Design,Synthesis And Application Of Coumarin—based Fluorescent Probe For Reactive Sulfur Species

Coumarins are widely exsited in many natural plants.Due to their excellent photophysicalproperties,such as high fluorescence quantum yield,good photo stability,easily tunable emission wavelength and good biocompatibility,coumarins are widely used in the fields of bioimaging and detection.In this work,a series of novel biologically reactive sulfur species?RSS?probes were developed through the modification and optimization of classical diethylaminocoumarin.The sensing properties as well as the application of these probes were investigated.Some major works were summarized as follows:?1?Traditional fluorescent probe for hydrogen polysulfides?H₂S_n?utilizing H₂S_n-induced benzodithione formation always suffer from the interference caused by hydrogen sulfide?H₂S?or biothiols.Herien,C-HPS,a novel H₂S_n-specific fluorescent probe was developed by taking advantage of a unique H₂S_n-induced substitution-cyclizationreaction cascade.C-HPS equipped a relatively poor leaving group,p-methoxyphenol,as a result,only H₂S_n can trigger the cleavage of this moiety.After that nucleopilic subsititution occurs,the unstable intermediate will slowly release the elemental sulfur to generate the corresponding sulfhydryl coumarin.The subsequent intramolecular cyclization would ultimately lead to the formation of a rigid product with larger?-conjugation system.The addition of H₂S_n to the C-HPS system can significantly increase the fluorescence intensity at 576 nm?approximately 97 folds enhancement?.Moreover,C-HPS displayed a fast response time?6 min?and sufficient sensitivity?LOD=20 n M?toward H₂S_n.Application of C-HPS to visualize H₂S_n in MGC-803 cells was also achevied.?2?The development of single-molecule fluorescent probes that can simultaneously distinguish H₂S,H₂S_n and biothiols with different fluorescent signals is of great importance for elucidation the complex functional roles of H₂S,H₂S_n and biothiols in physiological activities.We reported the first single-molecule fluorescent probe,MCP1,which could simultaneously detect H₂S,H₂S_nand biothiols from distinct emission channels.By taking advantage of the differences in nucleophilicities and molecular structures between H₂S,H₂S_n and biothiols,a novel fluorescent probe,MCP1,was developed for simultaneous detection of above three species.Free MCP1 was non fluorescence,however,addition of biothiols,H₂S,H₂S_n to the solution of MCP1 can induce signiciant fluorescence signals enhancement at 469 nm,508 nm and 576 nm,respectively.Moreover,the probe MCP1 shows perfect selectivity,the detection H₂S or H₂S_n could realized even high level of bakground biothiol exsit.Co-localization imaging experiments indicating the probe was mainly located in the lysosome.Notably,application of MCP1 to distinguish image of hydrogen sulfide and hydrogen sulfide in cells could be realized sucessfully.In order to study the reaction mechanism,between MCP1 and H₂S,purification of the reaction mixture was conducted and a green fluorescent product ND was successfully separated.The chemical structure of ND was characterized by NMR,HR-MS,FTIR,XRD,and it was proved to be a new coumarin-naphthyridine structure.Optimization of the reaction conditions between H₂S and MCP1 can increase the yield of ND to 97%.Moreover,theis reaction was quite universal and a series of coumarin-naphthyridine dyes with different electron donors were readily synthesized through the optimized reaction condition.Significantly,ND displayed excellent photophysical properties,such as high quantum yield?QY=0.508?,good stabilityas well as excellent two-photon performance?91.908GM?.Meawhile,the free amino and carboxyl group in its chemical structure can give this novel dye platform a great potential for biological applications.