Studies on the mechanism of coumarin-induced toxicity in rat hepatocytes: comparison with dihydrocoumarin and other coumarin metabolites.

Single doses of coumarin (125 mg/kg, ip) produced a depletion of hepatic nonprotein sulfhydryl groups (mainly reduced glutathione; GSH) in young male Sprague-Dawley rats after 2 hr and increased liver weight and produced hepatic centrilobular necrosis after 24 hr. Coumarin also produced time- and dose-dependent toxic effects in primary rat hepatocyte cultures. A marked reduction of GSH levels was also observed in vitro and this was not due either to the formation of oxidized glutathione (GSSG) or to the leakage of GSH and/or GSSG from the hepatocytes. Coumarin-induced toxicity in rat hepatocytes could be inhibited by the cytochrome P450 inhibitors ellipticine and metyrapone and potentiated by depleting hepatocyte GSH levels with diethyl maleate. In contrast to coumarin, dihydrocoumarin--which lacks the 3,4-double bond--produced little toxicity in rat hepatocytes either in vivo (127 and 254 mg/kg, ip) or in vitro. Similarly, coumarin was more toxic to rat hepatocytes than a number of known coumarin metabolites including 3- and 7-hydroxycoumarin and o-hydroxyphenylacetic acid. The results of these studies demonstrate a good in vivo/in vitro correlation for the effects of coumarin and dihydrocoumarin in rat hepatocytes. Furthermore, the data suggest that coumarin hepatoxicity in the rat is due to coumarin bioactivation by cytochrome P450-dependent enzymes to a toxic metabolite(s), which may be a coumarin 3,4-epoxide intermediate. GSH appears to protect against coumarin-induced toxicity possibly by the formation of conjugates with the toxic coumarin metabolite(s).