Simultaneous determination of guanine, uric acid, hypoxanthine and xanthine in human plasma by reversed-phase high-performance liquid chromatography with amperometric detection.
A reversed-phase high-performance liquid chromatographic method with amperometric detection is described for the separation and quantification of uric acid, guanine, hypoxanthine and xanthine. The isocratic separation of a standard mixture of the compounds was achieved in 5 min on a Spherisorb 5 C18 reversed-phase column, with a mobile phase of NaH2PO4 (300 mmol dm-3, pH 3.0)-methanol-acetonitrile-tetrahydrofuran (97.8 + 0.5 + 1.5 + 0.2). Uric acid, guanine, hypoxanthine and xanthine were completely separated, with detection limits in the range 2-20 pmol per injection. The effect of pH and the composition of the mobile phase on the separation are described. The hydrodynamic voltammograms of these compounds were recorded at a glassy carbon electrode. The linear range of the calibration graph for each compound was: uric acid, 1-5000 mumol dm-3; guanine, 0.5-2000 mumol dm-3; hypoxanthine, 0.1-500 mumol dm-3 and xanthine, 0.5-5000 mumol dm-3. The within- and between-day precision was good. The uric acid and hypoxanthine content in human plasma was measured using the proposed method. Good recoveries of uric acid (97.9-103%), hypoxanthine (98.0-99.2%), guanine (96.0-98.3%) and xanthine (96.0-102%) were obtained from human plasma. The results of electrochemical detection were in good agreement with those of UV detection.
Liu
Z
Z
Laboratory of Electroanalytical Chemistry, Changchun Institute of Applied Chemistry, Chinese Academy of Science.
Li
T
T
Wang
E
E
D002138:Calibration; D002851:Chromatography, High Pressure Liquid / Q000379:methods* / Q000706:statistics & numerical data; D004563:Electrochemistry; D006147:Guanine / Q000097:blood*; D006801:Humans; D006863:Hydrogen-Ion Concentration; D019271:Hypoxanthine; D007042:Hypoxanthines / Q000097:blood*; D012680:Sensitivity and Specificity; D014527:Uric Acid / Q000097:blood*; D019820:Xanthine; D014970:Xanthines / Q000097:blood*
D007042:Hypoxanthines; D014970:Xanthines; D019820:Xanthine; D014527:Uric Acid; D019271:Hypoxanthine; D006147:Guanine