Abstract: In order to further realize the high value utilization of agaro-oligosaccharides (AOS), antioxidant activities and mechanisms of AOS, 2, 2-azobis (2-methylpropylimidazole) dihydrochloride (AAPH) -induced zebrafish model were studied. The composition of AOS was determined by HPLC. The optimal AAPH induced zebrafish oxidative stress model was established, and the antioxidant activities of AOS with different concentrations were investigated by determination of embryo survival rate, reactive oxygen production rate, and cell death rate. The results showed that AOS was mainly composed of agarobiose (51.60%), agarotetraose (28.40%) and agarohexaose (14.50%). Zebrafish embryos with AOS concentration below 200 μg/mL had no teratogenic and lethal toxicity. AOS (25, 50, 100 μg/mL) had a protective effect on oxidative stress induced by AAPH (15 mmol/L) in a dose-dependent manner. The optimal concentration of 100 μg/mL AOS could significantly increase the survival rate of zebrafish embryos (96.88%, p<0.05), and could significantly reduce the reactive oxygen species production rate (100.53%, p<0.05) and cell death rate (101.69%, p<0.05) of AAPH-induced zebrafish.In this study, the zebrafish model was used to reveal for the first time. AOS had antioxidant activity in vivo by eliminating a large number of reactive oxygen species (ROS) production and preventing cell death and injury, which provided a theoretical basis for the application of AOS as a health food.