The in vitro transformation, under anaerobic conditions, of 3- and 7-monosulfated and unsulfated bile acids, was studied in incubates of fecal flora from three healthy subjects. Chenodeoxycholic acid 7 alpha-sulfate and ursodeoxycholic acid 7 beta-sulfate were recovered unchanged, in all cultures, at the end of the incubation time. 3-Sulfated bile acids were metabolized in a different way by the three stool specimens. During the transformation of chenodeoxycholic acid 3-sulfate, desulfation, 7-dehydroxylation and 3-epimerization were observed. In contrast, 3-epimerization was not noticed when ursodeoxycholic acid 3-sulfate and lithocholic acid 3-sulfate were metabolized, the latter being principally transformed into delta 3-cholenic acid, probably by a bacterially mediated trans-elimination of sulfate group. The results obtained seem to prove that the presence of a SO3H group in 7-position usually hinders microbial transformations, which are not affected by a sulfate group in 3-position. Moreover, the 3-sulfated bile acids proved to be less sensitive to the microbial action than the corresponding unsulfated acids, with exception of lithocholic 3-sulfate.