Design,Synthesis And Application Of Novel Fluorescent Probes For Nitroxyl And Peroxynitrite

Active nitrogen molecule?RNS?refers to the interaction between nitric oxide?NO?and compounds,including reactive oxygen species.A series of free radicals and nitro compounds with high oxidation activity including ONOO^-and its proton nitroxyl?HNO?were derived.Nitrogen oxygen free radical?HNO?is an acti ve nitrogen molecule?RNS?,produced by one electron reduction and protonation of nitric oxide?NO?,which is an important signal molecule in many physiological processes.Previous studies have shown that HNO has become a potential biomarker for the diagnosis and treatment of different diseases such as cardiovascular disease.Peroxynitrite anion?ONOO-?is a kind of reactive oxygen species?ROS?.ONOO-is produced by the interaction of nitric oxide?NO?and superoxide anion radical(.O^2-).ONOO-plays a very important role in signal transduction and antibacterial activity,which can damage proteins,lipids,many key biomolecules such as transition metalloenzyme centers and nucleic acids.Because of its unique and potentially important biological activity reports,RNS has received extensive attention,so it has developed a new type of high sensitivity.It is still of great significance to study the biological function of reactive nitrogen species by fast detection of the fluorescence probe of reactive nitrogen species.Nitroxyl?HNO?,as an electron reduced and protonated form of nitric oxide,is emerging as a potential diagnostic and therapeutic biomarker.It is still of great interest to develop probes of desirable properties to study its biological functions.Here we develop a near infrared fluorescence probe for detecting and visualizing exogenous and endogenous HNO in living cells.The probe is designed by coupling a HNO-responsive moiety,diphenylphosphinobenzoyl group,with a near infrared fluorophore with large of Stokes shift via an ester linker.The probe was initially nonfluorescent.HNO-catalysed oxidation reaction generates an aza-ylide,which intramolecularly attacks the carbonyl carbon,liberating the initial fluorophore with activated fluorescence signals.The probe is shown to exhibit high sensitivity,high selectivity and fast response towards HNO detection in vitro.Furthermore,imaging studies reveal that the probe is capable of detecting exogenous HNO with dose-dependent fluorescence signals.Its ability to image endogenous HNO without or with induction is also demonstrated in living cells.This turn-on fluorescence probe provides a useful tool for studying HNO in living cells.Pernitrite?ONOO^-?,as one of the most typical active nitrogen molecules,plays an important role in signal transduction through the immunogenicity of nitrotyrosine residues and invading disease resistance.This is due to the diffusion-controlled reaction of nitric oxide?NO?and superoxide anion radical(.O^2-)in the life system.Pernitrite has strong oxidation and nucleophilic properties,and can react with a variety of bioactive substances,such as protein,nucleic acid,lipid and so on.The imbalance of the production of ONOO^-can cause ischemia-reperfusion injury,Alzheimer's disease,autoimmunity,and so on,even cancer.However,the exact cellular mechanism of ONOO^-is still poorly known,largely due to the lack of appropriate methods to detect its distribution in living cells.Therefore,there is an urgent need to develop a method for in situ analysis of ONOO^-in biological systems.Pernitrite fluorescence probe was designed and synthesized.1,8-naphthalene dicarboxyimide,a typical intramolecular charge transfer?ICT?fluorescent group,was used to masking the chromophore to disable fluorescence.Pernitrite analytes were used to selectively conduct the response under a series of biological analysis conditions.The probe showed a 63-fold fluorescence enhancement and a detection limit of 30 nm.It was found that in probe BHI-DP,diphenylphosphonate blocked the ICT process,therefore,without ONOO^-,the probe did not show fluorescence and the fluorescence quantum yield decreased seriously.The selective cleavage of phosphite with ONOO^-resulted in the recovery of the ICT process,which resulted in an open fluorescence response.The selective detection of peroxynitrite with open response in hela cells was confirmed by confocal microscope imaging.This method can be used to design fluorescent probes for selective detection of ONOO^-based on the cleavage of phosphite groups.