ANALYSTS OF DBP DNA ADDUCTS

本项目将对体内形成的消毒液副产物（DBP） DNA加合物进行全面检测和部分结构解析。分析将依赖于荧光后标记，毛细管电泳-激光诱导荧光检测（CE-LIF），高效液相色谱和质谱（MS）技术的组合。CE-LIP检测低amol水平的核苷酸标准（与32P-标记后相同的超灵敏度）。将检测暴露于四种DBP化学品（溴二氯甲烷、氯酸钠、3-氯-4-（二氯甲基）-5-羟基-2 (5H)-呋喃酮和二溴乙酸）的动物的组织样本（由国家毒理学计划提供）。由于用于加合物标记的染料具有显著的质量，因此所得到的染料- dna加合物偶联物可以用基质辅助激光解吸电离(MALDI)-质谱法以高灵敏度（低fmol水平）检测。飞行时间和傅里叶变换质谱仪器都将被使用，后者提供精确的质量和（MS）n测量，以增强该项目中发现的未知DNA加合物的部分结构解析。按照建议，在这些暴露的动物中全面、明确地检测DNA加合物，同时监测致癌性，这是朝着了解暴露于这些化学品对人类造成的相应风险迈出的良好的第一步。CE-LIF的超灵敏度使得该方法很可能在需要时对未来的人类样本分析有用。

Comprehensive detection and some structural elucidation of disinfectant by-product (DBP) DNA adducts formed in vivo will be undertaken in this project. The analysis will rely on a combination of fluorescence post- labeling, capillary electrophoresis-laser induced fluorescence detection (CE-LIF), HPLC and mas spectroscopy (MS) techniques. CE-LIP detects nucleotide standards at the low amol level (same ultra-sensitivity as 32P- post-labeling). Tissue samples (furnished by NTP) from animals exposed to four DBP chemical (bromodichloromethane, sodium chlorate, 3- chloro-4-(dichloromethyl)-5-hydroxyl-2 (5H)-furanone and dibromoacetic acid) will be tested. Because the dye for adduct labeling has significant mass, the resulting dye-DNA adduct conjugates can be detected with high sensitivity (low fmol level) by matrix-assisted laser desorption ionization (MALDI)-MS. Both time-of-flight and Fourier transform MS instruments will be used, with the latter providing accurate mass and (MS)n measurements to enhance the partial structural elucidation of unknown DNA adducts to be discovered in this project. Comprehensive, definitive detection of DNA adducts in these exposed animals as proposed, with simultaneous monitoring of carcinogenicity, is a good, first step towards understanding the corresponding risk to humans from exposure to these chemicals. The ultra-sensitivity of CE-LIF makes it likely that the methodology will be useful, as needed, for the analysis of human samples in the future.