Non-digestive oligosaccharides are often added to infant formula to help formula-fed infants develop an intestinal microbiota composed predominantly of Bifidobacteria, similar to that of breast-fed infants. Because various types of oligosaccharides exhibit specific microbial metabolism, the combined usage of oligosaccharides is considered to provide additive or synergistic effects for Bifidobacteria growth in the intestinal microbiota. The aim of this study was to evaluate the combined effect of lactulose, raffinose, and galacto-oligosaccharide (GOS) on the growth of Bifidobacterium breve, one of the major Bifidobacteria found in infant intestinal microbiota, using an in vitro mixed culture model. Seven typical bacterial species found in infant intestinal microbiota, including B. breve, were selected and then co-cultured under anaerobic conditions to mimic the infant intestinal environment. Each oligosaccharide was added to the medium, alone or in combination with other oligosaccharides. At all times, the total amount of added oligosaccharides composed 1% of the medium. Cells were harvested after several hours of incubation, and bacterial genomic DNA was extracted. Bacterial cell numbers were determined using quantitative realtime-PCR, with specific primers targeting the 16s rRNA genes of different bacterial groups. The combination of lactulose, raffinose, and GOS promoted the growth of Bifidobacterium breve compared with any single oligosaccharide or the combination of lactulose and raffinose. The combined usage of lactulose, raffinose, and GOS may provide the benefit of promoting a Bifidobacteria-predominant intestinal microbiota in formula-fed infants.
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