【作者】 徐富；

【导师】 阮文权；

【作者基本信息】 江南大学 ， 环境工程， 2013， 博士

【摘要】 在生产规模的厌氧废水处理中，厌氧反应器的运行稳定性和高效能在一定程度上取决于能否培养出沉降性能好和产甲烷活性强的厌氧颗粒污泥。如果厌氧反应器内的污泥以松散的絮状体存在，则易出现污泥流失、有机负荷低、处理效果差等问题。成熟的厌氧颗粒污泥属于稀缺、生长慢、难培养、出厂价格高、运输费用大的商品。因而，研究厌氧颗粒污泥生产规模化培养技术，并解决工程应用的一些问题，是具有一定的实际意义和理论价值。本论文通过好氧剩余活性污泥启动生产规模厌氧反应器，研究厌氧颗粒污泥规模化培养，用灰色关联度研究颗粒化影响因素的重要程度，采用分子生物技术手段研究微生物种群结构的稳定性，投加无机阳离子Ca²⁺和聚丙烯酰胺（polyacrylamide, PAM）促进因子研究厌氧污泥颗粒化的生物强化作用，并比较厌氧颗粒污泥基质降解动力学，分析厌氧颗粒污泥生产规模化培养及其形成机制。主要研究结果如下：（1）通过控制有机负荷（organic loading rate, OLR）为8.8kg化学需氧量（chemicaloxygen demand, COD）·m^-3·d^-1、液体上升流速为5.4m·h^-1和水力停留时间（hydraulicretention time, HRT）为5.9h处理啤酒废水，好氧剩余活性污泥实现厌氧颗粒污泥规模化培养，颗粒污泥粒径达到2.5mm，粒径大于0.5mm的颗粒污泥占反应器生物总量的比率为57%。内循环（internal circulation, IC）厌氧反应器成熟的颗粒污泥粒径分布是第一反应室颗粒为2.0～2.5mm，第二反应室颗粒为0.8～1.5mm；粒径在1.74mm左右时比产甲烷活性（specific methanogenic activity, SMA）为1.95g CH4-COD·g volatile suspendedsolids （VSS）^-1·d^-1；粒径为1.0².5mm的颗粒污泥，比重为1.02¹.06，机械强度为1600³400Pa。粒径2.5mm左右成熟的厌氧颗粒污泥中紧密粘附的胞外聚合物（tightlybound-extracellular polymeric substances, TB-EPS）和松散附着的EPS（loosely bound-EPS,LB-EPS）分别是30.2mg total organic carbon（TOC）·g suspended solids （SS）^-1和15.2mgTOC·g SS^-1，是接种污泥的1.9倍和2.6倍。研究结果表明厌氧颗粒污泥规模化培养机理是微生物代谢产物EPS的内因和选择压的外因共同作用，EPS中的TB-EPS影响颗粒的大小，LB-EPS影响颗粒的粘结能力和结构强度。选择压影响生物量浓度、粒径分布和颗粒化率，同时也影响颗粒污泥比重、机械强度、沉降速度等性质。（2）通过灰色关联度分析，厌氧污泥颗粒化影响的重要程度：液体上升流速> HRT>OLR>进水COD。扫描电子显微镜（scanning electron microscope, SEM）、X射线能谱（energy dispersive spectrometer, EDS）和X射线衍射（X-ray diffraction, XRD）分析流失污泥和截留污泥，结果表明：液体上升流速使颗粒污泥受到颗粒碰撞摩擦力和水流剪切力作用，影响颗粒污泥的表面粗糙度、污泥形状和完整度；有机底物特性影响颗粒污泥的结构、组成和形成；底物的复杂性影响颗粒污泥的微生物多样性。（3）在不同OLR下，通过聚合酶链式反应和变性梯度凝胶电泳（polymerase chainreaction with denaturing gradient gel electrophoresis, PCR-DGGE）分析成熟的厌氧颗粒污泥中微生物种群结构的稳定性，结果表明：细菌种群结构的动态变化越来越显著，而古菌种群结构变化小。根据DGGE图谱计算细菌和古菌的Shannon-Wiene指数（H’）、Margalef丰富度指数和优势度指数S，表明多样性指数越高，生态优势度越小，丰富度越低。低负荷下优势古菌为甲烷八叠球菌（Methanosarcina sp.），高负荷下优势古菌为甲烷鬃菌属（Methanosaeta sp.）。（4）无机阳离子Ca²⁺浓度在300mg L^-1时，与对照组相比，厌氧颗粒污泥中EPS浓度和EPS中蛋白质含量达到最大值，增加了31.8%和53.1%，多糖含量下降了11.0%，蛋白质/多糖的比率从2.0增加到3.5，Zeta电位从-45mV增加到-30mV，颗粒污泥粒径从2.5mm增加到2.75mm，产甲烷速率提高了5.0%，颗粒污泥的脱氢酶活性、BAA-蛋白质水解酶活性、β-葡萄糖苷酶活性、碱性磷酸酶活性、辅酶F420活性分别提高了36.4%、28%、11.2%、29.8%和28.0%。PAM浓度为20mg L^-1时，与对照组相比，EPS及其含有的蛋白质、多糖和Zeta电位达到最大值，分别增加了54.0%、82.0%、3.9%和41.3%，蛋白质/多糖的比率从2.1增加到4.2，Zeta电位从-46mV增加到-27mV，颗粒污泥粒径从2.5mm增加到3.0mm，产甲烷速率提高了15.1%，颗粒污泥的上述五种酶分别提高了120%、40%、29.4%、40.4%和166.7%。研究表明Ca²⁺和PAM对厌氧污泥颗粒化起到生物强化作用。（5）外源厌氧颗粒污泥（R1反应器）历时12周完成启动，OLR为7.4kg COD·m^-3·d^-1、出水COD约为450mg·L^-1和COD去除率为80%。剩余活性污泥（R2反应器）历时30周颗粒化完成，OLR为8.8kg COD·m^-3·d^-1左右、出水COD约为240mg·L^-1和COD去除率为90%。在R1和R2中，挥发性脂肪酸（volatile fatty acids, VFA）分别是200mg·L^-1和100mg·L^-1左右（以乙酸计），VFA/碱度的比率分别为0.3和0.15，甲烷浓度分别是77.2%和78.6%，R1中EPS浓度从36.4mg·g SS^-1增加到36.8mg·g SS^-1，R2中EPS浓度从22.5mg·g SS^-1增加到46.1mg·g SS^-1。R1大部分颗粒为1.5².5mm，粒径分布单一，而R2中厌氧颗粒化表现出不同规格0.0⁰.3mm、0.3⁰.5mm、0.5¹.5mm和1.5².5mm的颗粒，颗粒粒径分布差异性缩小。通过计算和比较Grau second-order动力学模型和修正的Stover–Kincannon动力学模型，确定剩余活性污泥可以做接种污泥代替厌氧颗粒污泥来启动厌氧反应器，并且能够取得较高的运行效能和实现厌氧污泥颗粒化，从而建立一种高活性厌氧颗粒污泥的规模化培养系统。更多还原

【Abstract】 In full-scale anaerobic wastewater treatment, the operational stability and highperformance of an anaerobic reactor depends largely on the efficient settling properties andhigh methanogenic activity of anaerobic granular sludge. If loose sludge floc exists in theanaerobic reactor, the reactor becomes prone to sludge washout, low organic loading rate, andpoor performance. Mature granular sludge merchandise is scarce and expensive, grows slowly,involves difficult training, and has high transportation cost. Therefore, a study on full-scalecultivation of anaerobic granular sludge can solve engineering problems as well as providepractical knowledge and theoretical value.In this dissertation, a full-scale anaerobic reactor was inoculated with residual activatedsludge to investigate the following: sludge granulation process, factors influencing anaerobicgranulation by grey relational analysis, microbial population dynamics by molecularbiotechnology, enhanced granulation of anaerobic granular sludge by Ca²⁺andpolyacrylamide （PAM）, and comparison of the kinetic modeling of identical full-scale reactorswith residual activated sludge and anaerobic granular sludge for brewery wastewatertreatment. These concepts demonstrated the full-scale cultivation process of anaerobicgranular sludge and its mechanisms. The main results were described as follows:（1） Anaerobic sludge granulation with residual activated sludge could accomplishfull-scale cultivation for brewery wastewater treatment by a continuously average organicloading rate （OLR） of8.8kg chemical oxygen demand （COD）·m^-3·d^-1, upflow velocity of5.4m·h^-1, and hydraulic retention time （HRT） of5.9h. After granulation, the bioparticles largerthan0.5mm accounted for57%of all bioparticles. Mature particle size in the internalcirculation reactor was between2.0mm and2.5mm in the first reaction, whereas that in thesecond reaction chamber was between0.8mm and1.5mm. When the diameter size was1.74mm, the specific methanogenic activity could reach the maximum level which was1.95gCH4-COD·g volatile suspended solids （VSS）^-1·d^-1. The specific gravity was between1.02and1.06and the mechanical strength was between1600Pa and3400Pa in the diameter size of1.0mm to2.5mm. The tightly bound-extracellular polymeric substances （TB-EPS） andloosely bound-EPS （LB-EPS） were30.2mg total organic carbon （TOC）·g SS^-1and15.2mgTOC·g suspended solids （SS）^-1in the diameter size of2.5mm. The results showed that sludgegranulation mechanism could be attributed to EPS and selection pressure in full-scalecultivation of anaerobic granular sludge. TB-EPS affected the particle size and LB-EPSaffected the particles capability and structural strength. Selection pressure affected thebiomass concentration, particle size distribution, and the bioparticle rate, but also affected theproportion of the specific gravity, mechanical strength, sedimentation and other properties.（2） The grey relational analysis results showed that the factors that influenced anaerobicgranulation with significant effects on anaerobic granulation were as follows: liquid upflowvelocity> hydraulic retention time （HRT）> organic loading rate （OLR）> influent COD. The sludge in the effluent and in the reactor influenced by liquid upflow velocity was examinedusing a scanning electron microscope, energy dispersive spectrometer, and X-ray diffractionanalysis. The results revealed that the liquid upflow velocity affected the surface roughness,sludge shape, and integrity of the granular sludge through the collision friction andhydrodynamic shear force. The characteristics of the organic substrate could influence theformation, composition, and structure of anaerobic granules. The complexity of the substratecould also exert selection pressure on microbial diversity in anaerobic granules, which wouldconsequently influence the formation and microstructure of granules.（3） The main archaeal species in the sludge samples at different OLRs variedsignificantly compared with bacteria species using the polymerase chain reaction withdenaturing gradient gel electrophoresis （DGGE）. The Shannon–Wiener index （H’）, Margalefrichness index, and dominance index S of bacteria and archaea calculated by DGGE profilesshowed that a higher diversity index resulted in less ecological dominance and reducedrichness. The dominant archaeal species in the treated sludge at low OLRs wasMethanosarcina, whereas that at high OLRs was Methanosaeta.（4） Compared with the control group, the contents of EPS and proteins could reach themaximal values and respectively increased by31.8%and53.1%when the Ca²⁺concentrationwas300mg L^-1. However, polysaccharides content decreased by11.0%, the ratio of proteinand polysaccharide increased from2.0to3.5, and Zeta potential increased from-45mV to-30mV. The above variations enhanced granular sludge particle size from2.5mm to2.75mm,and methanogenic rate increased by5.0%. Compared with the control group, thedehydrogenase, BAA-protein hydrolase, β-glucosidase, alkaline phosphatase, and coenzymeF420activities increased by36.4%,28%,11.2%,29.8%, and28.0%respectively. Comparedwith the control group, the contents of EPS, proteins, polysaccharides, and Zeta potentialcould reach the maximal values and respectively increased by54.0%,82.0%,3.9%, and41.3%when the PAM concentration was20mg L^-1. The ratio of protein and polysaccharideincreased from2.0to4.2, and Zeta potential increased from-46mV to-27mV. The abovevariations enhanced granular sludge particle size from2.5mm to3.0mm, and methanogenicrate increased by15.1%. The above enzyme activities increased by120%,40%,29.4%,40.4%, and166.7.0%, respectively, compared with the control group. Anaerobic sludgegranulation enhanced by Ca2+and PAM.（5） Two identical full-scale biogas-lift reactors treating brewery wastewater wereinoculated with different types of sludge. One reactor （R1） started up with anaerobic granularsludge in12weeks and obtained a continuously average OLR of7.4kg COD·m^-3·d^-1, effluentCOD of approximately450mg·L^-1, and COD removal efficiency of80%. The other reactor（R2） started up with residual activated sludge in30weeks and granulation was accomplishedwhen the reactor reached an average OLR of8.8kg COD·m^-3·d^-1, effluent COD ofapproximately240mg·L^-1, and COD removal efficiency of90%. The difference between thevolatile fatty acids （VFA） concentrations of R1and R2were significant, which were approximately200mg·L^-1and100mg·L^-1as acetic acid, respectively. The VFA/alkalinityratios of R1and R2were approximately0.3and0.15, respectively. Methane contents in thebiogas of R1and R2were about77.2%and78.6%, respectively. The result suggested that thedominant methanogens in R2were able to maintain a higher level of methanogenesis becauseof achieved anaerobic granulation sludge. EPS production increased slightly from36.4mg·gSS^-1to36.8mg·g SS^-1in R1, whereas that increased from22.5mg·g SS^-1to46.1mg·g SS^-1inR2. The increased amount of EPS indicated that EPS could help in granule formation. A singleparticle size of R1was between1.5mm and2.5mm, whereas different particle sizedistribution of R2was0.0⁰.3mm,0.3⁰.5mm,0.5¹.5mm, and1.5².5mm. Differences insludge characteristics, methane content, VFA, and VFA/alkalinity ratio may be accounted forthe superior efficiency of the treatment performance of R2over R1. Grau second-order andModified Stover–Kincannon models were successfully used to develop kinetic parameters ofthe experimental data with high correlation coefficients. These findings demonstrated that theresidual activated sludge could be used effectively instead of anaerobic granular sludge toachieve higher performance and sludge granulation. These results indicated that the highactivity of anaerobic granular sludge full-scale cultivation system could be established.更多还原