Curcumin is a naturally occurring compound which is known to induce heme oxygenase 1 (HO-1), although the underlying mechanism has not been fully elucidated. This study investigates in detail the mechanism of HO-1 induction by curcumin in human hepatoma cells. There was increasing toxicity of curcumin at concentrations higher than 10 microM. Curcumin was found to induce HO-1 at doses of 10 to 25 microM. At both non-toxic and toxic doses, HO-1 induction was found to correlate with production of reactive oxygen species (ROS), suggesting a causative relationship. This was reinforced by the finding that pretreatment with the antioxidants N-acetylcysteine, vitamin E and catalase prevented HO-1 induction by curcumin. ROS production appeared to be mitochondrial in origin, and curcumin treatment resulted in depolarisation of the mitochondrial membrane potential. Nrf2 was induced by curcumin treatment, which was also partly ROS dependent. Using siRNA, Nrf2 was demonstrated to contribute to HO-1 induction. A panel of kinase inhibitors was used to examine the contribution of MAP kinases to the induction of HO-1 by curcumin. PKC and p38 MAPK activity are required for full induction of HO-1. Furthermore, curcumin also inhibited protein phosphatase activity. In conclusion, curcumin treatment results in ROS generation, activation of Nrf2 and MAP kinases and the inhibition of phosphatase activity in hepatocytes, and when curcumin is not administered in toxic doses, these multiple pathways converge to induce HO-1.