Establishment Of Tissue Culture System And Extraction Of Steroidal Saponins From Polygonatum Cyrtonema Hua

Polygonatum cyrtonema Hua is subordinate to Liliceae.It is neutral,sweet and is also a kind of tonic.It has the function of improving Kidney,lowering blood fat and sugar,lowering blood pressure,improving immunity,anti-aging,anti-tumor and other effects.It is the medicinal and edible perennial herbaceous plants for the traditional Chinese bulk medicine which has a good development and application value.Steroidal saponins from Polygonatum cyrtonema Hua are mainly yam saponins,diosgenin and so on,with hypoglycemic,anti-tumor,anti-HIV biological activity.Owing to low seed germination rate,long seedling period and other factors in artificial cultivation,the natural way of reproduction by Polygonatum cyrtonema Hua can not meet the market demand.Compared with the traditional cultivation methods,tissue culture is not affected by the season,the number of materials and other factors.It's a better way leading to a good seeding which can be stable in a large scale in the short term.Meanwhile,hairy roots have the advantages of fast growth rate,strong secondary metabolites and stability,and so on.Therefore,steroidal saponins can be produced by hairy root liquid culture technique.In this study,UPLC method was established for the simultaneous determination of yam saponins and diosgenin in wild plants and hairy roots of Polygonatum cyrtonema Hua,which provided a scientific basis for the utilization of Polygonatum cyrtonema Hua.The results are as follows:1.Different concentrations of hormone of seed are treated in low temperature sand.We find the higher germination rate was in the sandy soil at 0? for 100 d and inoculated into MS + 2.0 mg/L 6-BA medium.The germination rate of Polygonatum cyrtonema Hua was the highest,reaching 43.7%.2.Through the L9(34)design of experiment to screen out the optimal medium was MS + 1.0 mg/L 6-BA +0.5 mg/L NAA +0.3 mg/L IAA.The bud multiplication coefficient reached 3.2.The optimal rooting medium was 1/2MS + 1.0 mg/L NAA,and the average rooting rate was 86.92%.The optimum medium for hardening was peat soil and sand(2:1).3.The study used the orthogonal design experiments to filter out the optimal liquid culture conditions of the hairy root was liquid 1/2 MS,the pH was 5.8,shaking speed 150 r/min,dark culture at 26?,and the multiplication rate was 9.4.4.The extraction method of steroidal saponins was optimized by ultrasonic method.The extraction ratio was 1:15 by using 75%ethanol as extraction agent.After ultrasonic treatment for two hours,it was extracted with butyl alcohol.5.UPLC method was established for the simultaneous determination of yam saponins and diosgenin in wild plants and hairy roots of Polygonatum cyrtonema Hua.The contents of yam saponins and diosgenin in wild plant rhizome were 106.27?g/g and 84.95?g/g.The content of yam saponins and diosgenin in hairy roots were 89.13?g/g and 72.24?g/g.