A simplified procedure for simultaneous quantification of ceftiofur (CEF), fluoroquinolone (FQ) and sulfonamide (SA) antibacterials in bovine milk was developed. The reverse-phase liquid chromatography (RP-LC) multiclass method for analysis of eleven distinct compounds, from three antibacterial classes, was validated in line with Commission Decision 2002/657/EC. Confirmation of the analytes identities was performed by electrospray mass spectrometry detection. The analytes were extracted from milk matrix by liquid-liquid extraction with acidified ultrapure water and directly analyzed in the chromatograph. The SA compounds were pre-column derivatized with fluorescamine for fluorescence detection. The method provided good results regarding the analytical parameters of linearity, selectivity, sensitivity, precision, recovery, decision limit (CCα), detection capability (CCβ), limit of detection (LOD), limit of quantification (LOQ), stability and robustness. Analytes were extracted by liquid-liquid extraction in the fortified matrix and the compounds identity was confirmed by their precursor ion and fragments through tandem mass spectrometry analysis. Additionally, milk samples from two state capitals in the South Region of Brazil were analyzed by both the quantitative and confirmatory methods. The validation process showed correlation coefficients (r(2)) greater than 0.98 for all the analytes, with recovery rates up to 98% for all the studied drugs. LOD and LOQ limits ranged from 8.0 to 20.0 ng mL(-1) and 10.0 to 32.0 ng mL(-1), demonstrating good specificity of the method. The intra-day and inter-day precisions for all the analytes were below or equal to 7.40 and 10.13, respectively. The studied antibacterials were not detected in milk samples. The developed method represents an efficient alternative for multi-residue analysis in milk, being suitable and especially viable for monitoring in developing countries.
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