| Achyranthes bidentata Blume.is a perennial,deep roots,dicotyledonous plant,and is an important kind of traditional chinese bulk herbs.In this paper,we use the system that cell suspension culture of Achyranthes bidentata Blume.as tested material,and use cell suspension culture,atomic absorption spectrophotometry,spectrophotometric and high performance liquid chromatography technology,to study the growth of Achyranthes bidentata Blume.suspension cell,nutrient consumption,accumulation of cell oleanolic acid and polysaccharides,in addition to analysis the influence of Me JA and SA on the Achyranthes bidentata Blume.cell biomass and accumulation of major medicinal components,in order to provide theoretical basis for taking advantage of Achyranthes bidentata Blume.suspension cell in large-scale to product more main medicinal ingredient.The results of this study are as follows:1.Callus induction and subculture.The optimal medium of Achyranthes bidentata Blume.leaf callus induction was B5 + 2,4-D 2.0 mg·L-1 + 6-BA 0.5 mg·L-1 and the induction rate was 87.5%;The callus weredivided into fourdifferent types after subculture,which were A,B,C,D types respectively,and A-type callus have loose structure,rapid growth and were the most suitable callus for cell suspension culture.2.Establishment of cell suspension culture system of Achyranthes bidentata Blume.Select the fast-growing A type callus whose were color and quality were better,then,add them to sucrose medium(B5+ 2.0 mg·L-1 2,4-D + 0.5 mg·L-1 6-BA),3% sucrose,30 g·L-1（FW）inoculation,adjust the p H 5.5,a week subculture cycle,build fast growth,gooddispersion and stability Achyranthes bidentata Blume.cell suspension culture system.3.The study of growthdynamicsduring Achyranthes bidentata Blume.cell suspension culture.Measure the cell growthdynamics in the process of Achyranthes bidentata Blume.cell suspension culture.1)Achyranthes bidentata cell growth curve was like "S",and the 0-6d wasdelay period phase,7-21d was logarithmic growth phase and 21-24d was stable growth phase.2)The p H value of culture mediumdecreased first,then increased.The culture medium conductivity curve and cell biomass growth curve was mirror relationship,anddeclined firstly after rising;3)When cells were cultured 15d suspendly,sucrose is almost absorbed completely in medium,when 21d,nitrogen,phosphorus,potassium and magnesium ions in the medium were also consumed widely,73.75%、98.05%、91.14%and 79.01%,but calcium ion concentration had been adequate,and could meet the basic needs of cell growth;4)The accumulation curve of oleanolic acid in Achyranthes bidentata were similar to curve of cell growth,they belong to the growth-coupled relationship,when suspension cells were cultured 24d,the content of oleanolic acid was the highest,which was 1.418 mg·g-1;5)In the Achyranthes bidentata cell suspension culture,the cells polysaccharidein content increased to the peak at 18d,which was 6.56mg·g-1.The accumulation curve of polysaccharides and cell growth curvedoes not fully synchronized,and cell polysaccharide content reached to the highest point earlier than the cell biomass.4.The influence of Me JA on the suspension cell biomass and cell oleanolic acid and polysaccharides accumulation of Achyranthes bidentata Blume.1)When Achyranthes bidentata suspension cells were cultured after 9d,addingdifferent concentrations methyl jasmonate,and cultured coverage to 24d,which was added 0.01 mmol·L-1 methyl jasmonate,cell biomass,cell oleanolic acid and polysaccharides reached highest,respectively was 17.51 g·L-1（DW）、2.67 mg·g-1、3.47 mg·g-1.2)Cell suspension were cultured 0d,6d,12d,18d,21d,respectively,adding 0.01 mmol·L-1methyl jasmonate,the culture coverage to 24d,it was found when 0d add,cell biomass、cell oleanolic acid content reach the highest,17.05 g·L-1（DW）、3.11 mg·g-1,but 0d add,cells polysaccharide content reached the highest,were 3.24 mg·g-1.3)When suspension cells were cultured 0d,adding 0.01 mmol·L-1methyl jasmonate,and coverage after 15d,18d,21d,24d,27d,which subjects in 24d,cell biomass and cell oleanolic acid reached highest,14.79 g·L-1（DW）and 3.74 mg·g-1,but polysaccharides reached highest in 21d,3.89 mg·g-1.5.The influence of SA on the suspension cell biomass and cell oleanolic acid and polysaccharides accumulation of Achyranthes bidentata Blume.1)When Achyranthes bidentata suspension cells were cultured 9d,addingdifferent concentrations SA,and cultured coverage to 24d,which is added 1 mg·L-1 SA,cell biomass,cell oleanolic acid and polysaccharides reached highest,respectively was 16.13 g·L-1（DW）、3.27 mg·g-1 and 3.65 mg·g-1.2)Cell suspension were cultured 0d,6d,12d,18d,21d,respectively,adding 1 mg·L-1 SA,the culture coverage to 24d,it was found that cell biomass 、 cell oleanolic acid content and cells polysaccharide content reach the highest,16.91 g·L-1（DW）、2.82 mg·g-1 and 3.64 mg·g-1.3)When suspension cells were cultured 18d,adding 1 mg·L-1 SA,and coverage after 20d、22d、24d and 26d,which subjects in 24d,cell biomass、cell oleanolic acid and polysaccharides reached highest,18.14 g·L-1（DW）、3.36 mg·g-1 and 3.50 mg·g-1,respectively. |
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