Cereal Proteins Research Articles

Whole cereal protein-based Pickering emulsions prepared by zein-gliadin complex particles

Zein and gliadin are both cereal proteins, indicating that they are highly compatible. In this study, gliadin was firstly added to zein to achieve different zein/gliadin ratios in aqueous ethanol solution, then we synthesized zein-gliadin complex particles (ZGCP) via an antisolvent approach and investigated their efficiency as stabilizers to fabricate Pickering emulsions. The addition of gliadin affected the self-assembly, aggregation and wettability of ZGCP. Scanning electron microscope (SEM) showed that spherical zein particles were transformed into aggregation of nano-sized ZGCP, and FTIR results indicated that weak flocculation behavior of ZGCP involved in intermolecular β-sheet interactions. Furthermore, relatively hydrophilic gliadin was used to modify the hydrophobic property of zein, resulting in formation of near-neutral wettability of ZGCP. Compared with zein or gliadin, ZGCP tend to form a dense interfacial layer around droplet and smaller emulsion droplet size, endowing ZGCP-stabilized emulsions with higher viscoelastic attributes and much better stability against coalescence. This finding may develop theoretical and technical supports for the utilization of the whole cereal protein-based Pickering emulsions as novel natural ingredients in food industry.

Variable Immunogenic Potential of Wheat: Prospective for Selection of Innocuous Varieties for Celiac Disease Patients via in vitro Approach.

Celiac Disease (CD) is a multifactorial, autoimmune enteropathy activated by cereal proteins in genetically predisposed individuals carrying HLA DQ2/8 genes. A heterogenous gene combination of the cereal prolamins is documented in different wheat genotypes, which is suggestive of their variable immunogenic potential. In the current study, four wheat varieties (C591, C273, 9D, and K78) identified via in silico analysis were analyzed for immunogenicity by measuring T-cell proliferation rate and levels of inflammatory cytokines (Interferon-γ and Tumor Necrosis Factor-α). Peripheral Blood Mononuclear Cells and biopsy derived T-cell lines isolated from four CD patients in complete remission and two controls were stimulated and cultured in the presence of tissue transglutaminase activated pepsin-trypsin (PT) digest of total gliadin extract from test varieties. The immunogenicity was compared with PBW 621, one of the widely cultivated wheat varieties. Phytohaemagglutinin-p was taken as positive control, along with unstimulated cells as negative control. Rate of cell proliferation (0.318, 0.482; 0.369, 0.337), concentration of IFN- γ (107.4, 99.2; 117.9, 99.7 pg/ml), and TNF- α (453.8, 514.2; 463.8, 514.2 pg/ml) was minimum in cultures supplemented with wheat antigen from C273, when compared with other test varieties and unstimulated cells. Significant difference in toxicity levels among different wheat genotypes to stimulate celiac mucosal T-cells and PBMC's was observed; where C273 manifested least immunogenic response amongst the test varieties analyzed.

Cereal proteins in nanotechnology: formulation of encapsulation and delivery systems

Cereals are an abundant and relatively inexpensive source of plant-based proteins that play critical nutritional and functional roles in foods. However, their poor solubility in aqueous solutions limits their application in many food products. A number of strategies are, therefore, being developed to overcome this limitation. Physical, chemical, and enzymatic modification of cereals protein can be used to improve their solubility and functionality. For instance, modified cereal proteins can be used as emulsifiers to stabilize lipid nanodroplets. The low water-solubility of cereal proteins is an advantage in the fabrication of protein nanoparticles created using antisolvent precipitation methods. These nanoparticles can be used to encapsulate bioactive agents or to stabilize emulsions through a Pickering mechanism.

Cereal protein-based nanoparticles as agents stabilizing air–water and oil–water interfaces in food systems

There has been a recent surge of interest in the use of food-grade nanoparticles (NPs) for stabilizing food foams and emulsions. Cereal proteins are a promising raw material class to produce such NPs. Studies thus far have focused mostly on wheat gliadin and maize zein-based NPs. The former are effective interfacial stabilizing agents, while the latter due to their high hydrophobicity generally result in poor interfacial stability. Several strategies to modify the surface properties of wheat gliadin and maize zein NPs have been followed. In many instances, this resulted in improved foam or emulsion stability. Nonetheless, future efforts should be undertaken to gain fundamental insights in the interfacial behavior of NPs, to further explore NP surface modification strategies, and to validate the use of NPs in actual food systems.

The impact of alkaline conditions on storage proteins of cereals and pseudo-cereals

Alkaline conditions have different impacts on proteins. Firstly, they can impact on the protein structure which then influences their solubility and other techno-functional properties. The former generally increases with the difference between pH and pI, and generally increases protein extraction yield. Secondly, chemical reactions can lead to negative nutritional effects by loss of essential amino acids such as lysine. Chemical reactions can also lead to additional crosslinks and to (un)desired color and flavor components. (Pseudo)cereal proteins are often exposed to alkaline conditions, for example, during pretzel production and extraction of rice and pseudo-cereal protein.

Immunoreactive cereal proteins in wheat allergy, non-celiac gluten/wheat sensitivity (NCGS) and celiac disease

Wheat is one of the major sources of nutrients for the world population, but its consumption may cause inflammatory immune reactions such as wheat allergy, celiac disease or non-celiac gluten/wheat sensitivity in predisposed individuals. Various wheat proteins including gluten and non-gluten proteins have been identified as triggering factors. With mounting evidence pointing to an increasing prevalence of adverse reactions to wheat, recent research efforts aim at elucidating the underlying causes. Multidisciplinary approaches toward a better understanding of the complex interactions between immunoreactive components in cereals and the human gastrointestinal and immune systems, including the contribution of gut microbiota, will be key to devising novel preventive and therapeutic strategies for at-risk individuals.

Enzyme-assisted development of biofunctional polyphenol-enriched buckwheat protein: physicochemical properties, in vitro digestibility, and antioxidant activity.

During the last decade buckwheat was reported to have positive health effects. The present study investigated a high-polyphenol buckwheat protein (Fagopyrum esculentum Moench) prepared by enzyme-assisted processing, together with its physicochemical properties, in vitro digestibility, and antioxidant activity. Buckwheat protein prepared from the synergistic enzymatic action of α-amylase and amyloglucosidase (E-BWP) had much higher polyphenol content than buckwheat protein prepared by isoelectric precipitation (I-BWP) or salt extraction (S-BWP). Rutin degraded during the process, giving quercetin. The protein constituents and amino acid composition of E-BWP were very similar to those of native buckwheat and were able to meet the WHO/FAO requirements for both children and adults. During in vitro digestion, E-BWP showed anti-digestive behavior with a nitrogen release that was lower than that of I-BWP or S-BWP. The positive effect of the polyphenol content of E-BWP resulted in a higher 1,1-diphenyl-2-picrylhydrazyl (DPPH) content and greater reducing activity. Buckwheat protein with high polyphenol content was successfully developed by enzyme-assisted processing. It had a well-balanced amino acid profile, antidigestive behavior, and high antioxidant activities. The results suggest that enzyme-assisted processing is promising in the production of polyphenol-enriched cereal protein, contributing higher functionality with good nutritional and antioxidant properties. © 2018 Society of Chemical Industry.

Porovnanie vybraných genotypov pšenice, ovsa a pohánky na proteomickej úrovni

Cereals and pseudocereals are one of the most important crops. In recent years, number of people suffering from various food allergies and intolerances has grown and exactly, cereal proteins are the most common food allergen group. Pseudocereals are suitable alternative that have positive effect on human organism and do not induce allergies. The aim of this work was to analyze grains of winter wheat (Triticum aestivum L.), oat (Avena sativa) and buckwheat (Fagopyrum esculentum Moench.) and compare protein composition from the point of nutritional and technological quality. Wheat had the best technological quality based on the highest content of gluten proteins (63.42%) and HMW-GS (15.21%). Potentially allergenic wheat proteins with molecular weight in the range of 25 to 45 kDa gave strong signal in Western blot. Oat, due to the higher coefficient of nutritional quality (373.89%) and content of albumins and globulins (49.29%), had worse technological and better nutritional quality than wheat. The best nutritional quality was based on the highest coefficient of nutritional quality (6005.93%), content of albumins and globulins (60.81%) and essential amino acids in buckwheat. Buckwheat is suitable crop for gluten-free diet because it contained only 1.35% of prolamins that were not detected by Western blot analysis.

Конструирование рекомбинантных продуцентов ферментных препаратов для кормопроизводства с помощью экспрессионной системы на основе гриба Penicillium verruculosum

Using the expression system based on the recipient strain Penicillium verruculosum 537 (ΔniaD) and promoter of the cellobiohydrolase 1 gene, producing strains were created. That made it possible to obtain new enzyme preparations (EP) for feed production, which are characterized by a high content of exodepolymerases (endo-β-l,4-glucanases and endo-β-l,4-xylanase), degrading non-starch polysaccharides (NSP) of grain cereals used in feed. The content of the endodepolymerases in new EP was 44-68% of the total protein content (compared with 15% in the case of the recipient strain), the content of exodepolymerases (cellobiohydrolases) fell to 8-26% compared to 60% in the case of the EP, obtained using recipient strain. New EP had high specific activity of endodepolymerases, which is 1.8-4.6 times higher than for the EP obtained with the recipient strain; the specific activity of exodepolymerases decreased in 2,0-3,7 times. New EP showed endoglucanase and xylanase activity in a wide range of pH and temperature, including the physiological value of these parameters (pH 3 and 7 and Г=37-38 °C); characterized by a high stability of endodepolymerases under the influence of digestive proteases (pepsin and trypsin), as well as the stability of ndodepolymerases under granulation conditions of animal feed (80 °C). The xylanase of the new EP was not inhibited by the inhibitors of cereal proteins. An experimental batch of new EP was manufactured by the enzyme-producing plant "Agroferment" and tested in broiler and pig feeding. The input of new commercial EP into broiler diets has had a positive impact on the safety of chickens, increasing their live weight and reducing feed cost. The supplementation of commercial EP into pig ration also led to positive results during feeding, contributed to the increase in the average daily growth of pig biomass, reduced feed consumption, protein and metabolic energy per unit of live weight gain, improved digestibility and assimilability of feed nutrients, especially of raw fiber and NSP. Feed enzyme preparation, xylanase, endoglucanase, non-starch polysaccharides, Penicillium verruculosum Funding-The work was done with the financial support of the Ministry of Science and Higher Education (State Task 0104-2019-0009).

Gluten Immunogenic Peptides as Standard for the Evaluation of Potential Harmful Prolamin Content in Food and Human Specimen.

Gluten is a complex mixture of storage proteins in cereals like wheat, barley, and rye. Prolamins are the main components of gluten. Their high content in proline and glutamine makes them water-insoluble and difficult to digest in the gastrointestinal tract. Partial digestion generates peptide sequences which trigger immune responses in celiac and gluten-sensitive patients. Gluten detection in food is challenging because of the diversity, in various food matrices, of protein proportions or modifications and the huge number of immunogenic sequences with differential potential immunoactivity. Attempts to develop standard reference materials have been unsuccessful. Recent studies have reported the detection of a limited number of dominant Gluten Immunogenic Peptides (GIP) that share similarities to epitopes presented in the α-gliadin 33-mer, which showed to be highly proteolytic resistant and is considered to be the most immunodominant peptide within gluten in celiac disease (CD). GIP were detectable and quantifiable in very different kind of difficult to analyze food, revealing the potential immunogenicity by detecting T-cell activity of celiac patients. But GIP were also found in stool and urine of celiac patients on a supposedly gluten-free diet (GFD), showing the capacity to resist and be absorbed and excreted from the body, providing the first simple and objective means to assess adherence to the GFD. Methods to specifically and sensitively detect the most active GIP in food and biological fluids are rational candidates may use similar analytical standard references for determination of the immunopathological risk of gluten exposure in gluten-related diseases.

Growth and instability in agriculture - A case of pulses production in India

Pulses are an important commodity group of crops that provide high quality protein complementing cereal proteins for pre-dominantly substantial vegetarian population of the country. In India, pulses can be produced with a minimum use of resources and hence, it becomes less costly even than animal protein. Although this crop group is more important from the nutritional point of view, there has not been any significant development both in area and production during the last 50 years. In recent years, commercial and major millets are replacing the area under pulses and has led to several issues requiring attention of all the concerned. The study is based on time-series data for 60 years (1950-51 to 2009-10) on area, production and productivity of pulses in India. Exponential growth rate, instability index and Hazel’s decomposition analysis was used and it was found that, the growth rate of area under pulses during the study period was non-significant, while production growth rate was just 0.59. The variation around the trend (instability index 10.56) and the variation around mean (CV 14.45 %) was more in case of production. Increase in mean yield accounted for 99 per cent of the increase in production. Variance in production of pulses was mainly due to residual factors.

Occurrence of serum antibodies against wheat alpha-amylase inhibitor 0.19 in celiac disease.

The alcohol-soluble fraction of wheat gluten (gliadins) induces in genetically susceptible individuals immunologically mediated celiac disease (CLD). However, gliadins and related cereal proteins are not unique foodstuff targets of CLD patients´ immune system. Non-gluten wheat alpha-amylase inhibitor 0.19 (AAI 0.19) has been found to be capable of activating human monocyte-derived dendritic cells and inducing pro-inflammatory status in intestinal mucosa of patients with celiac disease (CLD). The possible contribution of this reactivity in incomplete remission of CLD patients on a gluten-free diet (GFD) is matter of contention. In an attempt to characterize the antigenicity of AAI 0.19 in patients with active CLD, patients on a GFD and healthy controls we developed ELISA employing wheat recombinant AAI 0.19. Using this test we revealed a significant (P<0.001) elevation of IgA anti-AAI 0.19 antibodies (Ab) in patients with active CLD (12 out of 30 patients were seropositive) but also in CLD patients on a GFD (15/46), in contrast to healthy controls (2/59). Anti-AAI 0.19 IgG Ab levels were increased (P<0.001) only in patients with active CLD (14/30) in contrast to the controls. Interestingly, the levels of anti-AAI 0.19 IgG Ab were decreased in CLD patients on a GFD (P<0.001, 1/46) compared to the controls (1/59). Notably, 20 out of 30 patients with active CLD were positive either for IgA or for IgG anti-AAI 0.19 Ab. Thus, the majority of CLD patients developed a robust IgA and IgG Ab response against AAI 0.19. These findings may contribute to the broadening of the knowledge about CLD pathogenesis.

Rice‐Endosperm and Rice‐Bran Proteins: A Review

AbstractRice protein is the most preferred protein among all other cereal proteins in terms of its amino acid structure, and it contains the second highest content of lysine, the limiting amino acid in cereals. Processing rough rice into edible rice yields different by‐products including rice bran. Rice‐endosperm and rice‐bran proteins are known to be hypoallergenic, nutritious, and healthful, which gives them high potential in the food‐ingredient industry, especially in gluten‐free preparations and in infant formulas. Multiple methods can be used for the extraction of rice‐endosperm and rice‐bran proteins, including chemical and enzymatic extraction methods. The functional properties, including emulsification, foaming, gelation, and water and oil absorption, of rice‐endosperm and rice‐bran proteins require enhancement before utilizing them in food applications because of their low solubilities, which affects the other functionalities. Enhancement methods include manipulating pH and salt concentration of extracting solvents and selecting specific extraction procedures. Rice‐endosperm and rice‐bran peptides are known to exhibit different bioactivities. These peptides can be generated by different hydrolysis approaches, including the enzymatic, chemical, or microbial assays. Some of the confirmed bioactivities of rice‐endosperm and rice‐bran peptides are the antioxidant, antihypertensive, anticancer, anti‐inflammatory, and antimicrobial activities. These bioactivities increase the potential of rice‐endosperm and rice‐bran proteins to be utilized in food applications, as natural preservatives, or in pharmaceutical applications.

Further analysis of barley MORC1 using a highly efficient RNA-guided Cas9 gene-editing system.

SummaryMicrorchidia (MORC) proteins comprise a family of proteins that have been identified in prokaryotes and eukaryotes. They are defined by two hallmark domains: a GHKL‐type ATPase and an S5‐fold. In plants, MORC proteins were first discovered in a genetic screen for Arabidopsis thaliana mutants compromised for resistance to a viral pathogen. Subsequent studies expanded their role in plant immunity and revealed their involvement in gene silencing and genome stabilization. Little is known about the role of MORC proteins of cereals, especially because knockout (KO) mutants were not available and assessment of loss of function relied only on RNAi strategies, which were arguable, given that MORC proteins in itself are influencing gene silencing. Here, we used a Streptococcus pyogenes Cas9 (SpCas9)‐mediated KO strategy to functionally study HvMORC1, one of the current seven MORC members of barley. Using a novel barley RNA Pol III‐dependent U3 small nuclear RNA (snRNA) promoter to drive expression of the synthetic single guide RNA (sgRNA), we achieved a very high mutation frequency in HvMORC1. High frequencies of mutations were detectable by target sequencing in the callus, the T0 generation (77%) and T1 generation (70%–100%), which constitutes an important improvement of the gene‐editing technology in cereals. Corroborating and extending earlier findings, SpCas9‐edited hvmorc1‐ KO barley, in clear contrast to Arabidopsis atmorc1 mutants, had a distinct phenotype of increased disease resistance to fungal pathogens, while morc1 mutants of either plant showed de‐repressed expression of transposable elements (TEs), substantiating that plant MORC proteins contribute to genome stabilization in monocotyledonous and dicotyledonous plants.

TaAAP6-3B, a regulator of grain protein content selected during wheat improvement

BackgroundThe content of grain protein (GPC) in cereals is an important part of total protein in human food. Exploring and utilizing new GPC genes is one of the most effective approaches for wheat quality breeding.ResultsThree homoeologues of TaAAP6(−3A, 3B, 3D)were cloned by homology cloning from OsAAP6.Temporal and spatial expression analysis showed that TaAAP6homoeologues were preferentially expressed in developing grains, and TaAAP6-3B may play a major role in regulating GPC in wheat. Association analysis indicated thatTaAAP6-3B-I is significantly correlated with higher GPC than that of TaAAP6-3B-II for 115 wheat lines in all five environments. TaAAP6-3B-I, the favored allele of TaAAP6-3B, was preferentially expressed in preliminary developing grain stage. Two functional markers were developed to discriminate 197F2populations and the result showed that TaAAP6-3B-I (high-protein content) was completely dominant. Two cis-regulatory elements appear to be associated with high GPC were found in the 5’UTR of TaAAP6-3B-I.The change of the TaAAP6-3B locus types indicated that the gene was subjected to selection pressures during long process of artificial selection.ConclusionsTaAAP6-3B is a regulator of GPC and its favored allele TaAAP6-3B-I exhibits an obvious potential application in wheat high-GPC breeding.

Identification and characterization of finger millet OPAQUE2 transcription factor gene under different nitrogen inputs for understanding their role during accumulation of prolamin seed storage protein.

In this study, we report the isolation and characterization of the mRNA encoding OPAQUE2 (O2) like TF of finger millet (FM) (Eleusine coracana) (EcO2). Full-length EcO2 mRNA was isolated using conserved primers designed by aligning O2 mRNAs of different cereals followed by 3' and 5' RACE (Rapid Amplification of cDNA Ends). The assembled full-length EcO2 mRNA was found to contain an ORF of 1248-nt coding the 416 amino acids O2 protein. Domain analysis revealed the presence of the BLZ and bZIP-C domains which is a characteristic feature of O2 proteins. Phylogenetic analysis of EcO2 protein with other bZIP proteins identified using finger millet transcriptome data and O2 proteins of other cereals showed that EcO2 shared high sequence similarity with barley BLZ1 protein. Transcripts of EcO2 were detected in root, stem, leaves, and seed development stages. Furthermore, to investigate nitrogen responsiveness and the role of EcO2 in regulating seed storage protein gene expression, the expression profiles of EcO2 along with an α-prolamin gene were studied during the seed development stages of two FM genotypes (GE-3885 and GE-1437) differing in grain protein content (13.8 and 6.2%, respectively) grown under increasing nitrogen inputs. Compared to GE-1437, the EcO2 was relatively highly expressed during the S2 stage of seed development which further increased as nitrogen input was increased. The Ecα-prolamin gene was strongly induced in the high protein genotype (GE-3885) at all nitrogen inputs. These results indicate the presence of nitrogen responsiveness regulatory elements which might play an important role in accumulating protein in FM genotypes through modulating EcO2 expression by sensing plant nitrogen status.

Identification of potential lipid binding regions in cereal proteins and peptides with the use of bioinformatics

In an era where sustainability becomes increasingly prominent it is clear that agricultural sources are playing an important role in this goal. Cereal science is a well-established research area when it comes to agricultural and food science related topics. Throughout the years a wealth of information has been gathered when it comes to cereals. Recent bioinformatics tools allow detailed scanning of cereal proteins and peptides. The approach presented in this paper will demonstrate a full identification and partial characterization of all potential helical lipid binding regions in a number of proteins and peptides present in cereals. These results will be linked to experimental findings as much as possible and where relevant the contribution to more mechanistic insight in the functionality of the proteins and peptides will be discussed. Finally the use of the approach in helping researcher to fine-tune their search for a better understanding of their protein or peptide of interest will be explained.

Using LC-MS to examine the fermented food products vinegar and soy sauce for the presence of gluten

A strict, lifelong gluten-free (GF) diet is currently the only treatment for coeliac disease (CD). Vinegar and soy sauce are fermented condiments that often include wheat and/or barley. During fermentation cereal proteins are partially degraded by enzymes to yield peptide fragments and amino acids. Whether these fermented products contain intact or degraded gluten proteins and if they are safe for people with CD remains in question. LC-MS offers the benefit of being able to detect hydrolysed gluten that might be present in commercial vinegar and soy sauce products. LC-MS revealed the presence of gluten in malt vinegar, wherein the identified peptides derived from B-, D- and γ-hordein from barley, as well as γ-gliadin, and HMW- and LMW-glutenins from wheat that are known to contain immunopathogenic epitopes. No gluten was detected in the soy sauces examined despite wheat being a labelled ingredient indicating extensive hydrolysis of gluten during soy sauce production.

Lysine Requirements of Healthy Pregnant Women are Higher During Late Stages of Gestation Compared to Early Gestation

Lysine is the first limiting amino acid in cereal proteins and is found mainly in animal-derived products. Current Dietary Reference Intake (DRI) recommendations extrapolate lysine requirements during pregnancy from nonpregnant adult data, and may underestimate true requirements. Our objective is to define a quantitative lysine requirement in healthy pregnant women and to determine whether requirements vary between 2 phases of gestation. Fourteen pregnant women in early (12-19 wk) and 19 women in late (33-39 wk) gestation were studied using the indicator amino acid oxidation technique. Individual lysine intakes (6-84 mg · kg-1 · d-1, deficient to excess) were tested on each study day as a crystalline amino acid mixture based on egg protein composition. Isonitrogenous diets maintained protein intake at 1.5 g · kg-1 · d-1 and calorie intake at 1.7 times resting energy expenditure during each study day. Phenylalanine and tyrosine intakes were held constant across all lysine intakes. Breath and urine samples were collected at baseline and isotopic steady state. Lysine requirements were determined by measuring the oxidation of L-[1-13C]-phenylalanine to 13CO2 (F13CO2). Biphase linear regression crossover analysis was used to determine a breakpoint (which represents the estimated average requirement, EAR) in F13CO2. The EAR for lysine during early gestation was determined to be 36.6 mg · kg-1 · d-1 (R2 = 0.484, upper 95% CI = 46.2 mg · kg-1 · d-1), similar to an earlier adult requirement of 36 mg · kg-1 · d-1. The EAR for lysine during late gestation was determined to be 50.3 mg · kg-1 · d-1 (R2 = 0.664, upper 95% CI = 60.4 mg · kg-1 · d-1), 23% higher than the current pregnancy DRI EAR recommendation of 41 mg · kg-1 · d-1. Our results suggest that lysine requirements are higher during late gestation compared to early gestation, and that current dietary lysine recommendations during late stages of pregnancy may be underestimated. The results have implications for populations consuming cereal-based diets as their primary source of protein. This trial was registered at clinicaltrials.gov as NCT01776931.

Effect of the dough mixing process on the quality of wheat and buckwheat proteins

The changes in the structure of cereal proteins during the mixing of flour into dough was described and evaluated. Wheat gliadins and glutenins (gluten proteins) have unique physical properties and play an important role in breadmaking. The effect of mixing time on the formation and the structure of the gluten network was determined using scanning electron microscopy (SEM). Buckwheat flour (gluten-free) was used to compare the development of structure during the mixing process.