Design,Synthesis And Applications Of Ratiometric Fluorescent Probes Based On FRET Mechanism

In contrast to the general instrumental analysis methods,fluorescence analyses have many advantages of high sensitivity,high selectivity,simple manipulations,and real-time detection.Among them,compared with nano fluorescent probes and fluorescent protein sensors,organic small molecular fluorescent probes,which have outstanding features such as their easily modifiable molecular structures,adjustable fluorescence performance,low biological toxicity,and good biocompatibility,have won the favor of many researchers.Reports about the detection of analytes such as metal ions,anions and active small molecules by using organic small molecular fluorescent probe have emerged one after another.According to the fluorescence emission signal changes of the probe after detecting the analyte,organic small molecular fluorescent probes are mainly divided into single emission,multi-channel and ratiometric type.After the ratiometric fluorescent probe responds to the analyte,it exhibits changes in two different fluorescent signals.We can calculate the ratio of the fluorescence intensity at different wavelengths to make the probe achieve ratiometric detection of the analyte.This feature enables ratiometric fluorescent probe to effectively eliminate interference from test instruments,test environments and test methods,and realize self-calibration of probe to avoid the influence of probe concentration.Forster Resonance Energy Transfer(FRET),which is often used in the construction of ratiometric fluorescent probes,is a nonradiative process.The excited FRET donor transfers energy to the FRET acceptor in the ground state through long-range dipole-dipole interactions.The main point of designing FRET probes is to ensure effective overlap between the donor emission spectrum with the acceptor absorption spectrum,thereby improving the energy transfer efficiency(ETE),and to ensure the two well-separated emission bands with comparable intensities,thus improve the accuracy of detection.Nevertheless,the larger the spectral overlap,the shorter the emission shifts of FRET systems.It is difficult to balance these two points.Therefore,it is still a big challenge to design FRET ratiometric fluorescent probes with high ETE and longer FRET emission shift.We are committed to the design,synthesis and application of ratiometric fluorescent probes based on the FRET mechanism.We designed and synthesized a SO2 derivative probe,a pH probe,and a thiophenol probe using 1,8-naphthalimide or coumarin as the FRET donor.The spectroscopic properties and application of the first two probes in biological detection and other aspects has been further explored.Chapter 1,we outline the common recognition mechanisms of fluorescent probes and the research progress on the fluorescent probe for detecting SO2 derivatives,pH,and thiophenol.Chapter 2,using 1,8-naphthalimide derivatives as the FRET donor and the benzoindole-based hemicyanine moiety as the FRET acceptor,a new endoplasmic reticulum(ER)targeted ratio fluorescent probe NBIS for detecting SO2 derivatives was developed.The probe has a high ETE of 89%and a detection limit of 16.2 nM for HSO3-/SO32-.In addition to its ER-targeting properties,NBIS has also been successfully applied to image exogenous and endogenous HSO3-/SO32-in living cells.Chapter 3,using coumarin derivatives as FRET donors,a new type of ratiometric fluorescence and colorimetric pH probe BDMNP based on the FRET-ICT mechanism was designed.The probe has the ETE of up to 96%and the clear spacing of two emission peaks(113 nm).It possesses good linear response in the pH ranges of 5.4-6.8(pKa=6.10),which can monitor lysosomal pH changes in living cells and detect the subtle pH changes of a variety of environmental water samples.Chapter 4,a new ratiometric thiophenol fluorescent probe DMNP was designed and successfully synthesized,in which coumarin derivatives was used as FRET donors.