Construction And Biological Imaging Study Of Fluorescent Probes For Detection Of Biothiols

Biothiols(glutathione,cysteine and homocysteine)are components of living organisms which play vital roles in many physiological processes such as protein synthesis,antioxidant in the cellular,redox,carbon dioxide fixation and so on.The abnormal levels of biothiols in living systems are associated with many major diseases.As the signal transfer molecule,endogenous hydrogen sulfide and polysulfide play positive roles in a series of physiological activities such as cardiovascular relaxation and neurotransmitter conduction.Therefore,it is significant to detect these thiols.Fluorescence probe has the advantages of high sensitivity,controllable structure,easy synthesis,low cost and high spatiotemporal resolution which has been widely used in biological and medical research.Although partial success has been obtained in construction of thiol fluorescent probes,there are still many limits in sensitivity,selectivity or responsiveness.On the other hand,there are few fluorescent probes have been reported which sensing of homocysteine,glutathione or polysulfide which is urgent need for further studies.Due to the mutual transformation and restriction of the equilibrium relationship between endogenous biothiol,it is facing great challenges for its dynamic monitoring.Therefore,the development of multi-functional fluorescent probes to achieve simultaneous detection of multiple target thiols provides a deeper understanding of the dynamic distribution and the transformation law of these thiols which has very important scientific value and practical significance.In this thesis,based on the integration of organic chemistry,analytical chemistry,and biological sciences,taking the structural innovation of small-molecule fluorescent probes as the starting point,and aiming at solving above bottlenecks in the development of biothiol fluorescent probes,a series of biothiol fluorescent probes were designed and synthesized.For this purpose,the work carried out in this paper can be divided into five parts as follows:(1)In this chapter,based on heavy atom effect and intramolecular charge transfer strategy,a turn-on fluorescent probe has been designed and synthesized for the detection of Hcy based on benzothiazole.The probe displays fast response to Hcy within 10 min,and presents high sensitivity with a low detection limit of 0.16 ?M in PBS buffer.Moreover,the probe has high selectivity and anti-interference ability which has been successfully applied to the fluorescence imaging of endogenous and exogenous homocysteine in Hela cells.(2)To further improve the existing deficiencies of fluorescent probes for glutathione detection,we have developed a novel anthraquinone-based "off-on" fluorescent probe for GSH detection with high selectivity and sensitivity in PBS buffer,and the detection limit was as low as 89 nM.The specific fluorescence response mechanism of the probe to glutathione is focused in this chapter,and it is completely different from the mechanism of the traditional nitrobenzothiazole fluorophore on biothiols.A new highly sensitive and highly selective biological imaging method for detection of endogenous and exogenous GSH in human breast cancer cells has been successfully established.(3)In this chapter,aiming at the bottleneck problem of simultaneous detection of multi-target thiols,a novel versatile fluorescent probe based on the structure of indole has been designed and synthesized for simultaneous detection of Cys/Hcy and H2S in mild conditions.The probe exhibited fast response to Cys/Hcy and H2S within 10 min and 12 min respectively and the detection limit were 0.18 ?M and 0.15 ?M respectively.Furthermore,the probe has also been successfully applied to the fluorescence imaging of endogenous and exogenous Cys in human breast cancer cells.(4)Based on the previous work,the coumarin dye with higher fluorescence quantum efficiency was used as luminophore of the fluorescent probe.In this chapter,A tri-site versatile fluorescent probe has been designed and synthesized for simultaneous sensing of hydrogen sulfide and glutathione in the different buffer solutions.The probe could fully response to hydrogen sulfide and glutathione within 10 min,and the detection limit was 75 nM and 280 nM respectively.In addition,the probe has also been successfully applied to the fluorescence imaging of GSH and H2S in human breast cancer cells.(5)With the advantages of penetrating and low background,near infrared fluorescent probe is especially suitable for biological samples analysis and detection.Based on the photoinduced electron transfer strategy,a novel near-infrared fluorescent probe which used nile red dye as luminophore has been designed and synthesized for highly sensitive detection of hydrogen polysulfides in PBS buffer.The probe could rapidly response to polysulfide within 5 min and the detection limit was 3.9 nM.In addition,the probe has also been successfully applied to the fluorescence imaging of endogenous and exogenous polysulfide in human breast cancer cells.