We have studied inactivation of four strains each of Escherichia coli and Listeria innocua in milk by the combined use of high hydrostatic pressure and the lactoperoxidase-thiocyanate-hydrogen peroxide system as a potential mild food preservation method. The lactoperoxidase system alone exerted a bacteriostatic effect on both species for at least 24 h at room temperature, but none of the strains was inactivated. Upon high-pressure treatment in the presence of the lactoperoxidase system, different results were obtained for E. coli and L. innocua. For none of the E. coli strains did the lactoperoxidase system increase the inactivation compared to a treatment with high pressure alone. However, a strong synergistic interaction of both treatments was observed for L. innocua. Inactivation exceeding 7 decades was achieved for all strains with a mild treatment (400 MPa, 15 min, 20°C), which in the absence of the lactoperoxidase system caused only 2 to 5 decades of inactivation depending on the strain. Milk as a substrate was found to have a considerable effect protecting E. coli and L. innocua against pressure inactivation and reducing the effectiveness of the lactoperoxidase system under pressure on L. innocua. Time course experiments showed that L. innocua counts continued to decrease in the first hours after pressure treatment in the presence of the lactoperoxidase system. E. coli counts remained constant for at least 24 h, except after treatment at the highest pressure level (600 MPa, 15 min, 20°C), in which case, in the presence of the lactoperoxidase system, a transient decrease was observed, indicating sublethal injury rather than true inactivation.