Study On Enhancment Of Effective Compound Accumulation In Orostachys Cartilaginous Calluses And Pathway With Related Signaling Molecules

Orostachys Cartilagineus is belongs to the Crassulaceae family,which has high medicinal value.At present,there is a shortage of wild resources and artificial cultivation techniques,which limits the source of raw materials and hinders their resource utilization and development.Therefore,the application of plant cell culture technology is another way to obtain raw materials.In this study,the callus of Orostachys cartilagineus was used as the material.In this study,the callus of Orostachys cartilagineus wa used as the experimental material,and two kinds of elicitors,methyl jasmonate(MeJA)and salicylic acid(SA),were selected and optimized.The best way to induce callus induction;at the same time,by studying the synthesis of nitric oxide(NO)and its related enzyme activities in the flavonoid synthesis pathway mediated by MeJA and SA,in order to improve the secondary metabolism by regulating the level of NO signal.The accumulation of products provides a theoretical basis for the future development of Orostachys Cartilagineus products.In order to promote the accumulation of effective substances in callus,SA and MeJA were used to induce the accumulation of active substances.After 25 days of reactor incubati-on,the concentration of the inducer was 100 μmol.When treated with SA for 48 h,it was beneficial to the accumulation of total polysaccharides and total flavornoids.SA treatment for 36 h was beneficial to the accumulation of total phenols;total accumulation of total pol-ysaccharides,total phenols and total flavonoids was best at 48 h after MeJA treatment.The results of the addition of elicitor culture in the reactor showed that the two elicitors signifycantly increased the accumulation of active substances in the callus,and the accumulation of active substances in the SA-treated callus was slightly higher than that of MeJA.To determine the changes of H2O2,NO and related enzymes in the synthesis of flavonoids induced by MeJA and SA,the activities of nitric oxide synthase(NOS),nitrate reductase(NR)and five flavonoids were determined.Catalase(CAT)analysis of H2O2-mediated MeJA and SA-induced flavonoid synthesis;compensation by nitrate reductase(NR)using nitrate reductase(NR)inhibitor tungstate(TUN)Type inhibitor glutamine(Gin),nitric oxi-de synthase(NOS)inhibitor PBITU,NO exogenous donor sodium nitroprusside(SNP)and NO quencher c-PTIO.The results showed that NO is key signal molecule in the synthesis of flavonoids induced by MeJA and SA.NR plays an important role in the synthesis of flavonoids under the participation of NOS,and the peak time of NO is earlier than flavonoids.Upstream,the peak time of H2O2 is later than NO,which is presumed to be a downstream molecule of NO.Therefore,adding MeJA and SA to treat callus can effectively increase the accumulation of effective substances,not only to meet market demand,but also to protect natural resources and maintain the ecological environment.