Impact of roasting, high-pressure homogenization, and alkaline pH-shifting on hemp seed protein functionality

• Roasting + homogenization + pH-shifting (RHP) of hemp protein was studied. • RHP reduced hemp protein ζ -potential and mean hydrodynamic diameter, D h . • RHP alters the secondary and tertiary structure of hemp protein. • RHP enhanced hemp protein functionality by 5 – 7 folds. Hemp seed proteins have garnered significant research attention in recent years; however, the poor functional properties of hemp protein limit their use in many industrial applications. In this study, we investigate the effects of roasting, high-pressure homogenization, and alkaline pH-shifting on hemp protein structure and functional properties. We utilize a complementary combination of electrophoresis, dynamic light scattering, electrophoretic mobilities, spectroscopy (infrared, fluorescence, and circular dichroism), and microscopy to study the multiscale structural changes occurring in hemp protein and relate it to the protein's functionality. Our results demonstrate that the combination of roasting with high-pressure homogenization and alkaline pH-shifting leads to a significant decrease in ζ -potential (ca. -22 mV) and mean hydrodynamic diameter, D h (ca. 150 nm), alongside notable redshift ( λ max , 327 nm → 336 nm) and reduced α-helix content (> 50%). We postulate that these structural changes drive the improved protein functionality, i.e., foaming properties by ca. 7-fold and emulsifying properties by ca. 5-fold, compared to the native protein. Novel insights from this study present a promising avenue for tuning the structure of hemp protein to mitigate current challenges associated with using the protein in food applications.