Impact of lipases with different substrate specificity in wheat flour separation on the properties of the resultant gluten

Wheat gluten was isolated in a laboratory dough-batter flour separation process in the presence or absence of lipases differing in hydrolysis specificity. The obtained gluten was blended with wheat starch to obtain gluten-starch (GS) blends of which the water and oil binding capacities were investigated. Furthermore, GS blends were mixed into dough and processed into model breads, of which dough extensibility and loaf volume were measured, respectively. In comparison to GS blends prepared with control gluten, oil binding capacity was higher when GS blends contained gluten isolated with Lecitase Ultra (at 5.0 mg enzyme protein/kg flour), a lipase hydrolyzing both non-polar and polar lipids. Additionally, dough extensibility and total work needed for fracture were lower for dough prepared from GS blends containing gluten isolated with Lipolase (at 5.0 mg enzyme protein/kg flour), a lipase selectively degrading non-polar lipids. In GS blend bread making, this resulted in inferior loaf volumes. Comparable GS blend properties were measured when using control gluten and gluten isolated with YieldMAX, a lipase mainly degrading N-acyl phosphatidylethanolamine. In conclusion, properties of GS blend model systems are altered when gluten prepared in the presence of lipases is used to a degree which depends on lipase specificity and concentration.     

A single amino acid substitution in puroindoline b impacts its self-assembly and the formation of heteromeric assemblies with puroindoline a

Puroindolines (PINs) A and B were purified from soft (Paledor) and hard (Recital, Courtot) wheat cultivars. Their purity and heterogeneity due to post-translational processing were characterized by SDS- and acid-PAGE, reversed-phase HPLC and mass spectrometry. By using dynamic light scattering (DLS), asymmetrical flow field-flow fractionation (AF4) and size exclusion chromatography (SEC), we showed that the size distributions of PINA are similar for the three varieties and that, in solution, they self-assembled into small aggregates, mainly dimers. Conversely, PINB isolated from hard varieties (PINB-D1b and PINB-D1d) are assembled into large aggregates while PINB-D1a formed small aggregates, mainly monomers. Mixed solutions of PINA and PINB formed heteromeric aggregates. The large PINB-D1b aggregates were retained even at a high (4:1) PINA/PINB weight ratio. Reversible dissociation of large aggregates into small aggregates suggested that weak interactions control the self-assembly of PINs. The aggregative properties of PINs have now to be taken into account when studying their interactions with other components to decipher the causal relationships between these proteins and grain hardness.     

Whole wheat bread: Effect of bran fractions on dough and end-product quality

Consumption of whole-wheat based products is encouraged due to their important nutritional elements that benefit human health. However, the use of whole-wheat flour is limited because of the poor processing and end-product quality. Bran was postulated as the major problem in whole wheat breadmaking. In this study, four major bran components including lipids, extractable phenolics (EP), hydrolysable phenolics (HP), and fiber were evaluated for their specific functionality in flour, dough and bread baking. The experiment was done by reconstitution approach using the 2⁴ factorial experimental layout. Fiber was identified as a main component to have highly significant (P < 0.05) and negative influence on most breadmaking characteristics. Although HP had positive effect on farinograph stability, it was identified as another main factor that negatively impacted the oven spring and bread loaf volume. Bran oil and EP seemed to be detrimental to most breadmaking characteristics. Overall, statistical analysis indicates that influence of the four bran components are highly complex. The bran components demonstrate multi-way interactions in regards to their influence on dough and bread-making characteristics. Particularly, Fiber appeared to have a high degree of interaction with other bran components and notably influenced the functionality of those components in whole wheat bread-making.     

The effect of heat treatment on the soluble protein content of oats

The effect of heat treatment on the soluble protein content in oat groats (Kerstin commercial variety) was evaluated using asymmetric flow field-flow fractionation (AF4) in combination with online multiangle light scattering (MALS) and UV detection. The AF4 method was used to separate the monomeric proteins from globulin hexamer and aggregate proteins and β-glucan polysaccharides in the soluble oat protein fraction. The total amount of soluble protein (with respect to total protein) was reduced to 35.7 ± 4.5 wt. % in heat treated oats from 74.6 ± 5.3 wt. % in non-heat treated oats. The ratio of monomeric to globulin hexamer and aggregate proteins was reduced from 1.82 to 1.48 as a result of heat treatment. Sodium-dodecyl-sulfate polyacrylamide gel electrophoresis (SDS-PAGE) analysis revealed the selective elimination of protein bands associated with the albumin and prolamin protein fractions as a result of heat treatment. These results were supported through amino acid analysis by cation exchange chromatography coupled with UV detection which revealed a reduction in amino acid residues associated with prolamin. The globulin proteins were found to be less sensitive to heat treatment.     

Interplay between starch and proteins in waxy wheat

Most of the unique properties of waxy wheat have been associated with the lack of amylose, that in turn may affect the mutual interactions between starch and proteins. To address this particular aspect, we carried out molecular, rheological, and calorimetric studies on flours from two waxy wheat lines that were compared with a non-waxy one. Dough thermal properties and water binding capacity were investigated by Differential Scanning Calorimetry (DSC) and by thermogravimetric analysis, respectively. Protein solvation, aggregation, and thiol accessibility were also investigated, together with dough mixing properties and stickiness. Proteins in waxy wheat samples needed more water to complete solvation, likely because of the water-retaining capacity of waxy wheat starch. In waxy wheat dough, water was tightly bound to starch, and DSC studies indicated an increase in gelatinization temperature. Moreover, the low water mobility in waxy wheat resulted in low and retarded gluten hydration and in high stickiness. In samples with the highest stickiness, protein aggregates were stabilized mainly by hydrophobic interactions. Differences between waxy wheat lines may be attributed to a different structural organization of components within each class of biopolymers.     

The impact of redox agents on further dough development,relaxation and elastic recoil during lamination and fermentation of multi-layered pastry dough

The role of gluten proteins during lamination and fermentation of multi-layered wheat flour pastry dough was examined by including oxidizing or reducing agents in the recipe to respectively strengthen or weaken the gluten protein network. Pastry burst rig textural measurements showed that dough strength increases during lamination up to 16 fat layers. However, further lamination up to 64 and 128 fat layers decreases the dough strength, most likely due to destruction of layer integrity. Redox agents strongly affect dough strength. Furthermore, fermentation and spread tests showed that they strongly influence elastic recoil immediately after lamination and during relaxation. Moreover, elastic recoil consistently occurs to a greater extent in the final direction of sheeting. None of the observed changes in dough strength and relaxation behaviour could be linked to changes in the levels of protein extractable in sodium dodecyl sulfate containing medium (SDS-EP). This suggests that changes occur preferentially either within the SDS-extractable or within the non-SDS-EP fraction and that they do not render non-extractable protein fractions extractable or vice versa. Furthermore, elastic recoil is most likely caused by reformation of inter- and intramolecular hydrogen bonds and hydrophobic interactions.     

Dough rheological properties and noodle-making performance of non-waxy and waxy whole-wheat flour blends

Dough rheological properties and noodle-making performance of non-waxy whole-wheat flour (WWF) with partial- or full-waxy (PW- or FW-) WWF substitution were studied. The substitution levels were 0, 250, 500, 750, and 1000 g/kg, respectively. FW-WWF reduced the peak viscosity and pasting temperature of WWF blends as its substitution level was increased due to its higher proportions of B-type starch granules and short amylopectin chains, while PW-WWF increased peak viscosity with the increasing substitution level because of its higher amylopectin content. As demonstrated by farinograph and rheometer measurements, FW-WWF interfered with gluten development because of the increased competition for water by arabinoxylans and amylopectin; however, PW-WWF enhanced dough strength due primarily to its increased protein content. Consequently, FW-WWF showed a detrimental effect on cooked noodle texture as the cooked noodle hardness was reduced by 50% at the 1000 g/kg substitution level. In contrast, PW-WWF enhanced noodle integrity and elasticity by increasing cooked noodle cohesiveness and resilience by 10.1% and 14.8%, respectively, at the 1000 g/kg substitution level. The results suggest that with waxy WWF substitution, the changes in starch composition, arabinoxylans, and protein content could modify the interactions among flour components and influence the quality characteristics of noodle products.     

Effects of post-anthesis fertilizer on the protein composition of the gluten polymer in a US bread wheat

Both genetic and environmental factors influence the types and amounts of wheat proteins that link together to form polymers essential for flour quality. To understand how plant growth conditions might influence gluten polymer formation, protein fractions containing small and large polymers were separated from flour from the US wheat Butte 86 grown in the absence or presence of post-anthesis fertilizer. Proteins in the polymer fractions were analyzed by quantitative two-dimensional gel electrophoresis (2-DE). The ratio of high molecular weight glutenin subunits (HMW-GS) to low molecular weight glutenin subunits (LMW-GS) increased in both fractions in response to fertilizer, due in part to small increases in the proportions of individual HMW-GS. There were also changes within the LMW-GS. In particular, omega and alpha chain terminators increased in proportion in both polymer fractions, but changes were more pronounced in the large polymer fractions. Serpins also increased in both polymer fractions. Additionally, the study revealed differences in the proportions of traditional LMW-GS in small and large polymer fractions. LMW-s type proteins were more abundant in the large polymers while LMW-i type proteins were more prevalent in the small polymers, suggesting that these proteins may play different roles in the gluten polymer.     

Thermal degradation kinetics of bioactive compounds from black rice flour (Oryza sativa L.) extracts

The effect of thermal processing on the degradation of the phytochemicals in black rice flour by means of fluorescence spectroscopy and degradation kinetics was investigated. In order to investigate the influence of food matrices, a comparative analysis between integral rice flour and different fractions was performed. The preliminary compositional results suggested a higher content in phytochemicals in fraction four of the seven fractions of black rice flour, which was sifting through a sieve with a diameter of 180 μm. The compositional complexity was highlighted by fluorescence spectroscopy. The heat-treatment caused structural changes that led to red- or blue-shifts in maximum emission. The first-order kinetic model was used to describe the mechanism of degradation. The activation energies were 10.07 ± 1.04 kJ/mol for total polyphenolic, 7.26 ± 0.58 kJ/mol for total monomeric anthocyanins and 6.71 ± 1.12 kJ/mol for antioxidant activity in case of integral flour extract. For fraction four extract obtained by, the E_a values were: 3.51 ± 0.53 kJ/mol, 11.49 ± 1.47 kJ/mol, 15.80 ± 1.50 kJ/mol and 19.91 ± 3.27 kJ/mol, respectively. The calculated values of the activation energy revealed higher temperature dependence of total polyphenols in integral flour and of antioxidant activity in fraction four, respectively.     

Influence of the cultivar,environment and management on the grain yield and bread-making quality in winter wheat

The genotype, environment and their interaction play an important role in the grain yielding and grain quality attributes. The main aim of this study was to determine the contributions of the genotype, environment and their interaction to the variation in bread-making traits. The data that were used for the analyses performed in this study were obtained from 3 locations in Poland from post-registration multi-environment trials with winter wheat in 2009 and 2010. The experimental factors were the cultivar (7 cultivars) and the crop management level (low input and high input). In the multi-environment trials, 17 traits were investigated that characterize grain, flour and dough quality. Most of the traits were affected much more strongly by environmental factors (i.e., year and location) than by genotype. The variance components revealed an especially strong effect of the year on the baking score, loaf volume and water absorption, as well a strong effect of the location on dough development and protein content. The obtained results demonstrate that the grain quality as measured by the parameters based on the protein content and quality may be substantially improved by crop management practices, especially by N fertilization level.     

Effect of allelic variation at the Glu-3/Gli-1 loci on breadmaking quality parameters in hexaploid wheat (Triticum aestivum L.)

Low molecular weight glutenin subunits (LMW-GS) encoded by the Glu-3 loci are known to contribute to wheat breadmaking quality. However, the specific effect of individual Glu-3 alleles is not well understood due to their complex protein banding patterns in SDS-PAGE and tight linkage with gliadins at the Gli-1 locus. Using DNA markers and a backcross program, we developed a set of nine near isogenic lines (NILs) including different Glu-A3/Gli-A1 or Glu-B3/Gli-B1 alleles in the genetic background of the Argentine variety ProINTA Imperial. The nine NILs and the control were evaluated in three different field trials in Argentina. Significant genotype-by-environment interactions were detected for most quality parameters indicating that the effects of the Glu-3/Gli-1 alleles are modulated by environmental differences. None of the NILs showed differences in total flour protein content, but relative changes in the abundance of particular classes of proteins cannot be ruled out. On average, the Glu-A3f, Glu-B3b, Glu-B3g and Glu-B3i_Man alleles were associated with the highest values in gluten strength-related parameters, while Glu-A3e, Glu-B3a and Glu-B3i_Chu were consistently associated with weak gluten and low quality values. The value of different Glu-3/Gli-1 allele combinations to improve breadmaking quality is discussed.     

Modification of protein structure and dough rheological properties of wheat flour through superheated steam treatment

Native (NF, 13.5% w.b) and moistened (MF, 27% w.b) wheat flours were treated with superheated steam (SS) at 170 °C for 1, 2 and 4 min, and their protein structure as well as dough rheological properties were analyzed. Confocal laser scanning microscopy (CLSM) and SDS-PAGE patterns indicated the formation of protein aggregates with reduced SDS extractability after treatment. Farinograph and dynamic rheometry measurements showed that the strength as well as elastic and viscous moduli of the dough made from SS-treated flours progressively increased with SS treatment time. And both the improvements were more pronounced for superheated steam-treated moistened flours (SS-MF) than for superheated steam-treated native flours (SS-NF). Size-exclusion high performance liquid chromatography (SE-HPLC) analysis demonstrated that dough rheological parameters have positive correlations with SDS unextractable polymeric proteins (UPP) contents. SS treatment on flours led to a transition of protein secondary structures to more ordered form (α-helix and β-sheet). Additionally, free sulfhydryl (SH) contents decreased after treatment, which implied that disulfide bonds accounted for protein extractability loss and dough rheological properties improvement. Elevated moisture level promoted the modification of both protein structure and dough behaviors of flours during SS treatment.     

Combined mutations in five wheat STARCH BRANCHING ENZYME II genes improve resistant starch but affect grain yield and bread-making quality

Increases in the proportion of amylose in the starch of wheat grains result in higher levels of resistant starch, a fermentable dietary fiber associated with human health benefits. The objective of this study was to assess the effect of combined mutations in five STARCH BRANCHING ENZYME II (SBEII) genes on starch composition, grain yield and bread-making quality in two hexaploid wheat varieties. Significantly higher amylose (∼60%) and resistant starch content (10-fold) was detected in the SBEII mutants than in the wild-type controls. Mutant lines showed a significant decrease in total starch (6%), kernel weight (3%) and total grain yield (6%). Effects of the mutations in bread-making quality included increases in grain hardness, starch damage, water absorption and flour protein content; and reductions in flour extraction, farinograph development and stability times, starch viscosity, and loaf volume. Several traits showed significant interactions between genotypes, varieties, and environments, suggesting that some of the negative impacts of the combined SBEII mutations can be ameliorated by adequate selection of genetic background and growing location. The deployment of wheat varieties with increased resistant starch will likely require economic incentives to compensate growers and millers for the significant reductions detected in grain and flour yields.     

Modeling the effects of microfluidization conditions on properties of corn bran

Corn bran was microfluidized through a 200-μm channel in the pressure range of 124.1–158.7 MPa for 1–5 passes following the central composite experimental design. Physicochemical properties and antioxidant properties of microfluidized bran samples were measured and fitted to the second order polynomial model. The response surface equations obtained showed that all the properties examined had a positive linear relationship with pressure and a negative quadratic relationship with number of passes except for ABTS radical scavenging activity which was quadratically related to both processing parameters. The number of passes generally had a more pronounced effect on the examined properties compared with pressure. Within the experimental range, the maximum values of swelling capacity, water-holding capacity, and oil-holding capacity were respectively 10.62 ml/g d.w. (at 158.7 MPa), 5.49 g water/g d.w. (at 158.7 MPa), and 4.61 g oil/g d.w. (at 124.1 MPa); the maximum values of surface reactive phenolic content, DPPH and ABTS radical scavenging activities were 148.80 mg/FAE g d.w. (at 158.7 MPa), 50.02 μmol TE/g d.w. (at 158.7 MPa), and 47.90 μmol TE/g d.w. (at 145.9 MPa), respectively. All maximum values of the properties occurred at 5 passes.     

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.     

Oat protein solubility and emulsion properties improved by enzymatic deamidation

Oat protein produced by a dry milling process was enzymatically deamidated by a food-grade protein-glutaminase (PG), and the effects on structure, solubility and emulsifying properties of oat proteins were studied. The reactions were conducted at neutral pH and low salt concentration conditions. Oat proteins were deamidated up to a deamidation degree of 59%. The solubility of proteins doubled. Oil-in-water emulsions prepared with the native and deamidated oat proteins differed as the emulsions prepared from oat proteins with high deamidation degree had a more uniform oil droplet particle size and longer stability. Fourier transform infrared (FT-IR) analysis demonstrated that oat protein secondary structure became more flexible by deamidation. Protein-glutaminase mediated deamidation appeared to be a promising technique to improve oat protein functionality such as emulsifying ability and solubility.     

Characterizing the pigmented traditional rice cultivars grown in temperate regions of Kashmir (India) for free and bound phenolics compounds and in vitro antioxidant properties

The purpose of the research was to identify the phenolic and flavonoid compounds of seven different traditional pigmented whole rice cultivars grown in the temperate regions of Kashmir so as to study their relationship with in vitro antioxidant capacities. The completely pigmented rice cultivars were found to have higher phenolic, flavonoid, anthocyanin contents and exhibited higher antioxidant capacities than the light colored and sparely colored rice cultivars. A total of 40 compounds had been identified in the analyzed rice cultivars that were found to be distributed in 6 major categories with 6-phenolics, 6-flavonoids, 11-hydroxycinnamic acid derivatives, 7-hydroxybenzoic acid derivatives, 3-anthocyanins and 7-flavonoid glucosides of different flavonoid compounds. Among the free and bound fractions for each cultivars the light and sparsely colored depicted higher content of phenolics and in vitro antioxidant properties in bound faction, while the completely pigmented cultivars showed higher antioxidant properties in free fractions. The anthocyanins quercetin-3-O-galactoside, cyanidin-3-O-rutinoside and pelargonidin-3-O-diglucoside had been identified by LC-MS existing in the free fractions of the analyzed rice cultivars whereas, the free fraction of acetone + H₂O possessed higher percentage of phenolic compounds as compared to methanolic extracts and bound fractions. The black colored cultivars possessed higher DPPH scavenging activity and lipid peroxidation inhibition.     

Dehydration of gluten matrix as a result of dietary fibre addition – A study on model flour with application of FT-IR spectroscopy

Nowadays, consumers demand dietary fibre-enriched products of appropriate taste, texture, smell and appearance. Unfortunately, addition of the dietary fibre supplements to bread significantly reduces its quality which is connected with changes in the structure of gluten proteins. Structural changes as well as changes in the water state of gluten matrix induced by eight dietary fibres were observed by using Fourier transform infrared spectroscopy. To facilitate this the difference spectra were calculated by subtraction of the control (gluten only) infrared spectrum from the spectra of gluten-fibre mixtures. The presence of positive bands at ca. 1597 and 1235 cm⁻¹ indicated aggregation of gluten proteins into hydrogen bonded β-sheets. These β-sheets can be formed by other β-sheets, antiparallel-β-sheets, β-turns and/or α-helices. The aggregation is probably induced by partial dehydration of gluten matrix due to competition for water molecules between gluten proteins and fibre polysaccharides. This assumption is confirmed by the presence of the negative band at 3237 cm⁻¹ and decrease in the intensity of the band at 3051 cm⁻¹. These bands are assigned to the weak and strong H-bonds in the gluten matrix, respectively. The results indicated that both weak and strong H-bonds are necessary to dough formation of adequate rheological properties.     

Effects of drought on the morphological and physicochemical characteristics of starch granules in different elite wheat varieties

In this study, scanning electron microscopy (SEM) revealed the formation of pits and pores on the surfaces of starch granules in response to drought stress, with substantially more pronounced effects in the ordinary yield potential wheat cv. Xindong 23 than the excellent yield potential wheat cv. Xindong 20. Drought induced a significant reduction in starch granule sizes in both wheat varieties, though the reduction observed in Xindong 23 was six times more pronounced than that observed for Xindong 20. Amyloglucosidase and α-amylase treatment of starch from wheat grown in drought conditions released significantly more reducing sugars compared with samples from irrigated controls. SEM and confocal laser scanning microscopy (CLSM) revealed that starch granules from the two wheat varieties grown under drought conditions had substantially increased fluorescence after treatment with proteolytic enzymes and staining with methanolic merbromin and 3-(4-carboxybenzoyl) quinoline-2-carboxaldehyde dyes. Analysis of pasting properties showed significant increases of peak viscosity, trough viscosity, break down, and setback following drought stresses. Furthermore, drought induced a significant reduction in the water binding capacity and increased damage to starch only in Xindong 23. These results provide insight into the potential mechanisms through which drought influences the ultrastructures and physicochemical properties of starch in wheat.