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New assay procedure for glutathione S‐transferase using analytical capillary isotachophoresis: A procedure for the simultaneous analysis of reduced and oxidized glutathione together with glutathione conjugates of electrophilic compounds
Author(s) -
Holloway Christopher J.,
Battersby Rüdiger V.
Publication year - 1986
Publication title -
electrophoresis
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 0.666
H-Index - 158
eISSN - 1522-2683
pISSN - 0173-0835
DOI - 10.1002/elps.1150070704
Subject(s) - glutathione , chemistry , isotachophoresis , chromatography , conjugate , capillary electrophoresis , electrophile , homologous series , glutathione s transferase , electrophoresis , quantitative analysis (chemistry) , dinitrobenzene , combinatorial chemistry , biochemistry , organic chemistry , enzyme , electrolyte , mathematical analysis , mathematics , electrode , catalysis
A procedure for the simultaneous analysis of reduced and oxidized glutathione together with glutathione conjugates of electrophilic organic compounds by capillary isotachophoresis is described. This method can be applied to quality control of such synthetic conjugates, which are commonly employed as affinity ligands in the chromatographic purification of glutathione S‐transferases (EC 2.5.1.18). Using this new assay procedure, it has been found that the glutathione conjugate of 1‐chloro‐2,4‐dinitrobenzene is unstable in neutral to alkaline solution, degrading to glutathione and 2,4‐dinitrophenol. Anomalies in the electrophoretic mobilities of a homologous series of alkylglutathione derivatives were observed. A possible explanation is provided on the basis of the formation of micelle‐like aggregates, this tendency increasing with increasing length of the hyrocarbon chain. A useful and widely applicable aspect of this analytical procedure is as a method of monitoring glutathione S‐transferase. The major advantages of this new assay technique can be summarized as follows: (1) the assay does not depend on optical properties of the acceptor, (2) the reaction can be monitored in the presence of unrefined biological material;aliquots can be injected without pretreatment and (3) the spontaneous oxidation of glutathione in the assay mixture can be monitored simultaneously.