Donor and Acceptor Engineering for BINOL based AIEgens with Enhanced Fluorescence Performance

A D–π–A strategy enhances aggregation-induced emission performance with a large Stokes shift over 200 nm.


General information
All chemicals and solvents were commercially available and were used without further purification. All commercially available reagents were used as received unless otherwise stated. 1 H NMR, 13 C NMR spectra were measured on a Agilent AV-400 NMR spectrometer. Proton Chemical shifts of NMR spectra were given in ppm relative to internals reference TMS (1H, 0.00 ppm). ESI-HRMS spectral data were recorded on a Finnigan LCQDECA mass spectrometer.
Fluorescence emission spectra were obtained using Hitachi F-7000 spectrometer at 298 K. Absorption spectra were recorded on a Hitachi PharmaSpec UV-1900 UV-Visible Spectrophotometer. The absolute fluorescence quantum yield was measured using a Hamamatsu quantum yield spectrometer C11347 Quantaurus_QY. The fluorescence lifetime was measured using a Hamamatsu Compact Fluorescence Lifetime Spectrometer C11367. The particle size was measured using a MAL-DLS Zetasizer Nano-ZS90. Single crystals were grown from isopropanol/ dichloroethane via solute solution diffusion method. Single crystal X-ray diffraction intensity data of 2b were collected on Agilent Technologies (Gemini), and the data of 1c were collected on Broker D8 venture with METAUET D2 X-ray source 153K, and the data of 2c were collected Bruker apex II DUO with microfocus Mo X-ray Source 153K. The ground-state geometries were optimized using the density function theory (DFT) method with B3LYP hybrid functional at the basis set level of 6-31G (d,p). All the calculations were performed using Gaussian 09 package. MTS method was used for testing the cell viability and described in the experimental section. HepG 2 cells were obtained from Shanghai Institute of Biochemistry and Cell Bioc emistry and Cell Biology, Chinese Academy of Science. Confocal lasing scanning microscopic (CLSM) images of single-photo were obtained using LSM 780 (Zeiss). Unless otherwise noted, materials were obtained from commercial suppliers and were used without further purification. All the solvents were dried according to the standard methods prior to use. All of the solvents were either HPLC or spectroscopic grade in the optical spectroscopic studies. S3

Synthesis of BIN-5, BIN-COM and BIN-COP
The synthesis methods of BIN-COM and BIN-COP can be found in our previous work. 1 The synthesis methods of BIN-5 can be found in reference. 2 All of them have been characterized by NMR and HRMS which are same as reference.

Cell culture and imaging
HepG 2 cells were cultured in Dulbecco's modified Eagle medium (DMEM) containing 10% fetal bovine serum and 1% Antibiotic-antimycotic at 37℃ in a 5% CO2/95% air incubator. For fluorescence imaging, cells (4 × 10 3 /well) were passed on a 6-well plate and incubated for 24 h. For fluorescence imaging, cells (4 × 10 3 /well) were passed on a 6-well plate and incubated for 24 h. Before the staining experiment, cells were washed twice with physiological saline, incubated with 5 μM probe and 2 ul oleic acid for different times at 37℃. Then washed 3-6 times with physiological saline. The confocal fluorescent images were captured with an excitation light at 405 nm. For co-localization experiment, cells were incubated with 5 μM probe and 2ul oleic acid for 6h, then washed 3-6 times with physiological saline, then incubated with 1μm BODIPY 493/503 for 15 min. The excitation light of BODIPY 493/503 is 488 nm.

Cytotoxicity study
Toxicity toward HepG 2 cells was determined by MTS (3-(4, 5-dimethylthiazol-2-yl)-5-(3-carboxymethoxyphenyl)-2-(4-sulfophenyl)-2H-tetrazoliu m reduction assay following literature procedures. About 10000 cells per well were seeded in 96-well plates and cultured overnight for 70-80% cell confluence. The medium was replaced with 100 μL of fresh medium with different concentration of probes, to which 100 μL complexes at 200 μL. 24 hours later, 100 μL of 20% MTS solution in PBS was replaced with the old medium in each well for additional 0.5h incubation. The metabolic activity of the probes treated cells was expressed as a relative to untreated cell controls taken as 100% metabolic activity. Figure S1. Views of crystal packing mode of 1c, 2b and 2c. Carbon, hydrogen, oxygen and nitrogen atoms are shown in gray, green, red and blue, respectively. The single-crestal data can be found on CCDC, the deposition number of 2b, 1c and 2c are 1907970, 1907963 and 1907968, respectively .  Table S1. The dihedral angle and the distance data of 1c, 2b, and 2c. The dihedral angle of purine core and donor group (Ø P-D ) or acceptor group (Ø P-A ).

Views of the molecular stacking structures in single crystals
b The distance of adjacent molecule's purine core (P), donor group (D), and acceptor group (A). c The vertical distance of adjacent molecule's purine core (P).

Crystallographic data of 1c, 2b, and 2c
Crystal data and structure refinements of 1c: