【作者】 赵良元；

【导师】 杨劭；

【作者基本信息】 华中师范大学 ， 植物学， 2012， 博士

【摘要】 十溴联苯醚(Decabromodiphenyl Ether, BDE-209)由于优异的阻燃效能而最为全球性普遍使用的的溴剂阻燃剂。近年来,BDE-209被发现在环境中普遍存在,甚至南极的沉积物和企鹅中都可检测到其存在,由于其持久性、易于生物富集并且具有内分泌干扰作用而。由于BDE-209的高疏水性,水环境沉积物是BDE-209的主要蓄积库,中国是世界上沉积物BDE-209检出率与检出值最高的国家之一。目前,对于环境中的BDE-209污染的修复方法主要有光降解、高温和微生物降解,但这些方法大都不适用于水环境中BDE-209污染的修复。绿色、经济、可持续性的生物修复技术-植物修复是解决沉积物有机污染物最有前景的修复技术,由于在水环境沉积物污染物修复中微生物修复技术难以有效应用,因此植物修复技术具有更好的应用前景,但是目前关于沉积物PBDEs污染的植物修复技术研究尚未有报道。本研究前期通过筛选实验发现水生植物水葱(Scirpus validus Vahl)对沉积物中的BDE-209具有良好的修复效果。因此,本研究进一步开展水葱来对沉积物中的BDE-209的抗性及修复机制研究,包括水葱对BDE-209的抗性、生物富集能力及修复效率,增效方法、BDE-209代谢变化、根际微生物群落及降解菌分析等,本研究结果对水环境多溴联苯醚污染的生物修复具有参考价值。通过比较我国常见的四种挺水植物对BDE-209的修复效果,选择水葱(Scirpus validus Vahl)用于进一步研究。为了解水葱对BDE-209的抗性,研究了水葱对沉积物中BDE-209的生理学响应。结果表明沉积物中的BDE-209在2-30 mg·kg-1dw内不会对水葱产生不利影响(p>0.05),但其作为一种OPs在高浓度时(30mg·kg-1)在修复初期对水葱产生了胁迫,表现在根部丙二醛(MDA)增加,叶绿素含量、叶绿素荧光、根系活力有所降低,60天内植物体内抗氧化酶系统超氧化物歧化酶(SOD)、过氧化物酶(POD)及过氧化氢酶(CAT)活性增强,但BDE-209的早期胁迫并没有对水葱的生长造成明显影响,其生物量无显著差异(p>0.05)。以上结果表明水葱能在BDE-209高达30 mg·kg-1的底泥中正常生长,对该污染物有很强生理学抗性。研究了水葱对三种典型沉积物(淤泥、黄泥、砂质沉积物)中BDE-209的修复效率及吸收富集能力。经过18个月修复实验,淤泥修复组、黄泥修复组、砂质沉积物修复组中BDE-209分别减少了92.84%、84.04%及72.22%,去除率比各自对照组分别提高了148%、197%、233%。结果表明水葱可有效修复三种典型沉积物中BDE-209污染。水葱不仅可以通过根部从沉积物中吸收富集BDE-209,而且还具有向地上部分传导的能力。但是水葱组织对沉积物中BDE-209的吸收富集过程是一个被动过程,与沉积物中BDE-209的浓度紧密相关。由于BDE-209的高疏水性,水葱对BDE-209的BCFs与TFs都相对较小,表明其对BDE-209的吸收富集传导作用有限。BDE-209主要富集在水葱根部,只有少量传输到地上部分。水生植物修复沉积物BDE-209污染过程中,光解及其他非生物因素对沉积物中BDE-209去除贡献比例较小,分别占8.44%及8.23%,水葱根部及地上部分对沉积物中BDE-209吸收富集对BDE-209的去除贡献作用有限,仅仅占0.5%。沉积物土著微生物及水葱根际微生物才是植物修复BDE-209的主要成员,对照组中土著微生物的降解占83.35%,实验组中水葱根际微生物的作用占94.66%。采用GC/MS对BDE-209在根际沉积物、水葱根部及地上部分的降解产物进行了研究,在根际沉积物中发现了16种PBDEs同系物,主要为Nona-BDEs、Octa-BDEs、Hepta-BDEs、Hex-BDEs、Penta-BDEs、Tetra-BDEs及Tri-BDEs7种溴代PBDEs,水葱根部组织中检测出16种BDE-209代谢产物,主要为Nona-BDEs、Hex-BDEs、Penta-BDEs、Tetra-BDEs、Tri-BDEs及Di-BDEs 6种溴代PBDEs,地上部分发现7种BDE-209代谢产物,主要Hepta-BDEs、Penta-BDEs、Tetra-BDEs及Di-BDEs 4种溴代PBDEs,以上结果表明BDE-209在沉积物-水生植物系统中可通过脱溴作用代谢成低溴代PBDEs.为提高水葱对沉积物中BDE-209污染的修复效率,分析了表面活性剂及β-环糊精强化植物修复的效果以及水葱对增效剂的耐受能力。结果表明沉积物中浓度范围为300-1000 mg-kg-1的四种增效剂CATB、SDS、Tween80、β-CD在整个修复过程中对水葱的生长(株高及茎干直径)没有明显影响,说明水葱对这四种高浓度增效剂具有较强抗性。沉积物中加入600-1000 mg-kg-1 CATB、SDS、Tween80都可有效提高水葱修复沉积物中BDE-209的效率；180天对照组BDE-209去除了34.93%,300-1000 mg-kg-1 CTAB对BDE-209修复效率提高了2.85%-11.78%; 300-1000 mg-kg-1 SDS的对BDE-209的修复效率提高了4.52%-14.25%; 300-1000mg-kg-1 Tween80对BDE-209的修复效率提高了7.37%-19.33%; p-CD在300-1000 mg-kg-1对BDE-209的植物修复效率只增加了1.03%～7.78%,其对沉积物中BDE-209的去除没有明显促进作用。以上结果表明三种表面活性剂都可以作为沉积物BDE-209水葱修复的增效剂,其中Tween80效率较好。为揭示水葱修复沉积物中BDE-209过程中根际微生物的作用,采用DAPI荧光染色计数、平板计数法、及沉积物脱氢酶与脲酶活性分析了根际及非根际沉积物中微生物(细菌、真菌、放线菌)数量和活性。结果表明水葱的种植可显著提高沉积物中微生物的数量,三种沉积物根际中细菌数量较非根际增加了2-12倍,根际效应明显,根际沉积物真菌数量较非根际增加了1-4倍,水葱的种植对沉积物中放线菌的影响较小,淤泥和黄泥根际沉积物中放线菌数量较非根际区别不明显,砂质根际沉积物中放线菌数量明显高于非根际。同时,由于微生物数量的增加,三种沉积物根际微生物酶活力(脱氢酶及脲酶)在修复过程中都显著大于非根际,说明水葱的种植可以提高沉积物中微生物的活力。沉积物酶活表现出淤泥>黄泥>砂质沉积物的趋势。以上结果显示微生物数量及沉积物酶活的差别是造成水葱对三种典型沉积物中BDE-209修复效率不同的主要原因之一。为了解水葱根际沉积物中细菌群落特征并辨别主要降解菌,采用限制性内切酶片段长度多态性(restriction fragment length polymorphism, RFLP)分析了水葱根际沉积物中细菌多样性及差异。结果表明,水葱根际沉积物中细菌多样性丰富,最多包含分布于12个已知门类的细菌和一些未被认知的序列：a-变形菌(Alphaproteobacteria)、β-变形菌(Betaproteobacteria)、δ-变形菌(Deltaproteobacteria)、γ-变形菌(Gammaproteobacteria)、浮霉菌门(Planctomycetes)和酸杆菌门(Acidobacteria)、优杆菌属(eubacterium)、拟杆菌门(Bacteroidetes)、黄杆菌属(Flavobacterium)、绿弯菌门(Chloroflexi)及未能归类的OTUs。BDE-209污染根际沉积物及未污染根际沉积物中优势菌群为β-变形菌(β-proteobacteria)、δ-变形菌(δ-Proteobacteria)、α-变形菌(α-proteobacteria)及酸杆菌(Acidobacteria)及绿弯菌(Chloroflexi),但是两种沉积物优势菌数量及细菌群落结构上有明显差异。本研究表明根际沉积物中优势菌β-Proteobacteria及α-Proteobacteria有可能是降解BDE-209的主要菌群。淡水河河道水质持续净化技术集成示范工程由沉淀塘、曝气、人工湿地和水生植物塘组成,目的是研究集成技术对河道有毒污染物和常规污染物的处理效果。由于植物修复是该工程的主要工艺环节,本文对该示范工程运行中有毒有机污染物的处理效果进行了监测分析。多次监测结果表明,示范工程对多环芳烃(polycyclic aromatic hydrocarbons, PAHs)处理效率达40%以上,其中5种同系物的去除率：萘(NAP) 44.13%-59.23%,芴(FLU) 44.1%-54.61%,菲(PHE)50.90%～54.70%,芘(PYR)61.88%～80.61%,荧葸(FLT)44.83%-56.64%。同时示范工程对淡水河中的十溴联苯醚(BDE-209)以及内分泌干扰物双酚A (bisphenol A, BPA)的处理效果均达到100%。结果表明淡水河河道水质持续净化技术集成示范工程的工艺对河水中有毒有机污染物具有良好的净化作用,具有推广价值。更多还原

【Abstract】 Decabromodiphenyl Ether (BDE-209) is the domainant polybrominated diphenylethers (PBDEs) that have been used extensively as flame retardants in a wide range of products such as plastics, textiles, and electric circuitry owing to their excellent Flame retardant efficiency and thermal stability. At present, BDE-209 was the ubiquitously detected contaminant in the environment and even its extence was found in the sediments and penguins of the Antarctic. Owing to high volume production, lipophilicity, bioaccumulation, persistence and endocrine disruptor of BDE-209, deca-BDE has become the most ubiquitously detected contaminants of major concern in the environment. Sediments are regarded as ultimate sink of decabromodiphenyl ether (BDE-209) in aquatic environment due to the high hydrophobicity and china is one of the countries where BDE-209 is the mostly detected regarding the concentrations and detection rates.The methods applied for remediation of PBDEs contaminated environment such as photodegradation, nanoparticles degradation and microbial degradation are expensive, disruptive to the environment and also involve high energy consumption, thus those were not suitable for remediation BDE-209 contaminated sediment in aquatic environment. The emerging technology of phytoremediation was a promising remediation method for organic contaminants cleanup thanks to its ecological and economic sustainability.However, only few phytoremediation reports has been applied for PBDEs remediation and the potential for any aquatic macrophytes for phytoremediaiton of decabromodiphenyl ether contaminated sediments has not been explored. The aquatic macrophyte, S. validus Vahl, was selected among the common emerging macrophytes by the high efficiency of remediation of BDE-209.In this research the phytoremediation of BDE-209 by S. validus Vahl including the BDE-209 resistance, remediation efficiency, bioaccumulation capability, remediation mechanisms, BDE-209 metabolic changes and microbial ecological changes was investigated at an environmentally relevant concentration in three typical sediments.In this study, S. validus Vahl seedlings were grown in BDE-209 contaminated sediment for determining the eco-physiological responses of S. validus Vahl to BDE-209 phytotoxicity at the increasing concentrations of BDE-209 (0,2,8,30 mg-kg-1). The results indicated that BDE-209 at the BDE-209 concentrations ranged from 2 to 30 mg-kg-1 did not inhibit the growth of S. validus Vahl. Stress effect was observed on plant height, stem diameter, chlorophyll content, MDA contents and root activity of S. validus Vahl at high contaminated levels in the initial stages. SOD, POD and CAT which constitute the antioxidant enzyme system were increased in 60 days. The above results clearly demonstrated growth and physiological function of S. validus Vahl was not adversely impacted by BDE-209 contaminated sediment and thus could be used in phytoremediation of BDE-209 contaminated sediment.Phytoremediation of BDE-209 by aquatic macrophyte, S. validus Vahl was investigated at an environmentally relevant concentration in three typical sediments (silt, clay and sand sediment).At the end of the experiment (18 months), the dissipation rates of BDE-209 in three typical sediments were significantly enhanced by S. validus Vahl compared with those in control sediments (p<0.01), and the average removal rates of BDE-209 were 92.84%(silt sediment),84.04%(clay sediment) and 72.22%%(sandy sediment), respectively, which were 148%(p<0.01),197%(p<0.01) and 233% (p<0.01) higher than those of the control sediments.The ability of S. validus Vahl to uptake and translocate BDE-209 was confirmed by the detection of BDE-209 accumulated at the root tissue with concentrations of 0-300μg kg-1dw and translocation to shoot tissue (0-64μg kg-1dw). BDE-209 uptake and translocation in tissue of S. validus Vahl may be influenced by residual BDE-209 concentration in sediments indicated that BDE-209 uptake was a passive process. The BDE-209 BCFs and RCFs of S. validus Vahl was comparatively lower because of high hydrophobicity. The uptake suggested that BDE-209 sorb strongly to the roots of S. validus Vahl and the translocation was limited.In the process of BDE-209 phytoremediation by S. validus Vahl, photolysis and other abotic losses were miner possible pathways of BDE-209 dissipation which accounted for 8.44%and 8.23%, respectively. The accumulation of BDE-209 in plant biomass only made a small contribution (0.5%) to the total removal of BDE-209. Through analysis of pathways of BDE-209 removal, this enhanced dissipation of BDE-209 by macrophyte-microbal association might be mainly the result of S. validus Vahl-promoted microbial degradation (83.35%in control sediments,94.66%in Treatment). Metabolites of BDE-209 in the sediment-macrophyte System were measured by GC/MS, A total of sixteen additional lower brominated PBDE congeners (di-to nona-) were detected in the soil and plant tissues after plant harvest, confirming metabolic debromination of BDE-209 in this system.Evidence of a relatiely higher proportion of penta-through di-BDE congeners in plant tissues than in the soil indicates that there is further debromination of PBDEs within plants or low brominated PBDEs are more readily taken up by plants.To increase the bioavailability of BDE-209 and facilitate its degradation in phytoremediation, plant-accelerated dissipation of BDE-209 in rhizosphere sediments of S. validus Vahl in the presence of a cation ionic-surfactant (CTAB), an anionic-surfactant (SDS), a nonionic-surfactant (Tween 80) and a Cyclodextrin (P-CD) at the concentrations ranged from 300 to 1000 mg·kg-1 were studied. Siginificantly negative changes were not observed for the biomass in terms of plant height and stem diatemeter of S. validus Vahl when the addition of 300 to 1000 mg-kg-1 CTAB, SDS, Tween 80 and P-CD. This indicated that it is preferable for CTAB, SDS, Tween 80 andβ-CD to be utilized as the BDE-209 phytoremediation amendment. Furthormore, the addition of CTAB, SDS, Tween 80 with initial concentrations of 300 to 1000 mg-kg-1 in rhizosphere sediments significantly enhanced the phytoremediation effiencies.As for the control,34.93%of BDE-209 was dissipated from the sediment. 2.85%-11.78%of increase in BDE-209 removal rates was obtained in the addition group of SDS. 4.52%-14.25%of increase in BDE-209 removal rates was obtained in the addition group of Tween 80.7.37%-19.33%of increase in BDE-209 removal rates was obtained in the addition group of Tween 80. Though the removal efficiency of BDE-209 was increased in the preasence ofβ-CD and 1.03%-7.78%of increase in BDE-209 removal rates was obtained, no significantly enhance of BDE-209 phytoremediation was not observed in the addedβ-CD concentration ranges. Results obtained from this study provided some insight with regard to the feasibility of phytoremediation for BDE-209 contaminated sediments with addition of suitable surfactants, especially Tween 80.During the process of phytoremediation, phytoremediation, the number of microorganisms including bacteria, fungi and actinomycetes counted by DAPI (4,6-diamidino-2-phenylidole) fluorescence direct count and plate count method in three typical rhizosphere and sediments were examinated. The results showed that the number of microorganisms in sediments was significantly increased by planting and the number of bacteria and fungi increased by 2-12 times and 1-4 times in the respective rhizosphere sediment. No obvious difference of quantity of actinomycetes was observed between rhizosphere and non-rhizosphere in silt and clay sediments expect for sand sediments. As the number of microorganisms increased, planting of S. validus Vahl enhanced the dehydrogenase activities and urease in three typical sediments. The increase of sediment dehydrogenase activities and urease in the rhizosphere signified the activities of microorganisms were enhanced and both were showed a trend of silt>clay>sand sediments.The results suggested that the numbers and activities of microorganisms is an important reason to explain why there existed the differences of BDE-209 dissipation rates in three typical sediments by the presence of S. validus Vahl.Diversity of bacteria was studied from rhizosphere BDE-209 contaminated and noncontaminated sediments of S. validus Vahl by PCR, RFLP and sequence analysis o f 16S rDNA and comparing with the published sequences in GenBank database. Based on the restriction fragment length polymorphism(RFLP) profile generated 200 clones from the 16 s rDNA library, Phylogenetic results indicated that 200 clones could be divided into 12 phylotypes, includingβ-Proteobacteria, 8-Proteobacteria, a-Proteobacteria, Acidobacteria and Chloroflexi, Planctomycetes, eubacterium, Flavobacterium and some unidentified OTUs.The predominant bacteria in contaminated and uncontaminated rhizosphere sediments wasβ-proteobacteria,δ-proteobacteria,α-proteobacteria, Acidobacteria and Chloroflexi, Bacterial community structure of BDE-209 contaminated rhizosphere sediment has distinguished feature and obviously different from the unpolluted rhizosphere sediment sample, which is mainly reflected in the dominant position ofβ-proteobacteria,δ-proteobacteria,α-proteobacteria, Acidobacteria and Chloroflexi in the bacterial flora, andβ-proteobacteria andα-proteobacteria may play important role in the BDE-209 degradation in the rhizosphere sediment of S. validus Vahl.Demonstration project of water quality for sustainable purification in Danshui river was constituted by four parts, precipitation pond, aeration pond, artificial wetland and aquatic plants pond with the aim to evaluate the removal efficiency of toxic pollutants and conventional pollutants by the integration technology. Monitoring Analysis of toxic pollutants removal and Conventional pollutants by the Demonstration project was made and preferable results were obtained in this research. More than 40%of polycyclic aromatic hydrocarbons (PAHs) were removed from water by demonstration project, and the removal rates of five kinds of homologues were, NAP,44.13%-59.23%; FLU,44.1%54.61%; PHE,50.90%-54.70%; PYR,61.88%-80.61%and FLT,44.83%-56.64%. Decabromodiphenyl ether(BDE-209) and bisphenol A(BPA) could be fully removed by this project.The results suggested that demonstration project in Danshui river could effectively remove the toxic organic pollutants from water, demonstrating its popularization value for purification of toxic organic pollutants.更多还原