The Synthesis Of An Acridone Derivative Fluorescence Probe And Its Imaging Application In Vitro And In Vivo

Acridone and its derivatives are a class of organic compounds with nitrogen heterocyclic,being of wide application prospects due to the intrinsic physiological and pharmacological activities,and excellent water solubility and photoelectrical properties improved by reasonable modification which can be combined with DNA,protein and other substances in the organism.Therefore,based on acridone nucleus,this article successfully synthesized a new acridone derivative（10-Methyl-2-Amino Acridone,MAA）for the imaging of cells,zebrafish and exosomes.This article includes two parts.Part one:a water-soluble acridone derivative MAA with excellent photoelectric properties was successfully synthesized by substitution and reduction reaction based on acridone acted as raw material.Its synthetic route mainly includes the following steps:Firstly,the hydrogen on the 10 position of acridone is easily deprotonated in the hot ethanol potassium solution,and then nucleophilic substitution reaction between deprotonated acridone and CH₃I solution occurs in N,N-Dimethylformamide（DMF）solution to modify the methyl group to form 10-methyl acridone.Secondly,10-methyl acridone is nitrified in the mixture solution of concentrated nitric acid to form 10-methyl-2-nitro acridone.Lastly,The reduction reaction between 10-methyl-2-nitro acridone and Na₂S to obtain 10-methyl-2-amino acridone（MAA）.Afterwards,the structure of the synthesized MAA was characterized by infrared spectroscopy,mass spectrum,carbon spectrum and hydrogen spectrum and further studied its optical and electrochemical properties.The study shown that:compared with the acridone,the water-soluble of MAA was significantly enhanced,the Stokes shift was increased,and showed green fluorescence under UV lamp.MAA has stable fluorescence performance with the fluorescence quantum yield of 49%.Notably,MAA has a outstanding two-photon activity under near infrared excitation.At the same time,MAA has obvious electrochemical activity on glassy carbon electrode and other substrate electrodes,such as gold electrode and Indium Tin Oxide（ITO）electrode,and interaction with salmon sperm DNA.In addition,MAA has visible light induced oxidase activity and can oxidize TMB（3,3’,5,5’-Tetramethylbenzidine,TMB）,resulting in color change of its solution.Therefore,MAA is expected to be used to build a photoelectric and visual biosensor device for biomedical applications.Part two:MAA serves as a one and two photon fluorescent probe for the imaging of cells,zebrafish and exosomes based on the first part of the study.The experimental results suggest that MAA has low cytotoxicity,excellent cellular permeability and biocompatibility,and can enter into the nucleus and cytoplasm at the same time.MAA can enter into the body of zebrafish by swallowing and skin absorption,mainly accumulated at the head,the abdominal cavity and tail,and can be observed the green fluorescence under the one and two-photon microscope.In addition,MAA is easy to be protonated and can be electrostatic adsorption with the negatively charged exosomes under the acid condition,leading to the MAA-labeled exosomes,realizing the imaging of the exosomes in the HepG2 cells.Therefore,MAA,as a two-photon fluorescent probe,has potential application value in the field of biological imaging.