Abstract: Waterborne polyurethane (WPU) antibacterial film was prepared by UV curing after introducing hydrophilic quaternary ammonium salts to polyurethane (PU) molecular chains via quaternization reaction between different alkyl chain haloalkanes and tertiary amine groups. PUs were bonded with tertiary amine group and unsaturated group, using 3-dimethylamino-1,2-propanediol as the chain extender and hydroxyethyl methacrylate as the capping agent. The structure and properties of WPU dispersion and light cured films were characterized by FT-IR, particle size analyzer, optical contact angle instrument, etc. The antibacterial properties of the materials were tested by shaking-flask method and zone of inhibition. And the effects of tertiary amine compounds on the properties of WPU were investigated. When the mass fraction of tertiary amine compound was 9% and the alkyl long chain was 12, the antibacterial rates of WPU against Gram-negative E. coli and Gram-positive S. aureus were more than 99.5%. Meanwhile, the longer the alkyl chain was, the better the effect was. Through UV-curable crosslinking, it could still maintain excellent antibacterial performance and reflect positive durability after 96 h water washing. The zone of inhibition test showed that the prepared quaternary ammonium salts WPU antibacterial film belonged to contact-killing material which avoided the pollution and adverse effects on the environment, human beings and other materials. With the increase of tertiary amine content, the quaternization degree of WPU system first increased and then decreased, and the variation of quaternization degree affected the antibacterial properties. Moreover, polyurethane material also exhibited good mechanical properties. The increase of hard segment content of the system enhanced the tensile strength of polyurethane. And when the alkyl long chain was 12, the waterborne polyurethane material had better toughness.