Development Of Green Fluorescent Probes Based On Coumarin And BODIPY And Their Biological Applications

Fluorescence imaging technology has been widely used in biological functional molecular markers and detection.Based on fluorescent dyes,the fluorescent labeling technology and fluorescent molecular probe have the advantages of simple operation,low cost,good reproducibility,high sensitivity and good membrane penetration.Combining with fluorescence imaging technology,the real-time imaging analysis and in situ non-invasive detection of target small molecules in organisms can be realized,so as to monitor the small-molecules level of living cells and study their physiological and pathological processes.The changes of redox state of cells are closely related to their physiological and pathological processes.Hydrogen sulfide(H2S),biothiols,human NAD(P)H:quinone acceptor oxidoreductase I(hNQO1),hydrogen peroxide(H2O2)and other small redox molecules widely exist in organisms,which play an important role in maintaining the homeostasis of intracellular redox balance.Detection of physiological level by fluorescent probe is helpful to the research,diagnosis and treatment of related diseases.Among them,there are many endogenous fluorescent molecules in plant cells,which will interfere with the detection of it.However,the green light is the best wavelength range for detecting small molecules in plants,it is necessary to develop green fluorescent probes to study redox reactions in plant cells.A new coumarin fluorescent dye and a green NBD-BOD fluorescent probe have been developed for detecting intracellular redox small moleculars,such as H2S,biothiols,hNQO1 enzyme and H2O2.The novel dye shows excellent properties and has the largest Stokes shift(Δλ=138 nm)known in coumarin fluorescent dyes.Importantly,the secondary amine in the coumarin dye enable it to further biologically connect with other detection groups to obtain a series of derivatives.Representative derivatives were showcased for various successful applications of fluorogenic enzyme assay,biolabelling and mitochondrial staining.A green NBD-BOD fluorescent probe was developed based on the thiolysis of NBD amines.It shows high sensitivity,good biocompatibility and can be used for real-time visualization of plant endogenous H2S under physiological conditions.As NBD-amine cracking probe has the best selectivity in the detection of biothiol and H2S,the detection limit of the probe for H2S is 56 nM,and the detection of H2S is not affected by other biothiols and active substances.