Orthogonal Test Optimization of Extraction Technology of Total Fla-vonoids from Fruits and Leaves of Prunus mume Sieb. et Zucc. and Its in Vitro Anti-oxidation Activity

[Objectives]To optimize the extraction technology of total flavonoids component,and investigate its in vitro anti-oxidation activity.[Methods]The single factor was inspected firstly. By orthogonal experiment,the best extraction conditions of total flavonoids from fruits and leaves of P. mume Sieb. et Zucc. were determined,and reducing ability of the extracted total flavonoids and its DPPH and ABTS scavenging abilities were explored. [Results] The best extraction technology conditions: solid-liquid ratio of 1∶ 50,ethanol concentration of 50%,extraction time of 2. 5 h,extraction temperature of 85 ℃,two-time extraction. By detecting DPPH and ABTS scavenging abilities of total flavonoids,the anti-oxidation activity of the total flavonoids from fruits and leaves of P. mume Sieb. et Zucc. was analyzed and evaluated. [Conclusions]Fruits and leaves of P. mume Sieb. et Zucc. had a certain in vitro anti-oxidation activity,and heat reflux extraction method of its total flavonoids had high extraction rate and simple and convenient operation,which had some practical value.

ISSR Analysis of Genetic Relationship of Germplasm Resource in Prunus mume Sibe. et Zucc

Using ISSR technique to analyze the diversity and genetic relationship of germplasm resources in 39 Prunus mume Sibe. et Zucc., the result showed that 10 primers were screened with high resolution from 51 primers, 120 fragments were amplified, of which 98 were polymorphic loci, accounting for 81.67% of total. Tested materials were divided into 3 classes, as was fundamentally accorded with the traditional classification base on horticulture. There was no obvious difference in geographic relationship among the clustering results.

青木香(Aristolochia debillis Sieb.et Zucc.)的消炎和镇痛活性(英文)

目的:检测青木香(AristolochiadebillisSieb.etZucc.)的块根水提取液的消炎和镇痛效果及毒性。方法:用蛋清诱导法、热板法和扭体法。结果:青木香的块根水提取液无毒性,1000mg/kg,i.g.的青木香块根水提取液比生理盐水的消炎和镇痛效果明显,比200mg/kg,i.g.乙酰水杨酸的消炎效果好,2000mg/kg,i.g.的青木香块根水提取液比乙酰水杨酸的消炎效果更加显著。结论:青木香具有较好消炎和镇痛活性,无毒性。

野生大豆(Glycine soja Sieb.et Zucc.)叶柄维管束分化观察

对野生大豆(Glycine sojaS ieb.et Zucc.)叶柄维管束分化过程进行了观察.结果表明,野生大豆从茎中分出的3个叶迹,在叶柄基部先愈合成维管柱,然后再分开进入叶柄;叶柄维管束在分出3小叶之前,再发生不同程度愈合,形成圆形维管柱进入2个侧小叶柄;剩余的维管束再分离进入中间小叶柄.同样,中间小叶柄维管束在进入叶片之前也要愈合成维管柱后,才逐渐进入叶片,形成叶脉.

野大豆(Glycine soja Sieb.et Zucc.)一新变种

命名发表了野大豆一新变种,即闭花野大豆Glycine soja Sieb.et Zucc.var.cleistogama C.S.Zhu et S.X.Zhu。

A PRELIMINARY STUDY ON THE PHENOLOGY OF TAXUS CUSPIDATA SIEB.ET ZUCC.

In this paper, the phenomena of the life cycle of Taxus cuspidata Sieb. et Zucc., such as germination, leafing, flowing seed mature,new shoot growth etc. were observed and measured. As a result, the phenological regularity of Taxus cuspidata was found initially. At the same time,according to its five-day mean increment and height growth curve, the growing process of new shoot of Taxus cupidata was divided into three periods:slow period peak period and decline period. The paper supplied scientific data for the introduction of the rare and imininent variety, domestication,cultivating, expoiting, utilizing and protecting of Taxus cuspdata.

青木香(Aristolochia debillis Sieb. et Zucc.)的抗菌活性(英文)

通过琼脂平板扩散试验和双倍稀释试验,对青木香(Aristolochia debillis Sieb. et Zucc.)的石油醚提取物、乙酸乙酯提取物和甲醇提取物进行了抗细菌和抗真菌作用的研究,在三种提取物中,只有甲醇提取物对试验的所有革兰氏阳性和革兰氏阴性细菌都表现出显著的抗菌作用,乙酸乙酯提取物只对测试的部分细菌显示出抗菌作用,石油醚提取物不具有抗菌活性.甲醇提取物比乙酸乙酯提取物具有更强的抗菌作用和更广的抗菌谱.三种提取物对所有试验的真菌都没有显示出抗菌作用.

Genetic Relationships among Prunus mume var. pendula Using AFLP Markers

Genetic relationships among Prunus mume var. pendula were studied by using AFLP markers. 18 accessions representing 14 cultivars of Prunus mume var. pendula were selected from the germplasm collection at the Research Center of China Mei Flower. Seven Mse I-EcoR I AFLP primer combinations revealed 450 legible bands, and 269 of which were polymorphic markers. A similarity matrix was prepared using the simple matching coefficient of similarity and Neis (72) distance coefficient. A UPGMA dendrogram demonstrated the genetic relationships of the cultivars. The information given by AFLP markers was basically consistent with the morphological classification and the evolutionary history of the morphotypes, and roughly supported the new revised classification system for Chinese Mei Cultivars. But there were still several exceptions: 1) the Guhong Chuizhi inserted between the Tiaoxue Chuizhi and the Danfen Chuizhi; 2) the Wufu Chuizhi kept off the Pink Pendant Form, and the Moshan Chuizhi was removed from Viridiflora Pendant Form; 3) the Danbi Chuizhi and the Shuangbi Chuizhi of Viridiflora Pendant Form got together well but fell within the Pink Pendant Form.

Structure Investigation of a New Alkaloid from Zanthoxylum Schinifolium Sieb et Zucc

There are more than 250 kinds of Zanthoxylum plants in the world, of which about 40 species grow in China. In last 30 years, nearly 80 kinds of the plants have been studicd intensively and the results show that they are of different chemical compositions and various physiological actions, such as having anthelmintic action, bacteriostasis,

Microscopic Identification of Zhuang Medicine Laportea bulbifera(Sieb. et Zucc.) Wedd.

[Objectives] This study was conducted to investigate the microscopic identification characteristics of Laportea bulbifera(Sieb. et Zucc.) Wedd. [Methods] The characteristics of L. bulbifera leaf, stem and powder were identified by microscopy. [Results] The main microscopic identification characteristics of the tissue structures of L. bulbifera leaf and stem at different positions and L. bulbifera powder were determined. [Conclusions] The microscopic identification results are characteristic, and could serve as the identification basis of L. bulbifera. These results could provide references for the quality control of L. bulbifera.

Optimization of extraction and determination of emodin from Polygonum cuspidatum Sieb. et Zucc. products by HPLC-DAD

A uniform experimental design procedure was used to investigate the effects of some operating parameters on the extraction of emodin from Polygonum cuspidatum Sieb. et Zucc. products. Variables tested were volume ratio of material to solvent, size of material, extraction time and temperature and composition of extraction solvent （mixtures of acetone-water）. Each variable was tested at seven levels; 7 experiments were performed in random order. Analyses of the extracts were performed by high-performance liquid chromatography with diode array detection（HPLC-DAD）. Analytical responses were processed by using a forward regression analysis, in order to find polynomial function describing the relationship between variables and responses. For all the analytes the experimental conditions for providing the highest extraction yield inside the experimental domain considered were found. Finally, a simple, rapid and accurate analytical method was developed for the determination of emodin by high performance liquid chromatography. The separation is achieved within 25 rain on an ODS column using methanol and water as gradient mobiles. Emodin can be quantified by using external standard method detecting at 436 nm. Good linearity is obtained with correlation coefficient exceeding 0.9986 and the detection limit and the quantification limit are 1.53 and 3.23 mg/L respectively. This method shows good reproducibility for the quantification of the emodin with intra-day and inter-day relative standard deviation less than 2.3% and 5.6% respectively. Under optimized extraction conditions, the recovery of the standard is 96.5%. The validated method is successfully applied to quantify the emodin in seven Polygonum cuspidatum sieb. Et zucc. products, which provided an idea for the pre-treatment of determination of active compounds in traditional Chinese medicines.

Study on Quality Standards of Laportea bulbifera(Sieb. et Zucc.) Wedd. in Yao’s Folk-medicine

[Objectives] This study was conducted to establish the quality control standards for Laportea bulbifera(Sieb. et Zucc.) Wedd. [Methods] Microscopic identification and thin layer chromatography were used to identify and determine the moisture, total ash and extract contents. [Results] According to the microscopic results, the stem transection is round and has five longitudinal edges at which the cell wall is obviously thickened. Bristles can be observed. A series of collenchyma cells are arranged closely inside the epidermis. There are several rows of parenchyma cells. The vascular bundles are of different sizes;and the ray part is wide, and at the junction of the ray and the cortex, there are often scattered secretory cells. The center is a broad medulla. The powder is brown. There are many starch grains. The bristles are unicelluar, and have warty protrusions and expanded base. The fibers are bundled, and the cells are large. The epidermal cells are in the shape of an elongated spindle, in a mosaic arrangement, and the cell wall is slightly thickened. From the thin layer identification chromatograms of tested L. bulbifera, it can be seen that spots were observed at the same positions as the control medicinal material, and the durability of the method was good. The extract content should not be less than 12.00% by hot extraction with water as solvent, and not be less than 3.00% by cold extraction with ethanol as solvent. The moisture content should not be more than 16.50%. The total ash content should not be more than 14.00%. The acid-insoluble ash content should not be more than 2.50%. [Conclusions] The microscopic identification results are reliable and can be used as the identification basis of L. bulbifera. The thin layer identification method and the test results can provide a basis for quality control of L. bulbifera.

DETERMINATION OF POLYDATIN IN POLYGONUM CUSPIDTUM SIEB. ET ZUCC. BY TLC-FLUORESCENCE SPECTROPHOTOMETRY

Objective A method of TLC-fluorescence spectrophotometry was established to assay the content of polydatin in polygonum cuspidatum sieb. et zucc. Methods: Polydatin was extracted by methanol and separated with chloroform-acetone-formic acid-water (4∶4∶0.5∶0.2) by thin layer chromatography. The excitation wavelength and emission wavelength were 284 nm and 384 nm, respectively. Results The linear regression equation of the calibration graph was y=7.02179x+4.5143, a linear regression correlative coefficient r=0.9936. Conclusion This method was proved simple, stable and sensitive. It can be used in quality control of herbs.

Anticancer Effects of Crude Extract from Melia toosendan Sieb.et Zucc on Hepatocellular Carcinoma In Vitro and In Vivo

Objective： To investigate the anti-cancer effects of crude extract from Melia toosendan Sieb. et Zucc and its possible molecular mechanisms in vitro and in vivo. Methods： Transonic alcohol-chloroform extraction method was used to extract toosendanin from the bark of Melia toosendan Sieb. et Zucc, and the content of toosendanin in the crude extract was measured by high performance liquid chromatography （HPLC）. Anti-cancer effects of crude extract from Melia toosendan Sieb. et Zucc were investigated in in vivo and in vitro studies. In the in vitro experiment, human hepatocellular carcinoma cell lines SMMC-7721 and Hep3B were co-incubated with toosendanin crude extract of different concentrations, respectively. In the in vivo experiment, BALB/c mice were subcutaneously inoculated with mouse hepatocellular carcinoma H22 cells and treated with crude extract. Results： HPLC revealed the content of toosendanin was about 15%. Crude extract from Melia toosendan Sieb. et Zucc inhibited cancer cells growth in a dose- and time-dependent manner. The 50% inhibitory concentration （IC50, 72 h） was 0.6 mg/L for SMMC-7721 cells and 0.8 mg/L for Hep3B cells. Both high-dose [0.69 mg/（kg·d）] and low-dose [0.138 mg/（kg·d）] crude extract could markedly suppress cancer growth, and the inhibition rate was greater than 50%. Hematoxylin and eosin staining showed necrotic area in cancers and transmission electron microscopy displayed necrotic and apoptotic cancer cells with apoptotic bodies. Immunohistochemistry showed that the expression of Bax and Fas increased and the expression of Bcl-2 reduced. Conclusions： Toosendanin extract has potent anti-cancer effects via suppressing proliferation and inducing apoptosis of cancer cells in vivo and in vitro. The mechanism of apoptosis involves in mitochondrial pathway and death receptor pathway.

梅花扦插繁殖研究进展

阐述了梅花(Prunus mume Sieb.)扦插繁殖中梅花树种的遗传差异、插穗生长程度、扦插繁殖时间的差异、基质选择以及不同类型植物生长调节剂对成活率的影响,讨论了种植后环境对扦插繁殖过程的影响,对扦插繁殖存在的问题提出了一些建议,旨在进一步促进梅花扦插繁殖的发展。

中国栽培和野生大豆农艺品质性状与SSR标记的关联分析 I.群体结构及关联标记

关联作图是一种利用连锁不平衡(linkage disequilibrium,LD)检测自然群体中基因位点及其等位变异的方法。利用60个SSR标记,对全国大豆地方品种群体(393份代表性材料)和野生大豆群体(196份代表性材料)的基因组变异进行扫描,分析两类群体的连锁不平衡位点、群体结构,并采用TASSEL软件的GLM(general linear model)方法对16个农艺、品质性状观测值进行标记与性状的关联分析。结果表明:(1)在公共图谱上不论共线性的或是非共线性的SSR位点组合都有一定程度的LD,说明历史上发生过连锁群间的重组;栽培群体的连锁不平衡成对位点数较野生群体多,但野生群体位点间连锁不平衡程度高,随距离的衰减慢。(2)群体SSR数据遗传结构分析发现,栽培群体和野生群体分别由9和4个亚群体组成,亚群的划分与群体地理生态类型相关联,证实地理生态类型划分有其遗传基础。(3)栽培群体中累计有27个位点与性状相关;野生大豆种质中累计有34个位点与性状相关。部分标记在两类群体中都表现与同一性状关联,检出的位点有一致性,也有互补性;一些标记同时与2个或多个性状相关联,可能是性状相关乃至一因多效的遗传基础;关联位点中累计有24位点(次)与遗传群体连锁分析定位的QTL一致。

中国栽培和野生大豆农艺及品质性状与SSR标记的关联分析--Ⅱ．优异等位变异的发掘

在前文研究已检出与农艺品质性状显著关联的SSR位点的基础上,本文进一步对与性状关联位点的等位变异作解析,通过将携带某等位变异的所有材料表型均值与携带无效等位基因(null allele)材料表型均值做比较,估计等位变异的潜在表型效应增量(减量),进一步利用该信息估计位点增效(减效)等位变异的平均效应,鉴别出一批农艺品质性状优异位点、等位变异及携带优异等位变异的载体材料。发现在栽培及野生种质中检出的优异等位变异有同、有异、有互补性。发现关联位点正、负效应等位变异均值间有差异,可根据育种目标性状选择要求,选取适合的位点及相应等位变异。同一标记位点可与多性状关联,其等位变异在不同性状间各有其表型效应的方向和大小;等位变异在相关性状效应上方向、大小的异同解释了性状间正、负相关的遗传原因。关联作图得到的信息可以弥补家系连锁法QTL定位信息的不足,并直接利用等位变异信息进行亲本选拔、组合选配及后代等位条带辅助选择以提高育种成效。

中国野生大豆群体农艺加工性状与SSR关联分析和特异材料的遗传构成

选用204对SSR标记对全国野生大豆群体(174份代表性样本)的基因组扫描,采用TASSEL软件的GLM(general linear model)方法对百粒重、开花期、成熟期、干豆腐得率、干豆乳得率和耐淹性性状值关联分析,解析与性状关联位点的优异等位变异,鉴别出一批与农艺、加工性状关联的优异等位变异及携带优异等位变异的载体材料;进一步分析极值表型材料的遗传构成。结果表明:(1)累计51个位点(次)与性状关联,有些标记同时与2个或多个性状相关联,可能是性状相关的遗传基础;关联位点中累计16位点(次)与连锁分析定位的QTL一致;(2)与地方品种群体和育成品种群体的关联位点比较,发现野生群体关联位点只有少数与之相同,群体间育种性状的遗传结构有明显差异。(3)与多性状关联的位点其等位变异对不同性状的效应方向可相同可不同,如GMES5532a-A332对百粒重和耐淹性的相对死苗率都是增效效应,而GMES5532a-A344对百粒重是减效效应,对相对死苗率是增效效应;(4)极值表型材料间的遗传构成有很大差异。表型值大的材料携带较多增效效应大的位点等位变异,例如N23349的百粒重是9.08g,含有4个增效效应较大的位点等位变异;表型值小的材料携带较多减效效应大的位点等位变异,如N23387的百粒重是0.75g,含有4个减效效应较大的位点等位变异。关联作图得到的信息可以弥补连锁定位信息的不足,尤其是全基因组位点上复等位变异的信息为育种提供了亲本选配和后代等位条带辅助选择的依据。

中国野生大豆群体特征和地理分化的遗传分析

【目的】从分子标记等位变异水平上探讨中国野生大豆群体的遗传特征、连锁不平衡特点和地理生态分化的遗传机制,并以重要生态性状全生育期为代表解析性状地理分化的遗传基础。【方法】从全国24个省区不同地理生态型的野生大豆材料中抽选174份组成代表性样本,选用204个SSR标记,利用TASSEL及STRUCTURE 2.2软件进行群体连锁不平衡(linkage disequilibrium,LD)和群体遗传结构分析。在此基础上对群体的地理分化、亚群体特异性及全生育期位点等位变异的地理分化进行遗传分析。【结果】中国野生大豆群体蕴含丰富的遗传变异,20条连锁群中,I和C2连锁群有相对较多的位点平均等位变异和遗传分化。不论是共线性组合,还是非共线性组合,都有一定程度的LD存在,说明历史上发生过连锁群间的大量重组;野生群体D′平均值为0.34,高值多,比栽培大豆高,说明野生群体发生过更多的重组,保留下较高的LD。采用H-W平衡模型将野生群体聚成4类,模型聚类亚群划分与地理生态分类相关、有交叉,推测各地理亚群体发生过材料的迁移。各地理亚群经长期自然选择,各位点等位基因的频率发生变化,还有新生的与绝灭的,因此,各地理亚群间产生明显的等位基因分化。与地理生态性状全生育期关联的有15个位点160个等位变异,其中,亚群特有等位变异共58个,来自14个位点,同一位点可以在多个亚群体产生不同的特有等位变异,其效应从南至北逐渐下降,这说明生态区间不仅有强烈的遗传分化,而且是有规律的遗传分化。【结论】中国野生大豆群体遗传多样性高,共线、非共线位点间连锁不平衡程度高,4个生态亚群体间位点高度遗传分化,产生大量地区特有等位变异,全生育期还表现由南向北降效的规律性遗传分化。

多环境下野生大豆染色体片段代换系群体农艺性状相关QTL/片段的鉴定

【目的】改进染色体片段代换系群体,挖掘野生大豆(Glycine soja Sieb.et Zucc.)中蕴藏的农艺性状优异等位变异,为拓宽栽培大豆(Glycine max(L.)Merr.)的遗传基础提供材料和依据。【方法】通过标记加密和剔除部分单标记型片段的方法,改进以野生大豆N24852为供体,栽培大豆NN1138-2为受体的染色体片段代换系(CSSL)群体Soja CSSLP1;对改进后的群体(Soja CSSLP2)进行3年2点田间试验,通过单标记分析、区间作图、完备复合区间作图和基于混合线性模型的复合区间作图等4种定位方法,结合与轮回亲本有显著差异的染色体片段代换系间相互比对,检测与大豆开花期、株高、主茎节数、单株荚数、百粒重和单株粒重相关的野生片段。【结果】改进后的群体(Soja CSSLP2)由150个CSSL构成,其中,有130个家系与Soja CSSLP1相同;在原遗传图谱上,新增40个SSR标记,相邻标记间平均遗传距离由16.15 c M变为12.91 c M,大于20 c M的区段由32个减少至17个,标记覆盖遗传距离总长度较原图谱(2 063.04 c M)增加103.52 c M;群体NN1138-2背景回复率变幅为79.45%—99.70%,平均为94.62%。利用Soja CSSLP2群体,分别鉴定到与开花期、株高、主茎节数、单株荚数、百粒重和单株粒重相关的4、5、5、7、14和3个工作QTL(working QTL)/片段,其中有15个工作QTL/片段能在多个环境下检测到,属共性工作QTL(joint working QTL);除片段Sct_190—Sat_293上的主茎节数位点外,野生等位变异具有的加性效应方向与双亲表型差异方向一致;单个位点分别能解释5%—64%的表型变异;同时,分别检测到3、2和2个与地点存在互作的株高、主茎节数和单株荚数QTL/片段,其中与凤阳环境的互作均具有增加表型的效应,这可能与凤阳较南京所处纬度高有关;这些位点/片段分布在26个染色体片段上,其中有7个片段与2个及以上性状相关,可能是性状相关的遗传基础;与前人结果比较,有3个开花期、3个株高、2个主茎节数、2个单株荚数、8个百粒重、2个单株粒重位点能在其他遗传背景栽培大豆中检测到,说明在这些位点上野生大豆和栽培大豆间及栽培大豆间均存在遗传差异;另外18个位点(片段)为本研究利用野生大豆的新发现。【结论】大豆开花期、株高和主茎节数的遗传基础较百粒重简单,前者均存在效应较大位点/片段,后者多由小效应位点控制,遗传基础极为复杂;野生大豆中蕴藏着新的等位变异,能拓宽栽培大豆遗传基础。