We have investigated the importance of nitronate formation from 2-nitropropane (2-NP) for the oxidative metabolism and the genotoxicity of 2-NP in 2sFou rat hepatoma cells. Treatment of the cells with 2-NP for up to 3 h resulted in the time-dependent appearance of nitrite in the culture medium and in a weak induction of DNA repair synthesis. Both nitrite formation and repair induction were markedly enhanced in cells exposed to the anionic form of 2-NP, propane 2-nitronate. These observations suggest that propane 2-nitronate, rather than 2-NP itself, is oxidized by the liver cells to yield a DNA-damaging product. The results also indicate that the nitro/nitronate equilibration in intact liver cells is slow, suggesting that nitronate formation represents the rate-limiting step in the metabolic activation of 2-NP.