5-氟尿嘧啶-β-环糊精结肠靶向前体药物的研究

5-氟尿嘧啶(5-Fluorouracil， 5-FU)是一种抗代谢药物，广泛用于临床治疗结直肠癌、胃癌、乳腺癌等多种癌症，但其首过代谢显著、亲脂性较低，选择性差、毒副作用大。为克服这些缺点人们对5-FU进行了大量的修饰工作，包括小分子修饰以及与各种载体形成微球、微囊、纳米粒、共价前药等。 环糊精(Cyclodextrin，简称CD)，可被结肠中的糖苷酶特异性地降解成小分子糖，而胃和小肠中由于缺乏相应的酶而使环糊精不被降解，这一特性在结肠药物的靶向输送及释放中有重要应用价值。环糊精中含有丰富的羟基，易进行化学修饰，将药物与环糊精通过共价键结合制成前药，使其在胃和小肠中不降解，而在盲结肠中被特异性的酶降解释出药物，达到结肠靶向释药的目的。研究表明，环糊精作为一种前药载体为结肠靶向释药和缓释、控释系统提供了一种有效的手段。 本工作选择5-氟尿嘧啶为模型药物、β-环糊精作为载体，通过中间体5-FU羧酸衍生物的制备及其与β-环糊精的偶联，合成了系列5-FU-β-CD前体药物，并利用紫外、红外、质谱、核磁、元素分析、热分析等手段对其进行结构表征。同时，还研究了前体药物的体外释药性质。具体内容包括： 1. 含有羧基的5-FU衍生物中间体的合成：(5-氟尿嘧啶-1-基)-乙酸(FUAC)、3-(5-氟尿嘧啶-1-基)-丙酸(FUPC)、5-(5-氟尿嘧啶-1-基)-戊酸(FUVC)的合成。 2. 中间体5-FU的羧酸衍生物与β-CD的偶联：分别通过以6-OTs-β-CD为中间体的取代法和活化酯法，合成了第一面取代和第二面取代的5-FU-β-CD大分子前体药物。在二面取代的前体药物制备中，通过改变原料的比例，合成了系列不同取代度(DS)的2-[(5-氟尿嘧啶-1-基)-乙酰基] -β-环糊精结合物。 3. 对上述前体药物进行体外释放研究：分别考察了前体药物在不同pH缓冲溶液中的水解行为及其在小鼠胃肠道人工体液中的酶解行为，并通过UV-Vis及HPLC对前体药物释放情况进行检测分析。 5-Fluorouracil(5-Fu), commonly known as a broad-spectrum antineoplastic drug, has been widely used in the treatment of various kinds of cancer including colon cancer for 40 years. However, this antitumor agent exhibits serious adverse effects, such as their marrow toxicity, gastrointestinal reaction and low selectivity in their clinical use. In order to improve its antitumor activity and reduce its toxicity, the compound was modified in various ways, including the formation of conjugated prodrugs with kinds of carrier, microsphere and nanoparticles etc. Cyclodextrins(CDs) are known to be barely capable of being hydrolyzed and only slightly absorbed in passing through the stomach and small intestine; however they are fermented into small saccharides by colonic microflora and thus absorbed as small saccharides in the large intestine. This biodegradation property of CDs may be useful as a colon-targeting carrier, and thus CD prodrugs may serve as a source of site-specific delivery of drugs to colon. It was demonstrated that prodrugs of CDs can provide a versatile means for construction of not only colon targeted delivery systems, but also delayed release systems. 5-Fluorouracil was taken as a model drug and β-CD as the carrier in this study. Series prodrugs of 5-FU was prepared through the preparation of reactive 5-FU derivatives containing carboxyl group and coupling to hydroxyl groups of CD. The structures of the conjugates were charactered by using IR, UV–vis, ESI-MS, 1H, 13C-NMR spectra, elemental analyses, and thermal analysis. In vitro hydrolysis behavior in aqueous solution and in rat gastrointestinal tract contents of the conjugates were also investigated. The main content of this dissertation includes following aspects: 1. The preparation of 5-FU derivatives containing carboxyl group: 5-Fluorouracil- acetic acid(FUAC)、3-(5-FU-1)-propionic acid (FUPC)、and 5-(5-FU-1)-valeric acid(FUVC). 2. The coupling of 5-FU derivatives to β-CD: 5-FU was selectively conjugated onto the primary or secondary hydroxyl groups of β-CD through an ester linkage, by the substitution of 6-OTs-β-CD and the activated ester method respectively. For the secondary face conjugation, the degree of substitution(DS) can be controlled by changing the mole ratio of the starting materials(FUAC and β-CD). 3. In vitro release behavior of the conjugates in aqueous solution and in rat gastro- intestinal tract contents of the conjugates were investigated, and the reaction was monitored and analyzed by using UV-Vis and HPLC methods.

若尔盖高原花湖湖滨湿地甲烷排放研究

若尔盖高原湿地位于青藏高原东北部地区，平均海拔3,400-3,600m，是长江和黄河的自然分水区，区内发育了大面积的草本沼泽以及高寒沼泽化草甸、高寒湖泊。由于它所处的位置海拔高、气候波动较大，并处于我国三大自然区的交错过渡带，因而被认为是我国最为典型的脆弱湿地生态系统之一。由于地处偏远、自然环境条件恶劣等多方面的原因，针对若尔盖湿地的科学研究资料一直以来还非常缺乏。本文对国内外近年来在湿地生态系统甲烷排放过程、研究方法，以及关于湿地生态系统甲烷排放的影响因素进行了综述，并采用静态箱－气相色谱法，从湿地环境格局、湿地甲烷排放等方面，对若尔盖高原典型高寒湖泊湖滨不同类型湿地甲烷排放特征进行了研究，并进一步探讨了影响若尔盖高原高寒湖泊湖滨带甲烷排放的因素。得到如下结果：1．若尔盖高原花湖湖滨湿地在植物生长季（6 至8 月），甲烷排放平均速率为0.315 mg·m-2·h-1；不同月份间甲烷排放速率存在差异，分别为：-0.054、0.471、0.493 mg·m-2·h-1。不同类型湿地甲烷排放速率亦表现出差异，两栖蓼（Polygonum amphibium）湿地、滩涂和藏嵩草（Kobresia tibetica）草甸甲烷排放速率分别为：0.464、0.477、0.005mg·m-2·h-1。2．若尔盖高原花湖湖滨湿地甲烷排放速率与土壤10cm 温度显著相关。土壤温度是影响若尔盖高原花湖湖滨不同类型湿地甲烷排放的重要因素之一。随着土壤温度的升高，土壤微生物活性增强，使土壤中的氧消耗加快，氧化还原电位下降，有利于产甲烷菌的生长，从而增加土壤的甲烷产生量。3．地表水位与若尔盖高原花湖湖滨湿地甲烷排放速率相关性不显著。地表水覆盖，使得湿地土壤缺氧状况得到加强，增强了土壤中产甲烷菌的活性，促进甲烷形成，再通过植物、气泡或扩散的形式释放出土壤。但水层的加深，也使土壤中已产生的甲烷在通过气泡或扩散形式穿越水层时，被氧化的量增加，从而减少了甲烷向大气中的排放。4．植被高度以及植被地上生物量与若尔盖高原花湖湖滨带甲烷排放速率的相关性不显著。植物主要通过凋落物以及根系分泌物的输入为产甲烷菌提供基质，并作为土壤与大气之间的甲烷气体交换的传输途径；与其他环境因素共同影响湿地生态系统甲烷排放。The Zoige wetland on the eastern fringe of Qinghai-Tibetan Plateau, with averagealtitude between 3,400 and 3,600m, is the watershed of Yangtze River and YellowRiver. There are large area of peatland, subalpine meadow and lakes in this region.Due to its high elevation, transitional topology and high fluctuation of climate, theZoige wetlands represent one of the most fragile wetland ecosystems in China. And asa result of remote location and harsh environment conditions, the researches on theZoige wetland are relatively rare, especially the researches on the methane emissionfrom littoral zone of alpine lakes. Variations of methane emission rates as measuredby the method of static chamber – gas chromatography (GC) were detected fromlittoral zone of alpine lake on the Zoige Plateau. Relationships between methaneemission rates and environmental factors were analyzed. It is concluded that:1．The average methane emission rate in the littoral zone of Huahu Lake, ZoigePlateau is 0.315 mg·m-2·h-1, with evident spatial and temporal variations. The littoralzone has different methane effluxes with -0.054, 0.471, and 0.493 mg·CH4·m-2·h-1in June, July and August respectively. Different types of wetland have differentmethane emission rates, with value of 0.464, 0.477, and 0.005 mg·CH4·m-2·h-1 forPolygonum amphibium wetland ( PA ), Shoal ( S ) and Kobresi tibetica meadow ( KT ), respectively.2． The soil temperature at 10cm is significantly correlated with the methane effluxesin littoral zone of Huahu Lake, Zoige Plateau, and which is one of the most important factors influencing the methane emission from this region. The activities of soilmicroorganisms rise under higher soil temperature and increases oxygen consumptionand decreases Eh, which is in favor of the methanogensis, and enhances theproduction of methane in soil.3． The correlation between the standing water and methane effluxes from littoralzone of Huahu Lake is not significant. Because of the standing water, the anaerobicconditions of wetland soil have been enhanced, and are favor to the decomposition oforganic matter. And the anaerobic conditions strengthen the methanogensis’ activities,thus the methane production, which release to the atmosphere by diffusion, ebullitionand aerenchymal plants. With the water level’s increase, more methane produced insoil which is transferred by ebullitions or diffusion are oxidated, thus reduce themethane release to the atmosphere.4． The height and aboveground biomass of vegetation are not significant related tothe methane effluxes from littoral zone of Huahu Lake, Zoige Plateau. The vegetationprovides substrates for methanogensis by litter and root exudates; act as thetransportation way of methane between soil and atmosphere; influence the methaneemission of wetland ecosystems with other environment factors.

厌氧复合菌系的筛选及含纤维素原料预处理复合菌剂的研究

本文从不同厌氧生境中获得7组(C-2、Y-2、L-2 、NZ、H-3、CZ、L-3)具有纤维素降解能力的复合菌系。经过不断传代、淘汰纤维素降解能力降低的菌系，最后得到一组高效、传代稳定的厌氧纤维素分解复合菌系L-3。该菌系可使滤纸在42 h内溃烂，并能在分解纤维素的同时产氢气。对L-3复合菌系的产酶条件进行了研究，结果表明，在实验范围内该菌系的产酶最适条件为：pH 6.5，温度37 ℃，接种量5 %，最佳碳源为滤纸，最佳氮源为硫酸铵。第10天测得羧甲基纤维素酶(CMCase)、滤纸酶(FPA)、外切葡聚糖酶(C1)、β-葡聚糖苷酶(β-glucodase)的酶活分别为0.216 U/ml、0.101 U/ml、0.132 U/ml、0.002 U/ml，滤纸失重率70.6 %。发酵代谢产物乙醇和丁酸含量分别可达1378 mg/L 、2695 mg/L，发酵产生的气体中氢气含量最高可达70.2 %。DGGE结果表明该菌系主要由14种菌组成，其中有三株菌在发酵前后菌数发生了明显的变化，说明在以滤纸为底物的降解过程中，这三株菌起到了重要作用，对这三株菌进行了分子生物学鉴定，初步定为Clostridium phytofermentans、Clostridium cellulovorans、Desulfovibrio sp。 利用实验室分离得到的纤维素降解菌，最终配制出由10、X-1、X-13、ST-13、L-3组成的好氧-厌氧纤维素降解复合菌剂。以秸秆为发酵底物，菌剂接种量1%，利用复合菌剂预处理后的秸秆，发酵总产气量相对于对照提高了71.62%，甲烷含量最高可达70.08%。 A group of microbial consortia L-3 was isolated from the anaerobic fermentation residue of corn stalk, which could degrade cellulose and produce hydrogen. The CMCase, FPA, C1 and β-glucosidase activity of L-3 could reach to 0.216 U/ml, 0.101 U/ml, 0.132 U/ml and 0.002 U/ml, respectively. In the filter degrading process, the filter paper collapsed in the liquid culture within 42 h and the filter degrading rate could reach to 70.6% in the 13 days, meanwhile, hydrogen was determined and the highest hydrogen content was 70.2%. The optimum cellulase-degrading conditions were filter papaer as the carbon source, (NH4)2SO4 as the nitrogen source, 37 ℃ and pH 6.5 in this experiment. DGGE results showed that the microbial consortia L-3 mainly included 14 strains. The amount of 3 strains were changed during the fermentation. These strains were identified as Clostridium phytofermentans、Clostridium cellulovorans、Desulfovibrio sp by 16S rDNA sequence analysis. The cellulose- degrading microbial agent was composed by 10, X-1, X-13, ST-13, L-3 which were isolated in the laboratory. The straw pretreated by cellulose-degrading microbial agent was used to ferment, the total biogas production increased by 72% comparing to the control. The content of methane could reach to 70.08%。

无介体双室微生物燃料电池产电性能初步研究

本文对无介体双室微生物燃料电池的产电性能进行了初步研究，并根据不同运行阶段产电性能的优劣，对其中微生物的差异性进行了比较分析。全文分为两个部分： 第一部分：以乙酸钠为阳极原料构建双室微生物燃料电池（MFC），研究不同阴极受体、外接电阻、乙酸钠浓度和pH等因素对电池产电性能的影响，研究结果表明：在500mL的阴阳极反应体系中，选用乙酸钠作为阳极底物，质量浓度为6.46 g/L， pH 7.0，接入500Ω外电阻，阴极电子受体选择高锰酸钾的情况下，微生物燃料电池产电性能最好，最大电功率密度达到294.72 mW/m2，库伦效率能达到25.87%。在确定最适外接电阻阻值的同时对MFC内阻进行测定，阻值为871.87Ω。 第二部分：微生物燃料电池运行中，比较以厌氧污泥作为接种源的第一阶段和只接入附着有大量微生物电极的第二阶段的产电性能，得出第二阶段产电性能优于第一阶段，最大电功率密度达到353.57mW/m2，比第一阶段提高58.85 mW/m2；库伦效率为39.35%，增幅达52％左右；针对微生物燃料电池运行过程中，底物CH3COONa可能存在其它的代谢途径，本实验进行了第二阶段产电性能与CH3COONa消耗率关系以及阳极液面上方气体成分和含量的研究，发现第二阶段50h前CH3COONa的大量消耗主要用于微生物的生长，在整个运行过程中，阳极液面上方含有CH4和CO2；对气体测定的同时还发现，振荡能增强电功率密度的输出；通过对电极上和污泥中微生物差异性分析得出，δ-变形菌纲、β-变形菌纲和拟杆菌门的菌种更适应微生物燃料电池的运行环境，能在电极上大量富集，提高电池的产电性能，只接入附着有大量微生物的电极能有效降低热袍菌纲的菌种数量，降低了CH3COONa的无为消耗，有效提高了电池的库伦效率。 Electricity production in the mediator-less two-chambered microbial fuel cell(MFC) was researched. Based on the result in the different operation phase in the MFC, the microbial diversity was analysed. The paper involved two parts: Part 1: A two-chambered microbial fuel cell (MFC) was constructed with high-concentration sodium acetate as fuel in the anode. The influence of different electron acceptors in the cathode, external resistance value, pH value and concentration of sodium acetate on electricity generation in MFC was investigated. The result showed that the maximum power density of 294.72 mW/m2 and the coulombic efficiency of 25.87% was achieved at sodium acetate concentration of 6.46 g/L, pH 7.0, external resistance 500Ωin the anode and when using potassium permanganate as electron acceptor in the cathode. While decided the value of resistor, we found that shaking has effect on electricity production in the MFC. Part 2: Comparing the electricity production in different operation phases when using anaerobic sludge as inoculum in the first phase and microbes in the anodic electrode as inoculum in the second phase, the result showed that electricity production in the second phase was more than that in the first phase, the maximum power density of 353.57 mW/m2 and the coulombic efficiency of 39.35% was achieved, 58.85 mW/m2 and 52％ more than that in the first phase, respectively. According to the fact that CH3COONa might be metabolized in other pathway in the running process in the MFC, we determining the relationship between electricity production and CH3COONa consumption, and the gas content in the anode, we found that CH3COONa was mainly used for microbe growth before 50h, and the anode contained CH4 and CO2. At the same time, we found that shaking could improve power density. The analysis on diversity of microbe in the anodic electrode and anaerobic sludge showed that δ-proteobacterium, β-proteobacterium and Bacteroidetes adapted themselves to the running environment of MFC. The anode could enrich them to improve the electricity production while reduced the quantity of Thermotogales, which were obligately anaerobic organotrophs with a fermentative metabolism, to increase the coulombic efficiency effectively.

厌氧产氢菌的分离筛选、诱变选育及其产氢特性的研究

本文从四川绵竹酒厂、成都市龙泉长安垃圾填埋场以及四川大学荷花池底的厌氧污泥中先后分离得到63株厌氧产氢菌，其中H-8、H-61、HC-10等16株产氢细菌产氢能力较高，HC-10的产氢能力最高，最大产氢量和最大产氢速率分别达到2840 ml H2/L培养基和25.39 mmol H2/g drycell·h，对HC-10进行生理生化鉴定和分子生物学鉴定，判定其为clostridium sp.，对HC-10的产氢条件进行了研究，结果表明，该菌的最适生长温度为35 ℃，最适生长初始pH为7，以葡萄糖为最佳碳源，以蛋白胨为最佳氮源，不利用无机氮源，其产氢发酵液相产物以乙醇和乙酸为主，其发酵类型属于乙醇型发酵。此外，以酒糟废液作为底物，进行了菌株HC-10的生物强化试验，研究表明，投加了HC-10的强化系统其产氢量比对照高出40.32%。 同时为了获得厌氧产氢菌的高效突变株，分别以产氢菌H-8和H-61为原始菌株进行微波诱变处理，对微波诱变参数进行了优化，考察了突变株的遗传稳定性、产氢特性及耐酸性。菌株H-8经过微波诱变得到5株高产氢突变株HW7、HW33、HW181、HW184、HW195，经多次传代表明HW195是稳定的高产突变株。突变株HW195具有较好的耐酸性，在pH值为2.8时仍能生长。通过间歇发酵实验，其最大产氢量和最大产氢速率分别达到2460 mL/L培养基和27.97 mmol H2/g drycell·h，比原始菌分别提高了50.75%和41.7%。菌株H-61经过微波诱变后选育得到的突变株HW-18，其最大产氢量和最大产氢速率分别达到2190 mL/L培养基和25.86 mmol H2/g drycell·h，比原始菌分别提高了23.03%和31.00%。 为了对比各种诱变方式对产氢菌产氢能力的影响，以厌氧产氢菌H-61为原始菌株，先后经亚硝基胍(NTG)、紫外(UV)诱变，选育得到1株高产突变株HCM-23。在葡萄糖浓度为10 g/L的条件下，其产氢量为3024 mL/L培养基，比原始菌株提高了69.89%；其最大产氢速率为33.19 mmol H2/g drycell·h，比原始菌株提高了68.14%。经过多次传代实验，稳定性良好。其发酵末端产物以乙醇和乙酸为主，属于典型乙醇型发酵。其最适产氢初始pH为6.5，最适生长温度为36 ℃，以蔗糖为最佳碳源。与原始菌株相比，突变株HCM-23的产氢特性发生了改变，如生长延滞期延长，可利用无机氮源等。 From anaerobic activated sludge, 16 strains of hydrogen producing bacteria were newly isolated. One of them named as HC-10 had the highest hydrogen producing capability, under the batch fermentative hydrogen production condition, the maximal hydrogen yield and hydrogen production rate was 2840 mL/L culture and 25.39 mmol H2/g drycell·h. It was identified as clostridium sp.HC-10 by 16S rDNA sequence analysis. Various parameters for hydrogen production, including substrates, initial pH and temperature, have been studied. The optimum condition for hydrogen producing of strain HC-10 were achieved as: initial pH 7.0, temperature 35 ℃, glucose as the favorite substrate, Moreover, using distiller's solubles wastewater as substrate, HC-10 strain was added in the biohydrogen producing system to research the bioaugmentation effection. The results showed that the hydrogen production of bioaugmentation system was 40.32% higher than the noaugmentation system. An anaerobic, hydrogen producing strain H-8 was irradiated by microwave to optimize the microwave mutagenesis condition, and to test the heredity, hydrogen-producing potential and aciduric of the mutants. An aciduric mutant named as HW195 with steady hydrogen-producing capability was obtained, which can grow at pH 2.8. Its capability of hydrogen production was tested in the batch culture experiments. The maximum hydrogen yield and hydrogen production rate was 2460 mL/L culture and 29.97 mmol H2/g drycell·h, which was 50.7% and 41.7% higher than those of the initial strain, respectively. When used the strain H-61 as original strain, a mutant named as HW18 was obtained. The maximum hydrogen yield and hydrogen production rate was 2190 mL/L culture and 25.86 mmol H2/g drycell·h, which was 23.03% and 31.00% higher than those of the initial strain, respectively. The results demonstrated that microwave mutagenesis could be used in the field of hydrogen producing microorganism. The hydrogen producing strain H-61 was used as an original strain which was induced by NTG and UV for increasing and the hydrogen production capability. One of the highest efficient H2-producing mutants was named as HCM-23 with its stable hydrogen production capability. which was tested in the batch culture experiments. With the condition of 10 g/L glucose, its cumulative hydrogen yield and hydrogen production rate was 3024 mL/L culture and 33.19 mmol H2/g drycell·h, 69.89％and 68.14％ higher than that of the original strain, respectively. The terminal liquid product compositions showed that the mutant HCM-23 fermentation was ethanol type, while the original strain H-61 fermentation was butyric acid type. Varieties of parameters of hydrogen production fermentation were studied, including time, carbon source, nitrogen source, glucose concentration, glucose utilization, initial pH and incubation temperature had been studied, indicated the optimum condition of hydrogen production for the mutant HCM-23 as initial pH6.5, temperature 36 ℃, and the favorite substrate was sucrose. The hydrogen production characters of the mutant and the original strain were different, such as, the growth lag phase and the utilization of inorganic nitrogen source, etc. This work shows a good application potential of NTG-UV combined mutation in the biohydrogen production. And the hydrogen production mechanism and metabolic pathway should be explored furthermore.

自养氨氧化混合种群特性和共存机理初步研究

组特殊自养氨氧化混合种群，表现：无机环境种群生长迅速、生物量高；在一个完全无机的自养生长环境中，不仅保持高氨氧化速率，并出现丰富的异养微生物种群；该种群置于异养、厌氧环境中，迅速表现出产氢特征。对于这样一个特殊的生态体系，研究其共生机理，以及联接这些种群之间的碳源和能源问题，将具有非常重要意义。我们拟从种群特征、细胞表面分泌产物、游离体系产物多糖、蛋白和脂肪酸方面开展研究。 第一部分，自养氨氧化混合种群的基本特征。采用氨氧化培养基，进行种群氨氧化特征研究；采用扫描电镜观察自养混合种群的微观特征；沉降、离心去除微生物种群，分析水相中的总有机碳、糖类等物质；利用LB培养基进行种群的分离、纯化，并采用DGGE手段对微生物种群结构进行分析。结果表明，接入菌种后（2/5000（V/V）），培养液中氨（200mg/L）在3－5天内快速降解；亚硝酸盐与氨氮变化呈负相关趋势，仅有少量硝酸盐含量（< 30mg/L）。氨氧化种群的生物量增长与氨氧化趋势一致，初始生物量7.75 mg/L(蛋白含量)，3-5天后生物量快速增长，并达到最高63.06 mg/L（蛋白含量）。电镜图片显示，种群外包裹一层粘液。离心除去菌体后，检测培养液总有机碳和糖的含量，同样表现出与生物量增长相似的特征，分别由初始的3.73、2.35 mg/L，3－5内天迅速增加，并分别达到最大值35.19、27.45 mg/L。经初步分离、纯化并对纯化菌株进行测序，获得了10株异养微生物分别为布鲁氏菌科苍白杆菌属、纤维单孢菌、类芽孢菌属、黄杆菌属、无色杆菌、鞘脂单胞菌、嗜麦芽寡养单胞菌、噬氢菌属、硫红球菌、假单胞菌；DGGE显示，约有20分条离带，我们对其中的两条优势条带进行切割回收测序，鉴定为欧洲亚硝化单胞菌（Nitrosomonas eur）。 第二部分：混合种群自养-异养菌共生的可能机制。在对微生物种群特征初步分析基础上，针对胞外糖类组分可能被微生物代谢分解，我们重点对微生物细胞蛋白质与糖类进行分析。采用超声结合RIPA裂解液裂解，SDS-PAGE电泳分析混合种群总蛋白种类，并通过氨基酸分析仪及红外光谱法分析氨基酸组成及蛋白红外特征。采用超声破碎结合反复冻融对细胞样品进行处理，提取液采用醇沉、Sevage脱氮白，凝胶过滤方法脱盐和分级分离。对提取物的糖分析包括：紫外扫描，红外光谱，核磁共振，单糖组成分析；扫描电镜观察菌群破裂现象。SDS-PAGE分析结果表明：氨氧化种群不同生长阶段都显示出42kD蛋白表达量很高，d4时42kD蛋白表达已经很强，4-7d内一直持续这种过量表达，直到d8后表达开始减弱。说明42kD蛋白可能与氨氧化密切相关。红外光谱分析显示：细胞提取物的特征峰分布在3427.42cm-1、1718.18 cm-1和1681.72 cm-1、1160.07和1086.74 cm-1，分别对应为OH、 C=O、C－O－C基团，表明具有蛋白的典型特征；氨基酸分析显示蛋白中的Gly,Asp,Ala,Glu含量相对较高。 提取物中胞外多糖分离谱图得到不均一组分，共得到6个收集峰；紫外扫描在201－213 nm处有多糖吸收峰，同样表明多糖成分不均一性；多糖红外光谱特征峰主要分别在3400.49 cm-1、2920.28 cm-1、1154.54和1087.52 cm-1，对应OH、－CH2－ or CH 、C－O－H or C－O－C等多糖特征基团；多糖提取物核磁共振1H d4.3～5.9之间出现强吸收峰，这是1H中，多糖存在的明显证据，1H NMR中，其中O-乙酰基的甲基上的氢信号为d1.1～1.3之间。糖肟全苯甲酸酯衍生物的HPLC测定中，得到单一的单糖峰，由于时间问题，还未进行更深入的试验；电镜图片显示，种群中的细胞有大量的破裂现象。 实验表明，自养氨氧化混合种群显示出快速的氨氧化速率，氨氧化过程生物量和有机质的增加明显。微生物种群包裹粘液层，并分离纯化出大量的异养菌；去除菌体后的游离培养液中存在有机质（包括多糖）说明无机自养生长体系中存在异养菌生长、繁殖的二次碳源；细胞提取物中蛋白条带数目多、种类丰富；细胞多糖提取物具有明显的多糖特征，以及单糖的存在。结合种群的显微特征和游离体系中的有机质的检测结果，我们认为，无机自养生长体系中，种群细胞生长过程中发生的破裂现象可能是导致大量的蛋白、多糖释放到游离胞外，并成为其他异养菌生长的碳源和氮源。这可能是自养体系中，大量异养菌共生的可能机制，至于是什么原因引起种群生长过程中产生的破裂现象，还有待下一步深入研究。 A group of mixed autotrophic ammonia oxidizing populations, having much biological characteristic tested by concerned personnel for pilot test: Performed rapid population growth and obtained high biomass in inorganic environment; Not only maintained a high rate of ammoxidation, promoted a wealth of heterotrophic microbial populations growth in a totally inorganic and autotrophic growth environment; Placed in heterotrophic and anaerobic environment,had the performance characteristics that could rapidly produce hydrogen.For such a special ecological system, Study its symbiotic mechanism and the connection between these populations of carbon and energy issues, will have a very important significance. We intended from the characteristics of the population, the secretion product of cell surface, free substance in the liquid medium like polysaccharide, protein and fatty acids carrying out research. Part I: The basic features of mixed autotrophic ammonia oxidizing populations . Use inorganic liquid medium, processed study for ammonia oxidation characteristics of the population; we used scanning electron microscopy to get micro-features of autotrophic ammonia oxidizing populations .The medium was carried out settlement and centrifugal then removed the microbial populations, after all of that we analysis the water phase for total organic carbon(TOC), carbohydrate and other substances; Solid ammonia oxidizing medium was adopted to separation and purification of population, DGGE means was for structure analysis of microbial population. The results showed that after the inoculum of bacteria (2 / 5000 (V / V)), ammonia in the culture medium (200 mg / L) was rapid degradation in 3-5 days; ammonia and nitrite have the negative correlation between changes in the trend, then only a small amount of nitrate content (<30mg / L). The biomass growth of ammoxidation population in line with the trend of ammonia oxidation, the initial volume of it was 7.75 mg / L (protein content), in 3-5 days upto 63.06 mg / L (protein content). Electron microscope image showed, the populations were wrapped in a layer of mucus, including the a large number ruptted micorbe , Centrifuge to remove bacteria, then detected the medium for total organic carbon and sugar content, result took on the same characteristics with biomass growth, that were from the initial 3.73、2.35 mg / L respectively, in 3-6 days achieved rapid increase in the maximum to 35.19、27.45 mg / L respectively. After initial separation、 purification ,then processed sequencing to strains purified and got the result that there were 10 heterotrophic microorganisms : Brucella Branch pale bacillus, Cellu lomonas, Bacillus species category, a Flavobacterium, colorless Bacteria, Aeromonas sheath fat, little support maltophilia Aeromonas, macrophages species hydrogen, sulphur-MI, Pseudomonas bacteria spores; DGGE display, there were 20 separation bands approximately. Part II: Mixed populations that autotrophic - heterotrophic bacteria symbiotic mechanism. On the basis of preliminary analysis of microbial population characteristics, aiming at extracellular carbohydrate components might be decomposition by microbial, we focused on microbial cell protein and carbohydrate analysis. Using ultrasound combined with RIPA lysis cracking the cells, SDS-PAGE electrophoresis analysis the total protein species of the population, and through the amino acid analyzer studied the compositions of amino acid and infrared spectroscopy analysis of a protein infrared characteristics. Using ultrasound combined with repeatedly freezing and thawing to treated the cell sample, then took the means that alcohol precipitation, deproteinization by Sevage, gel filtration aimed at desalination and grade separation to deal with the lysates . The extraction of sugar analysis included: UV scanning, IR, NMR, single-sugar composition analysis. SDS-PAGE analysis showed that: 42 kD protein expression was very high at different growth stages of mixed autotrophic ammonia oxidizing populations , on the fourth day, 42 kD protein expression had been very strong, 4-7d, it had continued this excessive expression, then started to weaken after 7 days. 42 kD protein that might be closely associated with ammonia oxidation. Infrared spectral analysis showed that: cell extracts with the characteristic that the peak distribution in 3427.42 cm-1、1718.18 cm-1 and 1681.72 cm-1、1160.07 cm-1 and 1086.74 cm-1 corresponding to OH、C = O、C-O-C Groups which had the typical characteristics of protein; and analysis showed that amino acids including Gly, Asp, Ala, Glu ,the content in the protein is relatively high. Exopolysaccharide in the extracts had the separation map that it was uneven, received a total of six collection peaks by the detection mode of phenol-sulphruic acid method ; ultraviolet scan in the 201-213 nm department had polysaccharide absorbing peak, the same ingredients that polysaccharide heterogeneity; infrared polysaccharide spectral characteristics of the main peak at 3400.49 cm-1, 2920.28 cm-1, 1154.54 and 1087.52 cm-1, corresponding OH,-CH2-or CH, C-O-H or C-O-C;and other characteristics of polysaccharide group; 1H NMR of polysaccharide extract appeared absorption peak between d4.3 ~5.9, which is the apparent evidence of polysaccharide, In 1H NMR, the hydrogen signal of one of O-acetyl was between 1.1 to 1.3. The determination of Sugar oxime whole benzoate derivatives by HPLC, there was a single-sugar peak, as a matter of time, yet more in-depth test. Summary: Mixed autotrophic ammonia oxidizing populations show us that it had the ability in ammonia oxidizing and it was great, organic matter and biomass increased significantly in the process of ammonia oxidation. Microbial populations was wrapped up slime layer, the phenomenon of cell breakdown obviously, and there were a lot of separation and purification of the heterotrophic bacteria; a lot of organic matter (including polysaccharides)remined in the medium that removal of cell indicated the inorganic system existed secondary carbon sources that could be used by the heterotrophic bacteria ; there were a large number proteins bands of cell extract, rich variety; cell extracts of polysaccharide had obvious characteristics of polysaccharide, and the existence evidence of single-sugar. Combined population of microscopic characteristics and free of organic matter in the test results, we believe that the health of inorganic system, population growth occurred in the course of the breakdown of the phenomenon is likely to lead to a lot of protein and polysaccharide released into the extracellular free, And other heterotrophic bacteria use them to the growth as carbon and nitrogen. This may be autotrophic system, the large number of heterotrophic bacteria symbiotic mechanism.

亚硝化—厌氧氨氧化处理猪场废水研究

猪场废水COD浓度高、氨氮浓度高、悬浮物浓度高，已成为农村面源污染的主要来源，并严重威胁到农村饮用水安全。猪场废水氨氮浓度高、处理难度大，如何采用经济高效的方法，去除氨氮使其达到排放标准，一直是猪场废水处理中面临的重要难题。 厌氧氨氧化是近年受到国内外水处理研究者广泛关注的新型生物脱氮技术，具有不需要外加有机碳源、节省供氧量、降低能耗等优点。虽然国内外研究者对厌氧氨氧化过程的脱氮机理、厌氧氨氧化菌的生理生化特性等进行了多方面的研究，但已有的报道大多以模拟废水为研究对象，以猪场废水为研究对象的报道，在国内外文献中极少有报导。 本论文以猪场废水为主要研究对象，考察了猪场废水的亚硝化过程、厌氧氨氧化的启动过程，并对亚硝化和厌氧氨氧化联合用于猪场废水脱氮进行了探索。 1．论文首先研究了猪场废水的亚硝化过程，考察了废水水质和主要运行条件对亚硝化过程的影响。实验表明：（1）亚硝化阶段反应时间为8到10h时，出水中氨氮和亚硝酸盐浓度比可达到1：1～1：1.23，满足厌氧氨氧化反应对二者比例的要求；达到前述要求时，氨氮去除率达到58.3～65.6 %，亚硝化率在整个过程均保持在97 %以上，COD去除率在59.2～68.6 %；（2）曝气量（溶解氧）对亚硝化过程影响显著，随着曝气量增大，达到厌氧氨氧化要求的氨氮与亚硝酸盐氮浓度比例所需水力停留时间τ越短，pH出现明显下降的时间越短；（3）τ对应的pH在7.8～8.1之间，无需进行pH调节即可满足厌氧氨氧化反应对pH的要求；（4）氨氮和COD降解过程遵循一级反应动力学，氨氮和COD降解的速率常数分别为0.0656～0.0724 1/h和0.0491～0.0664 1/h。 2．在进行亚硝化过程研究的同时，以模拟废水为试验对象，进行厌氧氨氧化启动研究。以反硝化污泥和养殖厂储水池厌氧底泥的混合污泥作为接种污泥，历时大约100天，培育出具有厌氧氨氧化活性的污泥，氨氮和亚硝酸盐氮最高进水浓度分别为223.8 mg/L和171.4 mg/L，去除率最高分别达48%和41.5%，此时二者消耗比例为1.33：1。 3．在猪场废水的亚硝化研究完成和厌氧氨氧化过程初步启动成功后，在模拟废水中逐步加入猪场废水的亚硝化处理出水，逐步实现亚硝化和厌氧氨氧化的组合。亚硝化出水添加到厌氧反应器后，厌氧氨氧化反应仍可继续进行，且去除效率逐步提高。研究发现添加的亚硝化出水中携带的亚硝化细菌在厌氧氨氧化菌膜外层生长并累积，增加了厌氧氨氧化反应基质的传质阻力，妨碍了厌氧氨氧化效率的提高。 4．亚硝化－厌氧氨氧化实际工程应用探索中，生物接触氧化池可在有效去除废水中的有机物的同时实现亚硝化，出水中氨氮和亚硝酸盐比例平均为1.10，可满足后续厌氧氨氧化的要求；在适宜的进水浓度和温度下，ABR池出现了厌氧氨氧化启动的迹象；研究同时发现，水质的波动和气温的变化是工程中影响厌氧氨氧化菌活性的重要因素。 论文的主要创新点在于：（1）以猪场废水为研究对象，以实现厌氧氨氧化为目标，对亚硝化过程进行了比较详细的考察，获得了亚硝化出水满足厌氧氨氧化要求的工艺条件，通过对其COD和氨氮降解过程的考察，得出亚硝化阶段COD降解和氨氮去除的动力学模型；（2）对亚硝化－厌氧氨氧化处理猪场废水进行了探索，确立了影响其污染物去除率稳定的重要因素。 论文的上述研究成果，为厌氧氨氧化技术的实用性研究提供理论依据。 Piggery wastewater, which is characterized by high concentration of COD、ammonium and suspend substance, has become a most important source of non-point source pollution and also severely threats drinking water security in rural area. How to discharge piggery wastewater with the ammonium concentration meeting standard by economical and effective method? This is the most urgent problem in piggery wastewater treatment. As a new biological nitrogen removal technology, Anammox process has been paid more and more attention by researchers all over the world. Anammox has advantages of no need of organic carbon addition, low oxygen consumption and energy consumption. Plenty of investigations have been carried out to the mechanism, physiological and biochemical characteristic of bacteria about Anammox. Most of researches focused on synthetic wastewater, there is rare report about its application in piggery wastewater. In this paper，experimental studies were performed to investigate Sharon process in treatment of piggery wastewater，the start up process of Annammox using synthetic wastewater were studied, the feasibility of applying Sharon-Anammox process in the nitrogen removal of piggery wastewater was evaluated. 1. Sharon process of piggery wastewater was firstly investigated to analyze the effects of water quality and main running parameters, which meet the NH4+-N to NO2--N ratio requirement of successive Anammox. Results showed: (1)During Sharon Process，after 8～10 hours’ reaction the NH4+-N to NO2--N ratio in effluent reached 1:1.0～1:1.23, when the removal percentage of NH4+-N was 58.3～65.6 %, a semi-nitration rate of above 97 % was achieved during the process; meanwhile 59.2～68.6 % of the COD was also removed. (2)The aeration rate（oxygen） had obvious effect on the hydraulic retention time(τ) which met the NH4+-N to NO2--N ratio requirement of Anammox. As aeration rate increased, the hydraulic retention time(τ) was shortened. (3) The pH corresponding to τ was between 7.8 and 8.1, thus it needed no artificial adjustment. (4) The reduction of ammonia and COD followed the first-order reaction kinetics. The velocity constants of ammonia and COD were 0.0656～0.0724 1/h and 0.0491～0.0664 1/h, respectively. 2. The startup of Anammox process using the artificial wastewater was performed simultaneously with Sharon. The aim was to investigate the running parameters of Anammox and make foundation for the combination stage. By using the mixture of denitrifying sludge and anaerobic sludge in tank of the breeding factory, sludge of Anammox activity was cultivated in UASB after 100 days. The removal percentage of NH4+-N and NO2-N were up to 48% and 41.5%, respectively, when the NH4+-N and NO2-N influent concentration were 223.8 mg/L and 171.4 mg/L, respectively, the NH4+-N and NO2-N removal rate was 1.33:1. 3. After investigation of Sharon and startup of Anammox, effluent of Sharon process was added into the synthetic wastewater to combine Sharon and Anammox step by step. It took some time after the addition of Sharon effluent that Anammox reaction continued and the removal rate kept increasing. It indicated that nitrifying bacteria were carried by the Sharon effluent cumulated in the outer layer of Anammox. This enhanced transfer resistance of Anammox reaction and the increasing removal rate was restrained. 4. In the bio-contact oxidation pond of practical project, Sharon process were carried out successfully and organic compounds were removed effectively. An average NO2-N/ NH4+-N rate of 1:1.0 was achieved in the effluent, which met the requirement of successive Anammox. Under condition of suitable influent concentration and temperature, there was evidence that Anammox could start up in ABR. The variety of wastewater and temperature had great affects on Anammox activity in practical engineering. Innovation of this paper: (1) The Sharon process for treating piggery wastewater was discussed in details. Technological parameters that met requirement of Anammox were obtained. The dynamic models of COD and ammonium removal in the process were educed. (2) Sharon-Ananmmox for treatment of piggery wastewater was investigated, and the primary influencing factors was studied. This paper could be a theoretical consult for research of Anammox utility.

厌氧好氧一体化生物反应器处理发酵废水的中试研究

一体化反应器由于投资少、占地小、管理运行方便等优点而备受青睐。但现有的一体化反应器大都适用于处理中低浓度废水，耐受负荷普遍偏低。本课题研制出新型高效的厌氧好氧一体化生物反应器，旨在通过反应器结构优化、高效微生物载体研制、配合高效微生物菌剂技术处理中高浓度有机废水，实现高效和低耗，降低设备造价，提高反应器运行稳定性。 首先开展了菌剂对废水的适配试验。采用15种不同的微生物菌剂，以葡萄糖配水、中药提取废水、啤酒废水、氨氮配水为基质，分别测定了微生物菌剂的耗氧速率和厌氧比产甲烷速率，以其为指标比较了各菌剂对废水的适配性。根据结果选择活性高的14＃、8＃、10＃菌剂，在试验室进行了菌剂对废水的连续处理试验，取得良好的处理效果，为菌剂在厌氧好氧一体化生物反应器的小试、中试中的应用奠定了基础。 经小试研究后，又对厌氧好氧一体化生物反应器进行了处理发酵废水的中试研究。试验结果表明，反应器启动快，系统有机负荷2.72 kgCODm-3d-1时整个反应器去除率保持在84.5％～93.19％，在30多天内一次启动成功。冲击负荷试验中，系统总有机负荷最高可达到8.88 kgCODm-3d-1，系统去除率稳定在88.10％～96.88%，说明反应器处理效率高，抗冲击能力强。稳定运行期间，COD去除率可达90％以上，各项指标都能达到国家排放标准。 此外，对反应器配套系统高效菌剂、高分子复合颗粒载体进行了研究。结果显示，菌剂与反应器适配良好，各功能区形成了丰富、高活性的微生物，厌氧区颗粒污泥TS高达83.9 gL-1，VS/TS为56.9％～57.4％，比产甲烷活性为280～350 mLCH4 gvss-1d-1；好氧区固定化微生物TS高达1.921 gL-1，VS/TS为94.02～94.30％。对载体性能的研究表明，此高分子复合颗粒载体密度适中，易于流化，不易流失；粗糙多空，易于挂膜；且无生物毒害作用，稳定安全，是一种优良的生物载体。反应器各功能区对废水的降解过程分析，说明了反应器、菌剂、载体适配良好，在其协同作用下，实现了污染物的高效降解。 The integrated reactors were popular because of their characteristics such as little investment, small occupation of land, convenient of manage and running etc. But the present integrated reactors were mostly applied for treating wastewater of low concentration, the load tolerance was generally on the low side. A new type integrated anaerobic-aerobic bio-reactor was developed, which was conducted to treating organic wastewater of middle or high concentration by optimization of reactor structure, development of efficient microbe carrier and adaptation of high active microbial blends, to achieve high efficiency and low consume, reduce equipment cost, enhance running stabilization of reactor. The adaptability test of microbial blends on different wastewater was carried on firstly. Oxygen consumption rate and anaerobic specific activity of methane producing of 15 different microbial blends were measured separately taking glucose artificial wastewater, Chinese herb extracting wastewater, brewery wastewater and ammonia nitrogen artificial wastewater as substrate, by which the adaptabilities of different microbial blends to wastewater were compared. According to the results high active microbial blends 14#, 8# and 10# were selected and used in the continuous treatment of wastewater in the laboratory and had obtained good effect, which had laid a foundation for application microbial blends to small scale test and pilot test of integrated anaerobic-aerobic bio-reactor. After the small scale test, the pilot test of the integrated anaerobic-aerobic bio-reactor treating fermentation wastewater was carried on. The test results showed fast initiation of the reactor. When system organic load reached 2.72 kgCODm-3d-1the COD removal rate of the reactor was stable between 84.5%～93.19% and it initiated successfully in more than 30 days at a time. In the load shock test the maximum organic load of system could reach to 8.88 kgCODm-3d-1 and the COD removal rate could be stable between 88.10%~96.88% which indicated that the reactor was efficient for treating wastewater and had strong resistance to shock load. At stable running period the COD removal rate of the reactor was over 90% and each index of wastewater could reach to the national discharge standards. In addition, the high active microbial blends and the macromolecule compound granule carrier, the matching system of the reactor was studied. It showed that the microbial blends adapted well to the reactor and abundant and high active microbes were formed in each functional field. The TS of granule sludge in anaerobic field was as high as 83.9 gL-1, the VS/TS was 56.9%~57.4%, the specific activity of methane producing was 280~350 mLCH4 gvss-1d-1. And the TS of immobilized biological granule was as high as 1.921 gL-1, the VS/TS was 94.02%~94.30%. Study on the carrier showed that the self-made macromolecular compound granule carrier was moderate of density, easy of fluidization, unease of running off, rough and porous, easy of films fixation, no bio-toxic, stable and safe, was a kind of superior carrier. Analysis of degradation process in each functional field confirmed the reactor, microbial blends and carriers were in good adaptation and wastewater was decontaminated by their cooperation.

泥生绿菌的分离鉴定及其在净化废水中作用的初步探讨

本文介绍了从厌氧间歇膨胀光合反应器内的活性污泥中分离并鉴定的泥生绿菌(Chlorobium limicola Nadson)S1,它属严格厌氧光能自养型细菌，在有硫化物和少量碳酸氢盐存在下，有广泛利用有机物的能力，它的最适生长温度为28-30℃,最适生长PH为6.5-7.0，且含有氢化酶。因此，它能与甲烷发酵菌共存而共同作用，达到废水净化之目的。通过光照(2#反应器)和黑暗(1#反应器)对比实验，表明了在光照条件下即有泥生绿菌S1存在下，反应系统能更好地降低CODcr、BOD5 和提高CH4 含量，在四个负荷段的运行中，2#反应器在后三个负荷段的甲烷含量能稳定在91.6%而1#反应器为87%，2#反应器的二氧化碳含量为4.5%而1#反应器为8.8%，于28.35g/l.d的负荷下，2#反应器CODcr去除率达83.4%，BOD去除率达74.53%，分别较1#反应器高10.8%,6.4%。COD去除率提高了14%，BOD去除率提高了9.3%。本试验的试验条件为：白天自然光照，晚上电源光照，光照强度为1000-2500lux，通过连续动态运转，并以恒定的流速将废液注入反应器中，进水PH控制在6.5-7.2，反应器厌氧，恒温室温度控制在30±1℃。为使整个试验中同一水质条件下进行，进水采用化学合成培养基。This paper reports a Chlorobium Liwicola S1's isolation and identification. It is a strictly anaerobic and photosynthetic autotrophic bacterium. Along with sulfidedepondent CO2 assiwilaton,a few simple organic compounds can be photoassimilated. Acetate is most effectively used. Its best conditons of growth are 28-30℃,PH 6.5-7.0, and it contains hydrogenase. So it can live with methanefermentative bacteria in order to treat wastewater. At the same time, the treatment of wastwater using Chlorobium Limicola S1 with methane-fermontative bacteria under dark anaerobic and light anaerobic conditions is studied. In contrast with 1# reactor-darken, 2# reactor-illuminated can lossen wastewater's CODcr, BOD5 and on hance CH4 content better. In the test, 2# reactor's CH4 content is stable at 91.6%, but 1# reactor's is 87%. The CO2 content of 2# reactor is 4.5%, but 1# reactor's is 8.8%. When the load of teatment is 28.35g/l.d, the COD removal effficiency is 83.4% and the BOD removal efficiency is 74.53% in 2# reactor. They are separately 10.8%, 6.4% higher than 1# reactor's.

碱法草浆造纸蒸煮黑液厌氧生物降解性及其主要微生物类群研究

造纸行业是造成我国水环境有机污染物的重要污染源之一，其水污染的特点是小厂多、草浆多、工艺落后、污染扩散面广、造成废 水排放量大，每年排放的废水量约39亿立方米，占全国工业废水排放量的1/6，其中有机污染物（以BOD5计）160万吨左右，约占全 国工业废水中有机污染物总量的1/4。尤以占全国制浆造纸行业90%以上的碱法草浆造纸厂的蒸煮黑液量大面广，除含有机物外，还 含有木质素、残碱、硫化物、氯化物等污染物，属于PH值高、色度深、难于治理的高浓度有机废水，对水体污染特别严重，各地要 求治理呼声很高，急待研究并尽快找出各种有效的治理途径。对于碱法草浆蒸煮，黑液高浓度废水的治理，有各种方法，根据国内 的研究进展和我们已有试验工作表明，最经济有效，具有实用价值，在生产上可获得成功是厌氧处理法。近10多年来，国外关于高 效厌氧处理技术研究进展迅速，并出现了多种多样的工艺设备，如高效厌氧生物反应器，并在实用化方面取得了很大成绩，建立了 生产性装置，达到了高负荷运行，效果良好。本试验是根据我们已有研究基础，针对我国国情，对小型制浆造纸厂水污染防治除了 开发碱回收及各种综合利用技术外，要特别加强废水(废液)实用技术研究的指导思想，本试验采用改进型的上流式厌氧污泥床反应 器，设计了两种试验方案，通过试验结果如下。1. 试验方案I—碱法草浆黑液酸化和厌氧发酵I号UASB反应器动态模型试验结果表 明：(1). 采用中温35℃±1℃高效厌氧反应器USAB内装有填料(陶粒)和三相分离器，具有保持高浓度生物量和防止污泥流失的特点 ，污泥浓度Vs 可达30%以上，因而具有高效、节能、产能、滞留期短的优点，当进水CODcr在7500-10000mg/l，HRT由7天缩短到3天 ，有机容积负荷在1.22gCODcr/l·d-3.43gCODcr/l·d时，CODcr平均去除率可达55%-45.5%，最高CODcr去除率可达60.2-63.5%， BOD5去除率可达75.9-83.2%,沼气容积产气率可达0.29-0.67l/l·d，每克CODcr转化为沼气产率达0.39-0.48l/gCODcr·d，CH4含量 65.8-75.5%。厌氧发酵出水再用化学法进行后处理脱除难降解的木质素，CODcr总去除率达80%以上。(2). 动态试验结果表明：采 用酸化—厌氧发酵处理黑液工艺合理，技术路线可行。2. 试验方案II—黑液用化学法(Hcl)去除木质素进行厌氧发酵，II号UASB反 应器动态模型试验结果表明：(1). 采用中温35℃±1℃高效厌氧反应器UASB(内有软填料)，当进水CODcr7000-13000mg/l左右，HRT 由6天缩短到1天，有机负荷由0.98gCODcr/l·d增加到11gCODcr/l·d时，COD平均去除率均可稳定在70-77%，BOD5去除率为87.3- 93.1%，沼气容积产气率0.21-2.6l/l·d，每克CODcr转化为沼气产率为0.39-0.48l/gCODcr·d，高的可达0.53l/gCODcr·d，转化 率较高，CH4含量63-70%。(2). 试验证明碱法草浆黑液物化预处理—厌氧发酵处理的技术路线也是可行的，工艺合理、效果较好。 在有条件的工厂可采用。3.厌氧发酵阶段几大类群微生物计数表明：(1). 当发酵工艺和运行处于相对稳定状态时，微生物群体的 组成也达到相对的稳定，各类微生物之间保持动态平衡关系。当产乙酸菌的数量为107-108个/ml时，产甲烷菌的数量为105-106 个/ml，当产乙酸菌数量为106-107个/ml时，产甲烷菌的数量为103-105个/ml。(2).稳态运行条件下，黑液预处理为甲烷发酵创造 了有利的生态环境，获得了较好的处理效果和较高的COD转化为沼气的产率0.39-0.48l/g·CODcr·d，反应器中形成较为稳定而数 量较下水污泥中高1-2个数量级的厌氧发酵微生物区系组成。这一结果为黑液厌氧发酵提供了微生物理论依据。Paper industry is one of the important pollution source of water environment in our country. Its character of water pollution is many small factories, much grass pulp, disadvantageous technique, large preading area of pullution. Its effluent makes up 1/6 of whole country's industry wastwater. Its organic pollutant accounts 1/4 of whole country's. Alkaline grass paper pulp effluent with pollutants such as ligoin, remaining alkali sulfide, chloride besides organic material, is a kind of high concentrate organic wastewater which has high PH walug, dark colour and is difficult in treatment. There is urgent require to find ways to treat the wastewater. There are different ways to treat alkaline paper grass pulp effluent. According to the research advances and our experiment work, the most economical and useful way is anaerobic degradation which was advanced quick in last ten years. In the control of waste water of small pulp paper mill, the study of wastewater utilization technology should be emphasized, besides alkaline retrieving and different kinds of comprehensive utilization technology. Our experiment used modified UASB(Upflow Anaerobic Sludge Blanket Reactor). Two kinds of plan were disgned. The results are lined below. 1. The first experiment plant-aciding black pulp effluent and methanogenic digestion. The dynamic model experiment results of I-UASB reactor showed: (1)The mesophilic(35℃±1℃)high effect UASB reactor having haydite and threee state seperation in it had the character of keeping high bioimass concentration and preventing losss of sludge. It had advantages of high effect, energe saving, energe prodcing and short HRT(Hydroulic retention time). When the influent COD was 7500-10000mg, HRT was shortened from 7 days to 3days, organic loading rate was 1.22g-3.43COD/l· d, the average COD removal efficiency was 55%-45%. The highest COD efficiency was 60.2-63.5%, BOD removal of 75.9 -83.4% was achieved. Biogass production rate were up to 0.29-0.67l/l·d. Biogass converted efficiency from every gram of COD could reach 0.39-0.48l/gCOD·d. Methane content was 65.0-75.5%. Chemical method was used to deplate lignin in anaerobic digestion effluent. Total COD removal efficiency could be more than 80%. (2)Using aciding annaerobic digestion to treat the black effluent was reseanable in technique and technology. 2. The second experiment plan-anaerobic digestion was used after the chemical method was used to deplate lignin in the black effluent. The result of dynamic experiment of II-UASB reactor showed: (1)High effect mesophilic (35℃±1℃)UASB reactor having soft slaffing in was used. When influent COD was about 7000-13000mg/l, HRT was shortened from 6 days to 1 day and organic loading rate was increased from 0.90 to 11g COD /l·d, average COD removal efficiency remained stable on 70-77%. BOD, removal efficiency was between 87.3-93.1%. Biogass production rate was 0.2-2.6l/l ·d .Biogass converted efficiency from a gram of COD was 0.39-0.481/gCOD·d with the high value of 0.53l/gCOD·d. Methane content was 63-70%. (2)The way that using physical, chemical Pre-treatment-anaerobic digestion to treat alkaline black effluent is feasible and can be used in some factories when the condition exists. 3. Counting of several class of microoganisms in anaerobic digestion stage showed: (1)As the disgestion was in stable motion, the compositon of microorganic colony could get relative stable. Dynamic balance was remaining among different kinds of microorganism such as methanogenic bacteria, Acidogenic bacteria, sulfate reducing bacteria, and heterotrophic bacteria. (2)Under stable motion, the pre-treatment of black effluent produced favourable eco-enviroment for methanegenic digestion. Good treatment effect and high biogass convertent efficiency from COD(0.39-0.48l/g·COD· d)were gotten. Some stable and high quantity(10-100times more than sewage sludge)microorganism colony were formed in the reactor. This result provided theoretical basis for anaerobic digestion of black effluent.

功能菌剂的混合发酵研究

随着化工行业的发展，大量有毒有害难降解有机物随工业废水的排放进入环境，这些物质能够在环境中长期存在、积累和扩散，通过食物链对动植物的生存及人类的健康造成不良影响。本文以苯酚、对氯硝基苯、氯苯和十六烷为模拟污染物，以前期研制的功能菌剂为对象，经过紫外线线诱变筛选出优于出发菌株的功能菌，对诱变后功能菌的理化性能进行了研究，对菌种进行了鉴定，在此基础上，就其相互之间的微生态关系进行研究，为混合发酵提供理论基础，并就其最佳发酵条件及发酵参数进行了研究，最后对发酵产品的性能进行了检测。目前，国内外有关功能菌剂的研究还存在多方面的不足，主要包括：①由于多菌种混合发酵过程较为复杂，各菌之间存在复杂的相互作用，影响因素较多，关于菌种之间的相互关系研究得很少，环境功能菌剂的发酵方法大多采用单独发酵后混合的方式。单独发酵对原材料、设备和能源的利用率较低，对于多菌种制剂发酵，在设备、能源和原材料的方面造成的浪费更大，将会大幅增加菌剂的生产成本，影响多菌种功能菌剂的发展；②功能菌剂生产过程的质量控制方面研究得较少；③功能菌剂产品的稳定性、抗冲击性能研究得较少，对环境微生物制剂的研究主要集中在菌种选育和培养条件优化方面。 通过本论文研究，得到以下主要结论。 （1）在紫外线诱变处理中，用紫外线对发生一定程度退化的出发菌株进行诱变处理后，六株具有高效降解性能的菌株被筛选出来，诱变筛选出的菌株形态和ERIC-PCR指纹图谱与出发菌株相比发生了明显改变；而且诱变后的菌株对目标难降解底物的降解能力均得到改善，其中，FPN、FCB、F14、FEm对目标底物的降解率提高了20%以上；诱变后菌株经过7次连续传代接种后，对目标难降解底物的降解率无显著变化，具有一定的遗传稳定性。并对诱变后的功能菌进行了初步的鉴定，这6株菌都分别是芽孢杆菌。 （2）对诱变后的功能菌相互之间的微生态关系进行了研究，通过抑菌实验、生长量以及基质消耗量的比较，确定它们之间的生长关系是无害共栖关系，可以进行混合发酵。 （3）对该功能菌剂进行发酵培养条件研究，结果表明发酵培养基的最佳成分（g/L）：葡萄糖 31.0g/L、玉米粉10.0g/L、磷酸氢二钾1.0g/L、硫酸铵1.1g/L、硫酸镁0.55g/L。通过研究不同的培养条件对菌体生长和降解性能的影响，确定了最佳培养条件：培养基初始pH7.5；最适温度32℃；培养基装液量125mL（250 mL三角瓶），以及培养时间对降解性能的影响，培养20 h的产物对降解最为有利。通过研究添加不同目标污染物对菌体生长和降解性能的影响，确定了添加目标污染物的最佳量以及最佳时间：苯酚投加量：1.125 g/L，对氯硝基苯投加量：0.1 g/L；最佳投加时间为发酵培养开始后4 h。 （4）以摇瓶分批发酵最优条件为基础，对FPN、F10、FCB、FNa、F14 和 FEm进行了摇瓶分批发酵试验。以摇瓶分批发酵试验数据为依据，对功能菌剂分批发酵动力学进行了研究，建立了菌体生长和基质消耗的动力学模型，拟合模型能较好的反映功能菌剂分批发酵过程。 （5）功能菌剂和活性污泥协同作用，可以提高系统的生物降解能力，功能菌剂投加量为2%，新鲜活性污泥3500 mg/L，降解24 h条件下，功能菌剂和活性污泥的协同作用对COD的去除率和对照组相比，最多的提高了36.8%。功能菌剂和活性污泥协同作用以及活性污泥的单独作用，其生物降解过程均符合一级反应动力学过程，功能菌剂和活性污泥协同作用的生物降解动力学方程为：，相关系数97%。采用SBR运行方式，引入功能菌剂的SBR系统明显能够改善和提高生物降解的效率。与仅有活性污泥的系统相比，系统对COD的平均去除率可以提高27.1%，同时，系统的耐负荷冲击以及耐毒害冲击的性能比仅有活性污泥的SBR系统强，特别是负荷冲击对引入功能菌剂的SBR系统影响很小。仅有活性污泥的SBR系统经过负荷冲击和毒害冲击之后，不能恢复到冲击之前的水平，而且系统有效作用时间的周期比引入功能菌剂的SBR系统相比大大缩短，而引入功能菌剂的SBR系统处理效果较为稳定，恢复能力很强。 Along with the development of industries, many recalcitrant organic chemicals have been discharged into natural environments together with wastewaters and can exist in waters, soil and sediments for a long time without degradation. These haz-ardous substances, their byporducts and metabolizabilities can be highly toxic, mu-tagenic and carcinogenic, thereby threatening animals, plants and human health through food chain. Consequently the removal of these compounds is of significant interest in the area of wastewater treatment. In this dissertation, the phenol, hydro-quinone, chlorobenzene and hexadecane treated as the model pollutants, the func-tional microorganism agent was used as the starting strains, they treated with ultra-violet light, and then the mutant strains with high degradation ability were screened out and identified primarily, the relationship between these stains were studied, the medium composition and fermentation conditions were optimized, the degradation ability of the fermented production was tested. The literature survey indicates that the study of the microorganism agent is far from complete and more information is re-quired on following problems. 1, Because of the complexity of relationship in mixed fermentation and the complicated factors, the study is hardly to process.2, There is a lack of information on the quality control of the producing process .3, And there is a lack of information on the stability about the microorganism agent. In this dissertation, the main results of the present study could be summarized as follows: (1)The degenerate starting strains were treated with the ultraviolet light, and six mutant strains with high biodegradation ability were screened out by using the me-dium with selective pressure of model pollutants. The mutant strains had great changes in colonialmorphology and ERIC-PCR fingerprinting. And the mutant strains got obvious advantages over the starting strains in degradation ability and over 20% improvement of removal rates was achieved for FPN、FCB、F14 and FEm. The de-gradation ability of the mutant strains was stable after seven generations. After that, the mutant strains were primarily identified as bacillus respectively. (2) The relationship between these mutant strains was studied. By the compari-son of antibiosis effect, biomass and consumption of substrate, the relationships were neutralism and they could be mixed fermented. (3) The optimized cultivation conditions were as follows: glucose 31.0 g/L, corn power 10 g/L, K2HPO4 1.0 g/L, (NH4)2SO4 1.1 g/L, MgSO4 0.55 g/L, initial pH7.5, temperature 32℃, working volume 125 mL/250 mL, and cultivation time 20h (con-sidering the time effect on degradation ability), adding pollutants phenol (1.125 g/L) and hydroquinone (0.1 g/L) into the broth at 4 h after cultivation. (4) Based on the above optimum condition, the batch fermentation was per-formed with strains FPN, F10, FCB, FNa, F14 and FEm in shake flask. The batch fermentation kinetics was studied based on the experimental data. Two kinetic models were constructed which could reflect the regularity of growth and substrate consump-tion in the process of batch fermentation. (5) The co-operation of functional microorganism agent and activated sludge could raise biodegradation of system by adding some microorganism agent and 3500 mg/L fresh activated sludge. Bioaugumentation by the addition of high effective deg-radation culture enhanced the treatment effect of SBR system and the COD removal rate was increased by 20%-36.8%. Its biodegradation matched first-order dynamical reaction equation, and the reaction equation was ln0.2327.391ct=−+. The micro-organism agent had the effect of optimization to activated sludge micro-ecosystem. The SBR system adding 2% microorganism agent, the average COD removal rate of that was increased by 27.1% and stronger anti-shock ability to load and toxicant were achieved (compared with SBR system just adding activated sludge). Especially the load-shock has barely effect to the SBR system adding microorganism agent. After the load and toxicant shock, the SBR system just adding activated sludge couldn’t come back to original level and the activated sludge micro-ecosystem was frustrated. The applying of microorganism agent increased biological activity and system’s re-sistance ability to load shock and toxicant shock.

利用含纤维素原料发酵产氢的研究

本文从新鲜大熊猫粪便和实验室保存的沼气发酵富集物中筛选得到 4 株厌氧纤维素分解菌B5、C3、D3-2、D4-1，利用这4 株菌预处理秸秆，然后将预处理后的秸秆用本实验室保存的厌氧产氢菌来发酵进行生物产氢。同时还比较研究了：○1 用1% H2SO4、25% NH3 · H2O和12% NaOH对秸秆进行化学预处理；○2 用厌氧纤维素分解菌对秸秆进行生物预处理；○3 化学与生物组合预处理对秸秆发酵生物产氢的影响。实验结果表明：12% NaOH和生物组合预处理后的秸秆发酵产氢效果最好，其产氢量为21.04 mL g-1，是未经预处理秸秆的75 倍；最高氢气浓度为57.3%，是未经预处理秸秆的96 倍；其产氢的最适pH 为4.5 ~ 6.0，最佳底物浓度为45 ~ 55 g L-1；其发酵过程中的挥发性脂肪酸（VFAs）以乙酸和丁酸为主。 本实验筛选到的 4 株厌氧纤维素分解菌株中，B5 和D4-1 在降解纤维素的同时还具有直接以纤维素为底物产氢的功能，因此本文分别对菌株B5 和D4-1 以及二者的组合菌株B5+D4-1 直接利用秸秆为基质发酵生物产氢做了初步探索研究。结果发现：组合菌株发酵产氢的效果以及对秸秆纤维素和半纤维素的降解率要比单菌株好。菌株B5+D4-1 发酵，秸秆的产氢量为11.4 mL g-1，分别是B5 和D4-1 单菌株的1.6 倍和3.1 倍；组合菌株B5+D4-1 发酵的最大氢气浓度为31.6%，分别是B5 和D4-1 单菌株的1.3 倍和2.4 倍。在发酵过程中，组合菌株B5+D4-1 对秸秆纤维素和半纤维素的最高降解率分别为35.0%和11.8%，分别是菌株B5 的1.2 倍和1.1 倍，是菌株D4-1的1.5 倍和1.3 倍。菌株B5，D4-1 以及组合菌株B5+D4-1 发酵过程产生的挥发性脂肪酸（VFAs）均以乙酸为主。菌株B5 单独发酵过程中只检测到乙酸和丁酸，菌株D4-1 单独发酵以及组合菌株B5+D4-1 发酵过程检测到有乙醇、乙酸和丁酸。 The fermentative bio-hydrogen production by anaerobic hydrogen bacteria preserved in our laboratory from the straw which had been pretreated by four anaerobic cellulolytic decomposition strains of B5, C3, D3-2, D4-1 which were isolated and screened from giant panda’s excrement and biogas fermentation enrichments conserved in our laboratory was studied. Besides, the impact of chemical(1% H2SO4、25% NH3·H2O and 12% NaOH), biological (cellulolytic strains of B5, C3, D3-2, D4-1) and chemical-biological combination pretreatment on bio-hydrogen production from straw by fermentation was also comparatively studied. The experiments showed that the best results of bio-hydrogen production were obtained from the straw with 12% NaOH-biological combination pretreatment method, its capability of bio-hydrogen production was 21.04 mL g-1, which was 75 times higher than the straw without pretreatment; the maximum concentration of H2 was 57.3%, which was 96 times higher than the straw without pretreatment; its optimum pH range was 4.5 ~ 6.0, and its optimum range of substrate concentration was 45 ~ 55 g L-1; In the process of fermentation, the main composition of VFAs were acetate and butyrate. Among the four strains of B5, C3, D3-2, D4-1, B5 and D4-1 have the function of hydrogen-producing by cellulose used as substrate when it decompose cellulose, so the preliminary exploration and research on fermentative bio-hydrogen production by B5, D4-1 and B5+D4-1 which directly used straw as substrate was carried out. The results showed that the combination strains of B5+D4-1 was strikingly better than either B5 or D4-1 strain in the fermentative hydrogen production. The hydrogen-production capability of B5+D4-1 was 11.4 mL g-1 which was respectively 1.6 times and 3.1times higher than B5 and D4-1; the maximum hydrogen concentration of B5+D4-1 was 31.6% which was respectively 1.3 times and 2.4 times higher than B5 and D4-1. In the process of fermentation, the maximum degradation rate of cellulose and hemicellulose in straw was respectively 35.0% and 11.8% by B5+D4-1, which was 1.2 times and 1.1 times higher than B5, and was 1.5 times and 1.3 times higher than D4-1 respectively. The Volatile Fattty Acids(VFAs) generated in the process of fermentation with strains of B5, D4-1 and B5+D4-1 were all mainly acetate. Acetate and butyrate were detected in the process of fermentation with B5, ethonal, acetate and butyrate were detected in the process of fermentation with D4-1 and B5+D4-1.

耐酸产甲烷菌的分离及其复合菌剂的研究

本文从成都龙泉垃圾填埋场和宜宾造纸厂分离到耐酸性能优良的高温产甲烷菌RY3和中温产甲烷菌SH4，并将其与实验室现有的利用不同底物的产甲烷菌配伍组合成了复合菌剂。采用活性污泥作为固体附着物，研制出了固体产甲烷菌复合菌剂。 菌株RY3的pH耐受范围为5.5～10.5，最适生长pH 6.0～8.0。菌株RY3为革兰氏阳性，长杆状，多数单生，不运动；菌落浅黄色，形状近圆形；利用H2+CO2或甲酸盐作为唯一碳源生长，不利用乙酸盐，对氯霉素非常敏感。该菌最适生长温度为55℃～65℃，最适NaCl浓度为0～2%。根据形态和生理生化特性及16S rDNA序列分析将其初步定为热自养甲烷热杆菌（Methanothermobacter thermautotrophicus）。添加RY3菌液与仅添加厌氧污泥作为接种物相比一周内可使达到最大产甲烷速率所需时间缩短三分之二，甲烷总产量提高约1.8倍。菌株SH4的生长pH范围5.5～9.5，其对酸碱具有良好的适应性，培养3天后，在初始pH值为6.0～8.0的培养基中甲烷产量相差不大，且基本达到最大产量。SH4革兰氏染色阳性，短杆状，多数单生，不运动；菌落近圆形，微黄；利用H2+CO2或甲酸盐作为唯一碳源生长，不利用乙酸盐，对氯霉素非常敏感。SH4最适生长pH 为7.0，最适生长温度为35℃，最适NaCl浓度为0～1.5%。实验表明，添加SH4菌液与仅添加厌氧污泥作为接种物相比可使产甲烷启动时间缩短三分之一，甲烷总产量亦有大幅提高。从形态和生理生化特征以及16S rDNA序列分析表明SH4为嗜树木甲烷短杆菌（Methanobrevibacter arboriphilus）。 以活性污泥为附着物，与培养基和菌种经搅拌后厌氧发酵可得产甲烷菌固体复合菌剂。固体复合菌剂的pH耐受范围为5.5～9.5，温度耐受范围为15℃～65℃，表明其对环境的适应性较强。以猪粪为底物进行厌氧发酵，接种复合菌剂进行试验，以接种实验室长期富集的产甲烷厌氧污泥作为对照，在20℃时，发酵甲烷浓度与对照基本一致，但每日产气量优于对照，第15天时接种复合菌剂的发酵瓶每日产气量是对照的1.59倍；50℃时达到最大甲烷含量所需时间比对照缩短三分之二，三周内总产气量约为对照的2.7倍，甲烷总产量约为2.8倍。以不加接种物为对照，接种复合菌剂20℃时发酵甲烷含量达到50%约需2周，对照2周内甲烷含量最高仅为4.3%；50℃时接种复合菌剂发酵仅需约1周甲烷含量便可达50%，对照则至少需要2周。 In this paper, high-temperature Methanogen RY3 and middle-temperature SH4 were isolated from Chengdu Longquan refuse landfill and Yibin paper mill. They could be used to make compound inoculum that producing methane with the existing Methanogens utilized different substrate. With using anaerobic activated sludge be solid fixture, the process had been designed to produce solid compound inoculum. Strain RY3 possessed excellent capacity of acid and alkali-tolerant. The pH-tolerant scale of RY3 was 5.5～10.5 and its optimum pH value for growth was 6.0～8.0. RY3 was G+, long-rod shape, monothetic and nonmotile, the colony was pale yellow with suborbicular-shape. Formate or H2+CO2 but not acetate was utilized by RY3 as sole C-source, and it was very sensitive to chloramphenicol. Besides, strain RY3 grew fastest at 55℃～65 and 0℃～2% NaCl. Characteristics of modality and physiology with sequence analysis of the 16s rDNA gene of strain RY3 preliminarily showed that it was Methanothermobacter thermautotrophicus. The experiments indicated that the time which began to produce methane with the highest velocity could be shortened two third by adding RY3 in one week, and the total methane production also was 1.8 times than before. Strain SH4 possessed wide scale of growing pH（5.5～9.5）and excellent ability of acclimatizing itself to acid-alkali. The methane production had no apparent difference among those cultivated in different initial pH（6.0～8.0）after three days and equaled to the maximum production basically. Cells of SH4 were G+, short-rod sharp, monothetic and nonmotile. The colony was pale yellow with suborbicular-shape. Formate or H2+CO2 but not acetate was utilized by SH4 as sole C-source, and it was very sensitive to chloramphenicol. Besides, it grew fastest at pH 7.0，55 ℃～65 and 0℃～2% NaCl concentration. The experiment indicated the time that began to produce methane could be shortening one third by adding SH4. And the total methane production also rose apparently. Characteristic of modality and physiology with sequence analysis of the 16S rDNA gene of strain SH4 demonstrated it was Methanobrevibacter arboriphilus. The activated sludge was utilized as fixture, mixed with culture medium and inocolum, that the solid compound inoculum could be produced by anaerobic fermentation. The compound inoculum could grow between pH 5.5～9.5, 15℃～65. It demonstrated the compound inoculum ha℃ve great ability of adapting to circumstance. In the experiment that making pig manure be substrate and taking the anaerobic sludge producing methane that cultured in long term in laboratory to be comparison, the concentration of methane in fermentation added compound inoculum almost equal to the comparison at 20℃, but the volume of gas production could be a little higher. The gas production everyday inoculated compound inoculum was 1.59 times to comparison. The time that the concentration of methane to maximum could be shortening by two third by adding compound inoculum, and the total gas production was 2.7 times to comprison while the total methane production was 2.8 times. If take the no inoculum be the comprasion, anaerobic fermentation added compound inoculum made the concentration of methane to 50% in 2 weeks but the comparison only to 4.3% at 20℃. The time that the concentration of methane to 50% by adding compound inoculum only need 1 week, but the comparison need 2 weeks at 50℃.

投加溶胞菌群对剩余污泥消化影响的初步研究

活性污泥法是目前世界上普遍应用的污水生物处理工艺，其在运行过程中产生大量的剩余污泥。由于剩余污泥处理费用巨大及污泥最终处置对环境具有潜在危害问题，污泥的处理和处置已经成为水处理领域关注的焦点。本文利用实验室筛选的溶胞菌群，在好氧消化的同时对污泥进行前处理，促进剩余污泥的破解与溶胞，再通过两相厌氧处理对污泥进行进一步消化，以研究投加溶胞菌对剩余污泥消化的影响。 本研究中溶胞菌污泥减量化技术分为两个部分，第一，污泥在溶胞菌作用下的好氧消化与污泥传统好氧消化的对比研究，利用取自成都三瓦窑污水处理厂剩余污泥，向好氧污泥消化反应器中投加溶胞菌，检测各项污泥指标，并通过同传统好氧污泥消化对比，以研究溶胞菌对污泥好氧消化的影响。第二， 经过溶胞菌处理后好氧消化的剩余污泥进行两相厌氧处理研究。通过建立好氧溶胞联合两相厌氧消化系统的来处理剩余污泥，并与相同条件运行的两相厌氧消化系统做对比，检测运行过程中系统中物质成分变化，研究了其处理能力和运行稳定性，探索了两相厌氧消化系统中的发酵类型差别，验证了好氧溶胞对剩余污泥的破解效果。 研究结果表明：污泥在溶胞菌作用下的好氧消化效果和消化效率均优于传统好氧消化。在溶胞菌群存在的情况下，剩余污泥的TSS和VSS去除率达到40%和53%，远高于传统好氧消化的12%和20%。污泥经过溶胞及好氧消化后，TCOD去除率达到54.4%。经过溶胞菌处理后的剩余污泥再进入两相厌氧处理系统，进入厌氧处理系统的剩余污泥的VSS／TSS比值约为0.62。在两相厌氧处理水力停留时间(HRT)为8d时，溶胞处理污泥厌氧消化后VSS去除率达到55.17%，对照组两相厌氧系统的VSS去除率平均值为18.53%。经过溶胞处理的两相厌氧系统的污泥减量了能力远高于对照组。两相厌氧系统的pH值和碱度说明系统运行较为稳定。产酸相的有机酸中乙酸含量高于丙酸和丁酸，说明发酵末端产物以乙酸为主。在20天的试验周期内，污泥溶胞处理后、两相厌氧系统产甲烷相产气量累积产气量为1.2L，对照组只有375ml。气体中甲烷含量都在55%左右。该研究结果表明，好氧溶胞对污泥有破解能力，溶胞处理对两相厌氧中产酸相水解污泥细胞有明显的促进作用，提高了产酸相的水解酸化能力和效率。该研究对于利用生物溶胞途径提高污泥消化效率具有重要意义。 The actived sludge process has been used more and more extensively, but the procedure will lead to a large quantity of excess sludge. The treatment of Excess activated sludge has becomes a focuses not only for it is a seriously negative effect on environment but also for the costly disposal comes subsequently. The cell lysing bacterium was keeped in our lab to joined in the digestion of the excess activated sludge which was carrying at the same time with pre-processing of sludge to investigated the influence of cell lysing bacterium on excess sludge. There are two part in the method of cell lysing bacterium digesting sludge technology, the first, comparison of excess sludge digestion between anaerobic Cell-lysing Pretreatment and Conventional Aerobic Process. The sludge which was collected from San Wanyao disposal plant in Chengdu was thrown into the aerobic process system with cell-lysing bacterium, then, the indexes were detected to compare the difference between the cell-lysing bacterium in aerobic process and the traditional method to determine the influence of cell-lysing bacterium on aerobic process ; The second, the research on the sludge which was pro-treated with cell-lysing and aerobic digestion in the diphase of anaerobic digestion system. The system of cell-lysing combined with diphase of anaerobic digesting was created to compare to the diphase of anaerobic digested system, the changes of mass constituent was detected to study the ability and steady of disposal. Moreover, the research explored the difference among the types of fermentation. The efficacious of aerobic process was been proved. The result shows that the digesting rate of aerobic process with cell-lysing bacterium was higher than the traditional process. The ratio of sludge is reach to 40%~53%, which was far more effectively than the traditional process rate of 12%~20%. The TCOD of sludge which was treated with cell lysing bacterium and Aerobic Process is reach to 54.4%. Then, the sludge was thrown into the diphase of anaerobic digesting system. VSS／TSS of sludge is 0.62, HRT is 6d, the reduction of VSS is reach to 40.8%. The pH and alkalinity indicate the steady running of the diphase anaerobic digest system. In the acerbity phasing, the content of acetic acid was more than butanoic acid and propanoic acid in organic acid, it is demonstrated that the main composition of final production of fermentation was Acetic Acid. During the 20d of experiment, methylhydride phasing of diphase anaerobic digest system produced 1.2L methylhydride, however, there is only 375ml in CK, the content of methylhydride in all gas phase was around the rate of 55%. The average ratio of VSS was 18.53% in CK diphase anaerobic digest system which was far more unavailable than the mass sludge rate of 55.17%. Results demonstrated that aerobic cell-lysing digested the sludge, the treat of cell-lysing could obviously promoted the hydrolyzeing of sludge cell in the acerbity phasing, which improved the ability and rate of hydrolization and acidification. This study is significant in inhenceing the rate of sludge digestion in the method of cell-lysing bacterium.