Quantification of the 2-Deoxyribonolactone and Nucleoside 5′-Aldehyde Products of 2-Deoxyribose Oxidation in DNA and Cells by Isotope-Dilution Gas Chromatography Mass Spectrometry: Differential Effects of γ-Radiation and Fe<sup>2+</sup>−EDTA
作者:Wan Chan、Bingzi Chen、Lianrong Wang、Koli Taghizadeh、Michael S. Demott、Peter C. Dedon
DOI:10.1021/ja910928n
日期:2010.5.5
The oxidation of 2-deoxyribose in DNA has emerged as a critical determinant of the cellular toxicity of oxidative damage to DNA, with oxidation of each carbon producing a unique spectrum of electrophilic products. We have developed and validated an isotope-dilution gas chromatography-coupled mass spectrometry (GC-MS) method for the rigorous quantification of two major 2-deoxyribose oxidation products:
DNA 中 2-脱氧核糖的氧化已成为 DNA 氧化损伤细胞毒性的关键决定因素,每个碳的氧化都会产生独特的亲电子产物谱。我们开发并验证了一种同位素稀释气相色谱-质谱联用 (GC-MS) 方法,用于严格定量两种主要的 2-脱氧核糖氧化产物:1'-氧化的 2-脱氧核糖内酯脱碱基位点和核苷 5' 5'-氧化化学的-醛。该方法需要消除这些产物分别为 5-亚甲基-2(5H)-呋喃酮 (5MF) 和糠醛,然后用五氟苯肼 (PFPH) 衍生化,添加同位素标记的 PFPH 衍生物作为内标,提取衍生物,并通过 GC-MS 分析进行定量。该方法的精密度和准确性通过含有 2-脱氧核糖内酯和核苷 5'-醛损伤的寡脱氧核苷酸进行验证。此外,在对照研究中利用了烯二炔抗生素、新制癌素和加利车霉素 γ(1)(I) 的明确定义的 2-脱氧核糖氧化化学,新制癌素每 10( 6) nt per microM 根据其已建立的次要