The effect of 2-chloroethyl phosphoric acid (CEPA) on aflatoxin biosynthesis, the expression of aflatoxin biosynthetic genes, reactive oxygen species (ROS) formation, cellular redox status, and enzymes involved in glutathione consumption and regeneration in Aspergillus flavus was investigated. The results demonstrated that CEPA dose dependently inhibited aflatoxin B(1) production. The expression of two typical genes involved in aflatoxin biosynthesis, aflR and aflD, was reduced after CEPA treatment at 7 d. Meanwhile, CEPA significantly reduced ROS production and thiobarbituric acid reactive substances (TBARS), increased the ratio of reduced glutathione (GSH) and oxidized glutathione (GSSG) at 5, 6 and 7 d. The activities of glutathione peroxidase (GPx), glutathione reductase (GR), as well as glucose-6-phosphate dehydrogenase (G6PDH) were significantly inhibited after CEPA treatment at 5, 6 and 7 d. The present study suggested that ethylene reduced aflatoxin production is due to oxidative stress alleviation of fungal cells and is related to glutathione redox state changes.