The physicochemical properties of plasma-activated water (PAW) generated under different process conditions were investigated, and their changes under different storage conditions were also studied. The results showed that increasing the processing time and power, and decreasing generated water volume, could cause an increase in the redox potential, conductivity, and temperature of PAW, and a decrease in its pH. A slower dissipation of the reactive oxygen and nitrogen species in PAW was found on storage at 4 °C in a sealed conical flask than on storage at room temperature. The inactivation ability of plasma-activated lactic acid (LA) to Listeria monocytogenes (L. monocytogenes) and Pseudomonas aeruginosa (P. aeruginosa) was higher than that of PAW or LA alone under the same experimental conditions. The results of this study may provide theoretical information for the application of PAW as a potential antimicrobial agent in the future.