Qingdao Institute of Bioenergy and Bioprocess Technology, Chinese Academy of Sciences
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NO reduction by CO over TiO2-gamma-Al2O3 supported In/Ag catalyst under lean burn conditions
No full textTiO2/gamma-Al2O3 supported In/Ag catalysts were prepared by impregnation method, and investigated for NO reduction with CO as the reducing agent under lean burn conditions. The microscopic structure and surface properties of the catalysts were studied by N-2 adsorption-desorption, X-ray diffraction, transmission electron microscopy, X-ray photoelectron spectroscopy, ultraviolet-visible spectroscopy, H-2 temperature-programmed reduction and Fourier transform infrared spectroscopy. TiO2/gamma-Al(2)O(3)supported In/Ag is a good catalyst for the reduction of NO to N-2. It displayed high dispersion, large amounts of surface active components and high NO adsorption capacity, which gave good catalytic performance and stability for the reduction of NO with CO under lean burn conditions. The silver species stabilized and improved the dispersion of the indium species. The introduction of TiO2 into the gamma-Al2O3 support promoted NO adsorption and improved the dispersion of the indium species and silver species. (C) 2016, Dalian Institute of Chemical Physics, Chinese Academy of Sciences. Published by Elsevier B.V. All rights reserved
N-Acylated chitosan bis(arylcarbamate)s: A class of promising chiral separation materials with powerful enantioseparation capability and high eluents tolerability
No full textIn order to comprehensively understand the influence of coordination of the substituent at 2-position with those at 3- and 6-positions on the properties of chitosan derivatives, a series of chitosan 3,6-bis(arylcarbamate)-2-(amide)s (CACAs) and the related chiral stationary phases (CSPs) were prepared and reported in the.present study. Specifically, chitosan was N-acylated with carboxylic acid anhydrides, and then further derivatized with various aryl isocyanates to afford CACAs, from which a class of coated type CSPs were prepared. When the substituent introduced on the acyl group at 2-position and those on the phenyl group of the carbamates at 3- and 6-positions were fittingly combined, these prepared CACAs based CSPs would exhibit powerful chiral recognition ability, further resulting in a class of promising chiral separation materials with excellent enantioseparation performance. Meanwhile, these newly developed materials with suitable molecular weight also bear a high tolerability towards organic solvents, even including pure tetrahydrofuran, thus broadening their application in enantiomeric separation. (C) 2016 Elsevier B.V. All rights reserved
Pyrolysis kinetics of invasive coastal plant Spartina anglica using thermogravimetric analysis
No full textThe pyrolysis kinetics of invasive plant Spartina anglica biomass was investigated via a thermogravimetric (TG) analyzer from 50 to 800 degrees C in an inert argon atmosphere at different heating rates of 5-30 degrees C/min, and the kinetic parameters were deduced by Starink and master-plots methods. The results showed that three stages appeared in the thermal degradation process, and the mean value of activation energy was calculated to be 291.57 kJ/mol by the Starink method. It suggested that the most probable pyrolysis kinetic model was g(a) = [-ln(1-a)](4) deduced by the master-plots method, which meant a random nucleation and nuclei growth mechanism. These results provided fundamental useful information for describing the pyrolysis process, and the subsequent designing and running of a pyrolytic processing system using S. anglica as feedstock
雨生红球藻再生培养模式的构建及相关生理学的基础研究
No full text虾青素具有良好的着色性能和抗氧化性能，在食品、饲料、药品等领域有广泛的应用。雨生红球藻是目前最安全、高效的虾青素来源，培养雨生红球藻生产虾青素具有极高的经济和社会效益。培养雨生红球藻用于虾青素生产已有数十年的历史，然而，目前工业规模生产的技术仍然停留在二十年前的水平。我国是雨生红球藻虾青素的新兴市场，国内产业化过程处在高速发展阶段，但是生产技术方面的发展落后于国外厂商，因此急需技术革新和发展自主知识产权的生产技术。
针对这一现状，本论文基于前期工作观察到的雨生红球藻红色游动细胞在形态、生化组分和细胞分化方面的特殊性，认识到这一形态的细胞在雨生红球藻生活周期中可能起着相当重要的作用，对其发生和发展的过程进行深入研究有可能提供新的培养模式。本论文首先通过研究孢囊化诱导条件、萌发过程的氮、光强、初始密度等环境因素对厚壁孢子萌发率和同步化程度影响的关系，建立雨生红球藻厚壁孢子萌发和子细胞再孢囊化过程的同步调控方法。随后，综合运用显微观察和生理生化性状测定等手段，阐明红色游动细胞形成过程中脂肪-蛋白质-碳水化合物的代谢过程和色素的变化规律。之后，基于建立的细胞周期同步调控方法制备红色游动细胞群，通过研究红色游动细胞在高光、极端温度等环境逆境下的光合作用强度、存活率和生长等生理指标，评价红色游动细胞对不利环境的耐受性。最后，发展基于厚壁孢子同步萌发的雨生红球藻再生培养新模式，并开展室内室外不同规模的培养验证和工艺优化。通过上述研究，本论文获得如下的主要结果：
首先，建立了雨生红球藻厚壁孢子萌发和子细胞再孢囊化过程的同步调控方法。在最优条件下，雨生红球藻厚壁孢子的48 h萌发率达到90%以上，萌发产生红色游动细胞的再孢囊化效率达到100%。这一调控效果可以在多个连续的世代培养过程中实现。研究结果表明吸收足够量氮是诱导雨生红球藻厚壁孢子萌发的必要条件，光照强度通过影响氮的吸收速率而调控厚壁孢子的萌发进程，并通过快速消耗培养基中的氮和结合光诱导共同作用来诱导新生的红色游动细胞进入再孢囊化过程。研究还发现雨生红球藻厚壁孢子成熟度与其萌发效率负相关，Car/Chl比值可作为衡量雨生红球藻厚壁孢子成熟度的一个简易指标。
其次，碳水化合物和脂肪可能是雨生红球藻厚壁孢子中同等重要的碳储存物质。在萌发过程中，首先代谢碳水化合物用于孢子囊形成阶段的能量需求，之后代谢脂肪用于游孢子释放阶段的能量需求，并为蛋白质和结构性脂肪的快速合成提供前体。光合作用只能延迟萌发过程的脂肪代谢，却不能阻止油脂储存物质的代谢。研究结果表明，雨生红球藻通过快速合成叶绿素和降解胞内虾青素生成叶黄素，提高细胞对光量子的吸收，并提供脂肪酸作为萌发过程的部分能量来源。虾青素的遮光效应和叶黄素循环的耗散作用共同对厚壁孢子萌发过程的孢子囊和子代游孢子行使光保护功能。
第三，本研究证明了一种以厚壁孢子为初始材料的循环再生培养新模式。在这一培养模式中，厚壁孢子萌发释放游孢子和新形成游动细胞的再孢囊化过程可以在同一环境条件下连续进行；同时再孢囊化形成的新一代厚壁孢子保留了再次萌发的能力，作为下一批次培养的接种物。此外，高光强和氮限制对维持这一生活周期的快速运转至关重要。
第四，雨生红球藻红色游动细胞较绿色游动细胞具有更强的高光耐受能力和更大的光合作用容量，这一特殊性质与其胞内虾青素储存物的遮光和ROS清除作用，以及其活跃的虾青素合成过程消耗光合作用产生的过量活性氧有关，同时红色游动细胞可以快速转化为抗逆性更强的中间态不动细胞palmella也对其具有更强的高光耐受能力有贡献。但是，本研究对红色游动细胞的不同光保护途径，比如光化学淬灭、非光化学淬灭，以及酶淬灭系统的贡献额度尚不清楚，需要更多的工作进行深入研究。研究还发现，雨生红球藻可在冰点温度下存活并保留复苏生长的能力，而对高温的耐受性，尤其是未萌发的子细胞对高温的耐受性较弱。
第五，本研究提供的再生培养新模式在室内室外实施可获得高虾青素含量的雨生红球藻生物质，在室内获得的最高生物质产率和虾青素产率分别为0.498 g/L/d和24.1 mg/L/d，在室外培养获得的最高生物质产率和虾青素产率分别为0.327 g/L/d和11.5 mg/L/d。氮消耗速率(SNI)决定再生培养模式进入再孢囊化阶段的时间点，这一指标取决于氮浓度、光照强度和初始接种密度三者的交互作用；再生培养模式的再孢囊化阶段中，虾青素产率仅取决于培养光强和细胞密度。研究还发现室外培养下的光能利用效率要比室内至少低50%，通过与新技术结合可能有助于提高室外环境下的生物质产率和虾青素产率。Astaxanthin has being widely used as food, feed and drugs, due to its superior performance as coloration feed additives and antioxidant activities. Microalga Haematococcus pluvialis (Chlorohyceae) is by far the most safe, efficient source of astaxanthin, therefore to culture this microalga for astaxanthin production has a very high economic and social benefits. Cultivation of H. pluvialis for astaxanthin production has a long history; however, the technology in industrial scale still remains at the level of 20 years ago. China is an emerging market for astaxanthin of H. pluvialis and the industrialization process stands in a stage of rapid development, but the production technologies lag behind foreign manufacturers. Therefore, production technologies of H. pluvialis, of a high level and with independent intellectual property rights, will be required.
Previously, we observed special characteristics in the morphology, biochemistry and cell transformation of H. pluvialis red zoospores. Based on this observation, we recognize the red zoospores may play an important role in its life cycle, and an in-depth study about the development process of red zoospores may help to build a novel cultivation mode. In this study, the effects of environmental factors related to encystment and germination, i.e. nitrate concentration, light intensity and initial cell density, on the germination rate and synchronization level of H. pluvialis hematocysts were investigated firstly, which may help to build an artificial control method for synchronous germination of hematocysts and synchronous encystment of red zoospores. Then, we try to clarify the metabolic patterns of cellular fat, protein, carbohydrate and pigments during the formation process of red zoospores, via a multiple observations of microscopic examination and physiological and biochemical study. Cultures, which contained synchronous red zoospores, were prepared through the aforesaid cell cycle synchronization method, and latterly were studied to evaluate tolerance to adverse environment, via analysis of photosynthesis, survival rate and growth under high light and/or adverse temperature. Finally, a novel mode for growing H. pluvialis, namely regeneration cultivation mode, was proposed, and the applications of this mode in outdoor and indoor conditions were evaluated in different scales. The achieved main results were as follows:
Firstly, an artificial control method for synchronous germination of hematocysts and synchronous encystment of red zoospores was developed. A 90% of hematocysts germination rate at 48 h and a 100% of red zoospores encystment rate were achieved at optimum conditions. The aforementioned efficiencies could be achieved in continues several generations. The results also suggested that absorbing a certain amount of nitrogen was required to induce hematocysts germination, which could be adjusted by light intensity. The transformation between germination of hematocysts and encystment of red zoospores was controlled by interaction of light intensity and nitrogen consumption. The results also suggested that the germination rate was negatively associated with the maturation degree of hematocysts, which could be simply reflected in ratio of total carotenoids to total chlorophyll (Car / Chl).
Secondly, both carbohydrate and fat were equally important as main carbon reserves of H. pluvialis hematocysts. Mobilization of carbohydrate firstly appeared to meet energy requirement for formation of sporangium, and then the mobilization of fat appeared subsequently to satisfy energy requirement for releasing red zoospores and to provide precursors for synthesizing proteins and structure lipids. The mobilization of fat could be delayed, but cannot be replaced, by photosynthesis. The results suggested the photosynthesis could be enhanced with increasing of chlorophyll and lutein, which was a product of degradation of astaxanthin, to provide fatty acid as energy for germination. Both sun-shading effect of astaxanthin and xanthophyll cycle play important functions to protect sporangium and red zoospores during germination of H. pluvialis hematocysts.
Thirdly, a novel regeneration cultivation mode to grow H. pluvialis with hematocysts as inoculums was proposed, in which the reproduction of red zoospores by hematocysts germination and the encystment of daughter cells sequentially took place in a same cultivation condition. The newly formed hematocysts retain ability to germination, and thus could be used as inoculums for next batch of cultivation. Moreover, conditions of high light and nitrogen limitation were critical to success of this cultivation mode.
Fourthly, red zoospores were more resistant than green zoospores to high light, which could be owe to sun-shading and ROS scavenging effect of cellular astaxanthin, and ROS consumption via active synthesis of astaxanthin, as well as the ability to transform into palmella, which had even higher resistant. However, a further study was required to clarify the distribution among different protection strategies, including photochemical quenching, non-photochemical quenching and scavenging enzyme system. Our results also suggested the H. pluvialis could be resistant to long period of frozen stress, but can not resistant to high temperature stress, especially for its sporangium and red zoospores.
Lastly, H. pluvialis biomass with high content of astaxanthin was achieved through this proposed regeneration cultivation mode in different scales of outdoor and indoor cultivations. The best biomass productivity and astaxanthin productivity from indoor cultivation were 0.498 g/L/d and 24.1 mg/L/d, respectively. The best biomass productivity and astaxanthin productivity from indoor cultivation were 0.327 g/L/d and 11.5 mg/L/d, respectively. The process for re-encystment was controlled by specific nitrogen input (SNI), which was adjusted by interaction of nitrate concentration, light intensity and cell density. Moreover, there is a positive correlation between the astaxanthin productivity and interaction of light intensity and cell density. Also, the results suggested the light energy efficiency is at least 50% less for outdoor condition than indoor condition
乙醇型发酵产氢菌特性及其生物强化技术研究
No full text随着资源短缺、能源危机和环境污染等问题的加剧，人们逐渐认识到厌氧发酵技术（既能消除污染又能产生可再生能源：氢气或沼气）的重要性。在厌氧生物处理过程中，微生物处于整个工艺的核心地位，直接决定着发酵效率的高低。因此，选取高效关键菌株，优化菌群结构，对提高微生物菌群的活性及秸秆厌氧发酵产氢产甲烷的效率具有重要意义。基于本实验室分离得到的一株糖类发酵产氢产乙醇乙酸菌Hydrogenispora ethanolica LX-B，本课题研究了该菌株的产氢生理特征，并将该菌株投加到秸秆沼气发酵系统中，即使用生物强化法提高了秸秆甲烷产量，实验结果表明，该菌株具有较高的产氢效率，投加到秸秆沼气发酵系统中，实现了在低能耗、无污染、条件温和的反应过程中提高秸秆降解率和产能效率的目的。其主要研究内容和结果如下：
1、 以一株乙醇型发酵产氢厌氧细菌Hydrogenispora ethanolica LX-B为研究对象，首先研究了其对底物葡萄糖浓度及其产物乙酸、乙醇浓度和氢分压的耐受性，实验结果表明，该菌株具有产氢速率高、生长速率快、对底物和产物耐受性高的优点。该菌株能在高达41.5 g/L的葡萄糖底物浓度下生长，在1.1 g/L葡萄糖，温度37 °C，初始pH 7.0条件下，最大产氢率为1.8 mol H2/mol glucose，同时产生乙醇1.4 mol /mol glucose和乙酸0.8 mol /mol glucose（电子转化率为 114 %）。能耐受100 mM产物乙酸和乙醇（抑制率＜ 20%），且在较高的氢分压（27 mmol/L）下，能维持正常的生长和产氢；其次，以木质纤维素典型糖类组分为碳源，评价了Hydrogenispora ethanolica LX-B利用不同糖类产氢的情况，结果表明，该菌株能利用戊糖、己糖和多糖等生长，其中，葡萄糖、木糖、阿拉伯糖、甘露糖和半乳糖是较适合的碳源；同时研究了该菌对不同浓度热-NaOH碱处理玉米秸秆的利用性，研究发现，使用0.25% NaOH预处理，最大产氢量为67.8 mL/g-corn straw，比对照组提高了21.9%（对照组产氢量为55.6 mL/g-corn straw)。
2、 在研究了Hydrogenispora ethanolica LX-B产氢特性的基础上，为提高玉米秸秆厌氧发酵产甲烷的效率，在发酵系统中投加该菌株，探究了其对玉米秸秆沼气发酵的生物强化作用。结果表明，H. ethanolica使玉米秸秆厌氧发酵的甲烷产量提高16-18%，当5%接种时，最大甲烷产量达到233.8 mL/g-corn straw。H. ethanolica使甲基纤维素和木聚糖甲烷产量分别提高了21.7%、14.2%。推测H. ethanolica最可能的强化机制为：H. ethanolica将更多的木质纤维素生物质的水解产物，像五碳糖和六碳糖，转化为乙醇、乙酸和氢气，一方面促进了秸秆中纤维素和木聚糖的降解，另一方面为产甲烷菌提供了更多的底物。With the increasing of resources shortage, energy crisis and environmental pollution, much attention has been focusing on anaerobic digestion (owing to its high pollution removal ability and renewable energy generation: hydrogen or biogas). Microbes are the main vehicle for anaerobic digestion, which determine the production efficiency. Therefore, it is significant to pay attention to exploitations of highly more efficient microorganisms and optimize the fermentation conditions, which is responsible for the efficiency of anaerobic technology. Previously, we isolated a hydrogen producing ethanol-type bacterium Hydrogenispora ethanolica LX-B. In this study, the characteristic about anaerobic biohydrogen of this bacterium, application the bioaugmentation technology into the anaerobic digestion of corn straw was investigated. Results showed that the strain possess high hydrogen production efficiency, application into the anaerobic digestion of corn straw could accelerate the degradation of straw and improve production efficiency in the condition of energy saving, environmental friendly and mild conditions. Main contents and results of the present study are as follows:
1. The biohydrogen production potential of Hydrogenispora ethanolica LX-B was studied. Firstly, the effect of glucose and products inhibition by acetate, ethanol and hydrogen on the growth and hydrogen production was assayed. Results showed that Hydrogenispora ethanolica is a promising strain for hydrogen production with relatively high hydrogen yields, high growth rate, a wide rage of substrate utilization and production tolerance. The bacterium grew well and kept high hydrogen yield in a wide range of initial glucose concentrations up to 41.5 g/L. The maximal yields of 1.8 mol H2/mol glucose and 1.4 mol ethanol/mol glucose were observed at 1.1 g/L glucose at 37 °C and pH 7.0. Neither acetate nor ethanol exhibited significant inhibition: the addition of ethanol or acetate up to 100 mM only had less than 20% inhibition on the growth and glucose consumption during two weeks of incubation. Besides, under high hydrogen partial pressure of 27 mmol/L culture, the growth and glucose consumption of the pure culture was not inhibited. Secondly, hydrogen production ability on various sugars and corn straw was also investigated. Results demonstrated that the strain could grow on pentose, hexose and polyose, in which glucose, xylose, arabinose, mannose and galactose are the preferred carbon source. Besides, the strain would be an ideal candidate for hydrogen production with cellulosic biomass. The maximum H2 yield of 6.8 mL/g was achieved by NaOH-pretreated corn straw, which was 21.9% higher than the control (5.6 mL/g-corn straw).
2. The bioaugmentation effect of H. ethanolica LX-B on anaerobic digestion of corn straw was studied. Results showed that bioaugmentation with the strain could increase 16-18% methane yield of corn straw degradation. The maximum methane yield was 233.8 mL/g-corn straw with 5% inoculation. H. ethanolica could improve the methane yields from methyl cellulose and xylan (models for cellulose and hemicelluloses, respectively) by 21.7% and 14.2%. A proposed mechanism is that H. ethanolica promotes more hydrolysates of lignocellulosic biomass (such as pentose and hexose) to ethanol, acetate and hydrogen, and provides a source of substrates for the methanogens. As a result, the degradation of cellulose of corn straw were improved
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No full text有机微孔材料是一类孔尺寸小于 2 nm的新型孔材料，其具有比表面积大， 化学稳定性高，比容高等优点，在存储分离中具有潜在的应用价值，引起了科学家的热切关注。有机多孔材料主要由 C，H，O，N，B等轻质元素组成，与传统的无机材料相比较，可以在较大范围内通过化学修饰的方法实现材料骨架结构的改变，从而可实现气体的高密度存储。本论文的工作主要是设计合成了两类共轭微孔聚合物，并通过固体核磁，氮气吸附等温线表征聚合物基本结构，通过CO2，H2，CH4，在不同温度下的吸附等温线研究各种聚合物对上述气体的吸附存储，及选择分离能力。论文主要包括以下内容：第一章：微孔有机聚合物的综述。第二章：四个咔唑基共轭微孔聚合物PCz-Cn-Cz(n=3-6)由FeCl3氧化偶联聚合得到。这四个聚合物的BET比表面积分别为862 m2 g-1、870 m2 g-1、768 m2 g-1和785 m2 g-1，有趣的是，尽管这四个聚合物的结构是由不同的柔性亚烷基链连接刚性的咔唑基团构成的，但是它们的孔径分布都集中在0.5 nm处，我们推测这可能是因为柔性的亚烷基链容易扭曲，导致分子内的夹层和孔隙填充。气体吸附曲线表明，在77 K/1 bar，最大的H2吸附量是1.33 wt%；273 K/1 bar，CO2的吸附量可达到16.8 wt%；273 K/1 bar，CH4的最大吸附量是2.11 wt%；298 K/1 bar，CO的吸附量可达到1.37 wt%。273 K，CO2 / N2和CO2 / CH4的最大分离比分别是47.7和14.0；298 K，CO2 / N2和CO2 / CH4的最大分离比分别是33.8和7.3。第三章：四种共价三嗪骨架聚合物（PCTF-1至PCTF-4）在ZnCl2的催化下用离子热反应聚合而成，前三种聚合物的BET表面面积分别为853 m2 g-1、811 m2 g-1、391 m2 g-1，与支链的长度相悖，这表明单体的支链越长，能够更有效堆积，从而导致材料的密度更高，表面积更低。PCTF-4与PCTF-2相比，仅仅是把PCTF-2支链上中间的苯换成了苯并噻二唑，然而N2吸附曲线测得PCTF-4的比表面积为1404 m2 g-1，接近PCTF-2的两倍。富氮的C3N3环与CO2的作用力强，增加了对CO2的吸附，特别是PCTF-4，在273 K/1 bar，CO2的吸附量是20.5 wt%，这个值在共价三嗪骨架聚合物中是最大的，这个结果表明，强极性基团(噻二唑)引入到聚合物骨架是一种有效的策略来增强微孔有机聚合物与CO2分子的作用力。另外，这些共价三嗪骨架聚合物有着很好的物理化学稳定性，在273 K显示了很高的CO2 / N2和CO2 / CH4分离比，最高分别是14-56，11-20。然而,在水蒸气的存在下,这些材料的CO2吸附量都减少了，这可能是由于聚合物与水形成了氢键作用，这个现象表明在干燥条件下表现良好的材料在更接近实际的条件下可能不是最有前途的材料。第四章：本论文总结。Microporous organic polymers (MOPs) can be defined as a series of porous materials with pore sizes smaller than 2 nm on average, which are comprised of light elements such as C, H, O, N, B and exhibit very high physical surface areas. To compare with inorganic pore materials the MOPs can be easily chemically modified the backbone of the bulk phase and show good performance in gas storage. Part I: a general over view of the microporous organic polymers.
Part II: Four carbazole-spacer-carbazole polymers PCz-Cn-Cz (n=3-6) with the similar topological model structures were designed and prepared by FeCl3 oxidative coupling polymerization. The Brunauer-Emmett-Teller (BET) specific surface areas of the obtained polymers are 862, 870, 768 and 785 m2 g-1, respectively, which are competitive with the reported conjugated microporous polymers. Interestingly, there are no obvious differences in the domain pore width (centred at 0.5 nm) and the pore size distribution among the four polymers, although they have different length soft alkylene chains to interlink same rigid backbone carbazole. This may be ascribed to the soft alkylene chains, which are easy to bend and result in intramolecular intercalation and pore filling. Gas (H2, CO2, CH4 and CO) adsorption isotherms show that the H2 storage of the polymers can be up to 1.33 wt% at 1.0 bar and 77 K, the uptake capacity for CO2 can reach 16.8 wt% at 1.0 bar and 273 K, the CH4 uptake can reach 2.11 wt% at 1.0 bar and 273 K and the CO uptake performance can be up to 1.37 wt% at 298 K and 1.0 bar. Selective adsorption of CO2/N2 and CO2/CH4 calculated using the initial gas uptake slopes shows that these networks display good selectivity with a maximum value of 47.7 (33.8) and 14 (7.3) at 273 K (298 K).Part III: Four covalent triazine-based frameworks (PCTF-1 to PCTF-4) were synthesized by a consolidated ionothermal reaction between aromatic nitriles under the catalysis of ZnCl2. The Brunauer-Emmett-Teller (BET) specific surface area values of PCTF-1 (853 m2 g-1), PCTF-2 (811 m2 g-1) and PCTF-3 (391 m2 g-1) are against with the increasing length of branched arm, indicating using monomers with longer branches are able to pack more efficiently, resulting in higher density materials with a lower surface area. PCTF-4, compared with PCTF-2, just changed the middle benzene of the branches to benzothiadiazole, however, N2 adsorption isotherms showed its BET specific surface area value (1404 m2 g-1) is the highest among the PCTFs, almost two times than that of PCTF-2. The nitrogen-rich characteristics of C3N3 triazine rings feature the frameworks strong affinity for CO2 and thereby high CO2 adsorption capacity. Especially for PCTF-4 with the benzothiadiazole, it exhibited the highest CO2 uptake (20.5 wt%) at 273 K and 1 bar, this value is one of the highest reported for covalent triazine-based frameworks. The results demonstrate that the introduction of strong polar groups (benzothiadiazole) into a polymer skeleton is an efficient strategy to produce CO2-philic microporous organic polymers with enhanced binding affinity with CO2 molecules. In addition, such PCTFs with high physical-chemical stability and comparable BET surface areas exhibited good ideal CO2/N2 selectivities (14-56) and CO2/CH4 selectivities (11-20) at 273 K, showing these materials are potential candidates for gas storage and separation. However, in the presence of water vapor, these performance of CO2 uptake all decreased due to via hydrogen bonding with water, suggesting materials that perform well in dry conditions may not always be the most promising materials under more practical conditions.
Part IV: the conclusion of the thesis
霉酚酸微生物合成和P450单加氧酶与还原伴侣蛋白的相互作用机制研究
No full text本报告围绕微生物生物合成霉酚酸（MPA）和 “微生物生物合成P450单加氧酶与还原伴侣蛋白的相互作用机制”研究课题展开工作，本报告为为下一步的课题进展提供了平台。
内容包括以下三个方面：
1. 在酿酒酵母宿主中表达来自短密青霉的霉酚酸合成相关基因
a) 利用酿酒酵母同源重组系统组装大片段DNA的技术路线，证明了该技术在理性设计组装外源DNA和代谢工程改造酿酒酵母的可行性、高效性和可重复性；
b) 分子克隆了来自短密青霉的霉酚酸（MPA）合成相关基因mpaA，mpaDE和mpaC的。针对mpaC突变株开展了发酵实验，通过HPLC未能检测到代谢产物5-MOA；三个蛋白均利用SDS-PAGE和Western Blot检测了蛋白表达情况。
c) 对MPA合成相关基因mpaA进行了密码子优化并蛋白纯化鉴定。
2. 国家优青项目“微生物生物合成P450单加氧酶与还原伴侣蛋白的相互作用机制”研究课题
a) 蓝细菌来源的6个还原伴侣Fdx的分子克隆和蛋白纯化。
b）克隆表达来自天然色链霉菌的18个P450酶基因和6个铁氧化还原蛋白Fdx、4个铁氧化还原蛋白还原酶FDR，对Fdx6密码子优化，并进行了蛋白纯化鉴定. P450和FDR进行了光谱分析。
c）分别采用黄体酮、睾酮、薯蓣皂苷元、丹参酮IIA、麦新米星IV(M-IV)和YC17等多种底物对这些P450和还原伴侣进行了酶催化反应筛选鉴（HPLC检测）。首次发现了天蓝色链霉菌的还原伴侣蛋白用于催化支撑PikC催化YC17合成产物。并且证明在支撑PikC的效力层面，不同组合的还原伴侣可以达到不同的催化效率，合成不同的目的产物。为进一步获得对“P450酶—Fdx—FdR”三蛋白系统相互作用和识别机制的规律性认识做出积累。
3. 其他：对序列未知的米曲霉表达载体pTAex3进行了全测序并获得了图谱等。The report is focused on the microbial biosynthesis of mycophenolic acid (MPA) and National Excellent Youth Project "The interaction mechanism of the microbial biosynthesized P450 oxygenase and reducing chaperones”. This report provides a platform for further research of the subjects.
There are mainly three aspects:
1. Expression of genes involved in MPA biosynthesis from Penicillium brevicompactum using Saccharomyces cerevisiae as a host
A) A homologous recombination based DNA assembler technology was used for construction of large DNA fragments in Saccharomyces cerevisiae. This method is proved feasible, efficient and reproducible for the rational design of metabolic engineering pathways.
B) The mpaA, mpaDE and mpaC gene in for MPA synthesis in Penicillium brevicompactum strain NRRL864 were molecularly clonned. Through fermentation experiments, the mpaC mutant strain showed no production of the 5-MOA through HPLC/LC-MS detection. To detect protein expression of the three proteins, SDS-PAGE and Western blot were carried out.
C) mpaA was codon optimized and the protein was purified and verified.
2. The projiect "The interaction mechanism of the microbial biosynthesized P450 oxygenase and reducing chaperones”.
A) Molecular cloning and protein purification of the 6 reduced chaperone Fdx from the cyanobacterium strain Synechococcus elongatus PCC7942.
B) Cloning and expression of the 18 P450 genes and 6 ferredoxins (Fdx), and 4 ferredoxin reductases (FDRs) from Streptomyces coelicolor A3(2). Fdx6 was codon optimized. The P450s were verified by CO-assay. The four FDR were analyzed using a spectral analysis.
C) Progesterone, testosterone, tanshinone IIA, diosgenin, mycinamicin IV (M-IV), and YC17 were used as substrates for the reaction by P450 and reducing partners (based on HPLC screening). It was discovered for the first time that the reductive chaperone protein of Streptomyces sp. are able to support PikC to catalyze YC17. And it is proved that the different combinations of the reducing partners have various catalytic efficiencies and can achieve different products. This helps to the understanding of the regularity of the interaction and recognition mechanism of the "Fdx - FdR - P450 system.
3. The expression vector in Aspergillus oryzae, pTAex3, was sequenced and the map was achieved
米糠资源高值化综合利用关键技术的研究
No full text本文采用双酶法水解米糠淀粉，并分别对液化工艺和糖化工艺进行了研究。通过对液化工艺进行单因素实验，考察了酶用量、醪浓度、pH、温度对液化工艺的影响，通过实验得到了较优的液化工艺条件为：酶用量为0.1%、醪浓度为25%、pH=6.0、温度为90℃。在单因素试验的基础上，利用Design Expert 8.0.6软件中的Box-Behnken试验设计对米糠液化工艺进行了进一步的研究及优化，得到了最优的液化工艺条件：酶用量为0.11%、醪浓度为25%、pH=6.0、温度为88℃。在此条件下，对模型的预测参数进行了三次平行验证试验，得到液化DX平均值为6.54%，与模型预测值较接近，说明Box-Behnken设计优化得到的液化工艺条件是可靠的。In this paper, the rice bran starch was hydrolyzed by double enzyme method, and the liquefaction and saccharification process were studied respectively. Through the study of the single factor experiment of liquefaction process, the effects of enzyme dosage, mash concentration, pH, temperature was studied. The optimum liquefaction process conditions as follows: 0.1% dosage of enzyme, mash concentration 25%, pH=6.0, 90℃. On the basis of single factor experiment, the liquefaction technology of rice bran starch was further researched and optimized by the box-Behnken design of design expert software 8.0.6, and the optimal liquefaction process conditions were obtained: 0.11% enzyme dosage, mash concentration 25%, pH = 6.0, temperature 88℃. Under this conditions, the forecast parameters of the model were three parallel experiment, and finally the liquefied DX average was 6.54%, and this valve was close to the model prediction, that the liquefaction conditions optimized by box-Behnken design was reliable
新型糖苷水解酶的结构和功能研究
No full text发展可再生能源是当前人类社会缓解能源危机，减少温室气体排放的重要应对策略之一。木质纤维素是一种重要的可再生资源，但是其存在一些固有的抗降解屏障，使其低成本的大规模应用目前还很难实现。木质纤维素等多糖类生物质资源的利用离不开糖苷水解酶的作用。糖苷水解酶是一类可以降解糖苷键的酶类,随着糖类生物学地位的日渐突出，糖苷水解酶的发现与研究也越发重要。为了阐明新型的糖苷水解酶的作用机理，本论文利用蛋白质晶体学的相关技术，对它们进行结构功能的研究。本实验室的前期工作已经通过核磁共振方法解析了CohA和DocA的单独溶液结构，通过表面等离子共振技术对CohA-DocA之间的相互作用强度进行了测定。本论文在此基础上进一步研究了CohA-DocA之间的相互作用对纤维小体组装与组装后功能的影响。以解纤维梭菌的粘连模块（CohC）和对接模块（DocC）作为对照组。本论文的工作主要包括对所需组件的基因克隆与蛋白表达纯化，实验分析相互作用对组装的影响以及对组装后复合物协同效应的影响。
（1）微拟球藻胞内糖苷水解酶的结构功能研究
微拟球藻是一种重要的产油微藻，作为微藻产油研究的模式藻株，研究者已经对其开展了大量的研究。此前的研究报道认为，微拟球藻胞内的物质存在转化通路，特别是多糖类物质可以转化为油脂储存。该过程中糖苷水解酶起着至关重要的作用。本论文选择了微拟球藻胞内转录水平最高的一种糖苷水解酶g6403进行了相关研究。生物信息学分析表明该酶是GH1家族的β-葡萄糖苷酶，该酶是定位于微拟球藻线粒体上的，与其物质能量代谢密切相关。通过蛋白质晶体学技术解析了g6403的蛋白结构，它有典型的糖苷水解酶的(β/α)8结构。其底物结合口袋与之前研究过的酶有一定的差异，可能与其独特的底物降解偏好性相适应。进一步对其酶学性质的研究表明其最适的酶活作用条件是与其细胞定位高度适应的。
（2）地衣聚糖酶F32EG5与底物复合物的结构研究
F32EG5是首次报道的GH5家族的地衣聚糖酶。为了揭示其独特的底物识别和催化机理，本研究论文解析了其无活性突变体E188Q和纤维四糖底物G4的复合物的晶体结构。结果表明，底物G4结合在酶的-3到+1位。F32EG5在-1和-2位对G4底物有严格的识别作用，在-3位和+1位的结合比较弱。在-1位的糖环呈现出扭曲的船式构象，糖链发生了较大角度的扭曲，这可能是其对β-1，3降解偏好性的重要原因之一。
（3）哈氏噬纤维菌纤维素内切酶CHU2103的结构研究
哈氏噬纤维菌可以实现对纤维素的高效降解，但是其纤维素降解机制既不同于厌氧菌的纤维小体机制，也不同于真菌的游离酶系，并且其编码的纤维素酶缺乏CBM结构域，这都预示着其有独特的纤维素降解机制。CHU2103是哈氏菌编码的一种进行性内切酶，本研究通过解析其晶体结构，发现它有一个很浅的底物结合裂隙，且其中的芳香族氨基酸的数目特别多，这可能是其缺乏的CBM的一种适应性机制。另外，我们发现其活性中心有一个较深的口袋结构，不同于常见的纤维素内切酶，我们推测其具有纤维素外切活性，导致其产物中可溶性还原糖明显多于不可溶性还原糖。
本论文对进行性纤维素内切酶CHU2103，地衣聚糖酶F32EG5+G4，以及β-葡萄糖苷酶进行了结构解析，阐述了它们独特的生化性质的结构基础，为对其进行进一步的性质研究和功能改造提供了重要的理论依据和基础。The global energy crisis and dioxide emission have demanded development of renewable energy. Lignocellulose is an important renewable resource, however it is hard to be used at a low cost due to its inherent anti-degration barrier. Glycoside hydrolases, which can hydrolyze the glycoside bond, have important applications in the utilization of lignocellulose. Glycoside hydrolyases are becoming more and more important according to the developments of carbohydrates biology. Protein crystallography is used to solve the structures of 3 novel glycoside hydrolases, and structure and function studies were done furtherly to elucidate the catalytic mechanisms of these enzymes in this dissertation.
(1) structure and function of a glucoside hydrolase from Nannochloropsis
Nannochloropsis spp. are emerging as a model system for oleaginous microalga on research and industrial production. Previous studies have shown that cabarhydrate could be converted to triacylglycerol (TAG) as storage material. Glycosidases play important roles during the transformation of carbohydrates. In this dissertation, glycosidase g6403 was taken as research object, whose transcriptional level was among the highest ones. Bioinformatics studies showed that this β-glucosidase was belonged to GH1 family and located at mitochondria. The crystal structure of this enzyme revealed that it had a typical (β/α)8 structure. There were some sructural differences between g6403 and homologous proteins at the substrate binding pocket which was an adaption of the substrate specificity. Characterization of g6403 showed that this enzyme adapted the marine environments well.
(2) structure of lichenase F32EG5 with ligand
F32EG5 was the first reported lichenase that belonged to GH5. The crystal structure of inactivated mutant F32EG5 (E188Q) with a ligand cellotetraose (G4) was solved to elucidate the catalytic mechanism of this enzyme. The substrate G4 binding in the catalytic cleft of F32EG5 fron positon -3 to +1. The recognition of substrate at -1 and -2 was quit strictly while relatively weak at subsite -3 and +1. The substrate adopts a skew-boat conformation at subsite -1 and the extend direction of the glycan change with a large angle. This might be the reason of the substrate preference of F32EG5.
(3) structure of a processive endo-cellulase CHU2103 from Cytophaga hutchinsonii
Cytophaga hutchinsonii can efficiently degrade cellulose by an unknown strategy, which is quite different from the two well-studied strategies of cellulose degradation for cellulolytic microorganisms. The aerobic fungi degrade cellulose by secreting a series of free cellulases, while most anaerobic cellulolytic microorganisms produce large multienzyme complexes, cellulosomes, bound to the outer surface of the organisms. Furthuemore, most of the cellulases in C. hutchinsonii do not contain any carbohydrate-binding modules (CBMs). The structure of CHU2103, which is a processive endo-cellulase from C. hutchinsonii, was solved in this study. The CHU2103 has a shallow substrate binding cleft while a deep active center pocket which is comparatively uncommon for endo-cellulases. CHU2103 maight also be an exoglucanases due to its special structure which lead to more soluable reducing sugars than insoluable reducing sugars.
The crystal structures of processisive endo-cellulase CHU2103, lichenase F32EG5 with a ligand G4 and β-glucosidases g6403 are solved in this dissertation and their structural basis for function are determined. This work will promote the further study of these enzymes and provide theoretical basis for the functional renovations. Our lab have purified CohA and DocA through Escherichia coli, studied their resolution structure through nuclear magnetic resonance (NMR), and tested their interaction through surface plasmon resonance (SPR), too. My work in this thesis dose further study about assembly CohA with DocA, and enzyme activity of assembled complexes. We select one cohesin(CohC) and one dockerin(DocC) from Clostridium cellulolyticum as control
WO3 with surface oxygen vacancies as an anode buffer layer for high performance polymer solar cells
No full text<p>
The exploration of inexpensive and efficient anode buffer layers is essential in large scale commercial applications of polymer solar cells (PSCs). Here, we report a simple way that can significantly enhance the power conversion efficiency (PCE) and extend the lifetime of PSCs. A solution-based tungsten oxide (WO3) layer with surface oxygen vacancies (V(O)s) is introduced as an efficient anode buffer layer between the active layer and indium tin oxide (ITO) glass. The PCEs of PSCs based on P3HT: PC61BM and PBDTTT-C: PC71BM active layers are improved by 24% (from 3.84% to 4.76%) and 27% (from 5.91% to 7.50%) with the introduction of the WO3 (V-O) anode buffer layer, respectively, compared to that of the conventional PEDOT: PSS layer. The excellent performance is ascribed to the greatly improved fill factor and enhanced short circuit current density of the devices, which are benefited from the surface with lots of VOs for better interfacial contact and excellent charge transport properties of the WO3(VO) layer. The impressive PCE, good stability, easy fabrication and compatibility with solution processed organic photovoltaic devices support this material's potential applications in PSCs for both wide bandgap and narrow bandgap polymers.</p