Fed-batch Culture And Anti-bacterial Property Of Adventitious Roots In Oplopanax Elatus Study

Oplopanaxs elatus is an endangered plant species with the high application values.However,the development and production of O.elatus have been restricted by the shortage of plant resources.The adventitious roots(ARs)of O.elatus can be rapidly produced using plant cell culture technology.Thus,the AR culture is an alternative method to obtain the plant material for this species.Based on the batch culture of O.elatus ARs,this paper cultured ARs in a fed-batch culture system and several factors of initial culture medium affecting AR biomass and bioactive compound accumulation were investigated.Furthermore,effect of extracts from ARs on anti-bacterial activity was identified and the anti-bacterial mechanism was also clarified.The final object of this study is to provide the theoretical basis for O.elatus ARs applying in the production of antibacterial drugs and natural preservatives.In the experiments of fed-batch culture,different volumes(2 L and 3 L)and strengths(1/2MS and MS)of the initial culture medium were designed to investigate their effects on AR biomass and bioactive compound accumulation.Results showed that the maximum AR biomass was found at the group of 3 L of 1/2MS medium,the AR fresh and dry weights reached 409.3 and 42.3 g,respectively.In this group,the contents of bioactive compounds including total polysaccharides,total saponins,total phenolics,and total flavonoids as well as flavonoid monomers of quercetin and kaempferol and their productivities were significantly enhanced;the yields of total polysaccharides,total saponins,total phenolics,and total flavonoids were 5.2-fold,2.4-fold,1.8-fold,and 2.4-fold more than the control group(batch culture);and the 2095.3 ?g-L-3 of quercetin and 761.2 ?g·L-1 of kaempferol were produced.Furthermore,the effects of sucrose concentrations(20?30?40 and 50 g·L-1)in the initial culture medium were investigated.The initial culture medium(3 L of 1/2MS)supplemented with 40 g·L-1 or 50 g·L-1 sucrose was favorable for AR biomass and bioactive compound accumulation.The maximum contents and productivities of total polysaccharides,total saponins,and total phenolics were determined at 40 g·L-1 or 50 g·L-1 sucrose.However,flavonoid accumulation at 40 g·L-1 sucrose was obviously higher than that at 50 g·L-1 sucrose,thereby suggesting that the initial culture medium can be added 40 G·-1sucrose during the fed-batch culture of O.elatus ARs in bioreactors.In anti-bacterial experiments,O.elatus ARs were extracted(OAE)and the effect of OAE on anti-bacterial activity was investi sated.Result showed that OAE had anti-bacterial activities against Escherichia coli,Staphylococcus aureus,Pseudomonas aeruginosa,and Bacillus subtilis,the higher inhibitory effects were exerted in E.coli and P.aeruginosa,in which the inhibition zone was greater than 14 mm.To clarify the anti-bacterial mechanism,the E.coli was used and 8 mg·L-1 of the minimum inhibitory concentration was confirmed.Furthermore,the changes of cell permeability and oxidative respiratory metabolism were investigated.The extracellular electric conductivity,protein concentration,leaked nucleic acids(oD260 velue),and alkaline phosphatase activity were significantly increased by the OAE treatment,indicating OAE altered cell permeability of E.coli.Moreover,the activity of glucose-6-phosphatase in the pentose phosphate pathway was distinctly decreased by the OAE treatment,demonstrating that the respiration rate of E.coli was inhibited.In conclusion,OAE inhibits E.coli growth through involving in regulating the cell permeability and inhibiting oxidative respiratory metabolism.