Design, Synthesis And Biological Application Of Fluorescent Probes For Sensitive And Rapid Detection Of Ozone

Depression is a mental illness that seriously affects the health of people around the world,leading to high rates of mortality and disability.The core symptoms include disappointment,lack of pleasure?decreased ability to experience pleasure from natural rewards?,irritability,inattention,and abnormal sleep and appetite?"neurasthenia"?.The chronic and depravation nature of depression has a major impact on the global burden of disease and disability.Although the incidence of depression is very common,its pathophysiological knowledge is not mature enough compared to other common chronic and potentially fatal multifactorial diseases,such as type?diabetes.The outbreak of reactive oxygen species?ROS?in the body will lead to an imbalance of redox reactions,resulting in diabetes,cancer and neurological diseases.As one of reactive oxygen species?ROS?,O₃ plays a key role in mental disorders?depression,Alzheimer's disease?through oxidative stress.Due to the complex biological environment in the brain and the blood-brain barrier,probes for measuring O₃ in living brains have not been reported.And the exact relationship remains unclear between O₃ and depression.Therefore,designation of the fluorescent probes for detecting O₃ in the brains of mice with depression-like behaviors,real-time monitoring of changes in O₃ content in mice,and exploring its relationship with the development of depression,is of great significance for the early diagnosis of depression patients.To this end,we designed and synthesized near-infrared and two-photon fluorescent probes for O₃ detection.Such probes facilitated imaging on tissue and in vivo.And it can be used for in situ visualization of the brain of mice with depression phenotypes.In this paper,we carried out the following two sections of the work:1.In situ visualization of ozone in the brains of mice with depression phenotypes by a new near-infrared fluorescence probe.We synthesized a near-infrared fluorescent imaging probe?ACy7?for tracking O₃ in the mouse brain.The specific cycloaddition reaction between O₃ and the probe greatly promoted conjugation of the"pre-"heptamethine system,which resulted in near-infrared fluorescence.This made the probe specificity,high sensitivity and deep tissue penetration.Using ACy7,we found that under glutamate stimulation,the O₃ content in PC12 cells was significantly higher than that in control.More importantly,we revealed for the first time that the levels of O₃ in the brains of mice with depression phenotypes were significantly higher than the levels in control.We further proved that excess O₃ in the mouse brains led to generation of pro-inflammatory cytokine IL-8,which in turn induced depression.This work provided strong direct evidence of a positive correlation between O₃ and depression,and would help to elucidate the oxidative stress-associated pathogenesis of depression.2.Two-photon fluorescence imaging for rapid detection of ozone in lysosomes.A two-photon fluorescent probe?Mor-AB?targeted lysosome was designed and synthesized.Mor-AB overcame the problem of slow reaction rates of probes with O₃,and could quickly responded to O₃ without the addition of bases.The probe used 4-amino-1,8-naphthalic anhydride as a fluorophore,3-butenyl group as an O₃ recognition group,and morpholine as a lysosomal targeting group.It reacted with O₃ via the recognition group 3-butenyl and undergo a series of transition states.This reaction caused a break in the C-O bond,and then the carbon dioxide molecule is removed to expose the amino group of the fluorophore.It is satisfactory that the exposed amino group catalyzed the progress of the reaction and enhanced the fluorescent signal.In summary,the probe was responsive,localizable lysosomes,and had the advantage of two-photon fluorescence imaging.The probe Mor-AB was expected to achieve rapid detection of O₃ in living organisms,which would be of great significance for exploring the pathogenesis of O₃ related diseases.