Abstract: Abstract: Soy Protein Isolate (SPI), a byproduct of the soybean oil industry, can widely be expected to serve as one of the best alternatives to synthetic plastics, due mainly to excellent film-forming properties, including biocompatible, biodegradable, environmentallyfriendly, and cheap to produce. Nevertheless, the SPI normally contains a large number of hydrogen, ionic, and hydrophobic bonds within the molecular structure. The SPI films presented relatively low water resistance, mechanical properties, and thermal stability, due to a large number of hydrophilic bonds in the structures (-OH, -NH2, -COOH, -SH), compared with conventional synthetic plastic films. Therefore, it is urgent to explore protein-based plastic films with improved functional properties. ZnO Nanoparticles (NPs) are highly functional nano-scale products with several attractive properties, such as non-toxic, large specific surface area, strong UV protection, antibacterial properties, and high thermal stability. As such, ZnO NPs can be incorporated into the SPI film matrix, further enhancing the physical and antibacterial properties of films. Furthermore, the presence of N-H groups in the protein chain can interact with ZnO NPs through hydrogen bonds. Auxiliary agents are also needed to integrate the SPI molecules and ZnO NPs, because the high energy, unsaturation, and instability of ZnO NPs can cause the surface roughness of the film. Alternatively, a by-product of grape processing, Grape-Skin Red (GSR) is generally used as a natural anthocyanin pigment. A typical C6-C3-C6 carbon skeleton structure in GSR also contains two benzoyl rings and oxygenated hexameric heterocycles with cations (a typical 2-phenyl-benzopyran cation structure). The structural homogeneity and surface roughness of film can be improved via the cross-linking of highly reactive structures with SPI molecules through hydrogen bonds and electrostatic interactions. These interactions also ensure the uniform mixing of ZnO NPs and SPI. GSR also contains rich and free phenolic hydroxyl structures that can offer the SPI film considerable antibacterial activity, which is extremely advantageous towards potential active packaging film applications. Therefore, this study aims to synthesize and characterize the SPI-based composite films with an SPI matrix and adjuvants (ZnO NPs and GSR). FTIR showed that the ZnO NPs and GSR were successfully dispersed in the SPI film matrix, interacting with the SPI molecules under hydrogen bonds and electrostatic interactions. SEM demonstrated that GSR promoted the dispersion of ZnO NPs in the SPI film matrix, where the compatibility of ZnO NPs and SPI resulted in a low level of roughness on the film. GSR significantly enhanced the mechanical and barrier properties, as well as the thermal stability of composite films, when combined with the ZnO NPs. The tensile strength of SPI composite films increased from 1.37 to 3.28 MPa. The melting point increased from 194 ℃ to 231 ℃, whereas, the water content decreased from 34.41% to 25.37%, and the water vapor permeability decreased from 5.57× 10-12 (g·cm)/(cm2·s·Pa) to 4.74 × 10-12(g·cm)/(cm2·s·Pa). Particle size and zeta potential indicated a uniform particle distribution in the solution (PDI value <0.3). GSR can be fixed inside the protein molecules or adsorbed on the surface by cross-linking resulting in changes in the secondary and tertiary protein structures and the charge distribution of the SPI. However, the charges on the film-forming solutions were highly stable (all >30). The composite films can widely be expected to apply in active packaging, due mainly to excellent antibacterial properties against Staphylococcus aureus and Escherichia coli. The finding can provide insightful ideas to design and produce biodegradable films.