Determination of flubendazole and its metabolites in eggs and poultry muscle with liquid chromatography-tandem mass spectrometry

The optimization of a quantitative and sensitive LC-MS/MS method to determine flubendazole and its hydrolyzed and reduced metabolites in eggs and poultry muscle is described. The benzimidazole components were extracted from the two matrices with ethyl acetate after the sample mixtures had been made alkaline. The HPLC separation was performed on an RP C-18 column with gradient elution, using ammonium acetate and acetonitrile as mobile phase. The analytes were detected after atmospheric pressure electrospray ionization on a tandem quadrupole mass spectrometer in MS/MS mode. The components were measured by the MS/MS transition of the molecular ion to the most abundant daughter ion. The overall extraction recovery values for flubendazole, the hydrolyzed metabolite, and the reduced metabolite in eggs (fortification levels of 200, 400, and 800 microg kg(-1)) and muscle (fortification levels of 25, 50, and 100 microg kg(-1)) were, respectively, 77, 78, and 80% and 92, 95, and 90%. The trueness (fortification levels of 400 and 50 microg kg(-1), respectively, for eggs and muscle), expressed as a percentage of the added values for these analytes, was, respectively, 89, 100, and 86 and 110, 110, and 98%. The proposed MS detection method operating in the MS/MS mode is very selective and very sensitive. The limits of detection for flubendazole and its hydrolyzed and reduced metabolites in egg and muscle were, respectively, 0.19, 0.29, and 1.14 microg kg(-1) and 0.14, 0.75, and 0.31 microg kg(-1). The limits of quantification were, respectively, 1, 1, and 2 microg kg(-1) and 1, 1, and 1 microg kg(-1). The discussed method was applied to a pharmacokinetic study with turkeys. Residue concentrations in breast and thigh muscle of turkeys orally treated with flubendazole were quantified. Medicated feed containing 19.9 and 29.6 mg kg(-1) flubendazole was provided to the turkeys for seven consecutive days. For the trial with the recommended dose of 19.9 mg kg(-1), one day after the end of the treatment, the mean sum of the flubendazole plus hydrolyzed metabolite residue values in thigh and breast muscle declined to below the maximum residue limit (50 microg kg(-1)) and were, respectively, 36.6 and 54.1 microg kg(-1). The corresponding values with the higher dose of 29.6 mg kg(-1) were, respectively, 101.7 and 119.7 microg kg(-1).