Purification and Characterization of the Glutenin Subunits of Triticum tauschii,Progenitor of the D Genome in Hexaploid Bread Wheat

N-terminal amino acid sequences and sodium dodecyl sulfate polyacrylamide gel electrophoresis (SDS-PAGE) molecular weights have been determined for high-performance liquid chromatography (HPLC)-purified high molecular weight (HMW) and low molecular weight (LMW) glutenin subunits (GS) of Triticum tauschii ssp. strangulata, contributor of the D genome to hexaploid bread wheat. The use of three different extraction procedures resulted in similar glutenin preparations. On the basis of N-terminal sequences, the same types of glutenin subunits that have been reported in bread and durum wheats (HMW-GS of both the x and y types and LMW-GS of the LMW-s, LMW-m, α-, and γ-types) were found in T. tauschii. However, the HMW-GS in T. tauschii were in greater proportion relative to LMW-GS when compared to reported values for a bread and durum wheat. Our results support the likelihood that differences in the proportions of the various subunits contributed by the A, B, and D genomes, rather than qualitative differences in the types of subunits, are responsible for the major differences in quality characteristics between bread wheat and durum wheat.     

Production of Flours with Reduced Epitope Content Using Milling Technology

Epitopes on the α‐gliadins are known to give rise to immune responses that may lead to the development of celiac disease in genetically predisposed individuals. The reduction of epitope levels in wheat‐based products would likely benefit this group of consumers and also consumers with non‐celiac gluten sensitivity. Conventional breeding of wheats with lowered epitope levels will take time, but in this study we show for the first time that milling technology can be used to produce flour mill streams that are depleted in α‐20 gliadin epitopes. Fifteen mill streams from two New Zealand wheat cultivars, Sapphire (a biscuit wheat) and Monad (a bread wheat), were tested with reversed‐phase HPLC and an α‐20 gliadin epitope ELISA kit. The level of α‐20 epitope measured in Sapphire gliadins was significantly less than that found in Monad gliadins, even taking into account differences in total protein content. For both cultivars, compared with the straight‐run flour, the break flours had similar or significantly higher proportions of α‐20 epitope per unit of protein, whereas most of the reduction streams had significantly lower proportions of α‐20 epitope per unit of protein. Theoretically, combining selected (mainly reduction) flour streams may produce flour with ∼75% of the epitope content of the straight‐run flour.     

Contribution of Sourdough Lactobacilli,Yeast, and Cereal Enzymes to the Generation of Amino Acids in Dough Relevant for Bread Flavor

The amino acid release was determined in wheat doughs supplied with salt, acid, dithiothreitol, or starter cultures to evaluate the relevance of the amino acid concentration on bread flavor. Wheat flour proteinases almost linearly released amino acids and the highest activity of wheat flour proteinases was found in acidified and reduced doughs. The effects of starter cultures on amino acid concentrations depended on their composition. Yeasts exhibited a high demand for amino acids, however, the total amino acid concentrations were not markedly affected by lactic acid bacteria. The individual amino acid contents were determined by the pH during fermentation and microbial metabolism. The formation of proline was favored by values higher than pH 5.5, whereas release of phenylalanine, leucine and cysteine mainly occurred at lower pH. Ornithine was found only in doughs fermented with Lactobacillus pontis. To determine effects of the amino acid concentration on bread aroma, fermented doughs were evaluated in baking experiments. An increased intensity of bread flavor was obtained by preferments prepared with lactic acid bacteria. The roasty note of wheat bread crust could be markedly enhanced by L. pontis. This results support the assumption that flavor of wheat bread is enhanced by increasing the concentration of free amino acids and especially ornithine in dough.     

Polymorphisms in two homeologous γ-gliadin genes and the evolution of cultivated wheat

The polymorphisms in two -gliadin genes GAG56D and GAG56B on the D- and B-genomes of polyploid wheat, respectively, were investigated by sequencing PCR products and by PCR-RFLP. Of GAG56D, two alleles fo and ok were previously known to occur in hexaploid wheat. Here, we found that 16 sequenced fragments of GAG56D from six recognized subspecies of Triticum aestivum, including 13 contributed by this study, were identical to either the fo or the ok allele. Considering published evidence, it was concluded that the investigated alleles of GAG56D stemmed from two different Aegilops tauschii plants and thus two independent origins of hexaploid wheat. Compared to GAG56D-sequences obtained from 10 accessions of Ae. tauschii, the fo and ok alleles clustered with fragments from three accessions collected in the Caspian region. By sequencing fragments of GAG56B, four distinct allelic groups were found among cultivated wheats, typical of bread wheat (p-aes), durum wheat of gliadin 45-type (a), durum wheat of gliadin 42-type (p-dur) and Timopheev's wheat (p-tim), respectively. Interestingly, the a allele found in gliadin 45-type durum wheat was shared by European spelt cultivars, which strongly supported the hypothesis that European spelt originated from a hybridization event between a tetra- and hexaploid wheat. The data also suggested that emmer might have been domesticated more than once. Phylogenetic analysis of GAG56-fragments obtained from putative B/G-genome donors excluded all candidate species as immediate donors of the B/G-genome, but instead indicated a monophyletic origin of all GAG56B alleles found in polyploid wheat, i.e. including T. timopheevii.     

The Inheritance of Morphological and Biochemical Traits Introgressed into Common Wheat (<Emphasis Type="Italic">Triticum aestivum</Emphasis> L.) from <Emphasis Type="Italic">Aegilops speltoides</Emphasis> Tausch

The genetic control of morphological characters and gliadin composition was studied in two bread wheat lines with introgressed segments from Aegilops speltoides Tausch. It was found that the transferred trait of leaf hairiness is controlled by one dominant gene, non-allelic to the known gene, Hl1. It was localized in 7B chromosome. Whole plant non-glaucousness is under the control of an inhibitor gene, allelic to the gene W1^I of wheat located on chromosome 2B. This gene was found to be epistatic to the gene controlling spike waxlessness. The introgressed gene for spike glume color was found to be allelic to the Rg1 gene located on 1BS of common wheat, but it was linked with another allele of the gliadin locus Gli-B1.     

Effect of spelt wheat flour and kernel on bread composition and nutritional characteristics

Spelt wheat seeds (Triticum aestivum subsp. spelta cv. Ostro) were used to obtain white spelt flour (64.5% yield), wholemeal spelt flour (100% yield), and scalded spelt wheat kernels. From these materials, white spelt wheat bread (WSB), wholemeal spelt wheat bread (WMSB), and spelt wheat bread with scalded spelt wheat kernels (SSKB) were made and were compared to the reference white wheat bread (WWB). The spelt wheat flours and breads contained more proteins in comparison to wheat flour and bread. Among the samples the highest rate of starch hydrolysis was noticed in WSB. During the first 30 min of incubation this particular bread was shown to have significantly more (P < 0.05) rapidly digestible starch than the WMSB and later on also more starch than in WWB and SSKB, respectively. The WMSB had the lowest hydrolysis index (HI = 95.7). However, the result did not differ significantly from that in the reference common wheat bread. On the other hand, the most refined spelt wheat flour resulted in a bread product (WSB) that was statistically withdrawn (P < 0.05) as one with the highest HI (112.6).     

Relationship Between the Qualitative and Quantitative Compositions of Gluten Protein Types and Technological Properties of Synthetic Hexaploid Wheat Derived from Triticum durum and Aegilops tauschii

The contribution of the diploid wheat species Aegilops tauschii (Coss.) Schmall to the technological properties of bread wheat (Triticum aestivum L.) was previously studied by the investigation of synthetic hexaploids derived from tetraploid durum wheat (T. turgidum L.) and three diploid Ae. tauschii lines. The results indicated that bread volume, gluten index, SDS‐sedimentation volume, and maximum resistance of gluten were significantly influenced by the Ae. tauschii lines. To determine the relationship between technological properties and qualitative and quantitative compositions of gluten proteins, the flours of parental and synthetic lines were extracted using a modified Osborne fractionation. Gliadin and glutenin fractions were then characterized by reversed‐phase (RP) HPLC on C₈ silica gel. The HPLC patterns revealed typical differences between synthetic and parental lines. The gliadin patterns of three synthetic lines and the glutenin patterns of two synthetic lines were more similar to that of the diploid Ae. tauschii parents involved in the hybrids. In the glutenin pattern of one synthetic line, characteristics from both Ae. tauschii and the durum wheat parents were observed. The amount of total gliadin and gliadin types of the synthetic lines was mostly intermediate between those of the durum and Ae. tauschii parents. The amounts of total glutenin and glutenin types (HMW and LMW subunits) of the synthetic lines were generally higher than those of the parental lines, and the ratio of gliadins to glutenins was significantly decreased. High positive correlations were found between the amount of total glutenins, HMW, and LMW subunits and bread volume, maximum resistance and extension area of gluten, and SDS‐sedimentation volume. The ratio of gliadins to glutenin subunits had a strong negative influence on these properties. The protein content of the flours and the amount of total gluten proteins were not correlated with any of the technological properties. Results on the relationship between biochemical characteristics and the breadmaking properties indicated that wheat prebreeding would benefit from studies on protein types and quantification in the choice of parents. In addition, the potential of the diploid Ae. tauschii for improvement of breadmaking quality should be further exploited.     

N‐Terminal Amino Acid Sequence Analysis of Endosperm Proteins in Japanese Hexaploid Wheat

Semidry electroblotting is convenient and allows a rapid and efficient protein transfer from two‐dimensional polyacrylamide gel electrophoresis (2D‐PAGE) gels onto sequencer stable supports for protein microsequence analysis in a gas‐phase sequencer. Using this technique, I determined the amino acid sequences of the endosperm proteins in Japanese hexaploid commercial wheats (Triticum aestivum). Based on sequence determination of the Japanese hexaploid wheats, the endosperm protein could be easily characterized. Wheat endosperm protein, extracted in the presence of 2‐mercaptoethanol and SDS, fractionated into many protein polypeptides using 2D‐PAGE under dissociating conditions. These components were grouped into HMW glutenin subunits, α‐, β‐ or γ‐gliadins, and novel protein polypeptides by using the N‐terminal amino acid sequences. The novel endosperm protein polypeptides were detected, and two new types of N‐terminal amino acid sequences have been found for protein poly‐peptides. These polypeptides have much faster electrophoresis mobility during 2D‐PAGE and are therefore probably a much smaller size than any other peptides of endosperm protein groups found in hexaploid wheat. Ten protein polypeptides have been purified from cultivars of Japanese wheat. Some differences in the contents of amino acids for four protein polypeptide spots were apparent in Japanese wheat.     

Studies of Partial Amino Acid Sequences of γ‐40k Secalins of Rye

Though γ‐40k secalins are a major protein type within rye storage proteins, total amino acid sequences are not as well known as the gluten proteins of wheat. Well‐reputed structural features such as amino acid compositions and molecular masses indicated a close relationship between γ‐40k secalins and γ‐gliadins of wheat, but the degree of homology of amino acid sequences and the positions of intramolecular disulfide bonds are unknown. Therefore, two major components of γ‐40k secalins (R1, R2) were analyzed for partial amino acid sequences. The R1 and R2, derivatized with 4‐vinylpyridine, were isolated from the prolamin fraction of rye cultivar Danko by means of a two‐step RP‐HPLC on C₁₈ silica gel. The proteins were digested in parallel with trypsin and thermolysin, and the partial hydrolyzates were separated by RP‐HPLC. Simultaneous measurement of UV absorbance at 210 and 254 nm allowed the detection of all peptides eluted as well as the specific detection of pyridylethylated cysteine peptides. Isolated peptides were characterized by sequence analysis, and in parts by mass spectrometry, and assigned to known sequences of γ‐gliadins. The results demonstrated that the N‐terminal domain of R1 and R2 remained undigested after tryptic hydrolysis; they were in agreement with the N‐terminal domain of γ‐gliadins in their molecular masses and in the absence of cysteine residues. Most of the isolated peptides originated from the C‐terminal domains, they covered 83% (R1) and 77% (R2), respectively, of the C‐terminal domain of a known γ‐gliadin (clone pW1020). Comparison of R1 and R2 revealed differences only in a few sequence positions. The degree of homology between the C‐terminal domains present in γ‐40k secalins and γ‐gliadins was ≈85%. All eight cysteine residues of γ‐gliadins were found in R1 and R2 sequences. Remarkably, sequences close to corresponding cysteine residues were identical for γ‐40k secalins and γ‐gliadins. Therefore, it can be assumed that the positions of intramolecular disulfide links are homologous.     

Molecular characterization of two novel Glu-D1-encoded subunits from Chinese wheat (Triticum aestivum L.) landrace and functional properties of flours possessing the two novel subunits

The high molecular weight glutenin subunits (HMW-GS) can be used for wheat quality improvement. Two novel alleles (designated 1Dx1.5* and 1Dy12.2*, respectively) at the Glu-D1 locus were identified in the Chinese wheat landrace variety ‘Jiuquanjinbaoyin’ by comparison of subunit mobility with that previously identified in several standard hexaploid wheats. The 1Dx1.5* and 1Dy12.2* genes were isolated using the allele-specific PCR primers and the complete open reading frames (ORFs) were obtained. Allele 1Dx1.5* consists of 2487 bp encoding a mature protein of 827 amino acid residues, whereas 1Dy12.2* consists 1980 bp encoding 658 residues. Comparisons of amino acid sequences analysis showed that 1Dx1.5* had higher similarity with the HMW-GS isolated from the wheat related species (Aegilops tauchi Coss.) than from the bread wheat varieties (Triticum aestivum L.). The 1Dy12.2* amino acid sequence showed a generally similar to the 1Dy12* isolated from Chinese endemic wheats. Meanwhile, the dough properties of the lines expressing (null, 7+8, 1.5*+10), (null, 7+8, 2+12.2*) and (null, 7+8, 2+12), respectively, were measured by a Mixograph, which demonstrates that the alleles 1.5*+10 can be considered as having positive impact on dough strength when compared with the alleles 2+12. In addition, the subunits 2+12.2* also showed a greater impact on dough strength than 2+12.     

Effects of Wheat Cultivar and Nitrogen Application on Storage Protein Composition and Breadmaking Quality

Influences of cultivar and nitrogen application on protein concentration and composition, and amount and size‐distribution of different protein components, were investigated in 10 spring wheat cultivars (Triticum aestivum L.) with widely varying gluten strength, grown under four nitrogen fertilizer conditions. The results showed that cultivar differences in gluten strength were determined by storage protein composition, differences in total amount of HMW glutenin subunits, the glutenin‐to‐gliadin ratio, and the relationship between SDS‐soluble and SDS‐insoluble protein polymers. Negative correlations were found between protein parameters related to gluten strength and bread volume. No cultivar stability for gluten strength in relation to differences in nitrogen application was found. Thus, the gluten strength was influenced by the nitrogen application in all the investigated cultivars. Increased nitrogen supply correlated significantly to an increase in all protein components containing gliadins and glutenins, but not to those containing albumins and globulins. The increase in protein components containing gliadins and glutenins correlated significantly with an increase in protein concentration and bread volume.     

Evaluation of genetic variation and grain quality of old bread wheat varieties introduced in north-western Italian environments

The increasing consumers’ request for new speciality wheat derivatives and the possible widening of marketing opportunities is giving to both landraces and old varieties, new chance of on farm survival providing a low cost way to safeguard these important genetic resources. A farmer’s consortium acting in north-western Italy, is attempting the introduction of some old bread wheat landraces in marginal environments of Piedmont and Liguria regions. The reason of this attempt is the local market demand of old bread varieties suitable for the preparation of traditional cakes and biscuits. Three Italian bread wheat landraces, a mixture of durum and bread wheat landraces and three modern varieties were evaluated. The entries were cropped in Val Borbera and Val d’Aveto (Piedmont and Liguria region, respectively) in the same growing season (2009–2010). The high variation of gliadin profile detected within the landraces indicates that all have retained the genetic heterogeneity typical of the old wheat landraces. In consequence of the health-promoting effects of whole grain consumption, eleven nutritional and technological traits of whole flours were investigated. The analysis of collected data revealed appreciable differences among the flours obtained from the landraces and the modern varieties. These differences can be mainly attributed to the intensive breeding carried out on bread wheat in the last century. The results of this study suggest that the old bread wheat landraces could have good chance of survive on farm when their characteristics fulfil the requirements of local communities.     

Effect of Nitrogen Fertilization on Quantity of Flour Protein Components,Dough Properties,and Breadmaking Quality of Wheat

Three winter wheat varieties with differing breadmaking quality were grown at two locations in two years at 0 or 3 × 60 kg of nitrogen application. The effect of nitrogen on amount of different components of gluten proteins was determined by reverse-phase HPLC. A high amount of nitrogen led generally to a significant increase of total protein content. However, this increase was obvious only for the gluten proteins; albumins and globulins remained nearly unaffected. The effect of increased protein content on gliadin to glutenin (gli-glu) ratio was inconsistent. While increased protein content increased the gli-glu ratio in the variety Capo, the opposite was true for the variety Renan. Gli-glu ratio of the variety Lindos showed no discernible tendency. As total protein content increased, the ratio of low molecular weight (LMW) to high molecular weight (HMW) glutenins decreased consistently, i.e., in all varieties, in both years and locations. Change of LMW to HMW ratio showed a significant negative correlation to sedimentation value and bread volume. There was no consistent change in the ratio between x- and y-type HMW subunits due to fertilization, as could be shown by densitometric measurements on SDS-PAGE gels. This ratio appeared to be dependent on the genotype and has decreased with decreasing quality. The amount of x-type subunits correlated closely with sedimentation value and bread volume. These results suggest that ratio of HMW glutenins, especially x-type subunits, to total protein content could be the best early detectable parameter with high predictive value for breadmaking quality.     

Gliadin Immunoreactivity and Dough Rheological Properties of Winter Wheat Genotypes Modified by Thioredoxin

Gliadins are among the most important protein fractions affecting wheat baking quality, but they are also plant allergens and a cause of celiac disease or food intolerance. Therefore, we investigated how gliadin immunoreactivity and dough rheological properties are influenced by thioredoxin, a regulatory disulfide protein that can reduce disulfide bonds, a typical motive in many allergenic proteins. Ten winter wheat genotypes of different qualities were analyzed. Reduction by thioredoxin strongly (>50%) decreased gliadin immunoreactivity as estimated by enzyme‐linked immunosorbent assay with immunoglobulin E (IgE) sera from allergic patients and standard antigliadin antibodies but did not significantly affect dough rheological properties. Most parameters from the Brabender extensigraph were only slightly lower. Simultaneously, the farinograph curve exhibited a drawdown dislocation, possibly due to increased water absorption by modified flour, and dough consistency visibly improved. Results suggest that thioredoxin may be a universal natural beneficial modifier, able to significantly decrease gliadin immunoreactivity (hence its potential allergenicity) without decreasing the unique technological properties of wheat flour.     

Characterization of the 1B‐Type ω‐Gliadins from Triticum aestivum Cultivar Butte

ω‐Gliadins were purified from wheat (Triticum aestivum L. ‘Butte’) flour and characterized. Although reversed‐phase HPLC (RP‐HPLC) separated the 1B‐encoded ω‐gliadins into two fractions, 1B1 and 1B2, these fractions had nearly identical amino acid compositions, three similar protein bands in SDS‐PAGE, 10 similar spots in two‐dimensional PAGE, and two similar N‐terminal amino acid sequences. The main components had a range in mass of 48,900–51,500 when estimated by mass spectrometry, significantly less than the mass estimated by SDS‐PAGE. The 1B fractions were digested with thermolysin, the peptides were separated by RP‐HPLC, the peptide mass was determined, and nine peptides from each fraction were sequenced with nearly identical results for the 1B1 and 1B2 digests. A possible consensus sequence of the 1B‐encoded ω‐gliadin internal repeat was QQQXP, where X was F, I, or L in descending order of occurrence. The 1D‐encoded ω‐gliadins were purified by RP‐HPLC as a single fraction that had one band in SDS‐PAGE, two spots in two‐dimensional PAGE, two components with mass of 41,923 and 42,770 detected by mass spectrometry, and two N‐terminal sequences. Circular dichroism (CD) spectra for the 1B and 1D ω‐gliadins were similar and were suggestive of mainly flexible random structure with a significant amount of the left‐handed polyproline II helical conformation in the 1D components.     

Transglutaminase treatment of wheat and maize prolamins of bread increases the serum IgA reactivity of celiac disease patients

Celiac disease (CD) is mediated by IgA antibodies to wheat gliadins and tissue transglutaminase (tTG). As tTG is homologous to microbial transglutaminase (mTG) used to improve foodstuff quality, it could elicit the immune response of celiac patients. This study evaluated the reactivity of IgA of celiac patients to prolamins of wheat and gluten-free (maize and rice flours) breads mTG-treated or not. Prolamins extracted from wheat and gluten-free breads were analyzed by ELISA and immunodetected on membranes with individual or pooled sera from nine celiac patients recently diagnosed. Sera pool IgA titers were higher against prolamins of mTG-treated wheat or gluten-free breads than against mTG-untreated, mainly due to two individual patients' sera. The electrophoretic pattern of gluten-free bread prolamins was changed by the mTG treatment, and a new 31000 band originated in maize was recognized by three CD patients' IgA.     

Syntheses of novel protein products (milkglyde, saliglyde, and soyglyde) from vegetable epoxy oils and gliadin

The aqueous alcohol-soluble fraction of wheat gluten is gliadin. This component has been implicated as the causative principle in celiac disease, which is a physiological condition experienced by some infants and adults. The outcome of the ingestion of whole wheat products by susceptible individuals is malabsorption of nutrients resulting from loss of intestinal vili, the nutrient absorption regions of the digestive system. This leads to incessant diarrhea and weight loss in these individuals. Only recently has this health condition been properly recognized and accurately diagnosed in this country. The culprit gliadin is characterized by preponderant glutamine side-chain residues on the protein surface. Gliadin is commercially available as a wheat gluten extract, and in our search for new biobased and environmentally friendly products from renewable agricultural substrates, we have exploited the availability of the glutamine residues of gliadin as synthons to produce novel elastomeric nonfood products dubbed "milkglyde", "saliglyde", and soyglyde from milkweed, salicornia and soybean oils. The reaction is an amidolysis of the oxirane groups of derivatized milkweed, salicornia, and soybean oils under neat reaction conditions with the primary amide functionalties of glutamine to give the corresponding amidohyroxy gliadinyl triglycerides, respectively. The differential scanning calorimetry, thermogravimetric analyses, and rheological data from a study of these products indicate properties similar to those of synthetic rubber.     

Reduced Gelatinization,Hydrolysis, and Digestibility in Whole Wheat Bread in Comparison to White Bread

Wheat, an important crop in North Dakota and the United States, is often used for bread. Health concerns related to chronic diseases have caused a shift toward consumption of whole wheat bread. There has been some indication that the rate and amount of starch digestibility of whole wheat breads may be lower than for their refined flour counterparts. This research investigated the components of whole wheat bread that may reduce starch digestibility and impact nutritional quality. Six formulations of flour were used, which included two refined flours, two whole wheat flours, and two whole wheat flours with added starch. The starch was added to whole wheat flours to increase the starch level to that of the refined flour so that we can determine whether or not the dilution of the starch in whole wheat bread was a factor in lowering the estimated glycemic index (eGI) of whole wheat bread. White and whole wheat flours and breads were evaluated for chemical composition, baking quality by 1 , and eGI by the Englyst assay. Whole wheat breads had significantly (P < 0.05) higher mineral, protein, arabinoxylan, and phenolic acid contents, as well as significantly (P < 0.05) lower eGI. The starch molecular weight was also significantly (P < 0.05) higher for whole wheat and whole wheat + starch breads compared with white breads. The eGIs of refined flour breads were 93.1 and 92.7, whereas the eGIs of whole wheat and whole wheat + starch breads ranged from 83.5 to 85.1. Overall, several factors in the whole wheat bread composition can be found to affect the quality and starch hydrolysis.     

Molecular Weight Distribution of Proteins in Hard Red Spring Wheat: Relationship to Quality Parameters and Intrasample Uniformity

Molecular weight distribution (MWD) of proteins extracted from hard red spring wheat was analyzed by size‐exclusion HPLC to investigate associations with wheat and breadmaking quality characteristics. Certain protein fractions were related to associations between wheat and breadmaking parameters, specifically when effect of quantitative variation of protein on those parameters was statistically eliminated by partial correlation analysis. SDS‐unextractable high molecular weight polymeric proteins had positive partial correlations with percent vitreous kernel content and breadmaking parameters, including mix time and bread loaf volume. SDS‐extractable protein fractions that were eluted before the primary gliadin peak had positive partial correlations with kernel hardness and water absorption parameters. The proportion of main gliadin fractions in total protein had a negative partial correlation with bread loaf volume and positive correlations with kernel hardness and water absorption parameters. Intrasample uniformity in protein MWD and kernel characteristics was estimated from three kernel subsamples that were separated according to single kernel protein content within individual wheat samples by a single‐kernel near‐infrared sorter. Wheat subsamples were significantly different in protein MWD. Intrasample uniformity in protein MWD did not differ greatly among wheat samples.     

Specificity analysis of anti-gliadin mouse monoclonal antibodies used for detection of gliadin in food for gluten-free diet

A gluten-free diet (GFD) is the sole effective long-lasting treatment of celiac disease. Four monoclonal antibodies (Abs) were prepared by immunization of animals kept on GFD with gliadin. The specificity of these Abs to decapeptides of alpha- and gamma-gliadin and omega-secalin was analyzed by the PEPSCAN technique. Repetitive sequences of alpha- and gamma-gliadin and omega-secalin containing the motifs QPFPXQ (X = Q, L, P) were recognized by all Abs tested. These Abs also frequently reacted with peptides containing the sequences QQSFPQQ, QQTFPQP, and QPFRPQ. On the basis of PEPSCAN results two Abs--8D4 and 7C6--were selected for the construction of a new ELISA kit for the detection of gliadin in food. The comparison of data obtained using the newly developed ELISA kit and commercially available ones indicated that Abs selection on the basis of their fine specificity to gliadin is useful for sensitive detection of gliadin in foods.