| Hypochlorous acid(HOCl),an important reactive oxygen species(ROS)in living organisms,has emerged as a critical signaling molecule for both physiological and pathological processes.Endogenous HOC1 is mainly produced by the enzyme myeloperoxidase(MPO)-catalyzed peroxidation of chloride ions in leukocytes,which is closely linked to the immune defense systems.Under the physiological environments,HOC1 is highly reactive and short-lived.However,an excess amount of HOCl can lead to tissue damage and diseases,such as atherosclerosis,osteoarthritis,rheumatoid arthritis and cancers.Therefore,it is crucial to investigate the distribution and functions of HOC1 for biological research and clinical diagnoses.Mitochondria,the organelle found in almost all eukaryotic cells,plays a vital role in the life and death of cells.The most prominent function of mitochondria is to produce ATP,the energy currency of the cell.Mitochondrial dysfunction is extensively involved in many types of human diseases including neurodegenerative and cardiovascular diseases,and thus prompts research on mitochondria-specific diagnoses and therapies.Consequently,development of efficient methods for monitoring mitochondrial morphology as well as membrane potential(??m)is of great importance for both biomedical research and early diagnosis of the related diseases.This work mainly covers the following two parts:1.We designed an ICT-based BD derivative fuorophore,7-(dimethylamino)b enzo-[c]-[1,2,5]-oxadiazole-4-carbaldehyde(DBDC),as the fuorescence reporting group.The HPBD probe for HOC1 was prepared via a simple one-step reaction:DBDC reacted with 2-mercaptoethanol in dry dichloromethane in the presence of a right amount of methanesulfonic acid to synthesize HPBD.Several function al groups,such as oxime and diaminomaleonitrile,have also been identifed as HOCl-reactive moieties.For comparison,hydroxylamine and diaminomaleonitrile were also synthesized(HPBD-1 and HPBD-2).The synthetic intermediates and HOCl probes were characterized by 1H NMR,13C NMR and high resolution ma ss spectrometry(HRMS).By comparsion,HPBD was chosen as the probe for H OC1 detection and its basic photophysical properties were investigated.Live cell s imaging studies demonstrate that the probe can be applied successfully to dete ct exogenous and endogenous HOCl.2.We designed an ICT-based BD derivative fuorophore,7-(dimethylamino)b enzo-[c]-[1,2,5]-oxadiazole-4-carbaldehyde(DBDC),as the fuorescence reporting group.Three fluorescent dyes Mitol,Mito2,Mito3 were abtained by reacting w ith three different cationic groups with strong electron withdrawing ability.The introduction of the electron withdrawing groups in fluorescent dyes not only ext ends the emission wavelength,but also has the ability to locate the mitochondri a.Through a series of in vitro experiments,we screened out the mitochondrial fluorescence probe Mito3 with the smallest external factors,stable optical proper ties,low toxicity and good membrane permeability.Then the probe directly appl ied to cell imaging experiments without washing.Fluorescence background is ve ry low.The staining of Mito3 in MCF-7 cells treated with a membrane potentia 1 uncoupler carbonyl cyanide m-chlorophenylhydrazone(CCCP)was examined.T he staining of Mito-3 was the same in the absence or presence of CCCP,sugge sting that Mito-3 was independent of the mitochondrial membrane potential. |
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