Frelon S, Douki T, Ravanat J L, Pouget J P, Tornabene C, Cadet J
Laboratoire Lésions des Acides Nucléiques, Service de Chimie Inorganique et Biologique, UMR 5046, Département de Recherche Fondamentale sur la Matière Condensée, CEA/Grenoble, F-38054 Grenoble Cedex 9, France.
Chem Res Toxicol. 2000 Oct;13(10):1002-10. doi: 10.1021/tx000085h.
A method involving high-performance liquid chromatography (HPLC) separation associated with tandem mass spectrometry (MS/MS) detection in the multiple-reaction monitoring mode was set up for the assessment of radiation-induced degradation products of DNA bases. This sensitive and specific assay is aimed at assessing six oxidized 2'-deoxyribonucleosides and two modified purine bases within both isolated and cellular DNA. For this purpose, stable isotopically labeled internal standards were prepared and used for isotope dilution mass spectrometry measurements. The latter method was validated through a comparison with two other assays, including HPLC associated with electrochemical detection and gas chromatography coupled to mass spectrometry. Using the specific and sensitive HPLC-MS/MS approach, 5,6-dihydroxy-5,6-dihydrothymidine, 5-hydroxy-2'-deoxyuridine, 5-(hydroxymethyl)-2'-deoxyuridine, 5-formyl-2'-deoxyuridine, 8-oxo-7,8-dihydro-2'-deoxyadenosine, 8-oxo-7,8-dihydro-2'-deoxyguanosine, 4, 6-diamino-5-formamidopyrimidine, and 2, 6-diamino-4-hydroxy-5-formamidopyrimidine were quantified within both isolated and cellular DNA upon exposure to gamma-radiation.
建立了一种在多反应监测模式下，结合高效液相色谱（HPLC）分离与串联质谱（MS/MS）检测的方法，用于评估辐射诱导的DNA碱基降解产物。这种灵敏且特异的分析方法旨在评估分离的和细胞中的DNA内的六种氧化2'-脱氧核糖核苷和两种修饰嘌呤碱基。为此，制备了稳定同位素标记的内标并用于同位素稀释质谱测量。通过与其他两种分析方法进行比较对后一种方法进行了验证，这两种方法包括与电化学检测联用的HPLC和与质谱联用的气相色谱法。使用特异且灵敏的HPLC-MS/MS方法，在分离的和细胞中的DNA暴露于γ辐射后，对5,6-二羟基-5,6-二氢胸苷、5-羟基-2'-脱氧尿苷、5-（羟甲基）-2'-脱氧尿苷、5-甲酰基-2'-脱氧尿苷、8-氧代-7,8-二氢-2'-脱氧腺苷、8-氧代-7,8-二氢-2'-脱氧鸟苷、4,6-二氨基-5-甲酰胺基嘧啶和2,6-二氨基-4-羟基-5-甲酰胺基嘧啶进行了定量分析。
