Design,Synthesis And Applications Of Mitochondrial-target Viscosity Fluorescence Probes

Abnormalities of mitochondrial viscosity are associated with many diseases and cell dysfunctions,such as cell malignancy,atherosclerosis,diabetes,and Alzheimer's disease.Therefore,it is of great significance to accurately measure the changes of local viscosity in the mitochondria of living cells.Compared with other analytical techniques,fluorescence imaging has become a powerful tool for detecting biological molecules and biological parameters in living systems due to its high sensitivity,non-invasive detection,high selectivity,and real-time imaging.Based on the research status of mitochondrial targeted viscosity fluorescent probes,three types of mitochondrial targeted-viscosity fluorescent probes were successfully designed and synthesized,include enhanced,ratio and double detection,and their spectral properties and biological applications were further studied.Firstly,two novel mitochondrial-targeted fluorescent probes MHC-V1 and MHC-V2were designed and synthesized using hemicyanine and carbazole as acceptors and donors,respectively.Because the probe molecule is easy to form the TICT excitation state when excited by light,the self-fluorescence of the probe is weak,and the fluorescence intensity was gradually enhanced with increasing the solution viscosity.Cell experiments showed that the probes were membrane permeable and had a high co-localization coefficient with Mitochondrial-Tracker,and suitable for the fluorescence imaging of mitochondrial viscosity in living cells.Secondly,through two ways to hinder the process of twisted intramolecular charge transfer,the first dual-detection fluorescent probe Mito-VH,which can image the viscosity and hydrogen peroxide in mitochondria,was designed and studied.Due to the presence of the excited state of TICT,the probe itself has almost no fluorescence.As the viscosity of the solution increased,the fluorescence of the probe at 607 nm was gradually increases.In addition,the probe can recognize hydrogen peroxide in the green channel.With the addition of H₂O₂,the fluorescence at 510 nm gradually increases.Cell experiments showed that the probe Mito-VH can be used for dual-channel real-time detection of viscosity and H₂O₂ in mitochondria with different fluorescence signals in living.Thirdly,by combining two fluorophores with different viscosity sensitivities,a novel mitochondrial-targeted two-photon fluorescence viscosity probe TM-V was designed and synthesized.The probe shows a good linear relationship between the fluorescence intensity ratio of the two fluorophores and the viscosity of the medium,enabling the quantitatively determination and imaging of the mitochondrial viscosity in living cells.The probe has been successfully used for two-photon fluorescence imaging of cell and tissue viscosities.It has been demonstrated that the excessive generation of ROS increased the viscosity of mitochondria by single-photon and two-photon fluorescence imaging.