An AIE-active hemicyanine fluorogen with stimuli-responsive red/blue emission: Extending the pH sensing range by "switch + knob" effect

a Division of Biomedical Engineering, The Hong Kong University of Science & Technology (HKUST), Clear Water Bay, Kowloon, Hong Kong, China, b Department of Chemistry, Institute of Molecular Functional Materials, HKUST, Clear Water Bay, Kowloon, Hong Kong, China, c Department of Physics, HKUST, Clear Water Bay, Kowloon, Hong Kong, China, d State Key Laboratory of Molecular Neuroscience, HKUST, Clear Water Bay, Kowloon, Hong Kong, China, e HKUST Fok Ying Tung Research Institute, Nansha, Guangzhou, China, f Department of Polymer Science & Engineering, Zhejiang University, Hangzhou 310027, China


Table and Figure Captions
Table S1.Crystal data and structure refinement for TPE-Cy Table S2.Bond lengths and angles for TPE-Cy single crystal structure Table S3.Sample preparation details for the PL measurement shown in Figure 1

Table S4. Sample preparation details for reversibly switching absorption and emission behaviors of TPE-Cy
High resolution mass spectra of TPE-Cy (A) before and (B) after addition of

Materials
Solvents such as THF, chloroform, dichloromethane (DCM), and hexane were distilled under normal pressure under nitrogen immediately prior to use.Milli-Q water was used as deionized water.4-Bromobenzaldehyde, tetrakis(triphenylphosphine)palladium(0), citric acid, and sodium hydrogen phosphate were purchased from Sigma-Adrich.Aniline was purchased from Int. Lab.All the chemicals were used as received without further purification.
Compound 1 1 and 4 2 were prepared according to the previously published procedures.
Buffer solutions with pH 113 were purchased from Merk, Riedel-de Haen and Sigma-Aldrich.Buffers with pH 5.107.13were prepared by citric acid and sodium hydrogen phosphate 3 and confirmed by pH meter.The aqueous mixtures of TPE-Cy for pH-responsive UV and PL tests were prepared by adding an aliquot of DMSO stock solution into aqueous buffers with specific pH values.

Instruments
pH measurements were performed on a Beckman 340 pH/Temp meter. 1 H and 13 C NMR spectra were measured on a Bruker ARX 400 NMR spectrometer using CDCl 3 , DMSO-d 6 , or D 2 O as solvent and tetramethylsilane (TMS) as internal reference.UV absorption spectra were taken on a Milton Ray Spectronic 3000 array spectrophotometer.Photoluminescence (PL) spectra were recorded on a Perkin-Elmer LS 55 spectrofluorometer.Solid state quantum efficiency was measured using an integrating sphere at an excitation wavelength of 325 nm.
High-resolution mass spectra (HRMS) were obtained on a GCT Premier CAB 048 mass Electronic Supplementary Material (ESI) for Chemical Science This journal is © The Royal Society of Chemistry 2012 spectrometer operated in MALDITOF mode.Elemental analysis was performed with an Elementar Vario Micro Cube.Particle sizes and Zeta potential of the nanoaggregates were determined using a ZETA-Plus potential Analyzer.Morphologies of nanoaggregates were studied by a JEOL 100CX transmission electron microscope.
Suitable single crystals of TPE-Cy were grown from dichloromethane and hexane with an aliquot of ethanol at room temperature in the dark.X-ray diffraction (XRD) intensity data were collected at 173 K on a Bruker-Nonices Smart Apex CCD diffractometer with graphite monochromated Mo Kα radiation.Processing of the intensity data was conducted using the SANT and SADABS routines, and the structure and refinement were carried out using the SHELTL suite of X-ray programs (version 6.10).The OPTEP drawing of TPE-Cy is given in Scheme 1 and its crystal data are summaried in Table S1.The powder XRD diffractogram was recorded on a Philips PW 1830 powder diffractometer using the monochromatized X-ray beam from a nickel-filtered Cu K  radiation ( = 1.5406Å).

Synthesis and Characterization Data of TPE-Cy
Preparation of 2. Into a 100 ml round bottom flask were added 1 1 (376 mg, 1 mmol), 4-bromobenzaldehyde (203 mg, 1.1 mmol) and tetrakis (triphenylphosphine)palladium(0) (3.5 mg, 3 μmol) were dissolved in 50 mL of degassed THF.7.2 mL of saturated sodium carbonate solution was then added to the mixture under stirring.After reflux for 12 h, the mixture was cooled to room temperature and filtered.The solvent was removed under reduced pressure, the residue was dissolved in DCM and the solution was washed with brine and water.The organic layer was separeted and dried over magnisium sulfate.After solvent evaporation , the crude product was purified by a silica-gel column using DCM/hexane Preparation of 3. Into a 250 ml Two necked flask was added compound 2 (327 mg, 0.75 mmol) in 40 mL of anhydrous ethanol under nitrogen.After all the solids were completely dissolved under heating, 0.4 mL (4.5 mmol) of aniline was added.After reflux overnight, the mixture was cooled using ice water.The precipites formed were filtered out, washed twice with cold ethanol, and vacuum-dried without further purification.The product was obtained as a yellow solid in 90% yield (346 mg). 1  Preparation of TPE-Cy.Into a 250ml flask were added compound 3 (256 mg, 0.5 mmol) and 4 (738 mg, 2.5 mmol).After vacuum-dried and refilled with nitrogen three times, 30 mL dry THF and 15 mL Ac 2 O were then injected to the mixture.After refluxed overnight, the mixture was cooled to room temperature, the solvent was removed by rotary evaportaion and the residues were extracted with chloroform and water.The organic layers were combined, Electronic Supplementary Material (ESI) for Chemical Science This journal is © The Royal Society of Chemistry 2012 further washed with brine, and dried over magnisium sulfate.After solvent evapration, the crude product was purified by silica-gel column chromatography using chloroform/methanol mixture as eluent.The product was obtained as a red solid in 73% yield (260 mg).

Procedures for Sample Preparation in TEM, pH-Switching, and NMR Studies
Sample Preparation for TEM Study 50 µL of 0.31 mM TPE-Cy stock solution in ethanol was added dropwise to 950 µL hexane while sonicating.The resultant mixture was droped onto copper 400-mesh carrier grids covered with carbon-coated Formvar films.The solvent was removed under vacuum at room temperture.

pH-Switched Absorbtion and Emission of TPE-Cy
Experimental details are shown in Table S4.To avoid dillution effect, parallelled samples were prepared.The "Cycle no.0" means that the dye molecules are first put in deionized water without any acid and base.This aqueous mixture is prepared by adding an aliquot of concentrated DMSO stock solution of TPE-Cy into a large volume of water, in which TPE-Cy molecules are well-dispersed as nanoparticles (Figure S2).From no. 0.5 to no. 4, base and acid were added alternatively to adjust the pH.Additional water, if needed, is added to ensure that the final dye concentrations of all samples are identical.F(000) 1622

Sample Preparation for in-situ NMR Study Shown in
Crystal size 0.38 x 0.12 x 0.05 mm 3 Theta range for data collection 5.01 to 66.99°.
Electronic Supplementary Material (ESI) for Chemical Science This journal is © The Royal Society of Chemistry 2012 Electronic Supplementary Material (ESI) for Chemical Science This journal is © The Royal Society of Chemistry 2012 mixture as eluent.A light yellow powder was obatined in 97% yield (422 mg, 0.967 mmol).

Table S1 .
Electronic Supplementary Material (ESI) for Chemical Science This journal is © The Royal Society of Chemistry 2012 Crystal data and structure refinement for TPE-Cy a

Table S3 .
Sample preparation details for the PL measurement shown in Figure1a The concentration of dye stock solution in ethanol is 0.31 mM.The calculated volume of stock solution is added to the premixed solvents of ethanol and hexane with different volume ratios under sonication or stirring. a

Table S4 .
Sample preparation details for reversibly switching absorption and emission behaviors of TPE-Cy