Quantification and pharmacokinetic study of entrectinib in rat plasma using ultra‐performance liquid chromatography tandem mass spectrometry

: To characterize the preclinical plasma pharmacokinetics of entrectinib, a reproducible and precise assay is necessary. In this study, we developed and validated a simple ultra-performance liquid chromatography tandem mass spectrometry (UPLC-MS/MS) method for the measurement of entrectinib using carbamazepine as the internal standard in rat plasma. Sample preparation was a simple protein precipitation with acetonitrile, then entrectinib was eluted on an Acquity UPLC BEH C18 column (2.1 × 50 mm, 1.7 μm) using a gradient elution with a mobile phase composed of acetonitrile (A) and 0.1% formic acid in water (B). Detection was achieved using multiple-reaction monitoring in positive ion electrospray ionization mode. The method showed good linearity over the concentration range of 1-250 ng/mL (r2  > 0.9951). The intra- and inter-day precision was determined with the values of 6.3-12.9 and 2.6-6.9%, respectively, and accuracy values of 0.5-11.6%. Matrix effect, extraction recovery, and stability data all met the acceptance criteria of US Food and Drug Administration guidelines for bioanalytical method validation. The method was successfully applied to a pharmacokinetic study. In this study, we developed the complete validated method for the quantification of entrectinib in rat plasma.