Kyoritsu University of Pharmacy, Minato-ku, Tokyo, Japan.
Bioorg Med Chem. 2010 Dec 1;18(23):8284-8. doi: 10.1016/j.bmc.2010.10.002. Epub 2010 Oct 8.
N-Nitrosodialkylamines are known to be potent indirect-acting mutagens/carcinogens, which are activated by cytochrome P450. The reaction product of N-nitroso-N-methylbutylamine (NMB) with modified Fenton's reagent supplemented with copper salt (Fe²(+)-Cu²(+)-H₂O₂) was reported to be mutagenic in Salmonella typhimurium TA1535 without S9 mix. In this study, the NMB activation mechanism was investigated by ESR spectroscopy with radical trapping agents to detect radical species and also by observing changes in mutagenic potency with a Salmonella strain in the Ames assay in the presence of radical trapping agents. In ESR spectroscopy experiments, the hydroxyl radical generated from the modified Fenton's reagent was detected using the hydroxyl radical trapping agent 5,5-dimethyl-1-pyrroline N-oxide (DMPO). Since the amount of the DMPO-OH adduct decreased with the addition of NMB, hydroxyl radical was presumed to react with NMB followed by the generation of nitric oxide (NO), which was detected as CarboxyPTI through reaction with 2-(4-carboxyphenyl)-4,4,5,5-tetramethylimidazoline-1-oxyl 3-oxide (CarboxyPTIO). The mutagenicity of the reaction extract decreased following the addition of DMPO or CarboxyPTIO. Furthermore, the mutagenicity of the reaction product in the presence of DMPO was enhanced by the addition of NO. The reaction product from NMB with Fe²(+)-Cu²(+)-NO in the absence of H₂O₂ was mutagenic, and this activity increased with the introduction of additional NO. These findings suggest that hydroxyl radical takes part in the generation of NO from NMB and that NO plays an important role in NMB activation in the presence of Fe²(+) and Cu²(+).
N-亚硝胺是已知的强间接致突变/致癌物质，可被细胞色素 P450 激活。有报道称，N-亚硝-N-甲基丁基胺（NMB）与改良芬顿试剂（补充铜盐（Fe²(+)-Cu²(+)-H₂O₂）的反应产物在没有 S9 混合物的情况下在鼠伤寒沙门氏菌 TA1535 中具有致突变性。在这项研究中，通过 ESR 光谱学使用自由基捕获剂来检测自由基种类，并通过在存在自由基捕获剂的情况下用沙门氏菌菌株在 Ames 测定中观察致突变能力的变化来研究 NMB 的激活机制。在 ESR 光谱学实验中，使用羟基自由基捕获剂 5,5-二甲基-1-吡咯啉 N-氧化物（DMPO）检测改良芬顿试剂产生的羟基自由基。由于 DMPO-OH 加合物的量随着 NMB 的加入而减少，因此假定羟基自由基与 NMB 反应，随后生成一氧化氮（NO），通过与 2-(4-羧基苯基）-4,4,5,5-四甲基咪唑啉-1-氧-3-氧化物（CarboxyPTIO）反应检测到一氧化氮。添加 DMPO 或 CarboxyPTIO 后，反应提取物的致突变性降低。此外，添加 NO 后，DMPO 存在下反应产物的致突变性增强。在没有 H₂O₂ 的情况下，NMB 与 Fe²(+)-Cu²(+)-NO 的反应产物具有致突变性，并且该活性随着引入更多的 NO 而增加。这些发现表明，羟基自由基参与 NMB 中 NO 的生成，并且在存在 Fe²(+)和 Cu²(+)的情况下，NO 在 NMB 激活中起重要作用。
