Bioequivalence and Analytical Validation of Linagliptin: A Comprehensive Study Using LC-MS/MS in Human Plasma

Abstract

This study provides a detailed bioequivalence analysis and analytical validation of Linagliptin 5 mg using human plasma samples. Employing a robust LC-MS/MS method with electrospray ionization (ESI) in positive ion mode, the study adheres to stringent guidelines to ensure reliability and reproducibility. The method validation followed the European Medicines Evaluation Agency (EMEA) and ICH M10 guidelines, encompassing parameters such as specificity, carry-over, lower limit of quantification (LLOQ), calibration, accuracy, precision, matrix effects, and stability. The LLOQ was determined at 0.25 ppb, with a signal-to-noise ratio exceeding 10. The calibration curve displayed linearity across 0.25 to 16 ppb with weighted regression. Intra- and inter-day accuracy and precision tests confirmed deviations below 5.8%. Matrix effect evaluations using plasma from six different donors demonstrated consistent results with RSDs below 12.5%. Stability tests under various conditions, including short-term, freeze-thaw, and long-term, revealed no significant degradation, supporting the robustness of the assay. Additionally, pharmacokinetic profiles from human volunteers were analyzed, confirming bioequivalence between Linagliptin and the reference formulation. These findings highlight the method’s applicability for regulatory compliance and its potential for clinical and pharmaceutical research.