Effects of Inoculation with Phosphate Solubilizing Bacteria on the Physiology,Biochemistry,and Expression of Genes Related to the Protective Enzyme System of Fritillaria taipaiensis P.Y.Li

Fritillaria taipaiensis P.Y.Li is a widely used medicinal herb in treating pulmonary diseases.In recent years,its wild resources have become scarce,and the demand for efficient artificial cultivation has significantly increased.This article is the first to apply phosphate solubilizing bacteria isolated from the rhizosphere soil of F.taipaiensis P.Y.Li to the cultivation process of F.taipaiensis P.Y.Li.The aim is to identify suitable reference strains for the artificial cultivation and industrial development of F.taipaiensis P.Y.Li by examining the effects of various phosphate solubilizing bacteria and their combinations on photosynthesis,physiological and biochemical properties,and gene expression related to the protective enzyme system in F.taipaiensis P.Y.Li.The experiment,conducted in pots at room temperature,included a control group(CK)and groups inoculated with inorganic phosphorussolubilizing bacteria:W1(Bacillus cereus),W2(Serratia plymuthica),W12(Bacillus cereus and Serratia plymuthica),and groups inoculated with organophosphorus-solubilizing bacteria:Y1(Bacillus cereus),Y2(Bacillus cereus),Y12(Bacillus cereus and Bacillus cereus),totaling seven groups.Compared to CK,most growth indices in the bacterial addition groups showed significant differences,with W12 achieving the highest values in all indices except the leaf area index.The content of photosynthetic pigments,photosynthetic parameters,and osmoregulatory substances increased variably in each bacterial treatment group.W12 exhibited the highest content of chlorophyll a and soluble protein,while W1 had the highest free proline content.The activities of peroxidase(POD),superoxide dismutase(SOD),and catalase(CAT)in all inoculated groups were higher than in CK,with significant changes in SOD and CAT activities.The malondialdehyde(MDA)content in all inoculated groups was lower than in CK,with Y12 being the lowest,at approximately 30%of CK.Gene expression corresponding to these three enzymes also increased variably,with POD expression in Y2 being the highest at 2.73 times that of CK.SOD and CAT expression in Y12 were the highest,at 1.84 and 4.39 times that of CK,respectively.These results indicate that inoculating phosphate solubilizing bacteria can enhance the growth of F.taipaiensis P.Y.Li,with the mixed inoculation groups W12 and Y12 demonstrating superior effects.This lays a theoretical foundation for selecting bacterial fertilizers in the cultivation process of F.taipaiensis P.Y.Li.

遮荫下高原濒危药用植物川贝母(Fritillaria cirrhosa)光合作用和叶绿素荧光特征

测量了川贝母遮荫和全光下叶片光合生理参数和叶绿素荧光参数,探讨了其遮荫下生理生态学指标的变化和强光下的光适应机制,且对光合生理的测量过程给出了全面系统的介绍。川贝母日光合曲线呈双峰变化,遮荫后川贝母光合效率(Pmax)、细胞内CO2浓度(Ci)分别增加了31.1%、10%(p<0.01),有效光反应光强域值(PFD)增加了331.5[μmol/(m2.s)],最大表观量子效率(AQY)和气孔导度(COND)也有所增加,暗呼吸速率(Rd)和叶片蒸汽压亏缺(Vpdl)明显降低,此均有利用光合产物的积累;F′v/F′m(光照下反应中心能量捕获效率)、qP(光化学淬灭)、ETR(电子传递效率)、Psips2(PSⅡ的效率)分别增加了14.7%(p<0.01)、8.8%(p<0.01)、10%(p<0.01)、24.2%(p>0.05),促进了川贝母叶片对光能的利用;Fv/Fm(光化学效率)变化不明显,说明"灯笼花"阶段,全光下川贝母没有受到到明显的胁迫,光合机构未遭到破坏。当自然光强超过光饱和点时,川贝母主要通过提高非化学淬灭,耗散过多吸收的热能以防止光合机构的破坏。当出现极端气候(高温干旱)时,川贝母繁育退化现象("树儿子"或"灯笼花"阶段退回到"一匹叶"生长阶段)可能是对环境适应的主要方式。另外川贝母叶片狭小、叶片倾角较大降低了强光辐射的有效面积,在形态结构上有利于避免过高光强对叶光合器官的损伤。

秋水仙素诱导川贝母(Fritillaria cirrhosa D.Don)愈伤组织多倍体的研究

应用组织培养技术对川贝母 (Fritillaria cirrhosa D.Don)进行多倍体诱导。结果表明 :组织培养条件下 ,将川贝母愈伤组织在培养基中添加一定浓度的秋水仙素处理一段时间 ,或者经一定浓度的秋水仙素浸泡一段时间后再培养 ,均可诱发川贝母多倍体的产生 ,但以前者效果较好 ,在秋水仙素浓度为 10 0 0 mg/ L ,处理 5 d的条件下 ,诱导率最高达 70 %。细胞染色体鉴定结果为 :四倍体染色体数为 2 n=4x=48,而二倍体的染色体数为 2 n=2 x=2 4。

甘肃马唧山地区榆中贝母(Fritillaria yuzhongensis)的生态学特性及种子发芽规律的初步研究

榆中贝母(Fritillaria yuzhongensis)生长于马啣山的高山地带,全生育期约100 d左右,是一种生育期较短的类短命植物。在果实成熟时,种子的胚发育不全,具有后熟作用的特点,包括形态后熟和生理后热两个阶段;因此,需要一定时期的低温处理,才能完成后熟过程,以解除休眠,促使种子发芽。本试验表明:榆中贝母完成形态后熟过程的最佳温度是7℃左右,经过100d以上的低温处理才能使原胚发育成熟,继续在7℃低温下处理70d以上,才能完成生理后熟过程。榆中贝母种子休眠时间长,需要经过170d以上的低温处理,种子才能萌发。本试验结果为榆中贝母种子的萌发提出了适宜的温度范围和低温处理时间,为提高种子的出苗率和缩短栽培年限提供了科学依据。

Cytotoxic effect of methanolic extracts of Fritillaria imperialis bulbs and Eryngium caucasicum leaves on hepatoma and colon cancer cells

Objective: To evaluate antitumor activities of Fritillaria imperialis and Eryngium caucasicum methanolic extracts on human hepatoma (HepG2) and colon cancer (HCT116) cell lines in comparison to human foreskin fibroblasts as the normal cells. Methods: Methanolic extracts of Fritillaria imperialis and Eryngium caucasicum were prepared by the maceration method. The effect of the extracts at various concentrations (100, 200, 400, 600, and 800 μg/mL) on cell survival was evaluated using the MTT method. Besides, fluorescence staining was used to evaluate death patterns of the cells. Results: MTT assay showed that Fritillaria imperialis significantly decreased the viability of all cell lines after 24 and 48 hours of treatments. However, Eryngium caucasicum extract did not show any significant cytotoxicity effect on the cell lines. Fluorescence staining revealed that Fritillaria imperialis induced apoptosis of HCT116 cells at 550 μg/mL. Conclusions: Fritillaria imperialis extract has antiproliferative and cytotoxic effects on HCT116 and HepG2 cancer cells and therefore, may serve as an anticancer agent.

A new veratramine alkaloid from the bulbs of Fritillaria hupehensis

A new highly conjugated alkaloid of veratramine type, 22S,25S,5α-veratramine-7(8),12(14)-diene-3β,13β,23β-triol-6-one (1), was isolated from the bulbs of Fritillaria hupehensis Hsiao et K.C. Hsia. Its structure was determined on the basis of spectroscopic evidences.

Analysis of Hupehenine in the Total Alkaloids from Fritillaria hupehensis by HPLC-ELSD

A reverse-phase high pressure liquid chromatography （HPLC） method with evaporative light scattering detection （ELSD） has been developed for the quantitative analysis of hupehenine in the total alkaloids from Fritillaria hupehensis. Samples were analyzed on a reverse-phase column （Hypersil C-18） with a mobile phase of methanol：water：chloroform： triethylamine （85：15：1：0.6）. The ELSD was set at the drift tube temperature of 68.3℃ and gas flow rate of 1.8 L/min. Hupehenine＇s retention time was 13.7 min with an asymmetry factor of 1.2. The validity of the method has been verified with linearity, limit of detection, accuracy and precision. The logarithmic linear curve was obtained from 8.936 to 134.04 μg/mL （r=0.9993）. The detection limit （S/N〉3） of hupehenine was 1.79 μg/mL on the column. Intra-day RSD was 1.42% and inter-day RSD was 2.26% （3 days within a week）. The average recovery of hupehenine was 101.50%, and RSD was 1.62%.

药用植物平贝母(Fritillaria ussuriensis)组织培养的研究——Ⅰ.愈伤组织的诱导和器官分化及细胞学观察

本文对平贝母(Fritillaria ussuriensis Maxim)根、鳞茎、叶等器官在 MS,N_6,B_5,WS 和 H5种基本培养基上的培养进行详细观察。通过球状体及不定芽的形态发生曲线及差异显著性测定表明,不同培养基与激素及不同激素浓度与组合,对平贝母器官发生的影响,筛选出诱导高频率球状体与不定芽发生的培养基 MS+1.0 ppmIBA+1.0 ppmZT,MS+0.5 ppmIAA+2.0ppmBA,MS+2.0ppm NAA+0.5 ppmKT。培养过程中球状体与不定芽的形态发生的组织细胞学观察结果表明,球状体发生经过鳞片原基阶段,而不定芽发生不经此阶段;球状体形成后发育成不定芽,但生长情况优于直接产生的不定芽。早期球状体的染色体数目为2n=24。

A new cycloartane triterpenoid from the leaves and stems of Fritillaria hupehensis

A new triterpenoid has been isolated from the leaves and stems of Fritillaria hupehensis Hsiao et K.C. Hsia. Its structure was established as （2329-9,19-cycloart-23-ene-3α,25-diol 1 through chemical and spectroscopic studies including 2D NMR. Another known triterpenoid 9,19-cycloart-25-ene-313,24ζ-diol 2 was also isolated.

Heavy collecting induces smaller and deeper Fritillariae Cirrhosae Bulbus in the wild

Fritillariae Cirrhosae Bulbus is a well-known traditional Chine medicinal herb. However, the demand for this herb is leading to over-collection and its decline in the wild. This study aims to investigate the ecological conditions of Fritillaria cirrhosa under the influence of human disturbance, biotic species interactions and climatic conditions. We established a total of 78 plots at 14 sites of F. cirrhosa in its distribution center, the Hengduan Mountains area. At each site, we estimated the abundance of F. cirrhosa at different distances from roads. The diameter and height of F. cirrhosa fruit and bulbs were measured and compared to underground bulb depth. We then analyzed the effects of environmental conditions and human disturbance on the abundance of F. cirrhosa using variance partitioning. We found that(1)abundance of F. cirrhosa and their underground bulb depth showed a significant linear increase with the distance from the main road;(2) the diameter/height of fruits and the diameter/height of fruits/bulbs showed significantly different responses to the human disturbance;(3) the community associates,climate and spatial conditions can explain 58%, 22% and 27%, respectively, of the variance in the F. cirrhosa abundance. These results highlight the fact that human disturbance and biotic factors have a great influence on the survival of F. cirrhosa, even more than climate conditions.

贝母属(Fritillaria)植物同物异名现象与思考

贝母属(Fritillaria)植物种类丰富,分布区域广泛。在国内外植物分类研究中,很多种类的分类学地位较为混乱,出现了大量同物异名的现象,为该属植物的研究造成巨大障碍。因此以《Flora of China》中记述的贝母属植物种类为基础,整理了我国近些年来不同学者对该属植物的新发现与新归并;并结合英国皇家植物园(邱园)的Plant List数据库中记录的贝母属植物种类,对该属植物的名称进行了详细的梳理,列举了存在"同物异名"问题的贝母属物种。为研究贝母属植物的分类及该属植物的开发与利用,提供科学参考。

Cloning of GF14 Gene from the Bulbs of Fritillaria crrishoa

GF14 widely occurs in plant species functioning as a regulatory protein in cell signal transdution, including monoamine synthesis, exocytosis and cell cycle regulation. A partial sequence, 413 bp in length, is cloned from the bulbs of F. crrihosa by means of RT PCR technique. Sequence alignment with the BLAST program shows that it is highly similar to GF14 gene. A new full length cDNA, 1133 bp, is obtained by RACE techniques. It contains a major open reading frame of 262 amino acids, 5’and 3’ untranslated regions and a long Poly (A) tail. Sequence analysis of deduced GF14 protein from F. crrihosa indicates several conserved motifs of GF14 proteins are also found in sequence of GF14 protein of F. crrihosa.

Current Situation of Fritillaria thunbergii Miq. Production in Yinzhou District of Ningbo City

Yinzhou District of Ningbo City is the home and main production area of Fritillaria thunbergii Miq. Based on the analysis of Fritillaria thunbergii Miq. production in Yinzhou District in 2015,this paper firstly introduced current situation of Fritillaria thunbergii Miq. planting. The competition of Fritillaria thunbergii Miq. products was in disorder,the market price in 2015 dropped to half in 2014,and many Fritillaria thunbergii Miq. growers suffered losses. To keep sustainable development of traditional Fritillaria thunbergii Miq. industry in old production areas and maintain brand and name of genuine materials of " Zhejiang 8 Famous Kinds Herbal Medicines",it is required to actively reduce the inventory,make up short board,strengthen advantages,implement the reform of supply font,keep up with the times and market,and succeed through improving the quality.

SIECHUANSINE,A NEW STEROIDAL ALKALOID FROM FRITILLARIA

From the bulbs of Fritillaria siechuanica S.C.Chen cultivated in Sichuan province,a new steroidal alkaloid,siechuansine,was isolated.Spectral methods demonstrated that it is a 13—epimer of wanpeinine A.

Two New Isosteroid Alkaloids from Fritillaria monatha Migo（Ⅲ）

Two new isosteroid alkaloids, named pengbeimine A and pengbeimine C, were isolated from the fresh bulbs of FritiUaria monatha Migo. Their structures were determined by spectral methods, and were proved by X-ray diffraction as well.

Effects of Different Seedling Raising Methods on Seedling Growth of Two Kinds of Fritillaria

[Objectives] To reveal the effects of film mulching and open ground seedling raising on the differences between Fritillaria przewalskii Maxim. and Fritillaria pallidiflora Schrenkf., find the optimal seedling raising method, and provide scientific basis for standard seedling raising technique and protecting wild resources of Fritillaria. [Methods] This experiment counted the seedling emergence number and calculated the seedling emergence indicator and the yield. [Results] The results showed that under the condition of film mulching and open ground seedling raising, the changes of Fritillaria seedlings showed an "S" curve, conforming to the Logistic equation, and the rapid growth period appeared 5-13 d after seedling emergence. Compared with the open ground seedling raising, the seedlings of F. przewalskii Maxim. and F. pallidiflora Schrenkf. raised through film mulching emerged 2-4 d earlier, and the yield increased by 56.8% and 21.71%, respectively, and the individual quality was excellent. [Conclusions] The film mulching of F. przewalskii Maxim. and F. pallidiflora Schrenkf. has great significance to protect the wild resources of Fritillaria in Eryin Mountain Area. It is recommended to adopt the film mulching method for Fritillaria seedling raising, to reduce the yield decline due to lodging, and adopt sun-shading net covering after sowing.

TWO NEW STEROIDAL ALKALOIDS,DONGBEININE AND DONGBEIRINE,FROM FRITILLARIA THUNBERGII MIO.VAR.CHEKIANGENSIS

Two new D/E trans-5(?)-cevanine alkaloids,Dongbernine and Dongbeirine,have been isolated from Fritillaria thunbergii Miq.var.chekiangensis together with six known compounds,peimine,peiminine,isoverticine,verticindiol,B—sitosterol daucoterol.

Somatic Embryogenesis and Plant Regeneration from Leaflets of Fritillaria ussuriensis M.

A method for the production of somatic embryos and subsequent plant regeneration for Fritillaria ussuriensis M. is described. Whole leaflet cxplants, derived from plantlets grown in vitro, formed light yellowish embryogenic calli within one month of culture in the dark. Somatic embryogenesis was obtained after a 28 d incubation on MS induction medium supplemented with 2 mg / L 2,4-D 0.5 mg / L BA, 0.5 mg / L KT and 500 mg / L CH followed by transfer to a second N medium containing 0.5 mg / L KT and 100 mg / L CH. Somatic embryos were transferred to MS medium with 0.1mg/ L NAA and placed in the light for regeneration. After two weeks, mature somatic embryo developed into whole plantlet.

中国贝母属(Fritillaria)7种植物的核型研究（英文）

通过对7种国产贝母属(Fritillaria)植物的染色体核型进行观察和研究,报道了小白花贝母(F. albidoflora Duan&Zheng)、川贝母(F. cirrhosa Don)、伊贝母(F. pallidiflora Schrenk ex Fischer&Meyer)、华西贝母(F. sichuanica Chen)、托里贝母(F. tortifolia Duan&Zheng)、新疆贝母(F. walujewii Regel)、裕民贝母(F. yuminensis Duan)等7种植物的染色体数目及核型,其中3种为首次报道。结果显示,7种国产贝母属植物的核型均具有高度不对称性。此外,小白花贝母与已报道的黄花贝母(F. verticillata Willdenow)的核型存在明显差异,提示Flora of China将小白花贝母归并入黄花贝母的分类处理可能并不恰当,二者的关系需进一步研究。