Development and validation of a selective and robust LC-MS/MS method for high-throughput quantifying rizatriptan in small plasma samples: application to a clinical pharmacokinetic study.

An analytical method based on liquid chromatography with positive ion electrospray ionization (ESI) coupled to tandem mass spectrometry detection (LC-MS/MS) was developed for the determination of a potent 5-HT(1B/1D) receptor agonist, rizatriptan in human plasma using granisetron as the internal standard. The analyte and internal standard were isolated from 100 microL plasma samples by liquid-liquid extraction (LLE) and chromatographed on a Lichrospher C18 column (4.6mm x 50mm, 5 microm) with a mobile phase consisting of acetonitrile-10mM aqueous ammonium acetate-acetic acid (50:50:0.5, v/v/v) pumped at 1.0 mL/min. The method had a chromatographic total run time of 2 min. A Varian 1200 L electrospray tandem mass spectrometer equipped with an electrospray ionization source was operated in selected reaction monitoring (SRM) mode with the precursor-to-product ion transitions m/z 270-->201 (rizatriptan) and 313.4-->138 (granisetron) used for quantitation. The assay was validated over the concentration range of 0.05-50 ng/mL and was found to have acceptable accuracy, precision, linearity, and selectivity. The mean extraction recovery from spiked plasma samples was above 98%. The intra-day accuracy of the assay was within 12% of nominal and intra-day precision was better than 13% C.V. Following a 10mg dose of the compound administered to human subjects, mean concentrations of rizatriptan ranged from 0.2 to 70.6 ng/mL in plasma samples collected up to 24h after dosing. Inter-day accuracy and precision results for quality control samples run over a 5-day period alongside clinical samples showed mean accuracies of within 12% of nominal and precision better than 9.5% C.V.