DEVELOPMENT OF AN ASSAY FOR THE SIMULTANEOUS DETERMINATION OF SILDENAFIL (VIAGRA) AND ITS METABOLITE (UK-103,320) USING AUTOMATED SEQUENTIAL TRACE ENRICHMENT OF DIALYSATES AND HIGH-PERFORMANCE LIQUID-CHROMATOGRAPHY

The development of the ASTED (automated sequential trace enrichment of dialysates) system to prepare plasma samples prior to high-performanc e liquid chromatography (HPLC) of sildenafil and its demethylated meta bolite (UK-103,320) is described. Investigations to elucidate potentia l pitfalls of the ASTED on-line sample preparation system prior to sep aration of the analytes by HPLC are presented. The procedure is shown to be selective for sildenafil and UK-103,320, and linear over the ran ge 1.00-250 ng/ml. The intra-batch imprecision (C.V.) of the method at plasma analyte concentrations of 1.00, 5.00, 50.0 and 200 ng/ml was 1 1.2, 3.10, 1.50, and 1.30%, respectively, and the corresponding inter- batch imprecision was estimated to be 13.5, 7.09, 3.69, and 5.43%. At these plasma analyte concentrations the overall inaccuracy (% bias) of the procedure ranged from 3.6 to 8.4%. The method showed similar assa y performance for the estimation of the metabolite, UK-103,320. The ap plication of the assay to a pharmacokinetic investigation during a cli nical study is presented. (C) 1997 Elsevier Science B.V.