Flaxseed Protein Hydrolysates Research Articles

Flaxseed- and chia seed-derived protein hydrolysates exhibiting enhanced in vitro antidiabetic, anti-obesity, and antioxidant properties

This study investigated in vitro antidiabetic, anti-obesity, and antioxidant activities of chia seed protein hydrolysates (CSPHs) and flaxseed protein hydrolysates (FSPHs) generated using three food-grade proteases at the hydrolysis at intervals of 120 min for 6 h. The inhibitory potentials of CSPHs and FSPHs on the enzymatic biomarkers related to diabetes (α-glucosidase and DPP-4) and obesity (pancreatic lipase and cholesteryl esterase) were determined. The antioxidant capacity of CSPHs and FSPHs was also assessed using ABTS, DPPH, and FRAP antioxidant assays. Increasing the proteolytic reaction time was shown to significantly increase the degree of hydrolysis values of CSPHs and FSPHs. Protein hydrolysates resulted in higher inhibitory potential against enzymatic biomarkers related to metabolic diseases (diabetes and obesity) as well as higher antioxidant activities compared with undigested proteins. Overall, the lowest IC50 inhibition values demonstrated in DPP-IV were observed by alcalase-catalyzed CSPH at 6 h (IC50 = 272.17 μg/mL) and bromelain-catalyzed FSPH at 6 h (IC50 = 338.70 μg/mL). The strongest inhibition of pancreatic lipase was observed in bromelain-catalyzed hydrolysates (IC50 = 292.43 μg/mL and IC50 = 307.62 μg/mL for CSPH and FSPH, respectively) after 6 h. These findings indicate that CSPHs and FSPHs exhibited enhanced antidiabetic, anti-obesity, and antioxidant properties. Therefore, these hydrolysates can be successfully used for their applications in the development of functional foods.

Investigation of cryoprotectants-treated surimi protein deterioration during chilled and frozen storage: Functional properties and kinetic modeling.

The relative cryoprotective effects of flaxseed protein hydrolysate and pectin in comparison with conventional cryoprotectant (sucrose + sorbitol + sodium tripolyphosphates) on stabilization of proteins in surimi of Capoor (Cyprinus carpio) were investigated during freezing (-20°C for 4 months) and chilling storage (4°C for 10 days). Although pectin caused to improve water-holding capacity (27.8%; 4°C and 21.5%; -20°C) on account of highly more inhibitory impact on the ice crystals growth, the protein denaturation may have occurred. It can be related to higher reduction in the amount of salt extractable protein (%) and the immeasurable value of thiol group in surimi formulation containing pectin compared with other cryoprotectants. The results of modeling surimi samples showed that salt extractable protein and sulfhydryl content were in good agreement with the first-order reaction model at -20°C and second-order kinetic model at 4°C. In comparison with other samples, samples treated with flaxseed protein showed the lowest reaction rate constant during chilled and frozen storage. The results confirmed that flaxseed protein with no sweetness and considerable caloric value had a cryoprotective effect similar to sucrose + sorbitol + polyphosphate and even better.

Production, characterization, and storage stability of nutritionally enriched flaxseed‐based spread

Flaxseed has recently gained interest as a functional food. The bioactive properties of deoiled flaxseed protein hydrolysates make them suitable for use as preservatives in natural food products. Spreads made from nuts and seeds have recently been used as a substitute for dairy butter. In this study, production, physical properties, oxidative stabilities, antioxidant activities, and storage stabilities of flaxseed-based spreads were investigated. Germinated deoiled flaxseed flour, raw deoiled flaxseed protein hydrolysate, and their combination may be regarded as novel functional food ingredients for spread production. Results showed that the raw deoiled flaxseed protein hydrolysate spread had higher oxidative stabilities. The total phenolic content (24.41 ± 0.17 mgGAE/g), total flavonoid content (18.34 ± 0.03 mgCE/g), and antioxidant activities of raw deoiled flaxseed protein hydrolysate spread had also significantly higher than the other spreadable products. These results indicated that the raw deoiled flaxseed protein hydrolysate could be a good nutritive ingredient to produce spread. Novelty impact statement In this study, germinated deoiled flaxseed flour, raw deoiled flaxseed flour protein hydrolysate, and a combination of both had been considered as novel functional food ingredients for spread production. This study provides detailed information about the oxidative stability and antioxidant activity of these plant-based spreads during storage conditions. In addition, this study also identified the functional group of the spreads by FTIR analysis.

Analysis of key precursor peptides and flavor components of flaxseed derived Maillard reaction products based on iBAQ mass spectrometry and molecular sensory science

Flaxseed derived Maillard reaction products (MRPs) have typical meaty flavor, but there is no report on comparison of their amino acids and peptides reactivity. The peptides and amino acids of flaxseed protein hydrolysates were separately collected by G-15 gel chromatography. Taste dilution analysis (TDA) showed that peptides-MRPs had high umami, mouthfulness, and continuity enhancement. Further, LC-MS/MS revealed that flaxseed protein hydrolysates consumed 41 peptides after Maillard reaction. Particularly, DLSFIP (Asp-Leu-Ser-Phe-Ile-Pro) and ELPGSP (Glu-Leu-Pro-Gly-Ser-Pro) accounted for 42.22% and 20.41% of total consumption, respectively. Aroma extract dilution analysis (AEDA) indicated that formation of sulfur-containing flavors was dependent on cysteine, while peptides were more reactive than amino acids for nitrogen-containing heterocycles. On the other hand, 11 flavor compounds with flavor dilution (FD) ≥ 64 were identified for flaxseed derived MRPs, such as 2-methylthiophene, 2-methyl-3-furanthiol, furfural, 2-furfurylthiol, 3-thiophenethiol, thieno[3,2-b] thiophene, 2,5-thiophenedicarboxaldehyde, 2-methylthieno[2,3-b] thiophene, 1-(2-methyl-3-furylthio)-ethanethiol, 2-methylthieno[3,2-b] thiophene, and bis(2-methyl-3-furyl)-disulfide. In addition, we further demonstrated the flavors formation mechanism of flaxseed derived MRPs.

Flaxseed protein: extraction, functionalities and applications

Flaxseed (Linum usitatissimum L.), one kind of common oilseeds, has many biologically active compounds including α-linolenic acid (ALA), lignan and dietary fiber. At present, the research of flaxseed is focused on biological and clinical studies of its compounds. However, the studies of flaxseed protein obtained generally from flaxseed cake, are relatively limited. As many other kinds of plant proteins, flaxseed protein presents many excellent functionalities which can be used for food applications. Many evidence have showed that flaxseed protein hydrolysates can provide health benefits to our body, such as anti-bacterial activity, antioxidant capacity and angiotensin-converting enzyme inhibition ability. Therefore, in order to make better use of flaxseed protein in foods, the extraction, functionalities and applications of flaxseed protein were reviewed in this paper. In addition, the preparation of flaxseed protein hydrolysates was discussed.

Ameliorative effect of enhanced Fischer ratio flaxseed protein hydrolysate in combination with antioxidant micronutrients on ethanol-induced hepatic damage in a rat model

Alcohol abuse causes severe metabolic abnormalities inducing hepatic damage and malnutrition. Since higher Fischer ratio proteins have therapeutic value in liver diseases, an investigation was undertaken to study the ameliorative effect of the enhanced Fischer ratio flaxseed protein hydrolysate (EFR-FPH) alone and in combination with antioxidant micronutrients on ethanol-induced hepatotoxicity in a rat model. The EFR-FPH was prepared by dual enzymatic hydrolysis and charcoal treatment of flaxseed protein. The ratio of the branched-chain:aromatic amino acids (Fischer ratio) was found to be 7·08. The EFR-FPH, characterised using LC-MS/MS, showed the abundance of free leucine and isoleucine compared with phenylalanine and tyrosine. The matrix-assisted laser desorption/ionisation-time of flight MS analysis revealed the larger peptides present in EFR-FPH with mass 2·3 kDa. The EFR-FPH improved the nutritional status, liver function and antioxidant defense in the ethanol hepatotoxicity-induced rat model. The hepatoprotective effect of EFR-FPH was significantly enhanced when combined with selenium or vitamin E. Ethanol-induced changes in the liver tissue were effectively suppressed in the groups receiving EFR-FPH. Flaxseed-based hepatoprotective dietary supplement was formulated incorporating an optimum level of EFR-FPH (10 %) based on sensory acceptability and was fortified with selenium and vitamin E. The hepatoprotective formulation significantly lowered aspartate transaminase, alanine transaminase, alkaline phosphatase and bilirubin by 47, 61, 55 and 78 %, respectively, and improved the antioxidant defense in the ethanol hepatotoxicity-induced rat model. The current investigation suggests that EFR-FPH in synergy with antioxidant micronutrients is potent in ameliorating ethanol-induced hepatotoxicity and has a potential to form a hepatoprotective dietary supplement.

Biological Potential of Flaxseed Protein Hydrolysates Obtained by Different Proteases.

Flaxseed meal, a byproduct of flaxseed oil extraction, was treated as low-value agrowaste for a long time despite its high protein content. Flaxseed meal has recently garnered increasing interest as a source of proteins and other bioactive compounds with positive impacts on human health. The aim of this study was to investigate the in vitro biological potential of flaxseed protein hydrolysates (FPH). Three FPHs were prepared using three hydrolytic enzymes: Alcalase, Neutrase and Protamex. The molecular weight profile of peptides contained in the hydrolysates was determined by size exclusion chromatography (SEC). The oxygen radical absorbance capacity (ORAC) assay was used to determine the peptide antioxidant capacity, while proliferative effects were studied in two cell lines: HeLa and HaCaT. The latter was also used to determine the protective effect of the FPH during induced oxidative stress. Alcalase showed the highest proteolytic activity, while the produced flaxseed protein hydrolysate (FPH-A) exhibited the strongest antioxidant potential. FPH-A had cytotoxic effects at 10mg/mL in HeLa cells, but it stimulated HaCaT cell growth. Moreover, a mild protective effect of FPH-A was detected in HaCaT cells after induction of oxidative stress. Flaxseed protein hydrolysates obtained by Neutrase (FPH-N) and Protamex (FPH-P) have less pronounced or no potential at all, with respect to their antioxidative or antiproliferative activity. Therefore, to increase value-added utilization of flaxseed meal we suggest further research on hydrolysate obtained by Alcalase.

Influence of spray drying encapsulation on the retention of antioxidant properties and microstructure of flaxseed protein hydrolysates.

In this research, bioactive peptides produced from flaxseed protein by alcalase, pancreatin, trypsin and pepsin, were encapsulated by spray drying. After analysis of amino acid composition and antioxidant properties of hydrolysates, the effect of spray-drying encapsulation via different maltodextrin (MD) to hydrolysate ratios (1:1, 2:1 and 3:1 w/w) on the production yield, physicochemical properties, functional activities, chemical structure, and morphology of final powder particles were evaluated. Among the hydrolysates, peptides produced with alcalase had the highest hydrolysis degree (38.2%), hydrophobic amino acids (255 mg/g) and antioxidants (126 mg/g). Among spray-dried samples, the powders obtained by 3:1 w/w ratio (MD: peptide) showed the highest radical scavenging activity for DPPH- (68.93%), ABTS+ (85.62%), hydroxyl (94.97%), nitric oxide (64.03%), reducing power (95.49%), total antioxidant activity (96.68%), and iron (95.31%) and copper (95.49%) chelating activity. Evaluation of chemical structure (FTIR) indicated that hydrolysates were coated and dispersed within maltodextrin matrix. SEM images showed the effect of different carrier ratios on the production of irregular and shrunk particles with different sizes and matrix-type structures.

Importance of Flaxseed and its Components in the Management of Hypertension.

This review paper describes the effects of flaxseed and its components (flax oil, secoisolariciresinol diglucoside [SDG], flax lignan complex [FLC], and flaxseed protein hydrolysate [FPH]) on blood pressure (BP) in Sprague Dawley rats (SDR), spontaneously hypertensive rats (SHR), and humans. Flaxseed, flax oil, and FLC had variable effects on BP in humans, while SDG and FPH significantly reduced the BP in SDR and SHR. The effect of SDG was dose-dependent and long lasting. The lowering of BP is mediated through inhibition of soluble epoxide by α -linolenic acid in flax oil, stimulation of guanylate cyclase and inhibition of angiotensin converting enzyme (ACE) by SDG, and inhibition of renin and ACE activity by FPH. Flaxseed, flax oil, and FLC have variable effects on BP (none, slight, and significant). They are effective in lowering BP in individuals with hypertension and metabolic syndrome but ineffective in healthy individuals' ineffectiveness of flaxseed and its compounds in lowering BP may be due to their low doses, long interval of dosing, short duration of consumption, and patient status. In conclusion, the data at present suggest that flaxseed, flax oil, and FLC cannot serve as therapeutic agents for the treatment of hypertension. However, they can be used as an adjunct in the treatment of hypertension. A clinical trial should be conducted of these agents with higher doses which would be given twice daily for long duration. Pure SDG and FPS may serve as therapeutic agents for the treatment of hypertension but they have not been tried in humans.

Color and flavor of flaxseed protein hydrolysates Maillard reaction products: effect of cysteine, initial pH, and thermal treatment

ABSTRACT The effects of cysteine, initial pH, and thermal treatment on the color and flavor derived from Maillard reaction of flaxseed protein hydrolysates were investigated. Browning and color of Maillard reaction products (MRPs) were inhibited at higher cysteine concentration and lower initial pH. Higher cysteine or initial pH suppressed the cross-linking of 128–1,000 Da peptides and accelerated the degradation of peptides with a higher molecular weight (MW) (>1,000 Da). The higher temperature and longer reaction time resulted into a marked increase in flavor formation in the Maillard reaction model. Partial least squares regression (PLSR) revealed that volatile compounds, MW distribution, and amino acids of MRPs had a significant influence on MRPs sensory characteristics. The generation of umami, mouthfulness, and continuity was observed at relatively lower temperatures and longer heating time. The sulfur- and nitrogen-containing compounds from the low MW peptides (<1,000 Da) may lead to meaty aroma in MRPs.

High hydrostatic pressure-assisted enzymatic hydrolysis improved protein digestion of flaxseed protein isolate and generation of peptides with antioxidant activity

Exploration of innovative high hydrostatic pressure (HHP)-assisted enzymatic hydrolysis of plant based food proteins may help improve peptide yield and bioactivity of hydrolysates. In this study, we performed enzymatic hydrolysis of flaxseed proteins using trypsin under HHP (100 and 300 MPa for 5 and 10 min) to evaluate the effect of presurization on protein denaturation, degree of hydrolysis (DH), and peptide profile and bioactivity of hydrolysate. Spectrofluorimetric analyses showed that 300 MPa induced the maximum destablization of flaxseed protein structures. The same pressure level drastically improved the DH by 1.7 times as compared to that of control. Applying HHP did not modify the peptide profiles of flaxseed protein hydrolysates but their concentrations increased with severity of treatment. Similarly, peptide molecular weight distributions were affected by pressurization parameters, increasing mainly the relative abundance of 500–1500 Da peptides. Finally, pressurization at 300 MPa for 5 and 10 min improved the antioxidant activity of flaxseed protein hydrolysates by 39 and 55%, respectively, compared to the control.

Impact of a high hydrostatic pressure pretreatment on the separation of bioactive peptides from flaxseed protein hydrolysates by electrodialysis with ultrafiltration membranes

High hydrostatic pressure (HHP) and electrodialysis with ultrafiltration membranes (EDUF) are two efficient technologies used respectively to improve protein enzymatic hydrolysis and recovery of bioactive peptides, but they have never been tested together. Hence, in this study, HHP pre-treatment was performed on defatted flaxseed protein isolate prior to enzymatic hydrolysis and the resulting peptides were separated by EDUF. HHP pretreatment influenced particle size, protein conformation, and degree of hydrolysis. After EDUF separation, peptide fractions (generated after enzymatic hydrolysis of control and pressure-treated protein isolate) recovered in KCl fraction were enriched in arginine and associated with a decrease in systolic blood pressure (SBP) in spontaneously hypertensive rats. Additionally, the final EDUF hydrolysate generated from pretreated protein and the initial EDUF hydrolysate from native protein were also associated with lower SBP. However, only the control KCl fraction obtained from native protein hydrolysate was associated with anti-diabetic activity.

Enzymatic hydrolysis of flaxseed (Linum usitatissimum L.) protein and sensory characterization of Maillard reaction products

We aimed to simplify the enzymolysis process for flaxseed protein hydrolysates production as Maillard reaction products (MRPs) to generate different flavor characteristics. More than 50% activity of immobilized enzymes (Alcalase and Flavourzyme) was retained after repeated use. Subsequently, after five weeks, the activities of the immobilized enzymes were also observed to be higher after storage at 4 °C. The optimum conditions for production of flaxseed protein hydrolysates using sequential enzymatic hydrolysis were as follow: 3,000 U/g of Alcalase at 60 °C and pH 8.0 for 2 h and 120 U/g of Flavourzyme at 50 °C and pH 6.5 for 2 h. Partial least squares regression analysis revealed that resulting peptides with the molecular weight (MW) higher than 1,000 Da could improve the mouthfulness and stability in umami soup; whereas, peptides with MW of 128–1,000 Da mainly contributed to the generation of meat-like flavor compounds with a significant effect on umami taste and bitterness.

Antioxidant Properties of Flaxseed Protein Hydrolysates: Influence of Hydrolytic Enzyme Concentration and Peptide Size

AbstractFlaxseed is an emerging source of protein raw materials for bioactive protein hydrolysate production; however, there is insufficient information on the structural properties of its antioxidative peptides. The aim of this work was to determine the peptide composition and relationships to antioxidant properties of flaxseed protein hydrolysates (FPH) obtained using two protease (thermoase) concentrations (2.5 and 3.0%). The FPH were passed through ultrafiltration membranes to yield fractions with &lt;1, 1–3, 3–5, and 5–10 kDa peptide sizes, which were also tested for antioxidant properties. Mass spectrometry data yielded several peptide sequences located within the flaxseed conlinin, but a lesser number was found in the 3.0% thermoase‐FPH. 2,2‐Diphenyl‐1‐picrylhydrazyl radical scavenging was lowest (54%) for the 3.0% thermoase‐FPH while membrane separation enhanced the values up to 65% for the &lt;1 kDa peptides. Metal chelation and iron‐reducing ability were also significantly (P &lt; 0.05) enhanced after membrane fractionation. In contrast, the hydroxyl radical scavenging ability was unchanged (2.5% thermoase‐FPH) or significantly (P &lt; 0.05) diminished (3.0% thermoase‐FPH) after membrane fractionation, except for the 5–10 kDa peptides that had notably (P &lt; 0.05) higher values. We conclude that the &lt;1 and 1–3 kDa peptides may be useful agents to suppress oxidative stress within living tissues while the 3–5 and 5–10 kDa could be suitable preservatives to enhance food shelf life.

Effect of Trypsin on Antioxidant Activity and Gel-Rheology of Flaxseed Protein

Abstract Enzymatic hydrolysis of flaxseed protein (FP) was carried out using trypsin in order to obtain flaxseed protein hydrolysates possessing better antioxidative property and modified rheological properties. The antioxidative properties of hydrolysates were much higher than the unhydrolyzed flaxseed protein. The hydrolysis also significantly reduced the hydrodynamic diameter of the magnitude of zeta potential of the dispersions. The gelling point of the hydrolysates occurred earlier than the unhydrolyzed sample while the duration of hydrolysis (30–120 min) did not affect gelling point of the hydrolysates. Considerable decrease in the gel strength and the frequency dependence of gel strength were observed in gels produced using hydrolyzed flaxseed protein. The above findings indicate that hydrolysates possessing high degree of antioxidative properties. The gels produces from these hydrolysates will have fast gelling property and will produce gels with reasonable strength. Thus, flaxseed protein hydrolysates obtained from trypsin hydrolysis can be used in applications that require proteins with higher antioxidative properties but softer texture.

Use of Different Proteases to Obtain Flaxseed Protein Hydrolysates with Antioxidant Activity.

The antioxidant activity of flaxseed protein hydrolysates obtained using five different enzymes was evaluated. Proteins were isolated from flaxseed cake and were separately treated with papain, trypsin, pancreatin, Alcalase and Flavourzyme. The degree of hydrolysis (DH) was determined as the percentage of cleaved peptide bonds using a spectrophotometric method with o-phthaldialdehyde. The distribution of the molecular weights (MW) of the hydrolysis products was profiled using Tricine-sodium dodecyl sulfate-polyacrylamide gel electrophoresis (Tricine-SDS-PAGE) and size exclusion-high performance liquid chromatography (SE-HPLC) separations. The antioxidant activities of the protein isolate and hydrolysates were probed for their radical scavenging activity using 2,2′-azino-bis-(3-ethylbenzothiazoline-6-sulfonate) radical cation (ABTS•+) and photochemiluminescence (PCL-ACL) assays, and for their ferric reducing antioxidant power (FRAP) and ability to bind Fe2+. The hydrolysates were more effective as antioxidants than the protein isolate in all systems. The PCL-ACL values of the hydrolysates ranged from 7.2 to 35.7 μmol Trolox/g. Both the FRAP and ABTS•+ scavenging activity differed among the hydrolysates to a lower extent, with the ranges of 0.20–0.24 mmol Fe2+/g and 0.17–0.22 mmol Trolox/g, respectively. The highest chelating activity (71.5%) was noted for the pancreatin hydrolysate. In general, the hydrolysates obtained using Alcalase and pancreatin had the highest antioxidant activity, even though their DH (15.4% and 29.3%, respectively) and the MW profiles of the peptides varied substantially. The O2•− scavenging activity and the ability to chelate Fe2+ of the Flavourzyme hydrolysate were lower than those of the Alcalase and pancreatin hydrolysates. Papain was the least effective in releasing the peptides with antioxidant activity. The study showed that the type of enzyme used for flaxseed protein hydrolysis determines the antioxidant activity of the hydrolysates.

Thermoase-derived flaxseed protein hydrolysates and membrane ultrafiltration peptide fractions have systolic blood pressure-lowering effects in spontaneously hypertensive rats.

Thermoase-digested flaxseed protein hydrolysate (FPH) samples and ultrafiltration membrane-separated peptide fractions were initially evaluated for in vitro inhibition of angiotensin I-converting enzyme (ACE) and renin activities. The two most active FPH samples and their corresponding peptide fractions were subsequently tested for in vivo antihypertensive activity in spontaneously hypertensive rats (SHR). The FPH produced with 3% thermoase digestion showed the highest ACE- and renin-inhibitory activities. Whereas membrane ultrafiltration resulted in significant (p < 0.05) increases in ACE inhibition by the <1 and 1–3 kDa peptides, only a marginal improvement in renin-inhibitory activity was observed for virtually all the samples after membrane ultrafiltration. The FPH samples and membrane fractions were also effective in lowering systolic blood pressure (SBP) in SHR with the largest effect occurring after oral administration (200 mg/kg body weight) of the 1–3 kDa peptide fraction of the 2.5% FPH and the 3–5 kDa fraction of the 3% FPH. Such potent SBP-lowering capacity indicates the potential of flaxseed protein-derived bioactive peptides as ingredients for the formulation of antihypertensive functional foods and nutraceuticals.

Anti-diabetic and antihypertensive activities of two flaxseed protein hydrolysate fractions revealed following their simultaneous separation by electrodialysis with ultrafiltration membranes

Flaxseed protein hydrolysate has been fractionated by electrodialysis with two ultrafiltration membranes (20 and 50kDa) stacked in the system for the recovery of two specific cationic peptide fractions (KCl-F1 and KCl-F2). After 360min of treatment, peptide migration increased as a function of time in KCl compartments. Moreover, the use of two different ultrafiltration membrane allowed concentration of the 300–400 and 400–500Da molecular weight range peptides in the KCl-F1 and KCl-F2 fractions, respectively, compared to the initial hydrolysate. After mass spectrometry analysis, higher amounts of low molecular weight peptides were recovered in the KCl-F2 compartment while relatively higher molecular weight peptides were more detected in the KCl-F1 compartment. Amino acid analysis showed that His, Lys and Arg were especially concentrated in the KCl compartments. Finally, glucose-transport assay demonstrated that the KCl-F2 fraction increased glucose uptake while oral administration of KCl-F1 and final FPH decreased systolic blood pressure.

Antioxidant Capacity of Flaxseed Products: The Effect of In vitro Digestion

This study evaluated the effect of in vitro digestion of flaxseed products on Folin-Ciocalteu reagent reducing substances (FCRRS), its antioxidant capacity and prevention of oxidative DNA damage in human monocyte cell line U937. Flaxseed protein isolate was obtained from defatted flaxseed meal and the protein hydrolysate with high antioxidant capacity was obtained from hydrolysis of the protein isolate with Alcalase in a two factor central composite rotatable design (pH 8.5 and enzyme: substrate 1:90, w/w). The FCRRS content and antioxidant capacity measured by FRAP and ORAC in aqueous and 70% methanol extracts were the highest in protein hydrolysate, followed by protein isolate, while the defatted meal showed the lowest values. After in vitro gastrointestinal digestion, the FCRRS content of protein isolate and hydrolysate reached similar values, however the hydrolysate had the highest antioxidant capacity, measured by FRAP while the isolate had the highest ORAC values. The defatted meal showed the lowest capacity in all assays (p < 0.05). The hydrolysate did not protect against DNA damage induced by H2O2 in U937 cells under the conditions of the present study. The results suggest that flaxseed protein isolate and hydrolysate are potential functional food ingredients with antioxidant capacity.

Multifunctional Cationic Peptide Fractions from Flaxseed Protein Hydrolysates

The aim of this work was to determine the multifunctional properties of flaxseed protein-derived cationic peptide fractions. Alcalase hydrolysis of flaxseed protein fractions liberated cationic peptides, which were separated into two major fractions (FI and FII) by chromatography using a cation-exchange column. Due to their cationic property, the peptide fractions bound and inactivated calmodulin (CaM, a negatively charged enzyme activator protein) with concomitant inhibition of CaM-dependent phosphodiesterase (CaMPDE); this activity was substantially reduced as CaM concentration increased. Enzyme kinetics studies showed competitive inhibition of CaMPDE by FI and FII with enzyme-inhibitor dissociation constants of 0.0202 and 0.0511mg/ml, respectively. Only the FII peptides showed multifunctional activities by inhibiting CaMPDE, angiotensin converting enzyme (ACE) and renin. Separation of FII peptides by reverse phase HPLC resulted in eight fractions (FII-2 to FII-9) that inhibited the activities of CaMPDE, ACE, and renin but this multifunctional activity was more pronounced in FII-6. From LC-MS analysis, identified peptides present in FII fraction had molecular size range of 330-735Da, which suggests potential for increased absorption. Potential peptide sequences were identified for each of the HPLC fractions and shown to contain either lysine or arginine as the positively charged amino acid residue. The multifunctional properties of the cationic peptide fractions can potentially enhance their use in targeting multiple symptoms of cardiovascular disease, considering that the excessive levels of CaM, CaMPDE, renin and ACE play important roles in enhancing progression and intensity of chronic human diseases.