Design And Syntheses Of Coumarin Derivatives-based Fluorescent Probes And Their Applications In Biological Imaging

Fluorescent probes have many advantages such as high sensitivity and selectivity,high spatiotemporal resolution,fast response,simple operation.Fluorescent probes have attracted many interests and show great potential in the field of environment science,medicine,and biology.The large Stokes shift is an ideal property of fluorescent probes,which can greatly reduce self-absorption and avoid the interference of incident light.More importantly,a larger Stokes shift can effectively minimize the background fluorescence of biological samples.Currently,there are only few reports on fluorescent probes with large Stokes shift,so it is urgent to develop a novel strategy to increase the Stokes shift of known fluorophores.Lipid droplets(LDs)are ubiquitous lipid-rich organelles that found in fungi,plants,and animals.LDs play a vital role in a variety of biological processes.However,abnormal LDs can lead to many human diseases.Staphylococcus aureus(S.aureus)is a common pathogen.S.aureus infection can cause a variety of diseases,and may also bring a significant burden to healthcare-related expenditures.In this paper,two types of coumarin derivatives with large Stokes shifts were developed for the imaging of lipid droplets and S.aureus.The specific research content is as follows:1.Based on the 4-position ester substitution strategy,a LDs targeting fluorescent probe 1 was designed through simply introducing the 4-substituted ester group into the classic coumarin fluorophore.Probe 1 possess a larger Stokes shift due to the steric hindrance and resonance effect of the 4-substituted ester group.The sensing properties of probe 1 were investigated by ultraviolet-visible absorption spectroscopy and fluorescence emission spectroscopy.Probe 1 showed a large Stokes shift(118-184 nm)in both weak polarity and polarity solvents,good biocompatibility,and excellent anti-bleaching performance.Probe 1 has been successfully used to specifically label the morphology and distribution of lipid droplets in living cells.Moreover,the probe could be employed in the imaging of tumor tissues in vitro and in vivo.In order to verify the design principle,the comparative compound 2 with 3-substituted ester group was designed and constructed.This comparative compound showed no obvious solvatochromic effect,small Stokes shift(50～80 nm),which limit its real application in bioimaging.Confocal imaging experiments show that the comparative compound did not stain LDs.These experiments showed that by changing the position of the functional group,the biological function of the fluorescent probe is greatly changed.This research was expected to promote the development of large Stokes shift fluorescent probes and their applications in the field of biomedicine.2.By employing the 4-carboxy substituted strategy,a coumarin based fluorescent probe 2 for the detection of Staphylococcus aureus with large Stokes shift was developed.Confocal imaging showed that the fluorescence of S.aureus incubated with probe 2 was significantly increased.By contrast,after incubated with probe 2,Gram-positive bacteria(B.subtilis,E.Faecium,and E.facecalis)and Gram-negative bacteria(Escherichia coli,and S.Maltophilia),as well as mammalian cells showed no obvious fluorescence changes.In addition,the probe does not recognize inactivated S.aureus.This research provides new ideas and methods for the detection of live S.aureus,which can be used for the detection of S.aureus in various fields such as medical and health,production,and life.