Quantitation glutathione

Accumulation of Ag on protein A-coated crystals to which Ab has been bound has been used to quantitate glutathione-5-transferase (GST) [133]. The piezoelectric (PZ) biosensor detects a decrease in the resonant frequency of the PZ oscillating crystal when antigens are adsorbed on it. 

The quantitative analysis for reduced glutathione in blood is complicated by the presence of many potential interferents. 

Soffers AEMF, Ploeman JHTM, Moonen MJH, Wobbes T, van Ommen B, Vervoort J, et al. Regioselectivity and quantitative structure-activity relationships for the conjugation of a series of fluoronitrobenzenes by purified glutathione S -transferase enzymes from rat and man. Chem Res Toxicol 1996 9 638-46. 

Rao et. al recently published another method for the quantitation of the MB fraction. The method is based on the selective activating capacity of dithiothreitol on CR isoenzyme MB, after isoenzyme MM is activated by glutathione (51). Apparently the MB isoenzyme is not activated by glutathione" Eut is activated by dithiothreitol. The difference between CR activities obtained in the presence of glutathione alone and those obtained with both glutathione and dithiothreitol represent MB activity. The correlation is excellent (r 0.998) between the activity of MB in the isoenzyme mixture determined by this method, and the activity of isolated MB. 

Simple etching of the capillary end served to decouple the electrophoretic current from that of amperometric detection, permitting quantitation of attomole levels of catecholamines from brain microdialyzates.24 A postcolumn reactor using bromine generated electrochemically in situ has been used in the detection of peptide thiols, such as glutathione and cysteine, separated by capillary electrophoresis.25... 

Gan, J. et al. 2005. Dansyl glutathione as a trapping agent for the quantitative estimation and identification of reactive metabolites. Chem. Res. Toxicol. 18 896. 

Carter JH, Richmond RE, Carter HW, et al. 1992. Quantitative image cytometry of hepatocytes expressing gamma-glutamyl transpeptidase and glutathione S-transferase in diethylnitrosamine-initiated rats treated with phenobarbital and/or phthalate esters. J Histochem Cytochem 40 1105-1115. 

An analogous assay using a radiolabeled soft nucleophile would also be required to complement the hard nucleophile radiolabeled cyanide trapping assay. Investigations into radiolabeled glutathione have proved unsuccessful since the material is unstable due to cross-reactions induced by beta radiation. Alternate soft nucleophiles such as cysteine, N-acetyl cysteine and P-mercaptoethanol all have promise as radiolabeled substances for quantitative trapping experiments since they are more stable than GSH and equally nucleophilic, although clearly these would not be substrates for GST. 

Lehrfeld, J. HPLC separation and quantitation of phytic acid and some inositol phosphates in foods. J Agr Food Chem 1994 42(12) 2726-2731. Obata, H., K. Imai, N. Inoue, and M. Umebayashi. A highly sensitive quantitative determination of glutathione in plant roots by high performance gel filtration chromatography and fluoro-metic detection after pre-column derivatization with N-(7-dimethyl-amino-4-methyl coumarinyl)-malei-mide. Phytochem Anal 1994 5(5) 239-242. 

Plagia D, Valentine W (1967) Studies on the quantitative and qualitative characterization of erythrocyte glutathione peroxidase. J Lab Chn Med 70 158-169... 

Sharer, J.E. Elfarra, A.A. (1992) A -(2-Hydroxy-3-buten-l-yl)glutathione and A -(l-hydroxy-3-buten-2-yl)glutathione are in vivo metabolites of butadiene monoxide detection and quantitation in bile. Chem. Res. Toxicol., 5, 787-790... 

These data suggest a quantitative difference in the formation of AMCC between species, most likely in the formation of the reactive carbamoylating intennediate which acylates glutathione. 

An HPLC method was developed and validated for the determination and quantitation of both DFC cysteine and glutathione conjugates. This method was based on a cleavage of the disulphide and/or thioester bonds between the metabolites and their conjugate sulphur-containing moiety using dithioerythritol to yield DFC, which was then stabilized by derivatizing to DFC acetamide (91). 

Zhang F, Bartels MJ, Stott WT Quantitation ofhuman glutathione S-transferases in complex matrices by liquid chromatography/tandem mass spectrometry with signature peptides. Rapid Commun. Mass Spectrom. (2004) 18 491 98. 

Substrate Solution Transfer 12.110 g of Tris[tri(hydroxy-methyl)aminomethane], 3.475 g of hydroxylamine hydrochloride, 1.624 g of glutathione, and 5.060 g of carbobenzyloxy-glutaminylglycine into a 500-mL beaker. Add 350 mL of water, and using a magnetic stirrer, mix well. Adjust the pH to 6.0 with appropriate concentrations (usually 1 N and 6 N) of hydrochloric acid. Quantitatively transfer this mixture into a 500-mL volumetric flask, and dilute to volume with water. 

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