Elsevier

Analytical Biochemistry

Volume 141, Issue 2, September 1984, Pages 481-487
Analytical Biochemistry

Fluorescent analysis of α-keto acids in serum and urine by high-performance liquid chromatography

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Abstract

A procedure for rapid separation and microquantitative determination of various α-keto acids in serum and urine was developed. The procedure used reverse-phase high-performance liquid chromatography of the α-keto acids after derivatization into fluorescent quinoxalines by reaction with o-phenylenediamine. Deproteinization of serum with tungstic acid or methanol facilitated a constant recovery of α-keto acids. The useful range of analysis of seven α-keto acids by isocratic chromatography was from 10 to 250 pmol. The fluorescence emission was measured at 410 nm with excitation at 350 nm. The data obtained from samples of patients with chronic pyruvic acidemia and maple syrup urine disease, confirmed the usefulness of the method in clinical applications. A slightly modified procedure was needed for the analysis of oxaloacetic acid and phenylpyruvic acid.

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