【作者】 王姗；

【导师】 王沫；

【作者基本信息】 华中农业大学 ， 特种植物育种与栽培， 2010， 硕士

【摘要】 石杉科植物是一种重要天然药物资源,我国科学家早在20世纪80年代初,首先从用于民间的草药“千层塔”(即蛇足石杉)中分离得到石杉碱甲(huperzineA,hup-A),经过一系列药理试验,结果显示:石杉碱甲具有高效.低毒.高选择性的抗乙酰胆碱酯酶活性,并且能提高学习效率、改善老年人记忆功能的作用；临床应用疗效明显,毒性作用低。由于石杉碱甲是一种低毒、高效、可逆和高选择性的乙酰胆碱酯酶(AChE)抑制剂,已被国际上列为第二代的AChE抑制剂之一。蛇足石杉是目前石杉碱甲主要来源植物,但仅靠此种野生资源远远不能满足需求,而大量的人工合成工作几乎没有得到活性比天然HupA更好的衍生物和类似物,因此,寻找到新的HupA天然植物资源意义重大。以植物的亲缘关系为根据,扩大药源,寻找新的植物来源,已被证实是可以采用的途径,因此在石杉属发掘其他同类植物显得尤为重要。皱边石杉、四川石杉与蛇足石杉同属于石杉属下蛇足石杉组。目前,对石杉科植物石杉碱甲含量的测定多集中于蛇足石杉即千层塔这一种植物,而对于蛇足石杉和其科属中其他植物的系统比较研究鲜有报道。石杉属植物三种植物主要是分布在郁闭度、空气相对湿度均较大的山地密林下、沟谷阴湿土中及潮湿背阴的岩石陡壁上；调查区内与其伴生的主要有石松(伸筋草)、水竹,柳杉的小植株等(出现频度高),这给石杉属植物生长提供了大的荫蔽度；同一品种呈团块状分布。本研究选择湖北恩施利川境内蛇足石杉、皱边石杉和四川石杉从形态学、组织学、孢粉学等方面对三种植物进行比较研究；同时研究了不同生长季节和不同生长年份的植物中石杉碱甲的含量差异,为物种鉴定、资源保护及开发利用提供参考依据。1.对原植物形态的观察比较发现：三种植物在外观形态上差别较大,主要表现在叶的形态上,据植株外部形态可将三种植物很好的区别,显微结构鉴定只能作为其分类的一个辅助手段。石杉属植物形态相对简单,叶的形态是分类鉴别的主要对象,在大小、叶缘、叶柄长度上有明显的不同。蛇足石杉微褶皱,有粗大而不整齐的尖齿,叶尖微下垂,叶柄明显长度为2-5mm；皱边石杉叶缘锯齿粗而密,波状褶皱明显,叶柄长度2-3.5mm；与前两种相比,四川石杉叶片较小,叶边缘平直,不皱曲,有稀疏不规则的锯齿,无明显叶柄,且易出现茎芽苞。茎芽孢也可作为重要的分类依据。2.植物组织显微比较显示,三种石杉属植物根、茎结构相似。根均具有轻微栓化的表皮细胞、发达的皮层、管状中柱以及呈外始式分化的木质部等。但在维管束在数目和筛胞排列方式上有所不同：蛇足石杉根部管胞孔径较大,弧形排列,维管柱鞘细胞排列整齐,紧凑；皱边石杉木质部管胞数目较多,弧形紧密排列；四川石杉木质部管胞孔径大,排列不紧密,大小管胞间隔排列。茎均由表皮、皮层、维管柱、木质部、韧皮部组成,韧皮部位于木质部之间,且木质部四周均有辐射状排列的倒三角形脊状突起,形成星散状；三种植物维管束木质部脊状突起数目、维管束相对面积和韧皮部的排列方式存在差异。3.对三种植物的孢子进行扫描电镜观察,结果显示：三种植物孢子在形态上较一致：均为四分孢子,辐射对称,且三裂缝明显,赤道面观均为扇形。但同时三者在微形态方面也表现出了多样性,特别是外壁纹饰及裂缝长度具有种的特异性：皱边石杉辐射区和辐射间区光滑,裂缝最长,达到赤道线,孢子最饱满；四川石杉远极面孔穴最深,靠近裂缝处纹饰近光滑；蛇足石杉辐射间区在三种之间凹陷程度最大,是进行同属植物孢子鉴定时的重要依据。4.三种植物中均含有石杉碱甲,且四川石杉中含量最高,因此,皱边石杉和四川石杉可以用来提取天然石杉碱甲。四川石杉不同植株高度含量测定显示,株高在Ⅱ级(10±1cm)时石杉碱甲含量最高,达0.6388mg/g,与Ⅰ级(5±1cm)、Ⅲ级(15±1cm)植物中石杉碱甲含量差异达极显著,为四川石杉高效合理采收提供理论指导。更多还原

【Abstract】 Huperziaceae is the most important natural pharmaceutical resources, Scientist of China separated and discovered huperzineA from folk herbal medicine, which called "QianCengta" at the beginning of 80’s in the 20th Century. A series study proved that huperzineA have antiacetylcholinesterase activity:high efficiency、low toxicity、high selectivity; can improve learning efficiency; ameliorate the memory of elderly people. Because of such advantage of hupA, it has determined as the second generation antiacetylcholinesterase (AChE).Huperzia serrata (Thunb.ex Murray) Trev. is the major plant as the source of hupA, but this plant can not meet the demand, and a lot Synthetic work not abtained better activity bioactive substances than the natural HupA,so it is significant to found new plant resources which contain HupA. According to the plant genetic relationship enlarge medicine source, searching for another new plant resources, have confirmed as a useful method, therefore it is better way to found other plant in Huperzia Bern.. Huperzia serrata (Thunb.ex Murray) Trev.、Huperzia crispate (Ching ex H.S.Kung) Ching.、Huperzia sutchueniana (Herter) Ching., this three plant belongs to the same group under Huperzia Bern.. At present, more research about the content of HupA focus on Huperzia serrata (Thunb.ex Murray) Trev., and less systematic report on the other plant among Huperzia Bern.The plant of Huperzia Bern. are mainly distribute under mountain dense forest, gully, dankness soil and wet rock steep face, those enviroment have higher canopy density and atmospheric relative humidity. In investigated area, there are many other plant grow with the Huperzia Bern., such as lycopod, Cyperus alternifolius, younger Cryptomeria fortunei Hooibrenk ex Otto et Dietr.(appear with high frequency),those plant can provide a higher canopy density, and the same species grow in a small area, appears concentration phenomena.We choose Huperzia serrata (Thunb.ex Murray) Trev., Huperzia crispate (Ching ex H.S.Kung) Ching., Huperzia sutchueniana (Herter) Ching. in EnShi area, Hubei, and compared the three plant from morphology, histology, palynology, at the same time, we study the content of HupA from different harvest time and different growing years of these plant, in order to provide reference to species identification, new resources exploitation.1. Comparative Observation on plant morphology, we found that the three plant have difference in appearance. Mainly showed in leaf morphology, according to morphology,we can identified the three plant, histology identified only as a auxiliary method of their classification. The morphology of Huperzia Bern. relatively simple, leaf morphology is the main object of classification and identification,but in leaf size, margin, petiole length were significantly different.2. Comparison of plant histology display:the there species of root, stem structure are similar. All have small bolt of root epidermal cells, developed cortex, siphonostele, and xylem in exarch development, but vascular bundle in the number and arrangement of sieve cells are different. Stem composed by the epidermis, cortex, vascular bundle, xylem and phloem. And radial xylem are arranged around the ridge-like protrusions inverted triangle, star formation bulk. Three kinds of vascular plants the number of xylem ridges processes, the relative area of vascular bundle and phloem are arranged different.3. The spores of 3 different plants were observed with scanning electron microscope. We found that different plant are similar in shape:all are tetraspore, radial symmetry, and three cracks are obviously, equatorial view are fan-shaped。But different in micro-morphology, especially in surface ornamentation and crack length. Every species has their own characteristics。Such as:the spore of Huperzia serrata (Thunb.ex Murray) Trev., interradial region level in the depression between the maximum of three. Huperzia crispate (Ching ex H.S.Kung) Ching. spore’s radiation zone and interradial region are smooth, crack up, reaching the equator line, the most full of spores. Huperzia sutchueniana (Herter) Ching. spore has deepest point in distal face, near the crack of ornamentation nearly smooth.4. Huperzine A is contained in three plants, in Huperzia sutchueniana (Herter) Ching. the concentration is the highest, so, Huperzia sutchueniana (Herter) Ching. and Huperzia crispate (Ching ex H.S.Kung) Ching. can as natural new resources. Huperzia sutchueniana (Herter) Ching. determination of different plant height showed that plant height in levelⅡ(10±1cm),the content is the highest,0.6388mg/g, with level (5±1cm),levelⅢ(15±1cm) highly significant differences in content, it can provide scientific basis for efficient harvesting of Huperzia sutchueniana (Herter) Ching.更多还原