Design,Synthesis And Bioimaging Study Of NIR Cyanostilbene Derivatives

The development of bioimaging technology established a solid foundation for biomedical researches.Fluorescence imaging with the advantages of noninvasive,high sensitivity,and low-cost,plays a key role in bioimaging field.Diverse fluorescent materials currently have been designed and developed for fluorescent imaging contrast agents,including quantum dots(Qdots),upconverted nanoparticles,fluorescent protein and organic dyes.Organic dyes are one kind of good exogenous contrast agents for fluorescence bioimaging due to their excellent biocompatibility and multifunctionality.But most conventional organic dyes have the problem of photobleaching and aggregation caused quenching(ACQ).Different from ACQ fluorescent dyes,a novel kind of aggregation-induced emission fluorogens(AIEgens)which were developed by Tang and co-workers in 2001,are weakly or nonemissive when molecules are uniformly dispersed in solution but have strong emission when forming aggregates.The fluorescence intensity of AIEgens can be prominent enhanced by increasing the concentration of fluorescent molecules inside,which is beneficial for improving the signal-to-noise ratio,photosensitivity,spatial resolution of imaging and resistance to photobleaching.It is well-known that near infrared(NIR)dyes with emission spectra within the“optical window”of biological tissues(i.e.,650-900 nm)can efficiently reduce the interference from tissue autofluorescence.In addition,NIR fluorescent dyes can also provide deep imaging/sensing penetration depth due to the reduced optical attenuation of biological tissues in the NIR wavelength range.Researchers have integrated magnetic contrast agent with AIEgens in a single probe(multimodality contrast agent)to further improve the bioimaging quality.AIEgens based on twisted intramolecular charge transfer(TICT)mechanism is a class of fluorescent probe is sensitive to the microenvironment(such as viscosity,polarity and temperature).TICT probe has been widely used to gas sensing and liquid sensing.Especially in biological,the fluorescence response characteristics of the TICT probe to the microenvironment parameters can be used to visualize the intracellular microenvironment,which is conducive to in-depth investigation of cell behavior and metabolic activities.Simultaneously,the design of TICT molecules with a strong electron donor-acceptor(D-A)structure can reduce the energy gap and achieve near-infrared emission.In this thesis,cyanostilbene-?-triphenylamine as the core part,a series of D-?-A-type cyanostyrene derivatives are designed by connecting donor groups with different electron-withdrawing abilities.These cyanostyrene derivatives have near infrared emission and TICT characteristics since the molecular structure with strong D-A effect.Through the single-molecule water-soluble modification and nanparticles fabrication by polymer coating,we applied our cyanostyrene derivatives in specific imaging of cell microenvironment and fluorescence and multimodal imaging in vivo.The specific research contents are as follows:1.We designed and synthesized a donor–?-accepter luminogen,2-([1,1'-biphenyl]-4-yl)-3-(4-((E)-4-(diphenylamino)styryl)phenyl)fumaronitrile(TBB)by Suzuki coupling reaction.TBB exhibited twisted intramolecular charge transfer based NIR emission,aggregation-induced emission and temperature-sensitive emission features.Ratiometric fluorescent thermometer was constructed by encapsulating thermo-sensitive NIR fluorophore TBB and Rhodamine 110 dye into an amphiphilic polymer matrix F127 to form TBB&R110@F127 nanoparticles(TRF NPs).TRF NPs showed good temperature sensitivity of 2.37%·?^-1,wide temperature response ranges from 25 to 65? and excellent temperature-sensitive emission reversibility.Intracellular thermometry experiments indicated that TRF NPs could monitor the cellular temperature change from 25 to 53? for Hep-G2 cells under photothermal therapy agent heating process,indicating the considerable potential applications of TRF NPs in biological thermometry field.2.We designed and synthesized a D-?-A cyanostyrene derivative TBPY with a pyridine group.TBPY has NIR emission,TICT and polarity sensitivity characteristics.We successfully obtained the water-soluble TBPY pyridine salt TBPYS through the water-soluble molecular modification of TBPY.Compared with TBPY,the emission of TBPYS is further red-shifted,the maximum emission peak is up to 774 nm and the fluorescence is almost quenched.We found that TBPYS exhibited significant fluorescence enhancement in the BSA solution.Compared with the aqueous solution of TBPYS,the fluorescence intensity was increased by 85 times,and the maximum emission peak was blue-shifted to 621 nm.Using the luminescence properties of TBPYS in solvents of different polarities,we designed a bright fluorescent probe based on polar response for cell imaging.Cell imaging experiments show that TBPYS-labeled cells do not need to be washed,and can be imaged directly without washing,and have a high signal-to-noise ratio(S/N=16).3.A new(D-?-A fluorogen(4-((E)-4-(diphenylamino)styryl)phenyl)-3-(4'-(1,2,2-triphenylvinyl)-[1,1'-biphenyl]-4-yl)fumaronitrile(TB-TPE)is designed and synthesized.TB-TPE exhibits TICT and AIE in NIR region with the emission peak at 714 nm and fluorescence quantum yield of 6.6%in solid state.Through encapsulating TB-TPE by polystyrene-polyethylene glycol(PS-PEG),water-soluble TB-TPE@PS-PEG nanoparticles(TP NPs)are fabricated,which displays polymer encapsulation enhanced emission with the Qy of 46.5%due to the strong restriction effect on TICT process and the destruction of H aggregation for TB-TPE by polymer matrix.Then Au coated Fe₃O₄(Fe₃O₄@Au)nanocrystal is embedded in TP NPs to form high fluorescent TB-TPE&Fe₃O₄@Au@PS-PEG nanoparticles (TFAP NPs)with the Qy of 39.7%.Cellular imaging of TP NPs for Hep-G2 cells and multimodality imaging of TFAP NPs for mouse liver tumor are successfully realized,demonstrating that polymer encapsulated TB-TPE holds great prospects as a multifunctional fluorescence probe for bioimaging.4.We design a high-efficient fluorescent and magnetic multi-modal probe by doping a fluorescent molecule 2-(4-bromophenyl)-3-(4-(4-(diphenylamino)styryl)phenyl)fumaronitrile(TB)with near-infrared(NIR)fluorescent emission(714 nm)and superparamagnetic iron oxide(SPIO)into polystyrene-poly(ethylene glycol)(PS-PEG)matrix to form TB/SPIO@PS-PEG nanoparticles(TSP NPs).The as-prepared TSP NPs exhibit red aggregation-induced emission(AIE)with the maximum wavelength at 655 nm and high fluorescence quantum yields(QYs)of 14.6%,which facilitates to improve the sensitivity and signal-to-background ratio for fluorescence molecular imaging(FMI).And the phase behavior investigation in TB/SPIO@PS-PEG probe system by Flory-Huggins lattice theory elucidates the high-efficient fluorescence of the multi-modality probe originated from the poor miscibility between TB and SPIO.Meanwhile,TSP NPs possess good superparamagnetism and relaxivity which can be used as appropriate magnetic contrast agents for magnetic resonance imaging(MRI)and magnetic particle imaging(MPI).In addition,the good biocompatibility and photostability of TSP NPs make them suitable for long-term monitoring.In vivo fluorescence imaging results indicate that TSP NPs can monitor subcutaneous tumor growth for more than 24 days in a real-time manner.Multimodality imaging consisting FMI,MRI and MPI reveals that TSP NPs can monitor liver tumor in situ with almost unlimited depth in tissues and high temporal-spatial resolution.As the multi-modal probes,TSP NPs manifest obvious synergistic advantages of long-term monitoring and high penetration depth and hold great prospects in preoperative diagnosis and intraoperative surgical navigation of carcinoma in situ.