Study On Building Of Tripterygium Wilfordii Hook. F. Immobilized Cell And Tirpolide Synthesis

Tripterygium wilfordii Hook.f.is the Euonymus Branch dicotyledonous woody vine plants,also known as graceful jessamine herb,yellow TengMu etc,mainly distributed in south of the Yangtze river basin in southwest China and all over.Tripterygium as a kind of traditional Chinese herbal medicine,is promoting blood circulation to remove blood stasis,real detumescence,insecticidal,hemostatic effect and so on.Tripterygium triptolide from the tripterygium plant tissue,high titer,an effective ingredient of the tripterygium.Tripterygium suspension cell as materials,the immobilized cell of Tripterygium wilfordii was constructed,and control the immobilized cell and suspension cells,compared with study the regulation of biological characteristics and the influence of secondary metabolism process.The main contents is as follows:1.The immobilized cell of Tripterygium wilfordii was first constructed by sodium alginate embedding method,and then its growth and tirpolide production were studied.The results showed that the cell growing could be divided into three phases:slow growth,exponential growth,and steady growth.The contents of total sugar,soluble protein,malondialdehyde decreased with culture time increasing.The pH value of cell supernate had a tendency of ascending first and descending in succession,however,the cell vitality exhibited an opposite tendency In addition,during the process of culture,the POD activity appeared a type V shape,the SOD activity displayed a smooth curve,and the PAL present a trend that it first increased sharply and then decreased slowly.The triptolide contents of extra-and intra-cellular showed the same changing trends and both reach the maximum yields on the 15th day of culture.2.Tripterygium immobilized cells and tripterygium endophytic fungi are elicitor control each other,immobilized cells in cell inside and outside and endophytic fungi(NS-13)outward release triptolide content,at the same time,the influence of affect tripterygium the peak production time.The results showed that the elicitor regulation,the immobilized cell intracellular a prime peak quality score for:the influence law of NS-13 inoculum elicitor than NS-13 mycelium elicitor is more advantageous to the immobilized cell intracellular triptolide synthesis,NS-13 inoculum elicitor add the earlier,for intracellular triptolide synthesis promoting the more obvious.Under the control of the immobilized cell extracellular a prime peak production accumulated rule is:NS-13 inoculum elicitor in 6d add more can promote the immobilized cell extracellular a triptolide production,its value is much higher than CK.And plant tissue as elicitor of endophytic fungi(NS-13),controls the triptolide peak production effects:plant cell tissue as elicitor to secondary metabolites,and promote the synthesis of endophytic fungi of the bacteria to release triptolide.3.Studied Tripterygium immobilized cells and suspension cells after the precursor feeding,the different concentration of citric acid,salicylic acid and phenylalanine for its biological characteristics and production process of triptolide influence.The results show that citric acid,salicylic acid and phenylalanine treatments in suspension cell growth have different degrees of promoting function,but immobilized cells the growth have no significant in promoting.Different precursor,different concentration of immobilized cells and suspended cells intracellular triptolide a peak mass fraction influences on difference,citric acid,SA and Phe have the ability of promoting to immobilized cells intracellular triptolide synthesis in the appropriate concentrations,and only in the following three concentrations,citric acid 0.5 mmol/l,SA 0.5 mmol/1 and Phe 0.01 mmol/1 to the suspension cell intracellular triptolide synthesis have promote role.Different precursor,different concentration of immobilized cells and suspended cells extracellular triptolide a peak mass fraction influences on difference,citric acid,SA and Phe in the appropriate concentrations have the ability of promoting to suspension cells to extracellular release of secondary metabolites,all of these can effectively promote the suspension cell extracellular accumulation of triptolide.Compared with suspension cells,three kinds of precursor things only in the appropriate concentration,can promote the immobilized cells to extracellular release triptolide.4.Adding inhibitor in tripterygium immobilized cells and suspension cells,the cell growth and cell inside and outside a prime biosynthesis pathways in the effects of inhibitors.Adding three kinds of inhibitors of different concentration in immobilized cells and suspended cells intracellular have effects of peak mass fraction of triptolide:monoterpenes synthetic metabolism inhibitors(alpha pinene,terpineol and camphor)in the plasmid DXP way,have the ability of promoting to synthesis of triptolide.Alpha pino dilute concentration is smaller,more can promote the immobilized cell intracellular triptolide synthesis,but terpineol and camphor,the higher of concentration,the immobilized cell intracellular triptolide synthesis effect is stronger.In the appropriate concentrations of the inhibitor have the ability of promoting to suspension cells to intracellular triptolide synthesis.Different inhibitors,different concentrations in immobilized cells and suspended cells extracellular a prime peak mass fraction effects:alpha pino dilute and camphor density is small,the immobilized cells have the ability of promoting to suspension cells to extracellular release of triptolide,and terpineol concentration is higher,the immobilized cells to promote to the extracellular release of triptolide.But,alpha pino dilute,terpineol and camphor is the higher concentration,the promoting suspension cells to extracellular release of secondary metabolites triptolide.