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Oct 24, 2016 - ABSTRACT: The tetrakisquinoline ligand HT(6-MeO8Q)-. HPN (N,N,N′,N′-tetrakis(6-methoxy-8-quinolylmethyl)-2-hy- droxy-1 ...
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Fluorescent Detection of Phosphate Ion via a Tetranuclear Zinc Complex Supported by a Tetrakisquinoline Ligand and μ4‑PO4 Core Yuji Mikata,*,†,‡,§ Risa Ohnishi,§ Risa Nishijima,§ and Hideo Konno∥ †

Department of Chemistry, Biology, and Environmental Science, Faculty of Science, ‡KYOUSEI Science Center, and §Department of Chemistry, Faculty of Science, Nara Women’s University, Nara 630-8506, Japan ∥ National Institute of Advanced Industrial Science and Technology (AIST), 1-1-1 Higashi, Tsukuba, Ibaraki 305-8565, Japan S Supporting Information *

ABSTRACT: The tetrakisquinoline ligand HT(6-MeO8Q)HPN (N,N,N′,N′-tetrakis(6-methoxy-8-quinolylmethyl)-2-hydroxy-1,3-propanediamine) exhibited Zn2+-induced fluorescence enhancement with high specificity and sensitivity (IZn/ I0 = 57 and ICd/IZn = 6% in the presence of 2 equiv of Zn2+; LOD (limit of detection) = 15 nM). This ligand also exhibited fluorescence enhancement specific to inorganic phosphate (PO43−) in DMF−HEPES buffer (50 mM HEPES, 100 mM KCl, pH = 7.5) (1:1) in the presence of 2 equiv of Zn2+. The structure of the unprecedented tetranuclear zinc complex with a μ4-PO4 bridge was elucidated by X-ray crystallography as the key species responsible for fluorescence enhancement.

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Previously, we reported the pyrophosphate (PPi, P2O74−)specific fluorescence enhancement of the dinuclear zinc complex of HTQHPN (N,N,N′,N′-tetrakis(2-quinolylmethyl)2-hydroxy-1,3-propanediamine, Chart 1).17 The structure of

INTRODUCTION Detection and quantification of specific anions is one of the most attractive subjects in analytical and environmental science.1−4 Particularly, inorganic phosphate (PO43−) as a target anion is important because of its abundance in living organisms and the environment.5 Phosphate pollution partly resulting from detergents and fertilizers from human activity is one of the current environmental problems discussed by the United States Environmental Protection Agency (EPA). In this context, methodologies for detection/quantification of PO43− have been studied extensively for a decade. Since the fluorescent detection strategy is the most popular, sensitive, and convenient approach, many fluorescent phosphate receptors utilizing hydrogen bonding or electrostatic interactions with metal complexes have been reported.5−12 The spatial recognition of the tetrahedral shape of phosphate would be an important objective in the development of metalcontaining fluorescent phosphate sensors. It affords high recognition specificity over other anions of different shapes. The stable complex formation with tetrahedral PO43− also allows comprehensive detection of all phosphate species including H2PO4−, HPO42−, and PO43− in aqueous media via deprotonation. Structurally determined phosphate-binding modes with metallohost molecules include monodentate, chelating, and bridging coordination modes. Crystal structures of multinuclear metal complexes with μ3- and μ4-PO4 cores are extremely rare. There have been only four examples for structurally elucidated μ4-PO4 complexes utilizing tetranuclear Fe3+,13 Cu2+,14,15 and Cr3+16 metal centers; however, none of these complexes have been applied to phosphate recognition machinery in optical signaling molecules. © XXXX American Chemical Society

Chart 1

Received: August 13, 2016

A

DOI: 10.1021/acs.inorgchem.6b01967 Inorg. Chem. XXXX, XXX, XXX−XXX

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(CD3CN, 75.5 MHz): δ (ppm) 158.0, 151.2, 149.8, 140.7, 140.5, 131.6, 127.4, 127.1, 123.3, 122.7, 107.8, 107.3, 60.3, 58.8, 58.5, 56.5, 56.3. Anal. Calcd for C49H49Cl2N6O15.5Zn2 ([Zn2(T(6-MeO8Q)HPN)(AcO)](ClO4)2·0.5H2O): H, 4.22; C, 50.23; N, 7.17. Found: H, 4.30; C, 50.22; N, 6.88. ESI-MS (positive mode, m/z): calcd mass for [Zn 2 (T(6-MeO8Q)HPN)(AcO)](ClO 4 ) + (C 49 H 48 ClN 6 O 11 Zn 2 ) 1059.16525, found 1059.20611; calcd mass for [Zn2(T(6-MeO8Q)HPN)(AcO)]2+ (C49H48N6O7Zn2) 480.10837, found 480.12891. [Zn4(T(6-MeO8Q)HPN)2(PO4)](ClO4)3·2THF·CH3OH·2.5H2O. To the THF solution (1.5 mL) of HT(6-MeO8Q)HPN (7.4 mg, 10 μmol) were added Zn(ClO4)2·6H2O (7.4 mg, 20 μmol) in methanol (1.0 mL) and Na3PO4·12H2O (1.9 mg, 5.0 μmol) in water (0.5 mL), and the solution was kept at 4 °C to give [Zn4(T(6-MeO8Q)HPN)2(PO4)](ClO4)3·2THF·CH3OH·2.5H2O as colorless crystals. Yield: 2.0 mg (18%). 1 H NMR (DMF-d7, 300 MHz): δ (ppm) 10.97 (dd, J1 = 5.0 Hz, J2 = 1.7 Hz, 2H), 10.47 (dd, J1 = 4.9 Hz, J2 = 1.8 Hz, 2H), 9.75−9.72 (m, 4H), 8.94 (dd, J1 = 8.5 Hz, J2 = 1.7 Hz, 2H), 8.67−8.62 (m, 4H), 8.45 (dd, J1 = 8.5 Hz, J2 = 1.8 Hz, 2H), 8.27−8.23 (m, 4H), 7.81−7.73 (m, 8H), 7.44 (d, J = 2.7 Hz, 2H), 7.35 (d, J = 2.4 Hz, 2H), 7.29 (d, J = 2.7 Hz, 2H), 7.23 (d, J = 2.4 Hz, 2H), 7.13 (d, J = 2.4 Hz, 2H), 7.10 (d, J = 2.7 Hz, 2H), 5.82 (d, J = 13.1 Hz, 2H), 4.86 (br., 2H), 4.56 (d, J = 13.7 Hz, 2H), 4.42 (d, J = 13.4 Hz, 2H), 4.27 (d, J = 13.1 Hz, 2H), 4.03 (s, 6H), 4.02 (d, J = 13.1 Hz, 2H), 3.93 (s, 6H), 3.86 (s, 6H), 3.78 (d, J = 14.6 Hz, 2H), 3.75 (s, 6H), 3.65 (d, J = 13.4 Hz, 2H), 3.55 (d, J = 13.4 Hz, 2H), 2.8−2.7 (br., 2H), 2.53 (d, J = 8.9 Hz, 2H), 2.29 (t, J = 11.6 Hz, 2H), 2.12 (t, J = 11.6 Hz, 2H). 13C NMR (DMF-d7, 75.5 MHz): δ (ppm) 163.3, 162.9, 162.7, 162.6, 158.1, 157.2, 156.1, 147.0, 146.3, 145.8, 145.4, 137.9, 137.5, 137.3, 137.3, 137.1, 137.0, 136.6, 136.2, 132.4, 132.0, 131.5, 128.9, 128.4, 127.7, 113.0, 112.7, 112.5, 66.7, 66.1, 65.8, 65.1, 64.0, 61.7, 61.5, 61.4. 31P NMR (DMF-d7, 121.5 MHz): δ (ppm) 4.98. Anal. Calcd for C94H94Cl3N12O28Zn4 ([Zn4(T(6-MeO8Q)HPN)2(PO4)](ClO4)3·2H2O): H, 4.23; C, 50.43; N, 7.51. Found: H, 4.13; C, 50.24; N, 7.40. ESI-MS (positive mode, m/z): calcd mass for [Zn 4 (T(6-MeO8Q)HPN) 2 (PO 4 )](ClO 4 ) 2 + (C94H90ClN12O18PZn4) 998.15440, found 998.24675; calcd mass for [Zn4(T(6-MeO8Q)HPN)2(PO4)]3+ (C94H90N12O14PZn4) 632.45343, found 632.55184. Spectroscopic Measurements. Absorbance and fluorescence spectra were measured in DMF−HEPES buffer (50 mM HEPES, 100 mM KCl, pH = 7.5) (1:1) at 25 °C unless otherwise indicated. For spectral titration, 1−4 μL aliquots of metal ion or anion solution in water were added to the ligand or zinc complex solution (3000 μL), maintaining the volume change within 1%. A Job’s plot (Figure 6) was performed by mixing of 51 μM solutions of Zn2+ complex and PO43− in different ratios, maintaining the total volume constant. ESI-MS was measured in diluted methanol solution. X-ray Crystallography. Single crystals of [Zn2(T(6-MeO8Q)HPN)(OAc)](ClO4)2·4CH3CN and [Zn4(T(6-MeO8Q)HPN)2(PO4)](ClO4)3·2THF·CH3OH·2.5H2O were covered by Paratone-N oil and mounted on a glass fiber. All data were collected at 153 K on a Rigaku Mercury CCD detector, with monochromatic Mo Kα radiation, operating at 50 kV/40 mA. Data were processed on a PC using CrystalClear Software (Rigaku). Structures were solved by direct methods (SIR-92) and refined by full-matrix least-squares methods on F2 (SHELXL-97). For [Zn2(T(6-MeO8Q)HPN)(OAc)](ClO4)2· 4CH3CN, the value for the Flack parameter is ambiguous (0.491 ± 0.013) probably because the partial orientation of solvent molecules (CH3CN) is an acentric source. For [Zn4(T(6-MeO8Q)HPN)2(PO4)](ClO4)3·2THF·CH3OH·2.5H2O, the removal of solvents by PLATON/SQUEEZE could improve the refinement (R = 0.0696 and Rw = 0.1898), but all solvents were assigned in the present report (electron count = 978 from PLATON/SQUEEZE; 984 for 2THF· CH3OH·2.5H2O). Crystal data and metric parameters are summarized in Tables S1−S3. CCDC-1479521 and 1479522 contain the supplementary crystallographic data for this paper. These data can be obtained free of charge from the Cambridge Crystallographic Data Centre via www.ccdc.cam.ac.uk/datarequest/cif.

HTQHPN18 is based on the fluorescent zinc sensor TQEN (N,N,N′,N′-tetrakis(2-quinolylmethyl)ethylenediamine, Chart 1).19 A crystallographic study using a PPi analogue reveals that the Zn2-TQHPN-PPi ternary complex exhibits characteristic intramolecular excimer formation between adjacent quinoline rings. Recently, an isoquinoline analogue of HTQHPN, 1-isoHTQHPN (N,N,N′,N′-tetrakis(1-isoquinolylmethyl)-2-hydroxy-1,3-propanediamine, Chart 1), was utilized as an OFF−ON−OFF fluorescent zinc sensor that specifically detects the mononuclear complex.20 In this Article, we describe the PO43−-specific fluorescence enhancement of the dinuclear zinc complex of HT(6-MeO8Q)HPN (N,N,N′,N′-tetrakis(6methoxy-8-quinolylmethyl)-2-hydroxy-1,3-propanediamine, Chart 1). The structure of the ternary complex with an unprecedented (μ4-PO4)Zn4 core was revealed by X-ray crystallography.

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EXPERIMENTAL SECTION

General Procedures. All reagents and solvents used for synthesis were from commercial sources and used as received. N,NDimethylformamide (DMF, Dojin) was spectral grade (Spectrosol). All aqueous solutions were prepared using Milli-Q water (Millipore). 1 H NMR (300 MHz), 13C NMR (75.5 MHz), and 31P NMR (121.5 MHz) spectra were recorded on a Varian GEMINI 2000 spectrometer and referenced to internal Si(CH3)4, solvent or external H2PO4 (20% in DMF-d7) signals. UV−vis and fluorescence spectra were measured on a Jasco V-660 spectrophotometer and Jasco FP-6300 spectrofluorometer, respectively. ESI-MS spectra were recorded on a JEOL JMS-T100LC mass spectrometer. Fluorescence quantum yields were measured on a HAMAMATSU Photonics C9920-02 absolute PL quantum yield measurement system. CAUTION: Perchlorate salts of metal complexes with organic ligands are potentially explosive. All due precautions should be taken. N,N,N′,N′-Tetrakis(6-methoxy-8-quinolylmethyl)-2-hydroxy1,3-propanediamine (HT(6-MeO8Q)HPN). To an acetonitrile solution (40 mL) of 6-methoxy-8-chloromethylquinoline (360 mg, 1.73 mmol) and 1,3-diamino-2-propanol (39 mg, 0.43 mmol) were added potassium carbonate (1.20 g, 8.67 mmol) and potassium iodide (288 mg, 1.73 mmol). The reaction mixture was refluxed for 2 days. The resultant solution was cooled to room temperature and filtered; then the solvent was evaporated. The residue was extracted with CH2Cl2−water, and the organic layer was dried and evaporated. The residue was recrystallized from methanol to give HT(6-MeO8Q)HPN as a white powder (260 mg, 0.33 mmol, 76%). 1 H NMR (CDCl3, Me4Si, 300 MHz): δ (ppm) 8.70 (dd, J1 = 4.3 Hz, J2 = 1.5 Hz, 4H), 7.90 (dd, J1 = 8.2 Hz, J2 = 1.8 Hz, 4H), 7.54 (d, J = 2.7 Hz, 4H), 7.27 (dd, J1 = 8.2 Hz, J2 = 4.3 Hz, 4H), 6.73 (d, J = 2.7 Hz, 4H), 4.53 (quint, J = 6.4 Hz, 1H), 4.45 (d, J = 15.3 Hz, 4H), 4.31 (d, J = 15.3 Hz, 4H), 3.68 (s, 12H), 2.85 (d, J = 6.4 Hz, 4H). 13C NMR (CDCl3, Me4Si, 75.5 MHz): δ (ppm) 157.0, 146.1, 142.6, 139.4, 134.7, 129.2, 121.1, 120.8, 104.0, 68.3, 59.8, 55.2, 54.9. Anal. Calcd for C47H46.4N6O5.2 (HT(6-MeO8Q)HPN·0.2H2O): C, 72.51; H, 6.01; N, 10.80. Found: C, 72.49; H, 5.74; N, 10.77. HRMS (ESI-MS): m/z calcd for C47H47N6O5 ([M + H]+) 775.36079, found 775.36562. [Zn2(T(6-MeO8Q)HPN)(OAc)](ClO4)2·4CH3CN. To the acetonitrile solution (0.4 mL) of HT(6-MeO8Q)HPN (7.8 mg, 10 μmol) were added Zn(OAc)2·2H2O (4.4 mg, 20 μmol) in methanol (0.2 mL) and NaClO4·H2O (2.8 mg, 20 μmol) in acetonitrile (0.4 mL), and the solution was kept at 4 °C to give [Zn2(T(6-MeO8Q)HPN)(OAc)](ClO4)2·4CH3CN as colorless crystals. Yield: 3.6 mg (31%). 1 H NMR (CD3CN, 300 MHz): δ (ppm) 9.67 (dd, J1 = 5.2 Hz, J2 = 1.8 Hz, 2H), 8.96 (dd, J1 = 4.9 Hz, J2 = 1.5 Hz, 2H), 8.50 (dd, J1 = 8.2 Hz, J2 = 1.5 Hz, 2H), 8.29 (dd, J1 = 8.2 Hz, J2 = 1.5 Hz, 2H), 7.75 (dd, J1 = 8.2 Hz, J2 = 4.9 Hz, 2H), 7.61 (dd, J1 = 8.5 Hz, J2 = 5.2 Hz, 2H), 7.55 (d, J = 2.4 Hz, 2H), 7.41 (d, J = 3.1 Hz, 2H), 7.18 (s, 4H), 5.28 (d, J = 14.0 Hz, 2H), 4.64 (br., 1H), 4.14 (d, J = 13.1 Hz, 2H), 3.94 (s, 6H), 3.84 (d, J = 13.4 Hz, 2H), 3.82 (s, 6H), 3.68 (d, 13.4 Hz, 2H), 2.61 (dd, J1 = 11.6 Hz, J2 = 2.7 Hz, 2H), 2.3−2.2 (m, 2H). 13C NMR B

DOI: 10.1021/acs.inorgchem.6b01967 Inorg. Chem. XXXX, XXX, XXX−XXX

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Figure 1. (a) UV−vis absorption and (b) fluorescence (λex = 341 nm) spectral changes of 34 μM HT(6-MeO8Q)HPN in DMF−HEPES buffer (50 mM HEPES, 100 mM KCl, pH = 7.5) (1:1) at 25 °C in the presence of 0−10 equiv of Zn2+.

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enhancement, indicating higher binding affinity with Cu2+ (Figure S5 in the SI). The selectivity of the ligand follows the Irving−Williams series. A large excess of Al3+, Cr3+, Fe2+, Fe3+, and Co2+ also prevented Zn2+-induced fluorescence due to the metal ion exchange. Biologically important metal ions, such as Na+, K+, and Ca2+, have no effect even in excess amount; however, fluorescence enhancement by added Mg2+ is unclear and under investigation. It is interesting that heavy metal ions including Ag+, Cd2+, and Hg2+ did not affect the fluorescence of the HT(6-MeO8Q)HPN-Zn2+ complex. The crystal structure for [Zn2(T(6-MeO8Q)HPN)(OAc)](ClO4)2 was determined by X-ray crystallography (Figure 3 and

RESULTS AND DISCUSSION Zn2+-Induced Fluorescence Enhancement of HT(6MeO8Q)HPN. HT(6-MeO8Q)HPN was synthesized from 6methoxy-8-chloromethylquinoline and 1,3-diamino-2-propanol and characterized by 1H/13C NMR and elemental analysis. The formation of the dinuclear zinc(II) complex was investigated by monitoring the absorbance and fluorescence spectral changes of HT(6-MeO8Q)HPN in DMF−HEPES buffer (50 mM HEPES, 100 mM KCl, pH = 7.5) (1:1) at 25 °C (Figures 1 and S1 in the SI). The titration curve reaches a constant maximum in the presence of ca. 10 equiv of Zn2+, indicating the moderate to weak metal binding of HT(6-MeO8Q)HPN. As the dizinc complex forms (see below), clear isosbestic points at 287 and 314 nm in the UV−vis spectra suggest the exclusive formation of a 1:2 (LM2) complex. ESI-MS measurements also support the direct formation of a LM2 complex (Figure S2 in the SI). This is completely different from the corresponding 1isoquinolylmethyl analogue 1-isoHTQHPN, which exhibits stepwise mono- and dinuclear complex formation.20 The limit of detection (LOD) for Zn2+ in DMF−H2O (1:1) was estimated to be 15 nM, which is significantly lower than the environmental allowance in Japan (0.15 μM (0.01 mg/L)) (Figure S3 in the SI). Figures 2 and S4 in the SI demonstrate the Zn2+-specificity of fluorescence enhancement of HT(6-MeO8Q)HPN. Cd2+ showed a slight increase in fluorescence intensity due to its Lewis acidity and d10 nature. No other metals responded to HT(6-MeO8Q)HPN; however, preincubation with 2 equiv of Cu2+ completely masked the Zn2+-induced fluorescence

Figure 3. X-ray crystal structure for [Zn2(T(6-MeO8Q)HPN)(OAc)](ClO4)2·4CH3CN with 50% probability ellipsoids. Hydrogen atoms, counteranions, and solvents were omitted for clarity.

Table S1 in the SI). The two zinc centers are five-coordinate and adopt intermediate geometry between trigonal bipyramid and square pyramid (τ = 0.48 and 0.49),21 each ligated by the three nitrogen atoms and an oxygen atom from the ligand, as well as an oxygen atom from the bridging acetate. No significant distortion derived from the ligand skeleton and substituents was observed (Table S2 in the SI). PO43−-Induced Fluorescence Enhancement of HT(6MeO8Q)HPN in the Presence of 2 equiv of Zn2+. The PO43−-induced absorbance and fluorescence changes of the dizinc complex were investigated (Figure 4). Absorption spectral changes were small (data not shown), but fluorescence spectra exhibited 2-fold enhancement in fluorescence intensity at 406 nm upon addition of 0.5 equiv of PO43− (ϕ = 0.159). In the presence of more than 0.5 equiv of PO43−, the fluorescence intensity decreased gradually (Figure 4b). This observation can be explained by the formation of a fluorescent [(PO4)(Zn2L)2]3+ ternary complex, followed by PO43−-induced

Figure 2. Relative fluorescence intensity of HT(6-MeO8Q)HPN at 406 nm in the presence of 2 equiv of metal ions in DMF−HEPES buffer (50 mM HEPES, 100 mM KCl, pH = 7.5) (1:1) at 25 °C. I0 is the emission intensity of the free ligand. C

DOI: 10.1021/acs.inorgchem.6b01967 Inorg. Chem. XXXX, XXX, XXX−XXX

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Figure 4. (a) Fluorescence spectral changes of 34 μM HT(6-MeO8Q)HPN in DMF−HEPES buffer (50 mM HEPES, 100 mM KCl, pH = 7.5) (1:1) at 25 °C in the presence of 2 equiv of Zn2+ and 0−10 equiv of PO43− (λex = 341 nm). (b) Fluorescence intensity changes at 406 nm.

Figure 5. (a) Partial ESI-MS spectrum for HT(6-MeO8Q)HPN in the presence of 2 equiv of Zn(OAc)2 and 0.5 equiv of Na3PO4 in CH3OH. (b) Simulated spectrum. See Figure S7 in the SI for full spectrum.

demetalation of a fluorescent complex. In the fluorescent anion detection using metal complexes, addition of excess anion often removes the metal ion from the fluorescent complex, resulting in a significant fluorescence decrease.17,22,23 In the present case, the demetalation was confirmed by the conversion to the free ligand in the UV−vis spectra (Figure S6 in the SI). In the presence of 0.5 equiv of PO43−, the formation of [(PO4)Zn4(T(6-MeO8Q)HPN)2]3+ (m/z = 632.8) was directly observed by ESI-MS spectrum as a tricationic species (Figures 5 and S7−

10 in the SI). A Job’s plot also supports the 1:2 stoichiometry for PO43− and [Zn2(T(6-MeO8Q)HPN)]3+ as a fluorescent ternary complex (Figure 6). In order to get detailed structural information on the fluorescent ternary complex, single crystals of a tetranuclear zinc complex were prepared from THF−CH3OH−H2O containing HT(6-MeO8Q)HPN, Zn(ClO4)2, and Na3PO4 (2:4:1 molar ratio) and analyzed by X-ray crystallography. Crystal data are summarized in Table S1 in the SI. Figure 7 D

DOI: 10.1021/acs.inorgchem.6b01967 Inorg. Chem. XXXX, XXX, XXX−XXX

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indication for intramolecular excimer formation between two quinolines attached to the same aliphatic nitrogen atom.24 Since no special feature was observed in the crystal structure of the ternary complex, the fluorescence enhancement in the presence of PO43− can be explained by phosphate-assisted zinc coordination.25 The negative charge of μ-PO4 reduces the electrostatic repulsion between neighboring zinc ions and facilitates the formation of a fluorescent ternary complex, [(μ4PO4)Zn4(T(6-MeO8Q)HPN)2]3+. Although only few PO43− species are actually present in the experimental condition (pH = 7.5), H2PO4− and HPO42− also assist the formation of the ternary complex accompanying the deprotonation. The addition of 0.5 equiv of PO43− actually promotes the Zn2+ complexation during the titration (Figure 8).

Figure 6. Job’s plot for fluorescence intensity of Zn2(T(6-MeO8Q)HPN)-PO43− complex in DMF−HEPES buffer (50 mM HEPES, 100 mM KCl, pH = 7.5) (1:1) at 25 °C. The sum of the concentrations of HT(6-MeO8Q)HPN and PO43− is 51 μM. Two equivalents of Zn2+ was added to the solution of HT(6-MeO8Q)HPN prior to mixing with PO43−.

Figure 8. Plot of the fluorescence intensity of HT(6-MeO8Q)HPN at 406 nm with increasing amounts of Zn2+ in the absence (blue triangles) and presence (red circles) of 0.5 equiv of PO43− in DMF− HEPES buffer (50 mM HEPES, 100 mM KCl, pH = 7.5) (1:1) at 25 °C.

On the basis of the fluorescent Zn2+ titration shown in Figure 1, approximately 50% of HT(6-MeO8Q)HPN binds with zinc in the presence of 2 equiv of Zn2+ under the present experimental conditions. The fluorescence feature and intensity of fully bound zinc complex in the presence of 10 equiv of Zn2+ closely resemble those of [(μ4-PO4)Zn4(T(6-MeO8Q)HPN)2]3+, by which the PO4-facilitated coordination mechanism is strongly supported. Since the unsubstituted HT(8Q)HPN exhibits similar but much smaller fluorescence changes with PO43−, the methoxy substitution is an effective strategy for signal enhancement. The fluorescence intensity changes of the dizinc complex in the presence of 0.5 equiv of various anions were investigated (Figure 9). No fluorescence wavelength shift was observed (Figure S11 in the SI), and PO43− exhibited the highest fluorescence enhancement. Other anions did not alter the fluorescence intensity except for HAsO42−, which displays a structural resemblance with PO43− exhibiting tetrahedral oxyanion character. Importantly, pyrophosphate, ATP, and other di- or triphosphate species quench the fluorescence due to the removal of Zn2+. These results highlight the high PO43− specificity of fluorescence enhancement via the formation of [(μ4-PO4)Zn4(T(6-MeO8Q)HPN)2]3+. The coexistence of other anions in the same concentration with PO43− does not impact the fluorescence of the ternary complex; however, a 10 times excess amount of PPi, ATP, ADP, AMP, and S2− prevents the fluorescent detection due to the Zn2+ removal mechanism discussed above.

Figure 7. X-ray crystal structure for [(μ4-PO4)Zn4(T(6-MeO8Q)HPN)2](ClO4)3·2THF·CH3OH·2.5H2O with 50% probability ellipsoids. Hydrogen atoms, counteranions, and solvents were omitted for clarity.

shows the ORTEP diagram for [(μ4-PO4)Zn4(T(6-MeO8Q)HPN)2]3+. Table S3 in the SI lists the bond distances around the metal center. The structure consists of two dizinc complexes assembled with a μ4-phosphate bridge. No μ-O monatomic bridge including a phosphate oxygen atom was formed. As mentioned, the tetranuclear complex with a μ4-PO4 bridge is extremely rare, and this is the first example of the (μ4PO4)Zn4 complex. All zinc centers are five-coordinate with highly distorted trigonal bipyramidal to square pyramidal geometries (τ = 0.54, 0.79, 0.67, and 0.39). There is an apparent π−π stacking interaction between one of the two quinoline moieties bound to Zn1 and Zn3 in the crystal structure, and this may stabilize the ternary complex in solution. All other quinolines were located separately, and there is no E

DOI: 10.1021/acs.inorgchem.6b01967 Inorg. Chem. XXXX, XXX, XXX−XXX

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present result will propose a new strategy for anion-sensing functionalities utilizing a simple fluorescent molecule system.

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ASSOCIATED CONTENT

* Supporting Information S

The Supporting Information is available free of charge on the ACS Publications website at DOI: 10.1021/acs.inorgchem.6b01967.

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Tables S1−S3 and Figures S1−S18 (PDF) Crystallographic data (CIF)

AUTHOR INFORMATION

Corresponding Author

*Tel/fax: +81-742-20-3095. E-mail: [email protected]. Figure 9. Relative fluorescence intensity of HT(6-MeO8Q)HPN at 406 nm in the presence of 2 equiv of Zn2+ and 0.5 equiv of anions (red bars), subsequent addition of 0.5 equiv of PO43− (blue bars), and further addition of 4.5 equiv of anions (green bars) in DMF−HEPES buffer (50 mM HEPES, 100 mM KCl, pH = 7.5) (1:1) at 25 °C. I0 is the emission intensity in the absence of anions.

Notes

The authors declare no competing financial interest.

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ACKNOWLEDGMENTS

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REFERENCES

This work was supported by the Research for Promoting Technological Seeds, JST, Adaptable and Seamless Technology Transfer Program through Target-driven R&D, JST, a Grant-in Aid for Scientific Research from the MEXT, Japan, and the Okumura Corporation public trust for constructer’s environment technology support fund.

The pH-dependence of PO43− detection of the dizinc complex was also monitored (Figure 10). Protonation of the

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Figure 10. Effect of pH on the fluorescence intensity of HT(6MeO8Q)HPN at 406 nm in the absence of metal and anions (green squares), in the presence of 2 equiv of Zn2+ (blue triangles), and in the presence of 2 equiv of Zn2+ and 0.5 equiv of PO43− (red circles) in DMF−H2O (1:1) at 25 °C.

nitrogen atoms of HT(6-MeO8Q)HPN at low pH prevents Zn2+ complexation, and the hydroxide anion removes Zn2+ from the ternary complex at high pH. In the presence of 0.5 equiv of PO43−, fluorescence was observed in the range of pH = 6−10, which is slightly extended at the high-pH region compared to that in the absence of PO43−.

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CONCLUSION HT(6-MeO8Q)HPN with 2 equiv of Zn2+ has been utilized for fluorescent detection of PO43−. X-ray crystallography reveals that the unprecedented formation of a tetranuclear zinc complex with a μ4-PO4 bridge is the key structure of fluorescence enhancement. The PO43− promotes the complex formation between HT(6-MeO8Q)HPN and Zn2+, which results in the increase of fluorescence intensity. A fine balance of moderate metal binding affinity and high fluorescence quantum yield of HT(6-MeO8Q)HPN upon Zn2+ binding exhibits a unique and specific sensing system for PO43−. The F

DOI: 10.1021/acs.inorgchem.6b01967 Inorg. Chem. XXXX, XXX, XXX−XXX

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DOI: 10.1021/acs.inorgchem.6b01967 Inorg. Chem. XXXX, XXX, XXX−XXX