Galacto-oligosaccharides as infant prebiotics: production, application, bioactive activities and future perspectives.

Galacto-oligosaccharides (GOS) are non-digestible oligosaccharides characterized by a mix of structures that vary in their degree of polymerization (DP) and glycosidic linkage between the galactose moieties or between galactose and glucose. They have enjoyed extensive scientific scrutiny, and their health-promoting effects are supported by a large number of scientific and clinical studies. A variety of GOS-associated health-promoting effects have been reported, such as growth promotion of beneficial bacteria, in particular bifidobacteria and lactobacilli, inhibition of pathogen adhesion and improvement of gut barrier function. GOS have attracted significant interest from food industries for their versatility as a bioactive ingredient and in particular as a functional component of infant formulations. These oligosaccharides are produced in a kinetically-controlled reaction involving lactose transgalactosylation, being catalyzed by particular ß-galactosidases of bacterial or fungal origin. Despite the well-established technology applied for GOS production, this process may still meet with technological challenges when employed at an industrial scale. The current review will cover relevant scientific literature on the beneficial physiological properties of GOS as a prebiotic for the infant gut microbiota, details of GOS structures, the associated reaction mechanism of ß-galactosidase, and its (large-scale) production.

Bifidobacterial ß-Galactosidase-Mediated Production of Galacto-Oligosaccharides: Structural and Preliminary Functional Assessments.

In the current study the ability of four previously characterized bifidobacterial ß-galactosidases (designated here as BgaA, BgaC, BgaD, and BgaE) to produce galacto-oligosaccharides (GOS) was optimized. Of these enzymes, BgaA and BgaE were found to be promising candidates for GOS production (and the corresponding GOS mixtures were called GOS-A and GOS-E, respectively) with a GOS concentration of 19.0 and 40.3% (of the initial lactose), respectively. GOS-A and GOS-E were partially purified and structurally characterized. NMR analysis revealed that the predominant (non-lactose) disaccharide was allo-lactose in both purified GOS preparations. The predominant trisaccharide in GOS-A and GOS-E was shown to be 3'-galactosyllactose, with lower levels of 6'-galactosyllactose and 4'-galactosyllactose. These three oligosaccharides have also been reported to occur in human milk. Purified GOS-A and GOS-E were shown to be able to support bifidobacterial growth similar to a commercially available GOS. In addition, GOS-E and the commercially available GOS were shown to be capable of reducing Escherichia coli adhesion to a C2BBe1 cell line. Both in vitro bifidogenic activity and reduced E. coli adhesion support the prebiotic potential of GOS-E and GOS-A.

Infant-Associated Bifidobacterial ß-Galactosidases and Their Ability to Synthesize Galacto-Oligosaccharides.

Galacto-oligosaccharides (GOS) represent non-digestible glycans that are commercially produced by transgalactosylation of lactose, and that are widely used as functional food ingredients in prebiotic formulations, in particular in infant nutrition. GOS consumption has been reported to enhance growth of specific bacteria in the gut, in particular bifidobacteria, thereby supporting a balanced gut microbiota. In a previous study, we assessed the hydrolytic activity and substrate specificity of seventeen predicted ß-galactosidases encoded by various species and strains of infant-associated bifidobacteria. In the current study, we further characterized seven out of these seventeen bifidobacterial ß-galactosidases in terms of their kinetics, enzyme stability and oligomeric state. Accordingly, we established whether these ß-galactosidases are capable of synthesizing GOS via enzymatic transgalactosylation employing lactose as the feed substrate. Our findings show that the seven selected enzymes all possess such transgalactosylation activity, though they appear to differ in their efficiency by which they perform this reaction. From chromatography analysis, it seems that these enzymes generate two distinct GOS mixtures: GOS with a relatively short or long degree of polymerization profile. These findings may be the stepping stone for further studies aimed at synthesizing new GOS variants with novel and/or enhanced prebiotic activities and potential for industrial applications.

Characterization of GH2 and GH42 ß-galactosidases derived from bifidobacterial infant isolates.

Bifidobacteria are among the first and most abundant bacterial colonizers of the gastrointestinal tract of (breast-fed) healthy infants. Their success of colonising the infant gut is believed to be, at least partly, due to their ability to metabolize available carbon sources by means of secreted or intracellular glycosyl hydrolases (GHs). Among these, ß-galactosidases are particularly relevant as they allow bifidobacteria to grow on ß-galactosyl-linked saccharidic substrates, which are present in copious amounts in the milk-based diet of their infant host (e.g. lactose and human milk oligosaccharides). In the present study we employed an in silico analysis to identify GH family 2 and 42 ß-galactosidases encoded by typical infant-associated bifidobacteria. Comparative genome analysis followed by characterisation of selected ß-galactosidases revealed how these GH2 and GH42 members are distributed among these infant-associated bifidobacteria, while their hydrolytic activity towards growth substrates commonly available in the infant gut were also assessed.