Development Of Lysosome-targeting Formaldehyde Fluorescent Donors And Highly Selective Fluorescent Probes For Glutathione

This thesis involves three different projects.The first project aims to develope lysosome-targeting formaldehyde fluorescent donors.Formaldehyde(FA)is a kind of reactive carbonyl species and cellular metabolites in biological system involved in growth and pathological processes.Development of small molecule tools capable of studying dose-effect of FA in biological system is of great significance for understanding its physiological and pathological roles.Two lysosome-targeting FA fluorescent donors,lyso-FAD-1 and lyso-FAD-2,were synthesized.Both can release FA with concurrent fluorescent turn-on response for tracking and quantification of FA release.In vitro FA release studies show that their FA release behaviors are modulable by pH,FA concentrations in the environment,and ring-strain in their structures,thus providing new research tools for FA dose-effect releationship study in lysosomes.We also verified the lysosome-targeted FA release and fluorescence generation of the donor lyso-FAD-1 in HeLa cells through fluorescence colocalization imaging studies with Lyso-Tracker Red.The second part involves development of analyte regeneration fluorescent probes for FA with increased emission wavelength.A new probe 3-1 was designed and synthesized,using boron-dipyrromethene(BODIPY)as the fluorophore.Unfortunately,probe 3-1 does not show the expected fluorescence response for FA,after reaction with FA,likely due to inefficient intramolecular charge transfer(ICT)and photo-induced electron transfer(PeT)fluorescence quenching of the probe.The third part refers to the development of glutathione(GSH)fluorescent probe with high selectivity.Malemide-based fluorescent probes generally cannot differentiate specific biothiols.However,we found that the BODIPY-malemide intermediate 3-8 in the synthesis of the probe 3-1 can detect GSH with fluorescence turn-on response,while almost no fluorescence turn-on response was identified for cysteine(Cys)and homocysteine(Hcy).We believe that the probe undergoes Michael addition-cyclization reaction with Cys/Hcy and form products with acceptor photo-induced electron transfer(a-PeT)fluorescence quenching effect,while the probe reacts with GSH and only forms Michael adducts with fluorescence turn-on response due to the removal of donor photo-induced electron transfer(d-PeT)fluorescence quenching effect of the maleimide.