High-performance liquid chromatographic determination of alpha-keto acids in human serum and urine using 1,2-diamino-4,5-methylenedioxybenzene as a precolumn fluorescence derivatization reagent.

Z J Wang, K Zaitsu, Y Ohkura

Index: J. Chromatogr. A. 430(2) , 223-31, (1988)

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Abstract

A simple and highly sensitive high-performance liquid chromatographic (HPLC) method for the determination of alpha-keto acids in human serum and urine is described. In an acidic solution, twelve species of alpha-keto acids examined were converted by reaction with 1,2-diamino-4,5-methylenedioxy-benzene into highly fluorescent derivatives. The derivatives were separated isocratically by reversed-phase HPLC on a TSK gel ODS-80TM column and detected fluorimetrically. Eight alpha-keto acids in human serum and eleven alpha-keto acids in human urine can be determined simultaneously. The detection limits (signal-to-noise ratio = 5) are 6-44 fmol in an injection volume of 5 microliters. The intra-assay relative standard deviations for both serum and urine sample analyses are usually ca. 5%.