Following the establishment of functionally active intestinal flora in three healthy Swedish children from birth up to 24 months of age, we investigated the development of different 24-carbon bile acids. The fecal bile acids were group-separated into unconjugated, glycine-conjugated, taurine-conjugated, and sulfated, so that we could follow the changes between the different fractions of conjugates. In meconium, most (55-63%) of the bile acids were conjugated with taurine; only 11-32% were conjugated with glycine. Deconjugation was the first sign of intestinal microbial activity on the bile acids. Already at 1 month of age, most of the bile acids were deconjugated; among the conjugated bile acids, the glycine-conjugated dominated over the taurine-conjugated. An unidentified conjugate of cholic and chenodeoxycholic acids (C, CDC) that separated with the sulfated bile acids was found. The unconjugated bile acids and those that arose from hydrolysis of existing conjugates were separated and identified by gas-liquid chromatography coupled to mass spectrometry (GC-MS). Twenty-nine different bile acids were identified. In meconium, 16 different bile acids were identified. C and CDC were identified in all samples. The bile acid pattern changed during the course of the study. Many of the identified bile acids were only found in one or a few of the analyzed samples, and sometimes only in samples from one child. 6 alpha-hydroxylated bile acids, probably not microbially synthesized, were present at high percentages in the children.(ABSTRACT TRUNCATED AT 250 WORDS)