Uric acid determinations: reversed-phase liquid chromatography with ultraviolet detection compared with kinetic and equilibrium adaptations of the uricase method

Abstract

A reversed-phase liquid-chromatographic procedure is presented for quantitation or uric acid in human serum, with absorbance measured at 292 nm. The mobile phase was sodium acetate (35 mmol/L, pH 5.0)/acetonitrile (9/1 by vol). Complete precipitation of serum proteins was obtained by mixing serum (50-500 microL) with an equal volume of acetonitrile, and the precipitate was removed by centrifugation. Aliquots (20 microL) of the supernate were injected directly into the liquid chromatograph, which was adjusted so that the absorbance reading of the uric acid peak was as high as possible. Routinely, a full-scale deflection of 1.28 absorbance units was used. The within-run precision (CV) was 0.6% for a serum uric acid concentration of 227 mumol/L and day-to-day precision over a 15-day period was 0.8% for uric acid of 345 mumol/L. No interferences from related compounds were observed. We compared results by this method with those by kinetic (aca, Du Pont) and equilibrium adaptations (Ames kit; Nyco-test, Nyegaard; and Monotest, Boehringer Mannheim) of uricase methods. The method we report is simple, and can be used in a fully automatic liquid-chromatographic system.