DETERMINATION OF ALIPHATIC THIOLS BY FLUOROMETRIC HIGH-PERFORMANCE LIQUID-CHROMATOGRAPHY AFTER PRECOLUMN DERIVATIZATION WITH 2-(4-N-MALEIMIDOPHENYL)-6-METHYLBENZOTHIAZOLE

A sensitive, fluorometric high-performance liquid chromatographic meth od for the detection of aliphatic thiols, following pre-column derivat ization with 2-(4-N-maleimidophenyl)-6-methylbenzothiazole, has been d eveloped. The N-maleimide, the acid and the methyl ester derivatives o f the commercially available 2-(4-aminophenyl)-6-methylbenzothiazole w ere synthesized and found to be equally effective for the precolumn de rivatization procedure. The resulting fluorescentic derivatives of ali phatic thiols were separated on a reversed-phase column (Ultrasphere-O DS) using 0.1% hexane-sulfonic acid in 10 mM potassium hydrogen phosph ate: acetonitrile (65:35) as a mobile phase and were detected fluorome trically (excitation 320 nm; emission 405 nm). The method is highly se nsitive (femtomole range) and is easily applied for determination of S H-containing drugs and endogenous thiols in biological samples.