Synthesis, Optical Properties And Bioimaging Application Of Fluorine And Nitrogen Co-doped Carbon Dots

Carbon dots?CDs?are a new type of carbon-based nanomaterials,which are generally less than 10 nm in size with fluorescent properties.Since the discovery of CDs in 2004,they have received the attention and research of various scholars.Since CDs have good biocompatibility,low biotoxicity,light stability,adjustable fluorescence emission and other excellent characteristics,they have broad potential application in the fields of photocatalysis,biosensors,optoelectronic devices,bioimaging and photodynamic therapy.However,due to the diversity of raw materials and the differences in synthesis conditions,the structure and optical properties of CDs that have been reported are also complicated.So far,the mechanism of CDs luminescence has not been studied clearly.A large number of research results have shown that the introduction of heteroatoms can significantly adjust the surface state and energy band gap of CDs,thereby affecting the optical properties such as the absorption spectra,emission wavelength and fluorescence lifetime of CDs.This thesis focuses on the effects of co-doping of fluorine?F?and nitrogen?N?elements on the ultraviolet-visible-near infrared absorption spectra,fluorescence emission spectra and nonlinear optical properties,and its application in bioimaging from deep red to near infrared region.It mainly consists of three parts:?i?the first case of F and N co-doped CDs?F,N-doped CDs?with photoactivation properties was synthesized by a simple microwave-assisted pyrolysis method.The synthetic F,N-doped CDs has a reversible piezochromic effect and pressure-triggered aggregation induced emission;?ii?using small molecule organics as the carbon source,F,N-doped CDs?N-CDs-F?with special donor-?-acceptor?D-?-A?structure were synthesized by solvothermal method.The N-CDs-F has the properties of full UV-visible-NIR spectral response,unique multi-photon excitation up-conversion fluorescence and large multi-photon absorption cross sections,and its absorption and emission wavelengths are red-shifted to the near-infrared region;?iii?specific tumor cells were selected as models to evaluate the cytotoxicity,in vitro and in vivo imaging effects of photoactivated F,N-doped CDs and near-infrared N-CDs-F.The main research contents and results of this thesis are as follows:?1?F and N co-doped CDs?F,N-doped CDs?were synthesized by one-step microwave-assisted pyrolysis using polyethyleneimine as the carbon and nitrogen source and levofloxacin as the fluorine source.TEM shows that the F,N-doped CDs have a uniform size distribution with an average size of 1.38 nm.The results of X-ray photoelectron spectroscopy?XPS?show that F,N-doped CDs have a high F doping amount?3.5%?and N doping amount?11.12%?,where F exists in the form of C-F bonds,and N exists in the form of pyrrole-N and amino-N.The aqueous solution of F,N-doped CDs showed bright blue fluorescence under UV light and the absolute quantum yield was 6.78%.At the same time,compared to most previously reported CDs,F,N-doped CDs exhibits a longer fluorescence lifetime?467 ns?.In addition,F,N-doped CDs showed fluorescence with excitation wavelength dependence.The effect of pressure on the fluorescence emission of F,N-doped CDs was studied by using a symmetrical diamond anvil box device with ultra-low fluorescence.The results show that F,N-doped CDs have a reversible piezofluorescent discoloration effect.Under the high pressure of 1.0 atm-10 GPa applied to F,N-doped CDs,as the pressure increases,the fluorescence intensity of F,N-doped CDs increases first and then decreases.Pressure release experiments show that the effect of pressure on the fluorescence of F,N-doped CDs is reversible.It is worth mentioning that in the pressure range of 1.0 atm-0.65 GPa,the F,N-doped CDs show pressure-induced aggregation-induced emission?AIE?behavior,.Most importantly,we found for the first time that CDs exhibited a photo-activated fluorescence enhancement effect.Whether under normal pressure?1.0 atm?or high pressure?0.1 GPa?conditions,through continuous UV?355 nm?irradiation,the F,N-doped CDs showed the phenomenon that the fluorescence intensity gradually increased with the increase of the irradiation time.These photoactivated F,N-doped CDs with piezochromic fluorescence enhancement greatly expand the applicability of CDs from normal pressure to high pressure,and their unexpected anti-photobleaching demonstrated their great potential in the fields of photodynamic therapy,long-term bio-imaging and optoelectronic devices.?2?Using anhydrous citric acid as the carbon source,urea and ammonium fluoride as the nitrogen and fluorine sources,the fluorine and nitrogen co-doped carbon dots?N-CDs-F?with a donor-?-acceptor?D-?-A?structure was prepared by a solvothermal method.Through a series of purification methods,obtained N-CDs-F showed an average size of 2.62 nm under TEM.The results of Fourier Transform Infrared Spectroscopy?FT-IR?and XPS showed that the N-CDs-F had a large number of nitrogen-containing,fluorine-containing and oxygen-containing functional groups,among which the fluorine content was 4.07%and the nitrogen content was15.41%.Ultraviolet absorption spectrum shows that N-CDs-F has full spectrum absorption characteristics in the ultraviolet-visible-near infrared range,and fluorescence emission spectrum shows that N-CDs-F has strong emission in the visible and NIR regions,and the fluorescence intensity of N-CDs-F has excellent temperature and pH response.Interestingly,N-CDs-F is a new type of CDs with a D-?-A conjugate structure and has large two-photon,three-photon,and four-photon absorption cross-sections.This is the first report that of the three-photon and four-photon absorption cross-sections of CDs;compared with the undoped CDs,the two-photon absorption cross-section of N-CDs-F is significantly larger than that of the undoped CDs.Importantly,the three-photon absorption cross section of N-CDs-F is three times for the three-photon absorption cross section of traditional organic compound with D-?-A structure.In addition,the co-doping of fluorine and nitrogen caused significant red shift in the fluorescence emission of N-CDs-F and obvious increase in NIR emission intensity.More importantly,N-CDs-F have strong up-conversion fluorescence at excitation wavelengths of 1060 nm,1600 nm and 2000 nm,which is extremely rare in the reports of CDs.In addition to its good nonlinear optical properties,N-CDs-F can also emit bright fluorescence in the far red-NIR?650-800 nm?region under 635 nm red light excitation.Under the excitation of 710 nm,the fluorescence quantum yield?QYs?of N-CDs-F is 9.8%higher than undoped CDs?8.1%?,so N-CDs-F can be used as an excellent NIR biological fluorescence probe.?3?Using CHO,HeLa,and HepG2 cells as models,the cytotoxicity and in vitro/in vivo imaging effects of photoactivated F,N-doped CDs and near-infrared N-CDs-F were evaluated,respectively.The results of the MTT and LDH kits show that the synthesized two fluorine and nitrogen co-doped carbon dots have low cytotoxicity and excellent biocompatibility.When the concentration of CDs reached 250?g mL^-1,the cell survival rate was still higher than 80%,and there was almost no damage to the cell membrane.The results of fluorescence inverted microscope and laser confocal microscope?CLSM?show that photoactivated F,N-doped CDs and near-infrared N-CDs-F all have good cell development effects.In addition,due to the ultraviolet-visible-near-infrared full-spectrum absorption characteristics of N-CDs-F,further research on the cell imaging of N-CDs-F.The results of Z-axis scanning and live cell workstations have proved that after N-CDs-F was endocytosed by the cell,the cytoplasm emitted bright crimson-NIR?650-800 nm?fluorescence under 635 nm red light excitation.At the same time,the N-CDs-F can also enter the nucleus of tumor cells and stain the nucleoli,which may be related to its affinity with the ribosome.The results of animal imaging experiments show that N-CDs-F also exhibits excellent imaging effects in mice under 735 nm excitation.Therefore,the above results indicate that the N-CDs-F is not only suitable for NIR live cell imaging,but also for in vivo imaging.