Abstract: Abstract: In this study, clove essential oil (CEO) concentrations ranging from 0 to 2.0%, incorporated in a soy protein isolate (SPI) film were used. Antibacterial activity, physical, mechanical, barrier, and antioxidant properties of SPI films were evaluated. The addition of clove essential oil to the film-forming emulsion led to an increase in the thickness of the films, which ranged between 47.37μm and 49.59μm. However, this effect was only significant at the highest level of CEO used (2%). Thus, clove essential oil might contribute to the formation of a loose film matrix with increased thickness. In general, the moisture content value increased as CEO was incorporated into the SPI film, which is attributed to the breakup of the film network. The addition of CEO at a level of 2% (v/v) decreased the moisture content value significantly (P<0.05), which is attributed to an increase in the hydrophobicity of films. The incorporation of clove essential oil decreased the water solubility of SPI films with respect to control. The addition of CEO at a level of 2% (v/v) decreased the water solubility value significantly (P<0.05). In our study, a lower moisture content with minimum solubility was achieved for films formulated with 2% clove essential oil. The water vapor permeability (WVP) of CEO incorporated films were slightly increased from 13.5 to 19.07 g.mm/(m2·d·kPa) as the concentrations of CEO (P<0.05). Film without clove essential oil had a tensile strength of 22.48MPa. Tensile strengths were weaker for films containing CEO than for the control film, significantly (P<0.05) decreasing as the oil concentration increased. Conversely, the elongation at break of SPI films increased significantly (P<0.05) from 25.50% to 36.68%. Additionally, CEO incorporated films became darker and showed a higher green value. The antioxidant activity of SPI films with and without incorporated clove essential oil was determined. The results showed that 1,1-Diphenyl-2-picrylhydrazyl radical (DPPH) scavenging activity and reducing power of the SPI films was significantly increased (P<0.05) with an increasing CEO concentration. The antibacterial activities of SPI films incorporated with clove essential oil against Gram-negative and Gram-positive bacteria were determined. This method is based on the measurement of the clear zone caused by the growth inhibition produced by a film disk containing the antimicrobial agent when put in direct contact with a bacterial culture. Films containing 1% of CEO were effective against all test microorganisms, and exhibited a strong inhibitory effect on the growth of Listeria monocytogenes and Lactobacillus sake, as evidenced by minimal growth around the film cuts. Inhibition was increased with an increasing concentration of essential oil. As expected, the films containing the highest oil content (2%) presented the greatest zone of inhibition (P<0.05). SPI films with CEO were significantly more effective against gram-positive bacteria (Listeria monocytogenes and Lactobacillus sake) than against gram-negative bacteria (Escherichia coli and Pseudomonas fluorescens). The lower antibacterial activity against Escherichia coli and Pseudomonas fluorescens could be due to the higher resistance of gram-negative microorganisms to these compounds. Our results pointed out that the incorporation of clove essential oil as a natural antibacterial agent has potential for using the developed film as an active packaging.