Preparation Of A Ratiometric Near-infrared Fluorescent Molecular Rotor And Its Application In Detecting Intracellular Viscosity And Protein Denaturation

In biological systems,changes in intracellular viscosity are closely related to some disease,and abnormalities in viscosity may lead to diabetes,atherosclerosis,and malignancy.The detection of viscosity is particularly important.The mechanism by which a molecular rotor fluorescent probe detects intracellular viscosity is that since the rotation of the molecular rotor is an effective quenching pathway in a non-viscous medium,the fluorescent compound coupled to the rotor portion causes weaker intrinsic fluorescence.The molecular rotor rotation resistance mechanism is suitable for the detection of denatured proteins.Many diseases(such as Alzheimer's disease and Parkinson's disease,type II diabetes,etc.)are produced by the transformation of protein structure,so the research progress on protein denaturation detection are important.The fluorescent probe designed in this paper is a ratiometric near-infrared molecular rotor based on heptamethine cyanine which can detect cell viscosity and protein denaturation.In the first part of our work,we prepared this ratiometric near-infrared fluorescent molecular rotor.We use heptamethine as a fluorophore,and introduce a six-membered ring on the conjugated chain of heptamethine to increase its photostability.The six-membered ring is replaced by a chlorine atom.An active site is added,and the ends of the heptamethyl phthalocyanine conjugated chain are respectively blocked with benzofluorene and hydrazine to enhance their stability.The electrons on the nitrogen atom in the crucible attack the chlorine atom on the six-membered ring and undergo further ring-closing reactions to produce the final product Mu1 molecular rotor.The molecular rotor uses benzopyrene as a strong electron acceptor group,and a new spiropyrrole-quinoline derivative is used as an electron donor group to design a rotor having a "D-?-A" molecular configuration.A molecular rotor is prepared by a one-step ring-closing reaction,which imparts viscosity responsiveness and detects protein denaturation.The second part of our work is the characterization of the near-infrared ratio fluorescent molecular rotor in detecting intracellular viscosity.Firstly,the optical properties in different viscosity solutions were tested by UV absorption spectroscopy,fluorescence emission spectroscopy and fluorescence lifetime spectroscopy.Secondly,through simulation calculation,it is demonstrated that the viscosity sensitivity of the molecular rotor is generated by rotation to reduce the energy barrier.Finally,the intracellular viscosity quantitative test was performed.The change in molecular rotor fluorescence was monitored by adding nystatin to the cells to change intracellular viscosity.The third part of our work is the characterization of the near-infrared ratio fluorescent molecular rotor in detecting protein denaturation.The optical properties of denatured/nondenatured insulin and bovine serum albumin solutions were tested by UV absorption spectroscopy and fluorescence emission spectroscopy.