High‐performance liquid chromatography/chemiluminescence determination of biological thiols with n‐[4‐(6‐dimethylamino‐2‐benzofuranyl)phenyl]maleimide

Publisher Website
Google Scholar
Abstract
The peroxyoxalate chemiluminescence detection of biological thiols combined with high-performance liquid chromatography (HPLC) is described. SH groups of the thiol compounds including glutathione (GSH), cysteine, N-acetylcysteine, cysteamine, and D-penicillamine were labelled with N-[4-(6-dimethylamino-2-benzofuranyl)phenyl]maleimide (DBPM), a specific fluorogenic reagent for SH group. The labelling reaction was carried out at 60°C for 30 min and at pH 8.5 and a sample of the resulting reaction mixture was subjected to HPLC. Five kinds of labelled thiols were separated within 12 min on ODS-80 column (150 × 4.6mm ID; 5 μm) and detected in the ranges from 500 fmol to 2 pmol/100 μL (cysteamine and N-acetylcysteine), to 3 pmol/100 μL (cysteine) and to 5 pmol/100 μL (GSH and D-penicillamine). The lower detection limits were from 7 fmol (cysteamine) to 113 fmol (GSH) per 100 μL (S/N = 2). The method was applied to the determination of thiols in a rat liver. The amounts of glutathione and cysteine were 1.23±0.15 μmol/g (n = 5) and 0.15±0.04 μmol/g (n = 5), respectively.

This publication has 32 references indexed in Scilit: