Development and validation of an HPLC confirmatory method for residue analysis of ten quinolones in tissues of various food-producing animales according to the European Union Decision 2002/657/EC

The aim of this work was to develop an HPLC method for the simultaneous determination of ten quinolones: enoxacin, ofloxacin, norfloxacin, ciprofloxacin, danofloxacin, enrofloxacin, sarafloxacin, oxolinic acid, nalidixic acid, and flumequine, in various tissues of food-producing animals. Separation was achieved on a PerfectSil(R) Target column (250 mm x 4 mm, ODS-3, 5 mu m), by MZ-Analysentechnik (Germany), at room temperature. The mobile phase consisted of 0.1% TFA-CH3OH-CH3CN and was delivered by a gradient program of 35 min. The detection and quantitation was performed on a photodiode array detector at 275 and 255 nm. Caffeine (7.5 ng/mu L) was used as the internal standard (IS). Analytes were isolated from tissue samples by 0.1% methanolic TFA solution. SPE, using LiChrolut RP-18 cartridges, was applied for further purification. The extraction protocol was optimized and the final recoveries varied between 92.0 and 107.4%. The method was fully validated according to Commission Decision 2002/657/EC. Limits of quantitation for the examined quinolones extracted from each tissue were much lower than the respective Maximum Residue Levels, ranging between 30 and 50 mu g/kg for bovine tissue, between 30 and 55 mu g/kg for ovine tissue, and between 40 and 50 mu g/kg for porcine tissue.