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Sourdough lactic acid bacteria and products for Celiac Susceptible People

by luciano

The lactic acid bacteria present in the sourdough have been shown to have significant abilities to hydrolyze gluten proteins; some strains of lactic bacteria used with specific temperatures, times and concentrations can also hydrolyse the peptides most resistant to gastro-intestinal digestion. Baked products made with sourdough can therefore be considered an excellent opportunity and a valid choice for people genetically predisposed to celiac disease.

Extract from the study “ Gluten-Free Products for Celiac Susceptible People”:
A – “ omissis…… The 33-mer peptide from α2-gliadin (amino acid sequence positions 56–88, LQLQPFPQPQLPYPQPQLPYPQPQLPYPQPQPF) contains three overlapping T-cell epitopes (3 × PQPQLPYPQ, 2 × PYPQPQLPY and PFPQPQLPY) for CD sensitive individuals. The human gastrointestinal enzymes pepsin, trypsin, and chymotrypsin were unable to hydrolyze the 33-mer peptide due to their inability to cleave before or after proline or glutamine, leaving the epitopes intact. Comparatively, large CD immunogenic peptides (≥9 amino acid residues) reach the small intestine (11) after crossing through the epithelial barrier and initiate immunogenic cascade in the lamina propria.

B – “omissis …Wheat flours modified to eliminate or reduce the immune toxicity of gluten have been used to prepare pasta and baked products. The large peptides of gluten need to be modified/converted into peptides of <9 amino acid residues to minimize the CD-induced immunoreactivity. This has been achieved through numerous approaches, including exogenous enzyme treatment, use of sourdough/lactic acid bacteria, use of genetically modified wheat, etc.”

C – “ omissis…The sourdough was prepared by fermenting flour with naturally occurring lactic acid bacteria (LAB) and yeasts. In the mature sourdoughs, the lactic acid bacteria were higher in number (> 10cfu/g) than the number of yeasts. Type I sourdough has a final pH of 4.0 at room temperature (20–30C) and is manufactured by continuous daily refreshments with the aim to maintain the microorganisms in an active state. It takes 2–5 (>30C) days of fermentation for developing type II sourdough as an acidifier with a pH that is <3.5 after 24 h of fermentation (131). The microorganisms were used in the late stationary phase of growth and exhibited restricted metabolic activity. The type III sourdough, as an acidifier supplement and aroma carrier in bread making, is a dried powder used for fermentation by certain starter cultures. A few reports are available about the use of sourdough for the preparation of gluten-free bread (84, 85). In one study it was reported that food processing by selected sourdough lactobacilli and fungal proteases may be considered an efficient approach for eliminating gluten toxicity, reducing the gluten level below 12 ppm (119). Further, sourdough fermentation, usually with a mixture of lactic acid bacteria (LAB) and yeasts, is traditionally used to produce leavened bread, especially from rye flour. Lactobacillus sp. are predominant among lactic acid bacteria (LAB) and they produce numerous mixed proteolytic enzymes, including metalloendopeptidases, such as PepO and PepF; aminopeptidases, such as PepN and PepC; dipeptidases, such as PepD; and dipeptidyl and tripeptidylpeptidases, such as the proline-specific Xaa-Pro dipeptidyl-peptidase (PepX) (132). The combination of wheat germination and sourdough fermentation with Lactobacillus brevis L62 extensively hydrolyzed wheat prolamin down to <5% of the initial content (133). A cell-free extract of two LABs, L. plantarum and Pediococcus pentosaceus, had a higher gliadin-degrading capacity (83%) in doughs than the corresponding cell suspension (70%), and complete gliadin degradation without using live LAB may be optimized (134). High molecular weight polymers, namely exopolysaccharides, are produced by lactic acid bacteria in presence of sucrose that mimics physiochemical properties of commercial hydrocolloids or gums, such as the ability to form a network and bind water. It counteracts the negative effects of excessive sourdough acidification and enhances loaf volume, shelf-life, the staling rate, and textural properties of products (129).”

Depeening
Gluten-Free Products for Celiac Susceptible People. Sweta Rai, Amarjeet Kaur and C. S. Chopra. Front. Nutriens 17 december 2018.

Quantitation of the immunodominant 33-mer peptide from α-gliadin in wheat flours

by luciano

In wheat there are multiple fractions able to activate the adverse response of the human immune system. Among these fractions the most active is that called 33-mer because it is the most resistant to human digestion and because it contains six copies of the three toxic epitopes and its intermolecular bonds are very strong. It is therefore important to know the quantity of this fraction in the grains. The study of which some parts are reported, examined 57 different types of wheat, ancient and modern, noting that the difference, in all soft wheat and spelt flour, of 33-mer is wide: from 90.9 to 602.6 μg / g made with flour. On the other hand, its presence in monococcum wheat and durum wheat was not detected. These results take on great importance because they allow grains to be chosen with limited or no presence of this important toxic fraction for products that are more suitable for non-celiac gluten sensitive people or those suffering from gluten disorders.

“All gluten protein fractions, namely the alcohol-soluble prolamins and the insoluble glutelins, contain CD-active epitopes3. The prolamin fraction is particularly rich in proline and glutamine and the numerous proline residues lead to a high resistance to complete proteolytic digestion by human gastric, pancreatic, and brushborder enzymes. Studies by Shan et al. (2002) showed that a large 33-mer peptide (LQLQPFPQPQLPYPQPQLPYPQPQLPYPQPQPF) from α2-gliadin (position in the amino acid sequence of α2-gliadin: 56–88) is resistant to cleavage by intestinal peptidases4,5. The 33-mer is widely called the most immunodominant gluten peptide4,6,7, because it contains three overlapping T-cell epitopes, namely PFPQPQLPY (DQ2.5-glia-α1a, one copy), PYPQPQLPY (DQ2.5-glia-α1b, two copies) and PQPQLPYPQ (DQ2.5-glia-α2, three copies)3, which result in the initiation of a strong immune response.

Einkorn, emmer and durum wheat

by luciano

Einkorn, emmer and durum wheat: they do not have the “33mer” fraction considered the most active in activating the adverse response of the immune system in celiac subjects. Also for this reason they are the most suitable genotypes for the researches whose aim is to “detoxify” the flours or to intervene with particular enzymes to hydrolyse the “toxic peptides”, however present; they are also more suitable for non-celiac gluten sensitive subjects.

“Quantitation of the immunodominant 33-mer peptide from α-gliadin in wheat flours by liquid chromatography tandem mass spectrometry.

Kathrin Schalk , Christina Lang , Herbert Wieser , Peter Koehler  & Katharina Anne Scherf. Scientific Reports volume 7, Article number: 45092 (2017)

Abstract

Coeliac disease (CD) is triggered by the ingestion of gluten proteins from wheat, rye, and barley. The 33-mer peptide from α2-gliadin has frequently been described as the most important CD-immunogenic sequence within gluten. However, from more than 890 published amino acid sequences of α-gliadins, only 19 sequences contain the 33-mer. In order to make a precise assessment of the importance of the 33-mer, it is necessary to elucidate which wheat species and cultivars contain the peptide and at which concentrations. This paper presents the development of a stable isotope dilution assay followed by liquid chromatography tandem mass spectrometry to quantitate the 33-mer in flours of 23 hexaploid modern and 15 old common (bread) wheat as well as two spelt cultivars. All flours contained the 33-mer peptide at levels ranging from 91–603 μg/g flour. In contrast, the 33-mer was absent (<limit of detection) from tetra- and diploid species (durum wheat, emmer, einkorn), most likely because of the absence of the D-genome, which encodes α2-gliadins. Due to the presence of the 33-mer in all common wheat and spelt flours analysed here, the special focus in the literature on this most immunodominant peptide seems to be justified……Omissis…..

Analysis of durum wheat, emmer and einkorn

The 33-mer peptide was also analysed in two durum wheat and two emmer cultivars (genome AABB) as well as two diploid einkorn cultivars (genome AA) (Table 1). In each of these wheat species, the 33-mer was not detected (<LOD). In comparison to hexaploid common wheat, durum wheat, emmer, and einkorn do not contain the D-genome, which originated from hybridisation of T. turgidum dicoccum (genome AABB) with Aegilops tauschii (genome DD)36. The absence of the 33-mer peptide can be explained by the fact that this peptide is encoded by genes located in the Gli-2 locus on chromosome 6D, which is missing in durum wheat, emmer, and einkorn. Studies by Molberg et al. showed clear variations in intestinal T-cell responses between common wheat and tetra- or diploid species due to different degrees of T-cell immunoreactivity between the gluten proteins encoded on the A-, B-, and D-genome. Einkorn cultivars were only recognized by DQ2.5-glia-α1a-specific T-cell clones, but not by DQ2.5-glia-α1b- and DQ2.5-glia-α2-specific T-cell clones. Emmer and durum wheat cultivars were all recognized by DQ2.5-glia-α1a-specific T-cell clones, but only two out of four emmer cultivars and three out of ten durum wheat cultivars activated DQ2.5-glia-α1b- and DQ2.5-glia-α2-specific T-cell clones37. Consistent with our results, Prandi et al.38 found that the 33-mer was not present in durum wheat. As a consequence, this peptide was used as a marker peptide to identify the presence of common wheat in durum wheat flours. One durum wheat cultivar was also analysed by van den Broeck et al.33 and the 33-mer peptide was not detected either”. https://creativecommons.org/licenses/by/4.0/deed.it