Design,Synthesis And Application Of Novel Near-infrared Fluorescent Probes For Detection Of Hydrogen Peroxide

As a typical reactive oxygen substance,hydrogen peroxide(H₂O₂)plays an important role in many biochemical processes in human body,such as cell migration,signal transduction and energy conversion.However,excessive accumulation of H₂O₂in human body may cause damages to proteins,lipids,nucleic acids and specific enzymes,leading to related neurodegenerative or cardiovascular diseases.Therefore,the accurate detection of H₂O₂ is of great significance to the elucidation of its biological effects and the investigation or prevention of related diseases.Routine methods for detecting H₂O₂ include chemiluminescence,chromatography and ultraviolet-visible absorption spectrophotometry.However,these methods are difficult to be applied to determination of H₂O₂ in vivo due to the complexity of chemical environment in organism and the limitations of derivative reagents and equipments.Compared with these methods,fluorescence spectroscopy exhibits the features of high sensitivity,high selectivity and low cost.Based on the modern synthetic technology,various types of organic fluorescent molecules and fluorescent nanomaterials with both high selectivity and good biocompatibility have been developed as probes for noninvasively sensing targets in organisms by turn-on,turn-off or ratiometric modes.In this work,two novel organic molecular fluorescent probes were designed and synthesized for highly sensitive and selective detection of H₂O₂ as well as fluorescent imaging of H₂O₂ distribution in cells.Molecular simulations and theoretical computations were performed to interpret the sensing mechanisms of the developed probes.Detailed research studies include the following parts:(1)An organic fluorescent probe HAA was developed for determination of H₂O₂.By using acetyl group as the recognition unit,this probe provides both colorimetric and fluorescence quenching response towards H₂O₂.Effect of intramolecular charge transfer(ICT)was proved to be the response mechanism by molecular simulation and theoretical computation.The near-infrared fluorescence emission(700 nm)and a large Stokes shift(150 nm)facilitate the application of this probe in clinical diagnosis.This probe exhibits high sensitivity for detecting H₂O₂,with a detection limit(LOD)of 0.85?mol/L.Common reactive oxygen species,metal ions or anions gave negligible interference on the response signals of H₂O₂.Fluorescent imaging of Hep G2 cells demonstrates the capacity of this probe for monitoring the distribution of both endogenous and exogenous H₂O₂ in cells.(2)A novel water-soluble near-infrared fluorescent probe EDHA was designed and synthesized.By respectively employing acetyl group and positively charged cyanine-like group as the recognition unit and the fluorophore,this probe possesses both mitochondrial targeting function and fluorescent turn-on response towards H₂O₂.Results of molecular simulation and theoretical computation proved that the enhanced ICT process in the product of the reaction between this probe and H₂O₂induced the response fluorescence signal.The features of near-infrared emission(640 nm)and good water solubility of this probe facilitate its application in bioanalysis.The LOD towards H₂O₂ in the PBS/Et OH(99:1,v/v)was 0.059?mol/L,and the interference induced by other reactive oxygen species,several types of metal ions or anions probably coexisting in real samples was negligible,indicating the superior detection performance and applicability of this probe.Performance of mitochondrial targeting and fluorescence imaging of this probe were evaluated by using A549 cells.Results demonstrated both the good mitochondrial targeting ability(Pearson's r=0.94)and the excellent fluorescent response towards H₂O₂ of this probe.These features were beneficial to the application of this probe in monitoring the distribution of endogenous and exogenous H₂O₂ in cells.