Immunonutrition, which focuses on specific nutrients in breast milk and post-weaning diets, plays a crucial role in supporting infants\' immune system development. This study explored the impact of maternal supplementation with Bifidobacterium breve M-16V and a combination of short-chain galacto-oligosaccharide (scGOS) and long-chain fructo-oligosaccharide (lcFOS) from pregnancy through lactation, extending into the early childhood of the offspring. The synbiotic supplementation\'s effects were examined at both mucosal and systemic levels. While the supplementation did not influence their overall growth, water intake, or food consumption, a trophic effect was observed in the small intestine, enhancing its weight, length, width, and microscopic structures. A gene expression analysis indicated a reduction in FcRn and Blimp1 and an increase in Zo1 and Tlr9, suggesting enhanced maturation and barrier function. Intestinal immunoglobulin (Ig) A levels remained unaffected, while cecal IgA levels decreased. The synbiotic supplementation led to an increased abundance of total bacteria and Ig-coated bacteria in the cecum. The abundance of Bifidobacterium increased in both the intestine and cecum. Short-chain fatty acid production decreased in the intestine but increased in the cecum due to the synbiotic supplementation. Systemically, the Ig profiles remained unaffected. In conclusion, maternal synbiotic supplementation during gestation, lactation, and early life is established as a new strategy to improve the maturation and functionality of the gastrointestinal barrier. Additionally, it participates in the microbiota colonization of the gut, leading to a healthier composition.

Galacto-oligosaccharides (GOS) are used as prebiotic ingredients in various food and pharmaceutical formulations. Currently, production of GOS involves the enzymatic conversion of lactose by transgalactosylation using β-galactosidase. The purity of the resulting product is low, typically limited to up to 55% GOS on total carbohydrate basis due to the presence of non-reacted lactose, and the formation of by-products glucose and galactose. In industrial practice high-purity GOS is manufactured by removing the unwanted mono- and disaccharides from raw GOS with simulated moving bed (SMB) chromatography. This purification step is associated with high processing cost that increases the price of pure GOS and limits its marketability. The last decades have witnessed a growing interest in developing competitive biotechnological processes that could replace chromatography. This paper presents a comprehensive review on the recent advancements of microbial GOS purification, a process commonly referred to as selective fermentation or selective metabolism. Purification strategies include: (i) removal of glucose alone or together with galactose by lactose negative yeast species, that typically results in purity values below 60% due to remaining lactose; (ii) removal of both mono- and disaccharides by combining the fast monosaccharide metabolizing capacity of some yeast species with efficient lactose consumption by certain lactose positive microbes, reaching GOS purity in the range of 60-95%; and (iii) the application of selected strains of Kluyveromyces species with high lactose metabolizing activity to achieve high-purity GOS that is practically free from lactose and monosaccharides.

Galacto-oligosaccharides (GOS) are prebiotic compounds widely used for their health-promoting effects. Conventionally, GOS is produced by the enzymatic conversion of lactose in stirred tank reactors (STR). The high operational costs associated with enzyme inactivation and removal might be reduced by the application of enzyme membrane reactors (EMR). In this study, we aimed to assess the potential of continuous GOS production by EMR using soluble Biolacta N5, a Bacillus circulans-derived commercial enzyme preparation. The steady-state performance of the EMR equipped with an ultrafiltration module was investigated as function of residence time (1.1-2.8 h) and enzyme load (17-190 U·g-1) under fixed operational settings of temperature (50 °C), pH (6.0), lactose feed concentration (300 g·kg-1), and recirculation flow-rate (0.18 m3·h-1). Results indicate that the yield of oligosaccharides with higher degree of polymerization (DP3-6) in STR (approx. 38% on total carbohydrate basis) exceeds that measured in EMR (ranging from 24% to 33%). However, a stable catalytic performance without a significant deterioration in product quality was observed when operating the EMR for an extended period of time (> 120 h). Approx. 1.4 kg of DP3-6 was produced per one gram of crude enzyme preparation over the long-term campaigns, indicating that EMR efficiently recovers enzyme activity.

Galacto-oligosaccharides (GOS) are prebiotic compounds, widely used as ingredients in various food, nutraceutical and pharmaceutical products. Enzymatic synthesis of GOS results in low-purity products that contain high amounts of glucose and lactose beside the valuable GOS. In this study, a systematic approach was used to develop yeast-based fermentation strategies to purify crude GOS. Potentially applicable yeast strains were identified based on an extensive search in literature databases followed by a series of laboratory-scale fermentation tests. Single- and two-step fermentation processes were designed for the removal of glucose alone or together with lactose from crude GOS syrup. Single-step fermentation trials with two strains of previously unreported species, Cyberlindnera jadinii NCAIM Y.00499 and Kluyveromyces nonfermentans NCAIM Y.01443, resulted in purified products free of both glucose and ethanol from a crude GOS syrup diluted to 15 and 10 w/v%, respectively. Simultaneous removal of glucose and lactose was achieved by Kluyveromyces marxianus DMB Km-RK in a single-step fermentation process with a yield of 97.5% and final purity of 100%. A two-step fermentation approach was designed to allow conversion of a glucose-free product into a high-purity GOS by removing glucose with C. jadinii Y.00499 in the first step, and lactose by Kluyveromyces lactis DMB Kl-RK in the second step, resulting in a final product with a yield of 100% and a final purity of 92.1%. These results indicate that the selected nonconventional yeasts are promising candidates for the removal of non-GOS components from commercial crude GOS products by selective fermentation.

A rational optimization for the synthesis of galacto-oligosaccharides (GOS) from lactose catalyzed by β-galactosidase from Aspergillus aculeatus, included in the commercial product Pectinex Ultra SP-L, has been performed by using experimental design and surface response methodology. This accurate tool optimized empirical production of the most desired high-GOS (tri-GOS and tetra-GOS) up to 16.4% under the following reaction conditions: 59 °C, 4 U/mL free enzyme concentration, pH 6.5, 250 g/L initial lactose concentration, and 20 h of reaction. The statistical analysis revealed temperature and initial lactose concentration as critical parameters. The successful immobilization of the enzyme on a glyoxyl-functionalized porous silica support slightly increased the yield toward high-GOS (17.6%), especially tri-GOS yield (15.3%), under the optimized reaction conditions as compared to the free enzyme. Furthermore, the promotion of the transgalactosylation reaction toward tri-GOS production increased 1.5-fold the productivity of high-GOS as compared to the free enzyme.

β-Galactosidase from Streptococcus thermophilus was overexpressed in a food-grade organism, Lactobacillus plantarum WCFS1. Laboratory cultivations yielded 11,000 U of β-galactosidase activity per liter of culture corresponding to approximately 170 mg of enzyme. Crude cell-free enzyme extracts obtained by cell disruption and subsequent removal of cell debris showed high stability and were used for conversion of lactose in whey permeate. The enzyme showed high transgalactosylation activity. When using an initial concentration of whey permeate corresponding to 205 g L-1 lactose, the maximum yield of galacto-oligosaccharides (GOS) obtained at 50°C reached approximately 50% of total sugar at 90% lactose conversion, meaning that efficient valorization of the whey lactose was obtained. GOS are of great interest for both human and animal nutrition; thus, efficient conversion of lactose in whey into GOS using an enzymatic approach will not only decrease the environmental impact of whey disposal, but also create additional value.

Like in many applications, solutions of high sugar content can cause serious problems due to microorganism contaminations. Hence, the main aim of this work was to study a nanofiltration process for GOS purification at 5 °C and 60 °C that may circumvent or reduce potential microbial growth. Process performances and rejection behaviors of monosaccharide as well as individual GOS components were compared. Operating at 5 °C is more advantageous especially with respect to the oligosaccharide (OS) recovery yield. Using a NF membrane (NP030) at 45 bar, a product purity of 85% (based on monosaccharide content) and an OS recovery yield of 82% could be achieved. However, a low average permeate flux of 3 L/m2 h had to be accepted. A diafiltration step improved product purity to 90% with 30% losses of OS. A qualitative theoretical discussion shows that a possible change of the pore radius distribution depending on temperature could play a role in solute rejection as well as selectivity.