HPLC assay of Lidocaine in plasma with solid phase extraction and UV detection

Abstract

A sensitive and reliable method based on solid-phase extraction and reversed-phase liquid chromatography was developed and validated for the quantitation of Lidocaine (Lid) in dog plasma. Phenacemide was used as an internal standard (IS) in the extraction which employed C 18 solid-phase extraction cartridges. The washing and eluting solutions were 2 ml acetonitrile-pH 9.0 phosphate buffer (10:90 v/v) and 0.5 ml acetonitrile-pH 4.0 phosphate buffer (40:60 v/v), respectively. The eluent obtained from the cartridge was directly analyzed on a reversed-phase ODS column with UV detection at 210 nm. A clean chromatogram and high sensitivity were achieved at this wavelength. The mobile phase was acetonitrile and pH 5.9 phosphate buffer (20:80 v/v). The retention times were 6.4 and 7.2 min for Lid and IS, respectively, at a flow rate of 1.0 ml min −1. The mean absolute recovery was 96.6% ( n=9) with a CV of 3.8% for Lid and 81.7% with CV of 2.5% ( n=3) for IS. The limit of quantitation was 20 ng ml −1, with the intra- and inter-day precisions ( n=5) of 4.4 and 3.4%, respectively, and the intra- and inter-day accuracies ( n=5) of −4.3 and −5.0%, respectively. For the analyses of Lid in spiked plasma samples at 20, 100 and 200 ng ml −1, the overall mean intra- and inter-day precisions ( n=15) were 3.9 and 4.9%, respectively, and the overall mean intra- and inter-day accuracies ( n=15) were −3.7 and −4.6%, respectively. The correlation coefficients for calibration plots in the range 20–1000 ng ml −1 in plasma were typically higher than 0.998. The suitability of the method was demonstrated by the study in a beagle dog receiving a low intravenous dose of Lid.