DOI: 10.1002/chem.201406453

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Pincer-Type Platinum(II) Complexes Containing N-Heterocyclic Carbene (NHC) Ligand: Structures, Photophysical and AnionBinding Properties, and Anticancer Activities Kai Li,[a, b] Taotao Zou,[a] Yong Chen,[a, c] Xiangguo Guan,[a] and Chi-Ming Che*[a, b] Abstract: Two classes of pincer-type PtII complexes containing tridentate N-donor ligands (1–8) or C-deprotonated N^C^N ligands derived from 1,3-di(2-pyridyl)benzene (10– 13) and auxiliary N-heterocyclic carbene (NHC) ligand were synthesized. [Pt(trpy)(NHC)]2 + complexes 1–5 display green phosphorescence in CH2Cl2 (F: 1.1–5.3 %; t: 0.3–1.0 ms) at room temperature. Moderate-to-intense emissions are observed for 1–7 in glassy solutions at 77 K and for 1–6 in the solid state. The [Pt(N^C^N)(NHC)] + complexes 10–13 display strong green phosphorescence with quantum yields up to 65 % in CHCl3. The reactions of 1 with a wide variety of anions were examined in various solvents. The tridentate Ndonor ligand of 1 undergoes displacement reaction with

Introduction Luminescent platinum(II) complexes with tridentate N-donor ligands such as terpyridine (trpy) have been extensively studied for their rich spectroscopic and luminescence properties.[1] Most of the studies on luminescent platinum(II)-terpyridyl complexes were focused on their photophysical and emission properties that are associated with intra- and intermolecular Pt···Pt and ligand–ligand interactions.[2] However, [Pt(trpy)Cl] + is non-emissive in solutions at room temperature; this has been attributed to the thermal population of the close-lying d–d states, which leads to quenching of the emissive excited states.[3] As a consequence, there have been extensive works [a] Dr. K. Li, Dr. T. Zou, Prof. Dr. Y. Chen, Dr. X. Guan, Prof. Dr. C.-M. Che State Key Laboratory of Synthetic Chemistry Institute of Molecular Functional Materials HKU-CAS Joint Laboratory on New Materials and Department of Chemistry, The University of Hong Kong Pokfulam Road, Hong Kong (P.R. China) Fax: (+ 852) 2915-5176 E-mail: [email protected] [b] Dr. K. Li, Prof. Dr. C.-M. Che HKU Shenzhen Institute of Research and Innovation Shenzhen 518053 (P.R. China) [c] Prof. Dr. Y. Chen Key Laboratory of Photochemical Conversion and Optoelectronic Materials, Technical Institute of Physics and Chemistry Chinese Academy of Sciences, Beijing 100190 (P.R. China) Supporting information for this article is available on the WWW under http://dx.doi.org/10.1002/chem.201406453. Chem. Eur. J. 2015, 21, 7441 – 7453

CN¢ in protic solvents. Similar displacement of the N^C^N ligand by CN¢ has been observed for 10, leading to a luminescence “switch-off” response. The water-soluble 7 containing anthracenyl-functionalized NHC ligand acts as a light “switch-on” sensor for the detection of CN¢ ion with high selectivity. The in vitro cytotoxicity of the PtII complexes towards HeLa cells has been evaluated. Complex 12 showed high cytotoxicity with IC50 value of 0.46 mm, whereas 1–4 and 6–8 are less cytotoxic. The cellular localization of the strongly luminescent complex 12 traced by using emission microscopy revealed that it mainly localizes in the cytoplasmic structures rather than in the nucleus. This complex can induce mitochondria dysfunction and subsequent cell death.

to vary the lowest-lying excited states as well as the luminescence properties of [Pt(trpy)L]n + complexes by using other ancillary ligand (L). For example, coordination of a strong sdonor such as aryl/alkyl acetylide ligand to PtII can raise the energy of 5dx2¢y2 orbital of PtII, thereby switch-on the metal-toligand charge transfer/ligand-to-ligand charge transfer (3MLCT/ 3 LLCT) emission in solutions at ambient temperature.[4] Due to the availability of the axial coordination site, PtII complexes are susceptible to nucleophilic attack through innersphere PtII-substrate binding interactions,[5] which could result in significant changes in absorption and emission energies. The non-covalent binding interactions between pincer-type PtII complexes and biomolecules have been used in the design of luminescent switch-on probes for the detection of biological cellular targets.[6] Nucleophilic attack at the axial coordination site can also result in ligand displacement reactions of PtII complexes. It is conceived that by rational molecular design, color, and luminescence changes could be integrated into the ligand exchange reactions, hence providing another approach to develop luminescent sensing and signaling.[7] In recent years, N-heterocyclic carbene (NHC) ligands, owing to their strong s-donating properties, have received an upsurge of interest in the design of luminescent PtII complexes.[8] Luminescent PtII-NHC complexes with tunable physical and chemical properties have been exploited for material applications such as organic light emitting diodes (OLEDs),[9, 10] vapor sensors,[11] and bio-probes, and for medicinal use as anticancer agents.[6c, 12] Previously, we reported luminescent PtII complexes containing a tridentate C-deprotonated C^N^N ligand and an-

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Scheme 1. Chemical structures of [Pt(C^N^N)(NHC)]PF6 and 1–13.

cillary NHC ligand (e.g., [Pt(C^N^N)(NHC)PF6] in Scheme 1).[6c] Coordination of NHC to the Pt(C^N^N) moiety significantly improves the emission properties of this class of complexes in solutions. Moreover, the strongly coordinating tridentate C^N^N and NHC ligands confer high stability of this type of complexes under physiological conditions, which imparts promising anticancer properties to the [Pt(C^N^N)(NHC)] + complexes. We have also reported on the highly phosphorescent [Pt(N^C^N)(NHC)] + complexes and demonstrated an interesting solid-state luminescence sensor for acidic vapor derived from this structure.[11a] Herein is described the synthesis and properties of a series of luminescent pincer-type platinum(II) complexes containing NHC ligand (1–8 and 10–13, Scheme 1). The strong s-donating NHC ligand destabilizes the 5dx2¢y2 orbital and d–d excited state(s) of PtII, resulting in enhanced/switch on effect of phosphorescence of platinum(II) terpyridyl complexes in solutions at room temperature. The dicationic [Pt(trpy)(NHC)]2 + scaffold imparts interesting substrate-binding properties. The treatment of 1 with various anions has been examined. By a rational molecular design, complex 7 is water-soluble and can be used as colorimetric and luminescence switch-on sensor for detection of CN¢ in aqueous media with good selectivity. Complex 12 was found to be highly cytotoxic towards cervical epithelioid carcinoma (HeLa) cells and the mechanism of action has been implicated by using fluorescence microscopic analysis.

Results and Discussion Synthesis For the preparation of 1–5, the corresponding imidazolium salt was firstly treated with tBuOK in MeOH heated at reflux to afford the NHC ligand, which was used for subsequent coordination to [Pt(trpy)(MeCN)](OTf)2 or [Pt(tBu3trpy)(MeCN)](OTf)2, which were both prepared following a modified literature method.[3d] Anion metathesis with NH4PF6 and subsequent diffusion of diethyl ether vapor into a MeCN (for 1–3) or a CHCl3 (for 4–5) solution afforded the products as light yellow crystals (for 1–3 and 5) or powder (for 4) with moderate yields of 36– 60 %. The water-soluble complexes 6–8 were obtained by using a silver-carbene transmetalation method in yields of 32– 46 %.[13] The preparation of 10 and 13 has been described elsewhere;[11a] complexes 11 and 12 were similarly prepared. Complexes 1–5 are readily soluble in MeCN, MeOH, and DMSO, but sparingly soluble in CHCl3 and CH2Cl2. Complexes 10–13 have good solubility in all these solvents. Complex 6 having two pendant SO3¢ moieties is soluble in water. Complex 7 is soluble in DMSO and water. Complex 8 is soluble in MeOH, DMSO, and water. All complexes were characterized by NMR and FABMS spectroscopies and satisfactory elemental analyses. 1H-1H COSY NMR spectra of 1 and 11 are provided in the Supporting Information (Figures S1 and S2). X-ray crystal structures The molecular structures of 1, 2, 5, and 12 were determined by using single-crystal X-ray analysis. All crystallographic data are given in Table S1 (the Supporting Information). The struc-

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Full Paper bly as a consequence of the trans influence of NHC ligand.[3a,c] The Pt1¢N1 and Pt1¢N3 distances of 1 are 2.026(7) and 2.049(6) æ, respectively. The NHC ligand in each crystal structure adopts a configuration that is markedly twisted with respect to the Pt-trpy plane with a dihedral angle of approximately 68–72 8. As expected in the crystal packing, there is no close intermolecular p–p or Pt···Pt interaction, attributed to the steric hindrance imposed by the aliphatic chains of the NHC ligand. The structure of 12, along with selected bond lengths and angles, is also shown in Figure 1. The structural parameters are comparable to that of 10.[11a] Electrochemical properties The cyclic voltammograms of 1, 5, and 10 in acetonitrile (0.1 m n-Bu4NPF6 as a supporting electrolyte) are shown in Figure S4 (the Supporting Information). Complexes of 1, 5, and 10 show two quasi-reversible reduction couples with E1/2 (vs. Cp2Fe + /0) at ¢1.14/¢1.65, ¢1.26/¢1.77, and ¢2.05/¢2.49 V, respectively. The cathodic shift of the reduction potentials of 5 with respect to those of 1 is attributed to electron-donating tBu substituents on 5, and this shift is consistent with the assignment of the reduction as terpyridine-centered. The much more negative potential for 10 is in line with the relative electron-rich Cdeprotonated N^C^N ligand. There is no anodic wave for each complex examined in this study within the electrochemical window. Photophysical properties

Figure 1. X-ray crystal structures of 1 (top) and 12 (bottom): Perspective view with thermal ellipsoids in 30 % probability and hydrogen atoms and counterions omitted for clarity. Selected bond lengths [æ] and angles [8] for 1: Pt1¢N1 2.026(7), Pt1¢N2 1.978(6), Pt1¢N3 2.049(6), Pt1¢C16 2.002(7), N1Pt1-N2 80.8(3), N2-Pt1-N3 79.8(2), C16-Pt1-N1 99.6(3), C16-Pt1-N3 99.9(3), N2-Pt1-C16 176.8(3), N1-Pt1-N3 160.5(3). Selected bond lengths [æ] and angles [8] for 12: Pt1¢N3 2.040(5), Pt1¢N4 2.038(4), Pt1¢C1 2.102(5), Pt1¢ C26 1.928(5), N3-Pt1-C26 80.3(2), N4-Pt1-C26 79.8(2), C1-Pt1-N3 98.43(19), C1-Pt1-N4 101.47(18), N3-Pt1-N4 160.03(18), C1-Pt1-C26 178.7(2).

ture of 1 together with selected bond lengths and angles are shown in Figure 1. The structures of 2 and 5 are shown in Figure S3 (the Supporting Information), and selected bond lengths and angles are listed in Table S2 (the Supporting Information). In each case, the Pt atom adopts a distorted squareplanar coordination geometry with a trans N-Pt-N bond angle of approximately 160–161 8, characteristic of pincer-type PtII complexes containing two five-membered-ring metallacycles.[14] The Pt¢N(middle) distances of approximately 1.98– 2.02 æ are longer than that in [Pt(trpy)Cl]ClO4 ( … 1.95 æ), possiChem. Eur. J. 2015, 21, 7441 – 7453

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Photophysical data of 1–8 and 10–13 including their absorption and emission maxima, emission lifetimes (t) and emission quantum yields (f) are summarized in Table 1. As depicted in Figure 2 a, complexes 1–6 in diluted solutions display intense vibronically structured absorptions at 240–350 nm (e … 1 Õ 104 m¢1 cm¢1) and less intense absorption at 350–400 nm, which are assigned to p–p*(trpy) transitions and an admixture of dp(Pt)!p*(trpy) metal-to-ligand charge transfer (1MLCT) and p(NHC)–p*(trpy) ligand-to-ligand charge transfer (1LLCT) transitions, respectively.[3a–c, 14a] It is noted that the absorption spectra of 1–4 and 6 are similar, being insensitive to the alkyl groups of NHC ligand. The solvent effect on the absorption of 1 has been examined. As shown in Figure 2 b, the absorption energy of 1 shows no significant change upon changing the solvent polarity. Installation of an anthracene group to the NHC ligand, as in 7, results in enhanced absorption at 350– 400 nm attributed to p–p*(anthracene) transitions (Figure 3). The less intense absorption tail extending to 450 nm is tentatively assigned to mixed dp(Pt)/p(NHC) to p*(anthrancene) transitions. For 8, the very intense absorption at … 300–400 nm is attributed to intra-ligand (IL) p–p*(2,6-bis-(1-(3-propylsulfonate)benzimidazol-2’-yl)pyridine (SO3-bzimpy)) transition and the weaker absorption at l > 400 nm is assigned to mixed dp(Pt)!p*(SO3-bzimpy) and p(NHC)–p*(SO3-bzimpy) transitions (Figure 3).[15] Emission properties of 1–6 were firstly examined in MeCN for 1–5 and in H2O for 6. Complexes 1–6 are non- or very

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Full Paper Table 1. Photophysical data of complexes 1–8 and 10–13. Absorption[a] lmax [nm] (e [103 m¢1 cm¢1]) 1 2 3 4 5 6 7 8 10 11 12 13

Solution[b]

242 (31.6), 260 (sh, 22.5), 273 (23.1), 286 (sh, 11.4), 298 (10.5), 329 (10.0), 342 (11.9), 373 (sh, 1.92), 391 (br, 0.73) 242 (28.8), 260 (sh, 21.0), 273 (21.4), 287 (sh, 10.0), 298 (9.17), 332 (9.37), 342 (10.6), 373 (sh, 1.41), 389 (br, 0.41) 243 (27.0), 261 (sh, 20.0), 273 (20.3), 288 (sh, 9.32), 299 (8.57), 333 (8.78), 341 (9.59), 370 (sh, 1.47), 388 (0.40) 243 (30.8), 260 (sh, 22.1), 273 (22.4), 287 (sh, 11.1), 298 (10.1), 331 (10.4), 343 (11.5), 369 (sh, 1.93), 388 (br, 0.65) 246 (39.5), 263 (29.8), 273 (26.0), 304 (12.5), 325 (14.7), 339 (16.0), 366 (sh, 1.86), 385 (br, 0.47) 241 (30.1), 259 (sh, 20.0), 271 (20.6), 285 (sh, 11.3), 296 (10.4), 328 (8.84), 340 (12.0), 368 (sh, 1.81), 385 (br, 0.60) 273 (sh, 15.4), 297 (8.75), 334 (6.82), 345 (9.27), 388 (br, 4.90), 403 (sh, 3.99), 440 (br, 0.88) 286 (sh, 12.9), 324 (20.8), 351 (23.7), 367 (sh, 21.4), 419 (sh, 1.39), 444 (sh, 1.09) 251 (24.6), 279 (21.1), 287 (21.9), 320 (8.56), 333 (9.15), 358 (6.99), 373 (7.33), 400 (br, 1.00) 251 (21.9), 279 (19.0), 287 (19.6), 321 (7.67), 333 (8.36), 358 (6.51), 373 (6.76), 400 (br, 0.80) 251 (20.9), 278 (18.6), 286 (18.3), 318 (7.66), 333 (7.32), 358 (5.57), 373 (5.49), 400 (br, 0.70) 235 (38.8), 273 (20.0), 284 (21.0), 317 (8.22), 327 (8.37), 354 (7.37), 371 (8.30), 390 (br, 1.16)

462 (0.4), 1.9 % 463 (0.3), 1.5 % 462 (0.3), 1.4 % 462 (0.4), 1.1 % 464 (1.0), 5.3 % –[c] –[c]

495, 526; 496, 524; 495, 526; 495, 525; 495, 525;

Emission lmax [nm] (t [ms]); F Solid (298 K) Solid (77 K) 468, 496 (1.7), 524 460, 493 (8.6), 522 466, 497 (0.2), 527 463 (2.5), 497, 524 465, 492 (12.6), 529 469, 503 (0.1), 534 –[d]

543 (5.4), 582; 13 % 534 (2.4), 577, 627 (sh) 480 (2.9), 512, 541; 521 (2.1) 18 % 480 (5.6), 512, 541 525 (0.2) (sh); 11 % 480 (2.2), 512, 540 486, 512 (1.2), (sh); 8.1 % 540 (sh) 471 (0.3),501, 533 469, 502 (1.5), (sh); 4.6 % 529 (sh)

495 (8.1) 457 (8.7), 493, 521 478 (5.6), 514, 540 (sh) 462 (16.8), 497, 527 462 (15.6), 498, 533 (sh) 467, 484 (1.0), 505 –[d] 533 (5.1), 575, 617 (sh) 514, 527 (2.6), 560 495 (1.2), 519, 562 502 (3.5) 469 (6.1), 506, 531 (sh)

Glassy solution[e] 461 (20.4), 496, 525 460 (22.2), 496, 522 461 (21.1), 496, 525 460 (22.3), 495, 525 453 (20.3), 487, 526 463 (21.3), 497, 537 462, 482 (15.0), 520, 560 528 (12.7), 569, 615 477 (16.9), 543 475 (15.7), 512, 543 475 (16.9), 508, 541 468 (19.0), 505, 535

[a] Determined in MeCN for 1–5 and 10–13 and in H2O for 6–8 ( … 2 Õ 10¢5 m); sh denotes shoulder and br denotes broad. [b] Determined in CH2Cl2 for 1– 5, in H2O for 8, and in MeCN for 10–13. [c] Not measured in CH2Cl2 due to solubility reasons. [d] No measurable emission. [e] Determined at 77 K in butyronitrile for 1–5 and 10–13, in EtOH/MeOH/DMF (5:5:1, v/v) for 6 and 7, and in EtOH/MeOH (4:1, v/v) for 8.

weakly emissive (f< 0.1 %) in these solutions. Examination of the solvent effect on the emission properties of 1 shows that 1 is emissive in CH2Cl2 (lmax = 462 nm, f= 1.9 %) but non-emissive in DMF, MeOH, and THF. This can be ascribed to the more pronounced non-radiative decay of 1 in highly polar solvents. The emission quantum yields and lifetimes of 1–5 in CH2Cl2 are in the range of 1.1–5.3 % and 0.3–1.0 ms, respectively. The vibronic spacings of 1.2–1.4 Õ 103 cm¢1 for 1–5 are indicative of the 3p–p*(terpyridine) nature of the emitting states. Similar to that observed for the absorption bands of 1–4, the emission energies of this series of complexes are not affected by the alkyl groups of NHC ligand (the Supporting Information, Figure S5). Emission properties of 1–6 in solid state and glassy solutions have been studied (Table 1 and the Supporting Information, Figure S6–S11). Complexes 1–6 show moderate-to-intense green emissions in the solid state at room temperature. There is no distinct red-shift of emission energy for these complexes in the solid state compared with those in solutions and this is consistent with the lack of significant Pt···Pt or p–p interactions as revealed by the crystal structures of 1, 2, and 5. Upon cooling to 77 K, complexes 1–6 exhibit increased emission intensity and lifetimes (Table 1). At 77 K, complexes 1–5 in frozen butyronitrile and 6 in frozen EtOH/MeOH/DMF (5:5:1, v/v) all exhibit strong green phosphorescence with lifetimes of 20.3– 22.3 ms. The well-resolved vibronic-structured emissions are indicative of significant intra-ligand (IL) (trpy) character of the emitting states. Chem. Eur. J. 2015, 21, 7441 – 7453

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Even with the presence of the anthracene moiety, complex 7 is non-emissive in degassed water at room temperature, which is attributed to intramolecular energy-transfer process. A strong phosphorescence at 482 nm is observed for 7 in 77 K EtOH/MeOH/DMF (5:5:1, v/v) glassy solution, which, compared to the emission data of 1–6, is attributable to the Pt-trpy moiety (Table 1 and Figure S12). Complex 8 exhibits an intense orange emission (lmax = 543 nm) with a quantum yield of 13 % in degassed aqueous solution (Figure 3). The vibronic-structured emission band together with a long lifetime of … 5.4 ms is consistent with an emitting state of 3IL p–p*(SO3-bzimpy) in nature.[15] Intense and slightly blue-shifted emissions are observed for 8 in the solid state (298 and 77 K) and in glassy EtOH/MeOH (4:1, v/v) solution (the Supporting Information, Figure S13). Complexes 10–13 display intense vibronically structured absorption bands at 350–380 nm arising from 1IL (N^ C^N) transitions. The weak absorption tailing to 420 nm can be tentatively assigned to dp(Pt)!p* (N^C^N) 1MLCT transition (the Supporting Information, Figure S14). A comparison between the absorption spectra of 10 and 1 is shown in Figure 3. The IL absorption of 10 is red-shifted from that of 1 by about 35 nm, attributed to destabilization of dp orbital by C-deprotonated carbon donor atom. In degassed MeCN, complexes 10–13 show vibronically structured emission with an emission maximum at 470–480 nm (f= 4.6–18 %, t = 0.3–5.6 ms). The vibronic spacings (1.3 Õ 103 cm¢1) and long emission lifetimes are consis-

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Figure 2. UV/Vis absorption spectra of (a) complexes 1–5 in MeCN and 6 in H2O and (b) complex 1 in various solvents (concentration … 2 Õ 10¢5 m).

tent with the emitting states to be triplet metal-perturbed 3IL in nature. The blue-shift of the emission maximum of 13 by 9 nm from that of 10 is attributed to the electron-withdrawing effect of the CF3 group. The effect of solvent polarity on the emission properties was also examined and the emission spectra of 10–13 in various solvents are depicted in Figures S15– S18 (the Supporting Information), showing negligible solvatochromic behavior. Upon decreasing the solvent polarity, their emission quantum yields increase to 34–65 % in CHCl3, which are comparable to that of [Pt(N^C^N)L] (L = Cl¢ or ¢CŽCPh) complexes in CH2Cl2.[14b, c] Complexes 10–13 in 77 K glassy butyronitrile solutions display intense, slightly blue-shifted (< 5 nm) green emission with lifetimes of … 15–20 ms. Their vibronic-structured emission bands and long emission lifetimes are indicative of N^C^N-based 3IL excited states. In the solid state at 298 K, complexes 10–13 are emissive with lmax at 469– 525 nm. Upon cooling to 77 K, the emissions of 10–13 in the solid state are observed at energies similar to those recorded at ambient temperature.

Figure 3. (a) UV/Vis absorption spectra of 1 (in MeCN), 7 (in H2O), 8 (in H2O), and 10 (in MeCN); (b) emission spectra of 1 (in CH2Cl2), 8 (in H2O), and 10 (in MeCN); concentration … 2 Õ 10¢5 m.

DFT and TDDFT calculations To gain insight into the electronically excited state properties of [Pt(trpy)(NHC)]n + complexes, density functional theory (DFT) and time-dependent density functional theory (TDDFT) calculaChem. Eur. J. 2015, 21, 7441 – 7453

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Figure 4. DFT calculated frontier orbitals of 1 at optimized S0 geometry.

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Full Paper tions on 1 have been performed (Figure 4). The optimized S0 structure of 1 is in good agreement with the crystal structure (the Supporting Information, Figure S19). The calculated bond lengths and angles are within 0.02 æ and 1 8 difference from the values determined by X-ray analysis. According to the results of TDDFT calculation (the Supporting Information, Table S3), the lowest energy absorption (S1 S0) of 1 is derived primarily ( … 99 % contribution) from the HOMO!LUMO transition. As depicted in Figure 4, the electron density in HOMO is localized on the p(NHC) and dp(Pt) orbitals and that in LUMO predominantly on the p*(trpy) orbital. Thus, the S1 state can be described to have mainly LLCT/MLCT character. The calculated S1 S0, S2 S0 and S3 S0 transitions are at … 400, … 353, and … 348 nm for 1, which match well with the experimental low-energy absorption bands in the 320–400 nm region.[16] !

!

!

!

Anion-binding properties In view of the dicationic nature of [Pt(trpy)(NHC)]2 + complexes, we examined the treatment of 1 with a wide variety of anions including F¢ , Cl¢ , Br¢ , I¢ , SO42¢, NO3¢ , HSO4¢ , H2PO4¢ , AcO¢ , CN¢ , and PF6¢ . As depicted in Figure S20 (the Supporting Information), in acetonitrile, an excess amount of F¢ , AcO¢ (100fold) or SO42¢ (50-fold) induces notable absorption spectral changes of 1. The absorption of 1 at … 310–400 nm disappears accompanied by an advent of absorption at … 280 nm in the presence of CN¢ (the Supporting Information, Figure S21). As there is no receptor group such as hydrogen-bonding donor present in 1, these anions are conceived to bind to PtII at the axial coordination site. The disappearance of the low-energy

absorption of 1 in the presence of CN¢ is attributed to the formation of [Pt(n-Bu2Im)(CN)3]¢ (9) through displacement of terpyridine by CN¢ (Figure 5 a). To testify this speculation, complex 9 was independently prepared and characterized. As depicted in Figure 5 b, the 1H NMR spectrum of the reaction mixture revealed the presence of 9 and free terpyridine. ESI-MS analysis of a solution of 1 in the presence of excess CN¢ also showed the expected m/z peak at 453.2 attributed to 9 (Figure 5 c). For comparison, we also examined the reaction of 10 with CN¢ in MeCN by UV/Vis absorption and 1H NMR spectroscopy. Figure S22 (the Supporting Information) shows the UV/ Vis absorption spectral change of 10 in the presence of CN¢ . A similar decrease of the absorbance at wavelength > 325 nm was observed. This absorption spectral change is attributed to dissociation of the two Pt¢N bonds in 10. The findings of 1 H NMR spectroscopy (Figure 6 b) and ESI-MS experiments (the Supporting Information, Figure S22) are indicative of the formation of [Pt(NCN)(NHC)(CN)2]¢ (14). The relatively strong Pt¢ C(NHC) bond remained intact in the presence of excess CN¢ . This ligand displacement reaction by CN¢ binding to PtII has resulted in complete quenching of the emission (Figure 6 c). As the nucleophilicity of anion is solvent dependent, we then examined the reaction of 1 with various anions in MeOH. The findings revealed that in MeOH (the Supporting Information, Figure S23), CN¢ can be distinguished from the other anions F¢ , AcO¢ , and H2PO4¢ .[17] The selectivity of 1 towards CN¢ was then examined in H2O/MeOH (9:1, v/v) solution. As depicted in Figure S24 (the Supporting Information), addition of 20 equivalents of the other anions except for CN¢ did not induce significant spectral changes of 1. Thus, [Pt(N^N^N)(NHC)]n + can be

Figure 5. (a) Schematic illustration of the reaction of 1 with CN¢ and characterization of the reaction mixture by using the (b) 1H NMR and (c) ESI-MS spectra. The reaction mixture contains about 3.5 equiv [n-Bu4N]CN. The 1H NMR spectra of 1, free terpyridine, and independently synthesized 9 are shown in (b) for comparison. The 1H NMR and ESI-MS spectra were recorded in CD3CN and MeCN, respectively. Chem. Eur. J. 2015, 21, 7441 – 7453

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Figure 6. (a) Schematic illustration of the reaction of 10 with CN¢ and the characterization of the reaction mixture by (b) 1H NMR spectra. The reaction mixture contains about 7 equiv [n-Bu4N]CN. The 1H NMR spectrum of 10 is shown in (b) for comparison. The 1H NMR spectra were recorded in CD3CN; (c) Photographs of 10 in MeCN in the absence and presence of CN¢ under daylight and UV-lamp (365 nm).

used as an irreversible chemodosimeter for detection of CN¢ in aqueous solution with high selectivity. Selective and sensitive detection of anions is an important area because anions play crucial roles in physiological and industrial processes.[18] Cyanide is known to display high toxicity to human health. As human has a ultra-low tolerance limit of CN¢ (< 0.2 ppm), selective detection of CN¢ with high sensitivity in aqueous media is of importance.[19] One common tactic for recognizing CN¢ , as well as other anions, relies on their bindings with functional systems through non-covalent interactions, such as hydrogen bonding.[20] Reaction-based cyanide sensors, that is, chemodosimeters, have also been reported for the selective detection of cyanide ions. For example, this approach has been realized by using organic dyes that can readily undergo reactions with CN¢ .[21] The strong affinity of CN¢ towards metal ions has also been utilized to induce striking color and emission changes through ligand displacement.[22] Inspired by the release of terpyridine ligand from the treatment of 1 with CN¢ in protic solvent (MeOH and H2O), we sought to tune the absorption and emission properties of the [Pt(N^N^N)(NHC)]n + system so that the sensing process can be visualized by changes in absorption (color) and emission in the UV/Visible spectral region. Complex 7 having an anthracene moiety grafted to NHC ligand was designed and prepared. As aforementioned, this complex does not show anthracenebased emission due to quenching by intramolecular energytransfer process. Upon displacement of the terpyridine ligand in 7 with the addition of three equivalents of CN¢ , the fluorescence of the anthracene moiety is switched on. Kinetic information of the treatment of 7 with CN¢ in water was obtained by monitoring the absorption spectral changes of 7 ( … 2 Õ 10¢5 m) in the presence of NaCN ( … 1 Õ 10¢4 m). The spectral change was completed within 10 min (the Supporting Information, Figure S25). Therefore, a reaction time of 10 min was set Chem. Eur. J. 2015, 21, 7441 – 7453

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for the absorption and emission titration studies and the results are shown in Figure 7. The disappearance of the absorption at l > 400 nm produced a visible color change of a solution of 7 from yellow to colorless. The attack of CN¢ to the PtII site switches on the anthracene fluorescence. The detection limit for CN¢ is evaluated to be < 1 ppm from the linear plot of emission intensity at lem = 414 nm versus [CN¢] (the Supporting Information, Figure S26). The specificity of 7 towards CN¢ detection has been examined. As depicted in Figure 7 c and Figure S26 (the Supporting Information), CN¢ can induce changes of absorption color and emission energy with high selectivity over the other anions. Treatment of 7 with CN¢ has also been examined by using 1H NMR spectroscopy in DMSO solution. The 1H NMR spectrum of the reaction mixture, as depicted in Figure S27 (the Supporting Information), revealed the presence of the free terpyridine ligand. A test paper for rapid detection of CN¢ was prepared by dropping an aqueous solution of 7 to a filter paper followed by drying. As depicted in Figure 8, the test paper shows strong blue luminescence only when CN¢ is present. Moreover, the interference of other anions for CN¢ testing was examined. The CN¢ could still react with 7 to give a blue emission in the presence of these anions. This result shows that 7 can be used for recognizing CN¢ in an easy-to-prepare method without resorting to a specific analysis instrument. The response of the water-soluble complex 8 towards CN¢ was also examined for comparison. As shown in Figure 9, upon addition of excess CN¢ , the strong orange phosphorescence of 8 is quenched accompanied by concomitant generation of deep-blue emission from free SO3-bzimpy ligand. ESI-MS spectrum of the reaction mixture of 8 with CN¢ showed the formation of 9 and the liberation of SO3-bzimpy ligand (the Supporting Information, Figure S28). The treatment of CN¢ with 8 was also monitored by absorption spectroscopy and, as shown in

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Figure 7. (a) Absorption and (b) emission spectral traces of 7 in H2O ( … 2 Õ 10¢5 m) upon addition of CN¢ (0 to 2.4 Õ 10¢4 m); (c) photographs of 7 in H2O in the absence and presence of various anions (40 equiv each) under 365 nm irradiation.

Figure 8. Photograph of a test paper coated with 7 and its responses to various anions in the absence (top) and presence (bottom) of CN¢ under a UVlamp (365 nm).

Figure S28 (the Supporting Information), there was only a slight change in the absorption spectrum of 8 one hour after the addition of 5 equivalents of CN¢ revealing a slower reaction when compared with that of CN¢ with 7. In vitro cytotoxicity

Figure 9. Emission spectra of 8 in H2O ( … 2 Õ 10¢5 m) in the absence and presence of NaCN ( … 2 Õ 10¢3 m) (emission spectrum of free SO3-bzimpy ligand in H2O is shown for comparison). Inset: photographs of 8 in H2O in the absence and presence of CN¢ under 365 nm irradiation.

In previous work, the [Pt(C^N^N)(NHC)] + complex with lipophilic alkyl chains on the carbene ligand was shown to be highly cytotoxic and specific towards HeLa cells.[6c] In the present study, we examined the in vitro anticancer properties of the PtII complexes towards HeLa cells by means of MTT (3-(4,5dimethylthiazol-2-yl)-2,5-tetrazolium bromide) assays. Figure 10 depicts the percentage cell survival of the HeLa cells (with an incubation period of 72 h) versus the concentrations of [Pt(N^C^N)(NHC)] + complexes 10–13, with cisplatin as reference. From the cytotoxicity profiles, the IC50 values of 10–13

were determined to be 0.46–2.45 mm, which are up to 22-fold more potent than cisplatin (IC50 = 10 mm). The trend in cytotoxicity follows 12 (IC50 = 0.46 mm) > 13 (IC50 = 1.70 mm) > 10 (IC50 = 2.45 mm) > 11 (IC50 = 3.00 mm), which could be correlated to the lipophilicity of the PtII complexes caused by alkyl groups on the NHC ligand. The correlation between in vitro anticancer activity and lipophilicity has been observed in the cases of anticancer AuIII porphyrin complexes and other pincer PtII-NHC complexes.[6c, 23] In contrast, complexes 1–4 and 6–8 are much

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Figure 10. Plot of cell viability [%] versus concentration after treatment with platinum(II) complex for 72 h.

less cytotoxic with IC50 values of > 55 mm (the Supporting Information, Table S4) except for 4, which contains two long hexadecyl chains (IC50 = 16.4 mm). The intense emission of the [Pt(N^C^N)(NHC)] + complexes allows direct monitoring of cellular uptake by fluorescence microscopy. After treatment of HeLa cells with 2 mm of 12 for only 10 min, bright-green emission could be detected in cytoplasm, indicative of efficient cellular uptake (Figure 11). A simi-

Figure 12. Effects of 12 on mitochondrial dysfunction: (a) HeLa cells treated with or without 12 were then stained with Mitotracker, and images were taken by using fluorescent microscopy (lex = 546 nm, lem > 580 nm); (b) HeLa cells treated with or without 12 were then stained with JC-1, and images were taken by using fluorescent microscopy (lex = 470 nm, lem > 515 nm).

stead, the cells treated with 2 mm of 12 for 1 h showed green fluorescence that could be attributed to monomeric form of JC-1, indicative of mitochondria depolarization. Thus, complex 12 efficiently accumulates in cytoplasm of cancer cells and induces mitochondrial dysfunction leading to cell death.

Conclusion

Figure 11. Fluorescence microscopic analysis of HeLa cells treated with 2 mm of 12 after 10 min. Complex 12 was excited at 340 nm using an emission filter of 510 nm (blue pseudo-color).

lar intracellular localization was observed for 10-treated HeLa cells that were co-stained with the nuclei specific dye Hoechst 33342 (the Supporting Information, Figure S29). Interestingly, the cytoplasmic location (not in the nucleus) of 10 and 12 is reminiscent of the case for our previously reported [Pt(C^N^N)(NHC)] + complex (Scheme 1), which is mainly localized in mitochondria.[6c] However, attempts to co-localize the platinum(II) complex with mitochondria specific MitoTracker did not reveal its accumulation in mitochondria. Instead, significant mitochondria disruption/swelling was identified (Figure 12 a). Since mitochondria is a well-known therapeutic target of cancer cells,[24] we then tested whether the complexes could induce mitochondria dysfunction by measuring the mitochondria membrane potential using the JC-1 marker, a cell permeable dye that could selectively accumulate in normal mitochondria inner membrane and form J-aggregates with orange emission.[12] As shown in Figure 12 b, the untreated HeLa cells with JC-1 staining exhibited orange fluorescence that is typical of JC-1 aggregate formation in mitochondria. InChem. Eur. J. 2015, 21, 7441 – 7453

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Two classes of pincer-type PtII complexes containing NHC ligands have been synthesized and characterized. The [Pt(trpy)(NHC)]2 + complexes 1–5 show IL (trpy) 3p–p* phosphorescence in CH2Cl2. In protic solvents, complex 1 reacts with CN¢ with high selectivity over other anions. A luminescence switch-on chemodosimeter 7 for CN¢ detection in water with high sensitivity and selectivity has been demonstrated. The [Pt(N^C^N)(NHC)] + complexes 10–13 display higher in vitro cytotoxicity than cisplatin towards HeLa cells. Fluorescence microscopic analysis revealed that 12 could readily enter into cancer cells and accumulate in cytoplasm, resulting in mitochondrial dysfunction and cancer cell death. This work shows that NHC ligand can be used as a handle for tuning of the physical and chemical properties of luminescent platinum(II) complexes to provide an entry to new functional materials with applications in biomedical science.

Experimental Section Materials and reagents All materials used were received from commercial sources unless stated otherwise. 1,3-bis-n-butylimidazolium bromide (nBu2ImBr),[25] 1,3-bis-cyclohexylimidazolium chloride (Cy2ImCl),[26] [nBu2ImAgBr],[13a] sulfonate pendant imidazolium salt Im(SO3)2Na,[27] 2,6-bis-(benzimidazol-2’-yl)pyridine (bzimpy),[28] 2,6-bis-(1-(3-propylsulfonate)benzimidazol-2’-yl)pyridine (SO3-bzimpy),[15b] [Pt(trpy)Cl]Cl,[3a] [Pt(tBu3trpy)Cl]Cl,[14a] and K[Pt(SO3-bzimpy)Cl][15b] were pre-

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Full Paper pared according to literature procedures. Synthesis of 1,3-bis-cetylimidazolium bromide (Cetyl2ImBr) and the synthetic schemes and experimental details for complexes 6–8 are provided in the Supporting Information. The syntheses of complexes 10 and 13 have been described elsewhere[11a] and complexes 11 and 12 were similarly prepared except that iPr2ImCl or Cy2ImCl was used instead of n-Bu2ImBr, respectively. For the synthesis of complexes 1--5, [Pt(trpy)(MeCN)](OTf)2 or [Pt(tBu3trpy)(MeCN)](OTf)2 were obtained by [Pt(trpy)Cl]Cl or [Pt(tBu3trpy)Cl]Cl in MeCN heated at reflux in the presence of AgOTf (2 equiv) in dark for 60 h. After cooling to room temperature, the mixture was filtered through Celite. The filtrate was evaporated to dryness by using a rotary evaporator and used in next step without further purification. [Pt(trpy)(n-Bu2Im)](PF6)2 (1): A mixture of n-Bu2ImBr (50 mg, 0.19 mmol) and tBuOK (47 mg, 0.42 mmol) in MeOH (15 mL) was heated at reflux for 1 h, followed by addition of [Pt(trpy)(MeCN)](OTf)2 (0.096 mmol). The mixture was heated at reflux for a further 24 h, then cooled to room temperature, and then stirred with NH4PF6 (160 mg, 1 mmol). After removal of the solvent by using a rotary evaporator and washing with water and chloroform, the residue was dissolved in acetonitrile. Yellow crystals were obtained by slow diffusion of diethyl ether vapor into the acetonitrile solution (35 mg, 41 %). 1H NMR (400 MHz, CD3CN): d = 8.54 (t, J = 8.1 Hz, 1 H), 8.41–8.35 (m, 6 H), 7.97 (d, J = 5.5 Hz), 7.65 (m, 2 H), 7.49 (s, 2 H), 4.35 (t, J = 7.2 Hz, 4 H), 1.83 (m, 4 H), 1.25 (m, 4 H), 0.78 ppm (t, J = 7.4 Hz, 6 H); 31P NMR (162 MHz, CD3CN): d = ¢144.5 ppm (sept, J = 706.3 Hz); 19F NMR (376 MHz, CD3CN): d = ¢72.9 ppm (d, J = 706.5 Hz); FAB-MS: m/z: 753.2 [M¢PF6] + ; elemental analyses calcd (%) for C26H31N5F12P2Pt: C 34.75, H 3.48, N 7.79; found: C 34.83, H 3.61, N 7.94. [Pt(trpy)(iPr2Im)](PF6)2 (2): The procedure is similar to that for 1, except that iPr2ImCl was used instead of n-Bu2ImBr (yield: 36 %). 1 H NMR (400 MHz, CD3CN): d = 8.54 (t, J = 8.2 Hz, 1 H), 8.41–8.37 (m, 6 H), 7.97 (d, J = 5.1 Hz, 2 H), 7.65 (m, 2 H), 7.60 (s, 2 H), 5.34 (m, 2 H), 1.46 ppm (d, J = 6.7 Hz, 12 H); 31P NMR (162 MHz, CD3CN): d = ¢148.9 ppm (sept, J = 706.3 Hz); 19F NMR (376 MHz, CD3CN): d = ¢72.9 ppm (d, J = 706.5 Hz); FAB-MS: m/z: 725.2 [M¢PF6] + ; elemental analyses calcd (%) for C24H27N5F12P2Pt: C 33.11, H 3.13, N 8.05; found: C 32.94, H 3.08, N 7.97. [Pt(trpy)(Cy2Im)](PF6)2 (3): The procedure is similar to that for 1, except that Cy2ImCl was used instead of n-Bu2ImBr (yield: 39 %). 1 H NMR (400 MHz, CD3CN): d = 8.56 (t, J = 8.2 Hz, 1 H), 8.41–8.37 (m, 6 H), 7.95 (d, J = 5.5 Hz, 2 H), 7.63 (m, 2 H), 7.56 (s, 2 H), 4.94 (m, 2 H), 1.77–1.10 ppm (20 H); 31P NMR (162 MHz, CD3CN): d = ¢144.5 ppm (sept, J = 706.3 Hz); 19F NMR (376 MHz, CD3CN): d = ¢72.9 ppm (d, J = 706.5 Hz); FAB-MS: m/z: 805.1 [M¢PF6] + ; elemental analyses calcd (%) for C30H35N5F12P2Pt: C 37.90, H 3.71, N 7.37; found: C 37.87, H 3.59, N 7.34. [Pt(trpy)(Cetyl2Im)](PF6)2 (4): The procedure is similar to that for 1, except that Cetyl2ImBr was used instead of n-Bu2ImBr. After stirring with NH4PF6 and evaporation of the solvent by using a rotary evaporator, the residue was washed with water and then dissolved in chloroform. Yellow powder was obtained by slow diffusion of diethyl ether vapor into a solution of chloroform (yield: 60 %). 1 H NMR (400 MHz, CD3CN): d = 8.55 (t, J = 8.2 Hz, 1 H), 8.40 (m, 6 H), 7.99 (d, J = 5.4 Hz, 2 H), 7.64 (d, J = 4.6 Hz, 2 H), 7.49 (s, 2 H), 4.34 (t, J = 6.9 Hz, 4 H), 1.84 (m, 4 H), 1.26–1.03 (m, 52 H), 0.86 ppm (t, J = 6.1 Hz, 6 H); 31P NMR (162 MHz, CD3CN): d = ¢144.5 ppm (sept, J = 706.3 Hz); 19F NMR (376 MHz, CD3CN): d = ¢72.9 ppm (d, J = 706.5 Hz); FAB-MS: m/z: 1089.6 [M¢PF6] + ; elemental analyses calcd (%) for C50H79N5F12P2Pt: C 48.62, H 6.45, N 5.67; found: C 48.87, H 6.04, N 5.74. Chem. Eur. J. 2015, 21, 7441 – 7453

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[Pt(tBu3trpy)(n-Bu2Im)](PF6)2 (5): The procedure is similar to that for 1, except that [Pt(tBu3trpy)(MeCN)](OTf)2 was used instead of [Pt(trpy)(MeCN)](OTf)2. After stirring with NH4PF6 and evaporation of the solvent by using a rotary evaporator, the residue was washed with water. The residue was then dissolved in chloroform. Greenish-yellow crystals were obtained by slow diffusion of diethyl ether vapor into the chloroform solution (36 mg, 38 %); 1H NMR (400 MHz, CD3CN): d = 8.43 (s, 2 H), 8.37 (d, J = 2.1 Hz, 2 H), 7.82 (d, J = 6.0 Hz, 2 H), 7.57 (dd, J1 = 6.1 Hz, J2 = 2.2 Hz, 2 H), 7.48 (s, 2 H), 4.32 (t, J = 7.4 Hz, 4 H), 1.83 (m, 4 H), 1.57 (s, 9 H), 1.44 (s, 18 H), 1.11 (m, 4 H), 0.78 ppm (t, J = 7.4 Hz, 6 H); 31P NMR (162 MHz, CD3CN): d = ¢144.5 ppm (sept, J = 706.5 Hz); 19F NMR (376 MHz, CD3CN): d = ¢72.9 ppm (d, J = 706.5 Hz); FAB-MS: m/z: 921.5 [M¢PF6] + , 388.2 [M¢2PF6]2 + ; elemental analyses calcd (%) for C38H55N5F12P2Pt: C 42.78, H 5.20, N 6.56; found: C 42.52, H 4.73, N 6.55. Complex 6: A mixture of sulfonate pendant imidazolium salt Im(SO3)2Na (70 mg, 0.23 mmol), silver oxide (59 mg, 0.25 mmol), and NaCl (15 mg, 0.26 mmol) in DMSO (3 mL) was stirred at 60 8C for 20 h. After cooling to room temperature, the mixture was filtered and the solution was used for next step. The in situ-prepared solution was mixed with [Pt(trpy)Cl]Cl (51 mg, 0.10 mmol). The mixture was then heated to 90 8C in the presence of AgPF6 (103 mg, 0.41 mmol) for 20 h. The reaction mixture was then cooled to room temperature and filtered. The residue was washed with methanol, redissolved in water and filtered. Slow evaporation of the water upon heating yielded a yellow powder (23 mg, 32 %). 1 H NMR (400 MHz, D2O): d = 8.52 (t, J = 8.0 Hz, 1 H), 8.44–8.36 (m, 6 H), 8.11 (d, J = 5.5 Hz, 2 H), 7.67 (m, 2 H), 7.63 (s, 2 H), 4.76 (t, J = 7.1 Hz, 4 H), 3.48 ppm (t, J = 6.7 Hz, 4 H); FAB-MS: m/z: 710.6 [M] + ; elemental analyses (%) calcd for C22H21N5O6PtS2·H2O: C 36.26, H 3.18, N 9.61; found: C 36.07, H 3.16, N 9.47. Complex 7: A mixture of An-Im-SO3 (73 mg, 0.20 mmol), silver oxide (57 mg, 0.25 mmol), and NaCl (13 mg, 0.22 mmol) in DMSO (3 mL) was stirred at 60 8C for 20 h. After cooling to room temperature, the mixture was filtered and the solution was used for next step. The in situ-prepared solution was mixed with [Pt(trpy)Cl]Cl (52 mg, 0.10 mmol). The mixture was then heated to 90 8C in the presence of AgPF6 (90 mg, 0.36 mmol) for 20 h. The reaction mixture was then cooled to room temperature and filtered to remove the solid. DMSO was removed from the filtrate under reduced pressure. The residue was washed with MeOH and then dissolved in water. Slow evaporation of the water upon heating yielded orange crystalline needles (45 mg, 46 %). 1H NMR (400 MHz, [D6]DMSO): d = 8.77 (s, 1 H), 8.67 (d, J = 5.9 Hz, 2 H), 8.47–8.38 (m, 8 H), 8.28 (d, J = 1.8 Hz, 1 H), 8.07 (d, J = 7.9 Hz, 2 H), 7.81 (d, J = 8.4 Hz, 2 H), 7.76 (t, J = 6.5 Hz, 2 H), 7.48–7.40 (m, 4 H), 4.76 (t, J = 7.1 Hz, 2 H), 2.63 (t, J = 6.9 Hz, 2 H), 2.32–2.39 ppm (m, 2 H); FABMS: m/z: 793.6 [M¢PF6] + ; elemental analyses calcd (%) for C35H28N5F6O3PSPt·3 H2O: C 42.34, H 3.45, N 7.05; found: C 42.33, H 3.48, N 6.98. Complex 8: AgPF6 (27 mg, 0.11 mmol) was added to a solution of K[Pt(SO3-bzimpy)Cl] (60 mg, 0.073 mmol) and [n-Bu2ImAgBr] (53 mg, 0.143 mmol) in DMSO (10 mL). The resulting yellow suspension was stirred at 85 8C for 18 h. The mixture was then cooled to room temperature and filtered. DMSO was removed from the filtrate under reduced pressure. The residue was redissolved in MeOH and filtered. The filtrate was then concentrated and a yellow powder was obtained by slow diffusion of diethyl ether vapor into the MeOH solution (28 mg, 41 %); 1H NMR (400 MHz, [D6]DMSO): d = 9.03 (d, J = 8.4 Hz, 2 H), 8.59 (t, J = 8.3 Hz, 1 H), 8.20 (d, J = 8.5 Hz, 2 H), 8.04 (s, 2 H), 7.63 (t, J = 7.8 Hz, 2 H), 7.47 (d, J = 7.7 Hz, 2 H), 5.91 (d, J = 8.3 Hz, 2 H), 5.15 (t, J = 7.5 Hz, 4 H), 4.33 (t, J = 6.8 Hz, 4 H), 2.70 (m, 4 H), 2.29 (m, 4 H), 1.67 (m, 4 H), 1.14 (m,

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Full Paper 4 H), 0.54 ppm (t, J = 7.3 Hz, 6 H); FAB-MS: m/z: 929.6 [M+ +H] + ; elemental analyses calcd (%) for C36H43N7O6S2Pt·3 H2O: C 43.98, H 5.02, N 9.97; found: C 44.12; H 4.84, N 9.99. Complex 9: A solution of [Pt(dmso)2Cl2] and n-Bu2ImBr (1 equiv) in DMSO was heated to 90 8C for 17 h in the presence of sodium hydrogen carbonate (2.5 equiv). After completion of the reaction, the mixture was filtered through Celite and the solvent was removed under vacuum. The residue was washed with H2O and H2O/acetone, affording a white powder. To an acetone solution (3 mL) of the white powder (44 mg, 0.085 mmol) was added tetrabutylammonium cyanide (71 mg, 0.26 mmol) and the mixture was stirred at room temperature overnight. The solution was then removed by using a rotary evaporator, and the residue was washed with water and diethyl ether, affording the product as a white powder (two-step overall yield: 50 %). 1H NMR (400 MHz, CD3CN): d = 7.10 (s, 2 H), 4.15 (t, J = 7.2 Hz, 4 H), 3.07 (t, J = 8.1 Hz, 8 H), 1.86 (m, 4 H), 1.59 (m, 8 H), 1.35 (m, 12 H), 0.96 ppm (m, 18 H); elemental analyses calcd (%) for C30H56N6Pt: C 51.78, H 8.11, N 12.08; found: C 51.66, H 8.18, N 12.07. Complex 11: Yellow crystals were obtained by slow diffusion of diethyl ether vapor into a chloroform solution (yield: 80 %). 1H NMR (400 MHz, CDCl3): d = 8.06 (t, J = 7.6 Hz, 2 H), 7.97 (d, J = 5.2 Hz, 2 H), 7.84 (d, J = 7.6 Hz, 2 H), 7.62 (d, J = 7.6 Hz, 2 H), 7.37 (s, 2 H), 7.34 (t, J = 7.6 Hz, 1 H), 7.30 (m, 2 H), 5.26 (m, 2 H), 1.47 ppm (d, J = 6.8 Hz, 12 H); 31P NMR (162 MHz, CDCl3): d = ¢144.3 ppm (sept, J = 712.7 Hz); 19F NMR (376 MHz, CDCl3): d = ¢73.5 ppm (d, J = 712.6 Hz); FAB-MS: m/z: 578.2 [M¢PF6] + ; elemental analyses calcd (%) for C25H27F6N4PPt·0.5 CHCl3 : C 39.10, H 3.54, N 7.15; found: C 39.05, H 3.39, N 7.02. Complex 12: Yellow crystals were obtained by slow diffusion of diethyl ether vapor into a chloroform solution (yield: 76 %). 1H NMR (400 MHz, CDCl3): d = 8.06 (t, J = 7.6 Hz, 2 H), 7.95 (d, J = 5.6 Hz, 2 H), 7.85 (d, J = 8.0 Hz, 2 H), 7.63 (d, J = 8.0 Hz, 2 H), 7.38 (t, J = 7.6 Hz, 1 H), 7.35 (s, 2 H), 7.30–7.27 (m, 2 H), 4.80 (m, 2 H), 2.00–1.19 ppm (20 H); 31P NMR (162 MHz, CDCl3): d = ¢144.2 ppm (sept, J = 712.7 Hz); 19F NMR (376 MHz, CDCl3): d = ¢73.5 ppm (d, J = 712.6 Hz); FAB-MS: m/z: 658.2 [M¢PF6] + ; elemental analyses calcd (%) for C31H35F6N4PPt·0.5 Et2O·0.5 H2O: C 46.63, H 4.87, N 6.60; found: C 46.49, H 5.22, N 6.70.

respectively, n is the refractive index of the solvents, D is the integrated intensity, and f is the luminescence quantum yield.[29] The quantity B is calculated by B = 1–10¢AL, in which A is the absorbance at the excitation wavelength and L is the optical path length. Errors for wavelength values (1 nm) and f (10 %) are estimated. Cyclic voltammetric measurements were performed with a Princeton Applied Research electrochemical analyzer (potentiostat/galvanostat Model 273A). n-Bu4NPF6 (TBAH; 0.1 m) in MeCN was used as a supporting electrolyte for the electrochemical measurements at room temperature. All solutions used in electrochemical measurements were deaerated with nitrogen gas. Ag/AgNO3 (0.1 m in MeCN), a glassy carbon electrode, and a platinum wire were used as reference electrode, working electrode, and counter electrode, respectively. Scan rates were 100 mV s¢1 and the potentials were reported with respect to the potential of a ferrocenium/ferrocene (Cp2Fe + /0) redox couple. For MTT assays, the absorbance was quantified using PerkinElmer Fusion Reader (Packard BioScience Company). Fluorescence images were examined in Axiovert 200 (Carl Zeiss) and in an Axio Vision Rel. 4.5 imaging system (Carl Zeiss).

X-ray diffraction measurement Single crystals of complexes 1, 2, 5, and 12 were grown by slow diffusion of Et2O into MeCN solutions of the complexes. The X-ray diffraction data were collected on a Bruker X8 PROTEUM singlecrystal X-ray diffractometer. Raw frame data were integrated by using the SAINT program. Multiscan SADABS was applied for absorption correction. The structures were solved by direct methods employing the SHELXS-97[30] program and refined by full-matrix least-squares using the program SHELXL-97.[31] The positions of the hydrogen atoms were calculated on the basis of the riding mode with thermal parameters equal to 1.2 times that of the associated C atoms, and these positions participated in the calculation of the final R indices. In the final stage of least-squares refinement, all non-hydrogen atoms were refined anisotropically. CCDC-966967 (1), CCDC-966968 (2), CCDC-966969 (5), and CCDC1049548 (12) 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/ data_request/cif.

Physical measurements and instrumentation The solvents used for photophysical measurements were of HPLC grade. Elemental analyses were performed by the Institute of Chemistry at the Chinese Academy of Sciences, Beijing. Fast atom bombardment (FAB) mass spectra were obtained on a Finnigan Mat 95 mass spectrometer. Electrospray Ionization (ESI) mass spectra were collected on a Finnigan LCQ quadrupole ion trap mass spectrometer (samples were dissolved in HPLC grade solvent). 1 H, 19 F, and 31P NMR spectra were recorded on DPX 300 or Bruker Avance 500, 400 FT-NMR spectrometers. UV/Vis absorption spectra were recorded on a PerkinElmer Lambda 19 UV/Vis spectrophotometer. Steady-state emission spectra at 298 K were obtained on a Spex 1681 Flurolog-2 Model F111 spectrophotometer equipped with a Hamamatsu R928 PMT detector. All solutions for photophysical measurements, except stated otherwise, were degassed in a high-vacuum line with at least four freeze–pump–thaw cycles. Emission lifetimes were measured with a Quanta-Ray Q-switch DCR-3 Nd:YAG pulsed laser system. Emission quantum yields of solutions were measured by using a degassed acetonitrile solution of [Ru(bpy)3](PF6)2 (bpy = 2,2’-bipyridine) (fr = 0.062) as the standard and calculated by fs = fr(Br/Bs)(ns/nr)2(Ds/Dr), in which the subscripts s and r refer to the sample and reference standard solution Chem. Eur. J. 2015, 21, 7441 – 7453

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Computational methods The density functional theory (DFT) and time-dependent density functional theory (TDDFT) calculations for complex 1 were carried out using PBE0 functional[32] with a basis set of 6-31 + G*[33] for C, H, and N atoms and a pseudo-potential Stuttgart/Dresden (SDD)[34] basis set for Pt atom. The Gaussian 09 package was used for the calculations.[35]

MTT cytotoxicity assay Drug-treated cells (seeding density: 4 Õ 103 cells per well) were incubated with MTT for 4 h at 37 8C in a humidified atmosphere of 5 % CO2 and were subsequently lysed in solubilizing solution. Cells were then maintained in a dark, humidified chamber overnight. The formation of formazan was measured by using a microtitre plate reader at 580 nm. Growth inhibition by a drug was evaluated by IC50 (concentration of a drug causing 50 % inhibition of cell growth). Each growth inhibition experiment was repeated at least three times and results were expressed as mean œ standard deviation (SD).

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Full Paper Fluorescence microscopic analysis HeLa cells (2 Õ 105 cells) were seeded in a one chamber slide (Nalgene; Nunc) with culture medium (2 mL per well) and incubated at 37 8C in a humidified atmosphere of 5 % CO2/95 % air for 24 h. After treating with platinum(II) complex and/or organelle specific dye, cells were directly exposed for fluorescent imaging without removing the old medium.

Acknowledgements This work was supported by the National Key Basic Research Program of China (no. 2013CB834802), the University Grants Committee (Area of Excellence Scheme AoE/P-03/08), and CASCroucher Funding Scheme for Joint Laboratories. Keywords: anions · cancer luminescence · platinum

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carbenes

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cyanides

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Full Paper [35] Gaussian 09, Revision C.01, M. J. Frisch, G. W. Trucks, H. B. Schlegel, G. E. Scuseria, M. A. Robb, J. R. Cheeseman, G. Scalmani, V. Barone, B. Mennucci, G. A. Petersson, H. Nakatsuji, M. Caricato, X. Li, H. P. Hratchian, A. F. Izmaylov, J. Bloino, G. Zheng, J. L. Sonnenberg, M. Hada, M. Ehara, K. Toyota, R. Fukuda, J. Hasegawa, M. Ishida, T. Nakajima, Y. Honda, O. Kitao, H. Nakai, T. Vreven, J. A. Montgomery, Jr., J. E. Peralta, F. Ogliaro, M. Bearpark, J. J. Heyd, E. Brothers, K. N. Kudin, V. N. Staroverov, R. Kobayashi, J. Normand, K. Raghavachari, A. Rendell, J. C. Burant, S. S. Iyengar, J. Tomasi, M. Cossi, N. Rega, J. M. Millam, M. Klene, J. E. Knox, J. B.

Chem. Eur. J. 2015, 21, 7441 – 7453

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Cross, V. Bakken, C. Adamo, J. Jaramillo, R. Gomperts, R. E. Stratmann, O. Yazyev, A. J. Austin, R. Cammi, C. Pomelli, J. W. Ochterski, R. L. Martin, K. Morokuma, V. G. Zakrzewski, G. A. Voth, P. Salvador, J. J. Dannenberg, S. Dapprich, A. D. Daniels, O. Farkas, J. B. Foresman, J. V. Ortiz, J. Cioslowski, and D. J. Fox, Gaussian, Inc., Wallingford CT, 2011.

Received: December 12, 2014 Published online on April 1, 2015

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Pincer-Type Platinum(II) Complexes Containing N-Heterocyclic Carbene (NHC) Ligand: Structures, Photophysical and Anion-Binding Properties, and Anticancer Activities.

Two classes of pincer-type Pt(II) complexes containing tridentate N-donor ligands (1-8) or C-deprotonated N^C^N ligands derived from 1,3-di(2-pyridyl)...
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