Are pyridinethiones reliable photochemical oxyl-radical sources for photobiological studies?: The importance of secondary photolysis products in the guanine oxidation of 2′-deoxyguanosine and cell-free DNA

The photolysis of the pyridinethiones 1a-d releases oxyl (hydroxyl, isopropyloxyl, tert-butoxyl, and benzoyloxyl) radicals, which damage DNA through strand cleavage and guanine oxidation. Unexpectedly, the N-hydroxy derivative 1a is significantly less effective in the oxidation of cell-free DNA than the alkoxy and benzoyloxy ones 1b-d. A similar trend was observed for the photooxidative consumption of 2'-deoxyguanosine (dG) by these pyridinethiones. Detailed mechanistic investigations have revealed that the guanine oxidation by the pyridinethiones 1b-d is not caused by oxyl radicals but by a type-I photooxidation process through the novel betain intermediate 8, which is formed in small amounts (ca. 5%) during the photolysis of these thiones from the intermediary photoproduct disulfide 3. This photosensitized guanine oxidation is effectively inhibited by the N,N'-dioxide 7, which is produced only in the photolysis of the N-hydroxypyridine-2-thione (1a) and not from the N-alkoxy and N-benzoyloxy derivatives 1b-d. Thus, for the N-hydroxy derivative 1a hydroxyl radicals are the main DNA-damaging species. The N-alkoxy and N-benzoyloxy derivatives 1b-d are more effective DNA-photooxidizing reagents than the N-hydroxypyridinethione 1a, because 1b-d oxidize DNA photocatalytically through sensitization by the betain, while 1a oxidizes DNA mainly through the stoichiometrically photogenerated hydroxyl radicals.