To assess the antibacterial activity of ZAO nanoparticles, DH 5α bacteria was cultured in Luria-Bertani (LB) liquid medium at 37 °C with shaking until the medium optical density (O.D.600) reached 0.3 to 0.6. Then the bacteria were collected by centrifuging at 5000 rpm for 10 min. The pellet was then resuspended gently with fresh LB medium (make O.D.600 = 0.5). Pipette 3 ml of the resuspended bacteria in a 15 ml tube, after adding ZAO nanoparticles, the bacteria was cultured at 37 °C with shaking, and O.D.600 was detected at different culturing time (3 h, 6 h, 9 h and 12 h) with spectrophotometer.