Peptide Fractions Research Articles

Separation and Purification of Antioxidant Peptide from Fermented Whey Protein by Lactobacillus rhamnosus B2-1.

In this study, a antioxidant activity peptide fraction was separated and purified from metabolites of whey protein fermented by Lactobacillus rhamnosus B2-1. The fermentation sample was separated by macroporous resin D101 and Sephadex G-15. The collected fractions were tested for antioxidant and antitumor activities. In order to test the antioxidant activity of fractions, Hydroxyl (·OH), 2,2'-azino-bis (3-ethylbenzothiazoline-6-sulfonic acid (ABTS), and Oxygen Radical Absorbance Capacity (ORAC) were used. The final purified peptide B11 showed highest ABTS and ·OH radical scavenging rate by 84.36±1.89% and 62.43±2.64%, respectively, and had an ORAC activity of 1,726.44± 2.76 μM Trolox equivalent/g. Further, the inhibitory effect of B11 on the proliferation of LoVo human colon cancer cells, KB and Cal-27 human oral cancer cells were enhanced with increasing concentrations of B11. B11 contains 51.421% amino acids, with Glu and Asp being the major constituents. In this study, we obtained peptide fraction B11 with antioxidant activity, which is promising for development.

The peptide fraction of Bothrops jararaca snake venom induces toxicological effects on the male reproductive system after local envenomation in mice

Bothrops envenomation is complex and provokes prominent local tissue damage and systemic disturbances, but little is known about their effects on the male reproductive system. After intratesticular injection, the bioactive peptide fraction (Bj-PF) obtained from Bothrops jararaca snake venom changes the structure of different stages of the seminiferous epithelium cycle in adult mice. For the first time, we investigated whether local envenomation of Bj-PF induces toxicological effects on the male reproductive system, particularly on the seminiferous epithelium and Sertoli cells. Male adult mice were treated with 0.24 mg.kg−1 by intramuscular (i.m.) injection for 24 h. The testes samples were collected for morphological and morphometric evaluation. The toxicological effects of Bj-PF were also analyzed on mitochondrial metabolism and nitrite (NO2) production in 15P-1 Sertoli cell culture. Bj-PF changed the structure and function of the seminiferous epithelium, particularly the disruption of the epithelium and the presence of degenerated germ cells in the adluminal compartment, but there were no alterations in the basal compartment. Bj-PF increased the thickness of the seminiferous epithelium and decreased the lumen diameter of the tubule. Semiquantitative histological assessment of the degree of tubule degeneration revealed that Bj-PF also increased the number of hypospermatogenic tubules compared to control. Bj-PF reduced NO2 levels in 15P-1 Sertoli cells without changing the mitochondrial metabolism. Overall, the fact that Bj-PF alters the structure and function of the seminiferous epithelium suggests that bioactive peptides found in B. jararaca snake venom can have toxicological effects on the reproductive systems of affected male mice, providing new insight into the biological characteristics of snake venom and therapeutic strategies for envenomation inflammation.

Myocardial Mechanical Dispersion Predicts Adverse Cardiac Remodeling in Patients with ST Segment Elevation Myocardial Infarction Who Underwent Primary Percutaneous Coronary Intervention.

The aim of the study was to investigate whether increased left ventricular mechanical dispersion is an early predictor for adverse cardiac remodeling in ST-segment elevation myocardial infarction patients who had post-percutaneous coronary intervention thrombolysis in myocardial infarction (TIMI) flow grade > 2. A total of 119 post-percutaneous coronary intervention ST elevation myocardial infarction patients with TIMI flow grade >2 were prospectively included in the study. Left ventricular global longitudinal strain was quantified by 2-dimensional speckletracking echocardiography, and left ventricular mechanical dispersion was determined at baseline and after 1 year to assess adverse cardiac remodeling. The levels of circulating biomarkers were measured at the baseline. TIMI score and the Global Registry of Acute Coronary Events score systems were used to evaluate the prognosis of patients. Patients with high quartile versus low quartile of left ventricular mechanical dispersion exerted higher Global Registry of Acute Coronary Events and TIMI score grades, left ventricular endsystolic volume, global longitudinal strain, and levels of the N-terminal fragment of brain natriuretic peptide and lower left ventricular ejection fraction. Multivariate log regression showed that N-terminal fragment of brain natriuretic peptide > 953 pg/mL, global longitudinal strain > -8%, and high quartile of left ventricular mechanical dispersion remained independent predictors for adverse cardiac remodeling. Addition of left ventricular mechanical dispersion to the N-terminal fragment of brain natriuretic peptide improved the discriminative potency of the whole model. Measurement of left ventricular mechanical dispersion might be useful in determining the risk of adverse cardiac remodeling in post-percutaneous coronary intervention ST elevation myocardial infarction patients.

Application of assisted extraction technologies to optimize the extraction of biologically active substances from plant materials

There is an increased interest in obtaining amaranth grain extracts enriched with biologically active substances by deep processing of raw materials with a further prospect of their use as part of specialized multi-purpose food products. The aim of this work was to evaluate the effectiveness of membrane filtration for the extraction of biologically active substances from amaranth grains. Amaranth grain extract was obtained by enzymatic hydrolysis using a proteolytic enzyme followed by membrane treatment. the developed approach made it possible to obtain an extract from amaranth grains - a source of biologically active peptides, 20-hydroxyecdysone and polyphenols (including saponins). In total, triterpene saponins were concentrated in the extract by 8 times, and the phytoecdysteroid 20-hydroxyecdysone was more than 30 times concentrated. The high content of low molecular weight peptide fractions (more than 80%) in the composition of the extract makes it promising to evaluate its antihypertensive and antidiabetic properties in vivo. The presence of 20-hydroxyecdysone in the extract, an adaptogen with a proven effect, opens up the possibility of its use as a functional food ingredient in specialized food products, including for athletes.

Inhibitory effect of casein and alpha-lactalbumin on Chryseobacteria spp. isolated from milk and dairy products

In this study, pathogenic and food spoilage bacteria Chryseobacteria spp. were isolated by a new method from 150 samples of raw milk (26/50), heat-treated milk (HTM) (5/50) and butter (2/50). The species with the highest prevalence was C. indologenes (10.67%) followed by C. bovis and C. gleum (4.67% each), and C. bernardetii (2%). The three new strains C. bernardetii, C. gleum, and C. indologenes were named MW703610, MW703611, and MW703612, respectively, with identities ranging from 97.8% to 98.9%. Gene sequencing was performed on some isolates and proved the highest sequence similarity for C. bernardetii WD1 (97.8%), C. gleum WD2 (98.9%) and C. indologenes WD3 (98.9%). Most of the isolated Chryseobacteria spp. showed proteolytic and lipolytic activity. Therefore, casein and alpha-lactalbumin were used as natural antibacterial substances against all isolates. Higher bioactivity against the isolates was obtained using casein rather than the alpha-lactalbumin. Based on these results, casein and alpha-lactalbumin may be promising natural bioactive agents against pathogenic Chryseobacteria spp. Further studies should be done to establish the natural peptide fractions associated with the observed activity.

Clinical Proteomics: Liquid Chromatography-Mass Spectrometry (LC-MS) Purification Systems.

Liquid chromatography/mass spectrometry (LC/MS) has become a routine powerful technology in clinical proteomic studies for protein identification, protein characterization, and the discovery of biomarkers. In this chapter, we describe two protocol methods to analyze clinical patient samples using a resin-based depletion column followed by either protein In-Gel enzymatic digestion or protein In-Solution enzymatic digestion using a simple kit-based approach (i.e., using the PreOmics iST sample preparation kit), followed by analysis using one-dimensional reverse-phase chromatography (RPC) or high pH reversed-phase peptide fractionation.

In vitro gastrointestinal digestion of buckwheat (Fagopyrum esculentum Moench) protein: release and structural characteristics of novel bioactive peptides stimulating gut cholecystokinin secretion.

Satiety hormone cholecystokinin (CCK) plays a vital role in appetite inhibition. Its secretion is regulated by dietary components. The search for bioactive compounds that stimulate CCK secretion is currently an active area of research. The objective of this study was to evaluate the ability of buckwheat (Fagopyrum esculentum Moench) protein digest (BPD) to stimulate CCK secretion in vitro and in vivo and clarify the structural characteristics of peptides stimulating CCK secretion. BPD was prepared by an in vitro gastrointestinal digestion model. The relative molecular weight of BPD was <10 000 Da, and peptides with <3000 Da accounted for 70%. BPD was rich in essential amino acids Lys, Leu, and Val but lacked sulfur amino acids Met and Cys. It had a stimulatory effect on CCK secretion in vitro and in vivo. Chromatographic separation was performed to isolate peptide fractions involved in CCK secretion, and five novel CCK-releasing peptides including QFDLDD, PAFKEEHL, SFHFPI, IPPLFP, and RVTVQPDS were successfully identified. A sequence length range of 6-8 and marked hydrophobicity (18-28) were observed among the most CCK-releasing peptides. The present study demonstrated for the first time that BPD could stimulate CCK secretion and clarify the structural characteristics of bioactive peptides having CCK secretagogue activity in BPD.

Offline Peptide Fractionation and Parallel Reaction Monitoring MS for the Quantitation of Low-Abundance Plasma Proteins.

Mass spectrometry (MS)-based protein quantitation is an attractive means for research and diagnostics due to its high specificity, precision, sensitivity, versatility, and the ability to develop multiplexed assays for the "absolute" quantitation of virtually any protein target. However, due to the large dynamic range of protein concentrations in blood, high abundance proteins in blood plasma hinder the detectability and quantification of lower-abundance proteins which are often relevant in the context of different diseases. Here we outline a streamlined method involving offline high-pH reversed-phase fractionation of human plasma samples followed by the quantitative analysis of specific fractions using nanoLC-parallel reaction monitoring (PRM) on a Q Exactive Plus mass spectrometer for peptide detection and quantitation with increased sensitivity. Because we use a set of synthetic peptide standards, we can more efficiently determine the precise retention times of the target peptides in the first-dimensional separation and specifically collect eluting fractions of interest for the subsequent targeted MS quantitation, making the analysis faster and easier. An eight-point standard curve was generated by serial dilution of a mixture of previously validated unlabeled ("light") synthetic peptides of interest at known concentrations. The corresponding heavy stable-isotope-labeled standard (SIS) analogues were used as normalizers to account for losses during sample processing and analysis. Using this method, we were able to improve the sensitivity of plasma protein quantitation by up to 50-fold compared to using nanoLC-PRM alone.

Tracking the Biostimulatory Effect of Fractions from a Commercial Plant Protein Hydrolysate in Greenhouse-Grown Lettuce.

Protein hydrolysate biostimulants are environmentally friendly options for the reduction of nitrogen input, but their plant growth-promoting mechanisms are still not completely unveiled. Here, to put the “signaling peptide theory” to the test, a greenhouse experiment was undertaken using low (1 mM) and optimal (8 mM) NO3-treated butterhead lettuce and three molecular fractions (PH1 (>10 kDa), PH2 (1−10 kDa) and PH3 (<10 kDa) fractions), in addition to the whole product Vegamin®: PH, in a randomized block design. PH1 and PH3 significantly increased fresh yield (+8%) under optimal (lighter leaves), but not under low (darker leaves) NO3 conditions. Total ascorbic acid, lutein and β-carotene increased with PH3, and disinapoylgentobiose and kaempferol-3-hydroxyferuloyl-sophorosie-7-glucoside content increased with PH (whole/fractions) treatments, particularly under low NO3 conditions. The complete hydrolysate and analyzed peptide fractions have differential biostimulatory effects, enhancing the growth and nutritional quality of lettuce.

Protease-Assisted Mild Extraction of Soluble Fibre and Protein from Fruit By-Products: A Biorefinery Perspective.

By-products from the fruit supply chain, especially seeds/kernels, have shown great potential to be valorised, due to their high content of macronutrients, such as lipids, protein, and fibre. A mild enzymatic assisted extraction (EAE) involving the use of a protease was tested to evaluate the feasibility of a cascade approach to fractionate the main fruit by-products components. Protease from Bacillus licheniformis (the enzyme used in the AOAC 991.43 official method for dietary fibre quantification) was used, and besides protein, the conditions of hydrolysis (60 °C, neutral pH, overnight) allowed us to dissolve a portion of soluble fibres, which was then separated from the solubilized peptide fraction through ethanol precipitation. Good protein extraction yields, in the range 35-93%, were obtained. The soluble fibre extraction yield ranged from 1.6% to 71% depending on the by-product, suggesting its applicability only for certain substrates, and it was found to be negatively correlated with the molecular weight of the fibre. The monosaccharide composition of the soluble fibres extracted was also diverse. Galacturonic acid was present in a low amount, indicating that pectin was not efficiently extracted. However, a predominance of arabinose and galactose monomers was detected in many fractions, indicating the isolation of a fruit soluble fibre portion with potential similarity with arabinogalactans and gum arabic, opening up perspectives for technological applications. The residual solid pellet obtained after protease assisted extraction was found to be an excellent fibre-rich substrate, suitable for being subjected to more "hard" processing (e.g., sequential pectin and hemicellulose extraction) with the objective to derive other fractions with potential great added economic value.

Antioxidant, Anti-Diabetic, Anti-Obesity, and Antihypertensive Properties of Protein Hydrolysate and Peptide Fractions from Black Sesame Cake.

A low-value by-product of cold-pressed sesame oil is defatted black sesame cake (DBSC). The remaining protein and essential amino acids may be utilized as a renewable biological source to produce bioactive products. The bioactivities of the protein hydrolysate from black sesame cake and its peptide fractions were examined in this study for in vitro antioxidant activity and inhibition of DPP-IV, ACE, α-amylase, α-glucosidase, and pancreatic lipase. By using Flavourzyme to hydrolyze DBSC, followed by ultrafiltration, fractions with peptide sizes of <3, 3−10, and >10 kDa were obtained. According to the findings, the products of DBSC could neutralize free radicals and prevent ferric ion redox reactions. The highest inhibitory effects were shown with low Mw peptides (<3 kDa) against ACE, DPP-IV, α-amylase, and α-glucosidase. DBSC has demonstrated potential as a nutraceutical or functional ingredient for preventing and treating disorders associated with free radicals, such as diabetes, hypertension, and hyperglycemia.

Bile acid-binding capacity of peptide extracts obtained from chicken blood hydrolysates using HPLC

Bile acids are involved in the modulation of various metabolic processes facilitating the biliary excretion of endogenous and exogenous cholesterol. The objective of this study was to determine the glycocholic acid binding capacity (BC) of chicken blood hydrolysates using an optimized RP-HPLC methodology. Samples were hydrolysed using a combination of five different enzymes. Alcalase and Protamex hydrolysates presented the highest BC, with mean values of 20.09% and 20.61%, respectively. Subsequently, both hydrolysates were ultrafiltered to obtain fractions >10 kDa, between 10 and 3 kDa, and <3 kDa, and the highest BC values were obtained for peptide fractions >10 kDa. Finally, the protein fragments (MW > 10 kDa) potentially responsible for BC were identified by LC-MS/MS. The results confirmed the relation of BC with the molecular weight of the peptides generated, suggesting that certain protein fragments generated from chicken blood could contribute to a positive impact on health by interfering with cholesterol metabolism.

Quantitative Proteomics Analysis of Purified Rat Liver Golgi.

The Golgi is the central organelle in the secretory pathway, essential for post-translational modifications, sorting and trafficking of secretory and membrane proteins and lipids in all eukaryotic cells. During mitosis, the mammalian Golgi membranes undergo continuous disassembly and reassembly processes which are critical for Golgi biogenesis during the cell division. To better understand the underlying molecular mechanism of this highly dynamic process, we analyzed the proteins that are in or associated with interphase and mitotic Golgi membranes using an in vitro Golgi assembly assay and quantitative proteomics. In this study, by combining an isobaric mass tag labeling strategy with OFFGEL peptide fractionation, LC-MS/MS analyses identified and quantified a total of 1193 Golgi-resident or -associated proteins. These proteins included Golgi structural proteins, Golgi-resident enzymes, Rab GTPases, and SNARE proteins. This systematic quantitative proteomic study revealed the comprehensive molecular machinery of the Golgi and the dynamic protein changes in its disassembly and reassembly processes. Here we describe the detailed procedures and protocols for this analysis.

Bioactive properties of peptide fractions from Brazilian soy protein hydrolysates: In silico evaluation and experimental evidence

Soybeans are a known source of dietary proteins and potential bioactive peptides. In this study, a protein hydrolysate from soybean protein concentrate was produced using papain. The peptides were separated by ultrafiltration (< and > 3 kDa, LMMH (low molecular mass) and HMMH (high molecular mass), respectively) and sequenced through LC-MS/MS. To obtain a thorough identification of the peptides in the hydrolysate, different analysis methods and bioinformatics techniques were applied, covering a molecular mass range from more than 480 Da up to small dipeptides. The antioxidant and the inhibitory α -glucosidase and lipase potentials were evaluated by different in vitro tests. Sixty-nine peptides were identified in the HMMH fraction and 32 in LMMH, but only 16 matched the 118 sequences obtained by in silico simulated hydrolysis. Unlike previous reports, the HMMH fraction showed higher antioxidant activity, by all 5 in vitro methods applied, which was not accompanied by the in silico evaluation. Both high and low molecular mass fractions showed similar inhibitory activities against α -glucosidase and pancreatic lipase. LMMH, however, showed better results for α -glucosidase inhibition (IC50 = 0,94), in agreement with the in silico evaluation. This combination of bioactivities makes the fractions of this hydrolysate potential food ingredients with the possible ability to delay the lipid peroxidation of meat products, limiting the digestion of lipids in the product and also with the potential to delay the digestion of carbohydrates ingested in the same meal.

A Novel Peptide with Antifungal Activity from Red Swamp Crayfish Procambarus clarkii.

The defense system of freshwater crayfish Procambarus clarkii as a diversified source of bioactive molecules with antimicrobial properties was studied. Antimicrobial activity of two polypeptide-enriched extracts obtained from hemocytes and hemolymph of P. clarkii were assessed against Gram positive (Staphylococcus aureus, Enterococcus faecalis) and Gram negative (Pseudomonas aeruginosa, Escherichia coli) bacteria and toward the yeast Candida albicans. The two peptide fractions showed interesting MIC values (ranging from 11 to 700 μg/mL) against all tested pathogens. Polypeptide-enriched extracts were further investigated using a high-resolution mass spectrometry and database search and 14 novel peptides were identified. Some peptides and their derivatives were chemically synthesized and tested in vitro against the bacterial and yeast pathogens. The analysis identified a synthetic derivative peptide, which showed an interesting antifungal (MIC and MFC equal to 31.2 μg/mL and 62.5 μg/mL, respectively) and antibiofilm (BIC50 equal to 23.2 μg/mL) activities against Candida albicans and a low toxicity in human cells.

Hatched Eggshell Membrane Can Be a Novel Source of Antioxidant Hydrolysates to Protect against H2O2-Induced Oxidative Stress in Human Chondrocytes.

Natural antioxidants derived from agricultural by-products have great promise and ecological advantages in the treatment of oxidative stress-related diseases. The eggshell membrane (ESM) from hatched eggs, i.e., the hatched ESM, is a globally abundant agricultural byproduct, and its high-value utilization has been rarely studied compared to the well-studied ESM from fresh eggs. In this research, we systematically characterized the hatched ESM as a novel source of antioxidant hydrolysates and explored their potential role in H2O2-induced human chondrocytes. The results showed that the hatched ESM is a protein-rich fibrous mesh material with a significantly different structure and composition from those of fresh ESM. Enzymatic hydrolysis of hatched ESM can produce antioxidant hydrolysates rich in low molecular weight (MW) peptides, which mainly derived from the Lysyl oxidase homolog by Nano-LC-MS/MS analysis. The peptide fraction with MW &lt; 3 kDa (HEMH-I) exhibited the highest DPPH radical scavenging, Fe2+-chelating, and Fe3+-reducing abilities. In H2O2-induced human SW1353 chondrocytes, HEMH-I treatment significantly increased the cell viability and ameliorated oxidative stress, inflammatory response, and cartilage matrix degradation by reducing the level of ROS, matrix metalloprotease 3 (MMP3), MMP13, and IL-6, and by promoting the expression of SOD and type II collagen, potentially through activating the cellular Keap1/Nrf2/HO-1 pathway. This study provides a theoretical basis for the value-added application of hatched ESM waste to produce antioxidant hydrolysates and indicates their potential as functional food and pharmaceuticals.

Novel Peptide Sequences with ACE-Inhibitory and Antioxidant Activities Derived from the Heads and Bones of Hybrid Groupers (Epinephelus lanceolatus × Epinephelus fuscoguttatus).

The heads and bones of hybrid groupers are potential precursors for angiotensin-converting enzyme (ACE)-inhibitory and antioxidant peptides. The aim of this study was to isolate the dual-action peptides from the Alcalase-treated head and bone hydrolysate of hybrid groupers followed by identification of the novel peptides. The stability of these peptides against stimulated in vitro gastrointestinal digestion (SGID) was also determined. Fraction HB-IV (less than 1 kDa) obtained from ultrafiltration showed the strongest ACE-inhibition ability (IC50: 0.28 mg/mL), which was comparable to the potency of the commercial supplement, PeptACE (IC50: 0.22 mg/mL). This fraction also demonstrated the highest hydroxyl radical scavenging and metal-chelating activities. However, further fractionation of HB-IV by a series of chromatography resulted in peptide fractions of reduced ACE-inhibitory and antioxidant activities. The hydroxyl radical scavenging and reduction potential of HB-IV were enhanced, whereas ACE-inhibitory and metal-chelating activities were reduced following SGID. A total of 145 peptide sequences were identified from HB-IV, of which 137 peptides were novel to the BIOPEP database. The results suggested that the bioactive peptides isolated from the heads and bones of hybrid groupers could be used as functional foods/ingredients with potential ACE-inhibitory and antioxidant effects.

The Emerging Role of uORF-Encoded uPeptides and HLA uLigands in Cellular and Tumor Biology.

Recent technological advances have facilitated the detection of numerous non-canonical human peptides derived from regulatory regions of mRNAs, long non-coding RNAs, and other cryptic transcripts. In this review, we first give an overview of the classification of these novel peptides and summarize recent improvements in their annotation and detection by ribosome profiling, mass spectrometry, and individual experimental analysis. A large fraction of the novel peptides originates from translation at upstream open reading frames (uORFs) that are located within the transcript leader sequence of regular mRNA. In humans, uORF-encoded peptides (uPeptides) have been detected in both healthy and malignantly transformed cells and emerge as important regulators in cellular and immunological pathways. In the second part of the review, we focus on various functional implications of uPeptides. As uPeptides frequently act at the transition of translational regulation and individual peptide function, we describe the mechanistic modes of translational regulation through ribosome stalling, the involvement in cellular programs through protein interaction and complex formation, and their role within the human leukocyte antigen (HLA)-associated immunopeptidome as HLA uLigands. We delineate how malignant transformation may lead to the formation of novel uORFs, uPeptides, or HLA uLigands and explain their potential implication in tumor biology. Ultimately, we speculate on a potential use of uPeptides as peptide drugs and discuss how uPeptides and HLA uLigands may facilitate translational inhibition of oncogenic protein messages and immunotherapeutic approaches in cancer therapy.

Integrated Evaluation of Dual-Functional DPP-IV and ACE Inhibitory Effects of Peptides Derived from Sericin Hydrolysis and Their Stabilities during In Vitro-Simulated Gastrointestinal and Plasmin Digestions.

Sericin, a byproduct of the silk industry, is an underutilized protein derived from the yellow silk cocoon. This research aimed to produce and characterize the bioactive peptides from sericin using various enzymatic hydrolysis methods. Alcalase, papain, neutrase, and protease were tested under their respective digestion conditions. Among the enzymes tested, neutrase-catalyzed sericin into specific peptides with the strongest dipeptidyl peptidase IV (DPP-IV) and angiotensin-converting enzyme (ACE) inhibitory properties. The peptides were subjected to a simulated in vitro gastrointestinal (GI) digestion in order to determine their stability. The GI peptides that were produced by neutrase hydrolysis continued to have the highest DPP-IV and ACE inhibitory activities. The neutrase -digested peptides were then fractionated via ultrafiltration; the peptide fraction with a molecular weight <3 kDa (UF3) inhibited DPP-IV and ACE activities. After being subjected to in vitro blood plasma hydrolysis, the UF3 was slightly degraded but retained its bioactivity. As a result of these findings, sericin peptides can be utilized as novel dietary ingredients that may alleviate some metabolic syndromes via the dual inhibitory properties of DPP-IV and ACE.

A peptide fraction from hardened common beans (Phaseolus vulgaris) induces endothelium-dependent antihypertensive and renal effects in rats

Beans reached the research spotlight as a source of bioactive compounds capable of modulating different functions. Recently, we reported antioxidant and oxidonitrergic effect of a low molecular weight peptide fraction (<3 kDa) from hardened bean (Phaseolus vulgaris) in vitro and ex vivo, which necessitate further in vivo assessments. This work aimed to evaluate the hypotensive effect and the involved physiological mechanisms of the hardened common bean peptide (Phaseolus vulgaris) in normotensive (Wistar) and hypertensive (SHR) animals. Bean flour was combined with a solution containing acetonitrile, water and formic acid (25: 24: 1). Protein extract (PV3) was fractioned (3 kDa membrane). We assessed PV3 effects on renal function and hemodynamics of wistar (WT–normotensive) and spontaneously hypertensive rats (SHR) and measured systemic arterial pressure and flow in aortic and renal beds. The potential endothelial and oxidonitrergic involvements were tested in isolated renal artery rings. As results, we found that PV3: I) decreased food consumption in SHR, increased water intake and urinary volume in WT, increased glomerular filtration rate in WT and SHR, caused natriuresis in SHR; II) caused NO- and endothelium-dependent vasorelaxation in renal artery rings; III) reduced arterial pressure and resistance in aortic and renal vascular beds; IV) caused antihypertensive effects in a dose-dependent manner. Current findings support PV3 as a source of bioactive peptides and raise the potential of composing nutraceutical formulations to treat renal and cardiovascular diseases.