对羟基苯甲酸酯雄激素受体干扰效应及邻苯二甲酸酯对青海弧菌生物毒性

发布时间：2018-07-07 10:45

本文选题：对羟基苯甲酸酯 + 邻苯二甲酸酯　；参考：《浙江大学》2017年硕士论文

【摘要】：对羟基苯甲酸醋类(Parabens)和邻苯二甲酸醋类(Phthalate esters,PAEs)物质作为抗菌防腐剂和塑化剂被广泛的添加到个人护理品、消费品中,其残留在多种环境介质和生物体内被频繁检出,目前关于其人体健康和生态环境风险相关研究相对匮乏。本文选取不同侧链结构和类型Parabens和PAEs物质为研究对象,分析了parabens对雄激素受体(Androgen receptor,AR)干扰效应,考察了污染物水解后干扰效应的变化,探讨了其雄激素受体干扰效应的分子机制。以青海弧菌(Vibrio qinghaiensis sp.-Q67)为模式生物,研究了PAEs与镉(Cadimum,Cd)的单一和二元联合毒性,分析了重金属Cd胁迫对PAEs急性毒性的影响,并利用经典模型预测了PAEs与Cd的联合毒性效应。从构效关系角度分析了parabens及PAEs的侧链长度和基团类型对其毒性的影响。主要获得如下研究结果:(1)采用酵母双杂交方法研究了10种芳氧基和烷氧基侧链parabens对AR的干扰效应。发现parabens的烷氧基侧链C原子数为1-4时,AR拮抗效应随侧链增长而显著降低;当侧链C原子数为7,8和12时,parabens无显著AR拮抗效应。相比烷氧基侧链,具有芳氧基侧链的parabens如对经基苯甲酸苄醋(Benzyl paraben,BzP)和对经基苯甲酸苯醋(Phenyl paraben,PnP)具有相对较高的AR拮抗效应。分子对接(Molecular docking)揭示与天然雄激素二氢睾酯(Dihydrotestosterone,DHT)具有相似的结合模式,parabens与AR受体配体结合域(Ligand binding domain,LBD)877号苏氨酸和705号天冬酰胺形成氢键。Parabens与AR LBD的结合能量与其AR拮抗效应显著相关(R~2=0.84,P0.05)。为进一步分析parabens侧链对AR拮抗活性的影响,采用猪肝醋酶将parabens水解,发现水解后parabens的AR拮抗活性完全消除。高效液相色谱串联紫外检测器(HPLC-UV)及超高效液相色谱串联质谱检测器(UPLC-MS/MS)分析发现parabens水解后酯键断裂,生成对羟基苯甲酸。相较于芳氧基侧链parabens,烷氧基侧链parabens是猪肝酯酶的最优底物,更易于被水解。(2)以Vibrio qinghaiensis sp.-Q67为模式生物,研究了PAEs与Cd的单一和二元联合毒性。发现六种PAEs包括邻苯二甲酸二甲醋(Dimethyl phthalate,DMP)、邻苯二甲酸二乙酯(Diethyl phthalate,DEP)、邻苯二甲酸丁苄酯(Benzyl butyl phthalate,BBP)、邻苯二甲酸二正丁酯(Dibutyl phthalate,DBP)、邻苯二甲酸二异辛醋(Diisooctyl phthalate,DIOP)和邻苯二甲酸二正辛酯(Di-n-octyl phthalate,DOP)对青海弧菌急性毒性的EC50值大于100mg/L(DMP:134.4mg/L;DEP:283.1 mg/L;DBP:895.1 mg/L),说明对青海弧菌急性毒性较弱;随PAEs侧链长度增加,其急性毒性显著减小(R~2=0.8961,P0.05)。进一步测定了PAEs与Cd的二元联合毒性,并结合独立作用(Independent action,IA)和浓度加和(Concentration addition,CA)模型进行了预测探讨。发现二元混合物DMP-DEP,DEP-DBP和DMP-DBP的毒性单位(Toxicunit,TU)值均大于1.2,联合毒性表现为拮抗效应;二元混合物DMP-Cd,DEP-Cd和DBP-Cd的TU值介于0.8-1.2,联合毒性表现为简单相加效应。重金属Cd胁迫显著增加了PAEs对青海弧菌急性毒性。
[Abstract]:Parabens and Phthalate esters, PAEs, as antiseptic preservatives and plasticizers, are widely added to personal care products. In consumer products, their residues are frequently detected in a variety of environmental media and organisms, and the relative research on human health and ecological environment risks is relative. In this paper, the interference effects of parabens on androgen receptor (Androgen receptor, AR) were analyzed by selecting different side chain structures and types of substance Parabens and PAEs. The changes of interference effect after the hydrolysis of pollutants were investigated, and the molecular mechanism of the interference effect of androgen receptor was discussed, with Vibrio Qinghai (Vibrio qinghaiensis sp.-Q6). 7) for model organisms, the single and two element joint toxicity of PAEs and Cadimum (Cd) was studied. The effects of heavy metal Cd stress on the acute toxicity of PAEs were analyzed, and the combined toxicity of PAEs and Cd was predicted by the classical model. The effects of side chain length and group type on the toxicity of parabens and PAEs were analyzed from the point of view of structure-activity relationship. The results are as follows: (1) the interference effect of 10 aryl oxygen groups and alkoxy side chain parabens on the AR was studied by yeast two hybrid method. It was found that when the number of parabens alkoxy side chain C atoms was 1-4, the AR antagonistic effect decreased significantly with the side chain growth. When the number of side chain C atoms was 7,8 and 12, parabens had no significant AR antagonistic effect. The oxygen group side chain, with the aryl side chain parabens, has a relatively high AR antagonism to the Benzyl Paraben, BzP and Phenyl paraben (PnP). Molecular docking (Molecular docking) reveals a similar binding pattern with the natural androgen two hydrotestosterone (Dihydrotestosterone, DHT). The binding energy of Bens with AR receptor ligand binding domain (Ligand binding domain, LBD) 877 and No. 705 asparagine is significantly related to the AR antagonistic effect of the hydrogen bond.Parabens and AR LBD (R~2=0.84, P0.05). The AR antagonistic activity of rabens was completely eliminated. High performance liquid chromatography tandem UV detector (HPLC-UV) and ultra high performance liquid chromatography tandem mass spectrometry detector (UPLC-MS/MS) analysis found that the ester bond breaks after parabens hydrolysis and produces P hydroxybenzoic acid. Compared to the aryl side chain parabens, the alkoxy side chain parabens is the best substrate for porcine liver esterase, and the alkoxy side chain parabens is the best substrate. It is easy to be hydrolyzed. (2) a single and two element joint toxicity of PAEs and Cd is studied with Vibrio qinghaiensis sp.-Q67 as a model organism. Six kinds of PAEs include two methyl acetic acid (Dimethyl phthalate, DMP), two (Diethyl phthalate, DEP), phthalic two formate, and phthalic two a. Acid two n-butyl (Dibutyl phthalate, DBP), phthalic acid two isooctyl vinegar (Diisooctyl phthalate, DIOP) and ortho benzoate two n-octyl (Di-n-octyl phthalate, DOP) are more acute toxicities of Vibrio Qinghai than 100mg/L (DMP:134.4mg/L; The acute toxicity of PAEs and Cd was significantly reduced (R~2=0.8961, P0.05). The combined toxicity of PAEs and Cd was further measured, and the prediction was carried out in combination with the independent action (Independent action, IA) and concentration addition (Concentration addition, CA). The combined toxicity of DMP-Cd, DEP-Cd and DBP-Cd was higher than 1.2, and the TU value of two yuan mixture was 0.8-1.2, and the combined toxicity showed simple additive effect. Heavy metal Cd stress significantly increased the acute toxicity of PAEs to Vibrio Qinghai.
【学位授予单位】：浙江大学
【学位级别】：硕士
【学位授予年份】：2017
【分类号】：X171.5

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