Development Of Two Novel Fluorescent Probes For Peroxynitrite And Their Application

Peroxynitrite（ONOO-） is one of the endogenous reactive oxygen species（ROS）,which plays a vital role in physiology and pathology.However,the precise pathogenic roles played by this substance in biological systems have not yet been elucidated completely owing to its short lifetime,high reactivity and low concentration.Thus,the development of novel techniques for detecting ONOO-,with excellent biocompatibility,sensitivities,and site-specificities,is the hotspots and difficulties of current research fields.Fluorescent probe,an efficient molecular tool for both analytical sensing and optical imaging,has attracted a great deal of attention due to the superiority of high sensitivity,nondestructive detection and well spatial resolution for imaging.In this thesis,two novel fluorescent probes were designed,synthesized and successfully employed in the detection of the exogenous and endogenous ONOO^-in living cells.（1）By virtue of fluorescein and heptamethine cyanine（Cy7）as started materials,two compounds Fd-P and Cy-P were rationally designed,synthesized and verified by MS,¹H NMR and ¹³C NMR.（2）By introducing the indoline-2,3-dione moiety as the recognition site,we proposed a novel fluorescence probe Fd-P with highly specific response toward ONOO^-.Optical experiments show that the linear response range of probe Fd-P to ONOO^-is 0～10?M,and the detection limit is 26 n M.Probe Fd-P shows specific recognition performance for ONOO^-and does not respond to other ROS/RNS,which allows the probe Fd-P to accurately detect ONOO^-both in buffer solutions and in vivo.The probe Fd-P has low cytotoxicity and has been successfully used to image the varied concentration of exogenous ONOO^-in living cells.And in the stress response to external stimuli,Fd-P can be used to track the endogenous production of ONOO^-in living cells.（3）The fluorescent probe Cy-P is utilized for the detection of exogenous and endogenous ONOO^-in living cells.Due to the strong oxidizing ability of ONOO^-,the?-conjugated system of Cy-P was destroyed and formed a new substance with blue fluorescence,leading to a 268 nm blueshift of fluorescent emission.The linear detection range of the probe Cy-P to ONOO^-is 0～15?M,the detection limit is 13 n M.The probe exhibits superior selectivity for ONOO^-over various potentially interfering biological analytes.The probe has low cytotoxicity and has been successfully used for imaging the exogenous and endogenous ONOO^-in living cells.