Abstract:
A selective, sensitive and high throughput liquid chromatography–tandem mass spectrometry (LC–MS/MS) method has been developed and validated for the chromatographic separation and quantitation of diastereomers of S002-333, a novel anti-thrombotic agent in rabbit plasma. Sample clean-up involved liquid–liquid extraction (LLE) of both the isomers and internal standard (β-carbolinamide) from 200µl of rabbit plasma. Both the analytes were chromatographically separated on a chiralcell OJ-RH column (150 ×4.6mm, 5µ particle size) using a gradient flow program comprising of 0.1% formic acid, methanol and acetonitrile as the mobile phase. The parent→product ion transitions (MRM) for both the isomers and IS were 386.4 →214.2 m/z and 216.1→144.2 m/z respectively, were monitored on a triple quadrupole mass spectrometer, operating in positive ion mode. The MS/MS response was linear over the concentration range from 1.56 ng/mL to 400 ng/ml, with a lower limit of detection (LOD) was 1.56 ng/ml. The intra- and inter-day precisions (% R.S.D.) between 3.96 to 13.80 for both analytes and the accuracies (% bias) were between -4.05 to 5.93. The validated method can be used in most or all stages of the screening and optimizing process for pharmacokinetic and toxicokinetics studies.
