Pyrophosphate-Induced Intramolecular Excimer Formation in

Jun 15, 2018 - The ligand library includes six HTQHPN derivatives with ... (IPPi/I0 = 12.5) and selectivity toward PPi sensing (IATP/IPPi = 20% and ...
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Article Cite This: Inorg. Chem. XXXX, XXX, XXX−XXX

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Pyrophosphate-Induced Intramolecular Excimer Formation in Dinuclear Zinc(II) Complexes with Tetrakisquinoline Ligands Yuji Mikata,*,†,‡,§ Risa Ohnishi,§ Risa Nishijima,§ Arimasa Matsumoto,† and Hideo Konno∥ †

Department of Chemistry, Biology, and Environmental Science, Faculty of Science, Nara Women’s University, Nara 630-8506, Japan KYOUSEI Science Center, Nara Women’s University, Nara 630-8506, Japan § 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 Downloaded via KAOHSIUNG MEDICAL UNIV on June 15, 2018 at 14:42:45 (UTC). See https://pubs.acs.org/sharingguidelines for options on how to legitimately share published articles.

‡

S Supporting Information *

ABSTRACT: Dinuclear Zn2+ complexes with HTQHPN (N,N,N′,N′tetrakis(2-quinolylmethyl)-2-hydroxy-1,3-propanediamine) derivatives have been prepared, and their pyrophosphate (PPi, P2O74−) sensing properties were examined. The ligand library includes six HTQHPN derivatives with electron-donating/withdrawing substituents, an extended aromatic ring, and six-membered chelates upon zinc binding. Complexation of ligand with 2 equiv of Zn2+ promotes small to moderate fluorescence enhancement around 380 nm, but in the cases of HTQHPN, HT(6-FQ)HPN (N,N,N′,N′tetrakis(6-fluoro-2-quinolylmethyl)-2-hydroxy-1,3-propanediamine), and HT(8Q)HPN (N,N,N′,N′-tetrakis(8-quinolylmethyl)-2-hydroxy-1,3-propanediamine), subsequent addition of PPi induced a significant fluorescence increase around 450 nm. This fluorescence enhancement in the longwavelength region is attributed to the conformational change of the bis(quinolylmethyl)amine moiety which promotes intramolecular excimer formation between adjacent quinolines upon binding with PPi. The structures of PPi- and phosphate-bound dizinc complexes were revealed by X-ray crystallography utilizing phenyl-substituted analogues. The zinc complex with HT(8Q)HPN exhibits the highest signal enhancement (IPPi/I0 = 12.5) and selectivity toward PPi sensing (IATP/IPPi = 20% and IADP/IPPi = 25%). The fluorescence enhancement turned to decrease gradually after the addition of more than 1 equiv of PPi due to the removal of zinc ion from the ligand−zinc−PPi ternary complex, allowing the accurate determination of PPi concentrations at the fluorescence maximum composition. The practical application of the present method was demonstrated monitoring the enzymatic activity of pyrophosphatase.

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(PeT) quenching, 15 chelation enhanced fluorescence (CHEF),22,23 internal charge transfer (ICT),12 excited state intramolecular proton transfer (ESIPT),29 and intermolecular14,19,21,28 and intramolecular25 excimer formation. Conformational change30 and restriction of rotational movement31 of fluorophore induced by PPi binding are also used as a reporter for fluorescent sensing. In addition to the monitoring of the fluorescence changes in a PPi-bound fluorophore, the indicator displacement assay (IDA) is another strategy to detect the interaction between PPi and host molecule via fluorescent9,11 and colorimetric32 readout. Instead of such metalloligand−PPi interactions, simple removal of quenching metal ions such as Cu2+ by external PPi also recovers the intrinsic fluorescence of fluorophores.33−35 Conversely, PPi also sequesters metal (mostly Zn2+, Cd2+, and Al3+) from fluorescent complexes, disrupting the fluorescence.36−40 In addition to the signal switching mechanism mentioned above, high specificity toward

INTRODUCTION

Fluorescent sensing of anions including phosphate species is one of the most important topics in analytical and environmental chemistry.1−4 Particularly, pyrophosphate (PPi, P2O74−) detection has been extensively studied because of its significant importance in biology.5,6 The detection of PPi is further utilized for applications such as DNA sequencing (pyrosequencing)7 and cancer diagnosis by monitoring telomerase activity via TRAP (telomeric repeat amplification protocol) connected with ELIDA (enzymatic luminometric inorganic pyrophosphate detection assay).8 Although a hydrogen-bonding interaction9−15 can be used to facilitate binding of fluorescent probes with PPi, most reported fluorescent PPi probes utilize a mononuclear 16−21 or dinuclear22−30 Zn2+ complex as the binding motif. The electrostatic interactions and intranuclear distance between the two cationic sites of the dinuclear Zn(II) complex provide an ideal environment for accommodation of PPi. Frequently used switching strategies for signal transduction upon PPi binding include inhibition of photoinduced electron transfer © XXXX American Chemical Society

Received: March 19, 2018

A

DOI: 10.1021/acs.inorgchem.8b00726 Inorg. Chem. XXXX, XXX, XXX−XXX

Article

Inorganic Chemistry PPi over other anions, especially phosphate analogues such as PO43− (Pi), ATP, and ADP, is an important prerequisite for PPi sensors. Recently, we have reported that the dinuclear zinc complex of HTQHPN (N,N,N′,N′-tetrakis(2-quinolylmethyl)-1,3-diamino-2-propanol (1), Chart 1)41 exhibits PPi-specific fluorescence

Chart 2. Derivatives of HTQHPN

Chart 1. HTQHPN and TQEN Structures

enhancement.30 The structure of HTQHPN was derived from fluorescent zinc sensor TQEN (N,N,N′,N′-tetrakis(2quinolylmethyl)ethylenediamine, Chart 1).42 The binding of PPi to the zinc centers of [Zn2(TQHPN)(OAc)]2+ induces the conformational change that promotes the formation of an intramolecular excimer between adjacent quinoline rings. Such an excimer formation in the ligand−zinc−PPi ternary complex was elucidated by X-ray crystallography with Ph2PPi, a diphenyl ester of PPi. In this study, we further examined the property of fluorescent PPi sensing with HTQHPN−Zn2 complexes. Six derivatives of HTQHPN with electron-donating (2) and -withdrawing (3) substituents, extended aromatic ring (4), and six-membered chelation upon zinc binding (5 and 6) have been investigated (Chart 2). By using the P1P2-diphenyl ester of PPi (Ph2PPi), the PPi-specific conformational changes that would induce excimer formation of dizinc complexes of ligands 1, 3, and 6 were elucidated by X-ray crystallography. Such an excimer was not expected for the complexes with acetate and monophosphate (PhOPO32−) analogues. The enzymatic activity of pyrophosphatase was successfully monitored by fluorescent changes of the HTQHPN−Zn2 complex.

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methoxyquinoline (1.38 g, 6.65 mmol), 1,3-diamino-2-propanol (150 mg, 1.66 mmol), potassium carbonate (4.60 g, 33.3 mmol), and potassium iodide (1.10 g, 6.63 mmol) in dry acetonitrile (40 mL) was refluxed for 2 days. After the reaction mixture was cooled to room temperature, solvent was evaporated and organic materials were extracted with chloroform−water. The organic layer was dried and evaporated to give HT(6-MeOQ)HPN (2) as yellow powder in quantitative yield. 1 H NMR (CDCl3, 300 MHz): δ (ppm) 7.90 (d, J = 9.2 Hz, 4H), 7.79 (d, J = 8.7 Hz, 4H), 7.39 (d, J = 8.5 Hz, 4H), 7.29 (dd, J1 = 9.2 Hz, J2 = 2.7 Hz, 4H), 6.93 (d, J = 2.7 Hz, 4H), 4.04 (d, J = 14.6 Hz, 4H), 3.96 (d, J = 14.3 Hz, 4H), 3.88 (s, 12H), 4.2−3.8 (br, 1H), 2.74− 2.72 (m, 4H). 13 C NMR (CDCl3, 75.5 MHz): δ (ppm) 157.2, 157.1, 143.1, 134.9, 130.0, 128.0, 121.7, 121.2, 105.0, 67.7, 61.8, 59.0, 55.5. Anal. Calcd for C47H48N6O6 (HT(6-MeOQ)HPN·H2O): C, 71.19; H, 6.10; N, 10.60. Found: C, 71.23; H, 5.92; N, 10.60. N,N,N′,N′-Tetrakis(6-fluoro-2-quinolylmethyl)-1,3-diamino-2propanol (HT(6-FQ)HPN (3)). The mixture of 2-chloromethyl-6fluoroquinoline (1.17 g, 6.00 mmol), 1,3-diamino-2-propanol (135 mg, 1.50 mmol), potassium carbonate (4.15 g, 30.0 mmol), and potassium iodide (996 mg, 6.00 mmol) in dry acetonitrile (30 mL) was refluxed for 2 days. After the reaction mixture was cooled to room temperature, the mixture was filtered and evaporated; then, organic materials were extracted with chloroform−water. The organic layer was dried and evaporated, and the residue was purified by alumina column chromatography (eluent: chloroform−methanol = 99:1) and recrystallized from ethanol to afford HT(6-FQ)HPN (3) as yellow powder (444 mg, 0.61 mmol, 41%). 1 H NMR (CDCl3, 300 MHz): δ (ppm) 7.98 (dd, J1 = 9.2 Hz, J2 = 5.2 Hz, 4H), 7.87 (d, J = 8.5 Hz, 4H), 7.47 (d, J = 8.5 Hz, 4H), 7.40 (m, 4H), 7.30 (dd, J1 = 8.9 Hz, J2 = 2.7 Hz, 4H), 6.10 (br, 1H), 4.2− 4.0 (br, 1H), 4.08 (d, J = 15.0 Hz, 4H), 4.02 (d, J = 14.7 Hz, 4H), 2.82−2.68 (m, 4H). 13 C NMR (CDCl3, 75.5 MHz): δ (ppm) 159.9 (d, J = 246.6 Hz), 159.0, 144.1, 135.4 (d, J = 4.9 Hz), 131.0 (d, J = 8.5 Hz), 127.6 (d, J =

EXPERIMENTAL SECTION

General. All reagents and solvents were purchased from commercial sources and used without further purification. Spectral grade N,N-dimethylformamide (DMF) (Spectrosol, Dojin) and MilliQ water (Millipore) were used for preparation of spectral measurement solutions. 1H/13C NMR (300/75.5 MHz) spectra were recorded on a Varian GEMINI 2000 spectrometer and referenced to Si(CH3)4 (TMS) or solvent signals. UV−vis spectra were measured on a Jasco V-660 spectrophotometer. Fluorescence spectra were measured on a Jasco FP-6300 spectrofluorometer. Fluorescence quantum yields were measured on a HAMAMATSU photonics C9920-02 absolute PL quantum yield measurement system. Fluorescence lifetimes were measured on a HORIBA fluorescence lifetime system Tempro. Caution! Perchlorate salts of metal complexes with organic ligands are highly explosive. These materials must be handled with care and appropriate precautions. Compounds. N,N,N′,N′-Tetrakis(2-quinolylmethyl)-1,3-diamino2-propanol (HTQHPN (1)),41 N,N,N′,N′-tetrakis(8-quinolylmethyl)1,3-diamino-2-propanol (HT(8Q)HPN (6)),30 and P1P2-diphenyl pyrophosphate (Ph2PPi)43 were prepared according to the literature method. The X-ray structure of HT(8Q)HPN (6) was obtained as a perchlorate salt in this work. N,N,N′,N′-Tetrakis(6-methoxy-2-quinolylmethyl)-1,3-diamino-2propanol (HT(6-MeOQ)HPN (2)). The mixture of 2-chloromethyl-6B

DOI: 10.1021/acs.inorgchem.8b00726 Inorg. Chem. XXXX, XXX, XXX−XXX

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Inorganic Chemistry

kept at 4 °C to give [Zn2(T(Phe)HPN)(OAc)](ClO4)2 as colorless crystals. Yield, 5.1 mg (4.1 μmol, 27%). 1 H NMR (DMSO-d6, 300 MHz): δ (ppm) 9.0−8.86 (m, 8H), 8.71 (d, J = 7.0 Hz, 2H), 8.36 (dd, J1 = 8.6 Hz, J2 = 7.6 Hz, 4H), 8.28 (d, J = 8.5 Hz, 2H), 8.18 (m, 2H), 8.1−8.0 (m, 4H), 8.0−7.8 (m, 6H), 7.65− 7.5 (m, 4H), 5.32 (d, J = 19.2 Hz, 2H), 5.22 (d, J = 18.3 Hz, 2H), 5.12 (d, J = 18.0 Hz, 2H), 4.96 (d, J = 18.3 Hz, 2H), 4.30 (m, 1H), 2.8−2.6 (m, 5H), 2.09 (br, 2H). 13 C NMR (DMSO-d6, 75.5 MHz): δ (ppm) 140.8, 140.3, 134.6, 134.3, 133.53, 133.47, 131.0, 129.4129.2, 128.7, 128.4, 127.1, 126.6, 125.2, 124.3, 123.8, 59.1, 56.5, 45.3. Anal. Calcd for C61H48Cl2N6O11Zn2 ([Zn2(T(Phe)HPN)(OAc)](ClO4)2): H, 3.97; C, 58.91; N, 6.76. Found: H, 4.10; C, 58.62; N, 6.97. [Zn2(T(8Q)HPN)(OAc)](ClO4)2 ([12·(OAc)](ClO4)2). The methanol solution (0.4 mL) of HT(8Q)HPN (6.6 mg, 10 μmol) was mixed with Zn(OAc)2·2H2O (4.4 mg, 20 μmol) in methanol (0.4 mL) and NaClO4·H2O (2.8 mg, 20 μmol) in methanol (0.6 mL); then, acetonitrile (0.2 mL) and DMF (0.4 mL) were added to dissolve the precipitated materials. The solution was kept at 4 °C under ether diffusion conditions to give [Zn2(T(8Q)HPN)(OAc)](ClO4)2 as colorless crystals. Yield, 7.8 mg (7.4 μmol, 74%). 1 H NMR (CD3CN, 300 MHz): δ (ppm) 9.89 (d, J = 4.9 Hz, 2H), 9.16 (d, J = 4.9 Hz, 2H), 8.63 (d, J = 8.2 Hz, 2H), 8.42 (d, J = 8.2 Hz, 2H), 8.09 (d, J = 8.2 Hz, 2H), 7.92 (d, J = 6.1 Hz, 2H), 7.83 (dd, J1 = 8.2 Hz, J2 = 4.3 Hz, 4H), 7.73−7.68 (m, 4H), 7.53 (d, J = 6.1 Hz, 2H), 7.48−7.43 (m, 2H), 5.36 (d, J = 13.7 Hz, 2H), 4.69 (m, 1H), 4.18 (d, J = 13.4 Hz, 2H), 3.94 (d, J = 13.7 Hz, 2H), 3.72 (d, J = 13.4 Hz, 2H), 2.67−2.59 (m, 5H), 2.31−2.23 (m, 2H). 13 C NMR (CD3CN, 75.5 MHz): δ (ppm) 154.1, 152.7, 145.0, 144.9, 142.2, 142.1, 135.1, 134.8, 130.8, 130.4, 130.4, 130.2, 129.9, 129.8, 128.1, 128.0, 123.1, 122.5, 60.5, 60.3, 59.2, 58.7, 25.3. Anal. Calcd for C45H42Cl2N6O12Zn2 ([Zn2(T(8Q)HPN)(OAc)](ClO4)2·H2O): H, 3.99; C, 50.96; N, 7.92. Found: H, 3.95; C, 50.88; N, 8.05. [Zn2(T(6-FQ)HPN)(PhOPO3)]ClO4 ([9·(PhOPO3)]ClO4). The THF solution (0.9 mL) of HT(6-FQ)HPN (11.0 mg, 15 μmol) was mixed with Zn(OAc)2·2H2O (6.6 mg, 30 μmol) in water (0.2 mL), Na2PhOPO3 (3.8 mg, 15 μmol) in water (0.2 mL), and NaClO4· H2O (2.1 mg, 15 μmol) in water (0.1 mL). The solution was kept at 4 °C to give [Zn2(T(6-FQ)HPN)(PhOPO3)]ClO4 as colorless crystals. Yield, 7.6 mg (6.7 μmol, 45%). 1 H NMR (DMSO-d6, 300 MHz): δ (ppm) 9.99 (br, 2H), 9.22 (br, 2H), 8.6−8.3 (m, 4H), 7.94 (br, 2H), 7.79 (br, 4H), 7.69 (br, 2H), 7.6−7.4 (m, 4H), 7.2−6.8 (m, 4H), 6.65 (br, 1H), 4.74 (d, J = 16.2 Hz, 2H), 4.7−4.2 (m, 5H), 4.11 (d, J = 18.6 Hz, 2H), 3.09 (br, d, 2H), 2.27 (br, 2H). 13 C NMR (DMSO-d6, 75.5 MHz): δ (ppm) 161.0, 157.9, 157.4, 142.3, 140.0, 139.7, 130.9, 130.1, 129.0, 128.0, 122.4, 122.1, 120.6, 119.9, 119.3, 110.9, 110.6, 61.6, 59.0, 55.6. Anal. Calcd for C49H40.8ClF4N6O10.4PZn2 ([Zn2(T(6-FQ)HPN)(PhOPO3)](ClO4)·1.4H2O): H, 3.57; C, 51.03; N, 7.29. Found: H, 3.41; C, 51.29; N, 7.00. [Zn2(T(6-FQ)HPN)(Ph2PPi)(CH3OH)]ClO4 ([9·(Ph2PPi)(CH3OH)]ClO4). The DMF solution (0.4 mL) of HT(6-FQ)HPN (7.3 mg, 10 μmol) was mixed with Zn(ClO4)2·6H2O (7.4 mg, 20 μmol) in methanol (0.3 mL) and Na2Ph2PPi (3.7 mg, 10 μmol) in water (0.1 mL). The solution was kept at room temperature to give [Zn2(T(6FQ)HPN)(Ph2PPi)(CH3OH)]ClO4 as colorless crystals. Yield, 5.3 mg (4.0 μmol, 40%). 1 H NMR (DMSO-d6, 300 MHz): δ (ppm) 8.97 (br, 2H), 8.64 (br, 2H), 8.41 (d, J = 8.5 Hz, 2H), 8.16 (d, J = 7.9 Hz, 2H), 7.75 (dd, J1 = 9.2 Hz, J2 = 2.7 Hz, 2H), 7.67 (br, 2H), 7.8−7.5 (m, 4H), 7.4−6.6 (br, 14H), 4.49−4.32 (m, 8H), 4.25−4.05 (m, 4H), 3.3−3.1 (m, 2H), 2.8− 2.6 (m, 2H). The proton of CH3OH was not assigned. 13 C NMR (DMSO-d6, 75.5 MHz): δ (ppm) 161.0, 160.7, 157.7, 157.5, 157.4, 156.9, 151.0, 141.8, 141.1, 139.1, 138.4, 129.0, 128.8, 128.6, 128.2, 128.1, 122.4, 120.9, 120.7, 120.4, 120.1, 119.7, 111.5, 111.2, 111.0, 64.6, 63.0, 62.4, 61.4, 59.0.

9.8 Hz), 121.5, 119.3 (d, J = 25.6 Hz), 110.4 (d, J = 22.0 Hz), 67.7, 61.8, 59.3. Anal. Calcd for C45H40F4N6O2 (HT(6-FQ)HPN·C2H5OH): C, 69.94; H, 5.22; N, 10.87. Found: C, 69.84; H, 5.22; N, 10.87. N,N,N′,N′-Tetrakis(6-phenanthridylmethyl)-1,3-diamino-2-propanol (HT(Phe)HPN (4)). The mixture of 6-chloromethylphenanthridine (455 mg, 2.00 mmol), 1,3-diamino-2-propanol (45 mg, 0.50 mmol), potassium carbonate (1.38 g, 10.0 mmol), and potassium iodide (531 mg, 2.00 mmol) in dry acetonitrile (20 mL) was refluxed for 36 h. After the reaction mixture was cooled to room temperature, solvent was evaporated and organic materials were extracted with chloroform−water. The organic layer was dried and evaporated, and the obtained yellow solid was purified by recrystallization from methanol to afford HT(Phe)HPN (4) as yellow powder (270 mg, 0.32 mmol, 63%). 1 H NMR (DMSO-d6, 300 MHz): δ (ppm) 8.65 (d, J = 8.2 Hz, 4H), 8.58 (dd, J1 = 7.9 Hz, J2 = 1.5 Hz, 4H), 8.18 (d, J = 7.9 Hz, 4H), 7.86 (dd, J1 = 7.6 Hz, J2 = 1.5 Hz, 4H), 7.74 (dd, J1 = 7.6 Hz, J2 = 7.3 Hz, 4H), 7.64−7.55 (m, 8H), 7.26 (dd, J1 = 7.6 Hz, J2 = 7.6 Hz, 4H), 5.41 (br, 1H), 4.26 (d, J = 13.7 Hz, 4H), 4.19 (d, J = 13.4 Hz, 4H), 3.82 (br, 1H), 2.6−2.4 (m, 4H). 13 C NMR (DMSO-d6, 75.5 MHz): δ (ppm) 158.2, 142.4, 132.0, 130.5, 129.1, 128.4, 127.0, 126.7, 124.5, 123.4, 122.3, 122.2, 60.6, 60.0. Anal. Calcd for C59H46N6O (HT(Phe)HPN): H, 5.42; C, 82.88; N, 9.83. Found: H, 5.32; C, 82.57; N, 9.57. N,N,N′,N′-Tetrakis(2-quinolylmethyl)-1,5-diamino-3-pentanol (HTQHPeN (5)). The mixture of 2-chloromethylquinoline hydrochloride (2.14 g, 10.0 mmol), 1,5-diamino-3-pentanol dihydrochloride (478 mg, 2.50 mmol), potassium carbonate (6.91 g, 50.0 mmol), and potassium iodide (1.66 mg, 10.0 mmol) in dry acetonitrile (80 mL) was refluxed for 3.5 days. After the reaction mixture was cooled to room temperature, solvent was evaporated and organic materials were extracted with chloroform−water. The organic layer was dried and evaporated, and the residue was purified by size exclusion chromatography (eluent: chloroform) to afford HTQHPeN (5) as yellow amorphous (853 mg, 1.25 mmol, 50%). 1 H NMR (CDCl3, 300 MHz): δ (ppm) 8.04 (d, J = 8.5 Hz, 4H), 7.93 (d, J = 8.5 Hz, 4H), 7.70−7.57 (m, 12H), 7.47 (t, J = 7.9 Hz, 4H), 4.1−3.9 (br, 1H), 4.02 (d, J = 14.6 Hz, 4H), 3.94 (d, J = 14.7 Hz, 4H), 2.89−2.72 (m, 4H), 1.75−1.64 (m, 4H). 13 C NMR (CDCl3, 75.5 MHz): δ (ppm) 159.8, 147.2, 136.2, 129.2, 128.7, 127.2, 127.1, 125.9, 120.7, 61.1, 52.0, 34.3. Anal. Calcd for C45H42N6O (HTQHPeN): C, 79.15; H, 6.20; N, 12.31. Found: C, 78.88; H, 6.25; N, 12.02. [Zn2(T(6-FQ)HPN)(OAc)](ClO4)2 ([9·(OAc)](ClO4)2). The chloroform solution (0.2 mL) of HT(6-FQ)HPN (7.3 mg, 10 μmol) was mixed with Zn(OAc)2·2H2O (4.4 mg, 20 μmol) in methanol (0.4 mL) and NaClO4·H2O (2.8 mg, 20 μmol) in methanol (0.4 mL). The solution was kept at 4 °C to give [Zn2(T(6-FQ)HPN)(OAc)](ClO4)2 as colorless crystals. Yield, 8.7 mg (7.8 μmol, 78%). 1 H NMR (CD3CN, 300 MHz): δ (ppm) 8.92 (dd, J1 = 9.5 Hz, J2 = 4.9 Hz, 2H), 8.85 (dd, J1 = 9.3 Hz, J2 = 5.0 Hz, 2H), 8.47 (d, J = 8.5 Hz, 2H), 8.40 (d, J = 8.5 Hz, 2H), 7.94 (ddd, J1 = 11.3 Hz, J2 = 7.4 Hz, J3 = 2.1 Hz, 2H), 7.69−7.57 (m, 8H), 7.43 (d, J = 8.5 Hz, 2H), 4.65 (d, J = 17.4 Hz, 2H), 4.42 (d, J = 8.5 Hz, 2H), 4.36 (d, J = 8.5 Hz, 2H), 4.16 (m, 1H), 4.07 (d, J = 17.4 Hz, 2H), 3.18 (dd, J1 = 11.9 Hz, J2 = 2.7 Hz, 2H), 2.4−2.3 (m, 5H). 13 C NMR (CD3CN, 75.5 MHz): δ (ppm) 162.7, 159.5, 159.2, 159.1, 159.1, 143.3, 143.0, 141.6, 141.6, 141.5, 141.4, 130.9, 130.8, 130.6, 130.5, 130.3, 129.0, 128.9, 123.4, 123.0, 122.6, 121.9, 121.5, 113.0, 112.7, 112.5, 112.2, 63.1, 60.0, 59.3, 56.6. Anal. Calcd for C45H39Cl2F4N6O12.5Zn2 ([Zn2(T(6-FQ)HPN)(OAc)](ClO4)2·1.5H2O): H, 3.44; C, 47.35; N, 7.36. Found: H, 3.38; C, 47.34; N, 7.34. [Zn2(T(Phe)HPN)(OAc)](ClO4)2 ([10·(OAc)](ClO4)2). The chloroform solution (0.8 mL) of HT(Phe)HPN (12.9 mg, 15 μmol) was mixed with Zn(ClO4)2·6H2O (11.1 mg, 30 μmol) in methanol (0.4 mL) and NaOAc (1.23 mg, 15 μmol) in methanol (0.2 mL); then, DMF (0.5 mL) was added to dissolve the precipitated materials. The solution was C

DOI: 10.1021/acs.inorgchem.8b00726 Inorg. Chem. XXXX, XXX, XXX−XXX

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Inorganic Chemistry

Figure 1. Fluorescence spectral changes of 34 μM (a) HTQHPN (1), (b) HT(6-MeOQ)HPN (2), (c) HT(6-FQ)HPN (3), (d) HT(Phe)HPN (4), (e) HTQHPeN (5), and (f) HT(8Q)HPN (6) in DMF−H2O (1:1) at 25 °C in the presence of increasing concentrations of Zn2+. The spectra with dotted lines and red marks indicate free ligand and ligand with 2 equiv of Zn2+, respectively.

Figure 2. Perspective views for (a) [9·(OAc)](ClO4)2, (b) [10·(OAc)](ClO4)2, and (c) [12·(OAc)](ClO4)2·0.5H2O in 50% probability. Hydrogen atoms, counteranions, and solvents are omitted for clarity. H NMR (DMSO-d6, 300 MHz): δ (ppm) 9.93 (br, 2H), 9.70 (br, 2H), 8.7−8.2 (m, 4H), 8.2−7.0 (m, 20H), 6.85 (br, 1H), 5.19 (br, 2H), 4.60 (br, 1H), 4.4−3.6 (m, 6H), 2.62 (br, 2H), 2.11 (br, 2H). 13 C NMR (DMSO-d6, 75.5 MHz): δ (ppm) 153.1, 143.8, 140.1, 133.4, 129.2, 128.6, 126.7, 121.7, 121.0, 119.8. Anal. Calcd for C49H 51ClN 6 O13.5PZn2 ([Zn 2(T(8Q)HPN)(PhOPO3)]ClO4·4.5H2O): H, 4.52; C, 51.75; N, 7.39. Found: H, 4.11; C, 51.50; N, 7.20. X-ray Crystallography. Single crystals of HT(8Q)HPN·2HClO4· THF (6·2HClO4·THF), [Zn2(T(6-FQ)HPN)(OAc)](ClO4)2 ([9· (OAc)](ClO4)2), [Zn2(T(Phe)HPN)(OAc)](ClO4)2 ([10·(OAc)](ClO4)2), [Zn2(T(8Q)HPN)(OAc)](ClO4)2·0.5H2O ([12·(OAc)]1

Anal. Calcd for C59H55ClF4N7O14.5P2Zn2 ([Zn2(T(6-FQ)HPN)(Ph2PPi)(CH3OH)]ClO4·DMF·0.5H2O): H, 3.96; C, 50.68; N, 7.01. Found: H, 3.76; C, 50.71; N, 7.02. [Zn2(T(8Q)HPN)(PhOPO3)]ClO4 ([12·(PhOPO3)]ClO4). The THF solution (1.0 mL) of HT(8Q)HPN (9.9 mg, 15 μmol) was mixed with Zn(OAc)2·2H2O (6.6 mg, 30 μmol) in water (0.2 mL), Na2PhOPO3 (3.8 mg, 15 μmol) in water (0.2 mL), and NaClO4· H2O (2.1 mg, 15 μmol) in water (0.01 mL). The solution was kept at 4 °C to give [Zn2(T(8Q)HPN)(PhOPO3)]ClO4 as colorless crystals. Yield, 4.8 mg (4.5 μmol, 30%). D

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Figure 3. Fluorescence spectral changes of 34 μM (a) HTQHPN (1), (b) HT(6-MeOQ)HPN (2), (c) HT(6-FQ)HPN (3), (d) HT(Phe)HPN (4), (e) HTQHPeN (5), and (f) HT(8Q)HPN (6) in DMF−H2O (1:1) at 25 °C in the presence of 2 equiv of Zn2+ and increasing amounts of PPi. The spectra with dotted pink lines, blue marks, and red marks indicate free ligand, ligand with 2 equiv of Zn2+, and ligand with 2 equiv of Zn2+ and 1 equiv of PPi, respectively.

Figure 4. Plot of fluorescence intensity of (a) HTQHPN (1) at 455 nm, (b) HT(6-MeOQ)HPN (2) at 478 nm, (c) HT(6-FQ)HPN (3) at 441 nm, (d) HT(Phe)HPN (4) at 460 and 386 nm, (e) HTQHPeN (5) at 455 nm, and (f) HT(8Q)HPN (6) at 455 nm in DMF−H2O (1:1) at 25 °C in the presence of 2 equiv of Zn2+ and increasing amounts of PPi. (ClO 4 ) 2 ·0.5H 2 O), [Zn 2 (T(6-FQ)HPN)(PhOPO 3 )]ClO 4 ([9· (PhOPO3)]ClO4), [Zn2(T(6-FQ)HPN)(Ph2PPi)(CH3OH)]ClO4· DMF ([9·(Ph2PPi)(CH3OH)]ClO4·DMF), and [Zn2(T(8Q)HPN)(PhOPO3)]ClO4 ([12·(PhOPO3)]ClO4) were coated with ParatoneN oil and mounted on a glass fiber or a Mounted CryoLoop. All crystallographic data were collected at 153 K on a Rigaku Mercury or Saturn CCD detector, with a monochromatic Mo Kα sealed tube source, operating at 50 kV/40 mA (Mercury) or 50 kV/24 mA (Saturn). CrystalClear (Rigaku) was used for data collection and processing. All structures were solved by direct methods (SIR-9244 or SIR-200845), expanded using Fourier techniques, and refined by fullmatrix least-squares methods on F2 (SHELXL Version 2016/646). For [9·(OAc)](ClO4)2, [10·(OAc)](ClO4)2, [9·(PhOPO3)]ClO4, and [12·(PhOPO3)]ClO4, the disordered solvent molecules were removed by PLATON/SQUEEZE.47 Crystal data are summarized in Tables S1−S4.

Monitoring of Pyrophosphatase Activity. The enzymatic reaction mixture (3 mL) containing 340 μM PPi·4Na, 17 μM MgCl2, and 1 unit of pyrophosphatase (PPase, Fluka) in HEPES buffer (50 mM HEPES, 100 mM KCl, pH = 7.5) was incubated at 20 °C. During the reaction, 300 μL aliquots were taken from the reaction mixture every 10 min. The aliquot was immediately mixed with fluorescence monitoring solution (3 mL) which contains 34 μM HTQHPN and 68 μM Zn(OAc)2 in DMF−HEPES buffer (1:1); then, the fluorescence intensity at 455 nm (λex = 317 nm) was monitored to quantitate the concentration of PPi. The control experiment without PPase was executed.

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RESULTS AND DISCUSSION Ligand Synthesis and Characterization. All ligands were synthesized from corresponding chloromethylquinolines with a E

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Figure 5. Fluorescence spectra of 34 μM (a) HTQHPN (1), (b) HT(6-MeOQ)HPN (2), (c) HT(6-FQ)HPN (3), (d) HT(Phe)HPN (4), (e) HTQHPeN (5), and (f) HT(8Q)HPN (6) in DMF−H2O (1:1) at 25 °C in the presence of 2 equiv of Zn2+ and 1 equiv of various anions. The spectra with dotted lines, blue marks, and red marks indicate free ligand, ligand with 2 equiv of Zn2+, and ligand with 2 equiv of Zn2+ and 1 equiv of PPi, respectively.

showed characteristic changes depending on the ligand structure (Figure 3). For most complexes, a new fluorescence band around 450 nm was developed by the addition of 1 equiv of PPi, but this emission was disappeared upon addition of excess amount of PPi (Figure 4). For ligands 1, 3, and 6 in the presence of 2 equiv of Zn2+, the fluorescence band with moderate intensity around 450 nm increased until addition of 1 equiv of PPi (Figure 3a,c,f). Then, addition of more than 1 equiv of PPi induced a gradual fluorescence decrease (Figure 4a,c,f). This observation indicates the formation of fluorescent ligand−Zn2+−PPi (1:2:1) ternary complex in the presence of 1 equiv of PPi, where the maximum fluorescence was observed, followed by the removal of Zn2+ from the ternary complex upon addition of excess PPi. The UV−vis spectra reveal the formation of free ligand in the presence of excess amount of PPi (Figure S3). This OFF− ON−OFF sequential fluorescent change serves an accurate determination of PPi concentration at the fluorescence maximum composition by titration experiment. For 5, a fluorescence spectral change similar to that of the above complexes was observed (Figure 3e and 4e); however, the fluorescence intensity around 450 nm is small compared to 1, which possesses the same chromophore with 5, due to the weak metal binding affinity of 1,5-pentanediamine skeleton that forms a Zn2+-containing six-membered chelate. For 2 (Figure 3b) and 4 (Figure 3d), the strong emission of dizinc complexes (8 and 10) at 350−450 nm decreased upon addition of PPi, and the emission around 450 nm was scarcely detected in the presence of 1 equiv of PPi. The formation of the ligand−Zn2+− PPi (1:2:1) ternary complex for 2, 4, and 5 was confirmed by ESI mass spectra using the diphenyl ester of PPi (Ph2PPi) as an alternative for PPi (Figures S4−S9), but the strong fluorescence of the dizinc complexes 8 and 10 at the short wavelengths prevents the monitoring of long-wavelength emission of PPibound species. For 4, the fluorescence intensity ratio (I460/I386) reveals the formation of ternary complex in the presence of 1 equiv of PPi (Figure 4d).

quarter of an equivalent of 1,3-diamino-2-propanol or 1,5diamino-3-pentanol. The structure of the product was confirmed by 1H/13C NMR and elemental analysis. For HT(8Q)HPN·2HClO4, X-ray crystallography reveals the structure in the solid state (Table S1 and Figure S1). Synthesis and Characterization of Dinuclear Zinc(II) Complexes. The formation of dinuclear zinc(II) complexes 7−12 (7, [Zn2(TQHPN)]3+; 8, [Zn2(T(6-MeOQ)HPN)]3+; 9, [Zn 2 (T(6-FQ)HPN)] 3+ ; 10, [Zn 2 (T(Phe)HPN)] 3+ ; 11, [Zn2(TQHPeN)]3+; 12, [Zn2(T(8Q)HPN)]3+) (Chart 2) was confirmed by monitoring the absorbance and fluorescence spectral changes of 1−6 in 50% aqueous DMF at 25 °C (Figure 1 and Figure S2). All ligands exhibited several isosbestic points on UV−vis spectra until saturation of the spectral change around 2 equiv of Zn2+, suggesting the formation of the 1:2 (LM2) complex directly from the free ligand. The mononuclear complex48 was not detected for the present quinoline-based ligands. Single crystals of dizinc complexes, [9·(OAc)](ClO4)2, [10· (OAc)](ClO4)2, and [12·(OAc)](ClO4)2·0.5H2O, suitable for X-ray crystallography were prepared from corresponding ligand containing 2 equiv of Zn2+ in the presence of acetate and perchlorate (Figure 2 and Tables S1−S2). These structures show that each metal center is supported by three nitrogen atoms and an alkoxo oxygen atom of the ligand, as well as an oxygen atom from the acetate bridge, constructing the fivecoordinate, dinuclear zinc centers with trigonal bipyramid geometry (τ49 = 0.72 and 0.55 for [9·(OAc)]2+; 0.72 and 0.57 for [10·(OAc)]2+; 0.58 and 0.27 for [12·(OAc)]2+). No significant changes in distances and angles around zinc centers derived from ligand structure were observed except for that of [12·(OAc)]2+, which exhibits significant distortion due to the six-membered chelate ring. PPi-Induced Fluorescence Spectral Changes of 1−6 in the Presence of Zn2+. The PPi-induced absorbance and fluorescence changes of 1−6 in the presence of 2 equiv of Zn2+ were next investigated. In all cases, absorption spectral changes were small (Figure S3) but fluorescence spectral changes F

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Figure 6. Fluorescence intensity of a 34 μM (a) HTQHPN (1) at 455 nm, (b) HT(6-MeOQ)HPN (2) at 478 nm, (c) HT(6-FQ)HPN (3) at 441 nm, (d) HT(Phe)HPN (4) at 460 nm/386 nm, (e) HTQHPeN (5) at 455 nm/360 nm, and (f) HT(8Q)HPN (6) at 455 nm in DMF−H2O (1:1) at 25 °C in the presence of 2 equiv of Zn2+ and (red bars) 1 equiv of anions, (blue bars) 1 equiv of anions + 1 equiv of PPi, and (green bars) 1 equiv of PPi and 10 equiv of anions.

Fluorescence Anion Specificity of 1−6 in the Presence of Zn2+. Figure 5 shows the fluorescence spectra of 1−6 in the presence of 2 equiv of Zn2+ and 1 equiv of various anions. In most cases, the fluorescence spectrum in the presence of PPi exhibits specific emission around 450 nm assigned as an excimer emission from its wavelength and fluorescence lifetime values (τ = 6−10 ns),48,50−52 where the conformational change around the Zn2+ center induced by the binding with PPi facilitates the parallel configuration of two chromophores attached to the same aliphatic nitrogen atom at the excited state (vide infra). Similar but much smaller emission around this region was observed for ATP and ADP. Other anions did not alter the fluorescence of the dizinc complex except for SO42− with HT(Phe)HPN (4), which exhibited a 1.5-fold fluorescence enhancement at 386 nm (Figure 5d). The ternary complex containing SO42− seems to be formed, but no excimer emission around 450 nm was observed due to the difference in the structure of the anion-bound complex. The PPi-induced fluorescence spectral changes are scarcely affected by the coexistence of 1 equiv of other anions (Figure 6, blue bars), but excess amounts of phosphate-related anions such as ATP, ADP, AMP, and PO4 3− prevented the fluorescence enhancement due to the removal of Zn2+ from the complex as observed in the presence of excess PPi (Figure 6a,c,f, green bars). These results demonstrate the high specificity of fluorescence enhancement and binding affinity of dizinc complexes 7−12 derived from ligands 1−6 with PPi. Characterization of Ternary Complexes Containing P1P2-Ph2PPi and PhOPO32−. In order to gain structural information for PPi-bound species, single crystals of ternary complex were prepared utilizing P1P2-Ph2PPi (Ph2PPi) as a crystallizable alternative for PPi. We have previously reported that the use of phenyl esters such as Ph2PPi and PhOPO32− is effective for preparation of single crystals of ternary complexes with PPi and PO43−.30 Ph2PPi also induced long-wavelength emission similar to that of PPi upon mixing with a Zn2+containing solution of 3 (Figure 7). The ternary complex with PhOPO32− was also prepared as a nonfluorescent control

Figure 7. Fluorescence spectra of 34 μM HT(6-FQ)HPN (3) in DMF−H2O (1:1) at 25 °C in the presence of 2 equiv of Zn2+ and 1 equiv of Ph2PPi (blue), PhOPO32− (red), PPi (light blue), and PO43− (orange).

complex (Figure 7). Crystal data for [Zn2(T(6-FQ)HPN)(PhOPO3)]ClO4 ([9·(PhOPO3)]ClO4), [Zn2(T(6-FQ)HPN)(Ph2PPi)(CH3OH)]ClO4·DMF ([9·(Ph2PPi)(CH3OH)]ClO4· DMF), and [Zn 2 (T(8Q)HPN)(PhOPO 3 )]ClO 4 ([12· (PhOPO3)]ClO4 are summarized in Tables S3 and S4. Figure 8 shows the perspective views of cationic portion of the complexes. All structures shown in Figure 8 exhibit (pyro)phosphatebridged dizinc complexes. As reported previously,30 complexation with PPi induced geometrical changes in the zinc center from trigonal bipyramid to octahedron by an additional oxygen atom coordination from nonbridging phosphate moiety or solvent molecule (Figure 8b). This structural change is specific for PPi complex. Two quinolines attached to the same aliphatic nitrogen atom are responsible to the intramolecular excimer emission48,50−52 induced by the coordination of PPi because the octahedral geometry is favorable for excimer formation in comparison to the trigonal bipyramid geometry.48 Such a conformational change to octahedron was not observed for monophosphate-bridged ternary complexes (Figure 8a,c). Thus, the PPi-specific fluorescence enhancement via excimer G

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Figure 8. Perspective views for (a) [9·(PhOPO3)]ClO4, (b) [9·(Ph2PPi)(CH3OH)]ClO4·DMF, and (c) [12·(PhOPO3)]ClO4 in 50% probability. Hydrogen atoms except for the hydrogen-bonding OH group (in part b), counteranions, and solvents are omitted for clarity. For part c, only one of the two crystallographically independent molecules is shown.

The electron-withdrawing substituent on the quinoline ring reduces the fluorescence intensity of dizinc and ternary complexes. The weakened metal binding affinity is also responsible for the diminished fluorescence of 3. The strong fluorescence intensity of the dizinc complex of methoxysubstituted derivative HT(6-MeOQ)HPN (2) around 410 nm diminished by the addition of PPi although the excimer emission band around 480 nm was faintly observed (Figure 3b). The methoxy substituent did not increase the intensity of excimer emission in comparison to the Zn2+-induced monomer emission. For HT(Phe)HPN (4), the fluorescence intensity at the short wavelength was quenched by PPi, but the fluorescence intensity ratio at long and short wavelength (I460/I386) exhibits PPi specificity (Figure 6d, red bars). Similarly, the I455/I360 value highlights the PPi specificity of HTQHPeN (5) over ATP and ADP (Figure 6e, red bars; IATP/ IPPi = 37% and IADP/IPPi = 26%) because the emission around 455 nm is specific to PPi (Figure 5e). Among compounds 1−6, HT(8Q)HPN (6) exhibits the highest signal enhancement (IPPi/IZn2 = 13) and PPi selectivity (IATP/IPPi = 20% and IADP/ IPPi = 25%) in the presence of 2 equiv of Zn2+ (Table 1). The values of the fluorescence lifetime of the ternary complexes including 1 (7.3 ns), 3 (6.2 ns), and 6 (10.0 ns) (Table 1 and Figure S10−S12) support that the excimer emission is the main component of the PPi-specific long-wavelength emission.48,52 Effect of pH on Fluorescent Detection of PPi with 1. Figure 9 displays the pH-dependence of fluorescent PPi detection with 1. In the presence of 2 equiv of Zn2+ and 1 equiv of PPi, strong fluorescence appeared at 455 nm in the range pH = 4−9, within which the fluorescent ternary complex formed. Protonation on the nitrogen atoms of 1 at low pH regions and the formation of Zn(OH)2 at high pH inhibits the

formation is well explained by the change in the coordination geometry around the zinc center in the PPi-bound ternary complex. Comparison of the Property of 1−6 as a Fluorescent Sensor for PPi. The fluorescent PPi sensing properties of ligands 1−6 were summarized in Table 1. As seen in the ϕZn2 and ϕPPi values in Table 1, the fluoro-substituted derivative HT(6-FQ)HPN (3) exhibited much smaller fluorescence intensity in dizinc and PPi complexes in comparison to HTQHPN (1), but PPi selectivity over other diphosphate species was slightly improved (IATP/IPPi = 33% and IADP/IPPi = 24% for 3; while IATP/IPPi = 50% and IADP/IPPi = 30% for 1). Table 1. Fluorescent Properties of HTQHPN (1), HT(6MeOQ)HPN (2), HT(6-FQ)HPN (3), HT(Phe)HPN (4), HTQHPeN (5), and HT(8Q)HPN (6) λex (nm) Zn2 λem (nm) IZn/I0 PPi λem (nm) IPPi/IZn2 IATP/IPPi IADP/IPPi ϕL ϕZn2 ϕPPi τPPi (ns)

1

2

3

4

5

6

317 379 43 455 7.5 50% 30% 0.003 0.009 0.029 7.3c

336 413 20 478 0.60 156% 139% 0.016 0.092 0.047

317 419 9 441 6.5 33% 24% 0.003 0.005 0.012 6.2d

352 386 53 460 1.6 45%a 21%a 0.006 0.097 0.038

317 371 9.2 455 4.3 37%b 26%b 0.006 0.008 0.017

317 455 7 455 13 20% 25% 0.004 0.006 0.025 10.0c

Ratio of I460/I386. bRatio of I455/I360. cMeasured at 460 ± 10 nm (λex = 331 nm). dMeasured at 430 ± 10 nm (λex = 331 nm). a

H

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skeleton. The intramolecular excimer emission upon binding with PPi was observed with the common octahedral geometry around the zinc center. Increasing the fluorescence intensity of the ligand itself by addition of methoxy substituent (2) or introduction of an extended π-system (4) resulted in significant fluorescence quenching in the PPi complex due to the high fluorescence intensity of the dizinc complex. The PPi selectivity over other diphosphate-related anions was slightly improved in 3 and 6. The fluorescent PPi detection by the dizinc complex of 1 was applied in monitoring the enzymatic activity of PPase. Although further improvement to enhance the fluorescence intensity and PPi selectivity is required, the present work demonstrates the importance of HTQHPN structure as a useful platform for versatile fluorescent PPi sensing devices. The present investigation provides a useful information for logical design strategy of fluorescent probe molecules with simple and basic structure.

Figure 9. Effect of pH on fluorescence intensity (at 455 nm) of 34 μM HTQHPN (1) (green), [Zn 2 (TQHPN)] 3+ (7) (blue), and [Zn2(TQHPN)]3+ (7) in the presence of 1 equiv of PPi (red) in DMF−H2O (1:1) at 25 °C.

formation of [7·(PPi)]−. This pH window is suitable for PPi detection under physiological condition. Monitoring PPase Activity with 1. The enzymatic activity of pyrophosphatase (PPase) was evaluated by monitoring the fluorescence using HTQHPN (1) (Figure 10). The hydrolysis

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ASSOCIATED CONTENT

S Supporting Information *

The Supporting Information is available free of charge on the ACS Publications website at DOI: 10.1021/acs.inorgchem.8b00726. Tables and figures with characterization details and data (PDF) Accession Codes

CCDC 1829558−1829564 contain the supplementary crystallographic data for this paper. These data can be obtained free of charge via www.ccdc.cam.ac.uk/data_request/cif, or by emailing [email protected], or by contacting The Cambridge Crystallographic Data Centre, 12 Union Road, Cambridge CB2 1EZ, UK; fax: +44 1223 336033.

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Figure 10. Fluorescence intensity change (at 455 nm) of 34 μM HTQHPN (1) 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 aliquot of PPi hydrolysis reaction solution of indicated time in the presence (red) or absence (blue) of PPase. See Experimental Section for details.

AUTHOR INFORMATION

Corresponding Author

*Phone/fax: +81-742-20-3095. E-mail: [email protected]. jp. ORCID

Yuji Mikata: 0000-0002-9450-0908 Notes

of PPi was conducted in the reaction mixture containing PPase and MgCl2 in HEPES buffer (50 mM HEPES, 100 mM KCl, pH = 7.5) at 20 °C. Every 10 min, the aliquots of the reaction solution were mixed with fluorescence monitoring solution which contains HTQHPN and 2 equiv of Zn(OAc)2 in DMF− HEPES buffer (1:1); then, the fluorescence intensity at 455 nm (λex = 317 nm) was evaluated. As shown in Figure 10, the fluorescence intensity attributed to the HTQHPN-Zn2−PPi ternary complex was gradually decreased as the PPase hydrolyses PPi. The control experiment that lacks PPase did not exhibit any decrease of fluorescence, and the rate of fluorescence decay was a function of PPase concentration. It should be noted that, at the late stage of the reaction, the hydrolyzed product PO43− sequesters Zn2+ from HTQHPN− Zn2 (7) and HTQHPN−Zn2−PPi complexes, lowering the sensitivity of detection of PPi as discussed above.

The authors declare no competing financial interest.

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ACKNOWLEDGMENTS This work was supported by the Research for Promoting Technological Seeds, JST, Adaptable and Seamless Technology Transfer Program through Target-driven R&D, JST, Grant-in Aid for Scientific Research provided by JSPS KAKENHI Grant Number JP15K05454 and the Nara Women’s University Intramural Grant for Project Research.

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CONCLUSIONS The heptadentate ligands 1−6 exhibit PPi-induced fluorescent response in the presence of 2 equiv of Zn2+, depending on the substituents on the quinoline ring, chromophore, and ligand I

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DOI: 10.1021/acs.inorgchem.8b00726 Inorg. Chem. XXXX, XXX, XXX−XXX

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DOI: 10.1021/acs.inorgchem.8b00726 Inorg. Chem. XXXX, XXX, XXX−XXX