To evaluate toxic effects of Diethylstibestrol (DES) on aquatic organism, zebrafish (Danio rerio) embryo exposed in DES solutions was investigated, and the method of high performance liquid chromatography (HPLC) for DES to zebrafish embryo was established. Meanwhile, an oligonucleotide-based DNA chip was used to investigate gene expression changes in zebrafish treated with DES. Results showed that even under different treatments (0hpf and 8hpf), the sub lethal endpoints were both with this order: 72h delayed hatch>48h reduced rate of heartbeat>24h no spontaneous movements>32h no heartbeat; however, while comparing with 0hpf exposure, the toxicity of DES at 8hpf exposure was obviously decreased. The early stage of zebrafish embryo was more sensitive to the exposure of DES. The exposal concentrations of DES (0-0.3mg�L-1) and durations (24h, 48h and 72h) were significantly positive correlated with the levels of DES in zebrafish embryo; While the embryo after 24h, 48h and 72h exposure of 0.2mg�L-1DES, the levels of DES in zebrafish embryo were 25.78�4.95, 54.88�7.10 and 71.93�10.55�g�g-1, separately. Among 14 genes selected to study, 13 genes including vg3 were respectively up-regulated, but cyp1A1 were down-regulated.