ANALYSTS OF DBP DNA ADDUCTS

本项目将对体内形成的消毒剂副产物(DBP)DNA加合物进行全面的检测和一些结构鉴定。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学(MS)技术的组合。CE-LIP检测低氨基水平的核苷酸标准(与32P标记后的超灵敏相同)。将对暴露于四种DBP化学物质(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NTP提供)进行测试。由于用于加合物标记的染料具有显著的质量，因此利用基质辅助激光解吸电离(MALDI)-MS可以高灵敏度(低fmol水平)检测得到的染料-DNA加合物。将同时使用飞行时间和傅里叶变换MS仪器，后者提供准确的质量和(MS)n测量，以加强该项目中发现的未知DNA加合物的部分结构阐明。如建议的那样，对这些暴露的动物中的DNA加合物进行全面、明确的检测，并同时监测致癌性，是了解暴露于这些化学品对人类的相应风险的良好的第一步。CE-LIF的超高灵敏度使该方法有可能在需要时用于未来人体样本的分析。

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.