真菌诱导子对新疆紫草悬浮培养细胞的生长和紫草素合成的影响

４种真菌诱导子诱导新疆紫草悬浮培养细胞对细胞紫草素的合成均有促进作用,其中以黑曲霉诱导子效果最高,而且促进细胞紫草素的外排,约３０％的紫草素存在于培养液中;诱导促使细胞苯丙氨酸解氨酶活性显著提高,培养液中紫草素的含量变化与苯丙氨酸解氨酶活性变化呈正相关性。     

真菌诱导子对悬浮培养西洋参细胞的生理效应

报道了不同真菌诱导子对悬浮培养的西洋参（Ｐａｎａｘｑｕｉｎｑｕｅｆｏｌｉｕｍ）细胞生长、皂甙和多糖合成,以及细胞内和培养液中过氧化物酶活性的生理效应。悬浮培养的西洋参细胞经刺盘孢菌（Ｃｏｌｌｅｔｏｔｒｉｃｈｕｍｎｉｃｏｌｔｉａｎａｅ）丝体诱导子处理后,总皂甙产率可由对照的２９６ｍｇ／Ｌ增加到６７９ｍｇ／Ｌ（约占细胞干重的（１６．３％）,比对照提高约１．３倍,而且总皂甙的８５％排放在培养液中;经黑曲霉（Ａｓｐｅｒｇｉｌｌｕｓｎｉｇｒａｎ）诱导子处理后,细胞多糖含量可达到１１．７９％（细胞干重）,比对照增加１倍多。初步纯化的刺盘孢菌丝体诱导子和尖孢镰刀菌（Ｆｕｓｕｒｉｕｍｏｘｙｓｐｏｒｕｍ）滤液诱导子在诱导处理前期能明显促进西洋参细胞生长,同时细胞内及培养液中过氧化物酶活性显著增加;随时间延长,细胞生长和酶活性逐步受到抑制。     

新疆紫草细胞悬浮培养过程研究

新疆紫草细胞生长和紫草素合成之间属非生长偶联型,所以采用二步培养法研究悬浮培养过程。新疆紫草细胞悬浮培养的生长周期约为21 d,紫草素合成周期约为16 d。新疆紫草细胞生长阶段培养液的电导率与生物量呈线性负相关,随着生物量的增加,培养液的电导率降低。因此,可以通过测量电导率来预测培养体系中生物量的变化情况。细胞生长过程中硝酸盐、铵盐和可溶性糖的消耗与生物量的变化具有很好的线性相关性,基于硝酸盐、铵盐和可溶性糖的细胞收率系数分别为8.64、104.3和0.68 g/g。     

内生真菌和诱导子对长春花悬浮细胞及生物碱合成的影响

目的:探讨内生真茵和诱导子对长春花悬浮细胞生长及生物碱合成的影响.方法:接种内生真茵与悬浮细胞的共同培养,添加诱导子到细胞培养液中对长春花悬浮细胞进行诱导处理,检测实验处理后长春花悬浮细胞各项生化指标.结果:培养液pH值上升,悬浮细胞MDA舍量增加,细胞抗氧化酶(POD、CAT)和生物碱合成的关键性酶(PAL、TDC)活性升高.生物碱产量得到提高,诱导组悬浮细胞生物碱产量达到770.36μg/gFW,共培养组生物碱产量为693.76 μg/gFW,分别比对照组提高了48%和32%.结论:真菌及其诱导子能改变长春花悬浮培养细胞的态势,导致细胞代谢结构的改变,使生物碱的产量提高.     

pH值、激素对新疆紫草悬浮培养细胞生长及紫草宁衍生物合成的影响

报道了不同ｐＨ值、激素对新疆紫草悬浮培养细胞生长及紫草宁衍生物合成的影响。结果表明,新疆紫草细胞具有自我调节其培养液ｐＨ值的功能。适合于细胞生长及紫草宁衍生物形成的ｐＨ值为５．６±０．４０。ＢＡＰ、２,４－Ｄ、ＮＡＡ或ＩＢＡ对细胞生长无显著的促进作用,且都会抑制紫草宁衍生物的形成。在生长培养基中添加１．０ｍｇ／ｌ　ＩＡＡ和０．５ｍｇ／ｌＫＴ可促进细胞生长,而在生产培养基中附加０．１ｍｇ／ｌＫＴ和０．７５─１．０ｍｇ／ｌＩＡＡ则有利于紫草宁衍生物含量及产量的提高。     

新疆紫草细胞生产阶段培养中细胞生长及产物合成的研究I：悬浮培养

新疆紫草细胞生产阶段培养时生物产量随接种量的增加而增大,但生物量的倍增数并不增大。色素产量只在一定范围内与接种量成正相关关系,超过一定范围反而下降。当选定适当接种量时,加大培养基的浓度可提高色素产率。扩大培养规模,色素产量下降。     

新疆紫草细胞生产阶段培养中细胞生长及产物合成的研究Ⅰ:悬浮培养

新疆紫草细胞生产阶段培养时生物产量随接种量的增加而增大,但生物量的倍增数并不增大.色素产量只在一定范围内与接种量成正相关关系,超过一定范围反而下降.当选定适当接种量时,加大培养基的浓度可提高色素产率.扩大培养规模,色素产量下降.     

真菌诱导物在滇紫草细胞培养中对紫草色素形成的影响

在滇紫草细胞悬浮培养中,真菌诱导物可抑制细胞生长,促进紫草色素的合成。将培养６ｄ的曲霉菌丝体的粗提物以６００μｇ碳水化合物／５０ｍｌ培养液的浓度加入到处于指数生长初期的滇紫草细胞悬浮培养物中,诱导物促进紫草色素合成的作用最大,紫草色素含量为对照的两倍。经高压锅处理２０ｍｉｎ到２ｈ不影响诱导物的活性。真菌诱导物还影响了紫草色素各衍生物的相对含量。     

Effects of Rare Earth Elements on the Growth of Arnebia euchroma Cells and the Biosynthesis of Shikonin

Nd³⁺, La³⁺ and Ce³⁺ at proper concentrations had positive effects on the cell growth of Arnebia euchroma and production of shikonin derivatives. A mixture of rare earth elements (MRE, La₂O₃:CeO₂:Pr₆O₁₁: Sm₂O₃ = 255:175:3:1, mol/mol) behaved the most remarkable effects. Two-stage culture was used for the cell proliferation and the biosynthesis of shikonin derivatives. After 20 days culture, 0.05 mM MRE gave the highest cell biomass (24.8 g dry weight l⁻¹), which was 98.0% higher than that without rare earth elements. Similarly, when 0.05 mM MRE was added to the biosynthesis medium, the highest content (8.9% dry weight) and production (571.1 mg l⁻¹) of shikonin derivatives were obtained, which were 89.4% and 165.3% higher than those without rare earth elements, respectively. The increase of the cell biomass and shikonin derivatives may due to increasing the activities of peroxidase and phenylalanine ammonia lyase caused by the addition of the rare earth elements.     

米根霉诱导因子对紫草细胞培养中紫草宁色素分泌的影响

在紫草细胞培养中,加入采根霉粗提物可显著提高紫草宁色素产量,并可加快胞内色素分泌到培养液中的速率和数量。在细胞培养的第6天加入米根霉诱导因子时,其促进紫草宁色素分泌的作用最大,培养液中紫草宁色素含量是对照的2．24倍。此外,同时加入正十六烷和米根霉诱导因子对紫草宁色素的分泌具有协同作用。     

Cell Growth and Shikonin Production of Arnebia euchroma in a Periodically Submerged Airlift Bioreactor

Arnebia euchroma was grown in a 2-l periodically submerged, airlift bioreactor (PSAB) in which the non-submerged (immobilization culture) and submerged (suspension culture) operations were controlled automatically. PSAB had advantages in improving cell growth, shikonin content, shikonin production and cell aggregation compared with suspension culture. Under the optimal submerged/non-submerged period of 10 min/15 h, the shikonin content (4.6%, w/w) and, cell dry mass (16.8 g/l) were 229 and 26% higher than those in suspension culture. Revisions requested 31 October 2005 and 6 December 2005; Revisions received 2 December 2005 and 13 January 2006     

真菌诱导子对茶条槭细胞没食子酸积累的影响

茶条槭（Acer ginnala Maxim.）叶片中含有没食子酸,但含量较低。真菌诱导子可以增加植物中一些次生代谢产物的含量,但其机理尚不十分清楚。本研究在茶条槭细胞悬浮培养的对数期加入内生真菌（Phomopsis sp.）诱导子,茶条槭细胞中没食子酸含量在24 h后开始增加,48 h时没食子酸含量达到峰值,最高含量为12.2 mg·g^-1 DW,是对照的1.58倍。茶条槭细胞对内生真菌诱导子的防御反应不同于对病原和非生物胁迫。真菌诱导子不提高培养液中pH值,也不明显增加胞内Ca²⁺浓度,但增大细胞膜通透性。培养液电导率差异显著,细胞核发生分裂,说明真菌诱导子可能促进茶条槭细胞核内有丝分裂,促使茶条槭细胞对培养液中的无机盐离子的吸收,以满足细胞生长的需要。PAL酶活性升高,在48 h时为对照的1.75倍,说明PAL酶可能参与了真菌诱导没食子酸的合成。     

Effects of Physical and Chemical Factors on Callus Growth and Shikonin Derivative Formation in the Callus Cultures of Arnebia euchroma

The effects of some physical and chemical factors on callus growth and shikonin derivative formation in the callus cultures of Arnebla euchroma were discussed. According to experiments, the optimum temperature for callus growth and shikonin derivative formation was 25℃, and the favorable initial pH of media was in the range of 5.3–5.8. Authors also found that both callus growth and shikonin derivative formation were strongly inhibited by white light. Callus growth was promoted when 0.2 mg/L IAA and 0.5 mg/L KT were added to the media, but IAA and KT did not promote shikonin derivative formation. Furthermore, the content and yield of shikonin derivatives in cultures decreased in company with the increase of IAA and KT concentration in the media.     

真菌诱导子对悬浮培养南方红豆杉细胞态势及紫杉醇合成的影响

在南方红豆杉细胞悬浮培养过程中,研究了在指数生长期的末期加入真菌诱导子(尖孢镰刀菌的胞壁组分粗提物)对细胞态势及紫杉醇合成的影响。结果表明,真菌诱导子能在短期内激发细胞的防御性应答反应而产生氧迸发,细胞的氧化还原态势发生了明显的变化,培养基发生碱化现象,表征酶含量的蛋白质含量明显提高,SOD、POD、CAT的活力出现波动性变化,并具有一定的时序性。同时,南方红豆杉悬浮培养体系中产次生代谢途径中重要的酶PAL的活力得到提高,紫杉醇的合成被加强,产量得到了显著提高,达到了对照组的5倍左右。     

硬紫草细胞悬浮培养和紫草宁及其衍生物的形成

硬紫草细胞悬浮培养形成紫草宁及其衍生物时．细胞生长曲线呈扁平的s形。细胞停止生长后,紫草宁及其衍生物大量形成,二者的动态变化呈负相关。测定丁此过程中培养液的无机元素和可溶性糖含量、溶氧、pH值以及细胞形态的变化情况,可作为硬紫草细胞大规模培养的参考。     

内生真菌培养液对东北红豆杉细胞生长及紫杉醇合成的影响

产紫杉醇的内生真菌（Fusarium mairei）先培养在B5液体培养基中,然后制备成内生真菌培养液。在东北红豆杉（Taxus cuspidata）细胞悬浮培养的不同阶段（5、10和15天）,用不同剂量的内生真菌培养液（2、4和6mL）分别进行处理。结果表明,在用4mL内生真菌培养液处理的植物细胞中可获得最高的紫杉醇产量（5.88mg·L^-1）与释放率（67%）,分别是对照的1.9倍与5.6倍。添加时间方面,在植物细胞培养周期的第5天添加4mL内生真菌培养液,可获得最佳效果,紫杉醇产量与释放率分别为6.1mg·L^-1与75%,分别是对照的2倍与6.8倍。与其它诱导子相比,4mL内生真菌培养液不仅可提高紫杉醇的释放率,而且不会引起东北红豆杉细胞膜的明显伤害,说明内生真菌发酵液激活了紫杉醇主动运输过程中的相关酶类。     

Physico-chemical factors influencing the shikonin derivatives production in cell suspension cultures of Arnebia euchroma (Royle) Johnston, a medicinally important plant species

Cell suspension cultures of Arnebia euchroma were raised from in vitro leaf-derived friable callus on liquid MS [Murashige and Skoog] medium supplemented with BAP (6-benzylaminopurine) (10.0 μM) and IBA (indole-3-butyric acid) (5.0 μM). A two-stage culture system was employed using growth and production medium for cell biomass and shikonin derivatives, respectively. Factors such as light, temperature, sucrose and pH (hydrogen ion concentration) were studied to observe their effect on the shikonin derivative production. Light conditions completely inhibited shikonin derivative production. Out of different temperature regimes tested, the highest yield (586.17 μg/g FW) was found at 25°C. Maximum production (656.14 μg/g FW) was observed in 6% sucrose. An alkaline pH (7.25-9.50) favoured shikonin derivative production. The results showed that physical and chemical factors greatly influence the production of shikonin derivatives in cell suspension cultures of A. euchroma. Therefore, by employing optimum culture conditions, it is possible to enhance the production of secondary compounds from the cells. The factors optimized for in vitro production of shikonin derivatives during the present study can successfully be employed for their large-scale production in bioreactors.