Serum uric acid determined by reversed-phase liquid chromatography with spectrophotometric detection.

We describe a method for determining uric acid in serum by reversed-phase liquid chromatography with spectrophotometric detection at 280 nm. Serum, 100 ul is mixed with 100 u1 of a solution containing, per liter, 70 ml of acetonitrile in sodium acetate (20 mmol/Iiter, pH 4.0) and 500 mg of the internal standard, adenine. The mixture is allowed to stand in an ice bath for 3 mm, then centrifuged. A 7.5tl portion of the supernate is chromatographed on a “ Bondpak C18” column, with a 35 mI/liter solution of acetonitrile in sodium acetate (10 mmol/liter, pH 4.0) as the mobile phase. For 10 runs of duplicates, the within-run CV was 1.2% and the day-to-day CV (10 days) was 2.5% for a uric acid concentration of 53 mg/liter. Sera from 100 patients were analyzed for uric acid by the proposed method, a continuous-flow (SMA 12/60) method, and a uricase method; mean values for the 100 sera were 64, 71, and 64 mg/liter, respectively. Correlations were as follows: r = 0.987 for proposed method vs. SMA 12/60 and r = 0.997 for proposed method vs. the uricase method. The proposed method is sensitive and specific and we think it will be useful for evaluating other i.ilc acid methods for interference and specificity.