Design,Synthesis And Bioimaging Applications Of Two-Photon Viscosity Fluorescent Probe

Intracellular viscosity plays a critical role in biological processes,such as the diffusion of proteins and other biomolecules across cell membranes.Abnormal changes in the local viscosity of cell may lead to abnormal cell function or serious diseases.Therefore,it is urgent to develop a powerful microviscosity measurement tool.In recent years,fluorescence technology has become a powerful tool for detecting microenvironmental viscosity changes in living organisms due to its excellent properties including high sensitivity,non-invasive detection,high selectivity and real-time monitoring.Moreover,two-photon fluorescence technology using near-infrared photon excitation has become an indispensable imaging tool in biomedical research.In this paper,based on the research status of two-photon viscosity fluorescent probes,three two-photon viscosity fluorescent probes based on carbazolebenzaldehyde structure,carbazole-thiazole structure and coumarin-indoles structure were designed and synthesized respectively and their biological applications were further studied.In chapter 2,the two-photon viscosity fluorescent probe CBA was designed and synthesized by using carbazole derivatives as fluorophore.Under the excitation of 365 nm light,CBA tend to form TICT excited state in non-viscous media,and the entire molecule is in a distorted state and release internal energy through a non-radiative transition,thereby emitting weak fluorescence.However,the fluorescence gradually increases with the increase of solution viscosity.Cell imaging experiments showed that CBA exhibited excellent membrane permeability and high co-location coefficient with Nile Red,indicating that the probe CBA could be utilized for imaging cellular lipid drops viscosity.In chapter 3,based on the design of carbazole-thiazole structure,a two-photon viscosity fluorescent probe CB-V was synthesized,which was used to detect the abnormal changes of microenvironmental viscosity in vivo.Spectral test experiments show that in viscous glycerol,CB-V emits strong fluorescence at 602 nm,and the fluorescence intensity is increased by 9.6-fold showing a well viscosity response.In addition,the probe is hardly affected by pH and has good selectivity and light stability.More importantly,CB-V is applied to cell and in vivo imaging,indicating good biocompatibility and capable of two-photon imaging viscosity changes of microenvironments in organisms.In chapter 4,a two-photon fluorescence probe Mito-V with coumarin as fluorophore and indole salt as mitochondrial targeting group was synthesized based on the structure design of coumarin-indoles.As the viscosity increased from 1.7 cP to 953 cP,the fluorescence emission of probe Mito-V at 634 nm was enhanced by about 22 times and showing a good viscosity response.In addition,the probe also exhibited the properties of good selectivity,strong mitochondrial localization ability and little biological toxicity.Biological imaging experiments show that the probe Mito-V can conduct two-photon imaging of living cells and zebrafish stimulated by exogenous stimuli with high resolution both the one-photon and two-photon imaging mode.More importantly,the probe has been successfully used for imaging viscosity in live mice with better results.In addition,inflamed mice demonstrated that the fluorescence intensity was about 2.1-fold higher than for the normal mice.