Although small nZNI nanoparticles are useful in various applications, the biosafety of this material needs to be evaluated. In this study, Escherichia coli cells were exposed to 0, 112, 560, 1120 mg/L of nano-Fe0particles, respectively. Experiments were carried out to examine the activities of lactate dehydrogenase (LDH), cellular superoxide dismutase (SOD), and malondialdehyde (MDA) after exposure to nano-Fe0 for 24h. The activities of LDH and the levels of MDA were significantly increased (P<0.05), respectively. However, the activities of SOD were significantly decreased (P<0.05). A dose dependent increase in lipid peroxidation product (MDA) contents was observed in treatment groups(r=0.945, P<0.05).The result demonstrated that the damage to cell membranes and oxidative stress were mechanisms of nano-Fe0 ecotoxicity.