Microquantitative Simultaneous Analysis of Lactate and Pyruvate in Blood Plasma and Urine by High-Performance Liquid Chromatography

Abstract

A novel method for the microquantitative simultaneous determination of lactate and pyruvate in blood plasma and urine is described. The principle of the assay is to determine lactate after its enzymatic conversion to pyruvate followed by the derivatization of the latter into fluorescent 2-hydroxy-3-methylquinoxaline (QXL) and determination of it by reversed-phase high-performance liquid chromatography (HPLC). The complete conversion reaction mixture was optimized, preincubated with lactate dehydrogenase for 5min at 37°C, and deproteinized by the tungstic acid precipitation method. The supernatant was subjected to derivatization into QXL and then analyzed by HPLC. The present method is accurate and reproducible, and lactate can be measured up to 1, 200nmol in the conversion mixture. We also confirmed that the method is applicable for use in the clinical diagnosis of patients with acidemia, giving results in reasonably good agreement with those obtained by other conventional methods.