuronidation for HLM was 11.6 nmol/min/mg. We also found that BPAF glucuronidation could be mediated through several human recombinant UDP-glucuronosyltransferases (UGTs) including UGT1A1, UGT1A3, UGT1A8, UGT1A9, UGT2B4, UGT2B7, UGT2B15 and UGT2B17, among which UGT2B7 showed the highest efficiency of glucuronidation. To explain the biological function of BPAF biotransformation, the estrogenic activities of BPAF and BPAF-G were evaluated in ER-positive breast cancer T47D and MCF7 cells. BPAF significantly stimulates ER-regulated gene expression and cell proliferation at the dose of 100 nM and 1 μM in breast cancer cells. However, BPAF-G did not show any induction of estrogenic activity at the same dosages, implying that formation of BPAF-G is a potential host defense mechanism against BPAF. Based on our study, biotransformation of BPAF to BPAF-G can eliminate BPAF-induced estrogenic activity, which is therefore considered as reducing the potential threat to human beings. PMID:24349450

Photoreduction of Hg(II) and photodemethylation of methylmercury: the key role of thiol sites on dissolved organic matter

USGS Publications Warehouse

Jeremiason, Jeffrey D.; Portner, Joshua C.; Aiken, George R.; Hiranaka, Amber J.; Dvorak, Michelle T.; Tran, Khuyen T.; Latch, Douglas E.

2015-01-01

This study examined the kinetics of photoreduction of Hg(II) and photodemethylation of methylmercury (MeHg+) attached to, or in the presence of, dissolved organic matter (DOM). Both Hg(II) and MeHg+ are principally bound to reduced sulfur groups associated with DOM in many freshwater systems. We propose that a direct photolysis mechanism is plausible for reduction of Hg(II) bound to reduced sulfur groups on DOM while an indirect mechanism is supported for photodemethylation of MeHg+ bound to DOM. UV spectra of Hg(II) and MeHg+ bound to thiol containing molecules demonstrate that the Hg(II)–S bond is capable of absorbing UV-light in the solar spectrum to a much greater extent than MeHg+–S bonds. Experiments with chemically distinct DOM isolates suggest that concentration of DOM matters little in the photochemistry if there are enough reduced S sites present to strongly bind MeHg+ and Hg(II); DOM concentration does not play a prominent role in photodemethylation other than to screen light, which was demonstrated in a field experiment in the highly colored St. Louis River where photodemethylation was not observed at depths ≥10 cm. Experiments with thiol ligands yielded slower photodegradation rates for MeHg+ than in experiments with DOM and thiols; rates in the presence of DOM alone were the fastest supporting an intra-DOM mechanism. Hg(II) photoreduction rates, however, were similar in experiments with only DOM, thiols plus DOM, or only thiols suggesting a direct photolysis mechanism. Quenching experiments also support the existence of an intra-DOM photodemethylation mechanism for MeHg+. Utilizing the difference in photodemethylation rates measured for MeHg+ attached to DOM or thiol ligands, the binding constant for MeHg+ attached to thiol groups on DOM was estimated to be 1016.7.

Influence of Torulaspora delbrueckii in varietal thiol (3-SH and 4-MSP) release in wine sequential fermentations.

PubMed

Belda, Ignacio; Ruiz, Javier; Beisert, Beata; Navascués, Eva; Marquina, Domingo; Calderón, Fernando; Rauhut, Doris; Benito, Santiago; Santos, Antonio

2017-09-18

In last years, non-Saccharomyces yeasts have emerged as innovative tools to improve wine quality, being able to modify the concentration of sensory-impact compounds. Among them, varietal thiols released by yeasts, play a key role in the distinctive aroma of certain white wines. In this context, Torulaspora delbrueckii is in the spotlight because of its positive contribution to several wine quality parameters. This work studies the physiological properties of an industrial T. delbrueckii strain, for the production of wines with increased thiol concentrations. IRC7 gene, previously described in S. cerevisiae, has been identified in T. delbrueckii, establishing the genetics basis of its thiol-releasing capability. Fermentations involving T. delbrueckii showed improvements on several parameters (such as glycerol content, ethanol index, and major volatile compounds composition), but especially on thiols release. These results confirm the potential of T. delbrueckii on wine improvement, describing new metabolic features regarding the release of cysteinylated aroma precursors. Copyright © 2017 Elsevier B.V. All rights reserved.

Allocation of Secondary Metabolites, Photosynthetic Capacity, and Antioxidant Activity of Kacip Fatimah (Labisia pumila Benth) in Response to CO2 and Light Intensity

PubMed Central

Jaafar, Hawa Z. E.; Karimi, Ehsan; Ghasemzadeh, Ali

2014-01-01

A split plot 3 by 4 experiment was designed to investigate and distinguish the relationships among production of secondary metabolites, soluble sugar, phenylalanine ammonia lyase (PAL; EC 4.3.1.5) activity, leaf gas exchange, chlorophyll content, antioxidant activity (DPPH), and lipid peroxidation under three levels of CO2 (400, 800, and 1200 μmol/mol) and four levels of light intensity (225, 500, 625, and 900 μmol/m2/s) over 15 weeks in Labisia pumila. The production of plant secondary metabolites, sugar, chlorophyll content, antioxidant activity, and malondialdehyde content was influenced by the interactions between CO2 and irradiance. The highest accumulation of secondary metabolites, sugar, maliondialdehyde, and DPPH activity was observed under CO2 at 1200 μmol/mol + light intensity at 225 μmol/m2/s. Meanwhile, at 400 μmol/mol CO2 + 900 μmol/m2/s light intensity the production of chlorophyll and maliondialdehyde content was the highest. As CO2 levels increased from 400 to 1200 μmol/mol the photosynthesis, stomatal conductance, f v/f m (maximum efficiency of photosystem II), and PAL activity were enhanced. The production of secondary metabolites displayed a significant negative relationship with maliondialdehyde indicating lowered oxidative stress under high CO2 and low irradiance improved the production of plant secondary metabolites that simultaneously enhanced the antioxidant activity (DPPH), thus improving the medicinal value of Labisia pumila under this condition. PMID:24683336

One-step fabrication of PEGylated fluorescent nanodiamonds through the thiol-ene click reaction and their potential for biological imaging

NASA Astrophysics Data System (ADS)

Huang, Hongye; Liu, Meiying; Tuo, Xun; Chen, Junyu; Mao, Liucheng; Wen, Yuanqing; Tian, Jianwen; Zhou, Naigen; Zhang, Xiaoyong; Wei, Yen

2018-05-01

Over the past years, fluorescent carbon nanoparticles have got growing interest for biological imaging. Fluorescent nanodiamonds (FNDs) are novel fluorescent carbon nanoparticles with multitudinous useful properties, including remarkable fluorescence properties, extremely low toxicity and high refractive index. However, facile preparation of FNDs with designable properties and functions from non-fluorescent detonation nanodiamonds (DNDs) has demonstrated to be challengeable. In this work, we reported for the first time that preparation of Polyethylene glycol (PEG) functionalized FNDs through a one-step thiol-ene click reaction using thiol containing PEG (PEG-SH) as the coating agent. Based on the characterization results, we demonstrated that PEG-SH could be efficiently introduced on DNDs to obtain FNDs through the thiol-ene click chemistry. The resultant FND-PEG composites showed high water dispersibility, strong fluorescence and low cytotoxicity. Moreover, FND-PEG composites could be internalized by cells and displayed good cell dyeing performance. All of these features implied that FND-PEG composites are of great potential for biological imaging. Taken together, a facile one-step strategy based on the one-step thiol-ene click reaction has been developed for efficient preparation of FND-PEG composites from non-fluorescent DNDs. The strategy should be also useful for fabrication of many other functional FNDs via using different thiol containing compounds for the universality of thiol-ene click reaction.

Characterization of thiol-based redox modifications of Brassica napusSNF1-related protein kinase 2.6-2C.

PubMed

Ma, Tianyi; Yoo, Mi-Jeong; Zhang, Tong; Liu, Lihong; Koh, Jin; Song, Wen-Yuan; Harmon, Alice C; Sha, Wei; Chen, Sixue

2018-04-01

Sucrose nonfermenting 1-related protein kinase 2.6 (SnRK2.6), also known as Open Stomata 1 (OST1) in Arabidopsis thaliana , plays a pivotal role in abscisic acid (ABA)-mediated stomatal closure. Four SnRK2.6 paralogs were identified in the Brassica napus genome in our previous work. Here we studied one of the paralogs, BnSnRK2.6-2C , which was transcriptionally induced by ABA in guard cells. Recombinant BnSnRK2.6-2C exhibited autophosphorylation activity and its phosphorylation sites were mapped. The autophosphorylation activity was inhibited by S-nitrosoglutathione (GSNO) and by oxidized glutathione (GSSG), and the inhibition was reversed by reductants. Using monobromobimane (mBBr) labeling, we demonstrated a dose-dependent modification of BnSnRK2.6-2C by GSNO. Furthermore, mass spectrometry analysis revealed previously uncharacterized thiol-based modifications including glutathionylation and sulfonic acid formation. Of the six cysteine residues in BnSnRK2.6-2C, C159 was found to have different types of thiol modifications, suggesting its high redox sensitivity and versatility. In addition, mBBr labeling on tyrosine residues was identified. Collectively, these data provide detailed biochemical characterization of redox-induced modifications and changes of the BnSnRK2.6-2C activity.

GC-MS-based metabolite profiling of Cosmos caudatus leaves possessing alpha-glucosidase inhibitory activity.

PubMed

Javadi, Neda; Abas, Faridah; Abd Hamid, Azizah; Simoh, Sanimah; Shaari, Khozirah; Ismail, Intan Safinar; Mediani, Ahmed; Khatib, Alfi

2014-06-01

Cosmos caudatus, which is known as "Ulam Raja," is an herbal plant used in Malaysia to enhance vitality. This study focused on the evaluation of the α-glucosidase inhibitory activity of different ethanolic extracts of C. caudatus. Six series of samples extracted with water, 20%, 40%, 60%, 80%, and 100% ethanol (EtOH) were employed. Gas chromatography-mass spectrometry (GC-MS) and orthogonal partial least-squares (OPLS) analysis was used to correlate bioactivity of different extracts to different metabolite profiles of C. caudatus. The obtained OPLS scores indicated a distinct and remarkable separation into 6 clusters, which were indicative of the 6 different ethanol concentrations. GC-MS can be integrated with multivariate data analysis to identify compounds that inhibit α-glucosidase activity. In addition, catechin, α-linolenic acid, α-D-glucopyranoside, and vitamin E compounds were identified and indicate the potential α-glucosidase inhibitory activity of this herb. GC-MS and multivariate data analysis was applied to discriminate Cosmos caudatus samples extracted with water and different ratio of ethanol. Orthogonal partial least-squares (OPLS) model developed was used to determine the major metabolites contributed to α-glucosidase inhibitory activity. This approach also has the ability to predict the bioactivity of a new set of extracts based on a developed validated regression model that is important for quality control of the herb preparation. © 2014 Institute of Food Technologists®

Production of Secondary Metabolites in Extreme Environments: Food- and Airborne Wallemia spp. Produce Toxic Metabolites at Hypersaline Conditions

PubMed Central

Frisvad, Jens C.; Kocev, Dragi; Džeroski, Sašo; Gunde-Cimerman, Nina

2016-01-01

The food- and airborne fungal genus Wallemia comprises seven xerophilic and halophilic species: W. sebi, W. mellicola, W. canadensis, W. tropicalis, W. muriae, W. hederae and W. ichthyophaga. All listed species are adapted to low water activity and can contaminate food preserved with high amounts of salt or sugar. In relation to food safety, the effect of high salt and sugar concentrations on the production of secondary metabolites by this toxigenic fungus was investigated. The secondary metabolite profiles of 30 strains of the listed species were examined using general growth media, known to support the production of secondary metabolites, supplemented with different concentrations of NaCl, glucose and MgCl2. In more than two hundred extracts approximately one hundred different compounds were detected using high-performance liquid chromatography-diode array detection (HPLC-DAD). Although the genome data analysis of W. mellicola (previously W. sebi sensu lato) and W. ichthyophaga revealed a low number of secondary metabolites clusters, a substantial number of secondary metabolites were detected at different conditions. Machine learning analysis of the obtained dataset showed that NaCl has higher influence on the production of secondary metabolites than other tested solutes. Mass spectrometric analysis of selected extracts revealed that NaCl in the medium affects the production of some compounds with substantial biological activities (wallimidione, walleminol, walleminone, UCA 1064-A and UCA 1064-B). In particular an increase in NaCl concentration from 5% to 15% in the growth media increased the production of the toxic metabolites wallimidione, walleminol and walleminone. PMID:28036382

Superhydrophobic hybrid inorganic-organic thiol-ene surfaces fabricated via spray-deposition and photopolymerization.

PubMed

Sparks, Bradley J; Hoff, Ethan F T; Xiong, Li; Goetz, James T; Patton, Derek L

2013-03-13

We report a simple and versatile method for the fabrication of superhydrophobic inorganic-organic thiol-ene coatings via sequential spray-deposition and photopolymerization under ambient conditions. The coatings are obtained by spray-deposition of UV-curable hybrid inorganic-organic thiol-ene resins consisting of pentaerythritol tetra(3-mercaptopropionate) (PETMP), triallyl isocyanurate (TTT), 2,4,6,8-tetramethyl-2,4,6,8-tetravinylcyclotetrasiloxane (TMTVSi), and hydrophobic fumed silica nanoparticles. The spray-deposition process and nanoparticle agglomeration/dispersion provide surfaces with hierarchical morphologies exhibiting both micro- and nanoscale roughness. The wetting behavior, dependent on the concentration of TMTVSi and hydrophobic silica nanoparticles, can be varied over a broad range to ultimately provide coatings with high static water contact angles (>150°), low contact angle hysteresis, and low roll off angles (<5°). The cross-linked thiol-ene coatings are solvent resistant, stable at low and high pH, and maintain superhydrophobic wetting behavior after extended exposure to elevated temperatures. We demonstrate the versatility of the spray-deposition and UV-cure process on a variety of substrate surfaces including glass, paper, stone, and cotton fabric.

Covalent Incorporation of Ionic Liquid into Ion-Conductive Networks via Thiol-Ene Photopolymerization.

PubMed

Tibbits, Andrew C; Yan, Yushan S; Kloxin, Christopher J

2017-07-01

Ene-functionalized ionic liquids with a range of different cationic groups and counteranions react stoichiometrically within a tetrathiol-divinyl ether formulation within 20 minutes to form thiol-ene polymers with measurable ionic conductivities via a photoinitiated polymerization and crosslinking reaction. Dynamic mechanical analysis indicates that these networks are more spatially heterogeneous and possess higher glass transition temperatures (T g ) compared with thiol-ene formulations without charge. While tuning the molar content of ionic liquid monomer is one method for adjusting the crosslink and charge densities of the thiol-ene polymeric ionic liquid networks, the presence of cation-anion interactions also plays a critical role in dictating the thermomechanical and conductive properties. Particularly, while cationic structure effects are not significant on the polymer properties, the use of a weakly coordinating hydrophobic anion (bistriflimide) instead of bromide-based networks results in an apparent decrease in hydrated ion conductivity (7.4 to 1.5 mS cm -1 ) and T g (-9.6 to -17.8 °C). © 2017 WILEY-VCH Verlag GmbH & Co. KGaA, Weinheim.

Effect of Thiols, Zinc, and Redox Conditions on Hg Uptake in Shewanella oneidensis

DOE PAGES

Szczuka, Aleksandra; Morel, Francois M. M.; Schaefer, Jeffra K.

2015-05-18

Mercury uptake in bacteria represents a key first step in the production and accumulation Of methylmercury in biota. Previous experiments with mercury methylating bacteria have shown that Hg uptake is enhanced by some thiols, in particular cysteine, and that it is an energy-dependent process through heavy Metal TA transporters. In this study, we examine Hg uptake in the nonmethylating facultative aerobe, Shewanella oneidensis, under both anaerobic and aerobic conditions. Our results demonstrate similar characteristics of the Hg uptake system to those of the Hg methylating strains: uptake is enhanced in the presence of some thiols but not others; uptake ismore » energy dependent as evidenced by inhibition by a protonophore; and uptake is inhibited by high Zn(II) concentrations. Initial cellular uptake rates in S. oneidensis were remarkably similar under aerobic and fumarate-reducing conditions. In conclusion, these data support a similar Hg(II) uptake mechanism within the proteobacteria of accidental Hg(II) transport through heavy metal transporters with similar rates of uptake but differences in the ability to take up Hg bound to different thiols.« less

The roles of protein disulphide isomerase family A, member 3 (ERp57) and surface thiol/disulphide exchange in human spermatozoa-zona pellucida binding.

PubMed

Wong, Chi-Wai; Lam, Kevin K W; Lee, Cheuk-Lun; Yeung, William S B; Zhao, Wei E; Ho, Pak-Chung; Ou, Jian-Ping; Chiu, Philip C N

2017-04-01

Are multimeric sperm plasma membrane protein complexes, ERp57 and sperm surface thiol content involved in human spermatozoa-zona pellucida (ZP) interaction? ERp57 is a component of a multimeric spermatozoa-ZP receptor complex involved in regulation of human spermatozoa-ZP binding via up-regulation of sperm surface thiol content. A spermatozoon acquires its fertilization capacity within the female reproductive tract by capacitation. Spermatozoa-ZP receptor is suggested to be a composite structure that is assembled into a functional complex during capacitation. Sperm surface thiol content is elevated during capacitation. ERp57 is a protein disulphide isomerase that modulates the thiol-disulphide status of proteins. The binding ability and components of protein complexes in extracted membrane protein fractions of spermatozoa were studied. The roles of capacitation, thiol-disulphide reagent treatments and ERp57 on sperm functions and sperm surface thiol content were assessed. Spermatozoa were obtained from semen samples from normozoospermic men. Human oocytes were obtained from an assisted reproduction programme. Blue native polyacrylamide gel electrophoresis, western ligand blotting and mass spectrometry were used to identify the components of solubilized ZP/ZP3-binding complexes. The localization and expression of sperm surface thiol and ERp57 were studied by immunostaining and sperm surface protein biotinylation followed by western blotting. Sperm functions were assessed by standard assays. Several ZP-binding complexes were isolated from the cell membrane of capacitated spermatozoa. ERp57 was a component of one of these complexes. Capacitation significantly increased the sperm surface thiol content, acrosomal thiol distribution and ERp57 expression on sperm surface. Sperm surface thiol and ERp57 immunoreactivity were localized to the acrosomal region of spermatozoa, a region responsible for ZP-binding. Up-regulation of the surface thiol content or ERp57 surface

Building thiol and metal-thiolate functions into coordination nets: Clues from a simple molecule

NASA Astrophysics Data System (ADS)

He, Jun; Yang, Chen; Xu, Zhengtao; Zeller, Matthias; Hunter, Allen D.; Lin, Jianhua

2009-07-01

The simple and easy-to-prepare bifunctional molecule 2,5-dimercapto-1,4-benzenedicarboxylic acid (H 4DMBD) interacts with the increasingly harder metal ions of Cu +, Pb 2+ and Eu 3+ to form the coordination networks of Cu 6(DMBD) 3(en) 4(Hen) 6 ( 1), Pb 2(DMBD)(en) 2 ( 2) and Eu 2(H 2DMBD) 3(DEF) 4 ( 3), where the carboxyl and thiol groups bind with distinct preference to the hard and soft metal ions, respectively. Notably, 1 features uncoordinated carboxylate groups and Cu 3 cluster units integrated via the thiolate groups into an extended network with significant interaction between the metal centers and the organic molecules; 2 features a 2D coordination net based on the mercapto and carboxylic groups all bonded to the Pb 2+ ions; 3 features free-standing thiol groups inside the channels of a metal-carboxylate-based network. This study illustrates the rich solid state structural features and potential functions offered by the carboxyl-thiol combination.

Decreased plasma thiol antioxidant barrier and selenoproteins as potential biomarkers for ongoing methylmercury intoxication and an individual protective capacity.

PubMed

Usuki, Fusako; Fujimura, Masatake

2016-04-01

Manifestation of methylmercury (MeHg) toxicity depends on individual susceptibility to MeHg, as well as MeHg burden level. Therefore, biomarkers that reflect the protective capacity against MeHg are needed. The critical role of oxidative stress in the pathogenesis of MeHg cytotoxicity has been demonstrated. Because MeHg has high affinity for selenohydryl groups, sulfhydryl groups, and selenides, and causes posttranscriptional defects in selenoenzymes, proteins with selenohydryl and sulfhydryl groups should play a critical role in mediating MeHg-induced oxidative stress. Here, plasma oxidative stress markers and selenoproteins were investigated in MeHg-intoxicated rats showing neuropathological changes after 4 weeks of MeHg exposure. The thiol antioxidant barrier (-SHp) level significantly decreased 2 weeks after MeHg exposure, which is an early stage at which no systemic oxidative stress, histopathological changes, or clinical signs were detected. Diacron reactive oxidant metabolite (d-ROM) levels significantly increased 3 weeks after MeHg exposure, indicating the occurrence of systemic oxidative stress. Rats treated with lead acetate or cadmium chloride showed no changes in levels of -SHp and d-ROM. Selenoprotein P1 abundance significantly decreased in MeHg-treated rats, whereas it significantly increased in rats treated with Pb or Cd. Plasma selenium-dependent glutathione peroxidase (GPx3) activity also significantly decreased after MeHg exposure, whereas plasma non-selenoenzyme glutathione reductase activity significantly increased in MeHg-treated rats. The results suggest that decreased capacity of -SHp and selenoproteins (GPx3 and selenoprotein P) can be useful biomarkers of ongoing MeHg cytotoxicity and the individual protective capacity against the MeHg body burden.

Trichoderma secondary metabolites that affect plant metabolism.

PubMed

Vinale, Francesco; Sivasithamparam, Krishnapillai; Ghisalberti, Emilio L; Ruocco, Michelina; Wood, Sheridan; Lorito, Matteo

2012-11-01

Recently, there have been many exciting new developments relating to the use of Trichoderma spp. as agents for biocontrol of pathogens and as plant growth promoters. Several mechanisms have been proposed to explain the positive effects of these microorganisms on the plant host. One factor that contributes to their beneficial biological activities is related to the wide variety of metabolites that they produce. These metabolites have been found not only to directly inhibit the growth and pathogenic activities of the parasites, but also to increase disease resistance by triggering the system of defence in the plant host. In addition, these metabolites are also capable of enhancing plant growth, which enables the plant to counteract the disease with compensatory vegetative growth by the augmented production of root and shoot systems. This review takes into account the Trichoderma secondary metabolites that affect plant metabolism and that may play an important role in the complex interactions of this biocontrol agent with the plant and pathogens.

Flavone Biotransformation by Aspergillus niger and the Characterization of Two Newly Formed Metabolites

PubMed Central

Assawah, Suzan W.; El-Sharkawy, Saleh H.; Abdel-Salam, Amal

2008-01-01

Aspergillus niger isolated from Allium sativum was used at large scale fermentation (150 mg flavone/200 ml medium) to obtain suitable amounts of the products, efficient for identification. Then spectral analysis (UV, IR, 1H-NMR, 13C-NMR) and mass spectrometry were performed for the two products, which contributed to the identification process. The metabolite (1) was identified as 2'-hydroxydihydrochalcone, and the metabolite (2) was identified as 2'-hydroxyphenylmethylketone, which were more active than flavone itself. Antioxidant activities of the two isolated metabolites were tested compared with ascorbic acid. Antioxidant activity of metabolite (1) was recorded 64.58% which represented 79% of the antioxidant activity of ascorbic acid, and metabolite (2) was recorded 54.16% (67% of ascorbic acid activity). However, the antioxidant activity of flavone was recorded 37.50% which represented 46% of ascorbic acid activity. The transformed products of flavone have antimicrobial activity against Pseudomonas aeruginosa, Aspergillus flavus and Candida albicans, with MIC was recorded 250 µg/ml for metabolite (2) against all three organism and 500, 300, and 300 µg/ml for metabolite (1) against tested microorganisms (P. aeruginosa, Escherichia coli, Bacillus subtilis, and Klebsiella pneumonia, Fusarium moniliforme, A. flavus, Saccharomyces cerviceae, Kluveromyces lactis and C. albicans) at this order. PMID:23990746

MERCURY(II) ADSORPTION FROM WASTEWATERS USING A THIOL FUNCTIONAL ADSORBENT

EPA Science Inventory

The removal of mercury(II) from wastewaters (coal-fired utility plant scrubber solutions) using a thiol functional organoceramic composite (SOL-AD-IV) is investigated. A simulant is employed as a surrogate to demonstrate the removal of mercury from real waste solutions. Equilibri...

Mutagenic activity and metabolites in the urine of workers exposed to trinitrotoluene (TNT).

PubMed Central

Ahlborg, G; Einistö, P; Sorsa, M

1988-01-01

Urine samples taken after work and after a free weekend from 50 workers employed in various activities in a chemical plant manufacturing explosives were analysed. On the basis of hygienic surveys, the subjects were divided into three categories of exposure to trinitrotoluene (TNT). The urine analyses consisted of gas chromatographic identification of TNT and its two metabolites, 4-ADNT and 2-ADNT, and a determination of the mutagenic activity. Two frame shift detector strains of Salmonella typhimurium were used, TA 98 and TA 98 NR, the latter being deficient in endogenous nitroreductase activity. On the basis of previous results on TNT mutagenicity, no exogeneous metabolic system was used to test the urine concentrates. Both tester strains showed that the mean urinary mutagenic activity was higher in the after work samples than in post weekend samples from the same subjects, showing that bacterial nitroreductase activity was not significantly responsible for the mutagenicity, although the response was higher with strain TA 98 than with TA 98 NR. The interindividual variation in urine mutagenicity was high, however, and the difference between the two sampling times was statistically significant (p less than 0.05) only for the high exposed group (workers in trotyl foundry and sieve house). Correlation between urinary mutagenicity and concentration of TNT in urine was poor; correlation was significant only with the urinary concentration of 4-ADNT. The correlation between urinary TNT and both metabolites was good (p less than 0.001). These results suggest that analysis of 4-ADNT in urine would be a sufficient biological measure for controlling exposure to TNT. PMID:3378017

Redox Reactivity of Cerium Oxide Nanoparticles Induces the Formation of Disulfide Bridges in Thiol-Containing Biomolecules.

PubMed

Rollin-Genetet, Françoise; Seidel, Caroline; Artells, Ester; Auffan, Mélanie; Thiéry, Alain; Vidaud, Claude

2015-12-21

The redox state of disulfide bonds is implicated in many redox control systems, such as the cysteine-cystine couple. Among proteins, ubiquitous cysteine-rich metallothioneins possess thiolate metal binding groups susceptible to metal exchange in detoxification processes. CeO2 NPs are commonly used in various industrial applications due to their redox properties. These redox properties that enable dual oxidation states (Ce(IV)/Ce(III)) to exist at their surface may act as oxidants for biomolecules. The interaction among metallothioneins, cysteine, and CeO2 NPs was investigated through various biophysical approaches to shed light on the potential effects of the Ce(4+)/Ce(3+) redox system on the thiol groups of these biomolecules. The possible reaction mechanisms include the formation of a disulfide bridge/Ce(III) complex resulting from the interaction between Ce(IV) and the thiol groups, leading to metal unloading from the MTs, depending on their metal content and cluster type. The formation of stable Ce(3+) disulfide complexes has been demonstrated via their fluorescence properties. This work provides the first evidence of thiol concentration-dependent catalytic oxidation mechanisms between pristine CeO2 NPs and thiol-containing biomolecules.

Kynurenine pathway metabolites are associated with hippocampal activity during autobiographical memory recall in patients with depression.

PubMed

Young, Kymberly D; Drevets, Wayne C; Dantzer, Robert; Teague, T Kent; Bodurka, Jerzy; Savitz, Jonathan

2016-08-01

Inflammation-related changes in the concentrations of inflammatory mediators such as c-reactive protein (CRP), interleukin 1β (IL-1), and IL-6 as well as kynurenine metabolites are associated with major depressive disorder (MDD) and affect depressive behavior, cognition, and hippocampal plasticity in animal models. We previously reported that the ratios of kynurenic acid (KynA) to the neurotoxic metabolites, 3-hydroxykynurenine (3HK) and quinolinic acid (QA), were positively correlated with hippocampal volume in depression. The hippocampus is critical for autobiographical memory (AM) recall which is impaired in MDD. Here we tested whether the ratios, KynA/3HK and KynA/QA were associated with AM recall performance as well as hippocampal activity during AM recall. Thirty-five unmedicated depressed participants and 25 healthy controls (HCs) underwent fMRI scanning while recalling emotionally-valenced AMs and provided serum samples for the quantification of kynurenine metabolites, CRP, and cytokines (IL-1 receptor antagonist - IL-1RA; IL-6, tumor necrosis factor alpha - TNF, interferon gamma -IFN-γ, IL-10). KynA/3HK and KynA/QA were lower in the MDD group relative to the HCs. The concentrations of the CRP and the cytokines did not differ significantly between the HCs and the MDD group. Depressed individuals recalled fewer specific AMs and displayed increased left hippocampal activity during the recall of positive and negative memories. KynA/3HK was inversely associated with left hippocampal activity during specific AM recall in the MDD group. Further, KynA/QA was positively correlated with percent negative specific memories recalled in the MDD group and showed a non-significant trend toward a positive correlation with percent positive specific memories recalled in HCs. In contrast, neither CRP nor the cytokines were significantly associated with AM recall or activity of the hippocampus during AM recall. Conceivably, an imbalance in levels of KynA versus QA

Baicalin and its metabolites suppresses gluconeogenesis through activation of AMPK or AKT in insulin resistant HepG-2 cells.

PubMed

Wang, Tao; Jiang, Hongmei; Cao, Shijie; Chen, Qian; Cui, Mingyuan; Wang, Zhijie; Li, Dandan; Zhou, Jing; Wang, Tao; Qiu, Feng; Kang, Ning

2017-12-01

Scutellaria baicalensis Georgi (S. baicalensis), as a traditional Chinese herbal medicine, is an important component of several famous Chinese medicinal formulas for treating patients with diabetes mellitus. Baicalin (BG), a main bioactive component of S. baicalensis, has been reported to have antidiabetic effects. However, pharmacokinetic studies have indicated that BG has poor oral bioavailability. Therefore, it is hard to explain the pharmacological effects of BG in vivo. Interestingly, several reports show that BG is extensively metabolized in rats and humans. Therefore, we speculate that the BG metabolites might be responsible for the pharmacological effects. In this study, BG and its three metabolites (M1-M3) were examined their effects on glucose consumption in insulin resistant HepG-2 cells with a commercial glucose assay kit. Real-time PCR and western blot assay were used to confirm genes and proteins of interest, respectively. The results demonstrate that BG and its metabolites (except for M3) enhanced the glucose consumption which might be associated with inhibiting the expression of the key gluconeogenic genes, including glucose-6-phosphatase (G6Pase), phosphoenolypyruvate carboxykinase (PEPCK) and glucose transporter 2 (GLUT2). Further study found that BG and M1 could suppress hepatic gluconeogenesis via activation of the AMPK pathway, while M2 could suppress hepatic gluconeogenesis via activation of the PI3K/AKT signaling pathway. Taken together, our findings suggest that both BG and its metabolites have antihyperglycemic activities, and might be the active forms of oral doses of BG in vivo. Copyright © 2017 Elsevier Masson SAS. All rights reserved.

EFFECTS OF METHOPRENE, ITS METABOLITES, AND BREAKDOWN PRODUCTS ON RETINOID-ACTIVATED PATHWAYS IN TRANSFECTED CELL LINES

EPA Science Inventory

Methoprene is a terpene-based insecticide designed to act as an agonist of insect juvenile hormone, which is essential for the transition from larval to adult forms in some metamorphic insects. Recent evidence suggests that a methoprene metabolite, methoprene acid, activates a ve...

Simultaneous determination of albumin and low-molecular-mass thiols in plasma by HPLC with UV detection.

PubMed

Borowczyk, Kamila; Wyszczelska-Rokiel, Monika; Kubalczyk, Paweł; Głowacki, Rafał

2015-02-15

In this paper, we describe a simple and robust HPLC based method for determination of total low- and high-molecular-mass thiols, protein S-linked thiols and reduced albumin in plasma. The method is based on derivatization of analytes with 2-chloro-1-methylquinolinium tetrafluoroborate, separation and quantification by reversed-phase liquid chromatography followed by UV detection. Disulfides were converted to their thiol counterparts by reductive cleavage with tris(2-carboxyethyl)phosphine. Linearity in detector response for total thiols was observed over the range of 1-40 μmol L(-1) for Hcy and glutathione (GSH), 5-100 μmol L(-1) for Cys-Gly, 20-300 μmol L(-1) for Cys and 3.1-37.5 μmol L(-1) (0.2-2.4gL(-1)) for human serum albumin (HSA). For the protein S-bound forms these values were as follows: 0.5-30 μmol L(-1) for Hcy and GSH, 2.5-60 μmol L(-1) for Cys-Gly and 5-200 μmol L(-1) for Cys. The LOQs for total HSA, Cys, Hcy, Cys-Gly and GSH were 0.5, 0.2, 0.4, 0.3 and 0.4 μmol L(-1), respectively. The estimated validation parameters for all analytes are more than sufficient to allow the analytical method to be used for monitoring of the total and protein bound thiols as well as redox status of HSA in plasma. Copyright © 2015 Elsevier B.V. All rights reserved.

Reaction Mechanisms of Metals with Hydrogen Sulfide and Thiols in Model Wine. Part 2: Iron- and Copper-Catalyzed Oxidation.

PubMed

Kreitman, Gal Y; Danilewicz, John C; Jeffery, David W; Elias, Ryan J

2016-05-25

Sulfidic off-odors arising during wine production are frequently removed by Cu(II) fining. In part 1 of this study ( 10.1021/acs.jafc.6b00641 ), the reaction of H2S and thiols with Cu(II) was examined; however, the interaction of iron and copper is also known to play an important synergistic role in mediating non-enzymatic wine oxidation. The interaction of these two metals in the oxidation of H2S and thiols (cysteine, 3-sulfanylhexan-1-ol, and 6-sulfanylhexan-1-ol) was therefore examined under wine-like conditions. H2S and thiols (300 μM) were reacted with Fe(III) (100 or 200 μM) alone and in combination with Cu(II) (25 or 50 μM), and concentrations of H2S and thiols, oxygen, and acetaldehyde were monitored over time. H2S and thiols were shown to be slowly oxidized in the presence of Fe(III) alone and were not bound to Fe(III) under model wine conditions. However, Cu(II) added to model wine containing Fe(III) was quickly reduced by H2S and thiols to form Cu(I) complexes, which then rapidly reduced Fe(III) to Fe(II). Oxidation of Fe(II) in the presence of oxygen regenerated Fe(III) and completed the iron redox cycle. In addition, sulfur-derived oxidation products were observed, and the formation of organic polysulfanes was demonstrated.

Depsides: Lichen Metabolites Active against Hepatitis C Virus

PubMed Central

Vu, Thi Huyen; Le Lamer, Anne-Cécile; Lalli, Claudia; Boustie, Joël; Samson, Michel

2015-01-01

A thorough phytochemical study of Stereocaulon evolutum was conducted, for the isolation of structurally related atranorin derivatives. Indeed, pilot experiments suggested that atranorin (1), the main metabolite of this lichen, would interfere with the lifecycle of hepatitis C virus (HCV). Eight compounds, including one reported for the first time (2), were isolated and characterized. Two analogs (5, 6) were also synthesized, to enlarge the panel of atranorin-related structures. Most of these compounds were active against HCV, with a half-maximal inhibitory concentration of about 10 to 70 µM, with depsides more potent than monoaromatic phenols. The most effective inhibitors (1, 5 and 6) were then added at different steps of the HCV lifecycle. Interestingly, atranorin (1), bearing an aldehyde function at C-3, inhibited only viral entry, whereas the synthetic compounds 5 and 6, bearing a hydroxymethyl and a methyl function, respectively, at C-3 interfered with viral replication. PMID:25793970

Oxidative stress and pathology in muscular dystrophies: focus on protein thiol oxidation and dysferlinopathies.

PubMed

Terrill, Jessica R; Radley-Crabb, Hannah G; Iwasaki, Tomohito; Lemckert, Frances A; Arthur, Peter G; Grounds, Miranda D

2013-09-01

The muscular dystrophies comprise more than 30 clinical disorders that are characterized by progressive skeletal muscle wasting and degeneration. Although the genetic basis for many of these disorders has been identified, the exact mechanism for pathogenesis generally remains unknown. It is considered that disturbed levels of reactive oxygen species (ROS) contribute to the pathology of many muscular dystrophies. Reactive oxygen species and oxidative stress may cause cellular damage by directly and irreversibly damaging macromolecules such as proteins, membrane lipids and DNA; another major cellular consequence of reactive oxygen species is the reversible modification of protein thiol side chains that may affect many aspects of molecular function. Irreversible oxidative damage of protein and lipids has been widely studied in Duchenne muscular dystrophy, and we have recently identified increased protein thiol oxidation in dystrophic muscles of the mdx mouse model for Duchenne muscular dystrophy. This review evaluates the role of elevated oxidative stress in Duchenne muscular dystrophy and other forms of muscular dystrophies, and presents new data that show significantly increased protein thiol oxidation and high levels of lipofuscin (a measure of cumulative oxidative damage) in dysferlin-deficient muscles of A/J mice at various ages. The significance of this elevated oxidative stress and high levels of reversible thiol oxidation, but minimal myofibre necrosis, is discussed in the context of the disease mechanism for dysferlinopathies, and compared with the situation for dystrophin-deficient mdx mice. © 2013 The Authors Journal compilation © 2013 FEBS.

Enzymatic improvement of mitochondrial thiol oxidase Erv1 for oxidized glutathione fermentation by Saccharomyces cerevisiae.

PubMed

Kobayashi, Jyumpei; Sasaki, Daisuke; Hara, Kiyotaka Y; Hasunuma, Tomohisa; Kondo, Akihiko

2017-03-15

Oxidized glutathione (GSSG) is the preferred form for industrial mass production of glutathione due to its high stability compared with reduced glutathione (GSH). In our previous study, over-expression of the mitochondrial thiol oxidase ERV1 gene was the most effective for high GSSG production in Saccharomyces cerevisiae cells among three types of different thiol oxidase genes. We improved Erv1 enzyme activity for oxidation of GSH and revealed that S32 and N34 residues are critical for the oxidation. Five engineered Erv1 variant proteins containing S32 and/or N34 replacements exhibited 1.7- to 2.4-fold higher in vitro GSH oxidation activity than that of parental Erv1, whereas the oxidation activities of these variants for γ-glutamylcysteine were comparable. According to three-dimensional structures of Erv1 and protein stability assays, S32 and N34 residues interact with nearby residues through hydrogen bonding and greatly contribute to protein stability. These results suggest that increased flexibility by amino acid replacements around the active center decrease inhibitory effects on GSH oxidation. Over-expressions of mutant genes coding these Erv1 variants also increased GSSG and consequently total glutathione production in S. cerevisiae cells. Over-expression of the ERV1 S32A gene was the most effective for GSSG production in S. cerevisiae cells among the parent and other mutant genes, and it increased GSSG production about 1.5-fold compared to that of the parental ERV1 gene. This is the first study demonstrating the pivotal effects of S32 and N34 residues to high GSH oxidation activity of Erv1. Furthermore, in vivo validity of Erv1 variants containing these S32 and N34 replacements were also demonstrated. This study indicates potentials of Erv1 for high GSSG production.

The Significance of Lichens and Their Metabolites

NASA Astrophysics Data System (ADS)

Huneck, S.

Lichens, symbiontic organisms of fungi and algae, synthesize numerous metabolites, the "lichen substances," which comprise aliphatic, cycloaliphatic, aromatic, and terpenic compounds. Lichens and their metabolites have a manifold biological activity: antiviral, antibiotic, antitumor, allergenic, plant growth inhibitory, antiherbivore, and enzyme inhibitory. Usnic acid, a very active lichen substance is used in pharmaceutical preparations. Large amounts of Pseudevernia furfuracea and Evernia prunastri are processed in the perfume industry, and some lichens are sensitive reagents for the evaluation of air pollution.

Insights into the mechanisms of Promysalin, a secondary metabolite with genus-specific antibacterial activity against Pseudomonas

USDA-ARS?s Scientific Manuscript database

Promysalin, a secondary metabolite produced by Pseudomonas putida RW10S1, has antibacterial activity against a wide variety of Pseudomonas sp., including both human and plant pathogens. Promysalin induces swarming and biofilm formation in the producing species, and inhibits growth of susceptible sp...

Comparison of inhibition of murine leukaemia cell growth by 9-isothiocyanatoacridine and its cytosine adduct: involvement of thiols.

PubMed

Bajdichova, M; Paulikova, H; Jakubikova, J; Sabolova, D

2007-01-01

Cytotoxicity of two fluorescent acridine derivatives - 9-isothiocyanatoacridine (AcITC) and N-(9-acridinylthiocarbamoyl) cytosine (AcTCC) - a novel acridine compound, were investigated. Both substances have cytotoxic activity against the L1210 cellular line, IC50 values were in the micromolar range. Despite the high reactivity of AcITC towards thiols, its effects on leukemia cells were similar to naturally occurring isothiocyanates. AcITC changed the intracellular level of glutathione (GSH), and induced apoptosis. Arrest of cell cycle (G2/M-phase) was also observed. AcITC primarily reacted with -SH groups on cellular surface, and the study of the interaction of the isotiocyanate with human erythrocyte ghosts confirmed that the plasma membrane was the first place where AcITC bound. AcTCC does not react with cellular thiols; images obtained with fluorescent microscopy confirmed interaction of AcTCC with chromatine. Although AcTCC induced cellular arrest in the G2/M phase, apoptosis was not confirmed.

Facile construction of macroporous hybrid monoliths via thiol-methacrylate Michael addition click reaction for capillary liquid chromatography.

PubMed

Lin, Hui; Ou, Junjie; Liu, Zhongshan; Wang, Hongwei; Dong, Jing; Zou, Hanfa

2015-01-30

A facile approach based on thiol-methacrylate Michael addition click reaction was developed for construction of porous hybrid monolithic materials. Three hybrid monoliths were prepared via thiol-methacrylate click polymerization by using methacrylate-polyhedral oligomeric silsesquioxane (POSS) (cage mixture, n=8, 10, 12, POSS-MA) and three multi-thiol crosslinkers, 1,6-hexanedithiol (HDT), trimethylolpropane tris(3-mercaptopropionate) (TPTM) and pentaerythritol tetrakis(3-mercaptopropionate) (PTM), respectively, in the presence of porogenic solvents (n-propanol and PEG 200) and a catalyst (dimethylphenylphosphine, DMPP). The obtained monoliths possessed high thermal and chemical stabilities. Besides, they all exhibited high column efficiencies and excellent separation abilities in capillary liquid chromatography (cLC). The highest column efficiency could reach ca. 195,000N/m for butylbenzene on the monolith prepared with POSS-MA and TPTM (monolith POSS-TPTM) in reversed-phase (RP) mode at 0.64mm/s. Good chromatographic performance were all achieved in the separations of polycyclic aromatic hydrocarbons (PAHs), phenols, anilines, EPA 610 as well as bovine serum albumin (BSA) digest. The high column efficiencies in the range of 51,400-117,000N/m (achieved on the monolith POSS-PTM in RP mode) convincingly demonstrated the high separation abilities of these thiol-methacrylate based hybrid monoliths. All the results demonstrated the feasibility of the phosphines catalyzed thiol-methacrylate Michael addition click reaction in fabrication of monolithic columns with high efficiency for cLC applications. Copyright © 2014 Elsevier B.V. All rights reserved.

Thiol dioxygenase turnover yields benzothiazole products from 2-mercaptoaniline and O2-dependent oxidation of primary alcohols.

PubMed

Morrow, William P; Sardar, Sinjinee; Thapa, Pawan; Hossain, Mohammad S; Foss, Frank W; Pierce, Brad S

2017-10-01

Thiol dioxygenases are non-heme mononuclear iron enzymes that catalyze the O 2 -dependent oxidation of free thiols (-SH) to produce the corresponding sulfinic acid (-SO 2 - ). Previous chemical rescue studies identified a putative Fe III -O 2 - intermediate that precedes substrate oxidation in Mus musculus cysteine dioxygenase (Mm CDO). Given that a similar reactive intermediate has been identified in the extradiol dioxygenase 2, 3-HCPD, it is conceivable that these enzymes share other mechanistic features with regard to substrate oxidation. To explore this possibility, enzymatic reactions with Mm CDO (as well as the bacterial 3-mercaptopropionic acid dioxygenase, Av MDO) were performed using a substrate analogue (2-mercaptoaniline, 2ma). This aromatic thiol closely approximates the catecholic substrate of homoprotocatechuate of 2, 3-HPCD while maintaining the 2-carbon thiol-amine separation preferred by Mm CDO. Remarkably, both enzymes exhibit 2ma-gated O 2 -consumption; however, none of the expected products for thiol dioxygenase or intra/extradiol dioxygenase reactions were observed. Instead, benzothiazoles are produced by the condensation of 2ma with aldehydes formed by an off-pathway oxidation of primary alcohols added to aqueous reactions to solubilize the substrate. The observed oxidation of 1º-alcohols in 2ma-reactions is consistent with the formation of a high-valent intermediate similar to what has been reported for cytochrome P450 and mononuclear iron model complexes. Copyright © 2017 Elsevier Inc. All rights reserved.

New Pioglitazone Metabolites and Absence of Opened-Ring Metabolites in New N-Substituted Thiazolidinedione.

PubMed

Campos, Michel Leandro; Cerqueira, Letícia Bonancio; Silva, Bruna Cristina Ulian; Franchin, Taísa Busaranho; Galdino-Pitta, Marina Rocha; Pitta, Ivan Rocha; Peccinini, Rosângela Gonçalves; Pontarolo, Roberto

2018-06-01

Thiazolidinediones (TZDs) are drugs used to treat type 2 diabetes mellitus; however, several safety concerns remain regarding the available drugs in this class. Therefore, the search for new TZD candidates is ongoing; metabolism studies play a crucial step in the development of new candidates. Pioglitazone, one of the most commonly used TZDs, and GQ-11, a new N -substituted TZD, were investigated in terms of their metabolic activity in rat and human liver microsomes to assess their metabolic stability and investigate their metabolites. Methods for preparation of samples were based on liquid-liquid extraction and protein precipitation. Quantitation was performed using liquid chromatography (LC)-tandem mass spectrometry, and the metabolite investigation was performed using ultraperformance LC coupled to a hybrid quadrupole-time of flight mass spectrometer. The predicted intrinsic clearance of GQ-11 was 70.3 and 46.1 ml/kg per minute for rats and humans, respectively. The predicted intrinsic clearance of pioglitazone was 24.1 and 15.9 ml/kg per minute for rats and humans, respectively. The pioglitazone metabolite investigation revealed two unpublished metabolites (M-D and M-A). M-A is a hydration product and may be related to the mechanism of ring opening and the toxicity of pioglitazone. The metabolites of GQ-11 are products of oxidation; no ring-opening metabolite was observed for GQ-11. In conclusion, under the same experimental conditions, a ring-opening metabolite was observed only for pioglitazone. The resistance of GQ-11 to the ring opening is probably related to N -substitution in the TZD ring. Copyright © 2018 by The American Society for Pharmacology and Experimental Therapeutics.

Facile fabrication of redox-responsive thiol-containing drug delivery system via RAFT polymerization.

PubMed

Zhuang, Yuanyuan; Su, Yue; Peng, Yu; Wang, Dali; Deng, Hongping; Xi, Xiaodong; Zhu, Xinyuan; Lu, Yunfeng

2014-04-14

A novel kind of redox-responsive polymeric drug delivery system has been designed and prepared successfully through the coupling of the multithiol branched polymers and thiol-containing drugs. The branched poly((S-(4-vinyl) benzyl S'-propyltrithiocarbonate)-co-(poly(ethylene glycol) methacrylate)) (poly(VBPT-co-PEGMA)) was synthesized by one-pot reaction via reversible addition-fragmentation chain transfer (RAFT) copolymerization. Subsequently, the hydrophobic thiol-containing anticancer drug 6-mercaptopurine (MP) was conjugated to poly(VBPT-co-PEGMA) by thiol-disulfide exchange reaction, resulting in the formation of poly(VBPT-co-PEGMA)-S-S-MP conjugate. Due to its amphiphilicity, poly(VBPT-co-PEGMA)-S-S-MP conjugate self-assembled into amphiphilic micelles in aqueous solution. Under a reductive environment, the disassembly of polymeric micelles resulted in the MP release. Flow cytometry and confocal laser scanning microscopy (CLSM) measurements demonstrated that the poly(VBPT-co-PEGMA)-S-S-MP micelles could be taken up by Raji cells (a Burkitt lymphoma cell line). The viability of the Raji cells incubated with the glutathione (GSH) mediated poly(VBPT-co-PEGMA)-S-S-MP micelles was investigated by Cell Counting Kit-8 (CCK-8) assay. The experimental results showed that the viability of the glutathione monoester (GSH-OEt) pretreated cells was lower than that without pretreatment, while the viability of the buthionine sulfoximine (BSO) pretreated cells was higher than that without pretreatment. The poly(VBPT-co-PEGMA)-S-S-MP micelles could induce the apoptosis of Raji cells, and the apoptosis behavior was dose-dependent. This redox-responsive polymer-drug conjugate provides a promising platform for the delivery of thiol-containing biological molecules.

Surface plasmon resonance based spectrophotometric determination of medicinally important thiol compounds using unmodified silver nanoparticles

NASA Astrophysics Data System (ADS)

Vaishnav, Sandeep K.; Patel, Kuleshwar; Chandraker, Kumudini; Korram, Jyoti; Nagwanshi, Rekha; Ghosh, Kallol K.; Satnami, Manmohan L.

2017-05-01

The determination of thiol based biological molecules and drugs, such as cysteine (Cys) (I), α-lipoic acid (II), and sodium 2-sulfanylethane sulphonate (Mesna (III)) in human plasma are becoming progressively more important due to the growing body of knowledge about their essential role in numerous biological pathways. Herein we demonstrate a sensitive colorimetric sensor for the determination of medicinally important thiol drugs based on aggregation of the citrate capped silver nanoparticles (Ag NPs). This approach exploited the high affinity of thiols towards the Ag NPs surface which could tempt replacement of the citrate shell by the thiolate shell of target molecules, resulting in aggregation of the NPs through intermolecular electrostatic interaction or hydrogen-bonding. Because of aggregation, the plasmon band at around 400 nm decreases gradually, along with the appearance of a new band connoting a red shift. The calibration curves are derived from the intensity ratios of A530/A400, which display a linear relation in the range of 1 μM-150 μM, 5 μM-200 μM and 10 μM-130 μM, respectively. The obtained detection limits (3σ) were found to be 1.5 μM, 5.6 μM and 10.2 μM for compound I-III, respectively. The proposed method has been successfully applied for the detection of thiol compounds in real samples.

Thiol-catalyzed formation of lactate and glycerate from glyceraldehyde. [significance in molecular evolution

NASA Technical Reports Server (NTRS)

Weber, A. L.

1983-01-01

The rate of lactate formation from glyceraldehyde, catalyzed by N-acetyl-cysteine at ambient temperature in aqueous sodium phosphate (pH 7.0), is more rapid at higher sodium phosphate concentrations and remains essentially the same in the presence and absence of oxygen. The dramatic increase in the rate of glycerate formation that is brought about by this thiol, N-acetylcysteine, is accompanied by commensurate decreases in the rates of glycolate and formate production. It is suggested that the thiol-dependent formation of lactate and glycerate occurs by way of their respective thioesters. Attention is given to the significance of these reactions in the context of molecular evolution.

Subcellular Distribution of O-Acetylserine(thiol)lyase in Cauliflower (Brassica oleracea L.) Inflorescence.

PubMed

Rolland, N; Droux, M; Douce, R

1992-03-01

The subcellular localization of O-acetyiserine(thiol)lyase (EC 4.2.99.8) in nongreen tissue from higher plants has been studied using purified proplastids, mitochondria, and protoplasts from cauliflower (Brassica oleracea L.) buds as a source of subcellular fractions. O-Acetylserine(thiol)lyase has been detected in both organelles (proplastids and mitochondria) and a cytosolic extract obtained by protoplast fractionation. We confirmed these observations, demonstrating that a form of the enzyme different in global charge and separated from others by anion-exchange chromatography corresponded to each subcellular location. Our observations are consistent with the need for cysteine biosynthesis in each subcellular compartment where the synthesis of proteins occurs.

Odorant Screening and Quantitation of Thiols in Carmenere Red Wine by Gas Chromatography-Olfactometry and Stable Isotope Dilution Assays.

PubMed

Pavez, Carolina; Agosin, Eduardo; Steinhaus, Martin

2016-05-04

The sensory impact of thiols in Vitis vinifera 'Carmenere' red wines was evaluated. For this purpose, aroma extract dilution analysis was applied to the thiols isolated from a Carmenere red wine by affinity chromatography with a mercurated agarose gel. Results revealed the presence of four odorants, identified as 2-furanylmethanethiol, 3-sulfanylhexyl acetate, 3-sulfanyl-1-hexanol, and 2-methyl-3-sulfanyl-1-butanol, with the latter being described here for the first time in Carmenere red wines. Quantitation of the four thiols in the Carmenere wine screened by aroma extract dilution analysis and in three additional Carmenere wines by stable isotope dilution assays resulted in concentrations above the respective orthonasal odor detection threshold values. Triangle tests applied to wine model solutions with and without the addition of the four thiols showed significant differences, thus suggesting that the compounds do have the potential to influence the overall aroma of red wine.

Liquid—liquid interface-mediated Au—ZnO composite membrane using ‘thiol-ene’ click chemistry

NASA Astrophysics Data System (ADS)

Ali, Mohammed; Ghosh, Sujit Kumar

2015-07-01

A nanoparticle-decorated composite membrane has been devised at the water/CCl4 interface based on the self-assembly of ligand-stabilized gold and zinc oxide nanoparticles, exploiting the ‘thiol-ene’ click chemistry between the thiol groups of 11-mercaptoundecanoic acid-stabilized ZnO nanoparticles and the ene functionality of cinnamic acid attached to gold nanoparticles. The interfacial assembly of ultrasmall particles leads to a multilayer film that exhibits charge-dependent permeability of amino acid molecules across the membrane.

Nonenzymatic formation of 'energy-rich' lactoyl and glyceroyl thioesters from glyceraldehyde and a thiol

NASA Technical Reports Server (NTRS)

Weber, A. L.

1984-01-01

The energy rich thioester, N-acetyl-S-lactoylcysteine, is formed under anaerobic conditions from glyceraldehyde and N-acetylcysteine at ambient temperature in aqueous solutions of sodium phosphate (pH 7.0). The conversion occurs at a rate of about 0.4 percent per day in reactions with 10 millimoles (mM) glyceraldehyde, 40 mM thiol, and 500 mM sodium phosphate (pH 7.0). Thioester formation proceeds at an estimated efficiency of 76 percent. The formation of lactoyl thioester most likely occurs by the phosphate catalyzed dehydration of glyceraldehyde to give pyruvaldehyde, which combines with thiol to form a hemithioacetal that rearranges to the thioester. A second energy rich thioester, N-acetyl-S-glyceroylcysteine, is also produced from glyceraldehyde when these reactions are carried out in the presence of oxygen and to a limited extent in the absence of oxygen. In the presence of oxygen, the formation of glyceroyl thioester continues until the thiol disappears completely by oxidation. The significance of these reactions to the energetics of the origin of life is discussed.

Nonenzymatic formation of energy-rich lactoyl and glyceroyl thioesters from glyceraldehyde and a thiol

NASA Technical Reports Server (NTRS)

Weber, A. L.

1983-01-01

The energy rich thioester, N-acetyl-S-lactoylcysteine, is formed under anaerobic conditions from glyceraldehyde and N-acetylcysteine at ambient temperature in aqueous solutions of sodium phosphate (pH 7.0). The conversion occurs at a rate of about 0.4% per day in reactions with 10 millimoles (mM) glyceraldehyde, 10 mM thiol, and 500 mM sodium phosphate (pH 7.0). Thioester formation proceeds at an estimated efficiency of 76%. The formation of lactoyl thioester most likely occurs by the phosphate catalyzed dehydration of glyceraldehyde to give pyruvaldehyde, which combines with thiol to form a hemithioacetal that rearranges to the thioester. A second energy rich thioester, N-acetyl-S-glyceroylcysteine, is also produced from glyceraldehyde when these reactions are carried out in the presence of oxygen and to a limited extent in the absence of oxygen. In the presence of oxygen the formation of glyceroyl thioester continues until the thiol disappears completely by oxidation. The significance of these reactions to the energetics of the origin of life is discussed.

Thiol surface complexation on growing CdS clusters

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

Swayambunathan, V.; Hayes, D.; Schmidt, K.H.

1990-05-09

The growth of small CdS colloidal particles has been initiated by pulse radiolytic release of sulfide from thiol (3-mercapto-1,2-propanediol, RSH) in the presence of Cd{sup 2+} ions. The kinetics and stoichiometry of the ensuring reactions were followed by conductivity, absorption spectroscopy, and light-scattering techniques. The final CdS product has been identified by electron diffraction. The formation of Cd-thiolate complexes at the surface of the particles is indicated by conductivity and by energy dispersive analysis of X-ray (EDAX) results. The rate of formation of CdS clusters is strongly pH dependent due to the pH effect on the stability of Dd{sup 2+}/HS{supmore » {minus}} complexes. At low pHs (4.0-5.3) the growth mechanism is proposed to be primarily a cluster-molecule process. At this pH range Cd{sup 2+} ions at the CdS particle surface complex with thiolate ions stronger than in the bulk of the solution. The size control of the particles by thiols is proposed to result from a competition of thiolate ions with HS{sup {minus}} ions for cadmium ions at the surface of the growing particles.« less

Formation of difluorothionoacetyl-protein adducts by S-(1,1,2,2-tetrafluoroethyl)-L-cysteine metabolites: Nucleophilic catalysis of stable lysyl adduct formation by histidine and tyrosine

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

Hayden, P.J.; McCann, D.J.; Stevens, J.L.

1991-06-18

{sup 19}F NMR spectorscopy was used in conjunction with isotopic labeling to demonstrate that difluorothionoacetyl-protein adducts are formed by metabolites of the nephrotoxic cysteine conjugate S(1,1,2,2-tetrafluoroethyl)-L-cysteine (TFEC). To determine which amino acid residues can be involved in adduct formation, the reactivity of TFEC metabolites with a variety of N-acetyl amino acids was also investigated. An N{sup {alpha}}-acetyl-N{sup {epsilon}}-(difluorothionoacetyl)lysine (DFTAL) adduct was isolated and characterized by {sup 19}F and {sup 13}C NMR spectroscopy and mass spectrometry. N{sup {alpha}}-Acetylhistidine and N-acetyltyrosine were found to act as nucleophilic catalysts to facilitate the formation of both the protein and DFTAL adducts. Adduct formation wasmore » greatly reduced when lysyl-modified protein was used as the substrate, indicating that lysyl residues are primary sites of adduct formation. However, N{sup a}-acetyllysine, at concentrations of >100-fold in excess compared to protein lysyl residues, was not effective in preventing binding of metabolites to protein. Therefore, nucleophilic catalysis at the surface of the protein may be an important mechanism for the binding of TFEC metabolites to specific lysyl residues in protein. TFEC metabolites were very reactive with the thiol nucleophiles glutathione and N-acetylcysteine. However, the predicted difluorodithioesters could not be isolated. Bothe stable difluorothioacetamide and less stable difluorodithioester protein adducts may play a role in TFEC-mediated enphrotoxicity.« less

Prediction of Losartan-Active Carboxylic Acid Metabolite Exposure Following Losartan Administration Using Static and Physiologically Based Pharmacokinetic Models.

PubMed

Nguyen, Hoa Q; Lin, Jian; Kimoto, Emi; Callegari, Ernesto; Tse, Susanna; Obach, R Scott

2017-09-01

The aim of this study was to evaluate a strategy based on static and dynamic physiologically based pharmacokinetic (PBPK) modeling for the prediction of metabolite and parent drug area under the time-concentration curve ratio (AUC m /AUC p ) and their PK profiles in humans using in vitro data when active transport processes are involved in disposition. The strategy was applied to losartan and its pharmacologically active metabolite carboxylosartan as test compounds. Hepatobiliary transport including transport-mediated uptake, canilicular and basolateral efflux, and metabolic clearance estimates were obtained from in vitro studies using human liver microsomes and sandwich-cultured hepatocytes. Human renal clearance of carboxylosartan was estimated from dog renal clearance using allometric scaling approach. All clearance mechanisms were mechanistically incorporated in a static model to predict the relative exposure of carboxylosartan versus losartan (AUC m /AUC p ). The predicted AUC m /AUC p were consistent with the observed data following intravenous and oral administration of losartan. Moreover, the in vitro parameters were used as initial parameters in PBPK permeability-limited disposition models to predict the concentration-time profiles for both parent and its active metabolite after oral administration of losartan. The PBPK model was able to recover the plasma profiles of both losartan and carboxylosartan, further substantiating the validity of this approach. Copyright © 2017 American Pharmacists Association®. Published by Elsevier Inc. All rights reserved.

Influence of Different Drying Treatments and Extraction Solvents on the Metabolite Profile and Nitric Oxide Inhibitory Activity of Ajwa Dates.

PubMed

Abdul-Hamid, Nur Ashikin; Abas, Faridah; Ismail, Intan Safinar; Shaari, Khozirah; Lajis, Nordin H

2015-11-01

This study aimed to examine the variation in the metabolite profiles and nitric oxide (NO) inhibitory activity of Ajwa dates that were subjected to 2 drying treatments and different extraction solvents. (1)H NMR coupled with multivariate data analysis was employed. A Griess assay was used to determine the inhibition of the production of NO in RAW 264.7 cells treated with LPS and interferon-γ. The oven dried (OD) samples demonstrated the absence of asparagine and ascorbic acid as compared to the freeze dried (FD) dates. The principal component analysis showed distinct clusters between the OD and FD dates by the second principal component. In respect of extraction solvents, chloroform extracts can be distinguished by the absence of arginine, glycine and asparagine compared to the methanol and 50% methanol extracts. The chloroform extracts can be clearly distinguished from the methanol and 50% methanol extracts by first principal component. Meanwhile, the loading score plot of partial least squares analysis suggested that beta glucose, alpha glucose, choline, ascorbic acid and glycine were among the metabolites that were contributing to higher biological activity displayed by FD and methanol extracts of Ajwa. The results highlight an alternative method of metabolomics approach for determination of the metabolites that contribute to NO inhibitory activity. The association between metabolite profiles and nitric oxide (NO) inhibitory activity of the various extracts of Ajwa dates was evaluated by utilizing partial least squares (PLS) model. The validated PLS model can be employed to predict the NO inhibitory activity of new samples of date fruits based on their NMR spectra which was important for assessing fruit quality. The information gained might be used as guidance for quality control, nutritional values and as a basis for the preparation of any food supplements for human health that employs date palm fruit as the raw material. © 2015 Institute of Food

Photoluminescence and time-resolved carrier dynamics in thiol-capped CdTe nanocrystals under high pressure

NASA Astrophysics Data System (ADS)

Lin, Yan-Cheng; Chou, Wu-Ching; Susha, Andrei S.; Kershaw, Stephen V.; Rogach, Andrey L.

2013-03-01

The application of static high pressure provides a method for precisely controlling and investigating many fundamental and unique properties of semiconductor nanocrystals (NCs). This study systematically investigates the high-pressure photoluminescence (PL) and time-resolved carrier dynamics of thiol-capped CdTe NCs of different sizes, at different concentrations, and in various stress environments. The zincblende-to-rocksalt phase transition in thiol-capped CdTe NCs is observed at a pressure far in excess of the bulk phase transition pressure. Additionally, the process of transformation depends strongly on NC size, and the phase transition pressure increases with NC size. These peculiar phenomena are attributed to the distinctive bonding of thiols to the NC surface. In a nonhydrostatic environment, considerable flattening of the PL energy of CdTe NC powder is observed above 3.0 GPa. Furthermore, asymmetric and double-peak PL emissions are obtained from a concentrated solution of CdTe NCs under hydrostatic pressure, implying the feasibility of pressure-induced interparticle coupling.

Endophytic Streptomyces in the traditional medicinal plant Arnica montana L.: secondary metabolites and biological activity.

PubMed

Wardecki, Tina; Brötz, Elke; De Ford, Christian; von Loewenich, Friederike D; Rebets, Yuriy; Tokovenko, Bogdan; Luzhetskyy, Andriy; Merfort, Irmgard

2015-08-01

Arnica montana L. is a medical plant of the Asteraceae family and grows preferably on nutrient poor soils in mountainous environments. Such surroundings are known to make plants dependent on symbiosis with other organisms. Up to now only arbuscular mycorrhizal fungi were found to act as endophytic symbiosis partners for A. montana. Here we identified five Streptomyces strains, microorganisms also known to occur as endophytes in plants and to produce a huge variety of active secondary metabolites, as inhabitants of A. montana. The secondary metabolite spectrum of these strains does not contain sesquiterpene lactones, but consists of the glutarimide antibiotics cycloheximide and actiphenol as well as the diketopiperazines cyclo-prolyl-valyl, cyclo-prolyl-isoleucyl, cyclo-prolyl-leucyl and cyclo-prolyl-phenylalanyl. Notably, genome analysis of one strain was performed and indicated a huge genome size with a high number of natural products gene clusters among which genes for cycloheximide production were detected. Only weak activity against the Gram-positive bacterium Staphylococcus aureus was revealed, but the extracts showed a marked cytotoxic activity as well as an antifungal activity against Candida parapsilosis and Fusarium verticillioides. Altogether, our results provide evidence that A. montana and its endophytic Streptomyces benefit from each other by completing their protection against competitors and pathogens and by exchanging plant growth promoting signals with nutrients.

Unusual thiol-based redox metabolism of parasitic flukes.

PubMed

Tripathi, Timir; Suttiprapa, Sutas; Sripa, Banchob

2017-08-01

Parasitic flukes are exposed to free radicals and, to a greater extent, reactive oxygen species (ROS) during their life cycle. Despite being relentlessly exposed to ROS released by activated immune cells, these parasites can survive for many years in the host. Cellular thiol-based redox metabolism plays a crucial role in parasite survival within their hosts. Evidence shows that oxidative stress and redox homeostasis maintenance are important clinical and pathobiochemical as well as effective therapeutic principles in various diseases. The characterization of redox and antioxidant enzymes is likely to yield good target candidates for novel drugs and vaccines. The absence of active catalase in fluke parasites offers great potential for the development of chemotherapeutic agents that act by perturbing the redox equilibrium of the cell. One of the redox-sensitive enzymes, thioredoxin glutathione reductase (TGR), has been accepted as a drug target against blood fluke infections, and related clinical trials are in progress. TGR is the sole enzyme responsible for Trx and GSH reduction in parasitic flukes. The availability of helminth genomes has accelerated the research on redox metabolism of flukes; however, significant achievements have yet to be attained. The present review summarizes current knowledge on the redox and antioxidant system of the parasitic flukes. Copyright © 2016 Elsevier Ireland Ltd. All rights reserved.

Thiol peroxidase deficiency leads to increased mutational load and decreased fitness in Saccharomyces cerevisiae.

PubMed

Kaya, Alaattin; Lobanov, Alexei V; Gerashchenko, Maxim V; Koren, Amnon; Fomenko, Dmitri E; Koc, Ahmet; Gladyshev, Vadim N

2014-11-01

Thiol peroxidases are critical enzymes in the redox control of cellular processes that function by reducing low levels of hydroperoxides and regulating redox signaling. These proteins were also shown to regulate genome stability, but how their dysfunction affects the actual mutations in the genome is not known. Saccharomyces cerevisiae has eight thiol peroxidases of glutathione peroxidase and peroxiredoxin families, and the mutant lacking all these genes (∆8) is viable. In this study, we employed two independent ∆8 isolates to analyze the genome-wide mutation spectrum that results from deficiency in these enzymes. Deletion of these genes was accompanied by a dramatic increase in point mutations, many of which clustered in close proximity and scattered throughout the genome, suggesting strong mutational bias. We further subjected multiple lines of wild-type and ∆8 cells to long-term mutation accumulation, followed by genome sequencing and phenotypic characterization. ∆8 lines showed a significant increase in nonrecurrent point mutations and indels. The original ∆8 cells exhibited reduced growth rate and decreased life span, which were further reduced in all ∆8 mutation accumulation lines. Although the mutation spectrum of the two independent isolates was different, similar patterns of gene expression were observed, suggesting the direct contribution of thiol peroxidases to the observed phenotypes. Expression of a single thiol peroxidase could partially restore the growth phenotype of ∆8 cells. This study shows how deficiency in nonessential, yet critical and conserved oxidoreductase function, leads to increased mutational load and decreased fitness. Copyright © 2014 by the Genetics Society of America.

Adaptive aneuploidy protects against thiol peroxidase deficiency by increasing respiration via key mitochondrial proteins.

PubMed

Kaya, Alaattin; Gerashchenko, Maxim V; Seim, Inge; Labarre, Jean; Toledano, Michel B; Gladyshev, Vadim N

2015-08-25

Aerobic respiration is a fundamental energy-generating process; however, there is cost associated with living in an oxygen-rich environment, because partially reduced oxygen species can damage cellular components. Organisms evolved enzymes that alleviate this damage and protect the intracellular milieu, most notably thiol peroxidases, which are abundant and conserved enzymes that mediate hydrogen peroxide signaling and act as the first line of defense against oxidants in nearly all living organisms. Deletion of all eight thiol peroxidase genes in yeast (∆8 strain) is not lethal, but results in slow growth and a high mutation rate. Here we characterized mechanisms that allow yeast cells to survive under conditions of thiol peroxidase deficiency. Two independent ∆8 strains increased mitochondrial content, altered mitochondrial distribution, and became dependent on respiration for growth but they were not hypersensitive to H2O2. In addition, both strains independently acquired a second copy of chromosome XI and increased expression of genes encoded by it. Survival of ∆8 cells was dependent on mitochondrial cytochrome-c peroxidase (CCP1) and UTH1, present on chromosome XI. Coexpression of these genes in ∆8 cells led to the elimination of the extra copy of chromosome XI and improved cell growth, whereas deletion of either gene was lethal. Thus, thiol peroxidase deficiency requires dosage compensation of CCP1 and UTH1 via chromosome XI aneuploidy, wherein these proteins support hydroperoxide removal with the reducing equivalents generated by the electron transport chain. To our knowledge, this is the first evidence of adaptive aneuploidy counteracting oxidative stress.

Solving the Jigsaw Puzzle of Wound-Healing Potato Cultivars: Metabolite Profiling and Antioxidant Activity of Polar Extracts

PubMed Central

2015-01-01

Potato (Solanum tuberosum L.) is a worldwide food staple, but substantial waste accompanies the cultivation of this crop due to wounding of the outer skin and subsequent unfavorable healing conditions. Motivated by both economic and nutritional considerations, this metabolite profiling study aims to improve understanding of closing layer and wound periderm formation and guide the development of new methods to ensure faster and more complete healing after skin breakage. The polar metabolites of wound-healing tissues from four potato cultivars with differing patterns of tuber skin russeting (Norkotah Russet, Atlantic, Chipeta, and Yukon Gold) were analyzed at three and seven days after wounding, during suberized closing layer formation and nascent wound periderm development, respectively. The polar extracts were assessed using LC-MS and NMR spectroscopic methods, including multivariate analysis and tentative identification of 22 of the 24 biomarkers that discriminate among the cultivars at a given wound-healing time point or between developmental stages. Differences among the metabolites that could be identified from NMR- and MS-derived biomarkers highlight the strengths and limitations of each method, also demonstrating the complementarity of these approaches in terms of assembling a complete molecular picture of the tissue extracts. Both methods revealed that differences among the cultivar metabolite profiles diminish as healing proceeds during the period following wounding. The biomarkers included polyphenolic amines, flavonoid glycosides, phenolic acids and glycoalkaloids. Because wound healing is associated with oxidative stress, the free radical scavenging activities of the extracts from different cultivars were measured at each wounding time point, revealing significantly higher scavenging activity of the Yukon Gold periderm especially after 7 days of wounding. PMID:24998264

Synthesis and Evaluation of Vitamin D Receptor-Mediated Activities of Cholesterol and Vitamin D Metabolites

PubMed Central

Teske, Kelly A.; Bogart, Jonathan W.; Sanchez, Luis M.; Yu, Olivia B.; Preston, Joshua V.; Cook, James M.; Silvaggi, Nicholas R.; Bikle, Daniel D.; Arnold, Leggy A.

2016-01-01

A systematic study with phase 1 and phase 2 metabolites of cholesterol and vitamin D was conducted to determine whether their biological activity is mediated by the vitamin D receptor (VDR). The investigation necessitated the development of novel synthetic routes for lithocholic acid (LCA) glucuronides (Gluc). Biochemical and cell-based assays were used to demonstrate that hydroxylated LCA analogs were not able to bind VDR. This excludes VDR from mediating their biological and pharmacological activities. Among the synthesized LCA conjugates a novel VDR agonist was identified. LCA Gluc II increased the expression of CYP24A1 in DU145 cancer cells especially in the presence of the endogenous VDR ligand 1,25(OH)2D3. Furthermore, the methyl ester of LCA was identified as novel VDR antagonist. For the first time, we showed that calcitroic acid, the assumed inactive final metabolite of vitamin D, was able to activate VDR-mediated transcription to a higher magnitude than bile acid LCA. Due to a higher metabolic stability in comparison to vitamin D, a very low toxicity, and high concentration in bile and intestine, calcitroic acid is likely to be an important mediator of the protective vitamin D properties against colon cancer. PMID:26774929

The glycolytic metabolite methylglyoxal activates Pap1 and Sty1 stress responses in Schizosaccharomyces pombe.

PubMed

Zuin, Alice; Vivancos, Ana P; Sansó, Miriam; Takatsume, Yoshifumi; Ayté, José; Inoue, Yoshiharu; Hidalgo, Elena

2005-11-04

Methylglyoxal, a toxic metabolite synthesized in vivo during glycolysis, inhibits cell growth. One of the mechanisms protecting eukaryotic cells against its toxicity is the glyoxalase system, composed of glyoxalase I and II (glo1 and glo2), which converts methylglyoxal into d-lactic acid in the presence of glutathione. Here we have shown that the two principal oxidative stress response pathways of Schizosaccharomyces pombe, Sty1 and Pap1, are involved in the response to methylglyoxal toxicity. The mitogen-activated protein kinase Sty1 is phosphorylated and accumulates in the nucleus following methylglyoxal treatment. Moreover, glo2 expression is induced by methylglyoxal and environmental stresses in a Sty1-dependent manner. The transcription factor Pap1 also accumulates in the nucleus, activating the expression of its target genes following methylglyoxal treatment. Our studies showed that the C-terminal cysteine-rich domain of Pap1 is sufficient for methylglyoxal sensing. Furthermore, the redox status of Pap1 is not changed by methylglyoxal. We propose that methylglyoxal treatment triggers Pap1 and Sty1 nuclear accumulation, and we describe the molecular basis of such activation mechanisms. In addition, we discuss the potential physiological significance of these responses to a natural toxic metabolite.

Diamond surface functionalization with biomimicry - Amine surface tether and thiol moiety for electrochemical sensors

NASA Astrophysics Data System (ADS)

Sund, James B.; Causey, Corey P.; Wolter, Scott D.; Parker, Charles B.; Stoner, Brian R.; Toone, Eric J.; Glass, Jeffrey T.

2014-05-01

The surface of conducting diamond was functionalized with a terminal thiol group that is capable of binding and detecting nitrogen-oxygen species. The functionalization process employed multiple steps starting with doped diamond films grown by plasma enhanced chemical vapor deposition followed by hydrogen termination and photochemical attachment of a chemically protected amine alkene. The surface tether was deprotected to reveal the amine functionality, which enabled the tether to be extended with surface chemistry to add a terminal thiol moiety for electrochemical sensing applications. Each step of the process was validated using X-ray photoelectron spectroscopy analysis.

Hmb(off/on) as a switchable thiol protecting group for native chemical ligation.

PubMed

Qi, Yun-Kun; Tang, Shan; Huang, Yi-Chao; Pan, Man; Zheng, Ji-Shen; Liu, Lei

2016-05-04

A new thiol protecting group Hmb(off/on) is described, which has a switchable activity that may be useful in the chemical synthesis of proteins. When placed on the side chain of Cys, Cys(Hmb(off)) is stable to trifluoroacetic acid (TFA) in the process of solid-phase peptide synthesis. When Cys(Hmb(off)) is treated with neutral aqueous buffers, it is cleanly converted to acid-labile Cys(Hmb(on)), which can later be fully deprotected by TFA to generate free Cys. The utility of Cys(Hmb(off/on)) is demonstrated by the chemical synthesis of an erythropoietin segment, EPO[Cys(98)-Arg(166)]-OH through native chemical ligation.

Biosynthesis of the active compounds of Isatis indigotica based on transcriptome sequencing and metabolites profiling

PubMed Central

2013-01-01

Backgroud Isatis indigotica is a widely used herb for the clinical treatment of colds, fever, and influenza in Traditional Chinese Medicine (TCM). Various structural classes of compounds have been identified as effective ingredients. However, little is known at genetics level about these active metabolites. In the present study, we performed de novo transcriptome sequencing for the first time to produce a comprehensive dataset of I. indigotica. Results A database of 36,367 unigenes (average length = 1,115.67 bases) was generated by performing transcriptome sequencing. Based on the gene annotation of the transcriptome, 104 unigenes were identified covering most of the catalytic steps in the general biosynthetic pathways of indole, terpenoid, and phenylpropanoid. Subsequently, the organ-specific expression patterns of the genes involved in these pathways, and their responses to methyl jasmonate (MeJA) induction, were investigated. Metabolites profile of effective phenylpropanoid showed accumulation pattern of secondary metabolites were mostly correlated with the transcription of their biosynthetic genes. According to the analysis of UDP-dependent glycosyltransferases (UGT) family, several flavonoids were indicated to exist in I. indigotica and further identified by metabolic profile using UPLC/Q-TOF. Moreover, applying transcriptome co-expression analysis, nine new, putative UGTs were suggested as flavonol glycosyltransferases and lignan glycosyltransferases. Conclusions This database provides a pool of candidate genes involved in biosynthesis of effective metabolites in I. indigotica. Furthermore, the comprehensive analysis and characterization of the significant pathways are expected to give a better insight regarding the diversity of chemical composition, synthetic characteristics, and the regulatory mechanism which operate in this medical herb. PMID:24308360

Allicin-inspired thiolated fluoroquinolones as antibacterials against ESKAPE pathogens.

PubMed

Sheppard, Jordan G; Long, Timothy E

2016-11-15

Thiolated fluoroquinolones were synthesized from ciprofloxacin and evaluated for antimicrobial activity against a panel of pathogenic bacteria. Gram-positive species including methicillin-resistant Staphylococcus aureus (MRSA) exhibited the highest level of increased sensitivity toward ciprofloxacin bound with a N-propylthio substituent. Evidence was found that the antibiotics form disulfides with low molecular weight thiols in bacteria and potentiate generation of cytosolic reactive oxygen species (ROS). In final analysis, the enhanced anti-MRSA activity of thiolated fluoroquinolones was attributed to increased cell permeability and reaction with cytosolic thiols that yields an inactive disulfide metabolite and the parent drug ciprofloxacin as an inhibitor of DNA synthesis. Copyright © 2016 Elsevier Ltd. All rights reserved.