Simultaneous determination of triclabendazole and its sulphoxide and sulphone metabolites in bovine milk by high-performance liquid chromatography

A simple method has been developed for the simultaneous determination of triclabendazole and its metabolites (sulphoxide and sulphone) in bovine milk by reversed-phase high-performance liquid chromatography (HPLC). A milk sample was homogenized with sodium sulfate anhydrous and acetonitrile, and centrifuged. The supernatant was isolated, rinsed with n-hexane saturated with acetonitrile, and evaporated. The residue was dissolved with 0.1 M potassium dihydrogenphosphate, and 0.1 M sodium hydrogencarbonate, and then cleaned up on a Bond Elut C18 cartridge. The three compounds were separated on a Capcell Pak C18 UG 120 (5 μm, 150×4.6 mm I.D.) column and determined by UV detection at 295 nm. The mobile phase was acetonitrile–0.05 M ammonium acetate (50:50), and the flow-rate was 0.8 ml/min at 40°C. The mean recoveries (n=4) were 89.1–95.0% with a relative standard deviation of 1.1–2.6%. The detection limits were 0.004–0.006 μg/g in milk. The proposed method was used to monitor raw milk samples for the market, and applied to the analysis of milk samples from 10 cows which had been administered with triclabendazole to control the liver fluke. The confirmation of the triclabendazole and its metabolites in the above milk sample has been achieved by electrospray LC–MS for the first time.