A small-scale method of sample preparation suitable for simultaneous HPLC-UV assay of capecitabine and its 5'-DFCR metabolite in mouse blood plasma

ABSTRACT

Abstract The objective of the study was to develop an easy, cheap, effective, and safe, small-scale method for sample preparation suitable for the simultaneous high-performance liquid chromatography (HPLC)-ultraviolet (UV) assay of capecitabine and its 5′-deoxy-5-fluorocytidine (5′-DFCR) metabolite in mouse blood plasma. The suitability of the proposed method of sample preparation was verified by the optimal effectiveness and efficiency achieved in the overall analytical workflow. The chromatographic separation of capecitabine and its first metabolite was performed on a Hypersil GOLD aQ column with a mobile phase consisting of 1% formic acid, methanol, and water, and run in a gradient elution mode. The absence of interfering endogenous components at the retention times of each analyte was confirmed by the chromatographic analysis of blank matrices and matrices spiked with the corresponding standards. The absence of any tactile matrix effect was also recorded. For the first time, the effect of the vacutainer's anticoagulant on the extraction efficiency of both analytes was evaluated. The method was found to be accurate, precise, and specific. The estimated mean "extraction" efficiencies were ≥90% for each analyte. The lower limit of quantitation for both capecitabine and 5′-DFCR was 0.05 μg/mL.