Effect of dual modification of sonication and γ-irradiation on structural, physicochemical, and functional properties of pumpkin seed protein isolate.

This study investigated the combined effects of sonication and γ-irradiation on the physicochemical, structural, and functional properties of pumpkin seed protein isolate (PSPI). Dual-modified PSPI exhibited significant improvements in functional attributes, including solubility (57.17%), emulsifying activity (71.62%), emulsifying stability (78.41%), surface hydrophobicity (3827.91), reduced turbidity (0.48), and enhanced in vitro digestibility (91.35%). Colour parameters indicated an improved visual profile (L* = 74.11; a* = 0.31; b* = 18.75). Antioxidant activity was also enhanced, as reflected by increased DPPH radical-scavenging activity (41.42%) and FRAP values (29.78 μmol Fe2+/g). Thermal analysis revealed modified transition temperatures (To = 51.5 °C, Td = 63.0 °C, Tc = 145.12 °C) and enthalpy change (ΔH = 28.23 J/g), indicating structural alterations and increased susceptibility to thermal unfolding. Circular dichroism analysis demonstrated conformational rearrangements characterized by reduced α-helix and β-sheet contents and increased β-turn and random coil structures, indicating loss of ordered secondary structure. These findings were further supported by SDS-PAGE, FTIR, and XRD analyses, which provided complementary evidence of structural modifications and reduced structural order following dual treatment. Morphological observations revealed smoother, porous, sponge-like structures, suggesting protein unfolding and structural reorganization. Overall, sequential sonication and γ-irradiation substantially altered the physicochemical and structural characteristics of PSPI and improved several functional and antioxidant properties under the conditions investigated. The findings provide a basis for further evaluation of dual-modified PSPI as a value-added plant-protein ingredient, while food-matrix validation, dose optimization, and scale-up studies remain necessary to establish its practical application.