The contribution of ingredient glycosylation and milk glycoproteins to infant milk formulations and bacterial microbiome activity
Dziembała, Iwona
Dziembała, Iwona
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Publication Date
2023-08-16
Keywords
glycosylation, milk glycoproteins, infant milk formulations, bacterial microbiome activity, Science and Engineering, Science, Microbiology, Biological and Chemical Sciences, infant milk formula, global glycosylation, lectin microarray, N-linked glycans, HPLC analysis, probiotes, gut microbiome, Bifidobacterium, Lactobacillus, antimicrobial activity
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Thesis
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Abstract
Infant milk formula (IMF) is a food that has a specific purpose in the feeding of infants at different stages of their development. It is modelled on human milk, which contains a number of unique ingredients to ensure the optimal development of the child. Bovine milk is the most widely used base in the world to create IMF. To ensure comparable biological effects in IMF-fed infants, bovine milk is adjusted to model the composition of human milk and the main goal of IMF manufacturers is understanding and simulating the composition of human milk in bovine milk-based formulas. The glycosylation of human milk contributes its complexity and bioactivities as a selective prebiotic and antibacterial agent, ensuring optimal development of the immune system and intestinal microbiome. Human milk oligosaccharides (HMOs) together with oligosaccharides conjugated to proteins (glycoproteins) and lipids (glycolipids) make up the global glycosylation of human milk. Approximately 70% of human milk proteins are glycosylated, and these glycoproteins contribute to the unique properties of milk. Bovine milk oligosaccharides (BMOs) are not such complex structures, but bovine milk glycoproteins expresses complex glycosylation which is structurally altered over the lactation period similar to HMOs. It has been suggested that glycans from bovine milk glycoproteins may functionally replace HMOs in bovine milk. We suggest that bovine milk glycoproteins are a potential source of components that structurally could contribute to increasing the similarity of global glycosylation of formulas to human milk. The infant gut microbiome and its proper development plays a key role in the acquisition of nutrients from food, contributes to the correct development of the immune system, and protect against pathogens and the development of infections. Feeding the intestinal bacteria with milk glycans and glycoproteins is strongly linked to the final composition of the infant gut microbiome, and HMOs have been particularly associated with promotion of Bifidobacterium growth. Lactobacillus strains present in breast milk are also present in the gut of infants, indicating the promotion of growth of this bacterial population by components of human milk. In addition to their prebiotic effects, milk glycans and glycoproteins show antimicrobial activity, especially those present in LF. Many studies have reported on the activity of HMOs, but there is a gap in the literature regarding the activity of milk glycoproteins on Lactobacillus growth and the effect on some of the most common infant gut pathogens, Staphylococcus aureus and enteropathogenic Escherichia coli (EPEC). In Chapter 2 of this thesis, the contribution of successive additions of raw ingredients to the global glycosylation of different IMF preparations was assessed and compared with human and bovine whole milks. Incubation on lectin microarrays allowed the determination of the global glycosylation profiles, including N- and O-linked glycosylation. HPLC analysis allowed the analysis of free OSs in addition to N-linked glycans. Hierarchical clustering of the resulting profiles clearly indicated that successive additions of raw components, initially oligosaccharides such as BMOs, 2’-fucosyllactose (2’-FL), fructo-oligosaccharides (FOS) and lacto-N-neotetraose (LNnT), altered the global glycosylation of the IMF base. However, the greatest similarity in global glycosylation to human milk was obtained with the addition of both BMOs and the glycoproteins lactoferrin (LF) and osteopontin (OPN). Thus, enriching the base IMF with these components can 'humanise' the basic formula. Chapter 3 reports the evaluation of the potential of milk glycoproteins as selective milk constituents in modulating the colonisation of the intestinal microbiome. The effects of the milk glycoproteins LF, OPN, and α-lactalbumin (α-Lac) on the growth of Lactobacilus rhamnosus, Lactobacillus paracasei and Lactobacillus brevis, and the pathogens S. aureus (MRSA) and EPEC was assessed at different concentrations, corresponding to their concentrations in human milk colostrum, mature human milk, mature bovine milk and standard IMF. The milk glycoproteins at various concentrations functioned as selective components important for long-term colonisation of intestinal bacteria, promoting the growth of different commensals at certain concentrations and inhibiting the growth of pathogens. In addition, the inhibitory effect of the culture supernatant of L. rhamnosus cultured with LF against MRSA was investigated. The greatest antagonistic effect was noted with supernatant from L. rhamnosus supplemented with LF at the concentration corresponding to human milk colostrum. The results of the study supports the idea of milk glycoproteins strategically modulating infant gut colonisation at critical periods of infant microbiome development and introduces the possibility of using products of microbial digestion of LF glycosylation by L. rhamnosus and its growth metabolites as an anti-MRSA therapy.
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NUI Galway