Influence of baking conditions on the detection and composition of gluten

Gluten network formation during processing plays a crucial role in determining the functional and quality aspects of wheat-based products. However, the underlying mechanism of cross-linking is not fully understood due to the complexity of the wheat proteome. Conformational changes of gluten proteins during processing of wheat flour also affect gluten analysis, which is important for determining a gluten-free status of food products. This study aimed to investigate the influence of processing on the immunochemical detection of gluten proteins as well as on the qualitative and quantitative protein composition of wheat-flour based products.

Different wheat flour-based and incurred baked goods (bread, crispbread, pretzel) made from one dough were characterized using immunological, chromatographic, gel- and mass spectrometry-based methods. The samples were considered to reflect different degrees of processing (heat, alkali treatment) and ranked according to an increasing degree of processing as follows: flour, dough, bread crumb, pretzel crumb, crispbread, bread crust, pretzel crust.

Different extraction protocols, including a modified Osborne fractionation, in combination with RP- and GP-HPLC analysis were employed to determine the protein recovery and solubility properties of the wheat proteins in the samples. Protein recovery referred to the protein content analysed relative to the total crude protein content. The results showed that the protein recovery decreased in the processed samples as the degree of food processing increased. For example, the protein recovery in dough was 90%, while in baked goods without alkali treatment, it ranged from 75% to 83%, and with alkali treatment (pretzel crust), it ranged from 58% to 61%. Proteome profiling using LC-MS/MS identified a total of 1488 peptides assigned to 93 protein groups. The ALGL fractions contained 71 protein groups, the gliadin fractions contained 27 protein groups, and the glutenin fractions contained 47 protein groups. Only 16 protein groups were found in all fractions. The majority of proteins in the ALGL fractions were meta-bolic and protective proteins (60%), while the gliadin and glutenin fractions were predominantly composed of storage proteins (≥ 58 %).

The involvement of specific gliadins and glutenins in gluten polymerization and network formation was observed through differential abundance analysis of proteins contained in the fractions of the bread samples compared to the flour. It revealed significant changes in the concentrations of 82 proteins, including 25 gluten proteins. In general, the changes of protein abundances in response to processing showed that glutenins are involved in gluten network formation via SS and non-SS crosslinks and most gliadins rather via non covalent bonds.

The results of the HPLC analyses related the reduced detectability of gliadin/gluten by ELISA to a decreased extractability of gluten, especially the glutenin fractions, from the baked goods. The LC-MS/MS analyses showed the presence of a considerable amount of gliadins, to which ELISA is sensitive, in the glutenin fractions.

Our findings are important for a better understanding of the influence of food processing and the detection of gluten in processed food. Moreover, specific gluten proteins were identified that contribute to gluten network formation and, consequently, bread-making quality.