MODULATION BY ACANTHOSPERMUM AUSTRALE EXTRACTS OF THE TUMOR INDUCED HEMATOPOIETIC CHANGES IN MICE

ABSTRACT

Previous studies on the Ehrlich ascites tumor (EAT) model indicate that tumor progression is associated with reduced myelopoiesis and increased extramedullar hematopoiesis. In order to investigate the in vivo antitumor activity of Acanthospermum australe, its hydroalcoholic extract was partitioned with different solvents and the resulting extracts were monitored by their effects on bone marrow and spleen hematopoietic progenitor cell proliferation and differentiation in EAT-bearing mice. Oral treatment of tumor-bearing mice with 3 doses of 100, 500 and 1000 mg/kg of the crude hydroalcoholic extract and its chloroformic, butanolic and aqueous fractions significantly stimulated myelepoiesis and brought extramedullar hematopoiesis back to near control values. In normal mice, stimulation of myelopoiesis was only observed with the crude and the butanolic extracts. All the extracts at 500 mg/kg significantly increased survival of tumor-bearing mice, however a clear survival advantage in the group treated with the butanolic extract was observed. These results suggest that A. australe may exert effects on myelopoiesis that may be implicated in antitumor immune responses.     

一株产长春新碱内生真菌的初步研究

目的 通过长春花叶内生真菌的分离，筛选产生长春新碱的菌株。方法 从长春花叶中分离内生真菌，对其发酵提取物进行TLC和HPLC分析。结果 从长春花叶中分离筛选到一株无孢菌群菌株：97CY3。它能产生长春新碱，HPLC测定其长春新碱含量约为0.205μg／L。结论 长春花内生真菌中，有的菌株可产生与宿主所产相同的抗癌物质长春新碱。     

长春花中长春碱含量测定方法的研究

研究了薄层扫描法及薄层－紫外分光光度法测定长春花中长春碱含量的两种方法，在硅胶Ｆ２５４－ＣＭＣ－Ｎａ碱性板上（ＮａＯＨ浓度为０．１ｍｏｌ／Ｌ），以氯仿－甲醇－石油醚（９：１：５）为展开剂，分离了长春花提取液中长春碱和长春新碱等１４种成分。前法采用双波长薄层扫描，λｓ＝２８０ｎｍ，λＲ＝３５０ｎｍ，线性范围在２￣１０μｇ。后法采用薄层分离后，样品带用甲醇洗脱后，紫外分光光度法测定含量。后法在５￣２５     

长春花及银杏植物细胞悬浮培养对青蒿素的生物转化研究

目的：对抗疟药物青蒿素（Ⅰ）进行了生物转化研究。方法：利用长春花及银杏植物细胞悬浮培养细胞进行生物转化。用硅胶柱色谱进行产物的分离，波谱方法鉴定产物的结构。结果：此两种植物悬浮细胞体系均能将青蒿素转化成3α-羟基去氧青蒿素（Ⅱ）。结论：此两种植物悬浮细胞体系均有有效转化青蒿素。     

Simultaneous quantitative determination of five alkaloids in Catharanthus roseus by HPLC-ESI-MS/MS

AIM： To establish a method to simultaneously determine the main five alkaloids of Catharanthus roseus for trace samples, a high-performance liquid chromatography–electrospray ionization-tandem mass spectrometry（HPLC-ESI-MS/MS） analysis method was developed. METHOD： The five Catharanthus alkaloids, vinblastine, vincristine, vinleurosine, vindoline, and catharanthine were chromatographically separated on a C18 HPLC column. The mobile phase was methanol-15 nmol？L–1 ammonium acetate containing 0.02% formic acid（65 ： 35, V/V）. The quantification of these alkaloids was based on the Multiple Reaction Monitoring（MRM） mode. RESULTS： This method was validated, and the results achieved the aims of the study. The intra- and inter-day precision and accuracy of the five alkaloids were within 1.2%-11.5%（RSD%） and-10.9%-10.5%（RE%）. The recovery rates of the five alkaloids of samples were from 79.9% to 91.5%. The five analytes were stable at room temperature for 2 h, at 4 °C for 12 h, and at-20 °C for two weeks. The developed method was applied successfully to determine the content of the five alkaloids in three plant parts of three batches of C. roseus with a minute amount collected from three regions of China. CONCLUSION： The HPLC-ESI-MS/MS method can be used for the simultaneous determination of five important alkaloids in trace C. roseus samples.     

提高长春花冠瘿细胞中吲哚生物碱含量的研究

研究了长春花冠瘿细胞接种物年龄与接种量对悬浮物培养细胞生物产量的影响,探讨了以大丽花轮枝孢菌的匀浆物为外源刺激物处理冠瘿细胞,对细胞生长和吲哚生物碱积累的作用,确立了外源刺激物产生效应的最适条件。     

长春花毛状根再生植株的获得及抗癌生物碱的产生

目的建立长春花转基因毛状根再生植株快速繁育体系,获得生物碱量提高的长春花再生植株。方法研究外源植物生长调节剂对长春花毛状根愈伤组织诱导、不定芽分化、不定根分化和再生植株移栽的影响,采用HPLC法测定长春花再生植株生物碱量,并采用定量PCR技术分析目的基因的表达情况。结果转基因长春花毛状根愈伤组织诱导和不定芽诱导的最佳培养基均是MS+6-BA 2.0 mg/L+NAA 0.3 mg/L,不定根分化的最适培养基是1/2 MS+NAA 0.3 mg/L。采用炼苗的改进方法可使长春花再生植株的移栽成活率达90%。HPLC分析结果表明,长春花再生植株中长春碱、长春新碱和阿吗碱量分别比对照提高4.0、5.8、1.8倍,其中,优良单株OG30-3的长春新碱和长春碱量比对照提高7倍和10倍。定量PCR分析结果表明,转基因植株orca3基因和g10h基因的表达量比非转基因植株的高。结论转基因长春花毛状根再生植株可促进抗癌生物碱的产生。     

长春花及银杏植物细胞悬浮培养对青蒿素的生物转化研究

目的：对抗疟药物青蒿素（Ⅰ）进行了生物转化研究。方法：利用长春花及银杏植物细胞悬浮培养细胞进行生物转化。用硅胶柱色谱进行产物的分离，波谱方法鉴定产物的结构。结果：此两种植物悬浮细胞体系均能将青蒿素转化成3α-羟基去氧青蒿素（Ⅱ）。结论：此两种植物悬浮细胞体系均有有效转化青蒿素。     

长春花中长春质碱含量与农艺性状的分析

目的:对国内外长春花主栽品种的长春质碱含量和农艺性状进行了分析.方法:记录长春花盛花期的农艺性状,反相高效认相色谱仪测定长春质碱含量.结果:发现不同品种的长春质碱含量和农艺性状均存在很大差异,太平洋系列(Pacifica)品种Polka Dot(PPD)中长春质碱质量分数最高,达到叶片干重的3.79 mg·g~(-1);清凉系列(cooler)品种Pink(CP)中长春质碱质量分数最低,仅为叶片干重的0.9 mg·g~(-1).长春花中长春质碱含量与农艺性状之间存在一定程度的相关性.通径分析表明,长春花主要农艺性状中,节间距对长春质碱含量的正影响达到了显著水平(P<0.05),通径系数为1.473.结论:本研究对高含量长春质碱长春花引种和育种具有参考价值.     

EMS诱变的长春花细胞系突变研究

目的 通过化学诱变的手段筛选生长快且吲哚碱含量高的长春花突变细胞系。方法 用不同浓度的甲基磺酸乙酯(EMS)处理长春花愈伤组织，挑选成活愈伤组织块继代扩大培养，比较其与对照愈伤组织生长速率、吲哚总碱积累等方面的差异。结果 用EMS处理的长春花愈伤组织与对照相比，不仅生长快，并且吲哚总碱含量高。结论 EMS处理的愈伤组织发生了变异，变异种是所期望的比较理想的细胞系。