Formation of 7,8-dihydro-8-oxoguanine in the 1,2-dioxetane-induced oxidation of calf thymus DNA: evidence for photosensitized DNA damage by thermally generated triplet ketones in the dark.
Isolated calf thymus DNA was treated with the 1,2-dioxetanes 3-acetoxymethyl-3,4,4-tri-methyl-1,2-dioxetane, 2,3-dimethylbenzofuran dioxetane, 3-hydroxymethyl-3,4,4-trimethyl-1,2-dioxetane (HTMD), 3,3,4,4-tetramethyl-1,2-dioxetane and 3,4,4-trimethyl-1,2-dioxetane (TrMD), which on thermal decomposition generate triplet-excited carbonyl products. To monitor quantitatively the formation of the mutagenic oxidation product 7,8-dihydro-8-oxoguanine (8-oxoGua), a sensitive and selective HPLC electrochemical assay was used after acidic hydrolysis (HF/pyridine) of the dioxetane-treated DNA. High yields of 8-oxoGua (up to ca 4% of the available guanine) were obtained for HTMD and TrMD. Both were investigated in detail with respect to effects of concentration, time and temperature. The oxidative reactivity of 1,2-dioxetanes was compared with several type I (benzophenone and riboflavin) and type II (methylene blue and rose bengal) photooxidants and disodium 1,4-etheno-2,3-benzodioxin-1,4-dipropionate as a chemical source of singlet oxygen. The persistence of 8-oxoGua towards oxidation by HTMD was examined in the reaction with 7,8-dihydro-8-oxo-2'-deoxyguanosine (8-oxodGuo) and with oxidized DNA. It was shown that, indeed, 8-oxoGua is consumed in the oxidized DNA on prolonged exposure to an excess of HTMD. The reaction of 8-oxodGuo with HTMD afforded the two 4R* and 4S* diastereomers of 9-(2-deoxy-beta-D-erythropentofuranosyl)-4, 8-dihydro-4-hydroxy-8-oxoguanine as main oxidation products. Trapping experiments with tert-butanol confirmed that hydroxyl radicals are not involved, whereas the use of the triplet quenchers sodium 9,10-dibromo-anthracene-2-sulfonate and 2,3-diazabicyclo[2.2.1]hept-2-ene established that triplet-excited states are mainly responsible for the observed DNA oxidation through type I action (electron transfer chemistry). The role of singlet oxygen was tested by means of deuterium isotope effects in D2O versus H2O, but no definitive conclusion could be reached in regard to the involvement of 1O2 in these oxidations.(ABSTRACT TRUNCATED AT 250 WORDS)
D000818:Animals; D002417:Cattle; D004247:DNA / Q000187:drug effects* / Q000378:metabolism / Q000528:radiation effects*; D004249:DNA Damage; D003624:Darkness; D006147:Guanine / Q000031:analogs & derivatives* / Q000378:metabolism; D006361:Heating; D006571:Heterocyclic Compounds / Q000633:toxicity*; D006573:Heterocyclic Compounds, 1-Ring; D007659:Ketones / Q000737:chemistry / Q000633:toxicity*; D010084:Oxidation-Reduction; D011836:Radiation Tolerance; D013950:Thymus Gland / Q000737:chemistry
D006571:Heterocyclic Compounds; D006573:Heterocyclic Compounds, 1-Ring; D007659:Ketones; C079142:1,2-dioxetane; C024829:8-hydroxyguanine; D006147:Guanine; D004247:DNA