Fluoroquinolone antibiotic determination in bovine milk using capillary liquid chromatography with diode array and mass spectrometry detection

A fast and sensitive method has been developed for the determination of five fluoroquinolones and a quinolone in commercial bovine milk by capillary-LC–DAD–MS after simple extraction method based on milk deproteinization with 15% trichloroacetic acid. Separation was carried out in a Zorbax SB-C18 column (150 mm × 0.5 mm, 5 μm) using a mixture of acetonitrile–5 mM ammonium formate (pH 3.7) as mobile phase in gradient elution mode. The sample was prepared in a 5 mM aqueous ammonium formate buffer for focusing purposes and 20 μL were injected. Flow rate and temperature were set at 20 μL min−1 and 25 °C, respectively. Method validation was performed according to the European Commission Decision 657/2002/EC. Sample detection limits were between 2.8 and 25 μg kg−1, and good linearity was observed up to 250 μg kg−1 for all analytes. Acceptable and constant recoveries of 96% for enrofloxacin, 77% for its metabolite ciprofloxacin, and 64% for flumequine, with RSDs (n = 4) lower than 8%, were obtained at the 100 μg kg−1 maximum level permitted in milk. Recoveries between 70% and 83% were obtained when difloxacin, sarafloxacin and ofloxacin (not allowed in animals that produce milk for human consumption) were added to milk samples at levels in the range 75–150 μg kg−1. Decision limit (CCα), detection capability (CCβ), repeatability, within-laboratory reproducibility and robustness were compliant with European Union regulations.