The acute toxicity test, genotoxicity and subchronic oral toxicity of hemp seed oil were investigated. Results indicated that LD₅₀ from the acute toxicity test of mouse was higher than 40 mL/kg. The negative results were showed in Ames test, micronucleus test of bone marrow and sperm abnormality test. No significant differences were found in body weight, food efficiency and blood endpoint values between testing groups and control group. Furthermore, the values of serum biochemical endpoints separately fell into the ranges of their normal specific concentrations. An absence of histological changes in rat organs was observed. It is concluded that no toxic effects of hemp seed oil were observed in this study.