The effect of change of the rheological properties of gluten with the addition of fractions with specific molecular weight was investigated. Fractions extracted from Hereward, Riband and Soissons flours were added to the dough prior to gluten extraction. Once extracted, the glutens were subjected to temperature sweeps and creep recovery rheological tests. In the temperature sweeps, Hereward fractions containing the larger polypeptides had a strengthening effect on the gluten, indicated by a decrease in tan delta and an increase in elastic creep recovery, while those fractions that comprised monomeric gliadins had a weakening effect. Adding total gluten also had a strengthening effect. For the biscuit-making flour Riband, the results were quite the reverse: all fractions appeared to strengthen the gluten network, while the addition of total gluten did not have a strengthening effect. For Soissons gluten, the addition of total gluten had a strengthening effect while adding any individual fraction weakened the gluten. The results were confirmed with creep-recovery tests.

Gluten was extracted from flours of several different wheat varieties of varying baking quality. Creep compliance was measured at room temperature and tanδ was measured over a range of temperatures from 25 to 95°C. The extracted glutens were heat-treated for 20min at 25, 40, 50, 60, 70 and 90°C in a water bath, freeze-dried and ground to a fine powder. Tests were carried out for extractability in sodium dodecyl sulphate, free sulphydryl (SH) groups using Ellman's method, surface hydrophobicity and molecular weight (MW) distribution (MWD) using field-flow fractionation and multi-angle laser light scattering. With increasing temperature, the glutens showed a decrease in extractability, with the most rapid decreases occurring between 70 and 90°C, a major transition in tanδ at around 60°C and a minor transition at 40°C for most varieties, a decrease in free SH groups and surface hydrophobicity and a shift in the MWD towards higher MW. The poor bread-making variety Riband showed the highest values of tanδ and Newtonian compliance, the lowest content of free SH groups and the largest increase of HMW/LMW with increasing temperature. No significant correlations with baking volume were found between any of the measured parameters.

Puroindoline proteins were purified from selected UK-grown hexaploid wheats. Their identities were confirmed on the basis of capillary electrophoresis mobilities, relative molecular mass and N-terminal amino acid sequencing. Only one form of puroindoline-a protein was found in those varieties, regardless of endosperm texture. Three allelic forms of puroindoline-b protein were identified. Nucleotide sequencing of cDNA produced by RT-PCR of isolated mRNA indicated that these were the 'wild-type', found in soft wheats, puroindoline-b containing a Gly -> Ser amino acid substitution (position 46) and puroindoline-b containing a Trp -> Arg substitution (position 44). The latter two were found in hard wheats. Microheterogeneity, due to short extensions and/or truncations at the N-terminus and C-terminus, was detected for both puroindoline-a and puroindoline-b. The type of microheterogeneity observed was more consistent for puroindoline-a than for puroindoline-b, and may arise through slightly different post-translational processing pathways. A puroindoline-b allele corresponding to a Leu -> Pro substitution (position 60) was identified from the cDNA sequence of the hard variety Chablis, but no mature puroindoline-b protein was found in this or two other European varieties known to possess this puroindoline-b allele. Wheats possessing the puroindoline-b proteins with point mutations appeared to contain lower amounts of puroindoline protein. Such wheats have a hard endosperm texture, as do wheats from which puroindoline-a or puroindoline-b are absent. Our results suggest that point mutations in puroindoline-b genes may confer hard endosperm texture through accumulation of allelic forms of puroindoline-b proteins with altered functional properties and/or through lower amounts of puroindoline proteins.

Flours from wheat varieties of differing bread-making quality were fractionated using a sequential salt precipitation technique. The gluten fractions in the different varieties varied in the proportion of HMW, LMW glutenins and gliadins. Their rheological behaviour was examined using constant strain (2%) small deformation oscillation tests over frequencies ranging from 0.005 to 10 Hz, before and after heating at 90 °C. The fractions containing a higher proportion of HMW glutenins were associated with a predominantly elastic character, whereas fractions containing mostly gliadins exhibited a viscous-like behaviour. The frequency dependent rheological behaviour of fractions containing HMW proteins was less susceptible to heat, and their elastic character was maintained after heating, whereas the rheology of intermediate fractions and fractions containing mostly gliadins was more susceptible to heating, indicating a rapid change from viscous to elastic behaviour after heating.

Creep and dynamic rheological tests indicated that addition of redox additives directly into gluten or into flour before gluten extraction resulted in differences in rheological parameters. Heating from 30 to 50 degrees C caused a decrease in elastic modulus whereas heating above 50 degrees C strengthened the structure as evidenced by an increase in elastic modulus. Higher elastic modulus and lower creep compliance values suggested that Hereward gluten was more highly cross-linked than Riband gluten. Glutenin seemed to be affected more than gliadin during temperature sweep experiments. Both oxidising and reducing reagents showed similar weakening effects on gluten irrespective of addition into flour or gluten. Treatment with oxidising reagents, particularly bromate, slightly delayed the temperature for the increase in elastic modulus as assessed by temperature sweep experiments. The identical effect of reducing and oxidising reagents may be attributable to the differences in mode of action and intrinsic material properties. (c) 2005 Society of Chemical Industry.

Protein-bound glutathione (PSSG) and protein-bound related thiol compounds, i.e. cysteine (PSSCys), glutamyl-cysteine (PSSGlu-Cys) and cysteinyl-glycine (PSSCys-Gly), were analysed in proteins of Osborne fractions, i.e. gliadin, glutenin and gliadin-, glutenin-subfractions separated by gel filtration chromatography, gel protein and the total gluten proteins separated from wheat varieties with varying breadmaking performances. The results showed that PSSG and some protein-bound related thiol compounds were found in monomeric gliadins, indicating that glutathione and some related thiol compounds are able to form disulphide bonds (SS) with sulphydryl group (SH) of those proteins and the formation of those disulphide bonds may prevent those monomeric proteins from binding to other proteins. It was also observed that a larger amount of PSSG in glutenin proteins was negatively correlated with the molecular weight (Mw) distribution of glutenin polymers, suggesting that PSSG and protein-bound related thiol compounds may play an important role in controlling polymerisation of glutenin. Furthermore, it was found that the level of PSSG in gel protein from flours with poor breadmaking performances was constantly higher and significantly different (p<0.05) from that of flours with good breadmaking performance. The same trend was observed with gluten samples from breadmaking and biscuitmaking flours.

The high pressure liquid chromatography method for determination of glutathione in free and protein-bound forms was re-established and has successfully been developed to measure glutathione related thiol compounds, i.e. l-cysteine, γ-l-glutamyl-l-cysteine and l-cysteinyl-l-glycine, in both free and protein-bound forms. The natural levels of those compounds in typical strong, weak flours, and flours from 36 wheat varieties grown in the UK were investigated. The total free and protein-bound glutathione compounds found in the 36 UK varieties was 358±51 and 190±17nmol/g, respectively. Multiple correlation analysis did not show a clear-cut relationship between the natural level of glutathione and any related thiol compound in either free or protein-bound forms and flour quality attributes, including rheological properties, baking performance, protein content and SDS sedimentation test values. Therefore, it can be suggested that glutathione and related thiol compounds at natural levels do not lead to significant differences in the rheological properties of dough and the baking performance of flour.

Glutens from poor breadmaking quality wheat, cv. Riband, had a higher SDS extractability than glutens from good quality cv. Hereward. Heating of gluten, especially above 70°C, caused a reduction in the amount of SDS-extractable gluten proteins. Treatment of gluten with redox additives (ascorbic acid, potassium bromate or glutathione) affected extractability, being highest for bromate treated glutens. The SH content of gluten was lower for poor breadmaking Riband and heating resulted in greater decrease in SH content of gluten from good breadmaking Hereward. Hereward gluten had a higher SS content than Riband. The alteration of SS content on heating was not significant and may indicate the heat-induced involvement of non-covalent interactions. SDS-PAGE revealed that oxidants, especially bromate, affect polypeptide composition leading to a more heat stable/tolerant protein structure.

ABSTRACTThe rheological properties of fresh gluten in small amplitude oscillation in shear (SAOS) and creep recovery after short application of stress was related to the hearth breadbaking performance of wheat flours using the multivariate statistics partial least squares (PLS) regression. The picture was completed by dough mixing and extensional properties, flour protein size distribution determined by SE‐HPLC, and high molecular weight glutenin subunit (HMW‐GS) composition. The sample set comprised 20 wheat cultivars grown at two different levels of nitrogen fertilizer in one location. Flours yielding stiffer and more elastic glutens, with higher elastic and viscous moduli (G′ and G″) and lower tan δ values in SAOS, gave doughs that were better able to retain their shape during proving and baking, resulting in breads of high form ratios. Creep recovery measurements after short application of stress showed that glutens from flours of good breadmaking quality had high relative elastic recovery. The nitrogen fertilizer level affected the protein size distribution by an increase in monomeric proteins (gliadins), which gave glutens of higher tan δ and flatter bread loaves (lower form ratio).

Chimpanzee immunoglobulins are virtually identical to human immunoglobulins and thus may have clinically useful applications. In a recent manuscript we described 4 chimpanzee monoclonal antibodies (MAbs) that neutralized hepatitis A virus (HAV) (1). The MAbs were isolated from a combinatorial cIDNA library of chimpanzee gamma1/kappa antibody genes using bacteriophage displaying Fab fragments on their surfaces. Three of the MAbs recognized the same or overlapping epitopes on the HAV capsid, whereas the fourth recognized a different, nonoverlapping epitope on the capsid. All 4 MAbs neutralized the homologous HAV strain, HM-175, in a radioimmunofocus assay and 2 of the 4 MAbs neutralized a heterologous simian HAV strain, AGM-27. From these data, we conclude that the MAbs must recognize at least 3 epitopes on the HAV capsid. Competition assays performed with neutralizing murine MAbs suggested that 3 of the chimpanzee MAbs recognized epitopes on the HAV capsid which have not been defined previously. It is hoped that these MAbs may be used directly in passive immunoprophylaxis to prevent hepatitis A in susceptible populations, e.g., travelers to regions where HAV is endemic or at risk individuals where epidemics are occurring.

Fractionation and reconstitution techniques were used to study the contribution of endogenous flour lipids to the quality of semisweet (Rich Tea-type) biscuits. Biscuit flour was defatted with chloroform and baked with bakery fat but without endogenous lipid addition. Semisweet biscuits baked from defatted flour were flatter, denser, and harder and showed collapse of gas cells during baking when compared with control biscuits. Defatted flour semisweet doughs exhibited a different rheological behavior from the control samples showing higher storage and loss moduli (G' and G' ' values), that is, high viscoelasticity. Functionality was restored when total nonstarch flour lipids were added back to defatted flour. Both the polar and nonpolar lipid fractions had positive effects in restoring flour quality, but the polar lipid fraction was of greatest benefit. Both fractions were needed for complete restoration of both biscuit quality and dough rheological characteristics.

Experiments in controlled environments examined the effects of the timing and severity of drought, and increased temperature, on grain development of Hereward winter wheat. Environmental effects on grain specific weight, protein content, Hagberg Falling Number, SDS-sedimentation volume, and sulphur content were also studied. Drought and increased temperature applied before the end of grain filling shortened the grain filling period and reduced grain yield, mean grain weight and specific weight. Grain filling was most severely affected by drought between days 1–14 after anthesis. Protein content was increased by stresses before the end of grain growth, because nitrogen harvest index was less severely affected than was dry matter harvest index. Hagberg Falling Number was increased to the greatest extent by stresses applied 15–28 days after anthesis. Treatment effects on grain sulphur content were similar to those on protein content, such that N:S ratio was not significantly affected by drought nor temperature stresses. The effects of restricted water on grain yield and quality were linearly related to soil moisture between 44 and about 73% field capacity (FC) from days 15–28. Drought stress (but not temperature stress) before the end of grain filling decreased SDS-sedimentation volume relative to drought applied later.

ABSTRACTThe rheological properties of dough and gluten are important for end‐use quality of flour but there is a lack of knowledge of the relationships between fundamental and empirical tests and how they relate to flour composition and gluten quality. Dough and gluten from six breadmaking wheat qualities were subjected to a range of rheological tests. Fundamental (small‐deformation) rheological characterizations (dynamic oscillatory shear and creep recovery) were performed on gluten to avoid the nonlinear influence of the starch component, whereas large deformation tests were conducted on both dough and gluten. A number of variables from the various curves were considered and subjected to a principal component analysis (PCA) to get an overview of relationships between the various variables. The first component represented variability in protein quality, associated with elasticity and tenacity in large deformation (large positive loadings for resistance to extension and initial slope of dough and gluten extension curves recorded by the SMS/Kieffer dough and gluten extensibility rig, and the tenacity and strain hardening index of dough measured by the Dobraszczyk/Roberts dough inflation system), the elastic character of the hydrated gluten proteins (large positive loading for elastic modulus [G′], large negative loadings for tan δ and steady state compliance [Je0]), the presence of high molecular weight glutenin subunits (HMW‐GS) 5+10 vs. 2+12, and a size distribution of glutenin polymers shifted toward the high‐end range. The second principal component was associated with flour protein content. Certain rheological data were influenced by protein content in addition to protein quality (area under dough extension curves and dough inflation curves [W]). The approach made it possible to bridge the gap between fundamental rheological properties, empirical measurements of physical properties, protein composition, and size distribution. The interpretation of this study gave indications of the molecular basis for differences in breadmaking performance.

ABSTRACTRelaxation behavior was measured for dough, gluten and gluten protein fractions obtained from the U.K. biscuitmaking flour, Riband, and the U.K. breadmaking flour, Hereward. The relaxation spectrum, in which relaxation times (τ) are related to polymer molecular size, for dough showed a broad molecular size distribution, with two relaxation processes: a major peak at short times and a second peak at times longer than 10 sec, which is thought to correspond to network structure, and which may be attributed to entanglements and physical cross‐links of polymers. Relaxation spectra of glutens were similar to those for the corresponding doughs from both flours. Hereward gluten clearly showed a much more pronounced second peak in relaxation spectrum and higher relaxation modulus than Riband gluten at the same water content. In the gluten protein fractions, gliadin and acetic acid soluble glutenin only showed the first relaxation process, but gel protein clearly showed both the first and second relaxation processes. The results show that the relaxation properties of dough depend on its gluten protein and that gel protein is responsible for the network structure for dough and gluten.

The relationships between wheat protein quality and baking properties of 20 flour samples were studied for two breadmaking processes; a hearth bread test and the Chorleywood Bread Process (CBP). The strain hardening index obtained from dough inflation measurements, the proportion of unextractable polymeric protein, and mixing properties were among the variables found to be good indicators of protein quality and suitable for predicting potential baking quality of wheat flours. By partial least squares regression, flour and dough test variables were able to account for 71–93% of the variation in crumb texture, form ratio and volume of hearth loaves made using optimal mixing and fixed proving times. These protein quality variables were, however, not related to the volume of loaves produced by the CBP using mixing to constant work input and proving to constant height. On the other hand, variation in crumb texture of CBP loaves (54–55%) could be explained by protein quality. The results underline that the choice of baking procedure and loaf characteristics is vital in assessing the protein quality of flours.

ABSTRACTFour hull‐less barley samples were milled on a Bühler MLU 202 laboratory mill and individual and combined milling fractions were characterized. The best milling performance was obtained when the samples were conditioned to 14.3% moisture. Yields were 37–48% for straight‐run flour, 47–56% for shorts, and 5–8% for bran. The β‐glucan contents of the straight‐run white flours were 1.6–2.1%, of which ≈49% was water‐extractable. The arabinoxylan contents were 1.2–1.5%, of which ≈17% was water‐extractable. Shorts and bran fractions contained more β‐glucan (4.2–5.8% and 3.0–4.7%, respectively) and arabinoxylan (6.1–7.7% and 8.1–11.8%, respectively) than the white flours. For those fractions, β‐glucan extractability was high (58.5 and 52.3%, respectively), whereas arabinoxylan extractability was very low (≈6.5 and 2.0%, respectively). The straight‐run white flours had low α‐amylase, β‐glucanase, and endoxylanase activities. The highest α‐amylase activity was found in the shorts fractions and the highest β‐glucanase and endoxylanase activities were generally found in the bran fractions. Endoxylanase inhibitor activities were low in the white flours and highest in the shorts fractions. High flavanoid, tocopherol, and tocotrienol contents were found in bran and shorts fractions.

ABSTRACT The rheological properties of gas cell walls in bread doughs are considered to be important in relation to their stability and gas retention during proof and baking. Large deformation rheological properties of gas cell walls were measured using biaxial extension for a number of doughs of varying breadmaking quality at constant strain rate and elevated temperatures of 25–60°C. Strain hardening and failure strain of cell walls both decreased with temperature, with cell walls in good breadmaking doughs remaining stable and retaining their strain hardening properties at higher temperatures (60°C), while the cell walls of poor breadmaking doughs became unstable at lower temperatures (45–50°C) and had lower strain hardening. Strain hardening measured at 50°C gave good correlations with baking volume, with the best correlations achieved between rheological measurements and baking tests that used similar mixing conditions. As predicted by the Considere failure criterion, a strain hardening value of 1 defines a region below which gas cell walls become unstable, and discriminates well between the baking quality of a range of commercial flour blends of varying quality. This indicates that the stability of gas cell walls during baking is strongly related to strain hardening properties, and that extensional rheological measurements can be used as indicators of baking quality.

Puroindolines (PINs) are very basic (pI>10), amphiphilic, highly surface active, low molecular weight, non-gluten proteins of bread wheat (Triticum aestivum). Despite being relatively minor components of wheat flour, PINs have been reported previously to play a key role in controlling wheat-endosperm texture. Some of the structural properties of PIN proteins purified by selective extraction, phase separation and cation-exchange chromatography from hexaploid wheat varieties grown in the UK, two with endosperm texture classified as soft, and four as hard, were characterized by capillary electrophoresis (CE) and mass spectrometry (MS). The same isoform. of PIN-a was detected in all varieties, regardless of their endosperm texture, while PIN-b was heterogeneous in terms of the isoforms present, both between the soft and hard varieties, and within the hard varieties. For the first time, the molecular masses of PIN-b proteins with different point mutations were reported. The alleles were also detectable by their migration times in CE. A CE-screening method was developed and used in a survey of UK-grown hexaploid wheats. Using the ratios of the peak areas and the migration times of PIN-a and PIN-b, not only could the soft and hard varieties be differentiated, but the different alleles of PIN-b within the hard varieties could also be distinguished.