Evaluation of the effect of apatinib (YN968D1) on cytochrome P450 enzymes with cocktail probe drugs in rats by UPLC-MS/MS

An accurate and validated liquid chromatography method and a triple quadrupole mass spectrometry method were developed and validated to simultaneously evaluate the cytochrome P450 (CYP) enzymes in vivo using the co-administration of these probes. Phenacetin, losartan, metoprolol and midazolam were used as the probe substrates for rat CYP1A2, CYP2C11, CYP2D4 and CYP3A1 enzymes, respectively. The purpose of the study was to investigate the effect of apatinib on these cytochrome P450 enzymes in vivo with co-administration of these probes. Plasma samples were prepared by precipitating protein with acetonitrile. The analytes were separated using a reversed-phase BEH C18 column (2.1 mm × 100 mm, 1.7 μm, Waters, USA) maintained at 40 °C. The mobile phase consisted of acetonitrile and water (containing 0.1% formic acid) with a gradient elution pumped at a flow rate of 0.4 mL/min. The analytes were detected with positive electrospray ionization in multiple reaction monitoring (MRM) mode for target fragment ions m/z 180.05 → 109.94 for phenacetin, m/z 423.1 → 207.2 for losartan, m/z 268.12 → 115.8 for metoprolol, m/z 326.02 → 290.99 for midazolam and m/z 285.1 → 193.1 for diazepam (IS). Good linearity was achieved to quantify the concentration ranges of 10-2000 ng/mL for phenacetin, 10-1000 ng/mL for losartan, 10-1000 ng/mL for metoprolol and 1-100 ng/mL for midazolam in rat plasma. The mean recoveries of phenacetin, losartan, midazolam and metoprolol from the plasma exceeded 77.07%. The intra-run and inter-run assay precisions were both less than 8.9%. This method was successfully applied to evaluate the effects of apatinib on the cytochrome P450 enzymes in rats.