Soluble arabinoxylans extracted from soft and hard wheat show a differential prebiotic effect in vitro and in vivo

Arabinoxylans (AX) are part of dietary fiber. They are currently under study due to their potential prebiotic effect. Wheat whole grain flours contain all the grain layers and, therefore, present a higher arabinoxylan content than white flour. It is known that the chemical structure of these compounds varies with the type of wheat cultivar and the tissue from which they are extracted. In this work, water soluble extractable arabinoxylans (WE-AX) from two types of wheat whole flours (hard and soft) were extracted. We characterized the molecular size distribution and the potential prebiotic effect of those extracts. The prebiotic effect was evaluated in vitro and confirmed in vivo. Bacterial group abundance (Lactobacillus, Bifidobacterium, Clostridium, Enterococcus, Bacteriodes and total bacteria) was determined by quantitative RT-PCR. The molecular size of AX from hard wheats was significantly higher than AX from soft wheats. Both extracts showed potential prebiotic activity by increasing the growth of beneficial bacteria in vitro and in vivo, decreasing the pathogens in the profile of intestinal microorganisms and increasing the amount of short chain fatty acids in the intestine. WE-AX from hard wheat showed a higher prebiotic activity. Prebiotic effect assessed in vitro and in vivo assays showed a significant correlation between both types of analysis. This finding suggests that the in vitro indices performed allow predicting the potential prebiotic effect in vivo.     

Benzoxazinoid and alkylresorcinol content,and their antioxidant potential,in a grain of spring and winter wheat cultivated under different production systems

The health-promoting properties of the grains of common wheat (Triticum aestivum L.) are associated with the presence of unique phytochemicals. This study determines the profile of alkylresorcinols (ARs) and benzoxazinoids (BXs) in T. aestivum spring and winter cultivars grown in Poland under the two different production systems: conventional and organic. Wheat grain extracts were subjected to qualitative and quantitative UPLC-UV-MS/MS analyses. The ARs profile consisted of five 5-n-alkylresorcinol derivatives, among which 5-n-heneicosylresorcinol (C21:0) and 5-n-nonadecanylresorcinol (C19:0) predominated; while six different BXs were determined in hydrothermally treated grains. Our research shows significant differences in the contents of ARs and BXs among wheat cultivars, as well as the two production systems used. Organically grown varieties had their total contents of ARs and BXs significantly higher than those grown conventionally. Another aim of the study was to determine the antioxidant capacity of alkylresorcinol extracts from tested wheat cultivars. The quantitative TLC-DPPH• method for determining the antiradical capacity of wheat ARs extracts was developed. We observed a positive relationship between the free radical scavenging activity of extracts and the total amount of ARs. The biological activity research is important for developing value-added wheat cultivars, having an improved profile and composition of nutritional substances.     

Application of an orcinol-ferric chloride colorimetric assay in barley and wheat accessions for water-extractable and total arabinoxylan

Arabinoxylan is an important hemicellulose potentially affecting wheat baking qualities, barley malt quality, and may impart prebiotic benefits. Water-extractable (WEAX) and total arabinoxylan (TOAX) were characterized in 204 wheat and barley accessions, respectively, using an orcinol-ferric chloride assay. WEAX measurement accuracy was optimum when glucose concentration was greater than 12–13 times pentose concentration. The modified method removed enough excess glucose spectral absorbance to make the corrected and uncorrected lines different, but still significantly correlated (p = 0.009, R = 0.94). Mean WEAX (expressed as percentage WEAX of TOAX) for the wheat accessions was 15.90%, ranging from 8.82% to 24.87%, and for barley accessions WEAX the mean was 7.10%, and ranged from 2.98% to 13.86%. Conclusions are 1) the assay is useful for a breeding program because of its semi-high throughput design for the simultaneous analysis of 16 (WEAX) to 40 (TOAX) barley lines or 24 (WEAX) to 40 (TOAX) wheat lines, 2) the trichromatic measurement reduces the impact of glucose, and 3) the broad range of WEAX measured showed that barley and wheat accessions vary in their extractable and unextractable components.     

GlutoPeak parameters of whole wheat flours for gluten quality evaluation in soft wheat breeding programs

In soft wheat breeding programs, the gluten strength of flours from specific genotypes is determined by various chemical and rheological tests. Based on such tests, the experimental wheat lines with very weak flour gluten are typically selected for the production of soft-dough biscuits, while the lines with medium gluten strength and extensibility are reserved for hard-dough biscuits. Often, the genotypes having high gluten strength are removed from such breeding programs. In the present study, the usability of the GlutoPeak tester on whole wheat flour samples was investigated for assessing the gluten strength of soft wheat breeding materials. In the study, 25 soft wheat genotypes, grown in seven locations for three years, were categorized by commonly used gluten-quality-related parameters. Based on the results of the study GlutoPeak whole wheat flour PMT values ranging from 30.0 to 50.0 s and AM values from 15.0 to 20.0 GPU were found to be suitable for soft-dough biscuit products, whereas the values between 40.0 and 60.0 s and 20.0 and 23.0 GPU were appropriate for hard-dough biscuit products. The genotypes exhibiting AM values > 24.0 GPU and PMT values > 60.0 s were judged to have too-strong gluten, and thus eliminated from the breeding program. The gluten aggregation energy (AGGEN), and the torque after the maximum torque (PM) values were only useful and applicable to flours for soft-dough products. The maximum torque (BEM) values were not effective in discriminating against the genotypes. The results of this study demonstrated that the GlutoPeak whole wheat PMT and AM parameters can be recommended as quick and accurate parameters especially for early generation screening with small-scale tests in soft wheat improvement programs.     

Variation in avenanthramide content in spring oat over multiple environments

Avenanthramides (AVNs), a group of phytochemicals which are unique to oats, provide health benefits through antioxidant activity and other bioactivities. In this study, we explored genotype-by-environment interactions and heritability for AVN concentrations in oats. Avenanthramide concentrations were quantified for 100 breeding lines and cultivars at three locations over two years. While year and environment had an influence on AVN concentrations, with the influence of year being more apparent than that of environment within a year, genotype had the largest impact on AVN concentration. All three major AVNs were found to be heritable. Two methods of calculating heritability on a line mean basis were used. The statistical method yielded heritability estimates of 0.34, 0.39, and 0.41 for AVN 2c, AVN 2p, and AVN 2f, respectively. By comparison the traditional plant breeding method yielded heritability estimates of 0.82, 0.88, and 0.89 for AVN 2c, AVN 2p, and AVN 2f, respectively, indicating that expected gains will be dependent on the scale of the breeding program, the number of target environments, and climate variability. The estimated heritabilities and the 11-fold range in AVN concentrations in the 100 genotypes studied provided evidence that variability for AVN concentration should allow breeding progress for higher AVN concentration.     

Quantitative LC-MS proteoform profiling of intact wheat glutenin subunits

Methodology for the quantitative analysis of intact glutenin proteins by ESI-LC-MS was developed with 1 Da mass resolution, constituting the first published application of proteoform profiling to plant biology. Two parent lines and 28 F5 crosses were analyzed in two blocks, 14 months apart. Control sample data were used to align retention times and normalize abundance between sample sets. A total of 4622 observations of 347 distinct proteoforms between 17 899 and 88 744 Da were observed. Proteoform abundances spanned a 1000-fold range and were linear (r² > 0.990) with dilution. A novel method for the objective elimination of low intensity, noise-dominated data using replicate variability within the dataset is presented. Two abundant PTMs were detected; one known but uncharacterized Bx and Dy high-molecular-weight glutenin subunit (HMWGS) PTM and the other in 24 low molecular weight proteoform pairs. Finally, 16 abundant proteoforms were detected in progeny but not in either parent. This application should increase the statistical power of correlations between gluten complement and functional data and drive the detection of novel PTMs that may indicate differential regulation of the cellular processes related to quality.     

Genetic responses in milling,flour quality,and wheat sensitivity traits to grain yield improvement in U.S. hard winter wheat

A rising global population necessitates continued genetic improvement of wheat (Triticum spp.), but not without monitoring of unintended consequences to processors and consumers. Our objectives were to re-establish trends of genetic progress in agronomic and milling traits using a generational meter stick as the timeline rather than cultivar release date, and to measure correlated responses in flour quality and human wheat-sensitivity indicators. Grain yield and kernel size showed stepwise increases over cycles, whereas wheat protein content decreased by 1.1 g/100 g. Reduced protein content, however, did not result in lower dough strength pertinent to bread baking. A novel method of directly testing gluten elasticity via the compression-recovery test indicated a general increase in gluten strength, whereas the ratio of total polymeric to total monomeric proteins remained stable. Also showing no change with genetic progress in yield were flour levels of gluten epitopes within the key immunotoxic 33-mer peptide. The oligosaccharide fructan, present in milled and wholemeal flours, increased with increasing grain yield potential. While yield improvement in U.S. bread wheat was not accompanied by a decline in gluten strength or systematic shift in a key wheat sensitivity parameter, the unanticipated rise in total fructans does implicate potentially new dietary concerns.     

Breeding and biotechnological efforts in Jatropha curcas L. for sustainable yields

Jatropha is a non-edible, important bioenergy plant, which can grow in marginalized land. The seeds possess about 36% oil and this would be converted into biodiesel or biojet-fuel. Jatropha provides an option for sustainable feed and fuel production due to its inherent qualities including hardy nature, drought tolerance and surviving with limited amount of water, tolerance to unfavorable conditions and excessive moisture. However, heterozygosity, low productivity and poor understanding of its genome are the major impediments to elite line development. Further, classical breeding and advanced technological investments remain limited owing to long juvenile phase and breeding cycles. Scientific technologies that lead to identification of elite genotypes and development of high yielding elite Jatropha lines and effective methods of detoxification of seeds needs immediate priority. Efficient tissue culture system, doubled haploids (DH) and genomic tools are increasingly made available to improve the seed yield and its oil quantity through the development of geminivirus disease resistant lines. The application of advanced, sequencing technologies has presented a repertoire of genomic information for this important yet orphan crop. In the present investigation, we highlight the achievements made in Jatropha towards development of high yielding, virus resistant elite lines and hybrids with yield potential ranging from 3 to 5 tons per hectare in a year, which is a first ever report in the world. We also developed potential biotechnological tools such as genetic transformation, genome editing and next generation genomics tools including linkage maps and QTLs for accelerating breeding efforts through marker assisted selection. Because of our concerted and continuous efforts during the past 15 years, we have overcome all the obstacles and developed high yielding, disease resistant hybrids/lines, advanced cultivation technologies with thorough knowledge on Agronomy of the Jatropha crop. The Jatropha cultivation technology developed for the first time in the world, could open up new avenues for higher yield productivity of commercial viability.     

An improved method for extraction of sorghum polymeric protein complexes

A method for fractionating sorghum proteins using extraction solvents and techniques designed to obtain polymeric protein structures (especially disulfide linked) was developed. Extraction and separation conditions were optimized in terms of completeness of protein extraction, sample stability, and analytical resolution. After pre-extraction of albumins and globulins, a 3-step sequential procedure involving no reducing agents was applied to ground whole sorghum flour. The three fractions obtained represented proportionally different protein polymer contents and molecular weight distribution as evidenced by comparative size exclusion chromatography. Protein composition also varied among the extracts with differences in kafirin composition and non-kafirin proteins detected in the fractions by RP-HPLC and SDS-PAGE analysis. The ability to quantify and further characterize sorghum polymeric protein complexes will be useful for additional studies linking protein structures with functionality and digestibility and variations for these properties within diverse sorghum germplasm.     

New angiotensin-converting enzyme inhibitory peptide from Coix prolamin and its influence on the gene expression of renin-angiotensin system in vein endothelial cells

Coix seed, which is a traditional Chinese medicine, has been used to treat hypertension for thousands of years. It has been shown that Coix prolamin peptides display high levels of angiotensin I converting enzyme (ACE) inhibitory activity. Hence, we purified the ACE inhibitory peptides from Coix prolamin hydrolysates and evaluated the influence of the most potent peptide on the renin-angiotensin system (RAS) genes expression in human umbilical vein endothelial cells (HUVECs). In this study, Coix prolamin peptides were sequentially separated by ultrafiltration, ion exchange chromatography, gel filtration chromatography and RP-HPLC, while the peptide structure was analyzed by mass spectrometry. Next, in silico proteolysis, pharmacophore and molecular docking were further applied to screen and optimize the structure of peptides. Finally, a novel ACE inhibitory peptide VDMF was obtained, in which its influence on the gene expression of RAS signaling pathway in AngⅡ-injury HUVECs was evaluated by quantitative real-time PCR. VDMF significantly down-regulated ACE, AngII type 1 receptor (AT1R) and ACE2 mRNA expression in comparation with model group, while up-regulating Mas gene expression. Hence, we obtained a novel antihypertensive candidate that was derived from the Coix peptides, which could involve a multi-modulation mechanism that regulates blood pressure.     

Mass spectrometry imaging of BARLEYmax fructooligosaccharides

BARLEYmax is a novel hull-less barley cultivar lacking activity of a key enzyme required for starch synthesis (SSIIa), resulting in a grain with reduced total starch, higher dietary fiber, and higher fructan content. In this study, we examined the quantitative differences in fructan among 2 hull-less barley lines (BARLEYmax and Mannenboshi) and 2 peeled barley lines (Glutinous barley rice and Hindmarsh). We found that BARLEYmax had a higher fructan content, particularly 2 types of fructooligosaccharides (FOS), namely kestose and nystose, the important components of fructan that have some beneficial effects for human health. We established a mass spectrometry imaging (MSI) method to further investigate the content and localization of FOS in the barley lines. We found that kestose, nystose, and fructosyl-nystose showed increased molecular ion intensities in BARLEYmax compared with those in other barley lines. These results confirmed that the loss of SSIIa activity in BARLEYmax leads to a decrease in starch in the cytoplasm and an increase in fructan in the vacuole as an energy store. Overall, these findings demonstrate that MSI analysis is helpful for understanding FOS localization in cereal grains and for visual comparison among matured grains.     

Enzymatic degradation of FODMAPS via application of β-fructofuranosidases and α-galactosidases- A fundamental study

Cereals and pulses often contribute to the intake of Fermentable Oligo-, Di-, Monosaccharides, and Polyols (FODMAPs) due to high amounts of fructans or galactooligosaccharides (GOS). FODMAPs can trigger symptoms of Irritable Bowel Syndrome (IBS) and therefore, the development of foods and beverages with a lower FODMAP-content are favourable for IBS patients. Enzyme technology is a promising tool to reduce the FODMAP-content in foods and to maintain product quality. This fundamental study investigates the efficiency of invertase, inulinase, and α-galactosidase as potential food additives to reduce the total FODMAP content of food ingredients. Extracts of high FODMAP ingredients, such as wheat and lentil, and standard solutions of various fructans and GOS were incubated with invertase, inulinase and α-galactosidase for 1 h and 2 h. Contents of oligosaccharides before and after treatment and related IBS-triggering reaction products were quantified using ion chromatography. Inulinase showed a high degradation yield (over 90% of degradation) for both GOS and fructans. For invertase only low degradation yields were measured. α-Galactosidase showed the highest efficiency in decomposing GOS (100% of degradation) and led to non-IBS triggering degradation products. This indicates a high potential for a combined inulinase/α-galactosidase treatment for products containing both fructans and GOS.     

Characterization and bioactivities of young rice protein hydrolysates

Immature rice was reported to contain higher quantities of bioactive compounds than mature rice. Young rice protein is easy to digest and has hypoallergenic potential, with protein content of 7.2–11.5% compared to rice bran at 9.8%. Few studies have reported on bioactivities and characterization of young rice proteins and their hydrolysates. Bioactivities of native protein and protein hydrolysates of two rice varieties (white rice and colored rice) were characterized and investigated for four development stages (flowery, milky, dough, and mature). Degree of hydrolysis of young rice protein was considerably higher than at the mature stage. Highest DPPH and iron chelating activity were found in alcalase® protein hydrolysate during the flowery-to-milky stage. Iron chelating activity was constant in all development stages because of the low polar amino acid content in rice. The ACE activity of alcalase® protein hydrolysate was higher than native protein at the same development stage, as observed in the milky and dough stages. Inhibitory activity of young rice hydrolysate HepG2 cells was concentration-dependent and not correlated with protein molecular size.     

Spatial distribution of functional components in the starchy endosperm of wheat grains

The starchy endosperm of the mature wheat grain comprises three major cell types, namely sub-aleurone cells, prismatic cells and central cells, which differ in their contents of functional components: gluten proteins, starch, cell wall polysaccharides (dietary fibre) and lipids. Gradients are established during grain development but may be modified during grain maturation and are affected by plant nutrition, particularly nitrogen application, and environmental factors. Although the molecular controls of their formation are unknown, the high content of protein and low content of starch of sub-aleurone cells, compared to the other starchy endosperm cells types, may result from differences in developmental programming related to the cells having a separate origin (from anticlinal division of the aleurone cells). The gradients within the grain may be reflected in differences in the compositions of mill streams, particularly those streams enriched in the central and outer cells of the starchy endosperm, respectively, allowing the production of specialist flours for specific end uses.     

Effects of wheat tempering and stone rotational speed on particle size,dough rheology and bread characteristics for a stone-milled weak flour

The poor technological performance of weak wheat flours means that they are usually considered difficult to be transformed into satisfactory bread. During milling, there are several settings that can affect flour characteristics. In this study, we tested two operative parameters that have the potential to affect flour quality – stone rotational speed and wheat tempering. Tempering moistures were set at 11%, 13%, 15%, and 17%, while stone rotational speeds were set at 173, 260, and 346 rpm. Both factors were found to affect operative milling parameters, notably flour yield, process productivity and specific energy consumption. Grain moisture had a significant effect on both dough rheology and bread characteristics (dough stability, tenacity, and extensibility). Dough stability was maximum at 13% moisture. Dough tenacity decreased as moisture increased, while extensibility increased as moisture increased. Bread specific volume and crumb specific volume were improved at 13% and 15% moisture. In conclusion, wheat tempering can be used to improve the potential of a weak flour and bread characteristics, while stone rotational speed affects operative parameters and white flour yield.     

Contributions of individual and combined Glu-B1x and Glu-B1y high-molecular-weight glutenin subunits to semolina functionality and pasta quality

Durum wheat is an important food crop used primarily for pasta production. High-molecular-weight glutenin subunits (HMW-GS) encoded by the closely linked genes Glu-B1x and Glu-B1y are known for their combined effects on pasta quality, but their individual contributions and interactions remain poorly understood. In this study, we show that individual loss-of-function mutants of Glu-B1x (ΔBx6) and Glu-B1y (ΔBy8) were associated with significant reductions in gluten strength compared to the wildtype, with stronger effects in the ΔBxy double mutant. Reductions in gluten strength were reflected in reduced mixograph and alveograph parameters, gluten index, faster extrusion flow rates and increased cooking loss. Interestingly, the Glu-B1x mutation was also associated with significant increases in grain and semolina protein content, increased pasta firmness, reduced starch viscosity and increased amylose in ΔBx6 and ΔBxy. In general, the ΔBx6 mutation had stronger effects than the ΔBy8 mutation, and significant interactions between the two genes were frequent. In addition to the basic knowledge gained on the individual effects of the Bx6 and By8 subunits and their interactions, the genetic stocks developed in this study provide useful tools to study the effects of natural or synthetic HMW-GS on pasta quality parameters in a background lacking endogenous HMW-GS.     

Effect of zein extrusion and starch type on the rheological behavior of gluten-free dough

Previous research has shown that zein, above its glass transition temperature, may adopt molecular structures that are able to form doughs with viscoelastic properties comparable to those of wheat gluten. It is hypothesized that extrusion can promote molecular changes in zein and favor interactions with starch that enhance dough viscoelasticity. Thus, the effects of extruding zein at 90–160 °C on the rheological properties of doughs prepared with potato, rice, and maize starches were determined.Formulations were optimized to provide similar mixing profiles to that of a standard wheat dough. For all zein samples, creep-recovery tests demonstrated that doughs prepared with maize and potato starches were less elastic when compared to doughs prepared with rice starch. Zein doughs produced using rice starch were comparable to wheat-dough. Extensional tests showed that zein extruded at 160 °C provided a larger increase in strain-hardening behavior, which is important for bread production. These samples also exhibited larger extensional stresses. Gel electrophoresis of zein extruded at 160 °C revealed an increase in protein aggregates and the presence of smaller peptides when compared to samples subjected at lower extrusion temperatures.Scanning electron micrographs of doughs containing zein showed starch granules embedded within an amorphous material and fibrous structures, which is attributed to elongated zein.     

Pilot scale production of a non-immunogenic soluble gluten by wheat flour transamidation with applications in food processing for celiac-susceptible people

Celiac disease (CD) is an immune-mediated disease triggered by wheat gluten and related prolamins. A lifelong gluten-free (GF) diet is mandatory to normalize the intestinal mucosa. We previously found that transamidation by microbial transglutaminase of gluten was effective in suppressing the gliadin-specific inflammatory response in CD patients without influencing the main technological properties of wheat flour or semolina. In this study, we produced on a pilot scale a soluble form of transamidated gluten (soluble protein fraction, spf), characterised by a high protein content (88 mg/ml), while native gluten was dramatically reduced (32 ± 2 ppm; R5-ELISA). Using HLA-DQ8 transgenic mice as a CD model, we found suppression of interferon-γ secretion in gliadin-specific CD4⁺ T cells challenged with spf-primed dendritic cells. In terms of functional properties, spf showed both solubility and emulsifying activity values within the range of commercial soluble glutens. Notably, dough prepared by mixing rice flour with spf could leaven. After baking, blended rice bread had a higher specific volume (2.9 ± 0.1) than control rice bread (2.0 ± 0.1) and acquired wheat-like sensory features. Taken together, our results highlighted the technological value of transamidated soluble gluten to improve both nutritional and sensory parameters of GF food.     

Characterization and comparison of predominant aroma compounds in microwave-treated wheat germ and evaluation of microwave radiation on stability

The present study was performed to evaluate the effects of microwave (MW) output power and treatment time on moisture content, lipase and lipoxygenase activities as well as colour changes of wheat germ (WG). In addition, the key aroma compounds in different MW-power-treated WG, which is of importance to the flavour of WG products, were also investigated. The obtained results showed that MW treatment maintained the inherent colour of WG and significantly reduced the moisture content (maximum reduction of 95%) and the activities of lipase and lipoxygenase (maximum reduction of 65% and 99%, respectively). In terms of aroma compounds, with the increase of the MW output power, the content of esters, alkanes, alcohols and acids decreased, while the content of heterocyclic compounds, nitrogen-containing compounds, aldehydes and ketones increased, providing more compounds with roasted flavour and less volatiles with grass-like flavour. Therefore, MW treatment was an effective stabilization method for WG utilization.