Ochratoxin A (OTA) is a widespread mycotoxin produced by several species of the genera Aspergillus and Penicillium. Since OTA is mainly found in foods such as rice and oats consumed all over the world, there are many researches on the reduction of OTA. Benzo [a] pyrene (B[a]P) and acrylamide (AA) may occur during the heat treatment process to reduce the OTA present in rice. In this study, we tried to identify the combined toxicity of the three substances (OTA, B[a]P, AA) in the liver. The cell viability of OTA, B[a]P and AA was checked to determine the concentration of IC20. The toxicity of the combination of the three substances showed synergistic effects on cell viability and NO production. To further elucidate the mechanism of toxicity in liver, real‐time PCR was used to confirm mRNA expression levels of enzymes involved in phase I and II pathway. In the case of cytochrome P450 (CYP) families (CYP1A1, CYP1A2, and CYP3A4) belonging to Phase I, the mRNA expression levels were significantly increased when all three of them were mixed with each other than those treated with OTA, B[a]P and AA, respectively. The mRNA expression levels of heme oxygenase‐1 (HO‐1) and glutamate cysteine ligase catalytic subunit (GCLC) belonging to Phase II were also similar to those of Phase I. Therefore, in all experiments, the combined toxicity of OTA, B[a]P, and AA was confirmed, indicating synergistic effects on hepatotoxicity.This abstract is from the Experimental Biology 2018 Meeting. There is no full text article associated with this abstract published in The FASEB Journal.