Raj Mehra

Stable monomeric intermediate with exposed Cys-119 is formed during heat denaturation of β-lactoglobulin

The role of the free sulfhydryl group of beta-lactoglobulin in the formation of a stable non-native monomer during heat-treatment of beta-lactoglobulin solutions was investigated. Two concomitant events occurred at the earlier stage of heating: unfolding of native globular monomer and intramolecular sulfhydryl/disulfide exchange reaction. Thus, two denatured monomeric species were formed: a non-native monomer with exposed Cys-121 (Mcys121) which became reversible after cooling, and a stable non-native monomer with exposed Cys-119 (Mcys119) which exhibited both a larger hydrodynamic conformation than native monomer and low solubility at pH 4.7. The results also show that the formation of these monomeric species throughout heat-induced denaturation of native beta-lg monomers is faster than their subsequent aggregation. A mechanism describing the behavior of beta-lg denaturation/aggregation during heat-treatment under selected conditions (5.8 mg/ml, low ionic strength, pH 6.6, 85 degrees C) is presented.

Neutral and acidic oligosaccharides in Holstein-Friesian colostrum during the first 3 days of lactation measured by high performance liquid chromatography on a microfluidic chip and time-of-flight mass spectrometry

Oligosaccharides (OS) from bovine milk are a class of bioactive molecules that are receiving increasing commercial attention for their potential health benefits. In the present work we measured, comprehensively and systematically, free milk OS in the colostrum of 7 Holstein-Friesian cows during the first 3 d of lactation in 12-h intervals by HPLC-chip/time-of-flight mass spectrometry to determine the biological variation of free milk OS in early lactation. The high sensitivity and resolution of the analytical technique made it possible to monitor all OS species, thus providing a comprehensive and quantitative analysis of OS variations during colostrum production. This study confirmed that although sialyllactose is the major OS in bovine colostrum, several neutral OS species are present in significant abundance even at the third day of lactation. Furthermore, variation in terms of OS species and relative abundances of OS between cows suggest individual animal variation. These variations are likely due to genetic factors because environmental factors such as nutrition, lactation number, and accommodation were the same for all cows. This investigation revealed that colostrum milk from Holstein-Friesian cows is a rich source of neutral and acidic OS for the food and pharmaceutical industries.

Milk immunoglobulins for health promotion

Abstract The biological function of bovine colostral immunoglobulins is to provide the newborn calf with adequate passive immune protection against microbial infections. Immunoglobulin preparations designed for farm animals are commercially available, and some colostrum-based products are marketed also for humans as dietary supplements. The concentration of specific antibodies against a certain pathogenic microorganism can be raised in colostrum and milk by immunizing cows with this pathogen or its antigen. Advances in bioseparation and chromatographic techniques have made it possible to fractionate and enrich these antibodies and formulate so-called hyperimmune colostral or milk preparations. Their efficacy in prevention and treatment of various microbial infections has been evaluated in numerous studies. Immune milk preparations have proven effective in prophylaxis against infections caused by a variety of gastrointestinal pathogens. Their therapeutic efficacy, however, seems more limited. A few commercial immune milk products are already on market and more applications can be expected in the coming years. This article reviews the recent progress made in isolation techniques of bovine immunoglobulins and the application of colostral and immune milk preparations in fighting various microbial infectious diseases in humans.

The food glycome: A source of protection against pathogen colonization in the gastrointestinal tract

Trillions of microbes inhabit the gastrointestinal tract of humans with significant differences in the composition and distribution of intestinal flora along its length. Normally there is a symbiotic relationship between the intestinal microflora and the host, with mutual advantages for both partners. When this relationship is altered, commensal bacteria can rapidly shift toward pathogenicity resulting in the onset and progression of gastrointestinal infection. Pathogen adhesion and colonization is often a prelude to infection, and intervention at this early stage can help prevent disease. Bacteria have evolved a multitude of adhesion mechanisms commonly targeting surface carbohydrate structures of the host. Here, we review the ability of various dietary carbohydrates to prevent adhesion of pathogens to host cells. Given their significance in disease, and their ability to cause chronic infection, we have focussed on 3 model pathogens, Helicobacter pylori, Campylobacter jejuni and Clostridium difficile, and dietary carbohydrates which can inhibit their adhesion. The discovery of novel anti-adhesive dietary carbohydrates, once developed as nutraceutical ingredients, may serve as a novel method for preventing infectious diseases in the human gastrointestinal tract. Anti-adhesive carbohydrates used in this context are not bactericidal. Therefore, the spread of pathogens with resistance to antibiotics is less likely to occur.

Spectroscopic characterization of heat-induced nonnative β-lactoglobulin monomers

Previous studies have shown that two altered monomeric species were formed in the early steps of thermal denaturation of bovine β‐lactoglobulin (β‐lg), the well‐known Cys121‐exposed intermediate (Mcys121), and a new, stable monomer with exposed nonnative Cys119 (Mcys119). In this study, circular dichroism and fluorescence spectroscopies were used to characterize the structural features of these molecules. The structural characteristics of MCys121 after heating and cooling cycles are similar to those of native β‐lg. In contrast, Mcys119 monomer exhibits some characteristics of the well‐known molten‐globule state. Combined with other published data, these results indicate that heating induces at least two molten globule‐like states of β‐lg, a highly reactive Mcys121 that returns to native state after cooling, and a less‐reactive Mcys119 that is trapped and stabilized in a molten globule‐like state by nonnative disulfide bond.

Effect of altering the daily herbage allowance to cows in mid lactation on the composition, ripening and functionality of low-moisture, part-skim Mozzarella cheese

Milk was collected from three spring-calving herds, on different daily herbage allowances (DHA) of perennial rye-grass (16, 20 or 24 kg dry matter (DM)/cow for a 17 week period. On five occasions, at weekly intervals in the middle of the period, the three different milks were converted into low-moisture part-skim Mozzarella cheese. Increasing the DHA resulted in significant increases in the concentrations of protein in the cheesemilk (P < 0.05) and cheese whey (P < 0.02). The moisture-adjusted cheese yield increased significantly (P < 0.01) on raising the DHA from 16 to 24 kg grass DM/cow. DHA had no significant effects on any of the gross compositional values of the cheese (although moisture and fat-in-DM levels tended to decrease and increase respectively with increasing DHA). The hardness of the uncooked cheese and functionality of cooked cheese (i.e. melt time, flowability, stretch and viscosity) were not significantly influenced by DHA over the 115 d ripening period at 4 degrees C.

Cheesemaking, compositional and functional characteristics of low-moisture part-skim Mozzarella cheese from bovine milks containing κ-casein AA, AB or BB genetic variants

Low-moisture part-skim Mozzarella cheeses were made, at pilot scale (450 kg), on five occasions at weekly intervals from milks containing κ-casein AA, AB or BB genetic variants. Compared with κ-casein A variant milks, the κ-casein B variant milks were associated with higher concentrations of casein ( P P <0·02) and total protein ( P <0·001), superior curd-forming properties ( P P <0·05). The moisture-adjusted (to 465 g/kg) cheese yields for the κ-casein AA, AB and BB cheeses were 91·5, 100·6, and 102·5 kg/1000 kg milk respectively. κ-Casein variant had no significant effect on the proteolysis and ripening of uncooked cheese or on the functionality (melt time, flowability and stretchability) of the cooked cheese during the course of a 90 d ripening period.

Effects of stocking density and concentrate supplementation of grazing dairy cows on milk production, composition and processing characteristics

The effects on milk composition and processing characteristics of varying grass supply by changing stocking density and of offering a concentrate supplement were investigated. The experiment was conducted over 28 weeks of the lactation (April-October) using 48 spring-calved Friesian-Holstein cows. Three herds each of 16 cows were offered a restricted grass supply, a standard grass supply and a standard grass supply with a supplement of 3 kg concentrate/d. Treatment groups were grazed separately with a residence time of 3 d/paddock. Milk production, composition and processing characteristics such as renneting properties, ethanol stability and plasmin activity were measured weekly. Increasing stocking density above the standard system resulted in significant reductions in milk fat and protein yields, the concentrations of total protein, casein and whey proteins, and a deterioration in most processing characteristics. Imposing concentrate supplementation on the standard system increased total protein, casein and whey protein concentrations but generally did not improve processing characteristics except for ethanol stability. These results suggest that the standard grass supply in a rotational grazing paddock system can support efficient production of quality milk, and concentrate supplementation will not improve processing characteristics when an adequate supply of good quality herbage is available.

Novel high-molecular weight fucosylated milk oligosaccharides identified in dairy streams.

Oligosaccharides are the third largest component in human milk. This abundance is remarkable because oligosaccharides are not digestible by the newborn, and yet they have been conserved and amplified during evolution. In addition to encouraging the growth of a protective microbiota dominated by bifidobacteria, oligosaccharides have anti-infective activity, preventing pathogens from binding to intestinal cells. Although it would be advantageous adding these valuable molecules to infant milk formula, the technologies to reproduce the variety and complexity of human milk oligosaccharides by enzymatic/organic synthesis are not yet mature. Consequently, there is an enormous interest in alternative sources of these valuable oligosaccharides. Recent research has demonstrated that bovine milk and whey permeate also contain oligosaccharides. Thus, a thorough characterization of oligosaccharides in bovine dairy streams is an important step towards fully assessing their specific functionalities. In this study, bovine milk oligosaccharides (BMOs) were concentrated by membrane filtration from a readily available dairy stream called “mother liquor”, and analyzed by high accuracy MALDI FT-ICR mass spectrometry. The combination of HPLC and accurate mass spectrometry allowed the identification of ideal processing conditions leading to the production of Kg amount of BMO enriched powders. Among the BMOs identified, 18 have high-molecular weight and corresponded in size to the most abundant oligosaccharides present in human milk. Notably 6 oligosaccharides contained fucose, a sugar monomer that is highly abundant in human milk, but is rarely observed in bovine milk. This work shows that dairy streams represent a potential source of complex milk oligosaccharides for commercial development of unique dairy ingredients in functional foods that reproduce the benefits of human milk.