The existence of antibiotic residues in edible products constitutes a health problem to the consumers. Reversed-phase high-performance liquid chromatography with diode array detection (HPLC–DAD) was optimized and validated for the simultaneous determination of florfenicol (FF), penicillin (PE), and tetracycline (TC) residues in dairy raw milk samples. The determination of these antibiotics was carried out on HP-ODS Hypersil C18 (5μm, 125*4 mm) column at a flow rate (1mL/min) and temperature (35 ⁰C). The extraction method includes deproteinization of the milk sample followed by a solid-phase extraction (SPE) clean-up. The method was validated according to the European Commission Decision 2002/657/EC and the International Conference of Harmonization Guidelines. The recoveries for the studied antibiotics ranged from 82–111.54 % making the method suitable for performing routine analysis. The proposed method was applied for the analysis of antibiotic residues in 50 dairy raw milk samples collected from many regions in Lebanon. The results showed the occurrence of these antibiotics residues in milk collected from different Lebanese regions. The numbers indicate that 22 % of milk samples were found to be positive for FF, 42 % for PE, and 28 % for TC residues.
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