A method of isolating chenodeoxycholic acid from hog bile is provided, which comprises the steps of treating the hog bile under saponifying conditions separating the bile acids from the water-soluble constituents of the saponified bile; esterifying the bile acids to obtain a mixture of bile acid esters; isolating from the mixture of bile acid esters the hyodeoxycholic acid ester as its adduct with a compound selected from the group consisting of benzene and toluene; treating the remaining bile acid esters under acetylating conditions; isolating the hyocholic acid ester triacetate; isolating the chenodeoxycholic acid ester diacetate; treating the chenodeoxycholic acid ester diacetate under saponifying conditions; isolating the chenodeoxycholic acid thus obtained; and, optionally purifying the chenodeoxycholic acid.The method described yields chenodeoxycholic acid of exceedingly high purity, which may be used in the treatment of gallstones by their dissolution in vivo.