Objective To establish a pre-column derivation of HPLC-UV and determine bile acid and hyodeoxycholic acid in artificial calculus bovis. Methods Bile acid and hyodeoxycholic acid were derived by 2-bromo-2’-acetonaphthoneat using triethylamine as the catalyst in 60 ℃ water bath. The effects of derivatizing agent usage

catalyst and derivatizing time were investigated to identify the optimal parameters. HPLC-UV parameters were established to determine the content of hyodeoxycholic acid in artificial calculus bovis. Results When the ratio of molar amount of derivatizing agents to that of cholic acid and hyodeoxycholic was over 30∶ 1

and the ratio of molar amount of catalyst to that of cholic acid and hyodeoxycholic ranged from 5∶ 1 to 60∶ 1

the reaction was completed in 60℃ water bath for 50 minutes. The linear relationship was found with agent concentration（0. 4

5. 0 μg） in cholic acid（ r = 0. 999 5） and the linear relationship was also present in hyodeoxycholic（0. 12

1. 5 μg

r = 0. 999 8）. The average recovery rate of bile acid（ n = 6） was 100% and that of the RSD was 2. 40%. The average recovery rate of hyodeoxycholic acid was 101% and that of the RSD was 1. 33%. In this sample

the content of cholic acid was 5. 55% and the content of hyodeoxycholic was 2. 55%. Conclusion The method is replicable

reliable and accurate to determine bile acid and hyodeoxycholic acid in artificial calculus bovis.