Quantitative determination of regorafenib and its two major metabolites in human plasma with high‐performance liquid chromatography and ultraviolet detection

A simple, highly sensitive and specific high‐performance liquid chromatography (HPLC) method was developed for the simultaneous quantitation of regorafenib, N‐oxidemetabolite (M‐2) and the desmethyl N‐oxide metabolite (M‐5) in human plasma. Regorafenib, M‐2, M‐5 and the internal standard sorafenib were separated using a mobile phase of 0.5% KH₂PO₄ (pH 3.5)–acetonitrile (30:70, v/v), on a Capcell PAK MG II column at a flow rate of 0.5 mL/min and measurement at UV 260 nm. The lower limits of quantification for regorafenib, M‐2 and M‐5 were 10 ng/mL for each analyte. A procedure using solid‐phase extraction required only a small amount of plasma (100 μL) for one analysis while providing high extraction recovery (>81% for all compounds) and good selectivity. Coefficients of variation for intra‐ and inter‐day assays were <12.2% for regorafenib, <12.3% for M‐2 and <15.1% for M‐5. Accuracies for intra‐ and inter‐day assays were <9.4% for regorafenib, <8.0% for M‐2 and <12.8% for M‐5 over a linear range from 10 to 10,000 ng/mL. This HPLC assay is suitable for clinical pharmacokinetic studies of regorafenib. The present HPLC method is currently in use for our observational studies of patients under treatment. Copyright © 2016 John Wiley & Sons, Ltd.