Effects And Regulation Of Mineral Elements On Accumulation Of Salvia Miltiorrhiza Secondary Metabolites

Danshen is one of Chinese traditional medicine, which comes from the root of Salvia miltiorrhiza Bunge and has been widely used as the bulk medicinal material. The curative effects of S.miltiorrhiza on cardio-cerebrovascular diseases and antioxidant are remarkable. Its pharmacological activity components have been basically clear, which contains rosmarinic acid, salvianolic acid, protocatechuic aldehyde, Danshensu and other water-soluble components, as well as liposoluble components-diterpene quinines tanshinones and royleanone.The addition of elicitor is widely known as one of the most effective ways to improve the synthesis of secondary metabolites in medicinal plants. In this paper, different kinds of elicitors were studied through tissue culture using S. miltiorrhiza hairy roots as experiment materials, to screen the effective elicitor which could promote the accumulation of active constituents in S. miltiorrhiza hairy roots. The selected elicitors contain heavy metal nickel, phosphorus necessary for plant growth, beneficial element-selenium and rare earth elements-cerium and neodymium. After ascertainment of the elicitor which could promote the accumulation of active constituents, the induction experiment in which this elicitor acted on S. miltiorrhiza hairy roots in time course was conducted. Through the induction experiment, the expression of relative genes in active constituent synthetic route and the activity of protective enzyme were measured for purpose of discussing the regulatory mechanism elicitor to the synthesis of S. miltiorrhiza active constituent. The main results were as followed: 1.The effect of various kinds of elicitors on accumulation of Salvia miltiorrhiza secondary metabolites, the results: under the induction of 10 μM Ni SO4, the synthesis of tanshinone II A, cryptotanshinone, dihydrotanshinone I and tanshinone I in S. miltiorrhiza hairy roots were all promoted; the synthetic amount of four tanshinones mentioned-above were increased under inductions of 0.1, 0.5, 1 m M Na2 Se O3; Under the Nd(NO3)3 concentration of 100 μM, the synthesis of lipophilic conponents of S. miltiorrhiza was promoted; under the treatment of 50 μM Ce(NO3)3, the synthesis of dihydrotanshinone I and cryptotanshinone were markedly promoted; under the induction of phosphorus concentration which was 1% of that of control group(0.0124 mmol/L), the synthetic amount of rosmarinic acid, salvianolic acid B, Danshensu, caffeic acid, ferulic acid and four tanshinones mentioned-above were all increased. 2. According to induction culture of S. miltiorrhiza hairy roots in time course with the treatment of 50 μM Ce(NO3)3, the results: compared with the control group, Ce(NO3)3 can promote the accumulation of dihydrotanshinone I and cryptotanshinone in S. miltiorrhiza hairy roots. Through the analysis of expressions of key genes in the synthetic route of tanshinone, it could be inferred that the metabolic pathways of dihydrotanshinone I and cryptotanshinone was probably different from tanshinone II A and tanshinone I. Besides, 50 μM Ce3+ could improve the activity of POD while has no obvious effect on SOD. 3.According to induction culture of S. miltiorrhiza hairy roots in time course with the treatment of phosphorus concentration which was 1% of that of control group(0.0124 mmol/L), the results: with low phosphorus in the culture environment, the accumulation of dry matter in S. miltiorrhiza hairy roots was impeded and the growth of S. miltiorrhiza hairy roots was restrained. And we could infer that the accumulation of dry matter in S. miltiorrhiza hairy roots was a quite long process in low-phosphorus condition. During the culture period on day 15-27, with the treatment of phosphorus which was 1% of that of control group, the accumulation of water-soluble components-rosmarinic acid, salvianolic acid B, Danshensu, caffeic acid, ferulic acid were effectively promoted. Meanwhile, the accumulation of liposoluble components-tanshinone II A, cryptotanshinone, dihydrotanshinone I and tanshinone I were promoted. 1% low-phosphorus induction significantly improve the activity of POD. 4.Through the comparison of transcriptomes of S. miltiorrhiza, we draw a conclusion that there were only the sequences of four SPX family genes-Sm SPX1, Sm SPX2, Sm SPX3 and Sm SPX4 in SPX domain. By using the online software-SOPMA, the structural information of four genes above was predicted through secondary structure prediction on the basis of primary sequence.