SYNTHESIS AND REACTIVITY OF COUMARIN 3,4-EPOXIDE

Coumarin is used widely as a fragrance constituent and is administered clinically in the treatment of certain lymphedemas and malignancies. Although toxicity occurs only rarely its humans treated clinically wit h high-dose coumarin, it is well established that coumarin is hepatoto xic in the rat. This species difference in susceptibility to toxicity reflects the disparate metabolic processes occurring in humans and rod ents. In humans, coumarin is converted extensively via cytochrome P450 2A6 to the nontoxic 7-hydroxycoumarin metabolite, In contrast, coumar in 3,4-epoxidation is thought to predominate in rodent species, result ing in the formation of several potentially toxic metabolites. Coumari n epoxide is thought to be highly unstable end has not been Isolated s ynthetically or as a microsomal product. To address this issue, coumar in 3,4-epoxide was synthesized, and its stability and fate have been d etermined. Coumarin 3,4-epoxide was prepared by reacting coumarin with dimethyldioxirane. The epoxide was stable in organic solvents and sur vived conditions required for analysis by gas chromotography. Its stru cture was confirmed via H-1-NMR and gas chromatography-mass spectromet ry-infrared spectroscopy (GC-MS-IR). In contrast, coumarin 3,4-epoxide was unstable in aqueous solution, converting within 20 sec to a ring- opened compound, Using GC-MS-IR analysis, the single coumarin 3,4-epox ide product was identified as o-hydroxyphenylacetaldehyde (o-HPA). Alt hough other investigators have suggested that 3-hydroxycoumarin is an intermediate in o-HPA formation from coumarin 3,4-epoxide, we have dem onstrated that 3-hydroxycoumarin, incubated in an aqueous system or wi th liver microsomal proteins, does not form o-HPA. Thus, the results o f the present work establish that coumarin 3,4-epoxide can be synthesi zed and that o-HPA, which has previously been shown to be a prominent coumarin metabolite in rat liver microsomal incubations, is formed dir ectly from coumarin 3,4-epoxide. These results suggest that both couma rin 3,4-epoxide and o-HPA may contribute to the hepatotoxicity of coum arin.