Abstract: AIM　To develop a new determination method of astrogaloside in anisic acid-sulfuric acid system by spectrophotofluorimetry. METHODS　Using 2% anisic acid anhydrous alcoholic solution as colour developing reagent in the presence of sulfuric acid to determine astragaloside in pig serum. The reaction was allowed to procced on a 60℃ water-bath for 20 min. RESULTS　The reaction product of astrogaloside and anisic acid has excitation and emission maxima at 320 and 387 nm, respectively. The linear range of determination is from 0.2 to 40 μg*mL^-1. The detection limit is 0.02 μg*mL^-1. The recoveries of samples are between 98.65% and 102.4%. CONCLUSION　The proposed method has advantages of high sensitivity, low detection limit and no interference. It has been used to determine several kinds of samples successfully.