Effects Of Culture Conditions On Content Of Active Compounds In Suspension Cells Of Salvia Miltiorrhiza

In this study, Salvia miltiorrhiza young leaves were used as explants for the induction ofcallus. Through optimization of culture medium, a kind of light-yellow, loose-texture, friableand fast growing callus was obtained and chosen to establish the Salvia miltiorrhizasuspension cells culture system. And then, the effects of several factors, including hormones,inoculation quantity, temperature, carbon sources, basical culture medium and elicitors, ongrowth and production of secondary metabolites in suspension cells were investigated. Theresults are as follows:1Salvia miltiorrhiza leaves were inoculated on solid MS base supplemented with2,4-D,6-BA and NAA or combinations of these phytohormones for callus induction. The resultsshowed that all the three regulators could induce callus in a range of concentration；themedium favored for callus induction was MS+2,4-D0.5mg/L, and for callus growth wasMS+6-BA0.5mg/L+2,4-D0.5mg/L. Lower concentration of2,4-D（0.5~1.0mg/L）issuitable for callus induction, however, higher concentration of2,4-D（≥1.5mg/L）inhibitcallus growth.2Screened callus was transferred to liquid medium and cultured for establishingsuspension cells, and then the effects of several factors on growth and production of depsidesand tanshinone were examined. The results indicated that the optimized medium forsuspension cells growth was MS+2,4-D0.5mg/L, and for accumulation of depsides was6,7-V+2,4-D0.5mg/L; in addition,60g/L inoculation quantity and24℃were better forsuspension cells preparation and growth. For both suspension cells growth and accumulationof depsides, sucrose was proved to be the most suitable carbon source. The cell growth periodwas approximately18-days long, and it could arrive at the peak point of both biomass andsecondary metabolite content by time of16days.3Some kinds of elicitors and combinations were fed on day10during culture. The resultsshowed that H2O2, oxalate, SA, yeast extract and the combinations of yeast extract withdifferent abiotic elicitors restrain callus growth. H2O2treatment can promote depsides andtanshinone production in cell suspension culture.H2O2treatment (10μmol/L) yielded the hightest depsides content (94.85mg/g), which was2.43-fold of the control. Oxalatetreatments promote depsides production and restrain tanshinone peoduction.SA treatment (1μmol/L) yielded the hightest tanshinone content (1.53mg/g). The combinations of yeastextract with different abiotic elicitors didn’t exhibit visibled effects on the secondarymetabolite content.