Comparison of historical and registered winter bread wheat varieties in terms of immunogenic epitopes causing celiac disease

of the Republic of Turkey, celiac disease has been reported in 154,027 cases in the country. This disease emerges as a result of an immune response to a protein complex called gluten. In this study, the levels of immunogenic epitopes that may trigger celiac disease were examined in modern and traditional wheat varieties grown in Turkey at different periods; the differences between the varieties and the effects of cultivation conditions on these levels were evaluated. Within the scope of the research, 24 different wheat varieties developed between 1930 and 2022 were cultivated under irrigated and rainfed conditions in Ilgın district of Konya using a randomized complete block design with three replications. Samples were classified into six groups based on registration periods as Historical (<1930), Pre-Green Revolution selection breeding (2nd Group), and varieties developed by hybridization and historically grouped as Pre-Green Revolution 1968–1970 (3rd Group), Post-Green Revolution 1979–2000 (4th Group), Post-Green Revolution 2001–2010 (5th Group), and Post-Green Revolution 2010-2022 onwards (6th Group). In the evaluation of quality parameters of wheat genotypes, weight, grain moisture, hectoliter weight, sedimentation, mixograph value, and wet gluten levels were analyzed. To determine immunogenic epitope levels, wheat samples were first milled to obtain the flour fraction. From this fraction, total protein and gliadin concentrations were measured, and the samples were subjected to sodium dodecyl sulfate polyacrylamide gel electrophoresis (SDS-PAGE) to separate gliadin fractions. The qualitative and quantitative analysis of gliadin fractions was performed using reverse-phase highperformance liquid chromatography (RP-HPLC). Based on the retention times obtained from chromatograms, four main gliadin regions were defined: ω5 gliadin (3–15 min), ω1,2 gliadin (18–24 min), α/β gliadin (25–39 min), and γ gliadin (40–65 min). Finally, the potential of gliadin protein fractions to elicit immune responses associated with celiac disease was weighted using immunogenic coefficients, and an epitope score analysis was performed. The findings from RP-HPLC analyses revealed that as the registration year progressed, especially under irrigated conditions, the immunogenic contribution of α-/β- gliadin and γ gliadin fractions associated with celiac disease increased, whereas under drought conditions, older wheat varieties exhibited higher epitope levels compared to modern ones. The epitope scores calculated within the scope of this study indicated that there were no significant differences in celiac-related epitope levels between historical and modern bread wheat varieties. Although the scores of modern groups were higher under irrigated conditions, the highest score under drought conditions was observed in the historical group, suggesting that the immunogenic load associated with celiac disease is not solely a consequence of modern breeding. Therefore, these data support that modern wheat breeding has not made a systematic contribution to the increase in celiactriggering epitopes and that historical and modern varieties should be evaluated independently in terms of celiac risk. High-resolution mass spectrometry data obtained through LC-QTOF analyses enabled a more advanced characterization of peptide structures belonging to gliadin fractions identified by HPLC. In these analyses, α-gliadinderived and similar celiac epitopes were screened, and peptides were evaluated by scanning the mass-to-charge (m/z) ratios with a ±0.8 tolerance based on values reported in the literature. To evaluate the gliadin protein profiles of wheat genotypes, the MALDITOF- MS technique was employed; peaks in the range of 600–5000 m/z were analyzed across 144 spectra. Overall, the MALDI-TOF-MS findings demonstrated that registration year, genetic background, and environmental conditions directly influence gliadin protein composition and the formation of the celiac-related immunogenic profile. Significant differences in terms of celiac-triggering epitopes were observed both in historical and landrace varieties that had not undergone any hybrid breeding process and in modern bread wheat cultivars developed through hybrid breeding. This is a significant indication that bread wheat breeding efforts have not contributed to the increase in celiac-triggering epitopes. According to the quality analysis results, differences were observed between wheat varieties grown under irrigated and rainfed conditions. In general, quality parameters such as grain yield, protein content, wet gluten amount, sedimentation, and mixograph energy reached higher values under irrigated conditions, whereas under rainfed conditions, particularly some ancient varieties stood out with lower gluten and protein content. Grain yield ranged between 301.80 and 956.90 kg/da, thousand kernel weight between 24.30 and 49.50 g, and hectoliter weight between 42.10 and 87.00 kg/hl. Wet gluten content ranged from 15.40% to 43.30%, protein content from 9.60% to 14.95%, and gluten index from 41.70% to 95.70%; sedimentation values ranged between 17.10 and 72.00 ml. Historical varieties generally stood out with lower protein, gluten, and energy values, while 5th and 6th group modern varieties exhibited superior quality parameters, showing advantage in terms of flour technology.