Analytical method development and validation for the determination of triazole antifungals in biological matrices

The World Health Organization released the fungal priority pathogen list (FPPL) in late 2022, identifying fungal pathogens in urgent need of research and intervention. Triazole antifungals, as first-line therapy, require therapeutic drug monitoring to optimize outcomes because of their narrow therapeutic range and interindividual pharmacokinetic variability. The bioanalysis process is challenging due to complex matrices. This study aimed to develop and validate an analytical method for quantifying voriconazole, itraconazole, and fluconazole in human plasma using high-performance liquid chromatography with an imprinted polymer sorbent. Optimization employed a C 18 column with isocratic elution (acetonitrile:water 70:30, v/v) at a flow rate of 1 mL/min and detection at 260 nm. Retention times were 2.467, 3.487, and 9.006 minutes for fluconazole, voriconazole, and itraconazole, respectively. System suitability testing met chromatographic requirements. The validated bioanalytical method complies with U.S. FDA guidelines, demonstrating linearity over the 0.5–5 mg/L range. Maximum percent bias was 13.4% for the calibration curve, 15.6% at the LLOQ, and 12.9% above the LLOQ. Precision showed a maximum coefficient of variation of 12.9%, confirming the method’s accuracy and reproducibility. The molecularly imprinted solid-phase extraction (MISPE) method showed superior selectivity and recovery compared with commercial C 18 SPE cartridges. This study establishes a reliable method for future clinical sample applications.