诱导子对野罂粟悬浮培养细胞中活性生物碱产生的作用考察

研究了真菌诱导子和金属诱导子对野罂粟培养细胞中生物碱(野罂粟碱和黑龙辛甲醚)产生的影响。在野罂粟悬浮培养细胞中加入真菌和金属诱导子以考察其对细胞生长和野罂粟生物碱产生的作用，并利用HPLC法测定了培养物中野罂粟碱和黑龙辛甲醚的含量。结果显示密环菌诱导子可使黑龙辛甲醚的含量达0．026mg/g，野罂粟碱的含量由0．0159提高到0．0178mg/g；灰葡萄孢菌可使黑龙辛甲醚含量达0．0346mg/g;Zn^2 和Cu^2 可使黑龙辛甲醚的含量由0．0252mg/g分别提高到0．0282和0．0395mg/g.本研究表明，添加适量的真菌和金属诱导子可使野罂粟碱和黑龙辛甲醚的含量有不同程度的提高。     

野罂粟总生物碱对急性气管-支气管炎止咳效应的探索性临床研究

目的 :评价野罂粟总生物碱制剂对急性气管 支气管炎肺气上逆证的止咳效应。方法 :随机、双盲、安慰剂平行对照、叠加试验 (add ondesign)设计。观察急性气管 支气管炎肺气上逆证共 89例 ,其中野罂粟总生物碱组 4 5例 ,安慰剂对照组 4 4例。结果 :试验组和安慰剂组服药后咳嗽明显减轻者分别占 75 .5 6 %和 77.2 7% (P =0 .84 87) ,咳嗽明显减轻者的减轻时间分别为 19.5 5± 18.5 7和 2 5 .2 7±19 .6 0h (P =0 .1993) ;d 3咳嗽消失率分别为33.33%和 36 .36 % (P =0 .76 42 ) ;咳嗽记分值平均等级分别下降 3.5 3和 4 .16 (P =0 .2 786 )。结论 :急性气管 支气管炎中度及重度咳嗽少痰肺气上逆证患者在使用头孢克洛缓释胶囊治疗的基础上 ,野罂粟总生物碱制剂没有明显的止咳、平喘作用。     

野罂粟总生物碱镇痛作用

目的：研究野罂粟总生物碱（total alkaloids of Papaver nudicaul，TAPN）的镇痛作用特点。方法：采用小鼠热板法和大鼠电刺激甩尾法两种致痛模型研究TAPN镇痛作用特点。结果：小鼠热板法和大鼠电刺激甩尾法致病实验结果表明：ipTAPN具有明显的镇痛作用，其作用强度约为吗啡的1／5～1/4。单次给药的镇痛作用达峰时间与吗啡相似，均在20min以内，但镇痛作用持续时间明显长于吗啡，可达4h以上。与吗啡明显不同的是．TAPN的镇痛作用无耐受性。结论：TAPN具有明显的镇痛作用，作用强度弱于吗啡．约为吗啡的1／5-1/4，但镇痛作用维持时间明显长于吗啡，可持续4h以上．且其镇痛作用无耐受性。     

从罂粟细胞悬浮培养获得可待因

本文叙述罂粟(Papaver somniferumL.)细胞悬浮培养合成可待因(Codeine)的研究结果。细胞悬浮培养:罂粟种子用20％Javex(5％次氯酸钙)消毒,盛于Petri碟中湿润的滤纸上,在黑暗中约25°下催芽。分离5～6天龄种苗的胚轴,移植于固体培养基上(含1毫克/拉升2,4-D(1-B_)的0.6％Difco-琼脂]培养。将诱导产生的愈伤组织移植于新鲜的培养基(含1-B_5)另加     

西洋参冠瘿组织培养条件对其人参皂苷Rb1含量的影响

考察了不同理化因子对西洋参冠瘿组织生长及其主要活性成分人参皂苷（ginsenosides)Rb1的影响，采用HPLC法测定了Rb1的含量。结果表明：在考察的6种培养基中，White培养基最适合人参皂苷Rb1的累积（0．755％）；培养25d时Rb1累积含量最高（0．732％）；接种量为6g FW/flask最有利于Rb1合成（0．618％）；培养基pH值在5．6时Rb1累积量最高（0．692％）；肌醇的浓度为0．05g/L时，能明显促进Rb1合成（1．881％）。     

B38-20包埋啤酒酵母的生理学和形态学研究

在精细合成培养基上连续20多批厌氧培养啤酒酵母CBS8066，研究包埋对其细胞形态学和生理学方面的影响。在此过程中，酒精产量由0．43g／g增至0．46g／g，生物质和甘油产量分别降低了58％和23％。第一批包埋细胞生长速率为0.13h^-1，经连续培养20批后，该速率逐步降至0.01h^-1。这些酵母细胞中总RNA含量由90．3mg／g降至55mg／g，总蛋白含量由460mg／g降至350mg／g，分别降低了39％和24％。     

不同诱导子对黄芩不定根生长及黄芩苷累积的影响

目的研究蔗糖、茉莉酸甲酯和水杨酸3种诱导子对黄芩不定根生长及黄芩苷累积的影响。方法向黄芩不定根悬浮培养液中添加不同种类及浓度的诱导子,观察不定根生长情况并测量不定根的生物量及黄芩苷的含量。结果培养基中蔗糖浓度为50 g·L^-1时可使不定根生物量达到对照组（30 g·L^-1）的1.62倍,黄芩苷的含量达到对照组的1.71倍为20.81 mg·g^-1。加入44.860 mg·L^-1茉莉酸甲酯可使黄芩苷的含量为对照组的3.22倍,达到最高39.14 mg·g^-1,但却大大影响黄芩不定根的产量。低浓度水杨酸对不定根生长及黄芩苷累积均有利。当其浓度为4 mg·L^-1时,不定根的生物量及黄芩苷含量均达到最大。结论在黄芩不定根悬浮培养中加入适量的蔗糖、茉莉酸甲酯及水杨酸,对不定根的生长及黄芩苷的合成有一定的促进作用。     

止泻木细胞悬浮培养生产生物碱研究(第二报) 加入前体并在搅拌下培养的影响

作者等考虑到在培养中加入前体胆固醇50mg/L,8天后的生物碱产量能达到143mg/L。放大到6L容器内,并掌握好培养基的成分、接种、通气及搅拌等要点,和前体物的加入步骤,可使生物碱生产水平在8天后达到0.88g/100g干燥细胞,相当于每天110mg/100g。和植物自然生长相比较,按4年长成的植物,每100g可含1g生物碱的含量计,等于每天0.68mg/100g的生长率,高     

天南星炮制过程中总生物碱的动态变化

目的：研究炮制过程中天南星总生物碱含量的动态变化,建立天南星总生物碱的含量测定方法。方法采用酸性染料比色法测定天南星总生物碱的含量。结果方法学考察结果,线性范围2．2～11μg? mL －1,r＝0．9995,平均加样回收率为99％,RSD％＝2．8％（n＝6）;天南星生品、中间品、制品总生物碱含量均值分别为1．286mg? g－1、0．806mg? g－1、0．727mg? g－1。结论总生物碱在炮制过程中含量逐步降低;该含量测定方法稳定可靠,重复性好。     

头孢霉AL031菌株液体培养工艺的研究

研究产生异香豆素类化合物的头孢霉（Cephalosporium sp.)AL031菌株的培养基和摇瓶培养条件。并通过正交试验进行优化。结果表明，其适宜的培养基组成（％）：马铃薯（煮汁）20，葡萄糖和蔗糖（1：2）2．0，KH2PO40．1，MgSO4.7H2O 0.05;Vitamin B10.01,摇瓶培养条件为：培养基的起始pH为7，500ml三角瓶装量200ml，接种量10％，24℃振汇培养4d，细胞生物量为10．8mg/ml发酵液。     

苯丙氨酸、蔗糖和甘露醇对杂种红豆杉细胞的生长及形成紫杉醇、巴卡亭III和10-去乙酰基巴卡亭III的影响

研究了苯丙氨酸、蔗糖和甘露醇对杂种红豆杉悬浮细胞的生长及形成紫杉醇、巴卡亭II和10-去乙酰基巴卡亭II的影响。结果表明,培养基中添加1.0mmol·L^-1或2.0mmol·L^-1苯丙氨酸和在培养28d时同时补加73.0mmol·L^-1蔗糖和137.3mmol·L^-1甘露醇能显著地促进细胞的生长和这3种紫杉烷的形成。同对照相比,有苯丙氨酸又补加蔗糖和甘露醇的细胞生物量增加了0.6～0.8倍,紫杉醇的产量增加了9～10倍,巴卡亭II的产量增加了2.5～3.0倍,10-去乙酰基巴卡亭II的产量增加了7倍。在培养28d时补加73.0mmol·L^-1蔗糖能显著地促进细胞的生长,但对细胞中这3种紫杉烷的含量没有显著的影响。     

Growth and production of camptothecin by cell suspension cultures of Nothapodytes foetida

Callus cultures were initiated from stem parts of Nothapodytes foetida on Murashige and Skoog's medium supplemented with different growth regulators. Suspension cultures were established and the cell biomass was higher in the presence of NAA in comparison with 2,4-D. Culture medium supplemented with NAA (10.74 microM) and BA (2.22 microM) attained 31.3 g/l DW during 20 days of cultivation in shake flasks. In the presence of NAA, maximum concentrations of camptothecin (0.035 mg/ml) and 9-methoxycamptothecin (0.026 mg/ml) were found in the medium. Alkaloid production was reduced in presence of 2,4-D in the culture medium. Cells contained trace amount of alkaloids. Alkaloids were detected and identified by means of TLC and HPLC.     

Production of eurycomanone from cell suspension culture of Eurycoma longifolia

Context: Eurycomanone is found in the Eurycoma longifolia Jack (Simaroubaceae) tree, exhibits significant antimalarial activity, improves spermatogenesis, suppresses expression of lung cancer cell tumour markers and regulates signalling pathways involved in proliferation, cell death and inflammation.

Objectives: Establishment of cell suspension culture of E. longifolia to determine the eurycomanone accumulation during cultures.

Materials and methods: Callus of E. longifolia was cultured in MS medium supplemented with 0.8% agar, 30/L sucrose, 1.25?mg/L NAA and 1?mg/L KIN for biomass production. Cell suspension culture was established by transferring friable calli to the same medium without agar. Eurycomanone content during cell culture was determined by HPLC with a C18 column, flow rate of 0.8?mL/min, run time of 17.5?min, detector wavelength of 254?nm. The stationary phase was silica gel and the mobile phase was acetonitric:H₂O. Roots of 5 year-old trees were used as the control.

Results: The cells from 3?g of inoculum increased in biomass with a maximum value of 16?g fresh weight (0.7?g dry weight) at 14th day of culture. The cell growth then decreased from day 14 to day 20. Eurycomanone was produced during culture from the beginning to 20th day, its highest content (1.7?mg/g dry weight) also obtained at 14th day (the control is 2.1?mg/g dry weight).

Discussion and conclusions: Cell suspension culture of E. longifolia is a suitable procedure to produce eurycomanone. The yield of eurycomanone biosynthesis in 14 days-old cells are relatively high, approximately 0.8 times the control.     

Withanolide production by root cultures of Withania somnifera transformed with Agrobacterium rhizogenes

Transformed root cultures of Withania somnifera Dunal (Solanaceae) were obtained by infecting shoots cultured in vitro with Agrobacterium rhizogenes LBA 9402. They grew axenically in the absence of exogenous plant growth regulators in Murashige and Skoog's medium containing 3% (w/v) sucrose. The root cultures synthesized several withanolides of which withanolide D was isolated and identified. Transformed root (clone HR (1)) showed a growth index of 20.07 and a withanolide D yield of 0.30 mg g(-1) DW. The productivity of withanolide D in transformed roots (0.181 mg 1(-1) d(-1)) was higher than in untransformed root cultures (0.026 mg 1(-1) d(-1)).     

Cultures ofGinkgo biloba,effect of nutritional and hormonal factors on the growth of cultured cells derived fromGinkgo biloba

Calli and suspension cultures were obtained following inoculation of the explant from leaves ofGinkgo biloba L. on the supplemented MS basal medium. The obtained calli and suspension cultured cells were able to produce detectable amounts of ginkgolides which are known as natural specific PAF antagonists. The production of ginkgolides in the calli and suspension cultured cells were identified using GC/MS, GC and HPLC with authentic compounds. Since the production of ginkglides A and B in the calli and suspension cultured cells had been confirmed, effects of types and concentration of plant growth regulators, media and illumination on the induction and growth of the callus were studied. The concentrations of growth regulators for optimal callus induction were 1.0 to 2.0 mg/L for NAA and 0.1 mg/L for kinetin. The growth of the callus seemed to be more stimulated with the combination of NAA and kinetin than NAA and BA with illumination at all concentration ranges of 1.0 to 4.0 mg/L for NAA and 0.1 to 1.0 mg/L for kinetin or BA studied. Among 8 different media used, the induction rate of callus on Anderson, Eriksson, and Schenk and Hildebrant at 4 weeks after the innoculation was almost the same as that of MS. However, callus was rarely induced on Heller or White medium. Suspension cultures were easily initiated with 3 g of callus (fresh weight) derived from ginkgo leaves on supplemented MS medium. A typical growth curve of suspension cultured cells could be obtained by measuring the fresh weight of the suspension cultured cells at every 3 days. To improve the growth of suspension cultured cells of ginkgo, effects of concentrations of NAA, sucrose, phosphate ions and molar ratio of NH₄ ⁺ to NO₃ ^? ions in the culture medium were studied. The maximum growth of the cells was achieved when the culture medium contained 1.0 mg/L of NAA, 30 g/L sucrose, 1.75 mM phosphate ions and 1∶5 molar ratio of NH₄ ⁺ to NO₃ ^? ions.     

Improved production of caffeic acid derivatives in suspension cultures ofEchinacea purpurea by medium replenishment strategy

The aim of this study was to produce caffeic acid derivatives from adventitious root cultures of Echinacea purpurea, which are of high pharmaceutical value. The effects of both media optimization and replenishment strategies were adopted to achieve improved production of E. purpurea adventitious roots and caffeic acid derivatives. Of the different media strengths (0.25 MS, 0.5 MS, 0.75 MS and 1 MS) tested for the culturing of adventitious roots in 5 L bioreactors, 0.5 MS medium was found to be most suitable for the growth of adventitious roots. The optima accumulation of biomass (73.6 g L(-1) FW and 10.03 g L(-1) DW), phenolics (61.14 mg g(-1) DW) and flavonoids (38.30 mg g(-1) DW) were achieved in this medium. Furthermore, fed batch cultivations (media replenishment with 0.25 MS, 0.5 MS, 0.75 MS and 1 MS at the end of 2nd and 3rd weeks) to further enhance the production of adventitious root biomass and metabolites were also attempted. High adventitious root biomasses (83.1 g L(-1) FW and 15.30 g L(-1) DW) were achieved with feeding of the 0.5 MS medium at the end of 2nd week. This led to slight decreases in the total production of phenolics and flavonoids; however, this feeding was responsible for increases in the accumulation of caftaric acid (5.76 mg g(-1) DW) and cichoric acid (26.12 mg g(-1) DW).     

Comparative study of the effects of various culture conditions on cell growth and Gagaminine synthesis in suspension culture of Cynanchum wilfordii (MAXIM.) HEMSLEY

Gagaminine, a steroidal alkaloid isolated from the roots of Cynanchum wilfordii, exhibited potent inhibitory effects on aldehyde oxidase activity and lipid peroxidation. To determine whether it would be possible to mass produce this active component, which would be useful for animal tests, we tried to synthesize it using in vitro cell culture methods with various growth conditions. In a previous study it was found that calli were easily induced from the stem of this medicinal plant and cultivated effectively on MS medium containing 2,4-dichlorophenoxyacetic acid (2,4-D) 2 mg/l. In this work we attempted to determine the effects of various culture conditions on cell growth and gagaminine synthesis in suspension culture. Gagaminine production was increased markedly when cell growth proceeded to the death phase. Cell growth was more effective with 5% (v/v) sucrose, in the light (at 38 microE/m(2) x s), on medium containing 2,4-D 2 mg/l, with 2.5 g/10 ml medium as the initial cell concentration. The concentration of gagaminine was optimal with 3% sucrose, in darkness on medium 2,4-D 1 mg/l, with 2.5 g/10 ml medium as an initial cell concentration. However, the highest growth rate was 0.18 d(-1), when the gagaminine concentration was seven- and three-fold (at 140 mu/ml) that of the plant stem and 10 ml of medium respectively, on the 50 ml of medium in suspension culture.     

Production of triterpenoids in liquid-cultivated hairy roots of Galphimia glauca

The production of three triterpenoids from Galphimia glauca hairy root cultures, the sedative principle galphimine E (2), the recently described glaucacetalin A (3), and maslinic acid (6), was quantified by HPLC in the biomass and the culture medium. Batch cultures of the hairy root line VYT, obtained through infecting cotyledons with Agrobacterium rhizogenes ATCC 15 834, were grown for 41 days in shake flasks containing B5 medium without phytohormones. A maximum biomass of 11 g/L DW was obtained on day 33, while the doubling time was 6 days. Throughout the growth cycle fresh and dry weights as well as triterpene production were registered. Glaucacetalin A (3), excreted into the culture media, reached a maximum amount of 2.14 mg/L after 21 days while galphimine E (2) and maslinic acid (6) were recovered from the root biomasses reaching maximum concentrations of 0.11 and 0.43 mg/g, respectively, on day 39.     

Growth and diosgenin production by Dioscorea deltoidea cells in batch and continuous cultures

This paper reports the kinetics of growth and diosgenin production in batch cultures, and the application of the continuous culture (chemostat) technique to DIOSCOREA DELTOIDEA cells. In batch cultures, biomass production was dependent on the concentration (up to 60 g/liter) of the carbon source (sucrose); the cellular yield value obtained was 0.4 g cell dry wt/g sucrose utilized. Diosgenin was synthesized only after growth had ceased; its synthesis proceeded for about 18 days and a concentration of 1.8% (of cell dry wt.) was obtained. D. DELTOIDEA cells were grown at steady state conditions, at a constant growth rate in a chemostat. Only small amounts of diosgenin were produced by growing cells in the chemostat.