Adam W, Saha-Möller C R, Schönberger A, Berger M, Cadet J
Institute of Organic Chemistry, University of Würzburg, Germany.
Photochem Photobiol. 1995 Aug;62(2):231-8. doi: 10.1111/j.1751-1097.1995.tb05263.x.
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)
将小牛胸腺DNA进行分离，然后用1,2 - 二氧杂环丁烷处理，这些二氧杂环丁烷包括3 - 乙酰氧基甲基 - 3,4,4 - 三甲基 - 1,2 - 二氧杂环丁烷、2,3 - 二甲基苯并呋喃二氧杂环丁烷、3 - 羟甲基 - 3,4,4 - 三甲基 - 1,2 - 二氧杂环丁烷（HTMD）、3,3,4,4 - 四甲基 - 1,2 - 二氧杂环丁烷和3,4,4 - 三甲基 - 1,2 - 二氧杂环丁烷（TrMD），它们在热分解时会生成三线态激发的羰基产物。为了定量监测诱变氧化产物7,8 - 二氢 - 8 - 氧代鸟嘌呤（8 - oxoGua）的形成，在二氧杂环丁烷处理过的DNA经酸性水解（氢氟酸/吡啶）后，采用灵敏且具选择性的高效液相色谱电化学分析法。用HTMD和TrMD可获得高产率的8 - oxoGua（高达约占可用鸟嘌呤的4%）。对二者在浓度、时间和温度影响方面进行了详细研究。将1,2 - 二氧杂环丁烷的氧化反应活性与几种I型（二苯甲酮和核黄素）和II型（亚甲蓝和孟加拉玫瑰红）光氧化剂以及作为单线态氧化学来源的1,4 - 乙烯基 - 2,3 - 苯并二恶英 - 1,4 - 二丙酸钠进行了比较。在与7,8 - 二氢 - 8 - 氧代 - 2'- 脱氧鸟苷（8 - oxodGuo）及氧化DNA的反应中，检测了8 - oxoGua对HTMD氧化作用的持久性。结果表明，长时间暴露于过量的HTMD时，氧化DNA中的8 - oxoGua确实会被消耗。8 - oxodGuo与HTMD反应生成了9 - (2 - 脱氧 - β - D - 赤藓戊呋喃糖基) - 4,8 - 二氢 - 4 - 羟基 - 8 - 氧代鸟嘌呤的两种4R和4S非对映异构体作为主要氧化产物。用叔丁醇进行的捕获实验证实不涉及羟基自由基，而使用三线态猝灭剂9,10 - 二溴蒽 - 2 - 磺酸钠和2,3 - 二氮杂双环[2.2.1]庚 - 2 - 烯确定三线态激发态主要通过I型作用（电子转移化学）导致观察到的DNA氧化。通过重水（D2O）与水（H2O）中的氘同位素效应测试了单线态氧的作用，但关于1O2是否参与这些氧化反应无法得出明确结论。（摘要截短于250字）
