IHB OpenIR  > 学位论文
六溴环十二烷(HBCDs)立体异构体的生物累积特征及其细胞毒性研究
Alternative TitleStudy on bioaccumulative characteristics and cytotoxicity of hexabromocyclododecane stereoisomers
张晓岭
Subtype博士
Thesis Advisor徐盈
2008-06-15
Degree Grantor中国科学院水生生物研究所
Place of Conferral水生生物研究所
Keyword六溴环十二烷 Uplc/ms 手性对映体 电子废物 细胞毒性
Abstract六溴环十二烷(HBCDs)是一种广泛存在于环境中的新型持久性有机污染物(POPs)。近年来,HBCDs的环境问题正日益引起国际上的高度关注。本文建立了环境样品中痕量HBCDs的超高效液相色谱质谱联用(UPLC/MS)定性定量分析方法,调查了我国典型电子废物拆解地区土壤、蔬菜、底泥和淡水生物样品中HBCDs的浓度水平及其立体异构体构成。同时,应用高效液相色谱法(HPLC)分别在反相C18柱和PM-β-环糊精柱上成功分离制备出高纯度的α-, β-和γ-HBCD三种非对映异构体及(+)和(-)-α-, β-和γ-HBCD六种手性对映体标样。在此基础上,以人肝癌细胞株Hep G2为离体模型,综合评价了HBCDs立体异构体的细胞毒性。所获得的主要研究结果如下: 1. 建立了环境样品中痕量HBCDs的UPLC/MS分析方法。样品经索氏抽提后,过混合硅胶和氧化铝色谱柱分离纯化,HBCDs用UPLC/MS在ESI模式下以选择离子监测(SIM)方式进行检测。方法的添加回收率为79.3±10.2%-109.9±13.9%,α-, β-和γ-HBCD的检出限分别为20、45 和 15 pg。 2. 利用UPLC/MS法对我国典型电子废物拆解地区土壤、蔬菜、底泥和淡水生物样品进行分析发现,HBCDs在土壤和蔬菜中的平均浓度分别为61.34±31.25和458.87±296.43 pg/g 干重;但在底泥、田螺、鲫鱼和泥鳅中的平均浓度分别达到了14.19 ±10.88 ng/g 干重,186.16±83.16、377.27±189.97和1791.27±897.77 ng/g 脂重,这些结果表明我国局部地区的淡水生物已受到HBCDs的严重污染。 3. 研究发现,HBCDs的生物累积具有非对映体和手性对映体的选择性。底泥样品中作为最主要优势异构体的γ-HBCD,约占总HBCDs的62.94%,而在生物样品中却以α-HBCD为主要优势异构体,约占总HBCDs的63.12-77.27%。这可能是因为γ-HBCD在生物体内的代谢速率更快所致;就手性对映体而言,田螺选择性地累积(-)-α-和-γ-HBCD,而鱼类样品则选择性地累积(+)-α-和-γ-HBCD,表明手性对映体生物累积的选择性累积具有物种的差异。 4. 首次利用高效液相色谱的反相C18柱和β-环糊精柱,从技术级HBCD混合物中成功制备出HBCDs手性对映体标样 α-, β-和γ-HBCD,及其相应的手性对映体。经UPLC/MS鉴定发现,各异构体纯度>98%,为下一步HBCDs的毒理学研究创造了条件。 5. 以Hep G2细胞为模型,首次开展了HBCDs异构体的毒性研究。不同细胞毒性终点的试验结果一致表明, HBCDs非对映异构体的细胞毒性大小顺序为γ-HBCD ≥ β- HBCD > α-HBCD,而其手性对映体的细胞毒性大小顺序为:右旋异构体((+) α-, (+) β- and (+) γ-HBCD) 大于其相应的 左旋异构体((-) α-, (-) β- and (-) γ-HBCD)。研究还发现,HBCDs异构体诱导的胞内活性氧(ROS)水平和LDH释放之间具有良好的线性关系,这预示着HBCD异构体的致毒机制可能介导于氧化损伤。 以上研究结果为深入了解和研究HBCDs立体异构体,尤其是其手性对映体的环境行为,生物毒性及其生态风险评价提供了重要的科学依据。
Other AbstractHexabromocyclododecanes (HBCDs) are ubiquitous and emerging persistent organic pollutants (POPs). In recent years, environmental problem on HBCDs are attracting tremendous attention. In the present study, a qualitative and quantitative Ultra-High Performance Liquid Chromatography-mass spectrometric (UPLC/MS) method was established to investigate the contamination of HBCD stereoisomers in soils, vegetable, sediments and freshwater species from a typical electronic and electrical waste (EEW) dismantling area in China. Meantime, pure HBCD diastereoisomers (α-, β- and γ-HBCD) and their enantiomers ((+) and (-)-α-, β- and γ-HBCD) were prepared on an RP-C18 column followed by a PM-β-CD column using high performance liquid chromatography (HPLC). Their cytotoxicities were then evaluated in Hep G2 cells. The main results are as below: 1. An UPLC/MS method for determination of trace HBCDs in environment samples was developed. The sample was Soxhlet extracted and the extract was then separated and cleaned up on a multilayer silica gel and alumina column. HBCDs in the extract were determined by UPLC/MS with selected ion monitoring (SIM). The recoveries of spiked samples ranged from 79.3±10.2% to 109.9±13.9%. The limit of detection for α-, β-and γ-HBCD was 20, 45 and 15 pg, respectively. 2. The samples of soil, vegetable, sediment and freshwater organism were collected from a typical EEW dismantling area to determine HBCDs by UPLC/MS. It was found that average concentrations of HBCDs were 61.34±31.25 and 458.87±296.43 pg/g dry weight in soils and vegetables, respectively and up to 14.19 ±10.88 ng/g dry weight, 186.16±83.16, 377.27±189.97 and 1791.27±897.77 ng/g lipid weight in sediments, winkle, crucian carp and loach, respectively. These results indicated that the freshwater organisms were seriously contaminated by HBCDs. 3. It was found in the present study, the bioaccumulative characterics of HBCDs was diastereoselective and enantioselective for freshwater organisms. In details, γ-HBCD was the dominant diastereoisomer in the sediments (62.94% of total HBCDs). However, α-HBCD was selectively accumulated in the biotic samples (63.12-77.27%). The reason could be that γ-HBCD was more rapidly metabolized than α-HBCD in organism. In view of HBCD enantiomers, an enrichment of (-)-enantiomers of α- and γ-HBCD was found in the winkle, but, (+)-α- and -γ-HBCD selectively accumulated in fish, indicating that the selective bioaccumulative of HBCDs enantionmers was different from different species. 4. To the best our knowledge, the enantiomerically pure α-, β-, and γ-HBCD were firstly prepared . Racemic mixtures of α-, β-, and γ-HBCD were firstly prepared from the technical grade HBCDs using high performance liquid chromatography (HPLC) on RP-C18 column. Enantiomers of HBCDs were then obtained from the racemic mixtures of α-, β-, and γ-HBCD by HPLC on a permethylated β-cyclodextrin phase column. According to UPLC/MS analysis, the purities of individual HBCDs separated by HPLC were found to exceed 98% for diastereomers and enantiomers, which made the further toxicity study on HBCD stereoisomers possible. 5. The cytotoxicities of HBCD stereoisomers were evaluated in Hep G2 cells. Results from different toxicity-endpoints tests showed a good agreement that the order of cytotoxicity was γ-HBCD ≥ β- HBCD > α-HBCD, and that all (+)-enantiomers ((+) α-, (+) β- and (+) γ-HBCD) were significantly more toxic than their corresponding (-)-forms ((-) α-, (-) β- and (-) γ-HBCD). These results provide scientific basis for better understanding and further study environmental behavior, biological toxicity and ecological risk assessment of HBCD stereoisomers, especially enantiomers.
Pages110
Language中文
Document Type学位论文
Identifierhttp://ir.ihb.ac.cn/handle/342005/12292
Collection学位论文
Recommended Citation
GB/T 7714
张晓岭. 六溴环十二烷(HBCDs)立体异构体的生物累积特征及其细胞毒性研究[D]. 水生生物研究所. 中国科学院水生生物研究所,2008.
Files in This Item:
File Name/Size DocType Version Access License
10001_20051801193705(1643KB) 限制开放--Application Full Text
Related Services
Recommend this item
Bookmark
Usage statistics
Export to Endnote
Google Scholar
Similar articles in Google Scholar
[张晓岭]'s Articles
Baidu academic
Similar articles in Baidu academic
[张晓岭]'s Articles
Bing Scholar
Similar articles in Bing Scholar
[张晓岭]'s Articles
Terms of Use
No data!
Social Bookmark/Share
All comments (0)
No comment.
 

Items in the repository are protected by copyright, with all rights reserved, unless otherwise indicated.