Antibiotic drug residues are crucial to ensure food safety and minimize risk to human health. Herein, a sensitive high-performance liquid chromatography-tandem mass spectrometry (HPLC-MS/MS) method was developed and validated for the determination of antibiotic residues (mainly amphenicols) consisting of chloramphenicol (CAP), thiamphenicol (TAP), florfenicol (FF), and florfenicol amine (FFA) in aquatic products. Amphenicols were well separated on a Kinetex F5 (100 mm × 3.0 mm, 2.6 µm) chromatographic column with the mobile phases of 1 mM ammonium acetate aqueous solution and methanol solution and measured after positive and negative electrospray ionizations using four internal standards. To our knowledge, it was the first time to report the good performance of F5 column and four internal standards for the determination of amphenicols. The established method featured a good linear relationship between chromatographic peak area ratios and the concentrations of amphenicols (R2 > 0.992), a wide and low detection matrix-based range of 0.01-5 μg/L, a low detection limit of 0.01 μg/kg, etc. The spiked assays evidenced the accuracy and reliability of the developed method with the recoveries between 84.0 and 105 %, the intraday relative standard deviations (RSDs) over the range of 0.769-13.7 %, and the interday RSDs over the range of 0.582-13.3 %. Finally, the proposed method was applied to investigate amphenicol residues in various aquatic products, including fish, shrimp, crab, shellfish, and other aquatic species.

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