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Determination of thiol by high-performance liquid chromatography and fluorescence detection with 5-methyl-(2-(m-iodoacetylaminophenyl)benzoxazole

Liang, Shu-Cai, Wang, Hong, Zhang, Zhi-Min, Zhang, Hua-Shan
Analytical and bioanalytical chemistry 2005 v.381 no.5 pp. 1095-1100
acetylcysteine, beta-mercaptoethanol, cysteine, derivatization, detection limit, fluorescence, glutathione, high performance liquid chromatography, humans, molecular weight, penicillamine, thiols, urine, wavelengths
A new high-performance liquid chromatographic (HPLC) method for measuring low molecular weight (LMW) thiol-containing compounds, including cysteine (CysH), glutathione (GSH), N-acetylcysteine (Nac), penicillamine (PA), and 2-mercaptoethanol (2-ME), has been developed by using 5-methyl-(2-(m-iodoacetylaminophenyl)benzoxazole (MIPBO) as fluorescence-labeling reagent. The derivatization and separation conditions have been investigated in detail. Detection limits ranging from 3.5 to 15.0 fmol were achieved for the thiols investigated in a 16 min separation with detection wavelengths 310 and 375 nm for the excitation and emission, respectively. The utility of the proposed method has been validated by measuring CysH in human urine samples.