Impact of dietary regime on the metabolomic profile of bovine buttermilk and whole milk powder.

INTRODUCTION: Bovine milk contains a rich matrix of nutrients such as carbohydrates, fat, protein and various vitamins and minerals, the composition of which is altered by factors including dietary regime. OBJECTIVES: The objective of this research was to investigate the impact of dietary regime on the metabolite composition of bovine whole milk powder and buttermilk. METHODS: Bovine whole milk powder and buttermilk samples were obtained from spring-calving cows, consuming one of three diets. Group 1 grazed outdoors on perennial ryegrass which was supplemented with 5% concentrates; group 2 were maintained indoors and consumed a total mixed ration diet; and group 3 consumed a partial mixed ration diet consisting of perennial ryegrass during the day and total mixed ration maintained indoors at night. RESULTS: Metabolomic analysis of the whole milk powder (N = 27) and buttermilk (N = 29) samples was preformed using liquid chromatography-tandem mass spectrometry, with 504 and 134 metabolites identified in the samples respectively. In whole milk powder samples, a total of 174 metabolites from various compound classes were significantly different across dietary regimes (FDR adjusted p-value ≤ 0.05), including triglycerides, of which 66% had their highest levels in pasture-fed samples. Triglycerides with highest levels in pasture-fed samples were predominantly polyunsaturated with high total carbon number. Regarding buttermilk samples, metabolites significantly different across dietary regimes included phospholipids, sphingomyelins and an acylcarnitine. CONCLUSION: In conclusion the results reveal a significant impact of a pasture-fed dietary regime on the metabolite composition of bovine dairy products, with a particular impact on lipid compound classes.

Pasture feeding improves the nutritional, textural, and techno-functional characteristics of butter.

There is an increasing consumer desire for pasture-derived dairy products, as outdoor pasture-based feeding systems are perceived as a natural environment for animals. Despite this, the number of grazing animals globally has declined as a result of the higher milk yields achieved by indoor TMR feeding systems, in addition to the changing climatic conditions and lower grazing knowledge and infrastructure. This has led to the development of pasture-fed standards, stipulating the necessity of pasture and its minimum requirements as the primary feed source for products advertising such claims, with various requirements depending on the region for which it was produced. This work investigates the differences in the composition and techno-functional properties of butters produced from high, medium and no pasture allowance diets during early, mid, and late lactation. Butters were produced using milks collected from 3 feeding systems: outdoor pasture grazing (high pasture allowance); indoor TMR (no pasture allowance); and a partial mixed ration (medium pasture allowance) system, which involved outdoor pasture grazing during the day and indoor TMR feeding at night. Butters were manufactured during early, mid, and late lactation. Creams derived from TMR feeding systems exhibited the highest milk fat globule size. The fatty acid profiles of butters also differed significantly as a function of diet and could be readily discriminated by partial least squares analysis. The most important fatty acids in such an analysis, as indicated by their highest variable importance projection scores, were CLA C18:2 cis-9,trans-11 (rumenic acid), C16:1n-7 trans (trans-palmitoleic acid), C18:1 trans (elaidic acid), C18:3n-3 (α-linolenic acid), and C18:2n-6 (linoleic acid). Increasing pasture allowances resulted in reduced crystallization temperatures and hardness of butters and concurrently increasing the "yellow" color. Yellow color was strongly correlated with Raman peaks commonly associated with carotenoids. The milk fat globule size of cream decreased with advancing stage of lactation and churning time of cream was lowest in early lactation. Differences in the fatty acid and triglyceride contents of butter as a result of lactation and dietary effects demonstrated significant correlations with the hardness, rheological, melting, and crystallization profiles of the butters. This work highlighted the improved nutritional profile and functional properties of butter with increasing dietary pasture allowance, primarily as a result of increasing proportions of unsaturated fatty acids. Biomarkers of pasture feeding (response in milk proportionate to the pasture allowance) associated with the pasture-fed status of butters were also identified as a result of the significant changes in the fatty acid profile with increasing pasture allowance. This was achieved through the use of 3 authentic feeding systems with varying pasture allowances, commonly operated by farmers around the world and conducted across 3 stages of lactation.

The impact of seasonal calving systems with varying pasture allowance on Cheddar cheese composition, nutritional quality and ripening characteristics.

The objective of this study was to examine the impact of stage of lactation (early, mid and late) and proportion of pasture in the cows diet (high: GRS, medium: PMR and no: TMR) on the composition and quality of Cheddar cheese. Triplicate trials were carried out in each stage of lactation, and milk protein and fat contents were standardized for Cheddar cheese manufacture at pilot scale. As cheese milks were standardized for milk fat and protein contents, gross composition did not differ as a result of diet. Fatty acid profiles of GRS cheese were significantly different from TMR, while PMR profiles were less distinct and more similar to both GRS and TMR profiles, as illustrated by partial least squares discriminatory analysis. Fatty acids including CLA C18:2 cis-9, trans-11, C22:1 n-9 and C18:3 n-3 were most influential in this separation of profiles. Fatty acid profiling revealed that GRS derived cheese contained higher proportions of nutrients considered beneficial for human health including higher proportions of unsaturated fatty acids and omega-3 fatty acids. A biomarker model utilizing the proportions of 5 fatty acids was constructed and was effective at distinguishing between cheese of GRS, TMR and PMR feeding systems. Proportions of ρ-κ-casein, αs2-casein and αs1-casein in cheese also differed between diets while proportions of ρ-κ-casein, αs1-casein and ß-casein were lowest in late lactation cheese. The impact of diet was less influential compared with that of stage of lactation on the ripening characteristics of cheese. An index of primary proteolysis was highest in late lactation cheese. The peptides derived from the proteolysis of κ-casein and ß-casein and levels of secondary proteolysis, in particular, the proportions of 12 free amino acids were most influenced by stage of lactation. Overall this study demonstrated the effects of increasing pasture allowance and stage of lactation on the nutritional quality and ripening properties of Cheddar cheese.

Impact of varying levels of pasture allowance on the nutritional quality and functionality of milk throughout lactation.

The objective of this study was to examine the impact of increasing proportions of grazed pasture in the diet on the composition, quality, and functionality of bovine milk across a full lactation. Fifty-four spring-calving cows were randomly assigned to 1 of 3 groups (n = 18), blocked on the basis of mean calving date (February 15, 2020 ± 0.8 d), pre-experimental daily milk yield (24.70 ± 3.70 kg), milk solids yield (2.30 ± 0.27 kg), lactation number (3.10 ± 0.13), and economic breeding index (182 ± 19). Raw milk samples were obtained weekly from each group between March and November 2020. Group 1 (GRS) consumed perennial ryegrass and was supplemented with 5% concentrates (dry matter basis); group 2 was maintained indoors and consumed a total mixed ration (TMR) diet consisting of maize silage, grass silage, and concentrates; and group 3 consumed a partial mixed ration diet (PMR), rotating between perennial ryegrass during the day and indoor TMR feeding at night. Raw milk samples consisted of a pooled morning and evening milking and were analyzed for gross composition, free amino acids, fatty acid composition, heat coagulation time, color, fat globule size, and pH. The TMR milks had a significantly higher total solids, lactose, protein, and whey protein as a proportion of protein content compared with both GRS and PMR milks. The GRS milks demonstrated a significantly lower somatic cell count (SCC), but a significantly higher pH and b*-value than both TMR and PMR milks. The PMR milks exhibited significantly lower total solids and fat content, but also demonstrated significantly higher SCC and total free amino acid content compared with GRS and TMR. Partial least squares discriminant analysis of fatty acid profiles displayed a distinct separation between GRS and TMR samples, while PMR displayed an overlap between both GRS and TMR groupings. Variable importance in projection analysis identified conjugated linoleic acid cis-9,trans-11, C18:2n-6 cis, C18:3n-3, C11:0, and C18:2n-6 trans as the largest contributors to the variation between the diets. Milk fats derived from GRS diets exhibited the highest proportion of unsaturated fats and higher unsaturation, health-promoting, and desaturase indices. The lowest proportions of saturated fats and the lowest atherogenic index were also exhibited by GRS-derived milk fats. This work highlights the positive influence of grass-fed milk for human consumption through its more nutritionally beneficial fatty acid profile, despite the highest milk solid percentages derived from TMR feeding systems. Furthermore, this study demonstrates the proportional response of previously highlighted biomarkers of pasture feeding to the proportion of pasture in the cow's diet.

Advances of plant-based structured food delivery systems on the in vitro digestibility of bioactive compounds.

Food researchers are currently showing a growing interest in in vitro digestibility studies due to their importance for obtaining food products with health benefits and ensuring a balanced nutrient intake. Various bioactive food compounds are sensitive to the digestion process, which results in a lower bioavailability in the gut. The main objective of structured food delivery systems is to promote the controlled release of these compounds at the desired time/place, in addition to protecting them during digestion processes. This review provides an overview of the influence of structured delivery systems on the in vitro digestive behavior. The main delivery systems are summarized, the pros and cons of different structures are outlined, and examples of several studies that optimized the use of these structured systems are provided. In addition, we have reviewed the use of plant-based systems, which have been of interest to food researchers and the food industry because of their health benefits, improved sustainability as well as being an alternative for vegetarian, vegan and consumers suffering from food allergies. In this context, the review provides new insights and comprehensive knowledge regarding the influence of plant-based structured systems on the digestibility of encapsulated compounds and proteins/polysaccharides used in the encapsulation process.

Dairy structures and physiological responses: a matter of gastric digestion.

Digestion and health properties of food do not solely rely on the sum of nutrients but are also influenced by food structure. Dairy products present an array of structures due to differences in the origin of milk components and the changes induced by processing. Some dairy structures have been observed to induce specific effects on digestion rates and physiological responses. However, the underlying mechanisms are not fully understood. Gastric digestion plays a key role in controlling digestion kinetics. The main objective of this review is to expose the relevance of gastric phase as the link between dairy structures and physiological responses. The focus is on human and animal studies, and physiological relevant in vitro digestion models. Data collected showed that the structure of dairy products have a profound impact on rate of nutrient bioavailability, absorption and physiological responses, suggesting gastric digestion as the main driver. Control of gastric digestion can be a tool for delivering specific rates of nutrient digestion. Therefore, the design of food structure targeting specific gastric behavior could be of great interest for particular population needs e.g. rapid nutrient digestion will benefit elderly, and slow nutrient digestion could help to enhance satiety.

Influence of desialylation of caseinomacropeptide on the denaturation and aggregation of whey proteins.

The effect of the addition of caseinomacropeptide (CMP) or desialylated CMP on the heat-induced denaturation and aggregation of whey proteins was investigated in the pH range 3 to 7 after heating at 80°C for 30 min. The rate and temperature of denaturation, the extent of aggregation, and the changes in secondary structure of the whey proteins heated in presence of CMP or desialylated CMP were measured. The sialic acid bound to CMP favored the denaturation and aggregation of whey proteins when the whey proteins were oppositely charged to CMP at pH 4. A transition occurred at pH 6, below which the removal of sialic acid enhanced the stabilizing properties of CMP against the denaturation and aggregation of the whey proteins. At pH >6, the interactions between desialylated CMP and the whey proteins led to more extensive denaturation and aggregation. Sialic acid bound to CMP influenced the denaturation and aggregation behavior of whey proteins in a pH-dependent manner, and this should be considered in future studies on the heat stability of such systems containing CMP.

Pilot-scale formation of whey protein aggregates determine the stability of heat-treated whey protein solutions-Effect of pH and protein concentration.

Denaturation and consequent aggregation in whey protein solutions is critical to product functionality during processing. Solutions of whey protein isolate (WPI) prepared at 1, 4, 8, and 12% (wt/wt) and pH 6.2, 6.7, or 7.2 were subjected to heat treatment (85°C × 30 s) using a pilot-scale heat exchanger. The effects of heat treatment on whey protein denaturation and aggregation were determined by chromatography, particle size, turbidity, and rheological analyses. The influence of pH and WPI concentration during heat treatment on the thermal stability of the resulting dispersions was also investigated. Whey protein isolate solutions heated at pH 6.2 were more extensively denatured, had a greater proportion of insoluble aggregates, higher particle size and turbidity, and were significantly less heat-stable than equivalent samples prepared at pH 6.7 and 7.2. The effects of WPI concentration on denaturation/aggregation behavior were more apparent at higher pH where the stabilizing effects of charge repulsion became increasingly influential. Solutions containing 12% (wt/wt) WPI had significantly higher apparent viscosities, at each pH, compared with lower protein concentrations, with solutions prepared at pH 6.2 forming a gel. Smaller average particle size and a higher proportion of soluble aggregates in WPI solutions, pre-heated at pH 6.7 and 7.2, resulted in improved thermal stability on subsequent heating. Higher pH during secondary heating also increased thermal stability. This study offers insight into the interactive effects of pH and whey protein concentration during pilot-scale processing and demonstrates how protein functionality can be controlled through manipulation of these factors.

Gastric digestion of α-lactalbumin in adult human subjects using capsule endoscopy and nasogastric tube sampling.

In the present study, structural changes in the milk protein α-lactalbumin (α-LA) and its proteolysis were investigated for the potential formation of protein-fatty acid complexes during in vivo gastric digestion. Capsule endoscopy allowed visualisation of the digestion of the test drinks, with nasogastric tubes allowing sampling of the gastric contents. A total of ten healthy volunteers had nasogastric tubes inserted into the stomach and ingested test drinks containing 50 g/l of sucrose and 25 g/l of α-LA with and without 4 g/l of oleic acid (OA). The samples of gastric contents were collected for analysis at 3 min intervals. The results revealed a rapid decrease in the pH of the stomach of the subjects. The fasting pH of 2·31 (SD 1·19) increased to a pH maxima of pH 6·54 (SD 0·29) after ingestion, with a subsequent decrease to pH 2·22 (SD 1·91) after 21 min (n 8). Fluorescence spectroscopy and Fourier transform IR spectroscopy revealed partial protein unfolding, coinciding with the decrease in pH below the isoelectric point of α-LA. The activity of pepsin in the fasting state was found to be 39 (SD 12) units/ml of gastric juice. Rapid digestion of the protein occurred: after 15 min, no native protein was detected using SDS-PAGE; HPLC revealed the presence of small amounts of native protein after 24 min of gastric digestion. Mirocam® capsule endoscopy imaging and video clips (see the online supplementary material) revealed that gastric peristalsis resulted in a heterogeneous mixture during gastric digestion. Unfolding of α-LA was observed during gastric transit; however, there was no evidence of a cytotoxic complex being formed between α-LA and OA.

Options for substantiating protein content claims for conventional foods.

In Canada and the United States, front-of-package protein content claims require data to support the quality of the protein. In general, protein quality reflects the product of the amino acid composition of the food protein relative to human amino acid requirements and a measure of digestibility. The currently accepted method in both jurisdictions is the protein digestibility-corrected amino acid score (PDCAAS) that requires the measurement of true fecal protein (nitrogen) digestibility. The latter must be measured in vivo using a rat model. This requirement for animal testing is inconsistent with international efforts to reduce the usage of animals in testing for regulatory purposes. The current commentary positions four options to remove the need to use animal testing for determining protein quality, when considering protein content claim substantiation. These options include (i) a focus on protein quantity alone; (ii) the use of the amino acid score alone, with no correction for digestibility; (iii) the use of a fixed digestibility coefficient to estimate protein quality; and (iv) the use of in vitro methods to measure protein and/or amino acid digestibility. The relative merits and deficiencies of the options are positioned with the goal of encouraging dialogue within the regulatory agencies to move towards alternative approaches for substantiating protein content claims on foods, including those derived from plant-based sources.

Current advances for in vitro protein digestibility.

Protein is an essential macronutrient in our diet, source of nitrogen and essential amino acids, but the biological utilization of dietary protein depends on its digestibility and the absorption of amino acids and peptides in the gastrointestinal tract. The methods to define the amount and the quality of protein to meet human nutritional needs, such as the Digestible Indispensable Amino Acid Score (DIAAS), require the use of animal models or human studies. These in vivo methods are the reference in protein quality evaluation, but they are expensive and long-lasting procedures with significant ethical restrictions. Therefore, the development of rapid, reproducible and in vitro digestion methods validated with in vivo data is an old demand. This review describes the challenges of the in vitro digestion methods in the evaluation of the protein nutritional quality. In addition to the technical difficulties to simulate the complex and adaptable processes of digestion and absorption, these methods are affected by similar limitations as the in vivo procedures, i.e., analytical techniques to accurately determine bioavailable amino acids and the contribution of the endogenous nitrogen. The in vitro methods used for the evaluation of protein digestibility, with special attention on those showing comparative data, are revised, emphasizing their pros and cons. The internationally harmonized digestion protocol proposed by the INFOGEST network is being adapted to evaluate protein and amino acid digestibility. The inter-laboratory reproducibility of this protocol was demonstrated for dairy products. The in vivo/in vitro comparability results obtained to date with this protocol for several plant and animal sources are promising, but it requires an extensive validation with a wider range of foods and substrates with known in vivo digestibility. These in vitro methods will probably not be applicable to all foods, and therefore, it is important to identify their limitations, not to elude their use, but to apply them within the limits, by using the appropriate standards and references, and always as a complementary tool to in vivo tests to reduce their number.

Microencapsulation of flaxseed oil in pea protein-gum arabic complex coacervates delays lipid digestion in liquid yoghurt.

Flaxseed oil coacervates were produced by complex coacervation using soluble pea protein and gum arabic as shell materials, followed by either spray or electrostatic spray drying and their incorporation to yoghurt. Three yoghurt formulations were prepared: yoghurt with spray-dried microcapsules (Y-SD); with electrospray-dried microcapsules (Y-ES); with the encapsulation ingredients added in free form (Y). The standardised semi-dynamicin vitrodigestion method (INFOGEST) was employed to study the food digestion. The structure was analysed by confocal laser scanning microscopy and particle size distribution. Protein and lipid digestion were monitored by cumulated protein/free NH2 release and cumulated free fatty acids release, respectively. Stable microcapsules were observed during gastric digestion, but there was no significant difference in protein release/hydrolysis among samples until 55 min of gastric digestion. Formulation Y showed less protein release after 74 min (40.46 %) due to the free SPP being available and positively charged at pH 2-4, resulting in interactions with other constituents of the yoghurt, which delayed its release/hydrolysis. The total release of protein and free NH2 by the end of intestinal digestions ranged between 46.56-61.15 % and 0.83-1.57 µmol/g protein, respectively. A higher release of free fatty acids from formulation Y occurred at the end of intestinal digestion, implying that coacervates promoted the delayed release of encapsulated oil. In summary, incorporating protein-polysaccharides-based coacervates in yoghurt enabled the delay of the digestion of encapsulated lipids but accelerated the digestion of protein, suggesting a promising approach for various food applications.

Electrostatic spray drying: A new alternative for drying of complex coacervates.

Complex coacervation can be used for controlled delivery of bioactive compounds (i.e., flaxseed oil and quercetin). This study investigated the co-encapsulation of flaxseed oil and quercetin by complex coacervation using soluble pea protein (SPP) and gum arabic (GA) as shell materials, followed by innovative electrostatic spray drying (ES). The dried system was analyzed through encapsulation efficiency (EE) and yield (EY), morphological and physicochemical properties, and stability for 60 days. Small droplet size emulsions were produced by GA (in the first step of complex coacervation) due to its greater emulsifying activity than SPP. Oil EY and EE, moisture, and water activity in dried compositions ranged from 75.7 to 75.6, 76.0-73.4 %, 3.4-4.1 %, and 0.1-0.2, respectively. Spherical microcapsules were created with small and aggregated particle size but stable for 60 days. An amount of 8 % of quercetin remained in the dried coacervates after 60 days, with low hydroperoxide production. In summary, when GA is used as the emulsifier and SPP as the second biopolymer in the coacervation process, suitable coacervates for food applications are obtained, with ES being a novel alternative to obtain coacervates in powder, with improved stability for encapsulated compounds. As a result, this study helps provide a new delivery system option and sheds light on how the characteristics of biopolymers and the drying process affect coacervate formation.

Effect of In Vitro Enzyme Digestion and Bile Treatment on Milk Extracellular Vesicles Stability.

SCOPE: Milk extracellular vesicles (EVs) are nanosized particles with potential immune bioactivities. This study examines their fate during in vitro infant gastrointestinal digestion (GI). METHODS AND RESULTS: Bovine milk is digested using the in vitro INFOGEST method, adjusted for the infant. To unravel the contribution of digestive enzymes from bile, milk is treated with digestive enzymes, bile, or a combination of both. EVs are collected posttreatment using differential ultracentrifugation. EVs characterization includes electrophoresis, immunoblotting, nanoparticle tracking analysis, and atomic force microscopy. EVs protein markers programmed cell death 6-interacting protein (ALIX), tumor susceptibility gene 101 (TSG101), cluster of differentiation 9 (CD9), and xanthine dehydrogenase (XDH) are detected after gastric digestion (G60), but their signal intensity is significantly reduced by intestinal conditions (p < 0.05). Enzyme digestion, compared to bile treatment (I60 + bile), results in a significant reduction of signal intensities for TSG101 and CD9 (p < 0.05). Nanoparticle tracking analysis shows a significant reduction (p < 0.05) of EV numbers at the end of the intestinal phase. EVs are detected by atomic force microscopy at the end of the intestinal phase, showing that intact EVs can survive upper gut digestion. CONCLUSION: Intact EVs can be found at the end of the intestinal phase. However, digestive enzymes and bile reduce the quantity and characteristics of EVs, with digestive enzymes playing a larger role.

An examination of the impact of unmelted, melted, and deconstructed cheese on lipid metabolism: a 6-week randomised trial.

Background: Evidence suggests cheese has a favourable or neutral effect on cardiometabolic health, compared to butter. To date, studies have only considered the cheese matrix in its unmelted form, while the effect of melted cheese remains unknown. Objective: To test the effect of 6-week daily consumption of ∼40 g dairy fat, eaten in either as unmelted cheese, melted cheese, or in a fully deconstructed form, on markers of metabolic health in overweight adults aged ≥50 years of age. Design: A 6-week randomised parallel intervention, where 162 participants (43.3% male) received ∼40 g of dairy fat per day, in 1 of 3 treatments: (A) 120 g full-fat Irish grass-fed cheddar cheese, eaten in unmelted form (n 58); (B) 120 g full-fat Irish grass-fed cheddar cheese eaten in melted form (n 53); or (C) the equivalent components; butter (49 g), calcium caseinate powder (30 g), and Ca supplement (CaCO3; 500 mg) (n 51). Results: There was no difference in weight, fasting glucose, or insulin between the groups post-intervention. Melted cheese, compared to unmelted cheese, increased total cholesterol (0.23 ± 0.79 mmol L-1vs. 0.02 ± 0.67 mmol L-1, P = 0.008) and triglyceride concentrations (0.17 ± 0.39 mmol L-1vs. 0.00 ± 0.42 mmol L-1, P = 0.016). Melted cheese increased total cholesterol concentrations by 0.20 ± 0.15 mmol L-1 and triglyceride concentrations by 0.17 ± 0.08 mmol L-1 compared to unmelted cheese. No significant differences were observed between the cheese forms for change in HDL, LDL or VLDL cholesterol. Conclusion: Compared to unmelted cheese, melted cheese was found to increase total cholesterol and triglyceride concentrations in middle-aged, overweight adults with no effect on weight or glycaemic control.

Infant milk formula, produced by membrane filtration, promotes mucus production in the upper small intestine of young pigs.

Human breast milk promotes maturation of the infant gastrointestinal barrier, including the promotion of mucus production. In the quest to produce next generation infant milk formula (IMF), we have produced IMF by membrane filtration (MEM-IMF). With a higher quantity of native whey protein, MEM-IMF more closely mimics human breast milk than IMF produced using conventional heat treatment (HT-IMF). After a 4-week dietary intervention in young pigs, animals fed a MEM-IMF diet had a higher number of goblet cells, acidic mucus and mucin-2 in the jejunum compared to pigs fed HT-IMF (P < 0.05). In the duodenum, MEM-IMF fed pigs had increased trypsin activity in the gut lumen, increased mRNA transcript levels of claudin 1 in the mucosal scrapings and increased lactase activity in brush border membrane vesicles than those pigs fed HT-IMF (P < 0.05). In conclusion, MEM-IMF is superior to HT-IMF in the promotion of mucus production in the young gut.

The Impact of Varying Pasture Levels on the Metabolomic Profile of Bovine Ruminal Fluid.

A pasture or concentrate-based dietary regime impacts a variety of factors including both ruminal health and function, and consequently milk production and quality. The objective of this study was to examine the effect of feeding differing pasture levels on the metabolite composition of bovine ruminal fluid. Ruminal fluid was obtained from rumen-cannulated spring-calving cows (N = 9, Holstein-Friesian breed, average lactation number = 5) fed one of three diets across a full lactation season. Group 1 (pasture) consumed perennial ryegrass supplemented with 5% concentrates; group 2 received a total mixed ration (TMR) diet; and group 3 received a partial mixed ration (PMR) diet which included pasture and a TMR. Samples were taken at two timepoints: morning and evening. Metabolomic analysis was performed using nuclear magnetic resonance (1H-NMR) spectroscopy. Statistical analysis revealed significant changes across the dietary regimes in both morning and evening samples, with distinct alterations in the metabolite composition of ruminal fluid from pasture-fed cows (FDR-adjusted p-value < 0.05). Acetate and butyrate were significantly higher in samples derived from a pasture-based diet whereas sugar-related metabolites were higher in concentrate-based samples. Furthermore, a distinct diurnal impact on the metabolite profile was evident. This work lays the foundation for understanding the complex interaction between dietary regime and ruminal health.

Impact of feeding regimes and lactation stage on sensory attributes of Cheddar cheese.

This study investigated the effects of diet and stage of lactation (SOL) on sensory profiles, texture, volatile profiles, and colour of Cheddar cheese. Cheddar cheese was manufactured from early-, mid-, and late-lactation milk obtained from seasonally calved cows (n = 54). Cows were assigned a diet; group 1: perennial ryegrass (GRS), group 2: total mixed ration (TMR), and group 3: partial mixed ration (PMR). Instrumental analysis was performed at 270 days (mature Cheddar). Sensory evaluation took place after 548 days (extra mature Cheddar). Toluene was the only volatile compound that was significantly influenced by diet. The trained panel rated early-lactation cheese as stronger than mid- and late- for cowy/barny flavour and late-lactation cheese as sweeter than early- and mid-lactation cheese. Mid-lactation cheese was liked least overall. Early-lactation cheeses were rated higher for 'crumbly' texture than mid- and late. Diet affected consumer ratings, with GRS and PMR cheese rated as more intense than TMR for flavour, aftertaste, and saltiness. Consumers reported that TMR cheese was lighter in colour compared to GRS cheese, which was supported by instrumental analysis. Consumers perceived GRS as more springy and less crumbly than TMR and PMR, while Texture Profile Analysis indicated that TMR was harder than GRS. Consumer segmentation was observed with two clear preference groups, one preferring GRS and one preferring TMR. For both groups, 'taste' seemed to be the main driver of liking, highlighting that consumer preference is most impacted by individual taste preferences.

ß-Lactoglobulin-linoleate complexes: In vitro digestion and the role of protein in fatty acid uptake.

The dairy protein ß-lactoglobulin (BLG) is known to bind fatty acids such as the salt of the essential longchain fatty acid linoleic acid (cis,cis-9,12-octadecadienoic acid, n-6, 18:2). The aim of the current study was to investigate how bovine BLG-linoleate complexes, of various stoichiometry, affect the enzymatic digestion of BLG and the intracellular transport of linoleate into enterocyte-like monolayers. Duodenal and gastric digestions of the complexes indicated that BLG was hydrolyzed more rapidly when complexed with linoleate. Digested as well as undigested BLG-linoleate complexes reduced intracellular linoleate transport as compared with free linoleate. To investigate whether enteroendocrine cells perceive linoleate differently when part of a complex, the ability of linoleate to increase production or secretion of the enteroendocrine satiety hormone, cholecystokinin, was measured. Cholecystokinin mRNA levels were different when linoleate was presented to the cells alone or as part of a protein complex. In conclusion, understanding interactions between linoleate and BLG could help to formulate foods with targeted fatty acid bioaccessibility and, therefore, aid in the development of food matrices with optimal bioactive efficacy.

Dairy and plant based protein beverages: In vitro digestion behaviour and effect on intestinal barrier biomarkers.

The consumer demand for protein-enriched food products continues to grow, in parallel with consumers' interest in plant based alternatives. The replacement of milk protein by plant protein is likely to be occur predominantly in prepared consumer foods such as nutritional beverages. This study aimed to compare and contrast powder beverages formulated with commercially available dairy versus plant ingredients in terms of protein digestion and gut barrier health. After simulated static in vitro gastrointestinal digestion, the release of free amino acids increased for all model beverages. In addition, the majority of peptides present in digested beverages were < 0.8 kDa in size. Gastrointestinal digestion did not increase the degree of protein hydrolysis in beverages formulated with prehydrolysed milk protein, whey or pea ingredients. A 2 h permeability assessment of digested beverages across the intestinal barrier, using Caco-2/HT-29/MTX co-cultures, revealed reduced transcription of tight junction protein 1, claudin-1 and mucus protein 2 albeit gut barrier impedance was unchanged. IL-8 mRNA levels in cell monolayers was significantly increased with digested fluids treatment but even more so with digesta from hydrolysed milk protein beverage. Overall, the response observed on intestinal biomarkers with digested plant beverages was similar to dairy based beverages supporting the replacement of dairy with plant proteins in powder beverage formulations.