Bile acid excretion was studied in 9 human subjects simultaneously by the Lindstedt (Lindstedt, S. 1957. Acta Physiol. Scand. 40:1-9) isotopic turnovermethod and by fecal chemical analysis during a balance study. The identities of the fecal bile acids were confirmed by combined gas-liquid chromatography/mass spectrometry. Under the steady state conditions of the patient studies, bile acid excretion values obtained by fecal analysis were lower (by 18.1 to 44.2%) than the values obtained by the isotopic turnover method. This difference persisted even in those patients given [14C]chenodeoxycholic acid instead of [3H]chenodeoxycholic acid. The fecal excretion values were similar when calculated using either beta-sitosterol or chromium sesquioxide as fecal flow markers. The fecal excretion values during the earlier part of the isotopic study were higher than those during the latter part of the study. The lower values of bile acid excretion obtained from fecal analysis could not be explained by the loss of bile acids as sulfate conjugates or by losses due to bile acid degradation in the intestine. This study suggests that bile acid turnover is consistently higher than bile acid excretion under experimental conditions. It is recommended that the data obtained from the isotopic turnover method should not be compared with fecal excretion data.