Synthesis And Application Of Fluorescent Probes For Detection Of Active Species

Biological active species,include active sulfur species?RSS,such as GSH,Hcy,Cys,H₂S,H₂S_n,SO₂ derivatives,etc.?,active oxygen species(ROS,such as O₂^·-,H₂O₂,HOCl,·OH,HOBr,etc.)and active nitrogen species?such as RNS,such as ONOO^-,NO,NO₂,etc.?.Among them,the active sulfur species?RSS?is defined as"redox-active sulfur molecules capable of oxidizing or reducing biological molecules under physiological conditions".As important active sulfur species?RSS?,SO₂/SO₃^2-/HSO₃^-play important roles in living organisms.Studies have shown that SO₂ is another basic signaling molecule in the body in addition to hydrogen sulfide?H₂S?and nitric oxide?NO?.Endogenous SO₂ has been thought to significantly reduce mitochondrial deformation and swelling,and is involved in regulating various physiological processes,such as relaxing blood vessels,increasing antioxidant capacity,and maintaining redox balance.When taking too much SO₂/SO₃^2-/HSO₃^-can cause a range of conditions,such as respiratory diseases,allergic reactions,mitochondrial dysfunction,antioxidant system disorders,glial reactions,neurological and cardiovascular diseases,etc.Hydrogen sulfide?H₂S?is one of the simplest biothiols,which is toxic and smells of rotten eggs.It is found as an important signal conducting substance together with gases such as CO and NO.At the same time,its abnormal levels are related to many diseases such as diabetes,Down syndrome,Alzheimer's disease,and liver cirrhosis.As important members of the active sulfur species,SO₂/SO₃^2-/HSO₃^-and S^2-/HS^-/H₂S play a very important role in physiological and pathological processes in vivo.In recent years,RSS biology has increasingly become the focus of attention.Hypochlorous acid?HOCl?/hypochlorite?Cl O^-?is one of the most important reactive oxygen species?ROS?,which is catalyzed by the enzyme in the human body.In innate immunity,HOCl is an oxidant with strong antibacterial activity.Although it contributes to antibacterials,oxidative stress caused by abnormal levels involves many diseases such as atherosclerosis,arthritis,cardiovascular disease and even cancer.In addition,it has been widely used as an effective substance in daily life,including bleaching of paper and textiles,and disinfection of domestic water.The World Health Organization?WHO?has stipulated that the content of HOCl in drinking water does not exceed 0.2 ppm.Therefore,the accurate measurement of HOCl/Cl O^-becomes an important research area.In recent years,fluorescence imaging using fluorescent probes has the characteristics of real-time,sensitivity,non-invasiveness,low cost,and simple operation,and has become a powerful analysis tool for visualizing biological molecules and environmental monitoring.Therefore,the design and synthesis of related fluorescent probes are of great significance to investigate the distribution and role of active sulfur species in organisms and the rapid and accurate determination of HOCl/Cl O^-in the environment.In the first part of the thesis,a macrocyclic polyamine fluorescent probe molecule MPAD was synthesized using L-proline as a raw material.This molecule can specifically bind to Cu²⁺(Binding constant 1.32×10⁴ M^-1)without interference from other cations to form an MPAD-Cu²⁺complex.Fluorescence titration in the solution showed that Cu²⁺can quench the yellow fluorescence of the probe MPAD and thus obtain a complex with a lower fluorescent background.Fluorescence titration of the in-situ complexes showed that S^2-can significantly enhance the fluorescence of the system without interference,and has a better ability to detect S^2-?LOD=6.1?M?.The in situ formed complexes have been successfully used for fluorescence imaging of Hella cells,so they have potential applications in the detection of H₂S in living cells.In the second part of the thesis,two kinds of fluorescent probes P-Py and P-An containing pyrene and anthracene were successfully synthesized by Knoevenagel condensation reaction for the detection of SO₂.The probe P-Py showed green fluorescence in the aqueous solution,and caused Michael addition reaction after adding HSO₃^-.The fluorescence shifts blue and changes from green to blue,realizing the ratio sensing of HSO₃^-.The probe P-An has blue fluorescence in an aqueous solution,and the blue fluorescence is enhanced after interacting with HSO₃^-,showing a fluorescence-on sensing behavior.The fluorescence titration of the probes P-Py and P-An in solution indicates that both of them have good detection ability for HSO₃^-,which have been successfully applied to the fluorescence imaging research of Hella cells.All these results suggest that they have potential value for biological fluorescence imaging.In the third part,a binaphthol and lipoic acid-derived fluorescent compound BINOL-LA was synthesized.By using the cleavage reaction of partial disulfide bonds of lipoic acid under ultraviolet light or sunlight,silver nanoparticles with the fluorescent molecule as a ligand can be synthesized in a green way.We use silver nanoparticles?Ag NPs?as a carrier to make BINO-LA aggregated on silver nanoparticles,resulting in quenching of fluorescence,and a lower fluorescence background signal.The strong oxidizing property of HOCl/Cl O^-causes the disaggregation of Ag NPs nanostructure and the release of fluorescent molecules,which realize the colorimetric and fluoremetric sensing of HOCl/Cl O^-.This method has the advantages of fast response,good selectivity,and real-time detection.It is successfully applied to the detection of HOCl/Cl O^-in lake water and tap water.