Black Bean Protein Research Articles

Low pH-shifting treatment would improve functional properties of black turtle bean (Phaseolus vulgaris L.) protein isolate with immunoreactivity reduction.

Low pH-shifting was firstly applied in the black turtle bean (Phaseolus vulgaris L.) protein isolate treatment by acidic (pH 1.0-3.0) buffer incubation for 8h, then was adjusted to pH 7.2 and kept 3h for protein stabilizing. Mild loss of secondary structure was confirmed in the protein isolate after low pH-shifting treatment by CD and FT-IR analyses. Intrinsic fluorescence, UV spectra, surface hydrophobicity, SH content and SDS-PAGE analyses indicated the protein conformation was unfolded with the exposure of much more buried hydrophobic residues, which would result in the enhancement of emulsifying properties, foaming properties and fat holding capacity, and lead to the reduction of solubility and water holding capacity. Furthermore, lower immunoreactivity was observed by the ELISA, and improved digestibility was found in in vitro digestion assay. Our results suggested the low pH-shifting treatments would broaden the application of bean protein isolate with better hydrophobic processing functions and safety.

Investigating the calcium binding characteristics of black bean protein hydrolysate.

The black bean protein has been widely utilized to prepare hydrolysates with different bioactive properties. Herein, we hydrolyzed the black bean protein to prepare hydrolysate with calcium binding activity and characterized its behavior. Our results showed that ficin was superior in obtaining hydrolysate with calcium binding capacity in comparison with trypsin, alcalase and bromelain. In particular, the optimal capacity of ficin hydrolysate reached 77.54 ± 1.61 μg mg-1, where the optimal hydrolysis conditions of ficin were a temperature of 70 °C, a pH value of 6.2, an enzyme concentration of 1.61% and a time of 3 h. This might be due to high proportions of aspartic acid and glutamic acid (35.59%). Further spectral analysis evidenced the formation of hydrolysate-calcium complexes, demonstrating that the interaction between hydrolysate and calcium ions primarily occur on carboxyl oxygen atoms and amino nitrogen atoms. These findings provide a possible utilization of black bean hydrolysate to serve as a calcium supplement nutraceutical to enhance the absorption and bioavailability.

Black bean protein concentrate ameliorates hepatic steatosis by decreasing lipogenesis and increasing fatty acid oxidation in rats fed a high fat-sucrose diet.

The black bean is a legume widely consumed in Latin America, however its consumption has decreased significantly in recent decades. There is evidence that its consumption generates beneficial health effects due in part to the type of protein, resistant starches and polyphenols. Thus, their use in food formulation could impact health status. Therefore, the purpose of the present work was to evaluate the effects of the consumption of a bean protein concentrate (BPC) and a whole cooked bean flour (WCB) on body composition, glucose metabolism and energy expenditure in Wistar rats fed a control diet or high-fat diets with 5% sucrose in the drinking water. With this aim, rats were fed the experimental diets for 10 weeks. The results showed that consumption of either BPC or WCB reduced weight gain and body fat despite the consumption of a high-fat diet. This change was associated with a significant increase in energy expenditure and the capacity to adapt fuel oxidation to fuel availability. As a result, rats fed a bean-based diet had lower circulating glucose and insulin concentrations and normal glucose tolerance, which was associated with decreased expression of lipogenic genes in the liver. These results suggest that the type of protein and bioactive compounds particularly phenolic and flavonoid compounds present in BPC are suitable to improve the formulations used in dietary strategies for subjects with obesity or type 2 diabetes. The addition of legumes to the diet of subjects with insulin resistance, including black beans, could improve their metabolic status.

Application of ultrasound treatment for modulating the structural, functional and rheological properties of black bean protein isolates

SummaryThe modulating effect of ultrasound treatments at varying powers and times on the structural and functional properties of black bean protein isolate (BBPI) was investigated. Compared with native BBPI, low‐power (150 W) and medium‐power (300 W) ultrasound treatments increased the solubility, foaming and emulsifying properties of BBPI, especially at 300 W, 24 min. This effect arises predominantly due to increased exposure of hydrophobic groups, which serve to increase the interactions between the protein and water molecules. Additionally, an increase in the protein surface activity improved the absorption of protein molecules at the oil–water and air–water interfaces. Rheology data showed that increased hydrophobic and hydrogen‐bonding interactions improved the water‐holding capacity of BBPI gels following ultrasound treatment. However, high‐power (450 W) ultrasound treatment weakened the functional properties of BBPI, and this was likely due to the formation of macromolecular BBPI aggregates. Overall, this study indicates that ultrasound treatment could be a promising approach for modulating other plant protein resources as well as expanding the application of black bean protein.

The Effect of (-)-Epigallocatechin-3-Gallate Non-Covalent Interaction with the Glycosylated Protein on the Emulsion Property.

The effect of (−)-epigallocatechin-3-gallate (EGCG) on protein structure and emulsion properties of glycosylated black bean protein isolate (BBPI-G) were studied and compared to native black bean protein isolate (BBPI). The binding affinity of BBPI and BBPI-G with EGCG belonged to non-covalent interaction, which was determined by fluorescence quenching. EGCG attachment caused more disordered protein conformation, leading to a higher emulsification property. Among the different EGCG concentrations (0.10, 0.25, 0.50 mg/mL), the result revealed that the highest level of the emulsification property was obtained with 0.25 mg/mL EGCG. Therefore, the BBPI-EGCG and BBPI-G-EGCG prepared by 0.25 mg/mL EGCG were selected to fabricate oil-in-water (O/W) emulsions. After the addition of EGCG, the mean particle size of emulsions decreased with the increasing absolute value of zeta-potential, and more compact interfacial film was formed due to the higher percentage of interfacial protein adsorption (AP%). Meanwhile, EGCG also significantly reduced the lipid oxidation of emulsions.

Physicochemical and antioxidative characteristics of black bean protein hydrolysates obtained from different enzymes

Black bean is an excellent protein source for preparing hydrolysates, which attract much attention due to their biological activity. The objective of this study was to characterize the physicochemical and antioxidant properties of black bean protein, hydrolyzed by ficin, bromelain or alcalase until 300 min of hydrolysis. Results showed that bromelain and alcalase hydrolysates possessed higher degree of hydrolysis (DH) than that of ficin, thereby presenting different ultraviolet absorption, fluorescence intensity and circular dichroism. Moreover, all hydrolysates possessed the capacity to scavenge DPPH radical with the lowest IC50 of 21.11 μg/mL, as well as to chelate ferrous ion (Fe2+) with the IC50 values ranging from 6.82 to 30.68 μg/mL. Intriguingly, the oxidation of linoleic acid, sunflower oil and sunflower oil-in-water emulsion was remarkedly retarded by the three selected protein hydrolysates, especially by bromelain-treated protein hydrolysate, which might attribute to their high hydrophobicity and emulsifying properties. These findings can provide strong support for black bean protein hydrolysates to be employed in food products acting as natural antioxidant alternatives.

Electrospun Ultrafine Fibers from Black Bean Protein Concentrates and Polyvinyl Alcohol

In this study, ultrafine fibers were produced from black bean protein concentrates (BPCs) and polyvinyl alcohol (PVA) by electrospinning. The BPC was denatured under acidic (pH 2) or basic (pH 11) conditions. Polymer solutions containing different PVA concentrations (11% or 21%, w/v) and different BPC: PVA ratios (50:50 or 75:25, v/v) were used for fiber production. The electrical conductivity and rheological properties of the fiber-forming solutions were evaluated, as well as the morphology, size distribution, infrared spectrum, and thermal properties of the electrospun fibers. The fibers showed a homogeneous morphology and diameters ranging from 115 to 541 nm. Fibers from the solution containing BPC denatured at pH 11, 11% PVA, and 75:25 (v/v) BPC: PVA presented the lowest diameter, and those from BPC denatured at pH 2 had less beads than the fibers obtained from BPC denatured at pH 11. The solution formulation affected the thermal properties of the fibers, with weight loss increases ranging from 39.0% to 60.9%. The polymeric solutions containing PVA and BPC (whether denatured under basic or acidic conditions) resulted in ultrafine electrospun fibers with highly favorable characteristics that could potentially be used for the encapsulation of bioactive compounds and food applications.

Bioactive Peptides from Black Bean Proteins Play a Potential Role in the Prevention of Adipogenesis (P06-119-19)

ObjectivesFood-based products such as common bean peptides may contribute to reduce the risk of complications associated with obesity. The objective of this research was to produce and characterize peptides from common bean protein isolates and evaluate their potential to prevent adipogenesis. MethodsProtein hydrolysates of raw common black bean (BBPH) were generated using a simulated gastrointestinal digestion by either pepsin/pancreatin (p/p) or a protease: alcalase. SDS-PAGE was performed for visualization of the protein profile and the generated peptides were characterized using LC-ESI-MS/MS. Bioactive peptides potential was evaluated using BIOPEP database. The antiadipogenic potential was evaluated by the pancreatic lipase (triglyceride lipases, EC 3.1.1.3) inhibition assay. Determination of lipid accumulation in 3T3-L1 adipocytes was evaluated with Oil Red lipid staining. ResultsHydrolysates either with pepsin/pancreatin or alcalase showed strong inhibitory activity against pancreatic lipase with no statistical differences between them (IC50 = 3.21 and 1.23 mg/mL, respectively). Results from the SDS-PAGE indicate both BBPH had an average size of <15 kDa. It was possible to identify a higher number of peptide sequences with alcalase BBPH than using pepsin/pancreatin. Physical properties of identified peptides from pepsin/pancreatin were: SGNGGGGGASM, pI = 6, net charge = +1; SKPGGGSPVA pI = 10.1, net charge = +1; VELVGPK, pI = 6.9, net charge = +1; KPTTGKGALA, pI = 10.6, net charge = +1; the main reported bioactivities were DPP-IV and ACE inhibitors. BPPH inhibited 3T3-L1 adipocyte differentiation in a dose-dependent manner. Hydrolysates generated by simulated gastrointestinal digestion prevented lipid accumulation on adipocytes in basal conditions (IC50 = 1.08 mg/mL). Pepsin/pancreatin and alcalase BBPH promoted an important decrease in fat accumulation (45.28% and 12.08%, respectively) in comparison to non-treated adipocytes. ConclusionsCommon black bean peptides generated by either simulated gastrointestinal digestion or alcalase play an influential role in lipid metabolism. They could be considered a promising alternative to be integrated as functional ingredients into a food matrix. Funding SourcesScientific Projects Grant “Para Atender problemas Nacionales” 2016 No. 2081. CONACYT, and USDA-NIFA-HATCH project 1,014,457.

Changes on the Structural and Physicochemical Properties of Conjugates Prepared by the Maillard Reaction of Black Bean Protein Isolates and Glucose with Ultrasound Pretreatment.

The conjugates of black bean protein isolate (BBPI) and glucose (G) were prepared via the wet heating Maillard reaction with ultrasound pretreatment. The physicochemical properties of UBBPI-G conjugates prepared by ultrasound pretreatment Maillard reaction had been compared with classical Maillard reaction (BBPI-G). The reaction rate between BBPI and glucose was speeded up by ultrasound pretreatment. A degree of glycation (DG) of 20.49 was achieved by 2 h treatment for UBBPI-G, whereas 5 h was required using the classical heating. SDS-PAGE patterns revealed that the BBPI-G conjugates with higher molecular weight were formed after glycosylation. The results of secondary structure analysis suggested that the α-helix and β-sheet content of UBBPI-G were lower than that of BBPI-G. In addition, UBBPI-G conjugates had exhibited bathochromic shift compared with BBPI by fluorescence spectroscopy analysis. Finally, UBBPI-G achieved higher level of surface hydrophobicity, solubility, emulsification property and antioxidant activity than BBPI and BBPI-G (classical Maillard reaction).

PEGylation may reduce allergenicity and improve gelling properties of protein isolate from black kidney bean (Phaseolus vulgaris L.)

Effects of PEGylation with methoxy polyethylene glycol (mPEG) on gelling properties of black kidney bean protein isolate (BKBPI) were investigated. PEG-protein conjugations were confirmed using SDS-PAGE, and a significant decrease of allergenicity was observed using an ELISA test. Gelling concentration and gelling properties of BKBPI and mPEG-BKBPI with various treatments, including the additions of NaCl and N-ethylmaleimide (NEM) as well as a heat pretreatment were evaluated. Results showed that the critical gelling concentration of PEGylated BKBPI was 75 mg/ml, and the PEGylation resulted in a higher gelling strength and a shorter gelling time compared with the native BKBPI. The increases in gel strengths were observed both in BKBPI and mPEG-BKBPI with the addition of NaCl, and no obvious variations in storage modulus (G') were found after PEGylation even at a high NaCl concentration of 1 mol/l. Meanwhile, retardation of the G' was observed in the PEGylated protein with the addition of NEM. Furthermore, the G' increased significantly after preheating. Therefore, the results suggested that PEGylation was beneficial to the safety and gelling properties of BKBPI and served to extend applications of BKBPI.

Effects of cornstarch on the gel properties of black bean protein isolate.

The effects of common starch (CS) and high amylopectin starch (HAS) from corn on the properties of heat induced black bean protein isolate (BBPI) gels prepared by heating at 95°C for 30 min were investigated by using dynamic oscillatory rheometer, texture analyzer, and scanning electron microscopy (SEM). Compared with BBPI alone, the presence of cornstarch (1-4%, wt/vol) could improve storage modulus (G') and textural properties of BBPI (10%, wt/vol) gels. The mixed system of BBPI and 4% (wt/vol) HAS exhibited the highest G' and formed the gel faster and more easily, which resulted in firmer and more elastic gel than BBPI-CS at all starch concentrations. It was possible that HAS had lower pasting temperature and higher viscosity than CS, which was beneficial to the formation of BBPI gel network and strengthened the stability of network structure. Moreover, it might also be related to the synergistic effect between protein and starch. The CS and HAS existed in the BBPI gel network could bind water, leading to the increase in the water-holding capacity (WHC) of mixed gels, especially 4% (wt/vol) HAS, which was related to homogeneous and compact microstructure with small pores.

Improving antioxidant activity of black bean protein by hydrolysis with protease combinations

SummaryThis study explored the use of a simplex centroid design to produce protein hydrolysates with antioxidant properties using Alcalase® 2.4L, Flavourzyme® 500L and Neutrase® 0.8L. Proteases kinetic parameters and the ultrafiltration of protein hydrolysates were also investigated. The highest antioxidant activity, in the studied conditions, was reached when the mixture of Alcalase® 2.4L and Flavourzyme® 500L was used in the hydrolysates production. The antioxidant power of the black bean proteins, measured by the total antioxidant capacity and reducing power assay, increased after hydrolysis by 31% and 70%, respectively. The black bean proteins hydrolysates fractions (3–30 kDa) showed an antioxidant activity decrease along with a reduction in molecular weight, demonstrating that a set of varied molecular weight peptides was responsible for the antioxidant characteristics of black bean protein hydrolysates.

PEGylation of black kidney bean (Phaseolus vulgaris L.) protein isolate with potential functironal properties.

In this work, we investigated PEGylated black kidney bean protein isolates (BKBPI) by PEG succinimidyl carbonate (PEG-SC), PEG succinimidyl succinate (PEG-SS) and PEG succinimidyl propionate (PEG-SPA) conjugation. The functional properties, thermodynamic stability, in vitro digestion stability, and hemagglutination activity of the modified products were evaluated. The degree of PEGylation was measured, and FTIR analysis revealed that protein-PEG conjugations were formed, and that no obvious changes in water- and fat-holding capacities were observed. The solubility, emulsifying property, and foaming property were all improved through the modification, while, higher thermodynamic stability was achieved with the increase in Td values and reduction of ΔH. The PEGylated proteins were found to be more resistant to in vitro digestion, and the hemagglutination activity was significantly (P < 0.05) decreased, indicating the higher safety of the protein isolate. The results showed that the functional properties, thermodynamic stability, and biological safety of BKBPI were improved by PEGylation, which could serve to increase the applications for this protein.

Effect of glucose glycosylation following limited enzymatic hydrolysis on functional and conformational properties of black bean protein isolate

The effect of glycosylation following limited enzymatic hydrolysis on conformational and functional properties of black bean protein isolate (BBPI) was investigated in this study. The black bean protein hydrolysate (HBBPI) was prepared using alcalase at pH 9.0 and then glycosylated with glucose (G) at 80 °C for different incubation times from 1 to 6 h. Grafted HBBPI with glucose (HBBPI-G conjugates) had a higher molecular weight than HBBPI by SDS-PAGE analysis. HBBPI-G conjugates had a higher content of β-turn and random coil, but a lower content of α-helix and β-sheet structure than BBPI. HBBPI-G conjugates had lower fluorescence intensity and exhibited bathochromic shift compared with BBPI. Subsequently, the functional properties were also evaluated. Results indicated that the emulsifying activity and solubility were obviously improved (P < 0.05) by HBBPI-G conjugates incubated for 4 h with 6.5% degree of hydrolysis compared to BBPI. Additionally, the glycosylation had positive effects on the reducing power and hydroxyl radical scavenging rate. Therefore, the combination of limited hydrolysis and glycation can be used as an effective method for BBPI modification to obtain enhanced functional properties.

Black bean peptides inhibit glucose uptake in Caco-2 adenocarcinoma cells by blocking the expression and translocation pathway of glucose transporters.

The objective was to evaluate the effect of black bean protein fraction (PFRA), and its derived peptides on glucose uptake, SGLT1 and GLUT2 expression and translocation on Caco-2 cells. The effect of treatments was evaluated on glucose uptake, protein expression and localization and gene expression on Caco-2 cells. PFRA (10 mg/mL) lowered glucose uptake from 27.4% after 30 min to 33.9% after 180 min of treatment compared to untreated control (p < 0.05). All treatments lowered GLUT2 expression after 30 min of treatment compared to untreated control (31.4 to 48.6%, p < 0.05). Similarly, after 24 h of treatment, GLUT2 was decreased in all treatments (23.5% to 48.9%) (p < 0.05). SGLT1 protein expression decreased 18.3% for LSVSVL (100 μM) to 45.1% for PFRA (10 mg/mL) after 24 h. Immunofluorescence microscopy showed a decrease in expression and membrane translocation of GLUT2 and SGLT1 for all treatments compared to untreated control (p < 0.05). Relative gene expression of SLC2A2 (GLUT2) and SLC5A1 (SGLT1) was downregulated significantly up to two-fold change compared to the untreated control after 24 h treatment. Black bean protein fractions are an inexpensive, functional ingredient with significant biological potential to reduce glucose uptake and could be used as an adjuvant in the treatment of colorectal cancer.

Bean peptides have higher in silico binding affinities than ezetimibe for the N-terminal domain of cholesterol receptor Niemann-Pick C1 Like-1

Niemann-Pick C1 like-1 (NPC1L1) mediates cholesterol absorption at the apical membrane of enterocytes through a yet unknown mechanism. Bean, pea, and lentil proteins are naturally hydrolyzed during digestion to produce peptides. The potential for pulse peptides to have high binding affinities for NPC1L1 has not been determined. In this study , in silico binding affinities and interactions were determined between the N-terminal domain of NPC1L1 and 14 pulse peptides (5≥ amino acids) derived through pepsin-pancreatin digestion. Peptides were docked in triplicate to the N-terminal domain using docking program AutoDock Vina, and results were compared to those of ezetimibe, a prescribed NPC1L1 inhibitor. Three black bean peptides (−7.2 to −7.0kcal/mol) and the cowpea bean dipeptide Lys-Asp (−7.0kcal/mol) had higher binding affinities than ezetimibe (−6.6kcal/mol) for the N-terminal domain of NPC1L1. Lentil and pea peptides studied did not have high binding affinities. The common bean peptide Tyr-Ala-Ala-Ala-Thr (−7.2kcal/mol), which can be produced from black or navy bean proteins, had the highest binding affinity. Ezetimibe and peptides with high binding affinities for the N-terminal domain are expected to interact at different locations of the N-terminal domain. All high affinity black bean peptides are expected to have van der Waals interactions with SER130, PHE136, and LEU236 and a conventional hydrogen bond with GLU238 of NPC1L1. Due to their high affinity for the N-terminal domain of NPC1L1, black and cowpea bean peptides produced in the digestive track have the potential to disrupt interactions between NPC1L1 and membrane proteins that lead to cholesterol absorption.

Black bean (Phaseolus vulgaris L.) protein hydrolysates: Physicochemical and functional properties

Black bean protein hydrolysates obtained from pepsin and alcalase digestions until 120min of hydrolysis were evaluated by gel electrophoresis, relative fluorescence intensity, emulsifying properties, light micrograph of emulsions and in vitro antioxidant activity. The emulsion stability of the bean protein hydrolysates were evaluated during 30days of storage. The pepsin-treated bean protein hydrolysates presented higher degree of hydrolysis than the alcalase-treated protein hydrolysates. The alcalase-treated bean protein hydrolysates showed higher surface hydrophobicity. Moreover, the protein hydrolysates obtained with alcalase digestion presented higher emulsion stability during 30-days than those obtained from pepsin digestion. The protein concentrate and especially the hydrolysates obtained from alcalase digestion had good emulsion stability and antioxidant activity. Thus, they could be exploited as protein supplements in the diet as nutritional and bioactive foods.

Antioxidant activity of black bean (Phaseolus vulgaris L.) protein hydrolysates

The objective of this work was to study the effect of enzymatic hydrolysis of black bean protein concentrate using different enzymes. Bean proteins were extracted and hydrolyzed over a period of 120 min using the enzymes pepsin or alcalase. The protein hydrolysates’ molecular weight was assayed by electrophoresis and the antioxidant activity was evaluated by the capturing methods of free radicals ABTS●+ and DPPH. Electrophoretic results showed that the bands above 50 kDa disappeared, when the beans protein was subjected to hydrolysis with pepsin. The bean protein hydrolysate obtained by hydrolysis with alcalase enzyme, showed higher antioxidant activity for inhibition of the radical ABTS●+. However, the hydrolysates obtained by hydrolysis with pepsin had higher antioxidant activity for inhibition of the radical DPPH. The use of pepsin and alcalase enzymes, under the same reaction time, produced black bean protein hydrolysates with different molecular weight profiles and superior antioxidant activity than the native bean protein.

Changes in antioxidant and antiinflammatory activity of black bean (Phaseolus vulgaris L.) protein isolates due to germination and enzymatic digestion

Germination is an inexpensive process to improve the nutritional properties of legumes. The effect of germinating black bean seeds on the production of cotyledon protein hydrolysates (CPH) with antioxidant and antiinflammatory activities was analyzed in this research. After simulated enzymatic digestion, the oxygen radical absorbance capacity (ORAC) of CPH obtained from germinated black beans was lower than that observed for raw cotyledons. There were no significant differences among CPH cellular antioxidant activities (CAA), except for the high CAA of the 120min hydrolysate obtained from one day germinated black bean cotyledons. The most significant changes due to germination and enzymatic hydrolysis were observed for the inhibition of nitric oxide (NO) production in macrophages. The NO synthesis inhibition observed for raw CPH was reduced after simulated gastrointestinal digestion but for germinated samples the inhibition was doubled. Peptides derived from cell wall proteins produced during germination could be responsible of antiinflammatory activity.

Optimization of enzymatic production of anti-diabetic peptides from black bean (Phaseolus vulgaris L.) proteins, their characterization and biological potential.

The aim was to optimize the production of bioactive peptides from black bean (Phaseolus vulgaris L.) protein isolate and to determine their biological potential using biochemical and in silico approaches. Protein fractions were generated using eight commercially available proteases after 2, 3 and 4 h and 1:20, 1:30 and 1:50 enzyme/substrate (E/S) ratios. The best combination of conditions to generate anti-diabetic peptides was with alcalase for 2 h and E/S of 1:20; with inhibition values for dipeptidyl peptidase IV (DPP-IV, 96.7%), α-amylase (53.4%) and α-glucosidase (66.1%). Generated peptides were characterized using LC-ESI-MS/MS. Molecular docking analysis was performed to predict individual peptide biological potential using DockingServer®. Peptides EGLELLLLLLAG, AKSPLF and FEELN inhibited DPP-IV more efficiently in silico through free energy interactions of -9.8, -9.6 and -9.5 kcal mol(-1), respectively, than the control sitagliptin (-8.67 kcal mol(-1)). The peptide TTGGKGGK (-8.97 kcal mol(-1)) had higher inhibitory potential on α-glucosidase compared to the control acarbose (-8.79 kcal mol(-1)). Peptides AKSPLF (-10.2 kcal mol(-1)) and WEVM (-10.1 kcal mol(-1)) generated a lower free energy interaction with the catalytic site of α-amylase in comparison with acarbose (-9.71 kcal mol(-1)). Bean peptides inhibited the tested enzymes through hydrogen bonds, polar and hydrophobic interactions. The main bindings on the catalytic site were with ASP192, GLU192 and ARG 253 on DPP-IV; TYR151, HIS201 and ILE235 on α-amylase; and ASP34, THR83 and ASN32 on α-glucosidase. For the first time, a systematic evaluation and characterization of the anti-diabetic peptides from black bean protein isolate is presented with the potential for inhibiting important molecular markers related to diabetes.