Study On The Design,Synthesis And Biological Application Of Thiol Fluorescent Probe

Biothiol is a kind of bioactive molecule,which plays an important role in the structure and function of cells and plays an important physiological role in the human body.Hydrogen sulfide is the simplest biological thiol,a toxic gas with a foul smell,and acts as an important gas signal transduction molecule in the body.Cysteine(Cys),homocysteine(Hcy)and glutathione(GSH)are the most abundant small molecules of sulfhydryl amino acids(biological thiol)in mammalian cells,which play an important role in cell growth or apoptosis.The disorder of intracellular thiol can affect cell function and even cause a variety of diseases.In addition,thiol are not uniformly distributed in the Subcellular organelles.Among them,the suborganelle lysosome is the place where biological macromolecules are degraded by hydrolases,and it is involved in many physiological functions such as cell metabolism,intracellular transport,cell membrane circulation and cell apoptosis to maintain cellular environmental homeostasis.The concentration of thiols has an important effect on the morphology and function of lysosom.Therefore,the detection of thiols in subcellular organelles is of great significance for revealing the subcellular function of thiols.Fluorescent probe has many advantages,such as low cost,simple operation,no damage,high sensitivity and real-time monitoring,etc,and has been widely used in biological and medical research.Therefore,it is very important to design and synthesize a high-sensitivity fluorescent detection specific probe for lysosomal thiols.Based on the research status of thiols,three fluorescent probes for detecting thiols were designed and synthesized,and their spectral properties and applications in biological imaging were studied.(?)Hydrogen sulfide(H₂S)plays a key role in lysosomal organelles.H₂S can induce cell death,which is related to the activation of calprotease,lysosomal instability and the release of lysosomal proteases.Therefore,high sensitivity and high selectivity detection of H₂S in lysosomes will help to understand the kinetics and mechanism of intracellular reactions and the physiological role of H₂S in biologic system.In this chapter,we report a new lysosomal-targeted fluorescent H₂S probe CMDN.As the concentration of H₂S increases,the fluorescence of the probe CMDN at 460 nm is greatly enhanced.The probe has high selectivity and remains stable in the presence of interfering substances in the lysosome,with no change in fluorescence emission.Fluorescence imaging showed that CMDN had membrane permeability.Compared with the lysosomal localization dyes,CMDN had a higher co-localization coefficient(>0.90),which could be used to analyze the change of H₂S in lysosomes.In addition,fluorescence imaging results indicate that it can be used as an endogenous and exogenous H₂S probe.The CMDN probe can also detect H₂S in zebrafish at the physiological level.(?)Biothiol is a part of reactive sulfur species,which is a kind of bioactive molecules that play an important physiological role in human body.However,due to their similarity in structure and reaction sites,it is difficult to identify and detect biological thiols simultaneously.In this chapter,a lysosomal-targeted fluorescent probe CM-NBD,was designed and synthesized by linking 7-nitrobenzofuran with aminomorphin-coumarin derivatives by a simple diethyl ether bond.In the presence of Cys and Hcy,the probe generates blue and green fluorescence emission at 480 nm and 550 nm under excitation of two independent wavelengths,respectively.However,the addition of GSH and H₂S to the probe only produced blue fluorescence at 480nm.The probe CM-NBD can be well localized in lysosomes and has a high co-localization coefficient due to its specific localization group.This probe can not only image exogenous biological thiols,but also distinguish Cys from biothiol(Hcy/GSH and H₂S)in cell and zebrafish models.(?)Cysteine(Cys)is a key amino acid involved in many physiological and pathological processes in the human body,and plays an important role in maintaining redox homeostasis of life system.The concentration of intracellular Cys is abnormal under oxidative stress,which leads to many diseases.The ratio fluorescence probe can detect the fluorescence intensity of two emission peaks,which can avoid the errors caused by probe concentration,instrument factors and environmental factors,so as to improve the accuracy of detection.Polymers have attracted much attention because of their advantages of good cytocompatibility and low toxicity.Therefore,we designed and synthesized a polymer ratio fluorescent probe,Nap Cys-PDM for selective detection of Cys.This probe uses naphthimide fluorophore as the fluorescence group and acrylate group as the recognition group of Cys.Biological imaging experiments showed that Nap Cys-PDM could not only detect Cys in organisms,but also monitor the changes of Cys concentration in cells and zebrafish under oxidative stress caused by H₂O₂/LPS.