Simultaneous Sensing of Aqueous Anions and Toxic Metal Ions by

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Simultaneous Sensing of Aqueous Anions and Toxic Metal Ions by Simple Dithiosemicarbazones and Bioimaging of Living Cells Sivalingam Suganya,† Hye Jin Zo,‡ Jong S. Park,‡ and Sivan Velmathi†,* †

Organic and Polymer Synthesis Laboratory, Department of Chemistry, National Institute of Technology, Tiruchirappalli-620 015, India Department of Organic Material and Polymer Engineering, Dong-A University, Busan 604-714, Korea

‡

S Supporting Information *

ABSTRACT: New thiophene (TSC-1) and bipyridine (TSC-2) anchored thiosemicarbazones were synthesized for the recognition of biologically active and toxic analyte. Both the receptors TSC-1 and TSC-2 commonly exhibited strong sensitivity toward anions such as F−, AcO−, and CN− ions over other anions, whereas TSC-1 and TSC-2 recognized Cd2+ and Hg2+ ions, respectively. Both anion and cation recognition were clearly differentiated from colorimetric, absorption, and emission responses. A significant shift with remarkable fluorescence enhancement at 540 nm was observed for TSC-1-Cd2+, which describes an internal charge transfer (ICT) mechanism. In contrast, anion complexes of TSC-1 show fluorescence enhancement at 570 nm. TSC-2 acts as a photoinduced electron transfer (PET) sensor during both anion and cation sensing accompanied by fluorescence quenching. Furthermore, TSC-1/2 preferentially recognizes cations in the presence of competitive anions. Interestingly, selective detection of the CN− ion using the Hg2+-TSC-2 ensemble also achieved through a relay recognition method. Job’s plot analysis reveals that TSC-1/2 binds with the selective anions in 2:1 mode, and 1:1 mode was determined for Cd2+-TSC-1 and Hg2+-TSC-2, respectively. TSC-1 can be utilized for practical applications such as the recognition of targeted F− ions in daily used household toothpaste, and Cd2+ ion detection was achieved in living cells using bioimaging.

1. INTRODUCTION The development of synthetic organic protocols for individual recognition of cations, anions, and neutral molecules has garnered significant interest in the fields of organic and supramolecular chemistry.1−3 However, limited success has been achieved in terms of realizing dual responsive nature of organic receptors for the simultaneous recognition of cations and anions.4,5 The demonstration of a single receptor for individual analyte recognition and specific analyte detection utilizing displacement approach with distinct color changes and readable signals at different wavelengths would have additional impact in this field. Human health relies on the help of both positive and negative ions. Regarding biological concerns, F− ions play a major role in dental care, bone health, and the nervous system6−8 while AcO− ions are the main component in enzyme based reactions and food preservatives.9−11 On the other hand, with higher concentrations of F− ions in biological systems and drinking water, the activity of F− ions becomes toxic, giving rise to severe health risks. Similarly, with respect to environmental concerns, toxic analytes such as cyanide ion,12−15 Hg(II)16−18 and Cd(II)19−21 ions pose major health risks to living creatures. These toxic ions mainly originate from industrial wastes, electrical batteries, oil production, and electroplating processes. Cyanide is the most toxic anion, and the strong nucleophilic nature of CN− causes it to interfere in the electron transfer process and restrain the functions of the respiratory system. Free CN− or HCN produced from industrial processes is able to pollute drinking water and the environment which causes human beings to die even at trace levels. U.S. Environmental Protection Agency (EPA) suggests that the maximum level of cyanide in drinking water should be 100 mg/m3. Similarly, all forms of Hg, including elemental, organic, and inorganic forms, © 2014 American Chemical Society

are highly poisonous, and the ingested ions impair biological functions, resulting in damage to or disease of neurological, cardiovascular, and reproductive systems. The World Health Organization (WHO) and EPA have strictly fixed the maximum levels of these toxic ions that can be present in drinking water. Protecting the environment from contamination by toxic ions is an important task with respect to human health as well as the health of other organisms. Selectivity is the most significant and interesting issue in specific analyte detection. Recently, many reports22−25 are available for selective analyte detection using a relay recognition mechanism which prompted us to accomplish the selectivity. Cyanide is a well-known strong nucleophilic anion, thus it can easily displace the heavy metal ions such as Hg2+ ion and form a new Hg(CN)2 complex.26 Colorimetric chemosensors27−29 are always welcome in the molecular recognition field, because they do not require sophisticated instruments and the results can be easily monitored by the naked eye. A colorimetric F− ion sensor could have practical applicability in the detection of F− ions from real samples such as toothpaste and mouthwash.30 In addition, fluorescent sensors31−33 can be used as powerful detectors for metal ions in living cells that can be monitored through bioimaging processes. In general, the sensing device should be neutral and provide both hydrogen bonding sites as well as coordination sites for the recognition of charged analytes. These considerations have led researchers to apply thiosemicarbazones34−37 as dual responsive receptors since they contain hydrogen bonding sites (−NH) and Received: Revised: Accepted: Published: 9561

February 20, 2014 May 6, 2014 May 17, 2014 May 17, 2014 dx.doi.org/10.1021/ie500737b | Ind. Eng. Chem. Res. 2014, 53, 9561−9569

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2.1.1. Synthesis of TSC-1. TSC-1 was prepared by reacting 1 mmol of 2,5-thiophenedicarboxaldehyde with 2 mmol of N-phenyl thiosemicarbazide in EtOH medium. A drop of acetic acid (AcOH) was added to the reaction mixture and allowed to reflux for 3 h at 70 °C. After cooling to r.t., the precipitate was filtered, washed with EtOH, and dried in a vacuum oven. Yield: 85%. M.pt: 220 °C. IR (cm-1, KBr): 3324, 3126, 2973, 1593, 1517, 1447, 1284, 1050. 1H NMR (400 MHz DMSO-d6): 12.02 (NH, 1H, s), 9.86 (NH, 1H, s) 8.37 (CHN, 1H, s), 7.60− 7.58 (Ar−H, 3H, t), 7.43−7.38 (Ar−H, 2H, d), 7.26−7.22 (Ar−H, 1H, s). 13C NMR (100 MHz DMSO-d6): 175.55, 140.58, 138.78, 137.49, 131.38, 128.09, 125.85, 125.46, 125.32. HRMS: 437.0671 (calculated), 437.0669 (found). 2.1.2. Synthesis of TSC-2. 2,2′-Bipyridine-5,5′-dicarbaldehyde was synthesized using reported procedure.38 TSC-2 was prepared by reacting 1 mmol of [2,2′-bipyridine]-5,5′dicarbaldehyde with 2 mmol of N-phenyl thiosemicarbazide in EtOH medium. A drop of AcOH was added to the reaction mixture and allowed to reflux for 3 h at 70 °C. After cooling to r.t., the precipitate was filtered, washed with EtOH, and dried in a vacuum oven. Yield: 70%. M.pt: 220 °C. IR (cm−1, KBr): 3366, 3450, 1609, 1593, 1488, 1457, 1284, 1050. 1H NMR (400 MHz DMSO-d6): 12.05 (NH, 1H, s), 10.26 (NH, 1H, s), 9.13 (CHN, 1H, s), 8.60−8.58 (Ar−H, 1H, d), 8.46−8.43 (Ar− H, 1H,d), 8.24 (Ar−H, 1H, s), 7.57−7.55 (Ar−H, 2H, d), 7.41−7.39 (Ar−H, 2H, t), 7.23−7.20 (Ar−H, 1H,t). 13C NMR (100 MHz DMSO-d6): 176.21, 155.17, 149.11, 139.38, 138.95, 135.11, 130.52, 128.07, 126.06, 125.49, 120.42. HRMS: 511.148 16 (calculated), 511.147 99 (found).

coordination sites (CHN). In continuation of our research, here we present two novel thiosemicarbazones for the simultaneous detection of anions as well as cations. Cation recognition is well distinguished by means of heteroatoms present in both the receptors, thus TSC-1 shows selective affinity toward Cd2+ ions and TSC-2 shows selective affinity toward Hg2+ ions. While the two thiosemicarbazones sense common anions such as F−, ACO−, and CN−. Beyond this multiple analyte detection, interestingly the CN− ion was selectively recognized by Hg2+-TSC-2 ensembles using the displacement approach and revived the free TSC-2.

2. EXPERIMENTAL PROCEDURES 2.1. Materials and Instruments Used. Starting materials used for the synthesis, solvents such as dimethyl sulfoxide (DMSO) and ethanol (EtOH) used for spectroscopic experiments, and all the anion and metal salts were received commercially and used as such without any further purification. All the anions were used in the form of tetra butyl ammonium salts, and metal ions used were in the form of chloride, nitrate, acetate, and sulfate. The molecular weights of the receptors were obtained using a Orbitrap Q exactive mass spectrometer. 1 H and 13C NMR spectra were obtained on a BRUKER AV III-400 MHz spectrometer using DMSO-d6 as a solvent. IR spectra were recorded on a NICOLET IS5 instrument using KBr plates. UV−vis spectra were recorded on a Shimadzu UV-2600 spectrophotometer with a quartz cuvette (path length = 1 cm) at room temperature (r.t). Fluorescence spectra were recorded on a Shimadzu RF-5301 PC spectrophotometer. Fluorescence imaging was performed with a Leica TCS-SP5-X AOBS confocal microscope. For all the spectroscopic titrations, 5 × 10−5 M solution of the receptors TSC-1 and TSC-2 in DMSO and 1.5 × 10−3 M of all anions in DMSO and H2O and aqueous DMSO and 1.5 × 10−5 M of cations in H2O were prepared and used. A synthetic scheme for TSC-1 and TSC-2 is given in the Supporting Information (Scheme-1).

3. RESULTS AND DISCUSSION Dual responsive heterocyclic linked dithiosemicarbazones (TSC-1/2) were synthesized and well characterized using spectroscopic and mass analysis (Supporting Information

Figure 1. 1H NMR spectrum of receptor TSC-2 recorded in DMSO-d6 in the (a) absence and presence of F− ion, (b) 0.5, (c) 1, (d) 1.5, and (e) 2 equiv. 9562

dx.doi.org/10.1021/ie500737b | Ind. Eng. Chem. Res. 2014, 53, 9561−9569


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Figure 2. 1H NMR spectrum of receptor TSC-2 recorded in DMSO-d6 in the (a) absence and presence of AcO− ion, (b) 0.5, (c) 1, (d) 1.5, and (e) 2 equiv.

shielding region was observed. Consequent addition of F− ions still causes upfield shifting of the aromatic protons present in TSC-2. In contrast, with gradual addition of 0−2 equiv of AcO− ions to TSC-2, broadening of NH proton takes place instead of complete deprotonation. Such differentiation is mainly due to the high basicity of the F− ions compared to AcO− ions. However, imine and aromatic protons are shifted in the upfield region in the presence of AcO− ions similar to F− ions. In addition to 1H NMR, UV−vis and fluorescence titrations were carried out for complete understanding of the sensing nature and stoichiometry of the newly formed complex. 3.2. Colorimetric Analysis: Anion and Cation Sensing. In the visual sensing analysis, the presence of 2 equiv of F−/AcO−/CN− ions made distinct color changes of TSC-1 and TSC-2 from fluorescent green to orange. The remaining anions, Cl−, Br−, AcO−, HSO4−, H2PO4−, and OH−, were found to be insensible toward both receptors, TSC-1 and TSC-2, even with excess addition (Figure 3).The immediate response as distinct color changes demonstrates the formation of anion complex with TSC-1 and TSC-2.

Figure.S1). Both the receptors exhibit highly selective and sensitive responses toward specific anionic and cationic species.

3.1. 1H NMR Titration. 1H NMR titration was done in DMSO-d6 to further support the binding nature of synthesized receptor (TSC-2) in the presence of F− and AcO− ions. Figure 1 and 2 demonstrates the titration with the gradual addition of 0.5−2 equiv of F− and AcO− ions to TSC-2. In the absence of any anions, TSC-2 showed two singlets for NH protons at 12.05 and 10.26 ppm and one singlet for a CHN proton at 9.13 ppm. We observed that in the presence of even 0.5 equiv of F− ions deprotonation of the NH signal and the signal corresponds to imine proton (CHN) shifting in the

Figure 3. Color changes observed upon the addition of 2 equiv of various anions (1.5 × 10−3 M in DMSO) to (a) TSC-1 and (b) TSC-2 (5 × 10−5 M in DMSO). From left to right: free sensor, F−, Cl−, Br−, AcO−, H2PO4−, OH−, HSO4−, and CN−. 9563



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Figure 4. Color changes observed upon the addition of 2 eq. of various cations (1.5 × 10−5 M in H2O) to (a) TSC-1 and (b) TSC-2 (5 × 10−5 M in DMSO). From left to right: R, R+ Cr3+, R + Mn2+, R + Fe3+, R + Co2+, R + Ni2+, R + Cu2+, R + Zn2+, R + Cd2+, R + Hg2+, R + Sn2+, R + Pb2+.

Simultaneously, an investigation of the aqueous cation recognition capability of TSC-1 and TSC-2 was carried out to validate their dual sensing nature. Here, naked-eye colorimetric experiments were successful and TSC-1 showed selective sensing of Cd2+ ion, as the fluorescent green color was changed into yellow (Figure 4a). TSC-2 meanwhile showed selective sensing of Hg2+ ion, as the light yellow color changed into intense yellow (Figure 4b). However, no significant changes were observed in the presence of other metal ions such as Cr3+, Mn2+, Fe3+, Co2+, Ni2+, Cu2+, Zn2+, Pb2+, and Sn2+. Such color changes suggest the formation of a ligand to metal charge transfer (LMCT) complex via coordination bonding.5,39,40 The fascinating observation of different cation recognition by two receptors even though they contain the same thiosemicarbazone binding sites suggests that the development of an organic scaffold with different hetero atoms can exhibit selective metal ion detection toward different toxic metal ions. 3.3. UV−vis Titrations. To examine the dual responsive nature of TSC-1 and TSC-2, UV−vis titrations were carried out by the individual addition of anions and cations as well as in the presence of both competitive analytes. TSC-1 showed a strong intramolecular charge transfer band at 410 nm and was redshifted at 494 nm with the addition of 200 μL of F−, AcO−, and CN− ions with an isosbestic point at 448 nm. Similar observations were made in the case of TSC-2. With the addition of 200 μL of the above ions to TSC-2, a strong band at 385 nm was red-shifted to the visible region at 476 nm with a single isosbestic point at 421 nm. The remaining anions such as Cl−, Br−, HSO4−, OH−, and H2PO4− show no significant absorption changes for both receptors (Figure 5). Incremental titrations were carried out for both TSC-1 (Supporting Information Figure.S2) and TSC-2 (Supporting Information Figure.S3) in the presence of F−, AcO−, and CN− ions. Uniform increase and decrease in the absorption spectrum at 493 and 411 nm reveal the anion recognition nature through the hydrogen bonding interaction connecting the NH binding site and the analytes. To address the effect of water on anion sensing, colorimetric titrations were carried out between TSC-1, and different ratios of aqueous anions (DMSO:H2O). Unlike other reversible receptors, TSC-1 showed a colorimetric (Supporting Information Figure.S4) and optical response for aqueous anion up to (0:10, DMSO:H2O), thus demonstrating aqueous anion detection nature (Supporting Information Figures.S5−S6). Likewise, the cation recognition nature of TSC-1 and TSC-2 was also examined. The manifestation of colorimetric titrations was observed in the absorption spectrum. TSC-1 showed

Figure 5. UV−vis spectra after the addition of 2 equiv of various anions (1.5 × 10−3 M in DMSO) to (a) TSC-1 and (b) TSC-2 (5 × 10−5 M in DMSO).

selective sensing of aqueous Cd2+ ion (Figure 6a), and TSC-2 showed selection of aqueous Hg2+ ion (Figure 6b) over other cations such as Cr3+, Mn2+, Fe3+, Co2+, Ni2+, Cu2+, Zn2+, Pb2+, and Sn2+even at higher equivalents. Upon the gradual addition of Cd2+ ions to TSC-1 (Supporting Information Figure.S7a), a new band at 456 nm was observed. Similarly, a new band at 440 nm was observed in the presence of 0−200 μL of Hg2+ ions to TSC-2 (Supporting Information Figure.S7b). The formation of new bands at two different wavelengths supports the anion and cation recognition by both receptors. The detection limit of Cd2+ and Hg2+ ions by TSC-1 and TSC-2 was achieved at lower concentrations on the 10−7 M level. 3.4. Fluorescence Titrations. Fluorescence probe is a powerful tool, offering a spontaneous means of monitoring 9564



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Figure 6. UV−vis spectra after the addition of 2 equiv of various cations (1.5 × 10−5 M in H2O) to (a) TSC-1 and (b) TSC-2 (5 × 10−5 M in DMSO).

Figure 7. Fluorescence spectra after the addition of 2 equiv of various anions (1.5 × 10−3 M in DMSO) to (a) TSC-1 and (b) TSC-2 (5 × 10−5 M in DMSO).

targeted analyte recognition via a bioimaging process of living cells, and simultaneous utilization of invitro and invivo assays can also be accomplished. To elucidate the emission properties of the receptors TSC-1 and TSC-2 alone and accompanied by particular metal ions or anions, emission titrations were carried out. Fluorescence responses in the presence of various anions, such as F−, AcO−, Cl−, Br−, HSO4−, OH−, CN−, and H2PO4− were studied. Among all these anions, only F−/AcO−/CN− showed significant fluorescence enhancement and quenching toward TSC-1 and TSC-2, respectively (Figure 7). With the gradual addition of F−/AcO−/CN− to TSC-1 and TSC-2, uniform enhancement in the fluorescence intensity at 570 nm (Supporting Information Figure.S8) and gradual quenching at 552 nm (Supporting Information Figure.S9) was observed. Simultaneously, the cation recognition nature of TSC-1 was studied and it was found that excluding Cd2+ ions, no cations showed a considerable shift or change (Figure 8). TSC-1 showed an emission band at 486 nm and the intensity is quenched with the addition of up to 100 μL of Cd2+ ions. Surprisingly, upon further addition of Cd2+ ions, the emission intensity is red-shifted to 540 nm with a significant enhancement (Figure 9a), which is attributed to an internal charge transfer mechanism (ICT).41 In the case of TSC-2, the strong emission centered at 552 nm is quenched exclusively in the presence of Hg2+ ions on the basis of a heavy atom effect (Figure 9b). This fluorescence response differentiation between TSC-1 and TSC-2 is ascribed to the bi pyridine N atom of TSC-2 involved in metal coordination through electron transfer, which leads to the observed fluorescence quenching. In contrast, thiophene S atom is not involved in metal coordination and leads to fluorescence enhancement. Table 1

Figure 8. Fluorescence spectra after the addition of 2 equiv of various cations (1.5 × 10−5 M in H2O) to TSC-1 (5 × 10−5 M in DMSO).

represent the calculated binding constants (Benesi−Hildebrand plot), detection limits, and quenching constant (Stern−Volmer plot) values from absorption and emission measurements. 3.5. Competitive Sensing of Anions and Cations. Since both receptors are respectively composed of two types of binding sites toward anions and cations, the binding efficiency of competing ions over other ions was examined for TSC-2. Two types of competitive titrations were carried out, and the output was measured as a readable signal using a UV−vis spectrophotometer. In the first experiment, TSC-2 was titrated with F− ions in the presence of Hg2+ ions. As shown in Supporting Information Figure.S10, the band at 440 nm is retained after the addition of F− ions and no new band is 9565



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metal ion can be displaced in the presence of specific anion. Hg2+-TSC-2 ensembles were utilized to examine the selective of detection CN− ion. Absorption titration reveals the presence of CN−, which displaces the Hg2+ ion. A strong band at 440 nm corresponding to Hg2+-TSC-2 was completely decreased upon the gradual addition of 2 equiv of CN− ion, restore the original absorption spectrum of free TSC-2 and form Hg(CN)2 complex. Whereas, the presence of other anions such as F−, AcO−, Cl−, Br−, HSO4−, OH−, and H2PO4− ions did not show any obvious changes (Figure 10a). Colorimetric titration also

Figure 9. Fluorescence spectra after the addition of 0−2 equiv of Cd2+ and Hg2+ ions to (1.5 × 10−5 M in H2O) to (a) TSC-1 and (b) TCS-2 (5 × 10−5 M in DMSO).

Figure 10. (a) UV−vis and (b) colorimetric titration after the addition of 2 equiv of various anions (1.5 × 10−3 M in DMSO) to the solution of the TCS-2-Hg2+ ensemble. From left to right: free TSC-2, Hg2+, F−, Cl−, Br−, AcO−, H2PO4−, OH−, HSO4−, CN−.

Table 1. Binding Constants (Ka), Detection Limit (LOD), and Quenching Constants (Ksv) Values Ka

host-guest −

TSC-1-F TSC-1-AcO− TSC-1-CN− TSC-1-Cd2+ TSC-2-F− TSC-2-AcO− TSC-2-CN− TSC-2-Hg2+ TSC-2-Hg2+-CN−

5.833 5.546 2.364 9.668 4.335 2.648 8.697 6.118 9.391

× × × × × × × × ×

3

10 104 103 105 103 104 103 104 103

± ± ± ± ± ± ± ± ±

0.36 0.11 0.11 0.08 0.68 0.29 0.08 0.03 0.23

LOD (μM)

stoichiometry (host:guest)

Ksv (104)

0.231 0.352 0.101 33.76 0.147 0.426 0.937 0.310 0.750

1:2 1:2 1:2 1:1 1:2 1:2 1:2 1:1 1:1

NA NA NA NA 5.80 3.072 3.419 3.841 NA

supports the above findings; the original color of the free TSC2 was retained after the consequent addition of Hg2+ and CN− ions (Figure 10b). The observation was further confirmed from UV−vis and fluorescence titration with the incremental addition of Hg2+ and CN− ions. A strong absorption at 440 nm of the TSC-2-Hg2+ ensemble was completely decreased upon the addition of 2 equiv of CN− ion, and this revived the original spectrum of free TSC-2 (Figure 11a). Likewise, while the emission spectrum of TSC-2 was quenched in the presence of Hg2+ ion, the addition of CN− ion replaced the Hg2+ ion and enhanced the emission spectrum of TSC-2 (Figure 11b). This result shows the selective sensing of CN− ion and also the effective recycling nature of TSC-2. Figure 12 describes the above competitive and sequential analyte detection findings. Meanwhile, the Cd2+-TSC-1 ensemble did not show any sensitivity with the CN− ion, which may be the weak interaction between CN− and Cd2+ ions. 3.7. Analytical Application. 3.7.1. Fluoride Ion Detection in Toothpaste and Mouthwash. To study the analytical applications of TSC-1 and TSC-2, detection of F− ions in toothpaste (TP)42 and mouthwash (MW) was successfully carried out, exploiting colorimetric and UV−vis titrations. Toothpaste was stirred in distilled water (20 mg/mL) and filtered, and the filtrate was used for the titration. Upon addition of this aqueous solution of toothpaste and mouthwash to TSC-1 (5 μM), the fluorescent green color of the receptor

observed at 476 nm. Subsequently, in the second titration, TSC-2 was titrated with Hg2+ ions in the presence of F− ions. Here, a strong reduction of the band at 476 nm and a new band at 440 nm were observed, indicating sequestration of F− ions from weak hydrogen bonding of a F−-TSC-2, complex and the formation of a strongly coordinated Hg2+-TSC-2 complex. The out coming F− ions again form anion pair with excess Hg2+ ions and yield a HgF2 complex. Binding constant values further support the results. Similarly, higher affinity of metal ions over anions was also observed in case of TSC-1 Supporting Information Figure.S11. 3.6. Sequential Detection of Hg2+ and CN− Ions. The reversible recognition of second analyte is a new and interesting concept by means of forming a coordination complex and the 9566



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spectroscopy. This analytical titration suggests that these colorimetric chemosensors can be utilized as a powerful device for the estimation of F− ion concentration in toothpaste and mouthwash (Figure 13). Similar results were observed in the case of TSC-2.

Figure 13. UV−vis spectrum of TSC-1 (5 × 10−5 M in DMSO), TSC1 + TP, TSC-1 + MW. (inset) Colorimetric analysis: TSC-1, TSC-1 + TBAF, TSC-1 + TP, TSC-1 + MW.

3.7.2. Bioimaging of Living Cells. Based on the fluorescence switch on response, TSC-1 was utilized to examine the cell permeability and the sensitivity of Cd2+ ions in the living cells using a fluorescence microscope. RAW264.7 cells were grown in H-DMEM (Dulbecco’s Modified Eagle’s Medium, high glucose) supplemented with 10% FBS (Fetal Bovine Serum) in an atmosphere of 5% CO2. The cells cultured in DMEM were treated with Cd2+ solutions (20 μM, pH 7.4) in sterilized PBS medium and incubated at 37 °C for 30 min. The treated cells were washed with PBS to remove the excess metal ions.

Figure 11. (a) UV−vis spectra and (b) fluorescence spectra after the addition of CN− ion (1.5 × 10−3 M in DMSO) to the solution of TCS-2-Hg2+ ensemble.

turned into an orange color. Simultaneously, the color change was confirmed by a new band at 494 nm in UV−vis

Figure 12. Competitive and sequential detection of analyte by TSC-2. 9567


Industrial & Engineering Chemistry Research DMEM (2 mL) was added to the cell culture, which was then treated with 20 μM DMSO solution of TSC-1. The samples were incubated at 37 °C for 30 min. The culture medium was removed, and the treated cells were washed with PBS before observation. RAW264.7 cells illustrate a week fluorescent image in the presence of only TSC-1 (20 μM) and a strong green fluorescence was visualized when the addition of Cd2+ ion (20 μM) solution which clearly suggests the TSC-1 is strong enough for the cell membrane permeable and can easily detect Cd2+ ion in the living system (Figure 14).

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AUTHOR INFORMATION

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REFERENCES

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4. CONCLUSION In conclusion, two different thiosemicarbazones with different heteroatoms (TSC-1, TSC-2) were prepared and characterized by various analytical techniques. Both receptors showed dual responsive behavior toward anions and transition metal ions. Among different kinds of anions, both receptors exhibited sensing nature toward F−, AcO−, and CN− ions and selective sensing of Cd2+ ions by TSC-1 and Hg2+ ions by TSC-2 were also achieved and confirmed by colorimetric assays, UV−vis, and fluorescence titrations. The Hg2+-TSC-2 ensemble was utilized for the sequential recognition of cyanide ion selectively. TSC-1 was successfully applied to recognize F− ions in toothpaste and mouthwash samples and also Cd2+ ion in living cell. ASSOCIATED CONTENT

ACKNOWLEDGMENTS

One of the authors, S.V., expresses her thanks to DRDO (ERIP/ER/1006004/M/01/1333 dated 23-05-2011) for financial assistance in the form of a major sponsored project. This research was partly supported by the Basic Science Research Program through the National Research Foundation of Korea funded by the Ministry of Education, Science and Technology (grant no. 2013004436).

Figure 14. RAW264.7 cells supplemented with TSC-1 and TSC-1 + Cd2+. (a and d) Bright field image. (b and e) Fluorescence image. (c and f) Merge cell image of a and b and d and e. The samples supplemented RAW264.7 cells were grown in H-DMEM medium and incubated at 37 °C for 30 min. Concentrations used for the bioimaging process: TSC-1 (20 μM, pH 7.4) and Cd2+ (20 μM, pH 7.4).

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S Supporting Information *

Reaction scheme and selected UV−vis and fluorescence graph. This material is available free of charge via the Internet at http://pubs.acs.org/. Corresponding Author

*E-mail: [email protected]. Tel.: 91-431-2503640. Fax: 91-431-2500133. Notes

The authors declare no competing financial interest. 9568



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Simultaneous Sensing of Aqueous Anions and Toxic Metal Ions by

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