The present study aimed to synthesize Zinc oxide (ZnO) nanoparticles and to analyze the dose-dependent effect on the oxidative stress and antioxidant enzyme activities in 3T3-L1 adipocytes. ZnO is widely used in the several cosmetic lotions and biomedical products. There are several studies that have reported the ZnO nanoparticle-mediated cytotoxicity on various tissues. However, there are no studies carried out on dose-dependent effect of ZnO nanoparticles in the adipose tissue. ZnO nanoparticle was synthesized by chemical pyrolysis method and characterized by the SEM. A cytotoxicity assay was carried out to determine 3T3-L1 cell viability. 3T3-L1 cell morphology was significantly altered, and most of the cells are dead at higher concentration of ZnO nanoparticles. ZnO nanoparticles increased reactive oxygen species (ROS), lipid peroxidation (MDA), and reduced glutathione (GSH) in 3T3-L1 adipocytes. In addition, antioxidant enzyme activity and its mRNA expression were also upregulated in 3T3-L1 adipocytes. In conclusion, the present study showed that ZnO nanoparticles significantly altered oxidative stress and antioxidant enzyme activity in 3T3-L1 adipocytes on a dose-dependent manner.