Inhibition properties of dipeptides from salmon muscle hydrolysate on angiotensin I-converting enzyme

We isolated Phe–Leu as an angiotensin I‐converting enzyme (ACE) inhibitor from hydrolysate of chum salmon muscle. The IC₅₀ value of this peptide was 13.6 μm , and it showed non‐competitive inhibition. The reverse sequence dipeptide Leu–Phe also showed ACE inhibitory activity. However, Leu–Phe is much less inhibitory than Phe–Leu with an IC₅₀ value of 383.2 μm . In addition, the inhibition mode was competitive. To investigate the relationship between dipeptide sequence and ACE inhibition properties, we further measured ACE inhibitory activity and inhibition mechanism using six Trp‐containing dipeptides, which had been identified from the same salmon muscle hydrolysate as ACE inhibitory peptides in a previous study. Peptides with Trp as the C‐terminal residue, Ala–Trp, Val–Trp, Met–Trp, Ile–Trp, Leu–Trp showed non‐competitive inhibition. On the other hand, reversed sequence peptides with Trp at the N‐terminal were competitive inhibitors, except Trp–Leu. These results indicate that the sequence of ACE inhibitory dipeptides can affect both inhibitory potency and the inhibition mechanisms.     

IDENTIFICATION OF ANGIOTENSIN I-CONVERTING ENZYME INHIBITORY PEPTIDES DERIVED FROM THE PEPTIC DIGEST OF SOYBEAN PROTEIN

Peptidic fractions which inhibit angiotensin I‐converting enzyme (ACE) were separated from peptic digests of soybean by ion exchange chromatography and gel filtration. Further separation of the peptidic fractions by ODS HPLC afforded active peptides, the amino add sequences of which were identified by Edman's procedure as: Ile‐Ata (inhibitory against ACE with an IC₅₀of 153 μM), Tyr‐Leu‐Ala‐Gly‐Asn‐Gln (14 μM), Phe‐Phe‐Leu (37 μM), Ile‐Tyr‐Leu‐Leu (42 μM), and Val‐Met‐Asp‐Lys‐Pro‐Gln‐Gly (39 μM). The antihypertensive activity of the soybean peptides was also investigated. Peptide fractions (2.0 g/kg body weight, oral administration) markedly towered the blood pressure of spontaneously hypertensive rats (SHRs).     

Purification,Identification, and In Vivo Activity of Angiotensin I‐Converting Enzyme Inhibitory Peptide,from Ribbonfish (Trichiurus haumela) Backbone

Ribbonfish (Trichiurus haumela) backbone is normally discarded as an industrial waste from fish processing. A method of developing angiotensin I‐converting enzyme inhibitory (ACEI) peptides from ribbonfish backbone was previously optimized. The purposes of the study were to characterize the active peptides in the hydrolysate and to evaluate its in vivo activity. Ribbonfish backbone protein hydrolysate prepared by acid protease was fractionated into 4 fractions (I, MW < 1 kDa; II, MW = 1 to 5 kDa; III, MW = 5 to 10 kDa; and IV, MW > 10 kDa) through ultrafiltration membranes. Fraction I, showing the highest ACEI activity, was further purified using consecutive chromatographic techniques including gel filtration and reversed phase high‐performance liquid chromatography. The purified ACE inhibitory peptide was determined to have a molecular weight of 317.25 Da, with a sequence of Leu‐Trp and an IC₅₀ value of 5.6 μM. Systolic blood pressure of spontaneously hypertensive rats was significantly decreased from 181 ± 2.0 to 161.3 ± 2.3 mm Hg after 4 h of oral administration of Leu‐Trp at a dose of 10 mg/kg of body weight. These results indicated that ribbonfish backbone protein could be used for development of antihypertensive agent.     

Isolation and characterization of angiotensin I‐converting enzyme inhibitory peptides from wheat gliadin hydrolysate

Angiotensin I‐converting enzyme (ACE) inhibitory peptide was isolated from wheat gliadin hydrolysate prepared with acid protease. Consecutive purification methods were used for peptide isolation including ion‐exchange chromatography, size‐exclusion chromatography, and reverse‐phase high‐performance liquid chromatography. The amino acid sequence of this peptide was identified as Ile‐Ala‐Pro, and the ACE inhibitory activity (IC₅₀ value) was 2.7 μM . The hypotensive activity of Ile‐Ala‐Pro on spontaneously hypertensive rats was investigated. This peptide inhibited the hypertensive activity of angiotensin I with intravenous injection, and decreased the blood pressure significantly with intraperitoneal administration.     

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

Angiotensin 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.     

Utilisation of chickpea protein isolates for production of peptides with angiotensin I‐converting enzyme (ACE)‐inhibitory activity

Chickpea protein isolates and the protease alcalase were used for the production of protein hydrolysates that inhibit angiotensin I‐converting enzyme (ACE). The highest degree of inhibition was found in a hydrolysate obtained by 30 min of treatment with alcalase. This hydrolysate was used as starting material for the purification of ACE‐inhibitory peptides. After Biogel P2 gel filtration chromatography and HPLC C₁₈ reverse phase chromatography, four peptides with ACE‐inhibitory activity were purified. Two of them were competitive inhibitors of ACE, while the other two were uncompetitive inhibitors. These results show that chickpea proteins are a good source of ACE‐inhibitory peptides when hydrolysed with the protease alcalase. © 2002 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     

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).     

Optimization of peptic hydrolysis parameters for the production of angiotensin I-converting enzyme inhibitory hydrolysate from Acetes chinensis through Plackett-Burman and response surface methodological approaches

BACKGROUND: Angiotensin I‐converting enzyme (ACE) plays an important physiological role in regulating blood pressure. The elevation of blood pressure could be suppressed by inhibiting ACE. ACE inhibitory peptides derived from food proteins could exert antihypertensive effects without side effects. Acetes chinensis is a marine shrimp suitable for the production of ACE inhibitory peptides. The principal objective of this study was to screen for the significant variables, and further to optimize the levels of the selected variables, for the enzymatic production of ACE inhibitory peptides from Acetes chinensis. RESULTS: Plackett–Burman design and response surface methodology were employed to optimize the peptic hydrolysis parameters of Acetes chinensis to obtain a hydrolysate with potent ACE inhibitory activity. The peptic hydrolysis variables were subject to a Plackett–Burman design for screening the main factors. The selected significant parameters such as pH, hydrolysis temperature and enzyme/substrate (E/S) ratio were further optimized using a central composite design. The optimized conditions were: pH 2.5, hydrolysis temperature 45 °C, E/S ratio 17 800 U kg^?1 shrimp and substrate concentration 200 g L^?1. The results showed that 3–5 h hydrolysis could result in a hydrolysate with ACE inhibition IC₅₀ of 1.17 mg mL^?1 and a high DH of 25–27%. CONCLUSION: Plackett–Burman design and RSM performed well in the optimization of peptic hydrolysis parameters of Acetes chinensis to produce hydrolysate with ACE inhibitory activity. A hydrolysate with potent ACE inhibitory activity and high degree of hydrolysis was obtained, so that the yield of ACE inhibitory peptides in it was high. Copyright © 2011 Society of Chemical Industry     

Chymotryptic hydrolysates of α-kafirin,the storage protein of sorghum (Sorghum bicolor) exhibited angiotensin converting enzyme inhibitory activity

Kafirin is the main storage protein (prolamin) in sorghum grains. α-Kafirin, the alcohol soluble fraction, was isolated from sorghum flour. Treatment of α-kafirin with chymotrypsin yielded a hydrolysate which on fractionation, using Sephadex G-25 column, yielded four fractions with significant angiotensin converting enzyme (ACE) inhibitory activity in vitro. The IC₅₀ values of these fractions ranged from 1.3 to 24.3 μg/ml. Two of the fractions were found to be competitively inhibiting the enzyme, while two other fractions were non-competitive inhibitors. These results demonstrate that chymotryptic hydrolysates of sorghum prolamin could serve as a good source of peptides with angiotensin I converting enzyme inhibitory activity.