Intact Whey Protein Research Articles

Survival of intact bovine whey proteins across in vivo and simulated static in vitro models of the adult gastrointestinal tract

Bovine whey contains bioactive proteins that could benefit consumer health, but their intact bioactivity depends on their ability to survive the digestive process and remain intact to their sites of action. Our study assessed whey protein degradation in the adult human jejunum after whey protein isolate ingestion and during static in vitro gastric and intestinal digestions, using LC-MS/MS-based proteomics and SDS-PAGE. We found that 53 % of the protein counts identified in the gastric digesta, including β-lactoglobulin and lactoferrin were stable under simulated gastric conditions. However, most proteins were degraded during both in vitro and in vivo intestinal digestion. The intact protein survival profiles were closely aligned between in vitro and in vivo digestion samples. Understanding the survival of specific whey proteins during digestion will help determine their biological relevancy in their intact forms within the gastrointestinal tract and may inspire strategies to enhance their stability for specific functions.

Characterization of some physicochemical, textural, and antioxidant properties of muffins fortified with hydrolyzed whey protein.

Whey protein hydrolysates, derived from enzymatic hydrolysis of whey protein isolates or concentrates, offer enhanced bioavailability and solubility compared to intact whey protein. In this study, whey protein hydrolysates (WPHs) having different hydrolysis degrees (5%, 10%, and 15%) were produced and muffin cakes were enriched with the addition of WPHs. In general, the addition of WPHs showed a significant effect on oil and protein content while the emulsion activity was improved with the increased hydrolysis degree (HD). The degree of hydrolysis increment resulted in a significant increase in both antioxidant power and antiradical activity of the WPHs. Ferric-reducing antioxidant power and ABTS radical scavenging activity ranged between 18.83-87.27 mg TE/100 g and 211.8-5063.1 mg TE/100 g, respectively. The highest FRAP and ABTS values were recorded for the 15% HD while the lowest was for the native whey protein isolate (WPI). The induction periods giving a clear information for the oxidative stability were 1593 min for the control muffins, and it was 1654for the muffin added with WPI. Rheological data revealed that all cake batter samples including WPHs showed viscoelastic behavior. WPHs could be efficiently used in muffin formulation to increase the biofunctional effects of the final products.

Preventive effect of fermented whey protein mediated by Lactobacillus gasseri IM13 via the PI3K/AKT/FOXO pathway in muscle atrophy

This study investigated the preventive effects of whey protein fermented with Lactobacillus gasseri IM13 (F-WP) against dexamethasone (DEX)-induced muscle atrophy. C2C12 muscle cells were treated with F-WP followed by DEX treatment. Dexamethasone treatment inhibited myotube formation and the expression of myogenic regulatory factors; however, pretreatment with F-WP attenuated DEX-induced damage. The F-WP significantly activated the phosphorylation of the IGF-1/PI3K/AKT pathway and improved muscle homeostasis suppressed by DEX. Moreover, F-WP alleviated the phosphorylation of mTOR, S6K1, and 4E-BP1 and enhanced muscle protein synthesis. Muscle-specific ubiquitin ligases and autophagy lysosomes, which were activated by the dephosphorylation of FOXO3a by DEX treatment, were significantly attenuated by F-WP pretreatment of myotubes. For peptidomic analysis, F-WP was fractionated using preparative HPLC (prep-HPLC), and the AA sequences of 11 peptides were identified using MALDI-TOF/MS/MS. In conclusion, fermentation of whey protein by the specific probiotic strain IM13 produced bioactive peptides with high antioxidant and anti-sarcopenic-sarcopenic effects, which markedly enhanced myogenesis and muscle protein synthesis while diminishing muscle protein degradation compared with intact whey protein.

Degalactosylated Whey Protein Suppresses Inflammatory Responses Induced by Lipopolysaccharide in Mice.

The effects of degalactosylated whey protein on lipopolysaccharide (LPS)-induced inflammatory responses in mice were observed in comparison with intact whey protein. Intraperitoneal administration of both intact and degalactosylated whey proteins for 5 days did not affect body weight and food intake in mice. On day 6, intraperitoneal administration of LPS induced a marked decrease in body weight 4 h later. The LPS-induced decrease in body weight was significantly suppressed by the administration of degalactosylated whey protein, but not intact whey protein. Administration of LPS also significantly increase plasma tumor necrosis factor-α (TNF-α) and interleukin-1β (IL-1β) levels, which were significantly suppressed by the administration of degalactosylated whey protein, but not intact whey protein. Moreover, the application of degalactosylated whey protein to RAW264.7 cells significantly reduced mRNA expression of toll-like receptor 4 (TLR4) and significantly increased mRNA expression of mitogen-activated protein kinase phosphatase-1 (MKP-1). The marked increased expression of TNF-α and IL-1β in response to LPS in RAW264.7 cells was significantly suppressed by the application of degalactosylated whey protein. These results suggest that degalactosylated whey protein suppresses the effects of LPS in part by decreasing in TLR4 and increasing in MKP-1.

Cow and camel milk-derived whey and casein protein hydrolysates demonstrated effective antifungal properties against selected Candida species

Bioactive peptides derived from milk proteins are widely known to possess antibacterial activities. Even though the antibacterial effects of milk-derived peptides are widely characterized, not much focus is given to their antifungal characterization. Therefore, in this study, we investigated the antifungal properties of camel and cow whey and casein hydrolysates against various species of pathogenic Candida. The hydrolysates were produced using 2 enzymes (alcalase and protease) at differing hydrolysis durations (2, 4, and 6 h) and tested for their antifungal properties. The results showed that intact cow whey and casein proteins did not display any anti-Candida albicans properties, whereas the alcalase-derived 2 h camel casein hydrolysate (CA-C-A2) displayed a higher percentage of inhibition against Candida albicans (93.69 ± 0.26%) followed by the cow casein hydrolysate generated by protease-6 h (Co-C-P6; 81.66 ± 0.99%), which were significantly higher than that of fluconazole, a conventional antifungal agent (76.92 ± 4.72%). Interestingly, when tested again Candida krusei, camel casein alcalase 2 and 4 h (CA-C-A2 and CA-C-A4), and cow whey alcalase-6 h (CO-W-A6) hydrolysates showed higher antifungal potency than fluconazole. However, for Candida parapsilosis only camel casein alcalase-4 h (Ca-C-A4) and cow casein protease-6 h (Co-C-P6) hydrolysates were able to inhibit the growth of C. parapsilosis by 19.31 ± 0.84% and 23.82 ± 4.14%, respectively, which was lower than that shown by fluconazole (29.86 ± 1.11%). Overall, hydrolysis of milk proteins from both cow and camel enhanced their antifungal properties. Camel milk protein hydrolysates were more potent in inhibiting pathogenic Candida species as compared with cow milk protein hydrolysates. This is the first study that highlights the antifungal properties of camel milk protein hydrolysates.

Efficient encapsulation of fish oil: Capitalizing on the unique inherent characteristics of whey cream and hydrolyzed whey protein

The effects of protein concentration and of blending a phospholipid-rich whey coproduct, Procream (Salibra 700 Procream, Glanbia Nutritionals), with intact or hydrolyzed whey protein concentrate, on fish oil microencapsulation efficiency and oxidative stability were assessed. Trypsin and protease M, from Aspergillus oryzae, were used to produce 2 unique hydrolysates. All microcapsules had excellent encapsulation efficiencies (>92%) and good physical properties, regardless of protein content and Procream inclusion. Intact α-lactalbumin and β-lactoglobulin and their peptides were involved in stabilizing oil droplets. Disulfide interchange resulted in formation of protein aggregates, which were more pronounced in samples containing Procream. Although all microcapsules had relatively good oxidative stability, most had better stability at 2 versus 0.5% protein. Protease M hydrolysate + Procream microcapsules had the highest stability, regardless of protein content. Results demonstrated that Procream, at a reduced protein inclusion level, can partially replace more expensive whey protein ingredients in microencapsulation, when blended with a select hydrolysate.

Identification and characterization of novel α-amylase and α-glucosidase inhibitory peptides from camel whey proteins

This study explores the inhibitory properties of camel whey protein hydrolysates (CWPH) toward α-amylase (AAM) and α-glucosidase (AG). A general full factorial design (3 × 3) was applied to study the effect of temperature (30, 37, and 45°C), time (120, 240, and 360 min), and enzyme (pepsin) concentration (E%; 0.5, 1, and 2%). The results showed that maximum degree of hydrolysis was obtained when hydrolysis was carried out at higher temperature (45°C; P < 0.05), compared with lower temperatures of 30 and 37°C. Electrophoretic pattern displays degradation of all protein bands upon hydrolysis by pepsin at various hydrolysis conditions applied. All the 27 CWPH generated showed significant AAM and AG inhibitory potential as indicated by their lower IC50 values (mg/mL) compared with intact whey proteins. In total 196 peptides were identified from selected hydrolysates and 15 potential peptides (PepSite score > 0.8; http://pepsite2.russelllab.org/) were explored via in silico approach. Novel peptides PAGNFLMNGLMHR, PAVACCLPPLPCHM, MLPLMLPFTMGY, and PAGNFLPPVAAAPVM were identified as potential inhibitors for both AAM and AG due to their high number of binding sites and highest binding probability toward the target enzymes. CCGM and MFE, as well as FCCLGPVPP were identified as AG and AAM inhibitory peptides, respectively. This is the first study that reports novel AG and AAM inhibitory peptides from camel whey proteins. The future direction for this research involves synthesis of these potential AG and AAM inhibitory peptides in a pure form and investigate their antidiabetic properties in the in vitro, as well as in vivo models. Thus, CWPH can be considered for potential applications in glycaemic regulation.

The Acute Effect of Bovine Whey Protein Fractions on Blood Glucose and Insulin in Rats

The ingestion of whey protein (WP) leads to the reduction of postprandial blood glucose (BG) response paralleled with an increased level of insulin. However, the role of individual WP fractions: glycomacropeptide (GMP), β‐Lactoglobulin (β‐LG), and α‐Lactalbumin (α‐LA) remain unclear.The objective of this study was to investigate the effect of WP, GMP, β‐LG, and α‐LA on BG and insulin response in rats.We hypothesized that due to the difference in their amino acid profile the effect of WP fractions on short‐term BG control may differ.MethodsA randomized repeated measures study was conducted in rats fitted with jugular vein catheters and vascular access buttons (VAB). Nine male 10‐week‐old 275–300g Wistar Han rats were gavaged 350mg (allometricaly scaled from a human dosage of 10g) of either intact WP, GMP, β‐LG, α‐LA, or a glucose control dissolved in 3ml of water after being fasted for 6h during daylight. The use of a VAB allowed for the same rat to receive all five treatments in a random order, with a 48h washout period between treatments. Blood was collected at 0, 15, and 30 min for insulin, and at 0, 15, 30, 60, 90, and 120 min for glucose. Whole blood was analyzed for glucose using a HemoCue 201 Glucose Analyzer, and plasma was analyzed for insulin using a wide range ELISA. The data were tested for normality and analyzed using Two‐Way Repeated Measures ANOVA for the effect of time, treatment and a treatment by time interaction. The data for the area under the curve (AUC) for 2h BG and 30 min insulin were analyzed with One‐Way Repeated Measures ANOVA. The differences between the treatments were assessed with Tukey‐Kramer post hoc test.ResultsThere was an effect of treatment (P&lt;0.0001), time (P&lt;0.0001) and a treatment by time interaction (P&lt;0.0001) over 120 min on BG response. Whey protein, GMP, β‐LG, and α‐LA resulted in significantly lower BG compared to glucose treatment at 15 and 30 min (P&lt;0.05). Blood glucose AUC over 120 min was lower for WP, GMP, and α‐LA compared to glucose treatment (P&lt;0.05). There was an effect of treatment (P=0.02), time (P&lt;0.0001) and a treatment by time interaction (P=0.0002) on insulin response over 30 min. Glycomacropeptide had a lower insulin response at 15 min compared to glucose and β‐LG (P&lt;0.05). Insulin AUC over 30 min was lower for GMP compared to the glucose treatment (P=0.003).ConclusionAt the dose of 350mg WP, GMP, β‐LG, and α‐LA attenuated BG response at 15 and 30 min, while only WP, GMP, and α‐LA attenuated AUC over 120 min compared to the glucose control. GMP attenuated insulin response at 15 min compared to β‐LG and the glucose control, and attenuated AUC over 30 min compared to the glucose control. These data suggest that WP fractions possess unique properties not observed in WP.Support or Funding InformationSupported by the Natural Sciences and Engineering Research Council of Canada (NSERC). Erik Vandenboer has received support from NSERC and Nova Scotia Health Research Foundation.

Inter-relationship between lactose crystallization and surface free fat during storage of infant formula

Inter-relationship between lactose crystallization (LC), the amount and composition of surface free fat (SFF); and their effect on physico-chemical properties of infant formula (IF) containing hydrolyzed and intact (non-hydrolyzed) whey protein in their composition were investigated at two temperatures (25 and 45 °C) and five RH (11–65%) conditions. Results varied with compositional variation of IF. LC increased exponentially with SFF in non-hydrolyzed IF powders. IF composition influenced LC and caused selective migration of triglycerides, resulting in higher proportion of unsaturated fats in SFF of powders with large lactose crystals and vice-versa. Increase in SFF with increased proportion of saturated fats in their composition, resulted in reduced wettability of powders. Overall, IF composition affects LC which influences the amount and type of fat migration to particle surface resulting in varying wettability of IF powders.

Effects of whey protein hydrolysate ingestion on post-exercise muscle protein synthesis compared with intact whey protein in rats

BackgroundIt is well known that ingestion of protein sources can stimulate muscle protein synthesis (MPS). The intake of whey protein is highly effective especially for accelerating MPS. Whey protein hydrolysate (WPH) can raise postprandial plasma concentration of amino acids, which impact stimulation of MPS more rapidly and highly than intact whey protein. However, it is unclear which is more effective for stimulating MPS, WPH or intact whey protein. The aim of the present study was to compare the effects of the WPH and whey protein on MPS in rats after exercise.MethodsRats were first subjected to a 2 h. swimming protocol. After this, in experiment 1, we evaluated time-dependent changes in the fractional synthetic rate (FSR) of the triceps muscle in Male Sprague-Dawley rats after ingestion of intact whey protein (30, 60, 90 or 120 min after ingestion). Then in experiment 2, at the time point that the results of Experiment 1 revealed postprandial FSR was highest (60 min after ingestion), we measured the FSR after ingestion of the WPH or whey protein at two different doses (0.5 or 2.0 g protein/kg body weight), or with deionized water (control), again after exercise. Plasma components and mammalian target of rapamycin (mTOR) signaling were also measured.ResultsIn experiment 1, postprandial FSR was highest 60 min after whey protein was administered. In experiment 2, the FSR 60 min after ingestion of the WPH was higher than that of whey protein (significant treatment main effect). Moreover, at a lower dose, only the WPH ingestion caused greater MPS and phosphorylated 4E-binding protein 1 (4E-BP1) levels compared with the control group.ConclusionThese results indicate that ingestion of the WPH was associated with greater post-exercise MPS compared with intact whey protein, especially at lower doses.

Alteration in propagating colonic contractions by dairy proteins in isolated rat large intestine

Gastrointestinal conditions in which the transit of contents is altered may benefit from nutritional approaches to influencing health outcomes. Milk proteins modulate the transit of contents along different regions, suggesting that they have varying effects on neuromuscular function to alter gastrointestinal motility. We tested the hypothesis that bovine whey and casein milk protein hydrolysates could have direct modulatory effects on colonic motility patterns in isolated rat large intestine. Casein protein hydrolysate (CPH), whey protein concentrate (WPC), whey protein hydrolysate (WPH), and a milk protein hydrolysate (MPH; a hydrolyzed blend of 60% whey to 40% casein) were compared for their effects on spontaneous contractile waves. These contractions propagate along the length of the isolated intact large intestine (22 cm) between the proximal colon and rectum and were detected by measuring activity at 4 locations. Milk proteins were perfused through the tissue bath, and differences in contraction amplitude and frequency were quantified relative to pretreatment controls. Propagation frequency was decreased by CPH, increased by MPH, and unaffected by intact whey proteins. The reduced motility with CPH and increased motility with MPH indicate a direct action of these milk proteins on colon tissue and provide evidence for differential modulation by hydrolysate type. These findings mirror actions on lower gastrointestinal transit reported in vivo, with the exception of WPH, suggesting that other factors are required.

In vitro-digested milk proteins: Evaluation of angiotensin-1-converting enzyme inhibitory and antioxidant activities, peptidomic profile, and mucin gene expression in HT29-MTX cells

Over the past decades, several studies investigated the health-promoting functions of milk peptides. However, to date many hurdles still exist regarding the widespread use of milk-derived bioactive peptides, as they may be degraded during gastrointestinal digestion. Thus, the aim of our study was to in vitro digest intact whey protein isolate (WPI) and casein proteins (CNP), mimicking in vivo digestion, to investigate their bioactive effects and to identify the potential peptides involved. Whey protein isolate and CNP were digested using a pepsin-pancreatin protocol and ultra-filtered (3-kDa cutoff membrane). A permeate (<3 kDa) and a retentate (>3 kDa) were obtained. Soy protein was included as a control (CTR). Angiotensin-1-converting enzyme inhibitory (ACE1-I) and antioxidant activity (AOX) were assessed and compared with those observed in undigested proteins and CTR. Furthermore, the permeate was characterized by nano-liquid chromatography electrospray ionization tandem mass spectrometry (LC-nano ESI MS/MS) using a shotgun peptidomic approach, and retentate was further digested with trypsin and analyzed by MS using a shotgun proteomic approach to identify potentially bioactive peptides. Further, the effects of WPI, CNP, and CTR retentate on cell metabolic activity and on mucus production (MUC5AC and MUC2 gene expression) were assessed in intestinal goblet HT29-MTX-E12 cells. Results showed that WPI permeate induced a significant ACE1-I inhibitory effect [49.2 ± 0.64% (SEM)] compared with undigested WPI, CNP permeate, and retentate or CTR permeate (10.40 ± 1.07%). A significant increase in AOX (1.58 ± 0.04 and 1.61 ± 0.02 µmol of trolox AOX equivalents per mg of protein, respectively) upon digestion was found in WPI. Potentially bioactive peptides associated with ACE1-I and antihypertensive effects were identified in WPI permeate and CNP retentate. At specific concentrations, WPI, CNP, and CTR retentate were able to stimulate metabolic activity in HT29-MTX-E12 cells. Expression of MUC5AC was increased by CNP retentate and unaltered by WPI retentate; MUC2 expression was significantly increased by 0.33 mg/g of CNP and reduced by 1.33 mg/g of CNP. Our results confirm that milk proteins may be rich sources of bioactive compounds, with the greatest beneficial potential of CNP at the intestinal goblet cell level.

Whey Protein Hydrolysate Increases Amino Acid Uptake, mTORC1 Signaling, and Protein Synthesis in Skeletal Muscle of Healthy Young Men in a Randomized Crossover Trial

Muscle protein synthesis (MPS) can be stimulated by ingestion of protein sources, such as whey, casein, or soy. Protein supplementation can enhance muscle protein synthesis after exercise and may preserve skeletal muscle mass and function in aging adults. Therefore, identifying protein sources with higher anabolic potency is of high significance. The aim of this study was to determine the anabolic potency and efficacy of a novel whey protein hydrolysate mixture (WPH) on mechanistic target of rapamycin complex 1 (mTORC1) signaling and skeletal MPS in healthy young subjects. Ten young men (aged 28.7 ± 3.6 y, 25.2 ± 2.9 kg/m2 body mass index [BMI]) were recruited into a double-blind two-way crossover trial. Subjects were randomized to receive either 0.08 g/kg of body weight (BW) of WPH or an intact whey protein (WHEY) mixture during stable isotope infusion experiments. Fractional synthetic rate, leucine and phenylalanine kinetics, and markers of amino acid sensing were assessed as primary outcomes before and 1-3 h after protein ingestion using a repeated measures mixed model. Blood leucine concentration, delivery of leucine to muscle, transport of leucine from blood into muscle and intracellular muscle leucine concentration significantly increased to a similar extent 1 h after ingestion of both mixtures (P<0.05). Phosphorylation of S6K1 (i.e. a marker of mTORC1 activation) increased equally by ∼20% 1-h postingestion (P<0.05). Ingestion of WPH and WHEY increased mixed MPS similarly in both groups by ∼43% (P<0.05); however, phenylalanine utilization for synthesis increased in both treatments 1-h postingestion but remained elevated 3-h postingestion only in the WPH group (P<0.05). We conclude that a small dose of WPH effectively increases leucine transport into muscle, activating mTORC1 and stimulating MPS in young men. WPH anabolic potency and efficacy for promoting overall muscle protein anabolism is similar to WHEY, an intact protein source. This trial was registered at clinicaltrials.gov as NCT03313830.

Plasma concentrations and ACE-inhibitory effects of tryptophan-containing peptides from whey protein hydrolysate in healthy volunteers

The tryptophan-containing dipeptides isoleucine-tryptophan (IW) and tryptophan-leucine (WL) are angiotensin-converting enzyme (ACE)-inhibitors in vitro. These peptides are released by enzymatic hydrolysis of bovine whey protein. To exhibit ACE inhibition in vivo, peptides need to be absorbed into the circulatory system. This study aimed to determine the in vivo ACE-inhibitory potency of a whey protein hydrolysate (MPH), containing IW and WL, and to quantify plasma concentrations of these peptides after oral administration of MPH in healthy volunteers. Additionally, changes in blood pressure were investigated. After intake of 5 and 50g MPH, plasma ACE activity was reduced to 86.4 ± 5.9 and 75.1 ± 6.9% of baseline activity, respectively. Although a clear ACE inhibition was measured, no effect on blood pressure was seen. Basal plasma concentrations of the tryptophan-containing dipeptides were 2.8 ± 0.7nM for IW and 10.1 ± 1.8nM for WL. After intake of 5-50g MPH, peptide concentrations were dose dependently elevated to values between 12.5 ± 8.4 and 99.1 ± 58.7nM for IW and 15.0 ± 4.3-34.9 ± 19.4nM for WL. Administration of intact whey protein showed a minor ACE inhibition, probably caused by release of inhibitory peptides during gastrointestinal digestion. The increase of WL in plasma after intake of intact protein was similar to that determined after intake of MPH. In contrast, resulting IW concentrations were much lower after intake of intact whey protein when compared to MPH administration. After intake of MPH, plasma ACE activity decreased in parallel to the increase of IW and WL plasma concentrations. However, the resulting peptide concentrations cannot fully explain the reduction of ACE activity in plasma with a direct enzyme inhibition. Therefore, this study points to a gap in the understanding of the inhibitory action of these peptides in vivo. Thus, to further develop innovative food additives with ACE activity diminishing capabilities, it appears mandatory to better characterize the mode of action of these peptides.

Regulation of GLUT4 translocation in an in vitro cell model using postprandial human serum ex vivo.

What is the research question? This study used a new experimental model, in which culture medium is conditioned with human serum ex vivo, to investigate nutrient-mediated regulation of GLUT4 translocation in skeletal muscle cells in vitro. What is the main finding and importance? Human serum stimulated GLUT4 translocation, an effect differentially modulated by whether the culture medium was conditioned with serum from fasted subjects or with serum collected after feeding of intact or hydrolysed whey protein. Conditioning cell culture medium with human serum ex vivo represents a new approach to elucidate the effects of ingesting specific nutrients on skeletal muscle cell metabolism. Individual amino acids, amino acid mixtures and protein hydrolysates stimulate glucose uptake in many experimental models. To replicate better in vitro the dynamic postprandial response to feeding in vivo, in the present study we investigated the effects of culture media conditioned with fasted and postprandial human serum on GLUT4 translocation in L6-GLUT4myc myotubes. Serum samples were collected from healthy male participants (n=8) at baseline (T0), 60 (T60) and 120min (T120) after the ingestion of 0.33g(kg body mass)-1 of intact (WPC) or hydrolysed (WPH) whey protein and an isonitrogenous non-essential amino acid (NEAA) control. L6-GLUT4myc myotubes were starved of serum and amino acids for 1h before incubation for 1h in medium containing 1% postprandial human serum, after which GLUT4 translocation was determined via colorimetric assay. Medium conditioned with fasted human serum at concentrations of 5-20% increased cell surface GLUT4myc abundance. Incubation with serum collected after the ingestion of WPH increased cell surface GLUT4myc at T60 relative to T0 [mean (lower, upper 95% confidence interval)]; [1.13 (1.05, 1.22)], whereas WPC [0.98 (0.90, 1.07)] or NEAA [1.02 (0.94, 1.11)] did not. The differential increases in cell surface GLUT4myc abundance were not explained by differences in serum concentrations of total, essential and branched-chain amino acids or insulin, glucagon-like peptide 1 (GLP-1) and gastric inhibitory polypeptide (GIP). Using a new ex vivo, in vitro approach, cell culture medium conditioned with postprandial serum after the ingestion of a whey protein hydrolysate increased GLUT4 translocation in skeletal muscle cells.

Role of carbohydrate conjugation on the emulsification and antioxidant properties of intact and hydrolysed whey protein concentrate

The conjugation of proteins with carbohydrates generates covalent bonds which may improve their techno- and biofunctional properties and therefore expand their applications in the food industry. In the present study, a design of experiments (DOE) approach was used to determine the effect of conjugation of whey protein concentrate (WPC) and WPC hydrolysates with carrageenan (CGN) on its emulsifying and antioxidant properties. The DOE was composed of 3 levels with 3 factors, i.e., WPC:CGN (1:1.0 1:3.5, 1:6.0), time of conjugation (6, 27 and 48 h) and degree of hydrolysis (DH; unhydrolysed, low DH and high DH). The conjugated samples were characterised for their oxygen radical absorbance capacity (ORAC) and emulsion activity (EA). Two samples, C3 (1:1.0, 6 h and unhydrolysed) and C4 (1:1.0, 6 h and low DH) with ORAC and EA values of 601.30 ± 14.71 and 709.32 ± 11.11 μmol Trolox equivalents g−1 freeze-dried powder and 0.51 ± 0.01 and 0.58 ± 0.01 absorbance units, respectively, were selected for further study. Emulsions were generated with WPC, non-conjugated (NC3 and NC4) and conjugated (C3 and C4) samples and their functional properties were compared. The NC3 and NC4 samples had higher viscoelastic moduli (G′ and G″) than C3, C4 and the WPC samples which indicated that more stable emulsions may be formed with non-conjugated samples. However, the NC3, NC4 and WPC samples showed low emulsion stability (ES) after 28 days storage with values ranging from 78.6 ± 3.5–85.6 ± 3.6% whereas the conjugated C3 and C4 samples had improved ES with significantly (p < 0.05) lower coalescence index values (64.2 ± 2.5 and 66.7 ± 3.7%, respectively). However, conjugation of the hydrolysate (C4) had a significant (p < 0.05) negative effect on lipid oxidation (0.62 ± 0.04 equivalents of 1,1,3,3-tetraethoxypropane (TEP)/kg emulsion) compared to the unhydrolysed conjugate C3 (0.42 ± 0.04 eq TEP/kg emulsion).

Inhibitory properties of camel whey protein hydrolysates toward liver cancer cells, dipeptidyl peptidase-IV, and inflammation

This report describes an investigation of camel whey protein hydrolysates (CWPH) produced by gastric and pancreatic enzymes for their in vitro antidiabetic, anticancer, and anti-inflammatory properties. Degree of hydrolysis (DH) ranged from 8.54 to 47.53%, with hydrolysates generated using chymotrypsin for 6 h displaying the highest DH. Reverse phase-HPLC analysis showed that α-lactalbumin underwent complete degradation, with no intact α-lactalbumin detected in CWPH. The CWPH displayed enhanced antidiabetic activity compared with intact whey proteins; with pepsin- and chymotrypsin-generated CWPH displaying greater inhibition of dipeptidyl peptidase IV (DPP-IV), α-glucosidase, and α-amylase compared with trypsin-generated CWPH. The highest antiproliferative effect was observed for CWPH generated by chymotrypsin for 3 h, with only 4.5 to 6.5% viable liver cancer cells (HepG2) remaining when tested at concentrations from 400 to 1,000 µg/mL. The highest anti-inflammatory activity was manifested by CWPH generated by pepsin at 6-h hydrolysis. We report enhanced antiproliferative, antidiabetic, and anti-inflammatory activities upon hydrolysis of camel whey proteins, indicating their potential utilization as bioactive and functional ingredients.

THE ANGIOTENSIN CONVERTING ENZYME-INHIBITORY EFFECTS OF THE PEPTIDE ISOLEUCINE-TRYPTOPHAN AFTER ORAL INTAKE VIA WHEY HYDROLYSATE IN MEN

Objective: Angiotensin-Converting Enzyme (ACE)-inhibitors are treatment of choice in hypertensive patients. In former studies we revealed the ACE-inhibitory activity and antihypertensive effect of the natural dipeptide isoleucine-tryptophan (IW) and its effect on cardiac remodelling and coronary flow reserve in spontaneously hypertensive rats. The aim in this project was to determine in volunteers the ACE-inhibiting effect and the bioavailability of IW after oral intake of whey hydrolysate. A second aim was to study the influence of combined intake of IW-containing hydrolysate with further food ingredients. Design and method: 8 fasted normotensive volunteers received IW containing whey hydrolysate in different doses, intact whey protein, or water. Compounds were applied alone or with parallel intake of specific food ingredients (fiber, protein, carbohydrates and fat). Blood samples were taken at fixed time points for analysis of ACE activity and assessment of IW concentration in plasma by LC-MS/MS method. Results: Baseline concentrations of IW differed individually ranging from 1.2 to 2.1 nM. The intake of 10 g hydrolysate (equivalent to 50 mg IW) induced a significant increase in plasma IW concentrations (maximum values 8.5–27.5 nM after 25–30 min). The activity of plasma ACE was significantly reduced to 72 – 87% of baseline activity during this time interval. The maximum of ACE inhibition coincided with the peak of IW concentration. Increasing the applied hydrolysate to 50 g caused IW concentrations to increase to 18–159 nM after 29–65 min. In parallel, ACE activity was decreased to 62–77%. However, after intake of intact whey protein IW concentrations increased only slightly to 2.3–4.9 nM and ACE activity was 88–91% of baseline activity. No changes in IW concentration and ACE activities were observed following placebo application. Combined intake of either protein, fiber or fat delayed IW resorption and extended IW rise of plasma concentration and ACE inhibition. Carbohydrates did not affect IW resorption, however, ACE inhibition was diminished compared to the single application of hydrolysate. Conclusions: In healthy volunteers oral intake of IW via whey hydrolysate shows a marked increase of IW plasma concentration and inhibition of plasma ACE activity. In addition, effects are retarded by parallel intake of food ingredients.

A small dose of whey protein co-ingested with mixed-macronutrient breakfast and lunch meals improves postprandial glycemia and suppresses appetite in men with type 2 diabetes: a randomized controlled trial

Large doses of whey protein consumed as a preload before single high-glycemic load meals has been shown to improve postprandial glycemia in type 2 diabetes. It is unclear if this effect remains with smaller doses of whey co-ingested at consecutive mixed-macronutrient meals. Moreover, whether hydrolyzed whey offers further benefit under these conditions is unclear. The aim of this study was to investigate postprandial glycemic and appetite responses after small doses of intact and hydrolyzed whey protein co-ingested with mixed-nutrient breakfast and lunch meals in men with type 2 diabetes. In a randomized, single-blind crossover design, 11 men with type 2 diabetes [mean±SD age: 54.9±2.3 y; glycated hemoglobin: 6.8%±0.3% (51.3±3.4 mmol/mol)] attended the laboratory on 3 mornings and consumed 1) intact whey protein (15 g), 2) hydrolyzed whey protein (15 g), or 3) placebo (control) immediately before mixed-macronutrient breakfast and lunch meals, separated by 3 h. Blood samples were collected periodically and were processed for insulin, intact glucagon-like peptide 1 (GLP-1), gastric inhibitory polypeptide (GIP), leptin, peptide tyrosine tyrosine (PYY3-36), and amino acid concentrations. Interstitial glucose was measured during and for 24 h after each trial. Subjective appetite was assessed with the use of visual analog scales. Total postprandial glycemia area under the curve was reduced by 13%±3% after breakfast following the intact whey protein when compared with control (P<0.05). Hydrolyzed whey attenuated early glucose after breakfast when compared with control (P<0.05). Glycemia was improved postlunch after the intact whey protein only when compared with control (P<0.05). Greater satiety was observed after the intact whey protein only after both meals when compared with control (P<0.05). Insulin concentrations increased after both the intact and hydrolyzed whey protein, showing strong positive correlations with increases in valine and isoleucine (P<0.05). Incretin and appetite regulatory hormone responses were similar across trials (P>0.05). The consumption of a small 15-g dose of intact whey protein immediately before consecutive mixed-macronutrient meals improves postprandial glycemia, stimulates insulin release, and increases satiety in men with type 2 diabetes. This trial was registered at www.clinicialtrials.gov as NCT02903199.

Growth, Gastrointestinal Tolerance and Stool Characteristics of Healthy Term Infants Fed an Infant Formula Containing Hydrolyzed Whey Protein (63%) and Intact Casein (37%): A Randomized Clinical Trial.

To investigate whether healthy term infants, fed an infant formula containing hydrolyzed whey protein (HWP-F, hydrolyzed whey/intact casein =63/37), differ in growth, gastrointestinal tolerance and stool characteristics from those fed an infant formula containing intact whey protein (IWP-F, intact whey/intact casein =61/39) or breast milk. Healthy term infants, born within 14 days of the study’s commencement, were randomly assigned to be fed IWP-F or HWP-F until 13 weeks of age, and breast-fed (BF) infants were enrolled as a reference group. Anthropometric measurements, gastrointestinal tolerance indexes and stool characteristics were assessed at baseline, and 7 and 13 weeks of age. There were no significant differences in any growth measurements and the occurrence of crying, spit-up and difficult defecation among the three feeding groups during the study period. However, daily feeding frequency was consistently lower in the formula-fed infants than in the BF group throughout the study (p < 0.05), and infants in the HWP-F group consumed more formula than those in the IWP-F group at 7 and 13 weeks of age (p ≤ 0.002). The HWP-F-fed infants had more similar stool characteristics to the breast-fed infants than infants in the IWP-F group at 13 weeks of age, regardless of frequency, volume, color or consistency of stool. This study demonstrates that the HWP-F could support the normal growth of healthy term infants, to a comparable extent to that of breast-fed infants during the first three months of life. Moreover, stool characteristics of HWP-F-fed infants are much closer to breast-fed infants than IWP-F-fed infants, but no significant gastrointestinal tolerance improvement was observed in HWP-F group.