Abstract: This study investigated the effects of different addition levels (high, middle, low) of blueberry anthocyanins (BA) on the interactions, physicochemical properties, and in vitro simulated digestion characteristics of complexes formed between BA and Vicia faba protein (VFP), designated as L-BA, M-BA, and H-BA. Particle size distribution was used to characterize the complexes. The mechanism underlying the influence of BA concentration on its interaction with VFP was explored using ultraviolet-visible (UV-Vis) absorption spectroscopy, fluorescence spectroscopy, and molecular docking. Furthermore, the color, water-holding capacity (WHC), oil-holding capacity (OHC), foaming properties, emulsifying properties, and in vitro simulated digestion characteristics of the complexes were studied. The results showed that as BA concentration increased, the particle size of the complexes decreased and the particle size distribution became more uniform. UV-Vis absorption spectra indicated a red-shift in the characteristic absorption peaks of the complexes. Additionally, the intrinsic fluorescence intensity of the protein decreased in a concentration-dependent manner (P < 0.05). Synchronous fluorescence spectroscopy further revealed that the hydrophobicity of the microenvironment around tryptophan and tyrosine residues increased significantly with increasing concentration of BA (P < 0.05). Molecular docking simulations suggested that anthocyanins bound to the protein’s active sites through hydrogen bonding, hydrophobic interactions, and π-cation interactions. BA concentration and pH cooperatively determined the color changes of the complexes. Moderate BA addition significantly enhanced the WHC and OHC of the complexes (P < 0.05), while excessive addition led to a reduction in the foaming capacity, foam stability, emulsifying capacity, and emulsion stability. In vitro simulated digestion experiments demonstrated that the complexes effectively maintained the digestive stability of anthocyanins. Specifically, the H-BA complex achieved a BA retention rate of 74.34%. Furthermore, the 2,2’-azino-bis(3-ethylbenzothiazoline-6-sulfonic acid) (ABTS) cation radical scavenging activity, 1,1-diphenyl-2-picrylhydrazyl (DPPH) radical scavenging activity, and ferric reducing power of the complexes were significantly higher than those of free BA (P < 0.05). Collectively, complexation with VFP effectively preserved BA stability and its antioxidant activity. This study provides a theoretical foundation for the application of BA-VFP complexes in functional foods.

Key words: blueberry anthocyanins; Vicia faba protein; complexes; interaction; physicochemical properties; in vitro digestion