Abstract: The present study aimed to elucidate the antibacterial effect and underlying mechanisms of plasma-activated water (PAW) combined with phenyllactic acid (PLA) against Escherichia coli O157:H7. The inactivation effects of PAW and PLA on E. coli O157:H7 and their effect on cell morphology, cell membrane integrity and intracellular reactive oxygen species (ROS) level were investigated by plate counting, scanning electron microscopy and fluorescence staining. The results showed that the population of E. coli O157:H7 did not change significantly after PLA treatment at 0.125～1.0 mg/mL for 8 min (P>0.05). E. coli O157:H7 decreased by 5.65 lg CFU/mL after PAW treatment combined with 1.0 mg/mL of PLA for 8 min, which was significantly higher than the 1.06 lg CFU/mL of PAW alone (P<0.05). As shown by the scanning electron microscope images, obvious changes in the morphology of E. coli O157: H7 cells were observed after PAW treatment combined with PLA. After the co-treatment of PAW and PLA (1.0 mg/mL) for 8 min, the extracellular protein content, cell membrane potential and intracellular ROS level of E. coli O157:H7 increased by 25.6-, 0.75- and 9.53-fold (P<0.05), respectively, as compared with the control cells. In summary, PAW combined with PLA could effectively inactivate E. coli O157:H7 cells, which might be associated with membrane disruption and oxidative damages. The results could provide a scientific basis for the application of PAW and PLA in food sterilization and preservation.