TITLE

Rapid determination of reduced and oxidized glutathione levels using a new thiol-masking reagent and the enzymatic recycling method: application to the rat liver and bile samples

AUTHOR(S)
Shaik, Imam H.; Mehvar, Reza
PUB. DATE
May 2006
SOURCE
Analytical & Bioanalytical Chemistry;May2006, Vol. 385 Issue 1, p105
SOURCE TYPE
Academic Journal
DOC. TYPE
Article
ABSTRACT
A microtiter plate assay for quantitation of reduced (GSH) and oxidized (GSSG) glutathione in the rat liver tissue and bile is described. The assay is based on the established enzymatic recycling method and a new thiol-masking reagent, 1-methyl-4-vinyl-pyridinium trifluoromethane sulfonate (M4VP). Samples were first processed by homogenization with (liver) or addition of (bile) sulfosalicylic acid. The total glutathione and GSSG were then determined before and after rapid (=2 min) and efficient (100%) masking of the GSH content of the samples with M4VP followed by the enzymatic recycling assay. The percentages of error and coefficient of variation of the assay were within the accepted guidelines, indicating the accuracy and precision of the assay in the range of 6.25�100 pmol GSH per microplate well and 2.17�140 pmol GSSG per well, with lower limit of quantitation of 6.25 and 2.17 pmol per well for GSH and GSSG, respectively. Furthermore, the recoveries of added GSH or GSSG from the liver and bile samples were accurate and precise. The assay was applied to measurement of GSH, GSSG, and GSH:GSSG ratio in the liver and serially collected bile samples in sham-operated and ischemic rat livers, demonstrating a depletion of glutathione and a decrease in the GSH:GSSG ratio as a result of ischemia. The developed assay is rapid, sensitive, accurate, and precise and is suitable for studies of the redox status of liver under physiologic and pathophysiologic conditions.
ACCESSION #
20535721

 

Related Articles

  • Prodrug Approach for Increasing Cellular Glutathione Levels. Cacciatore, Ivana; Cornacchia, Catia; Pinnen, Francesco; Mollica, Adriano; Di Stefano, Antonio // Molecules;Mar2010, Vol. 15 Issue 3, p1242 

    Reduced glutathione (GSH) is the most abundant non-protein thiol in mammalian cells and the preferred substrate for several enzymes in xenobiotic metabolism and antioxidant defense. It plays an important role in many cellular processes, such as cell differentiation, proliferation and apoptosis....

  • Troglitazone Thiol Adduct Formation in Human Liver Microsomes: Enzyme Kinetics and Reaction Phenotyping. Jinping Gan; Qinling Qu; Bing He; Shyu, Wen C.; Rodrigues, A. David; Kan He // Drug Metabolism Letters;Aug2008, Vol. 2 Issue 3, p184 

    Troglitazone (TGZ) induced hepatotoxicity has been linked to cytochrome P450 (CYP)-catalyzed reactive metabolite formation. Therefore, the kinetics and CYP specificity of reactive metabolite formation were studied using dansyl glutathione (dGSH) as a trapping agent after incubation of TGZ with...

  • Content of Total Homocysteine and Major Aminothiols in Rats with Experimental Renal Ischemia. Kuz'menko, N. V.; Alexandrova, L. A.; Chefu, S. G.; Blashko, E. L.; Zhloba, A. A.; Cirlin, V. A. // Bulletin of Experimental Biology & Medicine;May2011, Vol. 151 Issue 1, p22 

    The contents of total homocysteine, cysteine, and glutathione in blood plasma and tissue of rats with renal ischemia were measured by HPLC. Our study was performed on the 'two-kidney, one-clip (0.13 mm)' model. The concentrations of homocysteine and cysteine in blood plasma from treated rats...

  • Glutathione: a review on biotechnological production. Yin Li; Gongyuan Wei; Jian Chen // Applied Microbiology & Biotechnology;Jan2005, Vol. 66 Issue 3, p233 

    This Mini-Review summarizes the historic developments and technological achievements in the biotechnological production of glutathione in the past 30 years. Glutathione is the most abundant non-protein thiol compound present in living organisms. It is used as a pharmaceutical compound and can be...

  • Cadmium Accumulation and Antioxidative Responses in the Sesbania drummondii Callus. Israr, M.; Sahi, S. V.; Jain, J. // Archives of Environmental Contamination & Toxicology;Jan2006, Vol. 50 Issue 1, p121 

    The effect of cadmium (Cd) on growth, accumulation, and antioxidative response was studied in Sesbania drummondii callus, cultivated on different concentrations of Cd (0–250 μM) for four weeks. Callus growth was comparable to that of the control for concentrations up to 50 μM Cd;...

  • Purification, Characterization, and Effect of Thiol Compounds on Activity of the Erwinia carotovora L-Asparaginase. Warangkar, Suchita C.; Khobragade, Chandrahas N. // Enzyme Research;2010, p1 

    L-asparaginase was extracted from Erwinia carotovora and purified by ammonium sulfate fractionation (60-70%), Sephadex G-100, CM cellulose, and DEAE sephadex chromatography. The apparent Mr of enzyme under nondenaturing and denaturing conditions was 150 kDa and 37 ±0.5 kDa, respectively....

  • Enzymes of the Trypanothione Metabolism as Targets for Antitrypanosomal Drug Development. Schmidt, Armin; Krauth-Siegel, R. Luise // Current Topics in Medicinal Chemistry;Nov2002, Vol. 2 Issue 11, p1239 

    Trypanosomatids are the causative agents of African sleeping sickness, Chagas' disease and the three forms of leihmaniasis. New drugs against these parasitic protozoa are urgently needed since the currently available chemotherapy is not at all satisfying. One promising approach towards the...

  • Influence of Changes in Glutathione Concentration on Body Temperature and Tolerance to Cerebral Ischemia. Kolesnichenko, L. S.; Kulinsky, V. I.; Sotnikova, G. V.; Kovtun, V. Yu. // Biochemistry (00062979);May2003, Vol. 68 Issue 5, p534 

    Two compounds that deplete glutathione (buthionine sulfoximine and diethyl maleate) with different mechanisms of action decrease body temperature and increase tolerance to complete global cerebral ischemia, both correlating closely with the glutathione concentration decrease. Glutathione...

  • Pyridinium Oxime Reactivators of Cholinesterase Inhibited by Diisopropyl-Fluorophosphate (DFP): Predictive Value of In-Vitro Testing for In-Vivo Efficacy. Petroianu, G. A.; Lorke, D. E. // Mini Reviews in Medicinal Chemistry;Nov2008, Vol. 8 Issue 13, p1328 

    Poisoning with organophosphorus cholinesterase inhibitors (OPCs) poses a serious global threat. Therapy comprises the use of atropine and pyridinium oximes to reactivate acetylcholinesterase (AChE). Clinical experience with established oximes (pralidoxime and obidoxime) is disappointing and...

Share

Read the Article

Courtesy of THE LIBRARY OF VIRGINIA

Sorry, but this item is not currently available from your library.

Try another library?
Sign out of this library

Other Topics