地下水中阿特拉津污染的原位生物修复研究/

2019-05-11 07:20:05

The 降解 阿特拉 地下水 atrazine



阿特拉津,也称莠去津,是在世界范围内广泛使用的一种三嗪类除草剂,很多研究认为它是一种环境雌激素,它会对人和野生动物产生激素类的作用,或阻碍生殖功能,或引发癌症等。低剂量(0.1μg/L)的阿特拉津暴露也会引起蛙类体内雌性荷尔蒙激素分泌量增多,使其内分泌系统遭到破坏,致使雄性蛙类雌性化或者导致雌雄同体。由于其生产和连年施用,造成了土壤及地下水的大面积污染。地下水的污染具有复杂性、隐蔽性和难以恢复的特点,一旦土壤或地下水遭受了污染,那么,自然恢复和净化的过程是漫长的,因此有必要寻求一种经济有效的地下水原位修复方法。
开展对阿特拉津污染治理方法的研究,不仅可以消除环境污染,确保我国环境安全,促进我国社会经济的可持续发展,而且以提高我国农产品在国际贸易中的竞争力。
因此,本文以地下水中阿特拉津为治理目标物,研究阿特拉津污染地下水的有效处理方法。
发达国家早在20世纪70年代就开展了对污染地下水的修复研究工作,提出地下水的污染治理方法有:隔离法、泵提法、吸附法、化学栅栏法、电化学法和生物修复方法等,其中生物修复方法以其具有的投资低、效益好、应用简便等特点,被逐渐应用于地下水有机污染的治理中,现已成为一项清洁环境并有很大发展潜力的新兴技术。对于点源污染的治理,90年代加拿大化铁炉大学Gillim教授提出了可渗透反应墙(PRB)技术,由于其具有投资少,地面扰动小,基本无能耗,以及便于和其他处理技术结合应用等优点,近十年来得到迅速发展。在这项技术中主要是将零价铁作为反应介质来处理一些卤代有机物和一些重金属类无机物,同时也有人将一些吸附型材料(活性炭、沸石等),制成可渗透反应墙体进行地下水的原位修复,在这些研究中,利用固定化微生物作为反应介质并制成生物可渗透反应墙体的研究很少见。对于面源的污染的修复目前多采用自然修复的方法,靠地下水中的土著微生物降解和地下水的稀释作用来降低污染物的浓度。如果在地下水环境中不存在能够降解阿特拉津的微生物或者土著微生物只能部分降解阿特拉津。就要人为的注入人工筛选的优势菌群来进行地下水的修复。
本文主要研究了阿特拉津在包气带及地下水中的迁移转化规律,筛选出在低温条件下能快速降解阿特拉津的菌株,并对其进行降解特性及降解机理的研究。对于阿特拉津污染的下水的生物修复主要采用注入优势菌群进行原位生物修复,同时吸收国外原位生物修复及PRB等方法的成功经验,设计生物活性反应墙,进行地下水原位修复方法的研究。本论文的研究成果对我国阿特拉津污染地下水的修复具有重要的理论意义和应用价值。
本论文的主要研究成果主要分为以下三个方面:
1.本论文开展了阿特拉津在包气带和含水层中的迁移转化研究,可得出如下结论:
(1)农药在土壤中向下迁移进入地下水这一过程受许多复杂因素的制约。静态吸附实验表明,三种岩性土壤对阿特拉津的吸附量都不大,其快速吸附基本在24小时内完成;AT的吸附量随着离子强度的增加而增加;随着pH值的增加,AT在土壤中的吸附量减少;总有机碳(TOC)与AT吸附量呈正相关关系,且对吸附量影响较大。
阿特拉津在土壤和地下水中的自然降解情况表明,阿特拉津的半衰期较长,会长期存留在地下水和包气带中。
(2)实验室土柱实验结果表明:
三种岩性土壤均有阿特拉津的淋出,而且土壤粒径越大,越有利于阿特拉津的淋出,污染地下水的可能性就越强,这与静态实验结果一致;土壤有机质含量小,农药的淋溶性能增强;降雨量大而强度相对较小,或降雨距施药时间短,均有利于农药的淋溶;间歇降雨比连续降雨有利于阿特拉津的淋出。
(3)阿特拉津在地下水中的迁移模拟表明,阿特拉津在地下水中的迁移主要以侧向和垂向为主,符合二维均质对流弥散模型。由于各种物理、化学及生物作用,阿特拉津的迁移与其周围地下水运动产生明显差异,经计算可知阿特拉津在含水层中的阻滞因子为2.45,迁移速度为4.08cm/d。
2.本实验采用富集驯化的方式,从吉林市农药厂排污口、长春市裴家垃圾场渗滤液及吉林市污水处理厂的活性污泥中筛选出四株在低温条件下能够快速降解阿特拉津的菌株。通过生理生化和16S rRNA同源性分析,可确定菌株L1在分类学上属于真核界(Eukaryota);真菌门(Fungi);担子菌亚门(Basidiomycota);锈菌纲(Urediniomycetes);红酵母属(Rhodotorula),粘质红酵母种(mucilaginosa)。
L2与Leifsonin naganoensis的亲缘关系较近,共同组成一个分支,两者的16SrRNA序列的相似性为99%,可确定菌株L1在分类学上属于细菌界(Bacteria);放线菌门(Actinobacteria);放线菌纲(Actinobacteridae);放线菌目(Actinomycetales);微杆菌科(Micrococcineae);微杆菌属(Microbacteriaceae);Leifsonia种。
N8与Stenotrophomonas maltophilia的亲缘关系较近,共同组成一个分支,两者的16SrRNA序列的相似性为99%,可确定菌株N8在分类学上属于细菌界(Bacteria);变形菌门(Proteobacteria)γ-变形菌纲(Gammaproteobacteria),黄色单胞菌目(Xanthomonadales),萨顿氏菌属(Stenotrophomonas ),嗜麦芽糖寡养单胞菌(maltophilia)。
W2虽然与假单胞菌属中的恶臭假单胞菌种的序列同源性为99%,而W2与假单胞菌属中恶臭假单胞菌种的生理生化特性存在差异,因此我们认为W2可能为细菌界(Bacteria),变形菌门(Proteobacteria),γ-变形菌纲(Gammaproteobacteria),假单孢杆菌目(Pseudomonadales),假单孢杆菌科(Pseudomonadales),假单胞菌属(Pseudomonas)中的一个新种。上述四种菌在国内外文献中未见报道。
通过降解实验,优化了四种单菌和混菌的最佳降解条件;在低温条件下四种单菌对不同浓度阿特拉津的降解半衰期分别为L1:4.4天、L2:4.2天、W2:1.5天、N8:8.4天,优于已报道低温菌的降解半衰期。阿特拉津的初始浓度对混菌降解的效果无影响,48小时混菌对阿特拉津的降解率均达到96.5%以上,说明混菌对阿特拉津的降解速率高于单菌。
降解机理实验研究表明W2中存在AtzA、AtzB、AtzC和AtzD四条降解基因与阿特拉津降解模式菌株Pseudomonas sp ADP几乎完全一致,表明菌株W2的降解途径与Pseudomonas sp ADP是一致的。在菌株N8中只找到AtzA降解基因,说明菌株N8不能将阿特拉津完全矿化,只能将阿特拉津水解为2-羟基阿特拉津,而在L1和L2中未找到这四种降解基因,说明L1和L2对阿特拉津的降解不遵循氯水解途径。通过GC/MS定性分析,混菌能催化阿特拉津发生脱氯水解,产生对生物无毒的羟基产物,同时能催化阿特拉津发生脱烷基反应,产生代谢产物为2-氯-4-胺基-6-乙胺基-1,3,5-三嗪、2-氯-4-胺基-6-异丙胺基-1,3,5-三嗪和2-羟基-4-乙胺基-6-异丙胺基-1,3,5-三嗪。进一步证明W2和N8遵循的是氯水解途径,而L1和L2遵循脱烷基的降解途径。经过氯离子和氨根离子的定量测定可知混菌能够完全矿化阿特拉津产生CO2和NH4+。
3.在实验室内利用渗流槽模拟地下水的污染,采用注入优势菌种的方法进行阿特拉津污染的修复,结果表明将混合菌按等比例混合注入注菌井,各采样口阿特拉津的浓度迅速降低对于28#、29#、35#、41#采样口阿特拉津浓度在五天之内降解率均达到90%以上,试验运行21天后,各取样口阿特拉津的降解率均达到92%以上。增大地下水的流速各采样口阿特拉津的浓度变化不大。。同时在实验过程中利用生物反应墙修复阿特拉津污染的地下水,将固定微生物的活性炭作为反应墙的装填介质,制成生物反应柱,同时与单纯的活性炭柱作对照,结果表明生物柱可有效的降解阿特拉津,在运行60天的时间内降解率均达到99%以上,而单纯的活性炭柱在运行40天左右大部分吸附位点被占据,淋滤液中阿特拉津的浓度逐渐增加,说明采用注入优势菌种及模拟生物反应墙这两种方法可有效的修复阿特拉津污染的地下水。



Atranex, also called atrazine, is a kind of triazine herbicides which is widely used in the world. It has been considered in many reports as a kind of environmental estrogen, and it would work as hormones to human beings and wildlife, retarding their reproduction, or causing cancer etc. Exposuring to low dose atrazine(0.1μg/L)will lead to the increase of female hormone secretory volume in the body of frogs, destroying its endocrine system, bringing about the feminization of male frog or hermaphroditism. Since the atrazine has been used year by year, the soil and groundwater is polluted in a large sale. Owing to the complexity, invisibility and difficulty in restoring of the groundwater pollution, once polluted, its natural recovery and purification process will take a long time. Therefore, it is necessary to find an economical and efficient in situ remediation method for it.
To study the remediation method for atrazine will not only eliminate the pollution, insure the national environmental safety, promote the sustainable development of national social economy , but also improve the competitive power of national agricultural products in the international trade.
Therefore, atrazine in the groundwater is taken as the target substance,and its economic and efficient remediation method is studied in this paper.
The study on the remediation of groundwater pollution in developed countries began from 1970’s, the treatment methods included isolation, pump lifting, adsorption, chemistry fencing, electrochemical process and bioremediation,etc. Among them, the bioremediation method had been used in the remediation of groundwater pollution and been a clean and potential emerging technology for its low investment, better efficiency and convenience in use.In 1990’s, the Canadian professor Gillim put forward PRB technique to the remediation of point source pollution, and it has been developed rapidly in the latest decade owing to its low investment, light ground disturbance,almost no energy requirement,etc. The technique mainly take zero valent iron as its reactive media, to treat certain kind of halogenating organic matter and some heavy metals. And meanwhile add some adsorption materials such as absorbent carbon and zeolite etc, to make the PRB for the insitu remediation of groundwater pollution. All among these studies, it is seldom the case that taking microorganisms as the reactive media to make the PRB.For the non-point source pollution, the natural remediation was used as a main method, it used indigene microorganisms to degrade pollutants and also used diluting effect. If there is no microorganisms which have the ability to degrade atrazine or atrazine only can be degraded partly, the dominance strains must be screened and injected to the groundwater to degrade it.
The achievement of the text included following three parts:
1.The text studied on the regulation of atrazine's transformation in unsaturated zone and groundwater,and its conclusions are followings
(1)The process of pesticides transfering downward the soil into the groundwater were constrained by many complex factors. The static adsorption experiment showed that the three kinds of lithology soil didn’t absorb much atrazine, and the adsorption of atrazine completed in 24 hours.The fewer the organic matter and ionic strength in the soil, the lower the absorption of atrazine is.whereas adsorptive capacity decreased when the pH increased.
Natural degradation of atrazine in the soil and groundwater showed that atrazine had a long half-life and long-term presence in groundwater and unsaturated zone.
(2)The results of the soil column experiments showed:
For three types of lithologies soil the atrazine could be strained out.The fewer the organic matter in the soil, the lower the absorption of atrazine was, responsively,the leaching ability of the pesticides would increase.When the amount of rainfall was great and its intensity was low, or the time to apply the pesticides was short, were all in favor of pesticides leaching. Intermittent rainfall was more advantageous than continuous rainfall for the atrazine leaching.
(3)The transfer simulation of atrazine in the groundwater showed that it mainly transferred in the crossrange direction and vertical direction, correspond to that of two dimension contraflow dispersion model. Owing to different kinds of physical, chemical and biological function,atrazine and groundwater ' s migration velocity was different.The atrazine 's retardation efficient was 2.45 ,migration velocity was 4.08 cm/d.
2.After enrichment culture and domestication, four strains were screened for their high degradable abilities, and they were from pollution discharge port of pesticide factory in Jilin, leachate of Peijia landfill of Changchun, and activated sludge of sewage disposal plant in Jilin. L1 was identified as Eukaryota Fungi Basidiomycota Urediniomycetes Rhodotorula mucilaginosa.L2was identified as Bacteria Actinobacteria Actinobacteridae Actinomycetales Micrococcineae Microbacteriaceae Leifsonia.N8 was identified as Bacteria Proteobacteria Gammaproteobacteria Xanthomonadales Stenotrophomonas maltophilia.The four strains which could degrade atrazine were not reported at the home and abroad.W2 may be a new species which belonged to the Bacteria Proteobacteria γ-Gammaproteobacteria Pseudomonadales Pseudomonadales Pseudomonas.
Through the degradation experiment,the optimal degradation conditions of odd and mixed strains were got. In cold condition, atrazine’s degradation half-life of L1 is 4.4 day , L2 is 4.2day , W2 is 1.5day , N8 is 8.4day,these degradation half-life are shorter than other atrazine degradation strains which were reported.The starting concentration had no effect on the degradation of mixed bacteria,and the mixed strains degradation rate reached 96.5% in 48h.
Using PCR, the atrazine-degradation genes AtzABCD were identified in W2,AtzA was identified in N8.the AtzABCD genes revealed significant homology (>99%) with the atrazine degradation genes of Pseudomonas sp. strain ADP.This result indicated atrazine degradation started with a hydrolytic dechlorination by W2 and N8.There were no atrazine-degradation genes in strain L1 and L2.Atrazine can be hydrolyzed and dehalogenated by mixed strains.The biodegradation products of hydroxyatrazine ,deisopropylatrazine and deethylatrazine were comfirmed by GC/MS After quantitative determination of chloride ion and ammonia ion,it could be known that the atrazine were mineralized to CO2 and NH4+.
3.The seepage tank was used in the lab to simulate the pollution of groundwater and the bioargumentation method was adopted . The results showed that when the mixed strains at equal ration were injected into the pit,the atrazine’s concentration of each sampling port was decreased rapidly, and for 28#、29#、35#、41#, the degradation rate reached higher than 90%, after running for 21days, 28# reached 92.3%, 9# reached 94.9%, 35# reached 98.5% and 41# 99.79%,the atrazine concentration was 1μg/L,which reached national drinking water hygienic criterion. The atrazine degradation rate in the outlet reached 92.4%. With the increase of the flowrate, the concentration in the sample outlet didn’t changed so much, which proved that to inject dominance microorganisms is an effective method for the remediation of groundwater pollution caused by no point source pollution.
In the experiment , the bio-reactor was used for the rehabilitation of the groundwater pollution caused by atrzine, and activated carbon was adopted as immobilization carrier, and its optimal particle diameter is 0.5-1mm, pH is 8.The activated carbon immobilized with microorganisms were filled in the reactor, another filled with activated carbon for comparison, the result showed that the degradation rate reached higher than 99% in the 60 running days, however, the effluent concentration of the reactor filled with activated carbon increased day by day when the absorption points were taken up.