Abstract: Plasma-activated water (PAW) is an emerging non-thermal sterilization technology, which shows potential applications in food preservation. The present study aimed to elucidate the antibacterial effect and underlying mechanisms of PAW against S. typhimurium cells. PAW obtained by plasma discharge for 30, 60, and 90 s were recorded as PAW30, PAW60, and PAW90, respectively. The results showed that the bactericidal effect of PAW against S. typhimurium cells was gradually enhanced with the increasing plasma discharge time. The population of S. typhimurium was decreased by 4.22 lg CFU/mL after PAW60 treatment for 10 min, which was significantly (P<0.05) lower than that of the control group (7.91 lg CFU/mL). The results of scanning electron microscope (SEM) indicated that the PAW60 treatment resulted in obvious changes in the morphology of S. typhimurium cells. The PAW60 treatment also caused severe disruption of the membrane integrity and permeability of S. typhimurium cells, leading to the significant leakages of cytoplasmic nucleic acids and proteins. After the PAW60 treatment, the extracellular membrane integrity of S. typhimurium cells was destroyed and the intracellular reactive oxygen species levels were also significantly increased. In summary, PAW could effectively inactivate S. typhimurium cells, which might be related to the changes in the structure and membrane permeability of S. typhimurium cells. The results of this study provide theoretical basis for the application of PAW in food sterilization and preservation.