Thanshinone Production And Hairy Root Growth In Salvia Miltionrrhiza Hairy Root Induced By Elicitors

Salvia miltiorrhiza Bunge,which is the dry root and the rhizome of orthodon sage the plant salvia miltiorrhiza,is a kind of Chinese traditional herb medicine. Recently,with the development of salvia miltiorrhiza pharmacological research, salvia miltiorrhiza secondary metabolite research is a new hot spot.The content of effective components is low in the original plant, and the growth cycle is long.Under the traditional cultivation pattern,we also face the quality is irregularity,seriously degraded and so on.How to obtain salvia miltiorrhiza resources which have high content of the effective component and stable quality is a big challenge of salvia miltiorrhiza medicinal materials entering the international market. The hairy roots having own characteristic become the good experimental material to research salvia miltiorrhiza effective component.But Afterward research show that, using hairy root production secondary metabolite output was quite still low and affected the further expansion and industrialization.Under present research about the hairy root indicate that the induction has become an effective method that can enhance secondary metabolite output in the hairy root.This article examine the different type elicitors,including Alternaria carthami,Cladosporium cladosporioides,ABA and PEG, under different concentrations and treatment time,were used to influence the hairy root culture system. After that, the growth percentage of hairy roots and four kinds of tanshinone contents (DihydrotanshinoneⅠ,Cryptotanshinone,TanshinoneⅠand TanshinoneⅡA)detected during experiments. And this provides the research basic to the hairy roots.Hope to find suitable elicitors that can effectively enhance the production tanshinones and optimal induction condition,to make the foundation for the further production and studies.During the culture process of Salvia miltiorrhiza hairy root,we find that the hairy root grows well in 6,7-V culture medium and grows bad in MS (including NH4N03) culture medium,but the effective component content is high.So the research is also to study the effect of different nutrient elements in 6,7-V liquid medium and MS liquid medium on the growth and accumulation of tanshinones in hairy root of Salvia miltiorrhiza,then provides the reference for the quality improvement of medium.1.Macroelement affects the growth and concentration of tanshinones significantly;Reducing the content of MgSO4·7H2O in the MS medium may promote the growth of hairy root and the release of TanshinoneⅡA;However, in the macroelements excessive NH4N03 does not favor to the growth of hairy root, the accumulation and release of tanshinones.So in later experiments,we use MS (excluding NH4N03) liquid medium to culture Salvia miltiorrhiza hairy root.2.The growth of hairy root and the content of tanshinones under different concentrations of two fungal elicitors (Alternaria carthami and Cladosporium cladosporioides). The induction effects of lower concentration of elicitors are prior to those of higher concentration of elicitors. Two different concentrations of fungal elicitor have contributed to the growth of hairy roots,overall,the dry weight of hairy roots had little effect. Two fungal elicitors increase effectively the yield of tanshinones and there are differences among different elicitors'effects. The highest content of DihydrotanshinoneⅠand TanshinoneⅡA in hairy root are achieved by Alternaria carthami elicitor at 50μg/mL, which are 3.16-fold and 1.25-fold of the control.With Cladosporium cladosporioides elicitor at 250μg/mL giving the highest content of Cryptotanshinone and TanshinoneⅡA (5.24-fold and 1.43-fold of the control).So we determine induced concentration of Alternaria carthami and Cladosporium cladosporioides elicitors are 50μg/mL and 250μg/mL respective.3.Two fungal elicitors have the different effects for the growth and the tanshinone synthesis of hairy roots. The results are as follows: the two fungal elicitors can promote the growth of hairy root with time, also selectively promote different tanshinones accumulation in hairy root and in medium, and both reached the maximum at 6 days.The elicitation effects of Cladosporium cladosporioides elicitors induced tanshinones in hairy roots and medium are better than Alternaria carthami elicitor. Two elicitors notably increased the crypotanshinone content.4.Different concentrations of two abiotic elicitors ABA and PEG notably inhibit the growth of hairy root,but selectively increased the tanshinone content.With increasing the concentration,all tanshinone contents show the trend of first increasing and then decreasing.The elicitation effects of lower concentration of elicitors are prior to those of higher concentration of elicitors. Optimal concentrations of ABA and PEG elicitors are 200μmol/L and 2% respective.5.One day after inducing,ABA and PEG elicitors inhibit the growth of hairy root,but after inducing six days,both can promote the growth of hairy root.The two elicitors also promote tanshinones accumulation in hairy root.The accumulation of tanshinones are increased rapidly after the inducting of ABA,all reached the maximum at 6 days.ABA-induced Cryptotanshinone and TanshinoneⅡA contents in each harvest period, compared with the control,are able to significantly improve.PEG-induced DihydrotanshinoneⅠand Cryptotanshinone contents show the trend of first increasing and then decreasing,both reached the maximum at 2 days.Accumulation of TanshinoneⅠincreased volatility,in each harvest period, compared with the control,is able to significantly improve.After one day PEG-induced,the content of TanshinoneⅡA continue to rise,and in each harvest period, compared with the control,are able to significantly improve.