Use of spectroscopic and fluorescence techniques to assess heat-induced molecular modifications of gluten

Fresh gluten isolated from soft wheat flour was heated at varying temperatures for different times: from 45 degrees C for 1 hr, to 110 degrees C for 18 hr; it was then freeze-dried. The solubility of the untreated and heated glutens in different solvents under nonreducing and reducing conditions was determined, and the extracts were analyzed for protein composition by sodium dodecyl sulfate polyacrylamide gel electrophoresis (SDS-PAGE) and acid PAGE. The intrinsic fluorescence, the fluorescence developed on titration with 8-aniline-1-naphthalene sulfonate (ANS), the UV spectra with second derivatives and solution stability were measured on acetic acid extracts of the glutens. Titration with ANS revealed sites with high and low probe affinity (high and low hydrophobicity). The results show: 1) a hydrophobic modification at 45 degrees C, this in no way affected the electrophoretic patterns; 2) moderate changes at 65 degrees C ascribable to conformational modifications; 3) heating at 90 degrees C or above strongly affected protein solubility in acetic acid and produced disulfide supported aggregates. Conformational modifications were also evident. In these changes gliadins, except omega-gliadins, and low molecular weight albumins and globulins were the most involved. 4) Heating at 110 degrees C for 18 hr produced insolubilization not reversed by dithiothreitol.