Bioanalytical Method Validation and Bioequivalence Study of Ibrutinib in Human Plasma Using LC-MS/MS: An Advanced Approach

Abstract

This study presents a comprehensive bioanalytical method validation and bioequivalence analysis for ibrutinib, a Bruton’s tyrosine kinase inhibitor, in human plasma samples. The method employs liquid chromatography-tandem mass spectrometry (LC-MS/MS) for quantification, utilizing apixaban as an internal standard. Method validation adhered to ICH M10 guidelines, assessing parameters such as specificity, carry-over, lower limit of quantitation (LLOQ), calibration curve linearity, accuracy, precision, matrix effects, and stability under various conditions. Calibration curves exhibited strong linearity across a range of 0.5–48 ppb (R² > 0.99), with an LLOQ of 0.5 ppb demonstrating a signal-to-noise ratio exceeding 10. The method demonstrated robust specificity with minimal interference, as well as high precision and accuracy, with intra- and inter-day deviations within acceptable limits (<15%). Stability assessments, including freeze-thaw, short-term, and long-term analyses, confirmed analyte integrity under varied conditions. The bioequivalence study compared test and reference formulations of ibrutinib in 26 healthy volunteers over multiple time points, with pharmacokinetic parameters including C_max and AUC analyzed for equivalence. Results affirmed bioequivalence between formulations, meeting regulatory criteria. This validated method provides a reliable tool for therapeutic drug monitoring and pharmacokinetic studies of ibrutinib, contributing to optimized clinical outcomes and regulatory compliance.