Site-specific Spontaneous Fluorescence Modification Of HSA And Bioimaging

Human serum albumin(HSA)is the most abundant protein in blood plasma,its numerous structural domains can be widely combined with a variety of endogenous and exogenous substances to store and transport them to various parts of the body.Among them,albumin,as the carrier of drugs,affects the distribution and efficacy of drugs in vivo,and plays a key role in the pharmacokinetics and delivery of drugs.In addition,many organic small molecule fluorescent dyes can also combine with albumin,and their photophysical properties and biomarker properties can be effectively improved.This thesis focuses on the construction of high-performance living labeling fluorescent dyes and their innovative imaging labeling applications.A series of fluorescent dyes were synthesized with coumarin and cyanine as the matrix respectively,and the performance of organic small molecule fluorescent dyes was improved though the fixed-point and spontaneous combination of HSA with them,realizing the in situ imaging analysis of blood drug concentration in vivo and the in situ imaging of hydrogen peroxide level in vivo.The specific research contents are as follows:1.For in situ drug monitoring in vivo,we synthesized fluorescent dye 2-1 based on quinoxaline coumarin.The spontaneous and covalent combination of Lysine at 161 position in HSA with the carbonyl group at 2-1 formed imines,thus constructing a fluorescent albumin with drug response properties.Similar to HSA itself,2-1 labeled HSA can also interact with ibuprofen,warfarin,clopidogrel and camptothecin as a coupled fluorescent probe.These drugs induce a significant enhancement of the fluorescence signal of the probe system in a concentration-dependent manner.Therefore,based on the coupled fluorescence probe 2-1-HSA,we can monitor these invisible drugs in real time in vivo through fluorescence signals.In addition,2-1-HSA can be highly enriched in tumors and achieve high-contrast fluorescent labeling of tumors in vivo,which has great potential in clinical tumor labeling and surgical resection.2.The structure of the traditional cyanine dye(Cy-7)was modified,and the pyridine group was introduced asymmetrically to realize the construction of a large Stokes shift(≈230 nm)cyanine fluorescent dye.These dyes can spontaneously bind to the Ⅲb domain of HSA and show significant fluorescence signal enhancement.Using these fluorescent albumin dyes,we have achieved high signal to noise ratio in situ imaging analysis of mouse liver and kidney functions.In addition,these fluorescent dyes 3-1 showed similar changes in photophysical properties after being encapsulated by F-127 and combined with HSA.We encapsulated fluorescent dye 3-1 and bis [3,4,6-trichloro-2(pentoxycarbonyl)phenyl] oxalate(CPPO)in F-127 to prepare ultra-high signal-to-noise ratio nanoprobe NIR-Cl-NP with specific response to hydrogen peroxide.This nanoprobe can visualize the production and fluctuation of endogenous hydrogen peroxide in the emergency trauma model of mice.