Abstract: In order to enhance the fluorescence intensity of 7 β-agonists, the fluorescent derivatizer 2-(4-sulfoyl chloro-phenyl) benzimidazole reacted with β-agonists, a high performance liquid chromatography-fluorescence detection method for 7 β-agonists in animal-derived food was established. Effects of pH of carbonate buffer solution, reaction temperature, reaction time and concentration of 2-(4-sulfoyl chloro-phenyl) benzimidazole on the fluorescence derivatization were investigated. The results showed that 7 β-agonists had a good linearity in the range of 1.0~100.0 ng/mL and the correlation coefficient (R²) was higher than 0.999 under the optimal conditions. The limits of detection for 7 β-agonists were about 0.30 μg/kg, and the limits of quantitation were less than 1.0 μg/kg. The recoveries of blank pork, mutton and beef samples at three different concentrations of 1.0, 10.0 and 50.0 μg/kg were 81.3%~107.3%, 80.5%~110.7% and 80.2%~105.1%, the intra-day and the inter-day relative standard deviations were 1.48%~13.51%, 1.26%~12.33%, 1.23%~11.49% and 2.21%~11.54%, 1.82%~13.31%, 1.07%~13.21%, respectively. Therefore, the developed method is suitable for determination of 7 β-agonists in animal-derived food qualitatively and quantitatively with satisfactory sensitivity, selectivity, accuracy and precision.