Synergistic disinfection and removal of biofilms by a sequential two-step treatment with ozone followed by hydrogen peroxide

Synergistic disinfection and removal of biofilms by ozone (O-3) water in combination with hydrogen peroxide (H2O2) solution was studied by determining disinfection rates and observing changes of the biofilm structure in situ by confocal laser scanning microscopy (CLSM) using an established biofilm of Pseudomonas fluorescence. The sequential treatment with 03, 1.0-1.7 mg/L, followed by H2O2, 0.8-1.1%, showed synergistic disinfection effects, while the reversed treatment, first H2O2 followed by O-3, showed only an additive effect. The decrease of synergistic disinfection effect by addition of methanol (CH3OH), a scavenger of hydroxyl radical (center dot OH), into the H2O2 solution suggested generation of hydroxyl radicals on or in the biofilm by the sequential treatment with O-3 followed by H2O2. The primary treatment with O-3 increased disinfection rates of H2O2 in the secondary treatment, and the increase of O-3 concentration enhanced the rates. The cold temperature of O-3 water (14 degrees C and 8 degrees C) increased the synergistic effect, suggesting the increase of O-3 adsorption and hydroxyl radical generation in the biofilm. CLSM observation showed that the sequential treatment, first with O-3 followed by H2O2, loosened the cell connections and thinned the extracellular polysaccharides (EPS) in the biofilm. The hydroxyl radical generation in the biofilm may affect the EPS and biofilm structure and may induce effective disinfection with H2O2. This sequential treatment method may suggest a new practical procedure for disinfection and removal of biofilms by inorganic oxidants such as O-3 and H2O2. (C) 2014 Elsevier Ltd. All rights reserved.