Aflatoxins (AFs) are dangerous mycotoxins, which include a great number of lipophilic molecules produced by aerobic microscopic fungi belonging to the genus Aspergillus causes health hazard including death to human and livestock. The objective of this study was to evaluate the effectiveness of ozone gas treatment on the fungal growth and detoxification of AFs - contaminated wheat and corn grains. Ozone concentration treatments 40 mg / Kg wheat for 1 hour and 80 mg / Kg corn for 2 hours of exposure time respectively were applied to contaminated samples of wheat and corn grains. It was observed that completely inhibition of Aspergillus growth and consequently the total aflatoxin content was decreased. In vivo, the biosafety assessment for 72 male albino rats fed on diet containing 70% wt. of ozone treated AFs – contaminated grains were evaluated comparing to control groups. Results indicated that rats fed on AFs contaminated grains have significantly increased the serum enzymes activity of alanine aminotransferase (ALT), aspartate aminotransferase (AST), alkaline phosphatase (ALP), gamma glutamyl transferase (GGT), malondialdehyde (MDA) content as well as the serum levels of creatinine, urea, glucose, total cholesterol (TC) and triglycerides (TG). Also, it was observed that a significant decrease in the level of serum total protein (TP), albumin (Alb), reduced glutathione (GSH) and testosterone hormone comparing to control groups. However, the oral administration of ozonized groups ameliorated the biochemical parameters compared to rats fed on contaminated grains. Moreover, histopathological studies of liver, kidney and testis tissues of rats fed on contaminated grains that revealed different lesions and changes in tissues, inversely to that improving effects in tissues of ozonized contaminated grains – fed rats. It was concluded that ozonization treatment were most effective in reduction of mold count and degradation of aflatoxins content for grains during storage.