Determination of BMS-186318 in dog, rat and monkey plasma by liquid chromatography ionspray mass spectrometry

EMS-186318 is a member of the recently discovered ''aminodiol'' class of HIV protease inhibitors. A simple but sensitive method was developed for the determination of EMS-186318 in dog plasma and then applied to monkey and rat plasma. The compound was extracted from dog plasma with methyl tert-butyl ether at basic pH. The dried extract was reconstituted in mobile phase and injected into a 150 x 2.1 mm i.d. Zorbax Rx-C-18 HPLC column. A portion of the effluent was directed into the LC-ionspray MS system, where the [M + H](+) ion of the secondary amine compound was monitored. The HPLC conditions were chosen in order to achieve a short run time and large sample throughput, with both analyte and internal standard eluting within 1.5 min. The liquid-liquid extraction procedure provided very clean extracts so that sufficient signal-to-noise ratio was obtained with single-stage mass spectrometry instead of the more costly tandem mass spectrometry. The required lower limit of quantitation of 2.5 ng ml(-1) was easily achieved. The method has also been validated for EMS-186318 in monkey plasma without modification. The method has been modified for rat plasma. Owing to irreproducibility observed when applying the liquid-liquid extraction method to rat plasma, a solid-phase extraction method was developed. The addition of phenylmethylsulfonyl fluoride was necessary to stabilize EMS-186318 in rat blood and plasma.