Design,Synthesis And Biological Application Of Fluorescent Probes For Detecting Active Sulfur And Reactive Oxygen Species

Sulfur dioxide（SO₂）,homocysteine（Hcy）,cysteine（Cys）and glutathione（GSH）,as the four kinds of biothiols with the most abundant contents,play important role in the organism.Superoxide anion(O₂^·-),peroxynitrite（ONOO^-）,as the two most important reactive oxygen species,are closely related to many diseases.Fluorescent probe is a new detection method,which has attracted more and more attention in recent years.Due to the advantages of high detection sensitivity,non-invasive,convenient and high spatial-temporal resolution,etc.,it is widely used for real-time detection of many active small molecules in living organisms,and can achieve the purpose of diagnosing a variety of diseases.Therefore,it is of great significance to develop fluorescent probe that can realize the real-time discrimination and detection of active molecules of biological systems(such as O₂^·-,ONOO^-,Cys,Hcy,GSH,SO₂)in the fields of chemistry,biology,and medicine.The main conclusions of this paper are shown as follows:1.A multifunctional fluorescent probe（Probe 1）was synthesized,which can detect Cys/Hcy,GSH and SO₂ in living cells and zebrafish with high selectivity.The probe used benzoindole salt as the fluorophore,NBD-Cl as the specific recognition group for Cys/Hcy and GSH,the C=C bond as the reaction site of SO₂.The fluorescent probe has a highly specific reaction to Cys/Hcy,GSH and SO₂.After the reaction,the product is easy to dissolve in water and has good photostability.It has many advantages,such as good performance,large Stokes shift and wide linear range.It has been successfully applied to fluorescence imaging analysis of endogenous and exogenous Cys/Hcy,GSH and SO₂ in living cells and zebrafish.2.Two new fluorescent probes,probe 2-1 and probe 2-2 were designed and synthesized,which can detect O₂^·-with large Stokes shift.The Stoke shift can be increased by combining 6-hydroxy-2-naphthaldehyde with 4-pyridineacetonitrile and 2-（pyrimidin-4-yl）acetonitrile respectively.At the same time,the recognition site of O₂^·-was introduced into the phenolic hydroxyl group of the fluorophore,to construct two fluorescent probes,which can be used to detect O₂^·-.Through comparison and analysis,the best probe 2-2 is selected.3.A novel ratio fluorescent probe with good photostability（Probe 3）was synthesized,which can be used to image O₂^·-and ONOO^-in cells.The fluorescent probe uses the ether bond as the specific recognition site of O₂^·-,and C=C as the recognition site of ONOO^-,oxidation reaction is based on O₂^·-attacking the ether bond and ONOO^-attacking C=C to destroy the total conjugated system to distinguish and detect O₂^·-and ONOO^-at the same time.The fluorescent product have large Stokes shift(Δλ_O2·^-=155 nm,Δλ^-_ONOO=100 nm)and obvious blue and red light emission characteristics,respectively,which have been successfully used for the fluorescence imaging analysis of O₂^·-and ONOO^-in living cells.