厚朴中α-葡萄糖苷酶抑制活性成分研究

本论文由三章组成。 第一章是关于厚朴中具有α-葡萄糖苷酶抑制活性成分的研究。凹叶厚朴的乙醇提取物显示了较强的α-葡萄糖苷酶抑制活性。为了确定其活性成分，在活性测试的指导下，通过溶剂萃取、树脂吸附和反复硅胶柱层析等分离方法从凹叶厚朴乙醇提取物中分离得到6 个生物碱，并用质谱和核磁共振等波谱方法分别鉴定为：木兰箭毒碱，木兰花碱，鹅掌楸碱，蕃荔枝碱，罗默碱和Lysicamine。应用小肠α-葡萄糖苷酶模型测定了它们对α-葡萄糖苷酶的抑制作用。其中，番荔枝碱和木兰箭毒碱对α-葡萄糖苷酶相对抑制活性最好，分别为60%和62%；其它四个生物碱成分对α-葡萄糖苷酶的抑制活性几乎相当，鹅掌楸碱为46%，罗默碱为51%，Lysicamine 为49%，木兰花碱为51%。 第二章报道了厚朴酚的衍生物及其对α-葡萄糖苷酶的抑制活性。根据糖苷酶抑制剂的结构特点，设计合成了一系列厚朴酚的衍生物。厚朴酚经过Mannich 反应和环氧化及开环反应制备了一系列衍生物，经活性测试发现衍生物活性与取代基关系较大，其中5,5′-diallyl-3-((bis(2-hydroxyethyl)amino)methyl)biphenyl-2,2′-diol 的抑制活性最高，为72%。 第三章综述了厚朴的化学成分及药理活性两个方面的研究进展。 The dissertation consists of three chapters. The first chapter is about the study on the constituents with α-glycosidase inhibitory activity from Magnolia officinalis. The EtOH extracts of M. officinalis Rehd. et Wils showed good inhibitory activity against α-Glucosidase. In order to determine the active compounds, bio-assay was used to guide the isolation. Six known alkaloids were isolated by solvent extraction and repeated silica gel column chromatography, and their structures were identified as liriodenine, anonaine, roemerine, lysicamine, magnoflorine and magnocurarine by spectroscopic methods. The inhibitory activity against α-Glucosidase of these alkaloids was measured with alvine screening model of α-glucosidase. Among them, lysicamine and liriodenine have the best inhibitory activity at 60% and 62%, respectively. The other four alkaloids have close inhibitory activity, from 46% to 51%. The second chapter is about the derivation of magnolol and the inhibitory a ctivity of the derivatives. Seven derivatives of magnolol were prepared by Manni-ch reaction, epoxidation followed by ring-opening reaction. Biological activity as say indicated the inhibitory activity was related to substituting groups. Among them, 5,5′-diallyl-3-((bis(2-hydroxyethyl)amino)methyl)biphenyl-2,2′-diol had the highest activity at 72%. The third chapter is a review on the progress of M. officinalis including chemical constituents and pharmacological activity.

岷江上游人工针叶、阔叶林土壤可溶性有机物质特征及其对凋落物分解的响应

土壤可溶性有机物质（Dissolved organic matter，DOM）作为土壤有机质的活性组分，在陆地生态系统物质循环中扮演非常重要的角色。土壤DOM的主要成分可溶性有机碳（Dissolved organic carbon，DOC）和氮（Dissolved organic nitrogen，DON）参与C、N循环过程。为深入揭示全球C、N循环过程机制，在未开展DOC和DON的地区进行相关研究是有必要的。森林土壤（包括枯枝落叶层）DOC、DON动态及调控机理的研究是目前国际上森林生态系统C、N循环研究热点之一。本研究立足于暖温带岷江上游茂县地区人工林植被，对土壤DOC和DON的库容量，季节动态及其与其它养分之间的关系进行了系统研究，旨在了解DOC和DON在该区生态系统中的重要作用，并探讨作为DOM主要来源的叶凋落物对DOC和DON的动态影响，研究有助于更加详细地了解该区生态系统C和N循环过程。本论文主要研究结论如下： 1研究了岷江上游地区两大主要土壤类型（棕壤和褐土）不同植物群落下土壤的DOC和DON含量及特征，结果表明：DOC和DON在两种土壤类型中均有库容量存在，DOC在0-10cm和10-20cm土层的含量幅度分别111.96~159.95 mg kg-1和69.02~100.84 mg kg-1。DON在0-10cm和10-20cm土层的含量幅度分别11.88~23.08 mg kg-1和4.70~10.77 mg kg-1。游离氨基酸在0-10cm和10-20cm土层的含量幅度分别0.84~1.66 mg kg-1和0.39~0.73 mg kg-1。DOC、DON与土壤中的一些养分因子表现出了显著的相关关系，共同反映了土壤的状况和质量，在该区开展DOC和DON的系统研究是有必要的。 2 对油松与连香树林地土壤DOC、DON以及其它化学指标的季节动态进行了研究，结果表明：油松与连香树林地土壤DOC和DON的季节动态变化表现了类似的规律，DOC和DON的含量均以秋季最高。DOC和DON的季节动态变化主要受凋落物生物因素的影响，但其微生物活力的生物因素以及降雨、温度等非生物因素也是控制土壤DOC和DON含量的重要因素。土壤DON在土壤中的行为不同于矿质氮，其季节动态不同于NO3--N和NH4+-N的季节动态，在研究N循环过程中，应考虑DON的变化情况。 3 对油松与连香树林地分解层和表层土壤（0-10cm）氨基酸周转动态进行了研究，结果表明：油松林地和连香树林地均以分解层的氨基酸含量高于矿质表层土壤的含量。每个取样时期，油松林地内各层次的氨基酸含量高于连香树林地内相应层次的含量。两林地各层次无机氮含量均超过了氨基酸的含量，并且室内培养30天后无机N的含量仍然高于氨基酸的含量，所以可以认为该区立地条件下无论是在有机分解层还是矿质土层植物吸收利用的氮素仍是以无机N为主。 4 松林下松针凋落物易于累积，这与松针凋落物分解缓慢有关，从而导致松林内养分周转缓慢。通过用不同性质凋落物和灌丛地土壤构建微生态系统，比较油松、辐射松、连香树、灌丛虎榛子凋落物分解对C、N循环过程的影响，结果显示油松和辐射松针叶凋落物比连香树、虎榛子凋落物分解更慢，减缓了养分循环过程。然而将针叶凋落物与阔叶凋落物混合后，油松和辐射松针叶凋落物的分解加快，C、N元素的循环过程也加速。此结果表明在松林内维持具有高质量凋落物的灌丛植被或在松林内栽植一些阔叶树种如连香树对维持和增进松树人工林的土壤肥力有重大的作用。室内培养的结果还显示添加凋落物后土壤DOC和DON的含量显著增加，表明凋落物是土壤DOM的直接来源。然而不同物种凋落物处理下土壤DOC和DON的含量有所不同，并随时间发生改变。混合凋落物处理下土壤DOC和DON的含量均高于松针凋落物单独处理下土壤DOC和DON的含量。DON是一个主要的水溶性N库，随时间的变化趋势与无机N的变化趋势不同，在土壤N循环过程中起到了中间N库的作用。 As a labile fraction of soil organic matter, dissolved organic matter (DOM) plays a very important role in material cycling of terrestrial ecosystem. The turnover of DOM is now being considered as main components in nutrient cycling. DOM mainly includes dissolved organic carbon (DOC), -nitrogen (DON), -phosphorous (DOP) and –sulfur (DOS). Among these constituents, DOC and DON directly participate in C and N cycling. It is essential to study DOC and DON dynamics and their controlling factors in the areas where no related study has ever been carried out. Study about them can provide data supports on understanding the mechanism of the global C and N cycling. DOC and DON dynamics and their controlling factors have been focused on in the research of C and N cycling of forest ecosystems. Based on forest plantations of Maoxian, Minjiang River in warm temperate zone, soil DOC and DON pool size, their seasonal dynamics, and the correlation between DOC, DON and other nutrients were studied in order to understand the importance of DOC and DON in the study area. Soil DOC and DON dynamics induced by leaf litter decomposition were also studied. The study contributed to comprehensively understanding C and N cycling processes and providing baseline data for including DOC and DON into the indices system of evaluating nutrient conditions. The results were as follows: 1 Several different plant communities under brown soil and Cinnamon soil were chosen as sampling plots. The contents and features of soil DOC and DON were evaluated. The results showed that DOC and DON were present under the two soil types. DOC contents in the top soil (0-10 cm) and the subsoil (10-20 cm) respectively varied from 111.96 mg kg-1to 159.95 mg kg-1, and 69.02 mg kg-1 to 100.84 mg kg-1. DON contents in the top soil (0-10 cm) and the subsoil (10-20 cm) respectively varied from 11.88 mg kg-1to 23.08 mg kg-1, and 4.70 mg kg-1 to 10.77 mg kg-1. Free amino acid contents in the top soil (0-10 cm) and the subsoil (10-20 cm) respectively varied from 0.84 mg kg-1to 1.66 mg kg-1, and 0.39 mg kg-1 to 0.73 mg kg-1. Significant correlations were found between DOC, DON and some nutrient indices, which together reflected soil condition and quality. It was hence essential to study DOC and DON in the study area. 2 Seasonal dynamics of DOC, DON, inorganic N, microbial biomass C and N were studied under Pinus tabulaeformis and Cercidiphyllum japonicum plantation. The results indicated that seasonal dynamics of soil DOC and DON under the two plantations performed similar change pattern, with the highest values in autumn. The seasonal dynamics of soil DOC and DON were mainly influenced by the litterfall. However, biotic factors such as soil microbial activities and abiotic factors such as precipitation and temperature also controlled the dynamics of soil DOC and DON. The seasonal dynamic of DON was different from that of NO3--N and NH4+-N, which showed that the behavioral differences between DON and inorganic nitrogen. And hence, it was proposed to include DON into soil N cycling in the study area. 3 Amino acid dynamics in Oa and topsoil (0-10 cm) under P. tabulaeformis and C. japonicum plantation were studied. The results showed that amino acid content in Oa was significantly higher than that in mineral soil. At each sampling time, significantly higher amino acid contents were found in P. tabulaeformis plantation than in C. japonicum plantation. The content of inorganic nitrogen was much higher than the content of amino acid in each sampling layer at each sampling time. After a 30-days laboratory incubation the content of amino acid was still lower than the content of inorganic nitrogen. The results implicated that the form of N absorbed by plants in these study sites were mainly inorganic nitrogen. 4 Usually needle litter is more resistant to decomposition, which leads to needle litter accumulation in pure coniferous stands and slows down the rate of nutrient circulation. By constructing microcosms with local shrubland soil and containing the four single-species (P. tabulaeformis, P. radiata, C. japonicum, Ostryopsis davidiana) litters, the decomposition rates and related C and N dynamics of needle litters and broadleaved litters during the early stage were compared. The results showed that the decomposition rates of pine needles were lower than those of broadleaved litters, which descended C and N cycling processes. However, the presence of C. japonicum or O. davidiana litter into pine needles increased the decomposition rates of pine needles and also dramatically promoted C and N cycling processes. It should be appropriate for plantation managers to consider C. japonicum as an ameliorative species or remain O. davidiana in pine plantations to improve soil conditions and help maintain soil fertility. The laboratory incubation still showed that DOC and DON contents in all litter-amended treatments were significantly higher than no litter-amended treatment, which proved that litter could be a direct source of DOM in soils. Different species litters induced different soil DOC and DON contents, which correspondingly changed over time. DOC and DON contents in mixed litter treatments were higher than those in pine needle litter treatments. As a major soluble N pool, DON developed a different changing pattern over time compared with inorganic N and played a role of interim N pool in soil N cycling.

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.

微生物法沥取重金属污泥中铬的研究

本文研究了两种微生物及其组合沥取、回收用微生物法治理电镀铬废水产生的铬污泥中的铬。铬污泥富含C、N、O，含铬量为13%， 经X-光电子能谱分析铬以三价态(氢氧化铬)存在。二种微生物分别从一酸性矿水和酸性污泥中分离筛选得到，经鉴定为硫杆菌属 (Thiobacillus Beijerinek)的两个不同种，一为氧化亚铁硫杆菌(Thiobacillu ferrooxidans, TF), 另一为氧化硫硫杆菌 (Thiobacillus thiooxidans, TT)。研究并比较了不同微生物对污泥中铬的沥取能力，结果表明，TT菌沥取铬效率最高。振荡、动 态淋滤、静置等沥取方式经过研究表明动态淋滤为最佳，室温条件下(15-20℃)，污泥浓度为20g/L时，总铬沥出率达60%时所需时 间：动态淋滤为48.5h，振荡和静置方式分别为91.22，81.6h。研究了不同温度、不同起始PH、不同污泥浓度及非成熟菌液对微生 物沥取能力的影响：(1) 沥取前期，温度对铬的沥出影响较大；微生物沥取反应基本属一级反应；温度与反应速率的关系基本符合 Arrhenius方程，但沥取后期这一特点并不突出。(2) 沥取液最适起始PH为菌液自然PH；PH值的人为改变将使铬的沥出大大降低。 (3) 污泥浓度与铬的沥出呈正相关，但浓度高于30g/L时，铬的沥出量不再增加。(4) 非成熟菌液沥出铬的能力较差，但沥取液中 微生物生长繁殖较为活跃。总结微生物沥取反应最佳沥取条件为：TT成熟菌液、污泥浓度10g/L、温度25-36℃、动态淋滤方式，此 时铬几乎可100%从污泥中沥出。经扫描电镜分析，沥取开始时，微生物紧密吸附于污泥颗粒表面上，表面紧密吸附为微生物发挥功 能提供了基础。微生物沥取污泥中铬的反应机理推测为：硫细菌代谢产硫酸或氧化Fe2+成Fe3+，利用酸，Fe3+ 及自身氧化酶系统 氧化污泥中Cr3+为Cr6+,Cr6+溶出结晶为CrO3。This paper has studied bioleaching and recovery of Chronium(Cr)from electroplating sludge by two consortum of bacteria and their combination, with sludge produced by microbiological process treating electroplating wastewater containing Cr as material. The share of Cr is 13% and its state is Cr (OH)3 in the sludge. One of the bacteria in the paper was isolated from acid sewage sludge and the other was from acid mineral water. The former was tested and determined as Thiobacillus ferroxidans(TF) and the latter was Thiobacillus thiooxidans(TT). Different microorganisms, responsible for the metal leaching activity, have great influence on the efficiency of leaching. The results showed that TT has biggest power. Experiments were conducted to examined effects of three different ways of leaching(Shaking, Down-leaching, Static-leaching). When temperature was in-door's (15-20℃)and concentration of the sludge was 20g/L, the bioleaching time required to reach 60% of Cr solubilization with the above three ways were 91.2, 48.5, 81.6h respectively. Down-leaching was proved to be the most efficient. The influence of different temperature, initial PH, concentration of the sludge and non-mature inoculum had been studied. The results obtained reveal that: (1) The variation of temperature is important during the time from initial to middle of leaching. The reaction of bioleaching belongs to first-order. The relation between the bioleaching rate constant(In k)and temerature can be expressed by Arrhenius function. (2) The fittest initial PH is the nature PH of mature inoculum. Any alteration with it could cause clearly negative effection. (3) The concentration of the sludge can make strong influence on the bioleaching efficiency. But when the concentration is above 30g/L, the increasing of Cr in the solution is little. (4) If non-mature inoculum acts as the bioleachin microorganism, little quantity of Cr would be gained from the sludge. But the micormass in the solution is very active. The results from electron microscope showed that microorganisms adhered to the surface of the sludge and the adherence was the first stage of the bioleaching. Some salts of Cr can be obtained afer the water of the bioleaching solution being evaporated. By analysing the results of experiment with X-Ray spectroscopy, the salt was identified as CrO3. The recovery rate of Cr is 78.4%.