?-ecdysterone Accumulation And Metabolic Regulation In Ajuga Lobata D.Don Suspension Cultural

Researches of Ajuga were mainly focused on its extraction,separation,component identification,applications of active ingredient and applications of pharmaceutical.However,there are few reports on metabolic regulation about ecdysterone of A.lobata D.Don.Cell biology techniques can be used to produce secondary metabolites,which does not only meet the needs of phytoecdysterone,but also protect natural resources and the environment.These techniques are not limited by the field conditions,short producing cycle,have high efficiency and do not harm to the environment and the like.Based on these,this paper studied the kinetics of the process during Ajuga cell suspension cultural and the effect of precursors,elicitors and inhibitors on A.lobata D.Don cell lines growth and ecdysterone accumulation.Conclusions are as follows:1.The growth cycle was 19d.Its curve was "S" shape and matched with logistic equation.In the growth cycle,1-3d was lag phase,3-11d was exponential phase,11-17 was stationary phase and 17-19d was decline phase.The pH value of the liquid culture medium was found to vary from 4.5 to 6 during suspension culture.At the beginning,the pH value was declined,and then risen up gradually.The relation of cells growth and nutrient consumption in A.lobata D.Don cells suspension culture was obviously correlated.Nutrient(carbor,phosphorus,nitrogen)change curve matched with cells growth curve,which also experienced four periods.The electrical conductivity of the culture medium gradually descended,with dry weight and ecdysterone accumulation arising.Electrical conductivity contributed to determining the best harvest time.Kinetics models of A.lobata D.Don cells suspension culture were established,which focused on cells growth.sucrose consumption and ecdysterone synthesis.2.The cells' ability of ecdysterone production will be weakened by subculturing.The direct precursors mevalonic MVA and phenylalanine L-PLA had significantly effect on cells growth.Low concentration MVA can dramatically improve the ability of cells to synthesize ecdysterone.compared to the control.The content of ecdysterone in the treated group increased by 440%and did little harm to cells.L-PLA had no effect on ecdysterone synthesizing,but it can dramatically improve cells viability.which was 2.4 times higher than that of control.A-pinene and terpineol as a metabolic inhibitor could significantly improve the ability of synthetic of ecdysterone in cells,but the terpineol was toxic to the growth of cells.Its cells biomass decreased by 30%compared to the control.Eventually,it resulted in inhibition of ecdysterone production.As an elicitor,NO can not benefit to producing ecdysterone efficiently.On the contrary,it would inhibit synthetic of ecdysterone in a short time after treatment.However,this inhibition will gradually weaken over time.3.The optimal concentration of a-pinene was 50?l.L-1,MVA was 10mg.L-1,NO donor SNP was 80 ?mol.L-1,L-PLA optimal concentration was 0.2mmol.L-1.When the ?-pinene,MVA,NO donor SNP were combined,seven kinds of combinations can stimulate A.lobata D.Don cells to synthesis more ecdysterone.The stimulating efficiencies sorted by the concentration of ecdysterone in treated cells were as follows:MVA + SNP + a-pinene>MVA+ SNP>MVA>MVA + a-pinene = SNP + a-pinene>a-pinene>SNP>CK.A-pinene,MVA and SNP could affect the synthesis process of ecdysterone in A.lobata D.Don cells synergistically.