The objective of this study was to determine the effects of hemin and heating temperature on the mutagenicity and lipid oxidation of pork batter during in vitro human digestion with enterobacteria. The mutagenicity and lipid oxidation of pork batter increased in the presence of hemin and with increasing temperatures from 60 to 80°C by 17.9 and 23.0% and by 54.6 and 21.2%, respectively. However, the pork batter mutagenicity decreased by 6.09% during in vitro human digestion, whereas lipid oxidation increased by 19.4%. The antioxidant activities of pork batter decreased with hemin and increasing temperature but gradually increased during in vitro human digestion, especially in the large intestine, regardless of the type of enterobacteria (Escherichia coli and/or Lactobacillus sakei). These results indicate that hemin and temperature are closely related to mutagenicity in pork batter, and in vitro human digestion with enterobacteria could reduce mutagenicity possibly owing to the increase in antioxidant activity by antioxidative enzymes released by enterobacteria.