碱法提取喜树碱工艺的研究

通过正交试验设计,探讨了以稀NaOH溶液为溶剂从喜树果实中提取喜树碱的工艺条件。确定的最佳工艺条件为:NaOH溶液浓度为0.3%,NaOH溶液用量16ml/g原料·次,提取时间为3~4h,提取温度为室温。     

北京“松黑木吉丁”或“松迹地吉丁”学名考

近年来北京等地的一些油松由于生长衰弱,遭受梢小蠹和吉丁的为害。过去北京等地记录的为害油松的吉丁为"松黑木吉丁"、"松迹地吉丁"、"松黑吉丁"或"迹地吉丁",所用的学名均是Melanophila acuminata。通过对采集于北京油松上的吉丁标本进行鉴定,发现这是一个误订,正确的学名应为松阴吉丁Phaenops yin Kubáňet Bíly,2009,分布于我国北京、陕西、甘肃、河北和山西。本文描述了松阴吉丁的形态特征,及与相近种的区分,并提供了松阴吉丁及迹地吉丁Melanophila acuminata的彩图。     

组织培养条件下喜树叶片细胞壁酸性降解的pH值观察

喜树叶片外植体经组织培养第13 d和23 d后进行解剖学观察,同时比较正常叶片的解剖学特征,发现在pH值为5.8的酸性培养基中,喜树叶片外植体中的海绵组织和栅栏组织等薄壁细胞相继发生明显的细胞壁降解,而表皮细胞发生较微弱的细胞壁降解现象;进行组织培养前,正常叶片的各类细胞未观察到细胞壁降解现象发生。应用BCECF-AM pH荧光探标记并采用激光共聚焦在480 nm波长下进行pH值测定发现,喜树叶片外植体经组织培养第13 d和第23 d后的海绵组织和栅栏组织等薄壁细胞部位的pH值均为5.2,但表皮细胞部位的pH值则为5.7~5.8,而正常叶片各类细胞的pH值平均为5.7。这说明, pH值为5.8的酸性培养基和喜树叶片薄壁细胞内的酸性成分自泌可能共同诱导了其细胞壁的酸性降解。     

Trichomes in Camptotheca acuminata Decaisne (Nyssaceae): Morphology,distribution, structure,and secretion

Camptotheca acuminata is a main source of the anti-cancer drug camptothecin (CPT). In this species, several studies have observed non-glandular trichomes (NGTs) and glandular trichomes (GTs). It has been assumed that GTs contain CPT, yet this has not been proven and no information is available on the accumulation of other secondary metabolites. The objective of this study was to describe the morphology, distribution and structure of C. acuminata trichomes and to investigate the chemical nature of the substances secreted by GTs. Light and fluorescence microscope, scanning electron microscope (SEM) and transmission electron microscope (TEM) were used to determine the morphology, distribution and structure of GTs and NGTs. Thin layer chromatography (TLC) and high-performance liquid chromatography (HPLC) analyses were carried out to confirm the presence of CPT in GTs, and histochemical tests were performed to investigate the presence of other secondary metabolites. C. acuminata possesses two types of GTs (GT1 and GT2), which differ in terms of their morphology, pattern of distribution and accumulated substances. The chemical analyses demonstrated that both GT1 and GT2 accumulate CPT. Histochemical analysis showed that phenols accumulate in the vacuole of GT2s. No isoprenoids were detected in GTs.     

Metabolite profiling of alkaloids and strictosidine synthase activity in camptothecin producing plants

Camptothecin derivatives are clinically used anti-neoplastic alkaloids that biogenetically belong to monoterpenoid indole alkaloids. Camptothecin-related alkaloids from the methanol extracts of Ophiorrhiza pumila, Camptotheca acuminata and Nothapodytes foetida plants were profiled and identified using a reverse-phase high performance liquid chromatography coupled with on-line photodiode array detection and electrospray-ionization ion-trap mass spectrometry. A natural 10-glycosyloxy camptothecin, chaboside, was accumulated in tissues of O. pumila but not in C. acuminata and N. foetida. Anthraquinones regarded as phytoalexins were present in the extracts of hairy roots and calli but not in the differentiated plants of O. pumila. These findings demonstrated a remarkable difference in the constituents between the differentiated plants and the hairy roots or calli tissues. The activity of strictosidine synthase, a key enzyme of camptothecin biosynthesis, was detected in the protein extracts of stems and roots of O. pumila, being correlated with the pattern of strictosidine synthase mRNA expression.     

Improvement of growth and camptothecin yield by altering nitrogen source supply in cell suspension cultures of Camptotheca acuminata

Nitrate at 70m gave the highest biomass of Camptotheca acuminata in suspension culture in MS medium, but a NH₄ ⁺/NO₃ ^– molar ratio of 5:1 (giving a total of 40 m N) gave the maximum camptothecin yield. A two-stage flask culture system was established to improve culture efficiency; cell dry weight, camptothecin content and yield was increased by 30%, 280% and 340%, respectively when compared with those of control, reaching up to 36g l^–1, 0.36mgg^–1, and 12.8mgl^–1, respectively.     

喜树种子挥发油化学成分的研究

利用常压水蒸气蒸馏法提取喜树种子挥发油,得率０．３５％。并进行ＧＣ／ＭＳ／ＤＳ研究研究,鉴定出占挥发油总组成８３．７４％和５５种成分的化学结构,其主要成分为：反－芳樟醇氧化物、苯乙醇、顺－芳樟醇氧化物、大牦牛儿烯、等。     

增强UV-B辐射对喜树幼苗生物量和两种生物碱含量的影响

通过盆栽试验,以自然辐射为对照,研究人工增强UV-B辐射下(5.0 μW/cm2)喜树幼苗生物量和各器官中喜树碱、10-羟基喜树碱含量的变化,试验结果表明:(1 )UV-B辐射处理前20d,处理组幼苗的全株以及各器官的鲜重与对照相差很少.至辐射处理的40d,处理组幼苗的全株以及各器官的生物量均高于正常条件的幼苗.试验后期,处理组单株生物量降低,长时间的UV-B辐射使喜树植株矮化、基茎加粗,同时还改变了喜树生长过程中干物质的分配,较多的干物质分配到喜树的茎和根中,而较少进入叶中;(2)UV-B辐射增强能明显增加喜树地上器官中喜树碱的含量,而对10-羟基喜树碱含量影响不明显.(3)各器官中生物碱含量与生物量的积累速率具有一定的相关性,生物量增长过快时单位质量植物体中的生物碱含量下降.     

涝渍胁迫对喜树幼苗形态和生理的影响

为了解喜树对涝渍土壤条件的适应性, 通过温室土培盆栽试验, 从形态和生理两方面探讨了涝渍胁迫对1年生喜树(Camptotheca acuminata)实生苗的影响, 试验分对照、轻度渍水、渍水和淹水等4个处理, 处理时间为 21 d。结果表明, 喜树根系生长以轻度渍水最好, 其次为对照, 而渍水和淹水的处理初生根系逐渐腐烂、死亡, 与之相对应, 轻度渍水处理的根系活力与对照无显著差异, 但渍水和淹水处理随着处理时间的延长逐渐下降, 同时观察到渍水和淹水的喜树茎在水面以下部位出现较多的皮孔。随着处理时间的延长, 轻度渍水处理喜树叶片内POD和SOD活性一直保持较高水平, 渍水和淹水处理则表现出先升高后下降的趋势, 各个处理叶片中O₂^-· 、H₂O₂和MDA含量有逐渐升高的趋势, 且各处理之间均表现为淹水>渍水>轻度淹水>对照。各处理根系LDH活性在处理前期较对照低, 而在处理的中后期, LDH活性均比对照高, 以渍水和淹水处理最高。因此, 喜树在轻度渍水条件下, 一方面由于皮孔和不定根的增多, 根系能够获得更多的氧气, 另一方面由于POD和SOD抗氧化酶活性的增强, 降低了O₂^-·和H₂O₂对细胞的伤害, 喜树能够在轻度渍水的立地上正常生长。