Gluten Aggregation Research Articles

Small‐scale methods to assess the gluten properties of durum wheat

Background and objectivesDurum breeders are interested in being able to remove weak gluten genotypes from their crossing populations. Although numerous tests exist to measure gluten strength, methods that can be used in early generations, use minimal sample with little processing and have good discrimination are lacking. The purpose of this work was to evaluate several small‐scale methods to predict gluten strength in durum wheat with emphasis on the gluten aggregation behavior measured using a GlutoPeak device.FindingsUsing as little as 9 g of wholemeal (or semolina), a reproducible GlutoPeak aggregation curve is developed at the end of 5 min. Wholemeal gave higher precision than semolina and is more convenient to prepare when screening breeders’ material. The GlutoPeak Area and Peak Mixing Time (PMT) are good predictors of mixograph and gluten index (GI), which are traditional measures of gluten strength. Good correlations between these parameters were obtained using a diverse set of 145 durum genotypes. GlutoPeak Area gave the best prediction of GI from wholemeal samples allowing a breeder to remove weak‐moderate GI samples from a population using as little as 9 g of wholemeal. The GlutoPeak test showed higher correlations with mixograph and GI than the small‐scale biochemical methods, was easier to perform and uses no chemicals. However, GlutoPeak is not suitable when only mg quantities of sample are available.ConclusionsThe utility of the GlutoPeak device has been further validated in this work for screening durum wheat lines for gluten strength. Wholemeal is a suitable sample (9 g minimum) and provides good precision and reproducibility with a short analysis time of 5 min and requires minimal operator skill. It is suited to screening breeders’ populations for gluten strength and is preferred over SIG, HPLC determination of UPP and IP% determination using a turbidity assay.Significance and noveltyThe GlutoPeak can be used to effectively screen out low‐intermediate gluten strength in a durum wheat breeders’ population using 9 g of wholemeal with a 5 min run time.

Interactions between dietary fiber and ferulic acid changed the aggregation of gluten in a whole wheat model system

As the main phenolic acid in wheat bran, ferulic acid (FA) may impact the reactions between disulfide bonds (SS) and sulfhydryl groups (SH) to change gluten proteins aggregation. This study investigated effects of dietary fiber (DF) and FA on secondary structures, thermal properties, and microstructures of gluten proteins. Results showed that the major portion of gluten secondary structures in all test groups was β-sheets (3583–4498 cm2/m2) while α-helices (869–1628 cm2/m2) made up only a small part. The group with 20 g/kg FA and 60 g/kg DF had a similar β-sheet content (4462 cm2/m2) as the control group (4408 cm2/m2), which decreased the damage of gluten structures induced by FA or DF. The total equivalent thiol (SHeq) content, SS content, denaturation temperature (Tp), and enthalpy (△H) of gluten proteins were significantly decreased (P < .05) compared with the control group when DF or FA increased. DF and FA addition also led to a more disordered gluten network shown by scanning electron microscopy (SEM). When DF and FA were added together, the interaction between DF and FA prevented the SS decrease. However, thermal properties analysis of groups including different contents of FA and DF indicated that stability was not improved as expected.

Wheat Gluten Aggregates as a Reinforcement for Poly(vinyl alcohol) Films

Dispersion of fillers in polymers is problematic, particularly nanofillers that prefer to agglomerate. Here, in-situ filler formation is demonstrated where a protein aggregate forms in a biodegradable polymer matrix via a low-energy solution incubation method. Wheat gluten protein molecules are dispersed in aqueous poly(vinyl alcohol) (PVA), and then cooperatively self-assemble into anisotropic structures. The tensile modulus of the composite films is up to 241% higher than the PVA controls, and the composite glass transition temperature (Tg) is increased by up to 12.1 ± 1.6 °C. Thermal processing for long times under increasing pressure can encourage protein aggregation into well-defined structures. However, the PVA phase is degraded by severe thermal processing, as evidenced by thermogravimetric analysis (TGA), Fourier transform infrared spectroscopy (FTIR), and modulus reduction. β-Sheet formation in the protein phase is observed with FTIR, indicating that the protein structures are amyloids.

GlutoPeak method improvement for gluten aggregation measurement of whole wheat flour

A shear-based device, the GlutoPeak, was developed to measure the aggregation behavior of gluten. In this study, the GlutoPeak testing method was improved and applied for the gluten aggregation evaluation of whole wheat flour (WWF) of different particle size ranges. The sample/solvent weight ratio, mixing speed, and testing temperature were adjusted to different levels for obtaining more repeatable Peak Maximum Time (PMT) and Maximum Torque (MT) using two commercial WWF samples. The temperature had a greater effect on the repeatability of the PMT, while the sample/solvent weight ratio and mixing speed had lower impact on the repeatability of both PMT and MT. Another four commercial wheat samples were milled to WWFs of different particle size ranges. The gluten aggregation of the WWF was measured using the improved GlutoPeak parameters, which were 8-g WWF and 10-g 0.5 mol/L CaCl2 solvent with a mixing speed of 3000 rpm and test temperature of 20 °C. The PMT significantly decreased (with the exception of SW) and MT significantly increased with a decrease of WWF particle size. Compared to the Mixolab and Farinograph tests, the GlutoPeak method took less time and provided significantly different (p < 0.05) results for WWFs of different particle size ranges.

Flour from sprouted wheat as a new ingredient in bread-making

Despite the nutritional and sensory improvements associated with sprouted grains, their use in baking has been limited until recently. Indeed, severe and uncontrolled grain sprouting induces high accumulations of enzymatic activities that negatively affect dough rheology and baking performance. In this study, wheat was sprouted under controlled conditions and the effects of enrichment (i.e. 15%, 25%, 33%, 50%, 75% and 100%) of the related refined flour (SWF) on dough rheological properties, baking performances and starch digestibility were assessed. Adding SWF to flour significantly decreased dough water absorption, development time, and stability during mixing, which suggests a weakening of the gluten network. However, no significant changes in mixing properties and gluten aggregation kinetics were measured from 25 to 75% SWF. Regardless of the amount added, SWF improved dough development and gas production during leavening. Decreases in gas retention did not compromise bread-making performances. The best result – in terms of bread volume and crumb porosity – was obtained with 50% SWF instead of using SWF alone. Interestingly, in 100% SWF bread the slowly digestible starch fraction significantly increased.

GlutoPeak profile analysis for wheat classification: Skipping the refinement process

The GlutoPeak test can predict wheat flour quality by measuring gluten aggregation properties in a short time and using a small amount of sample; thus it has usefulness along the entire wheat delivery chain. However, no information on the suitability of this new test on whole grain flours is available. In this contest a multivariate approach was used to assess the GlutoPeak test ability to predict dough quality directly from whole grain flour. GlutoPeak test was performed on both refined and whole grain flours (22 samples), obtaining both profiles and calculated indices, which were subjected to data exploration through Principal Component Analysis. Results suggested a trend according to farinographic stability values. Furthermore, k-Nearest Neighbours classification models were developed using the GlutoPeak profiles to predict farinographic stability, leading to average prediction ability of 81.8% for both refined and whole grain flour data. The present outcome suggests the possibility of predicting farinographic stability by GlutoPeak test directly on whole grain flour, thus skipping the refinement process.

Gluten Aggregation Behavior in High‐Shear‐Based GlutoPeak Test: Impact of Flour Water Absorption and Strength

Gluten aggregation behaviors of wheat flour were evaluated using a high‐shear‐based method with the GlutoPeak instrument and related to flour functional properties. GlutoPeak peak time (PT) and peak area (PA) were positively associated with gluten strength but negatively affected by farinograph absorption (FAB). GlutoPeak maximum torque (Tmax) was highly positively (P &lt; 0.001) correlated with FAB regardless of gluten strength. PT and PA increased with the decrease of FAB. This could result in overestimation of gluten strength owing to water absorption. To account for the impact of FAB, a new parameter GlutoPeak strength index (GSI) was introduced for predicting gluten strength. GSI was obtained by multiplying Tmax and PA. This arithmetic product was found to provide greater correlation (r = 0.91) with dough strength than those of PA (r = 0.84) or PT (r = 0.57) based on the analysis of 56 advanced breeding lines with wide range of FAB. Moreover, significant relationships were found between GSI and flour mixing and baking properties. Using 8 g of flour and with a test time of less than 10 min, the GlutoPeak instrument shows great potential as a rapid tool for gluten strength selection.

Effect of extra cysteine residue of new mutant 1Ax1 subunit on the functional properties of common wheat

Subunit pair 1Dx5 + 1Dy10 was recognized as superior subunit combination in wheat and contained an extra repetitive-domain cysteine residue in 1Dx5 that was important for understanding the formation of dough viscoelasticity. In this research, one specific serine codon of the 1Ax1 gene corresponding to the extra cysteine residue of 1Dx5 was substituted by a cysteine codon through site-directed mutagenesis. Four homozygous transgenic lines (T4) expressing the mutant 1Ax1 gene (mut1Ax1) were produced. Their greater dough strength and stability were confirmed by mixograph and were associated with highly increased gluten index, larger amounts of gluten macropolymers, larger size distribution for glutenin macropolymer particles and varied sodium-dodecyl-sulfate sedimentation volumes, compared with those of the one line expressing wild 1Ax1 that had similar expression level of transgene. The contents of β-sheets in dough and disulfide groups in gluten of the mut1Ax1 transgenic lines were significantly increased. The microstructure of dough mixed to peak showed a more continuous gluten matrix in the mutant transgenic lines than the one line mentioned-above. It was concluded that the extra cysteine residue of mutant 1Ax1 subunit plays a positive role in contributing to dough strength and stability of wheat by cross-linking into gluten aggregates through inter-chain disulfide bonds.

Structural properties of gluten modified by ascorbic acid and transglutaminase

ABSTRACTThe effect of transglutaminase (TG) and ascorbic acid (AA) on the physicochemical properties of gluten was studied. The results showed that TG increased the gluten yield, storage modulus, and particle size distribution by forming a G-L bond in the protein structure. TG changes the three-dimensional structure of gluten by forming a G-L bond in the protein chains and partly preventing disulfide bond formation. However, in the high concentration of the enzyme, by reducing the ratio of lysine to glutamine, TG catalyzes the deamidation of glutamine residues and thus the K′, gluten yield, and particle size decrease significantly. The rheological behavior of glutenin macropolymer (GMP) gel was changed and the gluten extensibility increased. AA leads to an improvement in the aggregation of gluten and increases the particle size distribution. The results showed that AA leads to an increase in the disulfide bond formation and the strength of the gluten network. Adding AA reduced the delta value and increased G’ compared with the control.

Exploring Functionality of Hard and Soft Wheat Flour Blends for Improved End‐Use Quality Prediction

Blending wheat or flour to meet end‐use requirements is a critical part of the production process to deliver consistent quality products. The functionality of commercial Canadian hard red wheat flour (HWF) and soft red wheat flour (SWF) blends with ratios of 100:0, 75:25, 50:50, 25:75, and 0:100 (HWF/SWF, w/w) was investigated with new and standard methods to discern which functional properties may be indicators of bread quality and processing performance. Rheological characteristics including farinograph water absorption behavior, dough development time (DT), stability, extensigraph extensibility, and gluten aggregation of wheat flours were significantly influenced by the proportion of HWF in blends of SWF and HWF (P &lt; 0.05). The SWF content in the blends had negative linear relationships with the protein content, lactic acid solvent retention capacity, water absorption, and GlutoPeak peak torque. Polynomial relationships were observed for sodium dodecyl sulfate sedimentation volume, DT, stability, extensibility, resistance, GlutoPeak peak time, and bread loaf volume with the amount of SWF in blends. The results indicate that linear responses may be more closely tied to protein content, whereas polynomial responses may be more indicative of protein quality and baking performance. The GlutoPeak peak time was sensitive to the addition of HWF in the blends, showing a significant change in gluten aggregation kinetics between the 0 and 25% HWF samples. Principal component analysis (PCA) confirmed that GlutoPeak peak time was a significant factor in differentiating the 0% HWF. Protein secondary structures identified in the final baked bread were also PCA factors differentiating the 0% HWF sample. Although the 0% bread sample did not deviate from the observed polynomial trend for bread loaf volume, the differences in bread protein secondary structures may translate into differences in processing tolerance in commercial settings.

Prediction of bread wheat baking quality using an optimized GlutoPeak®-Test method

The GlutoPeak®-Test (GPT) as a rapid small-scale technique was optimized to evaluate the gluten aggregation properties and to predict the loaf volume, on the basis of a multiyear and multilocation analysis of wheat samples, using different solvents. 5 % lactic acid and 1 % sodium chloride displayed significant GPT responses. Relationships between protein content, sedimentation value, GPT parameters and loaf volume were investigated. With 1 % sodium chloride, the torque 15 s before maximum torque (AM) presented the highest correlation with loaf volume of samples from 2013 to 2014 (r = 0.77, r = 0.63, p < 0.001, respectively). A multiple regression analysis indicated that the best prediction of loaf volume was a linear function of protein content and AM, explaining the variation in loaf volume by 63 % and providing an uncertainty of ±39 ml. The accuracy of the validation of the linear function leads to 64 % correct and to 36 % incorrect predictions of the loaf volume. This emphasizes that the application of the linear function of protein content and AM cannot replace the actual measurement of loaf volume, but it could be a useful rapid screening test in breeding for improved baking quality in bread wheat.

Aggregation of gluten proteins in model dough after fibre polysaccharide addition

FT-Raman spectroscopy, thermogravimetry and differential scanning calorimetry were used to study changes in structure of gluten proteins and their thermal properties influenced by four dietary fibre polysaccharides (microcrystalline cellulose, inulin, apple pectin and citrus pectin) during development of a model dough. The flour reconstituted from wheat starch and wheat gluten was mixed with the polysaccharides in five concentrations: 3%, 6%, 9%, 12% and 18%. The obtained results showed that all polysaccharides induced similar changes in secondary structure of gluten proteins concerning formation of aggregates (1604cm−1), H-bonded parallel- and antiparallel-β-sheets (1690cm−1) and H-bonded β-turns (1664cm−1). These changes concerned mainly glutenins since β-structures are characteristic for them. The observed structural changes confirmed hypothesis about partial dehydration of gluten network after polysaccharides addition. The gluten aggregation and dehydration processes were also reflected in the DSC results, while the TGA ones showed that gluten network remained thermally stable after polysaccharides addition.

Sprouted wheat as an alternative to conventional flour improvers in bread-making

Sprouting is a natural process that enhances the nutritional and sensory profile of cereal-based foods. The present work addressed the possibility of using refined flour from sprouted wheat (SWF) to improve the bread-making performance of some flours in place of conventional improvers - i.e. enzymatic improver (EI) and malt (M). Either 0.5% EI or M was added to the control flour (CTRL), as conventionally used in bakeries, whereas SWF was used up to 2%. Unlikely EI and M, 1.5% SWF showed a gluten aggregation strength similar to that of the CTRL, suggesting no worsening of the protein network characteristics. As for the leavening properties, dough development increased, thanks to the enrichment with 1.5% SWF. In addition, presence of SWF improved the amount of gas production during leavening-resulting in bread with high specific volume - and the crumb softness during storage. Addition of SWF may represent a valid alternative to enzymatic improvers or malt for improving the technological performance of wheat flours.

Protein Structural Features in Winter Wheat: Benchmarking Diversity in Ontario Hard and Soft Winter Wheat

A set of 32 winter wheat lines and varieties was selected to benchmark Ontario winter wheat as a first step toward improving quality. Protein secondary structure, total and accessible thiols, rheological properties, gluten aggregation kinetics, and network forming capabilities of different polymers were determined for each wheat line. Results revealed that there were statistically significant differences among the lines selected (P &lt; 0.05). The differences between hard and soft wheat classes were not as large as would be expected, however, despite the range of quality parameters measured. Benchmarks revealed that several soft wheat lines outperformed hard wheat lines in standard breadmaking quality measures. Protein conformation changed significantly as the moisture content of the samples increased to mimic different model product systems: flour, dough, and batter. The conformation of the flour samples exhibited different patterns between hard and soft wheat classes, although these differences became narrower in the dough and batter states. Principal component analysis (PCA) factors included most quality parameters measured, with the notable exceptions of solvent retention capacity tests and total thiols. Protein conformation and accessible thiols were significant PCA factors that tended to override the rheological measures of quality they represented, suggesting that protein secondary structure and disulfide bonding patterns are fundamental aspects of rheological quality measures.

Evaluation of a New Viscometer Performance in Predicting the Technological Quality of Soft Wheat Flour

Gluten aggregation properties were investigated by means of the GlutoPeak device, a viscometer recently proposed as a rapid and sensitive test for measurement of wheat flour technological performance. In this study, 62 soft wheat flour samples of different quality and end use were utilized to evaluate if the GlutoPeak parameters could adequately predict chemical and rheological characteristics of soft wheat flour dough, that is, protein content measured by the Kjeldahl method, dough strength measured by a Chopin alveograph, and dough stability and water absorption measured by a Brabender farinograph. Linear correlation analysis showed that most GlutoPeak curve parameters were strongly correlated with protein content, dough strength, and water absorption. The statistical models, obtained by a stepwise multiple regression method, showed the GlutoPeak device to be a promising tool to characterize wheat flour (Radj2 = 0.84 for protein content, Radj2 = 0.71 for dough strength, and Radj2 = 0.67 for water absorption). The rather high accuracy of the prediction models for the three mentioned parameters confirmed that GlutoPeak parameters are well correlated with other frequently used flour quality parameters and are able to describe flour technological performance.

Empirical rheology and pasting properties of soft-textured durum wheat (Triticum turgidum ssp. durum) and hard-textured common wheat (T. aestivum)

Puroindoline (PIN) proteins are the molecular basis for wheat kernel texture classification and affect flour milling performance. This study investigated the effect of PINs on empirical rheology and pasting properties in Triticum turgidum ssp. durum and Triticum aestivum. Soft wheat (cv. Alpowa), durum wheat (cv. Svevo) and their derivatives in which PINs were deleted (Hard Alpowa) or expressed (cv. Soft Svevo). Presence of PINs affected flour particle size and damaged starch. PINs increased the pasting temperature and breakdown viscosity, while the effect on peak viscosity and setback were not consistent. Presence of PINs was negatively associated with GlutoPeak gluten aggregation energy and farinograph dough stability, suggesting a weakening of the gluten matrix. As regards dough extensibility, the role of PINs was evident only in common wheat: 5DS distal end deletion increased the resistance to extension, without affecting the dough extensibility. This study showed PINs to have different impact on pasting and rheological properties of T. aestivum and T. turgidum ssp. durum flours.

Correlations between gluten aggregation properties defined by the GlutoPeak test and content of quality-related protein fractions of winter wheat flour

The gluten aggregation properties of 19 winter wheat cultivars were investigated using a rapid small-scale (7 min, 8.5 g flour) technique (GlutoPeak test, GPT). Correlations between GPT indices and gliadin (GLIA), glutenin (GLUT), SDS-soluble protein (SDSS), and glutenin macropolymer (GMP) concentrations were established. Two groups of samples were distinguished based on the shape of their GPT curves. Flours of group 1 gave curves with a slower buildup in torque (measured as aggregation time) and a higher area under the curve at peak (maximum torque), compared to those of group 2. Group 1 was characterized by lower GLIA and higher GLUT and GMP contents than group 2. GLIA (r = 0.70, p ≤ 0.001) and SDSS (r = 0.59, p ≤ 0.01) concentrations were significantly correlated to peak torque. GLUT and GMP contents were correlated to the aggregation time (GLUT: r = 0.72, p ≤ 0.001; GMP: r = 0.77, p ≤ 0.001) and the area under the curve at peak (GLU: r = 0.68, p ≤ 0.01; GMP: r = 0.78, p ≤ 0.001), confirming the importance of GMP for gluten strength. GPT could be an alternative to the labor-intensive quantitation of quality related protein fractions of wheat flour.

Characteristics of Perennial Wheatgrass (Thinopyrum intermedium) and Refined Wheat Flour Blends: Impact on Rheological Properties

Intermediate wheatgrass (IWG) (Thinopyrum intermedium) is a perennial grass with desirable agronomic traits and positive effects on the environment. It has high fiber and protein contents, which increase the interest in using IWG for human consumption. In this study, IWG flour was blended with refined wheat at four IWG‐to‐wheat ratios (0:100, 50:50, 75:25, and 100:0). Samples were analyzed for proximate composition, microstructure features, pasting properties (Micro Visco‐Amylo‐Graph device), protein solubility, and total and accessible thiols. Gluten aggregation properties (GlutoPeak tester) and mixing profile (Farinograph‐AT device) were also evaluated. IWG flour enrichment increased the pasting temperature and decreased the peak viscosity of blended flours. IWG proteins exhibited higher solubility than wheat, with a high amount of accessible and total thiols. The GlutoPeak tester highlighted the ability of IWG proteins to aggregate and generate torque. Higher IWG flour enrichment resulted in faster gluten aggregation with lower peak torque, suggesting weakening of wheat gluten strength. Finally, the addition of IWG to refined wheat flour resulted in a decrease in dough development time and an increase in consistency, likely because of the higher levels of fiber in IWG. The 50% IWG flour enrichment represents a good compromise between nutritional improvement and maintenance of the pasting properties, protein characteristics, and gluten aggregation kinetics.

Toughness of Natural Rubber Composites Reinforced with Hydrolyzed and Modified Wheat Gluten Aggregates

The toughness of natural rubber can be improved by using fillers for various rubber applications. Dry wheat gluten is a protein from wheat flour and is sufficiently rigid for rubber reinforcement. The wheat gluten was hydrolyzed to reduce its particle size and microfluidized to reduce and homogenize the particle size and size distribution. The processed wheat gluten aggregates with a size of ~500 nm were used to improve the toughness of natural rubber. The crosslinked rubber composites showed an improvement in moduli, tensile strength and toughness compared to natural rubber; had a greater toughness and elongation than the carbon black filled composites at higher filler fractions. For the composites prepared under the alkali condition, the modified particles increased the moduli in the larger strain region. For the composites prepared under the acidic condition, the modified particles improved the tensile strength, elongation, and toughness. The reinforced composites were characterized with stress–strain, dynamic frequency sweep, and swelling experiments to understand their structure-properties.

Starch facilitates enzymatic wheat gluten hydrolysis

Wheat gluten can be hydrolyzed by either using (vital) wheat gluten or directly from wheat flour. This study investigates the influence of the presence of starch, the main component of wheat, on enzymatic wheat gluten hydrolysis. Wheat gluten present in wheat flour (WFG) and vital wheat gluten (VWG) were hydrolyzed at constant protein concentrations, but subsequently 5.6 times higher amounts of wheat flour. Nevertheless, WFG hydrolysis at 40% total solids resulted in significantly higher degrees of hydrolysis (DH%) than VWG hydrolysis at 7.2% solids. This difference increased to up to 4.5% in 6 h and diminished again for longer reaction times. Possible differences in the gluten composition and the presence of albumins and globulins in wheat flour could not explain the difference in DH% because the addition of starch to VWG increased the rate of hydrolysis similarly. Instead, it was concluded that starch granules impede gluten aggregation, which facilitates the hydrolysis. At higher solid concentrations of up to 70% wheat flour, the positive effect of starch disappeared, because WFG hydrolysis was hindered by mass transfer limitations and lower water activities.