Abstract: In order to evaluate the toxicity of acetochlor, zebrafish was used as targeted animal for the test of acute toxicity to adult and embryo, teratogenesis and expression of thyroid related genes in larvae of zebrafish after exposure to acetochlor by static method and quantitive real-time PCR method. The results showed that the 96 h-LC50 value of acetochlor to adult zebrafish and 120 h-LC50 value to larvae was 3.04 mg/L and 5.32 mg/L, respectively, suggesting acetochlor has moderate toxicity to zebrafish. Notochord deformation and pericardial edema were observed in larvae after exposure to 0.3 mg/L acetochlor for five days. The result of window phase exposure test indicated that thyroid related genes such as d1, d2, trα, trβ, s1c5a5, crh, tsh in zebrafish larvae were down-regulated after exposure to 0.06 mg/L acetochlor from 3 h post-fertilization to 10 d, 14 d and exposure to 0.3 mg/L acetochlor from 3 h post-fertilization to 10 d; however, after exposure to 0.3 mg/L acetochlor from 3 h post-fertilization to 14 d, no significant change in the expression of the above-mentioned genes was observed except that d1 was significant up regulated. The results suggested that acetochlor could disrupt the early development of zebrafish larvae, which depends on the exposure dosage and time.