ArticleDrug design, development and therapy2026
Simultaneous Determination of Fexinidazole and Its Major Metabolites in Rat Plasma by UPLC-MS/MS: Method Validation and Application to Pharmacokinetic Study.
Article in Drug design, development and therapy, 2026. The graph could read no effect estimate from its abstract, so it casts no vote on the map. Not yet cited in PubMed.
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Abstract
Introduction: Fexinidazole is an oral antitrypanosomal drug approved for the treatment of human African trypanosomiasis (HAT). Its therapeutic efficacy primarily depends on the in vivo formation of its active metabolites, sulfoxide fexinidazole (M1) and sulfone fexinidazole (M2). Accurate and sensitive quantification of fexinidazole and its major metabolites is essential for pharmacokinetic evaluation and preclinical research. Therefore, a reliable bioanalytical method for their simultaneous determination in plasma is required. Methods: An ultra-performance liquid chromatography-tandem mass spectrometry (UPLC-MS/MS) method was developed and validated for the simultaneous determination of fexinidazole, M1, and M2 in rat plasma, using fluconazole as the internal standard (IS). Chromatographic separation was achieved on an Acquity UPLC BEH C18 column with gradient elution consisting of 0.1% formic acid in water and acetonitrile. The method was validated in accordance with bioanalytical guidelines, including assessments of selectivity, linearity, lower limit of quantification (LLOQ), precision, accuracy, recovery, matrix effects, and stability. The validated method was subsequently applied to a pharmacokinetic study in rats. Results: The method demonstrated good linearity within the investigated concentration ranges for all analytes. The LLOQs were 1 ng/mL for fexinidazole, 10 ng/mL for M1, and 50 ng/mL for M2. Intra-day and inter-day precision (RSD%) and accuracy (RE%) were within ±15%. Recovery was consistent and reproducible, matrix effects were acceptable, and stability under various conditions met bioanalytical requirements. The method was successfully applied to the pharmacokinetic evaluation of fexinidazole and its metabolites in rats. Discussion: The developed UPLC-MS/MS method exhibited satisfactory sensitivity, accuracy, and reproducibility for the quantification of fexinidazole and its active metabolites in rat plasma. This method provides reliable analytical support for pharmacokinetic studies and contributes to the preclinical investigation of fexinidazole.
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