Liquid chromatography-tandem mass spectrometry assay of reduced and oxidized glutathione and main precursors in mice liver.

A liquid chromatography/tandem mass spectrometry assay of glutathione (GSH), glutathione disulfide (GSSG) and of precursors (gamma-glutamyl-cysteine, cysteinyl-glycine, cysteine, cystine, homocysteine and homocystine) was developed to study glutathione synthesis in mice liver. After iodoacetic acid derivatization, the analytes were analyzed using reversed-phase gradient HPLC and detected using multiple reaction monitoring. Linear calibrations were performed over the concentrations range of 100-10,000 ng/mL for the thiol-containing precursors and extended up to 100,000 ng/mL for GSH and GSSG. The method was validated for each compound with inter-day accuracy below 11.9% and with precision below 15%. The method showed low limits of quantitation of 100 ng/mL for each thiol-containing compound and GSSG and of 200 ng/mL for other disulfides.