The objective of this study was to investigate the physicochemical characteristics of bear, pig, and cow biles to determine their authenticities for safe food distribution. The main bile acids of bear bile were tauroursodeoxycholic acid and taurochenodeoxycholic acid, while glycochenodeoxycholic acid and taurochenodeoxycholic acid for pig bile and taurocholic acid and glycocholic acid for cow bile were majorly detected by HPLC analysis. HPLC analysis was effective in monitoring of several samples imported as bear bile if employed to determine the authenticity of each bile. After the analysis of fatty acid composition by GC analysis, the ratio of the oleic acid of bear bile was relatively low compared to pig and cow biles. The ratio of the linoleic acid of bear bile was also similar to pig bile, whereas it had a tendency to be high compared to cow bear. The genetic analysis of the imported bile samples was mostly in agreement with the results of HPLC and GC analysis to identify the origin of imported biles. Finally, this study on the identification of bile origin by physicochemical analysis can give basic information to monitor the origin of biles and further to establish a reliable system for bear bile distribution.