摘要
胞外聚合物（Extacellular Polymeric Substances, EPS）是一定环境条件下由细菌、真菌、藻类细胞分泌的高分子聚合物，对重金属具有良好的吸附效果。研究证明真菌菌体对重金属具有良好的吸附性，真菌易得易培养，可作为良好的生物吸附材料。细胞壁是生物吸附过程中金属离子的主要富集场所，真菌细胞壁上的附着物质，主要为胞外聚合物，占总附着物质含量的75%左右。一直以来，人们的研究主要集中在活性污泥EPS及细菌EPS方面，而较少涉及真菌EPS。但不同来源的EPS具有不同的结构和组分，其对重金属的吸附能力和作用机理不同。为此，本文以课题组筛选的烟曲霉菌株为研究对象，研究了不同提取方法对烟曲霉EPS组分的影响规律，分析了其主要成分及含量，采用极谱法测定其络合容量、结合位点数和条件稳定常数；通过批式实验确定了其吸附重金属（Cu~(2+)、Cd~(2+)）的影响因素及最佳吸附条件，并考察了解吸性能。同时，利用红外光谱、X射线能量散射光谱探讨了EPS吸附去除金属离子Cu~(2+)、Cd~(2+)的过程机理。
通过比较分析H_2SO_4法、NaOH法、EDTA法和离子交换法（CER法）提取EPS的成分得出离子交换法提取的烟曲霉EPS多糖含量较高，对细胞破坏程度较小。确定了优化的CER法提取烟曲霉EPS的条件为：树脂投加量为70g g~(-1)、振荡速度为200r min~(-1)、振荡时间2h、液相pH6-8。提取的烟曲霉EPS用截留分子量为3500Da的透析袋透析纯化，纯化后EPS中多糖、蛋白质、核酸和脂类的含量分别为392±8、17.6±2、18.3±2、25.7±2.8mg g~(-1)DWEPS。
用微分脉冲极谱DPP（Differential Pulse Polarography）法得到了不同pH下Cu~(2+)、Cd~(2+)和Pb~(2+)的滴定曲线。结果表明在pH值4.0-7.0范围内烟曲霉EPS结合Cu~(2+)和Cd2离子能力随着pH值的增加而增加，Pb~(2+)在pH6.0和7.0时与EPS反应产生白色凝胶状沉淀物。EPS对不同金属离子的结合能力也有差异，EPS对Cu~(2+)、Cd~(2+)和Pb~(2+)结合能力排序为Cu~(2+)>Pb~(2+)> Cd~(2+)。并用Ruzic方法对Cu~(2+)、Cd~(2+)和Pb~(2+)的滴定曲线进行拟合，得到m/(CM-m)对m的Ruzic拟合曲线，拟合的结果与DPP法是一致的。
以提纯的烟曲霉EPS为吸附剂，考察了pH值、EPS投加量、吸附时间、温度、盐度及共存离子等对EPS吸附水中Cu~(2+)、Cd~(2+)的影响规律、热力学及动力学吸附特征，结果表明，pH值、EPS浓度、金属离子的初始浓度、NaCl浓度及共存离子对EPS吸附Cu~(2+)、Cd~(2+)有明显影响。在pH7.0、温度25℃、Cu~(2+)和Cd~(2+)初始浓度5mg L~(-1)和7mg L~(-1)， EPS投加量为135mg L~(-1)时，Cu~(2+)和Cd~(2+)的吸附去除率分别为90.6%和96.7%。EPS对Cu~(2+)、Cd~(2+)的吸附过程均可分为快速吸附--慢速吸附--吸附平衡三个阶段，达到吸附平衡的时间为3h，Freundlich和Langmuir吸附模型均能较好地拟合烟曲霉EPS对Cu~(2+)和Cd~(2+)的等温吸附实验数据，由Langmuir吸附模型得到的EPS对Cu~(2+)和Cd~(2+)的的最大吸附容量分别为40mg g~(-1)和85.5mg g~(-1)。Freundlich常数n值分别为4.04和3.59。烟曲霉EPS对Cu~(2+)、Cd~(2+)的吸附为一个吸热的自发过程。烟曲霉EPS的吸附动力学符合拟二级动力学模型。
探讨了烟曲霉EPS中Cu~(2+)、Cd~(2+)的解吸性能。结果表明，pH对解吸烟曲霉EPS上的Cu~(2+)、Cd~(2+)的影响非常明显。随着pH的降低解吸率升高，最大解吸率都超过90%，确定解吸的最佳pH为1.0。而NaCl对解吸烟曲霉EPS上的Cu~(2+)、Cd~(2+)的效果不佳，NaCl浓度达到2.0mM时，Cu~(2+)和Cd~(2+)的解吸率分别只达到50.3%和53.8%。
红外光谱(FTIR)分析表明，烟曲霉EPS吸附Cu~(2+)、Cd~(2+)过程中多聚糖中的羟基、羧基和C-O-C等是起作用的主要官能团。X射线能量散射光谱（EDX）分析表明EPS主要元素包括C和O，还含有P、S、K、Ca、Mg、Al和Na等元素，当EPS与Cu~(2+)离子作用后，EPS中Ca、Mg元素峰消失；与Cd~(2+)作用后，EPS中Ca元素峰明显减弱，Mg元素峰消失。同时分别在8keV和3.1keV处出现了Cu、Cd元素峰，在吸附过程中Ca、Mg被Cd~(2+)、Cu~(2+)交换下来，说明离子交换是EPS吸附Cd~(2+)、Cu~(2+)的机理之一。烟曲霉EPS与金属离子相互作用的机理还包括络合作用和静电作用，是几种机理共同作用的结果，而以络合作用为主。
Extacellular polymeric substances (EPS), produced by many microorganisms, such asalgae, bacteria, and fungi, play a crucial role in the biosorption of heavy metals. A number ofstudies showed that fungi were very effective for heavy metal adsorption. Moreover, fungican be easily cultured and obtained from a number of industrial by-products. Heavy metalsare accumulated mainly in cell wall in the process of biosrption. About75%of thecomponents of fungi cell wall are EPS. In recent years, some researchers have focused onEPS extracted from activated sludge and pure bacterial strains and investigated differentcombinations of metal ions, microorganisms and experimental conditions. But the biosorptionproperties of EPS from fungi remain less explored. The components of EPS are differentfrom microorganisms and results in different biosorption capacity and mechanism. In thepresent study, Aspergillus fumigatus, an isolated fungus in our lab, was used for EPSextraction. This study aims to investigate the sorption behaviour of the EPS for Cu~(2+)and Cd~(2+)in aqueous solutions. The equilibrium isotherms and the effect of various factors, such as pHand initial metal concentration, on binding of these metals were studied. The presence offunctional groups in the EPS which may play a role in sorption process was confirmed byFTIR. And EDX analysis was used for further confirmation of the mechanism of the sorptionof heavy metals by the EPS.
The efficacies of extracting EPS from Aspergillus fumigatus using H2SO4, NaOH, EDTAand CER (cation exchange resin) under various conditions were compared. Results showedthat CER was most effective in extracting EPS from Aspergillus fumigatus. The content ofpolysaccharide in EPS extracted by CER was very high. While, nucleic acids were very low,suggesting the EPS extracted were not contaminated by intracellular substances. So, CERwas chosen as extracting reagent and the optimal conditions were as follows: CER dosage70g g~(-1), oscillating speed200r min~(-1), extracting time2h and pH6-8. The contents of thecarbohydrate, protein nucleic acid and lipids in EPS were392±8、17.6±2、18.3±2、25.7±2.8mg g~(-1)DWEPS, respectively.
Biosorption of Cu~(2+), Cd~(2+)and Pb~(2+)of EPS was investigated as a function of pH usingdifferential pulse polarography (DPP) and the Ruzic model. Results showed that the EPSbiosorption capacity determined using either the direct titration curves i=f (CM) or the methodproposed by Ruzic were coincident. Cu~(2+)had the highest affinity with EPS followed by Pb~(2+)and Cd~(2+). The total number of binding sites for Cu~(2+)and Cd~(2+)increased with pH in the range of4.0-7.0. Similar trend was observed for Pb~(2+)at pH4.0-5.0, while precipitates wereobserved at pH6.0and7.0.
Sorption of Cu~(2+)and Cd~(2+)onto the extracellular polymeric substances (EPS) producedby Aspergillus fumigatus was investigated for the initial pH of the solution, EPSconcentrations, contact time, NaCl concentration, initial metal ion concentration and thepresence of other ions in the solution. The results showed that the adsorption of metal ionswas significantly affected by pH, EPS concentrations, initial metal concentration, NaClconcentration and co-ions. When pH was7.0, temperature was25℃, the initial Cu~(2+)andCd~(2+)concentration was5mg L~(-1)and7mg L~(-1), EPS dosage was135mg L~(-1), the removalefficiency of Cu~(2+)and Cd~(2+)reached90.6%and96.7%, respectively. The sorption processesof Cu~(2+)and Cd~(2+)with EPS both have two steps: fast sorption and slow sorption, sorptionequilibrium time was about3hours. Both the Langmuir and Freundlich adsorption modelscould describe the thermodynamic sorption processes of Cu~(2+)and Cd~(2+). The maximumsorption capacities of A. fumigatus EPS were40mg g~(-1)EPS and85.5mg g~(-1)EPS. Values of ncalculated from the Freundlich model were4.04and3.59for Cu~(2+)and Cd~(2+), respectively.The sorption was a spontaneous endothermic process. The sorption kinetics fitted well topseudo-second-order kinetic model.
The desoption of Cu~(2+)and Cd~(2+)absorbed in EPS was investigated. Results revealed thatdesorption of Cu~(2+)and Cd~(2+)from EPS was significantly affected by pH. The desorptionpercentage was increased with decrease of pH. The maximum desorption percentage washigher than90%for both Cu~(2+)and Cd~(2+). The optimal desorption pH was1.0. However, thedesorption percentage for both Cu~(2+)and Cd~(2+)was low when using NaCl as desorptionreagent. When concentration of NaCl was2.0mM, the desorption percentage for Cu~(2+)andCd~(2+)was50.3%and53.8%, respectively.
Analysis of FTIR spectra demonstrated that hydroxyl, carboxyl and C-O-C ofpolysaccharide were the main functional groups in the reaction between metal and EPS.Energy dispersive X-ray (EDX) system analysis revealed that C and O were the dominantelements in EPS, and P, S, K, Ca, Mg, Al and Na were also included in. The EDX pattern forthe Cu~(2+)-loaded and Cd~(2+)-loaded EPS revealed distinctive peaks for the respective metals.However, Ca and Mg were not detected in the EDX spectrum of Cu~(2+)-loaded EPS.Furthermore, for the Cd~(2+)-loaded EPS, the Ca content was found to be lower and Mg was notdetected. The results showed that the ion-exchange was an important mechanism involved inthe Cu~(2+)and Cd~(2+)sorption process taking place on EPS. Complexation and electrostaticinteractions were also involved in. There were several mechanisms responsible for the sorption of Cu~(2+)and Cd~(2+)together. And complexation was the main mechanism.
引文
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