Characterization and ACE-Inhibitory Activity of Amaranth Proteins

ABSTRACT:  Amaranth seeds have been considered as an excellent alternative or complementary source of food protein due to their balanced amino acid composition. However, their potential as a source of bioactive peptides has not been explored. The present study is aimed at characterizing and evaluating the activity of the angiotensin converting enzyme inhibitor of the amaranth protein concentrate and of hydrolysates produced with Alcalase. The protein concentrate, after simulated gastrointestinal digestion, showed lower angiotensin converting enzyme-inhibitory activity (IC₅₀ of 0.439 ± 0.018 mg protein/mL and 0.475 ± 0.021 mg protein/mL, for untreated and heat treated protein concentrate, respectively) than the hydrolysates produced with Alcalase, before and after simulated gastrointestinal digestion (IC₅₀ 0.118 ± 0.009, 0.123 ± 0.007, 0.137 ± 0.002, and 0.176 ± 0.014 mg protein/mL, respectively). The simulated gastrointestinal digestion (pepsin–pancreatin) did not significantly alter the angiotensin-converting enzyme inhibiting activity of the Alcalase hydrolysates, suggesting that the peptides of the hydrolysates were resistant to gastrointestinal hydrolysis. These results highlight the angiotensin converting enzyme-inhibitory potential of amaranth proteins, which is an indication of their health-promoting potential.     

Physicochemical, functional and angiotensin converting enzyme inhibitory properties of amaranth (Amaranthus hypochondriacus) 7S globulin

BACKGROUND: Amaranth 7S globulin is a minor globulin component and its impact on the properties of an amaranth protein ingredient depends on its proportion in the variety of amaranth being considered. Some physicochemical, functional and angiotesin I‐converting enzyme (ACE) inhibitory properties of amaranth vicilin were studied in this work and compared with the 11S globulin. RESULTS: Fluorescence spectroscopy results indicated that 7S globulin tryptophans were more exposed to the solvent and, by calorimetry, the 7S globulin denaturation temperature (T_d) was found lower than the 11S globulin T_d, suggesting a more flexible structure. The 7S globulin surface hydrophobicity was higher than that of the 11S globulin, which is in agreement with the better emulsifying properties of the 7S globulin. The solubility in neutral buffer of the 7S globulin (851 ± 25 g kg^?1) was also higher than that of the 11S globulin (195 ± 6 g kg^?1). Bioinformatic analyses showed the presence of ACE inhibitory peptides encrypted in 7S tryptic sequences and peptides released after in vitro gastrointestinal digestion showed a high ACE‐inhibitory capacity (IC₅₀ = 0.17 g L^?1), similar to that of 11S globulin peptides. CONCLUSION: Compared with the 11S globulin, the 7S globulin presents similar ACE inhibitory activity and some functional advantages, better solubility and emulsifying activity, which suits some food requirements. The functional behavior has been related with the structural properties. Copyright © 2011 Society of Chemical Industry     

Fractionation of angiotensin I converting enzyme inhibitory activity from pea and whey protein in vitro gastrointestinal digests

In vitro gastrointestinal digestion of pea and whey protein produced high angiotensin I converting enzyme (ACE) inhibitory activity with IC₅₀ values of 0.070 and 0.041 mg protein ml^?1 respectively. Ultrafiltration/centrifugation using a membrane with a molecular weight cut‐off of 3000 Da decreased the IC₅₀ value to 0.055 mg protein ml^?1 for pea permeate and 0.014 mg protein ml^?1 for whey permeate. Further fractionation by reverse phase HPLC gave IC₅₀ values as low as 0.016 mg protein ml^?1 for pea and 0.003 mg protein ml^?1 for whey. Consequently, these purification steps enriched the ACE inhibitory activity of the pea digest more than four times and that of the whey digest more than 13 times. HPLC profiles after digestion and ultrafiltration indicate that high ACE inhibitory activity is due to short and more hydrophobic peptides. The results also suggest that potent ACE inhibitory peptides were present alongside low active peptides in whey hydrolysate, while all peptides had more or less the same ACE inhibitory activity in pea hydrolysate. In addition, the hydrolysates and enriched fractions will resist in vivo gastrointestinal digestion after oral administration. Hence these ACE inhibitory peptides, as part of functional foods, can play significant roles in the prevention and treatment of hypertension. Copyright © 2004 Society of Chemical Industry     

Anthocyanins,total polyphenols and antioxidant activity in amaranth and quinoa seeds and sprouts during their growth

Total antioxidant capacity, total phenolic contents (TP) and anthocyanins contents (ANT) were determined in Amaranthus cruentus and Chenopodium quinoa seeds and sprouts. Antioxidant activity of the investigated seeds decreased in the following order: quinoa, amaranth v. Rawa, amaranth v. Aztek for FRAP and quinoa, amaranth v. Aztek, amaranth v. Rawa for both ABTS and DPPH. Sprouts activity depended on the length of their growth, and the peak values were reached on the fourth day in the case of amaranth and on the sixth day in the case of quinoa. The data obtained by the three methods showed significant correlation between TP content in seeds and sprouts. In sprouts grown in the daylight and in the darkness we observed some significant changes of TP, ANT and antioxidant activity. Amaranth and quinoa seeds and sprouts can be used in food, because it is a good source of ANT and TP with high antioxidant activity.     

Identification of bioactive peptides after digestion of human milk and infant formula with pepsin and pancreatin

Seven human milks were subjected to an in vitro digestion with pepsin and pancreatin to identify the peptides released from human proteins. On the basis of their sequences, 11 of the 23 peptides were synthesised and their angiotensin converting enzyme (ACE)-inhibitory and antioxidant activities were measured. The β-casein peptides HLPLP and WSVPQPK showed potent ACE-inhibitory and antioxidant activity, with a protein concentration needed to inhibit 50% ACE activity (IC₅₀) of 21 μm and a Trolox Equivalent Antioxidant Capacity (TEAC) of 1.297 μmol 6-hydroxy-2,5,7,8-tetramethylchroman-2-carboxylic acid (Trolox) equivs μmol⁻¹ of peptide, respectively. These activities were determined after digestion of eight infant formulas and compared with those found in digested human milk. One of the infant formulas exhibited a low IC₅₀ value (60.11 μg protein mL⁻¹ of reconstituted formula) and a high TEAC value (1.7056 μmol Trolox equivs mg⁻¹ of protein) and was therefore selected to identify the peptides responsible of these activities.     

NaCl和CaCl2对箭筈豌豆芽苗菜酚类物质积累与抗氧化性的影响

目的：开发箭筈豌豆菜用价值,探究盐溶液对箭筈豌豆芽苗菜酚类物质积累与抗氧化能力影响。方法：通过超高效液相色谱分析NaCl和CaCl2 对箭筈豌豆酚类物质积累的影响,并与豌豆苗、黄豆芽和绿豆芽进行对比,探究盐溶液处理下PAL、C4H、4CL关键酶活力与基因相对表达量变化、抗氧化酶活性变化和抗氧化能力变化。结果：NaCl和CaCl2 共同处理后箭筈豌豆芽苗菜总酚含量由(10.63±0.40) mg GAE/g DW提升至(15.76±0.36) mg GAE/g DW 。箭筈豌豆芽苗菜中共检出16种酚酸成分,在两种盐离子共同作用下,芦丁含量由(22.50± 0.49) mg/kg DW增长至(57.38±0.87) mg/kg DW,分别为豌豆苗、黄豆芽、绿豆芽的79.69,260.82,5.89倍。Na+ 通过提高PAL、4CL和C4H基因相对表达量提高了酶活力,Na+ 和Ca2+ 共同处理下基因相对表达量下降或无显著变化而酶活力提高。结论：NaCl和CaCl2 混合培养箭筈豌豆芽苗菜能够富集酚类物质,提升抗氧化能力。     

The Impact of Ozone Treatment on Changes in Biologically Active Substances of Cardamom Seeds

The overall objective of this study was to develop a decontamination method against microorganisms in cardamom (Elettaria cardamomum (L.) Maton) seeds using ozone as a decontaminating agent. Ozone treatment was conducted 3 times, at 24‐h intervals, and the parameters of the process were determined assuring the least possible losses of biologically active substances (essential oils and polyphenols): ozone concentration 160 to 165.0 g/m³; flow rate 0.1 L/min; pressure 0.5 atm; time 30 min. After each step of decontamination, the microbiological profile of the cardamom seeds was studied, and the contaminating microflora was identified. Next to the microbiological profile, the total polyphenol content (TPC), composition of essential oils, free radical‐scavenging capacity, total antioxidant capacity, ferric‐reducing antioxidant power (FRAP), and LC–MS polyphenol analysis were determined. This study shows that extract from cardamom seeds after ozone treatment is characterized by a better radical scavenging activity (IC₅₀ = 24.18 ± 0.04 mg/mL) than the control sample (IC₅₀ = 31.94 ± 0.05 mg/mL). The extract from cardamom seeds after ozone treatment showed an improved FRAP activity as well (613.64 ± 49.79 mmol TE/g compared to 480.29 ± 30.91 mmol TE/g of control sample). The TPC and the total antioxidant capacity were negatively affected, respectively, 41.2% and 16.2%, compared to the control sample.     

Identification of Bioactive Peptide Sequences from Amaranth (Amaranthus hypochondriacus) Seed Proteins and Their Potential Role in the Prevention of Chronic Diseases

Amaranth (Amaranthus hypochondriacus) is a pseudocereal with higher protein concentration than most cereal grains. Enzymatic hydrolysis and food processing could produce biopeptides from amaranth proteins; however, there is limited information about the bioactivity of peptides from amaranth proteins. The objective of this comprehensive review was to determine bioactive peptide sequences in amaranth proteins that may prevent cardiovascular disease, cancer, and diabetes. Amaranth proteins, reported in UniProt database, were evaluated for potential bioactive peptide using BIOPEP database. The 15 main proteins present in amaranth seed are 11S globulin, 7S globulin, α‐amylase inhibitor, trypsin inhibitor, antimicrobial proteins, nonspecific lipid‐transfer‐protein‐1, superoxide dismutase, ring‐zinc finger protein, prosystemin, amaranth albumin 1, glucose‐1‐phosphate adenyltransferase, glucosyltransferase, polyamine oxidase, granule‐bound starch synthase 1, and acetolactate synthase. All proteins showed high occurrence frequencies of angiotensin‐converting enzyme‐inhibitor peptides (A = 0.161 to 0.362), as well as of dipeptidyl peptidase IV inhibitor (A = 0.003 to 0.087). Other proteins showed antioxidative (A = 0.012 to 0.063) and glucose uptake‐stimulating activity (A = 0.023 to 0.042), and also antithrombotic (A = 0.002 to 0.031) and anticancer sequences (A = 0.001 to 0.042). The results of this study support the concept that amaranth grain could be part of a “healthy” diet and thereby prevent chronic human diseases.     

Isolation of peptides from a novel brewers spent grain protein isolate with potential to modulate glycaemic response

The in vitro dipeptidyl peptidase‐IV (DPP‐IV) inhibitory activity of a Brewers’ spent grain protein‐enriched isolate (BSG‐PI) Alcalase? hydrolysate (AlcH), which had previously been identified as a relatively potent angiotensin‐converting enzyme (ACE) inhibitor, was determined. The half maximal DPP‐IV inhibitory concentration (IC₅₀) value of AlcH following 240‐min digestion was 3.57 ± 0.19 mg mL^?1. Ultrafiltration fractionation did not significantly increase the DPP‐IV inhibitory activity of the AlcH fractions. Subjection of AlcH to simulated gastrointestinal digestion (SGID), which yielded SAlcH, resulted in a significant increase in DPP‐IV inhibitory activity (P < 0.05), particularly after the intestinal phase of digestion. Following semi‐preparative reverse phase high performance liquid chromatography (RP‐HPLC) fractionation of SAlcH, fraction 28 was identified as having highest mean DPP‐IV inhibitory activity. Two novel DPP‐IV inhibitory peptides, ILDL and ILLPGAQDGL, with IC₅₀ values of 1121.1 and 145.5 μm , respectively, were identified within fraction 28 of SAlcH following ultra‐performance liquid chromatography (UPLC)‐tandem mass spectrometry (MS/MS). BSG protein‐derived peptides were confirmed as having dual ACE and DPP‐IV inhibitory activities.     

Effect of the consumption of amaranth seeds and their sprouts on alterations of lipids and glucose metabolism in mice

Amaranth (Amaranthus hypochondriacus) is appreciated for its nutritional and functional quality. This study evaluated the effects of consuming amaranth seeds and their sprouts on the metabolic markers in ICR male mice. Ungerminated amaranth seeds (A) and one-day (G1) and three-day (G3) germinated amaranth sprouts were dried and milled. The mice groups (n = 10) were fed standard (N) or hypercaloric (H) diets and 1 g A, G1 or G3/kg body weight for 28 days. The total cholesterol (TC), high- and low-density lipoprotein (HDL and LDL), atherogenic index (AI), triglyceride (TG) and glucose levels were determined in blood samples. Germination increased the dietary fibre, phenolic and flavonoid concentration and decreased ash and squalene concentration. A, G1 or G3 consumption also decreased body weight, TC, LDL, AI and glucose levels in H group mice. Thus, amaranth seed or sprout consumption could be beneficial for people with altered lipid metabolism.     

In vitro inhibition of dipeptidyl peptidase IV by peptides derived from the hydrolysis of amaranth (Amaranthus hypochondriacus L.) proteins

Bioactive compounds present in foods could potentially have beneficial effects on human health. In this study we report the in vitro inhibitory capacity of peptides released from amaranth seed proteins after enzymatic digestion, against dipeptidyl peptidase IV (DPPIV); an enzyme known to deactivate incretins, hormones involved in insulin secretion. Other seeds, such as soybean, black bean, and wheat were also tested. The highest inhibition of DPPIV was observed with amaranth peptides released after simulated gastrointestinal digestion, showing an IC₅₀ of 1.1 mg/mL in a dose-dependent manner. In silico tryptic digestion of amaranth globulins was carried out releasing peptides larger than 13 residues. Some of these peptides were used for the in silico prediction of their binding modes with DPPIV. Docking models showed that the possible mechanism of globulin peptides to inhibit DPPIV was through blocking the active dimer formation. Peptides were also found inside the major cavity where the natural substrates reach the catalytic site of the enzyme. This is the first report of the identification of inhibitory DPPIV peptides from amaranth hydrolysates and the prediction of their binding modes at the molecular level, leading to their possible use as functional food ingredients in the prevention of diabetes.     

Isolation and characterisation of a novel angiotensin I‐converting enzyme‐inhibitory peptide derived from douchi,a traditional Chinese fermented soybean food

BACKGROUND: Douchi, a traditional fermented soybean food, has recently attracted a great deal of attention owing to its superior physiological activity. In the present study the angiotensin I‐converting enzyme (ACE)‐inhibitory activity of typical douchi procured from various regions of China was analysed. An ACE‐inhibitory peptide derived from the most potent douchi was also isolated and characterised. The pattern of ACE inhibition and resistance to hydrolysis by gastrointestinal proteases of this peptide are described. RESULTS: ACE‐inhibitory activities were detected in all douchi samples, with IC₅₀ values ranging from 0.204 to 2.011 mg mL^?1. Among the douchi samples, a Mucor‐type douchi exhibited the most potent ACE‐inhibitory activity (IC₅₀ = 0.204 mg mL^?1). A novel ACE‐inhibitory peptide was then isolated from this Mucor‐type douchi using ultrafiltration followed by Sephadex G‐25 column chromatography and reverse phase high‐performance liquid chromatography. The amino acid sequence of the purified peptide was identified by Edman degradation as His‐Leu‐Pro (IC₅₀ = 2.37 µmol L^?1). The peptide is a competitive inhibitor and maintained its inhibitory activity even after incubation with some gastrointestinal proteases. CONCLUSION: The present study shows that peptides derived from soybean fermentation during douchi processing could be the main contributor to the ACE‐inhibitory activity observed. Copyright © 2009 Society of Chemical Industry     

Angiotensin-I-converting enzyme inhibitory,antimicrobial, and antioxidant effect of bioactive peptides obtained from different varieties of common beans (Phaseolus vulgaris L.) with in vivo antihypertensive activity in spontaneously hypertensive rats

Peptides obtained from three varieties of common beans (Phaseolus vulgaris L.) named plus black (PB), azufrado higuera (AH) and pinto Saltillo (PS) presented antimicrobial, antioxidant and antihypertensive activities. Peptides were obtained from these common beans protein concentrates after treatment with Alcalase^® followed by ultrafiltration using 1-, 3- and 10-kDa molecular weight cutoff membranes. Antioxidant activity was determined by the 2,2′-azino-bis (3-etilbenzotiazolin-6-sulfonic) acid method and was expressed as Trolox equivalent antioxidant capacity (TEAC). The highest antioxidant activity (630, 550 and 517 mM TEAC/mg protein) was observed in the group of peptides with a molecular weight lower than 1 kDa (F < 1) from PB, AH and PS bean varieties, respectively. Antibacterial activity was determined as susceptibility test in which ten of twelve bacteria strains showed growth inhibition with the total hydrolysates (TH) and the peptidic fraction 3–10 kDa; subsequently, the minimum inhibitory concentration was determined with the standard microdilution assay in a 96-well plates in which the peptidic fraction F < 1 kDa presented antimicrobial activity against Shigella dysenteriae with the three beans varieties at 0.1, 0.4 and 0.3 mg/mL (for PB, AH and PS, respectively). Additionally to the antimicrobial and antioxidant activities, the TH of the PB and AH bean varieties presented high inhibition of the angiotensin-I-converting enzyme (ACE-I) (IC₅₀ = 4.34 ± 0.29 and 4.82 ± 1.59 μg/mL, respectively). And, when the peptidic fraction F3–10 kDa was tested, the AH variety showed a significant increase in the ACE-I capacity (IC₅₀ = 1.09 ± 0.04 μg/mL). Importantly, this peptidic fraction decreased the systolic blood pressure in a spontaneously hypertensive rat model after 2 h of administration by a single interperitoneally dose.     

Bioaccessibility and antioxidant activity of phenolic compounds in cooked pulses

The aim of this study was to evaluate the bioaccessibility of polyphenols and antioxidant activity in cooked pulses and to study the effect of cooking on their total phenolic content (TPC) and antioxidant capacity. Cooked faba beans showed the highest TPC, followed by soybeans and lentils or peas. TPC ranged from 10.4 ± 0.2 to 52.9 ± 0.3 mg/100 g and was positively correlated with antioxidant activity. Cooking resulted in increased TPC and antioxidant activity of the methanolic extracts, caused by cell disruption and improved extraction of polyphenols. Although polyphenols were lost in the cooking water, boiled legumes had more polyphenols than those resulting cooking broths. In vitro gastrointestinal digestion resulted in increased TPC and antioxidant capacity of the extracts. Soybeans showed the highest amount of bioaccessible polyphenols. The release of phenolics from cooked legumes was mainly achieved during the intestinal phase. Literature data may underestimate the TPC and antioxidant capacity of pulses.     

Investigation of a wild pear species (Pyrus elaeagnifolia subsp. Elaeagnifolia Pallas) from Antalya,Turkey: polyphenol oxidase properties and anti-xanthine oxidase,anti-urease,and antioxidant activity

Polyphenol oxidase properties and anti-xanthine oxidase, anti-urease, and antioxidant activity were investigated in Pyrus elaeagnifolia subsp. elaeagnifolia Pallas harvested from Antalya, Turkey. Optimum pH and temperature were 7.0 and 30°C, respectively. Selected kinetic properties of polyphenol oxidase was evaluated.The K_m and V_max-values, using 4-methylcatechol as substrate, were calculated as 3.57 mM and 4781 U/mg protein, respectively. IC₅₀-values ranged from 0.0036 to 4.0231 mM against sodium metabisulphite, ascorbic acid, sodium azide, and benzoic acid, while ascorbic acid was the strongest inhibitor. Aquatic extracts of the sample exhibited strong antioxidant capacity and substantial xanthine oxidase and urease inhibition, with IC₅₀-values of 10.75 ± 0.11 and 0.97 ± 0.03 mg/mL, respectively.     

人心果叶不同极性组分的抗氧化与抗菌活性

通过体外DPPH自由基、ABTS自由基清除能力和总抗氧化能力评价人心果叶不同极性萃取物（石油醚相、乙酸乙酯相、正丁醇相、水相）的抗氧化活性,并探究各极性组分对金黄色葡萄球菌、枯草芽孢杆菌和大肠杆菌的抑制能力。结果表明：乙酸乙酯组分的体外抗氧化活性最强,清除DPPH自由基能力IC₅₀值为（0.019±0.010） mg/mL,总抗氧化能力以水溶性维生素E当量（TEAC）表示,值为（5.824±0.234） mmol/g。当质量浓度达到0.50 mg/mL时,乙酸乙酯组分对ABTS自由基清除率就已经超过了50%;且乙酸乙酯组分能够明显抑制金黄色葡萄球菌的生长,最小抑菌浓度（MIC）和最小杀菌浓度（MBC）分别为0.313,0.625 mg/mL。人心果叶乙酸乙酯组分具有显著的抗氧化和抗菌活性,可用于天然抗氧化剂和抗菌剂的开发。     

Antioxidant activity of amaranth protein or their hydrolysates under simulated gastrointestinal digestion

Amaranth proteins were subjected to a simulated gastrointestinal digestion to evaluate the antioxidant activity of the products. A protein isolate (I) was first hydrolyzed with pepsin (Pe) (pH 2, 37 °C) and then with pancreatin (Pa) (pH 6, 37 °C). Different hydrolysis conditions were assayed and control reactions (without enzymes) were performed. Hydrolysis degree (HD) determined by TNBS method ranged from 13 to 37%. Soluble fractions in 35 mmol/L phosphate buffer, pH = 7.8 were obtained from freeze-dried samples, and antioxidant activity was evaluated by the ABTS⁺·scavenging and the ORAC assays. Antioxidant activity increased significantly (p < 0.05) after simulated gastrointestinal digestion. According to the results, digestion conditions (Pe/protein: 1:10, 60 min; and Pa/protein: 1:10, 60 min) were selected and applied to an amaranth protein alcalase-hydrolysate (H) (HD = 29.2 ± 1.3). After pepsin and pancreatin action (Hpepa), HD was 42.0 ± 2.6, slightly higher than that of the digested isolate (Ipepa) (36.9 ± 0.5). The corresponding soluble fractions exhibited different electrophoretic profiles (tricine-SDS-PAGE) and gel filtration chromatograms, evidencing the presence of different molecular species. Previous hydrolysis with alcalase did not improve the antioxidant activity after simulated gastrointestinal digestion according to the methodologies assayed. Both the protein isolate and the alcalase-hydrolysate showed a potential capacity to scavenge free radicals after gastrointestinal digestion, appearing as promising ingredients to formulate functional foods with antioxidant activity.     

Nutritional constituents,phytochemical profiles, <Emphasis Type="Italic">in vitro</Emphasis> antioxidant and antimicrobial properties,and gas chromatography–mass spectrometry analysis of various solvent extracts from grape seeds (<Emphasis Type="Italic">Vitis vinifera</Emphasis> L.)

The present study revealed that the nutritive value of grape seeds (Vitis vinifera L.) was 383.55±0.13 Kcal/100 g, with magnesium as the most abundant mineral element (70.44±0.88 mg/L). The maximum phenolic (392.58±1.70mg of GAE/g), flavonoid (256.16±1.60 mg of QE/g), and tannin (30.95±0.17mg of CE/g) contents were also found in the ethanol, dichloromethane, and hexane extracts, respectively. The major phytochemical compounds in the ethyl acetate extract were identified via gas chromatography–mass spectrometry (GC–MS) analysis. The ethanol extract has the highest antioxidant activity (IC₅₀=140±1.20 μg/mL for DPPH, 145.28±0.45mg α-tocopherol/g for total antioxidant capacity, and EC₅₀=80±1.41 μg/mL for ferric-reducing power assays). For β-carotene test, the highest antioxidant activity was obtained in the hexane extract. A satisfactory antimicrobial activity was found against a panel of microorganisms with the ethyl acetate extract as the best antimicrobial agent. Additionally, it was found that the bactericidal concentration required for the grape seed extract to kill Listeria monocytogenes should be less than 12.50 mg/mL (minimum inhibitory concentration=4).     

Generation of phenolic-rich extracts from brewers' spent grain and characterisation of their in vitro and in vivo activities

Brewers' spent grain (BSG) is a co-product of the brewing industry, which is rich in phenolic acids. This study compared the in vitro bioactive properties of aqueous, aqueous pH-shift and enzyme-aided phenolic extracts from pale (unroasted) wet BSG. Overall, the pH-shift extracts showed higher oxygen radical absorbance capacity (ORAC), trolox equivalent antioxidant capacity (TEAC) and ferric-reducing antioxidant power (FRAP) values compared with the enzyme-aided extracts. A selected enzyme-aided extract showed highest in vitro ACE inhibitory activity (ACE IC₅₀: 0.133 ± 0.032 mg mL⁻¹) and mediated significant (p < 0.05) reductions in blood pressure markers in vivo following ingestion by spontaneously hypertensive rats (systolic blood pressure; −35.35 ± 9.04 and diastolic blood pressure; −28.55 ± 4.22 mmHg). The results demonstrated that the observed in vitro activity of phenolic-rich extracts may be translated into an in vivo hypotensive effect. Enzyme-assisted extraction has potential as an alternative solvent-free approach for generating phenolic-rich extracts from BSG that may find use in the management of hypertension.     

Identification of angiotensin converting enzyme inhibitory and antioxidant peptides in a whey protein concentrate hydrolysate produced at semi‐pilot scale

Antioxidant and angiotensin converting enzyme (ACE) inhibitory peptides were identified in a 5 kDa ultrafiltration permeate of a whey protein hydrolysate generated at semi‐pilot scale. Further laboratory scale ultrafiltration of this 5 kDa permeate resulted in a 0.65 kDa permeate with antioxidant, (1.11 ± 0.074 μmol TE per mg dry weight, oxygen radical absorbance capacity, ORAC) and ACE inhibitory (ACE IC₅₀ 0.215 ± 0.043 mg mL^?1) activities. Semi‐preparative (SP) reverse phase high‐performance liquid chromatography (RP‐HPLC) of the 0.65 kDa permeate resulted in a fraction (SP_F3) with a 4.4‐fold increase in ORAC activity (4.83 ± 0.45 μmol TE mg dry weight) and a 1.3‐fold increase in ACE inhibitory activity (84.35 ± 1.36% inhibition when assayed at 0.28 mg mL^?1). Peptides within SP_F3 were identified using UPLC‐ESI‐MS/MS. Met‐Pro‐Ile had the highest ORAC activity (205.75 ± 12.08 μmol TE per mmol peptide) while Met‐Ala‐Ala and Val‐Ala‐Gly‐Thr had the highest ACE inhibitory activities (IC₅₀:515.50 ± 1.11 and 610.30 ± 2.41 μm , respectively).