| Nonlinear optical multi-photon absorption（MPA）materials are of great significance in a series of applications such as micromachining,biological imaging,sensing,and photodynamic therapy,especially in the field of biological imaging.At present,people have carried out systematic research on inorganic,organic,inorganic-organic hybrid and complexes multi-photon absorption materials.Among the multi-photon absorption complex materials,iridium（Ⅲ）complexes have attracted much attention due to their high stability,high quantum yield,long fluorescence lifetime（up toμs）,and strong photobleaching resistance at room temperature and physiological conditions.In this dissertation,14 kinds of iridium（Ⅲ）complexes were synthesized with metal iridium（Ⅲ）as the center and biocompatible organic small molecules as ligands.The applications of multi-photon absorption iridium complexes in biological fields such as recognition of bioactive molecules,response to the biological microenvironment,dynamic tracking of suborganelles,fluorescence lifetime imaging and photodynamic activity were systematically studied.1.Multi-photon absorption activity and methylglyoxal specific recognition of thiazoles/amino-phenanthroline iridium complexesWith benzothiazole derivatives as the first ligand（C^N ligand）,1,10-phenanthroline derivatives as the second ligand（N^N ligand）,iridium（Ⅲ）as the center,cyclometalated iridium complexes BPN and BPN2 with multiphoton absorption activity were designed and synthesized.By introducing monoamine and diamino groups into the 5 and 6 positions of 1,10-phenanthroline,it was found that the iridium complexes with diamino phenanthroline had a specific recognition effect on methylglyoxal（MGO）.The fluorescence emission spectra showed that the complex BPN2 had an obvious fluorescence response to MGO.The results of mass spectrometry further proved that the reaction product of diamino and MGO was a stable structure with six-membered rings.Theoretical calculation showed that the electronic transition ability of complex BPN2 was improved after the reaction with MGO,thus realizing the fluorescence enhancement process.The nonlinear optical activity of the product after the reaction between BPN2 and MGO was also obviously enhanced,which was conducive to further exploring the application of biological imaging.BPN2 had low cytotoxicity and can be quickly aggregated into lysosomes through cell membranes in cell imaging experiments,so that MGO in lysosomes can be monitored by one-photon and two-photon fluorescence.2.Multi-photon absorption and mitochondrial two-photon fluorescence lifetime imaging of thiazole/phenanthroline（-Me/-CHO/-CH2OH）iridium complexesThree kinds of iridium complexes BPM,BPA,and BPH were obtained by introducing groups with different electron-donating abilities（-Me/-CHO/-CH2OH）to modify the 4,7 positions of the second ligand 1,10-phenanthroline,and the single crystal of BPM was obtained by solvent evaporation.The crystal structure analysis showed that the intramolecular electron ionization of the complexes was high,which was conducive to the generation of nonlinear optical activity,and there was a large number of C-H…πand C-H...F non-classical hydrogen bond interaction between the molecules,which can effectively avoid the fluorescence quenching caused by aggregation.Photophysical experiments showed that the fluorescence intensity and quantum yield of the complexes BPM,BPA,and BPH increased with the enhancement of the electron-donating ability of the second ligand,and the two-photon and three-photon absorption activities also increased gradually（BPM>BPH>BPA）.In vitro selective tests indicated that the three complexes had specific recognition of DNA,and the viscosity test combined with molecular docking simulation proved that the binding of the complexes with DNA limited the molecular rotation and vibration,thus improving the radiation transition ability.BPM with excellent luminescence properties was selected for cell imaging experiments.BPM could be located in mitochondria via the cell membrane and interact with mitochondrial DNA to light up mitochondria.One-and two-photon fluorescence imaging of BPM in tumor tissues and mouse tumor sites was realized.In addition,viscosity dynamics during mitochondrial damage were monitored and quantified in real-time by two-photon lifetime imaging（TP-FLIM）.3.Design and synthesis of pyrimidine bridging dinuclear iridium complexes and exploration of photodynamic therapeutic applicationsTwo mononuclear iridium complexes（B1,B2）,two symmetrical binuclear iridium complexes（BD1,BD2）,and two asymmetric binuclear iridium complexes（BD3,BD4）were designed and synthesized with benzothiazole and 2-phenyl pyridine derivatives as C^N ligands and 2-pyrimidine-benzimidazole as bridging ligands.The photophysical properties indicated that the fluorescence of the binuclear iridium complexes was significantly reduced compared with the mononuclear iridium complexes.B1,B2 with good luminescence properties,and BD4 with high singlet oxygen（1O2）yield were selected for biological application exploration.In light and dark conditions,cytotoxicity tests indicated that B1 and B2 had low phototoxicity and dark toxicity,while BD4 had high phototoxicity.Confocal imaging showed that B1 and BD4 were distributed in the cytoplasm,while B2 was distributed in the lysosome,and had a bright fluorescence signal,which could be used for one-and two-photon cell and tissue imaging.Imaging tests of living and dead cells demonstrated that BD4 could effectively produce 1O2 under light,thus inducing cell apoptosis.In addition,the experiment of tumor-bearing mice showed that BD4 could stay in the living tumor site of mice for a long time,inhibited the tumor growth of mice under light,and cause tumor ablation,which had an obvious photodynamic therapeutic effect.Iridium（Ⅲ）can be excreted in vivo through organs such as the liver and kidney,urine,and feces,with little damage to the living body.Therefore,BD4 can be used as an effective photosensitizer in photodynamic therapy.4.Multiphoton absorption and mitochondrial viscosity monitoring of thiophene-dipyrazolyl iridium complexesIn order to obtain iridium complexes with high fluorescence quantum yield and nonlinear optical activity,the thiophene-dipyrazole group with nonlinear optical effect was selected as the second ligand,and 2-phenyl pyridine derivatives as the first ligand to form three iridium complexes TB1,TB2 and TB3.The introduction of quaternary ammonium salt in complex TB3 improved the molecular charge number and water solubility,and the quantum yield of fluorescence in the aqueous solution of TB3 was as high as 52.7%.In addition,the introduction of quaternary ammonium salt in TB3improves intramolecular electron mobility and optimizes nonlinear optical activity,resulting in large two-photon and three-photon absorption cross-sections in aqueous solutions.The viscosity test of the complex TB3 showed that the molecular rotation was limited and the radiation energy was increased at high viscosity.TB3,which has excellent luminescent properties in an aqueous solution,was selected for cell testing.TB3 had low cytotoxicity and can target and light mitochondria through electrostatic interaction with RNA.Further application exploration demonstrated that TB3 could be used as a two-photon fluorescence lifetime imaging probe to monitor and quantify viscosity changes during mitochondrial cracking in cells.More importantly,the probe showed excellent ability to detect abnormal viscosity in tumorous mice and could be used for imaging viscosity abnormalities in tumorous mice... |
