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A sensitive, specific, and reliable liquid chromatography tandem mass spectrometry (LC-MS/MS) method was developed for detection and identification of zeranol in chicken or rabbit liver. A homogenized liver sample was hydrolyzed with β-glucuronidase/arylsulfatase, and the hydrolysate was extracted with ethyl ether. The supernatant was evaporated to dryness, and the residue was dissolved in chloroform and re-extracted with sodium hydroxide. After acidification, the extract was cleaned up on a C18 solid-phase extraction cartridge and analyzed by electrospray LC-MS/MS in the negative ion mode. The multiple reaction monitoring transition from both m/z 321 to 277 and m/z 321 to 303 was monitored for confirmation, and the product ion of 277 was used for quantitation. Separation was performed on a Waters XTettra™ C18 column (50 × 2.1 mm, 3.5 μm) combined with a safeguard column (Symmetry C18, 20 × 3.9 mm, 5 μm), using a gradient elution with acetonitrile and 20mM ammonium acetate. Calibration curves were prepared and good linearity was achieved over the concentration ranges tested. For all liver samples fortified at 3 different levels of 1, 5, and 50 μg/kg, the overall recoveries and relative standard deviations were in the range of 61–90 and 8–13%, respectively. The limit of quantitation based on the assay validation was 1 μg/kg. The method had been used on a routine basis for detection and identification of zeranol in liver samples.

Detection and Identification of Zeranol in Chicken or Rabbit Liver by Liquid Chromatography-Electrospray Tandem Mass Spectrometry