Fabrication Of Responsive Polymers-Based Fluorescent Chemosensors

Due to their environmental responsiveness responsive polymers have found broad range of applications in catalysis, sensors, drug/gene delivery, self-assembly, and so on. In the current thesis, we explored the application of responsive polymers in the design of fluorescent chemosensors. The first chapter gave a brief introduction concerning the development and challenges of responsive polymers and chemosensors. The second chapters reported the design of thermosensitive microgel-based Cu2+chemosensors and improved detection sensitivity was achieved by simply increasing detection temperature. The third chapter reported the design of dye-labeled thermoresponsive block copolymers and employed the hydrophilic-hydrophobic transition of the polymers for the design of temperature chemosensors. The forth chapter reported the design of fluorescent chemosensors for the detection of both Zn2+and temperature by introducing polarity sensitive and Zn2+sensitive quinoline derivative into thermosensitive block copolymers. Finally, fluorescent pH chemosensors applied for near neutral and acid circumstances were designed based on dye-labeled pH sensitive block copolymers. The dissertation can be further clarified as follows:1. Thermoresponsive microgel-based Cu2+chemosensors possessing tunable detection sensitivity via thermo-induced microgel collapse/swelling were fabricated. A novel phenanthroline-containing fluorescent monomer capable of Cu2+-binding and fluorescence sensing, PhenUMA, was synthesized at first by reacting5-amino-1,10-phenanthroline with2-(3-isocyanato-propionyloxy)ethyl methacrylate. Near-monodisperse Cu2+-sensing microgels were synthesized via emulsion polymerization of N-isopropylacrylamide (NIPAM) in the presence of N,N’-methylenebis(acrylamide)(BIS), an anionic surfactant, and PhenUMA at around neutral pH and70℃. At20℃, PhenUMA-labeled microgels in their swollen state can selectively bind Cu2+over other metal ions (Al3+, Mg2+, Zn2+Fe3+, Mn2+, Ni2+, Ag+, Cd2+, Hg2+, and Pb2+), leading to prominent quenching of fluorescence emission intensity. At a microgel concentration of0.25g/L, Cu2+detection limit can be down to-125nM. When heated above32℃, fluorescence intensity of PhenUMA-labeled microgels in the absence of Cu2+exhibits ca.33%increase due to their volume phase transition, which is reasonable considering that fluorescent PhenUMA moieties are now located in a nonpolar environment. Furthermore, Cu2+detection sensitivity of PhenUMA-labeled microgels can be dramatically enhanced via thermo-induced microgel collapse at elevated temperatures. At a microgel concentration of0.083g/L, detection limits of Cu2+ions can be drastically improved from～28nM at20℃to～8nM at40℃. A plausible mechanism for the thermo-induced enhancement of Cu2+detection sensitivity has been proposed.2. Well-defined double hydrophilic block copolymers (DHBCs), PEG-b-P(NIPAM-co-TrpMA) and PEG-b-P(NIPAM-co-PhenUMA), bearing fluorescent L-tryptophan and phenanthroline moieties, TrpMA and PhenUMA, in the thermo-responsive block were synthesized, respectively. Upon heating in the aqueous solution, the fluorescence of PEG-b-P(NIPAM-co-TrpMA) decreased while the fluorescence of PEG-b-P(NIPAM-co-PhenUMA) enhanced. By mixing the two copolymers via comicellization, a novel responsive polymer-based ratiometric fluorescent thermometer with an overall～21.6-fold intensity ratio change in the temperature range of20-60℃was constructed, exhibiting improved performance compared with that of single-dye containing DHBCs.3. Responsive DHBCs-based fluorescent thermometers were fabricated and their application in intracellular temperature imaging was explored. Coumarin-based blue-emitting monomer, CM A,7-nitro-2,1,3-benzoxadiazole (NBD)-based green-emitting monomer, NBDAE, and rhodamine B-based red-emitting monomer, RhBEA, were synthesized at first. Well-defined DHBCs bearing CMA in the thermo-responsive PNIPAM block, PEG-b-P(NIPAM-co-CMA), were synthesized via reversible addition-fragmentation chain transfer (RAFT) polymerization of NIPAM and CMA in the presence of PEG-based macroRAFT agent. Similarly, NBDAE-labeled PEG-b-P(NIPAM-co-NBDAE) and RhBEA-labeled PEG-b-P(NIPAM-co-RhBEA) were synthesized, respectively. Based on the good fluorescence resonance energy transfer (FRET) potentials between CMA and NBDAE as well as between NBDAE and RhBEA ratiometric fluorescent thermometers were constructed by simply mixing PEG-b-P(NIPAM-co-CMA) and PEG-b-P(NIPAM-co-NBDAE) or PEG-b-P(NIPAM-co-NBDAE) and PEG-b-P(NIPAM-co-RhBEA) in a suitable molar ratio. In the temperature range of20-44℃ratiometric fluorescent thermometer exhibited-10.3-fold ratio change for the FRET pairs of PEG-b-P(NIPAM-co-CMA) and PEG-b-P(NIPAM-co-NBDAE) and~3.4-fold for PEG-b-P(NIPAM-co-NBDAE) and PEG-b-P(NIPAM-co-RhBEA). In addition to the utilization of conventional one-step FRET method cascade FRET based on three fluorophores (CMA, NBDAE, and RhBEA) was also explored for the improvement of the performance of ratiometric fluorescent thermometer, in which NBDAE acted as a FRET mediator by transferring the energy from CMA to RhBEA. Through the optimization of the molar contents of the three components～8.4-fold ratio change occurred in the temperature range of20-44℃but the detection sensitivity improved a lot. As far as we concerned, this work represents the first example of ratiometric fluorescent thermometer employing thermo-responsive polymer-based cascade FRET.4. Responsive DHBCs-based dual fluorescent chemosensors for Zn2+ions and temperature were synthesized. A novel quinoline-based polarity-sensitive and Zn2+-recognizing fluorescent monomer (ZQMA) was synthesized at first. Well-defined DHBCs bearing ZQMA in the thermoresponsive block, PEG-b-P(MEO2MA-co-OEGMA-co-ZQMA), were synthesized via RAFT polymerization of2-(2-methoxyethoxy)ethyl methacrylate (MEO2MA), oligo(ethylene glycol) monomethyl ether methacrylate (OEGMA), and ZQMA in the presence of PEG-based macroRAFT agent. The OEGMA contents in the thermoresponsive block varied in the range of0-12.0mol%to tune their lower critical solution temperatures (LCSTs). At20℃, almost nonfluorescent PEG-b-P(MEO2MA-co-ZQMA) molecularly dissolved in water and can selectively bind Zn2+ions over other common metal ions, leading to prominent fluorescence enhancement. At a polymer concentration of0.2g/L, the Zn2+detection limit can be down to～3.0nM. Upon heating to above the LCST, PEG-b-P(MEO2MA-co-ZQMA) self-assembles into micelles possessing P(MEO2MA-co-ZQMA) cores and well-solvated PEG coronas, and the fluorescence intensity exhibit～6.0-fold increase due to the fact that ZQMA moieties are now located in a more hydrophobic microenvironment. Compared to the unimer state at20℃, although PEG-b-P(MEO2MA-co-ZQMA) micelles possess slightly decreased detection limit for Zn2+(～14nM), reversible binding between ZQMA moieties and Zn2+ions at37℃can be achieved, as evidenced by the on/off switching of fluorescence emission via sequential addition of Zn2+and EDTA. In vitro fluorescence imaging studies suggested that the micelles can effectively enter into living cells and sensitively respond to Zn2+ions. Besides, by introducing blue emitting coumarin derivative labeled thermoresponsive DHBCs, PEG-b-P(MEO2MA-co-Coum), and red emitting RhBEA-labeled block copolymer, PEG-b-P(St-co-RhBEA) into the detection system of PEG-b-P(MEO2MA-co-ZQMA) Zn2+ion induced pink-white-green three color transition was obtained. The concentration of Zn2+ion can be simply judged by simply discriminating the color of the polymer solutions.5. Ultrasensitive ratiometric fluorescent pH chemosensors based on pH-responsive DHBCs covalently labeled with green-emitting BTPE and blue-emitting CMA moieties were constructed. The pH-sensitive emitting nature of BTPE and emission turn-on of CMA residues triggered by pH-actuated micelle-to-unimer transition of diblock micelles, together with their complementary pH-sensing range difference, synergistically contribute to the observed～250-fold changes of BTPE/CMA emission intensity ratios in the whole pH range. Two-photon ratiometric fluorescent pH mapping of intracellular gradients subjected by pH-responsive micellar nanoparticles in their endocytic pathway has also been achieved.6. Ultrasensitive ratiometric fluorescent pH chemosensors based on endosomal escaping block copolymers was fabricated and its application in intracellular pH imaging was investigated. Dually labeled pH-responsive diblock copolymers, P(DMA-co-HCCME)-b-P(DEA-co-BMA-co-TMR), were synthesized by combining consecutive RAFT polymerizations and post-functionalization, where DMA, DEA, and BMA are N,N-dimethylacrylamide, N,N-diethylaminoethyl methacrylate, and butyl methacrylate, respectively. Blue fluorescent pH sensitive HCCME was labeled in PDMA block and red fluorescent pH-inert TMR was labeled in endosome escapable P(DEA-co-BMA) block. Intracellular pH imaging demonstrated that P(DMA-co-HCCME)-b-P(DEA-co-BMA-co-TMR) can effectively escape from endosomes/lysosomes to reach cytosol and exhibited excellent sensitivity to pH fluctuations under external stimuli. Meanwhile, it was indicated that both pH buffering capability and appropriate hydrophilicity-hydrophobicity balance play important roles in effective endosome escaping.