• Covalent labelling of β-casein and its effect on the microstructure and physico-chemical properties of emulsions stabilized by β-casein and whey protein isolate

      Li, Meng; Auty, Mark; O'Mahony, James A.; Kelly, Alan L.; Brodkorb, Andre; Irish Dairy Levy Research Trust; Teagasc Walsh Fellowship Programme (Elsevier, 2016-12)
      The objective of this work was to investigate the effect of covalent labelling on the physico-chemical properties of β-casein (β-CN) in solution and in emulsions stabilized by β-CN and whey protein isolate (WPI). β-CN was covalently labelled by 5-(and 6)-carboxytetramethylrhodamine, succinimidyl ester (NHS-Rhodamine). The effect of conjugating β-CN with NHS-Rhodamine on the spectroscopic properties of labelled β-CN (β-CNlabelled) was examined. No significant difference in interfacial tension (p > 0.05) was found between mixture of WPI and β-CNlabelled (0.5% w/w WPI/β-CNlabelled) and of WPI and β-CN (0.5% w/w WPI/β-CN) in 10 mM phosphate buffer (pH 7.0) at 20 °C. Oil-in-water emulsions stabilized with either WPI/β-CN or WPI/β-CNlabelled (0.5% w/w) were also investigated using laser-light scattering, analytical centrifugation, rheometry and CLSM. It was shown that labelling had no significant effect on the physico-chemical properties of emulsions (p > 0.05) in terms of droplet size, creaming stability, viscosity or zeta-potential. Confocal micrographs of emulsions made with WPI/β-CNlabelled showed that both β-CN and whey proteins could be observed simultaneously, and were co-localized at the surface of fat globules. Furthermore, it was found through image analysis that β-CN produced a thicker interfacial layer than WPI.
    • Formation of non-native β-lactoglobulin during heat-induced denaturation

      Kehoe, Joseph James; Wang, Lizhe; Morris, Edwin R; Brodkorb, Andre; Teagasc Walsh Fellowship Programme; Department of Agriculture, Food and the Marine (Springer, 01/12/2011)
      A mechanism describing the denaturation and aggregation behavior during heat-treatment of pure β-lactoglobulin and β-lactoglobulin in whey protein isolate (WPI) under selected conditions (20 to 90 gL−1 in water at pH 7.0, 78 °C) is presented. A combination of reversed-phase and gel permeation chromatography was used to study the disappearance of native β-lactoglobulin and the formation of non-native intermediates in the aggregation process. The mean reaction order for pure β-lactoglobulin and β-lactoglobulin in WPI were the same, 1.4. While the rate of β-lactoglobulin denaturation was greater in WPI there was less aggregation compared to that of pure β-lactoglobulin. More of the β-lactoglobulin in WPI remained in a non-native monomer intermediate state after 30 min of heating. After an initial lag period, during which non-native monomers appeared, aggregates formed and rapidly reached a plateau in terms of their size. These aggregates were visualized using atomic force microscopy. There was no significant effect of protein concentration on either aggregate size or the number of exposed sulfhydryls in the heated solutions.
    • Impact of caseins and whey proteins ratio and lipid content on in vitro digestion and ex vivo absorption

      Mulet-Cabero, Ana-Isabel; Torcello-Gómez, Amelia; Saha, Shikha; Mackie, Alan R.; Wilde, Peter J.; Brodkorb, Andre; Irish Dairy Levy Research Trust; Teagasc Walsh Fellowship Programme; BBSRC UK; MDDT6261; et al. (Elsevier, 2020-07-30)
      Caseins and whey proteins are known as ‘slow’ and ‘fast’ proteins, respectively, based on their amino acid absorption rate. However, there is limited understanding of the mechanisms controlling their behaviour during gastro-intestinal transit. A protein model system (8% total protein) with varying casein:whey protein ratios (0:100, 20:80, 50:50 and 80:20) were subjected to in vitro gastro-intestinal digestion using a semi-dynamic gastric model, a static intestinal model and an ex vivo absorption model (Ussing chambers). The casein-rich (≥50%) samples showed the formation of solid coagula that were persistent throughout gastric digestion, which caused a delay in nutrient emptying, slower digestion and leucine absorption kinetics. In contrast, whey proteins formed more soluble aggregates during the gastric phase, which led to faster gastric emptying, rapid intestinal hydrolysis, and higher and faster leucine absorption. This work shows the key role of the gastric restructuring for the overall digestive mechanism and kinetics of food, in particular proteins.
    • Invited review: Whey proteins as antioxidants and promoters of cellular antioxidant pathways

      Corrochano, Alberto R.; Buckin, Vitaly; Kelly, Philip; Giblin, Linda; Department of Agriculture, Food and the Marine, Ireland; Teagasc Walsh Fellowship Programme; 13 F 454; 13 F 454-WheyGSH (Elsevier for American Dairy Science Association, 28/03/2018)
      Oxidative stress contributes to cell injury and aggravates several chronic diseases. Dietary antioxidants help the body to fight against free radicals and, therefore, avoid or reduce oxidative stress. Recently, proteins from milk whey liquid have been described as antioxidants. This review summarizes the evidence that whey products exhibit radical scavenging activity and reducing power. It examines the processing and treatment attempts to increase the antioxidant bioactivity and identifies 1 enzyme, subtilisin, which consistently produces the most potent whey fractions. The review compares whey from different milk sources and puts whey proteins in the context of other known food antioxidants. However, for efficacy, the antioxidant activity of whey proteins must not only survive processing, but also upper gut transit and arrival in the bloodstream, if whey products are to promote antioxidant levels in target organs. Studies reveal that direct cell exposure to whey samples increases intracellular antioxidants such as glutathione. However, the physiological relevance of these in vitro assays is questionable, and evidence is conflicting from dietary intervention trials, with both rats and humans, that whey products can boost cellular antioxidant biomarkers.
    • Whey protein effects on energy balance link the intestinal mechanisms of energy absorption with adiposity and hypothalamic neuropeptide gene expression

      Nilaweera, Kanishka; Cabrera-Rubio, Raul; Speakman, John R.; O'Connor, Paula M.; McAuliffe, Ann Marie; Guinane, Caitriona M.; Lawton, Elaine M.; Crispie, Fiona; Aguillera, Monica; Stanley, Maurice; et al. (American Physiological Society, 21/03/2017)
      We tested the hypothesis that dietary whey protein isolate (WPI) affects the intestinal mechanisms related to energy absorption and that the resulting energy deficit is compensated by changes in energy balance to support growth. C57BL/6 mice were provided a diet enriched with WPI with varied sucrose content, and the impact on energy balance-related parameters was investigated. As part of a high-sucrose diet, WPI reduced the hypothalamic expression of pro-opiomelanocortin gene expression and increased energy intake. The energy expenditure was unaffected, but epididymal weight was reduced, indicating an energy loss. Notably, there was a reduction in the ileum gene expression for amino acid transporter SLC6a19, glucose transporter 2, and fatty acid transporter 4. The composition of the gut microbiota also changed, where Firmicutes were reduced. The above changes indicated reduced energy absorption through the intestine. We propose that this mobilized energy in the adipose tissue and caused hypothalamic changes that increased energy intake, acting to counteract the energy deficit arising in the intestine. Lowering the sucrose content in the WPI diet increased energy expenditure. This further reduced epididymal weight and plasma leptin, whereupon hypothalamic ghrelin gene expression and the intestinal weight were both increased. These data suggest that when the intestine-adipose-hypothalamic pathway is subjected to an additional energy loss (now in the adipose tissue), compensatory changes attempt to assimilate more energy. Notably, WPI and sucrose content interact to enable the component mechanisms of this pathway.
    • Whey protein effects on energy balance link the intestinal mechanisms of energy absorption with adiposity and hypothalamic neuropeptide gene expression

      Nilaweera, Kanishka; Cabrera-Rubio, Raul; Speakman, John R.; O'Connor, Paula M.; McAuliffe, Ann Marie; Guinane, Caitriona M.; Lawton, Elaine M.; Crispie, Fiona; Aguillera, Monica; Stanley, Maurice; et al. (American Physiological Society, 19/06/2017)
      We tested the hypothesis that dietary whey protein isolate (WPI) affects the intestinal mechanisms related to energy absorption and that the resulting energy deficit is compensated by changes in energy balance to support growth. C57BL/6 mice were provided a diet enriched with WPI with varied sucrose content, and the impact on energy balance-related parameters was investigated. As part of a high-sucrose diet, WPI reduced the hypothalamic expression of pro-opiomelanocortin gene expression and increased energy intake. The energy expenditure was unaffected, but epididymal weight was reduced, indicating an energy loss. Notably, there was a reduction in the ileum gene expression for amino acid transporter SLC6a19, glucose transporter 2, and fatty acid transporter 4. The composition of the gut microbiota also changed, where Firmicutes were reduced. The above changes indicated reduced energy absorption through the intestine. We propose that this mobilized energy in the adipose tissue and caused hypothalamic changes that increased energy intake, acting to counteract the energy deficit arising in the intestine. Lowering the sucrose content in the WPI diet increased energy expenditure. This further reduced epididymal weight and plasma leptin, whereupon hypothalamic ghrelin gene expression and the intestinal weight were both increased. These data suggest that when the intestine-adipose-hypothalamic pathway is subjected to an additional energy loss (now in the adipose tissue), compensatory changes attempt to assimilate more energy. Notably, WPI and sucrose content interact to enable the component mechanisms of this pathway.
    • Whey protein isolate decreases murine stomach weight and intestinal length and alters the expression of Wnt signalling-associated genes

      McAllan, Liam; Speakman, John R.; Cryan, John F.; Nilaweera, Kanishka (Cambridge University Press, 13/01/2015)
      The present study examined the underlying mechanisms by which whey protein isolate (WPI) affects energy balance. C57BL/6J mice were fed a diet containing 10 % energy from fat, 70 % energy from carbohydrate (35 % energy from sucrose) and 20 % energy from casein or WPI for 15 weeks. Mice fed with WPI had reduced weight gain, cumulative energy intake and dark-phase VO2 compared with casein-fed mice (P< 0·05); however, WPI intake had no significant effects on body composition, meal size/number, water intake or RER. Plasma levels of insulin, TAG, leptin, glucose and glucagon-like peptide 1 remained unchanged. Notably, the intake of WPI reduced stomach weight and both length and weight of the small intestine (P< 0·05). WPI intake reduced the gastric expression of Wingless/int-1 5a (Wnt5a) (P< 0·01) and frizzled 4 (Fzd4) (P< 0·01), with no change in the expression of receptor tyrosine kinase-like orphan receptor 2 (Ror2) and LDL receptor-related protein 5 (Lrp5). In the ileum, WPI increased the mRNA expression of Wnt5a (P< 0·01) and caused a trend towards an increase in the expression of Fzd4 (P= 0·094), with no change in the expression of Ror2 and Lrp5. These genes were unresponsive in the duodenum. Among the nutrient-responsive genes, WPI specifically reduced ileal mRNA expression of peptide YY (P< 0·01) and fatty acid transporter protein 4 (P< 0·05), and decreased duodenal mRNA expression of the insulin receptor (P= 0·05), with a trend towards a decreased expression of Na–glucose co-transporter 1 (P= 0·07). The effects of WPI on gastrointestinal Wnt signalling may explain how this protein affects gastrointestinal structure and function and, in turn, energy intake and balance.