The effects of three risk factors for valproate (VPA) hepatotoxicity (i.e., young age, polypharmacy, and high VPA serum level) on the metabolism of VPA to its monounsaturated metabolites [2-en-VPA (2-en), 3-en-VPA (3-en) and 4-en-VPA (4-en)] were investigated in 106 patients treated with VPA (56 cases of monotherapy and 50 cases of polytherapy). In the monotherapy group, there was a significant negative correlation between age and 4-en/VPA ratio. In the same group, the 4-en/VPA ratio showed a significant positive correlation with serum VPA level, while 3-en/VPA and 2-en/VPA ratios showed significant negative correlations. In patients greater than 10 years, the 4-en/VPA ratio was significantly higher, while the 2-en/VPA ratio was significantly lower in the polytherapy group than in the monotherapy group. Our results indicate that all three risk factors clearly increase the metabolic conversion of VPA to 4-en, the most toxic VPA metabolite, and that polytherapy and high VPA serum level result in the inhibited beta-oxidative metabolism of VPA to 2-en. These altered VPA metabolic profiles are strikingly similar to the abnormal VPA metabolism previously reported in cases with fatal hepatic failure. Although VPA-induced fatal hepatotoxicity has been regarded as an idiosyncratic reaction, it is possible that these three factors enhance susceptibility to VPA hepatotoxicity by altering the metabolism of VPA.