Design, Synthesis And Application Of Fluorescent Probes Based On Thioketone Structure

Reactive oxygen species(ROS),produced endogenously from oxygen molecule mainly through the mitochondrial respiration process,could participate in a wide range of physiological and pathological processes.ROS generated in normal cellular environments are essential for life.Moreover,they could regulate the immune system to resist the invasion of pathogens.Overproduced ROS could cause oxidative stress through the oxidation of biomolecules,such as proteins,lipids and DNA.In addition,excessive ROS could also lead to various diseases,such as inflammation,angiocardiopathy,neurodegenerative injury and carcinogenesis,etc.As one of the most important ROS,Hypochlorous acid(HClO)is generated during an oxidation reaction between chloride ions and H2O2 that is catalyzed by myeloperoxidase(MPO)secreted by activated phagocytes in the inflammatory state.HClO plays a vital role in biological processes.On one hand,HClO regulates cytokines and biological factors to promote healing and inhibit inflammatory in the biological immune system.On the other hand,it can also promote the ability of activated phagocytes to kill a variety of pathogens.However,excessive HClO could cause cells lysis,tissue damage and diverse diseases such as hepatic ischemia reperfusion injury,lung injury,arthritis,neuron degeneration and cancers,etc.Peroxynitrite(ONOO-)is generated from the reaction between nitric oxide(NO)and superoxide(O2·-),which is a potent biological oxidant involved in a wide range of physiological and pathological processes.ONOO-can affect tyrosine nitration and oxidation of sulfhydryls,nucleic acids and unsaturated fatty-acid-containing phospholipids,eventually resulting in mitochondrial dysfunction and cell death.Therefore,ONOO-has been related closely to varied diseases,including ischemic stoke,sepsis,diabetes and chronic heart failure,etc.In addition,ONOO-can resist to invading pathogens to regulate the body immune function through the cytotoxic effect.In addition,ONOO-could also participate in the processes of siganal transduction and normal cell apoptosis.In this thesis,utilizing a Lawesson’s reagent that can efficiently convert C=O double bond into C=S double bond,thioketone-containing two-photon fluorescence probes(TPFPs)were designed and syntheized.The probes have been used to fluorescently image HClO and ONOO-in live cells with high specificity and sensitivity.The particulars are shown below:(1)In this section,two ratiometric TPFPs basd on coumarin(HClO-Ratio-1)and xanthone(HClO-Ratio-2)have been constructed.Both probes respond to HClO with high specificity,sensitivity,and good rapidity because of the reactive moiety thioketone.The up-regulation of endogenous HClO level in RAW264.7 cells under PMA stimulation was imaged by HClO-Ratio-1 and HClO-Ratio-2.Based on this,the HClO level in normal mice and hyperglycemic mice was obtained by HClO-Ratio-2 with two-photon fluorescence mode and the results were compared.The results indicated an apparent up-regualtion of HClO in the hyperglycemic mice,giving a direct and visual evidence of positive correlation between HClO level and hyperglycemia degree.(2)In this section,a fluorescence probe(MTCA)with double reacting moieties,thioketone and a-ketoamide,has been fabricated.This two-reacting-moiety character enables MTCA to respond to ONOO-and HClO successively and specificly.In each reaction step,the fluorescence of MTCA increases apparently and quickly.It is important that the decomposition of α-ketoamide by ONOO-initiates the subsequent recognition towards HCIO,creating the conditons for simultaneous detetion of ONOO-and HClO.The results provide an efficient analysis method for understanding the biological relation between ONOO-and HClO.