【作者】 刘晓猛；

【导师】 俞汉青；

【作者基本信息】 中国科学技术大学 ， 应用化学， 2008， 博士

【摘要】 生物处理是目前水污染控制的主要技术手段。在废水处理反应器中起重要作用的微生物通常以絮体污泥、生物膜和颗粒污泥三种聚集体的形式存在。这些微生物聚集体的形态、结构、功能、特性影响并决定着废水生物处理系统的运行稳定性、处理效率和出水水质。另一方面,微生物聚集体的形成过程却极为复杂,不仅形成过程时间长,而且影响因素众多。因此,对微生物聚集体的相互作用以及形成机制的研究是十分必要的。本论文基于大量的试验研究结果和DLVO理论的拓展应用,系统研究了废水处理反应器中微生物聚集体的表面特性、相互作用以及形成机制,深入探讨了微生物胞外多聚物EPS在微生物相互作用中的作用机制,并成功构建了微观尺度下微生物聚集体的三维模型。主要研究内容和研究成果如下:1.基于表面热力学分析所反映的微生物表面性质的差异,以一株产氢光合细菌（Rhodopseudomonas acidophila）为对象,建立了利用DLVO理论来解析与预测微生物之间的吸附与絮凝行为的理论方法,并探索了导致细菌絮凝性能变化的本质原因。在0.1 mol L^-1电解质浓度下以及pH约为7.0时候,体系具有最优的絮凝性能;Ca²⁺的加入可以有效地提高体系菌液的絮凝性能;EPS对体系絮凝性能的变化影响很大;导致光合细菌絮凝性能较差的主要原因是光合细菌自身特殊的表面性质,即较小的Hamaker常数值(2.27×10^-23J)与正的界面吸附自由能(21.75 mJ m^-2)。2.通过对真氧产碱杆菌（Ralstonia eutropha）生长周期的研究,成功地对微生物生长周期中表面特性以及絮凝性能的变化进行了定量表征。在生长周期中真氧产碱杆菌的水接触角与甲酰胺接触角在24 h内迅速下降,其疏水性在对数生长期内迅速降低;细菌表面张力中非极性作用项与极性作用项的比值(γ_B^LW/γ_B^AB)在生长周期中先迅速下降,随之维持在一个较低的水平,这说明在生长周期中细菌表面由疏水性为主变为以亲水性为主,从而导致在培养过程中细菌的稳定性逐渐升高;Lewis酸-碱水合作用项在整个培养过程中对总位能贡献由最初的吸附转变为排斥是导致体系稳定性变化的主要因素。3.通过对真氧产碱杆菌不同生长条件的优化分析,提出了一种能够快速、有效地定性预测不同底物状态下细菌絮凝性能大小的新方法。在30 g/L葡萄糖为底物、初始pH约为8.0的较优生长条件下,R.eutropha的生长快速,絮凝性能良好,合成PHAs的量也较多;而在其他条件下,真氧产碱杆菌的生长反而出现抑制现象;同时,在不同底物类型下细菌具有不同的斥力势垒水准,其絮凝性能的大小可以通过预测其斥力势垒的高低来判断。4.利用快速、灵敏、选择性高的三维荧光光谱方法,表征了好氧活性污泥与厌氧产甲烷污泥EPS的特性。好氧污泥中存在两个荧光峰,位置分别在280-285/340-350 nm和340-350/430-450 nm处,分别对应于蛋白与NADH的特征吸收峰;而在厌氧产甲烷污泥中除去出现蛋白特征吸收峰外,还在Ex/Em=400-410/465-475 nm处出现辅酶F₄₂₀特征吸收峰,它是产甲烷菌所特有的物质;两种EPS的三维荧光光谱参数（如峰位置、峰强度、不同峰强度比）都有所不同,说明了两者在成分和结构上的差异性。5.通过好氧和厌氧污泥表面EPS的提取实验,观察到污泥絮凝性能的变化。研究结果表明:无论是松散结合EPS（Loosely bound EPS,LB-EPS）还是紧密结合EPS（Tightly bound EPS,TB-EPS）都对污泥絮凝性能有着有益的贡献,且随着EPS的不断剥离,污泥的絮凝性能逐渐变差。以DLVO理论为基础构建了定量表征EPS在微生物相互作用中作用机制的新方法,利用该方法对试验数据的解析结果表明,在活性污泥中TB-EPS对于微生物的絮凝性能变化的贡献较大。6.成功建立了微观尺度下微生物聚集体的三维全局优化模型,并利用建立的模型优化了微生物聚集体数目从3增加到100所有的稳定最优构型。该方法是基于分子力学手段,将DLVO理论作为优化体系的势能函数,从而利用全局优化算法建立三维构型的优化模型。通过对构型结果的分析,提出了微生物聚集体生长的模型,即微生物粒子以一种多层分布的结构聚集在一起,从外部来看,聚集体结构呈现一种近似球形的分布。更多还原

【Abstract】 Biological treatment is one of the most widely used wastewater treatment processes.The microbial aggregates,including flocs,biofilm and granules,are the key element in the bioreactors.Their structure,shape,function and surface characteristics may significantly influent the stability,efficiency and effluent quality of bioreactors,and play a crucial role in wastewater biological treatments.However, the formation of microbial aggregates is a time-consuming process,and can be influenced by many factors.Therefore,study on the interactions among the microbial aggregates is essential to understand the microbial aggregation process.In this work, based on experimental results and innovational analytical methods,the original and extended DLVO theories are employed for characterizing the surface characteristics, the microbial adhesion and flocculation behavior of microbial aggregates in biological wastewater treatment.The roles of extracellular polymeric substances（EPS）in microbial aggregates are also explored and quantified.Main contents and results are as follows:1.Based on the microbial surface thermodynamic approach,which reflects the bacterial surface characteristics,a simple and effective method for the evaluation and determination of microbial adhesion and flocculation was developed.Taking as a photosynthetic bacterium Rhodopseudomonas acidophila as example,the relationship between its surface characteristics and flocculability was evaluated.The appropriate electrolyte concentration and pH for the strain were found to be 0.1 mol L^-1NaCl solution and pH 7.0,respectively.The addition of Ca²⁺could improve its flocculability.In addition,the EPS produced by the cell were observed to have a significant effect on its flocculation.The effective Hamaker constant and the repulsive total interfacial free energy seemed to be responsible for the poor flocculability of the strain.2.The microbial surface and flocculability were quantitatively characterized through the combination of the surface thermodynamic and the extended DLVO approaches,with Ralstonia eutropha,a polyhydroxybutyrate-producing bacterium,as an example.The total interfacial free energy(△G_adh)was changed from -80 mJ m^-2to 28.5 mJ m^-2,and the ratio ofγ_B^LW/γ_B^ABdecreased significantly in its entire growth process.This suggests that the bacterial surface changed from hydrophobic into hydrophilic.As a result,the stability ratio of suspensions increased with the increasing cultivation time.The Lewis acid-base interaction(W_AB)in the extended-DLVO approach could provide an additional asset to the increase or decrease in the total energy barrier,and,therefore,seems play an important role on bacterial suspensions.3.The different cultivation conditions for R.eutropha could be optimized and evaluated based on the extended DLVO theory.It was found that glucose of 30 g/L and pH 8.0 was the proper cultivation condition.The strain also had a good flocculability under the conditions.The energy barrier levels with different substrates were highly related to the substrate type.This could provide a feasible and useful method to qualitatively describe the microbial flocculability based on its substrate type.4.As a rapid,sensitive and selective analytical method,3-dimensional excitation-emission matrix fluorescence spectroscopy was applied to characterize the EPS extracted from the aerobic and anaerobic sludge in wastewater treatment reactors. Two fluorescence peaks were identified at（excitation/emission）280-285/340-350 nm and 340-350/430-450 nm in the aerobic sludge EPS fluorescence spectra.The two peaks were attributed to the protein-like and NADH fluorophores,respectively.The anaerobic sludge EPS fluorescence spectra also had two fluorescence peaks.One peak was similar to that for the aerobic sludge EPS and attributed to the proteins.Another peak corresponded to the F₄₂₀fluorophore,which was the characteristic substance of the methanogens.The differences in the EPS fluorescence parameters,e.g.,peak locations,intensities and ratios of various peak intensities,indicate the difference in the chemical structures of the EPS from various origins.5.The change of sludge flocculability and the role of EPS were evaluated from the EPS extraction experimental results.Both.the loosely bound EPS EPS（LB-EPS） and tightly bound EPS（TB-EPS）could provide an effective contribution on the sludge flocculability.After the extraction of the LB-EPS and TB-EPS,the sludge flocculability decreased significantly.Thus,based on the DLVO theory,the quantification method on the role of EPS in the flocculation was proposed.The results shows the TB-EPS in the actived sludge played a more significant role in the microbial flocculation.6.A global optimization model,based on the molecular mechanics（MM）method, was developed and constructed to simulate the three-dimensional structural morphology of microbial aggregates at a microscopic scale.The DLVO theory, usually used to describe the interactions between particles in colloidal suspensions,is modified to explain the interactions among microbial particles.With the constructed model,some putative aggregate micro-motifs with the microbial particles from 3 up to 100 was acquired.Results suggest that the microbial clusters were built up with the same growth pattern,which were arranged with a multi-shell structural packing style. This model offers a rational platform to investigate the microbial aggregate formation process.更多还原