Highest ACE-inhibitory Activity Research Articles

Assessment on multilayer ultrafiltration membrane for fractionation of tilapia by-product protein hydrolysate with angiotensin I-converting enzyme (ACE) inhibitory activity

Abstract Ultrafiltration membrane is a convenient system to fractionate fish protein hydrolysate and recovery of peptide fraction with high ACE inhibitory activity. However, the limitation of applying membrane system in protein fractionation is its poor selectivity. This study aiming to assess the application of multilayer ultrafiltration in order to improve the selectivity of the tilapia by-product protein hydrolysate (TBH) separation by achieving higher amount of small peptides. Flat sheet regenerated cellulose (RC) membrane with molecular weight cut-off (MWCO) of 10 and 5 kDa were used for the separation of peptide mixtures. The membrane were arranged in the orientation of 10/5 kDa and 5/5 kDa, in which these two membrane were stacked together in one device. The performance of multilayer membrane were evaluated based on the permeate flux and peptide transmission, and compare with the single membrane system (5 and 10 kDa). The performance ultrafiltration membrane for fractionating TBH was studied under two different conditions (rotation speed and pH). The highest permeate flux and peptide transmission were obtained at membrane with pore size of 10 kDa (single), followed by 10/5 (multilayer), 5 (single) and 5/5 kDa (multilayer). Based on selectivity analysis, most permeate produced were composed of peptides lower than 1500 Da. When the smaller membrane’s pore size is used (5 kDa membrane and 5/5 multilayer membranes), the amount of small-sized peptide (

Isolation and Purification of Antioxidant and ACE-Inhibitory Peptides from Yak (Bos grunniens ) Skin

Yak (Bos grunniens) is a kind of cattle which can survive on Tibetan plateau. In addition, yak skin is an unused resource during production of meat. In this research, the potential angiotensin I-converting enzyme (ACE)-inhibitory and antioxidant bioactivities of yak skin gelatin hydrolysates (YSGH) were investigated. Orthogonal test was applied to optimize the pretreatment process. Then the gelatin was treated with combinations of different kinds of enzymes and YSGH was separated by ultrafiltration membranes. Interestingly, all hydrolysates below 3 kDa by different enzyme combination treatments showed excellent antioxidant activities, and the fraction (<3 kDa) hydrolyzed by Proteinase K exhibited high ACE-inhibitory activity with an IC50 value of 1.119 mg mL−1. Furthermore, the hydrolysis condition for better activity of ACE-inhibitory fraction was optimized, and then a novel ACE-inhibitory peptide B4P was obtained through purification, which exhibited the highest ACE-inhibitory activity (IC50 = 35 µg mL−1). The molecular mass of B4P was 1496.7 Da. Practical Applications This study introduces the antioxidant and ACE-inhibitory activities of the peptides isolated from yak skin. During this process, the purification methods were also optimized. The peptides showed excellent bioactivities, which indicated that yak skin offers the potential for the production of functional foods or medicines with antihypertensive and antioxidant activities.

Bioactive peptides from Atlantic salmon (Salmo salar) with angiotensin converting enzyme and dipeptidyl peptidase IV inhibitory, and antioxidant activities

The pH shift method was utilised for the recovery of proteins from salmon trimmings (ST), yielding 93% (w/w) protein. ST protein (STP) hydrolysates were generated with different enzyme preparations. STP incubated with Corolase PP for 1h (STP-C1) had the most potent angiotensin converting enzyme (ACE) and dipeptidyl peptidase IV (DPP-IV) inhibitory and oxygen radical absorbance capacity (ORAC) activities. Analysis of fractions of STP-C1 using UPLC-MS/MS identified sixteen peptides/amino acids. Tyr-Pro had the highest ACE inhibitory activity (ACE IC50=5.21±0.94μM). The highest DPP-IV inhibitory activity was found with the amino acid Tyr (DPP-IV IC50=75.15±0.84μM). Val-Pro had the highest ORAC activity (19.45±2.15μmol of TEg−1). To our knowledge, the peptides Gly-Pro-Ala-Val, Val-Cys, and Phe-Phe have not been previously identified to have the activities tested in this study. These results indicate that STP hydrolysates are potential sources of bioactive peptides.

A novel antioxidant and ACE inhibitory peptide from rice bran protein: Biochemical characterization and molecular docking study

Rice bran protein was hydrolyzed using trypsin. The hydrolysate (RBPH) was then further separated by membrane bioreactor system, gel filtration and reversed phase high-performance liquid chromatography (RP-HPLC). A novel antioxidant and angiotensin I-converting enzyme (ACE) inhibitory peptide named as F2-a, which exhibited high DPPH free radicals scavenging activity, reducing power and ACE inhibitory activity (IC50 of 76 μM) was isolated. The amino acid sequence, Tyr-Ser-Lys (Mw: 395.0 Da), was identified by Quardrupole Time-of-flight Mass Spectrometer (Q-TOF-MS) with an electro-spray ionization (ESI) source. The molecular docking study revealed that the ACE inhibition of Tyr-Ser-Lys was mainly attributed to forming very strong hydrogen bonds with the active pockets of human ACE. These results indicate that rice bran is a potential source of bioactive peptides possessing antioxidant and ACE inhibitory activities.

Investigation of lactic acid bacterial strains for meat fermentation and the product's antioxidant and angiotensin-I-converting-enzyme inhibitory activities.

In the lactic acid bacteria (LAB) strains screened from our LAB collection, Lactobacillus (L.) sakei strain no. 23 and L. curvatus strain no. 28 degraded meat protein and tolerated salt and nitrite in vitro. Fermented sausages inoculated strains no. 23 and no. 28 showed not only favorable increases in viable LAB counts and reduced pH, but also the degradation of meat protein. The sausages fermented with these strains showed significantly higher antioxidant activity than those without LAB or fermented by each LAB type strain. Angiotensin-I-converting-enzyme (ACE) inhibitory activity was also significantly higher in the sausages fermented with strain no. 23 than in those fermented with the type strain. Higher ACE inhibitory activity was also observed in the sausages fermented with strain no. 28, but did not differ significantly from those with the type strain. An analysis of the proteolysis and degradation products formed by each LAB in sausages suggested that those bioactivities yielded fermentation products such as peptides. Therefore, LAB starters that can adequately ferment meat, such as strains no. 23 and no. 28, should contribute to the production of bioactive compounds in meat products.

ACE-Inhibitory and Antioxidant Activities of Peptide Fragments Obtained from Tomato Processing By-Products Fermented Using Bacillus subtilis: Effect of Amino Acid Composition and Peptides Molecular Mass Distribution.

The effects of amino acid composition and peptide molecular mass on ACE-inhibitory and antioxidant activities of protein fragments obtained from tomato waste fermented using Bacillus subtilis were evaluated. The addition of B. subtilis increased the relative amounts of aromatic and positively-charged amino acids which have been described to influence the biological activities of peptide fragments. IC50 values of hydrolysates for ACE-inhibitory and 2, 2'-diphenyl-1-picrylhydrazyl (DPPH) scavenging activities were found to be 1.5 and 8.2mg/mL, respectively. Size-exclusion chromatography (SEC) pattern of the hydrolysate indicated the breakdown of parent proteins to smaller peptides with molecular weights mainly below 1400Da. MALDI-TOF mass spectrometry analysis revealed that the highest ACE-inhibitory activity was due to peptides showing molecular mass range 500-800Da, while the most active antioxidant peptides were found to be mainly at the two different peptide weight ranges 500-800Da and 1200-1500Da.

Changes of Composition and Angiotensin I-Converting Enzyme-Inhibitory Activity During Douchi Fermentation

Douchi is a fermented soybean food, which is popular in China. Composition and angiotensin I-converting enzyme-inhibitory activity of Douchi fermented with Mucor wutungkiao were studied in this work. The results showed that the nutritional potency of protein, fat, and soluble polysaccharide was improved in Douchi fermentation. The angiotensin I-converting enzyme-inhibitory activity of Douchi that ripening for 2 weeks showed the highest angiotensin I-converting enzyme-inhibitory activity. Then this sample was purified with Sephadex G-25 gel chromatography, and the amino acids and angiotensin I-converting enzyme-inhibitory activity of the purified fractions were studied. The results suggested that hydrophobic amino acids were assumed to cause the main increase of the angiotensin I-converting enzyme-inhibitory activity in Douchi.

Structural Properties of Phycoerythrin from DulsePalmaria palmata

We found that the red alga dulse (Palmaria palmata) contains a lot of proteins, which is mainly composed of phycoerythrin (PE) and the protein hydrolysates showed high angiotensin I converting enzyme (ACE) inhibitory activities. Therefore, we investigated the structure of dulse PE to discuss its structure-function relationship. We prepared the chloroplast DNA and analyzed the nucleotide sequences encoding PE by cDNA cloning method. It was clarified that dulse PE has α- and β-subunits and they are composed by 164 amino acids (MW: 17,638) and 177 amino acids (MW: 18,407), respectively. The dulse PE contained conserved cysteine residues for chromophore attachment site. On the alignment of amino acid sequences of dulse PE with those of other red algal PE, the sequence identities were very high (81–92%). In addition, we purified and crystallized the dulse PE, and its crystal structure was determined at 2.09 A resolution by molecular replacement method. The revealed 3D structure of dulse PE which forms an (αβ)6 hexamer was similar to other red algal PEs. Conversely, it was clarified that the dulse PE proteins are rich in hydrophobic amino acid residues (51.0%), especially aromatic amino acid and proline residues. The data imply that the high ACE inhibitory activity of dulse protein hydrolysates would be caused by the specific amino acid composition and sequence of dulse PE. Practical Applications Dulse is an abundant and underused resource, which contains a lot of phycobiliproteins. Then, the dulse protein hydrolysates strongly inhibited the activity of ACE. Therefore, it has the potential to be an ingredient of functional food.

Production of Angiotensin-I-Converting-Enzyme-Inhibitory Peptides in Fermented Milks (Lassi) Fermented by Lactobacillus acidophillus with Consideration of Incubation Period and Simmering Treatment

The lassi, fermented milks product containing angiotensin-I-converting-enzyme (ACE)-inhibitory peptides were produced by using selected Lactobacillus acidophilus NCDC-15 and the incubation period and simmering effect was also optimized for production of ACE-inhibitory peptides. The time–temperature combination for the heat treatment was optimized using RSM. The biological activity was measured in the supernatant of the fermented milk after centrifugation. The lowest IC50 values for the inhibition of angiotensin-converting enzyme (ACE) was found 28.9 ± 0.95 μg protein/ml in the supernatant of milk fermented by L. acidophilus and heated at 78 °C for 10 h. The fractions which showed the highest ACE-inhibitory indexes were further purified by different techniques including solid phase extraction, RP-HPLC and FPLC and the related peptides were identified by LC–MS/MS using the Ultimate 3000 nano HPLC system (Dionex) coupled to a 4000 Q TRAP electro-spray ionization mass spectrometry. The high ACE-inhibitory activity containing fractions of the milk fermented by L. acidophilus contained the sequences of b-casein (b-CN) fragment. The fraction-III showed minimum IC50 value i.e. 14.57 ± 0.72 μg/ml compared with fraction-I and fraction-II. Among these peptides 14 peptides have been identified from the fraction-I of the lassi prepared from L. acidophilus i.e. β-CN f47–56, β-CN f47–57, β-CN f199–209, β-CN f176–182, β-CN f176–183, β-CN f176–184, β-CN f1–7, β-CN f57–68, β-CN f166–175, β-CN f195–206, β-CN f195–207, β-CN f195–209, β-CN f94–106 and β-CN f169–176 showed partially or completely homology to that the milk protein bioactive peptides having ACE inhibitory. The two peptides KVLPVPQK (β-CN f169–176) and YQEPVLGPVRGPFPIIV (β-CN f193–209) have the same sequence as ACE inhibitory peptides (Maeno et al. in J Dairy Sci 79(8):1316–1321, 1996; Yamamoto et al. in J Dairy Sci 77:917–922, 1994b).

Purification and identification of an angiotensin I-converting enzyme inhibitory peptide from cauliflower by-products protein hydrolysate

Cauliflower by-products (stems and leaves) are rich in leaf protein. Making full use of cauliflower by-products can not only alleviate the shortage of protein resources but also be beneficial to the environmental protection. This study sought to prepare and purify an angiotensin I-converting enzyme (ACE) inhibitory peptide from cauliflower by-products protein hydrolysate. Our result showed that the highest ACE inhibitory activity was observed in the Alcalase 2.4L hydrolysate among tested hydrolysates (pepsin, pancreatin, and Alcalase 2.4L). After purification, a dipeptide (Val-Trp) was identified, with a molecular weight of 304.1665Da. This peptide had a potent ACE inhibitory activity with IC50 of 31.30μM. Based on this purification process, approximately 42mg of this dipeptide was obtained from 1kg of fresh cauliflower by-products. Overall, the results suggest that cauliflower by-products protein can be used as an economical source of functional food material against hypertension.

Angiotensin I‐converting enzyme inhibitory peptides FQPSF and LKYPI identified in Bacillus subtilis A26 hydrolysate of thornback ray muscle

SummaryAngiotensin I‐converting enzyme (ACE) inhibitory peptides have been searched in thornback ray (Raja clavata) muscle hydrolysed with Bacillus subtilis A26 proteases until a hydrolysis degree of 18.35%. The hydrolysate showed an IC50 of 0.83 mg mL−1. To identify peptides responsible for this activity, the extract was eluted through size‐exclusion chromatography and fractions collected. The highest ACE inhibitory activity was found for fractions F2 and F3 which had IC50 of 0.42 and 0.51 mg mL−1, respectively. These fractions were analysed by nano‐liquid chromatography coupled to tandem mass spectrometry (nLC‐MS/MS). A total of 131 and 108 peptide sequences mainly derived from actin, myosin heavy chain and procollagen alpha 1 chain proteins were identified in fractions F2 and F3, respectively. FQPSF and LKYPI showed the best results with an IC50 of 12.56 and 27.07 μM, respectively. These results prove the potential of thornback ray muscle hydrolysate as a source of ACE inhibitory peptides.

In Vitro and In Vivo Assessment of Angiotensin-Converting Enzyme (ACE) Inhibitory Activity of Fermented Soybean Milk by Lactobacillus casei Strains.

Angiotensin-converting enzyme (ACE) inhibitory activity of fermented soybean milk (FSM) by Lactobacillus casei strains in vitro was investigated in this study. Effects of fermented soybean milk administration by gavage on systolic blood pressure and diastolic blood pressure was also evaluated in spontaneously hypertensive rats (SHR) rats and Wistar-Kyoto (WKY) rats. Results showed that, CICC 20280 and CICC 23184 FSM showed high ACE inhibitory activity in vitro test and ACE inhibitory activity of CICC 23184 FSM was higher than CICC 20280 FSM. The bioactive substances of FSM were peptide and γ-aminobutyric acid (GABA). Their contents in CICC 20280 FSM and CICC 23184 FSM were 3.97±0.67mg/ml (peptide), 1.71±0.36mg/ml (GABA) and 5.17±0.22mg/ml (peptide), 1.57±0.21mg/ml (GABA), respectively. Moreover, CICC 20280 and CICC 23184 FSM administration by gavage could effectively lower the blood pressure of SHR to a normal level, while there was no effect on blood pressure of WKY rats. This result indicated that the bioactive substances could play an antihypertensive role when the blood pressure was not within the normal levels (high levels).

Influence of gas-liquid two-phase flow on angiotensin-I converting enzyme inhibitory peptides separation by ultra-filtration.

Membrane fouling is a major problem in ultra-filtration systems and two-phase flow is a promising technique for permeate flux enhancement. The objective of this research was to study the use of an ultra-filtration (UF) system to enrich angiotensin-I converting enzyme (ACE) inhibitory peptides from tilapia protein hydrolysate. To select the most appropriate membrane and operating condition, the effects of membrane molecular weight cut-off (MWCO), transmembrane pressure (TMP) and cross-flow velocity (CFV) on permeate flux and ACE inhibitory peptide separation were studied. Additionally, the gas-liquid two-phase flow technique was applied to investigate its effect on the process capability. The results showed that the highest ACE inhibitory activity was obtained from permeate of the 1 kDa membrane. In terms of TMP and CFV, the permeate flux tended to increase with TMP and CFV. The use of gas-liquid two-phase flow as indicated by shear stress number could reduce membrane fouling and increase the permeate flux up to 42%, depending on shear stress number. Moreover, the use of a shear stress number of 0.039 led to an augmentation in ACE inhibitory activity of permeates. Operating conditions using a shear stress number of 0.039 were recommended for enrichment of ACE inhibitory peptides. © 2016 Society of Chemical Industry.

Angiotensin I Converting Enzyme Inhibitory Peptides Derived from Phycobiliproteins of Dulse Palmaria palmata.

We examined the inhibitory activity of angiotensin I converting enzyme (ACE) in protein hydrolysates from dulse, Palmaria palmata. The proteins extracted from dulse were mainly composed of phycoerythrin (PE) followed by phycocyanin (PC) and allophycocyanin (APC). The dulse proteins showed slight ACE inhibitory activity, whereas the inhibitory activity was extremely enhanced by thermolysin hydrolysis. The ACE inhibitory activity of hydrolysates was hardly affected by additional pepsin, trypsin and chymotrypsin treatments. Nine ACE inhibitory peptides (YRD, AGGEY, VYRT, VDHY, IKGHY, LKNPG, LDY, LRY, FEQDWAS) were isolated from the hydrolysates by reversed-phase high-performance liquid chromatography (HPLC), and it was demonstrated that the synthetic peptide LRY (IC50: 0.044 μmol) has remarkably high ACE inhibitory activity. Then, we investigated the structural properties of dulse phycobiliproteins to discuss the origin of dulse ACE inhibitory peptides. Each dulse phycobiliprotein possesses α-subunit (Mw: 17,477–17,638) and β-subunit (Mw: 17,455–18,407). The sequences of YRD, AGGEY, VYRT, VDHY, LKNPG and LDY were detected in the primary structure of PE α-subunit, and the LDY also exists in the APC α- and β-subunits. In addition, the LRY sequence was found in the β-subunits of PE, PC and APC. From these results, it was suggested that the dulse ACE inhibitory peptides were derived from phycobiliproteins, especially PE. To make sure the deduction, we carried out additional experiment by using recombinant PE. We expressed the recombinant α- and β-subunits of PE (rPEα and rPEβ, respectively), and then prepared their peptides by thermolysin hydrolysis. As a result, these peptides showed high ACE inhibitory activities (rPEα: 94.4%; rPEβ: 87.0%). Therefore, we concluded that the original proteins of dulse ACE inhibitory peptides were phycobiliproteins.

Release of angiotensin converting enzyme-inhibitory peptides during in vitro gastro-intestinal digestion of camel milk

Angiotensin-converting enzyme (ACE)-inhibitory peptides released from camel milk after simulated gastro-intestinal digestion were identified. The hydrolysis degree increased during digestion. The highest ACE-inhibitory activity was found in the post-pancreatic <3 kDa fraction. Peptides responsible for the biological activity were isolated by reversed-phase high-performance liquid chromatography and identified by mass spectrometry. Among the identified sequences, 17 were identical to known bioactive peptides with ACE-inhibitory activity. Based on previous structure–activity relationship studies, the sequence of some peptides allowed us to anticipate the presence of biological activities. The anti-hypertensive tripeptide isoleucine-proline-proline (IPP) was identified and quantified in digested camel milk. The amount of released IPP was 2.56 ± 0.15 mg L−1 of milk. For the first time, we showed that IPP is released during the gastro-intestinal digestion of camel milk κ-casein. This research provides the basis to increase the exploitation of the health benefits of camel milk.

Utilization of Shrimp By-Products by Bioconversion with Medical Fungi for Angiotensin I-Converting Enzyme Inhibitor and Antioxidant

ABSTRACTA liquid fermentation process to bioconvert shrimp by-products was developed with two species of fungi, Boletus edulis and Suillus bovinus. Angiotensin I-converting enzyme (ACE) inhibitory, antioxidant activities of bioconverted products and the deodorization effect of fungal bioconversion were determined. Two species of fungi could grow quickly in shrimp by-products medium and deodorize shrimp by-products. Bioconverted products had higher ACE inhibitory and antioxidant activities than control medium. Water extracts without crude polysaccharides from mycelia of B. edulis had the highest ACE inhibitory activity (IC50 = 0.084 ± 0.011 mg/mL). Crude exopolysaccharides of B. edulis had higher antioxidant activity. Bioconversion with the fungus was proven to be a novel, practicable way to recycle shrimp by-products.

Separation and purification of angiotensin-I-converting enzyme (ACE) inhibitory peptides from walnuts (Juglans regia L.) meal

Angiotensin-I-converting enzyme (ACE) inhibitory peptides were isolated from walnut pepsin hydrolysates using ultrafiltration, Sephadex G-25 chromatography, reverse-phase high-performance liquid chromatography (RP-HPLC), and the AKTA preparative HPLC separation and purification system. ACE inhibitory activity was measured using in vitro HPLC analysis, and N-terminal amino acid sequencing was used for structural identification. Those fractions with the highest ACE inhibitory activity were further purified using RP-HPLC and the AKTA systems. The most active ACE inhibitory peptide had the sequence Tyr-Val-Pro-His-Trp-Asp-Leu and a molecular weight of 929 Da. Its activity, along with the activity of a synthetic ACE inhibitory peptide with the same sequence, was measured using HPLC following in vitro gastrointestinal digestion experiments. The IC50 values obtained before and after digestion were 0.136 and 0.173 μM/mL, respectively, which indicated that the peptide was relative stable during digestion and might have in vivo biological activity.

Antioxidant and ACE Inhibitory Bioactive Peptides Purified from Egg Yolk Proteins.

Protein by-products from the extraction of lecithin from egg yolk can be converted into value-added products, such as bioactive hydrolysates and peptides that have potential health enhancing antioxidant, and antihypertensive properties. In this study, the antioxidant and angiotensin converting enzyme (ACE) inhibitory activities of peptides isolated and purified from egg yolk protein were investigated. Defatted egg yolk was hydrolyzed using pepsin and pancreatin and sequentially fractionated by ultrafiltration, followed by gel filtration to produce egg yolk gel filtration fractions (EYGF). Of these, two fractions, EYGF-23 and EYGF-33, effectively inhibited the peroxides and thiobarbituric acid reactive substance (TBARS) in an oxidizing linoleic acid model system. The antioxidant mechanism involved superoxide anion and hydroxyl radicals scavenging and ferrous chelation. The presence of hydrophobic amino acids such as tyrosine (Y) and tryptophan (W), in sequences identified by LC-MS as WYGPD (EYGF-23) and KLSDW (EYGF-33), contributed to the antioxidant activity and were not significantly different from the synthetic BHA antioxidant. A third fraction (EYGF-56) was also purified from egg yolk protein by gel filtration and exhibited high ACE inhibitory activity (69%) and IC50 value (3.35 mg/mL). The SDNRNQGY peptide (10 mg/mL) had ACE inhibitory activity, which was not significantly different from that of the positive control captopril (0.5 mg/mL). In addition, YPSPV in (EYGF-33) (10 mg/mL) had higher ACE inhibitory activity compared with captopril. These findings indicated a substantial potential for producing valuable peptides with antioxidant and ACE inhibitory activity from egg yolk.

Production of the angiotensin I converting enzyme inhibitory peptides and isolation of four novel peptides from jellyfish (Rhopilema esculentum) protein hydrolysate.

Angiotensin I converting enzyme (ACE) plays an important role in regulating blood pressure in the human body. ACE inhibitory peptides derived from food proteins could exert antihypertensive effects without side effects. Jellyfish (Rhopilema esculentum) is an important fishery resource suitable for production of ACE inhibitory peptides. The objective of this study was to optimize the hydrolysis conditions for production of protein hydrolysate from R. esculentum (RPH) with ACE inhibitory activity, and to isolate and identify the ACE inhibitory peptides from RPH. Rhopilema esculentum protein was hydrolyzed with Compound proteinase AQ to produce protein hydrolysate with ACE inhibitory activity, and the hydrolysis conditions were optimized using response surface methodology. The optimum parameters for producing peptides with the highest ACE inhibitory activity were as follows: hydrolysis time 3.90 h, hydrolysis temperature 58 °C, enzyme:substrate ratio 2.8% and pH 7.60. Under these conditions, the ACE inhibitory rate reached 32.21%. In addition, four novel ACE inhibitory peptides were isolated, and their amino acids sequences were identified as Val-Gly-Pro-Tyr, Phe-Thr-Tyr-Val-Pro-Gly, Phe-Thr-Tyr-Val-Pro-Gly-Ala and Phe-Gln-Ala-Val-Trp-Ala-Gly, respectively. The IC50 value of the purified peptides for ACE inhibitory activity was 8.40, 23.42, 21.15 and 19.11 µmol L(-1) . These results indicate that the protein hydrolysate prepared from R. esculentum might be a commercial competitive source of ACE inhibitory ingredients to be used in functional foods. © 2015 Society of Chemical Industry.

Angiotensin-I-converting enzyme-inhibitory peptides in commercial Wisconsin Cheddar cheeses of different ages

Bioactive peptides, including angiotensin-I-converting enzyme-inhibitory (ACEI) peptides, were investigated in commercially produced Wisconsin Cheddar cheeses that ranged in age from ≤6d to more than 2 yr. The ACEI activity of cheese was determined in water-soluble extracts (WSE) that were fractionated for components with molecular weight (MW) ≤3,000 Da, and peptides identified using HPLC and tandem mass spectrometry. The number of types of bioactive peptides increased with an increase in ripening time. Six of the identified ACEI peptides, Ile-Pro-Pro (IPP), Val-Pro-Pro (VPP), Glu-Lys-Asp-Glu-Arg-Phe (EKDERF), Val-Arg-Tyr-Leu (VRYL), Tyr-Pro-Phe-Pro-Gly-Pro-Ile-Pro-Asn (YPFPGPIPN), and Phe-Phe-Val-Ala-Pro (FFVAP), with known high ACEI activity (low IC50 values, the concentration needed to inhibit ACE to 50% of its original activity) were synthesized and used to quantify the amounts of these peptides in various cheese extracts. The concentrations of these 6 ACEI peptides increased up to a certain stage of ripening. The maximum contents of IPP, VPP, and EKDERF were 2.8, 7.4, and 5.3mg/100 g of cheese, respectively, and these levels were found in a 1-yr-old Cheddar cheese sample. The maximum content of VRYL (7.5mg/100 g of cheese) was found in a 2-yr-old Cheddar cheese sample, whereas the maximum content of YPFPGPIPN (6.8mg/100 g of cheese) was found in a 6-mo-old Cheddar cheese sample. Trace amounts of FFVAP were found in these cheeses. Aged Cheddar cheese was found to be a rich source of ACEI peptides even though large differences exist between cheeses from different manufacturers.