基于污染谱解析的饮水有机污染物混合暴露遗传毒性研究

Genotoxicity of mixture exposure of pollutants in drinking water has received considerable attention. Since pollutants in water are extremely complicated, previous study only performed qualitative description of water extracts through certain mutagenicity test because of limitation of techneques and methods. With the development of techneques, it is possible to identify characteristics of mixture in drinking water through pollution profile analysis and investigate its genotoxicity and features using genotoxicity assessment methods based on multiple effect endpoints. The current study is to investigate pollution profile features and pollutant categories of source water and drinking water of Qingcaosha Reservior in Shanghai using GC-MS and LC-MS, and then to investigate genotoxicity and individual contribution to genotoxicity of mixture pollutants from full enrichment and combination of compounds belonging to same or different categories based on classification of structure similarity according to analysis results of pollution profile, using Ames test, cytokinesis-block micronucleus cytome assays and in vitro γH2AX assay with high content screening, and PIG-A gene mutation assay. To get accurate understanding of genotoxic pollutants in water, we are to separate pollutants of enrichment using preparative chromatography, select multiple peak intervals according to characteristics of peak time and peak value, collect pollutants in subsection, and detect their genotoxicity. For positive components, qualitative and quantitative analysis is performed using GC-MS and LC-MS, and bioinformatics techneque is used in order to determine pollutants causing genotoxicity. The current study will provide a basis for genotoxicity assessment of mixture exposure in drinking water.

饮水污染物混合暴露遗传毒性备受重视。受技术制约，以往研究仅以某种致突变测试方法对水中提取物进行定性描述。随着技术发展，通过污染物谱分析揭示饮水混合污染物特征，并以多效应终点遗传毒性评价方法研究其遗传毒性及特征成为可能。本研究以GC-MS和LC-MS技术研究上海青草沙水源水及其饮水污染谱特征。以Ames试验、胞质阻滞微核细胞组学和γH2AX高内涵筛选试验、PIG-A基因检测，研究完整富集污染物和根据污染物谱分析结果，将结构相似污染归类，研究相同/不同类别组合时，混合污染物的遗传毒性，并分析各类污染物对遗传毒性的贡献率。为准确认识水中遗传毒性污染物，以制备色谱分离富集污染物，据出峰时间和出峰量特征，选择多个出峰区间，分段收集污染物，检测遗传毒性。对阳性组份，以GC-MS和LC-MS定性/量分析，利用生物信息学等技术，明确引起遗传毒性的污染物。研究将为饮水污染物混合暴露遗传毒性研究提供依据。

准确评价饮水污染物混合暴露特征和混合暴露遗传毒性是国内外相关研究领域关注的重要问题。以往研究仅以某种致突变测试方法对水中提取物进行定性描述。随着技术发展，通过污染物谱分析揭示饮水混合污染物特征，并以多效应终点遗传毒性评价方法研究其遗传毒性及特征成为可能。本研究优化了基于XAD-2-COCA双柱串联SPE-GC-MS检测水中半挥发性有机污染物的方法，解析了枯水期和丰水期原水和出厂水半挥发性有机污染谱，分别利用GC-MS和UPLC-MS/MS技术分析了溴代阻燃剂和有机磷阻燃剂污染情况。结果显示，枯水期和丰水期原水和出厂水中各检出一定数量半挥发性有机污染物和有机磷阻燃剂，丰水期污染比枯水期严重，亚硝胺类和氯酚类浓度在出厂水中有所增加。原水和出厂水中各有一定数量物质纳入我国《生活饮用水卫生标准》（GB5749-2006），但均未超标，同时仍有一定数量列入我国和美国EPA水中优先控制污染物清单的物质被检出，需引起重视。分别利用Ames试验、CBMN Cyt试验、γH2AX试验和小鼠骨髓细胞体内微核试验评价枯水期和丰水期原水和出厂水水样有机提取物遗传毒性，结果表明致突变性为阴性，能够引起染色体损伤和DNA双链断裂，出厂水有机提取物遗传毒性强于原水有机提取物，丰水期强于枯水期；PAHs对原水和出厂水有机提取物引起的染色体损伤作用有贡献，NAms对出厂水有机提取物引起的DNA双链断裂作用有贡献。对制备色谱获得的遗传毒性阳性区段利用GC-MS-AMDIS技术进行解析，并结合毒理学数据库，确定了与原水和出厂水有机提取物混合暴露导致遗传毒性有关的物质。本研究以上海市以青草沙水库为水源的某水厂为研究对象，分析枯水期和丰水期原水和出厂水的有机污染谱，并研究有机提取物混合暴露的遗传毒性及其特征，利用制备色谱等技术明确青草沙水源的饮用水中具有遗传毒性的污染物，为揭示以青草沙水库为水源的饮用水水质状况和安全性提供了参考，为饮用水污染物混合暴露遗传毒性的评价和全面认识饮用水污染物混合暴露的健康效应提供了方法学参考。