High-performance liquid chromatography with amperometric detection applied to the screening of 1,4-dihydropyridines in human plasma

A high-performance liquid chromatographic method with electrochemical detection has been developed for the determination of six 1,4-dihydropyridines: nifedipine, nimodipine, nisoldipine, nicardipine, felodipine and lacidipine. The chromatographic separation was performed using a Supelcosil LC-ABZ+Plus C₁₈ column. A mobile phase of methanol–water (70:30), containing 2 mM CH₃COOH–CH₃COONa at a flow-rate of 1 ml/min and a pH of 5.0, was used. The temperature was optimized at 30±0.2°C. The amperometric detector, equipped with a glassy carbon electrode, was operated at 1000 mV versus Ag/AgCl in the direct current mode. The method was applied to the determination of these compounds at ng/ml concentrations, obtaining intra-day reproducibilities of lower than 5.0% in terms of relative standard deviations and detection limits ranging from 16 to 44 ng/ml. The method was applied to the screening of 1,4-dihydropyridines in spiked plasma samples, with a total elution time of lower than 18 min, obtaining the best recoveries for nimodipine and felodipine (91 and 88%, respectively). These recoveries together with the low detection limits achieved allow its application to the analysis of these drugs in human plasma.