Effects of partial hydrolysis and subsequent cross-linking on wheat gluten physicochemical properties and structure.

The rheological behavior and thermal properties of wheat gluten following partial hydrolysis using Alcalase and subsequent microbial transglutaminase (MTGase) cross-linking were investigated. The wheat gluten storage modulus (G') and thermal denaturation temperature (Tg) were significantly increased from 2.26 kPa and 54.43°C to 7.76 kPa and 57.69°C, respectively, by the combined action of partial hydrolysis (DH 0.187%) and cross-linking. The free SH content, surface hydrophobicity, and secondary structure analysis suggested that an appropriate degree of Alcalase-based hydrolysis allowed the compact wheat gluten structure to unfold, increasing the ß-sheet content and surface hydrophobicity. This improved its molecular flexibility and exposed additional glutamine sites for MTGase cross-linking. SEM images showed that a compact 3D network formed, while SDS-PAGE profiles revealed that excessive hydrolysis resulted in high-molecular-weight subunits degrading to smaller peptides, unsuitable for cross-linking. It was also demonstrated that the combination of Alcalase-based partial hydrolysis with MTGase cross-linking might be an effective method for modifying wheat gluten rheological behavior and thermal properties.

Nanostructural morphology of plasticized wheat gluten and modified potato starch composites: relationship to mechanical and barrier properties.

In the present study, we were able to produce composites of wheat gluten (WG) protein and a novel genetically modified potato starch (MPS) with attractive mechanical and gas barrier properties using extrusion. Characterization of the MPS revealed an altered chain length distribution of the amylopectin fraction and slightly increased amylose content compared to wild type potato starch. WG and MPS of different ratios plasticized with either glycerol or glycerol and water were extruded at 110 and 130 °C. The nanomorphology of the composites showed the MPS having semicrystalline structure of a characteristic lamellar arrangement with an approximately 100 Å period observed by small-angle X-ray scattering and a B-type crystal structure observed by wide-angle X-ray scattering analysis. WG has a structure resembling the hexagonal macromolecular arrangement as reported previously in WG films. A larger amount of ß-sheets was observed in the samples 70/30 and 30/70 WG-MPS processed at 130 °C with 45% glycerol. Highly polymerized WG protein was found in the samples processed at 130 °C versus 110 °C. Also, greater amounts of WG protein in the blend resulted in greater extensibility (110 °C) and a decrease in both E-modulus and maximum stress at 110 and 130 °C, respectively. Under ambient conditions the WG-MPS composite (70/30) with 45% glycerol showed excellent gas barrier properties to be further explored in multilayer film packaging applications.

Wheat dough microstructure: the relation between visual structure and mechanical behavior.

The microstructure of food matrixes, and specifically that of wheat-flour dough, determines mechanical behavior. Consequently, the analysis of such microstructure is both necessary and useful for understanding the physico-chemical and mechanical alterations during the production of cereal-based products such as breads. Confocal laser scanning microscopy (CLSM) is an established tool for the investigation of these matrix properties due to its methodical advantages such as easy preparation and handling, and the high depth resolution due to the optical sectioning of probes. This review focuses on the microstructure of wheat-flour dough from a mechanical and visual point of view. It provides an overview of the dependencies between the visibly detectable microstructural elements achieved by CLSM and the physical determined rheological properties. Current findings in this field, especially on numerical microstructure features, are described and discussed, and possibilities for enhancing the analytical methodology are presented.

On characterization of anisotropic plant protein structures.

In this paper, a set of complementary techniques was used to characterize surface and bulk structures of an anisotropic Soy Protein Isolate (SPI)-vital wheat gluten blend after it was subjected to heat and simple shear flow in a Couette Cell. The structured biopolymer blend can form a basis for a meat replacer. Light microscopy and scanning electron microscopy provided a detailed view of structure formation over the visible surfaces of the SPI-gluten blend. Protein orientation in the direction of the flow was evident and fibrous formation appeared to exist on the macro- and micro-scale. Furthermore, according to texture analysis, the structured biopolymer obtained from the Couette Cell after processing at 95 °C and 30 RPM for 15 min has high tensile stress and strain anisotropy indices (∼2 and ∼1.8, respectively), comparable to those of raw meat (beef). The novel element in this work is the use of the neutron refraction method, utilizing spin-echo small angle neutron scattering (SESANS), to provide a look inside the anisotropic biopolymer blend complementing the characterization provided by the standard techniques above. With SESANS, it is possible to quantify the number of fibre layers and the orientation distribution of fibres. For a specimen thickness of 5 mm, the obtained number of fibre layers was 36 ± 4 and the standard deviation of the orientation distribution was 0.66 ± 0.04 radians. The calculated thickness of one layer of fibres was 138 µm, in line with SEM inspection.

Thermal conductivity and combustion properties of wheat gluten foams.

Freeze-dried wheat gluten foams were evaluated with respect to their thermal and fire-retardant properties, which are important for insulation applications. The thermal properties were assessed by differential scanning calorimetry, the laser flash method and a hot plate method. The unplasticised foam showed a similar specific heat capacity, a lower thermal diffusivity and a slightly higher thermal conductivity than conventional rigid polystyrene and polyurethane insulation foams. Interestingly, the thermal conductivity was similar to that of closed cell polyethylene and glass-wool insulation materials. Cone calorimetry showed that, compared to a polyurethane foam, both unplasticised and glycerol-plasticised foams had a significantly longer time to ignition, a lower effective heat of combustion and a higher char content. Overall, the unplasticised foam showed better fire-proof properties than the plasticized foam. The UL 94 test revealed that the unplasticised foam did not drip (form droplets of low viscous material) and, although the burning times varied, self-extinguished after flame removal. To conclude both the insulation and fire-retardant properties were very promising for the wheat gluten foam.

Observation particle morphology of colloidal system by conventional SEM with an improved specimen preparation technique.

On the basis of our previous report that polymer emulsion with different viscosity can be investigated by conventional scanning electron microscopy (SEM), we have developed an improved specimen preparation technique for obtaining particle morphology and size of colloidal silver, collagen, glutin, and polymer microspheres. In this study, we expect to provide a means for charactering the three-dimensional surface microstructure of colloidal particles. Dilution of the samples with appropriate volatile solvent like ethanol is effective for SEM specimen preparation. At a proper ratio between sample and ethanol, the colloidal particles are dispersed uniformly in ethanol and then deposited evenly on the substrate. Different drying methods are studied to search a proper drying condition, in which the small molecule solvent is removed without destroying the natural particle morphology. And the effects of ethanol in the specimen preparation process are described by analyzing the physicochemical properties of ethanol. The specimen preparation technique is simple and can be achieved in common laboratory for charactering the particle morphology of colloidal system.

A novel globular protein electrospun fiber mat with the addition of polysilsesquioxane.

The aim of this work has been to elaborate well defined gliadin nanofibers with incorporation of inorganic molecules, such as polyhedral oligomeric silsesquioxane (POSS). Nanofibers were obtained by electrospinning processing, controlling the relevant parameters such as tip-to-collector distance, voltage and feed rate. The fiber mats were characterized by SEM, confocal images, DSC, viscosity, FTIR and conductivimetry analysis. FTIR spectra showed characteristic absorption bands related to the presence of POSS-NH(2) within the matrices. SEM micrographs showed that gliadin fibers decreased their dimensions as the amount of POSS-NH(2) increased in the spinning solution. The electrical conductivity of gliadin solutions diminished as the concentration of POSS-NH(2) was increased. Besides, confocal micrographs revealed that POSS-NH(2) might be dispersed as nanocrystals into gliadin and gluten fibers. The dimension of gluten nanofibers was also affected by the POSS-NH(2) concentration, but conversely, this dependence was not proportional to the POSS-NH(2) amount. Somehow, the interaction between gliadin and POSS-NH(2) in aqueous TFE affected the solution viscosity and, as a consequence, higher jet instabilities and thinner fiber dimensions were obtained.

Gluten gel and film properties in the presence of cysteine and sodium alginate.

Wheat flour has an ability of forming dough by mixing with water, which exhibits a rheological property required for making bread. The major protein is gluten, which is a valuable protein material for food industry. In this study, gluten protein gels and films were formed with cysteine and sodium alginate. Adding cysteine improved gel and film properties (stress relaxation behavior, bending strength). The gel containing 0.01 M cysteine had a longer relaxation time and was more rigid than the gel without cysteine. Although adding sodium alginate to the gluten suspension containing cysteine improved the water-holding ability and homogeneity of the gel network, the film from this gel was more brittle than the gluten film with cysteine alone. Microstructural observations of the gels and films with scanning electron microscopy suggested that water evaporation was more heterogeneous from the gel containing sodium alginate than from the gel with cysteine alone. Fourier transform-infrared (FT-IR) analysis during film formation suggested that the presence of cysteine encourages interaction between gluten molecules and results in intermolecular beta-sheet formation in earlier stages than in the no additive condition. FT-IR results also suggested that the combined effect of sodium alginate and cysteine on the protein secondary structure was remarkably different from that of cysteine alone. Our results suggest that addition of a suitable amount of cysteine (0.01 M) and heat treatment to 80 degrees C during gluten gel and film formation induces a homogenous network in the gel and film by regulating disulfide-sulfide interactions.

Trafficking of storage proteins in developing grain of wheat.

The processing properties of the wheat flour are largely determined by the structures and interactions of the grain storage proteins (also called gluten proteins) which form a continuous visco-elastic network in dough. Wheat gluten proteins are classically divided into two groups, the monomeric gliadins and the polymeric glutenins, with the latter being further classified into low molecular weight (LMW) and high molecular weight (HMW) subunits. The synthesis, folding and deposition of the gluten proteins take place within the endomembrane system of the plant cell. However, determination of the precise routes of trafficking and deposition of individual gluten proteins in developing wheat grain has been limited in the past by the difficulty of developing monospecific antibodies. To overcome this limitation, a single gluten protein (a LMW subunit) was expressed in transgenic wheat with a C-terminal epitope tag, allowing the protein to be located in the cells of the developing grain using highly specific antibodies. This approach was also combined with the use of wider specificity antibodies to compare the trafficking and deposition of different gluten protein groups within the same endosperm cells. These studies are in agreement with previous suggestions that two trafficking pathways occur in wheat, with the proteins either being transported via the Golgi apparatus into the vacuole or accumulating directly within the lumen of the ER. They also suggest that the same individual protein could be trafficked by either pathway, possibly depending on the stage of development, and that segregation of gluten proteins both between and within protein bodies may occur.

Surface-associated proteins of wheat starch granules: suitability of wheat starch for celiac patients.

Wheat starch is used to make baked products for celiac patients in several European countries but is avoided in the United States because of uncertainty about the amounts of associated grain storage (gluten) proteins. People with celiac disease (CD) must avoid wheat, rye, and barley proteins and products that contain them. These proteins are capable of initiating damage to the absorptive lining of the small intestine in CD patients, apparently as a consequence of undesirable interactions with the innate and adaptive immune systems. In this study, starch surface-associated proteins were extracted from four commercial wheat starches, fractionated by high-performance liquid chromatography and gel electrophoresis, and identified by tandem mass spectrometry analysis. More than 150 proteins were identified, many of which (for example, histones, purothionins, and glutenins) had not been recognized previously as starch-associated. The commercial starches were analyzed by the R-5 enzyme-linked immunosorbent assay method to estimate the amount of harmful gluten protein present. One of these starches had a low gluten content of 7 ppm and actually fell within the range proposed as a new Codex Alimentarius Standard for naturally gluten-free foods (maximum 20 ppm). This low level of gluten indicates that the starch should be especially suitable for use by celiac patients, although wheat starches with levels up to 100 ppm are deemed safe in the proposed Codex standards.

Improved tensile strength of glycerol-plasticized gluten bioplastic containing hydrophobic liquids.

The aim of the present work has been to study the influence of hydrophobic liquids on the morphology and the properties of thermo-molded plastics based on glycerol-plasticized wheat gluten (WG). While the total amount of castor oil and glycerol was remained constant at 30 wt%, castor oil with various proportions with respect to glycerol was incorporated with WG by mixing at room temperature and the resultant mixtures were thermo-molded at 120 degrees C to prepare sheet samples. Moisture absorption, morphology, dynamic mechanical properties, and tensile properties (Young's modulus, tensile strength and elongation at break) of the plastics were evaluated. Experimental results showed that the physical properties of WG plastic were closely related to glycerol to castor oil ratio. Increasing in castor oil content reduces the moisture absorption markedly, which is accompanied with a significant improvement in tensile strength and Young's modulus. These observations were further confirmed in 24 wt% glycerol-plasticized WG plastics containing 6 wt% silicone oil or polydimethylsiloxane (PDMS) liquid rubber.

Aging properties of films of plasticized vital wheat gluten cast from acidic and basic solutions.

In order to understand the mechanisms behind the undesired aging of films based on vital wheat gluten plasticized with glycerol, films cast from water/ethanol solutions were investigated. The effect of pH was studied by casting from solutions at pH 4 and pH 11. The films were aged for 120 days at 50% relative humidity and 23 degrees C, and the tensile properties and oxygen and water vapor permeabilities were measured as a function of aging time. The changes in the protein structure were determined by infrared spectroscopy and size-exclusion and reverse-phase high-performance liquid chromatography, and the film structure was revealed by optical and scanning electron microscopy. The pH 11 film was mechanically more stable with time than the pH 4 film, the latter being initially very ductile but turning brittle toward the end of the aging period. The protein solubility and infrared spectroscopy measurements indicated that the protein structure of the pH 4 film was initially significantly less polymerized/aggregated than that of the pH 11 film. The polymerization of the pH 4 film increased during storage but it did not reach the degree of aggregation of the pH 11 film. Reverse-phase chromatography indicated that the pH 11 films were to some extent deamidated and that this increased with aging. At the same time a large fraction of the aged pH 11 film was unaffected by reducing agents, suggesting that a time-induced isopeptide cross-linking had occurred. This isopeptide formation did not, however, change the overall degree of aggregation and consequently the mechanical properties of the film. During aging, the pH 4 films lost more mass than the pH 11 films mainly due to migration of glycerol but also due to some loss of volatile mass. Scanning electron and optical microscopy showed that the pH 11 film was more uniform in thickness and that the film structure was more homogeneous than that of the pH 4 film. The oxygen permeability was also lower for the pH 11 film. The fact that the pH 4 film experienced a larger and more rapid change in its mechanical properties with time than the pH 11 film, as a consequence of a greater loss of plasticizer, was presumably due to its initial lower degree of protein aggregation/polymerization. Consequently, the cross-link density achieved at pH 4 was too low to effectively retain volatiles and glycerol within the matrix.

Properties and microstructure of thermo-pressed wheat gluten films: a comparison with cast films.

Wheat gluten films were prepared by thermo-pressing, and their mechanical properties were compared to those of cast films. The stress-strain relationship was established for films with various amounts of glycerol. Both relationships were quite different, revealing a different network organization. Thermo-pressed films presented higher stress values than cast films, but the effect of the glycerol amount was similar in both cases, an increase of the glycerol amount leading to a decrease of both films stress. The glycerol influence on the strain at break of thermo-pressed films was very limited, with strain values reaching a maximum around 200%. The role of disulfide bridges on themomoulded films mechanical properties was investigated, and it was shown that some rearrangements and a significative protein insolubilization occurred during the process. The effective flow porosity of the protein network for thermo-pressed films was estimated by water capillary rise measurements to about 7%. Scanning electron microscopy was used to obtain some information about the microstructure of both cast and thermo-pressed films.

Interaction of maize zein with wheat gluten in composite dough and bread as determined by confocal laser scanning microscopy.

Protein body-free maize zein, when mixed at 35 degrees C (above its glass transition temperature range), significantly (p < 0.01) improved the rheological and leavening properties of sorghum-wheat composite flour dough, resulting in improved loaf volume. Confocal laser scanning microscopy was used to observe the structure of zein fibrils and the interaction between zein and gluten proteins in the composite dough and bread systems. Autofluorescence and immunolocalization techniques were used to locate gluten and zein, respectively. Optical sections were collected every 0.4 microm through the samples and digitally processed to produce reconstructed three-dimensional images. Results showed that zein fibrils form an outer layer that intermittently coats the gluten networks, thereby strengthening them. This type of microstructure is able to withstand the pressure exerted by gas cell expansion during yeast fermentation to increase loaf volume.

Physicochemical studies of caroubin: a gluten-like protein.

It has been reported that caroubin, a protein mixture obtained from carob seeds, has rheological properties similar to those of gluten. Comparative studies of the effects of hydration and temperature on caroubin and gluten were carried out with the aid of NMR, FTIR, scanning electron microscopy, and differential scanning calorimetry techniques. The results show that caroubin has a more ordered structure than gluten and that hydration has little effect on its secondary structure when compared to gluten. Caroubin is more easily accessible to water than gluten, suggesting that caroubin is more hydrophilic in nature. On hydration, caroubin, like gluten, forms fibrillar structures and sheets.

Structural characteristics of corn starches extruded with soy protein isolate or wheat gluten.

Commercially available corn starches containing 0, 25, 50 and 70% amylose were extruded with 10, 20 and 30% soy protein isolate (SPI) or wheat gluten (WG) at 22% moisture content (dry basis) in a C.W. Brabender single screw laboratory extruder using a 140 degrees C barrel temperature and a 140 rpm screw speed. True, solid and bulk densities; percent total, closed and open pores; and shear strengths of the extrudates were determined. The microstructures of the extrudates were studied by scanning electron microscopy (SEM). The total pores of the extrudates were affected significantly (p > F = 0.0001) by type of protein (SPI or WG) and starch amylose. The open or closed pores, were affected by protein type only. The interaction between amylose and protein contents was highly significant (p > F = 0.0001). In general, the total pores and bulk densities were higher for WG-starch extrudates compared to SPI-starch extrudates. These values decreased as amylose content increased from 0 to 25% and then increased thereafter. The open pores, on the other hand, increased with increasing protein content from 10 to 20% and then decreased. Extrudates containing WG had higher shear strengths than those containing SPI.

Conformational differences between two wheat (Triticum aestivum) 'high-molecular-weight' glutenin subunits are due to a short region containing six amino acid differences.

'High-molecular-weight' (HMW, high-Mr) glutenin subunits are protein constituents of wheat (Triticum aestivum) seeds and are responsible in part for the viscoelasticity of the dough used to make bread. Two subunits, numbered 10 and 12, are the products of allelic genes. Their amino acid sequences have been derived from the nucleic acid sequences of the respective genes. Subunit 10 has fewer amino acids than subunit 12, but migrates more slowly on SDS/PAGE (polyacrylamide-gel electrophoresis). This anomaly is due to between one and six of the amino acid differences between the subunits, localized towards the C-terminal end of the proteins. This has been established by making chimaeric genes between the genes for subunits 10 and 12, transcribing and translating them in vitro and analysing the products by SDS/PAGE. The postulated conformational differences between subunits 10 and 12 are discussed in relation to current hypotheses for the structure of HMW glutenin subunits.