多元氨基酸、肽体系中稀土存在形态及配位作用特点研究

本文研究了稀土在多元氨基酸小分子生物配体体系存在形态及其作用特点，探索了稀土元素在生物体内的分布、代谢及其生物效应的机理，同时研究了稀土与复杂的生物大分子作用及稀土氨基酸固体配合物。

Fabrication of Silicon Crystal-Facet-Dependent Nanostructures by Electron-Beam Lithography

Silicon crystal-facet-dependent nanostructures have been successfully fabricated on a (100)-oriented silicon-on-insulator wafer using electron-beam lithography and the silicon anisotropic wet etching technique. This technique takes ad-vantage of the large difference in etching properties for different crystallographic planes in alkaline solution. The mini-mum size of the trapezoidal top for those Si nanostructures can be reduced to less than 10nm. Scanning electron microscopy(SEM) and atomic force microscopy (AFM) observations indicate that the etched nanostructures have controllable shapes and smooth surfaces.

西藏产雪莲花的化学成分研究

本学位论文报道了作为传统藏药材广泛使用的西藏产雪莲花化学成分的研究。论文由五章组成，第一章是三种西藏产雪莲花的化学成分的系统分离纯化和结构鉴定；第二章为西藏产雪莲花化学成分的液-质及串联质谱联用分析；第三章提出了以HPLC和TLC为检测方法的雪莲花药材质量标准草案；第四章给出了对西藏产雪莲花挥发油化学成分的气-质联用分析结果；第五章概述了雪莲花的化学成分及药理研究进展。 第一章包括三个部分。第一部分报道了绵头雪莲花(Saussurea laniceps Hand.-Mazz.)全草乙醇提取物化学成分的分离鉴定。采用正相硅胶柱层析及凝胶柱层析等分离方法，从西藏产绵头雪莲花的乙醇提取物中共分离鉴定出15个化合物。其中11个化合物为首次从该植物中分离得到，当中2个化合物系在凤毛菊属植物中首次发现。第二部分报道了水母雪莲花(Saussurea medusa Maxim.)全草乙醇提取物的化学成分。采用正、反相硅胶柱层析及凝胶柱层析等分离方法，共分离鉴定出15个化合物，其中1个为新化合物，另有4个化合物为首次从该植物中分离得到。新化合物结构通过质谱和一维及二维核磁共振等波谱解析方法及碱水解反应确定为巴豆酰基-高车前苷(M-7)。第三部分报道了三指雪莲花 (Saussurea tridactyla Sch.-Bip. ex Hook. f.)全草乙醇提取物的化学成分。采用正相硅胶柱层析及凝胶柱层析等分离方法，共分离鉴定出7个化合物，其中1个化合物为首次从该植物中分离得到。 第二章也包括三个部分。首先是采用液-质联用(HPLC-DAD-ESI-MSn)分析方法，对7个西藏不同产地的三指雪莲花化学成分进行了分析，通过与标准品的 UV和MS数据比较，共鉴定出14个峰，并对其中8个共有成分进行了定量测定。其次是关于八种西藏产雪莲花化学成分的液-质联用(HPLC-DAD-ESI-MSn)分析，通过与标准品的UV和MS数据比较，共鉴定出15个峰，并对其中8个共有成分进行了定量检测。最后通过对八种西藏产雪莲花主要化学成分的多级串联质谱(ESI-MSn)分析，快速、灵敏地鉴定出10个黄酮和3个香豆素化学成分。 第三章同样包括三个部分。首先是以绵头雪莲花中主要香豆素成分东莨菪素和伞形花内酯为对照品，通过TLC定性检测和HPLC含量测定，草拟出较严谨的药材质量标准。其次是将绵头雪莲花、三指雪莲花和雪兔子作为一个药材看待，草拟了以东莨菪素和伞形花内酯的TLC检测为指标的药材质量标准。最后是针对水母雪莲花，以主要黄酮成分芹菜素-7-O-b-D-葡萄糖苷为对照品作TLC检测，并草拟出该药材的质量标准草案。 第四章报道了西藏产雪莲花挥发油的化学成分分析。采用传统水蒸气蒸馏法分别从八种雪莲花全草中提取挥发油，利用气相色谱-质谱联用技术分别从水母雪莲花、绵头雪莲花、槲叶雪莲花、云状雪兔子、拉萨雪兔子、小果雪兔子、雪兔子和三指雪莲花中分别鉴定出83、83、56、34、21、20、24和20个化学成分，分别占其挥发油总量的70.7%、76.0%、82.2%、55.4%、49.7%、70.4 %、76.2%和 76.7%。 第五章为综述，总结和概括了雪莲花的化学和药理研究进展。 The dissertation reports the investigation of the chemical constituents of the genus Saussurea. Quite a lot of species in this genus are traditional Tibetan medicinal plants, and hence have been widely used in traditional Tibetan medicine. This dissertation consisted of five chapters. The first chapter is on the chemical constituents of three Saussurea plants. The second section is about the analysis of chemical constituents of Saussurea plants using HPLC-MS and ESI-MS/MS. In the third chapter, we proposed quality-control standards for the Genus Saussurea based on TLC (thin layer chromatography) and HPLC. The fourth chapter is about chemical compositions of the essential oil from the whole plant of Saussurea plants. The last chapter reviews the research progress of the Genus Saussurea. The first chapter consists of three parts. The first part is about chemical constituents of ethanol extracts from whole plant of Saussurea laniceps Hand.-Mazz. Fifteen compounds were isolated by column chromatography on normal phase silica gel and Sephadex LH-20. Among them, eleven compounds were isolated from this plant for the first time, and two compounds were isolated from Genus Saussurea for the first time. The second part is about chemical constituents of ethanol extracts from whole plant of Saussurea medusa Maxim. Fifteen compounds were isolated by column chromatography on normal phase, reversed phase silica gel and Sephadex LH-20. Five of them were isolated from this plant for the first time, and there is one new flavonoid glucoside which was identified as 6″-O-crotonoyl-homoplantaginin (M-7) based on the evidence of one- and two-dimensional nuclear magnetic resonance, mass spectrometry analysis, and alkaline hydrolysis reaction. The last part is about chemical constituents of ethanol extracts from whole plant of Saussurea tridactyla Sch.-Bip. ex Hook. f.. Seven compounds were isolated by column chromatography on normal phase silica gel and Sephadex LH-20. There is one compound which was isolated from this plant for the first time. The second chapter consists of three parts. In the first part, we analyzed the chemical constituents of S. tridactyla collected from seven different places in Tibet using HPLC-DAD-ESI-MSn. Fourteen peaks in the HPLC were identified by comparison of UV and MS spectra with those of authentic compounds, among which eight common peaks were quantified. In the second part, we analyzed the chemical constituents of eight Saussurea species using HPLC-DAD-ESI-MSn method. Fifteen peaks in the HPLC were identified by comparison of UV and MS spectra with those of authentic compounds and eight main peaks of them were quantified. In the last part, we analyzed the chemical compounds of the above eight Saussurea plants directly by ESI-MS/MS. Thirteen major compounds, including 10 flavonoids and 3 coumarins were easily rapidly identified. The third chapter consists of three parts. In the first part, we proposed a comparative high quality-control standard for S. laniceps, based on quality detection by TLC and quantity analysis by HPLC using two major compounds (umbelliferone and scopoletin) as standard compounds. In the second part, in viewing S. laniceps, S. tridactyla and S. gossypiphora as the members of one family of medicinal herbs, we suggested a quality-control standard based on the TLC detection of the two major compounds (umbelliferone and scopoletin). In the last part, we proposed a quality-control standard for S. medusa based on the TLC detection of its major component (apigenin 7-O-glucoside). The four chapter analyzed the chemical constituents of essential oil of eight Saussurea species. The essential oils were extracted from the whole plants of these samples with water stream distillation. By GC-MS analysis, we identified eighty-three compounds from S. medusa, eighty-three from S. laniceps, fifty-six from S. quercifolia, thirty-four from S. aster, twenty-one from S. kingii, twenty from S. simpsoniana, twenty-four from S. gossypiphora, and twenty from S. tridactyla respetively, which accounted for 70.7%, 76.0%, 82.2%, 55.4%, 49.7%, 70.4 %, 76.2% and 76.7% of the total essential oil, respectively. The last chapter reviews the research progress of the Genus Saussurea.

两种银莲花属植物的化学成分研究

本学位论文由5 章组成。第1 章报道了两头尖中三萜皂苷类化合物的全扫描电喷雾多级质谱分析及银莲花素A 的电喷雾质谱裂解规律；第2 和3 章报道了两种银莲花属药用植物化学成分的研究结果；第4 章报道了银莲花素A 的化学结构修饰及其对一种蛋白酪氨酸磷酸酯酶（PTP-1B）的抑制活性；第5 章综述了电喷雾质谱在皂苷类化合物结构鉴定中的应用进展。 第1 章报道了运用全扫描电喷雾多级质谱对两头尖中三萜皂苷类化合物的快速定性检测，共检测出18 个准分子离子峰，根据多级质谱数据并结合文献报道，对其中的15 个准分子离子峰进行了归属，并区分了一些同分异构体；更有意思的是，发现了3 个未见文献报道的三萜皂苷类化合物。根据它们的多级质谱数据，对其结构分别进行了初步解析。本章同时对银莲花素A 特殊的质谱裂解途径通过衍生物制备及其质谱分析进行了确认。 第2 和3 章，分别报道了两头尖和打破碗花花中水溶性多糖苷的分离纯化和结构鉴定。采用D101 大孔树脂和反复硅胶柱层析等分离手段，从两头尖水溶性部分分离得到8 个三萜皂苷类化合物，通过红外、电喷雾质谱和核磁共振等现代谱学方法，并结合传统的化学手段鉴定了它们的结构，其中4 个化合物为新化合物，分别命名为多被银莲花皂苷19、多被银莲花皂苷20、多被银莲花皂苷21和多被银莲花皂苷22。从打破碗花花水溶性部位分离得到6 个三萜皂苷类化合物，通过现代谱学手段，并结合传统的化学方法对它们进行了鉴定，其中1 个为新化合物，命名为打破碗花花苷H。 第4 章报道了以银莲花素A 及其同系物为先导化合物，进行化学结构修饰并对修饰产物进行广泛的生物活性筛选，发现在银莲花素A 及其同系物的结构中引入一些酸性基团后，其生物活性发生了变化，一些修饰产物显示出很强的PTP-1B 抑制作用，提示这些化合物有可能用于治疗II 型糖尿病。 第5 章综述了电喷雾多级质谱用于皂苷类化合物结构鉴定的研究进展。 This dissertation composes of five chapters. The first chapter elaborates the detection and qualification of the triterpenoidal saponins in Anemone raddeana by positive and negative full scan ESI-MSn. This part also reports the special fragmentation pathway of Raddeanin A by ESI-MS/MS. The second and third chapters present the phytochemical investigation of two medicinal plants from Anemone. The fourth part dwells on the structure modification of Raddeanin A and their inhibitory activity against PTP-1B. The last part is a review on the progress in the application of ESI-MSn in the structure identification of saponins. The first chapter reports the application of full scan ESI-MSn for fast analysis of triterpenoid saponins in Anemone raddeana. Eighteen quasi- molecular ion peaks were detected in the positive full scan ESI-MS and fifteen of them were identified by analysis of their tandem mass spectral data in the negative ion mode. Several isomers were differentiated. More interesting, three unreported triterpenoid saponins in this medicinal plant were detected and their structures were deduced according to the dissociation pathway of the known triterpenoidal saponins. This chapter also confirms the special fragmentation pathway of Raddeanin A by its derivative and the mass spectral analysis. The second and third chapters expatiate on the isolation and identification of the chemical constituents from A. raddeana and A. hupehensis. Eight compounds were isolated from the roots and stems of A. raddeana by methanol extraction and repeated column chromatography (including D101 and silica gel), and their structures were determined on the basis of IR, ESI-MS, NMR and chemical methods (including acid hydrolysis and alkaline saponification). Among them, four are new triterpenoid saponins and named as Raddeanoside R19, Raddeanoside R20, Raddeanoside R21 and Raddeanoside R22. Six compounds were isolated from the whole plants of A. hupehensis by the same methods as above, and their structures were also determined with the same way. One of them was confirmed to be new triterpenoid saponins and named as hupehensis saponin H. In the fourth chapter, in order to look for new active compounds, the structure of Raddeanin A and its analogs were modified. It was found that the modified products exhibited obvious inhibitory activity against PTP-1B when several acid groups were introduced. The fifth chapter summarizes the progress on the application of ESI-MSn in the structure identification of saponins.

水分亏缺和凋落物水浸液对云杉幼苗生长和种子萌发的影响

在人类活动导致全球变暖的前提下，由于全球气温的升高，地表水分加速向空中蒸发。从20世纪70年代至今，地球上严重干旱地区的面积几乎扩大了一倍。这一增长的一半可归因于气温升高而不是降雨量下降，因为实际上同期全球平均降水量还略有增长。干旱对陆地植物和农林生态系统产生深远影响，并已成为全球变化研究的一个重要方面。位于青藏高原东部的川西亚高山针叶林是研究气候变暖对陆地生态系统影响的重要森林类型。森林采伐迹地、人工林下和林窗环境作为目前该区人工造林和森林更新的重要生境，其截然不同的光环境对亚高山针叶林更新和森林动态有非常重要的影响。凋落物产生的化感物质可通过影响种子萌发和早期幼苗的定居而影响种群的建立和更新，而人工林和自然林物种以及更新速度的差异性也都受凋落物的影响。 云杉是川西亚高山针叶林群落的重要树种之一,在维持亚高山森林的景观格局和区域生态安全方面具有十分重要的作用，其自然更新能力及其影响机制一直是研究的热点问题。本试验以云杉种子和2年生幼苗为研究对象，从萌发、根尖形态、幼苗生长、光合作用、渗透调节和抗氧化能力等方面研究了不同光环境下水分亏缺和凋落物水浸液对云杉种子和幼苗生长的影响。旨在从更新的角度探讨亚高山针叶林自然更新的过程，其研究成果可在一定程度上为川西亚高山针叶林更新提供科学依据，同时也可为林业生产管理提供科学指导。主要研究结论如下： 水分亏缺在生长形态、光合作用、抗氧化能力、活性氧化对云杉幼苗都有显著影响。总体表现为，水分亏缺导致了云杉幼苗的高度、地径、单株总生物量降低，增加了地下部分的生长；水分亏缺显著降低了云杉叶片中相对含水量、光合色素、叶氮含量，净光合速率和最大量子产量(Fv/Fm)，提高了幼苗叶片中膜脂过氧化产物（MDA）的含量；水分亏缺提高了幼苗叶片中过氧化氢(H2O2)含量，超氧荫离子（O2-）生成速率以及脯氨酸和抗氧化系统的活性（ASA, SOD, CAT, POD, APX和GR）。从这些结果可知，植物在遭受水分亏缺导致的伤害时，其自身会形成防御策略，并通过改变形态和生理方面的特性以减轻害。但是，这种自我保护机制依然不能抵抗严重水分亏缺对植物的伤害。 模拟林下低光照条件显著增加单株植物的地上部分生长，尤其是其叶片的比叶面积（叶面积/叶干重），同时其光合色素含量和叶片相对含水量也显著增加，这些改变直接导致植株光合速率和生物量的增加。同时，与高光照水平相比，低光照幼苗的膜脂过氧化产物（MDA）和活性氧物质均较低，显示出低光照比高光照水平对植物的更低的氧化伤害。尽管低光照也导致大部分抗氧化酶活性降低，但这正显示出植物遭受低的氧化伤害，更印证了前面的结论。 凋落物水浸液影响了云杉种子的萌发和根系的生长，更在形态、光合作用、抗氧化能力、活性氧物质以及叶氮水平上显著影响了云杉幼苗，其中，以人工纯林凋落物的影响更有强烈。具体表现在，种子萌发速率和萌发种子幼根的长度表现为对照>自然林处理>人工纯林；凋落物水浸液抑制种子分生区和伸长区的生长，人工林处理更降低了根毛区的生长，使根吸水分和养分困难。对2年生幼苗的影响主要表现在叶绿素含量、光合速率以及叶氮含量的降低；膜脂过氧化产物、活性氧物质和抗氧化酶系统的显著增加。同样的，人工纯林处理对云杉幼苗的影响显著于自然林处理。 在自然生态系统中，由于全球变暖气温升高导致的水分亏缺和森林凋落物都存在森林的砍伐迹地，林窗和林下环境中。我们的研究表明，与迹地或林窗强光照比较，林下的低光照环境由于为植物的生长营造了较为湿润的微环境，因此水分亏缺在林下对云杉幼苗造成的影响微弱。这可以从植物的形态、光合速率以及生物量积累，过氧化伤害和抗氧化酶系统表现出来。另一方面，凋落物水浸液在模拟林下低光照环境对植物的伤害也微弱于强光照环境，这与强光照环境高的水分散失导致环境水分亏缺有关；而人工纯林处理对云杉幼苗的伤害比对照和自然林处理显示出强烈的抑制作用。 Under the pre-condition of global warming resulted from intensive human activities, water in the earth’s surface rapidly evaporates due to the increase of global air temperature. From 1970s up to now, the area of serious drought in the world is almost twice as ever. This increase might be due to the increasing air temperature and not decreasing rainfall because global average rainfall in the corresponding period slightly is incremental. Drought will have profound impacts on terrestrial and agriculture-forest system and has also become the important issue of global change research. The subalpine coniferous forests in the eastern Qinghai-Tibet Plateau provide a natural laboratory for the studying the effects of global warming on terrestrial ecosystems. The light environment significantly differs among cutting blanks, forest gap and understory, which is particularly important for plant regeneration and forest dynamics in the subalpine coniferous forests. Picea asperata is one of the keystone species of subalpine coniferouis forests in western China, and it is very important in preserving landscape structure and regional ecological security of subalpine forests. The natural regeneration capacities and influence mechanism of Picea asperata are always the hot topics. In the present study, the short-term effects of two light levels (100% of full sunlight and 15% of full sunlight), two watering regimes (100% of field capacity and 30% of field capacity), two litter aqueous extracts (primitive forest and plantation aqueous extracts) on the seed germination, early growth and physiological traits of Picea asperata were determined in the laboratory and natural greenhouse. The present study was undertaken so as to give a better understanding of the regeneration progress affected by water deficit, low light and litter aqueous extracts. Our results could provide insights into the effects of climate warming on community composition and regeneration behavior for the subalpine coniferous forest ecosystem processes, and provide scientific direction for the forest production and management. Water deficit had significant effects on growth, morphological, physiological and biochemical traits of Picea asperata seedlings. Water deficit resulted in the decrease in height, basal diameter, total biomass and increase in under-ground development; water deficit significantly reduced the needle relative water content, photosynthetic pigments, needle nitrogen concentration, net photosynthetic rate and the maximum potential quantum yield of photosynthesis (Fv/Fm), and increased the degree of lipid peroxidation (MDA) in Picea asperata seedlings; water deficit also increased the rate of superoxide radical (O2-) production, hydrogen peroxide (H2O2) content, free proline content and the activities of antioxidant systems (ASA, SOD, POD, CAT, APX and GR) in Picea asperata seedlings. These results indicated that some protective mechanism was formed when plants suffered from drought stress, but the protection could not counteract the harm resulting from the serious drought stress on them. Low light in the understory significantly increased seedling above-ground development, especially the species leaf area (SLA), and photosynthetic pigments and relative needle content. These changes resulted in the increase in net photosynthetic rate and total biomass. Moreover, the lower MDA content and active oxygen species (AOS) (H2O2 and O2-) in low light seedlings suggested that low light had weaker oxidative damage as compared to high light. Lower antioxidant enzymes activities in low light seedlings indicated the weaker oxidative damage on Picea asperata seedlings than high light seedlings, which was correlative with the changes in MDA and AOS. Litter aqueous extracts affected seed germination and root system of Picea asperata seedlings. Significant changes in growth, photosynthesis, antioxidant activities, active oxygen species and leaf nitrogen concentration were also found in Picea asperata seedlings, and plantation treatment showed the stronger effects on these traits than those in control and primitive forest treatment. The present results indicated that seed germination and radicle length parameters in control were superior to those in primitive forest treatment, and those of primitive forest treatment were superior to plantation treatment; litter aqueous extracts inhibited the meristematic and elongation zone, and plantation treatment caused a decrease in root hairs so as to be difficult in absorbing water and nutrient in root system. On the other hand, litter aqueous extracts significantly decreased chlorophyll content, net photosynthetic rate and leaf nitrogen concentration of Picea asperata seedlings; MDA, AOS and antioxidant system activities were significantly increased in Picea asperata seedlings. Similarly, plantation treatment had more significant effect on Picea asperata seedlings as compared to primitive forest treatment. In the nature ecosystem, water deficit resulted from elevating air temperature and litter aqueous extract may probably coexist in the cutting blank, forest gap and understory. Our present study showed that water deficit had weaker effects on low light seedlings in the understory as compared to high light seedlings in the cutting blank and forest gap. The fact was confirmed from seedlings growth, gas exchange and biomass accumulation, peroxidation and antioxidant systems. This might be due to that low light-reduced leaf and air temperatures, vapour-pressure deficit, and the oxidative stresses can aggravate the impact of drought under higher light. On the other hand, litter aqueous extracts in the low light had weaker effects on the Picea asperata seedlings than those at high light level, which might be correlative to the water evapotranspiration under high light. Moreover, plantation litter aqueous extracts showed stronger inhibition for seed germination and seedling growth than control and primitive forest treatments.

青藏高原东缘几种树苗对增强紫外线-B和氮供应的响应

大气臭氧的损耗导致了地球表面具有生物学效应的紫外线-B(UV-B)辐射增强。同时，大气成分变化中除了UV-B辐射增强外，氮沉降是一个新近出现而又令人担忧的环境问题，其来源和分布正在迅速扩展到全球范围，并不断向陆地和水生生态系统沉降。本试验在四川省境内的中国科学院茂县生态站内进行，以云山、冷杉、色木槭和红椋子幼苗为模式植物，从生长形态、光合作用、抗氧化能力和矿质营养等方面研究了青藏高原东缘4种树苗对全球变化－增强UV-B辐射和氮供应(氮沉降)的响应。该试验为室外盆栽试验，包括四个处理：(1)大气UV-B辐射+无额外的氮供应(C)；(2)大气UV-B辐射+额外的氮供应(N)；(3)增强UV-B辐射+无额外的氮供应(UV-B)；(4)增强UV-B辐射+额外的氮供应(UV-B+N)。其目的：一方面有助于丰富我国对全球变化及区域响应研究的全面认识，进一步完善在全球气候变化条件下臭氧层削减和氮沉降对陆地生态系统影响的内容；另一方面，在一定程度上有助于我们更好的理解在全球变化下森林更新的早期过程。具体结果如下： 增强的UV-B辐射在生长形态、光合、抗氧化能力、活性氧和矿质营养方面对4种幼苗都有显著的影响。UV-B辐射增强对幼苗的影响不仅与物种有关，而且，还与氮营养水平相关。总体表现为，高的UV-B辐射导致了色木槭和红椋子幼苗叶片的皱缩和卷曲，并降低了色木槭幼苗的叶片数和叶重，在额外的氮供应下，云杉、冷杉和红椋子的叶重也显著地降低了；色木槭和红椋子幼苗叶片的解剖结构受到了增强的UV-B辐射的影响，增强的UV-B辐射显著地降低了色木槭叶片的栅栏组织厚度，提高了红椋子叶片的厚度；增强的UV-B辐射显著地降低了4种幼苗的单株总生物量、植物地下部分的生长、总叶绿素含量 [Chl (a + b)]、净光合速率和最大量子产量(Fv/Fm)，提高了4种幼苗叶片的膜脂过氧化(MDA含量)，改变了植物体不同器官中的矿质元素含量；增强的UV-B辐射提高了冷杉、色木槭和红椋子叶片中的过氧化氢含量(H2O2)、超氧负离子(O2-)生成速率，在额外的氮供应下，云杉叶片中的活性氧含量也显著地提高了；在无额外的氮供应条件下，增强的UV-B辐射显著地提高了4种幼苗叶片中的UV-B吸收物质、脯氨酸含量和抗氧化酶的活性(SOD、POD、CAT、GR和APX)。在额外的氮供应条件下，UV-B辐射的增强却显著地降低了冷杉叶片中脯氨酸含量和红椋子叶片中UV-B吸收物质含量，但是，在4种幼苗叶片中，5种抗氧化酶的活性对UV-B辐射的增强没有明显的规律性，增强的UV-B辐射显著地提高了云杉叶片中的POD、SOD和GR的活性，提高了冷杉叶片中的POD和GR活性，提高了色木槭叶片中的POD、SOD和CAT活性和红椋子幼苗叶片中的POD和SOD活性。从这些结果可知，植物在遭受高的UV-B辐射导致的过氧化胁迫时，植物体内形成了一定的保护机制，但是，这种保护不能抵抗高的UV-B辐射对植物的伤害。 额外的氮供应在生长形态、光合、抗氧化能力、活性氧和矿质营养方面对4种幼苗都有一定的影响，不同幼苗对额外的氮供应响应不同，并且受到UV-B辐射水平的影响。在当地现有的UV-B辐射水平下，额外的氮供应显著地提高了幼苗的单株总生物量、植物地下部分的生长、Chl (a + b)、净光合速率(红椋子除外)、UV-B吸收物质(冷杉除外)、脯氨酸含量(红椋子除外)和部分抗氧化酶的活性，降低了H2O2的含量、O2-的生成速率和MDA含量(红椋子除外)，改变了植物体内部分矿质元素含量，显著地提高了云杉和冷杉叶片中的Fv/Fm。这些指标总体表明，在当地现有大气UV-B辐射水平下，额外的氮供应对植物的生长和发育是有利的。在增强的UV-B辐射水平下，4种幼苗的生长形态和光合大部分指标都没有受到额外氮供应的影响，额外的氮供应提高了红椋子幼苗的单株总生物量和Chl (a + b)含量，提高了冷杉和色木槭叶片中的活性氧含量和MDA含量，却降低了红椋子叶片中的活性氧含量；额外的氮供应也提高了云杉、色木槭和红椋子叶片中UV-B吸收物质和脯氨酸含量，降低了冷杉叶片中UV-B吸收物质和脯氨酸含量；在抗氧化酶活性方面，额外的氮供应降低了云杉、冷杉叶片中5种抗氧化酶的活性和红椋子叶片中POD和GR的活性，提高了色木槭叶片中的POD和SOD的活性；4种幼苗植物体内的矿质元素含量对额外的氮供应没有显著的规律性。从这些结果可知，在高的UV-B辐射下，额外的氮供应提高了云杉、冷杉和色木槭幼苗对高的UV-B辐射的敏感性，然而，额外的氮供应却促进了红椋子幼苗的生长，原因可能是，在高的UV-B辐射下，额外的氮供应增加了红椋子叶片的厚度、叶重和叶片数，降低了叶片中活性氧含量的结果。表明在高的UV-B辐射水平下，额外的氮供应降低了红椋子幼苗对高的UV-B辐射的敏感性。 在全球变化的趋势下，UV-B辐射增强和氮沉降可能同时存在，我们的研究表明，与大气UV-B辐射+无额外的氮供应处理相比，增强UV-B辐射+额外的氮供应处理显著地降低了幼苗的单株总生物量(红椋子除外)、Chl (a + b)、净光合速率、Fv/Fm(冷杉除外)和MDA含量(红椋子除外)，提高了活性氧含量 (云杉除外)、UV-B紫外吸收物质含量(冷杉除外)、脯氨酸含量和部分抗氧化酶的活性，改变了植物体不同器官中的矿质元素含量。结果表明，在当地现有条件下，全球变化(UV-B辐射增强和氮沉降)对云杉、冷杉和色木槭幼苗的生长是不利，尽管植物体内一些抗氧化性指标提高了，然而，却对红椋子幼苗的单株总生物量的累积没有显著的影响。 The depletion of the ozone led to the increase of ultraviolet-B (UV-B) with biological effects in the earth’s surface. At the same time, except for enhanced UV-B radiation, nitrogen deposition was an anxious environmental problem at present, rapidly expanding to the global scope and continuously depositing to land and aquatic ecosystem. The experiment was conducted in Maoxian Ecological Station of Chinese Academy of Sciences, Sichuan province, China. Picea asperata, Abies faxoniana, Acer mono Maxim and Swida hemsleyi (Schneid. et Wanger.) Sojak seedlings were selected as model plants to assess the effects of enhanced UV-B radiation and supplemental nitrogen supply on growth, morphological, photosynthesis, antioxidant and mineral nutrient traits of 4 species seedlings in east Qinghai-Tibetan Plateau. The experiment was potted outdoor, including 4 treatments: (1) ambient UV-B without supplemental nitrogen (control, C); (2) ambient UV-B with supplemental nitrogen (N); (3) enhanced UV-B without supplemental nitrogen (UV-B); (4) enhanced UV-B with supplemental nitrogen (UV-B+N). One hand, it was helpful for enriching our country to comprehensive understanding of the researches in the global change and the region response, further perfecting the effects of the depleted ozone layer and nitrogen deposition on land ecosystem under the global change; the other hand, it was favorable for us to better understanding of the early process of forest renews under the global change. The results were as follows: Enhanced UV-B radiation had significant effects on 4 species seedlings in growth, morphological, photosynthesis, antioxidant and mineral nutrient traits of 4 species seedlings. The effects of enhanced UV-B on plants were not only related with species, but also related with nitrogen nutrient level. Generally, the increase of UV-B radiation led to the shrinkage and curl of leaves in Acer mono Maxim and Swida hemsleyi (Schneid. et Wanger.) Sojak seedlings, and reduced the number of leaf and leaf weight of Acer mono Maxim seedlings, under supplemental nitrogen supply, leaf weight of Picea asperata, Abies faxoniana and Swida hemsleyi (Schneid. et Wanger.) Sojak seedlings significantly also reduced; the anatomical features of leaf in Acer mono Maxim and Swida hemsleyi (Schneid. et Wanger.) Sojak seedlings affected by enhanced UV-B radiation, the increase of UV-B radiation markedly reduced the palisade tissue thickness of Acer mono Maxim leaf and enhanced the leaf thickness of Swida hemsleyi (Schneid. et Wanger.) Sojak seedlings; the enhanced UV-B radiation significantly reduced total biomass per plant of 4 species seedlings, the growth of the underground parts, Chl (a + b), net photosynthetic rate and maximum potential quantum yield of photosynthesis (Fv/Fm), and increased the degree of lipid peroxidation (MDA content) and changed the content of mineral elements in different parts of plants; the enhanced UV-B radiation also increased the rate of superoxide radical (O2-) production and hydrogen peroxide (H2O2) content in leaves of Abies faxoniana, Acer mono Maxim, Swida hemsleyi (Schneid. et Wanger.) Sojak seedlings, under supplemental nitrogen supply, the reactive oxygen species in leaves of Picea asperata seedlings also significantly increased by enhanced UV-B radiation; under without supplemental nitrogen supply, enhanced UV-B radiation evidently induced an increase in UV-B absorbing compounds, proline content and the activities of antioxidant enzymes (SOD, POD, CAT, GR and APX) of leaves in 4 species seedlings. Under supplemental nitrogen supply, enhanced UV-B radiation induced a decrease in proline content of leaves in Abies faxoniana seedlings and UV-B absorbing compounds of leaves in Swida hemsleyi (Schneid. et Wanger.) Sojak seedlings, but, there were no obvious rules in the activities of five antioxidant enzymes of 4 species seedling leaves to enhanced UV-B radiation, enhanced UV-B radiation significantly increased the activities of POD, SOD and GR in Picea asperata leaves, the activities of POD and GR in Abies faxoniana leaves and the activities of POD, SOD and CAT in Acer mono Maxim leaves. The results indicated that some protective mechanism was formed when plants were exposed to enhanced UV-B radiation, but the protection could not counteract the harm of high UV-B radiation on plants. Supplemental nitrogen supply had some effects on 4 species seedlings in growth, morphological, photosynthesis, antioxidant and mineral nutrient traits. The response of 4 species seedlings was different to supplemental nitrogen supply, and was affected by UV-B levels. Under local ambient UV-B radiation, supplemental nitrogen supply significantly increased the total biomass per plant, the growth of underground parts, Chl (a + b), net photosynthetic rate (except for Acer mono Maxim seedlings), UV-B absorbing compounds (except for Abies faxoniana seedlings), proline content (except for Swida hemsleyi (Schneid. et Wanger.) Sojak seedlings) and the activities of some antioxidant enzymes, and reduced H2O2 content, the rate of O2- production and MDA content (except for Swida hemsleyi (Schneid. et Wanger.) Sojak seedlings) and changed the content of mineral elemental in different parts; supplemental nitrogen supply also evidently increased Fv/Fm in Picea asperata and Abies faxoniana seedlings. These results indicated that supplemental nitrogen supply was favorable for the growth of plants under local ambient UV-B radiation. Under enhanced UV-B radiation, mostly parameters in growth and morphology of 4 species seedlings were not affected by supplemental nitrogen supply. Supplemental nitrogen supply increased the total biomass per plant and Chl (a + b) of Swida hemsleyi (Schneid. et Wanger.) Sojak seedling, increased the reactive oxygen species and MDA content in Abies faxoniana and Acer mono Maxim leaves, and reduced the reactive oxygen species in Swida hemsleyi (Schneid. et Wanger.) Sojak leaves; supplemental nitrogen supply also increased UV-B absorbing compounds and proline content in Picea asperata, Acer mono Maxim and Swida hemsleyi (Schneid. et Wanger.) Sojak leaves, decreased UV-B absorbing compounds and proline content in Abies faxoniana leaves; in the activities of antioxidant enzymes, supplemental nitrogen supply significantly reduced the activities of antioxidant enzymes in Picea asperata and Abies faxoniana leaves and the activities of POD and GR in Swida hemsleyi (Schneid. et Wanger.) Sojak leaves, and increased the activities of POD and SOD in Acer mono Maxim leaves; the content of mineral elements in 4 species seedlings was no significantly rule to supplemental nitrogen supply. We knew from the results, under enhanced UV-B radiation, supplemental nitrogen supply made Picea asperata, Acer faxoniana and Acer mono Maxim seedlings more sensitivity to enhanced UV-B radiation, however, accelerated the growth of Swida hemsleyi (Schneid. et Wanger.) Sojak seedlings. The reason was probably that supplemental nitrogen supply increased the leaf thickness, leaf weight and leaf number, reduced the reactive oxygen content of leaf in Swida hemsleyi (Schneid. et Wanger.) Sojak seedlings grown under high UV-B radiation. This showed that supplemental nitrogen supply reduced the sensitivity of Swida hemsleyi (Schneid. et Wanger.) Sojak seedlings to high UV-B radiation. Under the tendency of the global change, enhanced UV-B radiation and nitrogen deposition may probably coexist. The results showed, compared with the treatment of ambient UV-B radiation without supplemental nitrogen supply, the treatment of enhanced UV-B radiation with supplemental nitrogen supply significantly reduced the total biomass per plants (except for Swida hemsleyi (Schneid. et Wanger.) Sojak seedlings), Chl (a + b), net photosynthetic rate, Fv/Fm and MDA content (except for Swida hemsleyi (Schneid. et Wanger.) Sojak seedlings), and increased reactive oxygen content (except for Picea asperata seedlings), UV-B absorbing compounds (except for Abies faxoniana seedlings), proline content and part antioxidant enzymes, and changed the content of mineral elements of different parts. The results indicated that the global change (enhanced UV-B and nitrogen deposition) were not favorable for the growth of plants under local ambient UV-B radiation and nitrogen nutrient level,, though increased some antioxidant indexes, however, the treatment of enhanced UV-B with supplement nitrogen supply did not significantly affect on the biomass accumulation of Swida hemsleyi (Schneid. et Wanger.) Sojak seedlings.

青杨组(Populus section Tacamahaca Spach)不同种对增强UV-B的反应差异

人类向大气中排放的大量氮氧化合物和氟氯烃类化合物(CFC’s)引起臭氧分子的分解，导致到达地球表面的紫外辐射增加，特别是UV-B辐射增强。本项目以青杨组杨树为模式植物，从形态和生理方面研究了来自不同UV-B背景下的康定杨与青杨在增强UV-B下的反应及其反应差异,并探讨了干旱、施肥对它们抗UV-B能力的影响。杨树具有分布广、适应性强、在生态环境治理和解决木材短缺方面均占有重要位置，研究成果可为生态系统的恢复与重建提供理论依据和科学指导。主要研究结果有以下： 1. 在温室中经过增强UV-B处理，杨树的外部形态及生理活动受到了一定程度的影响。增强UV-B导致康定杨、青杨的生物量、叶面积及节间长度降低，叶片增厚，SOD活性升高，膜伤害增加，而对叶片数目、R/S、叶绿素A、叶绿素B及整个叶绿素含量没有影响。两种杨树对UV-B胁迫的响应存在差异：在增强UV-B条件下，青杨的植株高度、生物量、叶面积、脯氨酸含量、长期用水效率受到的影响大于康定杨，相比而言，康定杨在比叶面积、叶片厚度、可溶性糖含量、UV-B吸收物质的含量及SOD和GPX活性方面增加的程度大于青杨。这些区别说明，来自于高海拔的康定杨比来自于低海拔的青杨对增强UV-B 具有更强的耐性。我们认为二者在叶片厚度、比叶面积、UV-B吸收物质含量及SOD、GPX活性差异是导致对增强UV-B耐性不同的原因。 2. 干旱与增强UV-B对杨树的生长和生理特性均产生了影响，而且两种胁迫共同作用时干旱表现减弱或加剧了UV-B对杨树某些形态和生理特性的影响。 据试验结果，干旱显著地降低了杨树的株高、叶片数目、叶面积，增加了叶片厚度，促进ABA的积累，提高了CAT活性。对于干旱，两种杨树之间也表现出了一定的差异性。可溶性蛋白质和脯氨酸在青杨叶片中得到显著积累，而在康定杨中没有变化。此外，CAT、长期用水效率在康定杨中受到的影响更加明显。长期用水效率的不同变化趋势说明两种杨树对水分胁迫采用了不同的用水策略，康定杨采用的是节水用水策略，提高用水效率，而青杨采用的是耗水的用水策略。根据干旱对叶面积、脯氨酸、ABA含量、CAT活性及长期用水效率等方面的影响，我们认为来自高海拔地区的康定杨比来自低海拔的青杨有更大的耐旱性，这是对生长环境长期适应的结果。在高海拔地区，因霜冻常带来土壤水分不可利用，降低了根系对水分的吸收，树木容易受到的生理性干旱。另外，高海拔的地区低的气温使植株对严寒有较强的耐性，减少了水分的需要。 生长于增强UV-B下的康定杨和青杨植株表现为高度降低，叶面积缩小，比叶面积增加；叶片栅栏组织、海绵组织均受到增强UV-B的影响，其厚度的增加导致整个叶片变厚。增强UV-B还显著提高了杨树的APX活性、UV-B吸收物质含量，而对叶片数目、ABA、可溶性蛋白质含量及CAT活性没有产生影响。试验中也观察到了两种杨树对增强UV-B响应的差异：与康定杨相比，在增强UV-B下青杨株高、叶面积降低的程度更大一些，SOD活性显著提高。另外UV-B吸收物质受到的影响不同。根据这些差别，高海拔的康定杨(3500 m)比来自低海拔的青杨(1500 m)增强UV-B有较强的耐性。 与水分充足情况下UV-B对植株的影响相比，干旱对杨树抗增强UV-B产生了一定的影响，表现为加剧或减弱UV-B对植物的影响，但这种影响与形态、生理指标有关。当干旱与增强UV-B共同作用时，杨树植株的株高、叶面积进一步降低、叶片进一步增厚。就脯氨酸的积累的而言，在没有水分胁迫时，增强UV-B促使它显著增加，而在干旱处理下这种效果变得不明显。干旱对增强UV-B的影响还与杨树的种类有一定的关系。在康定杨中，干旱减弱了增强UV-B对栅栏组织与海绵组织的影响，且在植株高度、叶面积上表现出累加效应，而在CAT上交互作用显著；但在青杨中干旱则加剧增强UV-B对栅栏组织与海绵组织的影响，在植株高度、叶面积及比叶面积上表现出显著的交互作用。据碳同素分析，在水分充足的条件下，无论是康定杨，还是青杨，增强UV-B均导致其长期用水效率的提高，然而当两种胁迫共同作用时，长期用水效率则表现出差异，在青杨中，长期用水效率得到进一步增高，而康定杨中干旱的效应被增强UV-B所减轻。 3. 田间试验表明，杨树的生长、生理特征都受到养分和增强UV-B的影响。施肥对杨树的影响表现为：提高了叶面积、生物量及SOD的活性，降低了抗坏血酸含量。对于施肥作用，两种杨树的反应也有区别：在康定杨中施肥显著增加了的叶片长度、宽度及光合色素的含量，降低了净光合速率、气孔导度及胞间CO2浓度；在青杨中，则SOD、GPX、APX活性表现增加。从试验看出，施肥对来自于高海拔地区的康定杨(3500 m)的影响较大，对来自低海拔的青杨(1500 m)影响较小，这与它们对原产地的生境适应有一定关系。在康定杨生长的高海拔地区，低温度和湿度不能为地上凋落物或土壤中的根分解提供理想的条件，造成当地土壤的低养分状况，所以当肥料施用以后，效果显著。 经过增强UV-B处理，杨树叶片中UV-B吸收物质含量、GPX的活性得到提高，而脯氨酸、丙二醛、可溶性蛋白质、叶绿素及类胡萝卜素含量没有受到影响。对于增强UV-B两种杨树受到的影响也有所不同：在青杨中增强UV-B导致叶面积缩小，生物量、净光合速率降低，APX的活性及长期用水效率的提高，而对康定杨的这些指标没有产生显著影响，相反抗氧化酶的活性明显高于青杨。这些差异性是由于两种杨树对原产地不同UV-B背景的长期适应结果。康定杨长期生长在较高UV-B环境中，对UV-B有较强的耐性。而青杨适应于较低的UV-B环境，对增强UV-B较为敏感。 试验中施肥也影响了植株对增强UV-B的反应，不过这种影响与杨树的种类及测定指标有一定的相关性。例如，在缺肥的情况下，青杨的长期用水效率和康定杨的叶绿素含量都受到增强UV-B的显著影响，而施肥以后这种影响变得不显著。在缺肥的条件下，GPX、APX在青杨中的活性、GPX在康定杨中的活性对增加UV-B反应不敏感；而施肥以后则变化显著，同样胞间CO2浓度在康定杨也有类似的变化。 For past decades, Ultraviolet radiation, especially UV-B reaching the Earth’s surface increased because of depletion of ozone layer resulted from emission of NxO and CFC’s from human activities. In this experiment, different species of Populus section Tacamahaca Spach from different UV-B background were selected as a model plant to assess the effects of enhanced UV-B radiation. Morphological and physiological traits induced by enhanced UV-B were observed and the different responses between P. kangdingensis and P. cathayana were discussed, furthermore the influences of drought and fertilizer on responses induced by enhanced UV-B were studied. Since poplars play an important role in lumber supply, and are important component of ecosystems due to their fast growth and wide adaptation, the study could provide a strong theoretical evidence and scientific direction for the afforestation, and rehabilitation of ecosystem. The results are as follows: 1. The experiment conducted in a greenhouse indicated that morphological and physiological traits of two poplars were affected by enhanced UV-B radiation. Enhanced UV-B radiation not only reduced biomass, leave area and internode length, but also increased leaf thickness and SOD activity as well as MDA concentration and electrolyte rate. However, no significant changes in leaf numbers, root shoot ratio, and total chlorophyll and chlorophyll component were observed. There were different responses to enhanced UV-B radiation between two species. Compared with P. kangdingensis, cuttings of P. cathayana, exhibited lower height increment and smaller leaf area. In addition, there were significant differences in free proline, soluble protein, and UV-B absorbing compounds, and the activity of SOD and GPX, long-term WUE between them. Differences in plant height, biomass, leaf area, free proline concentration, and long-termed WUE showed that P. cathayana were more affected by enhanced UV-B radiation than P. kangdingensis. In contrast, more increase of specific leaf mass, leaf thickness, and soluble sugar, and UV-B absorbing compounds, and activity of SOD and GPX were observed in P. kangdingensis. According to these results, we suggested that P. kangdingensis from high elevation, which adapted to higher UV-B environments, had more tolerance to enhanced UV-B than P. cathayana from low elevation, which adapted to lower UV-B environment. We believe it was the difference of leaf thickness, specific leaf mass, and UV-B absorbing compounds as well as the activity of SOD and GPX resulted in lower adaptation of P. cathayana to enhanced UV-B radiation. 2. Growth and physiological traits of two poplars were affected by both drought and enhanced UV-B radiation. Moreover, it was observed that when two stresses applied together drought could exacerbate UV-B effects or decrease sensitivity to UV-B. In the experiment, drought significantly decreased plant height, leaf numbers, leaf area, and increased leaf thickness, and ABA, and CAT activity of two poplars. There were significant interspecific differences to drought stress. Exposed to drought, soluble protein and proline concentration were increased in P. cathayana but not in P. kangdingensis. However, more changes in CAT and long-term WUE were observed in kangdingensis. Different change in long-term WUE suggests that two poplars adapted different water-use strategies. P. kangdingensis employ a conservative water-use strategy, whereas P. cathayana employ a prodigal water-use strategy. Based on the differences in leaf area, accumulation of free proline and ABA, CAT activity as well as long-term WUE, we believed that P. kangdingensis from high elevation had a greater tolerance to drought than P. cathayana from low elevation，which is the result of adaptation to local environment. In high elevation area, trees are prone to suffer from physiological drought because of un-movable water caused by frost. Besides lower temperature enable the plants had greater adaptability to frost as a results the requirement of water is reduced Enhanced UV-B radiation decreased shoots height, leaf area, and increased specific leaf mass and thickness of palisade and sponge layer as well as APX activity and UV-B absorbing compounds in both species. Whereas, leaf numbers, ABA content, soluble protein and CAT activity showed no differences to enhanced UV-B radiation. Interspecific differences were also observed. Compared with P. kangdingensis, P. cathayana showed lower shoot height and smaller leaf area, higher SOD activity. Besides, variation in UV-B absorbing compounds was found. These differences suggested that P. kangdingensis from high elevation (3500 m) was more tolerant to enhanced UV-B radiation than P. cathayana from low elevation (1500 m). Compared with morphological and physiological changes induced by enhanced UV-B radiation under well-watered conditions, drought exacerbated or decreased these changes. However, these effects vary with parameters measured. When two stresses applied together, shoot height and leaf area further decreased while leaf thickness further increased. Under well-watered conditions, enhanced UV-B radiation significantly increased proline content, but such effect was not observed under drought conditions. The effect of drought on enhanced UV-B radiation was related to species. For example, drought reduced the effects of enhanced UV-B radiation on palisade parenchyma and sponge mesophyll in P. kangdingensis, and additive effects in shoot height and leaf area and interactive effect CAT activity were observed. In contrast, for P. cathayana drought significantly exacerbated the effects of enhanced UV-B radiation on palisade parenchyma and sponge mesophyll; there were noticeable interaction in shoot height, leaf area and specific leaf mass. As far as long-term WUE is concerned, it was increased by enhanced UV-B radiation under well-watered conditions in both species. While different effect was observed between two species in combination of two stresses. Long-term water use efficiency was further increased in P. cathayana whereas the effect was less significant in P. kangdingensis. 3. The field experiment showed that growth and physiological traits of poplars were affected by nutrition and enhanced UV-B radiation. Fertilization significantly increased leaf area, biomass and SOD activity, reduced Ascorbic acid concentration. There was interspecific difference in response to fertilization. For P. kangdingensis, fertilization significantly increased leaf width, leaf length and photosynthetic pigments content while net photosynthetic rate and stomatal conductance, intercellular CO2 concentration were significantly decreased. However, for P. cathayana, these parameters were unaffected except the increase of SOD, GPX and APX activity. From above, it could concluded that P. kangdingensis from high elevation was more affected by fertilization than P. cathayana, This difference was due to adaptation to local environment., The low temperature and moisture where P. kangdingensis was collected can not provided optimum to decompose roots and litter fall as a result the nutrition in soil was poor. Exposed to enhanced UV-B radiation, for both species UV-B absorbing compounds and GPX activity were significantly increased while proline, MDA, soluble protein, chlorophyll, carotenoids were not affected. Different responses were also observed between the two species. Enhanced UV-B radiation caused significant decreases in leaf area, biomass, net photosynthetic rate and increase in APX activity and long-term WUE in P. cathayana but not in P. kangdingensis. In addition, activity in antioxidant enzymes was much higher in P. kangdingensis than in P. cathayana. In the experiment fertilization affected responses of cuttings to enhanced UV-B radiation, but it concern species and parameters measured. Long-term WUE in P. cathayana and chlorophyll in P. kangdingensis were significantly increased by enhanced UV-B radiation under non-fertilization treatments while the increase was not found under fertilization treatment. In contrast, under no fertilization treatment enhanced UV-B radiation did not affected GPX and APX activity in P. cathayana and GPX in P. kangdingensis while significant increase appeared after application of fertilization. Similar effect of enhanced UV-B radiation on intercellular CO2 concentration in P. kangdingensis was observed.

干旱胁迫与施N条件下白刺花（Sophora davidii）幼苗生长及其适应机制

干旱环境常常由于多变的降水事件和贫瘠土壤的综合作用，表现出较低的生产力和较低的植被覆盖度。全球性的气候变暖和人类干扰必将使得干旱地区缺水现状越来越严竣。贫瘠土壤环境中已经很低的有效养分含量也将会随着干旱的扩大而越来越低。干旱与半干旱系统中不断加剧的水分与养分的缺失将严重限制植物的生长和植被的更新，必然会使得已经恶化的环境恶化速率的加快、恶化范围的加大。如何抑制这种趋势，逐步改善已经恶化的环境是现在和将来干旱系统管理者面临的主要关键问题。了解干旱系统本土植物对未来气候变化的适应机制，不仅是植物生态学研究的重要内容，也对人为调节干旱环境，改善干旱系统植被条件，提高植被覆盖度具有重要的实践意义。 本研究以干旱河谷优势灌木白刺花（Sophora davidii）为研究对象，通过两年大棚水分和施N控制实验和一个生长季野外施N半控制实验，从植物生长－生理－资源利用以及植物生长土壤环境特征入手，系统的研究了白刺花幼苗生长特性对干旱胁迫和施N的响应与适应机制，并试图探讨施N是否可调节干旱系统土壤环境，人工促进干旱条件下幼苗定居，最终贡献于促进植被更新实践。初步研究结论如下： 1）白刺花幼苗生长、生物量积累与分配以及水分利用效率对干旱胁迫和施N处理的适应白刺花幼苗株高、基径、叶片数目、叶面积、根长、生物量生产、相对含水量和水分利用效率随着干旱胁迫程度的增加而明显降低，但地下部分生物量比例和R/S随着干旱胁迫程度的增加而增加。轻度施N处理下幼苗株高、基径、叶片数目、叶片面积和生物量生产有所增加。但重度施N处理下这些生长指标表现出微弱甚至降低的趋势。严重干旱胁迫条件下，幼苗叶面积率、R/S、相对含水量和水分利用效率也以轻度施N处理为最高。 2）白刺花幼苗叶片光合生理特征对干旱胁迫和施N处理的适应叶片光合色素含量和叶片光合效率随着干旱胁迫程度的增加而显著降低，并且PS2系统在干旱胁迫条件下表现出一定程度的光损害。但是比叶面积随着干旱胁迫程度的增加而增加。在相对较好水分条件下幼苗净光合速率的降低可能是因为气孔限制作用，而严重干旱胁迫条件下非气孔限制可能是导致幼苗叶片光合速率下降的主要原因。叶片叶绿素含量、潜在光合能力、羧化效率、光合效率以及RUBP再生能力等在施N处理下得到提高，并因而改善干旱胁迫条件下光合能力和效率。虽然各荧光参数对施N处理并无显著的反应，但是干旱胁迫条件下qN和Fv/Fm在轻度施N处理下维持相对较高的水平，而两年连续处理后在严重干旱胁迫条件下幼苗叶片光合效率受到重度施N处理的抑制，并且Fv/Fm和qN也在重度施N处理下降低。 3）白刺花幼苗C、N和P积累以及N、P利用效率对干旱胁迫和施N处理的适应白刺花幼苗C、N和P的积累，P利用效率以及N和P吸收效率随干旱胁迫程度的增加而显著降低，C、N和P的分配格局也随之改变。在相同水分处理下，C、N和P的积累量、P利用效率以及N和P吸收效率在轻度施N处理下表现为较高的水平。然而，C、N和P的积累量和P利用效率在重度施N处理下不仅没有表现出显著的正效应，而且有降低的趋势。另外，在相同水分条件下白刺花幼苗N利用效率随着施N强度的增加而降低。 4）白刺花幼苗生长土壤化学与微生物特性对干旱胁迫和施N的适应白刺花幼苗生长土壤有机C、有效N和P含量也随干旱胁迫程度的增加而明显降低。干旱胁迫条件下土壤C/N、C/P、转化酶、脲酶和碱性磷酸酶活性的降低可能表明较低的N和P矿化速率。尽管微生物生物量C、N和P对一个生长季干旱胁迫处理无显著反应，但微生物生物量C和N在两年连续干旱胁迫后显著降低。土壤有机C和有效P含量在轻度施N处理下大于重度施N处理，但是有效N含量随着施N强度的增加而增加。微生物生物量C和N、碱性磷酸酶和转化酶活性也在轻度施N处理下有所增加。但是碱性磷酸酶活性在重度施N处理下降低。 5）野外条件下白刺花幼苗生长特征及生长土壤生化特性对施N的适应植物生长、生物生产量、C的固定、N、P等资源的吸收和积累、其它受限资源的利用效率（如P）在轻度施N处理下均有所增加，但N利用效率有所降低。幼苗生物生产量及C、N和P等资源的分配格局在轻度施N处理下也没有明显的改变。白刺花幼苗叶片数目、生物生产量和C、N、P的积累量在重度施N处理下虽然也相对于对照有所增加，但幼苗根系长度显著降低。生物量及资源（生物量、C、N、P）在重度施N处理下较多地分配给地上部分（主要是叶片）。另外，土壤有机C、全N和有效N含量随外源施N的增加而显著增加，土壤pH随之降低，但土壤全P含量并无显著反应。其中有机C含量和有效P含量以轻度施N处理最高。微生物生物量C、N和P在轻度施N处理下也显著增加，而微生物生物量C在重度施N处理下显著降低。同时，转化酶、脲酶、碱性磷酸酶和中性磷酸酶活性在施N处理下也明显的提高，但酸性磷酸酶和过氧化氢酶活性显著降低，其中碱性磷酸酶和中性磷酸酶活性以轻度施N处理最高。 综合分析表明，干旱河谷水分和N严重限制了白刺花幼苗的生长。施N不能完全改变干旱胁迫对白刺花幼苗的抑制的作用，但是由于施N增加土壤N有效性，改善土壤一系列生物与化学过程，幼苗的生长特性也对施N表现出强烈的反应，表现为植物结构与资源分配格局的改善，植物叶片光合能力与效率的提高，植物生长以及利用其他受限资源（如水分和P）的效率的增加，致使植物自身生长及其生长环境在干旱环境下得到改善。但是过度施N不仅不能起到改善干旱胁迫下植物生长环境、促进植物生长的作用，反而在土壤过程以及植物生长过程中加重干旱胁迫对植物的伤害。因此，建议在采用白刺花作为先锋种改善干旱河谷系统环境的实践中，可适当施加N以改善土壤环境，调节植物利用与分配资源的效率，促进植物定居，得到人工促进种群更新的目的。但在实践过程中也要避免过度施N。 Arid regions of the world are generally noted for their low primary productivity which is due to a combination of low, unpredictable water supply and low soil nutrient concentrations. The most serious effects of global climate change and human disturbances may well be those which related to increasing drought since drought stress has already been the principal constraint in plant growth. The decline in total rainfall and/or soil water availability expected for the next decades may turn out to be even more drastic under future warmer conditions. Nevertheless, water deficit is not the only limiting factor in arid and semiarid environments. Soils often suffer from nutrient (especially N and P) deficiencies in these ecosystems, which can also be worsened by climate change. How to improve the poor soil quality and enhance the vegetation coverage is always the problem facing ecosystem managers. The adaptive mechanisms of native plant to future climate change is always the focus in plant ecology, it also plays important roles in improving vegetation coverage by manual controlled programmes. Sophora davidii is a native perennial shrub of arid valleys, which is often predominant on eroded slopes and plays a vital role in retaining ecological stability in this region. It has been found that S. davidii was better adapted to dry environment than other shrubs, prompting its use for re-vegetation of arid lands. A two-years greenhouse experiment and a field experiment were conducted in order to understand the adaptation responses of Sophora davidii seedlings to different water and N conditions, and further explore if additional N supply as a modified role could enhance the adaptation ability of S. davidii seedlings to dry and infertile environment. Two-month old seedlings were subjected to a completely randome design with three water (80%, 40% and 20% water field capacity (FC)) and three N supply (N0: 0, Nl: 92 and Nh: 184 mg N kg-1 soil) regimes. Field experiment was arranged only by three N supplies in the dry valley. 1) The growth, biomass partitioning and water-use efficiency of Sophora davidii seedlings in respond to drought stress and N supply Seedlings height, basal diameter, leaf number, leaf area, root length, biomass production, relative water content (RWC) and WUE were decreased with increase of drought stress. An increase in below-ground biomass was observed indicating a higher root/shoot ratio (R/S) under drought stress conditions. Low N supply increased seedlings height, basal diameter, leaf number, leaf area, and biomass production, but decreased root length. In contrast, these growth characteristics showed little or negative effect to high N supply treatment. Leaf percentages increased with increase of N supply, but fine root percentages decreased. In addition, Low N supply rather than the other two N treatments increased leaf area ratio (LAR), leaf/fine root mass ratio (L/FR), R/S and RWC under severe drought stress (20%FC), even though these parameters could increase with the high N supply treatment under well-watered condition (80%FC). Moreover, Low N supply also increased WUE under three water conditions, but high N supply had little effect on WUE under drought stress conditions (40%FC and 20%FC). 2) Leaf gas exchange and fluorescence parameters of Sophora davidii seedlings in respond to drought stress and N supply Leaf area (LA), photosynthetic pigment contents, and photosynthetic efficiency were decreased with increase of drought stress, but specific leaf area (SLA) increased. Photodamage in photosystem 2 (PS2) was also observed under drought stress condition. The decreased net photosynthetic rate (PN) under relative well-watered water conditions might result from stomatal limitations, but the decreased PN under other hand, photosynthetic capacity by increasing LA, photosynthetic chlorophyll contents, Pnmax, CE, Jmax were increased with increase N supply, and photosynthetic efficiency was improved with N supply treatment under water deficit. Although N supply did a little in alleviating photodamages to PS2 caused by drought stress, low N supply enhanced qN and kept relative high Fv/Fm under drought stress condition. However, high N supply inhibited leaf photosynthetic efficiency, and declined Fv/Fm and qN under severe drought stress condition after two year continues drought stress and N supply. 3) Carbon accumulation, nitrogen and phosphorus use efficiency of Sophora davidii seedlings in respond to drought stress and N supply C, N and P accumulation, NUE , N and P uptake efficiency (NUtE and NUtE ) P N P were decreased with increase of drought stress regardless of N supply. On the other hand, the S. davidii seedlings exhibited strong responses to N supply, but the responses were inconsistent with the various N supply levels. Low N supply rather than the other two N treatments increased C, N and P accumulation, improved NUEP, NUtE and NUtE under corresponding water condition. In contrast, high N supply N P did few even depressed effects on C, N and P accumulation, and NUEP, although NUtEN and NUtEP could increase with high N supply under corresponding water conditions. Even so, a decrease of NUEN was observed with increase of N supply under corresponding water conditions. 4) Soil microbial and chemical characters in respond to drought stress and N supply The content of soil organic C, available N and P were decreased with increase of drought stress. Decreases in C/N and C/P, and invertase, urea and alkaline phosphatase activity were also observed under drought stress conditions, indicating a lower N and P mineralization rate. Although microbial biomass C, N and P showed slight responses to drought stress after one growth period treatment, microbial biomass C and N were also decreased with increase of drought stress after two year continuous treatment. The content of soil organic C and available P showed the stronger positive responses to low N supply than which to high N supply, although than the other two N treatments increased microbial biomass N and invertase activity under severe drought stress condition, even though invertase activity could increase with high N supply treatment under relative well-water conditions. Moreover, low N supply treatment also increased C/P and alkaline phosphatase activity which might result from higher P mineralization, but high N supply did negative effects on alkaline phosphatase activity. 5) The growth characteristics of Sophora davidii seedlings and soil microbial and chemical characters in respond to N supply under field condition Low N supply facilitated seedlings growth by increasing leaf number, basal diameter, root length, biomass production, C, N and P accumulation and absorption, and enhancing the use efficiency of other limited resources as P. Compared to control, however, low N supply did little effect on altering biomass, C, N and P portioning in seedlings components. On the contrary, high N supply treatment also increased leaf number, biomass and C, N and P accumulation relative to control, but significantly decreased root length, and altered more biomass and resources to above-ground, which strongly reduced the ability of absorbing water under drought condition, and thus which might deep the drought stress. In addition, N supply increased soil C, N and available N content, but declined pH and showed little effects on P content. Low N supply showed higher values of soil C and available P content. Low N supply also increased microbial biomass C, N and P, although high N supply decreased microbial biomass C. N supply significantly enhanced soil invertase, urea, alkaline and neutral phosphratase activity, while declined acid phosphratase and catalase activity. Low N supply exhibited higher alkaline and neutral phosphratase activity compared to the others. The results from this study indicated that both drought and N limited the growth of S. davidii seedlings and their biomass production. Regardless of N supply levels, drought stress dramatically reduced the seedlings growth and biomass production. Although plant growth parameters, including basal diameter, height, leaf number, and biomass and their components were observed to be positive responses to low N supply, N supply alone can not alter the diminishing tendency which is caused by drought. available N content increased with increase N supply. In addition, low N supply rather These findings imply that drought played a primary limitation role and N was only the secondary. Even so, appropriate N supply was seemed to enhance the ability that S. davidii seedlings adapted to the xeric and infertile environment by improving soil processes, stimulating plant growth, increasing recourses accumulation, enhancing use efficiency of other limited resources, and balancing biomass and resources partitioning. Appropriate N supply, therefore, would be recommended to improve S. davidii seedling establishment in this region, but excess N supply should be avoided.

增强UV-B辐射对粗枝云杉(Picea asperata Mast.)和青杨(Populus cathayana Rehd.)的影响

光是植物赖以生存的重要环境因子，但是植物在获得光的同时不可避免的会受到紫外辐射的伤害。尤其是近年来，人类向大气中排放的大量氮氧化合物和氟氯烃类化合物(CFC’s)引起臭氧分子的分解，导致到达地球表面的紫外辐射增加，特别是UV-B辐射增强。而另一方面，植物对UV-B辐射反应的敏感性在种间和品种间存在差异，主要受植物基因型，生态型和生活型的控制。本项目分别以粗枝云杉和青杨组杨树为模式植物，从形态和生理生化方面分别研究了来自不同水分背景下的粗枝云杉种群和来自不同UV-B背景下的青杨种群在增强UV-B下的反应及其反应差异,并探讨了干旱、喷施外源脱落酸(ABA)对它们抗UV-B能力的影响。研究成果可为生态系统的恢复与重建提供理论依据和科学指导。主要研究结果如下： 1. 粗枝云杉的两个种群，湿润种群(来自四川黑水)和干旱种群(来自甘肃迭部)在水分良好和干旱状况下表现出对增强UV-B的不同响应。同时，干旱对粗枝云杉抗UV-B能力的影响也得到研究：两种胁迫共同作用时，干旱表现出在一定程度上减弱了增强UV-B对粗枝云杉的生理特性的影响。 干旱胁迫显著降低了两个粗枝云杉种群的光合同化速率(A), 气孔导度(gs)和PSII的有效光量子产量(Y), 同时，提高了非光化学猝灭效率(qN)和超氧化物歧化酶(SOD)的活性。与湿润种群相比，干旱种群抗旱性更强，表现为干旱种群拥有更高的SOD和干旱进一步加剧了UV-B的胁迫效应。 本研究中，干旱胁迫单独作用时，显著降低了青杨两个种群的生物量积累和气体交换，具体包括A、gs、蒸腾速率(E)和光合氮利用效率(PNUE)，提高了两个种群的瞬时水分利用效率(WUEi)、长期水分利用效率(WUET)、碳同位素组分(δ13C)和氮含量(N)。同时，UV吸收物质和ABA含量也得到积累。另一方面，增强UV-B对青杨两个种群各个指标的影响，同干旱所引起的效应有着相似的趋势。同低海拔种群相比，高海拔种群有着更强的抗旱和抗UV-B能力，具体表现在高海拔种群有着更多的生物量积累，更强的气体交换和水分利用效率及更高水平的ABA和UV吸收物质含量。相比干旱诱导的生物量积累和气体交换的降低，在干旱和增强UV-B两个胁迫同时作用于青杨时，这种降低表现的更为明显。显著的干旱和UV-B的交互作用还表现在WUEi, WUET, δ13C, 可溶性蛋白含量, UV吸收物质含量, ABA, 叶片和茎中的N含量以及C/N比中。 3. 经过一个生长季的试验观察，增强UV-B、外源ABA及两因子共同作用对青杨的生物量积累、气体交换、内源ABA和UV吸收物质含量、抗氧化系统以及碳、氮含量和碳/氮比均产生显著影响。本试验中，青杨的两个种群分别来自中国西南部的不同海拔地区，高海拔种群来自青海大通而低海拔种群来自四川九寨。外源ABA的胁迫为直接喷施ABA到青杨叶片，而增强UV-B胁迫是利用平方波系统分别保证青杨苗暴露于外界UV-B强度和两倍于外界UV-B强度下。 研究结果显示，增强UV-B显著的降低了两个青杨种群的株高、基茎、总叶面积和总生物量等生长指标，同时也导致其A、gs、E和叶片中碳含量的减少。而显著增加了SOD和过氧化物酶(GPx)活性水平，诱导了过氧化氢(H2O2)和MDA的显著增加，促进了UV吸收物质和不同器官中内源ABA含量的显著积累。另一方面，外源ABA引起了青杨光合同化速率的下降，SOD和GPx酶活性的增强，H2O2 和 MDA含量也表现出显著增加，同时，内源ABA含量得到显著累积。同低海拔种群相比，高海拔种群具有更加抗UV-B和外源ABA的特性。显著的UV-B和ABA的交互作用表现在A, E, SOD和GPx活性，以及叶片和根部的内源ABA等一系列指标中。在所有胁迫下，叶片中的碳和氮含量同其在茎和根中的含量显著相关，另外，叶片和茎中的氮含量同茎中的碳含量显著相关。 Sunlight is an indispensable environment factor for plants survival and development. Meanwhile, photosynthetic organisms need sunlight and are thus, inevitably, exposed to UV radiation. Especially for recent years, ultraviolet radiation, especially UV-B reaching the Earth’s surface increased because of depletion of ozone layer resulted from emission of NxO and CFC’s from human activities. On the other hand, the sensitivity of plants to UV-B radiation depends on the species, developmental stage and experimental conditions. In this experiment, two populations of Picea asperata Mast from different water background and two populations of Populus cathayana Rehder from different altitude background were selected as model plants to assess the effects of enhanced UV-B radiation. Morphological and physiological traits induced by enhanced UV-B in each plant species were observed and the different responses were discussed, furthermore the influences of drought and exogenous ABA on responses induced by enhanced UV-B were studied. The study could provide a strong theoretical evidence and scientific direction for the afforestation and rehabilitation of ecosystem. The results are as follows: 1. Different responses of two contrasting Picea asperata Mast. populations to enhanced ultraviolet-B (UV-B) radiation under well-watered and drought conditions were investigated. And the effects of enhanced UV-B on tolerance of drought were also observed in our study that the UV-B exposure may have alleviated some of the damage induced by drought. Two contrasting populations, originating from a wet and dry climate region in China, respectively, were employed in our study. Drought significantly decreased CO2 assimilation rate (A), stomatal conductance (gs) and effective PSII quantum yield (Y), while it significantly increased non-photochemical quenching (qN) and the activity of superoxide dismutase (SOD) in both populations. Compared with the wet climate population, the dry climate population was more acclimated to drought stress and showed much higher activities of SOD and ascorbate peroxidase (APX), and much lower levels of malondialdehyde (MDA) and electrolyte leakage. On the other hand, enhanced UV-B radiation also induced a significant decrease in the chlorophyll (Chl) content in both populations under well-watered conditions, and a significant increase in UV-absorbing compounds in the wet climate population. After one growing season of exposure to different UV-B levels and watering regimes, the increases in MDA and electrolyte leakage, as induced by drought, were less pronounced under the combination of UV-B and drought. In addition, an additive effect of drought and UV-B on A and gs was observed in the wet climate population, and on the activity of APX and qN in the dry climate population. 2. The significant effects of drought, enhanced UV-B radiation and their combination on Populus cathayana Rehd. growth and physiological traits were investigated in two populations, originating from high and low altitudes in south-west China. Our results showed that UV-B acts as an important signal allowing P. cathayana seedlings to respond to drought and that the combination of drought and UV-B may cause synergistically detrimental effects on plant growth in both populations. In both populations, drought significantly decreased biomass accumulation and gas exchange parameters, including A, gs, E and photosynthetic nitrogen use efficiency (PNUE). However, instantaneous water use efficiency (WUEi), transpiration efficiency (WUET), carbon isotope composition (δ13C) and nitrogen (N) content, as well as the accumulation of soluble protein, UV-absorbing compounds and abscisic acid (ABA) were significantly increased by drought. On the other hand, cuttings from both populations, when kept under enhanced UV-B radiation conditions, showed very similar changes in all above-mentioned parameters, as induced by drought. Compared with the low altitude population, the high altitude population was more tolerant to drought and enhanced UV-B, as indicated by the higher level of biomass accumulation, gas exchange, water-use efficiency, ABA concentration and UV-absorbing compounds. After one growing season of exposure to different UV-B levels and watering regimes, the decrease in biomass accumulation and gas exchange, induced by drought, was more pronounced under the combination of UV-B and drought. Significant interactions between drought and UV-B were observed in WUEi, WUET, δ13C, soluble protein, UV-absorbing compounds, ABA and in the leaf and stem N, as well as in the leaf and stem C/N ratio. 3. During one growing season, significant effects induced by enhanced UV-B radiation, exogenous ABA and their combination on biomass accumulation, gas exchange, endogenous ABA and UV-absorbing compounds concentrations, antioxidant system as well as carbon (C) content, nitrogen (N) content and C/N ratio were investigated in two contrasting Populus cathayana populations, originating from high and low altitudes in south-west China. Exogenous ABA was sprayed to the leaves and enhanced UV-B treatment was using a square-wave system to make the seedlings under ambient (1×) or twice ambient (2×) doses of biologically effective UV-B radiation (UV-BBE). Enhanced UV-B radiation significantly decreased height, basal diameter, total leaf area, total biomass, A, gs, E and carbon (C) content in leaves, and significantly increased activities of SOD and guaiacol peroxidase (GPx), hydrogen peroxide (H2O2) and malonaldehyde (MDA) content as well as the accumulation of UV-absorbing compounds and endogenous ABA concentrations among different organs in both populations. In contrast, exogenous ABA showed significant decrease in A and significant increases in activities of SOD and GPx, H2O2, MDA content and the endogenous ABA concentrations. Compared with the low altitude population, the high altitude population was more tolerant to enhanced UV-B and exogenous ABA. Significant interactions between UV-B and ABA were observed in A, E, activities of SOD and GPx, as well as in endogenous ABA in leaves and roots of both populations. Across all treatments, C and N content in leaves was strongly correlated with those were in stems and roots, respectively. Additionally, leaf and stem N content were significant correlated with stem C content.

微生物酶法拆分消旋芳香族氨基酸的研究

通过单因子和多因子摇瓶正交试验，确定了米曲霉液态发酵产氨基酰化酶的最佳发酵条件。优化发酵培养基组成（ρ/g L-1）： 葡萄糖40,蔗糖10,可溶性淀粉20,蛋白胨2.5,马铃薯液1 000mL, pH自然。培养基装量50mL/250mL三角瓶，接种量4%。培养温度30℃，转速100 rmin-1，发酵时间42h。每50mL培养物的总酶活由优化前的2627U提高到7338U，是优化前的2.79倍。 研究了米曲霉氨基酰化酶的部分酶学性质，该酶催化反应的最适pH为7.0，最适温度为40℃，低浓度的Co2+（5×10-4mol/L）对酶活激活作用显著，催化反应过程中，底物浓度大于0.2 mol/L时，存在高浓度底物抑制酶活力现象。 初步探索了包埋法固定化米曲霉氨基酰化酶的载体，在实验的五种载体中，以海藻酸钠为载体包埋固定化米曲霉氨基酰化酶酶活保留率高，且操作简单，成本低廉。对包埋法固定化米曲霉氨基酰化酶酶学性质进行了研究，较游离米曲霉氨基酰化酶，最适温度未发生改变，最适pH向碱性范围偏移至8.0，对酸碱和热的稳定性增强，最适底物浓度增大到0.4 mol/L。 根据氨基酰化酶能立体专一水解L-氨基酰化物的特点，利用米曲霉氨基酰化酶对消旋苯丙氨酸进行了拆分。在米曲霉氨基酰化酶选择性的作用于底物N-乙酰-L-苯丙氨酸，得到L-苯丙氨酸后，通过732阳离子树脂和结晶法分别将L-苯丙氨酸和N-乙酰-D-苯丙氨酸分离，N-乙酰-D-苯丙氨酸通过酸水解脱去乙酰基得到D-苯丙氨酸，拆分得到光学纯度为98%的L-苯丙氨酸（收率84.8%）和光学纯度为92.3%的D-苯丙氨酸（收率89.5%）。 separate factors tests and orthogonal experiments,the optimum fermentation conditions of aminoacylase –producing Aspergillus oryzae were determined, as follows（ρ/g L-1）,glucose 40,sucrose 10,soluble starch 20,peptone 2.5，potato juice 1000ml, inoculation volume 4%and fermentation temperature 30℃,rotation speed 100rmin-1.The highest total enzyme activity ,7338μ,was obtained after fermentation for 42 h, increased by 279% compared with the original value of 2627μbefore optimization. We dicussed partial characteristics of aminoacylase. The optimal pH and temperature of aminoacylase were 7.0 and 40℃ respectively. Low- concentration Co2+ (5×10-4mol/L）activated the aminoacylase remarkably while high-concentration substrate lowered the aminoacylase . Five vectors has been used for immobolizing the enzyme and calcium alginate showed to be the best one for it had the slightest influence on the enzyme activity, easy to operate ,and low in price, comparing with other fours. The enzymatic charateristic study showed that its optimum temperature didn’t change, but the optimum pH and substrat concentration were higher after immobilization. The stability of immobolized enzyme to acid, alkaline and heat rised as well. The aminoacylse from Aspergillus oryzae was used to resolute racemic phenylalanine to obtain D-phenylalanine. After catalyzing process, we took two methods to separate D-phenylalanine .In end,L-phenylalanine was obtained with 98% optical purity in 84.8% yield, D-phenylalanine was obtained with 92.3% optical purity in 89.5% yield.

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.

Liquid-phase hydrodechlorination of chlorinated benzenes over active carbon supported nickel catalysts under mild conditions

Liquid phase hydrodechlorination of chlorinated benzenes was studied over Ni/active carbon (Ni/AC), Ni/gamma-Al2O3, Ni/SiO2 and Raney Ni. The complete dechlorination of chlorobenzene (ClBz) was realized at 333-343 K on Ni/AC under hydrogen atmosphere of 1.0 MPa in the presence of alkaline hydroxide. Dichloro- and trichlorobenzenes were also hydrodechlorinated with 50-95% yields of benzene under the similar conditions, as above. The reaction follows zero-order to ClBz concentration and 1.9 order to hydrogen pressure. The reaction does not proceed in the absence of alkaline hydroxide, suggesting the complete coverage of active nickel surface with produced chlorine and the removal of the chlorine ion with hydroxide ion as a rate-limiting step. The active catalysts were characterized by H-2 chemisorption and transmission electron microscopy techniques. The apparent activity strongly depends on the active area of nickel on catalyst surface. (C) 2004 Published by Elsevier B.V.