Rheological properties of partially denatured whey proteins
MetadataShow full item record
As one of the sources for food materials, whey protein concentrates (WPC) are widely processed such as by heat treatment for different application purposes, one of which is to produce protein-based fat replacers. Those fat replacers benefit from high nutritive value and generally recognized as safe (GRAS) status of whey proteins, and moreover, the heat-induced denaturation and aggregations of these proteins provide the desired texture and mouthfeel to the low-fat food systems. Two fat replacers, i.e., Simplesse and Hiprotal60 serial products, have been comprehensively characterized in this research with various techniques. The scope of this study extended from the molecular structure of the heat-induced denatured whey proteins (β-lactoglobulins) to the flowing and rheological behaviour of the protein aggregates. From the characterization of the fat replacer materials, molecular dynamics (MD) computer simulations were also performed on β-lactoglobulin proteins, which are the major proteins in WPC. It has been predicted that the β-lactoglobulin molecule lost α-helices and β-barrels during heat-induced unfolding, and the four solvent inaccessible cysteine (Cys) residues (Cys66, Cys106, Cys119 and Cys121) exposed to the molecular surface. Such changes contributed to the protein-protein interactions and aggregations through –SH/S-S exchange interactions. Denatured proteins were also found to have weak ability to form hydrogen bonds with hydrated water molecules, but the hydrogen bonds between those hydrated water molecules were improved by protein unfolding, which decreased the repulsions due to hydration shells between protein molecules. As for the materials, the microparticulated proteins, Simplesse, were found to be particles with a median diameter, D[0.5] = 1.72 ± 0.04 μm, which is larger than the WPC (Lacprodan87) with D[0.5] = 0.48 ± 0.04 μm. As for the partially denatured proteins, i.e., Hiprotal60 products, values of D[0.5] ranged from 5.484 ± 0.001 μm and 3.296 ± 0.001 μm for Hiprotal60 and Hiprotal60-TS0709, respectively, to around 17 μm for both of Hiprotal60-TS0710 and Hiprotal60-TS0712, depending upon the degree of the protein aggregations. The structures of different protein aggregates were also visualized through ESEM, where small particles were found to form separate aggregates while those larger ones were observed to have more continuous structures. However, there was no disulphide bond present in those protein based fat replacers as indicated by the results of normal and reduced SDS-PAGE and thus, the aggregates were formed and stabilized by non-bonded interactions, mainly hydrophobic interactions, between those denatured protein molecules. From the flowing properties, all the protein-based fat replacers and WPC in aqueous solutions exhibited exponential increases in apparent viscosity at 100 s-1 with protein concentrations from 6% to 21% because of hydrodynamic and protein-protein interactions. Higher exponential dependence of viscosity on protein concentrations was observed for protein-based fat replacers, suggesting stronger interactions between those modified protein molecules. According to the shear thinning and thixotropic analyses, Simplesse required more protein molecules to aggregate for a given increases in viscosity than WPC, while for Hiprotal60 products, the flowing behavior changed with the extent of the protein denaturation and aggregations. Compared with WPC and other protein-based fat replacers, Hiprotal60-TS0710 and Hiprotal60-TS0712 with largest aggregates had the best structuring properties with the lowest concentration requirement for increased viscosity and the strongest network structures with the largest viscosities. From the oscillation tests, intermolecular or interfloc interactions were found in Simplesse as indicated by the solid-like behaviour of this material at high concentrations. Hiprotal60 and Hiprotal60-TS0709 behaved as viscoelatic liquid, and the Cox-Merz rule was valid for such samples, indicating the absence of a network formed by these protein molecules or aggregates. Because of their small values of storage moduli (G’), the rheological properties of Simplesse and Hiprotal60 and Hiprotal60-0709 were believed to result from colloidal crystal structures of the protein molecules and their flocs. The lattice-like structures of these colloidal crystals were mainly stabilized by electrostatic repulsions between the proteins. Hiprotal60-TS0710 and Hiprotal60-TS0712, were found to form strong gels with self-similar or fractal structures at high concentrations. The polymeric chains were densely packed in the gels as revealed by large values of fractal dimensions (≈ 2.3) of the self-similar networks. By adjusting the pH to the pI (≈ 4.5) of the whey proteins, all the protein-based fat replacers exhibited cold-setting gelation behaviour. Fractal structures were found in the acid-induced gels formed by Simplesse but not in those from Hiprotal60 or Hiprotal60-0709. It has been observed that the protein molecules and flocs pack more densely at pI than at natural pH as indicated by the fractal dimensions of acid-induced cold gels formed by Simplesse (≈ 2.3) and Hiprotal60-0710 and Hiprotal60-0712 (≈ 2.4). From the rheological tests of sol-emulsion systems, emulsion droplets were found to affect the rheological properties of the protein-based fat replacers. Emulsion droplets disrupted the interactions of Simplesse as indicated by decreased elasticity, but the viscosity increased due to more hydrodynamic interactions. Due to their larger particle sizes, Hiprotal60 and Hiprotal60-TS0709 induced flocculation of the droplets, which was improved by shearing treatments. Droplet flocculation was also induced by Hiprotal60-TS0710 and Hiprotal60-TS0712, but there was no shear-improving effect. This can be attributed to the large viscosity of the continuous phase in such systems. There was no active filling effect of protein-coated droplets on the gelling behaviour of Hiprotal60-TS0710 and Hiprotal60-TS0712, indicating that thermal denaturation of the protein layers of the droplets played an important role in their active filling effects on protein gels.