Abstract:To explore the effects of covalent modification of locust bean gum (LBG) on the physicochemical properties, structural characteristics and Pickering emulsion stability of pine kernel protein (PKP), infrared spectroscopy and fluorescence spectroscopy were used to analyze the structural differences between conjugates and physical mixtures. The mechanism of interaction between PKP and LBG at different mass ratios was revealed by measuring particle size, Zeta-potential and surface hydrophobicity. Pickering emulsions were prepared using PKP-LBG conjugates as emulsifiers, and the effects of oil phase volume fraction on emulsion properties and stability were investigated. The results showed that compared with the physical mixtures of PKP and LBG, the content of α-helix and β-sheet in the conjugates decreased, and the maximum fluorescence emission wavelength redshifted, indicating that covalent binding caused the protein structure to unfold and the spatial conformation to become loose. Particle size and Zeta-potential measurements showed that the average particle size of conjugates(especially M-P-L-0.5 conjugate with PKP:LBG mass ratio of 1.0∶0.5) decreased and the surface charge increased, which could resist protein aggregation. The surface hydrophobicity and free thiol content of conjugates were significantly higher than those of physical mixtures (P<0.05), indicating that heating changed the tertiary structure of the protein and exposed hydrophobic groups, which was significantly related to the improvement of solubility and emulsifying properties of conjugates (M-P-L-0.5 increased by 0.41 times and 2.47 times, respectively, compared with PKP) (P<0.05). Pickering emulsions were prepared using conjugates as emulsifiers. The average particle size of the emulsion was positively correlated with the volume fraction of the oil phase. Microscopic structure observations showed that M-P-L-0.5 could adsorb on the surface of oil droplets to form an interface layer and stabilize the emulsion. The droplets of M-P-L-0.5 with a 60% oil phase were large and the gaps were small, and the emulsion stability was significantly higher than that of other emulsions (P<0.05). Moreover, this emulsion had good storage, acid-base, centrifugation, salt ion and temperature stability. This study clarified the mechanism by which LBG covalent modification improved the functional properties of PKP and successfully prepared highly stable Pickering emulsions, providing a basis for increasing the added value of pine kernel meal and expanding the application of PKP in food.