Antihypertensive and Antioxidant Activity of Chia Protein Techno-Functional Extensive Hydrolysates

Twelve high-quality chia protein hydrolysates (CPHs) were produced from chia protein isolate (CPI) in a pilot plant of vegetable proteins. To obtain functional hydrolysate, four CPHs were hydrolyzed by the action of Alcalase, an endoprotease, and the other eight CPHs were hydrolyzed by the action of Flavourzyme, an exoprotease. Alcalase-obtained CPHs showed significant antihypertensive properties particularly, the CPH obtained after 15 min of hydrolysis with Alcalase (CPH15A), which showed a 36.2% hydrolysis degree. In addition, CPH15A increased the antioxidant capacity compared to CPI. The CPH15A physicochemical composition was characterized and compared to chia defatted flour (CDF) and CPI, and its techno-functional properties were determined by in vitro experiments through the analysis of its oil absorption capacity, as well as the capacity and stability of foaming and emulsifying, resulting in an emulsifier and stabilizer better than the intact protein. Therefore, the present study revealed that CPH15A has potent antihypertensive and antioxidant properties and can constitute an effective alternative to other plant protein ingredients sources that are being used in the food industry.Spanish Ministry of Science, Innovation, and Universities grant CYTED-2019/119RT056

Obtención de hidrolizados proteicos del gluten de trigo con actividad inmunomoduladora

Motivación: Tener una dieta rica en proteínas puede llegar a ser muy importante ya que éstas intervienen en diversos procesos metabólicos y funciones biológicas de gran importancia. Además de aportarnos los aminoácidos esenciales de diversas rutas moduladoras sistémicas, las proteínas y los péptidos procedentes de su hidrólisis pueden ejercer diversas actividades biológicas, habiéndose descrito péptidos con actividad anticancerígena, antimicrobiana, hipocolesterolémica, hipotensora, antioxidante, antiinflamatoria, etc.(2).El objetivo de este proyecto fue la obtención de hidrolizados proteicos con diferentes grados de hidrólisis a partir del gluten de trigo para la caracterización de péptidos bioactivos de trigo con capacidad inmunomoduladora.Métodos:1) Obtención de aislados e hidrolizados proteicos de trigo. Los aislados proteicos de trigo se obtuvieron a partir del gluten de la harina de trigo comercial mediante solubilización alcalina de las proteínas y posterior precipitación al punto isoeléctrico de las mismas (4). Se calcularon las riquezas proteicas así como el rendimiento de extracción proteica de cada una de las etapas del proceso. A continuación, las enzimas comerciales Izyme AL y Alcalasa se adicionaron al aislado proteico para la obtención de hidrolizados proteicos de trigo (3) y se tomaron alícuotas a diferentes tiempos para determinar el grado de hidrólisis usando la titulación con TNBS en función del tiempo (1).2) Determinación de la actividad inmunomoduladora. Los hidrolizados obtenidos a diferentes tiempos se utilizaron para estudios de inhibición de enzimas que participan en la respuesta inflamatoria humana (fosfolipasa A2, ciclooxigenasa 2, trombina y transglutaminasa) (3).Resultados: En base al pH, primeramente se solubilizaron las proteínas del gluten de la harina de trigo a pH 12 y posteriormente se precipitaron a pH 6. Las riquezas proteicas tanto de la harina (82.82%) como de los aislados (90.35%) fueron altas y muy similares, y el porcentaje de pérdidas durante el proceso de extracción proteica fue del 27.7%. Por todo ello, se decidió realizar la hidrólisis directamente a partir de la harina de trigo en lugar de utilizar los aislados proteicos con la idea de ahorrar costes en el en el proceso global. El valor del grado de hidrólisis obtenido para estos hidrolizados fue en torno al 25%, tras 60 minutos de incubación

Anti-inflammatory activity of lupine (Lupinus angustifolius L.) protein hydrolysates in THP-1-derived macrophages

The effect of two different lupine protein hydrolysates (LPHs) on in vitro macrophage activation in a THP-1-derived macrophage model was investigated. THP-1-derived macrophages were exposed to RPMI medium containing two LPHs obtained by enzymatic hydrolysis using two different proteases: Izyme AL and Alcalase 2.4 L. Cytokine's expression was measured by quantitative PCR. THP-1-derived macrophages exhibited attenuated expression of proinflammatory cytokines (tumor necrosis factor (TNF), IL-6, IL-1β) and increased expression of anti-inflammatory marker genes (chemokine (C-C motif) ligand 18 (CCL18)) relative to control without LPH. The anti-inflammatory effect of both hydrolysates favored M2 polarization by quenching C-C chemokine receptor type 2 (CCR2) expression and migratory capacity. Furthermore, LPHs significantly decreased nitric oxide production. Moreover, LPHs promoted the survival of human THP-1-derived macrophages. Therefore, inclusion of LPHs in foods may help to prevent chronic diseases associated with chronic inflammation.Ministerio de Economía y Competitividad AGL2012–40247-C02-0

Purificación y caracterización parcial de proteínas de almacenamiento de semillas de Lupinus angustifolius

Lupinus angustifolius seed proteins have been purified by sequential dialysis and ion exchange chromatography, and their amino acid composition has been studied in order to determine their nutritional value as sources of essential amino acids. Albumins include a great variety of proteins. Globulins were resolved in α, β and δ conglutins. Conglutin α is the main protein in the seeds of L. angustifolius, representing 76.6% of the total. While lysine was found to be the limiting amino acid in L. Angustifolius seed proteins as a whole, tyrosine was the limiting amino acid in albumins, and methione and lysine were limiting in globulins. Lysine, methionine and histidine were limiting amino acids in α conglutin.Se ha realizado una purificación por diálisis secuencial y cromatografía de intercambio iónico de las proteínas de la semilla de L. angustifolius, y su composición en aminoácidos ha sido estudiada para determinar su valor nutricional como fuentes de aminoácidos esenciales. Las albúminas están compuestas por una compleja mezcla de proteínas. Las globulinas se resolvieron en conglutinas α, β y δ. La conglutina α es la proteína más abundante en las semillas de L. angustifolius representando el 76.6% del total. Las proteínas de la semilla de L. angustifolius son limitantes en lisina. Las albúminas son limitantes en tirosina y las globulinas en metionina y lisina. La α conglutina es limitante en lisina, metionina e histidina

A lupine (Lupinus angustifolious L.) peptide prevents non-alcoholic fatty liver disease in high-fat-diet-induced obese mice

Bioactive peptides are related to the prevention and treatment of many diseases. GPETAFLR is an octapeptide that has been isolated from lupine (Lupinus angustifolius L.) and shows anti-inflammatory properties. The aim of this study was to evaluate the potential activity of GPETAFLR to prevent non-alcoholic fatty liver disease (NAFLD) in high-fat-diet (HFD)-induced obese mice. C57BL/6J mice were fed a standard diet or HFD. Two of the groups fed the HFD diet were treated with GPETAFLR in drinking water at 0.5 mg kg-1 day-1 or 1 mg kg-1 day-1. To determine the ability of GPETAFLR to improve the onset and progression of non-alcoholic fatty liver disease, histological studies, hepatic enzyme profiles, inflammatory cytokine and lipid metabolism-related genes and proteins were analysed. Our results suggested that HFD-induced inflammatory metabolic disorders were alleviated by treatment with GPETAFLR. In conclusion, dietary lupine consumption can repair HFD-induced hepatic damage possibly via modifications of liver's lipid signalling pathways

Wheat gluten protein hydrolysates anti-inflammatory study

Motivation: It is known that some peptides have bioactive properties, anticarcinogenic, anti-inflammatory or antimicrobial among others. Lately, researches made in this area have been focused on vegetable sources peptides, which have shown a great capacity of having anti-inflammatory effects. Therefore, the aim of this investigation was to check wheat gluten hydrolysates anti-inflammatory capacities.Methods: 1) Wheat gluten was used as starting material, which was hydrolysated with two different enzymes each on their own, Alcalse 2.4L and Izyme AL during 1 and 2 hours respectively, taking samples each 15 minutes (named A0, A15, A30, A45, A60; I0, I15, I30, I45, I60, I75, I90, I105, I120). The hydrolysis grade was obtained using 2,4,6-trinitrobenzene sulfonic acid (TNBS) method. 2) In order to determine their immunommodulatory capacity, the inhibitory effect of each sample was tested among chronic inflammatory diseases pathways common enzymes: thrombin and angiotensin-converting enzyme (ECA). 3) Blood cells, monocytes, submitted to lipopolysaccharide (LPS) infection, were incubated for 24 hours with A45 and I90 at different concentrations (10µg/mL and 5µg/mL) for the purpose of seeing IL-1β, TNF-α (pro-inflammatory proteins) and IL-10 (anti-inflammatory protein), cytokines genes expression change; studied by Q-PCR.Results & Conclusions: 1) The more was the time that were being hydrolysed, the bigger was the hydrolysis grade, reaching a maximum of 36% of it with Alcalase 2.4.L and a 11,869% with Izyme. 2) The hydrolysates named as A45 and I90 were the ones that shown the most effective inhibitory effect above inflammatory enzymes; their physicochemical features should be studied for understanding it. 3)The hydrolysates chosen in the enzymatic tests (A45 and I90) showed to have a anti-inflammatory activity potential on cells, since in most cases they tend to repress pro-inflammatory genes expression (TNF-α and IL-1β) and enhanced the expression of other anti-inflammatory cytokines such as IL-10 (in all cases). Therefore, this study reveals that the inclusion of wheat gluten protein hydrolysates on our diets could help in the treatment of inflammatory diseases

High Stabilization of Enzymes Immobilized on Rigid Hydrophobic Glyoxyl-Supports: Generation of Hydrophilic Environments on Support Surfaces

© 2020 by the authors.Very rigid supports are useful for enzyme immobilization to design continuous flow reactors and/or to work in non-conventional media. Among them, epoxy-methacrylic supports are easily functionalized with glyoxyl groups, which makes them ideal candidates for enzyme stabilization via multipoint covalent immobilization. However, these supports present highly hydrophobic surfaces, which might promote very undesirable effects on enzyme activity and/or stability. The hydrophilization of the support surface after multipoint enzyme immobilization is proposed here as an alternative to reduce these undesirable effects. The remaining aldehyde groups on the support are modified with aminated hydrophilic small molecules (glycine, lysine or aspartic acid) in the presence of 2-picoline borane. The penicillin G acylase from Escherichia coli (PGA) and alcohol dehydrogenase from Thermus thermophilus HB27 (ADH2) were immobilized on glyoxyl-functionalized agarose, Relizyme and Relisorb. Despite the similar density of aldehyde groups displayed by functionalized supports, their stabilization effects on immobilized enzymes were quite different: up to 300-fold lower by hydrophobic supports than by highly hydrophilic glyoxyl-agarose. A dramatic increase in the protein stabilities was shown when a hydrophilization treatment of the hydrophobic support surface was done. The PGA immobilized on the glyoxyl-Relisorb hydrophilized with aspartic acid becomes 280-fold more stable than without any treatment, and it is even more stable than the PGA immobilized on the glyoxyl agarose.This research was funded by the Spanish Ministry of Economy, Industry and Competitiveness (projects BIO2012-36861 and CTQ2015-70348) and the EU FP7 project SuSy (Sucrose Synthase as Cost-Effective Mediator of Glycosylation Reactions, C-KBBE/3293). Javier Rocha-Martin is grateful for the Juan de la Cierva fellowship (IJCI-2014-19260) funded by the Spanish Ministry of Economy, Industry and Competitiveness.Peer reviewe

Antioxidant and anti-inflammatory properties of bioavailable protein hydrolysates from lupin-derived agri-waste

Agri-food industries generate several by-products, including protein-rich materials cur-rently treated as waste. Lupine species could be a sustainable alternative source of protein compared to other crops such as soybean or chickpea. Protein hydrolysates contain bioactive peptides that may act positively in disease prevention or treatment. Inflammatory responses and oxidative stress underlie many chronic pathologies and natural treatment approaches have gained attention as an alternative to synthetic pharmaceuticals. Recent studies have shown that lupin protein hydrolysates (LPHs) could be an important source of biopeptides, especially since they demonstrate anti-inflam-matory properties. However, due to their possible degradation by digestive and brush-border en-zymes, it is not clear whether these peptides can resist intestinal absorption and reach the blood-stream, where they may exert their biological effects. In this work, the in vitro cellular up-take/transport and the anti-inflammatory and antioxidant properties of LPH were investigated in a co-culture system with intestinal epithelial Caco-2 cells and THP-1-derived macrophages. The results indicate that the LPH crosses the human intestinal Caco-2 monolayer and exerts anti-inflam-matory activity in macrophages located in the basement area by decreasing mRNA levels and the production of pro-inflammatory cytokines. A remarkable reduction in nitric oxide and ROS in the cell-based system by peptides from LPH was also demonstrated. Our preliminary results point to underexplored protein hydrolysates from food production industries as a novel, natural source of high-value-added biopeptides.Ministerio de Ciencia, Innovación y Universidades CYTED-2019/119RT056

Nutraceutical Extract from Dulse (Palmaria palmata L.) Inhibits Primary Human Neutrophil Activation

Palmaria palmata L. (Palmariaceae), commonly known as "dulse", is a red alga that grows on the northern coasts of the Atlantic and Pacific oceans, and is widely used as source of fiber and protein. Dulse is reported to contain anti-inflammatory and antioxidant compounds, albeit no study has investigated these effects in primary human neutrophils. Implication strategies to diminish neutrophil activation have the potential to prevent pathological states. We evaluated the ability of a phenolic dulse extract (DULEXT) to modulate the lipopolysaccharide (LPS)-mediated activation of primary human neutrophils. Intracellular reactive oxygen species (ROS) were measured by fluorescence analysis and nitric oxide (NO) production using the Griess reaction. Inflammatory enzymes and cytokines were detected by ELISA and RT-qPCR. The results show that DULEXT diminished the neutrophil activation related to the down-regulation of TLR4 mRNA expression, deceased gene expression and the LPS-induced release of the chemoattractant mediator IL-8 and the pro-inflammatory cytokines IL-1β, IL-6 and TNF-α. ROS, NO, and myeloperoxidase (MPO) were also depressed. The data indicated that DULEXT has the potential to disrupt the activation of human primary neutrophils and the derived inflammatory and prooxidant conditions, and suggest a new role for Palmaria palmata L. in the regulation of the pathogenesis of health disorders in which neutrophils play a key role, including atherosclerosisSpanish Ministry of Science, Innovation and Universities grant CYTED-2019 119RT056