RP–HPLC Developed Method for Uric Acid Estimation in Human Serum

Abstract

A sensitive, efficient, reproducible and rapid method for the specific determination of uric acid as pure and in human serum has been developed using reverse phase high performance liquid chromatographic (RP - HPLC). This method involves separation of uric acid on reverse phase HPLC Shimadzu LC–20 A, Japan and Phenomenex C18 column (250 × 4.6 mm, 5µm). The elution was done using a mobile phase consisting of sodium acetate buffer: acetonitrile (ACN) as the ratio of (95:05 v/v with pH adjusted at 4.0 using acetic acid). The separation was monitored for 10 min. at 285 nm using a UV-visible detector and 1.2 mL/min flow rate. The optimum conditions such as the composition of the mobile phase, flow rate, wavelength and pH were studied. The obtained results revealed that the value of R2 is (0.9994), the detection limit (0.01), the quantitative limit (0.033), the linear ranges (0.05 – 30) µg / mL. Quantitative recoveries of pure uric acid and spiked serum samples were between 98.75 – 101.20%. Compared to the other methods, the current method is rapid, simple and economical for the determination of uric acid as pure and in human serum.