Large deformation stress relaxation and compression-recovery of gluten representing different wheat classes

Despite the great variety of physicochemical and rheological tests available for measuring wheat flour, dough and gluten quality, the US wheat marketing system still relies primarily on wheat kernel hardness and growing season to categorize cultivars. To better understand and differentiate wheat cultivars of the same class, the tensile strength, and stress relaxation behavior of gluten from 15 wheat cultivars was measured and compared to other available physicochemical parameters, including but not limited to protein content, glutenin macropolymer content (GMP) and bread loaf volume. In addition, a novel gluten compression–relaxation (Gluten CORE) instrument was used to measure the degree of elastic recovery of gluten for 15 common US wheat cultivars. Gluten strength ranged from 0.04 to 0.43 N at 500% extension, while the degree of recovery ranged from 5 to 78%. Measuring gluten strength clearly differentiated cultivars within a wheat class; nonetheless it was not a good predictor of baking quality on its own in terms of bread volume. Gluten strength was highly correlated with mixograph mixing times (r = 0.879) and degree of recovery (r = 0.855), suggesting that dough development time was influenced by gluten strength and that the CORE instrument was a suitable alternative to tensile testing, since it is less time intensive and less laborious to use.     

Stress–Relaxation Properties of Mixograph Semolina–Water Doughs from Durum Wheat Cultivars of Variable Strength in Relation to Mixing Characteristics,Bread- and Pasta-making Performance

Stress–relaxation behaviours of Mixograph semolina–water doughs prepared from Canadian durum wheat cultivars with diverse gluten strength were investigated and related to mixing characteristics, large deformation properties, and bread- and pasta-making quality. Semolina from «strong» (S) and «moderately strong» (MS) durum wheat cultivars required a longer Mixograph mixing time (4–5 min) and higher work input (140–196 Arbitrary Units) to mix to peak dough resistance (PDR) than «weak» (W) and «very weak» (VW) durum cultivars (2–3 min and 80–117 AU). Extensigraph maximum resistance to extension (R_max/E ratio) and Alveograph P/L (tenacity to length ratio) values were higher for doughs from S cultivars than for MS, W, and VW cultivars. Doughs from S cultivars exhibited higher storage modulus (G′) and lower tan δ values at all frequencies, and slower rates of stress relaxation as compared to MS, W, and VW cultivars. Stress relaxation (times to relax 50% (t₅₀) and 75% (t₇₅) of initial stress) indicated that stronger doughs, which had higher proportions of glutenins, took longer to reach these iso-relaxation states, regardless of their initial relaxation modulus value. The parameters t₅₀and t₇₅were also strongly correlated with dough mixing properties, Extensigraph R_max/E, Alveograph P/L, mixing energy, mixing time and loaf volume obtained by a long and a short bread-making process. However, for S cultivars loaf volume was 10 to 20% lower than that expected of bread wheat of comparable protein content. Stress relaxation data demonstrated no simple correlation to pasta cooking quality indicating that stronger gluten did not translate into a superior pasta cooking quality. Results are interpreted in the context of multimodal networks and transient networks with reversible crosslinks.     

Dynamic water vapour sorption in gluten and starch films

Water sorption of gluten and wheat starch films as a function of water activity was studied using gravimetric step-change sorption experiments. Films of different thicknesses were used with the aim to vary the characteristic diffusion time and to get insights in the contribution of the polymer-chain rearrangement in the sorption behaviour. It is shown that both starch and gluten are in the glassy state for a water activity a_w below 0.9. From comparison of the dynamical sorption curves with a Fickian diffusion model, it is shown that water diffusion in gluten films seems Fickian for a_w < 0.7, and non-Fickian for a_w > 0.7, while for starch films, non-Fickian sorption behaviour is observed for a_w > 0.1. The results show that polymer-chain rearrangement and the stress built up in the matrix play an important role in the sorption dynamics of these films. Even when the material is in the glassy state matrix relaxation phenomena play a role in the sorption behaviour of starch and gluten.     

Temperature dependence of relaxation spectra for highly hydrated gluten networks

In the present investigation, the temperature dependence (0–50 °C) of the relaxation spectrum of hydrated gluten was studied using novel numerical algorithms. Tikhonov regularization, in conjunction with the L-curve criterion for optimal calculation of the regularization parameter, was used to generate the relaxation spectrum from stress relaxation measurements on shear. The methodology used revealed six molecular events with baseline resolution that could be grouped into fast- and slow-relaxation regimes. The fast-relaxation regime exhibited strong temperature dependence whereas the slow one is temperature independent indicating on the whole two dominant mechanisms of interactions. The “loop and train” structural model for gluten interactions was found adequate to describe the relaxation events in this system, with the fast regime being assigned to interactions due to hydrogen bonding whereas the slow one to permanent cross-linking of the entire network. Findings of the present investigation provide fundamental understanding and give new insights into the complexity of interactions and relaxation modes of hydrated gluten.     

Linear viscoelasticity of gluten: Decoupling of relaxation mechanisms

The influence of water content on the relaxation dynamics of mesoporous gluten networks has been explored in a wide range of temperatures. The systems were investigated in the linear viscoelastic region by means of stress relaxation, creep and numerical analysis of data. Time-temperature superposition principle and sticky reptation dynamics have been used to provide molecular interpretation of gluten relaxation. Overall, hydration influences relaxation behaviour of the system, which can be linked to changes in the secondary structure of gluten proteins with increase in water content. Relaxation spectra calculated with Tikhonov regularization revealed the remarkable influence of water on the long times relaxation processes of the material. Creep measurements and extraction of dynamic data with direct conversion of creep data via Laplace transform augmented the experimental timeframe of observations to low frequencies unattainable by standard frequency sweeps. The predominance of loss modulus at long times is attributed to migration of water within the nanopores of the structure. Samples also exhibit self-similar relaxation a characteristic of systems existing at a critical state. Two relaxation mechanisms can be distinguished: one arising from viscoelastic relaxation of protein chains and an additional stemming from poroelastic relaxation owing to migration of water in the system.     

The effect of sodium chloride on gluten network formation and rheology

Gluten samples were obtained from two wheat flours with different levels of total protein in the presence or absence of sodium chloride (2% flour base). The dynamic oscillation rheology, large extensional deformation, confocal laser scanning microscopy (CLSM), transmission electron microscopy (TEM) and chemical analysis of disulfide bond linkages and the ratio of polymeric glutenins and monomeric gliadins were used to investigate the effect of salt on the structure and rheological properties of gluten. CLSM and TEM images showed that NaCl caused the gluten to form fibrous structure. The presence of NaCl increased non-covalent interactions and β-sheet structure, measured by FTIR, in gluten proteins. The gluten matrix formed with salt resulted in higher tan δ values corresponding to a less elastic network when measured using oscillatory rheometry. Large deformation extensional measurements showed that the maximum force to fracture were lower for the gluten samples prepared in the presence of NaCl. The results from this study indicate that changes in the solvent quality due to the presence of NaCl during dough mixing result in different molecular conformation and network structure of gluten proteins which contributed to the differences in the rheological properties.     

In situ observation of the freezing process in wheat dough by confocal laser scanning microscopy (CLSM): Formation of ice and changes in the gluten network

Freezing of bread dough is widely applied in food industry. However, freezing impairs the baking performance of dough, which is largely attributed to structural changes as induced by ice formation. The aim of the present investigation was to image ice formation during freezing of dough and to assess the structural changes in the gluten network. A confocal laser scanning microscope (CLSM) equipped with a freezing stage was used to follow ice formation in the reflection and transmission (bright field) mode. Wheat dough with air inclusions served as a model for fermented dough. The gas pores and the ice crystals could be imaged by confocal laser reflection. Ice formation was initiated at the gas pore interface, where large ice crystals were formed during a freezing time of 4 h at −15 °C. The freezing of gluten samples stained with rhodamin was followed in the fluorescence mode. The cryoconcentration of gluten could be observed, but no irreversible changes in the microstructure of gluten were detected upon thawing. It is concluded that the gas pore interfaces in dough are preferential sites for ice nucleation, favouring the growth of ice crystals in these regions and by this a freeze induced redistribution of water in dough.     

Observation of wheat flour doughs under mechanical treatment using confocal microscopy and classification of their microstructures

RheOptiCAD^® is a cell allowing observation of microstructures under thermo-mechanical treatments. Using the device fitted on a confocal microscope, wheat flour doughs containing 3 levels of water were studied. Dough were placed between 2 parallel plates and submitted to motion of the bottom one. Temperature was controlled and fixed at 20 °C. Methodology and experimental parameters, but also balance between optical and mechanical ones are described and justified to set a dynamic observation in real time. Rhodamine B labeled gluten network was observed under dynamic mechanical treatment, i.e. a 0.4 mm-0.3 Hz oscillation applied during 30 s. The evolution of the gluten network microstructure along time of deformation and depending on the recipe was studied. Image sequences are presented to visualize the qualitative differences in the network. Differences in microstructures were observed. A grey scale mathematical morphology is proposed to qualify and quantify these differences. Images can be separated according to the level of orientation of the network and to the local protein concentration in the gluten fibers.     

Observation of wheat flour doughs under mechanical treatment using confocal microscopy and classification of their microstructures

RheOptiCAD^® is a cell allowing observation of microstructures under thermo-mechanical treatments. Using the device fitted on a confocal microscope, wheat flour doughs containing 3 levels of water were studied. Dough were placed between 2 parallel plates and submitted to motion of the bottom one. Temperature was controlled and fixed at 20 °C. Methodology and experimental parameters, but also balance between optical and mechanical ones are described and justified to set a dynamic observation in real time. Rhodamine B labeled gluten network was observed under dynamic mechanical treatment, i.e. a 0.4 mm-0.3 Hz oscillation applied during 30 s. The evolution of the gluten network microstructure along time of deformation and depending on the recipe was studied. Image sequences are presented to visualize the qualitative differences in the network. Differences in microstructures were observed. A grey scale mathematical morphology is proposed to qualify and quantify these differences. Images can be separated according to the level of orientation of the network and to the local protein concentration in the gluten fibers.     

Facilitation of α-polylysine in TGase-mediated crosslinking modification for gluten and its effect on properties of gluten films

In this study, α-polylysine was used to enhance the cross-linking effect of TGase on gluten and its effects on properties of gluten films were investigated. The amount of free ammonia released from the cross-linking reaction of gluten induced by TGase at the presence of α-polylysine obviously increased, and more polymers with higher molecular weight were formed from the SDS-PAGE results, which indicated that the TGase-mediated cross-linking reaction ability of gluten was strengthened with the incorporation of α-polylysine. The tensile strength of the films from gluten modified with TGase (20 units/g wheat gluten) and 2% α-polylysine (g/g gluten) for 3 h increased from 4.02 ± 0.09 MPa to 5.28 ± 0.14 MPa, which was more effective than that treated with TGase alone (in which the tensile strength of the films was 4.49 ± 0.10 MPa). The TGase treatment with α-polylysine of gluten improved the water stability of the films much more than that treated with TGase alone. A rougher surface and a more compact cross-section structure were observed by SEM for the films from TGase-α-polylysine treated gluten. The contact angles between the gluten films surface and a water droplet increased because of TGase-mediated cross-linking modification.     

Surface properties and locations of gluten proteins and lipids revealed using confocal scanning laser microscopy in bread dough

Surface properties of gluten proteins were measured in a dilation test and in compression and expansion tests. The results showed that monomeric gliadin was highly surface active, but polymer glutenin had almost no surface activity. The locations of those proteins in bread dough were investigated using confocal scanning laser microscopy and compared with polar and nonpolar lipids. Added gluten proteins participated in the formation of the film or the matrix, surrounding and separating individual gas cells in bread dough. Gliadin was found in the bulk of dough and gas ‘cell walls’. Glutenin was found only in the bulk dough. Polar lipids were present in the protein matrix and in gas ‘cell walls’, as well as at the surface of some particles, which appeared to be starch granules. However, nonpolar lipid mainly occurred on the surface of particles, which may be starch granules and small lipid droplets. It is suggested that the locations of gluten proteins in bread dough depends on their surface properties. Polar lipid participates the formation of gluten protein matrix and gas ‘cell walls’. Nonpolar lipids may have an effect on the rheological properties by associating with starch granule surfaces and may form lipid droplets.     

Preparation and properties of wheat gluten based bioplastics with fish scale

The objective of this study was to prepare the wheat gluten based bioplastics with fish scale (FS) through compression molding. The tensile strength of the wheat gluten/FS composites (the range of 6.5–7.5 MPa) was higher than that of the neat wheat gluten-based bioplastic (3.40 MPa). There was a good dispersion of the fish scale powder embedded within the wheat gluten matrix. Dynamic mechanical analysis results showed that the tan delta max peak height and storage modulus of the wheat gluten-based bioplasic was reduced by adding the fish scale. Moreover, the addition of the fish scale caused a weight loss and the surface of the wheat gluten based bioplastic after 120 h of accelerated weathering were differed from the neat wheat gluten based bioplastic. These results may help to find a new applications for fish scale waste to control the degradation rate of a wheat gluten based bioplastic in the agricultural field.     

Effect of temperature, time and wheat gluten moisture content on wheat gluten network formation during thermomolding

A plastic-like material can be obtained by thermomolding wheat gluten protein which consists of glutenin and gliadin. We studied the effect of molding temperature (130-170 °C), molding time (5-25 min) and initial wheat gluten moisture content (5.6-18.0%) on the gluten network. Almost no glutenins were extractable after thermomolding irrespective of the molding conditions. At the lowest molding temperature, the extractable gliadin content decreased with increasing molding times and moisture contents. This effect was more pronounced for the α- and γ-gliadins than for the ω-gliadins. Protein extractabilities under reducing conditions revealed that, at this molding temperature, the cross-linking was predominantly based on disulfide bonds. At higher molding temperatures, also non-disulfide bonds contributed to the gluten network. Decreasing cystine contents and increasing free sulfhydryl and dehydroalanine (DHA) contents with increasing molding temperatures and times revealed the occurrence of β-elimination reactions during thermomolding. Under the experimental conditions, the DHA derived cross-link lanthionine (LAN) was detected in all gluten samples thermomolded at 150 and 170 °C. LAN was also formed at 130 °C for gluten samples containing 18.0% moisture. Degradation was observed at 150 °C for samples thermomolded from gluten with 18.0% moisture content or thermomolded at 170 °C for all moisture contents.     

Similarities and differences in secondary structure of viscoelastic polymers of maize α-zein and wheat gluten proteins

The secondary structure of a dough-like zein polymer was compared to the structure present in a wheat viscoelastic system using FT-IR spectroscopy. When zein was mixed at 35 °C, which is above its glass transition temperature (T_g), changes in its secondary structure suggested that the protein loses its native structure, mainly composed of α-helices (68%), and a viscoelastic system is formed by a structural rearrangement that favors β-sheet structures. This rearrangement is very similar to the structural changes observed in gluten viscoelastic polymers. Upon removal of shear stress, the zein polymer showed a rapid decrease in the proportion of β-sheet structures (from 48% to 28% after the first 3 min) in favor of unordered structures. At the same time, the viscoelasticity of the polymer decreased rapidly. In contrast, gluten, in a similar viscoelastic system and held at the same temperature, showed a fairly constant high content of β-sheet structures (49%) coinciding with the slow relaxation time typical of gluten networks after the removal of shear. We speculate that the addition of a protein capable of causing extensive and stable β-sheet formation in the zein–starch viscoelastic polymer could increase the stability and relaxation time of the zein system and, thereby, create the possibility of a zein dough with similar functionality to a wheat viscoelastic system.     

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.     

Improving the baking quality of bread wheat using rapid tests and genomics: The prediction of dough rheological parameters by gluten peak indices and genomic selection models

The measurement of baking quality by dough mixing and viscoelastic tests is often labor-intensive and time-consuming, thus the possibility to predict these rheological parameters either by rapid tests or genomic selection models based on molecular markers was investigated in this study. A winter wheat breeding population of 128 genotypes was measured for its rheological parameters, gluten peak indices and other rapid test variables in multiple environments. The prediction accuracy of the rheological parameters was assessed in different cross-validation schemes, reflecting scenarios encountered either in food-processing or by plant breeders. Predictions based on gluten peak indices (r = 0.41–0.82) outperformed the other investigated rapid tests in the food-processing scenario. The combination of gluten peak indices with sedimentation value and protein content gave furthermore the overall highest prediction accuracy (r = 0.70). Cross-validation results from the plant breeding scenario revealed that the prediction accuracy of genomic selection models was merely slightly lower (r = 0.42) than using a combination of sedimentation value and protein content (r = 0.44). Merging gluten peak indices with genomic estimated breeding values resulted in a higher accuracy than either method alone (r = 0.52), and was a promising strategy to select for baking quality in wheat breeding.     

Susceptibility of wheat gluten to enzymatic hydrolysis following deamidation with acetic acid and sensory characteristics of the resultant hydrolysates

The effect of acetic acid and hydrochloric acid (HCl) deamidation pretreatment on the susceptibility of wheat gluten to enzymatic hydrolysis by Pancreatin and sensory characteristics of the resultant hydrolysates was investigated. At two degrees of deamidation (24% and 60%, with or without moisture-heating, respectively), wheat gluten pretreated by acetic acid deamidation was more susceptible to be hydrolyzed as evaluated by the hydrolysis degree, nitrogen solubility index, titratable acid amount and free carbohydrate content of the hydrolysates. Wheat gluten pretreated by acetic acid deamidation at a degree of 24% exhibited the highest susceptibility to enzymatic hydrolysis. Moisture-heating (121 °C, 10 min) in the deamidation pretreatment decreased the susceptibility of wheat gluten to enzymatic hydrolysis and the peptide factions of ≤3000 Da in the hydrolysates due to the formation of larger molecule weight aggregates. The hydrolysates prepared from acetic acid deamidated wheat gluten showed more intense glutamate-like and sauce-scented taste and better nutritional characteristics.     

Characteristics of enzymatic hydrolysis of thermal-treated wheat gluten

Effect of thermal treatment at 50–90 °C on wheat gluten hydrolysis by papain was evaluated in this study. Thermal treatment decreased the amount of sodium dodecyl sulfate (SDS) extractable protein. The treatments at 80 and 90 °C had a strong impact on protein extractability. Thermal treatment for 30 min resulted in a significant reduction in SDS extractable glutenin level in wheat gluten. A significant drop in free sulphydryl level was found in wheat gluten treated at 70 °C for 30 min. It indicated that cross-linking of glutenin through S–S occurred during thermal treatment. The treatments at 70–90 °C led to significant decreases in soluble and nitrogen level, while significant increases in peptide nitrogen amount in the hydrolysates from treated gluten were found. A time-dependent effect was observed for the changes in soluble forms of nitrogen and PN. Thermal treatment resulted in molecular mass distribution change according to gel permeation chromatography analysis. Thermal treatment significantly increased the amount of fractions with molecular mass beyond 10 K (67.2%) in the hydrolysates and greatly decreased the amounts of fractions with MM of 10–5 K and below 5 K in hydrolysates.     

A major QTL for gluten strength in durum wheat (Triticum turgidum L. var. durum)

Gluten strength is an important characteristic, determining the end product quality of durum wheat semolina. To identify the genetic basis of gluten strength in North Dakota durum cultivars, a doubled haploid population was developed from the cross of a weak gluten cultivar ‘Rugby’ and a strong gluten cultivar ‘Maier’. A framework linkage map consisting of 228 markers was constructed and used with phenotypic data on gluten strength (measured by sedimentation volume) to conduct single- and two-locus QTL analyses. Only one consistent QTL (QGs.ndsu-1B) contributing up to 90% of the phenotypic or 93% of the genotypic variation was detected on 1BS. No QTL × QTL or QTL × environment interactions were observed. The QGs.ndsu-1B was flanked by two DArT markers which were converted to STS markers and used along with SSR and EST-SSRs to develop a map of 1BS. QTL analysis delineated QGs.ndsu-1B in a 7.3 cM region flanked by an STS marker (STS-wPt2395) and a SSR marker (wmc85). The adapted background of this material and availability of PCR-based markers closely associated with this locus represent invaluable resources for marker-assisted introgression of gluten strength into other durum wheat varieties. A single QTL segregating in this population also makes it an ideal target for map-based cloning.     

Production of gluten-free wheat starch by peptidase treatment

The potential of peptidase-containing bran extracts from germinated cereals (wheat, emmer, barley) and a peptidase preparation from Aspergillus niger (AN-PEP) to degrade gluten in wheat starch below the threshold for gluten-free foods of 20 mg/kg was compared. The gluten-specific peptidase activity of the peptidases was determined by using gliadin as a protein-based substrate as well as the two celiac-active peptides PQPQLPYPQPQLPY (α-gliadin) and SQQQFPQPQQPFPQQP (γ-hordein). The peptidase activity of AN-PEP exceeded the activities of bran from germinated cereals by a factor up to 690,000. Three wheat starches with initial gluten contents of 110, 1679, and 2070 mg/kg, respectively, were incubated with bran extracts and AN-PEP, lyophilized, and residual gluten was quantitated by a competitive ELISA. Unlike peptidases from bran extracts, AN-PEP was capable of degrading gluten below 20 mg/kg in all starches. The absence of gluten in AN-PEP-treated starches was confirmed by liquid-chromatography-mass spectrometry. The properties of gluten-free starches were comparable to the native starches with the exception of a reduced viscosity after AN-PEP treatment. This problem could be overcome by using higher enzyme concentrations and shorter incubation times or by optimizing AN-PEP production for lower residual α-amylase activity.