Pea Protein Hydrolysate Research Articles

Amino acid composition and antioxidant properties of pea seed ( Pisum sativum L.) enzymatic protein hydrolysate fractions.

The amino acid composition and antioxidant activities of peptide fractions obtained from HPLC separation of a pea protein hydrolysate (PPH) were studied. Thermolysin hydrolysis of pea protein isolate and ultrafiltration (3 kDa molecular weight cutoff membrane) yielded a PPH that was separated into five fractions (F1-F5) on a C(18) reverse phase HPLC column. The fractions that eluted later from the column (F3-F5) contained higher contents hydrophobic and aromatic amino acids when compared to fractions that eluted early or the original PPH. Fractions F3-F5 also exhibited the strongest radical scavenging and metal chelating activities; however, hydrophobic character did not seem to contribute to reducing power of the peptides. In comparison to glutathione, the peptide fractions had significantly higher (p < 0.05) ability to inhibit linoleic acid oxidation and chelate metals. In contrast, glutathione had significantly higher (p < 0.05) free radical scavenging properties than the peptide fractions.

The impact of pea protein hydrolysates on bacterial physiological activity—An in vitro study

So far, food proteins have been perceived hitherto purely as a source of nutrients indispensable for maintaining life. However, latest findings strongly indicate that food proteins may release biologically active peptides in a consequence of enzymatic degradation. Such hydrolysates may be used as food components in order to beneficially influence human health. Additionally, such modified peptides may affect the balance of bacteria inhabiting human gastrointestinal tract and thus bring about health complication of the host. Although pea seeds are of significant nutritional value due to their high contents of proteins, carbohydrates and fibre, they are also responsible for health inconveniences resulting from their susceptibility to digestion and occurrence of antinutritional as well as allergic compounds. The enzymatic degradation may pass over these nutritional obstacles by liberating hydrolysates empowered not only to exert their impact on the human physiology but also on bacterial intestinal ecosystem. Therefore, the aim of this study was to evaluate the influence of pea protein hydrolysates on bacteria typical for the small intestine. Pea protein hydrolysates have proved to diversely modulate physiological activity of bacteria existing in different states. The observed detrimental effect on planktonic bacteria was abolished in the case of bacteria immobilized to the solid surfaces, confirming the protective effect of biofilms. Additionally, Lactobacilli displayed adaptive properties enabling them to utilize pea protein hydrolysates regardless their state of existence.

Determination of Nutritional and Bioactive Properties of Peptides in Enzymatic Pea, Chickpea, and Mung Bean Protein Hydrolysates

Within the primary structure of many pea and mung bean proteins are peptide sequences that can potentially be used in the formulation of therapeutic products for the treatment and prevention of human diseases. However, these peptide sequences need protease treatments before they can be released free of the parent proteins. Unlike chemical hydrolysis, enzymatic treatment enables more efficient tailoring of peptide products without formation of toxic by-products or destruction of amino acids. This review provides information on current methods that have been used to convert inactive pea and mung bean proteins into bioactive peptides. It focuses on 3 main bioactive properties, such as inhibitions of (1) angiotensin converting enzyme (ACE) activity; (2) calmodulin (CaM)-dependent enzymes; and (3) copper-chelating activity. ACE is an established marker for hypertension, high levels of some CaM-dependent enzymes are risk factors for various human diseases including cancer and Alzheimer's disease, and high vascular copper concentrations may potentiate atherosclerosis. Also reviewed are the production and evaluation of activity of hypoallergenic peptides that may offer protection against anaphylactic reactions. The 3 main proteins discussed are chickpea, mung bean, and field pea.

Protein Hydrolysates Induce CCK Release from Enteroendocrine Cells and Act as Partial Agonists of the CCK1Receptor

Protein has been reported to be the most satiating of all macronutrients. Upon gastrointestinal digestion, peptides are generated that stimulate the release of satiety hormones such as cholecystokinin (CCK) from enteroendocrine cells. As such, bioactive peptides could be the target of Functional Food ingredients with satiating effects. We set up an in vitro assay system to investigate if different protein hydrolysates exhibit varying CCK-releasing properties. Soy, pea, potato, casein, and whey protein hydrolysates were incubated with the enteric endocrine cell line STC-1 that endogenously expresses and secretes CCK. Release of CCK was measured by ELISA. All hydrolysates induced CCK release at low concentrations (>0.1 mg.L -1)); however, no significant differences in CCK-releasing properties between the different protein hydrolysates were found, suggesting a generic, nonspecific peptide-sensing mechanism in the STC-1 cells on hydrolyzed protein. As the ELISA exhibits sensitivity to all CCK isoforms possessing the C-terminal CCK octapeptide but varying in biological activity at the CCK 1 receptor (CCK 1R), a secondary module was added to the STC-1 cell assay. Intracellular calcium measurements were performed in CHO-CCK 1R cells. Following exposure of the STC-1 cells to the protein hydrolysates, the medium was tested on the CCK 1R assay. Released CCK was measured with higher sensitivity and lower variability than in the ELISA. Surprisingly, we found that some protein hydrolysates (soy > potato >> casein) also directly stimulated CCK 1R-expressing cells, while whey and pea protein hydrolysates were inactive. As CCK 1R is expressed in the GI tract, direct interaction of CCK 1R with dietary peptides may contribute to their satiety effects. Future experiments developing bioactive ingredients for Functional Foods for weight management could involve isolation of the active, CCK 1R-activating peptides in, for example, soy protein hydrolysates.

Different proteins and biopeptides differently affect satiety and anorexigenic/orexigenic hormones in healthy humans.

We investigated the effects of whey protein (WP), pea protein hydrolysate (PPH), a combination of WP+PPH and control (milk protein (MP) which consists of 80% casein and 20% WP) on appetite ratings, postprandial changes in hunger/satiety hormones and energy intake (EI). A randomized, crossover design which consisted of 2 parts (experiment 1 and 2). The peptides to be tested were provided as part of a shake (1024 kJ; en% P/F/C: 25/33/42) which contained either 15 g WP, 15 g PPH, a combination of 7.5 g WP and 7.5 g PPH (WP+PPH) or 15 g MP. 39 subjects (BMI: 27.6+/-1.7 kg m(-2); age: 42.3+/-13.8 years). In experiment 1 (duration 4 h), appetite profile was measured and blood samples were taken for analysis of hunger/satiety hormones and glucose. In experiment 2 (duration 7 h), appetite profile and EI (180 min after consumption of the shake) were measured. Some indications of lower hunger (experiment 1), desire to eat (experiment 2) and thirst (experiment 1 and 2) were shown after consumption of PPH compared to MP or WP+PPH (P<0.05). A longer intermeal interval and a higher satiety index were suggested after consumption of PPH. Both PPH and WP lead to greater satiety (experiment 2) and fullness (experiment 2) compared to MP and WP+PPH (P<0.05). For WP, a positive correlation between insulin and both cholecystokinine (CCK) and glucagon like peptide-1 (GLP-1) was observed (P<0.05). However, both CCK and GLP-1 were increased by MP (P<0.05), peptide YY (PYY) was stimulated by WP+PPH, while the decline in ghrelin was larger (P<0.05). No effect on EI was seen. There was modest evidence with respect to satiety by PPH consumption. Different exogenous biopeptides produced differences in release of endogenous peptides that had inconsistent relationships with satiety. Therefore, evidence derived from a supposed biomarker for satiety does not guarantee the highest satiety.

The effect of different protein hydrolysate/carbohydrate mixtures on postprandial glucagon and insulin responses in healthy subjects

To study the effect of four protein hydrolysates from vegetable (pea, gluten, rice and soy) and two protein hydrolysates from animal origin (whey and egg) on glucagon and insulin responses. Eight healthy normal-weight male subjects participated in this study. The study employed a repeated-measures design with Latin square randomization and single-blind trials. Protein hydrolysates used in this study (pea, rice, soy, gluten, whey and egg protein hydrolysate) consisted of 0.2 g hydrolysate per kg body weight (bw) and 0.2 g maltodextrin per kg bw and were compared to maltodextrin alone. Postprandial plasma glucose, glucagon, insulin and amino acids were determined over 2 h. All protein hydrolysates induced an enhanced insulin secretion compared to maltodextrin alone and a correspondingly low plasma glucose response. A significant difference was observed in area under the curve (AUC) for plasma glucagon between protein hydrolysates and the maltodextrin control drink (P<0.05). Gluten protein hydrolysate induced the lowest glucagon response. High amino-acid-induced glucagon response does not necessarily go together with low insulin response. Protein hydrolysate source affects AUC for glucagon more profoundly than for insulin, although the protein load used in this study seemed to be at lower level for significant physiological effects.

Physicochemical and bitterness properties of enzymatic pea protein hydrolysates.

The effects of different proteolytic treatments on the physiochemical and bitterness properties of pea protein hydrolysates were investigated. A commercial pea protein isolate was digested using each of 5 different proteases to produce protein hydrolysates with varying properties. After 4 h of enzyme digestion, samples were clarified by centrifugation followed by desalting of the supernatant with a 1000 Da membrane; the retentates were then freeze-dried. Alcalase and Flavourzymetrade mark produced protein hydrolysates with significantly higher (P < 0.05) degree of hydrolysis when compared to the other proteases. Flavourzyme, papain, and alcalase produced hydrolysates that contained the highest levels of aromatic amino acids, while trypsin hydrolysate had the highest levels of lysine and arginine. Papain hydrolysate contained high molecular weight peptides (10 to 178 kDa) while hydrolysates from the other 4 proteases contained predominantly low molecular weight peptides (</= 23 kDa). DPPH (1,1-diphenyl-2-picrylhydrazyl) free radical scavenging activity of the Flavourzyme hydrolysate was significantly (P < 0.05) the highest while alcalase and trypsin hydrolysates were the lowest. Inhibition of angiotensin converting enzyme (ACE) activity was significantly higher (P < 0.05) for papain hydrolysate while Flavourzyme hydrolysate had the least inhibitory activity. Sensory analysis showed that the alcalase hydrolysate was the most bitter while papain and alpha-chymotrypsin hydrolysates were the least. Among the 5 enzymes used in this study, papain and alpha-chymotrypsin appear to be the most desirable for producing high quality pea protein hydrolysates because of the low bitterness scores combined with a high level of angiotensin converting enzyme inhibition and moderate free radical scavenging activity.

The thermogenic and metabolic effects of protein hydrolysate with or without a carbohydrate load in healthy male subjects

High-protein diets are beneficial in weight maintenance because of their satiating and thermogenic effects. These effects may be partly mediated by the hormonal effects of proteins. This study investigated the effect of soy protein hydrolysate (SPH) with and without a carbohydrate pre- and afterload on energy metabolism and hormonal secretion in 8 healthy nonobese subjects. In an additional trial, pea protein hydrolysate was compared to SPH, both with a carbohydrate afterload. The study had a single-blind crossover design. In all cases, 0.4g protein and/or carbohydrate per kilogram of body weight was tested. Diet-induced thermogenesis (DIT) was measured by ventilated hood measurements, and postprandial blood samples were drawn over 3 hours. Soy protein hydrolysate consumption induced a higher DIT than a carbohydrate (CHO) load. Both conditions induced similar insulin responses. Soy protein hydrolysate induced a glucagon, but no glucose, response; whereas CHO induced a glucose, but no glucagon, response. Soy protein hydrolysate with a CHO pre- or afterload induced similar DIT and insulin responses. No glucose response was found when SPH preceded the CHO load. Total glucagon responses were similar with CHO as pre- and afterload, but time courses were different. Pea protein hydrolysate with a CHO afterload induced both higher insulin and glucagon responses (area under the curve) than SPH with CHO afterload, but DIT was similar in both conditions. In conclusion, this study shows that the larger DIT after protein than after CHO may be related to the glucagon response that is induced by protein but not by CHO; that the protein-induced DIT and glucagon response are not influenced by a CHO pre- or afterload; and that protein ingestion can fully prevent the plasma glucose increase associated with CHO when CHOs are ingested after proteins.

Different relationships with respect to satiety between exogenous and endogenous peptides.

The protein source may be a determinant of the satiating efficacy of protein. We investigated the effects of whey protein (WP), pea protein hydrolysate (PPH), a combination of WP+PPH, and control (milk protein, MP) on appetite ratings, postprandial changes in hunger/satiety hormones and energy intake (EI). A total of 39 subjects (BMI: 27.6±1.7 kg/m 2 ; age 42.3±13.8 years) participated in the study which had a randomized, crossover design and consisted of 2 parts (Experiments 1 and 2). In Experiment 1, appetite profile was measured and blood samples were taken. In Experiment 2, appetite profile and EI were measured. Some indications of lower hunger (Experiment 1), desire to eat (Experiment 2) and thirst (Experiments 1 and 2) were shown after consumption of PPH compared to MP or WP+PPH ( p <0.05). A longer intermeal interval and a higher satiety index were suggested after consumption of PPH. Both PPH and WP lead to greater satiety (Experiment 2) and fullness (Experiment 2) compared to MP and WP+PPH ( p <0.05). For WP, a positive correlation between insulin and both CCK and GLP-1 was observed ( p <0.05). However, both CCK and GLP-1 were increased by MP ( p <0.05) and PYY and ghrelin were stimulated by WP+PPH ( p <0.05). No effect on EI was seen. In conclusion, there was modest evidence with respect to satiety by PPH consumption. Different exogenous biopeptides produced differences in release of endogenous peptides that had inconsistent relationships with satiety. Therefore, evidence derived from a supposed biomarker for satiety does not guarantee the highest satiety.

Intragastric infusion of pea protein hydrolysate reduces food intake more than pea protein.

Because protein hydrolysates are digested faster than the corresponding proteins, they may cause a more rapid release of satiating gut peptides and early satiation. We tested this hypothesis in adult male rats, surgically equipped with intragastric (IG) cannulas and adapted to a 30-min test meal after 15-h food deprivation. When IG infusions of equiosmotic 12 ml of saline (S), 13.6% pea protein (PP), or 13.6% pea protein hydrolysate (PPH; provided by DSM/DFS, Delft, The Netherlands) were administered at lights off, both PP and PPH reduced 30-min food intake, but PPH was 26% more effective (p

Structural modulation of calmodulin and calmodulin-dependent protein kinase II by pea protein hydrolysates

The effects of two fractions of pea protein hydrolysate with high levels of positively charged amino acids on the structural conformations of calmodulin (CaM) and CaM-dependent protein kinase II (CaMKII) were determined using fluorescence and circular dichroism methods. In the presence of Ca2 + , addition of the protein hydrolysates to CaM and CaM/CaMKII complex led to increased exposure of aromatic groups as measured by intrinsic and extrinsic fluorescence spectroscopy. Near-UV circular dichroism data revealed an increase in the tertiary structure of CaM in the presence of pea protein hydrolysates. Effect of the protein hydrolysates on the CaM structure was greater with the fraction that contained higher contents of arginine and lysine when compared with the fraction with lower levels of these two amino acids. We concluded that the presence of the pea protein hydrolysates led to rearrangement of the native protein structure and exposure of buried hydrophobic groups of CaM and/or CaMKII.

Requirements for non-food applications of pea proteins A Review

So far, limited research is performed on technical applications of pea proteins and no applications have yet been reported. At ATO, three technical applications were investigated: surfactants, films, and microspheres as encapsulation matrices. Pea protein hydrolysates are surfactants with good emulsifying and foaming properties. By variation of enzyme type and degree of hydrolysis, the surfactant properties can be tailored toward specific applications. Pea protein films could be prepared by casting from dispersions at pH 7 and 10, and by compression moulding at 140 degrees C. Opposite to many other proteins, pea protein films combine strength (5-7.5 MPa) with high elongation at break (150%). A protein isolate derived from peas was applied as matrix material for the microencapsulation of beta-carotene, intended for cosmetic applications. Supercritical CO2 technology appeared to be a promising encapsulation technique for beta-carotene in porous pea protein microspheres. Advantages of this method are that no organic solvents are used, and that encapsulation is achieved under mild conditions, thereby preventing the sensitive beta-carotene from degradation.

Requirements for non-food applications of pea proteins. A review.

So far, limited research is performed on technical applications of pea proteins and no applications have yet been reported. At ATO, three technical applications were investigated: surfactants, films, and microspheres as encapsulation matrices. Pea protein hydrolysates are surfactants with good emulsifying and foaming properties. By variation of enzyme type and degree of hydrolysis, the surfactant properties can be tailored toward specific applications. Pea protein films could be prepared by casting from dispersions at pH 7 and 10, and by compression moulding at 140 degrees C. Opposite to many other proteins, pea protein films combine strength (5-7.5 MPa) with high elongation at break (150%). A protein isolate derived from peas was applied as matrix material for the microencapsulation of beta-carotene, intended for cosmetic applications. Supercritical CO2 technology appeared to be a promising encapsulation technique for beta-carotene in porous pea protein microspheres. Advantages of this method are that no organic solvents are used, and that encapsulation is achieved under mild conditions, thereby preventing the sensitive beta-carotene from degradation.

Comparison of solubility of pea protein hydrolysate by three analytical methods

Pea protein hydrolysate was obtained by enzymatic hydrolysis with trypsin. The degree of hydrolysis (DH) was controlled by using the pH-stat method. Solubility of the trypsin-treated hydrolysate was tested at nine different pH values starting from 2 up to 10. Protein determinations were carried out using Kjeldahl, Lowry and modified Lowry methods. The results revealed that samples analysed with either the Kjeldahl or Lowry method gave similar values. However, systematic consistent differences existed for those results obtained by the Kjeldahl and the modified Lowry as well as between those results obtained by the Lowry and the modified Lowry.

The solubility of trypsin pea protein hydrolysates

The solubility of trypsin pea protein hydrolysates

Determination of α-amino nitrogen in pea protein hydrolysates: a comparison of three analytical methods

Three spectrophotometric methods, using 2,4,6-trinitrobenzenesulphonic acid (TNBS), o-phthaldialdehyde (OPA) or ninhydrin, for the determination of α-amino nitrogen in pea protein isolates and hydrolysates were compared. The determined amounts of α-amino nitrogen differed greatly, depending on the method used. The TNBS and OPA methods produced comparable results, whereas the data obtained with the ninhydrin method were only half of the TNBS or OPA values. Colour stability, recovery of the standard from the protein matrix and reproducibility of the results were determined. The methods showed good accuracy (SE 1–3%) with recovery values of the standard ( l-leucine) from the protein matrix of 91, 111 and 75% for the OPA, ninhydrin and TNBS method, respectively.

Production and separation of the methionine rich fraction from chick pea protein hydrolysate generated by proteases of Bacillus amyloliquefaciens

Crude proteases produced by Bacillus amyloliquefaciens in solid substrate fermentation (SSF) were utilised for the production of a protein hydrolysate using chick pea as the protein source. These enzymes were found to be better hydrolytic agents than fungal enzymes. The hydrolysate generated was separated on an immobilised metal ion affinity chromatography (Cu 2+) column using a volatile elution buffer. Two fractions were rich in the amino acid methionine and could be used as a peptide fortifier in different food products.