We wished to characterize the carbohydrate fermentation by intestinal flora in formula-fed infants and in breast-fed infants. We also wished to compare the carbohydrate fermentation process in the two groups to determine whether differences that existed between groups could help explain the observation that breast-fed infants usually have milder forms of acute gastroenteritis. We performed in vitro incubations of fecal samples from nine formula-fed and 14 breast-fed infants and examined the samples for fermentation characteristics, the effect of acid pH on bacterial fermentation, and changes in carbohydrate fermentation in relation to the age of the infant. Fecal samples were incubated, with and without the addition of lactose, at a pH of 6.8 and at a pH of 5.5. Volatile fatty acids and carbohydrates were determined in the incubates. The addition of lactose to the incubate at pH 6.8 resulted in significantly increased volatile fatty acid production and larger amounts of lactose, glucose, and galactose compared with the values observed in 1-h incubates to which no lactose was added. At pH 5.5, volatile fatty acid production was significantly lower in both groups compared with that at pH 6.8, and the accumulation of monosaccharides in the incubate of feces of formula-fed infants increased significantly (p<0.05). In contrast, in stools from breast-fed infants, incubation at pH 5.5 resulted in a greater proportion of saccharides as lactose,i.e.a decrease in the amount of lactose hydrolyzed (p<0.01), accompanied by a nonsignificant increase in the amount of hexose. The decrease in lactose hydrolysis in breast-fed infants results in a lower luminal osmolar load, which may provide a partial explanation for the fact that acute gastroenteritis is milder in this population.