Ratio-Dependent Regulation of Butyrate Production by 2′-Fucosyllactose and Fructooligosaccharides in a Bifidobacterium bifidum-Faecalibacterium prausnitzii Co-Culture

An appropriate 2′-fucosyllactose (2′-FL) and fructooligosaccharides (FOS) mixing ratio may enhance butyrate production through substrate complementarity and metabolite-mediated interactions; however, the substrate balance required for this effect remains unclear. This study investigated how different 2′-FL/FOS ratios affect butyrate production in a two-species co-culture of Bifidobacterium bifidum and Faecalibacterium prausnitzii. 2′-FL and FOS were mixed at different weight ratios and added to monocultures and two-species co-cultures. The bacterial growth, relative abundance, and organic acid production were also evaluated. In monoculture, F. prausnitzii grew under FOS-containing conditions, and B. bifidum growth increased as the proportion of 2′-FL increased. In the co-culture of F. prausnitzii and B. bifidum, total OD600 and the production of lactate and acetate generally increased under 2′-FL-rich conditions. In contrast, butyrate production showed a nonlinear response and peaked at a 2′-FL/FOS ratio of 10:90 (w/w), although total co-culture growth was not maximized under this condition. These findings suggest that butyrate production in this co-culture system is regulated in a ratio-dependent manner and is not solely explained by total bacterial growth or acetate accumulation. Rather, the observed butyrate profile likely reflected the combined effects of substrate complementarity, maintenance of the F. prausnitzii population, and metabolite exchange between the two species.