Photochemistry and Photobiology of Furocoumarin Hydroperoxides Derived from Imperatorin: Novel Intercalating Photo-Fenton Reagents for Oxidative DNA Modification by Hydroxyl Radicals

Photochemical and photobiological properties of the imperatorin-derived furocoumarin hydroperoxides 1a, 1a′, 2a and 2a′ have been investigated. Irradiation (350 nm) of the hydroperoxide 2a′ afforded the alcohol 2b (2%), a diastereomeric mixture of the hydroxy epoxide 2c (40%; diastereomeric ratio = 80:20) and the epoxide 2d (8%). The formation of these products was rationalized in terms of homolysis of the hydroperoxide bond initiated by intramolecular energy transfer from the photoexcited furocoumarin chromophore. The quantum yields for the photolytic decomposition of hydroperoxides were estimated to be in the range of 0.03-0.85 and decreased in the order 2a ≫ 2a′ ≫ 1a′ ≥ 1a. The involvement of hydroxyl radicals in these reactions was established by trapping experiments with benzene and spectroscopic evidence was obtained by EPR spin trapping with 5,5-dimethylpyrroline-N-oxide. Fluorescence titration, DNA melting and linear dichroism studies of furocoumarins indicated that these compounds undergo efficient complexation and also intercalation into the DNA. The binding parameters K (intrinsic binding constant) and 1/n (frequency of binding sites) of complexes between furocoumarin derivatives and DNA were determined to be in the range of 3900-23900 M-1 and 0.017-0.045. The photoreaction of 1a′ and 1b′ with 2′-deoxyguanosine (dGuo) afforded exclusively 7,8-dihydro-8-oxo-2′-deoxy-guanosine (8-oxodGuo), presumably through singlet oxygen, which was formed in a type II photooxidation process. In contrast, the hydroperoxide 2a oxidized dGuo to oxazolone as major and 8-oxodGuo as minor products through hydroxyl radicals, which were generated from 2a under photolytic conditions. Interestingly, the photoreactions of furocoumarins with salmon testes DNA showed that the highly reactive (φ = 0.85) hydroperoxide 2a is also most efficient in inducing the mutagenic DNA oxidation product 8-oxodGuo. Hence, the novel furocoumarin hydroperoxide 2a constitutes the first intercalating photo-Fenton reagent and serves as convenient hydroxyl radical source for genotoxicity studies.