Gluten from some wheat varieties: a comparative study

Knowledge of the gluten composition of soft, durum and spelt wheat is relevant for the success of final baked products (especially salty baked product) and/or for the production of pasta. The characteristics of gluten are also fundamental if the aim is to create products suitable for people genetically predisposed to celiac disease, for those who are sensitive to gluten without celiac disease and, extensively, for those who suffer from intestinal inflammation. For all these people it is important to make products that are as digestible and tolerable as possible. Of all the known grains, einkorn wheat is the one that is considered the most suitable for this purpose.
The study “Comparative Study on Gluten Protein Composition of Ancient (Einkorn, Emmer and Spelt) and Modern Wheat Species (Durum and Common Wheat). Sabrina Geisslitz et al. Published: 12 September 2019 in Foods (MDPI)” analyzes some characteristics of the gluten of some grains (300) highlighting the differences; it also analyzes the effect on them of the use of nitrogenous fertilizers in cultivation.

The reason for the interest in the search for varieties of einkorn, emmer and spelt:
The “ancient” wheats einkorn (Triticum monococcum L., diploid), emmer (T. dicoccum L., tetraploid) and spelt (T. aestivum ssp. spelta, hexaploid) have been cultivated in very low amounts compared to the “modern” wheat species common wheat (T. aestivum L., hexaploid) and durum wheat (T. durum L., tetraploid) in the 20th century. The reasons for the low cultivation of ancient wheats are 30–60% lower grain yields, the presence of husks and poor baking properties compared to common wheat [1]. Nevertheless, ancient wheats have been rediscovered in the last 20 years, because a growing number of consumers associate their consumption with sensory and health benefits due to their comparatively higher contents of e.g., ferulic acid, vitamins, alkylresorcinols and lutein [2–8].
Common wheat is most suitable for bread making, because the flour forms a viscoelastic dough with a high gas holding capacity when it is mixed with water. In contrast, flours of ancient wheats yield softer dough with low elasticity and high extensibility because of their poor gluten quality [1,9–11].
This latter feature translates into a less “strong”[A] and therefore more digestible gluten. In addition, einkorn and emmer do not contain the gluten fraction (33mer” : Quantitation of the immunodominant 33-mer peptide from α-gliadin in wheat flours by liquid chromatography tandem mass spectrometry. Kathrin Schalk et al. 2017. Scientific Reports.) which is considered the one that most activates the immune response in celiac subjects as well as being among the least digestible. This last feature makes these grains, especially einkorn[B], the main candidates for decreasing exposure to celiac disease in genetically predisposed subjects.

Featured in the study:
The total protein content was equally influenced by location and wheat species, however, gliadin, glutenin and gluten contents were influenced more strongly by wheat species than location. Einkorn, emmer and spelt had higher protein and gluten contents than common wheat at all four locations. However, common wheat had higher glutenin contents than einkorn, emmer and spelt resulting in increasing ratios of gliadins to glutenins from common wheat (< 3.8) to spelt, emmer and einkorn (up to 12.1). With the knowledge that glutenin contents are suitable predictors for high baking volume, cultivars of einkorn, emmer and spelt with good predicted baking performance were identified. Finally, spelt, emmer and einkorn were found to have a higher nitrogen partial factor productivity than common and durum wheat making them promising crops for a more sustainable agriculture.

….omissis.
It is generally accepted that gluten proteins are one of the most important factors determining the baking quality of wheat flours. Gluten proteins are storage proteins and classified into gliadins (GLIA) soluble in aqueous alcohol and glutenins (GLUT) soluble in aqueous alcohol only after reduction of disulfide bonds. Not only the amount, but the ratio between GLIA and GLUT (GLIA/GLUT) has been shown to be responsible for good baking quality. GLIA/GLUT of common wheat is typically 1.5–3.1 [12,13], but a recent study showed that the GLIA/GLUT of ancient wheats was much higher (spelt: 2.8–4.0; emmer: 3.6–6.7; einkorn: 4.2–12.0)
Quantitation of GLIA, GLUT, Gluten and Total Protein Contents

….omissis….
As expected, common wheat had the lowest protein content (mean 96.1 mg/g) compared to the other four wheat species and durum wheat the highest one (mean 120.7 mg/g).

…..omissis
In accordance with the protein content, common wheat had the lowest GLIA (mean 41.2 mg/g) and gluten contents (mean 80.4 mg/g) compared to the other four wheats.

……omissis

The GLUT content decreased significantly from common wheat (mean 16.6 mg/g) and spelt (mean 19.0 mg/g) to einkorn (mean 10.3 mg/g), with durum wheat (mean 16.0 mg/g) and emmer (mean 12.8 mg/g) in between.

…omissis….

The differences in GLIA and GLUT contents consequently led to significant differences in the GLIA/GLUT ratios. A significant increasing trend was seen for GLIA/GLUT of the five wheat species. Common wheat had the lowest GLIA/GLUT (mean 2.5) and einkorn the highest (mean 6.5) with spelt (mean 3.3), durum wheat (mean 4.0) and emmer (mean 4.9) in between.
….omissis
Usually, the gluten content corresponds to 80–90% [C] of the protein content, but here it was between 60–80%. One reason for the lower proportion is that wholemeal flours were analyzed, and thus, more proteins of the ALGL (Fractions of albumins/globulins) fraction were present than in white flour. This confirmed our previous study, in which ALGL corresponded to 17–32% of the total protein content and gluten to 68–83% [11]. The variations in protein, gluten, GLIA and GLUT contents observed in the sample set are according to expectations, because the contents are known to be influenced by species and cultivars as well as growing conditions [18–21].

….omissis. Protein yield efficiency. Due to the fact that not only high kernel yields, but also high protein contents are important in wheat, the protein yield efficiency was calculated as a ratio of protein yield, which is the product of protein content and grain yield and quantity of fertilizer. It was remarkable that the ancient wheats spelt, emmer and einkorn had higher protein yield efficiency than the modern wheats common wheat and durum wheat. Thus, regarding either only grain yield or protein yield, it appears that ancient wheats have a better potential to use nitrogen more efficiently than modern wheats, underlining their potential as alternative crops for sustainable agriculture. Agronomic treatments including fertilization were performed according to the standard practices for the individual sites based on minimal amount of nitrogen required and local experience, as is common and has been reported earlier [19]. Thus, the fertilizer quantities were adapted to the respective needs of the wheat species and avoid lodging of spelt, emmer and einkorn (common wheat and durum wheat, 95–125 kg/ha; spelt, 60–95 kg/ha; emmer and einkorn, 0–75 kg/ha) [1], while still ensuring good comparability of the samples as has been demonstrated earlier [5,7].
….omissis. Independent of the growing location, common wheat generally had the lowest total protein, GLIA and gluten content and the lowest GLIA/GLUT compared to the other four wheat species. The protein contents of spelt, durum wheat, emmer and einkorn did not differ clearly, because the growing location had a more pronounced influence on emmer and einkorn than on common wheat, spelt and durum wheat.

…..omissis.
In general, spelt had the highest gluten content, but no difference was observed in the gluten contents of durum wheat, emmer and einkorn. All four species had significantly higher gluten contents then common wheat. However, the gluten composition of all wheat species was significantly different. Independent of the growing location einkorn had the lowest glutenin content, followed by emmer. These differences resulted in significant differences in GLIA/GLUT, which increased independent of growing location from common wheat to einkorn, with spelt, durum wheat and emmer in between. The GLIA/GLUT was influenced by the growing location, as already reported due to environmental conditions (temperature) and amount and time of fertilization [20].

….omissis. We identified a significant genetic variance for almost all wheat species regarding protein, gluten, GLUT and GLIA contents as well as GLIA/GLUT ratio indicating that cultivars exist within eeach wheat species with different expressions of the respective traits.
This is a prerequisite for breeding, which was fulfilled here
.
…omissis. Finally, a low GLIA/GLUT and a high GLUT content are important for good baking quality. It seems to be most promising to select for reduced GLIA/GLUT good baking quality. It seems to be most promising to select for reduced GLIA/GLUT and high GLUT contents in further breeding of spelt, emmer and einkorn.

…..omissis. Conclusions
The Bradford assay adapted for wheat proteins is a suitable tool to quantitate quality-related proteins and protein fractions of common wheat, spelt, durum wheat, emmer and einkorn. The method is easy to use and allows routine analysis of a high number of samples in a short time. Furthermore, we used isolated GLIA and GLUT fractions from common wheat as reference material for calibration, which were suitable for all five wheat species. This compensated one big disadvantage of the Bradford assay, because the absorbance and the dye binding depends on the amino acid sequence of the analyzed proteins.

For the first time, the protein and gluten contents and the gluten protein composition (GLIA and GLUT) of 15 cultivars each of common wheat, spelt, durum wheat, emmer and einkorn, which were grown at four different locations in Germany, were analyzed. To our knowledge, this is the first study comparing the five wheat species cultivated at four locations. Thus, this study represents a robust dataset for all five wheat species that shows the effect of the wheat species, but also the effects of different locations on protein and gluten contents and gluten protein composition.

The key protein parameters related with baking quality are high GLUT content and a relatively low GLIA/GLUT ratio. This enables the identification of cultivars within wheat species with predicted good baking performance independent of the growing location. To improve dough and baking properties of einkorn and emmer, breeding programs should select for low GLIA/GLUT and high GLUT content. Furthermore, common wheat had the lowest protein, gluten and GLIA contents, which is actually contrary to consumer expectations who typically think that ancient wheats have lower gluten contents. A side effect of increasing the cultivation of spelt, emmer and einkorn is their higher nitrogen PFP compared to common wheat. This makes these ancient wheats interesting crops for a more sustainable agriculture.

Keywords: baking quality; gliadin; glutenin

References:

[A] Perchè il glutine del monococco è meno forte di quello del grano duro e del grano tenero: la struttura del glutine è composta da glutenine che formano una struttura chiamata macropolimero (lo scheletro del glutine) che ingloba le gliadine. Le glutenine sono collegate tra loro da legami disolfuro che sono molto resistenti alla scissione. Il monococco ha più gliadine che glutenine rispetto agli altri grani, ha, quindi, uno “scheletro” meno sviluppato che rende il suo glutine meno “forte” . La maggiore presenza di gliadine rende, inoltre, gli impasti viscosi e soffici.

[B]“ Recent studies suggested that gliadin proteins from the ancient diploid einkorn wheat Triticum monococcum retained a reduced number of immunogenic peptides for celiac disease patients because of a high in vitro digestibility with respect to hexaploid common wheat. In this study, we compared the immunological properties of gliadins from two Triticum monococcum cultivars (Hammurabi and Norberto-ID331) with those of a Triticum durum cultivar (Adamello). Gliadins were digested by mimicking the in vitro gastrointestinal digestion process that includes the brush border membrane peptidases. Competitive ELISA, based on R5 monoclonal antibody, showed that gastrointestinal
digestion reduced the immunogenicity of Triticum monococcum gliadins; conversely, the immunogenic potential of Triticum durum gliadins remained almost unchanged by the in vitro digestion. The immune stimulatory activity was also evaluated by detecting the IFN-γ production in gliadin-reactive T-cell lines obtained from the small intestinal mucosa of HLA-DQ2+ celiac disease patients. Interestingly, gastrointestinal digestion markedly reduced the capability of Triticum monococcum gliadins (p < 0.05) of both cultivars to activate T cells, while it slightly affected the activity of Triticum durum. In conclusion, our results showed that Triticum durum was almost unaffected by the in
vitro gastrointestinal digestion, while Triticum monococcum had a marked sensibility to digestion, thus determining a lower toxicity for celiac disease patients.” Comparative Analysis of in vitro Digestibility and Immunogenicity of Gliadin Proteins From Durum and Einkorn Wheat. Luigia di Stasio et al. Frontiers in Nutrition maggio 2020).

[C] I chicchi di grano sono costituiti da diversi tessuti e la macinazione viene utilizzata per separare l’endosperma amidaceo (83% in peso secco (dw) come farina bianca dall’embrione (3% in dw), aleurone e strati di grano esterni (14% in dw) che insieme formano la crusca [5]. L’endosperma amidaceo, e quindi la farina bianca, contiene il 75-85% di amido e il 10-15% di proteine per dw. (Characterisation of Grains and Flour Fractions from Field Grown Transgenic Oil-Accumulating Wheat Expressing Oat WRI1; Per Snell et al. Published: 26 March 2022. Plants.).

References in square brackets refer to the full text of the study