In Iran, wheat is one of the most important sources of diet. Unfortunately, it is vulnerable to contamination with fungi consisting of aflatoxigenic moulds. Since 1997, ozone has been recognized as an effective oxidant and disinfection agent. This study demonstrates the benefits derived from ozone in the inhibition of Aspergillus parasiticus growth and aflatoxin production in wheat. Wheat samples inoculated with A. parasiticus spore suspensions (10,10 2 and 10 4 spores/g of wheat) were treated with ozonated water at various concentrations of 0, 1, 2, 2.5 mg ozone/L at different temperatures. Fungal mycelium was weighed to evaluate the effect of ozone on A. parasiticus growth. Moreover, the rate of aflatoxin production was determined using high-performance liquid chromatography. Our results indicated that ozonated water can inhibit A. parasiticus growth as well as the accumulation of aflatoxins effectively. The best result was obtained when samples were treated with 2.5 mg/L of ozonated water and at fungal spore concentrations of 10 and 10 2 for inhibition of fungal growth and aflatoxin production, respectively (P < 0.001). In conclusion, the usage of ozonated water during wheat tempering is a promising strategy for inhibition of A. parasiticus growth and for reducing aflatoxin production.