The Syntheses And Applications Of Small-molecule Fluorescent Probes Based On Stilbene Fluorophore

Fluorescent technology has many advantages,such as high sensitivity,good reproducibility,easy operation,fast response,low detection limit,low dosage,etc.It has been widely used in biology,clinical testing,environmental chemistry,food analysis,materials science,and drug design.Stilbene fluorophores have been received extensive attention in the synthesis and application of fluorescent probes,due to their excellent fluorescent properties,and the characteristics of optical regulatability and expansibility,which can be obtained by tailored synthesis.This thesis designed and developed multi-functional small molecule fluorescent probe based on stilbene to label peptides,proteins and oligosaccharide,and to detect various types of amines.?1?The new fluorescent noncanonical amino acid 4 with a stilbene backbone was synthesized,and its structure was confirmed by NMR,MS and IR.The excitation and emission wavelengths of compound 4 are 312 nm and 359 nm,respectively.The Stokes shift of it is 47 nm,and the quantum yield is 0.09.The photo-induced cis/trans isomerization of compound 4 in methanol solution was observed.The trans to cis isomerization rate is determined to be 0.046 min^-1.These properties enable compound4 to be used for structural and other related biological studies of proteins.?2?The Meldrum's acid derivative 5 which contains phenylene vinyl backbone was synthesized.The structure of compound 5 was fully characterized by standard ¹H NMR,¹³C NMR,COSY,HSQC,IR,MS and X-ray crystallography.Compound 5 is nearly non-fluorescent.However,it can react with a variety of amines to form fluorescent product AM-5,which can further react with thiols to afford AM-5-SH,leading to the linkage of the amine with thiol.Upon the addition of dithiothreitol?DTT?,both amine and thiol can be declicked,with the formation of 5-DTT.The HPLC-FD analysis of the derivatives of several representative amines coupled with 5demonstrates that this fluorogenic probe can be applied in the quantification of amines in various systems.The new derivatization method can provide sensitive analysis of ethylamine with high precision.Click reactions of compound 5 with peptides and chitohexaose indicate that it is very useful in biological studies involving amines.The formation of macrocyclic peptides via click reaction with 5further proved its great value in biological applications.The adjusted R² of the fitted hyperbolic curves of the click reactions between 5 and amines are very close to 1,which indicated that they can be well categorized as second-order reactions.The rate constants reflect the nucleophilicity of different amines studied in the click reaction.