Synthesis And Application Of PH-activatable Near-infrared Indocyanine Fluorescent Probes

As H~+ regulates a number of cell metabolic processes such as cell proliferation and apoptosis,ion transportation and neural signaling conduction etc..Abnormal intracellular pH(pHi)usually causes cell disfunction and some severe diseases.Thus,sensing p Hi plays an important role in investigating physiological and pathological cell processes.Compared with other methods,fluorescence imaging has several advantages like high sensitivity,non-invasiveness,simplicity etc..Based on these,we designed and synthesized a series of pH-activated fluorescence probes for intracellular pH recognition and detection,and then screened out the probes with suitable p H response range to tumor microenvironment and further researched on their application.We firstly constructed two dissymmetry cyanine probes 5 and 6,and investigated their absorption and fluorescence spectrum on different pH.It was found that the absorption and emission peaks are both in near-infrared region in weakly acid condition,which has advantage of avoiding optical absorption and autofluorescence interference from biological tissues.Moreover,probes 5 and 6 have high pH-sensitivity,and can make rapid response to pH fluctuation.By introducing hyaluronic acid and quaternary ammonium cations(with targeting tumor mitochondria),two kinds of tumor-targeted and specificity imaging probes were constructed by modification of the pH-activated near-infrared reversible fluorescence probe 6.The pKa values of probes 9 and 10 modified by quaternary ammonium were just within the pH range of tumor acid organelles,and their fluorescence signal both achieved reversible“off”and“on”.Probes 9 and 10 had good photostablity and high selectivity.Verified by ~1H NMR spectra,pH-sensitive spectral response was possibly attributed to the protonation and deprotonation of the indole nitrogen atoms.The ICT process was enhanced upon the protonation of the non-alkylated nitrogen atoms,which leaded to a bathochromic shift of absorption maxima and remarkable enhancement of the fluorescence intensity.The fluorescence imaging result demonstrated that probes 9and 10 had low cytotoxicity and good cell-membrane permeability,and both probes could be used for imaging pH fluctuations under acidic condition in living cells.