ANALYSTS OF DBP DNA ADDUCTS

本项目将对体内形成的消毒剂副产物（DBP）DNA加合物进行全面检测和部分结构解析。该分析将依赖于荧光标记后、毛细管电泳-激光诱导荧光检测（CE-LIF）、HPLC和质谱（MS）技术的组合。CE-LIP检测低水平的核苷酸标准品（与32 P-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（5 H）-呋喃酮和二溴乙酸）的动物的组织样本（由NTP提供）。因为用于加合物标记的染料具有显著的质量，所以可以以高灵敏度检测所得的染料-DNA加合物缀合物（低fmol水平）通过基质辅助激光解吸电离（MALDI）-MS。将使用飞行时间和傅里叶变换MS仪器，后者提供精确的质量和（MS）n测量，以提高未知的DNA加合物的部分结构说明将在这个项目中发现。按照建议，全面、明确地检测这些暴露动物体内的DNA加合物，同时监测致癌性，是了解暴露于这些化学品对人类造成的相应风险的良好的第一步。CE-LIF的超灵敏度使得该方法可能在未来根据需要用于人体样本的分析。

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.