Integration of targeted metabolome and transcript profiling of Pseudomonas syringae-triggered changes in defence-related phytochemicals in oat plants.

MAIN CONCLUSION: A gene-to-metabolite approach afforded new insights regarding defence mechanisms in oat plants that can be incorporated into plant breeding programmes for the selection of markers and genes related to disease resistance. Monitoring metabolite levels and changes therein can complement and corroborate transcriptome (mRNA) data on plant-pathogen interactions, thus revealing mechanisms involved in pathogen attack and host defence. A multi-omics approach thus adds new layers of information such as identifying metabolites with antimicrobial properties, elucidating metabolomic profiles of infected and non-infected plants, and reveals pathogenic requirements for infection and colonisation. In this study, two oat cultivars (Dunnart and SWK001) were inoculated with Pseudomonas syringae pathovars, pathogenic and non-pathogenic on oat. Following inoculation, metabolites were extracted with methanol from leaf tissues at 2, 4 and 6 days post-infection and analysed by multiple reaction monitoring (MRM) on a triple quadrupole mass spectrometer system. Relatedly, mRNA was isolated at the same time points, and the cDNA analysed by quantitative PCR (RT-qPCR) for expression levels of selected gene transcripts associated with avenanthramide (Avn) biosynthesis. The targeted amino acids, hydroxycinnamic acids and Avns were successfully quantified. Distinct cultivar-specific differences in the metabolite responses were observed in response to pathogenic and non-pathogenic strains. Trends in aromatic amino acids and hydroxycinnamic acids seem to indicate stronger activation and flux through these pathways in Dunnart as compared to SWK001. A positive correlation between hydroxycinnamoyl-CoA:hydroxyanthranilate N-hydroxycinnamoyl transferase (HHT) gene expression and the abundance of Avn A in both cultivars was documented. However, transcript profiling of selected genes involved in Avn synthesis did not reveal a clear pattern to distinguish between the tolerant and susceptible cultivars.

Pharmacological and In Silico Analysis of Oat Avenanthramides as EGFR Inhibitors: Effects on EGF-Induced Lung Cancer Cell Growth and Migration.

Avena sativa L. is a wholegrain cereal and an important edible crop. Oats possesses high nutritional and health promoting values and contains high levels of bioactive compounds, including a group of phenolic amides, named avenanthramides (Avns), exerting antioxidant, anti-inflammatory, and anticancer activities. Epidermal growth factor receptor (EGFR) represents one of the most known oncogenes and it is frequently up-regulated or mutated in human cancers. The oncogenic effects of EGFR include enhanced cell growth, angiogenesis, and metastasis, and down-regulation or inhibition of EGFR signaling has therapeutic benefit. Front-line EGFR tyrosine kinase inhibitor therapy is the standard therapy for patients with EGFR-mutated lung cancer. However, the clinical effects of EGFR inhibition may be lost after a few months of treatment due to the onset of resistance. Here, we showed the anticancer activity of Avns, focusing on EGFR activation and signaling pathway. Lung cancer cellular models have been used to evaluate the activity of Avns on tumor growth, migration, EMT, and anoikis induced by EGF. In addition, docking and molecular dynamics simulations showed that the Avns bind with high affinity to a region in the vicinity of αC-helix and the DGF motif of EGFR, jeopardizing the target biological function. Altogether, our results reveal a new pharmacological activity of Avns as EGFR tyrosine kinase inhibitors.

Effect of Germination on the Avenanthramide Content of Oats and Their in Vitro Antisensitivity Activities.

In this study, a method, based on an ultraperformance liquid chromatography coupled with high-field quadrupole orbitrap high-resolution mass spectrometry (UHPLC-QE-HF-HRMS) platform, was established for the trace determination of three major avenanthramides (AVNs). The MS conditions for determining the AVNs were optimized, and the cracking methods of avenanthramides were analyzed. The linear range of the results and the correlation coefficient were 1−2000 µg/L and >0.996, respectively. Further, the established method was employed for the determination of the AVN contents of oats at different germination times, and the results indicated that the AVN contents of Zaohua and Bayou oats increased 19.26 and 6.09 times, respectively, after germination. The total AVN content of both oat varieties reached a maximum on the fifth day of germination (153.51 ± 4.08 and 126.30 ± 3.33 µg/g for the Zaohua and Bayou oats, respectively). Furthermore, this study investigated the antiallergic and antioxidant activities of the germinated oats via hyaluronidase inhibition and 2,2-diphenyl-1-picrylhydrazyl (DPPH)-scavenging assays. The antiallergic and DPPH-scavenging abilities of the ungerminated forms of both oat varieties were weaker. However, on the fifth day of germination, the inhibition rate of anthranilamide hyaluronidase reached 72.7% and 67.3% for the Zaohua and Bayou oat varieties, respectively. The antiallergic abilities of the oats increased significantly on the fifth day of germination in terms of their antiallergic capacities and DPPH clearance (82.67% and 77.64% for the Zaohua and Bayou oats, respectively), and the two indicators exhibited similar trends. These findings demonstrated that AVNs exhibit good antisensitivity and antioxidation properties, and the antisensitivity effect correlated positively with the AVN content.

Facile Amidation of Non-Protected Hydroxycinnamic Acids for the Synthesis of Natural Phenol Amides.

Phenol amides are bioactive compounds naturally present in many plants. This class of compounds is known for antioxidant, anti-inflammatory, and anticancer activities. To better understand the reactivity and structure-bioactivity relationships of phenol amides, a large set of structurally diverse pure compounds are needed, however purification from plants is inefficient and laborious. Existing syntheses require multiple steps, including protection of functional groups and are generally overly complicated and only suitable for specific combinations of hydroxycinnamic acid and amine. Thus, to facilitate further studies on these promising compounds, we aimed to develop a facile general synthetic route to obtain phenol amides with a wide structural diversity. The result is a protocol for straightforward one-pot synthesis of phenol amides at room temperature within 25 h using equimolar amounts of N,N'-dicyclohexylcarbodiimide (DCC), amine, hydroxycinnamic acid, and sodium bicarbonate in aqueous acetone. Eight structurally diverse phenol amides were synthesized and fully chemically characterized. The facile synthetic route described in this work is suitable for a wide variety of biologically relevant phenol amides, consisting of different hydroxycinnamic acid subunits (coumaric acid, ferulic acid, and sinapic acid) and amine subunits (agmatine, anthranilic acid, putrescine, serotonin, tyramine, and tryptamine) with yields ranging between 14% and 24%.

Oat polyphenol avenanthramide-2c confers protection from oxidative stress by regulating the Nrf2-ARE signaling pathway in PC12 cells.

Accumulating evidence has demonstrated that cellular antioxidant systems play essential roles in retarding oxidative stress-related diseases, such as Parkinson's disease. Because nuclear factor erythroid 2-related factor 2 (Nrf2) is a chief regulator of cellular antioxidant systems, small molecules with Nrf2-activating ability may be promising neuroprotective agents. Avenanthramide-2c (Aven-2c), avenanthramide-2f (Aven-2f) and avenanthramide-2p (Aven-2p) are the most abundant avenanthramides in oats, and they have been documented to possess multiple pharmacological benefits. In this work, we synthesized these three compounds and evaluated their cytoprotective effect against oxidative stress-induced PC12 cell injuries. Aven-2c displayed the best protective potency among them. Aven-2c conferred protection on PC12 cells by scavenging free radicals and activating the Nrf2-ARE signaling pathway. Pretreatment of PC12 cells with Aven-2c efficiently enhanced Nrf2 nuclear accumulation and evoked the expression of a set of cytoprotective molecules. The mechanistic study also supports that Nrf2 activation is the molecular basis for the cellular action of Aven-2c. Collectively, this study demonstrates that Aven-2c is a potent Nrf2 agonist, shedding light on the potential usage of Aven-2c in the treatment of neuroprotective diseases.

Rapid UHPLC-MS metabolite profiling and phenotypic assays reveal genotypic impacts of nitrogen supplementation in oats.

INTRODUCTION: Oats (Avena sativa L.) are a whole grain cereal recognised for their health benefits and which are cultivated largely in temperate regions providing both a source of food for humans and animals, as well as being used in cosmetics and as a potential treatment for a number of diseases. Oats are known as being a cereal source high in dietary fibre (e.g. ß-glucans), as well as being high in antioxidants, minerals and vitamins. Recently, oats have been gaining increased global attention due to their large number of beneficial health effects. Consumption of oats has been proven to lower blood LDL cholesterol levels and blood pressure, thus reducing the risk of heart disease, as well as reducing blood-sugar and insulin levels. OBJECTIVES: Oats are seen as a low input cereal. Current agricultural guidelines on nitrogen application are believed to be suboptimal and only consider the effect of nitrogen on grain yield. It is important to understand the role of both variety and of crop management in determining nutritional quality of oats. In this study the response of yield, grain quality and grain metabolites to increasing nitrogen application to levels greater than current guidelines were investigated. METHODS: Four winter oat varieties (Mascani, Tardis, Balado and Gerald) were grown in a replicated nitrogen response trial consisting of a no added nitrogen control and four added nitrogen treatments between 50 and 200 kg N ha-1 in a randomised split-plot design. Grain yield, milling quality traits, ß-glucan, total protein and oil content were assessed. The de-hulled oats (groats) were also subjected to a rapid Ultra High Performance Liquid Chromatography-Mass Spectrometry (UHPLC-MS) metabolomic screening approach. RESULTS: Application of nitrogen had a significant effect on grain yield but there was no significant difference between the response of the four varieties. Grain quality traits however displayed significant differences both between varieties and nitrogen application level. ß-glucan content significantly increased with nitrogen application. The UHPLC-MS approach has provided a rapid, sub 15 min per sample, metabolite profiling method that is repeatable and appropriate for the screening of large numbers of cereal samples. The method captured a wide range of compounds, inclusive of primary metabolites such as the amino acids, organic acids, vitamins and lipids, as well as a number of key secondary metabolites, including the avenanthramides, caffeic acid, and sinapic acid and its derivatives and was able to identify distinct metabolic phenotypes for the varieties studied. Amino acid metabolism was massively upregulated by nitrogen supplementation as were total protein levels, whilst the levels of organic acids were decreased, likely due to them acting as a carbon skeleton source. Several TCA cycle intermediates were also impacted, potentially indicating increased TCA cycle turn over, thus providing the plant with a source of energy and reductant power to aid elevated nitrogen assimilation. Elevated nitrogen availability was also directed towards the increased production of nitrogen containing phospholipids. A number of both positive and negative impacts on the metabolism of phenolic compounds that have influence upon the health beneficial value of oats and their products were also observed. CONCLUSIONS: Although the developed method has broad applicability as a rapid screening method or a rapid metabolite profiling method and in this study has provided valuable metabolic insights, it still must be considered that much greater confidence in metabolite identification, as well as quantitative precision, will be gained by the application of higher resolution chromatography methods, although at a large expense to sample throughput. Follow up studies will apply higher resolution GC (gas chromatography) and LC (reversed phase and HILIC) approaches, oats will be also analysed from across multiple growth locations and growth seasons, effectively providing a cross validation for the results obtained within this preliminary study. It will also be fascinating to perform more controlled experiments with sampling of green tissues, as well as oat grains, throughout the plants and grains development, to reveal greater insight of carbon and nitrogen metabolism balance, as well as resource partitioning into lipid and secondary metabolism.

Overview of the Anticancer Profile of Avenanthramides from Oat.

Cancer represents one of the leading causes of death worldwide. Progresses in treatment of cancer have continued at a rapid pace. However, undesirable side effects and drug resistance remain major challenges for therapeutic success. Natural products represent a valuable starting point to develop new anticancer strategies. Polyphenols, well-known as antioxidant, exert anticancer effects through the modulation of multiple pathways and mechanisms. Oat (Avena sativa L., Poaceae) is a unique source of avenanthramides (AVAs), a group of polyphenolic alkaloids, considered as its signature compounds. The present review aims to offer a comprehensive and critical perspective on the chemopreventive and chemotherapeutic potential of AVAs. AVAs prevent cancer mainly by blocking reactive species. Moreover, they exhibit potential therapeutic activity through the modulation of different pathways including the activation of apoptosis and senescence, the block of cell proliferation, and the inhibition of epithelial mesenchymal transition and metastatization. AVAs are promising chemopreventive and anticancer phytochemicals, which need further clinical trials and toxicological studies to define their efficacy in preventing and reducing the burden of cancer diseases.

Synthesis of avenanthramides using engineered Escherichia coli.

BACKGROUND: Hydroxycinnamoyl anthranilates, also known as avenanthramides (avns), are a group of phenolic alkaloids with anti-inflammatory, antioxidant, anti-itch, anti-irritant, and antiatherogenic activities. Some avenanthramides (avn A-H and avn K) are conjugates of hydroxycinnamic acids (HC), including p-coumaric acid, caffeic acid, and ferulic acid, and anthranilate derivatives, including anthranilate, 4-hydroxyanthranilate, and 5-hydroxyanthranilate. Avns are primarily found in oat grain, in which they were originally designated as phytoalexins. Knowledge of the avns biosynthesis pathway has now made it possible to synthesize avns through a genetic engineering strategy, which would help to further elucidate their properties and exploit their beneficial biological activities. The aim of the present study was to synthesize natural avns in Escherichia coli to serve as a valuable resource. RESULTS: We synthesized nine avns in E. coli. We first synthesized avn D from glucose in E. coli harboring tyrosine ammonia lyase (TAL), 4-coumarate:coenzyme A ligase (4CL), anthranilate N-hydroxycinnamoyl/benzoyltransferase (HCBT), and anthranilate synthase (trpEG). A trpD deletion mutant was used to increase the amount of anthranilate in E. coli. After optimizing the incubation temperature and cell density, approximately 317.2 mg/L of avn D was synthesized. Avn E and avn F were then synthesized from avn D, using either E. coli harboring HpaBC and SOMT9 or E. coli harboring HapBC alone, respectively. Avn A and avn G were synthesized by feeding 5-hydroxyanthranilate or 4-hydroxyanthranilate to E. coli harboring TAL, 4CL, and HCBT. Avn B, avn C, avn H, and avn K were synthesized from avn A or avn G, using the same approach employed for the synthesis of avn E and avn F from avn D. CONCLUSIONS: Using different HCs, nine avns were synthesized, three of which (avn D, avn E, and avn F) were synthesized from glucose in E. coli. These diverse avns provide a strategy to synthesize both natural and unnatural avns, setting a foundation for exploring the biological activities of diverse avns.

Antiproliferative activity of vitexin-2-O-xyloside and avenanthramides on CaCo-2 and HepG2 cancer cells occurs through apoptosis induction and reduction of pro-survival mechanisms.

PURPOSE: CaCo-2 colon cancer cells and HepG2 liver cancer cells represent two malignant cell lines, which show a high resistance to apoptosis induced by the conventional anticancer drugs. Vitexin-2-O-xyloside (XVX) and avenanthramides (AVNs) are naturally occurring dietary agents from Beta vulgaris var. cicla L. and Avena sativa L., respectively. The aim of this work was to evaluate the antiproliferative effects and the reduction of the pro-survival mechanisms exerted by XVX and AVNs, used individually and in combination, in CaCo-2 and HepG2 cancer cells. METHODS: XVX and AVNs were isolated by liquid chromatography and characterized by HPLC-PDA-MS. The XVX and AVN antiproliferative effects were evaluated through sulforhodamine B method, while their pro-apoptotic effects through caspase activity assays. RTqPCR was used to investigate the modulation of the pro-survival factors baculoviral inhibitor of apoptosis repeat-containing 5 (BIRC5), hypoxia inducible factor 1 A (HIF1A), and vascular endothelial growth factor A (VEGFA). Cellular antioxidant activity (CAA) was investigated by means of DCFH-DA assay, whereas chemical antioxidant capacity was evaluated by the ORAC method. RESULTS: XVX and AVNs, both individually and in combination, inhibited the proliferation of CaCo-2 and HepG2 cancer cells, through activation of caspases 9, 8, and 3. XVX and AVNs downregulated the pro-survival genes BIRC5, HIF1A, and VEGFA. The CAA assay showed that AVNs exhibited strong antioxidant activity inside both CaCo-2 and HepG2 cells. CONCLUSIONS: The antiproliferative activity of the XVX + AVNs mixture represents an innovative treatment, which is effective against two types of cancer cells characterized by high resistance to the conventional anticancer drugs.

Avenanthramide-C reduces the viability of MDA-MB-231 breast cancer cells through an apoptotic mechanism.

BACKGROUND: Avenanthramides (AVN) are a relatively unstudied family of phytochemicals that could be novel chemotherapeutics. These compounds, found in oats, are non-toxic to healthy cells and have been shown to reduce viability of human colon and liver cancers in vitro. However, these studies do not elucidate a molecular mechanism for individual AVN. In this study we aim to see the effects of AVN on MDA-MB-231 breast cancer cells. METHODS: An MTT assay was used to determine cell viability. Staining and analysis with a flow cytometer was used to identify cell cycle progression and apoptosis. FloJo software was used to analyze the cytometric data. In all experiments, statistical significance was determined by a two-tailed t test. RESULTS: This study demonstrates that AVN-A, B, and C individually reduce viability in the MDA-MB-231 breast cancer cell line. AVN-C has the most potent decrease in tumor cell viability, decreasing viable cells to below 25% at 400 µM when compared to control after 96 h. We demonstrate that treatment with AVN-C causes DNA fragmentation and accumulation of over 90% of cells into a sub G1 cell cycle population. Further, we conclude that AVN-C treated cells activate apoptosis because 97% of treated cells stain positive for annexin V while 91% have caspase-3/7 activity, a late marker of apoptosis. CONCLUSIONS: Breast cancer cells treated with AVN-C have a decrease in cell viability, an increase in the sub G1 population, and stain positive for both annexin V and caspase activity, indicating that AVN-C induces apoptosis in breast cancer cells. These compounds may be able to act as chemotherapeutics as demonstrated through future in vivo studies.

The effect of mechanical processing on avenanthramide and phenol levels in two organically grown Italian oat cultivars.

Avenanthramides (AVNs), free and bound phenols and their antioxidant capacities (ORAC) were evaluated in two Avena sativa L. cultivars, Donata and Flavia. The cultivars (cvs.) were grown in loamy and medium texture soils and assessed after industrial dehulling and milling. Total dietary fiber, ß-glucan, starch and proteins were also evaluated. Cv. Donata showed 2.8 fold higher AVN storage as compared to cv. Flavia, which was linked with genotype. The accumulation of AVN content was also influenced by the texture of the soil. Dehulling resulted in a 75 and 37% AVN decrease in cv. Donata and Flavia, respectively. The dehulled grains of cv. Donata showed 40% reduction in free phenolic content, whereas the dehulled grains of both cvs. showed 67% reduction in bound phenols. Milling affected the bound phenolics and their antioxidant capacity. Cv. Flavia showed 1.3 fold higher ß-glucan than that of cv. Donata. Total dietary fiber was reduced by 50 and 12% after dehulling and milling, respectively, while marginal changes in proteins were observed after milling. The results suggest that the choice of genotype and the kind of dehulling processes that are employed are essential considerations in the production of oat-based products with high AVN content and extra health benefits.

Oat-based milk alternatives: the influence of physical and chemical properties on the sensory profile.

Introduction: Oat-based milk alternatives (OMAs) have become increasingly popular, perhaps due to their low allergenicity and preferred sensory attributes when compared to other milk alternatives. They may also provide health benefits from unique compounds; avenanthramides, avenacosides, and the dietary fibre beta-glucan. This has led to a variety of commercial options becoming available. Being a fairly new product, in comparison to other plant-based milk alternatives (PBMAs), means little research has been undertaken on the sensory profile, and how it is influenced by the physical and chemical properties. Methods: This study investigated the sensory, physical and chemical profiles of current commercially available OMAs, that varied in fortification, use of stabilisers, and oat content. The volatile compounds and their respective aromas were analysed using solid phase microextraction followed by gas chromatography mass spectrometry (GC-MS) and gas chromatography-olfactometry (GC-O). Liquid chromatography mass spectrometry (LC-MS) was used for identification of avenanthramides and avenacosides. Particle size and polydispersity index (PDI) were analysed using a Mastersizer and Zetasizer, respectively, with colour analysis carried out using a colourimeter, and viscosity measurements using a rheometer. Descriptive sensory profiling was used to assess the impact on the sensory characteristics of the different samples and the sensory data acquired were correlated with the instrumental data. Results: Samples with smaller particle size appeared whiter-both instrumentally and perceptually. The only clear plastic packaged product differed substantially in volatile profile from all other products, with a higher abundance of many volatile compounds, and high overall perceived aroma. Avenanthramides and avenacosides were present in all samples, but differed significantly in abundance between them. Discussion: The results suggested smaller particle size leads to whiter colour, whilst differences in processing and packaging may contribute to significant differences in aroma. Astringency did not differ significantly between samples, suggesting that the variation in the concentrations of avenacosides and avenanthramides were below noticeable differences.

Fluorescent Molecularly Imprinted Polymers Loaded with Avenanthramides for Inhibition of Advanced Glycation End Products.

Encapsulating bioactive avenanthramides (AVAs) in carriers to respond to the environmental changes of food thermal processing allows the controlled release of AVAs for the effective inhibition of biohazards. In this study, fluorescent molecular imprinted polymers (FMIPs) loaded with AVAs were prepared by reverse microemulsion. The fluorescent signal was generated by carbon dots (CDs), which were derived from oat bran to determine the load of AVAs. The FMIPs were uniformly spherical in appearance and demonstrated favorable properties, such as thermal stability, protection of AVAs against photodegradation, high encapsulation efficiency, and effective scavenging of free radicals. After consideration of the different kinetics models, the release of AVAs from the FMIPs matched the Weibull model and followed a Fickian diffusion mechanism. The FMIPs exhibited good inhibition of pyrraline in a simulated casein-ribose system and in milk samples, indicating the release of AVAs could inhibit the generation of pyrraline.

The Adapted POM Analysis of Avenanthramides In Silico.

POM analysis and related approaches are significant tools based on calculating various physico-chemical properties and predicting biological activity, ADME parameters, and toxicity of a molecule. These methods are used to evaluate a molecule's potential to become a drug candidate. Avenanthramides (AVNs) are promising secondary metabolites specific to Avena spp. (oat). They comprise the amides of anthranilic acid linked to various polyphenolic acids with or without post-condensation molecule transformation. These natural compounds have been reported to exert numerous biological effects, including antioxidant, anti-inflammatory, hepatoprotective, antiatherogenic, and antiproliferative properties. To date, almost 50 various AVNs have been identified. We performed a modified POM analysis of 42 AVNs using MOLINSPIRATION, SWISSADME, and OSIRIS software. The evaluation of primary in silico parameters revealed significant differences among individual AVNs, highlighting the most promising candidates. These preliminary results may help coordinate and initiate other research projects focused on particular AVNs, especially those with predicted bioactivity, low toxicity, optimal ADME parameters, and promising perspectives.

The Progress of Nomenclature, Structure, Metabolism, and Bioactivities of Oat Novel Phytochemical: Avenanthramides.

Oats are among the most commonly consumed whole grains and are widely grown worldwide, and they contain numerous nutrients, including proteins, lipids, vitamins, minerals, ß-glucan, and unique phytochemical polyphenol avenanthramides (Avns). Recent studies have indicated that Avns play essential roles in mediating the health benefits of oats. This review systemically summarized the nomenclature and structures of Avns, effect of germination on promoting Avns production, and in vivo metabolites produced after Avns consumption. The classical functions and novel potential bioactivities of Avns were further elucidated. The classical functions of Avns in cancer prevention, antioxidative response, anti-inflammatory reaction, and maintaining muscle health were expounded, and the internal mechanisms of these functions were analyzed. The potential novel bioactivities of Avns in modulating gut microbiota, alleviating obesity, and preventing chronic diseases, such as atherosclerosis and osteoporosis, were further revealed. This review may provide new prospects and directions for the development and utilization of oat Avns.

Simplified Analysis and Expanded Profiles of Avenanthramides in Oat Grains.

Uniquely, oats contain avenanthramides (AVAs), a group of phenolic alkaloids, exhibiting many health benefits. AVA analysis involves extraction with alcohol-based solvents and HPLC separation with UV and/or mass spectrometer detectors. There are many reported methods to extract AVAs. Almost all entail multiple extractions. The whole procedure is time- and labor-intensive. Furthermore, most quantifications are limited to three common AVAs (2f, 2p, 2c). The present study compared three extraction methods (all at 50 °C) for their effects on AVA concentrations and composition (% relative to total AVA) of oat grains. These included triplicate extractions with 80% ethanol containing 10 mM phosphate buffer (pH 2.0) (A), triplicate extractions with 80% ethanol (B), and a single extraction with 80% ethanol (C), while keeping solid/total solvent ratio at 1/60 (g/mL) and total extraction time of 60 min. Results showed that 80% buffered ethanol gave significantly lower AVA contents than 80% ethanol, while single and triplicate extractions with 80% ethanol produced the same extractability. However, the extraction method had no effect on AVA composition. Using 0.25 g sample size instead of 0.5 g saved extractants by half, without affecting AVA measurements. Consequently, a simplified method of extraction was developed, featuring Method C. The present study also expanded profiling individual AVAs beyond AVA 2c, 2p and 2f. Other AVAs identified and semi-quantified included 5p, 4p, 3f/4f, and 2pd. The simplified analysis was validated by measuring 16 selected oat grain samples. Some of these grains had relatively high contents of 4p, 3f/4f and 2pd, which have been considered minor AVAs previously.

Inhibitory Mechanism of Advanced Glycation End-Product Formation by Avenanthramides Derived from Oats through Scavenging the Intermediates.

As a special polyphenolic compound in oats, the physiological function of oat avenanthramides (AVAs) drives a variety of biological activities, and plays an important role in the prevention and treatment of common chronic diseases. In this study, the optimum extraction conditions and structural identification of AVAs from oats was studied. The inhibitory effect of AVAs from oats on advanced glycation end-products (AGEs) in a glucose-casein simulation system was evaluated, and this revealed dose-dependent inhibitory effects. The trapping capacity of AVAs to the α-dicarbonyl compounds of AGE intermediate products was determined by HPLC-MS/MS, and the results indicate that AVA 2c, AVA 2p, and AVA 2f exhibited the ability to capture α-dicarbonyl compounds. More importantly, AVA 2f was found to be more efficient than AVA 2p at inhibiting superoxide anion radical (O2-), hydroxyl radical (OH), and singlet oxygen (1O2) radical generation, which may be the main reason that AVA 2f was more efficient than AVA 2p in AGE inhibition. Thus, this research presents a promising application of AVAs from oats in inhibiting the food-borne AGEs formed in food processing.

Polyphenols and avenanthramides extracted from oat (Avena sativa L.) grains and sprouts modulate genes involved in glucose and lipid metabolisms in 3T3 L1 adipocytes.

This study aimed to evaluate the effect of polyphenol (PE) and avenanthramide (AE) extracts from oat grains (OG) and sprouts (OS) on genes related to glucose and lipid metabolisms in 3T3 L1 adipocytes. The AE-OS exerted the greatest effect on genes involved in glucose metabolism, increasing Glut4, Irs1, and Pi3k expression by 3.0- to 3.9-fold. Conversely, the PE-OS exerted the greatest effect on genes involved in lipid metabolism, decreasing Fasn and Acaca expression by 0.2- to 0.3-fold, and increasing Cpt1a and Acadm expression by 2.7- to 3.0-fold. These effects were mainly related to their high content of avenanthramides A (2p), B (2f), and C (2c), quercetin 3-O-rutinoside, kaempferol, sinapoylquinic acid, and apigenin and luteolin derivatives according to the chemometric analysis. In conclusion, this study demonstrated that oat sprouts extract exerts a greater effect than oat grains on the regulation of genes involved in glucose and lipid metabolisms in adipocytes. PRACTICAL APPLICATIONS: This study demonstrates that polyphenols and avenanthramides extracted from oat (Avena sativa L.) grains and sprouts modulate key genes involved in glucose and lipid metabolisms in adipocytes and that oat sprouts exert a greatest health beneficial effect than oat grains due to their higher content of bioactive compounds. In addition, the chemometric analysis identified the bioactive compounds that can be associated with the beneficial effects of oat grains and sprouts, which can be further used for the identification of oat varieties and oat-derived products with high content of these bioactive compounds and, thus, with high nutraceutical potential.

Characterization and antioxidant activity of avenanthramides from selected oat lines developed by mutagenesis technique.

From a mutagenized oat population, produced by ethyl methanesulfonate mutagenesis, hulled grains from 17 lines with elevated avenanthramide (AVN) content were selected and their AVN structures, concentrations and antioxidant potentials were determined by HPLC-MS2 and HPLC equipped with an on-line ABTS+ antioxidant detection system. The data obtained showed qualitative and quantitative differences in the synthesis of AVNs in the different lines, with a total AVN concentration up to 227.5 µg/g oat seed flour in the highest line, compared with 78.2 µg/g seed in the commercial line, SW Belinda. In total, 25 different AVNs were identified with avenanthramide B structures being among the most abundant, and AVN C structures having the highest antioxidant activity. The findings indicate the potential of oat mutagenesis in combination with a high precision biochemical selection method for the generation of stable mutagenized lines with a high concentration of total and/or individual AVNs in the oat seed grain.

Avenanthramides: Unique Bioactive Substances of Oat Grain in the Context of Cultivar, Cropping System, Weather Conditions and Other Grain Parameters.

Our study was focused on the evaluation of the content of a wider spectrum of eight avenanthramides (AVNs) as unique components of oat grain under the effects of four selected factors (cultivar, locality, cropping system, and year). The weather effects on changes in the AVN content and their relationship to other important parameters of oat grain were further evaluated in more detail. A sensitive UHPLC system coupled with a QExactive Orbitrap mass spectrometer was used for AVN quantification. AVNs confirmed a high variability (RDS = 72.7-113.5%), which was dominantly influenced by the locality and year factors. While most AVN types confirmed mutually high correlations (r = 0.7-0.9), their correlations with the other 10 grain parameters were lower (r ≤ 0.48). Their significant correlations (0.27-0.46) with ß-D-glucan could be used in perspective in breeding programs for the synergetic increase of both parameters. PCA analysis and Spearman correlations based on individual cultivars confirmed a significant effect of June and July precipitation on the increase of Σ AVNs. However, the results also indicated that higher precipitation can generate favorable conditions for related factors, such as preharvest sprouting evoking a direct increase of AVNs synthesis in oat grain.