Casein Functionalization Using High-Pressure Homogenization and Emulsifying Salts

In milk, casein proteins orientate themselves into spherical micellar structures with hydrophobic casein subtypes concentrated in the core, while hydrophilic casein subtypes populate the exterior. Previous research demonstrated that milk with the addition of emulsifying salts coupled with high-pressure homogenization induced an unprecedented amount of casein micelle dissociation. This research aims to quantify the extent of casein micelle dissociation in diluted skim milk and evaluate the functionality of these proteins following emulsifying salt treatment coupled with high-pressure homogenization. To evaluate the extent of micellar dissociation, dilute skim milk solutions (20% <i>v</i>/<i>v</i>) were prepared with a varying amount of treatment: no processing (control), just emulsifying salts (Treatment E, 100 mM sodium hexametaphosphate), just high-pressure homogenization (Treatment H, at 300 MPa), and EH (a combination of E and H treatments). Samples were then put through varying filter sizes (0.22 µm, 0.05 µm), and the permeates were analyzed using sodium dodecyl sulfate-polyacrylamide gel electrophoresis. In the control group (20% skim milk), 9.35% ± 2.53% casein protein permeated through a 0.05 µm filter. Alternatively, 93.2% ± 7.71% casein protein was present in EH samples post-filtration through a 0.05 µm filter, demonstrating a significant processing-induced dissociation of casein micelles. A potential benefit to this casein micelle size reduction is the exposure of highly functional hydrophobic subunits from the core of the micelle. In agreement, compared to the control samples, the EH samples had higher foam expansion index values (138.3% ± 12.58% vs. 33.33% ± 14.43% at 0 h), foam stability (113.3% ± 5.774% vs. 21.67% ± 2.887% after 8 h), emulsifying activity (ca. two-fold higher), and interaction with caffeine. These data demonstrate that E, coupled with H, enhances skim milk system functionality, and these changes are likely due to micellar dissociation and protein conformational changes. This work has direct applications in dairy systems (e.g., dairy foams, dairy ingredients) as well as implications for potential processing strategies for other protein-rich systems.