HPLC assay for albendazole and metabolites in human plasma for clinical pharmacokinetic studies

A sensitive and selective HPLC chromatography method using UV detection (295 nm) was developed for the determination of albendazole, albendazole sulfoxide (ABZSO), and albendazole sulfone (ABZSO2) in human plasma. Albendazole, ABZSO, ABZSO2, and the internal standard, oxibendazole, were extracted from human plasma by loading onto a conditioned C₁₈ SPE cartridge, rinsing with 15% methanol, and eluting with 90% methanol. Samples were evaporated under a stream of nitrogen, reconstituted with mobile phase, 1.25% triethylamine in water-methanol-acetonitrile (72:15:13, v/v/v) (pH* 3.1), and injected onto a Waters μBondapak™ Phenyl 3.9×300 mm HPLC column. Mobile phase flow rate was 1.0 ml/min. The retention times of albendazole, ABZSO, ABZSO2, and the internal standard were approximately 24.4, 7.9, 13.4, and 11.3 min, respectively. Total run time was 30 min. The assay was linear for concentration ranges in human plasma of 20-600 ng/ml for albendazole, 20-1000 ng/ml for ABZSO, and 20-300 ng/ml for ABZSO2. The analysis of quality control samples demonstrated excellent precision. Coefficients of variation for albendazole (20, 400, 600 ng/ml) were 6.7, 8.1 and 7.0%; ABZSO (20, 400, 800 ng/ml) were 6.0, 8.5 and 5.9%; ABZSO2 (20, 150, 300 ng/ml) were 3.1, 3.9 and 2.3%, respectively. The method appears to be robust and has been applied to a pharmacokinetic study of albendazole in healthy volunteers.