Transgalacto-oligosaccharides are a mixture of oligosaccharides consisting of glucose and galactose; they are not digested in the human small intestine. In vitro, they specifically stimulate the growth of bifidobacteria. The aim of the present work was to assess tolerance of transgalacto-oligosaccharides and the effects of their prolonged administration on bifidobacteria and fermentative activity of colonic flora. Eight healthy subjects were given 10 g of transgalacto-oligosaccharides per day for 21 d in two daily doses. A breath test and stool sample collection were carried out on d 1, 7, 14 and 21 of transgalacto-oligosaccharides ingestion. The stools of three subjects were collected and mixed before the study, and then inoculated in vitro into a fermentor to which 10 g transgalacto-oligosaccharides was added daily for 14 d. In the eight volunteers, administration of transgalacto-oligosaccharides led to a significant decrease in breath hydrogen excretion (P < 0.01) and a significant increase in fecal concentrations of bifidobacteria from (means +/- SEM) 8.6 +/- 0.6 to 9.7 +/- 0.5, 9.7 +/- 0.6 and 9.5 +/- 0.6 log colony-forming units (CFU)/g on d 1, 7, 14 and 21, respectively (P < 0.05). Fecal concentrations of enterobacteria, as well as stool weight, fecal water and pH did not change during the study. In vitro, transgalacto-oligosaccharides fermentation became more efficient and faster with time. In addition, metabolic alterations such as a rise in acetate proportion and lactate formation after 7 d of fermentation were observed, indicating the transformation of the inoculated fecal flora into an acid-resistant lactic flora. Prolonged administration of transgalacto-oligosaccharides, at a dose which does not induce digestive symptoms, increases the number of bifidobacteria and alters the fermentative activity of colonic flora in humans.