High-performance liquid chromatographic method with amperometric detection employing boron-doped diamond electrode for the determination of sildenafil, vardenafil and their main metabolites in plasma

Abstract

A simple, fast and sensitive HPLC method with electrochemical detection employing boron-doped diamond electrode (BDD) for the determination of sildenafil (Viagra™), vardenafil (Levitra™) and their main metabolites, N-desmethyl sildenafil and N-desethyl vardenafil in human plasma is presented. The assay involved drug extraction by tert-butyl methyl ether and isocratic reversed-phase liquid chromatography with amperometric detection. Complete separation of all analytes was achieved within 12min. The mobile phase consisted of 20mM sodium dihydrogen phosphate with 40mM sodium perchlorate/acetonitrile (70:30, v/v), pH 3.5. The electrode working potential was +1520mV (vs. Pd/H2). Calibration curves were linear over the concentration range of 10-400ngmL-1. Phloretin was used as an internal standard. The limits of detection (LOD) and quantification (LOQ) for the studied analytes were within the range of 2-4ngmL-1 and 7.0-13.4ngmL-1, respectively. The developed method was applied to human plasma samples spiked with analytes at therapeutic concentrations. The study confirms the method's suitability for both pharmacokinetic studies and therapeutic monitoring. © 2011 Elsevier B.V.