Human milk oligosaccharides (HMO) are the third most abundant solid component of human milk. It is likely that they are responsible for at least some of the benefits experienced by breast-fed infants. Until recently HMO were absent from infant formula, but 2'-fucosyllactose (2'-FL) and lacto-N-neoteraose (LNnT) have recently become available as ingredients. The development of formula containing these HMO and the quality control of such formula require suitable methods for the accurate determination of the HMO. We developed two different approaches for analysis of 2'-FL and LNnT in formula; high performance anion exchange chromatography with pulsed amperometric detection (HPAEC-PAD) and hydrophilic interaction liquid chromatography with fluorescence detection (HILIC-FLD). In lab trials using blank formula spiked with the two oligosaccharides, both approaches worked well with recoveries of 94⁻111% (HPAEC-PAD) and 94⁻104% (HILIC-FLD) and RSD (iR) of 2.1⁻7.9% (HPAEC-PAD) and 2.0⁻7.4% (HILIC-FLD). However, when applied to products produced in a pilot plant, the HPAEC-PAD approach sometimes delivered results below those expected from the addition rate of the ingredients. We hypothesize that the oligosaccharides interact with the formula matrix during the production process and, during sample preparation for HPAEC-PAD those interactions have not been broken. The conditions required for labeling the HMO for detection by the FLD apparently disrupt those interactions, and result in improved recoveries. It is likely that both analytical approaches are appropriate if a suitable extraction process is used to recover the HMO.

Odd-chain fatty acids (OCFAs), with potential value for growing infants, have been reported in breast milk. The association of location and lactation stage with the profile and content of OCFAs in breast milk was studied. We analyzed 1487 breast milk samples collected from 12 areas in China, and 102 infant formulas from different brands were purchased from the local supermarket. The content of sn-2 C15:0 significantly decreased from the colostrum to the mature stage, while that of C17:0 was not significantly increased by the lactation stage (p > 0.05). The content of C15:0 and C17:0 significantly decreased dramatically after the colostrum period, while the content of C13:0 was highest in the mature stage. The level of C15:0 and C17:0 in human milk from Gansu and Xinjiang was significantly higher than that from other areas. Similar trends were observed on the level of sn-2 C15:0 and C17:0, whereas the content of sn-2 C11:0 and C13:0 was significantly higher in breast milk from Shandong. Based on the PDS-LA analysis, the difference among infant formulas, each stage of human milk and human milk from different locations were different. Research is needed to determine if there are health benefits associated with OCFAs.

Human milk oligosaccharides (HMOs) signify a unique group of oligosaccharides in breast milk, which is of major importance for infant health and development. The functional benefits of HMOs create an enormous impetus for biosynthetic production of HMOs for use as additives in infant formula and other products. HMO molecules can be synthesized chemically, via fermentation, and by enzymatic synthesis. This treatise discusses these different techniques, with particular focus on harnessing enzymes for controlled enzymatic synthesis of HMO molecules. In order to foster precise and high-yield enzymatic synthesis, several novel protein engineering approaches have been reported, mainly concerning changing glycoside hydrolases to catalyze relevant transglycosylations. The protein engineering strategies for these enzymes range from rationally modifying specific catalytic residues, over targeted subsite -1 mutations, to unique and novel transplantations of designed peptide sequences near the active site, so-called loop engineering. These strategies have proven useful to foster enhanced transglycosylation to promote different types of HMO synthesis reactions. The rationale of subsite -1 modification, acceptor binding site matching, and loop engineering, including changes that may alter the spatial arrangement of water in the enzyme active site region, may prove useful for novel enzyme-catalyzed carbohydrate design in general.

The use of antibiotics to treat diarrhea and other diseases early in life can lead to intestinal disorders in infants, which can cause a range of immune-related diseases. Intestinal microbiota diversity is closely related to dietary intake, with many oligosaccharides impacting intestinal microorganism structures and communities. Thus, oligosaccharide type and quantity are important for intestinal microbiota construction. Galactooligosaccharides (GOS) are functional oligosaccharides that can be supplemented with infant formula. Currently, information on GOS and its impact on intestinal microbiota diversity and disorders is lacking. Similarly, GOS is rarely reported within the context of intestinal barrier function. In this study, 16S rRNA sequencing, gas chromatography, and immunohistochemistry were used to investigate the effects of GOS on the intestinal microbiota and barrier pathways in antibiotic-treated mouse models. The results found that GOS promoted Bifidobacterium and Akkermansia proliferation, increased short-chain fatty acid levels, increased tight junction protein expression (occludin and ZO-1), increased secretory immunoglobulin A (SIgA) and albumin levels, significantly downregulated NF-κB expression, and reduced lipopolysaccharide (LPS), interleukin-IL-1β (IL-1β), and IL-6 levels. Also, a high GOS dose in ampicillin-supplemented animals provided resistance to intestinal damage.

Origanum vulgare L. (oregano) is an aromatic plant with wide applications in the food and pharmaceutical industries. Cronobacter sakazakii, which has a high detection rate in powdered infant formula, adversely impacts susceptible individuals. Oregano essential oil (OEO) is a natural antibacterial agent that can be used to fight bacterial contamination. Here, OEO chemical compounds from eight oregano varieties were analyzed by gas chromatography-mass spectrometry and their antibacterial properties were assessed. The eight OEOs were clustered into two groups and were more diverse in group 2 than in group 1. Six compounds, including p-cymene, 3-thujene, γ-terpinene, thymol, carvacrol, and caryophyllene, were shared by eight OEOs. Among the eight oregano varieties, OEOs from O. vulgare sc2 had the strongest antibacterial activity against C. sakazaki, with the inhibition zone of 18.22mm. OEOs from O. vulgare jx, O. 'Nvying', O. vulgare 'Ehuang', and O. vulgare ssp. virens were also potent. Moreover, the antibacterial activity of OEOs was positively correlated with the relative content of thymol. As the main OEO antibacterial compound, thymol affected the normal growth and metabolism of C. sakazakii cells by destroying the bacterial membrane and decreasing the intracellular ATP concentration. Thus, in light of the antibacterial activity detected in the OEOs from the eight oregano varieties, this study provides a theoretical foundation for oregano cultivar management and development.