Improved functional properties for soy–wheat doughs due to modification of the size distribution of polymeric proteins

Physical modification of soy flour was shown to greatly improve the dough and baking qualities of soy–wheat (1:1) composite doughs, compared to raw soy flour, giving better stability and R_max, although extensibility was still below that of the wheat dough.Reasons for improvements caused by the physical-modification process were sought by determining the relative size distribution of proteins in the soy–wheat composite doughs by size-exclusion high-performance liquid chromatography (SE-HPLC). Results were expressed as the proportion of ‘unextractable polymeric protein’ (%UPP)—the proportion of the protein that is over 100,000 Da and only extractable after sonication. Protein extracts from the soy–wheat dough were sampled at different stages of dough mixing and fermentation, and their molecular-size distributions evaluated.Unextractable soy proteins were lower in raw soy flour (only 8% UPP) than in two physically-modified soy flours (19 and 34% UPP, respectively). Unextractable polymeric protein was much greater for wheat flour (57% UPP). After mixing a 1:1 soy–wheat composite dough, the %UPP was 36 and 22 (for the two types) when made from physically modified soy flours, compared to 8 for a composite dough using raw soy flour, and 43 for a wheat-only dough. The higher proportion of UPP for the wheat-modified soy doughs was taken as a reason for this composite dough providing better dough and baking qualities. Prolonged fermentation time caused a decrease in UPP percentages for all composite doughs and for the wheat-only dough.     

Winter wheat genotypes under different levels of nitrogen and water stress: Changes in grain protein composition

Hard white winter wheat with superior and consistent quality is preferable for Asian markets. This study investigated the combined influences of moisture deficit during grain-fill and N management on protein quality, dough rheological properties, and protein molecular weight distributions in soft and hard winter wheats. Genotypes were grown under an irrigation gradient and two N-fertilization levels. Grain polyphenol oxidase (PPO) activity, SDS sedimentation, and Mixograph analyses were evaluated. Flour protein composition was characterized using SE-HPLC. Moisture stress during grain-fill increased flour protein content. N fertilization increased flour protein content. No significant correlation was found between flour protein and PPO. Changes in protein composition were related to general increases in protein content, regardless if the result of reduced irrigation or increased fertilization rate. The percentage of monomeric proteins increased more than the polymeric proteins as flour protein increased. Similarly, SDS sedimentation volume increased as a function of protein content. As expected, subunit GluD1 5+10 was associated with larger sedimentation volume and higher dough strength in genotypes as compared to those with subunit GluD1 2+12. Biplot analyses showed that genotypes of similar protein quality and composition responded similarly to N and irrigation treatments.     

Predicting wheat quality – consequences of the ascorbic acid improver effect

Genotype and environment sets of wheat cultivars and breeding lines were tested for bread making, dough mixing, dough rheology, protein composition and thiol/disulphide composition to find methods that could identify wheat with high baking quality (high-BQ) and moderate work input (moderate-WI) requirement using the mechanical dough development system. Wheat with these properties generally had a high baking response to ascorbic acid (AA) and a GluD1a (HMW-GS 2 + 12) allelic composition. Strong wheat with high WI and high-BQ generally had low baking response to AA and a GluD1d (HMW-GS 5 + 10) allelic composition. Using protein composition data to identify wheat of high-BQ (with AA) and moderate-WI, it was best to select wheat with as high as possible percent of SDS-unextractable polymeric protein (%UPP) in flour and as low as possible %UPP in total polymeric protein. Using a dough extension test for identifying wheat of high-BQ (with AA) and moderate-WI, it was best to select wheat with intermediate values for maximum resistance to extension (Rmax) and for values of extension at Rmax as high as possible within the intermediate Rmax range. Cysteine content of protein fractions and glutathione content of flour gave mostly poor to weak correlations with all baking and mixing properties.     

Quality of shear fractionated wheat gluten – Comparison to commercial vital wheat gluten

The functional properties of gluten obtained with a shear-induced separation process, recently proposed by Peighambardoust et al. (2008), are compared with a commercially available vital wheat gluten. Two tests were performed. First, a relatively strong wheat flour, Soissons, was enriched with gluten protein. The resulting dough was then evaluated on its kneading performance. Second, a weak flour, Kolibri, was enriched to evaluate the baking properties. The wheat flour enriched with gluten protein obtained via the shear-induced separation process (SCG) showed comparable to improved gluten functionality relative to commercial available vital wheat gluten protein (CVWG). The differences in functionality cannot be directly related to the composition as analyzed with SE-HPLC, because the composition of the gluten materials was rather comparable. The differences in functionality may therefore be related to the different drying techniques used or to the inherent mildness of the shear-induced separation technique.     

Introgression of transgenes into a commercial cultivar confirms differential effects of HMW subunits 1Ax1 and 1Dx5 on gluten properties

Transgenes encoding the HMW subunits 1Ax1 and 1Dx5 have been transferred from “model” wheat lines into the commercial French bread wheat cultivar Soissons, using three backcrosses. Five pairs of BC3 expressing and null lines were isolated from each cross and multiplied to provide grain for functionality studies. Analysis of white flour samples confirmed the expression of the transgenes. SE-HPLC and Reomixer studies showed that the two transgenes had differential effects on dough functional properties. Thus, subunit 1Dx5 resulted in detrimental effects on dough development which were associated with decreased extractability of large glutenin polymers. In contrast, lines expressing subunit 1Ax1 contained increased proportions of extractable large glutenin polymers with three lines showing higher torque at similar mixing times (i.e. increased dough strength). This confirms the results obtained with the model wheat lines and shows that the 1Ax1 transgene can be used to increase dough strength in commercial cultivars.     

Incorporation of soy proteins into the wheat–gluten matrix during dough mixing

Farinograph methodology was used to evaluate the possible incorporation of soy proteins into a glutenin–soy complex during mixing and to study the contribution of soy proteins to the chemical and physical properties of the dough. To facilitate the interaction of soy and wheat proteins, a redox process was used, which allowed the partial reduction (using dithiothreitol, DTT) and subsequent reoxidation (using potassium iodate) of glutenin without changing its functionality in the dough (a composite of equal parts of wheat and soy flours, 300 g in total). Either raw soy flour (RSF) or physically modified soy flour (PMSF) was used as the soy component. Dough samples were taken at peak mixing time and at break time during mixing, and these were freeze dried for SE-HPLC analysis and capillary electrophoresis (Lab-on-a-chip).     

Extractability and chromatographic separation of rye (Secale cereale L.) flour proteins

A size exclusion – high performance liquid chromatography (SE-HPLC) method originally developed for separating wheat, barley or rice proteins was applied to study the extractability and molecular weight (MW) distribution of rye flour proteins. These were extracted with 50 mmol/l sodium phosphate buffer (pH 6.8) containing 2.0% (w/v) sodium dodecyl sulfate (SDS) and, optionally, 1.0% (w/v) dithiothreitol (DTT). About 95% of the proteins were extracted in buffer containing 2.0% SDS. Addition of 1.0% DTT to such buffer increased the protein extractability to 100%, indicating that rye flour contains some proteins cross-linked by disulfide (SS) bonds. The SE-HPLC profiles revealed that rye flour contains SS-linked HMW-secalins and 75 k γ-secalins which elute in specific peaks. Upon reduction, these SS-linked protein aggregates dissociate and some entrapped albumins, globulins and/or ω-secalins are released. Rye flour albumins and globulins elute over the entire SE-HPLC profile. In contrast, the monomeric ω-secalins and 40 k γ-secalins are detected in specific well resolved SE-HPLC peaks. The applied fast and reproducible method can be used to characterise and quantify rye flour proteins and to determine changes as a result of processing.     

Relationships Between Mixograph Parameters and Indices of Wheat Grain Quality

A 10-gram Mixograph was used to analyse 39 bread wheat cultivars having various agricultural characteristics and origins. Eleven parameters obtained during the first 8 minutes of the mixing procedure were selected using a repeatability analysis in order to investigate variations between genotypes. Several flour protein fractions characterised by their extractability properties and size distributions were quantified. In addition grain hardness, water-extractable arabinoxylan contents and relative arabinoxylan viscosity were also determined and correlated with dough formation during mixing. Dough consistency, as measured by the height of the Mixogram, was correlated with grain hardness, as expressed by the starch damage content and the proportion (%) of flour particles larger than 37·8 μm. No significant relation was found with the water-extractable arabinoxylan content or the relative arabinoxylan viscosity. The width and height of the Mixogram was related to the proportion of unaggregated proteins before the mixing peak and to polymeric proteins after the dough consistency reached a maximum. The Mixograph proved to be a powerful tool to investigate indices of bread making quality.     

Studies on green gram (Phaseolus aureus) protein concentrate and flour

Protein isolate from green gram (Phaseolus aureus) was prepared and the chemical composition was determined. It contained 64.04% protein, 1.8% total lipids, 27.64% total carbohydrates, 1.68% crude fibre and 4.84% ash. Iron, calcium, magnesium, copper, zinc, potassium and sodium were determined. The limiting amino acid in the protein isolate was lysine. In vitro digestibility pepsin followed the pancretion was the highest and the lowest was the digestion by pepsin alone. Water absorption, oil absorption, emulsion capacity and nitrogen solubility index (NSI) of the protein isolate were 2.26g/g, 1.24g/g, 31.4g/g and 6.8g/g, respectively. For comparison the same functional properties were determined for the flour of green gram. Replacing 5 and 10% of the wheat flour with green gram flour improved the mixing properties of dough and produced good acceptable bread. However, the addition of 15% green gram flour weakened the dough and lowered the quality of bread. Replacing 2.5, 5 and 7.5% of wheat flour with protein concentrate also weakened the mixing properties of the wheat dough and decreased the bread quality.     

Effects of vacuum mixing and mixing time on the processing quality of noodle dough with high oat flour content

The effects of different mixing parameters (vacuum mixing and mixing time) on oat (70% oat flour) and wheat noodle dough were investigated on the basis of textural properties and gluten formation. The results showed that at a vacuum degree of −0.06 MPa and mixing time of 10 min, oat and wheat dough sheets exhibited the highest resistance to extension and glutenin macropolymer (GMP) content, and had the most compact and uniform gluten network. Compared with wheat noodle dough, oat dough had lower resistance to extension, lower tightly bound water content, and higher GMP content. Microstructural examination showed that oat noodle dough had a more aggregated distribution of gluten protein compared with wheat noodle dough under the optimum mixing parameters. Furthermore, the poor binding ability of vital wheat gluten with water molecules caused the indexes of oat noodle dough to be more strongly affected by the changes in mixing parameters than wheat noodle dough.     

Phytate negatively influences wheat dough and bread characteristics by interfering with cross-linking of glutenin molecules

The influence of added phytate on dough properties and bread baking quality was studied to determine the role of phytate in the impaired functional properties of whole grain wheat flour for baking bread. Phytate addition to refined flour at a 1% level substantially increased mixograph mixing time, generally increased mixograph water absorption, and reduced the SDS-unextractable protein content of dough before and after fermentation as well as the loaf volume of bread. The added phytate also shifted unextractable glutenins toward a lower molecular weight form and increased the iron-chelating activity of dough. It appears that phytate negatively affects gluten development and loaf volume by chelating iron and/or binding glutenins, and consequently interfering with the oxidative cross-linking of glutenin molecules during dough mixing. Phytate could be at least partially responsible for the weak gluten network and decreased loaf volume of whole wheat flour bread as compared to refined flour bread.     

Wheat flour non-starch polysaccharides and their effect on dough rheological properties

Wheat (Triticum aestivum L.) flour is able to form dough with unique rheological properties that allow bread making. It is well known that wheat protein content affects dough rheological properties, but there is not enough evidence about the role of other minor flour constituents. One such minor constituent is non-starch flour polysaccharides, which are mainly pentosans formed by a xylopyranosyl linear chain branched with arabinofuranosyl residues. Their spatial distribution and branching pattern can affect their relationship with gluten forming proteins and thus influence their functional properties, the dough rheological properties, and thereby the flour baking quality. In this study the content and structural characteristics of non-starch polysaccharides were investigated, as well as their influence on some dough physicochemical parameters. Five different wheat flours samples milled from Uruguayan wheat varieties with diverse rheological and breadmaking properties were used in this study. Water soluble flour polysaccharides were extracted and the amount of pentosans was determined by the orcinol-HCl method. The pentoses composition was determined before and after acidic hydrolysis of the water soluble polysaccharide fractions by GC. No free pentoses were detected in any of the assayed flour samples, so the pentoses composition found in the hydrolyzed samples was attributed to the non-starch water soluble polysaccharides. Water unextractable non-starch polysaccharides were determined by difference between the total and the soluble non-starch polysaccharides flour content.An improved method for the quantification of water extractable and non-extractable non-starch polysaccharides, using baker's yeast, was developed. Using this method, total and soluble non-starch polysaccharides content could reliably be determined both in whole flour and in pentosans enriched fractions. Free monosaccharide content was in the range from 0.03% to 0.06% (w/w), while the arabinose/xylose (Ara/Xyl) ratios varied from 0.8 to 1.4 in soluble non-starch polysaccharides and from 0.7 to 0.9 in total non-starch polysaccharides. The different Ara/Xyl ratios found for water extractable and unextractable arabinoxylans clearly indicates different substitution degrees in the polymers. Analysis of the dough rheological parameters in relation to the water soluble and non-soluble non-starch polysaccharides and the Ara/Xyl ratios from different wheat varieties was performed. A clear relation between some of these parameters could be inferred, since a direct relationship between total unextractable (AXi) content and resistance of dough to extension (P), as well an inverse relationship between the same parameter and dough extensibility (L) were observed. These results suggest that the flour non-starch polysaccharide content, as well the Ara/Xyl ratios may be used as additional parameters to estimate some of the wheat flours dough properties.     

The influence of different nitrogen treatments on the size distribution of protein fractions in hard and soft wheat

The relative quantity of specific proteins, protein subunits, as well as amount and size-distribution of polymeric proteins in wheat kernels may vary due to environmental conditions. In this study, the effect of different nitrogen treatments on polymeric and monomeric proteins in wheat was determined. Two soft white biscuit wheat cultivars, a cracker wheat and a hard red bread wheat were planted under irrigation in a randomized complete block design with three replications in two successive years at six different nitrogen treatments. SE-HPLC was used to determine the amount of monomeric and polymeric proteins, and various quality characteristics were measured after harvesting. The large and small SDS-extractable polymeric proteins were not influenced by different N levels. There was a strong cultivar influence, where some cultivars had a larger reaction to N treatments than others. The later application (at flag leaf stage) of nitrogen did not increase the protein fractions. The total amount of N given seemed to have a greater influence on the protein fractions than the timing of the fertilizer application. The lowest N treatment consistently gave the lowest flour protein content value. There was a strong correlation between flour protein content and large monomeric proteins.     

The influence of abiotic stress conditions on dough mixing characteristics of two hard red spring wheat cultivars

Two commercial hard red spring wheat cultivars were exposed to high and low temperatures, as well as drought stress when the main tiller kernels were at the soft dough stage. The trial was done in the greenhouse for two consecutive seasons to determine the effects of these stress conditions on protein content, SDS sedimentation and selected Mixsmart characteristics. Heat stress had the largest effect on mixing characteristics. Heat and drought stress caused a significant increase in flour protein content of both cultivars and had similar effects on mixing characteristics. The Mixsmart characteristics associated with dough strength were increased by heat and drought stress. Cold stress caused a slight increase in protein content of the cultivars, but in general caused a reduction in dough strength as measured with Mixsmart characteristics. The reaction of Mixsmart characteristics to heat and drought stress was much larger in Duzi than in Kariega, confirming that there is a large genotype effect in rheological characteristics.     

A parametric and microstructural study of the formation of gluten network in mixed flour–water batter

The mechanism of gluten network development is still unclear and remains difficult to study since gluten network formation in bread dough is a rather quick process. In order to better characterize this dynamic event, we slowed down its kinetics by increasing the dough water content. During mixing, performed with a planetary mixer at variable mixing speeds and flour/water ratios, the torque was recorded. Common flours from wheat cultivars Orvantis, Caphorn and Isengrain, similar in composition and Farinograph parameters, were studied.     

Relationship of Protein Quantity,Quality and Dough Properties with Chinese Steamed Bread Quality

Using selected Chinese and Australian wheats, flour protein content and composition of high-molecular-weight (HMW) glutenin subunits were studied in relation to northern style Chinese steamed bread quality. Flour protein content had a significant impact on Chinese steamed bread quality. The Chinese wheats were characterised by shorter Farinograph dough development time and stability in comparison with the Australian wheats. Dough stickiness in the Chinese wheat cultivars was a significant factor deteriorating Chinese steamed bread quality. A significant negative correlation was found between Farinograph stability time and steamed bread quality in Australian wheats while a significant and positive correlation existed in Chinese wheats. It would be necessary to increase the dough strength of Chinese wheat cultivars in order to improve their steamed bread making quality.     

Mixing behaviour of a zero-developed dough compared to a flour–water mixture

The Z-blade mixing behaviour of zero-developed (ZD) doughs from the flours of two wheat cultivars of different gluten strength was compared to that of conventionally mixed dough made from the same flours. In farinograph experiments, use of ZD dough led to shorter development time (with less energy requirement), less stability time, and consequently earlier breakdown compared to conventional mixing of the corresponding flour–water mixture. Mixing of ZD doughs led to an almost similar decrease of glutenin macro-polymer (GMP) wet weight as that of doughs prepared from flour–water mixtures. However, comparison of wet weight of re-assembled GMP revealed that until time-to-peak (TTP) mixing, there was no difference in GMP recovery with respect to the starting material used in the z-blade mixing experiments. Beyond TTP, recovery of GMP in doughs prepared from both starting materials was reduced. The results of large-strain deformation rheology showed strong visco-elastic behaviour as characterised by the highest values of fracture properties (except ε_H), followed by a decline in those properties upon further mixing for doughs mixed from both flour–water mixture and ZD dough from both types of wheat cultivars. It was concluded that at mixing regimes before TTP, there was no difference between ZD doughs and flour–water mixtures in the mixer. When ZD dough is used as a starting material for dough preparation instead of flour, extra care should be taken not to over-mix the developing dough.     

Effect of incorporation of an i-type low-molecular-weight glutenin subunit and a modified γ-gliadin in durum and in bread wheat doughs as measured by micro-mixographic analyses

The effects of incorporation of an i-type low-molecular-weight glutenin subunit (LMW-i) and of a modified γ-gliadin showing an additional cysteine residue, on 2 g Mixograph parameters of durum (biotypes 42 and 45 of the Italian cv. Lira) and bread wheat (Australian cv. Kukri) doughs were studied. In bread wheat flour incorporation of the modified γ-gliadin resulted in a significant decrease in dough strength (decreased mixing time and peak resistance), but at the same time it produced a slight increase in dough stability (decreased resistance to breakdown). The incorporation of the LMW-i type into bread wheat dough had minimal effects on dough mixing requirements. The incorporation of both LMW-i type and modified γ-gliadin in durum wheat doughs produced a significant decrease in the overall dough strength, especially in Lira 45 biotype doughs. Reversed phase high-performance liquid chromatography (RP-HPLC), size exclusion high-performance liquid chromatography (SE-HPLC) and two-dimensional gels analyses of control and reconstituted semolina doughs showed that the two polypeptides were in the polymeric fraction. The effect of the incorporation of the two polypeptides in durum and bread wheat doughs showed remarkable differences and the reasons for this is discussed in terms of both intrinsic differences between wheat flour and durum semolina and in methodological approaches.     

Understanding the link between GMP and dough: from glutenin particles in flour towards developed dough

Clear correlations exist for glutenin macropolymer (GMP) quantity and rheological properties vs. wheat quality and dough rheological properties, but real insight in understanding these links is still missing. The observation that GMP consists of glutenin particles opens up new possibilities to reveal the underlying mechanism linking glutenin network properties with dough preparation. GMP was isolated from flour of three wheat varieties: Estica, Soissons and Baldus, strongly varying in their mixing requirements (expressed as time-to-peak, TTP). Decrease of GMP quantity and G′ vs. mixing energy was confirmed. More detail was obtained by studying the changes in GMP particles when mixing flour into dough. Mixing leads to a decrease in the average size of the particles. Interestingly, the TTP coincided with the work-input at which all particles just became soluble in SDS. At TTP, the average size of the GMP particles was the same for each variety. During mixing particles lost their globule shapes and appeared ruptured. Particle size analysis confirmed that particles were still present near TTP. Analysis of the change in particle size vs. energy input using physical principles revealed the following: (1) mixing energy is the predominant actuator in decreasing GMP particle size; (2) the initial GMP particle size in flour strongly determines the practical mixing requirements; and (3) the derived mixing energy vs. GMP particle size relationship was shown to be applicable for both Mixograph and Farinograph mixing. Our results demonstrate that, for the flour samples used, glutenin particle size determines TTP and GMP rheology, showing that glutenin particle properties could be a new key to understand the link between GMP and dough properties.     

Development of a standard test for dough-making properties of oat cultivars

Bread is consumed all over the world. However, so far, production of large volume bread is only possible with wheat. Alternatives, such as oats, are less suitable but this is partly due to the lack of knowledge about their functionality for other purposes than porridge, which is their most common use. Existing standard tests for the dough making characteristics of wheat flour are not suitable for oat flour, hampering research to optimize oats for bread-making purposes. We therefore set out to develop a test to evaluate oat in relation to mixing and dough making properties using wheat as a model. It was possible to reproduce the profile of various qualities of wheat flour using mixtures of oat flour and gluten in different proportions. Our standard test was based on a dough system composed of 87.2% oat flour and 12.8% gluten and it presented similar properties to a wheat flour with regard to resistance to extension. This dough system was sensitive and reliable (coefficient of variation lower than 10%) for detecting differences among oat cultivars, and it can be used to screen oat varieties and individual oat components in relation to relevant properties for bread-making purposes.