【作者】 OSAMA ABDALLA ABDELSHAFI MOHAMAD；

【导师】 韦革宏；

【作者基本信息】 西北农林科技大学 ， 环境科学， 2012， 博士

【摘要】 本论文利用从中国甘肃省金属尾矿分离到的根瘤菌作为生物吸附剂材料去除污染水体环境中的铜，系统的研究了铜耐受菌株的生物吸附特性和吸附机理，拟为重金属污染环境的修复与治理提供新的材料和方法。从甘肃省金属尾矿区豆科植物刺槐根瘤中共分离得到73株根瘤菌，其中菌株CCNWGS0123对铜的抗性较高，它可在pH4.0和pH10.0的培养基上生长，耐6%浓度的氯化钠，可利用琥珀酸钠、丙酮酸钠、丙二酸钠、D-木糖、半乳糖、D-果糖、丙酸钠、L-阿拉伯糖、D-核糖、鼠李糖、甘露糖、麦芽糖、柠檬酸钠、醋酸钠、葡萄糖、蔗糖、乳糖和肌醇为唯一碳源；利用半胱氨酸、L-天门冬酰胺、DL-α-氨基酸、L-甲硫氨酸、DL-组氨酸、天冬氨酸、丙氨酸、L-胱氨酸和L-谷氨酸为唯一氮源；能够抗300μg/ml氨苄西林、300μg/ml氯霉素、100μg/ml卡那霉素、100μg/ml氨苄西林钠、300μg/ml林可霉素、50μg/ml硫酸链霉素和5μg/ml丁胺卡那霉素；菌株CCNWGS0123可在含有2.2mM铜、0.1%甲基橙、0.2%溴酚蓝、0.2%刚果红和0.2%中性红的培养基上生长；可在YMA上产过氧化氢；16S rRNA、nodA、nodC和nifH基因序列分析，确定该菌株属于Mesorhizobium amorphae。在菌株CCNWGS0123对铜离子的吸附特性研究中发现，其活细胞和死细胞均可作为一种生物吸附剂去除水溶液中铜污染；在pH为5.0，转速为150rpm，温度为28°C时，铜的去除率最高，30分钟到达吸附平衡。吸附等温线拟合表明，Langmuir方程比Freundlich吸附式更适于拟合吸附过程。在菌体接种量为100mg/l和铜离子初始浓度为0.5g/l时，铜离子的去除率最高。傅立叶红外光谱(FT-IR)分析表明，细胞壁上O-H、N-H、C-H、C=O、-NH、-CN、C-N、C-O、酰胺（-I、-II、-III）、不饱和烯烃、烷基和芳香团等官能团参与了菌体和铜的相互作用。扫描电镜（SEM）显示，铜胁迫下细胞出现变形、聚集及细胞表面受损等现象。X-射线扫描能谱（EDX）分析结果显示，在1.1keV,8.1keV和8.9keV处出现了三个明显的吸收峰，表明细胞表面有铜的吸附。此外，铜的富集与细胞生长和生物量的增加有关，菌株是可能通过产生胞内多糖，将铜离子固定在细胞内，或通过在细胞外和细胞壁上分泌化合物螯合铜离子。本研究是用M. amorphae CCNWGS0123作为铜离子吸附剂的首次研究，该菌株不仅可与刺槐作为共生固氮促进植物生长，改善土壤营养条件，还可以作为生物修复体系去除污染土壤中的铜。该研究系统的阐明了CCNWGS0123作为吸附剂移除铜离子特性和机制，发现使用少量的菌体（活细胞和死细胞）进行生物吸附去除水体中的铜是一个经济、环保的途径，可用于水体污染处理材料。更多还原

【Abstract】 A total of76rhizobial strains were isolated from the nodules of Robinia pseudoacaciagrown in the mine tailings of Gansu Province, China. The strain CCNWGS0123was isolatedscreened and many physiological and biochemical characteristics were carried out. The resultsshowed that strain CCNWGS0123can grow in YMA medium at pH4.0to10.0or in6%NaCl. It can use the following materials as sole carbon sources: sodium succinate, sodiumpyruvate, sodium malonate, D-xylose, galactose, D-fructose, sodium propionate, L-arabinose,D-ribose, rhamnose, mannose, maltose, sodium citrate, sodium acetate, glucose, sucrose,lactose and inositol. L-cysteine, L-asparagines, DL-α-amino acid, L-methionine, DL-histidine,L-aspartic acid, L-alanine, L-cystine and L-glutamic acid could be used as sole nitrogensources. CCNWGS0123can grow on YMA plates supplemented with the followingantibiotics and chemical compounds:300μg/ml Ampicillin,300μg/ml chloramphenicol,100μg/ml kanamycin,100μg/ml ampicillin sodium,300μg/ml lincomycin,50μg/mlStreptomycin sulfate and5μg/ml Amikacin2.2mMcopper,0.1%methyl orange,0.2%Bromo-thymol blue,0.2%congo red and0.2%Neutral red. CCNWGS0123can producehydrogen peroxide in YMA. Based on the analysis of16S rRNA, nodA, nodC and nifH genessequences, the strain CCNWGS0123was identified as Mesorhizobium amorphae.Our studies showed that both dead and live cells of M. amorphae CCNWGS0123areeffective absorbents for copper removal from aqueous solution. The removal of copperincreased significantly by increasing the pH up to5.0, agitation speed150rpm andtemperature28°C. Equilibrium was reached within30min. The absorption isotherms could bewell fitted by the Langmuir equation followed by the Freundlich equation. The highestremoval of initial copper concentration was achieved at a dosage of100mg/L and the highestbiosorption capacity was found at an initial concentration of0.5g/L Cu (II). Fouriertransform-infrared spectroscopy (FT-IR) analysis indicated that many functional groups, suchas O-H, N-H, C-H, C=O,-NH,-CN, C-N, C-O, amide (-I,-II,-III), and unsaturated alkenes,alkyls and aromatic groups on the cell surface, were involved in the interaction betweenCCNWGS0123and Cu ion. Scanning electron microscope (SEM) results showed deformation,aggregation and cell-surface damage due to the precipitation of copper on the cell surface.Energy Dispersive X-ray Scanning (EDX) showed three distinct peaks at1.1keV,8.1keVand8.9keV, implying that copper ions have been absorbed on the cells. Moreover, copperaccumulation was related to cell growth or increased biomass. The CCNWGS0123strain could have produced the intracellular polysaccharide and immobilized the copper ions in theircells, or secreted compounds onto the extracellular space and cell wall. In addition, our resultssuggested that Robinia pseudoacacia and M. amorphae CCNWGS0123could be a system toextract copper from contaminated soils. To our knowledge, this is the first time that M.amorphae CCNWGS0123has been used as an absorbent of copper ions. Based on our results,we foresee the possibility of using Mesorhizobium amorphae CCNWGS0123for the removalof copper ions from aqueous solution compared with other biomaterials, thus reducingaqueous environmental pollution. It appears that the most economical way to use it would beto use small batches of biomass to remove toxic concentrations of copper ions from aqueoussolution. Dead and live cells of CCNWGS0123showed potential as an efficient biosorbent forthe removal of copper from aqueous solutions. Taking present findings into consideration,dead cells of CCNWGS0123were proven to be more efficient and low-cost biosorbents thanlive ones, and can be utilized as an alternative in the treatment of wastewater.更多还原