Design,Synthesis And Biological Imaging Study Of Several Reactive Fluorescent Probes

In recent years, the application of functional fluorescent dyes in biological and medical fields has become one of the most popular areas. Typically, fluorescent probes possess controllable structures, properties of low-cost synthesis, based on the recognition of irreversible chemical reaction. As a result, they can be used for real-time cell imaging. However, the imaging or detection efficiency of the organic fluorescent probes will usually be subject to some limitations in practical biological applications (e.g., sensitivity, specificity, response time). To overcome the above disadvantages, ten novel fluorescent probes have been designed and synthesized in the thesis. The main contents are as follows:1. By modifying the structure of the parent dye, three reactive fluorescent probes were synthesized using novel long-wavelength coumarin dyes as signal groups based on cyclization reaction, oxidation reaction and Michael reaction.1) A novel red fluorescent probe of P1with the emission wavelength of616nm has been synthesized. The probe PI could be used for monitoring and imaging F-in aqueous media based on the F-triggered Si-O bond cleavage and cyclization reaction. Such a probe was highly reactive for F-with high sensitivity and specificity.2) Based on the desulfurization of the thiocoumarin reaction, probe P2which shows fast response toward OCl-has been synthesized. The probe P2was highly selective for OCl-and the influence from other oxidants, free radicals and ions was negligible. Thus, the probe was suitable for detection of OCl-in real samples.3) Based on the Michael reaction of SO32-and α,β-unsaturated double bond, a colorimetric and "turn-on" fluorescent probe (P3) toward SO32-has been produced. Upon addition of SO32-, the absorption spectra were blue-shifted by90nm and the fluorescent intensity was enhanced10-fold. The probe P3has been successfully used for assay of SO32-in real samples.2. Using flavone dye as the matrix, a red fluorescent dye with emission wavelength longer than600nm was synthesized for the first time. Note that the conjugated system of the dye was increased by incorporating4-(dimethylamino)-cinnamaldehyde. The dye exhibited good photostability, large Stokes shift and reasonable fluorescent quantum yield. Using specific recognition groups as the receptor of the dye, two novel "turn-on" fluorescent probes were synthesized:1) A red fluorescent probe toward thiols was synthesized when2,4-dinitrobenzenesulfonate moiety was used as the recognition group. The probe displayed remarkable fluorescence enhancement (560-fold) toward thiol-containing amino acids. Furthermore, the probe has been successfully utilized for thiol imaging in living cells.2) Using2,4-dinitrophenyl ether as the recognition group, a sensitive and selective red fluorescent probe toward H2S was synthesized. By initiating the thiolysis reaction of2,4-dinitrophenyl ether by H2S, the fluorescence of the probe was remarkably increased112-fold. This novel probe was used for assay and imaging of H2S in RAW264.7cells.3. Based on the fact that the ratiometric fluorescent can eliminate the influence from the species concentration, excitation intensity, ambient temperature and other external factors through built-in correction mode, two ratiometric fluorescent probes were synthesized relying on the intermolecular charge transfer process:1) a ratiometric fluorescent probe toward SO32-was synthesized using4-hydroxynaphthylimide. The mechanism involved the sulfite-triggered intramolecular cleavage of the levulinate moiety, thus leading to a remarkable red shift of the absorption and fluorescence of the probe.2) a ratiometric fluorescence probe toward HSO3-was designed based on the blockage of the intramolecular charge transfer process induced by HSO3-.The probe displayed significant ratiometric fluorescence response and137nm blue shift of fluorescence emission.4. A bodipy-based thiol probe was constructed, which exhibited high selectivity toward biothiols. The high specificity of the probe was ascribed to the blockage of the PET and ICT processes caused by the Michael addition reaction of thiols with α,β-unsaturated double bonds. Thus, slight blue shift of absorption and significant enhancement in fluorescence intensity (40-fold) were obtained. This probe was capable of imaging biothiols in living cells. 5. Based on F" triggered Si-O bond cleavage reaction, two colorimetric and fluorescent probes toward F-have been synthesized:1) Using6-acetyl-2-naphthol as the fluorophore, probe P9was designed in terms of the ICT mechanism. Due to the strong nucleophilicity between F" and Si, the probe showed high selectivity toward F-. The9963-fold fluorescence enhancement led to THE assay of F" with concentration lower than40nM.2) Using2-hydroxy-1-naphthaldehyde as the raw material, probe P10was designed. With the addition of F-, the absorption of the probe was red-shifted by95nm and the color of solution was changed from colorless to yellow. This probe showed high sensitivity and fast response toward F".