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peer-reviewed Membrane separation processes used in the concentration and isolation of micellar casein-based milk proteins from skim milk rely on extensive permeation of its soluble serum constituents, especially lactose and minerals. Whereas extensive literature exists on how these processes influence the gross composition of milk proteins, we have little understanding of the effects of such ionic depletion on the core structural unit of micellar casein [i.e., the casein phosphate nanocluster (CPN)]. The 31P nuclear magnetic resonance (NMR) is an analytical technique that is capable of identifying soluble and organic forms of phosphate in milk. Thus, our objective was to investigate changes to the 31P NMR spectra of skim milk during microfiltration (MF) and diafiltration (DF) by tracking movements in different species of phosphate. In particular, we examined the peak at 1.11 ppm corresponding to inorganic phosphate in the serum, as well as the low-intensity broad signal between 1.5 and 3.0 ppm attributed to casein-associated phosphate in the retentate. The MF concentration and DF using water caused a shift in the relevant 31P NMR peak that could be minimized if orthophosphate was added to the DF water. However, this did not resolve the simultaneous change in retentate pH and increased solubilization of micellar casein protein. The addition of calcium in combination with orthophosphate prevented micellar casein solubilization and simultaneously contributed to preservation of the CPN structure, except for overcorrection of retentate pH in the acidic direction. A more complex DF solution, involving a combination of phosphate, calcium, and citrate, succeeded in both CPN and micellar casein structure preservation while maintaining retentate pH in the region of the original milk pH. The combination of 31P NMR as an analytical technique and experimental probe during MF/DF processes provided useful insights into changes occurring to CPN while retaining the micellar state of casein.

The Nitrogen solubility, turbidity, emulsion and foaming properties, and the heat stability of a brewers' spent grain protein-enriched isolate (BSG-PI) was studied between pH 2.0–12.0. The effect of enzymatic hydrolysis using Alcalase, Corolase PP, Flavourzyme and Promod 144MG on these properties was also determined. BSG-PI demonstrated low solubility below pH 6.0, however, hydrolysis with all enzymes significantly increased BSG-PI solubility between pH 2.0–12.0. Flavourzyme, Corolase PP and Promod 144MG hydrolysates displayed an improved emulsion activity index (EAI) with an EAI of 466.69 m2g−1 obtained for the Flavourzyme hydrolysates at pH 9.0. Improved foam expansion (FE) was observed above pH 9.0 in all samples while hydrolysis significantly reduced foam drainage stability (FS) of BSG-PI. BSG-PI was highly heat stable at 140 °C above pH 6.0 even after >300 min heating. Hydrolysis of BSG-PI resulted in increased turbidity when aqueous suspensions were stored overnight at 4 or 20 °C. The results demonstrate the potential of the alkaline extracted BSG-PI and its associated hydrolysates as a cereal derived source of technofunctional ingredients for the food industry.

This study investigates the ability of milk protein-derived peptides to specifically activate the serotonin 2C (5-HT2C) receptor, a key receptor in central regulation of food intake. A dose dependent 5-HT2C receptor activation by the 1 kDa ultrafiltration permeates of a sodium caseinate (NaCNH-1 kDa permeate) and a whey protein hydrolysate (WPH-1 kDa permeate) was demonstrated using an intracellular calcium mobilisation assay in human embryonic kidney (Hek) cells expressing the 5-HT2C receptor. Both samples activated the 5-HT2C but not the 5-HT2A and 5-HT2B receptors. NaCNH-1 kDa permeate significantly (p

BACKGROUND Dietary mineral deficiency, hypertension and diabetes have become serious human health problems. Dietary approaches are increasingly being investigated to address these issues. Identification of food-derived biological peptides has become an important approach to control such diseases. Peptides generated from aquatic byproducts have been shown to possess biological activities. RESULTS Significantly higher copper-chelating activity was observed on simulated hydrolysis of intact collagen. The collagen hydrolysate generated in the gastric stage exhibited moderate angiotensin-converting enzyme (ACE)-inhibitory activity with an IC50 value of 2.92 ± 0.22 mg mL−1, which significantly decreased to 0.49 ± 0.02 mg mL−1 after intestinal digestion. The dipeptidyl peptidase (DPP) IV-inhibitory potency of the collagen hydrolysate generated directly following simulated gastrointestinal digestion (SGID) (IC50 2.59 ± 0.04 mg mL−1) was significantly lower than that of the collagen tryptic hydrolysate (CTH) (IC50 1.53 ± 0.01 mg mL−1). The antioxidant activities of collagen and CTH using the ferric-reducing antioxidant power (FRAP) assay were 0.87 ± 0.10 and 1.27 ± 0.03 µmol Trolox equivalent (TE) g−1 respectively after SGID. CONCLUSION This study identifies collagen as a good and inexpensive substrate for the generation of biologically active peptides with potential applications as functional ingredients in the management of chronic illness and mineral deficiency problems. © 2014 Society of Chemical Industry

The effect of starting protein composition, and the origin and time of harvesting on the in vitro tyrosinase, dipeptidyl peptidase (DPP) IV and angiotensin converting enzyme (ACE) inhibitory and antioxidant activity of Palmaria palmata protein hydrolysates were investigated. Electrophoretic profiles showed significant differences in aqueous protein extracts obtained from the red macroalga Palmaria palmata harvested at different times of the year. No significant difference was observed in aqueous protein profiles extracted from wild and aquacultured samples of Palmaria palmata. Protein extracts from Palmaria palmata samples harvested from wild plants in April, July and October, and samples cultivated on longlines and harvested in April were hydrolysed with Alcalase 2.4 L and Corolase PP. The hydrolysates, when tested at 10 mg/ml, were shown to inhibit tyrosinase by 37–56%. The oxygen radical absorbance capacity (ORAC) and ferric reducing antioxidant power (FRAP) values of the hydrolysates ranged from 323–494 and 8.9–19.9 μmol trolox equivalents per gram, respectively. The Palmaria palmata hydrolysates inhibited DPP-IV (IC50: 1.60–4.24 mg/mL) and ACE (IC50: 0.14–0.35 mg/mL) activities. The starting protein composition had a significant effect on the tyrosinase inhibitory activity, while protein composition and hydrolytic enzyme preparation had a significant effect on DPP-IV inhibitory and antioxidant activities. In general, the origin of the samples, wild or cultivated, had no effect on the in vitro biological activity of the protein hydrolysates. The results show that Palmaria palmata hydrolysates may have potential applications as health enhancing ingredients and as food preservatives due to their antioxidant and tyrosinase inhibitory effects.

Whey proteins are rich sources of bioactive peptides which may play a role in the dietary management of chronic diseases. Fractionation via ultrafiltration (UF) was investigated for the enrichment of antioxidant, dipeptidyl peptidase IV (DPP-IV) and angiotensin converting enzyme (ACE) inhibitory activity in a tryptic hydrolysate of β-lactoglobulin (β-Lg TH). UF processing selectively enhanced the biofunctional properties of β-Lg TH with the permeate obtained using a 1 kDa polyethersulfone (HFP-1) membrane having highest in vitro multifunctional bioactivity. Compared to β-Lg TH, the antioxidant activity was 1.7-fold higher (46,765 ± 2504 vs. 77,251 ± 5124 µmol Trolox equivalent/100 g dw; P

BACKGROUND The use of Palmaria palmata (PP) as a natural ingredient in farmed Atlantic salmon diets was investigated. The effect of salmon diet supplementation with P. palmata (0, 5, 10 and 15%) or synthetic astaxanthin (positive control, PC) for 16 weeks pre-slaughter on quality indices of fresh salmon fillets was examined. The susceptibility of salmon fillets/homogenates to oxidative stress conditions was also measured. RESULTS In salmon fillets stored in modified atmosphere packs (60% N2/40% CO2) for up to 15 days at 4 °C, P. palmata increased surface −a* (greenness) and b* (yellowness) values in a dose-dependent manner, resulting in a final yellow/orange flesh colour. In general, the dietary addition of P. palmata had no effect on pH, lipid oxidation (fresh, cooked and fillet homogenates) and microbiological status. ‘Eating quality’ sensory descriptors (texture, odour and oxidation flavour) in cooked salmon fillets were not influenced by dietary P. palmata. Salmon fed 5% PP showed increased overall acceptability compared with those fed PC and 0% PP. CONCLUSION Dietary P. palmata was ineffective at providing red coloration in salmon fillets, but pigment deposition enhanced fillets with a yellow/orange colour. Carotenoids from P. palmata may prove to be a natural pigment alternative to canthaxanthin in salmon feeds. © 2014 Society of Chemical Industry

Enzymatic hydrolysis of whey protein concentrate (WPC) was performed using Alcalase, Neutrase, Corolase PP or Flavourzyme. The hydrolysates were filtered through membranes having different molecular mass cut-offs. The hydrolysate fractions had significantly (P

The next generation sequencing revolution has enabled rapid discovery of genetic markers, however, development of fully functioning new markers still requires a long and costly process of marker validation. This study reports a rapid and economical approach for the validation and deployment of polymorphic microsatellite markers obtained from a 454 pyrosequencing library of Atlantic cod, Gadus morhua, Linnaeus 1758. Primers were designed from raw reads to amplify specific amplicon size ranges, allowing effective PCR multiplexing. Multiplexing was combined with a three-primer PCR approach using four universal tails to label amplicons with separate fluorochromes. A total of 192 primer pairs were tested, resulting in 73 polymorphic markers. Of these, 55 loci were combined in six multiplex panels each containing between six and eleven markers. Variability of the loci was assessed on G. morhua from the Celtic Sea (n = 46) and the Scotian Shelf (n = 46), two locations that have shown genetic differentiation in previous studies. Multilocus F(ST) between the two samples was estimated at 0.067 (P = 0.001). After three loci potentially under selection were excluded, the global F(ST) was estimated at 0.043 (P = 0.001). Our technique combines three-primer and multiplex PCR techniques, allowing simultaneous screening and validation of relatively large numbers of microsatellite loci.

Minerals such as calcium, zinc, iron and copper are important elements for human health. Deficiencies of dietary minerals can lead to numerous diseases. Mineral chelating peptides have shown potential application in the management of mineral deficiencies. An increasing number of chelating peptides with the ability to facilitate and enhance the bioavailability of minerals are being discovered and identified. This review outlines the current food protein sources, analytical methods and the purification schemes of mineral chelating peptides, and discusses their structure–activity relationship and the bioavailability. The potential of mineral chelating peptides as functional food ingredients is also described.

Total phenolic content (TPC) and antioxidant activity of crude and enriched aqueous, ethanol/water and methanol/water extracts from four species of Irish macroalgae Laminaria digitata, Fucus serratus, Gracilaria gracilis and Codium fragile were assessed. The antioxidant activity and TPC of crude and enriched extracts were assessed using the DPPH, FRAP and Folin-Ciocalteu assays. Extracts of F. serratus were significantly higher (p 100 kDa) and hydrophobic with a significant increase in antioxidant activity and TPC (p

Summary Two types of brewers' spent grain (BSG), pale and black, were studied employing ultrafiltration (UF cut-off, 10 kDa) to obtain high-molecular-weight (HMW) and low-molecular-weight (LMW) phenolic fractions from one pale (P2) and one black (B2) BSG extracts. Of the four UF fractions obtained, the HMW B2 fraction had highest level of polyphenols (5.73 ± 0.25 mg GAE g−1 BSG dw) and protein (18.82 ± 0.41 mg protein g−1 BSG dw). Metal-chelating ability and antioxidative properties were also identified. Antioxidant activity evaluated using the DPPH and FRAP assays, indicated that activity in B2 extracts was associated mainly with the HMW fraction (3.10 ± 0.10 and 2.49 ± 0.09 TE g−1 BSG dw, respectively), whereas in P2 samples, antioxidant activity was highest in LMW fractions (0.58 ± 0.01 and 0.92 ± 0.03 TE g−1 BSG dw, respectively). The high antioxidant activity and metal-chelating ability observed for black BSG extracts may be attributed to the presence of coloured melanoidinic structures.

Summary The efficiency of solid–liquid extraction (SLE) and pressurised liquid extraction (PLE) for the recovery of antioxidant and polyphenols from the Irish macroalgae, Fucus serratus, Laminaria digitata, Gracilaria gracilis and Codium fragile, was assessed using the 2,2-diphenyl-1-picrylhydrazyl (DPPH) and ferric reducing antioxidant power (FRAP) assays and the Folin–Ciocalteu total phenol content (TPC) assay. Fucus serratus had TPC and antioxidant activities thirty times higher than the other species. Solid–liquid extraction cold water (CWSLE) had the highest TPC (81.17 μg GAE mg−1 sample) derived from F. serratus, compared with the TPC of 61.12 μg GAE mg−1 sample for the corresponding PLE extract. For both SLE and PLE extracts, low TPC levels were observed in L. digitata, G. gracilis and C. fragile. The majority of SLE extracts possessed higher FRAP and DPPH activities compared with their PLE counterparts. This study indicates that the high temperatures and pressures in PLE did not enhance the antioxidant activities relative to conventional SLE extraction.

The in vitro α-glucosidase, α-amylase, dipeptidyl peptidase IV (DPP-IV) and angiotensin converting enzyme (ACE) inhibitory activities of pale brewers' spent grain protein enriched isolate (BSG-PI) hydrolysates were studied. Eleven commercially available enzyme preparations derived from papaya, porcine pancreas, Aspergillus oryzae and Bacillus licheniformis were used to generate the hydrolysates. The hydrolysates had degree hydrolysis (DH) values ranging from 1.15 to 14.67% and the DH values obtained correlated well with hydrolysate molecular mass distribution profiles. Reverse-phase HPLC analysis of the BSG-PI and its hydrolysates demonstrated considerable variation in peptide composition. While tryptic digests of BSG-PI resulted in the greatest α-glucosidase inhibitory activity, no significant change in α-amylase inhibition was observed with the hydrolysates. Digestion with Corolase PP resulted in the highest DPP-IV inhibition (75 ± 3.06% at 3.5 mg mL− 1) while the Prolyve 1000 hydrolysate displayed highest ACE inhibition (89.25 ± 2.46% at 1 mg mL− 1). The results obtained highlight the potential role of the BSG-PI hydrolysates as functional food ingredients in the management of type II diabetes and hypertension.

peer-reviewed Whey protein isolate (WPI) solutions (100 g L−1 protein) were subjected to a heat-treatment of 80 °C for 10 min. Unheated and heat-treated WPI solutions were hydrolysed with Corolase® PP at pilot-scale to either 5 or 10% degree of hydrolysis (DH). Hydrolysates were subsequently processed via cascade membrane fractionation using 0.14 μm, and 30, 10, 5 and 1 kDa cut-off membranes. The compositional and molecular mass distribution profiles of the substrate hydrolysates and membrane processed fractions were determined. Whole and fractionated hydrolysates were assayed for both angiotensin-I-converting enzyme (ACE) inhibitory activity and ferrous chelating capabilities. A strong positive correlation (P < 0.01) was established between the average molecular mass of the test samples and the concentration needed to chelate 50% of the iron (CC50) in solution. The lowest ACE inhibition concentration (IC50 = 0.23 g L−1 protein) was determined for the 1 kDa permeate of the heat-treated 10% DH hydrolysate The work herein was funded by Enterprise Ireland as part of the Food for Health Ireland project, grant number; CC20080001. I. B. O’Loughlin was funded by Enterprise Ireland under the Teagasc Walsh Fellowship Scheme.

Summary Marine minerals are a potential source of calcium and magnesium for nutritional supplementation. This study analysed the solubilisation of calcium and magnesium from the skeletal remains of Lithothamnion calcareum. Scanning electron microscopy analysis demonstrated a nonporous microstructure. Spectrophotometric determination showed that the calcium and magnesium contents were 30.01 and 6.22% (w/w), respectively. Solubilisation of calcium and magnesium was highly pH dependent. The temperature-dependent solubilisation of calcium fitted the shrinking core model. The apparent activation energy for calcium solubilisation was 28.6 kJ mol−1. Inclusion of caseinophosphopeptides (CPPs), casein-derived mineral binding peptides, during the solubilisation of calcium and magnesium appeared to decrease the extent of calcium solubilisation at pH 6.0 and 8.0. The results herein have implications for the choice of optimal pH conditions for the sustained release of calcium and magnesium from marine mineral sources.

The hydroxycinnamic acid (HA) content of starting barley for brewers' spent grains (BSG), whole BSG and phenolic extracts from BSG was measured using high performance liquid chromatography (HPLC) and correlated with antioxidant potential. The effect of BSG phenolic extracts on antioxidant activity of fruit beverages was also assessed (using the total phenolic content (TPC), 2,2-diphenyl-1-picrylhydrazyl (DPPH) and ferric reducing antioxidant power (FRAP) assays). The concentration of HA present in barley extract and BSG was in the order of ferulic acid (FA), p-coumaric acid (p-CA) derivatives, FA derivatives, p-CA, caffeic acid (CA) and CA derivatives. Results suggested that brewing and roasting decreased the HA content. Antioxidant activity was significantly (P0.05) correlated with caffeic acid (R(2)=0.8309) and total HA (R(2)=0.3942) concentrations. Addition of extracts to fruit beverages resulted in a significant (P0.05) increase in antioxidant activity of cranberry juice, measured by the FRAP assay. In vitro digestion significantly (P0.05) reduced TPC, DPPH and FRAP activity of the fruit beverages.

Peer-reviewed. This article may be used for non-commercial purposes in accordance with Wiley Terms and Conditions for Self-Archiving., EIRCOD project, funded under the Sea Change initiative with the support of the Marine Institute and the Marine Research Sub-Programme of the National Development Plan 2007–2013 and co-funded by the European Regional Development Fund.

Xanthine oxidase (XO) and dipeptidyl peptidase IV (DPP-IV) inhibition by amino acids and dipeptides was studied. Trp and Trp-containing dipeptides (Arg-Trp, Trp-Val, Val-Trp, Lys-Trp and Ile-Trp) inhibited XO. Three amino acids (Met, Leu and Trp) and eight dipeptides (Phe-Leu, Trp-Val, His-Leu, Glu-Lys, Ala-Leu, Val-Ala, Ser-Leu and Gly-Leu) inhibited DPP-IV. Trp and Trp-Val were multifunctional inhibitors of XO and DPP-IV. Lineweaver and Burk analysis showed that Trp was a non-competitive inhibitor of XO and a competitive inhibitor of DPP-IV. Molecular docking with Autodock Vina was used to better understand the interaction of the peptides with the active site of the enzyme. Because of the non-competitive inhibition observed, docking of Trp-Val to the secondary binding sites of XO and DPP-IV is required. Trp-Val was predicted to be intestinally neutral (between 25% and 75% peptide remaining after 60 min simulated intestinal digestion). These results are of significance for the reduction of reactive oxygen species (ROS) and the increase of the half-life of incretins by food-derived peptides.

Sodium caseinate (NaCN) was hydrolysed with Prolyve 1000, a Bacillus licheniformis derived proteolytic preparation to a degree of hydrolysis of 21.57 ± 1.2%. An aliquot of this hydrolysate was cross-linked with transglutaminase (TGase). Furthermore, TGase cross-linked NaCN was also hydrolysed with Prolyve 1000. These three hydrolysate samples had broadly similar peptide and molecular mass distribution profiles while the reduction in residual antigenicity was not statistically significant ( P > 0.05) between the hydrolysates. Jurkat T cell viability studies showed that the hydrolysate cross-linked prior to hydrolysis was the most cytotoxic of the three hydrolysates studied. There was no significant difference ( P > 0.05) between the hydrolysates in catalase activity and glutathione content in Jurkat T cells following 24 h incubation. However, the production of interleukin-2 and interleukin-10 in Con-A stimulated Jurkat T cells was significantly decreased ( P

BACKGROUND Large quantities of brewers' spent grain (BSG), a co-product of the brewing industry, are produced annually. BSG contains hydroxycinnamic acids, and phenolic-rich extracts from BSG have previously demonstrated the ability to protect against oxidant-induced DNA damage. The present study investigated the anti-inflammatory potential of eight phenolic extracts from BSG: four pale (P1–P4) and four black (B1–B4) extracts. RESULTS BSG extracts were more cytotoxic in Jurkat T than U937 cells, with lower IC50 values in Jurkat T cells, measured using the (3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide (MTT) assay. Pale BSG extracts P2 and P3 showed the greatest anti-inflammatory potential, significantly (P

The prevalence of type 2 diabetes mellitus (T2DM) is increasing and it is estimated that by 2030 approximately 366 million people will be diagnosed with this condition. The use of dipeptidyl peptidase IV (DPP-IV) inhibitors is an emerging strategy for the treatment of T2DM. DPP-IV is a ubiquitous aminodipeptidase that cleaves incretins such as glucagon like peptide 1 (GLP-1) and glucose-dependent insulinotropic polypeptide (GIP), resulting in a loss in their insulinotropic activity. Synthetic DPP-IV drug inhibitors are being used to increase the half-life of the active GLP-1 and GIP. Dietary intervention is accepted as a key component in the prevention and management of T2DM. Therefore, identification of natural food protein-derived DPP-IV inhibitors is desirable. Peptides with DPP-IV inhibitory activity have been identified in a variety of food proteins. This review aims to provide an overview of food protein hydrolysates as a source of the DPP-IV inhibitory peptides with particular focus on milk proteins. In addition, the proposed modes of inhibition and structure–activity relationship of peptide inhibitors are discussed. Milk proteins and associated peptides also display insulinotropic activity and help regulate blood glucose in healthy and diabetic subjects. Therefore, milk protein derived peptide inhibitors may be a unique multifunctional peptide approach for the management of T2DM.

The kinetics and thermodynamics of the thermal inactivation of Corolase PP in two different whey protein concentrate (WPC) hydrolysates with degree of hydrolysis (DH) values of ~10 and 21%, and at different total solids (TS) levels (from 5 to 30% w/v), were studied. Inactivation studies were performed in the temperature range from 60 to 75 °C, and residual enzyme activity was quantified using the azocasein assay. The inactivation kinetics followed a first-order model. Analysis of the activation energy, thermodynamic parameters, and D and z values, demonstrated that the inactivation of Corolase PP was dependent on solution TS. The intestinal enzyme preparation was more heat sensitive at low TS. Moreover, it was also found that the enzyme was more heat sensitive in solutions at higher DH.

peer-reviewed The influence of different substrates and the effect of pH regulation during enzymatic hydrolysis of whey protein on glucose-stimulated insulin secretion by BRIN-BD11 pancreatic beta cells were studied. No significant differences (P >= 0.05) were detected in terms of glucose-stimulated insulin secretion by BRIN-BD11 pancreatic beta cells exposed to whey protein hydrolysates (WPH1 and WPH2) obtained with two different whey protein substrates. The whey protein hydrolysate (WPH3) obtained without pH regulation during hydrolysis, had a significantly lower insulinotropic potential in BRIN-BD11 cells than the WPH1 hydrolysate that was manufactured with pH regulation. Fractionation of WPH1 was carried out to enrich bioactive peptides. Comparing the different fractionation techniques studied (solid-phase extraction and semi-preparative reverse phase-high performance liquid chromatography), the most potent insulinotropic fraction was enriched in free amino acids and contained relatively hydrophilic peptides. This indicates that amino acids and hydrophilic peptides may be involved in the insulinotropic effect of WPH1. (C) 2013 Elsevier Ltd. All rights reserved. ACCEPTED peer-reviewed

Dipeptides with a C terminal Pro inhibit dipeptidyl peptidase IV (DPP-IV), a key enzyme in incretin hormone processing. It was hypothesised that tri- and tetrapeptides with a proline at the C-terminus may also be DPP-IV inhibitors. Therefore, an in silico hydrolysis approach was used to release short (4 ⩽ amino acids) C terminal Pro peptides from the individual caseins which constitute Pro rich substrates. This was achieved using theoretical digestion of caseins with a prolyl oligopeptidase activity. Fifteen peptides were subsequently selected for in vitro DPP-IV inhibitory analysis. Stability of these peptides to gastrointestinal enzymes was also evaluated in silico and the predicted breakdown peptides were assessed for their DPP-IV inhibitory and antioxidant potential. New DPP-IV inhibitors were identified, the most potent being Phe-Leu-Gln-Pro (IC50 65.3 ± 3.5 μM). A low in vitro antioxidant (2,2-diphenyl-1-picrylhydrazyl (DPPH) scavenging) activity was also associated with the peptides studied. The strategy presented highlights the utility of employing an in silico approach for the prediction of food-derived peptides with a potential role in glycaemic management for subsequent development of functional foods.

The in vitro dipeptidyl peptidase IV (DPP-IV) inhibitory activity of a whey protein hydrolysate (WPH) generated with a food-grade pancreatic enzyme preparation was studied. The 50% inhibitory concentration (IC50) value in the presence of WPH was 1.34 ± 0.11 mg mL−1. Ultrafiltration (UF) fractionation of WPH allowed enrichment in DPP-IV inhibitory peptides. The permeates generated by UF through 5 and 2 kDa membranes along with the hydrophilic fraction isolated by solid-phase extraction were significantly more potent DPP-IV inhibitors than WPH. Simulated gastrointestinal digestion of WPH resulted in an increased DPP-IV inhibitory potency (IC50 value of 1.02 ± 0.05 mg mL−1). Competitive inhibition of DPP-IV was observed with WPH and all its fractions, indicating a direct interaction of the bioactive peptides therein with the active site of DPP-IV. Combinations of sitagliptin, a conventional drug-inhibitor of DPP-IV, and whey-derived peptides resulted in an additive effect on DPP-IV inhibition.

BACKGROUND Membrane separation technologies seem to be one of the best available techniques to fractionate protein hydrolysates containing bioactive peptides and obtain peptide fractions with increased functionality. The influence of the molecular weight cut-off (MWCO) of the membrane and its hydrophilic/hydrophobic characteristics on the fractionation of a β-lactoglobulin tryptic hydrolysate were studied in this research work in order to obtain more purified peptidic fractions. RESULTS The molecular weight of the solutes, their charge and, ultimately, their hydrophobicity influenced peptide transmission. As regards the composition of the active layer of the membrane, high permeate flux values were maintained throughout the pH working range (from pH 4.0 to pH 10.0) when an extremely hydrophilic membrane was employed, even in the presence of peptide aggregates at acidic pH values. In addition, the decrease in the MWCO of the membrane (from 5 kDa to 1 kDa) did not improve the selectivity of the fractionation process, however, it assumed a significant decrease of the permeate flux through the membrane. CONCLUSION The low efficiency of the process in terms of peptide recovery and the poor separation observed between bioactive and non-bioactive peptides work against the use of extremely hydrophilic membranes to fractionate peptide hydrolysates. © 2013 Society of Chemical Industry

The use of herbs, spices, and whey proteins as natural alternatives to current flavor enhancers in reduced-salt ready-meals was investigated. Individual addition of garlic, rosemary, oregano, and sage at varying concentrations (0.1–0.5%) significantly improved the acceptability of each ready-meal and thus they are recommended as natural flavor enhancers in similar formulations (p < 0.05). However, both clove and pimento proved unacceptable as individual additions to ready-meals and therefore are not advised for individual use. Acceptability tests were subsequently utilized to formulate spice blends for inclusion in ready-meals. Incorporation of a spice blend consisting of garlic, rosemary, oregano, sage, and pimento in cottage pie and a blend of garlic, rosemary, and sage in chicken supreme significantly increased their acceptability (p < 0.05) and hence they are recommended for use in comparable formulations. Two lactoferrin whey proteins significantly increased the mean acceptability for each ready-meal...

Whey protein hydrolysates (WPHs) represent novel antidiabetic agents that affect glycemia in animals and humans, but little is known about their insulinotropic effects. The effects of a WPH were analyzed in vitro on acute glucose-induced insulin secretion in pancreatic BRIN-BD11 β cells. WPH permeability across Caco-2 cell monolayers was determined in a 2-tiered intestinal model. WPH effects on insulin resistance were studied in vivo following an 8-wk oral ingestion (100 mg/kg body weight) by ob/ob (OB-WPH) and wild-type mice (WT-WPH) compared with vehicle control (OB and WT groups) using a 2 × 2 factorial design, genotype × treatment. BRIN-BD11 cells showed a robust and reproducible dose-dependent insulinotropic effect of WPH (from 0.01 to 5.00 g/L). WPH bioactive constituents were permeable across Caco-2 cell monolayers. In the OB-WPH and WT-WPH groups, WPH administration improved glucose clearance after a glucose challenge (2 g/kg body weight), as indicated by differences in the area under curves (AUCs) (P ≤ 0.05). The basal plasma glucose concentration was not affected by WPH treatment in either genotype. The plasma insulin concentration was lower in the OB-WPH than in the OB group (P ≤ 0.005) but was similar between the WT and WT-WPH groups; the interaction genotype × treatment was significant (P ≤ 0.005). Insulin release from pancreatic islets isolated from the OB-WPH group was greater (P ≤ 0.005) than that from the OB group but did not differ between the WT-WPH and WT groups; the interaction genotype × treatment was not significant. In conclusion, an 8-wk oral administration of WPH improved blood glucose clearance, reduced hyperinsulinemia, and restored the pancreatic islet capacity to secrete insulin in response to glucose in ob/ob mice. Hence, it may be useful in diabetes management.

Four size groups of juvenile farmed Celtic Sea cod (2.7–41.8 g) were reared at a range of constant temperatures (8–19°C). The optimum temperature for growth (Topt.G) decreased from 15.1°C for ~3 g fish to 12.5°C for ~42 g fish. A comparison of these results with those published for a more northerly population (Icelandic) suggests that there is no significant difference in the optimal temperature for growth of cod stocks within the size range studied. In contrast, the growth rates of Celtic Sea cod were lower than those derived from these established models (for a northerly stock) for small juveniles ( 40 g). Thus, while Topt.G appears fixed across the range, there may be high plasticity in local growth performance throughout the Holarctic distribution. Some possible explanations for these differences are considered.

Summary Protein-enriched isolates and co-product fractions were obtained from sheared, pale and black brewers' spent grain (BSG) using sequential aqueous and alkaline (110 mm NaOH) extraction, followed by isoelectric precipitation at pH 3.8. A recovery of 59% of the original pale BSG protein and 15% of the black BSG protein was obtained for the final isolates. Gel permeation HPLC (GP-HPLC) revealed that 59% of the extracted pale BSG protein and only 6% of black BSG protein had a molecular mass >10 kDa. Glutamine/glutamate and proline were the most abundant amino acids present in both isolates. Analysis of four co-product fractions obtained during fractionation from both pale and black BSG revealed the presence of phenolics, with higher concentrations in the black BSG extracts. These fractions possessed antioxidant and free radical scavenging activity when tested using the ferric reducing ability of plasma (0.16 ± 0.01 to 4.33 ± 0.11 mg Trolox equivalents g−1 BSG dry weight) and diphenylpicrylhydrazyl (12.85 ± 1.16% to 59.50 ± 3.47% DPPHsc) assays, respectively. The protein-enriched isolates and the phenolic-rich extracts may find use as value-added ingredients for incorporation into conventional and functional foods.

The contribution of protein fraction and proteolytic enzyme preparation to the in vitro cardioprotective, anti-diabetic and antioxidant activity of Palmaria palmata protein hydrolysates was investigated. Aqueous, alkaline and combined aqueous and alkaline P. palmata protein fractions were hydrolysed with the food-grade proteolytic preparations, Alcalase 2.4 L, Flavourzyme 500 L and Corolase PP. The hydrolysates had angiotensin converting enzyme (ACE) and dipeptidyl peptidase (DPP) IV inhibitory activity with IC50 values in the range 0.19–0.78 and 1.65–4.60 mg mL−1, respectively. The oxygen radical absorbance capacity (ORAC) and ferric reducing antioxidant power (FRAP) values ranged from 45.17 to 467.54 and from 1.06 to 21.59 μmol trolox equivalents/g, respectively. Furthermore, hydrolysates (1 mg mL−1) were show to inhibit renin within the range 0–50 %. In general, Alcalase 2.4 L and Corolase PP hydrolysates of aqueous protein displayed the highest in vitro activity. The results indicate that protein fraction and enzyme preparation used have significant effects on in vitro biofunctional activity of the hydrolysates. This study demonstrates the potential of P. palmata protein hydrolysates as multifunctional functional food ingredients for the prevention/control of hypertension and type II diabetes.

14 páginas, 3 figuras, 3 tablas.-- Frederico Almada ... et al.-- Published by the Royal Society under the terms of the Creative Commons Attribution License http://creativecommons.org/licenses/by/4.0/, which permits unrestricted use, provided the original author and source are credited, The distribution and demographic patterns of marine organisms in the north Atlantic were largely shaped by climatic changes during the Pleistocene, when recurrent glacial maxima forced them to move south or to survive in northern peri-glacial refugia. These patterns were also influenced by biological and ecological factors intrinsic to each species, namely their dispersion ability. The ballan wrasse (Labrus bergylta), the largest labrid fish along Europe's continental margins, is a target for fisheries and aquaculture industry. The phylogeographic pattern, population structure, potential glacial refugia and recolonization routes for this species were assessed across its full distribution range, using mitochondrial and nuclear markers. The existence of a marked population structure can reflect both recolonization from three distinct glacial refugia and current and past oceanographic circulation patterns. Although isolated in present times, shared haplotypes between continental and Azores populations and historical exchange of migrants in both directions point to a common origin of L. bergylta. This situation is likely to be maintained and/or accentuated by current circulation patterns in the north Atlantic, and may lead to incipient speciation in the already distinct Azorean population. Future monitoring of this species is crucial to evaluate how this species is coping with current environmental changes, This study was funded by the Eco-Ethology Research Unit Strategic Plan (PEst-OE/MAR/UI0331/2011)—Fundação para a Ciência e a Tecnologia—FCT (partially FEDER funded), now included in MARE (UID/MAR/04292/2013). F.A. (SFRH/BPD/63170/2009) and S.M.F. (SFRH/BPD/84923/2012) were supported by FCT grants. C.S.L. was supported by MARE-ISPA/BI/004/2015. D.V.-R. was supported by a Marie Curie Intra European Fellowship within the 7th European Community Framework Programme (grant no. 625852). Sampling in the Azores was supported by CLIPE (PRAXIS/3/3.2/EMG/1957/95) and in continental Iberia by the European Science Foundation's MarinERA project ‘Marine phylogeographic structuring during climate change: the signature of leading and rear edge of range shifting populations'

peer-reviewed Wheat gluten, a Pro-rich dietary protein, was investigated for its potential to produce dipeptidyl peptidase IV (DPP-IV) inhibitory peptides during enzymatic hydrolysis with Debitrase HYW20. Nine gluten hydrolysates (H1–H9) were generated using a 2 factor × 3 level design of experiments (DOE) including the incubation temperature (40, 50 and 60 °C) and the enzyme: substrate ratio (E:S, 0.5, 1.0 and 1.5% (w/w)). Their DPP-IV half maximal inhibitory concentration (IC50) ranged from 0.24 ± 0.02 (H9) to 0.66 ± 0.06 mg mL−1 (H2A and H7) and their degree of hydrolysis (DH) from 31.7 ± 0.9 (H7) to 62.2 ± 3.0% (H6). Gluten and H9, the most potent DPP-IV inhibitory hydrolysate, were subjected to simulated gastrointestinal digestion (SGID), yielding Gluten_CorPP and H9_CorPP, respectively. H9_CorPP had a higher DPP-IV inhibitory potency than Gluten_CorPP (i.e., DPP-IV IC50 values of 0.33 ± 0.03 vs. 1.45 ± 0.26 mg mL−1, respectively). H9 and H9_CorPP both contained relatively potent DPP-IV inhibitory peptides such as Val-Pro-Leu, Trp-Leu and Trp-Pro which were identified by liquid chromatography tandem mass spectrometry (LC-MS/MS). In addition, several sequences possessing features of DPP-IV inhibitory peptides, mostly consisting of a penultimate or C-terminal Pro, were identified within H9. The presence of Pro-containing peptides within H9 may contribute to its stability to digestive enzymes. Gluten hydrolysates may have antidiabetic potential for humans

By combining next-generation sequencing technology (454) and reduced representation library (RRL) construction, the rapid and economical isolation of over 25 000 potential single-nucleotide polymorphisms (SNP) and >6000 putative microsatellite loci from c. 2% of the genome of the non-model teleost, Atlantic cod Gadus morhua from the Celtic Sea, south of Ireland, was demonstrated. A small-scale validation of markers indicated that 80% (11 of 14) of SNP loci and 40% (6 of 15) of the microsatellite loci could be amplified and showed variability. The results clearly show that small-scale next-generation sequencing of RRL genomes is an economical and rapid approach for simultaneous SNP and microsatellite discovery that is applicable to any species. The low cost and relatively small investment in time allows for positive exploitation of ascertainment bias to design markers applicable to specific populations and study questions.

► Protein rich isolate and hydrolysates were prepared from brewers' spent grain (BSG). ► The bioactivity of BSG protein preparations was measured in vitro. ► BSG protein preparations do not possess antioxidant potential. ► BSG protein preparations show selective immunomodulatory potential, reducing IFN-γ.

Glutamyl endopeptidase (GE) from Alcalase™ 2.4 L was purified using hydrophobic interaction (HIC) and ion-exchange (IEX) chromatography. The yield of GE obtained was approximately 42%. Bovine α-casein (containing α(s1)- and α(s2)-casein) was digested with GE at 37 and 50°C for 4h. Samples were withdrawn at various time intervals and the peptides generated were analysed using mass spectrometry. GE activity was highly specific and hydrolysed the peptide bond predominantly on the carboxy side of Glu residues while hydrolysis on the carboxyl side of Asp residues was also observed. Hydrolysis did not occur when Pro was at the P(1)' position. In Glu-Glu-X (X=Arg, Asn, Ile and Ser) and Glu-Glu-Glu-Lys sequences, hydrolysis of Glu-X and Glu-Lys was preferred. The results are relevant to our understanding of the hydrolytic specificity of Alcalase, a food-grade proteolytic preparation containing GE activity which is used in the generation of casein hydrolysates.

The tryptic digestion of β-lactoglobulin leads to the release of a large range of biologically active peptides. Ultrafiltration/nanofiltration technology can be used to fractionate protein hydrolysates in order to obtain permeate products with increased functionality and free from intact proteins and enzymes. The influence of the pH in the fractionation of the hydrolysate through a polyethersulfone membrane (MWCO 5 kDa) was investigated in this work. In this case peptide transmission was mainly governed by charge mechanisms and reached its maximum value when the pH value is close to the peptide isoelectric point. Almost complete rejection of acidic peptides was achieved at basic pH values due to electrostatic repulsive forces with the negatively charged membrane and, in agreement with Donnan theory, positively charged peptides shown lower transmission than the neutral species. The highest peptide recovery and the best separation factor between bioactive and non-bioactive peptides were obtained following nanofiltration at pH 8.0.

Enzymatic hydrolysis of sodium caseinate (NaCas) may lead to the development of bitterness. Careful selection of hydrolysis conditions (i.e. enzyme preparation and duration) yielding different degrees of hydrolysis (DH) may aid in the development of low bitterness.Eighteen NaCas hydrolysates were generated with four enzyme preparations (Alcalase 2.4L, Prolyve 1000, FlavorPro Whey and pepsin) to different DH values. Hydrolysate bitterness score, assessed using a trained panel (ten assessors), generally increased at higher DH values for Alcalase, Prolyve and pepsin hydrolysates. However, all FlavorPro Whey hydrolysates (DH 0.38-10.62%) displayed low bitterness score values (26.0%) comparable to that of intact NaCas (13.8 ± 2.0%, P0.05).Enzyme preparation and DH affect the bitterness of NaCas hydrolysates. The results are relevant for the generation of NaCas hydrolysates with reduced bitterness. © 2017 Society of Chemical Industry.