Synthesis Of Functional Carbon Dots And Their Applications In Fluorescent Imaging Of Cellular Microenvironment

Carbon dots(CDs),the emerging fluorescent nanomaterials,have the advantages of abundant precursors,simple synthesis processes,excellent photo-property,strong stability,and low cytotoxicity,and have gained increasing attention and played important roles in chem/biosensing and biomedical imaging.However,the synthesis of CDs is still semi-experienced,which causes the fact that the prediction of luminescent properties and bio-sensing/imaging applications of CDs are still challenging.Herein,oriented with demands on the functions of CDs in specific biological imaging application,we explored the relationship between the structure and function of CDs,and developed a controllable preparation method to creat a kind of CDs with the long-term mitochondrial tracking ability and proposed the the genetic effect of functional units in CDs.On the basis of the preparation strategy,we created a series of functional CDs that could differentiate cancer cells from normal cells,monitor the lysosomal ATP in situ and visualize the early cytotoxicity of triclosan in vivo based on the pushing the application of CDs in the fluorescence imaging of cellular microenviroment.The main content of this thesis includes:1.The synthesis method of fluorescent CDs based on structure-activity relationshipAt present,the properties and functions of CDs are unpredictable due to the lack of theoretical guidance in synthesis.In order to solve this problem,based on the luminescent mechanism of molecular state and the regulating effect of functional units on the properties of CDs,a synthesis method based on the structure-activity relationship was developed.According to the requirements in application,the CDs synthetic route can be determined through designing the functional units and precursors.In this chapter,fluorescent CDs ranged from green to red were prepared with the different funtional groups,where the lipophilic cation of rhodamine-like structure was taken as the functional units,and the citric acid and the derivatives of m-aminophenol were used as precursors.It is indicated that the mitochondrial targeting ability of MitoTCDs can be inherited from the lipophilic ability of the cationic rhodamine subunits,which demonstrates that functional units with genetic effects can directly regulate and adjust the photophysical properties and bioimaging functions of CDs.The developed preparation method based on structure-activity relationship provides a good foundation for the application of CDs in chem/bio-sensing and biomedical imaging.2.The synthesis of CDs and their applications in distinguishing cancer cells from normal cellsIn order to distinguidsh cancer cells from normal cells,considering that the lysosomes in cancer cells are numerous and large with the acidic environment that induced the nanoparticles aggregation,CDs with functional units of piperazine substituted rhodamine-like were developed using the precursors of citric acid and m-hydroxyphenyl piperazine,with the functions of cancer cells identification.These CDs have strong stability and good biocompatibility.The luminescent properties and functions of the CDs can inherit from the piperazine substituted rhodamine.These CDs can realize the specific long-term recognition of lysosomes and maintain strong and stable fluorescence in the acidic environment of lysosomes in cancer cells,thus distinguishing cancer cells from normal cells.Furthermore,these CDs can be applied for the fluorescence visualization of tumor in living mice.This work provides a new tool for the early diagnosis of cancer and promotes the application of CDs in biological imaging.3.The sythesis and application of CDs with the function of in situ and real-time monitoring the lysosomal ATP levelIn order to monitor lysosomal adenosine triphosphate(ATP)level in and real-timely,while effectively avoiding the interference of cytoplasm and other organelles,according to lysosomal acidic microenvironment and ATP,CDs with the functional units of amine and rhodamine 6G with the spirolactam ring were synthesized using citric acid,triethylenetetramine,and rhodamine 6G as precursors through microwave irradiation.A linear relationship between the fluorescence intensity of CDs and the concentration of ATP can be obtained from 0.01 to 0.3 mM with a detection limit of 0.3 μM.The results demonstrate that the fluorescence of CDs is "off" but increase when they enter the lysosomes through the cell membrane and cytoplasm,in which a chemical reacition between CDs and ATP was triggered under the acidic environment.The results indicat that the fluorescence of CDs inherited from the functional units.After encountering with the acidic environment,the spirolactam ring of rhodamine was broken by ATP,so that the fluorescence of rhodamine increased.Furthermore,based on the reversibility of fluorescence signals,CDs can monitor lysosomal ATP level in real-time under drug stimulation,such as chloroquine,etoposide,and oligomycin.This method is expected to be further used to study cell processes,enzyme activities,and drug mechanisms.4.Sythesis and application of functional CDs for early visual evaluation of low concentration triclosan in organismsIn order to early evaluate the broad-spectrum antibacterial triclosan(TCS)in organisms,considering that TCS,a uncoupler,can induce the microenvironmental changing,such as mitochondrial membrane potential decrease and fission,a new kind of CDs with the functional units of strong positive charged rhodamine-like structure were synthesized using L-(+)tartaric acid and 8-hydroxyjulolidine as precursors through microwave irradiation.With the strong stability and good biocompatibility,these CDs as the indicator can visual monitor the changes of mitochondrial microenvironments induced by TCS through the changes of fluorescence intensity and the fine structure.Monitoring the TCS in zebrafish was further realized based on these CDs.This method extends the function of CDs and provides an effective strategy for monitoring other broad-spectrum antibacterial.