Abstract: In this study, flaxseed protein (FSP) was modified by ultrasonic-assisted covalent grafting of tannic acid (TA) under alkaline conditions. The aim was to explore the synergism between ultrasonic and TA in protein modification and its effect on protein structure and function. The results indicated that ultrasound significantly increased the binding capacity and efficiency of polyphenols (P < 0.05). The cavitation effect disrupted the internal structure of the protein, promoting the exposure of sulfhydryl groups and accelerating the covalent coupling of FSP with TA. With increasing TA addition from 2% to 4%, the content of free sulfhydryl groups in the sample decreased markedly, the absolute value of the zeta potential increased and the particle size rose, indicating that the surface charge density of the system enhanced. At the structural level, the content of α-helix decreased while the proportion of β-fold increased, causing the overall conformation to shift from dense to loose. The molecular docking results indicated that TA could adsorb onto the protein surface through multiple hydrogen bonds and hydrophobic interactions, which might provide potential active sites for subsequent covalent reactions. At the functional level, the sample grafted with 4% TA under ultrasound treatment performed the best in terms of solubility, emulsifying, foaming and antioxidant properties. In the sample grafted with 6% TA, some properties decreased due to excessive crosslinking. In conclusion, ultrasonic-assisted covalent grafting is an efficient and green modification strategy that can markedly improve the structural and functional characteristics of proteins.

Key words: flaxseed protein; tannic acid; ultrasound treatment; covalent conjugation; emulsifying properties; antioxidant activity