ABSTRACT The prevalence of aflatoxins (AFB1) in Iranian wheat flours was determined by immunoaffinity chromatography (HPLC). A total of 180 wheat flour and 60 wheat bran samples including three types of flour and wheat bran were collected from different manufacturers. Also, human health risk assessment of aflatoxins due to consumption of wheat flours by children and adults was calculated using the Margin of Exposure (MOE) approach in the Monte Carlo (MC) algorithm. Considering the limit of detection of the current study, 144 out of 180 wheat flour samples and 54 out of 60 wheat bran samples were positive for aflatoxin B1. The amounts of AFB1 in wheat flour and wheat bran samples were detected to be 0.01–0.5 ng/g and 0.06–0.99 ng/g, respectively. The highest mean values of AFB1 were observed in Sangag, Barbary and Tafton flours from different manufacturers with amount of 0.5, 0.4 and 0.3 ng/g, respectively. Despite the contamination of wheat flour and wheat bran samples, AFB1 values were not higher than the limit of Institute of Standards & Industrial Research of Iran (5 ng/g) and European Union (2 ng/g). The mean of chronic daily intake in the adults and children on the basis of consumption of AFB1 in total samples was estimated to be as 0.26 and 0.04 mg/kg-d, respectively. The percentile 95 of MOE in total samples of flours in the adult and children were calculated as 1072 and 647, respectively. However, human health evaluation showed that both children and adult in the Alborz State are at remarkable cancer risk. Hence, national program is necessary to focus on this problem and firm inspection of wheat flours by the regulatory organs.