Design And Synthesis Of Cyanine Small Molecule Fluorescent Probe Based On ICT Mechanism And Its Application In Bioanalysis

In recent years,with the interdisciplinary combination of chemistry and biology,many new detection methods have been developed and they provided some new technical support for the exploration and understanding of biological and pathological changes in the diagnosis and treatment of diseases.Among these detection technologies,fluorescence probe has attracted extensive attention due to its simple synthesis and low cost,high sensitivity and good selectivity for the detection of target analyte,as well as its ability for in-situ and real-time dynamic detection.In this paper,based on the Intramolecular charge transfer（ICT）sensing mechanism,two fluorescent chemical sensors were designed and synthesized with Cyanine derivatives as fluorophores.Their spectral properties,monitoring and biological imaging applications were studied.The main contents of this paper are as follows:1.A double-mode optical probe（McyA）based on Merocyanine has been developed for detection of hydrazine and it’s in vivo and in vitro biological imaging application.In this work,the probe McyA for hydrazine（N₂H₄）was designed and synthesized with McyOH as the fluorophore and acetyl as the recognition group.Since the quaternary ammonium salt structure in the McyOH molecule has a strong electron-withdrawing effect（pulling）,and its phenate ion has a potential electron-donating effect（push）,which constitute aπ-conjugated system with a’push-pull’structure（ICT）.This’push-pull’conjugate system is capable of stimulating strong fluorescence.When the acetyl group is combined with the phenolic hydroxyl group,the formed probe McyA blocks the ICT process,which causes the initial fluorescence of the fluorophore to be quenched.However,upon addition of N₂H₄,N₂H₄ will attack the carbonyl group in McyA,which causes the electrons on the oxygen to transfer and form acetyl hydrazide.Furthermore,the fluorophore McyOH is released andthe fluorescence is restored.Simultaneously,the color of the probe changed from faint yellow to pink under natural light and it enables the naked eye detection of N₂H₄.The linear range is 0-5.0μΜand the detection limit is 0.042μΜ,which is much lower than the government requirement for the N₂H₄ content in drinking water（<10 ppb）.In order to explore the practical application,bioimaging experiments were conducted and demonstrated that the probe McyA has good biocompatibility and can be applied for tracking detection of N₂H₄ in vivo.2.A water-soluble near infrared fluorescent probe for ultrasensitive detection and imaging of dipeptidyl peptidase IV in biological sample has been developed.In this work,the probe（HCA-D）was designed and synthesized via Cyanine as the fluorophore and Boc-Gly-Pro-OH as the recognition group of dipeptidyl peptidase IV（DPP-IV）,respectively.Based on the property of fluorophore cyanine,the synthesized probe HCA-D has prominent optical properties,long wave emission,large conjugate plane and larger Stocks displacement base.In addition,the probe HCA-D has excellent water solubility due to the introduction of hydrophilic sulfonate group.Upon addition of DPP-IV under physiological conditions,the probe HCA-D emitted a significant NIR fluorescence signal at 690 nm with obvious red shift.In concentration range of 0-10 ng mL^-1,HCA-D reached an ultrasensitive response(LOD=0.19 ng mL^-1)for DPP-IV.According to above properties and the high selectivity of the probe,HCA-D can be utilized for accurate and dynamic monitoring of DPP IV activity in vitro and in vivo.Finally,it was used to determine the activity of dipeptidyl peptidase IV（DPP-IV）in human serum from actual samples and its bioimaging applications in living cells.