Simultaneous Determination of Thioridazine and Its Metabolites by UHPLC–MS/MS With Application to a Pharmacokinetic Study in Rats

ABSTRACT To develop an LC–MS/MS method for simultaneous quantification of thioridazine and its metabolites in rat plasma, samples were prepared by protein precipitation and filtration using thioridazine‐d3 as an internal standard. Chromatographic separation was achieved on a reversed‐phase C18 UPLC column with 0.1% formic acid in acetonitrile and 10‐mM ammonium acetate in water at 0.7 mL/min. Quantification was performed using multiple reaction monitoring in positive ion mode with the following precursor–product ion transitions: m/z 371.1 → 126.1 (thioridazine), m/z 387.1 → 126.1 (mesoridazine), m/z 403.1 → 126.1 (thioridazine‐2‐sulfone), m/z 387.2 → 126.1 (thioridazine‐5‐sulfoxide), and 374.1 → 129.1 (thioridazine‐d3). This method showed good linearity: 0.1–1000 ng/mL for thioridazine ( r 2 = 0.9980) and 0.5–1000 ng/mL for mesoridazine ( r 2 = 0.9981), thioridazine‐2‐sulfone ( r 2 = 0.9976), and thioridazine‐5‐sulfoxide ( r 2 = 0.9967). The accuracy of the LLOQ and QC samples for thioridazine and its metabolites was within 85%–115%, with intrabatch and interbatch precisions (CV%) below 15%. The average recovery ranged from 98% to 109%, and no significant matrix effect was observed, with values ranging from 93% to 110%. All analytes were stable under the conditions used in this study. This method was applied in a pharmacokinetic study following the intravenous administration of thioridazine to rats.