Design And Synthesis Of Ratiometric H₂S And PH Fluorescent Probes With Near-infrared Excitation And Its Application In Imaging

Fluorescence amaging based on fluorescent probes has become one of the most important methods in biological imaging applications due to its high sensitivity,excellent selectivity,less cell photodamage and acile preparation.However,most of fluorescent probes reported so far for biological imaging exhibited some limitations:Firstly,most of fluorescent probes are excited in ultraviolet-visible area,where they could be seriously suffered from autofluorescence interference of matrix.Meanwhile,there are also serious photobleaching,phototoxicity,and imperfect tissue penetration for bioimaging.Secondly,fluorescent probes response to an analyte of interest with optical signal changes only in fluorescent intensity.Single wavelength emission intensity-based probes are susceptible to interferences from various factors,such as photobleaching,surrounding microenvironment(pH,polarization,temperature,etc.),probe concentration and detection devices,which resulting in measuring the signal distortion;(3)Although ratiometric fluorescent probes have gradually increased in recent years,most of them are low-resolution due to serious overlap with a short shift of emission.Therefore,it is of great importance to develop novel ratiometric fluorescence probes with large emission shift based near infrared excitation.In this thesis,two types of fluorescence probes were designed and synthesized for imaging of H2S and pH in living cells and tissues.The concrete contents are as follows:(1)In Chapter II,We have designed and synthesized a two-photon ratiometric fluorescence probe based on principles of fluorescence resonance energy transfer(FRET)for detecting H2S in living cells and tissues.The small molecule ratiometric two-photon fluorescent probe,TR-H2S,comprised of a well-known acedan(2-acetyl-6-dialky laminonaphthalene)moiety as a two-photon excitation energy donor and a H2S-responsive anthocyanidin-analogue dye as an energy acceptor,linked by a rigid spacer.Experiment results indicated that TR-H2S displays a remarkable emission shift(up to 125 nm)with two well-resolved emission bands.Upon selective reaction with H2S,the conjugated structure of acceptor is interrupted by nucleophilic addition reaction,thus FRET process is suppressed,and green emission increase on excitation of the donor as well as crimson emission of the receptor weakened.The rapid response and ratiometric detection for H2S was realized by comparing the two emission band signals.It was also successfully applied to the two-photon ratiometric imaging detection of H2S in cells and tissues.(2)In Chapter III,We have designed and synthesized a near-infrared fluorescence probe based on principles of intramolecular charge transfer(ICT)for detecting the pH in mitochondria.The four pH-sensitive near-infrared fluorescent dyes BTCS1-4 we achieved by introducing 2-(2-hydroxyphenyl)benzothiazole group into the near infrared dye structure of CC.Under acidic conditions,the BTCS dyes emit fluorescence with short wavelength,and with the increase of pH of the solution,the hydroxyl group is deprotonated to give the O-with strong electron donating capacity,as a result,the fluorescent absorption/emission spectra of the BTCS dyes show red shift.Among the BTCS dyes,BTCS1 showed well-resolved emission spectrum in various pH conditions,and its derivatives(BTCS-pHmito)can mainly targeted in the mitochondria,indicating the near infrared fluorescent probe based on BTCS1 has potential applications in the detection of mitochondrial pH.