Temperature-induced Fluorescence Enhancement Of Biomimic Fluorescent Polymers And Its Application

Green fluorescent protein (GFP), due to its special optical properties,has been widely used as biological marker in molecular biology, cell biologyand biochemistry. Fully fluorescent GFP is efficiently formed at lowtemperature. When subjected to high temperature, denaturalization of GFPresults in the complete lose of fluorescence, thus greatly limiting its wideapplication. Therefore, searching for GFP mutants that have better emissionproperty at high temperature is necessary. We designed and fabricated a novelthermo-responsive block copolymer containing GFP-chromophore, in orderto improve the fluorescence property of GFP chromophore based on thepolymers’ thermosensibility. This dissertation includes the following threeparts:1. GFP chromophore was synthesized via2,3-cycloaddition. Thechromophore’s fluorescence is very weak due to its conformational rotation.In addition, with the increase of temperature, the fluorescence intensity of theGFP chromophore reduced rapidly.2. Thermo-sensitive block copolymer poly(ethylene glcol)-chromophore-poly(N-isopropylacrylamide)(PEG-c-PNIPAM) with a chromophore locatedin the middle of polymer backbone was successfully synthesized. In the firststep, macromolecular chain transfer agent with an azide group was prepared,which was further used to fabricate PEG-c-PNIPAM copolymer viareversible addition fragmentation chain transfer polymerization (RAFT) and“click” reaction. Because of the lower critical solution temperature (LCST) ofPNIPAM, the chromopore’s fluorescence property could be improved by theself-assembly of PEG-c-PNIPAM through changing the temperature of the copolymer aqueous solution.3. Two block thermo-sensitive copolymers poly(ethyleneglcol)-b-poly(N-isopropylacrylamide)-chromophore (PEG-b-PNIPAM-c)with a chromophore in the end of hydrophobilc block was successfullysynthesized. In the first step, PEG-macroinitiator was synthesized andemployed to prepare PEG-b-PNIPAM-Cl with different molecular weights.Then, PEG-b-PNIPAM-Cl copolymers were modified with azide group,which reacted with chromophore through “click” reaction. When thetemperature was above LCST, the self-assembly of PEG-b-PNIPAM-crestricted the chromophore’s conformational rotation, resulting in a greatenhancement of the copolymer’s fluorescence. The chromophore-basedcopolymers with temperature-induced emission enhancement property wereused for bacterial detection, showing high Bacillus thermophilus detectionsensitivity.