Developing The Synthesis Of Fluorescent Probes Based On Enviornment-Sensitive And Their Applications

Environment-sensitive dyes are a series of molecules whose fluorescent properties vary with the physical and chemical properties of their surroundings. Generally, these dyes affords very weak fluorescence emission in polar solvent, but fluoresces intensively in nonpolar solvent. In general, environment-sensitive fluorescent probes are poorly emissive in aqueous solution, while become highly fluorescent when bound to hydrophobic sites of proteins. Introducing groups of protein recognition to these environment-sensitive fluorescent molecules can pave a new way for the detection of targeting proteins with high sensitivity and selectivity. Therefore, these fluorpophores are attracted much more attention of researchers in recent years.Vicinal dithiol-containing proteins (VDPs) are proteins which contain a pair or more space-closed sulyfhdryl groups. In general, the vicinal thiols in biological systems are derived from cysteines that are sequence proximal. In oxidation condition, VDPs can form intramolecular or intermolecular disulfide bonds, and undergo reversible oxidative conversion to the intra- or interprotein disuldes in the redox environment. Thus, the detection of vicinal dithiol groups in proteins is still a challenge for scientists.Trivalent arsenic was reported as the recognition unit of vicinal thiols in proteins, but it is unstable and vulnerable oxidized to be pentavalent arsenic. Generally, trivalent arsenic can react with 1,2-ethanedithiol (EDT) to form a 5-membered dithiarsolane ring, which is more stable than the 6- and 7-membered ones. The 5-membered dithiarsolane ring can selectively discriminate vicinal dithiols from other forms of thiols. Bsed on the aboved mechanism, we combined the special recognition function of arsenical with the properties of environment-sensitive fluorophores to successfully design a fluorescent light-up probe for highly selective and sensitive detection of VDPs. The paper mainly divides into three chapters, which are listed as following:Chapter ?:The types of environment-sensitive fluorescent probes and applications on the detection of proteins are mainly introduced.Chapter ?:2-(4-dimethylaminophenyl)-4-(2-carboxyphenyl)-7-diethylamin-1-benzopyrylium (F1) possesses a D1-A-D2 system, and its fluorescence intensity increases obviously with the decrease of solvent polarity. In addition, dimethylaminophenyl and diethylamino groups in F1 can rotate with the solvent flexibility, and emissive intensity of F1 is greatly enhanced as the solvent viscosity increases. Thus, compound F1 is an environment-sensitive dye.In the paper, compound F1 was selected as fluorophore, and 2-(4-aminophenyl)-1,3,2-dithiarsenolane (PAO-EDT) was chosen as the recognition unit, to construct a fluorescence turn-on probe FAsH for detecting VDPs. Here, reduced bovine serum albumin (rBSA) was selected as the model protein and the fluorescence response of FAsH toward rBSA was examined. The experiment results showed that once introducing rBSA to the solution of FAsH, a dramatic increase in the fluorescence intensity was observed at 651 nm within 2.5 min, which then reached a plateau as the reaction proceeded. The fluorescence intensity of FAsH is directly proportional to the concentration of rBSA over the range of 0.06-0.9 ?M, with a detection limit of 0.015 ?M.In chapter ?:We combined 7-(diethylamino) coumarin derivatives as fluorophores with PAO-EDT as the recognition group to construct a ratiometric fluorescent probes CAsH2. It exhibited two emissions in water, which caused by intramolecular charge transfer (ICT,475 nm) and twisted intramolecular charge transfer (TICT,550 nm), respectively. However, once adding rBSA to the solution of CAsH2, the TICT was restricted by the protein microenvironment, which leads to the decrease of the fluorescence intensity at 550 nm. Meanwhile, the ICT was also restricted by the lower polarity of the detection system, resulting in higher fluorescence intensity at 468 nm. Based on this mechanism, the ratiometric fluorescent probe for VDPs was developed. As a control, compound CAsH1 and CAsH3 were synthesized to demonstrate the effect of stric hindrance and intramolecular rotation on detecting VDPs. Probe CAsH2 shows a good selectivity and sensitivity toward VDPs. The emission ratio (I468/I550) shows a linear relationship with VDPs concentration in the rang of 0-1.2 ?M. The detection limit was calculated to be 2.6 nM.