TANDEM MASS SPECTROMETRY FOR ASSESSING DNA DAMAGE

用于评估 DNA 损伤的串联质谱法

• 批准号: 6493950
• 负责人: MICHAEL L GROSS
• 金额: $ 16.87万
• 依托单位: UNIVERSITY OF NEBRASKA MEDICAL CENTER
• 依托单位国家: 美国
• 财政年份: 1988
• 资助国家: 美国
• 起止时间: 1988-12-01 至 2006-06-30
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/6493950
• 关键词: DNA damage; adduct; carbopolycyclic compound; chemical conjugate; electrospray ionization mass spectrometry; estrogens; liquid chromatography mass spectrometry; matrix assisted laser desorption ionization; oligonucleotides

A unifying goal for this research is to understand the mechanisms of fragmentations so that interpretations of production-ion spectra can be placed on a firm foundation. Therefore, the first aim is to evaluate various spectrometry methods to distinguish isomeric adducts and potential biomarkers. New to our effort will be a major activity to understand isomerically distinct fragmentations by using molecular orbital calculations. Six classes of fragmentations are identified for study on the basis of our experience in this area. In parallel with developing an understanding of ion chemistry, we will develop and validate electrospray liquid chromatography (LC/MS) and LC-tandem-MS methods for the trace determination of estrogen metabolites, conjugates, and nucleobase adducts. We are proposing two approaches: a low-throughput method that posses maximal certainty for identification of analytes and a high- throughput method for rapid screening of tissues. These methods will then be used to support research in other subprojects. Emphasis will be on analysis of tissues for estrogen biomarkers and on biological fluids for polycyclic aromatic hydrocarbon (PAH) adducts. We will also emphasize a biophysical aspect of MS in which the instrumentation is used to evaluate the binding of carcinogens to DNA by investigating non- covalent complexes of small duplex oligodeoxynucleotides and estrogens or PAH. Following the study of interactions, we focus on the modification reactions themselves by devising and utilizing MS methods for following directly the reactions of carcinogens with model single and double-stranded oligonucleotides. Sufficient preliminary results support the launch of these latter, relatively new projects.

这项研究的一个统一目标是了解碎裂机制，以便为生产离子光谱的解释奠定坚实的基础。因此，首要目标是评估各种光谱测定方法来区分异构加合物和潜在的生物标志物。我们的新努力将是通过使用分子轨道计算来了解异构不同碎片的主要活动。根据我们在该领域的经验，确定了六类碎片进行研究。在加深对离子化学的了解的同时，我们将开发和验证电喷雾液相色谱 (LC/MS) 和 LC-串联 MS 方法，用于痕量测定雌激素代谢物、缀合物和核碱基加合物。我们提出两种方法：一种对分析物鉴定具有最大确定性的低通量方法，以及一种对组织进行快速筛选的高通量方法。这些方法将用于支持其他子项目的研究。重点是分析组织中的雌激素生物标志物和生物体液中的多环芳烃 (PAH) 加合物。我们还将强调 MS 的生物物理方面，其中仪器用于通过研究小双链寡脱氧核苷酸和雌激素或多环芳烃的非共价复合物来评估致癌物与 DNA 的结合。在研究相互作用之后，我们通过设计和利用 MS 方法直接跟踪致癌物与模型单链和双链寡核苷酸的反应，重点关注修饰反应本身。足够的初步结果支持了后者相对较新的项目的启动。