Development and Validation of a LC-MS/MS Analytical Method for Quantification of Paclitaxel and Everolimus in Human Plasma: Application to Pharmacokinetic Studies

This study presents the development and validation of a sensitive and robust liquid chromatography-tandem mass spectrometry (LC-MS/MS) method for the simultaneous quantification of Paclitaxel and Everolimus in human plasma. Analytical method validation followed the International Conference on Harmonization (ICH) and European Medicines Evaluation Agency (EMEA) guidelines. Key validation parameters, including specificity, carry-over, lower limit of quantification (LLOQ), calibration curve linearity, accuracy, precision, matrix effect, and stability, were comprehensively evaluated. The method employed an Agilent Zorbax SB-C۱۸ column and a mobile phase of ۰.۳% formic acid in water and methanol under gradient conditions, achieving optimal separation at a flow rate of ۰.۵ mL/min. Mass spectrometry detection utilized a Waters Quattro Micro with electrospray ionization (ESI) in positive ion mode. The LLOQ for Paclitaxel was determined to be ۶.۲۵ ng/mL with a signal-to-noise ratio exceeding ۱۰, ensuring high sensitivity. Calibration curves demonstrated excellent linearity (R۲> ۰.۹۹) across a concentration range of ۶.۲۵-۴۰,۰۰۰ ng/mL for Paclitaxel and ۴۰-۴,۰۰۰ ng/mL for Everolimus. The intra- and inter-day precision and accuracy were within the acceptable limits of ≤۱۵% for all quality control levels (LQC, MQC, HQC). Matrix effect studies indicated minimal interference, with matrix effect factors (MEF) below ۱۵%. Short-term, freeze-thaw, and long-term stability tests confirmed the stability of analytes under experimental conditions. This validated method was successfully applied to pharmacokinetic studies involving volunteers administered Paclitaxel and Everolimus. The findings underscore the method's robustness and reliability for therapeutic drug monitoring and pharmacokinetic profiling. This work contributes significantly to advancing personalized medicine approaches for drugs with narrow therapeutic windows, such as Paclitaxel and Everolimus.

کلیدواژه ها:

LC-MS/MS ، Paclitaxel ، Everolimus ، Analytical Method Validation ، Human Plasma ، Pharmacokinetics ، Therapeutic Drug Monitoring ، Matrix Effect ، Stability Studies ، ICH Guidelines