High‐performance liquid chromatography/chemiluminescence determination of biological thiols withn‐[4‐(6‐dimethylamino‐2‐benzofuranyl)phenyl]maleimide
作者:
Kenichiro Nakashima,
Chiemi Umekawa,
Shin'Ichi Nakatsuji,
Shuzo Akiyama,
Richard S. Givens,
期刊:
Biomedical Chromatography
(WILEY Available online 1989)
卷期:
Volume 3,
issue 1
页码: 39-42
ISSN:0269-3879
年代: 1989
DOI:10.1002/bmc.1130030110
出版商: John Wiley&Sons, Ltd.
数据来源: WILEY
摘要:
AbstractThe 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 withN‐[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 andN‐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
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