| The pH value is the concentration index of hydrogen ions,which is used to analyze the pH of the solution.In the fields of chemical production,environmental governance,and chemical research and development,pH plays an important role.At the same time,in life,pH as a physiological parameter also plays a role that cannot be ignored;cells are the basic unit of life Each organelle maintains an appropriate pH value to perform different metabolic tasks.For example,the pH value of lysosomes is between 4.00 and 6.00,while the pH value of mitochondria is about 8.00.Apoptosis,which in turn causes Alzheimer's disease,cancer and other diseases.Therefore,real-time and accurate monitoring of the pH of organelles can provide an in-depth understanding of physiological and pathological processes.Fluorescence technology has significant advantages such as high sensitivity,no harm to cells,good biocompatibility,and simple operation.It has become a powerful tool for detecting the value of intact cells and subcellular organelles.In recent years,pH probes for different organelles have been reported successively.Many pH-responsive fluorescent probes have been reported,but some probes have complex structures and low yields;the penetrating ability of the probes is not strong enough to achieve deep tissue and living body imaging;the photobleaching of the probes may interfere with the cell imaging.Based on the above reasons,a new type of pH fluorescent probe was developed by targeting mitochondria and lysosomes,which successfully achieved fluorescence ratio imaging of mitochondria pH and fluorescence imaging of lysosome pH under stimulation of various apop to tic agents.The main research contents of this paper are as follows:1.A novel ratio-type fluorescent probe M-pH targeting mitochondria was designed and synthesized.The ratio type pH fluorescent probe based on ICT mechanism,the intensity ratio I584/I552 shows a good linear relationship with pH in the range of pH 6.00-7.80,and its pKa is 6.72.The lipophilic cation benzyl in the M-pH structure of the probe enables the probe to be targeted to the mitochondria of the cell.This performance is well demonstrated by colocalization experiments(R2=0.92).The probe's low cytotoxicity,high selectivity to H+in metal ions and amino acids,good stability and excellent reversibility,successful application in HeLa cells,Experimental data shows that the probe M-pH can monitor the pH change in living cells in real time.2.A novel two-photon fluorescent probe 1-ph targeting lysosomes was designed and synthesized.The 1,8-naphthimide group in the probe L-pH based on the PET mechanism serves as a two-photon fluorophore,piperazine serves as the reaction site,and benzyl chloride and morpholine target the lysosome.The fluorescence intensity and pH value show a good linear relationship in the range of 5.00-7.30.In SMMC-7721 cells,the co-staining experiment of L-pH and Lyso Tracker Green gives a co-localization coefficient of 0.91.The probe has good lysosomal specificity,indicating that L-pH can visually monitor lysosomal pH.At the same time,the probe L-pH was used to study the changes of lysosomal pH during apoptosis and mitochondrial autophagy,and revealed that the lysosomal pH increased during apoptosis and mitochondrial phagocytosis.Finally,the pH difference of mice was observed using L-pH.All these data indicate that L-pH can be put to practical use under various cellular stress conditions and in vivo monitoring of changes in lysosomal pH.3.Two-photon fluorescent probe DP-lys was designed and synthesized.Morpholine targets lysosomes in DP-lys probe based on PET mechanism,morpholine and N,N-dimethylethylenediamine as pH reaction sites,and 1,8-naphthalimide as fluorescence off recognition section.The probe DP-lys has a good linear relationship in the pH range of 7.00-9.40,has high selectivity to H+ in metal ions and amino acids,good stability and excellent reversibility.Experimental data show that the probe DP-lys shows a sensitive and reversible response to pH. |
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