【作者】 彭克俭；

【导师】 沈振国；

【作者基本信息】 南京农业大学 ， 植物学， 2006， 博士

【摘要】 植物修复（phytoremediation）是指利用植物清除土壤和水体中的污染物或降低污染物的毒性，使被污染环境得到恢复的一种技术。植物采矿（phytomining）是指利用超量积累植物从富含金属的土壤中提取金属并通过焚烧获得“生物矿砂”进一步提炼金属的技术。超量积累植物是指能超量吸收并积累重金属的植物。超量积累植物是植物修复和植物采矿的基础。本研究的主要目的是筛选出具有重金属耐性或超积累特性的植物，并研究其耐性或超积累机理，为重金属污染土壤和水体的植物修复提供理想的植物材料和可靠的理论依据。为了全面了解湖南湘西矿区土壤的重金属污染程度及污染区的植被组成和植物对重金属的积累特性，并筛选出理想植物修复材料，我们采用样方法于2003年秋、冬两次在湘西矿区进行调查采样。共调查4个地点，采集土壤样品20份，植物样品363份，植物样品分别属于125种、42科、102属。结果表明，植物叶片（或地上部）镉、铅、锌和铜的平均含量分别为19、81、637和8 mg kg^-1。镉含量最高的是半边莲（Lobelia chinensis）地上部，达到287 mg kg^-1，其次为龙葵（Solanum nigrum）、满天星（Hydddrocotyle sibthorpioides）和美洲商陆（Phytolacca americana）叶片的镉含量分别达到99、75和71 mg kg^-1。在调查区内，一些重金属含量较高的优势和次优势植物如美洲商陆、马兰（Kalimeris indice）、龙葵等可以用于重金属污染土壤的植物提取；一些重金属含量较低的优势和次优势植物如白茅（Imperata cylindrical）、五节芒（Miscanthus floridulus）等因为有发达的根系可以用于污染土壤的植物固定。为了进一步证实半边莲、龙葵和美洲商陆对镉的积累能力，我们于2004年7-10月再次在湘西地区重金属污染严重的矿业废弃地，对这三种植物和相应的土壤样品进行了全面调查和元素分析，共采集美洲商陆、龙葵、半边莲样品数量分别为43、26、11份，相应的土壤样品65份。分析结果表明，美洲商陆叶、龙葵叶和半边莲地上部镉的平均含量分别为55.2、36.9和141 mg kg^-1。所有样品中镉含量最高的是大田湾D17号美洲商陆叶的样品，镉含量达到402 mg kg^-1。三种植物叶或地上部镉含量与土壤中镉含量呈显著正相关。美洲商陆、龙葵、半边莲植株中镉的平均富积系数分别为1.19、2.63、1.63。室内溶液培养的结果显示，美洲商陆叶、龙葵叶及半边莲地上部镉含量最高分别达到了1150 mg kg^-1、1110 mg kg^-1、414 mg kg^-1。野外采集和室内水培结果都表明美洲商陆、龙葵和半边莲是三种新的镉超量积累植物。相比而言，三种植物中美洲商陆生物量大、镉含量高、耐性也强，而且对锌、锰、铅、铜也有很强的耐性和积累能力，更重要的是美洲商陆具有肉质化的宿根，能进行营养繁殖，种植一季可收获多茬，可能更适合于重金属复合污染土壤的植物修复。对野外34个样点的美洲商陆根、茎和叶及相应土壤样品锰含量进行分析，发现美洲商陆对锰也有很强的耐性和积累能力。34个不同采样点的美洲商陆叶片锰的平均含量为2020 mg kg^-1，其中大田湾Ⅱ样区的D26号美洲商陆样品叶片中的锰含量最高，为13400 mg kg^-1。相关性分析的结果表明，美洲商陆叶片中锰含量与相应土壤中总锰的含量在对数范围内呈显著正相关。此外，美洲商陆叶片对锰的平均积累因子为0.58，变化范围为0.60-3.90。水培试验结果也表明，在锰浓度为10 mM的营养液中生长15天的美洲商陆幼苗其叶片中锰含量可达到11600 mg kg^-1。再生苗水培试验也表明，在锰浓度为5mM的营养液中培养90天的美洲商陆，其叶片锰含量也可达11600mg kg^-1，而且刈割后培育出的再生苗在锰浓度同样为5mM的处理液中生长24天，其叶片中锰含量还高达9700 mg kg^-1，该处理的三生苗叶片中的锰含量仍然高达10200 mg kg^-1。这些结果说明，美洲商陆也是一种锰的超量积累植物。由于美洲商陆同时具有镉和锰超积累特性，并能进行营养繁殖再生新植株，因此在植物修复实践中具有重要的应用价值，并且也是一种研究植物对镉和锰吸收及积累机理的好材料。为了探讨美洲商陆对镉的吸收方式，我们采用水培法，比较了吉首和怀化两美洲商陆种群对镉处理的响应。同时还采用不同浓度的外源ABA和代谢抑制剂NaN₃分别与50μM镉对两种群美洲商陆进行复合处理，并分析了植物根、茎、叶的干重及镉含量。结果表明，镉处理15天显著抑制吉首和怀化两种群的生长，并且相同浓度镉处理下，两种群根、叶的干重没有显著差异，吉首种群根的镉含量显著低于怀化种群，而叶的镉含量显著高于怀化种群。外源ABA和NaN₃的加入，显著抑制了美洲商陆对镉的吸收，暗示美洲商陆对镉的吸收可能包含了被动吸收和主动吸收两个过程。为了弄清美洲商陆吸收和积累镉、锰之间的关系，以及镉／锰交互处理对美洲商陆生长和镉、锰吸收的影响，对野外美洲商陆叶片中元素含量进行相关性分析。结果表明，Cd含量跟Zn和Cu的含量成显著正相关，而跟Mg成显著负相关；Mn含量与Mg、Ni含量成显著正相关而与Ca成显著负相关。采用水培法，进行镉／锰交互处理，分析了美洲商陆生长及镉、锰的含量。结果表明锰能缓解镉对美洲商陆生长的抑制，缓解镉对叶绿素a的破坏，降低美洲商陆对镉的吸收和积累。这说明Cd、Mn和Mg在美洲商陆中可能共用了某些吸收或运输系统。美洲商陆叶片中镉、锰含量与硫含量在对数范围内成显著正相关，这说明美洲商陆叶片中可能存在某种含硫化合物参与了镉、锰的螯合解毒。采用水培法分别进行镉、锰处理，分析了美洲商陆根、叶中游离氨基酸的含量，但没有显著变化。随镉处理浓度增加，美洲商陆根和叶中非蛋白巯基和蛋白巯基含量都显著增加，而还原型谷胱甘肽和氧化型谷胱甘肽含量显著减少。此外，随锰处理浓度增加美洲商陆中根、叶中游离氨基酸含量没有显著变化，蛋白巯基逐渐减少，非蛋白巯基和GSH含量逐渐增加。推测镉、锰在美洲商陆中的解毒可能与某种含巯基的化合物有关，并且二者的解毒机制是有差异的。此外，我们在吉首市峒河下游污染河段发现了以龙须眼子菜（Potamogeton pectinatus）和竹叶眼子菜（Potamogeton malaianus）为优势植物的沉水植物群落。2004年夏季，分别在10个不同样点采集了该两种植物、河床底泥和水的样品并进行了元素含量分析。结果表明，调查区内河水和底泥的重金属含量均超过了国家环保总局规定的土壤和水的生态质量标准。植物中重金属含量与河水和底泥中重金属含量变化趋势一样，从上游至下游有增加的趋势。两种植物相比，龙须眼子菜叶中镉、锰的平均含量高于竹叶眼子菜，而铜、铅和锰的含量没有显著差异。除铅以外，植物中重金属的含量与水中的重金属含量呈显著相关，而与底泥则没有显著的相关性。室内浸提实验结果表明，龙须眼子菜对镉、铅、铜、锌和锰的平均转移效率分别达到84％、90％、91％、77％和84％。龙须眼子菜具有很强的转移水中重金属的能力，可以作为水体重金属污染的生物监测器，也可直接用于污染水源的植物修复。为了进一步评价龙须眼子菜在水污染控制与治理中的应用价值，我们采用室内浸提的方法研究了龙须眼子菜对镉、铅吸附的一般特征。结果表明，龙须眼子菜对镉、铅的吸附速率很快，大约20分钟就能达到吸附平衡，并且对镉、铅的最大吸附量分别达到32368、24776 mg kg^-1。龙须眼子菜对镉、铅吸附的时间动力学特征符合假二次方程，吸附的浓度动力学特征符合Langmuir方程。不同温度、不同pH值对龙须眼子菜吸附镉、铅离子的影响结果表明，与5℃、10℃、30℃处理相比，20℃时龙须眼子菜对镉、铅的吸附能力最强。在pH值3-7范围内，龙须眼子菜对镉的吸附随pH值升高而减少，对铅的吸附随pH值升高而增加。在0.5-10 g／L的盐度范围内，随着盐度的增加龙须眼子菜对镉吸附减少，但对铅的吸附却增加。由于龙须眼子菜对温度、酸碱度、盐度适应范围也很广，因此适宜用于各种复杂的污染水体的植物修复。更多还原

【Abstract】 Recently, there is considerable interest in developing cost—effective,environmentally friendly and plant-based technologies for the remediation of soils andwaters contaminated with heavy metals. This technology is termed phytoremediation.The technique of phytomining involves the use of hyperaccumulator plants to growand concentrate a metal. Subsequently, the crop is harvested and the metal exteracted.Hyperaccumulator plants were originally defined by Brooks et al. as taxa containing＞1000 mg kg^-1（ppm） Ni in their dry biomass. The extreme level of metal tolerance invascular plants is hyperaccumulation. Hyperaccumulators are defined as higher plantspecies whose shoots contain＞100 mg kg^-1 Cd,＞1000 mg kg^-1Ni, Pb and Cu, or＞10 000 mg kg^-1Zn and Mn when grown in metal-rich soils. Tolerance and metalaccumulation are two essential characteristics required for phytoremediation andphytomining. Hyperaccumulator is the most ideal plant used for phytoremediation andphytomining. So the present investigation aims to identify plant species that have anability to successfully grow and reproduce in adversely impacted mining environment,and to asses their metal accumulation capacities, and then to study the mechanisms ofmetal accumulation in plants. This information would be helpful in determining ifthose plant species are suitable for ecosystem recovery on mining areas andphytoremediation of metal-contaninated soils and water.To investigate the plant species composition and their ability to accumulateheavy metals at mining area in Xiangxi area, Southern China, we sampled plants andsoil samples twice at four contaminated sites in the Xiangxi area respectively in 2003autumn and winter. The concentrations of Cd, Pb, Zn and Cu in more than 363samples of 125 plant species and 20 soil samples were analyzed in the present study.The average concentrations of Cd, Pb, Zn and Cu in the plants were 19, 81,637 and 8mg kg^-1, respectively. The highest concentration of Cd in above-ground plant tissueswas found to be 287 mg kg^-1in the leaves of Lobelia chinensis Lour. at the Datianwansite, followed by Solamim nigrum L., Hydddrocotyle sibthorpioides and Phytolaccaamericana with 99, 75 and 71 mg kg^-1 Cd in the leaves. They might be potential Cdhyperaccumulators. At the three contaminated sites, some dominant and relative dominant species with high accumulation potential of metals, such as Kalimeris indice（L.） Sch.-Bip. and Solanum nigrum L., might be suitable for use in thephytoextraction of contaminated soils. The dominant and relative dominant specieswith low accumulation of metals and dense fibrous root systems, such as lmperatacylindrical （L.） Beauv. var. major C. E. and Miscanthus floridulus （Labill.） Warb.,might be suitable for stabilizing such metal contaminated sites.To validate Cd accumulation capacities of the three plants, Lobelia chinensisLour., Solanum nigrum L. and Phytolacca americana L., in the present study, theconcentrations of Cd in the three plant species, and the corresponding soil samplesfrom the mining and smelting contamination sites in the Xiangxi area of HunanProvince, China were further studied. The results showed that the averageconcentrations of Cd were 55.2, 36.9 and 141 mg kg^-1in P. americana leaf,, S. nigrumleaf and L. chinensis shoot, respectively. The highest Cd concentration of 402 mg kg^-1was found in the leaves of P. americana. Significant relationships were observedbetween the concentrations of Cd in the aerial tissues of the three plants and those ofcorresponding soils on a logarithmic scale. The average bioaccumulation factors ofCd in P americana （31 samples）, S. nigrum （22 samples） and L. chinensis （10samples） were 1.19, 2.96 and 1.63, respectively. Hydroponic experiments were alsoconducted to investigate the Cd uptake ability and mechanism of the three plants insolutions. The maximum Cd concentration in the aerial tissues of P. americana, S.nigrum and L. chinensis were 1150, 1110 and 414 mg kg^-1 under the laboratoryhydroponic conditions, respectively. The results suggested that P. americana, L.chinensis and S. nigrum may be considered to be Cd hyperaccumulators. Among thethree plant species under study, P. americana might be a suitable candidate for use inthe phytoremediation of metal contaminated soils due to its fast growth rate, highaboveground biomass, and elevated adaptability to a wide range of soil Cdconcentrations.The concentrations of Mn in P. americana and the corresponding soil samplesfrom the mining and smelting contamination sites in the same sites were furtherstudied. The results showed that the average concentration of Mn was 2020 mg kg^-1inP. americana leaf. The highest Mn concentration of 13400 mg kg^-1was found in theDatianwanⅡsite. Significant relationships were observed between the concentrationsof Mn in the aerial tissues of the plant species and those of corresponding soils on alogarithmic scale. The bioaccumulation factors of Mn in P. americana averaged 0.58, and ranged from 0.60-3.90. Hydroponic experiments were also conducted toinvestigate the Mn uptake ability and mechanism in solutions. The maximum Mnconcentration in the leaf of P americana was 11600 mg kg^-1in the 10000μM Mntreatment for 15 days. The results of regeneration seedling experiment showed thatMn concentration in the leaf P. americana was also up to 11600 mg kg^-1in the 5000μM Mn treatment for 90 days, besides the regeneration seedling of P americana alsocontained high concentration Mn in the leaf up to 9700 mg kg^-1in the 5000μM Mntreatment for 24 days after first harvested. These results suggested that P americanamay be considered to be a manangese hyperaccumulator. As a manangese andcadmium hyperaccumulator, P. americana with high biomass may have a greatpotential for future use in phytoremediation of heavy metals compound contaminatedsoil and studies on the mechanisms of rhizosphere interaction, uptake, transport andsequestration of Cd and Mn.To understand Cd adsorption manner by P americana, hydroponics experimentwas conducted. The growth and Cd bioaccumulation were compared between two Pamericana populations respectively from Huaihua and Jishou. The growth wasremarkably inhibited and Cd contents in leaf and root were significantly increasedwith the Cd concentration increasing in the nutrition solution. No significantdifference in the growth between the Huaihua and Jishou P americana populations,but Cd content in the root of Huaihua population was markedly higher than thepopulation of Jishou. Extrinsic ABA and metabolic inhibitor NaN₃ drastically reducedthe extent of Cd bioaccumulation in the leaf of two P. americana populations. ABAcan induce stomata closure and reduce the stomata transpirations. Besides, NaN₃ isthe inhibitor of the H⁺-ATPase in the plasma membrane and vacuole membrane.Therefore, these results suggested that Cd adsorption manner by P americanainvolved passive and active adsorption two processes.We examined the relationships between the concentrations of Cd and Mn andother transition metal in leaves of forty P americana samples from the field surveys.The concentrations of Cd significantly positively related with the concentrations of Znand Cu, but remarkably negatively correlated with the concentrations of Mg. On theother hand, marked positive relationships were found between the contents of Mn andthe contents of Mg and Ni, but significantly negative relationship between thecontents of Mn and Ca. Under hydroponics condition, the effect of the interactionbetween Cd and Mn on the growth and the concentrations of Cd and Mn were studied. The results showed that Cd toxicity to P. americana was weakened and Cdconcentrations in leaf, stem and root all decreased due to addition of Mn. Theconcentrations of Mn in leaf, root and stem decreased slightly due to the competitionof between Cd and Mn. These results suggested that Cd²⁺, Mn²⁺ and Mg²⁺may beabsorbed and transported via common pathway.We examined the relationships between the concentrations of Cd and Mn andsulfate in leaves of forty P. americana samples from the field surveys. Theconcentrations of Cd and Mn significantly positively correlated with theconcentrations of sulfate. The result indicated that Cd and Mn detoxification involvedin chelation of Cd and Mn in the cytosol by some thiol compounds. We furtherinvestigated the effect of Cd on the amino acid, glutathione and protein andnon-protein thiol contents of leaf and root in P americana under hydroponicscondition. The data showed that no significant deference was found after Cd and Mntreatment in the contents of amino acid in leaves and roots. But the contents of proteinand non-protein thiol markedly increased and the contents of GSH and GSSG in leafand root observably decreased with the Cd concentration increasing in the nutritionsolution. GSH is the precursor for the enzymatic synthesis of phytochelatins （PC）.Besides, the protein thiol in leaf of P. americna slightly decreased, non-protein andglutathione increased with the Mn concentrations in the solutions. All the resultssuggested that the phytochelatins was the potential ligands with Cd in leaf of P.americana, and the detoxification mechanisms were different between Cd and Mn.In addition, we also investigated in Donghe River in Jishou City located innorthwest Hunan Province, China and found the submerged plant community with thedominant species Potamogeton pectinatus and Potamogeton malaianus. In the study,heavy metal concentrations in the leaves of aquatic plants P. pectinatus and P.malaianus and the corresponding water and sediment samples from the Donghe Riverwere studied. Results showed that the elevated levels of metals were found in thesediments, especially Cd, Mn and Pb in comparison with the ecotox threshold valuesof soil and water developed by the Chian. Environmental Protection Agency. Thehighest concentrations of Cd, Pb, Cu, Zn and Mn in the leaves of P. pectinatus were596, 318, 62.4, 6590 and 16000 mg kg^-1（DW）, respectively. Significant relationshipswere observed between the concentrations of Zn, Cu and Mn in the leaves of bothaquatic plants and those in water. Laboratory experiments were also conducted toinvestigate the abilities of P. pectinatus to remove heavy metals from contaminated water. The average removal efficiencies were 84％, 90％, 91％, 77％and 84％for Cd,Cu, Pb, Zn and Mn, respectively, within the 5 h treatment. The results showed that P.pectinatus had high capability of removing heavy metals directly from water.To further estimate the application worth of P pectinatus in the water pollutioncontrol and treatment, biosorption of metals by P. pectinatus in the laboratoryexperiment was carried out to investigate Cd and Pb adsorption properties and theeffects of water temperature, pH value and salinity on Cd and Pb adsorption of Ppectinatus. Results were compared with other materials. Data obtained from the intialadsorption studies indicated that P. pectinatus was capable of removing Cd and Pbfrom solution. The metal biosorption was fast and equilibrium was attained within 20min. Overall adsorption processes of Cd and Pb with the increasing time were all bestdescribed by pseudo second-order kinetics. Data obtained from further batchadsorption with the metal concentration increasing studies conformed well to theLangmuir Model. Maximum adsorption capacities of Cd and Pb onto P. pectinatuswere 32368, 24776 mg kg^-1. The metal concentrations in the leaf of P. pectinatus werehighest at the 20^C compared to 5, 10 and 30℃. Other condition parameter notchanging, the concentration of Cd in leaf of P. pectinatus decreased but theconcentration of Pb increased with the pH value from 3 to 7 and salinity from 0.5 g/Lto 10 g/L increasing. P pectinatus more often has a high biomass and tolerates verywide ranges of temperature, pH value and salinity. Therefore, P. pectinatus can beused as a biological indicator while determining aquatic environmental pressures andused for phytoremediation of multiple metals contaminated water.更多还原