Extrusion Improves the Antihypertensive Potential of a Kabuli Chickpea (Cicer arietinum L.) Protein Hydrolysate.

Chickpea hydrolysates could have antihypertensive potential, but there are no evaluations in vivo. Thus, the antihypertensive potential of a chickpea protein hydrolysate obtained before and after extrusion (a process that modifies protein digestibility) was evaluated. Protein precipitates were obtained from extruded and unextruded chickpea flours by isoelectric precipitation and hydrolyzed (α-amylase/pepsin/pancreatin). Chemical composition was determined (standard methods). ACE-I inhibition assays were carried out using a colorimetric test. For antihypertensive effect evaluations, spontaneously hypertensive rats (n = 8) received the treatments intragastrically (extruded or unextruded hydrolysate (1.2 g/kg), captopril (25 mg/kg), or water only). Fat, ash, and carbohydrate contents were lower in extruded chickpea flour (p < 0.05 versus unextruded). The protein content varied between protein precipitates (91.03%/78.66% unextruded/extruded (dry basis)) (p < 0.05). The hydrolysates' IC50 values (mg/mL) were 0.2834 (unextruded)/0.3218 (extruded) (p > 0.05). All treatments lowered the blood pressure (p < 0.05 vs. water). The extruded hydrolysate showed a more potent antihypertensive effect than the unextruded one (p < 0.05), an effect similar to captopril (p > 0.05). The results suggest that protein extrusion can be used to generate protein hydrolysates with improved health benefits. The findings have implications for the design and production of functional foods that could help to prevent hypertension or serve as an adjunct in its treatment.

Anti-inflammatory and antioxidant effects of peptides released from germinated amaranth during in vitro simulated gastrointestinal digestion.

Amaranth (Amaranthus hypochondriacus) is an ancestral nutritional grain and good source of bioactive compounds as peptides. In this study, the effect of in vitro simulated gastrointestinal digestion (SGD) of germinated amaranth on the release of antioxidant and anti-inflammatory peptides was evaluated. The germinated amaranth peptides generated during SGD were released after 90 min of incubation with pancreatin and fractioned to F1 (>10 kDa), F2 (3-10 kDa), and F3 (<3 kDa). Among germinated amaranth peptides fractions tested, F2 had the highest antioxidant activity, while F1 and F2 exhibited a high anti-inflammatory response caused by lipopolysaccharide-induced in RAW 264.7 macrophages. A total of 11 peptides sequences were identified in the fractions evaluated, and they exhibit potential biological activity against non-communicable diseases. The findings from this study showed first time report on bioactive peptides, especially anti-inflammatory, from germinated amaranth released by in vitro gastrointestinal digestion.

Phytochemical Compounds and Antioxidant Activity Modified by Germination and Hydrolysis in Mexican Amaranth.

Amaranth (Amaranthus spp.) grains have become essential for human health and nutrition; due to the presence of bioactive compounds that have shown some biological activities. This study aimed to evaluate the effect of germination, enzymatic hydrolysis, and its combination on the phytochemical compounds and antioxidant activity in Mexican amaranth. Germinated amaranth flours (GAF) exhibited increases in the concentrations of soluble protein (SP), total phenolic content (TPC), total flavonoid content (TFC), total anthocyanin content (TAC) and antioxidant activity (AOX) by 35.7, 17.2, 163.0, 1472.2, and 54.3%, respectively, compared with ungerminated amaranth flours (UAF). In SDS-PAGE, both hydrolysates of UAF and GAF exhibited low molecular weight bands (< 10 kDa). The hydrolysates of UAFH and GAFH had the highest degree of hydrolysis at 205 min of sequential hydrolysis (pepsin with pancreatin) time with 73.4 and 60.3%, respectively. Both hydrolysates obtained from GAF and UAF released significantly SP, TPC, TFC after sequential enzymatic hydrolysis (up 205 min), which led to a remarkable improvement of AOX when compared to nonhydrolyzed amaranth samples. The UAFH and GAFH had the best AOX at 270 min of enzymatic hydrolysis with 983.1 and 1304.9 µmol TE/mg SP, respectively. Hence, the combination of germination and enzymatic hydrolysis could be used to produce functional ingredients for food product development.

Phenolic Acids Profiles and Cellular Antioxidant Activity in Tortillas Produced from Mexican Maize Landrace Processed by Nixtamalization and Lime Extrusion Cooking.

Phenolic acids profiles, chemical antioxidant activities (ABTS and ORAC), as well as cellular antioxidant activity (CAA) of tortilla of Mexican native maize landraces elaborated from nixtamalization and lime cooking extrusion processes were studied. Both cooking procedures decreased total phenolics, chemicals antioxidant activity when compared to raw grains. Extruded tortillas retained 79.6-83.5%, 74.1-77.6% and 79.8-80.5% of total phenolics, ABTS and ORAC values, respectively, compared to 47.8-49.8%, 41.3-42.3% and 43.7-44.4% assayed in traditional tortillas, respectively. Approximately 72.5-88.2% of ferulic acid in raw grains and their tortillas were in the bound form. Regarding of the CAA initially found in raw grains, the retained percentage for traditional and extruded tortillas ranged from 47.4 to 48.7% and 72.8 to 77.5%, respectively. These results suggest that Mexican maize landrace used in this study could be considered for the elaboration of nixtamalized and extruded food products with nutraceutical potential.

Characterization of peptides found in unprocessed and extruded amaranth (Amaranthus hypochondriacus) pepsin/pancreatin hydrolysates.

The objectives of this study were to characterize peptides found in unprocessed amaranth hydrolysates (UAH) and extruded amaranth hydrolysates (EAH) and to determine the effect of the hydrolysis time on the profile of peptides produced. Amaranth grain was extruded in a single screw extruder at 125 °C of extrusion temperature and 130 rpm of screw speed. Unprocessed and extruded amaranth flour were hydrolyzed with pepsin/pancreatin enzymes following a kinetic at 10, 25, 60, 90, 120 and 180 min for each enzyme. After 180 min of pepsin hydrolysis, aliquots were taken at each time during pancreatin hydrolysis to characterize the hydrolysates by MALDI-TOF/MS-MS. Molecular masses (MM) (527, 567, 802, 984, 1295, 1545, 2034 and 2064 Da) of peptides appeared consistently during hydrolysis, showing high intensity at 10 min (2064 Da), 120 min (802 Da) and 180 min (567 Da) in UAH. EAH showed high intensity at 10 min (2034 Da) and 120 min (984, 1295 and 1545 Da). Extrusion produced more peptides with MM lower than 1000 Da immediately after 10 min of hydrolysis. Hydrolysis time impacted on the peptide profile, as longer the time lower the MM in both amaranth hydrolysates. Sequences obtained were analyzed for their biological activity at BIOPEP, showing important inhibitory activities related to chronic diseases. These peptides could be used as a food ingredient/supplement in a healthy diet to prevent the risk to develop chronic diseases.

Identification of Bioactive Peptide Sequences from Amaranth (Amaranthus hypochondriacus) Seed Proteins and Their Potential Role in the Prevention of Chronic Diseases.

Amaranth (Amaranthus hypochondriacus) is a pseudocereal with higher protein concentration than most cereal grains. Enzymatic hydrolysis and food processing could produce biopeptides from amaranth proteins; however, there is limited information about the bioactivity of peptides from amaranth proteins. The objective of this comprehensive review was to determine bioactive peptide sequences in amaranth proteins that may prevent cardiovascular disease, cancer, and diabetes. Amaranth proteins, reported in UniProt database, were evaluated for potential bioactive peptide using BIOPEP database. The 15 main proteins present in amaranth seed are 11S globulin, 7S globulin, α-amylase inhibitor, trypsin inhibitor, antimicrobial proteins, nonspecific lipid-transfer-protein-1, superoxide dismutase, ring-zinc finger protein, prosystemin, amaranth albumin 1, glucose-1-phosphate adenyltransferase, glucosyltransferase, polyamine oxidase, granule-bound starch synthase 1, and acetolactate synthase. All proteins showed high occurrence frequencies of angiotensin-converting enzyme-inhibitor peptides (A = 0.161 to 0.362), as well as of dipeptidyl peptidase IV inhibitor (A = 0.003 to 0.087). Other proteins showed antioxidative (A = 0.012 to 0.063) and glucose uptake-stimulating activity (A = 0.023 to 0.042), and also antithrombotic (A = 0.002 to 0.031) and anticancer sequences (A = 0.001 to 0.042). The results of this study support the concept that amaranth grain could be part of a "healthy" diet and thereby prevent chronic human diseases.

Pepsin-pancreatin protein hydrolysates from extruded amaranth inhibit markers of atherosclerosis in LPS-induced THP-1 macrophages-like human cells by reducing expression of proteins in LOX-1 signaling pathway.

BACKGROUND: Atherosclerosis is considered a progressive disease that affects arteries that bring blood to the heart, to the brain and to the lower end. It derives from endothelial dysfunction and inflammation, which play an important role in the thrombotic complications of atherosclerosis. Cardiovascular disease is the leading cause of death around the world and one factor that can contribute to its progression and prevention is diet. Our previous study found that amaranth hydrolysates inhibited LPS-induced inflammation in human and mouse macrophages by preventing activation of NF-κB signaling. Furthermore, extrusion improved the anti-inflammatory effect of amaranth protein hydrolysates in both cell lines, probably attributed to the production of bioactive peptides during processing. Therefore, the objective of this study was to compare the anti-atherosclerotic potential of pepsin-pancreatin hydrolysates from unprocessed and extruded amaranth in THP-1 lipopolysaccharide-induced human macrophages and suggest the mechanism of action. RESULTS: Unprocessed amaranth hydrolysate (UAH) and extruded amaranth hydrolysate (EAH) showed a significant reduction in the expression of interleukin-4 (IL-4) (69% and 100%, respectively), interleukin-6 (IL-6) (64% and 52%, respectively), interleukin-22 (IL-22) (55% and 70%, respectively). Likewise, UAH and EAH showed a reduction in the expression of monocyte-chemo attractant protein-1 (MCP-1) (35% and 42%, respectively), transferrin receptor-1 (TfR-1) (48% and 61%, respectively), granulocyte-macrophage colony-stimulating factor (GM-CSF) (59% and 63%, respectively), and tumor necrosis factor-α (TNF-α) (60% and 63%, respectively). Also, EAH reduced the expression of lectin-like oxidized low-density lipoprotein receptor-1 (LOX-1) (27%), intracellular adhesion molecule-1 (ICAM-1) (28%) and matrix metalloproteinase-9 (MMP-9) (19%), important molecular markers in the atherosclerosis pathway. EAH, led to a reduction of 58, 52 and 79% for LOX-1, ICAM-1 and MMP-9, respectively, by confocal microscopy. CONCLUSIONS: Extruded amaranth hydrolysate showed potential anti-atherosclerotic effect in LPS-induced THP-1 human macrophage-like cells by reducing the expression of proteins associated with LOX-1 signaling pathway.

Extrusion improved the anti-inflammatory effect of amaranth (Amaranthus hypochondriacus) hydrolysates in LPS-induced human THP-1 macrophage-like and mouse RAW 264.7 macrophages by preventing activation of NF-κB signaling.

SCOPE: The objective was to compare the anti-inflammatory potential of unprocessed and extruded amaranth pepsin/pancreatin hydrolysates in LPS-induced human THP-1 macrophages-like and mouse RAW 264.7 macrophages focusing on their anti-inflammatory mechanism of action related to NF-κB signaling pathway. METHODS AND RESULTS: Amaranth hydrolysates were characterized by MS-MS and tested for anti-inflammatory effects on human and mouse macrophages. Peptides found in extruded amaranth hydrolysates displayed antioxidant capacity, angiotensin converting enzyme-inhibitor activity, and dipeptidyl peptidase-IV inhibitor activity. Gly-Pro-Arg peptide was present and reported as antithrombotic. Extruded amaranth hydrolysates (1 mg/mL) significantly reduced tumor necrosis factor alpha secretion in THP-1 and RAW 264.7 cells by 36.5 and 33.5%, respectively; with concomitant reduction in PGE2 (15.4 and 31.4%), and COX-2 (38.1 and 67.6%), respectively. Phosphorylation of IKK-α was significantly reduced by 52.5 and 88.2% leading to reduced phosphorylation of IκB-α (86.1 and 66.2%), respectively; resulting in a reduction in the expression of p65 NF-κB subunits in the nucleus by 64.2% for THP-1 and 70.7% for RAW 264.7 cells. CONCLUSION: Amaranth hydrolysates inhibited LPS-induced inflammation in human and mouse macrophages by preventing activation of NF-κB signaling. Extrusion improved anti-inflammatory effect of amaranth hydrolysates in both cells, which might be attributed to the production of bioactive peptides during processing.