The interactions between B. bifidum N4 and E. coli K-12 were investigated in their mixed cultures. Under conditions in which both bacteria grew well in their pure culture, B. bifidum inhibited the growth of E. coli even when the latter was inoculated at 104-fold and preincubated for 41 h. The inhibition in the mixed cultures appeared when the pH values were reduced below 4.6. When the lowering of pH was prevented by the addition of NaOH, no inhibition was observed. At the same initial pH of 6.5, lactic acid and acetic acid, metabolites of B. bifidum, had more inhibitory effect on the growth of E. coli than other aliphatic fatty acids. In the mixed cultures with E. coli, B. bifidum grew in the absence of its essential vitamins, riboflavin and pantetheine and, furthermore, aerobically.