Design And Synthesis New Types Of Fluorescence Probes For Intracellular Anions (chloride, Hypochlorite Anion) And Their Application In Vivo

There were various micro or trace amounts of active species (free radicals, anions and cations) in living systems. They hold a unique position in life processes and are of central importance for many physiological functions. These active species are important mediators for the pathological conditions of various diseases, such as carcinogenesis, inflammation, ischemia-reperfusion injury, and signal transduction. In order to further explore the close relation between these small biological molecules and various physiological or pathological processes, there is an exigent need for researchers to develop fast-respond probes to realize the accurate detection of these important species. However, as a result of the low concentrations of these active species, and the diversities and complexity of the biological environment, methods that can detect the bio-relative species with high selectivity are few. Take full advantage of benefits provided by confocal laser scanning microscopy (CLSM), spectrofluorimetry can serve as a powerful method due to its high sensitivity, simplicity in data collection, and high spatial resolution in imaging techniques. They are considered as the unique method that can realize the"real-time, dynamic, visualized"analysis of fluctuations of various bio-relative species in cells and in tissue. In a word, it is of great value for the researchers to explore fast and convenient probes to investigate the levels of the trace molecules in bio-system, and furthermore realize the real-time imaging in living cells.As one important element, chlorine distributes widely in cells in forms of chloride (Cl-). It holds a unique position in a wide range of many cellular functions, including regulation of cells volume and pH, apoptosis and immune response, intracellular trafficand enzyme catalysis reactions. Exceptional fluctuation in concentration of Cl- can result in some diseases, such as cystic fibrosis, myotonia, arrhythmia and myocardial ischemia. Thus, there is exigent challenge for researchers to develop fast-respond probes to trap Cl- for investigation of the mechanisms of the physiological and pathological processes that the chloride anions participate.As one of the biologically important ROS, which is produced in living organisms from hydrogen peroxide and chloride ions in a chemical reaction catalyzed by the enzyme myeloperoxidase (MPO) in activated neurophils, hypochlorite anion ( OCl- ) is known to be essential to several biological functions , for example, because OCl- can behave as a strong nucleophilic nonradical oxidant, it is regarded as an important agent with strong antibacterial properties that is used for natural defense; its efficacy lies in the fact that neither bacteria nor mammalian cells can neutralize its toxic effects-they lack the enzymes required for its catalytic detoxification. However, excessive or misplacedprod uction of OCl- can lead to tissue damage and diseases, such as atherosclerosis, arthritis, and cancers. Therefore, more and more attention has been paid to the studies of the biological functions.All in all, based on the crucial function of various anions during the physiological and pathological processes, researches about recognition and detection of the bio-relative anions have attracted drastic attention. Up to now, considering the external influence factors that will disturb the recognition process between the anion and the acceptor, it is of vital importance to develop and synthesis probes that are exclusive to specific anions.Considering the development status of anions and the excellent rate, sensitivity and selectivity, spectrofluorimetry combined with CLSM was applied successfully. It realized the application both in vivo and in clinic. The following are the main content of dissertation:First, we chose the low-toxic 5-Aminofluorescein as the chloride insensitive group. Simultaneously, 6-Methoxyquinoline was chosen as the chloride reactive group, whose fluorescence was quenched linearly with the increased chloride concentration. It was synthesized conveniently by only two steps. Structure of the MQAF was characterized by 1H NMR and 13C NMR, and fluorescence properties of the reaction between MQAF and Cl- were optimized in chemical system. It showed that the probe possessed potent selectivity to Cl- and could respond instantaneously to changes of chloride concentration. During experiments, MQAF was proved to exhibit potent selectivity toward Cl- and respond to changes in chloride concentration instantaneously. Owing to the outstanding performance of the MQAF presented here, we successfully applied it to monitor chloride concentration fluctuations between normal and ischemic conditions.Second, in this paper, we design a NIR OCl- fluorescent probe based on tricarbocyanine (Cy), whose maximum emission wavelength is within near-infrared region and choosing hydroxylamine hydrochloride as the reactive group. After addition of the OCl- anion, there undergoes an obvious fluorescence enhancement. Experiment results showed that the probe exhibited wide excitation and emission spectrum, besides, it could detect the hypochlorite anion sensitively and selectively in the near-infrared region.