Premium
Rapid liquid chromatographic assay of glutathione in cultured cells
Author(s) -
Leroy Pierre,
Nicolas Alain,
Thioudellet Christine,
Oster Thierry,
Wellman Maria,
Siest Gérard
Publication year - 1993
Publication title -
biomedical chromatography
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 0.4
H-Index - 65
eISSN - 1099-0801
pISSN - 0269-3879
DOI - 10.1002/bmc.1130070208
Subject(s) - chemistry , glutathione , chromatography , derivatization , repeatability , detection limit , quantitative analysis (chemistry) , high performance liquid chromatography , reversed phase chromatography , biochemistry , enzyme
A rapid, sensitive and selective method for the assay of glutathione in cultured cells has been developed using ion‐pair reversed phase rapid high performance liquid chromatography. The use of a 4 μm particle, 5 cm long column (Superspher 100 RP 18 end‐capped) allowed complete analysis of glutathione within 3 min. A postcolumn derivatization reaction with o ‐phthaladehyde and fluorometric detection made the assay fully selective with regard to other endogenous thiols and sensitive (the detection limit was 0.5 ng of glutathione injected). The linearity range was between 0.1 and 2.0 μg/mL with good repeatability (relative standard deviation less than 5% for the lowest concentration quantitated). Recoveries of GSH from cultured cell samples were above 98%. The rapid analysis enabled the processing of a large number of samples in a short time (up to 20 per hour). The method was applied to the measurement of the intracellular glutathione amount in V79 fibroblasts along cell growth in culture.