Construction And Application Of Small Molecule Fluorescent Probes For Detecting Oxidative Stress Process

The redox equilibrium is important in preserving the correct functionality of cellular vital functions.When the imbalance between oxidation and antioxidant action in the body and the tendency to oxidation may cause oxidative damage to cellular components,which is called"oxidative stress".At present,there is a variety of evidence that oxidative stress processes are closely related to the development of various diseases,so it is essential to develop a simple,rapid,and objective measure of oxidative stress that is useful to both researchers and clinicians.Due to the characteristics of high sensitivity,high selectivity and non-invasive analysis,fluorescent probes show enormous potential in the diagnosis of diseases relate to oxidative stress processes and real-time analysis of related biomarkers.In this thesis,two novel multifunctional fluorescent probes were constructed to achieve high sensitivity detection of biomolecules during oxidative stress with minimal interference to biological systems.The main research contents are as follows:（1）Inflammation,a common disease triggered by oxidative stress,could produce reactive high levels of reactive oxygen species（ROS）in tissues with a wide p H range.In this work,a novel p H sensitive fluorescent probe,Hcy-OH,for ratiometric detection and imaging of ROSs was developed,exhibiting specific responses to different ROSs at varying p H values.Hcy-OH exhibited selectivity for peroxynitrite（ONOO^-）under acidic condition,hypochlorite（Cl O^-）under neutral condition and singlet oxygen（¹O₂）under alkaline condition,and was successfully applied to visualize exogenous and endogenous ROS in cells.In addition,this sensing scheme can be used to image inflammatory tissue,confirming its application for accurate monitoring of arthritis at organism level,which also means that Hcy-OH could serve as a powerful tool to reveal the relationship between the physiology and pathology of inflammation.（2）Cellular senescence is a complex physiological process triggered by a range of factors,and cellular damage caused by ROS generated by incomplete aerobic metabolism is one of the well-known triggers of senescence.Herein,a novel multifunctional fluorescent probe（SA-HCy-1）for the detection of senescence was constructed.The glycosidic bond of the probe was broken after the interaction withβ-galactosidase（β-gal）,and then undergone a spontaneous 1,6-elimination reaction of 4-hydroxybenzyl under medium or basic environments to release a fluorophore（SA-HCy-OH）with near infrared fluorescence signal.This"and"logical model design is consistent with the changes ofβ-gal accumulation and lysosome deacidification during aging,avoiding the false positive signal caused by a single biomarker.Subsequently,in the presence of ROS,-C=C-bond of the near-infrared fluorophore was destroyed to generate xanthene derivative,accompanied by the increasing of a new emission peak of 468 nm.The ROS content in aging process can be further detected by the change of the ratio of two-channel fluorescence after reaction with ROS.This design of programming multiple detection programs into a single sensing platform has greatly improved the diagnostic accuracy.SA-HCy-1 has been successfully used to track cell senescence and skin tissue senescence,and was expected to provide ideas for the efficacy evaluation of anti-aging drugs.