Study On Synthesis And Application Of Fluorescent Dyes Based On Naphthamide Derivatives

Naphthalimide,as a kind of common fluorescent dye,has been widely concerned because of its stable structure and high fluorescence quantum yield since it was reported.4-amino-1,8-naphthalimide（ANI）is one of the most representative and widely studied dyes.With the continuous development of biological imaging technology,it plays an important role in the field of fluorescence imaging because of its excellent luminescent performance,easy modification and large Stokes shift,especially as a tool for disease diagnosis.Although ANI plays an important role in medicine,its synthesis method is still relatively simple.As far as we know,the current synthetic route of ANI is not only complicated,but also it is inevitable to use some dangerous drugs.Therefore,if we can design a safe,simple and efficient route to synthesize ANI,it will help to promote its further development in the fields of industry and biomedicine.Organelles are specialized subunits in cells,which play a specific and indispensable role in cell process.Fluorescence imaging technology is often used to study subcellular organelles because of its special features such as super-temporal resolution,non-invasive and real-time monitoring.However,the application of common naphthalamide fluorescent dyes in cells is often limited by poor water solubility or aggregation-caused quench（ACQ）.However,small molecular fluorescent dyes with aggregation-induced emission（AIE）and high molecular fluorescent dyes with good water solubility can be completely applied to specific cell microenvironment with high water content.Therefore,it is of great practical significance to develop fluorescent dyes that can target organelles and are perfectly suitable for cell-specific microenvironment.Based on the above background,this thesis innovatively proposed and optimized the synthetic route of ANI,and on this basis,developed a series of small molecules and polymer fluorescent dyes which can target different organelles and are suitable for cell microenvironment.The main contents are divided into the following three parts:PartⅠ:An innovative method of synthesizing ANI from 4-bromo-1,8-naphthalimide by classical Suzuki coupling reaction under the catalysis of palladium acetate was proposed,which avoids the use of explosive and dangerous chemicals.In addition,by optimizing the experimental conditions,the reaction yield was greatly improved and the substrate platform was broadened.A series of fluorescent dyes with different functions were prepared through this scheme to construct the fluorescent platform of ANI.Among them,dyes NNAP-LD（3e）,NNAP-ER（3i）and NNAP-Lyso（3j）not only have good luminescent properties,but also have different targeting groups,so they can accurately target the lipid droplets,endoplasmic reticulum and lysosomal subcellular organelles in He La cells.Finally,based on the above research,a new fluorescent probe NNAP-Lyso-Sec was designed and synthesized for targeted detection of selenocysteine（Sec）in lysosomes.Part II:In view of the fact that AIE fluorescent probe is not only suitable for cell microenvironment with high water content,but also has the advantages of good brightness,long-term in-situ retention,light stability and low cytotoxicity.In this part,a series of new dyes（NR-Lyso-I,NR-Lyso-II,NR-Lyso-III,NR-Lyso-IV）were designed and synthesized by alkyl chain engineering on the fluorescence platform of ANI.It is found that the alkylation reaction of ANI can transform the dye from ACQ effect to AIE effect.In addition,the AIE probe NR-Lyso-IV not only showed the characteristics of large Stokes shift（>100 nm）,excellent selectivity,light stability and low cytotoxicity,but also could simultaneously visualize lipid droplets and lysosomes in He La cells and zebrafish in vivo imaging.Part III:Water-soluble polymer dyes with different molecular weights（Nap NH₂-PDM-A,Nap NH₂-PDM-B,Nap NH₂-PDM-C,Nap NH₂-PDM-D）were accurately constructed by using the ANI fluorophore constructed above and poly（N,N-dimethylaminoethyl methacrylate）by RAFT polymerization,and their luminescent properties were studied and compared.The test results showed that all of them had the advantages of large Stokes shift,excellent light stability and good biocompatibility,but there were significant differences in fluorescence intensity and self-assembly performance in aqueous solution.In addition,the diameter of nanospheres is the key factor affecting the permeability of dye cells,so the difficulty of four polymer probes entering cells was further studied.The polymer dye Nap NH₂-PDM-C with good cell permeability was successfully screened by confocal fluorescence imaging technology.It not only has good cell permeability,but also can label lysosomes in cells quickly and accurately.Therefore,this work provides an important reference for constructing related polymer fluorescent dyes with good cell permeability in the future.