- Email: [email protected]
A highly selective turn-on chemosensor for Zn2+ in aqueous media and living cells
Accepted Manuscript Title: A highly selective turn-on chemosensor for Zn2+ in aqueous media and living cells Authors: Jae Min Jung, Seong Youl Lee, Eunju Nam, Mi Hee Lim, Cheal Kim PII: DOI: Reference:
S0925-4005(17)30082-5 http://dx.doi.org/doi:10.1016/j.snb.2017.01.076 SNB 21601
To appear in:
Sensors and Actuators B
Received date: Revised date: Accepted date:
9-11-2016 10-1-2017 10-1-2017
Please cite this article as: Jae Min Jung, Seong Youl Lee, Eunju Nam, Mi Hee Lim, Cheal Kim, A highly selective turn-on chemosensor for Zn2+ in aqueous media and living cells, Sensors and Actuators B: Chemical http://dx.doi.org/10.1016/j.snb.2017.01.076 This is a PDF file of an unedited manuscript that has been accepted for publication. As a service to our customers we are providing this early version of the manuscript. The manuscript will undergo copyediting, typesetting, and review of the resulting proof before it is published in its final form. Please note that during the production process errors may be discovered which could affect the content, and all legal disclaimers that apply to the journal pertain.
A highly selective turn-on chemosensor for Zn2+ in aqueous media and living cells Jae Min Jung,a Seong Youl Lee,a Eunju Nam,b Mi Hee Lim,b* Cheal Kima* a
Department of Fine Chemistry and Department of Interdisciplinary Bio IT Materials, Seoul National University of Science and Technology, Seoul 139-743, Republic of Korea. Fax: +822-973-9149; Tel: +82-2-970-6693; E-mail: [email protected] b Department of Chemistry, Ulsan National Institute of Science and Technology (UNIST), Ulsan 44919, Republic of Korea. Fax: +82-52-217-5409; Tel: +82-52-217-5422; E-mail: [email protected]
1
Graphical abstract
Highlights
A simple and easy-to-make fluorescence sensor 1 for Zn2+ was designed and synthesized. Sensor 1 could detect Zn2+ ions at much lower concentration than WHO guideline. 1 was applied to quantify and image Zn2+ in water samples, test kit and living cells. Sensing mechanism of Zn2+ by 1 via ICT was explained by theoretical calculations.
2
Abstract A new simple quinoline-based chemosensor 1 was synthesized for Zn2+. 1 showed the selective fluorescence enhancement in the presence of Zn2+ with a 1:1 stoichiometry in a nearperfect aqueous solution (bis-tris buffer:DMSO = 999:1), which was reversible with the addition of ethylenediaminetetraacetic acid (EDTA). The detection limit (0.6 μM) of 1 for Zn2+ was much lower than World Health Organization guideline (76 μM) in drinking water. 1 was successfully applied to quantify and image Zn2+ in water samples, test kit and living cells. The sensing mechanism of Zn2+ by 1 via the intramolecular charge transfer was explained by theoretical calculations.
Keywords: fluorescent chemosensor, zinc ion, cell imaging, test kit, theoretical calculations
3
Introduction Much attention has been recently focused on the design and synthesis of highly selective fluorescent chemosensors for sensing and monitoring heavy and transitional metal ions due to their easy monitoring, low cost and high selectivity. Zinc ion is one of the most abundant metal ions present in living organism, because of its rich coordination chemistry [1–4]. Although most of the Zn2+ ions in a cell are strongly bound to metalloproteins, free forms are also found throughout the cell. In the brain, 5-20% of the total Zn2+ is stored in presynaptic vesicles, and the highest intracellular free Zn2+ is found in the hippocampus. Zn2+ modulates brain excitability and plays a key role in synaptic plasticity [5–8]. On the other hand, the deficiency of zinc causes unbalanced metabolism, which in turn can result in retarded growth in children, brain disorders and high blood cholesterol, and also be connected in numerous neurodegenerative disorders, such as Alzheimer’s disease, epilepsy, ischemic stroke, and infantile diarrhea [9–18]. In addition, an excess of zinc in the environment may reduce the soil microbial activity which results in phytotoxic effect [19–24]. Therefore, selective detection and quantification for zinc ion is very important object of increasing investigation [25–28]. Several traditional methods, such as inductively coupled plasma atomic emission spectrometry, atomic absorption spectroscopy, and electrochemical methods, have been applied for analysis of zinc in environmental samples and for diagnosis of its roles in body tissue [29– 32]. However, these methods require sophisticated instrumentations, tedious sample preparation procedures, and trained operators. By contrast, chemosensors with high selectivity and sensitivity are effective tools for the selective recognition of chemicals and biological species in environmental chemistry and biology[33–35], as they allow nondestructive and prompt detection of metal ions by simple fluorescence enhancement (turn-on) responses [26– 28]. In addition, the development of chemosensors that can distinguish Zn2+ from Cd2+ is still a huge subject as they are in the same group of the Periodic table and have similar properties [36–39]. Thus, low cost and easily-prepared Zn2+ fluorescence chemosensors are needed for convenience [40–48]. Based on the above-mentioned requirements, we attempted to design and synthesize a new 4
chemosensor for the detection of Zn2+. The quinoline moiety is a well-known fluorogenic chelator for Zn2+ among transition metal ions [39]. In addition, 2,2’-oxybis(ethan-1-amine) group, being hydrophilic in nature, would make water-solubility of a chemosensor increase. Therefore, we planned to connect two quinoline moieties by using the 2,2’-oxybis(ethan-1amine) group as a linker (Scheme 1) [26,49–52]. The resulting compound 1 with two quinoline moieties was expected to bind more effectively to Zn2+ ions and show unique photophysical and sensing properties toward it. Herein, we report a new chemosensor 1, consisting of the two quinoline moieties connected by 2,2’-oxybis(ethan-1-amine) as a linker. 1 showed an intense fluorescence enhancement in the presence of zinc ions in aqueous solution, discriminated Zn2+ from Cd2+, and sensed quantitatively Zn2+ in water samples, living cells and test kit. Additionally, the sensing mechanism of Zn2+ by 1 was explained by theoretical calculations.
Experimental section Materials and instrumentation All the solvents and reagents (analytical grade and spectroscopic grade) were obtained commercially and used as received. NMR spectra were recorded on a Varian 400 spectrometer. Chemical shifts (δ) were reported in ppm, relative to tetramethylsilane Si(CH 3)4. Absorption spectra were recorded at 25 °C using a Perkin Elmer model Lambda 2S UV/Vis spectrometer. The emission spectra were recorded on a Perkin-Elmer LS45 fluorescence spectrometer. Electrospray ionization mass spectra (ESI-MS) were collected on a Thermo Finnigan (San Jose, CA, USA) LCQTM Advantage MAX quadrupole ion trap instrument. Elemental analysis for carbon, nitrogen and hydrogen was carried out by using a MICRO CUBE elemental analyzer (Germany) in laboratory Center of Seoul National University of Science and Technology, Korea. 3-Chloro-N-(quinolin-8-yl)propanamide was obtained from the previous study [40].
Synthesis of sensor 1 2,2’-Oxybis(ethan-1-1amine)
(0.195mL,
2mmol)
and
3-chloro-N-(quinolin-8-
yl)propanamide (0.98 g, 4.4 mmol) and triethylamine (TEA, 0.56 mL, 4 mmol) were dissolved in acetonitrile (MeCN, 30 mL), stirred and refluxed for 2 d (Scheme 1). The oily residue was 5
dissolved in methylene chloride and then this solution was washed six times with water. Organic layer was dried over anhydrous Na2SO4. The solvent was removed under reduced pressure to obtain yellow oil, which was purified by silica gel column chromatography (9 : 4 : 4 : 1 v/v CH2Cl2-C6H14-CHCl3-CH3OH). Yield: 0.47 g (49.5 %). 1H NMR (400 MHz, DMSOd6, 25 °C): δ = 11.39 (s, 2H), 8.88 (d, J = 6 Hz, 2H), 8.70 (d, J = 8.8 Hz, 2H), 8.37 (d, J = 8 Hz, 2H), 7.60 (d, J = 8.4 Hz, 2H), 7.58-7.53 (m, 4H), 3.58 (t, J = 5.2 Hz, 4H), 3.36 (s, 4H), 2.94 (s, 2H), 2.78 (t, J = 5.2 Hz, 4H), 13C NMR (100 MHz, DMSO-d6, 25 °C): δ = 170.7 (2C), 148.9 (2C), 138.0 (2C), 136.4 (2C), 134.1 (2C), 127.8 (2C), 126.9 (2C), 122.0 (2C), 121.5 (2C), 115.3 (2C), 70.2 (2C), 53.3 (2C), 48.9 (2C) ppm. ESI-mass: m/z calcd for C26H27N6O3-+Zn2+: 535.14; found 535.10. Elemental analysis calcd (%) for C26H28N6O3: C, 66.09; H, 5.97; N, 17.78; O, 10.16 found: C, 66.59; H, 5.57; N, 17.49. Fluorescence titration of 1 with Zn2+ Sensor 1 (2.3 mg, 0.005 mmol) was dissolved in DMSO (1 mL) and 12 μL of the sensor 1 (5 mM) was diluted to 2.988 mL bis-tris buffer solution (10 mM, pH 7.0) to make the final concentration of 20 μM. Zn(NO3)2 (0.01 mmol) was dissolved in bis-tris buffer solution (1 mL) and 0.6-12.6 μL of the Zn2+ solution (10 mM) were added to the sensor 1 solution (20 μM, 3 mL) prepared above. After mixing them for a few seconds, fluorescence spectra were obtained at room temperature. UV-vis titration of 1 with Zn2+ Sensor 1 (2.3 mg, 0.005 mmol) was dissolved in DMSO (1 mL) and 12 μL of the sensor 1 (5 mM) was diluted to 2.988 mL bis-tris buffer solution (10 mM, pH 7.0) to make the final concentration of 20 μM. Zn(NO3)2 (0.01 mmol) was dissolved in bis-tris buffer solution (1 mL) and 1.5-15 μL of the Zn2+ solution (10 mM) were transferred to separate sensor solutions (20 μM, 3 mL). After mixing them for a few seconds, UV-vis spectra were taken at room temperature. Job plot measurement of 1 with Zn2+ Sensor 1 (47.3 mg, 0.1 mmol) was dissolved in DMSO (1 mL) and 320 μL of the sensor 1 (5 mM) was diluted to 39.68 mL bis-tris buffer solution (10 mM, pH 7.0) to make the final concentration of 40 μM. 2.7, 2.4, 2.0, 1.8, 1.5, 1.2, 1.0, 0.6 and 0.3 mL of the 1 solution were 6
taken and transferred to fluorescence cells. Zn(NO3)2 (0.02 mmol) was dissolved in bis-tris buffer solution (1 mL) and 80 μL of Zn2+ (20 mM) was diluted to 39.92 mL bis-tris buffer solution (10 mM, pH 7.0). 0.3, 0.6, 1.0, 1.2, 1.5, 1.8, 2.0, 2.4 and 2.7 mL of the Zn(NO3)2 solution were added to each sensor 1 solution. Each cell had a total volume of 3 mL. After mixing them for a few seconds, fluorescence spectra were taken at room temperature.
Competitive experiments Sensor 1 (2.3 mg, 0.005 mmol) was dissolved in DMSO (1 mL) and 12 μL of the sensor 1 (5 mM) was diluted to 2.988 mL bis-tris buffer solution (10 mM, pH 7.0) to make the final concentration of 20 μM . MNO3 (M = Na and K, 0.01 mmol) or M(NO3)2 (M = Mn, Co, Ni, Cu, Zn, Cd, Mg, Ca and Pb; 0.01 mmol) or M(NO3)3 (M = Al, Ga, In, Fe and Cr; 0.01 mmol) or M(ClO4)2 (M = Fe, 0.01 mmol) were dissolved in bis-tris buffer solution (1 mL). 12 μL of each metal solution (10 mM) was taken and added to 3 mL of the solution of 1 (20 μM). Then, 12 μL of the Zn2+ solution (10 mM) was taken and added to the mixed solution of each metal ion. After mixing them for a few seconds, fluorescence spectra were taken at room temperature. 1
H NMR titration of 1 with Zn2+ Four NMR tubes of 1 (0.92 mg, 0.002 mmol) dissolved in DMSO-d6 (0.7 mL) were prepared,
and four different equivalents (0, 0.2, 0.5, and 1 equiv) of zinc nitrate dissolved in DMSO-d6 (0.3 mL) were added separately to the solutions of 1. After shaking them for a few seconds, the 1
H NMR spectra were taken.
Methods of cell test of 1 with Zn 2+ HeLa cells (ATCC, Manassas, USA) were maintained in media containing DMEM, 10% fetal bovine serum (FBS, GIBCO, Grand Island, NY, USA), 100 U/mL penicillin (GIBCO), and 100 mg/mL streptomycin (GIBCO). The cells grew in a humidified atmosphere with 5 % CO2 at 37 °C. Cells were seeded onto 6 well plate (SPL Life Sciences Co., Ltd., South Korea) at a density of 150,000 cells per 1 mL and then incubated at 37 °C for 24 h. Cells were first treated with 1 (dissolved in DMSO; 1 % v/v final DMSO concentration; 20 μM; at room temperature) for 30 min. Prior to addition of various concentrations of zinc nitrate (dissolved in water; 1 % v/v), cells were washed with 2 mL of media. Imaging was performed with an EVOS FL fluorescence microscope (Life technologies) using a DAPI light cube (DAPI, 2-(47
amidinophenyl)-1H-indole-6-carboxamidine; excitation 357 (± 22) nm; emission 447 (± 30) nm). Determination of Zn2+ in water samples Fluorescence spectral measurement of water samples containing Zn2+ were carried out by adding 12 μL of 5 mmol/L stock solution of 1 and 0.30 mL of 100 mmol/L bis-tris buffer stock solution to 2.688 mL sample solutions, respectively. After well mixed, the solutions were allowed to stand at 25 ◦C for 2 min before the test.
Fluorescence test kit. Sensor 1-test kits were prepared by immersing filter papers into sensor 1 solution (50 mM), and then dried in air. Then, Zn(NO3)2 of different concentration ranges (10, 50, 70 µmol) was separately dissolved in bis-tris buffer (1 mL). The test kits prepared above were added into different concentrated solutions, and then dried at room temperature. Method for theoretical calculations of 1 and 1-Zn2+ complex All DFT/TDDFT calculations based on the hybrid exchange correlation functional B3LYP [53][54] were carried out using Gaussian 03 program [55]. The 6-31G** basis set [56,57] was used for the main group elements, whereas the Lanl2DZ effective core potential (ECP) [58–60] was employed for Zn. In vibrational frequency calculations, there was no imaginary frequency for the optimized geometries of 1 and 1-Zn2+, suggesting that these geometries represented local minima. For all calculations, the solvent effect of water was considered by using the Cossi and Barone’s CPCM (conductor-like polarizable continuum model) [61,62]. To investigate the electronic properties of singlet excited states, time-dependent DFT (TDDFT) was performed in the ground state geometries of 1 and 1-Zn2+. The 25 singlet-singlet excitations were calculated and analyzed. The GaussSum 2.1 [63] was used to calculate the contributions of molecular orbitals in electronic transitions.
Results and discussion The sensor 1 was synthesized by the substitution reaction of 2,2’-oxybis(ethan-1-1amine) and 3-chloro-N-(quinolin-8-yl)propanamide in acetonitrile (Scheme1), and analyzed by 1H NMR, 8
13
C NMR, ESI-mass, spectrometry and elemental analysis.
Fluorescence and absorption spectroscopic studies of 1 toward Zn 2+ We examined the fluorescent behavior of 1 toward various metal ions in different solvents such as MeOH, DMSO, MeCN and bis-tris buffer solution (Fig. S1). In DMSO and bis-tris buffer solution, 1 showed selectively a fluorescence enhancement at 504 nm with only Zn2+. For practical applications, the detailed fluorescent behavior of 1 toward various metal ions was studied in bis-tris buffer solution (10 mM, pH 7.0). When excited at 340 nm, 1 exhibited a weak fluorescence emission (max = 504 nm) compared to that (39 folds) in the presence of Zn2+ (Fig. 1). On the contrary, upon addition of other metal ions such as Na+, K+, Mg2+, Ca2+, Al3+, Ga3+, Cr3+, Mn2+, Fe2+, Fe3+, Co2+, Ni2+, Cu2+, Zn2+, Ga3+ and Pb2+, no or slight increase in intensity was observed. For example, 1 showed slight fluorescent enhancement to Cd2+, but 1 can clearly distinguish Zn2+ from Cd2+. These results showed that sensor 1 could be used as a fluorescence detector for Zn2+. To study the chemosensing properties of 1, fluorescence titration of the sensor 1 with Zn2+ ion was performed. As shown in Fig. 2, the emission intensity of 1 at 504 nm steadily increased until the amount of Zn2+ reached at 2.0 equiv. The quantum yields (Ф) of 1 and 1-Zn2+ complex were found to be 0.009 and 0.1, respectively. The photophysical properties of 1 were also examined using UV-vis spectrometry. UV-vis absorption spectrum of 1 showed an absorption band at 300 nm (Fig. S2). Upon the addition of Zn2+ ions to a solution of 1, the band has redshifted to 355 nm. Meanwhile, clear isosbestic points were observed at 279 nm and 333 nm, suggesting that only one product was produced from the binding of 1 with Zn2+. The Job plot [64] showed a 1:1 complexation stoichiometry between 1 and Zn2+ (Fig. 3), which was further confirmed by ESI-mass spectrometry analysis (Fig. 4). The positive-ion mass spectrum of 1 upon addition of 1 equiv of Zn2+ showed the formation of the 1-(H+)+Zn2+ [m/z: 535.14; calcd, 535.10]. From the fluorescence titration data, the association constant for 1 with Zn2+ was determined as 8.6 (±0.1) x 103 M-1 using Benesi-Hildebrand equation (Fig. S3) [65]. This value was within the range of those (1.0 ~ 1.0 x 1012) reported for Zn2+ sensing chemosensors [66–69]. The detection limit was estimated as 0.6 μM based on the 3σ/slope (Fig. S4) [70], which was about 1/130th of the WHO guideline (76 μM) for Zn2+ ions in drinking water [41,71]. 9
To check the practical applicability of 1 as Zn2+ selective sensor, fluorescence competition experiments were carried out in the presence of Zn2+ mixed with various metal ions (Fig. S5). When 1 was treated with 2.0 equiv of Zn2+ in the presence of the same concentrations of other metal ions, other background metal ions had no obvious interference with the detection of Zn2+ ion, except for Ca2+, Co2+, Cu2+ and Pb2+. The interferences with Co2+ and Cu2+ might be due to the strong quenching properties to fluorescence [72,73]. These results showed that 1 could be a good Zn2+ sensor which could discriminate Zn2+ from Cd2+ commonly having similar properties. The 1H NMR titration experiments were studied to further examine the binding mode between 1 and Zn2+ ion (Fig. S6). Upon addition of Zn2+ (1 equiv) to sensor 1, the integral of the protons H5 and H16 with upfield shift decreased to half. Moreover, all of the aromatic and aliphatic protons showed downfield shift with separation of each peak group into a pair, indicating that a Zn2+ ion might bind to only one of two quinolone moieties. These results suggested that the oxygen atom of the ether moiety and the three nitrogen atoms might coordinate to Zn2+ ion as shown in Scheme 2. This coordinative behavior of five-membered ring between 1 and a zinc ion was previously observed in the similar type of zinc complexes [42]. There was no shift in the position of proton signals on further addition of Zn2+ (>1.0 equiv), indicating that 1-Zn2+ complex might have a 1:1 binding mode. To investigate the practical applicability of 1, the effect of pH on the fluorescence response of Zn2+ was conducted at various pH (2-12) (Fig. 5). The fluorescence intensity of 1 in the presence of Zn2+ showed a significant response between pH 7 and 12. These results showed that Zn2+ could be distinctly detected by the fluorescence spectral measurement using 1 over the environmentally and physiologically relevant pH range (pH 7.0-8.4) [74], especially for monitoring Zn2+ in water samples and living cells. To examine the reversibility of sensor 1 toward Zn2+ in buffer solution, ethylenediaminetetraacetic acid (EDTA, 2.0 equiv) was added to the complexed solution of sensor 1 and Zn2+. As shown in Fig. 6, a fluorescence signal at 504 nm was immediately quenched. Upon addition of Zn2+ again, the fluorescence was recovered. The fluorescence emission changes were almost reversible even after several cycles with the sequentially alternative addition of Zn2+ and EDTA. These results indicated that sensor 1 could be recyclable simply through treatment with a proper reagent such as EDTA. Such reversibility and regeneration could be important for the fabrication of chemosensor to sense Zn2+. 10
To further demonstrate the potential of 1 to monitor Zn2+ in living matrices, fluorescence imaging experiments were carried out in living cells (Fig. 7). HeLa cell were first incubated with various concentrations of aqueous Zn2+ solutions (0, 100 and 200 μM) for 24 h and then exposed to 1 for 1 h before imaging. The experimental results showed that the HeLa cells without either Zn2+ or 1 showed negligible intracellular fluorescence, while those cultured with both Zn2+ and 1 exhibited fluorescence. In order to examine the practical properties of the chemosensor 1 in environmental samples, the chemosensor 1 was applied for the determination of Zn2+ in water samples, using a calibration curve of 1 toward Zn2+ (Fig. S7). Tap water and drinking water samples were chosen. Each sample was analyzed for three replicates. As shown in Table 1, suitable recoveries and R.S.D. values of the water samples were obtained. These results indicated that the sensor 1 can apply to determination of Zn2+ concentrations in environmental samples. For practical application, fluorescent test kits were prepared by immersing strips of filter paper in buffer solution of 1 and then dried in air. As shown in Fig. 8, when the strips were exposed to various concentrations of Zn2+ (20, 50 and 70 μM), fluorescence turned on. These results showed the potential use of sensor 1 for the detection of Zn2+ in water.
Theoretical calculations In order to get detailed insight into the sensing mechanism of 1 toward Zn2+, we conducted density functional theory (DFT) calculations with the B3LYP/6-31G (d, p) method basis set using the Gaussian 03 program. The major bond lengths and angles of 1 and 1-Zn2+ are shown in Fig. 9. The four-coordinated structure of 1-Zn2+ complex was optimized. Zn2+ was coordinated to 1N, 5N, 6N and 7O atoms of 1 and the bond lengths of Zn-1N, Zn-5N, Zn-6N and Zn-7O were similar to those of the reported Zn complexes [75,76]. Time-dependent density functional theory (TD-DFT) calculations were performed with the optimized geometries (S 0). In the case of 1, the main molecular orbital (MO) contribution of the first lowest excited state was determined for HOMO → LUMO + 1 transition (330.57 nm, Fig. S8). For the 1-Zn2+ complex, the main molecular orbital (MO) contribution of the first lowest excited state was determined for HOMO → LUMO transition (396.90, Fig. S9). The calculated HOMO → LUMO excitation of 1-Zn2+ complex indicated ICT transition from the carbonyl group to the quinolone group. Based on the molecular orbitals (MOs), the chelation of Zn2+ with 1 rendered the HOMO to LUMO energy gap of 1 decrease (4.277 eV → 3.727 eV) (Fig. S10), which is 11
consistent with the red shift in the UV-Vis spectrum of 1-Zn2+ complex. On the basis of UVvis titration, Job plot, 1H NMR titration, ESI-mass spectroscopy analysis, and theoretical calculations, we proposed the structure of a 1:1 complex of 1 and Zn2+ as shown in Scheme 2.
Conclusion We have developed a new quinoline-functionalized chemosensor 1, which detected Zn2+ ion in aqueous solution. 1 showed a highly selective fluorescence response to Zn2+ with the detection limit of 0.6 μM. Importantly, 1 can clearly distinguish Zn2+ from Cd2+, while such distinguishment of Zn2+ from Cd2+ is a well-known challenge. The binding of the sensor 1 and Zn2+ was chemically reversible with EDTA. In addition, 1 could be successfully applied to real water samples, living cells and test-kits. Moreover, the sensing mechanism of Zn2+ by 1 via ICT mechanism was explained by DFT calculations. Therefore, we believe that sensor 1 will be an excellent prototype for monitoring Zn2+ concentrations in biological and environmental environments.
Acknowledgements Financial support from Basic Science Research Program through the National Research Foundation of Korea (NRF) funded by the Ministry of Education, Science and Technology (NRF-2014R1A2A1A11051794,
NRF-2015R1A2A2A09001301,
and
NRF-
2014S1A2A2028270) are gratefully acknowledged. We thank Nano-Inorganic Laboratory, Department of Nano & Bio Chemistry, Kookmin University to access the Gaussian 03 program packages.
Appendix A. Supplementary data Supplementary data related to this article can be found at http://????.
12
References [1]
K. Komatsu, Y. Urano, H. Kojima, T. Nagano, Development of an iminocoumarin-
based zinc sensor suitable for ratiometric fluorescence imaging of neuronal zinc, J. Am. Chem. Soc. 129 (2007) 13447–13454. [2]
Y.H. Lee, M.H. Lee, J.F. Zhang, J.S. Kim, Pyrene excimer-based calix[4]arene FRET
chemosensor for mercury(II), J. Org. Chem. 75 (2010) 7159–7165. [3]
R.K. Pathak, V.K. Hinge, A. Rai, D. Panda, C.P. Rao, Imino-phenolic-pyridyl
conjugates of calix[4]arene (L1 and L2) as primary fluorescence switch-on sensors for Zn2+ in solution and in HeLa cells and the recognition of pyrophosphate and ATP by [ZnL2], Inorg. Chem. 51 (2012) 4994–5005. [4]
A. Ajayaghosh, P. Carol, S. Sreejith, A ratiometric fluorescence probe for selective
visual sensing of Zn2+, J. Am. Chem. Soc. 127 (2005) 14962–14963. [5]
Y. Wu, X. Peng, B. Guo, J. Fan, Z. Zhang, J. Wang, A. Cui and Y. Gao, Boron
dipyrromethene fluorophore based fluorescence sensor for the selective imaging of Zn(II) in living cells, Org. Biomol. Chem. 3 (2005) 1387–1392. [6]
P. Jiang, L. Chen, J. Lin, Q. Liu, J. Ding, X. Gao and Z. Guo, Novel zinc fluorescent
probe bearing dansyl and aminoquinoline groupsElectronic supplementary information (ESI) available: NMR spectra and assignment, UV titration details, crystal structure and competitive fluorescent experiments of L, Chem. Commun. (2002) 1424–1425. [7]
A. Loas, R.J. Radford, S.J. Lippard, Addition of a second binding site increases the
dynamic range but alters the cellular localization of a red fluorescent probe for mobile zinc, Inorg. Chem. 53 (2014) 6491–6493. [8]
K. Tayade, B. Bondhopadhyay, H. Sharma, A. Basu, V. Gite, S. Attarde, N. Singh and
A. Kuwar, “Turn-on” fluorescent chemosensor for zinc(II) dipodal ratiometric receptor: application in live cell imaging, Photochem. Photobiol. Sci. 13 (2014) 1052–1057. [9]
J.H. Weiss, S.L. Sensi, J.Y. Koh, Zn2+: a novel ionic mediator of neural injury in brain
disease, Trends Pharmacol. Sci. 21 (2000) 395–401. [10]
B.K.Y. Bitanihirwe, M.G. Cunningham, Zinc: the brain’s dark horse, Synapse. 63
(2009) 1029–1049. [11]
X.M. Xie, T.G. Smart, A physiological role for endogenous zinc in rat hippocampal
synaptic neurotransmission, Nature. 349 (1991) 521–524. [12]
C.E. Outten, T. V O’Halloran, Femtomolar sensitivity of metalloregulatory proteins 13
controlling zinc homeostasis, Science. 292 (2001) 2488–2492. [13]
Z. Xu, G.-H. Kim, S.J. Han, M.J. Jou, C. Lee, I. Shin and J.Y Yoon, An NBD-based
colorimetric and fluorescent chemosensor for Zn2+ and its use for detection of intracellular zinc ions, Tetrahedron. 65 (2009) 2307–2312. [14]
K. Li, A. Tong, A new fluorescent chemosensor for Zn2+ with facile synthesis: “Turn-
on” response in water at neutral pH and its application for live cell imaging, Sens. Actuators B. 184 (2013) 248–253. [15]
H.J. Kim, S.Y. Park, S. Yoon, J.S. Kim, FRET-derived ratiometric fluorescence sensor
for Cu2+, Tetrahedron. 64 (2008) 1294–1300. [16]
M. Shellaiah, Y.-H. Wu, H.-C. Lin, Simple pyridyl-salicylimine-based fluorescence
“turn-on” sensors for distinct detections of Zn2+, Al3+ and OH- ions in mixed aqueous media., Analyst. 138 (2013) 2931–2942. [17]
G. J. Park, H. Kim, J. J. Lee, Y. S. Kim, S. Y. Lee, S.Y. Lee, I.S Noh and C. Kim, A
highly selective turn-on chemosensor capable of monitoring Zn2+ concentrations in living cells and aqueous solution, Sens. Actuators B. 215 (2015) 568–576. [18]
C. Gao, X. Jin, X. Yan, P. An, Y. Zhang, L. Liu, H. Tian, W. Liu, X. Yao and Y. Tang,
A small molecular fluorescent sensor for highly selectivity of zinc ion, Sens. Actuators B, 176 (2013) 775–781. [19]
Y.J. Na, I.H. Hwang, H.Y. Jo, S.A. Lee, G.J. Park, C. Kim, Fluorescent chemosensor
based-on the combination of julolidine and furan for selective detection of zinc ion, Inorg. Chem. Commun. 35 (2013) 342–345. [20]
S. Cui, G. Liu, S. Pu, B. Chen, A highly selective fluorescent probe for Zn2+ based on
a new photochromic diarylethene with a di-2-picolylamine unit, Dye. Pigm. 99 (2013) 950– 956. [21]
Y.-P. Li, Q. Zhao, H.-R. Yang, S.-J. Liu, X.-M. Liu, Y.-H. Zhang, T.-L. Hu, J.-T. Chen,
Z. Chang and Z.-H. Bu, A new ditopic ratiometric receptor for detecting zinc and fluoride ions in living cells, Analyst. 138 (2013) 5486–5494. [22]
M. Yan, T. Li, Z. Yang, A novel coumarin Schiff-base as a Zn(II) ion fluorescent sensor,
Inorg. Chem. Commun. 14 (2011) 463–465. [23]
H. Kim, J. Kang, K.B. Kim, E.J. Song, C. Kim, A highly selective quinoline-based
fluorescent sensor for Zn(II), Spectrochim. Acta. A. Mol. Biomol. Spectrosc. 118 (2014) 883– 887. 14
[24]
V. Bhalla, H. Arora, A. Dhir, M. Kumar, A triphenylene based zinc ensemble as an
oxidation inhibitor., Chem. Commun. 48 (2012) 4722–4724. [25]
S.Y. Park, J.H. Yoon, C.S. Hong, R. Souane, J.S. Kim, S.E. Matthews and Jacques
Vicens, A pyrenyl-appended triazole-based calix[4]arene as a fluorescent sensor for Cd2+ and Zn2+, J. Org. Chem. 73 (2008) 8212–8218. [26]
Z. Liu, C. Zhang, Y. Chen, W. He, Z. Guo, An excitation ratiometric Zn2+ sensor with
mitochondria-targetability for monitoring of mitochondrial Zn2+ release upon different stimulations, Chem. Commun. 48 (2012) 8365–8367 [27]
K. Tayade, S.K. Sahoo, B. Bondhopadhyay, V.K. Bhardwaj, N. Singh, A. Basu and R.
Bendre, Highly selective turn-on fluorescent sensor for nanomolar detection of biologically important Zn2+ based on isonicotinohydrazide derivative: application in cellular imaging, Biosens. Bioelectron. 61 (2014) 429–433. [28]
V.K. Gupta, N. Mergu, A.K. Singh, Fluorescent chemosensors for Zn2+ ions based on
flavonol derivatives, Sens. Actuators B. 202 (2014) 674–682. [29]
A. Rajabi Khorrami, A.R. Fakhari, M. Shamsipur, H. Naeimi, Pre-concentration of
ultra trace amounts of copper, zinc, cobalt and nickel in environmental water samples using modified C18 extraction disks and determination by inductively coupled plasma–optical emission spectrometry, Int. J. Environ. Anal. Chem. 89 (2009) 319–329. [30]
M. Ghaedi, F. Ahmadi, A. Shokrollahi, Simultaneous preconcentration and
determination of copper, nickel, cobalt and lead ions content by flame atomic absorption spectrometry, J. Hazard. Mater. 142 (2007) 272–278. [31]
K. Sreenivasa Rao, T. Balaji, T. Prasada Rao, Y. Babu, G.R.K. Naidu, Determination
of iron, cobalt, nickel, manganese, zinc, copper, cadmium and lead in human hair by inductively coupled plasma-atomic emission spectrometry, Spectrochim. Acta Part B. 57 (2002) 1333–1338. [32]
R. gulaboski, V. mireski, F. scholz, An electrochemical method for determination of
the standard Gibbs energy of anion transfer between water and n-octanol, Electrochem. Commun. 4 (2002) 277–283. [33]
Z. Li, M. Yu, L. Zhang, M. Yu, J. Liu, L. Wei, and H. Zhang, A “switching on”
fluorescent chemodosimeter of selectivity to Zn2+ and its application to MCF-7 cells, Chem. Commun. 46 (2010) 7169–7171. [34]
U.C. Saha, B. Chattopadhyay, K. Dhara, S.K. Mandal, S. Sarkar, A.R. Khuda-Bukhsh, 15
M. Mukherjee, M. Helliwell and P. Chattopadhyay, A Highly Selective Fluorescent Chemosensor for Zinc Ion and Imaging Application in Living Cells, Inorg. Chem. 50 (2011) 1213-1219. [35]
G.J. Park, Y.J. Na, H.Y. Jo, S.A. Lee, A.R. Kim, I. Noh and C. Kim, A single
chemosensor for multiple analytes: fluorogenic detection of Zn2+ and OAc− ions in aqueous solution, and an application to bioimaging, New J. Chem. 38 (2014) 2587-2594. [36]
V. Bhalla, Roopa, M. Kumar, Pentaquinone based probe for nanomolar detection of
zinc ions: chemosensing ensemble as an antioxidant., Dalton Trans. 42 (2013) 975–80. [37]
Y. Ma, F. Wang, S. Kambam, X. Chen, A quinoline-based fluorescent chemosensor for
distinguishing cadmium from zinc ions using cysteine as an auxiliary reagent, Sens. Actuators B. 188 (2013) 1116–1122. [38]
K.M.K. Swamy, M.-J. Kim, H.-R. Jeon, J.-Y. Jung, J.-Y. Yoon, New 7-
Hydroxycoumarin-Based Fluorescent Chemosensors for Zn(II) and Cd(II), Bull. Korean Chem. Soc. 31 (2010) 3611–3616. [39]
Z. Xu, J.-Y Yoon, D.R. Spring, Fluorescent chemosensors for Zn2+, Chem. Soc. Rev.
39 (2010) 1996–2006. [40]
E.J. Song, J. Kang, G.R. You, G.J. Park, Y. Kim, S.-J. Kim, C. Kim and Roger G.
Harrison, A single molecule that acts as a fluorescence sensor for zinc and cadmium and a colorimetric sensor for cobalt, Dalton Trans. 42 (2013) 15514–20. [41]
H. Gyu Lee, J. Hoon Lee, S. Pyo Jang, H. Min Park, S. Jin Kim, Y. Mee Kim, Cheal
Kim, Zinc selective chemosensor based on pyridyl-amide fluorescence, Tetrahedron. 67 (2011) 8073–8078. [42]
H.G. Lee, J.H. Lee, S.P. Jang, I.H. Hwang, S.-J. Kim, Y. Kim, C. Kim, Zinc selective
chemosensors based on the flexible dipicolylamine and quinoline, Inorganica Chim. Acta. 394 (2013) 542–551. [43]
E. J. Song, G. J. Park, J. J. Lee, S.Y. Lee, I. Noh, Y. C. Kim and Roger G. Harrison, A
fluorescence sensor for Zn2+ that also acts as a visible sensor for Co2+ and Cu2+, Sens. Actuators B. 213 (2015) 268–275. [44]
H. Kim, G.R. You, G.J. Park, J.Y. Choi, I. Noh, Y. Kim, S.J. Kim, C. Kim and Rogger
G. Harrion, Selective zinc sensor based on pyrazoles and quinoline used to image cells, Dye. Pigm. 113 (2015) 723–729. [45]
S. Sinha, T. Mukherjee, J. Mathew, S.K. Mukhopadhyay, S. Ghosh, Triazole-based 16
Zn2+-specific molecular marker for fluorescence bioimaging, Anal. Chim. Acta. 822 (2014) 60–68. [46]
K. Hanaoka, Y. Muramatsu, Y. Urano, T. Terai, T. Nagano, Design and synthesis of a
highly sensitive off-on fluorescent chemosensor for zinc ions utilizing internal charge transfer., Chemistry. 16 (2010) 568–572. [47]
D. Wang, X. Xiang, X. Yang, X. Wang, Y. Guo, W. Liu and W. Qin, Fluorescein-based
chromo-fluorescent probe for zinc in aqueous solution: Spirolactam ring opened or closed?, Sens. Actuators B. 201 (2014) 246–254. [48]
L. Zang, H. Shang, D. Wei, S. Jiang, A multi-stimuli-responsive organogel based on
salicylidene Schiff base, Sens. Actuators B. 185 (2013) 389–397. [49]
A. Ojida, Y. Mito-oka, K. Sada, I. Hamachi, Molecular recognition and fluorescence
sensing of monophosphorylated peptides in aqueous solution by bis(zinc(II)-dipicolylamine)based artificial receptors, J. Am. Chem. Soc. 126 (2004) 2454–2463. [50]
C. Lakshmi, R.G. Hanshaw, B.D. Smith, Fluorophore-linked zinc(II)dipicolylamine
coordination complexes as sensors for phosphatidylserine-containing membranes, Tetrahedron. 60 (2004) 11307–11315. [51]
B.A. Wong, S. Friedle, S.J. Lippard, Subtle modification of 2,2-dipicolylamine lowers
the affinity and improves the turn-on of Zn(II)-selective fluorescent sensors, Inorg. Chem. 48 (2009) 7009–7011. [52]
H. Kim, Y. Liu, D. Nam, Y. Li, S. Park, J.-Y Yoon, M. H. Hyun, A new phosphorescent
chemosensor bearing Zn-DPA sites for H2PO4−, Dye. Pigm. 106 (2014) 20–24. [53]
A.D. Becke, Density-functional thermochemistry. III. The role of exact exchange, J.
Chem. Phys. 98 (1993) 5648–5652. [54]
C. Lee, W. Yang, R.G. Parr, Development of the Colle-Salvetti correlation-energy
formula into a functional of the electron density, Phys. Rev. B. 37 (1988) 785–789. [55]
C.G. and J.A.P. M. J. Frisch, G. W. Trucks, H. B. Schlegel, G. E. Scuseria, M. A. Robb,
J. R. Cheeseman, J. A. Montgomery Jr., T. Vreven, K. N. Kudin, J. C. Burant, J.M.Millam, S. S. Iyengar, J. Tomasi, V. Barone, B. Mennucci, M. Cossi, G. Scalmani, N. Rega, G. A. Peters, GAUSSIAN 03 (Revision B.02), Gaussian, Inc., Wallingford CT. (2004). [56]
P.C. Hariharan, J.A. Pople, The influence of polarization functions on molecular
orbital hydrogenation energies, Theor. Chim. Acta. 28 (1973) 213–222. [57]
M.M. Francl, W.J. Pietro, W.J. Hehre, J.S. Binkley, M.S. Gordon, D.J. DeFrees and 17
J.A. Pople, Self-Consistent Molecular Orbital Methods. 23. A polarization-type basis set for 2nd-row elements, J. Chem. Phys. 77 (1982) 3654–3665. [58]
P.J. Hay, W.R. Wadt, Ab initio effective core potentials for molecular calculations.
Potentials for the transition metal atoms Sc to Hg, J. Chem. Phys. 82 (1985) 270–283. [59]
W.R. Wadt, P.J. Hay, Ab initio effective core potentials for molecular calculations.
Potentials for main group elements Na to Bi, J. Chem. Phys. 82 (1985) 284–298. [60]
W.R. Wadt, P.J. Hay, Ab initio effective core potentials for molecular calculations.
Potentials for K to Au including the outermost core orbitals, J. Chem. Phys. 82 (1985) 299– 310. [61]
V. Barone, M. Cossi, Quantum calculation of molecular energies and energy gradients
in solution by a conductor solvent model, J. Phys. Chem. A. 102 (1998) 1995–2001. [62]
M. Cossi, V. Barone, Time-dependent density functional theory for molecules in liquid
solutions, J. Chem. Phys. 115 (2001) 4708–4717. [63]
N.M. O’Boyle, A.L. Tenderholt, K.M. Langner, cclib: a library for package-
independent computational chemistry algorithms, J. Comput. Chem. 29 (2008) 839–845. [64]
P. Job, Formation and stability of inorganic complexes in solution, Ann. Chim. 9 (1928)
113–203. [65]
H.A. Benesi, J.H. Hildebrand, A Spectrophotometric Investigation of the Interaction
of Iodine with Aromatic Hydrocarbons, J. Am. Chem. Soc. 71 (1949) 2703–2707. [66]
H.-Y. Lin, P.-Y. Cheng, C.-F. Wan, A.-T. Wu, A turn-on and reversible fluorescence
sensor for zinc ion, Analyst. 137 (2012) 4415–4417. [67]
J.H. Kim, I.H. Hwang, S.P. Jang, J. Kang, S. Kim, I. Noh, Y. Kim, C. Kim and Roger
G. Harrison, Zinc sensors with lower binding affinities for cellular imaging, Dalton Trans. 42 (2013) 5500–5507. [68]
W.H. Hsieh, C.-F. Wan, D.-J. Liao, A.-T. Wu, A turn-on Schiff base fluorescence
sensor for zinc ion, Tetrahedron Lett. 53 (2012) 5848–5851. [69]
Y. Zhou, Z.-X. Li, S.-Q. Zang, Y.-Y. Zhu, H.-Y. Zhang, H.-W. Hou and T. C. W. Mak,
A novel sensitive turn-on fluorescent Zn2+ chemosensor based on an easy to prepare C3symmetric Schiff-base derivative in 100% aqueous solution, Org. Lett. 14 (2012) 1214–1217. [70] Y.-K. Tsui, S. Devaraj, Y.-P. Yen, Azo dyes featuring with nitrobenzoxadiazole (NBD) unit: A new selective chromogenic and fluorogenic sensor for cyanide ion, Sens. Actuators B. 161 (2012) 510–519. 18
[71]
Y.P. Kumar, P. King, V.S.R.K. Prasad, Zinc biosorption on Tectona grandis L.f. leaves
biomass: Equilibrium and kinetic studies, Chem. Eng. J. 124 (2006) 63–70. [72]
Z. Liu, Z. Yang, T. Li, B. Wang, Y. Li, D. Qin, M. Wang and M. Yan, An effective
Cu(II) quenching fluorescence sensor in aqueous solution and 1D chain coordination polymer framework, Dalton Trans. 40 (2011) 9370–9373. [73]
R. Homan, M. Eisenberg, A fluorescence quenching technique for the measurement of
paramagnetic ion concentrations at the membrane/water interface. Intrinsic and X537Amediated cobalt fluxes across lipid bilayer membranes, Biochim. Biophys. Acta - Biomembr. 812 (1985) 485–492. [74]
R.M. Harrison, D.P.H. Laxen, S.J. Wilson, Chemical associations of lead, cadmium,
copper, and zinc in street dusts and roadside soils, Environ. Sci. Technol. 15 (1981) 1378–1383. [75]
C. Bergquist, G. Parkin, Modeling the Catalytic Cycle of Liver Alcohol
Dehydrogenase:
Synthesis and Structural Characterization of a Four-Coordinate Zinc
Ethoxide Complex and Determination of Relative Zn−OR versus Zn−OH Bond Energies, Inorg. Chem. 38 (1999) 422–423. [76]
J.J. Lee, S.A. Lee, H. Kim, L. Nguyen, I. Noh, C. Kim, A highly selective CHEF-type
chemosensor for monitoring Zn2+ in aqueous solution and living cells, RSC Adv. 5 (2015) 41905–41913.
19
Biographies
Jae Min Jung earned the BS degree in 2016 at Seoul National University of Science and Technology. He is currently a Master Student at Seoul National University of Science and Technology. His scientific interest includes chemical sensors, DNA cleavage by the transition metal complexes and synthesis of catalyst.
Seong Youl Lee received the BS degree in 2015 at Seoul National University of Science and Technology. He is currently a master student at Seoul National University of Science and Technology. His research interest includes chemical sensors, synthesis of catalyst and inorganic medicine.
Eunju Nam received her BS degree in 2015 at Ulsan National Institute of Science and Technology (UNIST). She is currently a master student at UNIST. Her research interests lie in understanding how small molecules can control pathological factors and pathways in human diseases. Mi Hee Lim is currently an associate professor at UNIST. She received her PhD degree at Massachusetts Institute of Technology (MIT) in USA (2016) and conducted her postdoctoral research at California Institute of Technology (Caltech) (2016-2018). Since 2008, Dr. Lim and her group have been identifying the roles of metals, proteins, and reactive oxygen species in human neurodegenerative disorders.
Cheal Kim is currently a professor at Seoul National University of Science and Technology. He received the PhD degree in 1993 at University of California, San Diego in USA. He is now interested in development of chemical sensors, reactivity study of the transition metal complexes, DNA cleavage by their metal complexes and MOF.
20
Figure Captions Figure 1. Fluorescence spectral changes of 1 (20 µM) in the presence of different metal ions (2.0 equiv) such as Na+, K+, Mg2+, Ca2+, Al3+,Ga3+, Cr3+, Mn2+, Fe2+, Fe3+, Co2+, Ni2+, Cu2+, Zn2+, Ga3+ and Pb2+ with an excitation of 340 nm in buffer solution (10 mM bis-tris, pH 7.0). Figure 2. Fluorescence spectral changes of 1 (20 μM) in the presence of different concentrations of Zn2+ ions in buffer solution (10 mM bis-tris, pH 7.0). Inset: Fluorescence intensity at 504 nm versus the number of equiv of Zn2+ added.
Figure 3. Job plot for the binding of 1 with Zn2+. Intensity at 504 nm was plotted as a function of the molar ratio [Zn2+]/([1] + [Zn2+]). The total concentration of zinc ions with sensor 1 was 4.0 x 10-5 M. Figure 4. Positive-ion electrospray ionization mass spectrum of 1 (100 μM) upon addition of Zn(NO3)2 (1 equiv). Figure 5. Fluorescence intensity (at 504 nm) of 1 in the presence of Zn2+ at different pH values (2-12) in buffer solution (10 mM bis-tris, pH 7.0).
Figure 6. Reversible changes (at 504 nm) of 1 (20 μM) upon the sequential addition of Zn2+ and EDTA.
Figure 7. Fluorescent responses of 1 to Zn(II) in HeLa cells. Cells were preincubated with 1 for 30 min prior to addition of various concentrations of Zn(II). Conditions: [Zn(II)] = 0, 100, and 200 μM; [1] = 20 μM; 37 °C; 5% CO2. The scale bar is 50 μm. Figure 8. Detection of Zn2+ by test strips coated with 1. Dark blue color indicated fluorescenceoff and light blue color did fluorescence-on. Conditions: [Zn(II)] = 20, 50 and 70 μM; [1] = 50 mM.
21
Figure 9. The energy-minimized structures of (a) 1 and (b) 1-Zn2+ complex.
22
Fig. 1.
23
Fig. 2.
24
Fig. 3.
25
Fig. 4.
26
Fig. 5.
27
Fig. 6.
28
Fig. 7.
29
Fig. 8.
30
(a)
(b)
Fig. 9.
31
Scheme 1. Synthesis of 1.
32
Scheme 2. Fluorescence enhancement mechanism and proposed structure of 1-Zn2+ complex.
33
Table 1. Determination of Zn(II) in water samples Sample
Zn(II) added (μmol/L)
Zn(II) found (μmol/L)
Tap water
0.00
0.00
4.00a
3.8
0.00
0.00
4.00b
4.2
Drinking water
Recovery (%)
R.S.D. (n = 3) (%) 2.17
95
0.2 0.33
105
0.83
Conditions: [1] = 20 μmol/L in 10 mM bis-tris buffer-DMSO solution (999:1, pH 7.0). a. 4.00 μmol/L of Zn2+ ion was artificially added into tap water. b. 4.00 μmol/L of Zn 2+ ion was artificially added into drinking water.
34
S0925-4005(17)30082-5 http://dx.doi.org/doi:10.1016/j.snb.2017.01.076 SNB 21601
To appear in:
Sensors and Actuators B
Received date: Revised date: Accepted date:
9-11-2016 10-1-2017 10-1-2017
Please cite this article as: Jae Min Jung, Seong Youl Lee, Eunju Nam, Mi Hee Lim, Cheal Kim, A highly selective turn-on chemosensor for Zn2+ in aqueous media and living cells, Sensors and Actuators B: Chemical http://dx.doi.org/10.1016/j.snb.2017.01.076 This is a PDF file of an unedited manuscript that has been accepted for publication. As a service to our customers we are providing this early version of the manuscript. The manuscript will undergo copyediting, typesetting, and review of the resulting proof before it is published in its final form. Please note that during the production process errors may be discovered which could affect the content, and all legal disclaimers that apply to the journal pertain.
A highly selective turn-on chemosensor for Zn2+ in aqueous media and living cells Jae Min Jung,a Seong Youl Lee,a Eunju Nam,b Mi Hee Lim,b* Cheal Kima* a
Department of Fine Chemistry and Department of Interdisciplinary Bio IT Materials, Seoul National University of Science and Technology, Seoul 139-743, Republic of Korea. Fax: +822-973-9149; Tel: +82-2-970-6693; E-mail: [email protected] b Department of Chemistry, Ulsan National Institute of Science and Technology (UNIST), Ulsan 44919, Republic of Korea. Fax: +82-52-217-5409; Tel: +82-52-217-5422; E-mail: [email protected]
1
Graphical abstract
Highlights
A simple and easy-to-make fluorescence sensor 1 for Zn2+ was designed and synthesized. Sensor 1 could detect Zn2+ ions at much lower concentration than WHO guideline. 1 was applied to quantify and image Zn2+ in water samples, test kit and living cells. Sensing mechanism of Zn2+ by 1 via ICT was explained by theoretical calculations.
2
Abstract A new simple quinoline-based chemosensor 1 was synthesized for Zn2+. 1 showed the selective fluorescence enhancement in the presence of Zn2+ with a 1:1 stoichiometry in a nearperfect aqueous solution (bis-tris buffer:DMSO = 999:1), which was reversible with the addition of ethylenediaminetetraacetic acid (EDTA). The detection limit (0.6 μM) of 1 for Zn2+ was much lower than World Health Organization guideline (76 μM) in drinking water. 1 was successfully applied to quantify and image Zn2+ in water samples, test kit and living cells. The sensing mechanism of Zn2+ by 1 via the intramolecular charge transfer was explained by theoretical calculations.
Keywords: fluorescent chemosensor, zinc ion, cell imaging, test kit, theoretical calculations
3
Introduction Much attention has been recently focused on the design and synthesis of highly selective fluorescent chemosensors for sensing and monitoring heavy and transitional metal ions due to their easy monitoring, low cost and high selectivity. Zinc ion is one of the most abundant metal ions present in living organism, because of its rich coordination chemistry [1–4]. Although most of the Zn2+ ions in a cell are strongly bound to metalloproteins, free forms are also found throughout the cell. In the brain, 5-20% of the total Zn2+ is stored in presynaptic vesicles, and the highest intracellular free Zn2+ is found in the hippocampus. Zn2+ modulates brain excitability and plays a key role in synaptic plasticity [5–8]. On the other hand, the deficiency of zinc causes unbalanced metabolism, which in turn can result in retarded growth in children, brain disorders and high blood cholesterol, and also be connected in numerous neurodegenerative disorders, such as Alzheimer’s disease, epilepsy, ischemic stroke, and infantile diarrhea [9–18]. In addition, an excess of zinc in the environment may reduce the soil microbial activity which results in phytotoxic effect [19–24]. Therefore, selective detection and quantification for zinc ion is very important object of increasing investigation [25–28]. Several traditional methods, such as inductively coupled plasma atomic emission spectrometry, atomic absorption spectroscopy, and electrochemical methods, have been applied for analysis of zinc in environmental samples and for diagnosis of its roles in body tissue [29– 32]. However, these methods require sophisticated instrumentations, tedious sample preparation procedures, and trained operators. By contrast, chemosensors with high selectivity and sensitivity are effective tools for the selective recognition of chemicals and biological species in environmental chemistry and biology[33–35], as they allow nondestructive and prompt detection of metal ions by simple fluorescence enhancement (turn-on) responses [26– 28]. In addition, the development of chemosensors that can distinguish Zn2+ from Cd2+ is still a huge subject as they are in the same group of the Periodic table and have similar properties [36–39]. Thus, low cost and easily-prepared Zn2+ fluorescence chemosensors are needed for convenience [40–48]. Based on the above-mentioned requirements, we attempted to design and synthesize a new 4
chemosensor for the detection of Zn2+. The quinoline moiety is a well-known fluorogenic chelator for Zn2+ among transition metal ions [39]. In addition, 2,2’-oxybis(ethan-1-amine) group, being hydrophilic in nature, would make water-solubility of a chemosensor increase. Therefore, we planned to connect two quinoline moieties by using the 2,2’-oxybis(ethan-1amine) group as a linker (Scheme 1) [26,49–52]. The resulting compound 1 with two quinoline moieties was expected to bind more effectively to Zn2+ ions and show unique photophysical and sensing properties toward it. Herein, we report a new chemosensor 1, consisting of the two quinoline moieties connected by 2,2’-oxybis(ethan-1-amine) as a linker. 1 showed an intense fluorescence enhancement in the presence of zinc ions in aqueous solution, discriminated Zn2+ from Cd2+, and sensed quantitatively Zn2+ in water samples, living cells and test kit. Additionally, the sensing mechanism of Zn2+ by 1 was explained by theoretical calculations.
Experimental section Materials and instrumentation All the solvents and reagents (analytical grade and spectroscopic grade) were obtained commercially and used as received. NMR spectra were recorded on a Varian 400 spectrometer. Chemical shifts (δ) were reported in ppm, relative to tetramethylsilane Si(CH 3)4. Absorption spectra were recorded at 25 °C using a Perkin Elmer model Lambda 2S UV/Vis spectrometer. The emission spectra were recorded on a Perkin-Elmer LS45 fluorescence spectrometer. Electrospray ionization mass spectra (ESI-MS) were collected on a Thermo Finnigan (San Jose, CA, USA) LCQTM Advantage MAX quadrupole ion trap instrument. Elemental analysis for carbon, nitrogen and hydrogen was carried out by using a MICRO CUBE elemental analyzer (Germany) in laboratory Center of Seoul National University of Science and Technology, Korea. 3-Chloro-N-(quinolin-8-yl)propanamide was obtained from the previous study [40].
Synthesis of sensor 1 2,2’-Oxybis(ethan-1-1amine)
(0.195mL,
2mmol)
and
3-chloro-N-(quinolin-8-
yl)propanamide (0.98 g, 4.4 mmol) and triethylamine (TEA, 0.56 mL, 4 mmol) were dissolved in acetonitrile (MeCN, 30 mL), stirred and refluxed for 2 d (Scheme 1). The oily residue was 5
dissolved in methylene chloride and then this solution was washed six times with water. Organic layer was dried over anhydrous Na2SO4. The solvent was removed under reduced pressure to obtain yellow oil, which was purified by silica gel column chromatography (9 : 4 : 4 : 1 v/v CH2Cl2-C6H14-CHCl3-CH3OH). Yield: 0.47 g (49.5 %). 1H NMR (400 MHz, DMSOd6, 25 °C): δ = 11.39 (s, 2H), 8.88 (d, J = 6 Hz, 2H), 8.70 (d, J = 8.8 Hz, 2H), 8.37 (d, J = 8 Hz, 2H), 7.60 (d, J = 8.4 Hz, 2H), 7.58-7.53 (m, 4H), 3.58 (t, J = 5.2 Hz, 4H), 3.36 (s, 4H), 2.94 (s, 2H), 2.78 (t, J = 5.2 Hz, 4H), 13C NMR (100 MHz, DMSO-d6, 25 °C): δ = 170.7 (2C), 148.9 (2C), 138.0 (2C), 136.4 (2C), 134.1 (2C), 127.8 (2C), 126.9 (2C), 122.0 (2C), 121.5 (2C), 115.3 (2C), 70.2 (2C), 53.3 (2C), 48.9 (2C) ppm. ESI-mass: m/z calcd for C26H27N6O3-+Zn2+: 535.14; found 535.10. Elemental analysis calcd (%) for C26H28N6O3: C, 66.09; H, 5.97; N, 17.78; O, 10.16 found: C, 66.59; H, 5.57; N, 17.49. Fluorescence titration of 1 with Zn2+ Sensor 1 (2.3 mg, 0.005 mmol) was dissolved in DMSO (1 mL) and 12 μL of the sensor 1 (5 mM) was diluted to 2.988 mL bis-tris buffer solution (10 mM, pH 7.0) to make the final concentration of 20 μM. Zn(NO3)2 (0.01 mmol) was dissolved in bis-tris buffer solution (1 mL) and 0.6-12.6 μL of the Zn2+ solution (10 mM) were added to the sensor 1 solution (20 μM, 3 mL) prepared above. After mixing them for a few seconds, fluorescence spectra were obtained at room temperature. UV-vis titration of 1 with Zn2+ Sensor 1 (2.3 mg, 0.005 mmol) was dissolved in DMSO (1 mL) and 12 μL of the sensor 1 (5 mM) was diluted to 2.988 mL bis-tris buffer solution (10 mM, pH 7.0) to make the final concentration of 20 μM. Zn(NO3)2 (0.01 mmol) was dissolved in bis-tris buffer solution (1 mL) and 1.5-15 μL of the Zn2+ solution (10 mM) were transferred to separate sensor solutions (20 μM, 3 mL). After mixing them for a few seconds, UV-vis spectra were taken at room temperature. Job plot measurement of 1 with Zn2+ Sensor 1 (47.3 mg, 0.1 mmol) was dissolved in DMSO (1 mL) and 320 μL of the sensor 1 (5 mM) was diluted to 39.68 mL bis-tris buffer solution (10 mM, pH 7.0) to make the final concentration of 40 μM. 2.7, 2.4, 2.0, 1.8, 1.5, 1.2, 1.0, 0.6 and 0.3 mL of the 1 solution were 6
taken and transferred to fluorescence cells. Zn(NO3)2 (0.02 mmol) was dissolved in bis-tris buffer solution (1 mL) and 80 μL of Zn2+ (20 mM) was diluted to 39.92 mL bis-tris buffer solution (10 mM, pH 7.0). 0.3, 0.6, 1.0, 1.2, 1.5, 1.8, 2.0, 2.4 and 2.7 mL of the Zn(NO3)2 solution were added to each sensor 1 solution. Each cell had a total volume of 3 mL. After mixing them for a few seconds, fluorescence spectra were taken at room temperature.
Competitive experiments Sensor 1 (2.3 mg, 0.005 mmol) was dissolved in DMSO (1 mL) and 12 μL of the sensor 1 (5 mM) was diluted to 2.988 mL bis-tris buffer solution (10 mM, pH 7.0) to make the final concentration of 20 μM . MNO3 (M = Na and K, 0.01 mmol) or M(NO3)2 (M = Mn, Co, Ni, Cu, Zn, Cd, Mg, Ca and Pb; 0.01 mmol) or M(NO3)3 (M = Al, Ga, In, Fe and Cr; 0.01 mmol) or M(ClO4)2 (M = Fe, 0.01 mmol) were dissolved in bis-tris buffer solution (1 mL). 12 μL of each metal solution (10 mM) was taken and added to 3 mL of the solution of 1 (20 μM). Then, 12 μL of the Zn2+ solution (10 mM) was taken and added to the mixed solution of each metal ion. After mixing them for a few seconds, fluorescence spectra were taken at room temperature. 1
H NMR titration of 1 with Zn2+ Four NMR tubes of 1 (0.92 mg, 0.002 mmol) dissolved in DMSO-d6 (0.7 mL) were prepared,
and four different equivalents (0, 0.2, 0.5, and 1 equiv) of zinc nitrate dissolved in DMSO-d6 (0.3 mL) were added separately to the solutions of 1. After shaking them for a few seconds, the 1
H NMR spectra were taken.
Methods of cell test of 1 with Zn 2+ HeLa cells (ATCC, Manassas, USA) were maintained in media containing DMEM, 10% fetal bovine serum (FBS, GIBCO, Grand Island, NY, USA), 100 U/mL penicillin (GIBCO), and 100 mg/mL streptomycin (GIBCO). The cells grew in a humidified atmosphere with 5 % CO2 at 37 °C. Cells were seeded onto 6 well plate (SPL Life Sciences Co., Ltd., South Korea) at a density of 150,000 cells per 1 mL and then incubated at 37 °C for 24 h. Cells were first treated with 1 (dissolved in DMSO; 1 % v/v final DMSO concentration; 20 μM; at room temperature) for 30 min. Prior to addition of various concentrations of zinc nitrate (dissolved in water; 1 % v/v), cells were washed with 2 mL of media. Imaging was performed with an EVOS FL fluorescence microscope (Life technologies) using a DAPI light cube (DAPI, 2-(47
amidinophenyl)-1H-indole-6-carboxamidine; excitation 357 (± 22) nm; emission 447 (± 30) nm). Determination of Zn2+ in water samples Fluorescence spectral measurement of water samples containing Zn2+ were carried out by adding 12 μL of 5 mmol/L stock solution of 1 and 0.30 mL of 100 mmol/L bis-tris buffer stock solution to 2.688 mL sample solutions, respectively. After well mixed, the solutions were allowed to stand at 25 ◦C for 2 min before the test.
Fluorescence test kit. Sensor 1-test kits were prepared by immersing filter papers into sensor 1 solution (50 mM), and then dried in air. Then, Zn(NO3)2 of different concentration ranges (10, 50, 70 µmol) was separately dissolved in bis-tris buffer (1 mL). The test kits prepared above were added into different concentrated solutions, and then dried at room temperature. Method for theoretical calculations of 1 and 1-Zn2+ complex All DFT/TDDFT calculations based on the hybrid exchange correlation functional B3LYP [53][54] were carried out using Gaussian 03 program [55]. The 6-31G** basis set [56,57] was used for the main group elements, whereas the Lanl2DZ effective core potential (ECP) [58–60] was employed for Zn. In vibrational frequency calculations, there was no imaginary frequency for the optimized geometries of 1 and 1-Zn2+, suggesting that these geometries represented local minima. For all calculations, the solvent effect of water was considered by using the Cossi and Barone’s CPCM (conductor-like polarizable continuum model) [61,62]. To investigate the electronic properties of singlet excited states, time-dependent DFT (TDDFT) was performed in the ground state geometries of 1 and 1-Zn2+. The 25 singlet-singlet excitations were calculated and analyzed. The GaussSum 2.1 [63] was used to calculate the contributions of molecular orbitals in electronic transitions.
Results and discussion The sensor 1 was synthesized by the substitution reaction of 2,2’-oxybis(ethan-1-1amine) and 3-chloro-N-(quinolin-8-yl)propanamide in acetonitrile (Scheme1), and analyzed by 1H NMR, 8
13
C NMR, ESI-mass, spectrometry and elemental analysis.
Fluorescence and absorption spectroscopic studies of 1 toward Zn 2+ We examined the fluorescent behavior of 1 toward various metal ions in different solvents such as MeOH, DMSO, MeCN and bis-tris buffer solution (Fig. S1). In DMSO and bis-tris buffer solution, 1 showed selectively a fluorescence enhancement at 504 nm with only Zn2+. For practical applications, the detailed fluorescent behavior of 1 toward various metal ions was studied in bis-tris buffer solution (10 mM, pH 7.0). When excited at 340 nm, 1 exhibited a weak fluorescence emission (max = 504 nm) compared to that (39 folds) in the presence of Zn2+ (Fig. 1). On the contrary, upon addition of other metal ions such as Na+, K+, Mg2+, Ca2+, Al3+, Ga3+, Cr3+, Mn2+, Fe2+, Fe3+, Co2+, Ni2+, Cu2+, Zn2+, Ga3+ and Pb2+, no or slight increase in intensity was observed. For example, 1 showed slight fluorescent enhancement to Cd2+, but 1 can clearly distinguish Zn2+ from Cd2+. These results showed that sensor 1 could be used as a fluorescence detector for Zn2+. To study the chemosensing properties of 1, fluorescence titration of the sensor 1 with Zn2+ ion was performed. As shown in Fig. 2, the emission intensity of 1 at 504 nm steadily increased until the amount of Zn2+ reached at 2.0 equiv. The quantum yields (Ф) of 1 and 1-Zn2+ complex were found to be 0.009 and 0.1, respectively. The photophysical properties of 1 were also examined using UV-vis spectrometry. UV-vis absorption spectrum of 1 showed an absorption band at 300 nm (Fig. S2). Upon the addition of Zn2+ ions to a solution of 1, the band has redshifted to 355 nm. Meanwhile, clear isosbestic points were observed at 279 nm and 333 nm, suggesting that only one product was produced from the binding of 1 with Zn2+. The Job plot [64] showed a 1:1 complexation stoichiometry between 1 and Zn2+ (Fig. 3), which was further confirmed by ESI-mass spectrometry analysis (Fig. 4). The positive-ion mass spectrum of 1 upon addition of 1 equiv of Zn2+ showed the formation of the 1-(H+)+Zn2+ [m/z: 535.14; calcd, 535.10]. From the fluorescence titration data, the association constant for 1 with Zn2+ was determined as 8.6 (±0.1) x 103 M-1 using Benesi-Hildebrand equation (Fig. S3) [65]. This value was within the range of those (1.0 ~ 1.0 x 1012) reported for Zn2+ sensing chemosensors [66–69]. The detection limit was estimated as 0.6 μM based on the 3σ/slope (Fig. S4) [70], which was about 1/130th of the WHO guideline (76 μM) for Zn2+ ions in drinking water [41,71]. 9
To check the practical applicability of 1 as Zn2+ selective sensor, fluorescence competition experiments were carried out in the presence of Zn2+ mixed with various metal ions (Fig. S5). When 1 was treated with 2.0 equiv of Zn2+ in the presence of the same concentrations of other metal ions, other background metal ions had no obvious interference with the detection of Zn2+ ion, except for Ca2+, Co2+, Cu2+ and Pb2+. The interferences with Co2+ and Cu2+ might be due to the strong quenching properties to fluorescence [72,73]. These results showed that 1 could be a good Zn2+ sensor which could discriminate Zn2+ from Cd2+ commonly having similar properties. The 1H NMR titration experiments were studied to further examine the binding mode between 1 and Zn2+ ion (Fig. S6). Upon addition of Zn2+ (1 equiv) to sensor 1, the integral of the protons H5 and H16 with upfield shift decreased to half. Moreover, all of the aromatic and aliphatic protons showed downfield shift with separation of each peak group into a pair, indicating that a Zn2+ ion might bind to only one of two quinolone moieties. These results suggested that the oxygen atom of the ether moiety and the three nitrogen atoms might coordinate to Zn2+ ion as shown in Scheme 2. This coordinative behavior of five-membered ring between 1 and a zinc ion was previously observed in the similar type of zinc complexes [42]. There was no shift in the position of proton signals on further addition of Zn2+ (>1.0 equiv), indicating that 1-Zn2+ complex might have a 1:1 binding mode. To investigate the practical applicability of 1, the effect of pH on the fluorescence response of Zn2+ was conducted at various pH (2-12) (Fig. 5). The fluorescence intensity of 1 in the presence of Zn2+ showed a significant response between pH 7 and 12. These results showed that Zn2+ could be distinctly detected by the fluorescence spectral measurement using 1 over the environmentally and physiologically relevant pH range (pH 7.0-8.4) [74], especially for monitoring Zn2+ in water samples and living cells. To examine the reversibility of sensor 1 toward Zn2+ in buffer solution, ethylenediaminetetraacetic acid (EDTA, 2.0 equiv) was added to the complexed solution of sensor 1 and Zn2+. As shown in Fig. 6, a fluorescence signal at 504 nm was immediately quenched. Upon addition of Zn2+ again, the fluorescence was recovered. The fluorescence emission changes were almost reversible even after several cycles with the sequentially alternative addition of Zn2+ and EDTA. These results indicated that sensor 1 could be recyclable simply through treatment with a proper reagent such as EDTA. Such reversibility and regeneration could be important for the fabrication of chemosensor to sense Zn2+. 10
To further demonstrate the potential of 1 to monitor Zn2+ in living matrices, fluorescence imaging experiments were carried out in living cells (Fig. 7). HeLa cell were first incubated with various concentrations of aqueous Zn2+ solutions (0, 100 and 200 μM) for 24 h and then exposed to 1 for 1 h before imaging. The experimental results showed that the HeLa cells without either Zn2+ or 1 showed negligible intracellular fluorescence, while those cultured with both Zn2+ and 1 exhibited fluorescence. In order to examine the practical properties of the chemosensor 1 in environmental samples, the chemosensor 1 was applied for the determination of Zn2+ in water samples, using a calibration curve of 1 toward Zn2+ (Fig. S7). Tap water and drinking water samples were chosen. Each sample was analyzed for three replicates. As shown in Table 1, suitable recoveries and R.S.D. values of the water samples were obtained. These results indicated that the sensor 1 can apply to determination of Zn2+ concentrations in environmental samples. For practical application, fluorescent test kits were prepared by immersing strips of filter paper in buffer solution of 1 and then dried in air. As shown in Fig. 8, when the strips were exposed to various concentrations of Zn2+ (20, 50 and 70 μM), fluorescence turned on. These results showed the potential use of sensor 1 for the detection of Zn2+ in water.
Theoretical calculations In order to get detailed insight into the sensing mechanism of 1 toward Zn2+, we conducted density functional theory (DFT) calculations with the B3LYP/6-31G (d, p) method basis set using the Gaussian 03 program. The major bond lengths and angles of 1 and 1-Zn2+ are shown in Fig. 9. The four-coordinated structure of 1-Zn2+ complex was optimized. Zn2+ was coordinated to 1N, 5N, 6N and 7O atoms of 1 and the bond lengths of Zn-1N, Zn-5N, Zn-6N and Zn-7O were similar to those of the reported Zn complexes [75,76]. Time-dependent density functional theory (TD-DFT) calculations were performed with the optimized geometries (S 0). In the case of 1, the main molecular orbital (MO) contribution of the first lowest excited state was determined for HOMO → LUMO + 1 transition (330.57 nm, Fig. S8). For the 1-Zn2+ complex, the main molecular orbital (MO) contribution of the first lowest excited state was determined for HOMO → LUMO transition (396.90, Fig. S9). The calculated HOMO → LUMO excitation of 1-Zn2+ complex indicated ICT transition from the carbonyl group to the quinolone group. Based on the molecular orbitals (MOs), the chelation of Zn2+ with 1 rendered the HOMO to LUMO energy gap of 1 decrease (4.277 eV → 3.727 eV) (Fig. S10), which is 11
consistent with the red shift in the UV-Vis spectrum of 1-Zn2+ complex. On the basis of UVvis titration, Job plot, 1H NMR titration, ESI-mass spectroscopy analysis, and theoretical calculations, we proposed the structure of a 1:1 complex of 1 and Zn2+ as shown in Scheme 2.
Conclusion We have developed a new quinoline-functionalized chemosensor 1, which detected Zn2+ ion in aqueous solution. 1 showed a highly selective fluorescence response to Zn2+ with the detection limit of 0.6 μM. Importantly, 1 can clearly distinguish Zn2+ from Cd2+, while such distinguishment of Zn2+ from Cd2+ is a well-known challenge. The binding of the sensor 1 and Zn2+ was chemically reversible with EDTA. In addition, 1 could be successfully applied to real water samples, living cells and test-kits. Moreover, the sensing mechanism of Zn2+ by 1 via ICT mechanism was explained by DFT calculations. Therefore, we believe that sensor 1 will be an excellent prototype for monitoring Zn2+ concentrations in biological and environmental environments.
Acknowledgements Financial support from Basic Science Research Program through the National Research Foundation of Korea (NRF) funded by the Ministry of Education, Science and Technology (NRF-2014R1A2A1A11051794,
NRF-2015R1A2A2A09001301,
and
NRF-
2014S1A2A2028270) are gratefully acknowledged. We thank Nano-Inorganic Laboratory, Department of Nano & Bio Chemistry, Kookmin University to access the Gaussian 03 program packages.
Appendix A. Supplementary data Supplementary data related to this article can be found at http://????.
12
References [1]
K. Komatsu, Y. Urano, H. Kojima, T. Nagano, Development of an iminocoumarin-
based zinc sensor suitable for ratiometric fluorescence imaging of neuronal zinc, J. Am. Chem. Soc. 129 (2007) 13447–13454. [2]
Y.H. Lee, M.H. Lee, J.F. Zhang, J.S. Kim, Pyrene excimer-based calix[4]arene FRET
chemosensor for mercury(II), J. Org. Chem. 75 (2010) 7159–7165. [3]
R.K. Pathak, V.K. Hinge, A. Rai, D. Panda, C.P. Rao, Imino-phenolic-pyridyl
conjugates of calix[4]arene (L1 and L2) as primary fluorescence switch-on sensors for Zn2+ in solution and in HeLa cells and the recognition of pyrophosphate and ATP by [ZnL2], Inorg. Chem. 51 (2012) 4994–5005. [4]
A. Ajayaghosh, P. Carol, S. Sreejith, A ratiometric fluorescence probe for selective
visual sensing of Zn2+, J. Am. Chem. Soc. 127 (2005) 14962–14963. [5]
Y. Wu, X. Peng, B. Guo, J. Fan, Z. Zhang, J. Wang, A. Cui and Y. Gao, Boron
dipyrromethene fluorophore based fluorescence sensor for the selective imaging of Zn(II) in living cells, Org. Biomol. Chem. 3 (2005) 1387–1392. [6]
P. Jiang, L. Chen, J. Lin, Q. Liu, J. Ding, X. Gao and Z. Guo, Novel zinc fluorescent
probe bearing dansyl and aminoquinoline groupsElectronic supplementary information (ESI) available: NMR spectra and assignment, UV titration details, crystal structure and competitive fluorescent experiments of L, Chem. Commun. (2002) 1424–1425. [7]
A. Loas, R.J. Radford, S.J. Lippard, Addition of a second binding site increases the
dynamic range but alters the cellular localization of a red fluorescent probe for mobile zinc, Inorg. Chem. 53 (2014) 6491–6493. [8]
K. Tayade, B. Bondhopadhyay, H. Sharma, A. Basu, V. Gite, S. Attarde, N. Singh and
A. Kuwar, “Turn-on” fluorescent chemosensor for zinc(II) dipodal ratiometric receptor: application in live cell imaging, Photochem. Photobiol. Sci. 13 (2014) 1052–1057. [9]
J.H. Weiss, S.L. Sensi, J.Y. Koh, Zn2+: a novel ionic mediator of neural injury in brain
disease, Trends Pharmacol. Sci. 21 (2000) 395–401. [10]
B.K.Y. Bitanihirwe, M.G. Cunningham, Zinc: the brain’s dark horse, Synapse. 63
(2009) 1029–1049. [11]
X.M. Xie, T.G. Smart, A physiological role for endogenous zinc in rat hippocampal
synaptic neurotransmission, Nature. 349 (1991) 521–524. [12]
C.E. Outten, T. V O’Halloran, Femtomolar sensitivity of metalloregulatory proteins 13
controlling zinc homeostasis, Science. 292 (2001) 2488–2492. [13]
Z. Xu, G.-H. Kim, S.J. Han, M.J. Jou, C. Lee, I. Shin and J.Y Yoon, An NBD-based
colorimetric and fluorescent chemosensor for Zn2+ and its use for detection of intracellular zinc ions, Tetrahedron. 65 (2009) 2307–2312. [14]
K. Li, A. Tong, A new fluorescent chemosensor for Zn2+ with facile synthesis: “Turn-
on” response in water at neutral pH and its application for live cell imaging, Sens. Actuators B. 184 (2013) 248–253. [15]
H.J. Kim, S.Y. Park, S. Yoon, J.S. Kim, FRET-derived ratiometric fluorescence sensor
for Cu2+, Tetrahedron. 64 (2008) 1294–1300. [16]
M. Shellaiah, Y.-H. Wu, H.-C. Lin, Simple pyridyl-salicylimine-based fluorescence
“turn-on” sensors for distinct detections of Zn2+, Al3+ and OH- ions in mixed aqueous media., Analyst. 138 (2013) 2931–2942. [17]
G. J. Park, H. Kim, J. J. Lee, Y. S. Kim, S. Y. Lee, S.Y. Lee, I.S Noh and C. Kim, A
highly selective turn-on chemosensor capable of monitoring Zn2+ concentrations in living cells and aqueous solution, Sens. Actuators B. 215 (2015) 568–576. [18]
C. Gao, X. Jin, X. Yan, P. An, Y. Zhang, L. Liu, H. Tian, W. Liu, X. Yao and Y. Tang,
A small molecular fluorescent sensor for highly selectivity of zinc ion, Sens. Actuators B, 176 (2013) 775–781. [19]
Y.J. Na, I.H. Hwang, H.Y. Jo, S.A. Lee, G.J. Park, C. Kim, Fluorescent chemosensor
based-on the combination of julolidine and furan for selective detection of zinc ion, Inorg. Chem. Commun. 35 (2013) 342–345. [20]
S. Cui, G. Liu, S. Pu, B. Chen, A highly selective fluorescent probe for Zn2+ based on
a new photochromic diarylethene with a di-2-picolylamine unit, Dye. Pigm. 99 (2013) 950– 956. [21]
Y.-P. Li, Q. Zhao, H.-R. Yang, S.-J. Liu, X.-M. Liu, Y.-H. Zhang, T.-L. Hu, J.-T. Chen,
Z. Chang and Z.-H. Bu, A new ditopic ratiometric receptor for detecting zinc and fluoride ions in living cells, Analyst. 138 (2013) 5486–5494. [22]
M. Yan, T. Li, Z. Yang, A novel coumarin Schiff-base as a Zn(II) ion fluorescent sensor,
Inorg. Chem. Commun. 14 (2011) 463–465. [23]
H. Kim, J. Kang, K.B. Kim, E.J. Song, C. Kim, A highly selective quinoline-based
fluorescent sensor for Zn(II), Spectrochim. Acta. A. Mol. Biomol. Spectrosc. 118 (2014) 883– 887. 14
[24]
V. Bhalla, H. Arora, A. Dhir, M. Kumar, A triphenylene based zinc ensemble as an
oxidation inhibitor., Chem. Commun. 48 (2012) 4722–4724. [25]
S.Y. Park, J.H. Yoon, C.S. Hong, R. Souane, J.S. Kim, S.E. Matthews and Jacques
Vicens, A pyrenyl-appended triazole-based calix[4]arene as a fluorescent sensor for Cd2+ and Zn2+, J. Org. Chem. 73 (2008) 8212–8218. [26]
Z. Liu, C. Zhang, Y. Chen, W. He, Z. Guo, An excitation ratiometric Zn2+ sensor with
mitochondria-targetability for monitoring of mitochondrial Zn2+ release upon different stimulations, Chem. Commun. 48 (2012) 8365–8367 [27]
K. Tayade, S.K. Sahoo, B. Bondhopadhyay, V.K. Bhardwaj, N. Singh, A. Basu and R.
Bendre, Highly selective turn-on fluorescent sensor for nanomolar detection of biologically important Zn2+ based on isonicotinohydrazide derivative: application in cellular imaging, Biosens. Bioelectron. 61 (2014) 429–433. [28]
V.K. Gupta, N. Mergu, A.K. Singh, Fluorescent chemosensors for Zn2+ ions based on
flavonol derivatives, Sens. Actuators B. 202 (2014) 674–682. [29]
A. Rajabi Khorrami, A.R. Fakhari, M. Shamsipur, H. Naeimi, Pre-concentration of
ultra trace amounts of copper, zinc, cobalt and nickel in environmental water samples using modified C18 extraction disks and determination by inductively coupled plasma–optical emission spectrometry, Int. J. Environ. Anal. Chem. 89 (2009) 319–329. [30]
M. Ghaedi, F. Ahmadi, A. Shokrollahi, Simultaneous preconcentration and
determination of copper, nickel, cobalt and lead ions content by flame atomic absorption spectrometry, J. Hazard. Mater. 142 (2007) 272–278. [31]
K. Sreenivasa Rao, T. Balaji, T. Prasada Rao, Y. Babu, G.R.K. Naidu, Determination
of iron, cobalt, nickel, manganese, zinc, copper, cadmium and lead in human hair by inductively coupled plasma-atomic emission spectrometry, Spectrochim. Acta Part B. 57 (2002) 1333–1338. [32]
R. gulaboski, V. mireski, F. scholz, An electrochemical method for determination of
the standard Gibbs energy of anion transfer between water and n-octanol, Electrochem. Commun. 4 (2002) 277–283. [33]
Z. Li, M. Yu, L. Zhang, M. Yu, J. Liu, L. Wei, and H. Zhang, A “switching on”
fluorescent chemodosimeter of selectivity to Zn2+ and its application to MCF-7 cells, Chem. Commun. 46 (2010) 7169–7171. [34]
U.C. Saha, B. Chattopadhyay, K. Dhara, S.K. Mandal, S. Sarkar, A.R. Khuda-Bukhsh, 15
M. Mukherjee, M. Helliwell and P. Chattopadhyay, A Highly Selective Fluorescent Chemosensor for Zinc Ion and Imaging Application in Living Cells, Inorg. Chem. 50 (2011) 1213-1219. [35]
G.J. Park, Y.J. Na, H.Y. Jo, S.A. Lee, A.R. Kim, I. Noh and C. Kim, A single
chemosensor for multiple analytes: fluorogenic detection of Zn2+ and OAc− ions in aqueous solution, and an application to bioimaging, New J. Chem. 38 (2014) 2587-2594. [36]
V. Bhalla, Roopa, M. Kumar, Pentaquinone based probe for nanomolar detection of
zinc ions: chemosensing ensemble as an antioxidant., Dalton Trans. 42 (2013) 975–80. [37]
Y. Ma, F. Wang, S. Kambam, X. Chen, A quinoline-based fluorescent chemosensor for
distinguishing cadmium from zinc ions using cysteine as an auxiliary reagent, Sens. Actuators B. 188 (2013) 1116–1122. [38]
K.M.K. Swamy, M.-J. Kim, H.-R. Jeon, J.-Y. Jung, J.-Y. Yoon, New 7-
Hydroxycoumarin-Based Fluorescent Chemosensors for Zn(II) and Cd(II), Bull. Korean Chem. Soc. 31 (2010) 3611–3616. [39]
Z. Xu, J.-Y Yoon, D.R. Spring, Fluorescent chemosensors for Zn2+, Chem. Soc. Rev.
39 (2010) 1996–2006. [40]
E.J. Song, J. Kang, G.R. You, G.J. Park, Y. Kim, S.-J. Kim, C. Kim and Roger G.
Harrison, A single molecule that acts as a fluorescence sensor for zinc and cadmium and a colorimetric sensor for cobalt, Dalton Trans. 42 (2013) 15514–20. [41]
H. Gyu Lee, J. Hoon Lee, S. Pyo Jang, H. Min Park, S. Jin Kim, Y. Mee Kim, Cheal
Kim, Zinc selective chemosensor based on pyridyl-amide fluorescence, Tetrahedron. 67 (2011) 8073–8078. [42]
H.G. Lee, J.H. Lee, S.P. Jang, I.H. Hwang, S.-J. Kim, Y. Kim, C. Kim, Zinc selective
chemosensors based on the flexible dipicolylamine and quinoline, Inorganica Chim. Acta. 394 (2013) 542–551. [43]
E. J. Song, G. J. Park, J. J. Lee, S.Y. Lee, I. Noh, Y. C. Kim and Roger G. Harrison, A
fluorescence sensor for Zn2+ that also acts as a visible sensor for Co2+ and Cu2+, Sens. Actuators B. 213 (2015) 268–275. [44]
H. Kim, G.R. You, G.J. Park, J.Y. Choi, I. Noh, Y. Kim, S.J. Kim, C. Kim and Rogger
G. Harrion, Selective zinc sensor based on pyrazoles and quinoline used to image cells, Dye. Pigm. 113 (2015) 723–729. [45]
S. Sinha, T. Mukherjee, J. Mathew, S.K. Mukhopadhyay, S. Ghosh, Triazole-based 16
Zn2+-specific molecular marker for fluorescence bioimaging, Anal. Chim. Acta. 822 (2014) 60–68. [46]
K. Hanaoka, Y. Muramatsu, Y. Urano, T. Terai, T. Nagano, Design and synthesis of a
highly sensitive off-on fluorescent chemosensor for zinc ions utilizing internal charge transfer., Chemistry. 16 (2010) 568–572. [47]
D. Wang, X. Xiang, X. Yang, X. Wang, Y. Guo, W. Liu and W. Qin, Fluorescein-based
chromo-fluorescent probe for zinc in aqueous solution: Spirolactam ring opened or closed?, Sens. Actuators B. 201 (2014) 246–254. [48]
L. Zang, H. Shang, D. Wei, S. Jiang, A multi-stimuli-responsive organogel based on
salicylidene Schiff base, Sens. Actuators B. 185 (2013) 389–397. [49]
A. Ojida, Y. Mito-oka, K. Sada, I. Hamachi, Molecular recognition and fluorescence
sensing of monophosphorylated peptides in aqueous solution by bis(zinc(II)-dipicolylamine)based artificial receptors, J. Am. Chem. Soc. 126 (2004) 2454–2463. [50]
C. Lakshmi, R.G. Hanshaw, B.D. Smith, Fluorophore-linked zinc(II)dipicolylamine
coordination complexes as sensors for phosphatidylserine-containing membranes, Tetrahedron. 60 (2004) 11307–11315. [51]
B.A. Wong, S. Friedle, S.J. Lippard, Subtle modification of 2,2-dipicolylamine lowers
the affinity and improves the turn-on of Zn(II)-selective fluorescent sensors, Inorg. Chem. 48 (2009) 7009–7011. [52]
H. Kim, Y. Liu, D. Nam, Y. Li, S. Park, J.-Y Yoon, M. H. Hyun, A new phosphorescent
chemosensor bearing Zn-DPA sites for H2PO4−, Dye. Pigm. 106 (2014) 20–24. [53]
A.D. Becke, Density-functional thermochemistry. III. The role of exact exchange, J.
Chem. Phys. 98 (1993) 5648–5652. [54]
C. Lee, W. Yang, R.G. Parr, Development of the Colle-Salvetti correlation-energy
formula into a functional of the electron density, Phys. Rev. B. 37 (1988) 785–789. [55]
C.G. and J.A.P. M. J. Frisch, G. W. Trucks, H. B. Schlegel, G. E. Scuseria, M. A. Robb,
J. R. Cheeseman, J. A. Montgomery Jr., T. Vreven, K. N. Kudin, J. C. Burant, J.M.Millam, S. S. Iyengar, J. Tomasi, V. Barone, B. Mennucci, M. Cossi, G. Scalmani, N. Rega, G. A. Peters, GAUSSIAN 03 (Revision B.02), Gaussian, Inc., Wallingford CT. (2004). [56]
P.C. Hariharan, J.A. Pople, The influence of polarization functions on molecular
orbital hydrogenation energies, Theor. Chim. Acta. 28 (1973) 213–222. [57]
M.M. Francl, W.J. Pietro, W.J. Hehre, J.S. Binkley, M.S. Gordon, D.J. DeFrees and 17
J.A. Pople, Self-Consistent Molecular Orbital Methods. 23. A polarization-type basis set for 2nd-row elements, J. Chem. Phys. 77 (1982) 3654–3665. [58]
P.J. Hay, W.R. Wadt, Ab initio effective core potentials for molecular calculations.
Potentials for the transition metal atoms Sc to Hg, J. Chem. Phys. 82 (1985) 270–283. [59]
W.R. Wadt, P.J. Hay, Ab initio effective core potentials for molecular calculations.
Potentials for main group elements Na to Bi, J. Chem. Phys. 82 (1985) 284–298. [60]
W.R. Wadt, P.J. Hay, Ab initio effective core potentials for molecular calculations.
Potentials for K to Au including the outermost core orbitals, J. Chem. Phys. 82 (1985) 299– 310. [61]
V. Barone, M. Cossi, Quantum calculation of molecular energies and energy gradients
in solution by a conductor solvent model, J. Phys. Chem. A. 102 (1998) 1995–2001. [62]
M. Cossi, V. Barone, Time-dependent density functional theory for molecules in liquid
solutions, J. Chem. Phys. 115 (2001) 4708–4717. [63]
N.M. O’Boyle, A.L. Tenderholt, K.M. Langner, cclib: a library for package-
independent computational chemistry algorithms, J. Comput. Chem. 29 (2008) 839–845. [64]
P. Job, Formation and stability of inorganic complexes in solution, Ann. Chim. 9 (1928)
113–203. [65]
H.A. Benesi, J.H. Hildebrand, A Spectrophotometric Investigation of the Interaction
of Iodine with Aromatic Hydrocarbons, J. Am. Chem. Soc. 71 (1949) 2703–2707. [66]
H.-Y. Lin, P.-Y. Cheng, C.-F. Wan, A.-T. Wu, A turn-on and reversible fluorescence
sensor for zinc ion, Analyst. 137 (2012) 4415–4417. [67]
J.H. Kim, I.H. Hwang, S.P. Jang, J. Kang, S. Kim, I. Noh, Y. Kim, C. Kim and Roger
G. Harrison, Zinc sensors with lower binding affinities for cellular imaging, Dalton Trans. 42 (2013) 5500–5507. [68]
W.H. Hsieh, C.-F. Wan, D.-J. Liao, A.-T. Wu, A turn-on Schiff base fluorescence
sensor for zinc ion, Tetrahedron Lett. 53 (2012) 5848–5851. [69]
Y. Zhou, Z.-X. Li, S.-Q. Zang, Y.-Y. Zhu, H.-Y. Zhang, H.-W. Hou and T. C. W. Mak,
A novel sensitive turn-on fluorescent Zn2+ chemosensor based on an easy to prepare C3symmetric Schiff-base derivative in 100% aqueous solution, Org. Lett. 14 (2012) 1214–1217. [70] Y.-K. Tsui, S. Devaraj, Y.-P. Yen, Azo dyes featuring with nitrobenzoxadiazole (NBD) unit: A new selective chromogenic and fluorogenic sensor for cyanide ion, Sens. Actuators B. 161 (2012) 510–519. 18
[71]
Y.P. Kumar, P. King, V.S.R.K. Prasad, Zinc biosorption on Tectona grandis L.f. leaves
biomass: Equilibrium and kinetic studies, Chem. Eng. J. 124 (2006) 63–70. [72]
Z. Liu, Z. Yang, T. Li, B. Wang, Y. Li, D. Qin, M. Wang and M. Yan, An effective
Cu(II) quenching fluorescence sensor in aqueous solution and 1D chain coordination polymer framework, Dalton Trans. 40 (2011) 9370–9373. [73]
R. Homan, M. Eisenberg, A fluorescence quenching technique for the measurement of
paramagnetic ion concentrations at the membrane/water interface. Intrinsic and X537Amediated cobalt fluxes across lipid bilayer membranes, Biochim. Biophys. Acta - Biomembr. 812 (1985) 485–492. [74]
R.M. Harrison, D.P.H. Laxen, S.J. Wilson, Chemical associations of lead, cadmium,
copper, and zinc in street dusts and roadside soils, Environ. Sci. Technol. 15 (1981) 1378–1383. [75]
C. Bergquist, G. Parkin, Modeling the Catalytic Cycle of Liver Alcohol
Dehydrogenase:
Synthesis and Structural Characterization of a Four-Coordinate Zinc
Ethoxide Complex and Determination of Relative Zn−OR versus Zn−OH Bond Energies, Inorg. Chem. 38 (1999) 422–423. [76]
J.J. Lee, S.A. Lee, H. Kim, L. Nguyen, I. Noh, C. Kim, A highly selective CHEF-type
chemosensor for monitoring Zn2+ in aqueous solution and living cells, RSC Adv. 5 (2015) 41905–41913.
19
Biographies
Jae Min Jung earned the BS degree in 2016 at Seoul National University of Science and Technology. He is currently a Master Student at Seoul National University of Science and Technology. His scientific interest includes chemical sensors, DNA cleavage by the transition metal complexes and synthesis of catalyst.
Seong Youl Lee received the BS degree in 2015 at Seoul National University of Science and Technology. He is currently a master student at Seoul National University of Science and Technology. His research interest includes chemical sensors, synthesis of catalyst and inorganic medicine.
Eunju Nam received her BS degree in 2015 at Ulsan National Institute of Science and Technology (UNIST). She is currently a master student at UNIST. Her research interests lie in understanding how small molecules can control pathological factors and pathways in human diseases. Mi Hee Lim is currently an associate professor at UNIST. She received her PhD degree at Massachusetts Institute of Technology (MIT) in USA (2016) and conducted her postdoctoral research at California Institute of Technology (Caltech) (2016-2018). Since 2008, Dr. Lim and her group have been identifying the roles of metals, proteins, and reactive oxygen species in human neurodegenerative disorders.
Cheal Kim is currently a professor at Seoul National University of Science and Technology. He received the PhD degree in 1993 at University of California, San Diego in USA. He is now interested in development of chemical sensors, reactivity study of the transition metal complexes, DNA cleavage by their metal complexes and MOF.
20
Figure Captions Figure 1. Fluorescence spectral changes of 1 (20 µM) in the presence of different metal ions (2.0 equiv) such as Na+, K+, Mg2+, Ca2+, Al3+,Ga3+, Cr3+, Mn2+, Fe2+, Fe3+, Co2+, Ni2+, Cu2+, Zn2+, Ga3+ and Pb2+ with an excitation of 340 nm in buffer solution (10 mM bis-tris, pH 7.0). Figure 2. Fluorescence spectral changes of 1 (20 μM) in the presence of different concentrations of Zn2+ ions in buffer solution (10 mM bis-tris, pH 7.0). Inset: Fluorescence intensity at 504 nm versus the number of equiv of Zn2+ added.
Figure 3. Job plot for the binding of 1 with Zn2+. Intensity at 504 nm was plotted as a function of the molar ratio [Zn2+]/([1] + [Zn2+]). The total concentration of zinc ions with sensor 1 was 4.0 x 10-5 M. Figure 4. Positive-ion electrospray ionization mass spectrum of 1 (100 μM) upon addition of Zn(NO3)2 (1 equiv). Figure 5. Fluorescence intensity (at 504 nm) of 1 in the presence of Zn2+ at different pH values (2-12) in buffer solution (10 mM bis-tris, pH 7.0).
Figure 6. Reversible changes (at 504 nm) of 1 (20 μM) upon the sequential addition of Zn2+ and EDTA.
Figure 7. Fluorescent responses of 1 to Zn(II) in HeLa cells. Cells were preincubated with 1 for 30 min prior to addition of various concentrations of Zn(II). Conditions: [Zn(II)] = 0, 100, and 200 μM; [1] = 20 μM; 37 °C; 5% CO2. The scale bar is 50 μm. Figure 8. Detection of Zn2+ by test strips coated with 1. Dark blue color indicated fluorescenceoff and light blue color did fluorescence-on. Conditions: [Zn(II)] = 20, 50 and 70 μM; [1] = 50 mM.
21
Figure 9. The energy-minimized structures of (a) 1 and (b) 1-Zn2+ complex.
22
Fig. 1.
23
Fig. 2.
24
Fig. 3.
25
Fig. 4.
26
Fig. 5.
27
Fig. 6.
28
Fig. 7.
29
Fig. 8.
30
(a)
(b)
Fig. 9.
31
Scheme 1. Synthesis of 1.
32
Scheme 2. Fluorescence enhancement mechanism and proposed structure of 1-Zn2+ complex.
33
Table 1. Determination of Zn(II) in water samples Sample
Zn(II) added (μmol/L)
Zn(II) found (μmol/L)
Tap water
0.00
0.00
4.00a
3.8
0.00
0.00
4.00b
4.2
Drinking water
Recovery (%)
R.S.D. (n = 3) (%) 2.17
95
0.2 0.33
105
0.83
Conditions: [1] = 20 μmol/L in 10 mM bis-tris buffer-DMSO solution (999:1, pH 7.0). a. 4.00 μmol/L of Zn2+ ion was artificially added into tap water. b. 4.00 μmol/L of Zn 2+ ion was artificially added into drinking water.
34