Physical measurement
NMR spectra were recorded on a Varian VR400-MHz spectrometer with TMS as an internal standard. The melting points of the compound were determined on a Beijing XT4-100X microscopic melting point apparatus. The UV-Vis spectra were recorded on a Perkin-Elmer Lambda-35 UV-Vis spectrophotometer. Fluorescence spectra were obtained on a Cary Eclipse spectrophotometer at room temperature. The fluorescence image of HeLa cells was tested using olympus fv1300 laser confocal fluorescence microscopy. The SEM images was tested by F250 thermal field emission scanning electron microscopy. The binding activity was calculated by Gaussian16 software and the weak interaction was dealed with Grimme's D3BJ dispersion correction. HR-ESI-MS was tested by microTOF-Q II 10410 instrument. The XPS quantum were tested by FLS980 series of fluorescence spectrometers.
Detection of Zn 2+ in Water Solutions.
All spectroscopic measurements were performed in water solution, the pH (7.2) was controlled by HEPES buffer. In the titration experiments, each time 2 mL aqueous solution containing 100 µL solution of L (1 mmol/L) was filled in a quartz optical cell of 1 cm optical path length. Then, 10 µL Zn2+ stock solution (1.0×10− 3 M) was added to the compound solution with a micro-pipette. The spectra were collected after the addition of Zn2+ into the solution for 30 s. The excitation wavelength of fluorescence spectrum was 365 nm.
The detection limit (qmin) P-β-CD + L for Zn2+ in pure water environment was calculated by the equation.
In the equation, qmin was the detection limit, sb was the standard deviation of a blank signal, S was the sensitivity of fluorescence method for analyzing Zn2+ at 475 nm. According to the ratio equation, the detection limit could be obtained.
The synthesis process of organic small molecule L, Poly β-Cyclodextrin (P-β-CD) and host-guest supramolecular system P-β-CD + L are shown in Fig. 1. By SEM, EDS, HRTEM and solid state fluorescence spectrum, the compounds were characterized. And the corresponding element mapping (the presences of N element) demonstrated preliminary the construction of host-guest supramolecular system P-β-CD + L.
Detection of Zn2+ in Water Solutions
All spectroscopic measurements were performed in water solution, the pH (7.2) was controlled by HEPES buffer. In the titration experiments, each time 2 mL aqueous solution containing 100 µL solution of L (1 mmol/L) was filled in a quartz optical cell of 1 cm optical path length. Then, 10 µL Zn2+ stock solution (from H2O solution, 1.0×10− 3 M) was added to the compound solution with a micro-pipette. The spectra were collected after the addition of Zn2+ into the solution for 30 s. The excitation wavelength of fluorescence spectrum was 365 nm.
Cell imaging experiments
HeLa were cultured in a flask in Dulbecco’s modified Eagle’s medium supplemented with a heat-inactivated bovine serum (10%), 100 U mL− 1 penicillin and 100 U mL− 1 streptomycin and were maintained at 37 ℃ in a humidified atmosphere (5% CO2 and 95% air). The cells were seeded in six orifice plate and allowed to adhere for 12 h before treatment. Then, the cells were incubated with fresh media containing 100 uL P-β-CD + L (dissolved in H2O, pH = 7.2, 1.0 mmol/L), After 0.5 h, the growth medium was removed, and the cells were washed with saline solution for several times. The cover glass was then mounted on a microscope glass slide and was studied under a microscope. Then the water solution (100 uL, CZn2+ = 1.0 mmol/L) of was added to the HeLa cells handled with solution of P-β-CD + L. And the cells were incubated for 0.5 h, and the cells were washed with saline solution for several times, and the cell was studied under a microscope. Imaging was performed using an olympus fv1300 laser confocal fluorescence microscopy. Excitation wavelength was 365 nm and the emission collection wavelength was 422–522 nm (both blue fluorescence and green fluorescence were collected).
The cytotoxicity (HeLa cells) of P-β-CD + L
The cytotoxicity (HeLa cells) of Cou-In was tested by CCK8 methods. The experiment was divided into control group and experimental group. The control group was added with 100 uL/well complete medium. Diluted P-β-CD + L to 2%, 4%, 6%, 8%, 10% working solution, then added 100 uL/well, respectively. Three holes were set for each concentration.
HeLa cells in logarithmic growth phase were taken for cell counting and cell concentration was adjusted. According to the sample, cells were co-cultured with the cells for 24 h. The number of cells in each well was 4×103, and the cells were spread into 96-well plates. Cultured overnight in a 37 ℃ incubator with 5% CO2. According to the above group processing and training 24 h. Removed the medium. Clean the wells three times with PBS, then add medium containing 10% CCK-8, 5% CO2 and culture in an incubator at 37℃ for 2 hours. The absorbance value at 450 nm was detected by enzyme plate analyzer.
Synthesis and characterization of organic small molecule L
The preparation process of L was showed in Fig. 1. An ethanol solution (10 mL) of 2-Hydroxy-5-methylisophthalaldehyde (0.1 mol, 0.164 g) was added to another methanol (10 mL) containing 8-hydroxy-2-aminoquinoline (0.1 mol, 0.327g), Then the solution was reflux for 10 h and cooled to room temperature, then red precipitate appeared. The precipitation was filtered and dried under vacuum. Recrystallization from CH3OH/H2O (V:V = 1:2 gave the target product L which was dried under vacuum. Yield, 68.5%, mp: 260–261°C. 1HNMR (DMSO–d6, 40 0MHz, Fig. S1): δ 10.46 (2H, s, –OH), δ 10.22 (2H, s, –OH), δ 10.04 (2H, s, –OH), δ 7.70–7.91 (2H, m, ph-H from quinoline), δ 7.45 (1H, s, –CH = N), δ 7.11–7.16 (2H, m, ph-H from quinoline), δ 7.02–7.03 (2H, m, ph-H from quinoline), δ 6.89–6.91 (1H, d, ph-H from quinoline), δ 6.80–6.82 (2H, d,ph-H from quinoline), δ 6.55 (3H, s, ph-H), δ 2.31 (3H, s, –CH3). 13CNMR (DMSO–d6, 100 MHz, Fig. S2): 192.63, 157.26, 150.16, 140.97, 138.04, 137.66, 128.48, 124.22, 123.26, 122.24, 118.19, 112.25, 113.19, 114.41, 20.02. HRESI-MS (Fig. S3) for C26H22O3N4: 439.1749 (molecular ion peak of L). In addition, the Uv-vis spectra (Fig. S5) of L in DMF/C2H5OH was tested, and its extinction coefficient was calculated to be κmax = 2.19 × 104 L∙mol− 1∙cm− 1.
P-β-CD + L was prepared by host-guest supramolecular self-assembly method (Fig. 1), 10 mg L was dissolved in 10 mL DMF, then added to 20 mL H2O solution contained 100 mg P-β-CD. Ultrasound for five minutes, and the mixed solution was stirred at 80℃ for 12 h, then filtrated and evaporated excess solvent, light yellow solid was obtained. The light yellow solid was dissolved in pure water again and filterated, then freeze drying for 24 h. P-β-CD + L (light yellow) was obtained. The 1HNMR data in D2O confirmed the existence of L in the assembled system (Fig. S5). Simulatanelously, we confirmed that the Uv-vis spectrum of self-assembly system exhibited obvious difference (Fig. S4), which was concluded from the supramolecular interaction. And the supramolecular system was stable over a longer period of time (Fig. S6). In addition, it needed to be characterized by SEM, TEM, IR, fluorescence spectrum and XPS.