Triphenylethylene- and Tetraphenylethylene-Functionalized 1,3-Bis

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Article Cite This: ACS Omega 2018, 3, 16424−16435

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Triphenylethylene- and Tetraphenylethylene-Functionalized 1,3Bis(pyrrol-2-yl)squaraine Dyes: Synthesis, Aggregation-Caused Quenching to Aggregation-Induced Emission, and Thiol Detection Ming Hui Chua,† Hui Zhou,† Ting Ting Lin,† Jishan Wu,*,‡ and Jianwei Xu*,† †

Institute of Materials Research and Engineering, Agency for Science, Technology and Research (A*STAR), 2 Fusionopolis Way, Innovis, #08-03, Singapore 138634 ‡ Department of Chemistry, National University of Singapore, 3 Science Drive 3, Singapore 117543

ACS Omega 2018.3:16424-16435. Downloaded from pubs.acs.org by 178.57.68.102 on 12/07/18. For personal use only.

S Supporting Information *

ABSTRACT: Three novel pairs of 1,3-bis(pyrrol-2-yl)squaraine dyes, Nalkylated SQ1a−1b, and N-phenylated SQ2a−2b in which triphenylethylene moieties functionalized at 5-position of pyrrole, as well as SQ3a−3b with tetraphenylethylene (TPE) moieties attached at N-position of pyrrole, were synthesized. All six dyes were found not to exhibit aggregation-induced emission (AIE) properties. Spectrophotometric studies showed that N-TPEfunctionalized SQ3a−3b exhibited much larger molar extinction coefficients (ε: 1.36−2.14 × 105 M−1 cm−1) than 5,5′-triphenylethylene-functionalized SQ1a−2b (ε: 2.17−8.22 × 104 M−1 cm−1). Surprisingly, SQ2b showed a remarkable red-shifted maximum absorption (λmax: 723 vs 631−652 nm) compared to that of other squaraine dyes. All six squaraine dyes selectively responded to the addition of thiol-containing biomolecules, such as cysteine and gluthatione, with the disappearance of λmax in the near-infrared region in their respective absorption spectra. Interestingly, the thiolated species of SQ3a−3b were AIE active, with the characteristic AIE emission of TPE at λmax = 484−490 nm upon addition of water. Further thiol sensing on solid supports was examined, indicating the potential applications of TPE-functionalized squaraine dyes as bioprobes for the detection of important thiol-containing biomolecules, with a clear change from aggregation-caused quenching to AIE.

1. INTRODUCTION Squaraine dyes are widely reported for a range of applications, particularly in the area of organic photovoltaics, chemosensing, supramolecular recognition, and self-assembly.1−7 Squaraine dyes often exhibit intense emission in the far-red to nearinfrared region, which is highly desirable for bioimaging applications due to the advantages of low phototoxicity, low autofluorescence background, and ability for deeper tissue penetration.8−12 In fact, there were several reports on squaraine dyes for bioimaging applications, including the use of its strong two-photon absorption properties.13−21 Similar to other common luminogens, 1,3-bis(aryl)squaraine dyes exhibit aggregation-caused quenching (ACQ) properties due to favorable π−π interaction between the planar squaraine core. This may thus limit their performance in fluorescencebased applications such as bioimaging.22 The limitations, however, may be overcome and application performances may be even enhanced with the use of aggregation-induced emission (AIE) luminogens, as widely reported.22−27 Many conventional ACQ luminogens have been successfully conferred the highly desirable AIE properties via intelligent structural modifications and functionalization.28−34 However, there are still no reports of AIE-active 1,3-bis(aryl)squaraine © 2018 American Chemical Society

dyes to date. This thus prompted us to attempt to synthesize 1,3-bis(aryl)squaraine dyes with AIE properties. Triphenylethylene and tetraphenylethylene (TPE) are AIEactive moieties useful in the structural design and synthesis of novel AIEgens.35−37 As such, we herein report the synthesis of a series of six novel 1,3-bis(pyrrol-2-yl)squaraine dyes, SQ1a− 3b, with triphenylethylene and TPE units functionalized at different positions. However, none of these were found to be AIE active. The central four-membered squarylium core of 1,3bis(aryl)squaraine dyes is reactive toward nucleophiles, as evident in reported applications of cyanide and biological thiol chemosensing.38−44 As such, the potentials of SQ1a−3b for cysteine (Cys) and glutathione (GSH) detection were investigated. Biological thiols such as Cys and GSH play important roles in many biological processes, and so their detection in biological samples can be crucial in the detection or indication of any physiological abnormality.45−51 To date, numerous novel molecular bioprobes were developed for the detection of Cys and GSH.52−60 Our synthesized squaraine Received: September 22, 2018 Accepted: November 19, 2018 Published: December 3, 2018 16424

DOI: 10.1021/acsomega.8b02479 ACS Omega 2018, 3, 16424−16435

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Scheme 1. Synthesis and Chemical Structures of Squaraine Dyes SQ1a−SQ3ba

a

Inserted: the single-crystal X-ray structure of compound SQ1a. Reagents and Conditions: (a) NaH, dimethylformamide (DMF) at room temperature for 20 min then 80 °C for 2 h; (b) NaO(t-Bu), Pd(OAc)2, P(t-Bu)3, toluene, reflux for 6 h; (c) toluene, 1-butanol, reflux 6−18 h; (d) (i) K2CO3, Pd(PPh3)4, toluene, H2O, ethanol, 100 °C for 18 h; then (ii) tetrabutylammonium fluoride (TBAF), tetrahydrofuran (THF), reflux for 18 h.

Figure 1. Normalized UV−vis absorption and emission spectra of SQ1a−SQ3b.

method,35 was first alkylated with n-hexyl and tri(ethylene glycol) monomethyl ether chains to give intermediates 5a and 5b, followed by condensation reaction with squaric acid to afford squaraine dyes SQ1a and SQ1b, respectively. For the synthesis of the pair of sterically more congested dyes, SQ2a and SQ2b, with additional phenyl groups attached at the Nposition of the pyrrole moieties, Buchwald−Hartwig amination reactions were performed on 4 to afford N-phenylated intermediates 6a and 6b, which then underwent condensation reaction with squaric acid to obtain SQ2a and SQ2b, respectively. For SQ1a−2b, triphenyethylene groups that are attached to the 5-positions of the pyrrole moieties, were in conjugation with the 1,3-bis(pyrrol-2-yl)squaraine core.

dyes were found to be reactive to thiol compounds, and showed great selectivity toward Cys and GSH. In addition, the thiolated species of N-TPE-functionalized SQ3a−3b were found to be AIE active, which could be used to further authenticate the presence of these biomolecules and thus additionally improve detection reliability and accuracy.

2. RESULTS AND DISCUSSIONS 2.1. Synthesis and Structures of Squaraine Dyes. The chemical structures and synthesis of the six novel 1,3bis(pyrrol-2-yl)squaraine dyes, SQ1a−SQ3b, are shown in Scheme 1. In this work, triphenylethylenyl-functionalized intermediate 4, which was prepared according to our reported 16425


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Table 1. Optical Properties of SQ1a−SQ3b Obtained from UV−vis Absorption and Emission Spectroscopy λabs (nm)a SQ1a SQ1b SQ2a SQ2b SQ3a SQ3b

640 652 648 723 631 639

ε (M−1 cm−1)c 76 160 42 930 21 710 82 220 213 580 135 490

λem (nm)b

Δν (cm−1)d

Φ (%)e

τ (ns)f

689 697 695 766 662 670

× × × × × ×

11.9 35.1 1.5 4.5 6.1 5.9

1.29 1.64 0.34 1.27 0.41 0.47

2.04 2.22 2.13 2.33 3.23 3.23

5

10 105 105 105 105 105

a Absorption maximum wavelength. bFluorescence maximum wavelength. cMolar extinction coefficient. dStokes’ shift. eQuantum yield in dilute THF solution, measured in reference to rhodamine B in ethanol (Φref = 97%), measurements subjected to ±10% error. fLifetime of fluorescence.

Figure 2. Response of fluorescence maxima intensities of SQ1a, SQ2a, and SQ3a (10 μM) in THF/water of different proportions and that of SQ1b, SQ2b, and SQ3b (10μM) in DMSO/water of different proportions. The corresponding sets of fluorescence spectra are provided in SI Figures S5 and S6.

The final pair squaraine dyes synthesized, SQ3a and SQ3b, have AIE-active TPE functionalized at the pyrrole units’ nitrogen atom (1-position) instead. Suzuki couplings were first performed to attach phenyl groups containing n-hexyl and tri(ethylene glycol) monomethyl ether chains to the 2-position of N-Boc pyrroles, followed by deprotonation with tetrabutylammonium fluoride (TBAF) to afford intermediates 7a and 7b. Buchwald−Hartwig amination reactions were then performed to attach TPE moieties to the N-position, hence giving intermediates 8a and 8b that underwent subsequent condensation reactions with squaric acid to obtain SQ3a and SQ3b, respectively. As compared with SQ1a−2b, the TPE groups attached to N-position of pyrroles of SQ3a−3b were not in the conjugation with the central squaraine core. The single-crystal structure of compound SQ1a was obtained as shown in Scheme 1 (inserted). Squaraine SQ1a adopts a trans configuration in the crystalline state with pyrrole moieties oriented in an opposite direction. The two pyrrole moieties lie in the same plane as the central squarylium unit, which is associated with and contributes to the high extent of electronic conjugation across the 1,3-bis(pyrrol-2-yl)squaraine core. Because of their steric bulkiness, the CC bonds of the triphenylethylenyl moieties were positioned out of the plane with a torsional angle of approximately 52° about the pyrrole units. 2.2. Optical Properties of Squaraine Dyes. The Ultraviolet−visible (UV−vis) absorption and emission spectra of the six squaraine dyes were measured in THF and shown in Figure 1, and the data of optical properties are summarized in Table 1. The absorption and emission profiles of N-TPE squaraines SQ3a and SQ3b were generally slightly blue-shifted

compared with other four squaraine dyes. In addition, SQ3a and SQ3b also exhibited significantly larger molar extinction coefficient about their absorption maxima (ε: 1.36−2.14 × 105 M−1 cm−1) whereas Stokes’ shift values were generally narrower at 31 nm, compared with the other triphenylethylenyl-functionalized squaraine dyes SQ1a−SQ2b (ε: 2.17− 8.22 × 104 M−1 cm−1; Δν: 2.04−2.33 × 105 cm−1). Surprisingly, SQ2b exhibited significantly red-shifted absorption and emission profiles compared to those of SQ2a, as well as of the other squaraine dyes. SQ1b with tri(ethylene glycol) chains shows a bigger absorption λmax than SQ1a with long alkyl chains (Δλmax = 12 nm) although the difference is much less significant in comparison with the counterparts of SQ2a and SQ2b (absorption Δλmax = 75 nm). However, the reason for such a huge difference is not clear. The six dyes were found to be emissive in organic solvents. The fluorescence quantum yields of the N-alkylated SQ1a and SQ1b were amongst the highest at 11.9 and 35.1%, respectively, whereas the quantum yields of the N-phenylated SQ2a and SQ2b were amongst the lowest, at 1.5 and 4.5%, respectively. The relatively lower fluorescence quantum yields for the latter pair are due to the presence of an extra phenyl ring, which with intramolecular rotation in solution contributes to nonradiative decay of excited state. On the other hand, the N-phenylated attachments also deteriorated the co-planarity of the whole molecule and thus further lowered the fluorescence quantum efficiency. Similarly, intramolecular rotation of the Nfunctionalized TPE moieties of SQ3a and SQ3b also contribute much to the lower quantum yields of 6.1 and 5.9%, respectively. 16426


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Figure 3. DFT-calculated optimized structures of compounds SQ1a−SQ3b. Extended portions of hexyl and tri(ethylene glycol) monomethyl ether chains were omitted for simplicity.

2.3. ACQ Properties of SQ1a−SQ3b. Investigation into possible AIE properties in the six synthesized squaraine dyes was performed in both THF/water and dimethyl sulfoxide (DMSO)/water solvent combinations. In spite of the presence of AIE-active triphenylethylene and TPE groups, none of the squaraine dyes synthesized exhibit AIE properties, as shown in Figure 2. Although twisted intramolecular charge transfer might partially be responsible for fluorescence quenching at low-to-moderate proportions of water, dynamic light scattering studies confirmed the formation of aggregates at high proportions of water where dye fluorescence was effectively quenched, thus confirming their ACQ properties (Supporting Information (SI), Figures S7 and S8). From the single-crystal packing structure obtained for squaraine SQ1a, it was observed that the presence of bulky triphenylethylene groups managed to prevent close π−π interactions of the planar 1,3-bis(pyrrol-2-yl)squaraine cores, with a sufficiently large interplanar distance of 9.174 Å measured (SI Figure S1). Even so, this appears to be insufficient to confer the dye AIE properties. Nonetheless, it would be hard to explain the ACQ nature of these squaraine dyes based on the single-crystal packing structure of SQ1a alone. 2.4. Density Functional Theory (DFT) Calculations. Density functional theory (DFT) calculations at B3LYP/631G level of theory, implemented in Gaussian 09 program, were performed for SQ1a−SQ3b.61 The calculated optimized structures of the six compounds (Figure 3) suggest the planarity about the squaraine core, i.e., two pyrrole centers and the central squarylium moieties, whereas the bulky triphenylethylene and TPE moieties deviate from the central squaraine plane. In addition, phenyl rings functionalized at the pyrroles’ nitrogen atoms (SQ2a−SQ3b) were also calculated to be almost orthogonal to the planar core, which translates to minimal electronic communication with the central squaraine core.

Both the highest occupied molecular orbital (HOMO) and lowest unoccupied molecular orbital (LUMO) profiles for all six dyes were calculated to be evenly delocalized across the central 1,3-bis(pyrrol-2-yl)squaraine core (SI Figure S9). The orbital profiles were extended to the ethylene bonds of triphenylethylene moieties for SQ1a−SQ2b and the phenyl rings at the 5,5′-positions of pyrrole moieties of SQ3a−3b, whereas the N-functionalized phenyl rings and TPE units of SQ2a−2b and SQ3a−3b, respectively, were not involved. Time-dependent-DFT calculations performed for all six dyes in THF solvent (SI Figure S11) suggest that the NIR absorption bands from 600−725 nm measured were mainly attributed to squaraine-centered HOMO → LUMO transitions although the calculated λmax appeared blue-shifted to a different extent compared with actual measurements. 2.5. Reaction of SQ1a−SQ3b with Thiol Compounds. The central squarylium moiety of 1,3-bis(aryl)squaraine dyes is known to be susceptible to nucleophile attacks with applications such as cyanide and thiol chemosensors.38−44 Scheme 2 shows the mechanism of reaction between 1,3bis(pyrrol-2-yl)squaraines with thiol compounds. Thiol nucleophiles can undergo nucleophilic addition to the 2position of the central squarylium core, effectively breaking the electronic conjugation with one of the pyrrole units. A corresponding color change (often bleaching of intense coloration) will prevail. Associated fluorescence changes may be observed as well. Scheme 2. Mechanism of Reaction Between Thiol Compounds and 1,3-Bis(pyrrol-2-yl)squaraine Dyes

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Figure 4. (a) Normalized absorption spectra of SQ3a and SQ3b before and after thiolation with inserted photos of the sample solutions. Fluorescence spectra (λex = 300 nm) of thiolated solutions of SQ3a (b) and SQ3b (c) in DMSO/water of different proportions with inserted photos of the respective series of solutions taken under UV irradiation (302 nm). The responses of the respective fluorescence maximum intensities with percentage of water are plotted in (d).

proportion of water (SI Figures S14 and S15). Interestingly, AIE properties were observed for thiolated SQ3a and SQ3b solutions, as shown in Figure 4b−d, with the emergence of 484 and 490 nm emission peaks, respectively, as the proportion of water was increased. The peaks emerged at 20 and 30% water for SQ3a and SQ3b, respectively, thereafter dipping slightly at 40% water, probably attributable to change in aggregate particle size. Fluorescence of thiolated SQ3a continued to increase after 40% water, whereas fluorescence of SQ3b remains relatively steady. The origins of these two peaks were believed to be due to N-functionalized TPE units, whose optimized structures (SI Figure S12) are orthogonal to the attached pyrrole units. As shown, the enhancement of blue emission for thiolated SQ3a was more pronounced than that for SQ3b, probably due to the former bearing hydrophobic hexyl chains, hence favoring a greater extent of aggregation, whereas the latter contains more hydrophilic tri(ethylene glycol) chains, hence a lesser extent of aggregation. 2.6. Selectivity Test and Thiol Sensing on a Solid Support. The selectivity of SQ1b, SQ2b, and SQ3b toward biologically active thiol compounds was investigated. A large excess (100 molar equivalence) of all 20 primary amino acids and also GSH was added to the three dyes (10 μM) in DMSO/phosphate buffer solution (PBS) solution 1:1 v/v (pH = 7.4). Delightfully, all three squaraine dyes were found to selectively react with Cys and GSH, of which obvious

Despite the lack of AIE properties, the potentials of SQ1a− SQ3b for thiol sensing were investigated. More importantly, we predict that the breaking of the central planar 1,3bis(pyrrol-2-yl)squaraine core due to nucleophilic attack by thiol would disrupt the π−π stacking interactions (although weak but may still prevail in the compounds), which may be responsible for their ACQ behavior. As a result, the resultant nonplanar thiolated species may thus exhibit AIE properties. To begin with, compounds SQ1a−SQ3b were reacted with a large excess (100 molar equivalence) of decanethiol in DMSO solvent at 1 mM concentration. Stirring at 55 °C and under constant monitoring by thin-layer chromatography (TLC), SQ1a−SQ3b was found to fully react with decanethiol between 0.3 and 4 h. The absorption profile of the thiolated species of SQ1a−SQ3b was measured with clear disappearance of the intense absorption peak in the region of 600−725 nm (SI Figure S13). Figure 4a shows the normalized absorption spectra for SQ3a and SQ3b before and after thiolation. Without further purifications, the thiolated solutions were then diluted 10-fold with DMSO/water solvents of different proportions. Fluorescence spectroscopy was performed and the responses of maximum fluorescence intensities in relation to proportion of water were plotted (SI Figures S14 and S15). The species in solutions of thiolated SQ1a−SQ2b show ACQ behavior with fluorescence quenching on increasing 16428


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Figure 5. Absorption spectra of N-TPE squaraine SQ3b solutions (10 μM, in DMSO/PBS solution 1:1 v/v, pH = 7.4) in the presence of 100 molar equivalence of the 20 amino acids and glutathione. Inserts are photos of the respective solutions.

Figure 6. Cysteine and glutathione-coated TLC plates after dipping into 0.1 mM solutions of SQ1b, SQ2b, and SQ3b observed under visible light and under UV irradiation (365 nm). Water was used as a blank.

SQ3b, by extending the π-conjugation backbone, hence further red-shifting the emission wavelength to the NIR region and/or replacing TPE with other AIEgens with higher aggregation quantum yield.

decolorizations were visually observed and significant decreases in absorption intensities were recorded (SI Figure S19). Figure 5 shows the selectivity test for SQ3b. Interestingly, it was also observed that the presence of excess glutamic acid, tryptophan, and tyrosine consistently caused redshift and broadening of absorption λmax of the three squaraine dyes (Figure 5 and SI Figure S19). To demonstrate the possibility of the dyes, particularly NTPE SQ3b, for thiol sensing in solid biological samples, a simple study involving Cys- and GSH-coated TLC plates was performed. The thiol-coated TLC plates were briefly dipped into 0.1 mM solutions of SQ1b, SQ2b, and SQ3b and thereafter dried over warm air. Although the dyes stained the TLC plates, decolorization was observed on the spots where thiols were applied, as shown in Figure 6. Under UV irradiation (365 nm), SQ3b-stained TLC plates emitted red fluorescence, whereas thiol-coated spots emitted yellow fluorescence. Neither decolorization nor fluorescence was observed on water-coated (blank) spots. TLC plates were coated with Cys and GSH solutions with different concentrations, dried, and then dipped into SQ3b solution. Decolorization was observed to become fainter with decreasing thiol concentration (SI Figure S20). Under UV light, spots coated with higher concentrations of thiols (≥1 mM) were observed to exhibit pale yellow emission whereas spots coated with lower concentrations (≤0.5 mM) of thiols exhibited no fluorescence. However, the red background fluorescence resulting from SQ3b appeared to be emitted brighter than the yellow fluorescence on thiol-coated spots. Nonetheless, this could be circumvented and hence emission contrast improved, with possible modifications to the structural framework of

3. CONCLUSIONS In summary, six novel 1,3-bis(pyrrol-2-yl)squaraine dyes were synthesized with AIE-active triphenylethylenyl and TPE moieties functionalized at the pyrroles’ 2- and N-positions, respectively. Amongst them, SQ3a−3b exhibited an extremely large molar absorptivity of 1.36−2.14 × 105 M−1 cm−1, whereas SQ2b exhibited surprisingly red-shifted absorption and emission λmax compared with the rest. None of the dyes were found to exhibit AIE properties. The dyes were, however, found to react with decanethiol in DMSO with observable color changes supported by a diminishing of absorption λmax in spectrophotometric studies, and only the thiolated species of N-TPE SQ3a and SQ3b were found to exhibit AIE properties. Selectivity studies performed also confirmed that SQ1b, SQ2b, and SQ3b were selective toward biologically active thiols, Cys, and GSH. The dyes also showed promising results for the detection of thiols coated on solid support. More importantly, through these preliminary findings, we have presented a novel approach of a bioprobe system that triggers AIE from ACQ by a reaction with thiol compounds, which could be further explored and extended to different applications in the future. 4. EXPERIMENTAL SECTION 4.1. General Methods and Instrumentations. All reagents, solvents, and starting materials were obtained from commercial suppliers and used without further purifications 16429


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resolution mass spectra, HRMS (APCI, m/z): [M]+ calculated for C30H32N1, 406.2529; found, 406.2537. 4.2.2. Compound 5b: 1-(2-(2-(2-Methoxyethoxy)ethoxy)ethyl)-2-(1,2,2-triphenylvinyl)-1H-pyrrole. Compound 5b was synthesized via the same procedure as described above in compound 5a synthesis, with compound 4 (100 mg, 0.31 mmol) deprotonated by sodium hydride (60% w/w in grease, 15.6 mg, 0.39 mmol), followed by a nucleophilic substitution reaction with tri(ethylene glycol) monomethyl ether tosylate (148.5 mg, 0.47 mmol). The crude product was purified via column chromatography using EA/hexane (2:3 v/v) as eluent to obtain the product as a yellow gel (125 mg, 86% yield). 1 H NMR (400 MHz, CDCl3, δ): 3.29 (m, 2H), 3.37 (s, 3H), 3.47 (m, 2H), 3.53 (m, 2H), 3.58−3.64 (m, 6H), 5.78 (d, J = 4 Hz, 1H), 6.02 (d, J = 4 Hz, 1H), 6.63 (d, J = 4 Hz, 1H), 6.98− 7.13 (m, 15H). 13C NMR (100 MHz, CDCl3, δ): 46.57, 59.04, 70.42, 70.55, 70.57, 71.93, 107.92, 111.82, 121.43, 126.48, 126.53, 127.54, 127.62, 127.73, 130.35, 130.89, 131.50, 132.02, 134.66, 142.19, 142.65, 143.17, 144.01. HRMS (APCI, m/z): [M]+ calculated for C31H34N1O3, 468.2533; found, 468.2528. 4.2.3. Compound 6a: 1-(4-Hexylphenyl)-2-(1,2,2-triphenylvinyl)-1H-pyrrole. In a flame-dried Schlenk tube, compound 4 (100.0 mg, 0.31 mmol), 1-bromo-4-hexylbenzene (113 mg, 0.47 mmol), sodium tert-butoxide (60 mg, 0.62 mmol), and palladium acetate (57 mg, 0.062 mmol) were added, followed by being evacuated and backfilled with argon gas three times. Anhydrous toluene (5.0 mL) was then added, followed by tri-tert-butylphosphine (10 wt % in hexane, 56 μL, 0.19 mmol). The reaction mixture was allowed to stir at reflux conditions for 6 h. The reaction mixture was then cooled to room temperature and filtered via vacuum filtration to remove insoluble black by-products. The filtrate was washed with water and then extracted with ethyl acetate. The combined organic layer was then dried over magnesium sulfate and evaporated using a rotary evaporator. The crude product was purified via column chromatography using EA/hexane (1:50−3:50 v/v) as eluent to obtain the product as a yellow solid (115 mg, 77% yield). 1 H NMR (400 MHz, CDCl3, δ): 0.85 (t, J = 8 Hz, 3H), 1.25−1.30 (m, 6H), 1.51 (m, 2H), 2.49 (t, J = 8 Hz, 2H), 6.02 (d, J = 4 Hz, 1H), 6.22 (t, J = 4 Hz, 1H), 6.49 (d, J = 8 Hz, 2H), 6.67 (d, J = 4 Hz, 1H), 6.67 (d, J = 8 Hz, 2H), 6.89−7.42 (m, 17H). 13C NMR (100 MHz, CDCl3, δ): 14.07, 22.59, 28.73, 31.51, 31.68, 35.25, 108.92, 114.07, 122.11, 123.76, 125.90, 126.26, 126.42, 127.15, 127.31, 127.59, 128.23, 130.32, 131.29, 131.37, 134.51, 137.67, 140.30, 143.21, 143.57, 143.78. HRMS (APCI, m/z): [M]+ calculated for C36H36N1, 482.2842; found, 482.2838. 4.2.4. Compound 6b: 1-(4-(2-(2-(2-Methoxyethoxy)ethoxy)ethoxy)phenyl)-2-(1,2,2-triphenylvinyl)-1H-pyrrole. Compound 6b was synthesized via the same procedure as described above in compound 6a synthesis, from compound 4 (250 mg, 0.78 mmol), 1-bromo-4-(ethoxy-2-tri(ethylene glycol) monomethyl ether)benzene (370 mg, 1.17 mmol), sodium tert-butoxide (150 mg, 1.56 mmol), tri-tert-butylphosphine (10 wt % in hexane, 1.4 mL, 0.47 mmol), and palladium acetate (143.0 mg, 0.16 mmol). The crude product was purified via column chromatography using EA/hexane (1:1− 3:5 v/v). The pure product, however, could not be separated from unreacted 1-bromo-4-(ethoxy-2-tri(ethylene glycol) monomethyl ether)benzene. A yellow gel was obtained (122.0 mg), and HRMS confirms presence of the product. HRMS (APCI, m/z): [M]+ calculated for C37H38N1O4,

unless stated otherwise. Anhydrous toluene and tetrahydrofuran (THF) used for synthesis were purified via distillation under nitrogen atmosphere over sodium with benzophenone. Preparative separations and purifications of synthesized products were performed using column chromatography on silica gel grade 60 (Merck 0.040−0.063 mm, 230−400 mesh) purchased from Sigma-Aldrich. 1H and 13C nuclear magnetic resonance (NMR) spectra of compounds were recorded in deuterated solvents purchased from a commercial supplier with tetramethylsilane as internal standard. 1H and 13C NMR spectra were recorded using a Bruker 400 MHz NMR Spectrometer at 25 °C, and chemical shifts are expressed with a positive sign, in parts per million, relative to residual solvent signals as reference. Electron impact mass spectra were recorded using a Finnigan TSQ 7000 mass spectrometer, whereas electrospray ionization and atmospheric pressure chemical ionization (APCI) mass spectra were recorded from a Bruker micrOTOF-Q II. Both NMR and mass spectroscopy were used to identify and confirm structures of intermediates synthesized. Ultraviolet−visible (UV−vis) absorption spectra were recorded using a Shimadzu UV3600 UV−vis−NIR spectrophotometer, whereas photoluminescence (fluorescence) spectra and lifetime were obtained using a HORIBA Jobin Yvon Fluorolog spectrofluorometer where readings were recorded in units of counts-per-second per amperes (CPS/Amp). Dynamic light scattering of aggregate solutions was performed using Malvern Zetasizer Nano ZS. HPLC grade solvent, purchased from a commercial supplier, as well as freshly purified deionized (DI) water, was used to prepare sample solutions for the above photophysical characterizations. 4.2. Synthetic Procedures and Characterization Data. Intermediate compound, 1-bromo-4-(ethoxy-2-tri(ethylene glycol) monomethyl ether)benzene, was synthesized from 4bromophenol and tri(ethylene glycol) monomethyl ether with K2CO3 base, in DMF solvent, in accordance to reported procedures.62 4.2.1. Compound 5a: 1-Hexyl-2-(1,2,2-triphenylvinyl)-1Hpyrrole. In a flame-dried Schlenk tube, compound 4 (100.0 mg, 0.3 1 mmol) was dissolved in 5 mL of anhydrous DMF under inert argon gas atmosphere. Under stirring at room temperature, sodium hydride (60% w/w in grease, 15.6 mg, 0.39 mmol) was added and the reaction mixture was allowed to stir for 20 min. 1-Bromohexane (77 mg, 0.47 mmol) was dissolved in 1 mL of anhydrous DMF, and the solution added dropwise to the reaction mixture on stirring. The reaction mixture was then heated to 80 °C and allowed to stir for another 2 h, with constant monitoring by TLC. The reaction mixture was then cooled to room temperature and quenched by adding dropwise to a beaker of ice water. The mixture was then extracted with ethyl acetate (EA). The combined organic layer was washed with brine solution, dried over magnesium sulfate, and evaporated using a rotary evaporator. The crude product was purified via column chromatography using dichloromethane (DCM)/hexane (1:3 v/v) as eluent to obtain the product as a yellow gel (115 mg, 91% yield). 1 H NMR (400 MHz, CDCl3, δ): 0.85 (t, J = 8 Hz, 3H), 1.08−1.26 (m, 6H), 1.45 (quint. J = 8 Hz, 2H), 3.35 (t, J = 8 Hz, 2H), 5.77 (d, J = 4 Hz, 1H), 6.01 (d, J = 4 Hz, 1H), 6.98− 7.14 (m, 16H). 13C NMR (100 MHz, CDCl3, δ): 14.00, 22.53, 26.51, 30.96, 31.39, 47.27, 107.51, 112.01, 120.04, 126.45, 127.50, 127.57, 127.50, 127.57, 127.72, 130.33, 130.94, 131.54, 132.34, 134.43, 141.72, 142.62, 143.36, 133.24. High16430


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(150 mg, 0.66 mmol), 1-(4-bromophenyl)-1,2,2-triphenylethylene (407 mg, 0.99 mmol), sodium tert-butoxide (127 mg, 1.32 mmol), tri-tert-butylphosphine (10 wt % in hexane, 120 μL, 0.40 mmol), and palladium acetate (121 mg, 0.13 mmol). The crude product was purified via column chromatography using DCM/hexane (1:10−1:5 v/v) as eluent, then washed with n-hexane to obtain the product as a yellow solid (125 mg, 34% yield). 1 H NMR (400 MHz, CDCl3, δ): 0.89 (t, J = 8 Hz, 3H), 1.33 (m, 6H), 1.61−1.63 (m, 2H), 2.57 (t, J = 8 Hz, 2H), 6.31 (d, J = 4 Hz, 1H), 6.37 (d, J = 4 Hz, 1H), 6.89 (d, J = 8 Hz, 2H), 6.96 (d, J = 8 Hz, 2H), 7.00−7.12 (m, 20H). 13C NMR (100 MHz, CDCl3, δ): 14.10, 22.62, 29.09, 31.31, 31.75, 35.68, 109.01, 110.14, 123.76, 124.94, 126.56, 127.69, 127.97, 128.12, 130.23, 131.28, 131.80, 133.85, 138.69, 140.09, 140.88, 141.43, 142.07, 143.33, 143.46, 143.54. HRMS (APCI, m/z): [M]+ calculated for C42H40N1, 558.3155; found, 558.3151. 4.2.8. Compound 8b: 2-(4-(2-(2-(2-Methoxyethoxy)ethoxy)ethoxy)phenyl)-1-(4-(1,2,2-triphenylvinyl)phenyl)1H-pyrrole. Compound 8b was synthesized via the same Buchwald−Hartwig amination procedure as that for the synthesis of compound 6a, from compound 7b (600 mg, 1.96 mmol), 1-(4-bromophenyl)-1,2,2-triphenylethylene (822 mg, 1.98 mmol), sodium tert-butoxide (360 mg, 0.39 mmol), tri-tert-butylphosphine (10 wt % in hexane, 1.6 mL, 1.18 mmol), and palladium acetate (359 mg, 0.39 mmol). The crude product was then purified via column chromatography using EA/DCM (1:100−1:20 v/v) as eluent to obtain the product as a yellow gel (170 mg, 14% yield). 1 H NMR (400 MHz, CDCl3, δ): 3.37 (s, 3H), 3.55 (m, 2H), 3.65−3.70 (m, 4H), 3.76 (m, 2H), 3.88 (t, J = 4 Hz, 2H), 4.13 (t, J = 4 Hz, 2H), 6.31 (m, 2H), 6.76 (d, J = 9 Hz, 2H), 6.87 (d, J = 9 Hz, 2H), 6.96−7.13 (m, 20H). 13C NMR (100 MHz, CDCl3, δ): 59.03, 67.37, 69.79, 70.60, 70.70, 70.85, 71.95, 108.94, 109.70, 114.14, 123.42, 124.92, 125.50, 126.58, 127.68, 129.51, 131.26, 131.78, 133.10, 138.63, 141.47, 142.06, 143.28, 143.44, 143.55, 157.39. HRMS (APCI, m/z): [M]+ calculated for C43H42N1O4, 636.3108; found, 636.3104. 4.2.9. Compound SQ1a: 1,3-Bis(1-hexyl-5-(1,2,2-triphenylvinyl)-1H-pyrrol-2-yl)squaraine. In a flame-dried Schlenk tube, compound 5a (115 mg, 0.28 mmol) and squaric acid (16.2 mg, 0.14 mmol) were added and the tube was evacuated and backfilled with argon gas three times. Anhydrous toluene (3 mL) was added, followed by 1-butanol (1 mL), and the reaction mixture was allowed to stir at reflux for 6 h. After cooling to room temperature, the solvent was then removed via a rotary evaporator and the crude product was purified via column chromatography using EA/hexane (1:6 v/v) as eluent to isolate the product as a shiny green solid (103 mg, 41% yield). 1 H NMR (400 MHz, CDCl3, δ): 0.83 (t, J = 7 Hz, 6H), 1.16 (m, 10H), 1.52−1.57 (m, 6H), 4.15 (m, 4H), 6.07 (d, J = 4 Hz, 2H), 6.99−7.16 (m, 30H), 7.63 (d, J = 4 Hz, 2H). 13C NMR (100 MHz, CDCl3, δ): 14.06, 22.59, 26.26, 31.57, 31.88, 47.79, 120.09, 123.17, 127.49, 127.89, 128.14, 128.26, 129.50, 130.11, 130.44, 130.69, 131.44, 140.05, 141.93, 142.85, 146.77, 150.44. HRMS (APCI, m/z): [M] + calculated for C64H61N2O2, 889.4728; found, 889.4737. 4.2.10. Compound SQ1b: 1,3-Bis(1-(2-(2-(2methoxyethoxy)ethoxy)ethyl)-5-(1,2,2-triphenylvinyl)-1Hpyrrol-2-yl)squaraine. Compound SQ1b was synthesized via the same condensation reaction procedure as that described for the synthesis of compound SQ1a, using compound 5b (120

560.2795; found, 560.2796. Compound 6b was used as such in the subsequent synthesis step. 4.2.5. Compound 7a: 2-(4-Hexylphenyl)-1H-pyrrole. 4.2.5.1. Step 1: Suzuki Coupling. A flame-dried two-neck round bottom flask (RBF) fitted with a condenser was charged with N-Boc-pyrrole-2-boronic acid (500 mg, 2.37 mmol), 1bromo-4-hexylbenzene (474 mg, 1.97 mmol), potassium carbonate (1.64 g, 12 mmol), and palladium(0) tetrakistriphenylphospine (Pd(PPh3)4) catalyst (137 mg, 0.12 mmol) and evacuated and backfilled with argon gas three times. Toluene (30 mL), ethanol (5 mL), and deionized water (5 mL) and the reaction mixture were degassed thoroughly. The reaction mixture was then allowed to stir at 100 °C for 18 h. On cooling to room temperature, the reaction was quenched by adding saturated aqueous sodium hydrogen carbonate solution. The mixture was extracted with ethyl acetate, and the combined organic layer dried over magnesium sulfate and then evaporated using a rotary evaporator. The crude product was purified via column chromatography using DCM/hexane (1:4 v/v) as eluent to isolate the product as a colorless gel (500 mg, 65% yield). 4.2.5.2. Step 2: Deprotection. A flame-dried two-neck RBF fitted with a condenser was charged with the intermediate (500 mg, 1.53 mmol) isolated, as mentioned above, and evacuated and backfilled with argon gas three times. Anhydrous THF (10 mL) was added to dissolve the reactant, followed by tetrabutylammonium fluoride (1 M solution in THF, 7.63 mL, 7.63 mmol). The reaction mixture was allowed to stir at reflux condition for 18 h. On cooling to room temperature, the solvent was removed via a rotary evaporator and the crude product was purified via column chromatography using DCM/ hexane (1:1 v/v) as eluent to obtain the product as a white solid (275 mg, 79% yield). 1 H NMR (400 MHz, acetone-d6, δ): 0.88 (t, J = 8 Hz, 3H), 1.32 (m, 6H), 1.61 (t, J = 8 Hz, 2H), 2.59 (t, J = 8 Hz, 2H), 6.14 (dd, J = 4 Hz, 1H), 6.46 (t, J = 4 Hz, 1H), 6.82 (dd, J = 4 Hz, 1H), 7.18 (d, J = 8 Hz, 2H), 7.53 (d, J = 8 Hz, 2H), 10.42 (broad, 1H). 13C NMR (100 MHz, acetone-d6, δ): 14.85, 23.80, 29.77, 32.79, 32.97, 36.64, 106.39, 110.53, 119.94, 124.89, 130.08, 132.42, 133.22, 141.48. HRMS (APCI, m/z): [M]+ calculated for C16H22N1, 228.1747; found, 228.1748. 4.2.6. Compound 7b: 2-(4-(2-(2-(2-Methoxyethoxy)ethoxy)ethoxy)phenyl)-1H-pyrrole. Compound 7b was synthesized via the same procedure as described above for the synthesis of compound 7a, first via Suzuki coupling with NBoc-pyrrole-2-boronic acid (875 mg, 4.15 mmol), 1-bromo-4(ethoxy-2-tri(ethylene glycol) monomethyl ether)benzene (1.1 g, 3.46 mmol), potassium carbonate (2.39 g, 17.3 mmol), and Pd(PPh3)4 catalyst (200 mg, 0.17 mmol), of which the crude product was purified via column chromatography using EA/ hexane (1:1 v/v) as eluent to isolate the intermediate as a colorless liquid (1.0 g, 71% yield). Next, by deprotecting the intermediate isolated (1 g, 2.47 mmol) with tetrabutylammonium fluoride (1 M solution in THF, 12.5 mL, 12.5 mmol), the crude product was briefly purified via flash column chromatography using EA as eluent to obtain the product as a colorless sticky solid. Because of its chemical instability, the product was used as such for the next step without any further purification or characterization (600 mg, 79% yield). 4.2.7. Compound 8a: 2-(4-Hexylphenyl)-1-(4-(1,2,2triphenylvinyl)phenyl)-1H-pyrrole. Compound 8a was synthesized via the same Buchwald−Hartwig amination procedure as that for the synthesis of compound 6a, from compound 7a 16431


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35.81, 115.79, 124.26, 126.60, 127.68, 127.75, 128.35, 128.47, 131.37, 131.67, 132.19, 136.41, 140.23, 141.28, 143.33, 143.65, 143.74, 144.02, 144.95, 148.19. HRMS (APCI, m/z): [M]+ calculated for C88H77N2O2, 1193.5980; found, 1193.5984. 4.2.14. Compound SQ3b: 1,3-Bis(5-(4-(2-(2-(2methoxyethoxy)ethoxy)ethoxy)phenyl)-1-(4-(1,2,2triphenylvinyl)phenyl)-1H-pyrrol-2-yl)squaraine. Compound SQ3b was synthesized via the same condensation reaction procedure as that described for the synthesis of compound SQ1a, using compound 8b (170 mg, 0.27 mmol) and squaric acid (15 mg, 0.13 mmol). The reaction time was 18 h. The crude product was purified slowly via column chromatography using EA/DCM (0:1−1:1 v/v) as eluent to isolate the product as a dark blue solid (46 mg, 13% yield). 1 H NMR (400 MHz, CDCl3, δ): 3.38 (s, 6H), 3.55 (m, 4H), 3.66 (m, 4H), 3.70 (m, 4H), 3.77 (m, 4H), 3.89 (t, J = 5 Hz, 4H), 4.16 (t, J = 5 Hz, 4H), 6.70 (d, J = 4 Hz, 2H), 6.79 (d, J = 9 Hz, 4H), 7.02−7.17 (m, 42H), 8.00 (d, J = 4 Hz, 2H). 13C NMR (100 MHz, CDCl3, δ): 59.05, 67.50, 69.45, 70.61, 70.70, 70.71, 71.95, 114.44, 115.42, 123.12, 124.21, 126.52, 126.61, 127.75, 128.40, 130.02, 131.37, 131.67, 131.74, 132.20, 136.44, 140.22, 141.33, 143.32, 143.66, 143.74, 144.92, 147.85, 159.29. HRMS (APCI, m/z): [M]+ calculated for C90H81N2O10, 1349.5886; found, 1349.5875. 4.3. Reaction of SQ1a−SQ3b with Thiols. 4.3.1. Procedure for Reaction of SQ1a−SQ3b with Decanethiol in DMSO Solvent. Each of the six dyes was reacted with a large excess (100 molar equivalence) of decanethiol in DMSO. In 4 mL glass vials, 250 μL of each squaraine dye solution in DMSO (1 mM) was added with 250 μL of predissolved decanethiol solution (100 mL in DMSO) and then topped up with 2000 μL of solvent. The solutions were stirred using a magnetic stirrer at 55 °C, under constant monitoring via thinlayer chromatography (TLC), until the reactions were complete. Solution of SQ2b changed from green to dull pale orange, whereas decolorization of the intense blue coloration was observed for the rest of the dyes on complete reaction with decanethiol. The time taken for SQ1a, SQ1b, SQ2a, SQ2b, SQ3a, and SQ3b to completely react with decanethiols in DMSO was 4, 3, 0.3, 1, 2, and 2 h, respectively. 4.3.2. Procedure for Investigation of AIE Properties in Thiolated Solutions of SQ1a−SQ3b. The solution mixtures containing the thiolated species were diluted, without further purification, with DMSO and water, at different proportions (to a 10 μL concentration, 2 mL volume) following standard AIE-property investigation procedures. To a clean glass vial, 200 μL of concentrated thiolated solution was added using a micropipette. Next, DMSO was added and then under stirring condition, water was added slowly for aggregation to occur. The eventual solution was stirred and mixed thoroughly with the use of a vortex. The volume of water (in microliters) to be added is related to the proportion of water, f w (in %), via the relationship: the volume of water = 2000 (f w/100). The emission spectra of the series of solutions were thus measured. UV−vis absorption spectra of the diluted thiolated solutions were recorded as well. 4.4. Detection of Biologically Active Thiols. 4.4.1. Selectivity Test. All 20 primary amino acids and glutathione (GSH) used were purchased from Sigma-Aldrich and used without any further purification. They were predissolved as stock solutions of 10 mM concentration in deionized water, with the addition of catalytic amount of trifluoroacetic acid when necessary to aid in dissolution. PBS (1.0 M, pH = 7.4)

mg, 0.26 mmol) and squaric acid (14.6 mg, 0.13 mmol). The reaction time was 6 h. The crude product was purified via column chromatography using EA/hexane (3:2 v/v) as eluent to isolate the product that exists as a dark turquoise solid (115 mg, 43% yield). 1 H NMR (400 MHz, CDCl3, δ): 2.90−3.29 (m, 30H), 5.70 (d, J = 4 Hz, 2H), 6.66−6.78 (m, 30H), 7.26 (d, J = 4 Hz, 2H). 13 C NMR (100 MHz, CDCl3, δ): 47.65, 58.97, 70.28, 70.38, 70.51, 71.87, 120.25, 127.37, 127.46, 127.52, 127.59, 127.71, 127.82, 128.03, 128.09, 130.30, 130.32, 130.48, 130.66, 131.14, 131.52, 142.13, 142.65, 146.44, 152.07. HRMS (APCI, m/z): [M] + calculated for C 66 H 65 N 2 O 8 , 1013.4735; found, 1013.4731. 4.2.11. Compound SQ2a: 1,3-Bis(1-(4-hexylphenyl)-5(1,2,2-triphenylvinyl)-1H-pyrrol-2-yl)squaraine. Compound SQ2a was synthesized via the same condensation reaction procedure as that described for the synthesis of compound SQ1a, using compound 6a (112 mg, 0.23 mmol) and squaric acid (13 mg, 0.12 mmol). The reaction time was 24 h. The crude product was purified via column chromatography using EA/hexane (1:6 v/v) as eluent to isolate the product as a dark green solid (86 mg, 36% yield). 1 H NMR (400 MHz, CDCl3, δ): 0.85 (t, J = 8 Hz, 6H), 1.51−1.58 (m, 12H), 1.85 (m, 4H), 2.54 (t, J = 8 Hz, 4H), 6.16 (d, J = 4 Hz, 2H), 6.73−7.16 (m, 38H), 7.65 (d, J = 4 Hz, 2H). 13C NMR (100 MHz, CDCl3, δ): 14.10, 22.61, 28.83, 31.44, 31.75, 35.62, 119.31, 123.70, 126.74, 127.17, 127.32, 127.62, 128.09, 129.84, 130.11, 130.39, 130.63, 131.20, 135.32, 140.69, 142.69, 142.03, 142.50, 142.61, 146.33, 151.27. HRMS (APCI, m/z): [M]+ calculated for C76H69N2O2, 1041.5354; found, 1041.5347. 4.2.12. Compound SQ2b: 1,3-Bis(1-(4-(2-(2-(2methoxyethoxy)ethoxy)ethoxy)phenyl)-5-(1,2,2-triphenylvinyl)-1H-pyrrol-2-yl)squaraine. Compound SQ2b was synthesized via the same condensation reaction procedure as that described for the synthesis of compound SQ1a, using compound 6b (100 mg, 0.18 mmol) and squaric acid (10 mg, 0.09 mmol). The reaction time was 18 h. The crude product was purified slowly via column chromatography using EA/DCM (1:5 v/v) as eluent to isolate the product as a dark green solid (23 mg, 11% yield). 1 H NMR (400 MHz, CD2Cl2, δ): 3.31 (s, 6H), 3.48 (m, 4H), 3.56−3.66 (m, 12H), 3.80 (t, J = 4 Hz, 4H), 4.12 (t, J = 4 Hz, 4H), 5.94 (s, 2H), 6.85 (d, J = 8 Hz, 4H), 7.08−7.34 (m, 36H). 13C NMR (100 MHz, CD2Cl2, δ): 59.04, 67.92, 70.06, 70.88, 70.95, 71.21, 72.35, 114.42, 118.83, 124.79, 126.32, 127.70, 128.10, 128.70, 129.17, 130.74, 131.25, 131.49, 131.85, 140.29, 142.89, 143.13, 146.55, 146.90, 159.59. HRMS (APCI, m/z): [M]+ calculated for C78H73N2O10, 1197.5260; found, 1197.5270. 4.2.13. Compound SQ3a: 1,3-Bis(5-(4-hexylphenyl)-1-(4(1,2,2-triphenylvinyl)phenyl)-1H-pyrrol-2-yl)squaraine. Compound SQ3a was synthesized via the same condensation reaction procedure as that described for the synthesis of compound SQ1a, using compound 8a (110 mg, 0.2 mmol) and squaric acid (11 mg, 0.1 mmol). The reaction time was 18 h. The crude product was purified slowly via column chromatography using EA/hexane (1:19−1:4 v/v) as eluent to isolate product as a dark blue solid (8.6 mg, 4% yield). 1 H NMR (400 MHz, CDCl3, δ): 0.90 (t, J = 8 Hz, 6H), 1.34 (m, 12H), 1.63 (m, 4H), 2.62 (t, J = 8 Hz, 4H), 6.75 (d, J = 4 Hz, 2H), 7.03−7.17 (m, 46H), 8.01 (d, J = 4 Hz, 2H). 13C NMR (100 MHz, CDCl3, δ): 14.09, 22.60, 29.02, 31.13, 31.71, 16432


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Author Contributions

was prepared by dissolving PBS tablet purchased from SigmaAldrich into the stipulated amount of deionized water. To prepare the sample solutions for selectivity test, 200 μL of 10 mM amino acid/GSH stock solutions was diluted with 800 μL of PBS. Then, 200 μL of 0.1 mM squaraine dye stock solution in DMSO and 800 μL of DMSO were added. After vigorous mixing using a spin vortex, the sample solutions were left to stand for an hour prior to spectrophotometric measurements. The results and corresponding photos of color changes are shown in SI Figures S19 and 4. 4.4.2. Detection on Solid Support. To demonstrate the applicability of the squaraine dyes for sensing thiol over solid biological samples, a simple study involving the use of silicabased thin-layer chromatography (TLC) plates as solid support for thiol compounds was performed. Using capillary tubes, 10 mM cysteine and glutathione were separately introduced onto TLC plates and briefly dried over warm air using a hand-held heat gun. Water was used as a blank. The thiol-coated TLC plates were briefly dipped into 0.1 mM solution of squaraine SQ1b, SQ2b, and SQ3b, thereafter dried over warm air using a hand-held heat gun. The plates were observed both visually under light, as well as under UV irradiation at 365 nm. The results are shown in the main manuscript, Figure 5. Next, cysteine and GSH of different concentrations (10−0.05 mM) were coated on separate TLC plates, which were then dipped in SQ3b solution (0.1 mM). The results are shown in SI Figure S20.

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The manuscript was written through contributions of all authors. All authors have given approval to the final version of the manuscript. Notes

The authors declare no competing financial interest.

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ACKNOWLEDGMENTS This work is supported by IMRE, A*STAR. DFT calculation was supported by the A*STAR Computational Resource Centre through the use of its high-performance computing facilities. The authors would also like to acknowledge the Chemical, Molecular and Materials Analysis Centre (CMMAC) at NUS Department of Chemistry for mass spectrometry and X-ray diffraction characterization.

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ASSOCIATED CONTENT

S Supporting Information *

The Supporting Information is available free of charge on the ACS Publications website at DOI: 10.1021/acsomega.8b02479. (i) Single-crystal XRD data and packing structures for SQ1a (CCDC No. 1861906); (ii) fluorescence decay curves for lifetime measurements and thin-film fluorescence spectra; (iii) fluorescence spectra depicting ACQ properties of SQ1a−SQ3b; (iv) DFT (B3LYP/631G)-calculated optimized structures, HOMO and LUMO molecular orbital profiles and energy levels of SQ1a−SQ3b, as well as time-dependent DFT-calculated energies, oscillator strength, and compositions of major electronic transitions; (v) UV−vis absorption and fluorescence spectra and AIE-property studies of SQ1a−SQ3b after reaction with decanethiol; (vi) absorption spectra and photos of SQ1b and SQ2b from selectivity test with amino acids and photos of detection on solid support; (vii) 1H and 13C NMR and high-resolution mass spectra (HRMS) of all synthesized intermediates and final compounds reported in Section 4.2 (PDF)

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REFERENCES

(1) Sreejith, S.; Carol, P.; Chithra, P.; Ajayaghosh, A. Squaraine Dyes: A Mine of Molecular Materials. J. Mater. Chem. 2008, 18, 264− 274. (2) Beverina, L.; Salice, P. Squaraine Compounds: Tailored Design and Synthesis towards a Variety of Material Science Applications. Eur. J. Org. Chem. 2010, 2010, 1207−1225. (3) Ian Storer, R.; Aciro, C.; Jones, L. H. Squaramides: Physical Properties, Synthesis and Applications. Chem. Soc. Rev. 2011, 40, 2330−2346. (4) Hu, L.; Yan, Z.; Xu, H. Advances in Synthesis and Application of Near-Infrared Absorbing Squaraine Dyes. RSC Adv. 2013, 3, 7667− 7676. (5) Bricks, J. L.; Kachkovskii, A. D.; Slominskii, Y. L.; Gerasov, A. O.; Popov, S. V. Molecular Design of near Infrared Polymethine Dyes: A Review. Dyes Pigm. 2015, 121, 238−255. (6) Xia, G.; Wang, H. Squaraine Dyes: The Hierarchical Synthesis and Its Application in Optical Detection. J. Photochem. Photobiol., C 2017, 31, 84−113. (7) Philips, D. S.; Ghosh, S.; Sudheesh, K. V.; Suresh, C. H.; Ajayaghosh, A. An Unsymmetrical Squaraine-Dye-Based Chemical Platform for Multiple Analyte Recognition. Chem. - Eur. J. 2017, 23, 17973−17980. (8) Frangioni, J. V. In Vivo Near-Infrared Fluorescence Imaging. Curr. Opin. Chem. Biol. 2003, 7, 626−634. (9) Escobedo, J. O.; Rusin, O.; Lim, S.; Strongin, R. M. NIR Dyes for Bioimaging Applications. Curr. Opin. Chem. Biol. 2010, 14, 64−70. (10) Kobayashi, H.; Ogawa, M.; Alford, R.; Choyke, P. L.; Urano, Y. New Strategies for Fluorescent Probe Design in Medical Diagnostic Imaging. Chem. Rev. 2010, 110, 2620−2640. (11) Luo, S.; Zhang, E.; Su, Y.; Cheng, T.; Shi, C. A Review of NIR Dyes in Cancer Targeting and Imaging. Biomaterials 2011, 32, 7127− 7138. (12) Schäferling, M. The Art of Fluorescence Imaging with Chemical Sensors. Angew. Chem., Int. Ed. 2012, 51, 3532−3554. (13) Chung, S. J.; Zheng, S.; Odani, T.; Beverina, L.; Fu, J.; Padilha, L. A.; Biesso, A.; Hales, J. M.; Zhan, X.; Schmidt, K.; et al. Extended Squaraine Dyes with Large Two-Photon Absorption Cross-Sections. J. Am. Chem. Soc. 2006, 128, 14444−14445. (14) Wang, W.; Fu, A.; Lan, J.; Gao, G.; You, J.; Chen, L. Rational Design of Fluorescent Bioimaging Probes by Controlling the Aggregation Behavior of Squaraines: A Special Effect of Ionic Liquid Pendants. Chem. - Eur. J. 2010, 16, 5129−5137. (15) Ahn, H. Y.; Yao, S.; Wang, X.; Belfield, K. D. Near-InfraredEmitting Squaraine Dyes with High 2PA Cross-Sections for Multiphoton Fluorescence Imaging. ACS Appl. Mater. Interfaces 2012, 4, 2847−2854. (16) Podgorski, K.; Terpetschnig, E.; Klochko, O. P.; Obukhova, O. M.; Haas, K. Ultra-Bright and -Stable Red and Near-Infrared Squaraine Fluorophores for In Vivo Two-Photon Imaging. PLoS One 2012, 7, No. e51980.

AUTHOR INFORMATION

Corresponding Authors

*E-mail: [email protected] (J.W.). *E-mail: [email protected] (J.X.). ORCID

Jishan Wu: 0000-0002-8231-0437 Jianwei Xu: 0000-0003-3945-5443 16433


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(17) Belfield, K. D.; Bondar, M. V.; Haniff, H. S.; Mikhailov, I. A.; Luchita, G.; Przhonska, O. V. Superfluorescent Squaraine with Efficient Two-Photon Absorption and High Photostability. ChemPhysChem 2013, 14, 3532−3542. (18) Zhang, Y.; Yue, X.; Kim, B.; Yao, S.; Bondar, M. V.; Belfield, K. D. Bovine Serum Albumin Nanoparticles with Fluorogenic Near-IREmitting Squaraine Dyes. ACS Appl. Mater. Interfaces 2013, 5, 8710− 8717. (19) Wu, I. C.; Yu, J.; Ye, F.; Rong, Y.; Gallina, M. E.; Fujimoto, B. S.; Zhang, Y.; Chan, Y. H.; Sun, W.; Zhou, X. H.; et al. SquaraineBased Polymer Dots with Narrow, Bright near-Infrared Fluorescence for Biological Applications. J. Am. Chem. Soc. 2015, 137, 173−178. (20) Jurek, K.; Kabatc, J.; Kostrzewska, K. Fluorescent AminoSubstituted Squaraine Probes for Bovine Serum Albumin. Color. Technol. 2017, 133, 170−177. (21) Sun, C. L.; Lv, S. K.; Liu, Y. P.; Liao, Q.; Zhang, H. L.; Fu, H.; Yao, J. Benzoindolic Squaraine Dyes with a Large Two-Photon Absorption Cross-Section. J. Mater. Chem. C 2017, 5, 1224−1230. (22) Hong, Y.; Lam, J. W. Y.; Tang, B. Z. Aggregation-Induced Emission: Phenomenon, Mechanism and Applications. Chem. Commun. 2009, 4332−4353. (23) Hong, Y.; Lam, J. W. Y.; Tang, B. Z. Aggregation-Induced Emission. Chem. Soc. Rev. 2011, 40, 5361. (24) Mei, J.; Hong, Y.; Lam, J. W. Y.; Qin, A.; Tang, Y.; Tang, B. Z. Aggregation-Induced Emission: The Whole Is More Brilliant than the Parts. Adv. Mater. 2014, 5429−5479. (25) Hu, R.; Leung, N. L. C.; Tang, B. Z. AIE Macromolecules: Syntheses, Structures and Functionalities. Chem. Soc. Rev. 2014, 43, 4494−4562. (26) Mei, J.; Leung, N. L. C.; Kwok, R. T. K.; Lam, J. W. Y.; Tang, B. Z. Aggregation-Induced Emission: Together We Shine, United We Soar! Chem. Rev. 2015, 115, 11718−11940. (27) Wang, H.; Zhao, E.; Lam, J. W. Y.; Tang, B. Z. AIE Luminogens: Emission Brightened by Aggregation. Mater. Today 2015, 18, 365−377. (28) Zhao, Z.; Chen, S.; Lam, J. W. Y.; Lu, P.; Zhong, Y.; Wong, K. S.; Kwok, H. S.; Tang, B. Z. Creation of Highly Efficient Solid Emitter by Decorating Pyrene Core with AIE-Active Tetraphenylethene Peripheries. Chem. Commun. 2010, 46, 2221−2223. (29) Chen, S.; Zhao, Z.; Tang, B. Z.; Kwok, H. S. Non-Doped White Organic Light-Emitting Diodes Based on Aggregation-Induced Emission. J. Phys. D: Appl. Phys. 2010, 43, No. 095101. (30) Zhao, Q.; Li, K.; Chen, S.; Qin, A.; Ding, D.; Zhang, S.; Liu, Y.; Liu, B.; Sun, J. Z.; Tang, B. Z. Aggregation-Induced Red-NIR Emission Organic Nanoparticles as Effective and Photostable Fluorescent Probes for Bioimaging. J. Mater. Chem. 2012, 22, 15128−15135. (31) Zhao, Q.; Zhang, X. A.; Wei, Q.; Wang, J.; Shen, X. Y.; Qin, A.; Sun, J. Z.; Tang, B. Z. Tetraphenylethene Modified Perylene Bisimide: Effect of the Number of Substituents on AIE Performance. Chem. Commun. 2012, 48, 11671−11673. (32) Hu, R.; Gómez-Durán, C. F. A.; Lam, J. W. Y.; BelmonteVázquez, J. L.; Deng, C.; Chen, S.; Ye, R.; Peña-Cabrera, E.; Zhong, Y.; Wong, K. S.; et al. Synthesis, Solvatochromism, AggregationInduced Emission and Cell Imaging of Tetraphenylethene-Containing BODIPY Derivatives with Large Stokes Shifts. Chem. Commun. 2012, 48, 10099−10101. (33) Zhao, Z.; Chen, B.; Geng, J.; Chang, Z.; Aparicio-Ixta, L.; Nie, H.; Goh, C. C.; Ng, L. G.; Qin, A.; Ramos-Ortiz, G.; et al. Red Emissive Biocompatible Nanoparticles from TetraphenyletheneDecorated BODIPY Luminogens for Two-Photon Excited Fluorescence Cellular Imaging and Mouse Brain Blood Vascular Visualization. Part. Part. Syst. Charact. 2014, 31, 481−491. (34) Gomez-Duran, C. F. A.; Hu, R.; Feng, G.; Li, T.; Bu, F.; Arseneault, M.; Liu, B.; Pena-Cabrera, E.; Tang, B. Z. Effect of AIE Substituents on the Fluorescence of Tetraphenylethene-Containing BODIPY Derivatives. ACS Appl. Mater. Interfaces 2015, 7, 15168− 15176.

(35) Chua, M. H.; Ni, Y.; Garai, M.; Zheng, B.; Huang, K. W.; Xu, Q. H.; Xu, J.; Wu, J. Towards Meso-Ester BODIPYs with Aggregation-Induced Emission Properties: The Effect of Substitution Positions. Chem. - Asian J. 2015, 10, 1631−1634. (36) Chua, M. H.; Zhou, H.; Lin, T. T.; Wu, J.; Xu, J. W. Aggregation-Induced Emission Active 3,6-Bis(1,2,2-Triphenylvinyl)Carbazole and Bis(4-(1,2,2-Triphenylvinyl)Phenyl)Amine-Based Poly(Acrylates) for Explosive Detection. J. Polym. Sci., Part A: Polym. Chem. 2017, 55, 672−681. (37) Chua, M. H.; Zhou, H.; Lin, T. T.; Wu, J.; Xu, J. Triphenylethylenyl-Based Donor-Acceptor-Donor Molecules: Studies on Structural and Optical Properties and AIE Properties for Cyanide Detection. J. Mater. Chem. C 2017, 5, 12194−12203. (38) Sancenón, F.; Martínez-Máñez, R.; Soto, J. A Selective Chromogenic Reagent for Nitrate. Angew. Chem., Int. Ed. 2002, 41, 1416−1419. (39) Ros-Lis, J. V.; García, B.; Jiménez, D.; Martínez-Máñez, R.; Sancenón, F.; Soto, J.; Gonzalvo, F.; Valldecabres, M. C. Squaraines as Fluoro-Chromogenic Probes for Thiol-Containing Compounds and Their Application to the Detection of Biorelevant Thiols. J. Am. Chem. Soc. 2004, 126, 4064−4065. (40) Sreejith, S.; Divya, K. P.; Ajayaghosh, A. A Near-Infrared Squaraine Dye as a Latent Ratiometric Fluorophore for the Detection of Aminothiol Content in Blood Plasma. Angew. Chem., Int. Ed. 2008, 47, 7883−7887. (41) Hewage, H. S.; Anslyn, E. V. Pattern-Based Recognition of Thiols and Metals Using a Single Squaraine Indicator. J. Am. Chem. Soc. 2009, 131, 13099−13106. (42) Anees, P.; Joseph, J.; Sreejith, S.; Menon, N. V.; Kang, Y.; Wing-Kwong, Yu S.; Ajayaghosh, A.; Zhao, Y. Real Time Monitoring of Aminothiol Level in Blood Using a Near-Infrared Dye Assisted Deep Tissue Fluorescence and Photoacoustic Bimodal Imaging. Chem. Sci. 2016, 7, 4110−4116. (43) Diehl, K. L.; Bachman, J. L.; Anslyn, E. V. Tuning Thiol Addition to Squaraines by Ortho -Substitution and the Use of Serum Albumin. Dyes Pigm. 2017, 141, 316−324. (44) Anees, P.; Sreejith, S.; Ajayaghosh, A. Self-Assembled NearInfrared Dye Nanoparticles as a Selective Protein Sensor by Activation of a Dormant Fluorophore. J. Am. Chem. Soc. 2014, 136, 13233−13239. (45) Hwang, C.; Sinskey, A. J.; Lodish, H. F. Oxidized Redox State of Glutathione in the Endoplasmic Reticulum. Science 1992, 257, 1496−1502. (46) Dalton, T. P.; Shertzer, H. G.; Puga, A. Regulation of Gene Expression By Reactive Oxygen. Annu. Rev. Pharmacol. Toxicol. 1999, 39, 67−101. (47) Kleinman, W. A.; Richie, J. P. Status of Glutathione and Other Thiols and Disulfides in Human Plasma. Biochem. Pharmacol. 2000, 60, 19−29. (48) Shahrokhian, S. Lead Phthalocyanine as a Selective Carrier for Preparation of a Cysteine-Selective Electrode. Anal. Chem. 2001, 73, 5972−5978. (49) Seshadri, S.; Beiser, A.; Selhub, J.; Jacques, P. F.; Rosenberg, I. H.; D’Agostino, R. B.; Wilson, P. W. F.; Wolf, P. A. Plasma Homocysteine as a Risk Factor for Dementia and Alzheimer’s Disease. N. Engl. J. Med. 2002, 346, 476−483. (50) Zhang, S.; Ong, C. N.; Shen, H. M. Critical Roles of Intracellular Thiols and Calcium in Parthenolide-Induced Apoptosis in Human Colorectal Cancer Cells. Cancer Lett. 2004, 208, 143−153. (51) Poole, L. B. The Basics of Thiols and Cysteines in Redox Biology and Chemistry. Free Radical Biol. Med. 2015, 80, 148−157. (52) Peng, H.; Chen, W.; Cheng, Y.; Hakuna, L.; Strongin, R.; Wang, B. Thiol Reactive Probes and Chemosensors. Sensors 2012, 12, 15907−15946. (53) Jung, H. S.; Chen, X.; Kim, J. S.; Yoon, J. Recent Progress in Luminescent and Colorimetric Chemosensors for Detection of Thiols. Chem. Soc. Rev. 2013, 42, 6019−6031. 16434


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Article

(54) Yin, C.; Huo, F.; Zhang, J.; Martínez-Máñez, R.; Yang, Y.; Lv, H.; Li, S. Thiol-Addition Reactions and Their Applications in Thiol Recognition. Chem. Soc. Rev. 2013, 42, 6032−6059. (55) Chen, X.; Shen, X. Y.; Guan, E.; Liu, Y.; Qin, A.; Sun, J. Z.; Tang, B. Z. A Pyridinyl-Functionalized Tetraphenylethylene Fluorogen for Specific Sensing of Trivalent Cations. Chem. Commun. 2013, 49, 1503−1505. (56) Lou, X.; Hong, Y.; Chen, S.; Leung, C. W. T.; Zhao, N.; Situ, B.; Lam, J. W. Y.; Tang, B. Z. A Selective Glutathione Probe Based on AIE Fluorogen and Its Application in Enzymatic Activity Assay. Sci. Rep. 2014, 4, No. 4272. (57) Wang, K.; Peng, H.; Wang, B. Recent Advances in Thiol and Sulfide Reactive Probes. J. Cell. Biochem. 2014, 115, 1007−1022. (58) Yin, J.; Kwon, Y.; Kim, D.; Lee, D.; Kim, G.; Hu, Y.; Ryu, J. H.; Yoon, J. Cyanine-Based Fluorescent Probe for Highly Selective Detection of Glutathione in Cell Cultures and Live Mouse Tissues. J. Am. Chem. Soc. 2014, 136, 5351−5358. (59) Yin, J.; Kwon, Y.; Kim, D.; Lee, D.; Kim, G.; Hu, Y.; Ryu, J. H.; Yoon, J. Preparation of a Cyanine-Based Fluorescent Probe for Highly Selective Detection of Glutathione and Its Use in Living Cells and Tissues of Mice. Nat. Protoc. 2015, 10, 1742−1754. (60) Yin, C. X.; Xiong, K. M.; Huo, F. J.; Salamanca, J. C.; Strongin, R. M. Fluorescent Probes with Multiple Binding Sites for the Discrimination of Cys, Hcy, and GSH. Angew. Chem., Int. Ed. 2017, 56, 13188−13198. (61) Frisch, M. J.; Trucks, G. W.; Schle-gel, H. B.; Scuseria, G. E.; Robb, M. A.; Cheeseman, J. R.; Scalmani, G.; Barone, V.; Mennucci, B.; Petersson, G. A.; Nakatsuji, H.; Caricato, M.; Li, X.; Hratchian, H. P.; Izmaylov, A. F.; Bloino, J.; Zheng, G.; Son-nenberg, J. L.; Hada, M.; Ehara, M.; Toyota, K.; Fukuda, R.; Hasegawa, J.; Ishida, M.; Nakajima, T.; Honda, Y.; Kitao, O.; Nakai, H.; Vreven, T.; Montgomery, J. A., Jr.; Peralta, J. E.; Ogliaro, F.; Bearpark, M.; Heyd, J. J.; Brothers, E.; Kudin, K. N.; Staroverov, V. N.; Kobayashi, R.; Normand, J.; Raghavachari, K.; Rendell, A.; Burant, J. C.; Iyengar, S. S.; Tomasi, J.; Cossi, M.; Rega, N.; Millam, J. M.; Klene, M.; Knox, J. E.; Cross, J. B.; Bakken, V.; Adamo, C.; Jaramillo, J.; Gomperts, R.; Stratmann, R. E.; Yazyev, O.; Austin, A. J.; Cammi, R.; Pomelli, C.; Ochterski, J. W.; Martin, R. L.; Morokuma, K.; Zakrzewski, V. G.; Voth, G. A.; Salvador, P.; Dannenberg, J. J.; Dapprich, S.; Daniels, A. D.; Farkas, O.; For-esman, J. B.; Ortiz, J. V.; Cioslowski, J.; Fox, D. J.. Gaussian 09, revision A.02; Gaussian, Inc.: Wall-ingford CT, 2009. (62) Maeda, H.; Ito, Y.; Haketa, Y.; Eifuku, N.; Lee, E.; Lee, M.; Hashishin, T.; Kaneko, K. Solvent-Assisted Organized Structures Based on Amphiphilic Anion-Responsive π-Conjugated Systems. Chem. - Eur. J. 2009, 15, 3706−3719.

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Triphenylethylene- and Tetraphenylethylene-Functionalized 1,3-Bis

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