Process-driven modulation of underutilized safflower protein functionality through pH-shifting, high-pressure homogenization and ultrasound processes

This study investigates the process-driven modulation of underutilized safflower protein (SFP) functionality through sequential pH-shifting in combination with ultrasonication (US) or high-pressure homogenization (HPH). Native SFP exhibited extremely poor solubility and negligible foaming properties, which limited its applicability in food formulations. pH-shifting alone partially improved solubility and foaming performance by inducing conformational loosening. When combined with US, further enhancements were observed in solubility, foaming capacity, and foam stability, demonstrating the ability of cavitation forces to disrupt aggregates and expose functional groups. The sequential application of pH-shifting and HPH produced the most remarkable improvement in solubility and digestibility at moderate pressure levels, although excessive pressure caused reaggregation. Comprehensive structural analyses supported these functional trends. SDS-PAGE indicated intensified low-molecular-weight bands, suggesting partial dissociation and unfolding. FTIR revealed red-shifted Amide I bands, indicative of increased β-sheet formation, while fluorescence spectroscopy confirmed greater solvent exposure of tryptophan residues. Particle size analysis showed substantial size reduction and improved homogeneity, whereas zeta potential measurements suggested enhanced colloidal stability. Morphological examination by SEM further confirmed fragmentation and surface modification after treatment. Overall, the combination of pH-shifting with US or HPH effectively enhanced solubility, interfacial properties, and in vitro digestibility of SFP. These findings highlight a promising strategy for unlocking the functional potential of safflower protein, advancing its utilization as a sustainable alternative protein ingredient in future food systems.