The removal efficiency of electrolyzed oxidizing water on biofilm of Staphylococcus aureus was studied. Electrolyzed oxidizing water showed obvious damage on the Staphylococcus aureus biofilm，including substantially destroyed external matrix, severely destroyed cells and decreased bacterial number observed with SEM（scanning electron microscope）on 5 000-times and 40 000-times. The removal efficiency could be reduced significantly by prolonging storage time of oxidation potential acidic water , raising preservation temperature and the presence of organic interfering substances. The removal efficiency of sealing preservation was superior to openning preservation. The available chlorine, pH value and redox potential had a synergistic effect on the removal efficiency. When the amount of available chlorine was low, the removal efficiency of concentration variation was significant. When the amount of chlorine was high, the removal efficiency of concentration variation was not significant. Under low pH value conditions, the removal efficiency was comparatively better. The redox potential has a significant positive correlation with the removal efficiency.