Simple method for determination of the active metabolite of the inotropic drug pimobendan in rat liver microsomes

A rapid and sensitive high-performance liquid chromatographic method for qu antitation of the O-demethylated active metabolite formed in a liver micros omal assay system has been developed. The metabolite was separated on an In ertsil ODS-2 column and quantitated by fluorescence detection (excitation a t 338 nm, emission at 405 nm). The retention times for pimobendan and its m etabolite were 5.6 and 2.8 min, respectively. The intra- and inter-assay re lative standard deviations in the measurement of pimobendan O-demethylase a ctivity at the substrate concentrations of 1 mu M and 500 mu M were 2.0%, 6 .8%, 2.1% and 5.6%, respectively, and the limit of detection was 0.1 ng for the demethylated metabolite. (C) 2000 Elsevier Science B.V. All rights res erved.