组合白腐真菌预处理木质纤维素原料的研究

木质纤维素原料种类多、分布广、数量巨大，通过燃料乙醇生产技术、厌氧沼气发酵技术将其转化成乙醇、沼气等二次能源，一定程度上可以缓解化石能源的不断消耗所带来的能源危机，也解决了农林废弃物引起的环境污染问题。其中以木质纤维素原料生产燃料乙醇，还可以避免以淀粉类和糖类原料生产燃料乙醇时带来的“与人争粮”等一系列问题。因此具有重要的经济效益、环境效益和社会效益。 然而，木质纤维素原料结构致密，木质素包裹在纤维素、半纤维素外围，导致其很难被降解利用，必须进行适当的预处理，去除木质素，打破原有的致密结构，利于原料的后续利用。因此，预处理成为木质纤维素原料能源化利用的关键。而目前预处理环节的费用过于昂贵，于是寻找一种高效、低成本的预处理方法是当今研究的热点。 本论文采用组合白腐真菌对木质纤维素原料进行生物预处理研究，与其他物理化学法相比，该法有着专一性较强、反应温和、不造成环境污染、成本低等优势。白腐真菌主要通过分泌木质素降解酶对木质素进行降解，从而破坏原料的致密结构，提高后续利用效率。所以木质素降解酶酶活的高低是影响原料预处理效果的一个关键因素。于是本论文首先通过将白腐真菌进行组合的方式提高木质素降解酶（漆酶，Lac）酶活；接着对组合菌的菌株相互作用机理进行研究，阐明组合菌Lac 酶活提高的原因，为菌株组合提高Lac 酶活这种方法的应用提供理论依据，同时也为后续组合白腐真菌预处理木质纤维素原料提供指导；进一步采用固态发酵和木质素降解酶两种方式对木质纤维素原料进行预处理研究，最大化去除木质素成分，破坏原料的致密结构；最终对预处理后原料的酶解糖化进行初步研究，为原料后续的能源化应用奠定基础。具体研究结果如下： （1） 以实验室保存的三株主要分泌Lac 的白腐真菌为出发菌株，筛选得到一组Lac 酶活明显提高的组合菌55+m-6，其中菌株55 为Trametes trogii sp.，m-6 为Trametes versicolor sp.，组合后Lac 酶活较单菌株分别提高24.13倍和4.07 倍。组合菌的最适产酶条件为pH 6.5、C/N 16:1、Tween 80 添加量为0.01%，在该条件下组合菌的Lac 酶活峰值比未优化时提高4.11倍。 （2） 对组合菌55+m-6 菌株间相互作用机理进行研究，发现菌株之间不存在抑制作用；平板培养时，菌丝交界处Lac 酶活最高并分泌棕色色素；液体培养时，菌株m-6 对组合后Lac 酶活的提高起着更为重要的作用：菌株m-6的菌块、过滤灭菌胞外物以及高温灭菌胞外物均能明显刺激菌株55 的Lac产生；菌株55、m-6 进行组合后，同工酶种类未发生增减，但有三种Lac同工酶浓度有所提高；对菌株胞外物进行薄层层析和质谱分析，结果表明组合前后菌株胞外物中各物质在浓度上存在较大的变化。推测组合菌Lac酶活的明显提高，主要是由于菌株m-6 胞外物中的一些物质能刺激菌株55 分泌大量Lac 进行代谢，且这些刺激物质并非菌株m-6 特有，菌株55自身也可以代谢生成，但是适当的浓度才能刺激Lac 的大量分泌。 （3） 将组合菌55+m-6 用于固态发酵预处理木质纤维素原料，发现其对玉米秆的降解程度最大，在粉碎度40 目、含水率65%的最优处理条件下，处理至第15d，秸秆失重率为41.24%，其中木质素、纤维素、半纤维素均有降解，且Lac 和纤维素酶（CMC）酶活以及还原糖量均达到峰值。 （4） 对玉米秆进行木质素降解酶预处理，发现Lac/1-羟基苯并三唑（HBT）系统对玉米秆木质素的降解效果最好，在最优处理条件时，即HBT 用量0.2%、处理时间1d、Lac 用量50U/g，木质素降解率可达12.60%。预处理后玉米秆的致密结构被破坏，比表面积增大，利于后续酶与纤维素、半纤维素成分的结合。 （5） 对预处理后的玉米秆进行酶解糖化，其中组合菌固态发酵预处理后玉米秆的糖化率比对照高4.33 倍；Lac/HBT 系统预处理后玉米秆的糖化率比对照高2.99％，糖化液中主要含有木糖、葡萄糖两种单糖。 There are many kinds and large quantities of lignocellulosic biomass widely distributed on the earth. They can be converted into secondary energy such as fuel ethanol, biogas, et al., which can relieve the energy crisis caused by consumption of fossil energy resources and solve the problem of environmental pollution caused by agriculture and forestry waste. Meanwhile, the production of fuel ethanol from lignocellulosic biomass can ensure food supply to human kind instead of starch- and sugar-containing raw materials. So the energy conversion of lignocellulosic biomass contributes considerable economic, environment and social benefits. However, lignocellulosic biomass has the compact structure, in which lignin surrounds cellulose and hemicellulose, so it must be pretreated before energy usage and pretreatment is one of the most critical steps in the energy conversion of lignocellulosic biomass. At present, the cost of pretreatment is too expensive, so looking for an efficient and low-cost pre-treatment method is one of recent research hot spots. In this research, combined white rot fungi pretreatment method was used, which had some advantages in low cost, high specificity, mild reacting conditions and friendly environmental effects compared with the other physical and chemical methods. White rot fungi secrete lignin degrading enzymes to degrade the content of lignin and damage the contact structure of lignocellulosic biomass, so the activity of the lignin degrading enzymes is the key factor to the degradation effect of raw materials. Firstly, the combined fungi with high laccase activity were screened; secondly, the interaction mechanism between strains was studied, and the cause of higher laccase activity after strains combination was also preliminary clarified; under the guidance of the mechanism, lignocellulosic biomass was pretreated by the combined fungi; lastly, the enzymatic hydrolysis of pretreated lignocellulosic biomass was also preliminary studied; all of the researches could lay the foundation for the energy application of lignocellulosic biomass. The specific research results were as follows: (1) The combined fungi 55+m-6 with significant higher laccase activity were screened from the three white rot fungi stored in our lab which mainly secreted laccase. Strain 55 and strain m-6 were Trametes trogii sp. and Trametes versicolor sp., respectively. The laccase activity of combined fungi was 24.13 and 4.07-fold than strain 55 and strain m-6, respectively. The optimized condition for laccase production of the combined fungi in liquid medium was pH 6.5, C/N 16:1 and Tween 80 0.01%. In this optimized condition, the laccase activity of combined fungi was 4.11-fold higher comparing with which in non-optimized medium. (2) The interaction mechanism between strain 55 and strain m-6 was further studied, and no inhibition effect was observed. Brown pigment was secreted on the junction of the two strains on the plate, where the highest laccase activity was detected. Strain m-6 was much important to boost laccase activity of combined fungi in liquid medium, and strain 55 was stimulated by fungal plug, filter sterilized extracellular substances and high temperature sterilized extracellular substances of strain m-6 to produce laccase. The types of laccase isozymes did not change after combining strain 55 and strain m-6, but the concentrations of three types increased. Mass Spectrometry and TLC analysis of extracellular substances of each strain showed that concentration of some substances considerably changed after strains were combined. It was supposed that the cause of higher laccase activity of combined fungi was mainly due to some extracellular substances of strain m-6 with the appropriate concentration which stimulated laccase secretion of strain 55 and generated not only by strain m-6 but also by strain 55. (3) Combined fungi 55+m-6 were used to lignocellulosic biomass pretreatment with the type of solid-state fermentation. The highest degree of degradation of corn straw was obtained, including the rate of weight loss was 41.24% and the lignin, cellulose and hemicellulose were degraded partially under the optimized condition of 40 mesh, 65% water content on 15th day. Laccase, CMCase activities and content of reducing sugar reached the maximum value on that day. (4) Lignin degrading enzymes from combined fungi 55+m-6 were used for corn straw pretreatment. The most remarkable degradation of lignin in corn straw with Lac/1-hydroxybenzotriazole (HBT) system was observed, and the 12.60% lignin degradation was obtained under the optimized condition of 0.2% HBT, 50 U/g laccase for 1 d. After pretreated by Lac/HBT, the tight structure of corn straw was demolished and specific surface area increased, which had advantages for accessible of enzyme to cellulose and hemicellulose. (5) The corn straws pretreated by combined fungi 55+m-6 with the type of solid-state fermentation and Lac/HBT were used for enzymatic hydrolysis, and the saccharification rates of each pretreatment type were 4.33 times and 2.99% higher than CK, respectively. The enzymatic hydrolysis liquid of corn straw pretreated by Lac/HBT mainly contained xylose and glucose.

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℃.

一株丝状真菌对半纤维素利用的初步研究

本文筛选出一株能利用木糖产乙醇的丝状真菌Z7，对其利用木糖和半纤维素水解产物产乙醇的发酵条件进行了研究，并对Z7 利用玉米芯产木聚糖酶的条件进行了优化。全文分为三部分： 第一部分：目标微生物筛选、纯化及系统发育分析。以木糖为唯一碳源，采用梯度稀释和平板化线法从高温、中温酒曲中分离到16 株能利用木糖良好生长的丝状真菌；通过发酵试验复筛，获得一株能产乙醇的丝状真菌Z7；综合形态学和ITS 序列分析，初步鉴定为Aspergillus flavus。 第二部分：Z7 的乙醇发酵条件研究。以木糖为碳源，通过单因素试验确定最佳氮源和发酵温度；通过正交试验及SPSS 软件分析得到了不同N、P、K 成分对乙醇、残糖和菌体干重的影响。获得最佳的发酵条件为：（g/L）木糖50，尿素1, NH4NO3 1, K2HPO4 2 , KCl 0.5 , MgSO4.7H2O 0.5 , NaNO3 1 , pH 自然，培养温度33 ℃。以玉米芯半纤维素稀酸水解液为底物进行乙醇发酵，根据稀酸水解的单糖释放量和乙醇产量，确定115 ℃，1 h 为最佳玉米芯预处理条件；结合最佳发酵条件，添加1 g/L 的吐温20 能获得最大的乙醇浓度8.31 g/L。因此，Aspergillus flavus Z7 能利用半纤维素水解产物产乙醇，其中木糖的利用率80%以上。 第三部分：Z7 利用玉米芯产木聚糖酶条件优化。Aspergillus flavus Z7 在具有产乙醇能力的同时还具有产木聚糖酶的能力。本文通过单因素和正交试验得到最佳产酶培养基组分为：(g/L)玉米芯20，尿素2, 酵母膏2.5, K2HPO4 5，NaNO31, MgSO4.7H2O 1。单因素试验表明，用纱布代替塑料布密封摇瓶封口能显著提高产酶量；Z7 在碱性条件下具有更强的产酶性能。在最优条件下发酵，能产生最大木聚糖酶活122.23IU/mL。通过薄层分析，验证了Z7 产生的木聚糖酶具有水解木聚糖生成木糖及木寡糖的能力。 A strain of filamentous fungus which can produce ethanol by using the xylose was isolated in this research. The ethanol fermention conditions from xylose and dilute-acid hydrolyzate of the corn core were studied. The conditions of xylanase production by Z7 were also optimized. The paper involved three parts. Part1: Isolation, purification and phylogenetic analysis of the microbe. By using xylose as the single carbon source and the pla te streaking method, several filamentous fungi were isolated from the wine starter; through the fermentation test, a filamentous fungus Z7 which can produce ethanol was further recognized; furthermore, according to the morphologic observation and ITS seque nces analysis, Z7 was identified as Aspergillus flavus at the first step. Part2: Research on the condition of ethanol fermentation by Z7. By single factor experiment, the optional nitrogen resource and temperature of the fermentation were fixed; meanwhile, through the orthogonal array tests and the analysis of statistic software SPSS, the optional component of the culture medium and the fermentation condition were organized as follows: (g/L) xylose 50, urea 1, NH4NO3 1, K2HPO4 2, KCl 0.5 , MgSO4.7H2O 0.5, NaNO31, pH nature, temperature 33℃. Based on these optimal parameters, the fermentation of dilute-acid hydrolyzate of the corn core was carried on by Z7. According to the quantities of released sugar monomers and content of the ethanol, 115℃ in 1h is the best pretreatment condition; the maximal ethanol content can be obtained when 1g/L Tween 20 was added to. Therefore, the filamentous fungus Aspergillus flavus can use the hydrolysate of hemicellulose to produce ethanol, and the rate of xylose utilization was over 80%. Part3: Optimization of Z7’s xylanase producing condition from corn core. Aspergillus flavus Z7, which can utilize xylose or the hydrolysate of hemicellulose to produce ethanol, also had the ability of xylanase production. The optional component of the culture medium were fixed by the single factor experiment and the orthogonal array tests, and they were organized as follows: (g/L) corn core 20, Urea 2, Yeast extract 2.5, K2HPO4 5, NaNO31, MgSO4.7H2O 1; it was testified by the single factor experiment that sealing the shaking flasks with pledget other than plastic paper can obviously increase the xylanase activity; moreover, Z7 showed better xylanase production ability when in the alkali environment. Under the optional fermentation condition, the maximal xylanase activity 122.23IU/mL was proved. Through the analysis of thin- layer chromatography (TLC), the ability of xylanase from Z7, which can hydrolyze xylan to xylose monomer and oligomer, was vividly displayed.

泸州老窖古酿酒作坊空气微生物分布特征研究

本文主要研究了泸州老窖古酿酒作坊内外环境空气真菌和空气细菌的群落结构和分布特征。结果如下： 作坊内外环境空气微生物浓度差别显著，并随季节变换而变化，春、夏季微生物浓度较高，秋、冬季较低，空气真菌在夏季达到最高，细菌在春季最高。 古作坊内外环境检测到的真菌均为16 属，但优势菌属不同，作坊外的优势菌属为青霉属(Penicillium)、曲霉属(Aspergillus)、无孢菌(non-sporing)、枝孢霉属(Cladosporium)和链格孢属(Alternaria)；而作坊内优势菌属为曲霉属、青霉属、酵母菌(Yeast)、无孢菌，作坊内还含有较高浓度的根霉属（Rhizopus）、毛霉属（Mucor）、短梗霉属（Aureobasidiu），枝孢霉属和链格孢属等，曲霉属、酵母菌、根霉属、毛霉属为古酿酒作坊重要的酿酒真菌，青霉属、链格孢属为酿酒不利菌群。对古作坊内曲霉属进行了初步鉴定，主要是小冠曲霉（A.cristatellus）、米曲霉（A.oryzae）、黑曲霉（A.niger）和白曲霉（A.cadidus）。 空气细菌10 属21 种，作坊内外环境的优势菌属均为芽孢杆菌属（Bacillus）、微球菌属(Micrococcus)、葡萄球菌属(Staphylococcus)、假单胞菌属(Pseudomonad)，其中芽孢杆菌属在作坊内占有绝对的优势，浓度比在40℅以上，是古酿酒作坊重要的酿酒细菌，另外还检测到较高浓度的乳酸杆菌（lactobucillus），这类菌容易使酒味发涩发苦，为酿酒不利菌。 作坊内外环境空气微生物表现出明显的交流现象。作坊内，青霉属、枝孢霉属、链格孢属、葡萄球菌属等杂菌占有一定比例；而在作坊外,芽孢杆菌属、曲霉属、根霉属(Rhizopus)、酵母菌等处于相对较高水平，绿化环境较好的营沟头作坊内的短梗霉属，枝孢霉属和链格孢属等杂菌含量低于什字头和新街子作坊。 The community structure and distribution characteristic of airborne microbes was investigated in ancient brewage workshops of luzhoulaojiao. The results are as follows: The concentration of airborne microbes was different in interior and exterior environment of ancient workshops, and also varied by seasons. microbial concentration was higher in spring and summer, and lower in fall and winner. The highest levels of airborne bacteria was in spring, but the fungal’s in summer. The identified genus of fungi were 16 in interior and exterior environment of the ancient workshops. But the dominant genus were different , The advantage genus in the interior were Aspergillus, Yeasts, Penicillum and Nonsporing and in the exterior were Penicillum, Nonsporing, Cladosporium, Aspergillus and Aureobasidiu. Rhizopus ,mucor, Aureobasidiu, Cladosporium, Alternaria and all also were at a higher level. Among these, Aspergillus, Yeasts, Rhizopus ,mucor are important vintage flora . Penicillum, Alternaria do harm to vintage. Aspergillus of ancient workshops was identified , the preponderant aspergillus species were A.cristatellus, A.oryzae, A.niger and A.cadidus in ancient brewage workshops. 10 genus 21 species bacteria were identified, the advantage genuses among the interior and exterior of the three workshops were bacillus, microccus, Staphylococcus Pseudomonas. Bacillus, which account for beyond 40℅ of the total bacteria concentration in all sampling pots, was the most dominant genus. Lactobacillus was identified at a high level in ancient workshops, it makes spirit taste bitter and astringent. So it is not a kind of good bacterium for vintage. The fungus in the interior and exterior atmosphere characterized intercommunion phenomenon. Obviously, the concentration of profitless fungus such as Penicillum, Cladosporium, Alternaria appeared in the interior, and the fungus such as Bacillus, Aspergillus, Rhizopus and Yeasts in the exterior were at a relatively high level. the harmfull fungus in yinggoutou workshops such as Aureobasidiu, Cladosporium, Alternaria and all were lower than shenzitou and xinjiezi workshops.

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.

制革废水生物强化处理及中水回用试验研究

制革行业是轻工行业中仅次于造纸业的高耗水、重污染行业，作为劳动密集型行业，在解决大量人口就业问题的同时，也对所在地区环境造成了严重污染。目前我国制革行业每年排放废水8,000~12,000万吨，废水中含铬约3,500 t，SS为1.2×105 t，COD为1.8×105 t，BOD为7×104 t，对水体污染严重。 本研究在对厌氧酸化工艺进行研究、一级好氧处理段进行工艺比选研究的基础上，获得了匀质调节—SBBR—BAF的生物处理工艺，并依托该工艺进行了生物强化处理的研究，考察了菌剂的强化运行效果及其处理水回用的可行性。 研究表明，在进水COD＞3,000 mg/L，厌氧酸化具有很好的抗冲击作用，保证了好氧工艺出水COD＜200 mg/L；在进水COD＜3,000 mg/L，可只通过好氧处理实现出水COD＜200 mg/L。厌氧酸化停留时间选择不当，会导致厌氧出水硫化物浓度升高，严重影响好氧系统，会使好氧活性污泥因中毒而解絮。 研究表明，当进水COD为2,000~2,500 mg/L，NH4+-N为130~146 mg/L时，COD、NH4+-N去除率SBBR分别为93.8%~96.6%和14.5%~55.9%，SBR分别为88.8%~94.9%和13%~50.7%，表明SBBR优于SBR。同时，研究发现SBBR污泥增长率为0.05 kgVSS/kgCOD，仅为SBR0.57 kgVSS/kgCOD的8.8%。此外，研究发现SBBR在停止运行后经3个运行周期可回复原油能力，而SBR池经9个周期培养也不能恢复，说明SBBR恢复能力明显优于SBR。 研究表明，以匀质调节—SBBR—BAF为主的制革废水处理工艺，出水水质稳定，进水COD 801~2,834 mg/L、NH4+-N 87~203 mg/L，出水COD＜80 mg/L、NH4+-N＜10 mg/L，基本达到中水回用标准；操作简单灵活，没有污泥回流系统，污泥产率低，污泥处理费用低；工艺基本不需要添加化学药剂，既节约成本、又避免了二次污染；两级生物膜使得该工艺具有很强的耐冲击负荷能力，特别适合制革废水水质水量波动大的特点。 研究表明，高效菌对系统的启动具有一定的促进作用，强化系统生物膜6天可以成熟，对照系统生物膜9天可以成熟。同时高效菌能加速COD降解，缩短停留时间，强化系统6~8 h可使COD＜200 mg/L，对照系统8~10 h可使COD＜200 mg/L。长期运行表明，强化系统的SBBR在COD和NH4+-N的去除率都优于对照系统的SBBR。最终出水COD强化系统平均为53 mg/L、对照系统为74 mg/L。在模拟循环过程中，强化系统均有更高的稳定性。可实现8次理论循环，而对照系统只能实现4次理论循环。 研究表明，通过合理的工艺设计，可以实现猪皮制革废水达到《污水综合排放标准GB8976-1996》一级标准，同时满足工厂部分用水要求。通过添加高效微生物，可提高生物处理系统处理能力，使处理水能够满足工厂的多次回用。 As a labour-intensive industry, tanning has created large amount of working opportunities as well as caused severe contamination to environment. And it is one of the highest water-consuming and polluting industry, only second to manufacturing. At present time, Chinese leather industry emits wastewater about 80,000,000~120,000,000 t annually, which contains chromium about 3,500 t, SS 1.2×105 t, COD 1.8×105 t, BOD 7×104 t and ambient riverhead has been polluted greatly. Based on the research of anaerobic acidification and comparison of SBBR and SBR, biotreatment process (Homogenization—SBBR—BAF) had been established to amend the disadvantages of traditional sewage treatment such as too much sludge, high cost of advanced treatment and NH4+-N can not reach the emission standard. Research on the bioaugmentation was also been carried out. Researches showed, when COD of influent was beyond 3,000 mg/L, anaerobic acidification could resist strong impact, thus COD of effluent was less than 200 mg/L; when COD of influent was less than 3,000 mg/L, only throughout aerobic sewage treatment could COD of effluent beless than 200 mg/L. False residence tiome of anaerobic acidification would lead to the higher effluent concentration of sulfide and disintegration of aerobic activated sludge. Researches showed SBBR worked a better than SBR: when influent between 2,000 and 2,500 mg/L, NH4+-N between 130 mg/L and 146 mg/L, COD, NH4+-N removal rate of SBBR was 93.3%~96.6%, 14.5%~55.9% respectively while COD, NH4+-N removal rate of SBR was 88.8%~94.9%, 13%~50.7% respectively. Sludge growth rate of SBBR was 8.8% of that of 0.05 kgVSS/kgCOD. Besides, SBBR could recovered after 3 operating periods while SBR worked no better after 9 operating periods.Therefore, SBBR excelled SBR. Researches showed, effluent quantity of tannery wastewater treatment process (Homogenization—SBBR—BAF) was stable. When COD of influent was between 801 and 2,834 mg/L, NH4+-N was between 87 mg/L and 203 mg/L, COD of effluent was less than 80 mg/L, NH4+-N was less than 10 mg/L, which achieved the standard of reuse. This biotreatment was featured in low cost, easy and flexible management, less sludge, no inverse sludge system. Besides, this technique required no chemical, which could lower the cost and avoid secondary pollution. Great resistant of impact due to two membranes and was suitable for tannery wastewater which was featured by fluctuation of influent quality and quantity. Researches showed effective microorganisms promotes the startup of the process.Biofilm in the bioaugmentation process matured with 6 days while biofilm in normal process matured with 9 days. Effective microorganisms could accelerate the degradation of COD and shorten the residence time. Aggrandizement system could make COD<200 mg/L with 6 to8 hours while cntrolling system could make COD<200 mg/L with 8 to 10 hours. Long-term operating shows that SBBR in the bioaugmentation system worked better than the normal system in the treatment of COD and NH4+-N. The average COC of effluent in bioaugmentation system was 53 mg/L, normal system was 74 mg/L. In the simulative circulation process,aggrandizement process, which could fulfill 8 times theoretical circulation, works more stably than controlling process which could only fulfill 4 times theoretical circulation. Researches showed that reasonable design could make the wastewater meet the first grade of discharging standard of National Integrated Wastewater Discharge Standard (GB8976-1996), and partially meet the demand of water using of the factory. Adding effective microorganisms could enhance the biotreatment and make the effluents reuse many times.

功能菌强化应急处理苯胺突发污染事故研究

近年来各种环境污染事故频发，据统计仅2001~2003年间，发生的各类环境污染事故就高达5606次，其中水污染事故3235次，占全部的57.7%。这些事故不仅给人民生命财产造成巨大损失，也给生态环境造成严重的破坏。因此开发安全高效的应急处理技术迫在眉睫。本研究以筛选高效苯胺降解菌为基础，通过对高效菌降解性能的研究指导将高效菌作为功能郡主投加到已有生物处理系统强化应急处理苯胺突发污染事故废液，取得了良好的效果。 苯胺高效降解菌AN-P1为红球菌（Rhodococcus sp.），其通过间位途径降解苯胺，AN-P1利用苯胺生长和降解的最佳pH为6，最适浓度为2000 mg/L，最适温度为30 ℃，最佳接种量为0.3‰。AN-P1降解含500 mg/L、1000 mg/L、2000 mg/L苯胺的培养物分别经过28 h、24 h、32 h降解，出水苯胺含量能达到《污水综合排放标准》（GB8978-1996）一级标准。但由于苯胺降解过程中释放了大量氨氮，出水氨氮仍较高未能达标排放。而常规SBR系统应急处理效果较差，苯胺和COD去除率均低于10%，出水未能达标排放。活性碳吸附后的回收和后续处理也会带来操作不变和二次污染问题，且处理后出水往往难于达标排放，尚需进行进一步处理。 生物处理系统应急处理后恢复运行处理效果监测和PCR-DGGE图谱分析显示，用AN-P1菌强化应急处理系统后不仅能快速高效的去除苯胺，而且可以有效保障处理系统对污染物的净化性能，有效的保护系统中的功能微生物免受苯胺毒害。 研究结果表明，从实际处理效果、对原有生物系统性能保护及实际应用操作等多方面考虑，用AN-P1菌强化应急处理苯胺突发污染事故在技术上都是可行的。本研究为应急处理苯胺突然污染事故废液提供了新的方法。 Recent years, environment pollution accidents happened frequently, the data showed that there are 5606 accidents between 2001 and 2003, including 3235 water environment accidents, which is 57.7% of all. These accedents not only caused money lost and life lost but also caused serious damage to the ecologicl environment. So exploring highly-effective and secure methods to solve these accidents is an urgent mission. We screened a highly-effective aniline-degrading bacterium and did some researches on its ability to degrade aniline, in order to guide the emergency treatment of aniline containing wastewater that caused by sudden accident pollution with bioaugmentation. A highly-effective aniline-degrading bacterium AN-P1 was isolate and characterized as Rhodococcus sp. It degrades aniline through meta-cleavage pathway. The optimal pH and temperature for cell growth and aniline degradation were 6 and 30 ℃, respectively, and the opitimal concentration of aniline was 2000 mg/L, the optimal inoculation amount was 0.3‰.It took bacterium AN-P1 only 18 h, 24 h and 32 h, respectively, for the treatment of MSB containing 500 mg/L, 1000 mg/L, 2000 mg/L aniline to meet the first grade of national some of the NH4+-N which caused by aniline degradation. It took bacterium AN-P1 only 10 h, 20 h and 32 h, respectively, for the treatment of wastewater containing 500 mg/L, 1000 mg/L, 2000 mg/L aniline to meet the first grade of national integrated wastewater discharge standard. The bacterium AN-P1 can also remove some of the NH4+-N which caused by aniline degradation. It took bacterium AN-P1 only 10 h, 20 h and 32 h, respectively, for the treatment of wastewater containing 500 mg/L, 1000 mg/L, 2000 mg/L aniline to meet the first grade of national integrated wastewater discharge standard. By combing AN-P1 with regular SBR system, it took only 36 h for the emergency treatment of wastewater containing 2000 mg/L aniline under simulating engineering conditions to meet the discharge standard. While the NH4+-N of effluent can not meet the standard because of the high amount NH4+-N caused by aniline degradation. The regular SBR system was not good at aniline and COD removal. The removal efficiency of which are less than 10%. It cost 67.8 g activated carbon to absorbed 1000 mg aniline. It is inconvenient to transport and use it for the emergency treatment of aniline when the sudden pollution accident happened. Meanwhile, it was complex ad hard to recycle the activated carbon and treat the aniline wastewater get from activated carbon recycling too. Hard to meet the effluent standard was also a problem of activated carbon absorption method. According to the PCR-DGGE profile and removal efficiency of pollutants and COD when the systerm recover from emergency treatment, AN-P1 can efficiently protect the microbial community of regular activated sludge system against the aniline. It proved that combing AN-P1 with regular biological system is a feasible strategy for emergency treatment of aniline sudden pollution accident. The research offered a new way for emergency treatment of aniline sudden pollution accident.

重离子束在微生物诱变育种及生物能源开发中的应用

作为一种新型的辐射诱变源,重离子束在辐射诱变育种中的优越性已经显现。在此基础上综述了重离子束用于微生物诱变育种的基本原理、独特优点、所取得的成果及研究进展,并对其在新型生物能源开发中的潜力进行了展望。

逆运动学弹性共振散射方法在非束缚核结构研究中的应用

简要介绍了近几年发展起来的厚靶逆运动学弹性共振散射方法在非稳定核结构测量中的应用。它是研究非束缚态核结构的实验方法之一。通过测量反冲轻核的激发函数,提取共振态的能量、自旋宇称和衰变宽度等。主要用于研究非稳定核素的结构、核天体物理中相关核的阈能共振态的能级参数测量等。

进一步指认近质子滴线核~(129)Pm的衰变

20 0 4年曾报道过利用4 0 Ca + 92 Mo融合蒸发反应产生了近质子滴线新核素12 9Pm ,并首次观测到它 (EC + β+)衰变产生的能量为 99keV的γ射线 .为了进一步确认上述指认 ,以下提供了三方面的新证据 :1)在 16 4— 190MeV能量范围内测量了 99keV衰变γ射线的激发函数 ;2 )进行了交叉反应36 Ar + 96 Ru的实验 ,观察到了相同的 99keV衰变γ射线 ;3)用Woods SaxonStrutinsky方法计算了12 9Pm的核势能面 ,其基态自旋宇称被预言为 5 / 2 - ,所以12 9Pm的 (EC + β+)衰变有利于馈送到子核12 9Nd的 5 / 2 - 的低位态 ,这也与前期报道相符 .

14MeV中子反应和远离核研究

指出了中子源的重要性,扼要地说明了加速器中子源的特点。简单地介绍了14 MeV中子引起的核反应,发现了14 MeV中子可以引起重核的奇异(n,2p)反应,并以此为基础,形成了合成和研究重丰中子新核素的一条物理思想和生成、分离鉴别技术路线;先后合成和研究了~(185)Hf、~(237)Th、~(175)Er和~(197)Os等四种重丰中子新核素。

2003年秋季西安大气中黑碳气溶胶的演化特征及其来源解析

IEECAS SKLLQG

一台300kV电子帘加速器辐照箱屏蔽物理设计

介绍一台电子能量为 30 0keV、束流强度为 6 0mA的电子帘加速器辐照箱的屏蔽物理设计。箱体厚度为 1.5cm的铅。

The stripper design and test at HIRFL-CSR

Charge stripping is employed to produce multi-charged ions for injecting the cooling storage ring After penetrating through the carbon foil, the widened distribution of ion charge states poses a limit to the ion injection Therefore, the carbon foil plays a key role in the charge snipping injection In this paper, foul strippers for Heavy Ion Research Facility at Lanzhou (HIRFL) and Cooling Sun age Ring (CSR) are introduced The charge state distribution of the stripped ions is measured and the stripping efficiency of the foils is investigated The experimental results are consistent with the theoretical values