Distribution and location of ethanol soluble proteins (Osborne gliadin) as a function of mixing time in strong wheat flour dough using quantum dots as a labeling tool with confocal laser scanning microscopy

摘要

Gliadin is the more fluid of the two major classes of proteins, gliadins and glutenins, in wheat flour dough and flows best among these two protein classes. Glutenins because of their high elasticity and high hydrophobicity are not likely to disperse as well as gliadins especially if gliadins are not well dispersed or are not present in the dough matrix. The role of native gliadin in this networking process is not well understood and has not been studied by prior researchers due to unavailability of molecular tools to specifically probe gliadin behavior until now. Gliadins contribute to the viscosity and extensibility in dough. In this study, the distribution and location of ethanol soluble proteins (Osborne gliadins) as a function of mixing time in a Brabender Farinograph for model wheat flour dough were investigated for the first time using confocal laser scanning microscopy (CLSM). Gliadin proteins were tagged with water soluble, biocompatible amine derivatized polyethylene glycol functionalized quantum dots to increase the clarity and specificity of imaging. The effect of different mixing conditions on the distribution of ethanol soluble proteins (Osborne gliadins) and their role in building dough structure was investigated. The chosen mixing times were arrival time (AT), peak time (PT), departure time (DT) and breakdown time (10 min after departure time). Location and distribution of gliadins were investigated in AT, PT, DT and 10 min after departure time. Antibody-quantum dot (QP) mixture successfully bonded to gliadins located on dough sections. The specificity of gliadin antibody to ethanol soluble proteins (Osborne gliadins) was shown successfully with a Western Blot experiment excluding binding to all other hard wheat flour proteins. The images obtained from dough sections were bright and clear and allowed to distinguish gliadin easily. Mixing led to considerable changes in the distribution, average particle size, and number of particle count of gliadins in dough microstructure. The QDs were found to be localized not only around the air cells as indicated by higher intensities but also in the bulk dough. Quantum dots can be used as fluorophore probes to tag and track molecules of interest in food microstructure. This combined with immunohistochemistry techniques offers a better understanding of gliadin distribution in dough during mixing.