Carthamus Tinctorius Cell Suspension Culture And Research Of Bioreactor Scale Up

Plant Carthamus tinctorius cell suspension culture for natural secondary metabolites is a promising technology.However,the large-scale cell culture and efficacious product fermentation rate are under restrictions due to plant cell low rate of growth and long period culture and intense sensitivity to environment factors during cultivation.In this study,we establish the stable suspension culture system of Carthamus tinctorius cell callus.Through the system experimental design of medium composition and culture conditions,we investigate the effect of some factors on cell growth including fluid volume,inoculation quantity,pH,shear force,amino acids,carbon source,iron salt and inositol,and we find the key factors affecting cell growth including initial inoculation amount,pH,shear force,carbon source.Finally,the optimization of culture conditions show as follows:0-50%fluid volume,20%inoculation quantity,5.5-6.5pH,30%sucrose concentration,0.050-0.1mmol/L iron salt concentration,75 ± 15mg/L inositol concentration.We find that shear force is an important factor to influence cell physiological status during the process of cell scale-up culture in bioreactor.So this study establish rapid methods to detect living cell biomass and physiological characteristic state on-line by On-line b iomass monitor.And we also research the effect of different shearing rates on cell growth and metabolism.By investigating physiological parameters of metabolism CER,density of surviving cells,pH,and relationship between medium nitrogen source consumption and change of extracellular pyruvic concentration,we conclude cells are able to tolerate the shear force in mechanically stirred bioreactor with ?ave not more than 0.8W/kg which was determined by empirical formula.In order to understand intracellular physiological metabolic process,this study have initially build quantitative analysis method of intracellular metabolites by using GC-MS.We inspect different solute and ex traction methods of measuring intracellular organic.The result showed that-20?40%methanol as quencher and 75%ethyl alcohol as extractant under 90?water bath were beneficial to it.Plant cell cultured in environment could make a metabolic defense response,organic acid concentration declined and TCA metabolic pathways changed compared to normal cultured cells.