A kind of fluorescent imaging probe and its preparation method and application
A fluorescent imaging agent and probe technology, which is applied in the field of fluorescent imaging, can solve the problems of easy transmembrane transport into tumor cells, tumor cell residence time, difficult to predict cancer cells, easy aggregation, etc., to achieve good fluorescent imaging effect, transmembrane The effect of strong membrane transport and long residence time
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Embodiment 1
[0063] Example 1 Compound G 1 and G 2 Synthesis
[0064] 1 Compound G 0 Synthesis
[0065] According to the reported method (Chemical communications2017,53,6117-6120), G 0, specific method: KI (5.8g, 34.93mmol, 6eq) was dissolved in acetonitrile (25mL); then 5-chloro-1-pentyne (1841 μL, 17.45mmol, 3eq) was added. The reaction solution was stirred at 50°C for 40 minutes. To this was added 2,3,3-trimethylindoline (933 μL, 5.82 mmol, 1 eq). The reaction mixture was heated at 85°C for 24 hours. The reaction liquid was filtered, and then acetonitrile was removed by distillation under reduced pressure. The obtained solid was purified by column chromatography (mobile phase: MeOH and DCM volume ratio: 2-5%) to obtain compound 1 in a yield of 17% (224mg, 0.991mmol). Compound 1 (221 mg, 0.977 mmol, 1 eq) and 2-chloro-1-formyl-3-(hydroxymethylene)-1-cyclohexene (84.1 mg, 0.489 mmol, 0.5 eq) were dissolved in anhydrous acetonitrile (3 mL). Acetic anhydride (123 μL) and sodium ac...
Embodiment 2
[0078] Determination of photochemical and photophysical properties of embodiment 2 compounds G1 and G2
[0079] 1 Determination of absorption spectrum and fluorescence spectrum
[0080] Methanol was chosen as the solvent to prepare G 0 ,G 1 and G 2 The stock solution (20mM) was used to measure the spectrum. Dilute G with methanol and water, respectively 0 , G 1 and G 2 The stock solutions were diluted to different concentrations (4 μM, 3 μM, 2 μM, 1 μM, 0.5 μM) to obtain absorption spectra. Absorbance fit straight lines at λmax and concentration were obtained. Prepare 4 µM G 0, G 1 and G 2 solution to measure the fluorescence spectrum. The present invention uses methanol and water as solvents respectively to compare the fluorescence intensity of probes in different solvents.
[0081] 2 Determination of photon quantum yield
[0082] Prepare G with methanol and water as solvents, respectively 1 , G 2 and indocyanine green (ICG) solution (1 μM); ICG was used as a r...
Embodiment 3
[0095] Example 3 Tumor cell-specific imaging in vitro
[0096] 1 cell culture
[0097] To verify the specific accumulation of the probe at the tumor site, HeLa cells were selected as BR-positive tumor cells, and LO2 cells were selected as BR-negative normal cells. HeLa and LO2 cells were placed in high glucose DMEM medium containing 10% (v / v) fetal bovine serum (FBS) in 5% (v / v) CO 2 Cultivated in a humid and stable environment. The incubation temperature was 37°C.
[0098] 2 Cytotoxicity assay
[0099] Perform MTT assay to evaluate probe G 1 and G 2 cytotoxicity. HeLa and LO2 cells were seeded into 96-well plates, and the density was controlled at 10,000 cells per well. Cells were incubated at 37°C for 24h to allow cell attachment. Will G 1 and G 2 respectively dissolved in DMF to obtain 20mM G 1 and G 2 Stock solution, G 1 and G 2 The stock solution was diluted to the target concentration. After removing the medium, probe solutions at different concentrations ...
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