Potent ACE-inhibitory Activity Research Articles

Investigation of the regulatory effect of α-chymotrypsin-assisted hydrolysate from Sebastes schlegelii on blood pressure through in vitro, in silico ACE inhibitory activity, and in vivo spontaneously hypertensive rat hypertensive model

This study aimed to screen peptides with antihypertensive effects from the α-chymotrypsin hydrolysate of Sebastes schlegelii (SSA). SSA demonstrated ACE inhibitory activity with an IC50 value of 0.062 ± 0.001 mg/mL. The SSA significantly reduced body weight and systolic blood pressure in the SHR spontaneously hypertensive rat (SHR) model and improved hypertension-induced vasopathy and interstitial fibrosis in heart and vascular tissues, indicating the presence of ACE inhibitory peptides contributing to cardiovascular health. Peptide composition analysis identified fractions of SSA with potent ACE inhibitory activity, particularly those with molecular weights of 5 kDa or less. Further evaluation of ACE activity using Sephadex G-10 separated SSA fractions led to the amino acid sequence analysis of the most effective fraction (SSA-F1). Molecular docking simulations predicted that peptides from SSA-F1 inhibit ACE activity by binding to its active sites. This research suggests the potential of peptides from S. schlegelii for clinical hypertension treatment.

In Vitro and In Silico Studies on Angiotensin I-Converting Enzyme Inhibitory Peptides Found in Hydrophobic Domains of Porcine Elastin.

One of the most striking aspects of the primary structure in the hydrophobic domains of the tropoelastin molecule is the occurrence of the VAPGVG repeating sequence. Since the N-terminal tripeptide VAP of VAPGVG showed a potent ACE inhibitory activity, the ACE inhibitory activity of various derivatives of VAP was examined in vitro. The results showed that VAP derivative peptides VLP, VGP, VSP, GAP, LSP, and TRP exhibited potent ACE inhibitory activities, while the non-derivative peptide APG showed only weak activity. In in silico studies, the docking score S value showed that VAP derivative peptides VLP, VGP, VSP, LSP, and TRP had stronger docking interactions than APG. Molecular docking in the ACE active pocket showed that TRP, the most potent ACE inhibitory peptide among the VAP derivatives, had a larger number of interactions with ACE residues in comparison with APG and that the TRP molecule appeared to spread widely in the ACE pocket, while the APG molecule appeared to spread closely. Differences in molecular spread may be a reason why TRP exhibits more potent ACE inhibitory activity than APG. The results suggest that the number and strength of interactions between the peptide and ACE are important for the ACE- inhibitory potency of the peptide.

Yogurt Fortification by the Addition of Microencapsulated Stripped Weakfish (Cynoscion guatucupa) Protein Hydrolysate.

The aim of the present work was to fortify yogurt by adding a stripped weakfish (Cynoscion guatucupa) protein hydrolysate obtained with the enzyme Protamex and microencapsulated by spray drying, using maltodextrin (MD) as wall material. The effects on the physicochemical properties, syneresis, texture, viscoelasticity, antioxidant and ACE inhibitory activities of yogurt after 1 and 7 days of storage were evaluated. In addition, microbiological and sensory analyses were performed. Four yogurt formulations were prepared: control yogurt (without additives, YC), yogurt with MD (2.1%, YMD), with the free hydrolysate (1.4%, YH) and the microencapsulated hydrolysate (3.5%, YHEn). Yogurts to which free and microencapsulated hydrolysates were added presented similar characteristics, such as a slight reduction in pH and increased acidity, with a greater tendency to present a yellow color compared with the control yogurt. Moreover, they showed less syneresis, the lowest value being that of YHEn, which also showed a slight increase in cohesiveness and greater rheological stability after one week of storage. All yogurts showed high counts of the microorganisms used as starters. The hydrolysate presence in both forms resulted in yogurts with antioxidant activity and potent ACE-inhibitory activity, which were maintained after 7 days of storage. The incorporation of the hydrolysate in the microencapsulated form presented greater advantages than the direct incorporation, since encapsulation masked the fishy flavor of the hydrolysate, resulting in stable and sensorily acceptable yogurts with antioxidant and ACE inhibitory activities.

Bioactivities and ACE-inhibitory peptides releasing potential of lactic acid bacteria in fermented soy milk

This study was designed to evaluate the bioactivities such as β-glucosidase activity, α-galactosidase activity, and the growth behavior of the Lactobacillus cultures in soy milk medium. Ten Lactobacillus cultures were considered in this study. L. fermentum (M2) and L. casei (NK9) were selected due to their better α-galactosidase, β-glucosidase activity and growth behavior in soy milk medium during fermentation. Further, soy milk fermented with M2 showed higher proteolytic activity (0.67 OD) and ACE-inhibitory (48.44%) than NK9 (proteolytic activity: 0.48 OD and ACE-inhibitory activity: 41.33%). Bioactive peptides produced during the fermentation of soy milk using the selected Lactobacillus cultures were also identified with potent ACE-inhibitory activity by MALDI-TOF spectrometry, and the identified ACE inhibitory peptide sequences from fermented soy milk were characterized using Biopep database.Graphical abstract

Identification of Angiotensin I-Converting Enzyme-Inhibitory and Anticoagulant Peptides from Enzymatic Hydrolysates of Chicken Combs and Wattles.

Peptides from protein hydrolysate of a mixture of chicken combs and wattles (CCWs) were obtained through enzymatic hydrolysis, and their anticoagulant and inhibitory effects on angiotensin I-converting enzyme (ACE) were investigated. The protein hydrolysate exhibited anticoagulant capacity by the intrinsic pathway (activated partial thromboplastin time) and potent ACE-inhibitory activity. The peptides were sequenced by LC-MS to identify those with higher inhibitory potential. From the pool of sequenced peptides, the following three peptides were selected and synthesized based on their low molecular weight and the presence of amino acids with ACE-inhibitory potential at the C-terminus: peptide I (APGLPGPR), peptide II (Piro-GPPGPT), and peptide III (FPGPPGP). Peptide III (FPGPPGP) showed the highest ACE-inhibitory capacity among the peptides selected. In conclusion, a peptide (FPGPPGP) of unknown sequence was identified as having potent ACE-inhibitory capacity. This peptide originated from unconventional hydrolysates from poultry slaughter waste, including combs and wattles.

New ACE-inhibitory peptides derived from α-lactalbumin produced by hydrolysis with Bromelia antiacantha peptidases

Abstract Angiotensin converting enzyme (ACE) plays a key role in regulating blood pressure and ACE inhibitors are first-line drugs for treating hypertension. Food protein-derived peptides with ACE inhibitory activity are natural and safe alternatives for both prevention and treatment of hypertension. The second most abundant whey protein, α-lactalbumin (ALA) was hydrolysed with proteases from Bromelia antiacantha Bertol., a native plant. Two new peptides with potent ACE-inhibitory activity: TTFHTSGY (IC50 = 142 μM) and GYDTQAIVQ (IC50 = 1.0 mM) were purified by affinity chromatography and sequences were determined by mass spectrometry. In silico analysis indicated that neither of these peptides was toxic (ToxinPred). Inhibition kinetic analysis showed that TTFHTSGY was a fully functional ACE competitive inhibitor, whereas GYDTQAIVQ caused only partial ACE inhibition. Our results indicate that cysteine peptidases from B. antiacantha hydrolyse ALA to produce novel ACE inhibitory peptides that could be useful even as functional food ingredients.

Identification and characterization of a novel angiotensin I-converting enzyme inhibitory peptide (ACEIP) from silkworm pupa

The angiotensin I-converting enzyme inhibitory peptide (ACEIP) was isolated and characterized from silkworm pupae and purified using Sephadex G-25 gel filtration. The structure and physicochemical properties of pupa ACEIP were analyzed. The α-P3 fraction exhibited the most potent ACE inhibitory activity. After purification via semi-preparative reverse-phase HPLC (RP-HPLC) and HPLC, the α-P3-6-b component was revealed to have the highest ACE inhibitory activity (IC50=28.3 μg/mL). Edman degradation revealed a Val-Glu-Ile-Ser amino acid sequence in which novel active sequences were identified. Physicochemical property testing showed that purified pupa ACEIP exhibits good solubility, heat resistance, and acid resistance that all indicate ACEIP derived from silkworm pupa is an excellent food-derived ACEIP.

Resistance of casein-derived bioactive peptides to simulated gastrointestinal digestion

The resistance of six casein-derived peptides, including antihypertensive peptides RYLGY, AYFYPEL and YQKFPQY, to simulated gastrointestinal digestion and the effect on angiotensin-converting enzyme (ACE)-inhibitory activity were evaluated. After digestion, peptides RYLGY, AYFYPEL, and YQKFPQY were partly hydrolysed by the digestive enzymes. RYLGY and AYFYPEL maintained potent ACE-inhibitory activity, with IC50 values as low as 9.3 and 4.7 μg mL−1, respectively. Digestion fragments were sequenced and then synthesised to evaluate their activity. Several showed potent ACE-inhibitory activity, which could explain the in vitro activity of the digests. A notable antioxidant activity was also observed. Since AYFYPEL was less susceptible to digestion, we focused on the antihypertensive activity in spontaneously hypertensive rats of the main digestion fragments of RYLGY. Interestingly, these peptides showed moderate effects in vivo. This suggests that the undigested fraction could also contribute to the in vivo effects of RYLGY and AYFYPEL, and other minor fragments may also participate.

Optimization of the Extraction of Coconut Protein and the Evaluation of Antioxidant and ACE Inhibitory Activities of its Enzymatic Hydrolysates

The extraction of proteins from coconut meat was performed utilizing alkaline extraction. On the basis of single factor experiment, an orthogonal experiment was designed to optimize the extraction conditions (including extraction time, temperature, NaOH concentration and liquid to solid ratio). The highest extraction rate (7.3%) was obtained at 120 min, 45 °C, 0.1 M and liquid-solid ratio 25:1. Coconut protein was consequently hydrolyzed with four commercial enzymes (alcalase, trypsin, pepsin, papain). Furthermore, activities of hydrolysates on DPPH radical scavenging and ACE inhibition were assayed respectively. The results indicated that pepsin hydrolysate exhibited the highest DPPH radical scavenging activity (100%, 1 mg protein/mL), and trypsin hydrolysate possessed the most potent ACE inhibitory activity (40.2%, 1 mg protein/mL).

Phytochemical screening and evaluation of in vitro angiotensin-converting enzyme inhibitory activity of Artocarpus altilis leaf

This study investigates the effect of Artocarpus altilis leaf extracts on angiotensin-converting enzyme (ACE) activity. Among the extracts tested, hot ethanol extract exhibited a potent ACE-inhibitory activity with an IC50 value of 54.08 ± 0.29 µg mL−1 followed by cold ethyl acetate extract (IC50 of 85.44 ± 0.85 µg mL−1). In contrast, the hot aqueous extracts showed minimum inhibition with the IC50 value of 765.52 ± 11.97 µg mL−1 at the maximum concentration tested. Further, the phytochemical analysis indicated the varied distribution of tannins, phenolics, glycosides, saponins, steroids, terpenoids and anthraquinones in cold and hot leaf extracts. The correlation between the phytochemical analysis and ACE-inhibitory activity suggests that the high content of glycosidic and phenolic compounds could be involved in exerting ACE-inhibitory activity. In conclusion, this study supports the utilisation of A. altilis leaf in the folk medicine for the better treatment of hypertension. Further studies on isolation and characterisation of specific ACE-inhibitory molecule(s) from ethyl acetate, ethanol and methanol extracts of A. altilis leaf would be highly interesting.

Cloning, Soluble Expression, and Production of Recombinant Antihypertensive Peptide Multimer (AHPM-2) in Escherichia coli for Bioactivity Identification

Recombinant antihypertensive peptide multimer (AHPM-2, 8kDa/68AA), a new designed polypeptide with potential antihypertensive effect in vivo, is composed of 15 low-molecular-weight antihypertensive peptides tandemly linked up according to the restriction sites of gastrointestinal proteases. After gene optimization, the DNA fragment encoding AHPM-2 was chemically synthesized, cloned into the pET32a, and successfully expressed in E.coli, above 90% in a soluble form. After chromatographic purification, the expressed fusion protein Trx-AHPM-2 was subject to the simulated gastrointestinal digestion, and the hydrolysate showed potent ACE inhibitory activity with an IC50 value of 4.5±0.3 μg ml-1. The active fragments from the AHPM-2 were identified by UPLC-MS/MS. This method will be useful in obtaining an appreciable quantity of recombinant AHP at low cost, and the intact AHPM-2 is expected to be developed into functional food for preventing hypertension as well as for therapeutic.

Antihypertensive Properties of Dried Radish Leaves Powder in Spontaneously Hypertensive Rats

The study aim was to investigate the antihypertensive effect after oral supplementation of dried radish leaves powder (DRLP). Angiotensin-converting enzyme (ACE) activity was measured by spectrophotometric assay. The systolic blood pressure (SBP) was measured in spontaneously hypertensive (SHR) and normotensive rats (Wistar) by the tail cuff method after a 4-week diet with DRLP at the level of 2.5% or 5%. The supplementation of DRLP decreased SBP of SHR although the 5% supplementation level did not show any more pronounced effect than the 2.5% level did. The decrease in the SBP observed for both 2.5% and 5% DRLP was accompanied by significant increases of the urinary Na and K excretion. The DRLP supplementation showed a potent ACE-inhibitory activity in pulmonary tissue from both hypertensive and normotensive rats. However, the DRLP supplementation did not affect the SBP in normotensive rats. These results indicated that DRLP exerted an antihypertensive effect in SHR due to the decreased ACE activity and increased urinary Na excretion. (Korean J Nutr 2010; 43(6): 561 ~ 569)

ACE-inhibitory and antihypertensive properties of a bovine casein hydrolysate

The aim of this study was to investigate the potential angiotensin converting enzyme (ACE)-inhibitory activity and the antihypertensive effect, after a single oral administration, of a pepsin hydrolysed bovine casein (HBC) and a fraction with molecular mass lower than 3000 Da (HBC < 3000). ACE-inhibitory activity was measured by spectrophotometric assay. These products were orally administered by gastric intubation. The systolic (SBP) and the diastolic blood pressure (DBP) were measured in spontaneously hypertensive rats by the tail cuff method before administration and also 2, 4, 6, 8, and 24 h post-administration. HBC showed a potent ACE-inhibitory activity. This activity was 10 times higher in HBC < 3000. HBC and HBC < 3000 decreased the arterial blood pressure of the rats. The decrease in the SBP observed for HBC (400 mg/kg) or HBC < 3000 (200 mg/kg) was less pronounced than that caused by 50 mg/kg of captopril (antihypertensive positive control). However, the maximal decreases in DBP caused by HBC or HBC < 3000 were as high as the maximum decrease observed for captopril. The antihypertensive effect of these products was transient and reverted 24 h after the administration. HBC and HBC < 3000 exert antihypertensive effect caused by small peptides with ACE-inhibitory activity.

Casein hydrolysates as a source of antimicrobial, antioxidant and antihypertensive peptides

The aim of this work was to investigate the presence of antioxidant and ACE-inhibitory activity in ovine αs2-casein and bovine κ-casein hydrolysates with antibacterial activity. Several pep- tides which had been previously identified in these hydrolysates were selected in order to fulfil cer- tain structural requirements and they were chemically synthesised to evaluate their antioxidant and ACE-inhibitory activity. Hydrolysates of ovine αs2-casein and bovine κ-casein with pepsin strongly inhibited ACE activity, with IC50 values of 41.8 and 9.97 μmol·L −1 , respectively. The κ-casein hy- drolysate also exhibited a significant oxygen radical absorbance capacity, seven times higher than that of Trolox. From the chemically synthesised peptides, two of them, LKKISQ and PYVRYL, both from ovine αs2-casein, exerted potent ACE-inhibitory activity in the range of the most po- tent food-derived antihypertensive peptides described to date (IC50 values of 2.6 and 2.4 μmol·L −1 , respectively). The latter sequence, corresponding to the C-terminal hexapeptide of the ovine αs2- casein molecule, also had antioxidant activity. The activity found is discussed in relation to the peptide sequences. αs2-casein / κ-casein / antibacterial activity / antioxidant activity / angiotensin-converting enzyme-inhibitory activity / bioactive peptide

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 (IC 50) of 21 μ m 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 IC 50 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.

Production of the blood pressure lowing peptides from brown alga (Undaria pinnatifida)

Brown alga (Undaria pinnatifida) was treated with alginate lyase and hydrolyzed using 17 kinds of proteases and the inhibitory activity of the hydrolysates for the angiotensin-I-converting enzyme (ACE) was measured. Four hydrolysates with potent ACE-inhibitory activity were administered singly and orally to spontaneously hypertensive rats (SHRs). The systolic blood pressure of SHRs decreases significantly after single oral administration of the brown alga hydrolysates by protease S ‘Amano’ (from Bacillus stearothermophilus) at the concentration of 10 (mg protein) (kg body weight)−1. In the 17 weeks of feeding experiment, 7-week-old SHRs were fed standard diet supplemented with the brown alga hydrolysates for 10 weeks. In SHRs fed 1.0 and 0.1% brown alga hydrolysates, elevating of systolic bloodpressure was significantly suppressed for 7 weeks. To elucidate the active components, the brown alga hydrolysates were fractionated by 1-butanol extraction and HPLC on a reverse-phase column. Seven kinds of ACE-inhibitory peptides were isolated and identified by amino acid composition analysis, sequence analysis, and LC-MS with the results Val-Tyr, Ile-Tyr, Ala-Trp, Phe-Tyr, Val-Trp, Ile-Trp, and Leu-Trp. Each peptide was determined to have an antihypertensive effect after a single oral administration in SHRs. The brown alga hydrolysates were also confirmed to decrease the blood pressure in humans.

Isolation and antihypertensive effect of angiotensin I-converting enzyme (ACE) inhibitory peptides from spinach Rubisco.

Four new inhibitory peptides for angiotensin I-converting enzyme (ACE), that is, MRWRD, MRW, LRIPVA, and IAYKPAG, were isolated from the pepsin-pancreatin digest of spinach Rubisco with the use of HPLC. IC(50) values of individual peptides were 2.1, 0.6, 0.38, and 4.2 microM, respectively. MRW and MRWRD had an antihypertensive effect after oral administration to spontaneously hypertensive rats. Maximal reduction occurred 2 h after oral administration of MRW, whereas MRWRD showed maximal decrease 4 h after oral administration at doses of 20 and 30 mg/kg, respectively. IAYKPAG also exerted antihypertensive activity after oral administration at the dose of 100 mg/kg, giving a maximum decrease 4 h after oral administration. IAYKP, IAY, and KP, the fragment peptides of IAYKPAG, also exerted antihypertensive activity. LRIPVA [corrected] did not show any antihypertensive effect at a dose of 100 mg/kg despite its potent ACE-inhibitory activity.

食品タンパク質由来機能性ペプチドによる血圧降下作用 イワシペプチド（Ｖａｌ‐Ｔｙｒ）による降圧食品の開発を中心として

The procedures for preparing a food material with antihypertensive efficacy from sardine muscle have been reviewed. Hydrolysis of sardine muscle by pepsin yielded angiotensin I-converting enzyme (ACE)-inhibitory peptides. Further studies aimed at preparing a hydrolyzate with potent ACE-inhibitory activity as well as having good flavor were carried out using Bacillus licheniformis alkaline protease. After 17h of hydrolysis of 0.3wt% sardine protein, the 10wt% ethanol fraction obtained with ODS resin showed some excellent properties, including strong ACE-inhibitory activity (IC50=0.015mg protein/mL), high digestive resistance, no bitterness, and no antigenicity. In addition, this fraction was confirmed to have a significant in vivo depressor effect in mildly hypertensive volunteers (n=29), who showed a 9.3mmHg reduction in systolic blood pressure and 5.2mmHg reduction in diastolic blood pressure after 4 weeks of oral administration of this peptide drink (4g of the ODS fraction/200mL/day).

Antihypertensive effects of angiotensin fragments in SHR.

When angiotensin fragments, Val-Tyr and Angiotensin III (ANG III), with potent ACE inhibitory activity were intravenously administered to spontaneously hypertensive rat (SHR), a significant reduction of diastolic blood pressure was observed. After incubation of ANG III with SHR plasma, four fragments with ACE inhibitory activity, Val-Tyr (ANG (3-4)) (IC50 = 26.0 microM), Ile-His-Pro-Phe (ANG (5-8)) (11.6 microM), Tyr-Ile-His-Pro-Phe (ANG (4-8)) (457.5 microM), and Val-Tyr-Ile-His-Pro-Phe (ANG (3-8)) (6.55 microM), were confirmed to generate in SHR plasma. Compared the metabolic behavior of ANG II in SHR plasma with that in normotensive Wistar plasma, the initial degradation rate (3.07 nmol/ml/min) in Wistar plasma was about 2-fold higher than that in the SHR one (1.75 nmol/ml/min).