Factors that affect the concentration of secondary bile acids in the aqueous phase of stool may have a greater impact on colon carcinogenesis than those that only modify the total fecal bile acid concentration. This hypothesis was tested using stool samples of a subset of participants enrolled in a Phase III colorectal adenomatous polyp prevention trial, which documented the inability of a 13.5 g/day wheat bran fiber (WBF) supplement to reduce polyp recurrence. Stool was collected from 68 consecutively consented participants who were enrolled in a Phase III clinical trial of WBF for the prevention of adenomatous polyp recurrence. Nineteen (27.9%) of these fecal bile acid substudy participants were on the low fiber (2.0 g/day) intervention group, whereas 49 (72.7%) were on the high fiber (13.5 g/day) intervention group for approximately 3 years. Sixty-four participants had both the aqueous and solid phases of stool samples analyzed for bile acid content. Bile acid concentrations, measured in microg/ml for fecal water and microg/mg for dry feces, were determined for lithochilic, deoxycholic, chenodeoxycholic, cholic, ursodeoxycholic, isodeoxycholic, isoursodeoxycholic, ursocholic, 7-ketolithocholic, and 12-ketolithocholic acids. There were no significant differences between the low and high fiber groups concerning mean or median aqueous phase concentrations of lithocholic or deoxycholic bile acids. In contrast, the median concentrations of deoxycholic acid and other secondary bile acids (including lithochilic, isodeoxycholic, ursodeoxycholic, isoursodeoxycholic, ursocholic, 7-ketolithocholic, and 12-ketolithocholic acids) were significantly lower for the high fiber group in the solid-phase stool (P < 0.05). These results document that a high WBF intervention, taken for a median of 2.4 years, does not significantly reduce aqueous-phase concentrations of secondary bile acids in stool, although their concentrations in solid-phase stool were suppressed. Thus, the inability of the high WBF intervention to reduce colorectal adenoma recurrence may be a consequence of its lack of effect on fecal aqueous-phase secondary bile acid concentrations.