Development and Analytical Validation of a RP-HPLC–LC-MS/MS Method for Simultaneous Quantification of Fexofenadine and Losartan in Plasma: Application to Bioequivalence Studies

Abstract

This study presents the development and validation of a highly sensitive and selective analytical method for the simultaneous quantification of fexofenadine (FEX) and losartan (LOS) in human plasma, using reverse-phase high-performance liquid chromatography (RP-HPLC) coupled with liquid chromatography-tandem mass spectrometry (LC-MS/MS). The method was optimized for precise quantification, ensuring high specificity and reliability under stringent conditions. Calibration curves were constructed for FEX and LOS in plasma samples with concentration ranges from 1.575 ppb to 2000 ppb. A series of tests for specificity, linearity, accuracy, precision, and matrix effects were performed in accordance with ICH M10 guidelines, ensuring robustness and reproducibility. The method demonstrated minimal interference, as shown by the specific retention times of FEX and LOS in plasma, even in the presence of complex biological matrices. Calibration was achieved through weighted linear regression (1/X), with calculated regression coefficients consistently greater than 0.99. Method validation included assessment of the lower limit of quantification (LLOQ) and intra- and inter-day precision, which were found to be within acceptable limits. The matrix effects were minimal, indicating a high degree of method specificity. Additionally, the method was applied to bioequivalence studies, demonstrating its suitability for clinical pharmacokinetic applications. The proposed analytical method provides a reliable and efficient approach for quantifying FEX and LOS in plasma, with significant potential for routine pharmacokinetic and bioequivalence studies.