DEHP对土壤蚯蚓氧化胁迫及DNA损伤的研究

Oxidating Stress and DNA Damage of DEHP to Soil Earthworms

土壤环境中的酞酸酯污染日益严重,为了探讨和分析典型酞酸酯邻苯二甲酸二(2-乙基己基)酯(Di(2-ethylhexyl)phthalate,DEHP)对土壤动物的生态毒理效应,以赤子爱胜蚓为指示生物,暴露于DEHP浓度为CK、0.1、1、10、50 mg kg^(-1)人工土壤中,并于染毒后的7、14、21、28d取样测定。通过蚯蚓体内的超氧化物歧化酶(SOD)、过氧化氢酶(CAT)、过氧化物(POD)、谷胱甘肽转移酶(GST)等指标反映DEHP对蚯蚓的氧化激活程度,通过活性氧自由基(ROS)的含量反映DEHP对蚯蚓的细胞毒性,通过丙二醛(MDA)含量和Olive尾矩反映DEHP对蚯蚓的遗传毒性,从多个角度评价DEHP对蚯蚓的生态毒理效应。结果表明:(1)在DEHP的刺激下,各浓度组蚯蚓的SOD、CAT、POD、GST活性均呈激活状态,表明DEHP对蚯蚓的抗氧化酶有诱导作用;(2)DEHP影响蚯蚓的ROS含量,各染毒处理组与对照组相比均升高且差异性明显,表现出明显的剂量—效应关系;(3)对比各染毒处理组之间的数据,DEHP对蚯蚓MDA含量的影响无明显规律;(4)DEHP可使蚯蚓GST呈激活状态,表明中高浓度的DEHP对蚯蚓GST具有诱导作用;(5)DEHP能够引起蚯蚓体腔细胞DNA的损伤,且随着浓度的增加,Olive尾矩值随之增加,说明DNA损伤程度与DEHP浓度之间具有剂量-效应关系。从实验结果可以看出,DEHP可以对蚯蚓机体和DNA造成一定程度的损伤,表现出较强的生态毒理效应。

[Objective] Phthalate acid esters （ PAEs ） , also known as phthalate esters or ＂plasticizer＂ , are the reaction product of phthalic anhydride and alcohol. PAEs are widely used in the production of a huge number of products such as agricultural plastic film, toys, medical bags and hoses for transfusion, food packaging, cleansers, vinyl floor and wallpaper, and personal care products. Among them, Di （2-ethylhexyl） phthalate （DEHP） is one PAE, the highest in output and consumption and also a toxic organic compound highly hazardous to human health, disturbing endocrine systems and causing cancers, mutation, oxidative damage and genetic toxicity. As DEHP is weakly bonded to the plastic matrix by hydrogen bond and van der Waals force, it is easily detached and diffused into the air, water and soil environments. Therefore, its pollution and hazard to human health has aroused more and more attention from the people. So far studies have proven that DEHP can enter organisms via a variety of ways, causing poisonous damage to tissues and organs of the organisms. As DEHP is extensively used in agriculture and hence exists in large volumes in the soil system, it is necessary to investigate its ecological and toxicological effects, such as oxidative stress and DNA damage, on typical soil animals, in the hope that the study may serve as scientific basis for reasonable assessment of soil ecological risk of DEHP. [ Method ] To explore eco-toxicological effects of DEHP on soil animals, Earthworms （Eisenia foetida ） , used as bio-indicator, were exposed to DEHP varying in concentration （ 0, 0.1, 1, 10 and 50 mg kg^-1 ） in artificial soil. Earthworms were sampled after 7, 14, 21 and 28 days of exposure for analysis of superoxide dismutase （ SOD ） , catalase （ CAT ） , peroxidase （ POD ） and glutathione （ GST ） , which are the indices reflecting the degree of DEHP activating oxidation of earthworms, reactive oxygen species （ ROS ） , which is the indicator of cytotoxicity of DEHP to earthworms, and malondiadehyde （ MDA ） and olive tail moment, which are the indices reflecting genotoxicity of the substance to earthworms. [Resuh] （ 1 ） DEHP, above 0.1 mg kg-1 in concentration, significantly enhanced the activities of SOD, CAT, POD and GST in Eisenia foetida by a degree varying with the concentration of DEHP, which demonstrates that DEHP induces antioxidant enzymes in earthworms; （ 2 ） DEHP increased ROS concentration in earthworms as compared with control and the effect varied between treatments, displaying an apparent dose-effect relationship; （ 3 ） Comparison between the treatments shows that DEHP did have some effect on content of MDA, but the effect did not follow any specific rule and was the most obvious in earthworms exposed to DEHP, 10 mg kg-1 and 50 mg kg-1 in concentration; （ 4 ） In the presence of DEHP. GST in the earthworms was apparently activated, except for the treatment of 0.1 mg kg-1, where the earthworms did not have much difference from those in control. The observation demonstrates that DEHP, higher than 0.1 mg kg-1 in concentration induces GST in the earthworms; （ 5 ） Single cell gel electrophoresis shows that coelomocyte DNA of the earthworms was damaged in the presence of DEHP, regardless of concentration and Olive tail moment increased with the rising DEHP concentration, which suggests that DNA damage caused by DEHP is dose-dependent. [Conclusion] Once DEHP enters earthworms, it activates the defense system of antioxidant enzymes （ SOD, CAT and POD ） and GST detoxification enzyme in the earthworms to protect the organisms from injury. Nevertheless, DEHP still raises ROS content in the earthworms, and excessive ROS triggers peroxidation of lipids, producing MDA. ROS and MDA act jointly damaging cell DNA of the earthworms. To sum up, DEHP may cause a certain degree of damage to organisms and DNA of the earthworms, manifesting fairly strong eco-toxicological effects. Therefore, adequate attention should be paid to DEHP disposal.