Design,Synthesis And Bioactivity Of Brassinosteroid Mimetics Based On Bikinin

Brassinosteroids(BRs)are a kind of steroidal hormones synthesized in plants.The content of BRs in plants is scarce,but showing the significant regulation on growth,development,reproduction,senescence and response to stress.However,BR and its active derivatives are not economical for large-scale application in agriculture because of their high costs.4-((5-bromopyridin-2-yl)amino)-4-oxobutanoic acid(bikinin,Bik)is a novel highly active and nonsteroidal BR mimetics reported recently.Unlike BR,bikinin directly inhibit BIN2 that belongs to a kinase in the downstream of BR signaling,and transcription factors BZR1/BESlwere activated due to dephosphorylating,then expression of target genes were regulated.However,bioactivity of bikinin is only 1/30 of BR.To develop more potent BR mimetics,the study designed and synthesized 9 series of analogues based on bikinin by means of active substructure stitching and reasonable derivation design,then the compounds were subjected to systematic evaluation of bioactivity.The main results were as follows:1.Based on the binding conformation of bikinin with receptor and drug design method,we reasonably designed 9 series,a total of 36 compounds.Among them,21 compounds were new structure reported for the first time.All the structure of target compounds was confirmed by NMR and HRMS.The synthetic method was simple,easy and economic.Most of post-processing could be completed by recrystallization or washing.2.BR related activity of target compounds were evaluated on the Arabidopsis det2-1 mutant and rice for potent bikinin analogues.The results showed that various derivation of bikinin had different effects on activity.A compound lx had a methylthiazole group bearing an ethyl ester in the carboxyl group,which could result in 57%elongation of WT at 20 ?M and 64%at 40 ?M.In rice lamina inclination,the degree that induced by lx at 50 ?M was the same as the bikinin at 200 ?M.1x performed more efficiently than bikinin,but there was a certain gap between lx and EBL(24-epibrassinolide).Combining with all the results of structure-activity,it suggested that the methylthiazole group has a positive effect on activity in comparison to a halogen and methyl-substituted pyridine group,also better than pyrazole and quinoline groups.Methyl and ethyl ester in the carboxyl group provides better cell permeability,making it more efficient than compounds with carboxyl group.Compounds 1c,1v,1w and 1x,sharing a common thiazole group,exhibited significant differences in activity.This indicates that methyl and size of substituent group in the thiazole ring is crucial for biological activity,just like the halogen for bikinin.Compounds with aromatic ring replacing adipose chain structure could also maintain certain bioactivity.3.The mechanism was confirmed by Arabidopsis mutant growth and molecular docking.Unlike EBL,potent compound(1x(ethyl 4?((5?methylthiazol?2?yl)amino)?4?oxobutanoate,brazide)shares the common receptor BIN2 with bikinin that belongs to a kinase in the downstream of BR signaling.Also,applying lx and bikinin to maize seedling grown in darkness,the elongation of mesocotyl and primary root were observed depending on the low concentration,however,high concentration could significantly inhibit the length of root.Their effects were similar with EBL.4.Effects of brazide(Bra)as BR mimetics on alleviating the herbicide toxicity in susceptible maize(Zhengdan 958)were investigated.By application of NS,maize seedlings were observed growth retardation or even die when the concentration was 400 ?M.Pretreating EBL 0.1?M,Bik 10 ?M and Bra 10 ?M 24 h before NS treatment to seedlings were able to improve NS(100 ?M)induced growth inhibition and damage to photosynthetic system compared with plants only applying NS after 5-7 days.The significantly increased H2O2 and MDA levels in maize leaves comfirmed that NS induced an oxidative stress.However,pretreatment of Bra significantly improved the activities of antioxidant enzymes for scavenging ROS when compared with NS alone.Also,increasement of activities of detoxification-related enzymes,content of GSH and expression levels of GST1,ABC-2,ALS1 significantly alleviated the herbicide toxicity.The rate of NS degradation caused by Bra and Bik were higher than EBL on the third day.However,no significant difference was observed in NS residues induced by EBL,Bik and Bra on the seventh day.