Quantitative determination of cis-3,4-dichloro-N-methyl-N-(2-(1-pyrrolidinyl)-cyclohexyl)-benzamide and three of its metabolites in plasma by high-performance liquid chromatography.

A sensitive HPLC method was developed for simultaneous quantitation of cis-3,4-dichloro-N-methyl-N-[2-(1-pyrrolidinyl)-cyclohexyl]-benzamide and three of its metabolites in dog plasma. The method involved selective solid-phase extraction of the compounds of interest from dog plasma and HPLC separation of the analytes on a cyano column. Absorbance of the column effluent was monitored at 230 nm by a UV detector. The analytical procedure has a linear range of 10 ng/mL to 20 micrograms/mL, with a low limit of quantitation of 10 ng/mL for each analyte. The accuracy and intra- and interassay precision for each compound were < or = 11% in the concentration range evaluated. Applicability of this method to the quantitation of 1 and its metabolites was assessed in a preclinical pharmacokinetic study.