New Strategies for Molecular Recognition Based on Amines-derived Fluorescent Nanoprobes and Their Applications in Chemical and Biological Analysis

Author:Liu Shi Gang

Supervisor:luo hong qun


Degree Year:2019





The development of highly sensitive and selective,rapid,and economical analytical methods is very important for biomedical analysis,food quality and safety control,environmental monitoring,and forensic identification.Fluorescence analysis has found widespread applications in various fields because of its high sensitivity,easy operation,and cost-effectiveness.Fluorescent probes are very important for fluorescence analysis.In recent years,the preparation of various fluorescent probes and their application in biochemical analysis have attracted a great deal of attention.The nanotechnology has brought new perspectives and progress to many disciplines and fields.Similarly,fluorescent nanoprobes have brought new opportunities for fluorescence analysis technology.For example,fluorescent nanoprobes have been increasingly researched and applied in fluorescence imaging,biochemical sensing,medical diagnosis,and disease treatment,which promotes the rapid integration of different disciplines and the development of new theories and technologies.Amine is a type of nitrogen-containing organic substance and can be regarded as a hydrocarbon-based derivative of ammonia.The amine molecule has an unpaired electron pair on the nitrogen atom,and thereby the amine generally has a strong electron-donating ability,and is easy to form a hydrogen bond,making water-solubility well.In addition,many amine compounds are important substances for life-sustaining activities,which indicates that amine-based materials may have good biocompatibility.When using amine compounds as precursors to prepare photoluminescent nanomaterial,the amino group with strong electron donating ability can be used as a fluorescent auxochromic group or nitrogen can act as a doping element to improve the optical properties.Meanwhile,the existence of amino groups on the surface of the prepared nanomaterial which serves as a probe is beneficial to the specificity for target detection.In addition,the surface amino groups are also conducive to the water-solubility of the nanomaterial and facilitate its surface modification and functionalization,which can improve the analytical performance of a fluorescent probe.In this dissertation,several photoluminescent nanomaterials,including fluorescent polymer nanoparticles,graphite phase carbon nitride nanosheets,and carbon dots,were prepared by using amine compounds(polyethyleneimine,melamine,and ethylenediamine)as main precursors based on different preparation methods.Their chemical and optical properties were studied by various characterization techniques such as electron microscopy,spectroscopy and surface analysis.Furthermore,on the basis of molecular spectroscopic analysis techniques,the prepared photoluminescent nanomaterials were used as fluorescent probes to construct new methods for recognition of some ions and molecules.The analysis performance and mechanism of these assay methods were investigated,and also their practical applications in biochemical analysis were evaluated.The main contents of this dissertation are summarized as follows:1.Polyethyleneimine-formaldehyde fluorescent polymer nanoparticles as a nanoprobe for sensing copper ions in aqueous medium and constructing an IMPLICATION logic gateA fluorescence approach for rapid,environment-friendly,and selective detection of Cu2+ in aqueous media was developed using a kind of autofluorescent polymer nanoparticles as a nanoprobe.The polymer nanoparticles were synthesized by crosslinking hyperbranched polyethyleneimine(PEI)with formaldehyde in water solution,and they emitted strong intrinsic fluorescence without the conjugation to any external fluorescent agent.The Cu2+ assay was on the basis of the principle that Cu2+ can be adsorbed on the surface of the polyethyleneimine-formaldehyde nanoparticles(PEI-F NPs),resulting in the fast and remarkable fluorescence quenching through electron transfer.A low limit of detection(62 nM) for copper ion was obtianed.The developed sensor was applied to analysis of Cu2+ in environmental water samples with satisfactory results.Furthermore,the fluorescence of the PEI-F NPs-Cu2+ system can restore upon addition of cysteine(Cys) because of its ability to remove Cu2+ from the surface of PEI-F NPs.Based on the fluorescence “on-off-on”conversion,a reversible IMPLICATION logic gate was designed by using Cu2+ and Cys as the two inputs,and fluorescence intensity of the PEI-F NPs as the output signal.2.Preparation and optical properties of polyethyleneimine-glucose polymer nanoparticles and their application for sensitive detection of nitro-explosive picric acid in aqueous mediumWater-soluble nonconjugated polymer nanoparticles with strong fluorescence emission was prepared by hyperbranched polyethyleneimine(PEI) and D-glucose via Schiff base reaction and self-assembly in aqueous phase.The preparation of the PEI-D-glucose nanoparticles(PEI-G NPs)was facile and environmentally friendly under mild conditions.Also,the PEI-G NPs showed a high fluorescence quantum yield in aqueous solution,and the fluorescence properties(such as concentration-dependent and solvent-dependent fluorescence) and the origin of intrinsic fluorescence was investigated and discussed.The PEI-G NPs were then used to develop a fluorescent probe for fast,selective,and sensitive detection of nitro-explosive picric acid(PA)in aqueous medium,because the fluorescence can be easily quenched by PA whereas other nitro-explosives and structurally similar compounds only caused negligible quenching.A wide linear range(0.05-70μM)and a low detection limit(26 nM) were obtained.The fluorescence quenching mechanism was carefully explored.It was due to a combined effect of resonance energy transfer,static quenching,as well as inner filter effect between PA and the PEI-G NPs,which resulted in the good sensitivity for PA.Finally,the developed nanoprobe was successfully applied to detection of PA in environmental water samples.3.A fluorescent and colorimetric dual-signal readout pH nanoprobe and logic gate operation based on polyethyleneimine-derived fluorescent polymer nanoparticlesAutofluorescent polymer nanoparticles were synthesized by hyperbranched PEI and salicylaldehyde via environmentally friendly imine crosslinking and self-assembly in aqueous phase.Both fluorescence and UV-vis absorption of the PEI-salicylaldehyde nanoparticles(PEI-S NPs)showed a sensitive and reversible response to pH.The mechanism of pH-dependent optical properties was investigated.The existence of two proton-responsive functional groups(phenolic hydroxyl groups and amine groups) on the PEI-S NPs surface is crucial.The pH-dependent fluorescence is ascribed to electron transfer,which is controlled by the protonation degree of amine groups,while the pH-dependent UV-vis absorption is induced by ionization of phenolic hydroxyl groups.Based on the pH-mediated optical properties,colorimetric and fluorescent dual-signal pH sensor and multiple logic gates were developed.The fluorescent PEI-S NPs with easy preparation and sensitive response to pH are very promising for applications in biological,pharmaceutical,and material science fields.4.A ratiometric fluorescent strategy for alkaline phosphatase activity assay based on g-C3N4/CoOOH nanohybrid via target-triggered competitive redox reactionIn this chapter,fluorescent graphitic carbon nitride(g-C3N4) nanosheets are prepared by using melamine as a precursor.The g-C3N4 nanosheets and cobalt oxyhydroxide(CoOOH) nanoflakes are exploited for ratiometric fluorescence assay of alkaline phosphatase(ALP) activity.In the g-C3N4/CoOOH nanohybrid,g-C3N4 nanosheets serve as a signal unit and the CoOOH nanoflakes function as a recognition element,and initially,the fluorescence of g-C3N4 is quenched by the CoOOH nanoflakes.In the absence of target,the CoOOH nanoflakes of the nanohybrid system are able to oxidize o-phenylenediamine(OPD),and the resultant oxidation product(OxOPD) quenches the blue emission of g-C3N4 and meanwhile emits orange fluorescence which acts as another signal element.However,an efficient redox reaction between ascorbic acid(AA)and CoOOH can cause decomposition of the CoOOH nanoflakes,and additionally,ALP can catalytically hydrolyze L-ascorbic acid-2-phosphate(AAP) to generate AA.Thus,in the presence of target,the CoOOH nanoflakes are destroyed by AA preferentially and the OPD is rarely oxidized to OxOPD,accompanied with strong blue emission of g-C3N4 and weak orange fluorescence from OxOPD.Target-dependent dual-signal change makes the ratiometric assay possible,and also AA-induced signal variation was investigated and attributed to the stronger reducing capacity of AA than OPD.The ratiometric sensing platform for ALP activity assay provides a new perspective for the applications of two-dimensional nanomaterials to develop novel and sensitive biosensors.5.A new strategy for development of ratiometric sensing based on combined fluorescence and light scattering:The ratiometric detection of ascorbic acid using CDs/CoOOH nanosystem as a proof of conceptThe development of ratiometric fluorescent strategy is very significant and challenging in biochemical analysis.Simultaneous response of fluorescence and light scattering can be obtained by using nanomaterials with size-and shape-dependent physicochemical properties.On basis of the principle,a new strategy to design ratiometric sensor by combining fluorescence and light scattering,two different and independent signals,is proposed.To obtain fluorescence and scattering signals simultaneously under a same excitation,two signal collection strategies are proposed based on the principles of fluorescence,light scattering,and diffraction.One is to collect normal(down-conversion) fluorescence and second-order scattering(SOS) signals,and the other is to record the fluorescence excited by the second-order diffraction light of excitation wavelength λ/2(SODL-fluorescence) and first-order scattering(FOS) or frequency doubling scattering(FDS) signals.A proof of concept study has been performed by using fluorescent carbon dots(CDs,prepared using PEI and citric acid) and CoOOH nanoflakes system for ascorbic acid(AA) sensing.Apart from construction of ratiometric sensors,the combined fluorescence and scattering can also act as a useful technique to monitor aggregation-induced fluorescence quenching or enhancement.6.Oxidation etching induced dual-signal response of CDs/AgNPs nanosystem for ratiometric optical sensing of H2O2 and H2O2-related bioanalysisA new ratiometric optical sensing strategy for H2O2 detection was developed by etching silver nanoparticles(AgNPs)to deactivate fluorescence resonance energy transfer(FRET)and reduce Rayleigh scattering based on a hyphenated technique of fluorescence and second-order Rayleigh scattering(SOS).The ratiometric detection of H2O2 is achieved through exploiting a hybrid system fabricated by fluorescent carbon dots and silver nanoparticles(CDs/AgNPs).The CDs were obtained using ethylenediamine and citric acid via hydrothermal method.In the CDs/AgNPs system,the fluorescence of CDs was quenched because of FRET,and the scattering is strong due to the intrinsic high light-scattering power of AgNPs.With the introduction of H2O2,the AgNPs were etched and the CDs were released from the AgNP surface,resulting in the fluorescence enhancement and scattering decline.As a result,ratiometric sensing of H2O2 can be achieved based on the CDs/AgNPs system by simultaneous collection of fluorescence and SOS signals.The sensing system is further used for H2O2-generation bioanalysis,and as a proof-of-concept,ratiometric assay of glucose and evaluation of glucose oxidase activity are performed successfully.