Characterization and Bile Acid Binding Capacity of Dietary Fiber Obtained from Three Different Amaranth Products.

Amaranth is a dicotyledonous plant, now considered a health-promoting food. It has been rediscovered by the worldwide food industry, which is increasingly becoming aware of the many uses and benefits provided by amaranth in various food preparations. Amaranth dietary fibers, soluble and insoluble fractions, obtained from flour, protein isolate, and beverage were physicochemically characterized and their potential bile acid binding capacity was evaluated. Primary bile acids binding to fiber might contribute to a hypocholesterolemic effect, while the binding of secondary bile acids could minimize the cytotoxic effect that these metabolites exert on the colon. Amaranth fiber fractions were capable of sequestering cholate, taurocholate, deoxycholate, and bovine bile, with a percentage depending not only on the origin and the type of amaranth fiber evaluated but also on the bile acid studied. Flour fiber and the protein isolate insoluble fractions were the most efficient for binding bile and bile acids with uptake values between 29 and 100% relative to cholestyramine. Moreover, deoxycholate, a hydrophobic secondary bile acid, was the most captured by all the fractions, reaching 100% uptake with total and insoluble fibers of the three amaranth products. These results would suggest that the main effect through which amaranth fiber binds bile acids corresponds to an adsorptive effect mediated by hydrophobic interactions.

Amaranth as a Source of Antihypertensive Peptides.

Amaranth is an ancestral crop used by pre-Columbian cultures for 6000 to 8000 years. Its grains have a relevant chemical composition not only from a nutritional point of view but also due to the contribution of components with good techno-functional properties and important potential as bioactive compounds. Numerous studies have shown that amaranth storage proteins possess encrypted sequences that, once released, exhibit different physiological activities. One of the most studied is antihypertensive activity. This review summarizes the progress made over the last years (2008-2020) related to this topic. Studies related to inhibition of different enzymes of the Renin-Angiotensin-Aldosterone system, in particular Angiotensin Converting Enzyme (ACE) and Renin, as well as those referring to potential modulation mechanisms of tissue or local Renin-Angiotensin-Aldosterone system, are analyzed, including in silico, in vitro, in vivo, and ex vivo assays. Furthermore, the potential use of these bioactive peptides or products containing them, in the elaboration of functional food matrices is discussed. Finally, the most relevant conclusions and future requirements in research and development of food products are presented.

Development of a High Protein Beverage Based on Amaranth.

The objective of this study was to formulate a beverage based on amaranth proteins, stable and nutritious. The process of obtaining the beverage was based on the existing knowledge about starch separation techniques and techno-functional properties of the amaranth proteins. Gums, gellan and xanthan were added to the protein extract and it was heat-treated at 80 °C during 20 min. A beverage with a composition similar to skim cow's milk was obtained (3.42 ± 0.08; 0.60 ± 0.06; 1.9 ± 0.4; 0.43 ± 0.01; 3 and 90.58 ± 0.01% for proteins, lipids, fiber, ashes, carbohydrates and water, respectively). Thermal treatment caused the denaturation and aggregation of the proteins, while the addition of gums induced a decrease in the sensitivity to heat treatment of the proteins. Formation of protein aggregates and gum-protein complexes was characterized by electrophoresis, differential scanning calorimetry, and particle size distribution. Heat treatment and addition of gums generated macrocomplexes with enhanced absolute value of ζ-potential, which contributed to the high colloidal stability of amaranth-based beverage. This beverage is suitable for vegans, celiac patients, and lactose intolerants.

Data set on effect of amaranth proteins on the RAS system. In vitro, in vivo and ex vivo assays.

Data set presented in this article is related to the research paper entitled "Effect of amaranth proteins on the RAS system. In vitro, in vivo and ex vivo assays", available in Food Chemistry [1]. In this article, we evaluated the effect on systolic blood pressure of spontaneously hypertensive rats (SHR) of different samples with amaranth proteins/peptides. The effect of these samples on RAS system was evaluated using in vitro and ex vivo assays. The concentration of renin and angiotensin converting enzyme (ACE) was evaluated using two commercial ELISA kits. Renin concentration was estimated through a direct immunoassay and ACE concentration with an immunoassay based on a competitive inhibition. In addition, the ACE inhibitory activity in plasma was evaluated using a spectrophotometric assay according to [2]. Ex vivo experiments were done with thoracic aorta extracted during the surgical procedure employed to obtain blood samples according to [3]. Data presented in this article recollect a very extensive work on how can be affect the RAS system in SHR model using amaranth protein/peptides as potential antihypertensive samples. These data could be useful to design novel functional foods for hypertensive individuals.

Effect of amaranth proteins on the RAS system. In vitro, in vivo and ex vivo assays.

The aim of this work was to analyse the hypotensive effect of amaranth protein/peptides on spontaneously hypertensive rats (SHR). The mechanism of action of these peptides was studied in vivo and ex vivo. We also tested the effect of protection against gastrointestinal digestion (GID) exerted by an O:W emulsion on the integrity of the antihypertensive peptides. All samples tested produced a decrease in blood pressure (SBP). The animals treated with emulsion (GE) and emulsion + peptide (GE+VIKP) showed the most significant reduction in the SBP (42 ±â€¯2 mmHg and 35 ±â€¯2 mmHg, respectively). The results presented suggest that after GID, a variety of peptides with biological activities were released or were resistant to this process. These peptides play a role in the regulation of the SBP by acting on plasma ACE, plasma renin and the vascular system. These results support the use of amaranth protein/peptides in the elaboration of functional foods for hypertensive individuals.

Effect of the Incorporation of Amaranth (Amaranthus Mantegazzianus) into Fat- and Cholesterol-Rich Diets for Wistar Rats.

The hypocholesterolemic effect of amaranth was studied in male Wistar rats fed a high-fat diet that was supplemented with amaranth flour, AF, or isolated protein, AI. Likewise, an in vitro test was carried out, in which the capacity of the AI, AF, the digested isolate, DAI, and the digested amaranth flour, DAF, to displace the cholesterol of the model micelles was evaluated. The in vivo results showed an increase in the excretion of cholesterol through feces (77% for AF7; 23% and 108% for AI30 and AF30, respect control) and a decrease in the content of hepatic cholesterol (98% for AF7; 96% and 53% for AI30 and AF30 respect control); whereas in vitro it was shown that both AF and DAF have greater power to displace cholesterol than the AI and DAI (IC50 0.1, 0.71, 0.2, and 2.1 for AF, DAF, AI, and DAI, respectively). These evidences show that the proteins and fibers of amaranth have an effect on cholesterol metabolism. PRACTICAL APPLICATION: Nowadays, consumers give great importance to the effect that food has on health. The results shown in this work evidence the potential hypocholesterolemic activity presented by amaranth, this is of great importance due to the increase in the incidence of dyslipidemia in the world population and the importance of amaranth as a nonextensive crop of excellent agronomic, nutritional, and bioactive properties suitable for preparation of functional foods.

Antiproliferative Effect of Amaranth Proteins and Peptides on HT-29 Human Colon Tumor Cell Line.

Antiproliferative effect of Amaranthus mantegazzianus proteins and peptides released after simulated gastrointestinal digestion (DH% 37.8 ± 3.8) was investigated on human colon cancer cell line HT-29. Inhibition of proliferation of HT-29 cells was exhibited after a 24 h treatment with different concentrations of amaranth protein isolate (API) and the peptides released after digestion (DGS), presenting IC50 values of 1.35 ± 0.12 and 0.30 ± 0.07 mg soluble protein/mL, respectively. Lactate dehydrogenase assay indicated that both samples caused the loss of membrane integrity and cell lysis over HT-29 cells, and DAPI fluorescence microscopies evidenced typical apoptotic features. Moreover, Annexin V-FITC flow cytometry showed a significant increase of early apoptotic and late apoptotic/necrotic HT-29 cells compared to untreated ones, and caspase-3 assay confirmed the apoptosis induction with a 43.0 ± 10.3 and 65.8 ± 12.7% increase of caspase-3 activity produced by a 2 mg/mL treatment of API and DGS, respectively. In conclusion, amaranth peptides successfully released after simulated gastrointestinal digestion would exert a potential antiproliferative activity over HT-29 tumor cells. This effect was linked to the induction of cell necrosis and apoptosis, supporting the idea of using amaranth proteins as a potential food alternative ingredient for functional foods.

Amaranth peptides decreased the activity and expression of cellular tissue factor on LPS activated THP-1 human monocytes.

The effect of amaranth peptides on the activity and expression of tissue factor (TF) on THP-1 activated cells was evaluated in vitro. An active anticoagulant peptide fraction (AF) was found to inhibit TF expression (IC50 = 0.39 mg mL-1) and activity. Immunocytochemical fluorescence confocal microscopy analysis showed that treated monocytes decreased TF membrane translocation by 49.0% and increased two-fold in nuclei compared to a positive control, indicating a decrease of active TF to initiate the coagulation cascade. Moreover, a cytokine array suggested that the AF mechanism of action implied the inhibition of the NF-κB pathway. Expression of MIP-3α, interleukin-1ß, interleukin-1α, TARC, pentaxin 3, and PDGF-AA cytokines was highly suppressed by AF peptides, producing reductions of 78.8%, 61.8%, 54.1%, 42.6%, 37.9% and 37.8%, respectively, compared to a positive control. The results suggest a potential mechanism for the antithrombotic and anti-inflammatory effect of AF, by showing that amaranth peptides play a negative feedback regulatory role over the NF-κB pathway. In this research, we link for the first time the immunomodulatory activity of amaranth peptides with the inhibition of TF expression and therefore their antithrombotic potential.

Antithrombotic and Antioxidant Activity of Amaranth Hydrolysate Obtained by Activation of an Endogenous Protease.

Ingestion of diets with antithrombotic and antioxidant components offer a convenient and effective way to prevent and reduce the incidence of cardiovascular diseases. The aim of the present work was to obtain an amaranth hydrolysate by the activation of an endogenous aspartic protease, to establish adequate experimental conditions, and to evaluate its antithrombotic and antioxidant activity in order to assess its potential application as an ingredient in functional foods. The results obtained not only confirmed the presence of an endogenous protease in the amaranth isolate, but also allowed us to select an adequate incubation conditions (pH 2, 40 °C, 16 h). The hydrolysate obtained (degree of hydrolysis 5.3 ± 0.4 %) showed potential antithrombotic activity (IC50 = 5.9 ± 0.1 mg soluble protein/mL) and had more antioxidant activity than the isolate, indicating that the activation of the protease released bioactive peptides from amaranth proteins. Decreasing the pH is a simple and cheap process and is another way to obtain potential functional ingredients with bioactive compounds.

Antithrombotic Effects of Amaranthus hypochondriacus Proteins in Rats.

Cardiovascular disease (CVD) is a major cause of disability and premature death throughout the world. Diets with antithrombotic components offer a convenient and effective way of preventing and reducing CVD incidence. The aim of the present work was to assess in vivo and ex vivo effects of Amaranthus hypochondriacus proteins on platelet plug formation and coagulation cascade. Amaranth proteins were orally administrated to rats (AG, 8 animals) and bleeding time was determined showing no significant difference compared with control rats (CG, 7 animals). However, results show a strong tendency, suggesting that amaranth proteins are involved in the inhibition of thrombus formation. Non-anticoagulated blood extracted from animals was analyzed with the hemostatometer, where AG parameters obtained were twice the values showed by CG. The clotting tests, thrombin time (TT) and activated partial thromboplastin time (APTT), presented a 17 and 14% clotting formation increase respectively when comparing AG with CG. The ex-vivo assays confirm the hypothesis inferring that amaranth proteins are a potential antithrombotic agent.

Effect of amaranth flour (Amaranthus mantegazzianus) on the technological and sensory quality of bread wheat pasta.

The technological and sensory quality of pasta made from bread wheat flour substituted with wholemeal amaranth flour (Amaranthus mantegazzianus) at four levels, 15, 30, 40 and 50% w/w was investigated. The quality of the resulted pasta was compared to that of control pasta made from bread wheat flour. The flours were analyzed for chemical composition and pasting properties. Cooking behavior, color, raw and cooked pasta texture, scanning electron microscopy and sensory evaluation were determined on samples. The pasta obtained from amaranth flour showed some detriment of the technological and sensory quality. So, a maximum substitution level of 30% w/w was defined. This is an equilibrium point between an acceptable pasta quality and the improved nutritional and functional properties from the incorporation of amaranth flour.

Potential antitumor properties of a protein isolate obtained from the seeds of Amaranthus mantegazzianus.

BACKGROUND: Amaranth is a crop that can be grown in different soils and climates, being resistant to high temperatures, drought, and some pests. The amaranth plant has nutritional qualities and desirable biological properties. AIM OF THE STUDY: The aim of the study is to investigate the potential antitumor properties of Amaranthus-mantegazzianus-protein isolate (MPI) and to elucidate the possible mechanism of action. METHODS: We use four different tumor-derived and in vitro-transformed cell lines with different morphology and tumorigenicity (MC3T3E1, UMR106, Caco-2, and TC7). RESULTS: The MPI showed an antiproliferative effect on four cell lines with different potencies. The tumor-cell line UMR106 was the most sensitive (IC(50): 1 mg/ml). This antiproliferative effect of the MPI was enhanced by protease treatment (IC(50): 0.5 after 30% hydrolysis). In addition, the MPI produced morphological changes and caused a rearrangement of the cytoskeleton in UMR106 cell line. In an attempt to elucidate the mechanism of action, we observed that the MPI inhibited cell adhesion and induced apoptosis and necrosis in the UMR106 cell line. In reversibility studies, we were able to observe both temporary and permanent cytostatic and cytotoxic effects on the part of the MPI, depending on its concentration. CONCLUSIONS: we report a protein isolate from the seeds of Amaranthus mantegazzianus that exhibit potential antitumor properties and propose a putative mechanism of action.

Mature Amaranthus hypochondriacus seeds contain non-processed 11S precursors.

Amaranth is a dicotyledonous plant whose major seed storage proteins are globulins and glutelins. An unique feature of amaranth seeds is the presence of a fraction named albumin-2, that is extractable with water only after an exhaustive extraction of globulins and albumin-1. In this work, we tested the hypothesis that albumin-2 fraction could be constituted by a non-processed 11S globulin (proglobulin). To this end, the gene encoding the amaranth 11S subunit was cloned and expressed in Escherichia coli. Subsequently, the recombinant proglobulin and albumin-2 purified from seeds were treated with a sunflower vacuolar processing enzyme (VPE). A 55 kDa component of albumin-2 was specifically cleaved into 38 and 17-15 kDa polypeptides, as a consequence of this endoproteolytic cleavage a change of the oligomeric state from trimeric to hexameric was observed. Amaranth 11S globulin fraction was not modified under these proteolysis conditions. Using VPE-specific antibodies, it was shown that amaranth expresses a 57 kDa VPE, and that both developing and mature amaranth seeds have VPE activity, although the increase of this activity during amaranth seed development is higher than that observed for sunflower seeds. These results confirm the presence of unprocessed 11S precursors in mature amaranth seeds; this phenomenon cannot, however, be attributed to low VPE activity during developing of amaranth seeds.

Effect of pH and ionic strength modifications on thermal denaturation of the 11S globulin of sunflower (Helianthus annuus).

Helianthinin, the main storage protein of sunflowers, has low water solubility and does not form a gel when heated; this behavior is different from other 11S globulins and limits its food applications. To understand this particular behavior, changes on helianthinin association-dissociation state induced by modifications in pH and ionic strength were analyzed. The influence of these different medium conditions on its thermal stability and tendency to form aggregates was also studied. Helianthinin behavior at different pH values and ionic strengths is similar to other 11S globulins except that it remains in a trimeric form at pH 11. Helianthinin thermal stability is higher than other 11S globulins but is lower than oat 11S globulin. Alkaline pH produces a 10 degrees C decrease of its denaturation temperature and also of the cooperativity of denaturation process, but it does not affect the denaturation activation energy. The decrease in thermal stability with the pH increase is also manifested by its tendency to form aggregates by SH/SS interchange reactions. When thermal treatments at alkaline pH are performed, all helianthinin subunits form aggregates, characterized by a higher proportion of beta-polypeptides than alpha-polypeptides, which is an indication that aggregation is accompanied by dissociation. Treatments at 80 degrees C are sufficient to induce aggregation but not to produce denaturation, and in these conditions hexameric forms remain after the treatment.

Dynamic properties of soy globulin adsorbed films at the air-water interface.

In this paper we present surface dilatational properties of soy globulins (beta-conglycinin, glycinin, and reduced glycinin with 10 mM of dithiothreitol (DTT)) adsorbed onto the air-water interface, as a function of adsorption time. The experiments were performed at constant temperature (20 degrees C), pH (8.0), and ionic strength (0.05 M). The surface rheological parameters were measured as a function of protein concentration (ranging from 1 to 1x10(-3)% wt/wt). We found that the surface dilatational modulus, E, increases, and the phase angle, phi, decreases with time, theta, which may be associated with protein adsorption. These phenomena have been related to protein adsorption, unfolding, and/or protein-protein interactions (at long-term adsorption) as a function of protein concentration in solution. From a rheological point of view, the surface viscoelastic characteristics of soy globulin films adsorbed at the air-water interface are practically elastic. The main conclusion is that the dilatational properties of the adsorbed films depend on the molecular structure of the protein.

Hydration properties of soybean protein isolates

Estudaram-se as propriedades de hidrataçäo de isolados de soja com diferentes condiçöes de processamento (trratamentos térmicos, pH e concentraçäo de proteínas). Para diferentes amostras determinaram-se e correlacionaram-se o grau de desnaturalizaçäo, a solubilidade em água, em 0,2mol/L NaCl e em diferentes concentraçöes de dodecil sulfato de sódio, viscosidade e capacidade de absorçäo de água. Os tratamentos a temperaturas superiores aos 80§C desnaturaram as fracçöes 11S e 7S, provocando a exposiçäo de gruipos hidrofóbicos os que produziram agregados insolúveis, em água como em soluçäo com alta força ionica. Estes isolados possuíam alta capacidade de absorçäo de água e dispersöes com alta viscosidade. Achou-se uma correlaçäo significativa entre as propriedades de hidrataçäo e o coeficiente m, calculado através da funçäo de potência que relaciona a viscosidade com a concentraçäo proteica da dispersäo. Este coeficiente m também correlacionou com a entalpia dos isolados. Sobre a base destes resultados poderia-se sugerir que o coeficiente m dependente do comportamento hidrodinâmico das partículas foi um bom estimador do grau de desnaturaçäo proteica

Desarrollo de un ELISA de alta detectabilidad y especificidad para la cuantificación de gliadinas en alimentos destinados a enfermos celíacos / Development of a quantitative ELISA to determination of gladins in food aimed to coeliac patients

La enfermedad celíaca (EC) es una enfermedad gastrointestinal crónica de muy alta incidencia en nuestro país. Se estima que puede afectar 1 de cada 300 habitantes. En individuos susceptibles, la patología es provocada por la ingestión de mínimas cantidades de prolaminas de los cereales: trigo, triticale, cebada y centeno. Su único tratamiento es una estricta dieta libre de dichas proteínas. La Organización Mundial de la Salud (OMS) establece que un alimento puede ser considerado como apto para consumo por enfermos celíacos sólo si su contenido de gluten es inferior a 1 mg/100 g de producto seco. La detección precisa de estas proteínas requiere entonces de métodos de alta detectabilidad y especificidad para discriminar entre las proteínas nocivas y las de otros vegetales frecuentemente usados como reemplazo en la formulación de los alimentos para estos pacientes. En este trabajo, se muestra el desarrollo de un ELISA con anticuerpos policlonales, para la cuantificación de gliadinas en alimentos destinados a enfermos celíacos. Se empleó un diseño de ELISA competitivo secuencial con un antisuero obtenido en conejos que detecta selectivamente las prolaminas tóxicas. Este inmunoensayo presenta un nivel de detección de 0,1 mg de gluten/100 g de producto y cumple con los niveles de detectabilidad aconsejados por la OMS. Mediante el ELISA descripto se han analizado una gran variedad de muestras comerciales, pudiendo cuantificar las prolaminas incluso en alimentos que sufrieron tratamientos térmicos durante su fabricación

Desarrollo de un ELISA de alta detectabilidad y especificidad para la cuantificación de gliadinas en alimentos destinados a enfermos celíacos / Development of a quantitative ELISA to determination of gladins in food aimed to coeliac patients

La enfermedad celíaca (EC) es una enfermedad gastrointestinal crónica de muy alta incidencia en nuestro país. Se estima que puede afectar 1 de cada 300 habitantes. En individuos susceptibles, la patología es provocada por la ingestión de mínimas cantidades de prolaminas de los cereales: trigo, triticale, cebada y centeno. Su único tratamiento es una estricta dieta libre de dichas proteínas. La Organización Mundial de la Salud (OMS) establece que un alimento puede ser considerado como apto para consumo por enfermos celíacos sólo si su contenido de gluten es inferior a 1 mg/100 g de producto seco. La detección precisa de estas proteínas requiere entonces de métodos de alta detectabilidad y especificidad para discriminar entre las proteínas nocivas y las de otros vegetales frecuentemente usados como reemplazo en la formulación de los alimentos para estos pacientes. En este trabajo, se muestra el desarrollo de un ELISA con anticuerpos policlonales, para la cuantificación de gliadinas en alimentos destinados a enfermos celíacos. Se empleó un diseño de ELISA competitivo secuencial con un antisuero obtenido en conejos que detecta selectivamente las prolaminas tóxicas. Este inmunoensayo presenta un nivel de detección de 0,1 mg de gluten/100 g de producto y cumple con los niveles de detectabilidad aconsejados por la OMS. Mediante el ELISA descripto se han analizado una gran variedad de muestras comerciales, pudiendo cuantificar las prolaminas incluso en alimentos que sufrieron tratamientos térmicos durante su fabricación (AU)