Size Characterisation of Wheat Proteins, Particularly Glutenin, by Asymmetrical Flow Field-Flow Fractionation

Abstract

Asymmetrical flow field-flow fractionation was demonstrated to fractionate the most high molecular weight proteins of glutenin found in flour from wheat cultivars with excellent bread-making performances. The fractionation occurs in a flow of liquid on the basis of differences in diffusion coefficients, and therefore hydrodynamic diameters, and applies to large molecules and colloidal particles. Microgram quantities of glutenin were analysed and shown to have hydrodynamic diameters of 5 to 45 nm. Much larger sizes can be analysed because the method does not have an upper size limit as do gel electrophoresis and gel filtration. Fractionations can often be done in less than 5 minutes. Sonication in combination with a surfactant, SDS, dissolved proteins from freeze-dried extracts of flour obtained by sequential extraction using progressively increasing concentrations of dilute HCl. An early fraction showed a response around 8 nm hydrodynamic diameter, which corresponds to a molecular weight range of 23 000–128 000. This indicated the presence of gliadins. Later fractions gave responses up to 35 nm, which translated to the molecular weight range 440 000–11 million, which is consistent with the fact that these fractions contain glutenin. Reduction of a late-fraction sample shifted the band to 8 nm, which corresponds to glutenin subunits. The method appears to be a viable procedure to obtain rapid size fractionations of unreduced glutenin.