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[DOI] [PMC free article] [PubMed] [Google Scholar] Muttenthaler M
Ascorbic Acid
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The present invention provides a method of stabilising a biological sample comprising glutathione (GSH) and glutathione disulfide (GSSG), the method comprising the following steps: a) providing the biological sample comprising GSH and GSSG, b) contacting GSH and GSSG of the biological sample with a maleimide under conditions which allow alkylation of the sulfhydryl group of GSH with the maleimide to obtain maleimide-alkylated GSH, c) separating excess maleimide from maleimide-alkylated GSH and GSSG, d) contacting maleimide-alkylated GSH and GSSG with a reducing agent, selected from the group consisting of phosphine-based reducing agents such as tris(2-carboxyethyl)phosphine (TCEP) or tributylphosphine, and thiol-based reducing agents such as dithiothreitol (DTT), dithioerythritol (DTE) or β-mercaptoethanol, under conditions which allow reduction of GSSG by the reducing agent to obtain further GSH, and e) contacting maleimide-alkylated GSH and GSH with a heavy isotopologue of the maleimide under conditions which allow alkylation of the sulfhydryl group of GSH with the heavy isotopologue of the maleimide to obtain a heavy isotopologue of the maleimide-alkylated GSH
