【作者】 丁峰；

【导师】 宋文华；

【作者基本信息】 上海交通大学 ， 环境工程， 2009， 博士

【摘要】 利用定量结构-活性关系（quantitative structure-activity relationship, QSAR）方法开展的环境定量构效关系研究,是对环境中日益增多的各种化学品进行环境效应评价的重要手段。在有机污染化学和生态毒理学研究中,环境定量构效关系研究具有两个基本的功能:探求化学品的环境效应作用机制,为降低化学品对环境的风险提供理论指导;对未知化学品的环境行为和生态毒性进行预测、评价和筛选。因此理想的定量构效关系模型应该既具有较高的预测能力,又能够对化学品的环境效应作用机制做出合理的阐述。本论文以2-羟基-3-烷基-1,4-萘醌植物光系统II（photosystem II, PSII）抑制剂和三唑类杀菌剂为研究对象,通过QSAR研究方法,定量研究了萘醌类化合物的PSII抑制作用机制、活性位点以及三唑类杀菌剂对水生生物的生物致毒机制与活性位点和可能导致的环境效应,研究结果将有助于今后构建能应用于农药环境风险分析的环境定量构效关系模式并开展农药环境效应评价。利用二维定量构效关系（two-dimensional quantitative structure- activity relationship,2D-QSAR）和全息定量构效关系（holographical quantitative structure-activity relationship,HQSAR）法对23个2-羟基-3-烷基-1,4-萘醌化合物的PSII抑制活性进行了研究。并以此为基础,分别利用比较分子场分析（comparative molecular field analysis,CoMFA）和比较分子相似性指数分析（comparative molecular similarity indicesanalysis,CoMSIA）两种三维定量构效关系（three-dimensional quantitative structure-activity relationship,3D-QSAR）方法,对PSII抑制活性进行了研究。所建QSAR模型具有较高的拟合程度和预测能力,能够合理解释该类化合物PSII抑制作用机制。与2D-QSAR相比,HQSAR和3D-QSAR模型具有更高的拟合度和预测能力。QSAR研究表明,该类化合物对PSII抑制可能分为两步:首先化合物从水相迁移到生物相,穿过细胞膜到达作用靶点;接着3-烷基可能与D1蛋白非极性基团发生疏水作用,同时羰基氧原子与D1蛋白氨基酸残基发生氢键作用,从而替代质体醌,阻碍了植物光合作用过程中的电子传递,对PSII产生抑制作用。而3D-QSAR研究进一步指出,3-烷基取代基的形状/大小以及疏水性能够综合影响该类化合物穿透细胞膜的能力,并且4-羰基氧原子更易形成氢键。3-烷基和4-羰基很可能是该类抑制剂与D1蛋白的活性反应中心。为研究三唑类杀菌剂进入水环境后的水生生态毒性,并且为农药的QSAR研究提供基础数据,利用静态实验法分别测定了17种三唑类杀菌剂的48小时大型溞急性运动抑制毒性(EC₅₀（48h）)和96小时斑马鱼急性毒性(LC₅₀（96h）),并对该类农药对水生生态安全的影响进行了初步评价。利用2D-QSAR、CoMFA和CoMSIA两种3D-QSAR方法,对三唑类杀菌剂对斑马鱼和大型溞急性毒性进行研究≦SAR模型具有较高的拟合程度和预测能力,能够合理解释该类化合物对斑马鱼和大型溞的生物致毒机制。与2D-QSAR相比,3D-QSAR模型具有更高的拟合度和预测能力。2D-QSAR研究表明,该类化合物对斑马鱼和大型溞产生毒性效应可能为化合物首先穿过细胞膜在生物体内积聚,由疏水性参数（ClogP）和分子拓扑指数（ShpC和Dia）控制,化合物具有一定麻醉毒性;接着与生物大分子间发生电子交换,或亲核取代反应等,可用电性参数(E_LUMO、QC1和EQN1)描述,化合物具有反应性毒性并且高于麻醉作用。而3D-QSAR研究进一步表明,三唑环N1原子可能提供电子与细胞色素P450（cytochrome P450,CYP450）卟啉环活性中心亚铁原子形成配位键;苯环和烷基取代基可能与CYP450芳香受体部分发生疏水叠合,阻碍CYP450正常生理功能,从而对斑马鱼和大型溞产生毒性，这可能是该类化合物众多致毒机理中的一种。经过对2D-QSAR和3D-QSAR研究结果比较发现,三唑类杀菌剂对大型溞和斑马鱼的致毒机制类似，并且三唑环、苯环对位以及与硅原子相连的烷基可能是该类化合物与斑马鱼和大型溞生物大分子的活性反应中心。为验证3D-QSAR研究中三唑类杀菌剂对斑马鱼致毒机制的推测,并从分子水平预测该类农药对水生生物的影响与潜在的环境效应,以多效唑为例,测定了不同浓度多效唑对斑马鱼肝脏超氧化物歧化酶（superoxide dismutase,SOD）和过氧化氢酶（catalse,CAT）活性的影响。结果表明,SOD和CAT对多效唑的响应机制可能是由于CYP450酶系与污染物发生结合,阻碍了CYP450正常的催化氧化循环过程,使得活性氧（reactive oxygen species, ROS）生成量增加,从而影响了SOD和CAT活性。更多还原

【Abstract】 Environmental QSAR studies are the important means to evaluate the environmental effects caused by more and more chemicals existing in the environment. In the researches of organic chemistry about pollutants and ecotoxicology, environmental QSAR studies play two roles. One is to analyse the mechanism of environmental effects to offer the theoretical guidance of pollutants’environmental risk’s decreasing. The other is to predict, evaluate and screen of environmental behavior and eco-toxicity of unknown compounds. So, the ideal QSAR models not only have high predictive ability, but also can explain the mechanisms of environmental effects reasonably.In this paper, 2-hrdroxy-3-alkyl-1,4-naphthoquinones as inhibitors to PSII and triazole fungicides were regarded as research objects. The inhibitory mechanism and active sites of naphthoquinones to PSII, the toxic mechanism and active sites of triazoles to aquatic organisms and the environmental effects induced by triazoles were studied by using QSAR technique. The objective of the paper is to establish the environmental QSAR pattern to analyse and evaluate the environmental risk and effect of pesticides.2D-QSAR and HQSAR methods were used to study the PSII inhibitory activity of 23 2-hydroxy-3-alkyl-1,4-naphthoquinone derivatives. Based on it, 3D-QSAR including CoMFA and CoMSIA were employed to study the inhibitory activity. The QSAR models having the high fitting degrees and predictive abilities could interpret the PSII inhibitory mechanism reasonably. Compared with 2D-QSAR, HQSAR and 3D-QSAR models had the higher predictive abilities. The results showed that the inhibitory process might include two steps. Firstly, the inhibitors migrated from water to biological phase, arrived at the target through the membrane. Secondly, the inhibitors were possible to combine with the target via 3-alkyl’s hydrophobic interaction with the non-polar groups of D1 and acyl-oxygen’s H-bond interaction with the amino acid residues of D1. Thus, naphthoquinones competed with QB to block the electron’s transfer process of plant photosynthesis. 3D-QSAR results showed that the shape, size and hydrophobicity of 3-alkyl could affect the ability of compound through the membrane, and the 4-alcyl-oxygen could combine with D1 using H-bond easily. 3-alkyl and 4-carbonyl were probably the active sites of inhibitory reaction.BTo analyse the aquatic ecological toxicity of 17 triazole fungicides and provide the data for QSAR study, the acute immobilization test for the crustacean （Daphnia magna） at 48h (expressed as EC₅₀（48h）) and acute toxicity for the zebrafish （Danio rerio） at 96h (expressed as LC₅₀（96h）) were determined by static system. At the same time, the infections of traizoles to the safety of aquatic ecology were basically assessed.2D-QSAR and 3D-QSAR methods including CoMFA and CoMSIA were used to study the toxicities of triazoles to D. magna and D. rerio. The QSAR models having the high fitting degrees and predictive abilities could interpret the mechanism of toxicity to D. magna and D. rerio reasonably. Compared with 2D-QSAR, 3D-QSAR models had the higher predictive abilities. 2D-QSAR results showed that the pollutants firstly might accumulate in organisms through the membrane. The process could be controlled by the hydrophobic parameters （ClogP） and molecular topological indices （ShpC and Dia）. The pollutants might have the anaesthetic effect. Then, the pollutants could change the electron or nucleophilically displace with the biological macromolecules which could be affected by the electrical parameters. The pollutants might also have the reactive toxicity, higher than the anaesthetic toxicity. Moreover, 3D-QSAR studies indicated that the nitrogen atom in triazole ring might provide the lone pair electrons to form coordinate bond with the iron atom in the active center of CYP450 porphyrin ring. Phenyl and alkyl groups could form the hydrophobic interaction with the aromatic receptor of CYP450 and block the normal physiological function of CYP450. Therefore, triazole fungicides are toxic to D. magna and D. rerio. Comparing the results of 2D-QSAR and 3D-QSAR studies, one phenomenon that the toxic mechanisms of triazoles to D. magna were similar to D. rerio was found. Additionally, triazole ring, para positions of benzene ring and the substituted group linked with silicon atom were probably the active sites of toxic reaction.In order to validate the conjecture about the mechanism of triazoles on D. rerio in 3D-QSAR study and forecast the infection and potential environmental effects of triazoles to aquatic organisms on molecular level. The activities of SOD and CAT in the livers of zebrafish exposed to the various concentrations of paclobutrazole were measured, paclobutrazole as an example. The results indicated that the responses of SOD and CAT’s activities to triazoles were likely to be because of the interaction between CYP450 and triazoles which resulted in the interruption of natural cyclic catalytic reactions and then, the ROS increasing and the activities of SOD and CAT affected.更多还原