Chlorpyrifos is the most common insecticide in freshwater ecosystems, and detected in agricultural and fishery product. In this study, Oreochromis niloticus were exposed to 5, 10 and 15 ppb sublethal concentrations of chlorpyrifos in order to determine the oxidative stress response in liver. Acetylcholinesterase (AChE) activity was significantly inhibited. Superoxide dismutase activity (SOD) increased after 15 days of chlorpyrifos treatments at all concentrations (146.95%, 53.04%, 208.70%, respectively). Malondialdehyde levels were higher than that of the control level after 15 days of 5 ppb (95.65%), 10 ppb (69.56%) and 15 ppb (252.17%) chlorpyrifos treatments. Malondialdehyde levels were also increased ranging from 59.09%, 113.63% to 195.46% after 30 days of 5, 10 and 15 ppb chlorpyrifos exposures. Glutathione S-transferase activity decreased except for 15 days low concentration exposure. Catalase (CAT) activity decreased while there is no significant alteration in glutathione peroxidase activity. After recovery period, the low concentration group of chlorpyrifos provided a protection in AChE activity during recovery, but fish were observed to be unable to overcome the inhibition of AChE activity at high concentration groups. CAT activity remained reduced, SOD activity increased whereas the other biochemical parameters recovered to control levels. Results of this study suggest that chlorpyrifos induces oxidative stress in the liver of O. niloticus and this effect is not related with anti-acetylcholinesterase activity of pesticide.