DETERMINATION OF THE AROTINOID MOFAROTENE IN HUMAN, RAT AND DOG PLASMA BY HIGH-PERFORMANCE LIQUID-CHROMATOGRAPHY WITH AUTOMATED COLUMN-SWITCHING AND ULTRAVIOLET DETECTION

A sensitive and specific high-performance liquid chromatographic metho d was developed and validated for the determination of the third-gener ation retinoid (arotinoid) mofarotene (Ro 40-8757) in human, rat and d og plasma, using direct injection of deproteinated plasma samples, aut omated column switching (on-line solid-phase extraction) and ultraviol et detection. Plasma (0.5 ml) was deproteinated by adding ethanol (1 m l) containing the internal standard Ro 42-8659 (200 ng/ml). After cent rifugation, 019 ml of the supernatant were directly injected onto a pr ecolumn packed with C-18 Corasil 37-50 mu m. Polar plasma components w ere washed out from the precolumn using 1% ammonium acetate-acetic aci d-acetonitrile (900:9:100, v/v/v). After valve switching, the pre-conc entrated compounds were transferred to the analytical column (C-18) in the backflush mode, separated by gradient elution and detected at 300 nm. The retention times (total run times) were approximately 15 and 2 0 min for the internal standard and mofarotene, respectively. The meth od was linear in the range 10-1000 ng/ml with a limit of quantificatio n of 10 ng/ml. The mean recoveries were 80.4%, 81.7% and 77.8% (range 10-1000 ng/ml) and the inter-assay precision was 2.7% (range 20-1000 n g/ml), 1.5% and 2.0% (both range 100-1000 ng/ml) for human, rat and do g plasma, respectively. Mofarotene was found to be stable in human, ra t and dog plasma stored at -20 degrees C for 3 months and at 22 degree s C for 24 h. The method was successfully applied to clinical, pharmac okinetic and toxicokinetic studies.