Design,Synthesis And Biological Application Of Fluorescent Probes With Long Lifetime

Fluorescence lifetime imaging is one kind of fluorescence imaging method,which can directly provide the location,binding site and heterogeneity of the measured fluorophore by virtue of its fluorescence life-time characteristic.Fluorescence lifetime imaging has been widely used in many fields,such as cell biology,biophysics and biomedicine.The pure organic thermally activated delayed fluorescence(TADF)molecules,with intrinsic high emission quantum efficiency of 100%,long fluorescence lifetime up second levels low cytotoxicity,are different from many other luminescent materials for fluorescence imaging and can be used as excellent probes for fluorescence lifetime imaging applications.However,many TADF molecules have serious triplet-triplet annihilation(TTA)drawbacks limiting their emission in solid states when directly precipitated into nano-particles,which severely limits the application as active materials for bioimaging.So,it is of great importance to address this issue to pave the way towards their design,synthesis and applications in devices and biological cases.In this theme,a series of D-A-D type high-efficiency TADF molecules were designed and synthesized.Either,a new"preparation of host-guest co-doping,then co-precipitation"nanoparticles preparation method was proposed for the first time.This method can effectively weaken the TTA defects of TADF molecules,to a large extent,resulting into improvement in the stability of nanoparticles in all aspects,achievement of excellent life-time imaging and photodynamic efficacy.The details are as follows:1)Design and synthesis of a series of D-A-D red-emission TADF molecules centered with anthraquinone:A1,A2,A3,A4(A1 is a known molecule).According to their optical and thermodynamic properties and the symmetric structure of A3,the addition of the bulky TPE moitif,A3 has the longest emission wavelength(613 nm)and the smallest?E_STvalue(0.03 e V),and a best thermal stability(thermal decomposition temperature of 530 ^oC),suggesting A3 is the the best TADF among the foure designed molecules.2)Compound A3 was selected as the fluorogen for fluorescence lifetime imaging and therapeutic applications.By developing a new"host-guest co-doping,then co-precipitation"nanoparticle preparation method to reduce the TTA defects of TADF molecules,we obtained several A3-based nanoparticles.Comparatively,it was found that A3(CBP)NPs possessing the largest fluorescence intensity,the best particle morphology and stability.Thus,A3(CBP)NPs was used as nanoprobes for biological applilcations.After A3(CBP)NPs enter the cell,the fluorescence lifetime is only attenuated by 0.4 times,and the ¹O₂ yield reaches 28%,achieving excellent life-time imaging and photodynamic therapy effect.The results show that the preparation of TADF nanoparticles by this method can effectively improve the stability of nanoparticles and further expand the application range of TADF molecules.3)The chlorinated A3 with heavy atoms of Br and I gave two compounds of 2Br A3 and 2IA3.Nanoparticles were prepared using the same method and ratio as A3(CBP)NPs to obtain 2Br A3NPs and 2IA3NPs.Theoretical simulation,and physical properties measurements were conducted.It was found that 2Br A3NPs have a strong fluorescence emission,a fluorescence lifetime of 163?s,and a¹O₂yield of up to 62.5%,and the performance far exceeding A3(CBP)NPs.The results show that the addition of matching halogen heavy atoms can effectively enhance the spin-orbit coupling of electrons and achieve more excellent photodynamic efficacy and time-resolved imaging.