Based on the increasing importance and application of nanoparticles and observing their toxic effects, it is essential to investigate the effects of these substances on physiological traits. This study aimed to examine the effects of different concentrations of CuO nanoparticles (0, 10, 100, and 1000 mgL^-1) on the growth and biochemical parameters of flaxseed seedlings (Linum usitatissimum L.). This study was carried out in the Biology Department of Urmia University based on randomized design with three replications in the glass bottles.  Exposure of flaxseed seedlings to CuO nanoparticles resulted in a significant decrease in rootlets at 1000 mgL^-1 concentration, while hypocotyl measurement indicated the positive effect of increased concentrations of CuO nanoparticles on this parameter. Comparison of the mean data showed increment trend in the amounts of phenol, soluble sugar and lipid peroxidation with rising concentration of CuO, compared to control state. Exposure to CuO nanoparticles enhanced he activity of catalase enzyme, whereas superoxide dismutase and ascorbate peroxidase enzymes exhibited downregulation. The soluble protein and total flavonoid contents displayed significant decrements at 1000 mgL^-1 of CuO nanoparticles. Evidence from biochemical experiments indicates a strategy of flaxseed seedlings using pathways leading to the increase of these factors in order to prevent plant cells from oxidative stress.