Determination of the new monoamine oxidase inhibitor moclobemide and three of its metabolites in biological fluids by high-performance liquid chromatography

A sensitive high-performance liquid chromatographic method has been developed for the determination of moclobemide and three of its metabolites in plasma and urine. The four substances and the internal standard were extracted from basified plasma (pH 11) with dichloromethane, with recoveries of generally more than 70%. A column packed with 5-microns Spherisorb hexyl phase and an eluent consisting of acetonitrile and aqueous phosphate buffer 30:320 (v/v) with a pH of 3.9 were found most suitable for the chromatographic separation of the four compounds of interest. Non-ideal interactions of the basic compounds and the N-oxide metabolite occurred with many other reversed-phase Materials, as indicated by broad and tailing peaks. Limits of quantitation for moclobemide and its three metabolites in the range 20-30 ng/ml and a good intra-assay reproducibility ranging between 2 and 5% for concentrations over 100 ng/ml could be achieved for plasma, which increased to ca. 8-10% at a concentration of 50 ng/ml.