Alcalase-Based Chickpea (Cicer arietinum L.) Protein Hydrolysates Efficiently Reduce Systolic Blood Pressure in Spontaneously Hypertensive Rats.

Studies on antihypertensive chickpea protein hydrolysates have rarely performed in vivo evaluations, limiting the entry of such hydrolysates into functional food development and clinical trials. Thus, our aim was to optimize the hydrolysis conditions to produce an alcalase-based chickpea hydrolysate with a hypotensive effect in vivo at convenient oral doses. The hydrolysis reaction time, temperature, and alcalase/substrate concentration were optimized using a response surface analysis (RSA). ACE-I inhibition was the response variable. The optimized hydrolysis conditions were time = 0.5 h, temperature = 40 °C, and E/S concentration = 0.254 (U/g). The IC50 of the optimized hydrolysate (OCPH) was 0.358 mg/mL. Five hydrolysates from the RSA worksheet (one of them obtained after 5 min of hydrolysis (CPH15)) had an ACE-I inhibitory potential similar to that of OCPH (p > 0.05). At 50 mg/kg doses, OCPH and CPH15 promoted a clinically relevant hypotensive effect in spontaneously hypertensive rats, up to -47.35 mmHg and -28.95 mmHg, respectively (p < 0.05 vs. negative control). Furthermore, the hypotensive effect was sustained for at least 7 h post-supplementation. Overall, OCPH and CPH15 are promising ingredients for functional food development and as test materials for clinical trials.

"NeuroVanguard": a contemporary strategy in neuromonitoring for severe adult brain injury patients.

Severe acute brain injuries, stemming from trauma, ischemia or hemorrhage, remain a significant global healthcare concern due to their association with high morbidity and mortality rates. Accurate assessment of secondary brain injuries severity is pivotal for tailor adequate therapies in such patients. Together with neurological examination and brain imaging, monitoring of systemic secondary brain injuries is relatively straightforward and should be implemented in all patients, according to local resources. Cerebral secondary injuries involve factors like brain compliance loss, tissue hypoxia, seizures, metabolic disturbances and neuroinflammation. In this viewpoint, we have considered the combination of specific noninvasive and invasive monitoring tools to better understand the mechanisms behind the occurrence of these events and enhance treatment customization, such as intracranial pressure monitoring, brain oxygenation assessment and metabolic monitoring. These tools enable precise intervention, contributing to improved care quality for severe brain injury patients. The future entails more sophisticated technologies, necessitating knowledge, interdisciplinary collaboration and resource allocation, with a focus on patient-centered care and rigorous validation through clinical trials.

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.

Functional, immunological characterization, and anticancer activity of BaMtx: A new Lys49- PLA2 homologue isolated from the venom of Peruvian Bothrops atrox snake (Serpentes: Viperidae).

Bothorps atrox is responsible for most of the ophidism cases in Perú. As part of the envenoming, myotoxicity is one of the most recurrent and destructive effects. In this study, a myotoxin, named BaMtx, was purified from B. atrox venom to elucidate its biological, immunological, and molecular characteristics. BaMtx was purified using CM-Sephadex-C-25 ion-exchange resin and SDS-PAGE analysis showed a unique protein band of 13 kDa or 24 kDa under reducing or non-reducing conditions, respectively. cDNA sequence codified a 122-aa mature protein with high homology with other Lys49-PLA2s; modeled structure showed a N-terminal helix, a ß-wing region, and a C-terminal random coil. This protein has a poor phospholipase A2 enzymatic activity. BaMtx has myotoxic (DMM = 12.30 ± 0.95 µg) and edema-forming (DEM = 26.00 ± 1.15 µg) activities. Rabbit immunization with purified enzyme produced anti-BaMtx antibodies that reduced 50.28 ± 10.15% of myotoxic activity and showed significant cross-reactivity against B. brazili and B pictus venoms. On the other hand, BaMtx exhibits mild anti-proliferative and anti-migratory effects on breast cancer cells, affecting the ROS and NADH levels, which may reduce mitochondrial respiration. These results contribute to the understanding of B. atrox Lys49-PLA2 effects and establish the anticancer potential de BaMtx.

Efficacy of the adjuvanted subunit protein COVID-19 vaccine, SCB-2019: a phase 2 and 3 multicentre, double-blind, randomised, placebo-controlled trial.

BACKGROUND: A range of safe and effective vaccines against SARS CoV 2 are needed to address the COVID 19 pandemic. We aimed to assess the safety and efficacy of the COVID-19 vaccine SCB-2019. METHODS: This ongoing phase 2 and 3 double-blind, placebo-controlled trial was done in adults aged 18 years and older who were in good health or with a stable chronic health condition, at 31 sites in five countries (Belgium, Brazil, Colombia, Philippines, and South Africa). The participants were randomly assigned 1:1 using a centralised internet randomisation system to receive two 0·5 mL intramuscular doses of SCB-2019 (30 µg, adjuvanted with 1·50 mg CpG-1018 and 0·75 mg alum) or placebo (0·9% sodium chloride for injection supplied in 10 mL ampoules) 21 days apart. All study staff and participants were masked, but vaccine administrators were not. Primary endpoints were vaccine efficacy, measured by RT-PCR-confirmed COVID-19 of any severity with onset from 14 days after the second dose in baseline SARS-CoV-2 seronegative participants (the per-protocol population), and the safety and solicited local and systemic adverse events in the phase 2 subset. This study is registered on EudraCT (2020-004272-17) and ClinicalTrials.gov (NCT04672395). FINDINGS: 30 174 participants were enrolled from March 24, 2021, until the cutoff date of Aug 10, 2021, of whom 30 128 received their first assigned vaccine (n=15 064) or a placebo injection (n=15 064). The per-protocol population consisted of 12 355 baseline SARS-CoV-2-naive participants (6251 vaccinees and 6104 placebo recipients). Most exclusions (13 389 [44·4%]) were because of seropositivity at baseline. There were 207 confirmed per-protocol cases of COVID-19 at 14 days after the second dose, 52 vaccinees versus 155 placebo recipients, and an overall vaccine efficacy against any severity COVID-19 of 67·2% (95·72% CI 54·3-76·8), 83·7% (97·86% CI 55·9-95·4) against moderate-to-severe COVID-19, and 100% (97·86% CI 25·3-100·0) against severe COVID-19. All COVID-19 cases were due to virus variants; vaccine efficacy against any severity COVID-19 due to the three predominant variants was 78·7% (95% CI 57·3-90·4) for delta, 91·8% (44·9-99·8) for gamma, and 58·6% (13·3-81·5) for mu. No safety issues emerged in the follow-up period for the efficacy analysis (median of 82 days [IQR 63-103]). The vaccine elicited higher rates of mainly mild-to-moderate injection site pain than the placebo after the first (35·7% [287 of 803] vs 10·3% [81 of 786]) and second (26·9% [189 of 702] vs 7·4% [52 of 699]) doses, but the rates of other solicited local and systemic adverse events were similar between the groups. INTERPRETATION: Two doses of SCB-2019 vaccine plus CpG and alum provides notable protection against the entire severity spectrum of COVID-19 caused by circulating SAR-CoV-2 viruses, including the predominating delta variant. FUNDING: Clover Biopharmaceuticals and the Coalition for Epidemic Preparedness Innovations.

Profiling modifications in physicochemical, chemical and antioxidant properties of wild blackberry (Rubus sp.) during fermentation with EC 1118 yeast.

Mexico is an extensively diverse country with a wide variety of wild species of blackberries (Rubus spp.), which are rich in bioactive compounds, however, these fruits are underutilized. Fermentation is a process that transforms the chemical compounds of fruits and increases nutraceutical properties. This study aimed to determine the physicochemical changes and the bioactive compounds profile that take place during the fermentation of wild blackberries using yeast EC 1118 and to evaluate its relationship with antioxidant activity (AOx). The results indicated that after 96 h of fermentation the content of carbohydrates (56%), total phenolic compounds (37%), and anthocyanins (22%), decreased, respectively. The physicochemical parameters showed statistic differences (p ≤ 0.05) at the endpoint of fermentation. The diversity of fatty acids was increased (55%), compared with unfermented blackberries. The modification of carbohydrates, anthocyanins, catechin, gallic and ellagic acid profiles were also monitored performing chromatographic techniques. The AOx, determined by ORAC and DPPH assays, showed the highest results for ORAC at 96 h increased a 140.2%, while DPPH values enhanced a 36.6% at 48 h of bioprocessing. Strong positive correlations were found between fermentation time and DPPH values (r = 0.8131), between ORAC and gallic acid content (r = 0.8688), and between anthocyanin content and pH (r = 0.9126). The fermentation of wild blackberries with EC 1118 yeast represents an alternative for development and formulation of potential ingredients for functional foods. SUPPLEMENTARY INFORMATION: The online version contains supplementary material available at (10.1007/s13197-020-04953-x).

In vitro gastrointestinal digestion impact on stability, bioaccessibility and antioxidant activity of polyphenols from wild and commercial blackberries (Rubus spp.).

Gastrointestinal digestion (GID) is a physiological process that transforms the stability, bioaccessibility and antioxidant activity (AOX) of polyphenols from blackberries (Rubus spp.). This study aimed to investigate the effect of the INFOGEST® GID protocol on the phenolic stability, bioaccessibility and AOX of Mexican wild (WB) and commercial (CB) blackberries. After GID, the total phenolic and anthocyanin contents in blackberries decreased by ≥68% and ≥74%, respectively. More than 40 phenolics were identified during GID; most of them degraded completely during digestion. GID had a negative effect on the AOX of both fruits (>50%), but WB showed the highest antioxidant activities, as assessed by the ORAC, DPPH, reducing power and ß-carotene bleaching methods. In Caco-2 cells, the cell-based antioxidant activity of digested blackberries (p < 0.05) decreased by 48% in WB and by 56% in CB. The capacity to inhibit intracellular ROS decreased by 50% in WB and by up to 86% in CB, after digestion. GID is a complex process that impacts on the bioactive properties of food nutrients, especially phenolics. In vitro and cellular AOX of WB polyphenols withstood the gastrointestinal environment better than CB phenolics. The in vitro assays results suggest that phenolics from underutilized WB have a higher bioaccessibility and antioxidant capacity than the polyphenols from the most frequently consumed CB. However, whether this corresponds to a better bioaccessibility in humans remains to be determined in future work.

Impact of in vitro gastrointestinal digestion on peptide profile and bioactivity of cooked and non-cooked oat protein concentrates.

Oat (Avena sativa) is one of the most cultivated and consumed cereals worldwide. Recognized among cereals for its high protein content (12%-24%), it makes it an excellent source of bioactive peptides, which could be modified during processes such as heating and gastrointestinal digestion (GID). This work aims to evaluate the impact of heat treatment on the proteolysis of oat proteins and on the evolution of antioxidant peptide released during in vitro static GID, in terms of comparative analysis between cooked oat protein concentrate (COPC) and non-heated oat protein concentrate (OPC) samples. The protein extraction method and cooking procedure used showed no detrimental effects on protein quality. After GID, the proportion of free amino acids/dipeptides (<0.2 â€‹kDa) reached >40% for both samples (OPC and COPC), thus producing peptides with low molecular weight and enhanced bioactivity. Furthermore, during GID, the amino acid profile showed an increase in essential, positively-charged, hydrophobic and aromatic amino acids. At the end of GID, the reducing power of OPC and COPC increased >0.3 and 8-fold, respectively, in comparison to the non-digested samples; while ABTS•+ and DPPH• showed a >20-fold increase. Fe2+ chelating capacity of OPC and COPC was enhanced >4 times; similarly, Cu2+ chelation showed a >19-fold enhancement for OPC and >10 for COPC. ß-carotene bleaching activity was improved 0.8 times in OPC and >9 times in COPC; the oxygen radical antioxidant capacity assay increased 2 times in OPC and >4.7 times in COPC, respectively. This study suggests that OPC after cooking and GID positively influenced the nutritional and bioactive properties of oat peptides. Thus, COPC could be used as a functional food ingredient with health-promoting effects, as hydrothermal treatment is frequently used for this type of cereals.

Chikungunya encephalitis, a case series from an endemic country.

BACKGROUND: The Chikungunya Virus (CHIKV) was introduced into Honduras in 2015. Since then the WHO has reported more than 14,000 suspected cases in the country. OBJECTIVE: To describe the clinical, laboratory, neuroimaging, and pathological features of CHIKV encephalitis. PATIENTS AND METHODS: We evaluated all consecutive cases of CHIKV infection meeting encephalitis criteria at Hospital Escuela Universitario at Tegucigalpa, Honduras, during 2015. Who case definition was used: patient with neurological manifestations meeting clinical criteria (fever >38.5 °C, joint pain); resident/visitor in the last 15 days to an endemic area; laboratory confirmation with IgM/ELISA. Other etiologies were excluded by ancillary studies. RESULTS: Out of 95 cases with suspected CHIKV infection, 7 (7%) cases with CHIKV encephalitis were identified; mean age was 56 years and four were men. The mean latency from onset of symptoms to diagnosis was 5 five days. Clinical manifestations were: fever/arthralgia, headache/alteration of consciousness and status epilepticus. The EEG demonstrated slow background activity and generalized epileptiform discharges in three patients. Brain MRI showed bilateral white matter hyperintensities and one with focal encephalitis; CSF analysis demonstrated lymphocytic pleocytosis and hyperproteinorrachia. Two patients died. Postmortem brain examination of one patient revealed lymphocytic infiltrates with focal necrosis in hippocampus, frontal lobes and medulla oblongata. CONCLUSIONS: Neurological complications of CHIKV are infrequent, but may be severe. In this case series, the neurological manifestation was encephalitis. Predominant symptoms and signs were fever, behavioral abnormalities, headache and seizures. Because of the potential morbidity and mortality of CHIKV encephalitis, these patients should be admitted to hospital urgently.

Improving Polyphenolic Compounds: Antioxidant Activity in Chickpea Sprouts through Elicitation with Hydrogen Peroxide.

Elicitation appears to be a promising alternative to enhance the bioactive compound content and biological activities of legume sprouts. Multi-response optimization by response surface methodology (RSM) with desirability function (DF) was used to optimize the elicitor concentration (hydrogen peroxide (H2O2)) and germination time in order to maximize total phenolic content (TPC), total flavonoids content (TFC), and antioxidant activity (AOX) of chickpea sprouts. Chemical, antinutritional, and nutraceutical properties of optimized chickpea sprouts (OCS) were also determined. The predicted regression models developed were efficiently fitted to the experimental data. The results of the desirability function revealed that optimum attributes in chickpea sprouts can be achieved by the application of 30 mM H2O2 and 72 h of germination time, with global desirability value D = 0.893. These OCS had higher (p < 0.05) TPC (7.4%), total iso-flavonoids (16.5%), AOX (14.8%), and lower phytic acid (16.1%) and saponins (21.8%) compared to H2O2 non-treated chickpea sprouts. Optimized germination conditions slightly modified the flavonoid profile in chickpea; eight iso-flavonoids were identified in OCS, including formononetin and biochanin A, which were identified as the major compounds. Results from this study support elicitation with H2O2 as an effective approach to improve phytochemical content and antioxidant activity in chickpea sprouts.

High level of pre-treatment and acquired HIV drug resistance in Honduras: a nationally representative survey, 2016-17.

BACKGROUND: Pre-treatment HIV drug resistance (HIVDR) to NNRTIs has consistently increased in low-/middle-income countries during the last decade. OBJECTIVES: To estimate the prevalence of pre-treatment HIVDR and acquired HIVDR among persons living with HIV (PLHIV) on ART for 12 ±âŸ3 months (ADR12) and ≥48 months (ADR48) in Honduras. PATIENTS AND METHODS: A nationwide cross-sectional survey with a two-stage cluster sampling was conducted from October 2016 to November 2017. Twenty-two of 54 total ART clinics representing >90% of the national cohort of adults on ART were included. HIVDR was assessed for protease and reverse transcriptase Sanger sequences using the Stanford HIVdb tool. RESULTS: A total of 729 PLHIV were enrolled; 26.3% (95% CI 20.1%-33.5%) ART initiators reported prior exposure to antiretrovirals. Pre-treatment HIVDR prevalence was 26.9% (95% CI 20.2%-34.9%) to any antiretroviral and 25.9% (19.2%-33.9%) to NNRTIs. NNRTI pre-treatment HIVDR was higher in ART initiators with prior exposure to antiretrovirals (P = 0.001). Viral load (VL) suppression rate was 89.7% (85.1%-93.0%) in ADR12 and 67.9% (61.7%-73.6%) in ADR48. ADR12 to any drug among PLHIV with VL ≥1000 copies/mL was 86.1% (48.9%-97.6%); 67.1% (37.4%-87.5%) had HIVDR to both NNRTIs and NRTIs, and 3.8% (0.5%-25.2%) to PIs. ADR48 was 92.0% (86.8%-95.3%) to any drug; 78.1% (66.6%-86.5%) to both NNRTIs and NRTIs, and 7.3% (1.8%-25.1%) to PIs. CONCLUSIONS: The high prevalence of NNRTI pre-treatment HIVDR observed in Honduras warrants consideration of non-NNRTI-based first-line regimens for ART initiation. Programmatic improvements in HIVDR monitoring and adherence support may also be considered.

Fibrinogen-clotting enzyme, pictobin, from Bothrops pictus snake venom. Structural and functional characterization.

A thrombin-like enzyme, pictobin, was purified from Bothrops pictus snake venom. It is a 41-kDa monomeric glycoprotein as showed by mass spectrometry and contains approx. 45% carbohydrate by mass which could be removed with N-glycosidase. Pictobin coagulates plasma and fibrinogen, releasing fibrinopeptide A and induces the formation of a friable/porous fibrin network as visualized by SEM. The enzyme promoted platelet aggregation in human PRP and defibrination in mouse model and showed catalytic activity on chromogenic substrates S-2266, S-2366, S-2160 and S-2238. Pictobin interacts with the plasma inhibitor α2-macroglobulin, which blocks its interaction with fibrinogen but not with the small substrate BApNA. Heparin does not affect its enzymatic activity. Pictobin cross reacted with polyvalent bothropic antivenom, and its deglycosylated form reduced its catalytic action and antivenom reaction. In breast and lung cancer cells, pictobin inhibits the fibronectin-stimulated migration. Moreover, it produces strong NADH oxidation, mitochondrial depolarization, ATP decrease and fragmentation of mitochondrial network. These results suggest by first time that a snake venom serinprotease produces mitochondrial dysfunction by affecting mitochondrial dynamics and bioenergetics. Structural model of pictobin reveals a conserved chymotrypsin fold ß/ß hydrolase. These data indicate that pictobin has therapeutic potential in the treatment of cardiovascular disorders and metastatic disease.

Spatial distribution of dengue in Honduras during 2016-2019 using a geographic information systems (GIS)-Dengue epidemic implications for public health and travel medicine.

BACKGROUND: After serious epidemics of chikungunya (CHIKV) and Zika (ZIKV) in the Americas, dengue (DENV) have reemerged in most countries. We analyzed the incidence, incidence rates, and evolution of DENV cases in Honduras from 2015 to 2018 and the ongoing 2019 epidemic. METHODS: Using epidemiological weeks (EW) surveillance data on the DENV in Honduras, we estimated incidence rates (cases/100,000 population), and developed maps at national, departmental, and municipal levels. RESULTS: From January 1, 2016 to July 21, 2019, a total of 109,557 cases of DENV were reported, 28,603 in 2019, with an incidence rate of 312.32 cases/100,000 population this year; 0.13% laboratory-confirmed. The highest peak was reached on the EW 28°, 2019 (5299 cases; 57.89 cases/100,000 population). The department with the highest number of cases and incidence rate was Cortes (8404 cases, 479.68 cases/100,000 population in 2019). DISCUSSION: The pattern and evolution of DENV epidemic in 2019 in Honduras has been similar to that which occurred for in 2015. As previously reported, this epidemic involved the north and central areas of the country predominantly, reaching municipality incidences there >1000 cases/100,000 population (or 1%). Studies using geographical information systems linked with clinical disease characteristics are necessary to obtain accurate epidemiological data for public health systems. Such information is also useful for assessment of risk for travelers who visit specific areas in a destination country.

Biochemical and molecular characterization of the hyaluronidase from Bothrops atrox Peruvian snake venom.

Snake venoms are a rich source of enzymes such as metalloproteinases, serine proteinases phospholipases A2 and myotoxins, that have been well characterized structurally and functionally. However, hyaluronidases (E.C.3.2.1.35) have not been studied extensively. In this study, we describe the biochemical and molecular features of a hyaluronidase (Hyal-Ba) isolated from the venom of the Peruvian snake Bothrops atrox. Hyal-Ba was purified by a combination of ion-exchange and gel filtration chromatography. Purified Hyal-Ba is a 69-kDa (SDS-PAGE) monomeric glycoprotein with an N-terminal amino acid sequence sharing high identity with homologous snake venom hyaluronidases. Detected associated carbohydrates were hexoses (16.38%), hexosamines (2.7%) and sialic acid (0.69%). Hyal-Ba selectively hydrolyzed only hyaluronic acid (HA; specific activity = 437.5 U/mg) but it did not hydrolyze chondroitin sulfate or heparin. The optimal pH and temperature for maximum activity were 6.0 and 40 °C, respectively, and its Km was 0.31 µM. Its activity was inhibited by EDTA, iodoacetate, 2-mercaptoethanol, TLCK and dexamethasone. Na+ and K+ (0.2 M) positively affect hyaluronidase activity; while Mg2+, Br2+, Ba2+, Cu2+, Zn2+, and Cd2+ reduced catalytic activity. Hyal-Ba potentiates the hemorrhagic and hemolytic activity of whole venom, but decreased subplantar edema caused by an l-amino acid oxidase (LAAO). The Hyal-Ba cDNA sequence (2020 bp) encodes 449 amino acid residues, including the catalytic site residues (Glu135, Asp133, Tyr206, Tyr253 and Trp328) and three functional motifs for N-linked glycosylation, which are conserved with other snake hyaluronidases. Spatial modeling of Hyal-Ba displayed a TIM-Barrel (α/ß) fold and an EGF-like domain in the C-terminal portion. The phylogenetic analysis of Hyal-Ba with other homologous Hyals showed the monophyly of viperids. Further, Hyal-Ba studies may extend our knowledge of B. atrox toxinology and provides insight to improve the neutralizing strategies of therapeutic antivenoms.

Assessing the Sensitizing and Allergenic Potential of the Albumin and Globulin Fractions from Amaranth (Amaranthus hypochondriacus) Grains before and after an Extrusion Process.

Background: The first cases of food allergy to amaranth grain have recently been published. This pseudocereal is considered hypoallergenic, and there is scarce information about the allergenic potential of amaranth proteins, either before or after food processing. Objective: To evaluate, in a mouse model of food allergy, the sensitizing and allergenic potential of extruded and non-extruded albumin and globulin fractions from amaranth grains. Materials and Methods: Amaranth (Amaranthus hypochondriacus) flour was obtained and the albumin and globulin fractions isolated. These protein fractions were also obtained after flour extrusion. An intraperitoneal 28-day protocol was carried out to evaluate the sensitizing and allergenic potential of the proteins. The common and rarely allergenic proteins ovalbumin and potato acidic phosphatase were utilized as reference. Specific IgE and IgG antibodies were evaluated for all the proteins tested. Mast cell protease-1 (mMCP-1) responses were evaluated in serum samples collected after intragastric challenges with the proteins of interest. All serological evaluations were carried out using ELISA. Results: Mice were sensitized to the non-extruded albumin fraction from amaranth grains and to ovalbumin (p = 0.0045). The extrusion process of amaranth proteins abrogated the IgE responses triggered under non-extruded conditions (p = 0.0147). mMCP-1 responses were significantly detected in the group of mice sensitized to ovalbumin (p = 0.0138), but not in others. Conclusions: The non-extruded albumin fraction from amaranth has the potential to sensitize BALB/c mice, but this sensitizing potential fails to induce detectable serum levels of the mast cell degranulation marker mMCP-1 after intragastric challenges. Furthermore, the extrusion process abolished the sensitization potential of the amaranth albumins.

Germination in Optimal Conditions as Effective Strategy to Improve Nutritional and Nutraceutical Value of Underutilized Mexican Blue Maize Seeds.

Germination of grains is a bioprocess of emerging interest to improve nutritional and nutraceutical profile of cereals in a natural way. The aim of this work was to identify optimal germination conditions (temperature/duration) for producing a functional blue maize flour with maximum values of protein content (PC), antioxidant activity (AoxA), and total phenolic and anthocyanin contents (TPC, TAC). A central composite rotatable experimental design (response surface methodology) with two factors [Germination temperature (Gtemp, 20-40 °C) / Germination duration (Gdur, 12-220 h)] in five levels was used (13 treatments). Blue maize seeds were soaked in distilled water (25 °C / 12 h) before germination. The sprouts were dried, tempered (25 °C), and ground to obtain germinated blue maize flours (GBMF). The prediction models developed for each response variable showed high coefficients of determination, demonstrating their adequacy to explain the variations in experimental data. Maximum values of PC, AoxA, TPC, and TAC were attained at Gtemp = 26.9 °C / Gdur = 207.7 h. Optimized germinated blue maize flour (OGBMF) presented higher PC (+38.48%), AoxA (ABTS: +192%, ORAC: +160%, DPPH: +148%), TPC (+79%), and TAC (+9.9%) than unprocessed blue maize flour (UBMF). Germination at optimal conditions is an effective strategy to increase the nutritional/nutraceutical quality of blue maize seeds, thus the flour of these germinated seeds could be used for the development of functional foods.

Amaranth Protein Hydrolysates Efficiently Reduce Systolic Blood Pressure in Spontaneously Hypertensive Rats.

Alcalase is the enzyme of choice to release antihypertensive peptides from amaranth proteins, but the hydrolysis conditions have not been optimized yet. Furthermore, in vivo assays are needed to confirm such a hypotensive effect. Our aim was to optimize the hydrolysis of amaranth protein with alcalase and to test in vivo the hypotensive effect of the hydrolysates. A response surface analysis was carried out to optimize the hydrolysis reaction. The response variable was the Angiotensin Converting Enzyme (ACE-I) inhibition. The hydrolysis degree was determined (free alpha-amino groups measurement). The optimized hydrolysate bioavailability was assessed in the sera of mice and the hypotensive effect was assessed in spontaneously hypertensive rats. Control groups were administered captopril or water. The optimized hydrolysis conditions were: pH = 7.01, temperature = 52 °C, enzyme concentration 0.04 mU/mg, and time = 6.16 h. The optimized hydrolysate showed a 93.5% of ACE-I inhibition and a hydrolysis degree of 74.77%. After supplementation, the hydrolysate was bioavailable in mice from 5 to 60 min, and the hypotensive effect started at 4 h in spontaneously hypertensive rats (p < 0.05 vs. water group). This effect was similar to the captopril hypotensive effect for the next 3 h (p > 0.05). The use of amaranth-optimized hydrolysates as hypotensive supplements or ingredient for functional foods seems feasible.

Biochemical, biological and molecular characterization of an L-Amino acid oxidase (LAAO) purified from Bothrops pictus Peruvian snake venom.

An L-amino acid oxidase from Peruvian Bothrops pictus (Bpic-LAAO) snake venom was purified using a combination of size-exclusion and ion-exchange chromatography. Bpic-LAAO is a homodimeric glycosylated flavoprotein with molecular mass of ∼65 kDa under reducing conditions and ∼132 kDa in its native form as analyzed by SDS-PAGE and gel filtration chromatography, respectively. N-terminal amino acid sequencing showed highly conserved residues in a glutamine-rich motif related to binding substrate. The enzyme exhibited optimal activity towards L-Leu at pH 8.5, and like other reported SV-LAAOs, it is stable until 55 °C. Kinetic studies showed that the cations Ca2+, Mg2+ and Mn2+ did not alter Bpic-LAAO activity; however, Zn2+ is an inhibitor. Some reagents such as ß-mercaptoethanol, glutathione and iodoacetate had inhibitory effect on Bpic-LAAO activity, but PMSF, EDTA and glutamic acid did not affect its activity. Regarding the biological activities of Bpic-LAAO, this enzyme induced edema in mice (MED = 7.8 µg), and inhibited human platelet aggregation induced by ADP in a dose-dependent manner and showed antibacterial activity on Gram (+) and Gram (-) bacteria. Bpic-LAAO cDNA of 1494 bp codified a mature protein with 487 amino acid residues comprising a signal peptide of 11 amino acids. Finally, the phylogenetic tree obtained with other sequences of LAAOs, evidenced its similarity to other homologous enzymes, showing two well-established monophyletic groups in Viperidae and Elapidae families. Bpic-LAAO is evolutively close related to LAAOs from B. jararacussu, B. moojeni and B. atrox, and together with the LAAO from B. pauloensis, form a well-defined cluster of the Bothrops genus.