Synthesis And Biological Application Of Viscosity-responsive Fluorescent Probes

Intracellular viscosity has highly sensitive properties and the ability to maintain stable changes in the cellular environment,playing a crucial role in regulating physiological processes such as diffusion,transportation,and signal transduction between biomolecules.Abnormal viscosity will affect the complex system balance within cells,leading to physical dysfunction and multiple concurrent diseases（such as fatty liver,inflammation,hyperlipidemia,Alzheimer’s disease,etc.）.As an important cellular messenger,SO₂ is of great significance in regulating the normal operation of cell function.SO₂ often exists in the form of anion(SO₃^2-)in human body,while the effect of SO₂/SO₃^2-on viscosity change is not clear.Hence,it is important to study the relationship between viscosity and SO₂/SO₃^2-to understand the relevant biological and pathological processes.We expect this work to provide a significant impetus for the development of optical imaging tools suitable for biological viscosity environments and promotion of basic research on viscosity related diseases.In this thesis,a series of fluorescent probes connected by single bond,double bond and extended double bond were designed,which based on the different connection bonds between electron donor and acceptor in molecular rotor fluorescent probes,and their different response results with viscosity changes were described.From the perspective of biological field,this project also studied the impact of SO₃^2-/SO₂ on viscosity changes in mitochondria and its application in the diagnosis of related diseases.Herein,we have not only prepared fluorescent probes 2-2,2-3 that are sensitive to viscosity,but also designed and synthesized a dual-functional fluorescent probes 3-1,which can respond to SO₃^2-/SO₂and viscosity simultaneously on the basis of 2-2.The specific research works are summarized as follows:1.In response to the current problems of single structure of intracellular viscosity imaging tool and less research on functional diseases related to viscosity abnormalities,a series of near-infrared fluorescence probes 2-1,2-2,2-3,2-4,2-5 with single bond,double bond and extended double bond between fluorophore and molecular rotor were designed and synthesized.According to the experimental results,with the increase of viscosity,2-2and 2-3 showed high sensitivity to viscosity response and strong red fluorescence at 740nm;in addition,2-3 had excellent mitochondrial targeting ability,which can monitor the viscosity changes of mitochondria in living cells in real time;more importantly,2-3 had been used for imaging abnormal viscosity of fatty liver at the organ levels.In this work,we found that improving the connection bonds of molecular rotor fluorescent probes can be used as a general strategy for regulating the viscosity sensitive response of molecular rotors.2.In view of the difficulties in the design of the unclear relationship between viscosity and SO₂/SO₃^2-,fluorescence probes that can respond to SO₂/SO₃^2-and viscosity at the same time.Based on the modification of 2-2 with high sensitivity response viscosity in the previous research work,a dual-functional fluorescence probe 3-1 with SO₂/SO₃^2-and viscosity response was designed and synthesized,in which benzopyran unit was used as the reaction site of SO₂/SO₃^2-.In the spectral experiments,with the increase of SO₃^2-concentration,the fluorescence intensity of 3-1 at 740 nm decreased,indicating that there was a negative correlation between SO₃^2-concentration and the fluorescence intensity of3-1.In the cell experiments,it was observed that the mitochondrial viscosity gradually increased during the treatment with SO₃^2-,and finally induced cell apoptosis through the loss of MMP（mitochondrial membrane potential）.Treatment with SO₂ gas led to a obvious decrease in mitochondrial viscosity,while MMP level did not change clearly in these processes.In this work,we found that SO₂ and SO₃^2-have significantly different biological effects in regulating mitochondrial related biological processes for the first time.