Abstract:

A high performance liquid chromatographic method with fluorescence detection for the determination of chloramphenicol (CAP) residues in milk was developed. Although CAP itself is non-fluorescent the aromatic nitro group of CAP could be reduced to aromatic primary amino group and fluorescamine can be used as a selective reagent for primary amines. Therefore, the CAP was derivatized with fluorescamine prior to injection in this project. The HPLC method was performed on a Diamond C18 column (250 mm×4.6 mm i.d., 4.0 μm) with the mobile phase composed of sodium acetate buffer (0.02 mol/L, pH 6.0)-acetonitrile-tetrahydrofuran (76∶16∶8, v/v/v). The flow rate of the mobile phase was set at 1.0 mL/min, and the column was maintained at 40 ℃. Analytes were detected by a fluorescence detector at 410 nm excitation and 508 nm emission wavelength. The standard curve was linear in the range from 0.4 μg/L to 800 μg/L. The limit of detection (LOD) was 0.2 μg/L, and the limit of quantitation (LOQ) was 0.4 μg/L. Overall recoveries were between 66.6% and 92.8% with relative standard deviations between 4.5% and 9.4%. The procedure provides a rapid, reliable and sensitive method for the determination of CAP in milk.

Key words: chloramphenicol, fluorescamine, milk , 　pre-column derivatization, residues, high performance liquid chromatography-fluorescence detection