Spectrochimica Acta Part A: Molecular and Biomolecular Spectroscopy 141 (2015) 16–26

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Spectrochimica Acta Part A: Molecular and Biomolecular Spectroscopy journal homepage: www.elsevier.com/locate/saa

New mixed ligand palladium(II) complexes based on the antiepileptic drug sodium valproate and bioactive nitrogen-donor ligands: Synthesis, structural characterization, binding interactions with DNA and BSA, in vitro cytotoxicity studies and DFT calculations Leila Tabrizi, Hossein Chiniforoshan ⇑, Hossein Tavakol Department of Chemistry, Isfahan University of Technology, Isfahan 84156-83111, Iran

h i g h l i g h t s

g r a p h i c a l a b s t r a c t

 Two Pd(II) complexes have been

synthesized and characterized.  DFT calculations of Pd(II) complexes

were studied.  The interaction of the Pd(II)

complexes with CT-DNA were investigated.  The Pd(II) complexes can bind to bovine serum albumin proteins.  Evaluation of cytotoxic activity of the Pd(II) complexes were carried out.

a r t i c l e

i n f o

Article history: Received 27 October 2014 Received in revised form 9 January 2015 Accepted 15 January 2015 Available online 28 January 2015 Keywords: Palladium(II) complex Sodium valproate DNA and BSA binding study Cytotoxicity in vitro Theoretical calculation

a b s t r a c t The complexes [Pd(valp)2(imidazole)2] (1), [Pd(valp)2(pyrazine)2] (2) (valp is sodium valproate) have been synthesized and characterized using IR, 1H NMR, 13C{1H} NMR and UV–Vis spectrometry. The interaction of complexes with CT-DNA has been investigated using spectroscopic tools and viscosity measurement. In each case, the association constant (Kb) was deduced from the absorption spectral study and the number of binding sites (n) and the binding constant (K) were calculated from relevant fluorescence quenching data. As a result, a non-covalent interaction between the metal complex and DNA was suggested, which could be assigned to an intercalative binding. In addition, the interaction of 1 and 2 was ventured with bovine serum albumin (BSA) with the help of absorption and fluorescence spectroscopy measurements. Through these techniques, the apparent association constant (Kapp) and the binding constant (K) could be calculated for each complex. Evaluation of cytotoxic activity of the complexes against four different cancer cell lines proved that the complexes exhibited cytotoxic specificity and significant cancer cell inhibitory rate. Moreover, density functional theory (DFT) calculations were employed to provide more evidence about the observed data. The majority of trans isomers were supported not only by energies, but also by the similarity of its calculated IR frequencies, UV adsorptions and NMR chemical shifts to the experimental values. Ó 2015 Elsevier B.V. All rights reserved.

Abbreviations: DNA, deoxyribonucleic acid; CT-DNA, calf thymus DNA; BSA, bovine serum albumin; HAS, human serum albumin; EB, ethidium bromide; IC50, half maximal inhibitory concentration; SV, Stern–Volmer; Valp, sodium valproate; Pd, palladium; DFT, density functional theory; TD-DFT, time-dependent density functional theory; NBO, natural bond orbital; FT-IR, Fourier transform infrared spectroscopy; UV–Vis, ultraviolet visible; NMR, nuclear magnetic resonance; calc., calculated; DMSO, dimethyl sulfoxide; FMOs, frontier molecular orbitals; HOMO, highest occupied molecular orbitals; LUMO, lowest unoccupied molecular orbitals; RMS, root mean square; GIAO, GaugeIndependent Atomic Orbital. ⇑ Corresponding author. Tel.: +98 3133913261; fax: +98 3133912350. E-mail address: [email protected] (H. Chiniforoshan). http://dx.doi.org/10.1016/j.saa.2015.01.027 1386-1425/Ó 2015 Elsevier B.V. All rights reserved.

L. Tabrizi et al. / Spectrochimica Acta Part A: Molecular and Biomolecular Spectroscopy 141 (2015) 16–26

Introduction Over the decades, there has been broad research in pharmaceutical chemistry to design effective anticancer drugs, potentially valuable in the treatment of diverse cancers [1–4]. Cisplatin is one of the leading chemotherapeutic drugs among actinomycin, anthracycline antibiotics and extensively used metal-based anticancer drugs for cancer therapy, but it possesses inherent limitations as serious side effects such as neurotoxicity, tissue toxicity, nausea, nephrotoxicity, emetogenesis, gastrointestinal, bone marrow toxicity and acquired drug resistance [5–8]. As a result, significant attempts are being made to replace this drug with appropriate alternatives and various transition metal complexes have been synthesized and tested for their anticancer activities [9–11]. Over the past decades, the investigations on the interaction of metal complexes with deoxyribonucleic acid (DNA) are of great interest of researchers due to their potential applications as anticancer medications and stereo selective probes of nucleic acid structures [12,11,13–25]. Bovine serum albumin (BSA) is soluble protein that has the ability to transport a multitude of endogenous and exogenous ligands such as fatty acids, amino acids, steroids, metal ions and drugs in blood stream. It is used to investigate the interaction of protein–drug complex in the circulatory system due to structural similarities with human serum albumin (HSA) [26]. Valproic acid (valp) is an extensive spectrum anti-epileptic drug which is efficient against all seizure varieties and that is increasingly used in the treatment of other diseases, including bipolar disorder, migraine, and neuropathic pain [27]. Furthermore, valproic acid was shown to improve the effect of chemotherapy on EBVpositive tumors, and to take a multitude of anti-tumor properties in vitro and in clinically appropriate animal models [28,29]. Esiobu and Hoosein [30], studied the effect of sodium valproate on the growth of a broad spectrum of microorganisms and they understood that it is selectively potent against yeast strains and Mycobacterium smegmatis. Though synthesis, characterization and biological activity of mixed ligands metal complexes of valproate with different nitrogen based ligands have been studied for copper [31–37], rhodium [38], zinc [39,40], and platinum [41], the biological application of palladium complexes with valproate is not well explored. Herein, we are reporting the synthesis, characterization and potential DNA/protein binding abilities and in vitro cytotoxicity studies of new valproate complexes of palladium with two nitrogen based ligands of pyrazine and imidazole, [Pd(valp)2(imidazole)2] (1) and [Pd(valp)2(pyrazine)2] (2). The structure of ligand and its complexes were clarified by elemental analysis, FT-IR, 1H NMR, 13C NMR, UV/Vis spectroscopies. Theoretical calculations using density functional theory (DFT) were done in order to correlate between the theoretical and experimental results. In this line, natural bond orbital (NBO) analysis was performed to present details about the type of hybridization and the nature of bonding in the studied complexes.

Experimental section General Starting materials and solvents were purchased from Sigma–Aldrich or Alfa Aesar and used without further purification. CT-DNA and BSA were purchased from Sigma–Aldrich and were used as supplied. Cisplatin was gifted from Isfahan University of Medical Sciences. Infrared spectra were recorded on a FT-IR JASCO 680 spectrophotometer in the spectral range 4000–400 cm1 using

17

the KBr pellets technique. NMR spectra were recorded on Bruker spectrometer at 400.13 MHz for 1H measurements and 75 MHz for the 13C{1H} measurements using TMS as an internal standard in DMSO-d6 solvent at 298 K. Elemental analysis was performed on a Leco, CHNS-932 apparatus. Molar conductivity measurements were carried out with a Crison Basic 30 conductometer. UV–Vis spectra were recorded on a JASCO 7580 UV–Vis-NIR double-beam spectrophotometer using a quartz cell with a path length of 10 mm. The fluorescence spectra complex bound to DNA were obtained at an excitation wavelength of 522 nm on a Perkin–Elmer LS55 fluorescence spectrofluorometer. Viscosity experiments were conducted on an Ostwald’s viscometer, immersed in a thermostated water-bath maintained at 25 °C. Synthesis of [Pd(valp)2(imidazole)2] (1) An ethanolic solution (15 cm3) of sodium valproate (0.33 g, 2 mmol) was added slowly and simultaneously with a ethanolic solution (15 cm3) of imidazole (0.14 g, 2 mmol) to a stirred ethanolic solution (25 cm3) of Pd(OAc)2 (0.23 g, 1 mmol). The mixture was stirred for 4 h at room temperature and then filtered through a plug of MgSO4. The filtrate was concentrated to ca. 5 cm3 and to this concentrated solution; n-hexane (15 cm3) was added to precipitate a white solid, which was collected and air-dried. The compound is soluble in water, methanol, ethanol, acetone, chloroform, dichloromethane, diethyl ether and ethyl acetate. Yield: 75%. Elemental analysis results were in good agreement with the for C22H38N4O4Pd stoichiometry. Found (calc.%): C, 49.86 (49.95); N, 10.41 (10.59); H, 7.14 (7.24). 1 H NMR (DMSO) (Fig. S1): d (ppm) 0.84 (t, 12H, H-5), 1.29 (m, 8H, H-4), 1.40 (m, 8H, H-3), 2.33 (m, 2H, H-2), 7.09 (s, 2H, H-8), 7.37 (s, 2H, H-9), 8.15 (s, 2H, H-6), 9.53 (s, 2H, H-7). 13C{1H} NMR (DMSO) (Fig. S2): d (ppm) 14.32 (C-5), 20.93 (C-4), 34.67 (C-3), 47.51 (C-2), 108.32 (C-9), 124.13 (C-8), 153.45 (C-6), 185.14 (C-1). IR (KBr, cm1) (Fig. S5): 3584vs, 3122vs, 2962s, 2973s, 1620s, 1520s, 1515m, 1489m, 1441s, 1346w, 1326m, 1268m, 1259m, 1221w, 1197w, 1115w, 1108w, 1091m, 1063vs, 1015w, 970w, 951w, 925w, 915w, 902w, 890w, 785vs 737w, 651m. Conductivity (KM, mho cm2 mol1) in water: 42. UV–Vis in water, kmax (log e): 282 (3.91), 302 (3.84). Synthesis of [Pd(valp)2(pyrazine)2] (2) Complex 2 was prepared in a similar way to 1 with the use of pyrazine (0.16 g, 2 mmol) instead of imidazole. The compound is soluble in water, methanol, ethanol, acetone, chloroform, dichloromethane, diethyl ether and ethyl acetate. Yield: 72%. Elemental analysis results were in good agreement with the for C24H38N4O4Pd stoichiometry. Found (calc.%): C, 49.97 (54.08); N, 10.49 (10.51); H, 7.15 (7.18); 1H NMR (DMSO) (Fig. S3): d (ppm) 0.81 (t, 12H, H-5), 1.21 (m, 8H, H-4), 1.42 (m, 8H, H-3), 2.28 (m, 2H, H-2), 8.49 (d, 4H, H-11,12), 8.87 (d, 4H, H-10,13). 13C{1H} NMR (DMSO) (Fig. S4): d (ppm) 14.28 (C-5), 20.83 (C-4), 33.87 (C-3), 47.34 (C-2), 143.42 (C-11, 12), 147.42 (C-10, 13), 184.91 (C-1). IR (KBr, cm1) (Fig. S6): 3115s, 3009m, 2960s, 2970s, 1631s, 1560vs, 1508s, 1489m, 1451s, 1341w, 1260m, 1220w, 1196w, 1181vs, 1110m, 1027w, 974w, 927w, 882m, 791m, 713w, 642w. Conductivity (KM, mho cm2 mol1) in water: 38. UV–Vis in water, kmax (log e): 275 (3.71), 310 (3.74). DNA binding studies of palladium(II) complexes The binding properties of the complex of CT-DNA have been studied using electronic absorption spectroscopy, competitive binding experiments, fluorescence spectroscopy and viscosity measurements. All the experiments involving the interaction of the

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complexes with CT-DNA were carried out in MilliQ water containing tris–HCl buffer (pH 8.02). The solution of CT-DNA in the buffer gave a ratio of UV absorbance of ca. 1.8–1.9:1 at 260 and 280 nm, indicating that the CT-DNA was sufficiently free of protein [42]. Stock solution of DNA was always stored at 4 °C in the dark and used within four days. The CT-DNA concentration per nucleotide was determined spectrophotometrically by employing an extinction coefficient of 6600 M1 cm1 at 260 nm [43]. The complexes were dissolved in a solvent 99% tris–HCl buffer at 1.0  104 M1 concentration. Absorption spectral titration experiment was performed by keeping constant the concentration of the complex (10 lM) and varying the CT-DNA concentration. While measuring the absorption spectra, an equal amount of CT-DNA was added to both the complex solutions and the reference solution to take into account the absorbance of DNA itself. In the emission quenching experiment, ethidium bromide (EB) was used as a common fluorescent probe for the DNA in order to examine the mode and process of metal complex binding to the double-helix [44]. A 5.0 lL of the EB tris–HCl buffer solution (1.0 mmol L1) was added to 1.0 mL of DNA solution (at saturated binding levels) [45], stored in the dark for 2 h. Then the solution of each of the Pd(II) complexes was titrated into the DNA/EB mixture and diluted in tris–HCl buffer to 5.0 mL to get the solution with the appropriate complex/CT-DNA mole ratio. After the incubation at room temperature for 30 min, the fluorescence spectra of EB bound to DNA were recorded (kex = 522 nm) in Perkin–Elmer LS55 fluorescence spectrofluorometer. All measurements were performed at ambient temperature. The binding interaction of the metal complexes with DNA was studied by the well known method employing the Ostwald’s viscometer. The CT-DNA solution (5 lM) was titrated with Pd(II) complexes (0.5–3.5 lM), following the change of the viscosity in each case. Data are presented as (g/g0)1/3 versus the ratio of the concentration of the compound and CT-DNA, where g is the viscosity of CT-DNA in presence of the compound and g0 is the viscosity of CT-DNA alone. Viscosity values were calculated from the observed flow time of CT-DNA-containing solution corrected from the flow time of buffer alone (t0), g = t  t0 [46]. Protein (BSA) binding experiments of palladium(II) complexes The binding study with bovine serum albumin (BSA) for complexes 1 and 2 were done dissolving the BSA in MilliQ water (1.0  105 M1) and the stock solutions of each of the complexes were prepared in H2O at 1.0  105 M1 concentration. Both the absorption and fluorescence quenching experiments (kex = 280 nm) were performed by gradually increasing the complex concentration, keeping fixed the concentration of BSA. All the experimental sets were carefully degassed purging pure nitrogen gas for 5 min. Cytotoxicity assay in vitro Cell proliferation was evaluated by using a system based on the tetrazolium compound [3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide, MTT] which is reduced by living cells to yield a soluble formazan product that can be assayed colorimetrically [47,48]. The four well-characterized cell lines, the HeLa cells, the Hep-G2 cells, the KB cells and the AGZY-83a cells were grown in 25 cm2 tissue culture flasks in an incubator at 37 °C in a humidified atmosphere consisting of 5% CO2 and 95% air, being maintained in a continuous logarithmic culture in RPMI 1640, 10% (v/ v) heat-inactivated fetal calf serum, 20.0 mM Hepes, 0.112% bicarbonate, and 2.0 mM glutamine. Each of the four cell lines (100.0 lL) were seeded at a density of 2.0  105 cells/mL into sterile 96-well culture medium. After incubation for 24 h, cells were exposed to the tested complexes of serial concentrations. The com-

plexes were dissolved in DMF and diluted with RPMI 1640 or DMEM to the required concentrations prior to use (0.1% DMF final concentration). After 72 h of incubation, the cells were treated with 20.0 mL MTT (5.0 mg/mL) for 4 h for further cultivation. The media with MTT were removed, and 100.0 lL of DMSO was added to dissolve formazan crystal at room temperature for 30 min. The absorbance of each cell was measured at 450 nm. All experiments were made in quadruplicate. The half-maximum inhibitory concentration (IC50) values were obtained from the results of quadruplicate determinations of at least three independent experiments. In another trial the effect on cell growth for the four complexes was studied by culturing the cells in medium alone for 24 h, followed by 72 h treatment with 3.0 lg/mL concentrations of complexes (dissolved in DMF and diluted with distilled water, 0.1% DMF final concentration). The viable cells remaining at the end of treatment period were determined by MTT assay and calculated as % of control, treated with vehicle alone (DMSO) under similar conditions. Computational details All calculations were performed using GAUSSIAN 09 program suites [49]. All calculations have performed by density functional theory (DFT) [50], at the B3LYP level using the effective core potential (ECP) basis set (LANL2DZ) for all atoms [51,52]. The frequency calculations on these structures verified that the optimized structures are stationary points corresponding to global minima. In addition, natural bond orbital (NBO) analysis [53,54] was carried out on all of the optimized geometries (local minimum and TS) to obtained atomic charges. Vibrational band assignments were made using the Gauss-View molecular visualization program [55]. The electronic absorption spectra were calculated using the time-dependent density functional theory (TD-DFT) method [56– 58] with the same basis set. The nuclear magnetic resonance (NMR) chemical shift calculations were performed using Gauge-Independent Atomic Orbital (GIAO) method [59,60] and the 1H and 13C isotropic chemical shifts were referenced to the corresponding values for TMS, which was calculated at the same level of theory. The effect of solvent was examined using polarized continuum model (PCM). Dimethylsulfoxide (DMSO) with relative permittivity (e) 46.7 was used as a solvent in PCM calculations. Results and discussion Synthesis and characterization The complexes 1 and 2, were obtained in good yield via the reaction of 1:2:2 M ratio of Pd(OAc)2 with sodium valproate and N-donor ligands (imidazole or pyrazine) in ethanol. The preparation process of the complexes is very simple, and only one type of compound could be detected by the occurrence of only one precipitate, after the mixture of reactants was stirred at room temperature. Both complexes were isolated as white solids, stable at room temperature, soluble in water, methanol, ethanol, acetone, chloroform, dichloromethane, diethyl ether and ethyl acetate. Formation of isolated complexes has been confirmed on the basis of characteristic bands in the IR spectra, elemental analysis, resonance signals in the 1H-, and 13C{1H} NMR. The conductivity measurement of complexes in water showed the conductance values KM = 38 and 42 mho cm2 mol1 at 300 K. These values suggest that the complexes exist as non-electrolytes in solution [61]. The magnitude of the difference between the symmetric and asymmetric carboxylate stretching frequencies, [D = mas (COO)  ms(COO)], is often used as spectroscopic criteria to determine

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the mode of carboxylate binding [62]. The assignments of IR frequencies for the asymmetric stretch, mas(COO), the symmetric stretch, ms(COO), and the difference between these two values of valproic group in complexes 1, 2 and sodium valproate are shown in Table S1. The separation of the frequencies, Dm (COO) = 179 cm1, for complex 1 and Dm (COO) = 180 cm1, for complex 2 are significantly higher than that found in sodium valproate, Dm (COO) = 137 cm1. This result could indicate monodentate coordination mode of the carboxylate group, since [Dm (COO)1 or 2 > Dm (COO)Navalp] [63]. In the IR spectrum of imidazole ligand in complex 1, the strong bands at 1520 and 1489 cm1 have been assigned to m (C@C) and m (C@N) stretching vibrations of the imidazole ring. The very strong band at 1057 cm1 and a medium band at 1084 cm1 are assigned to imidazole C–N stretching vibrations. Upon coordination the imidazole ligand m (C–N) and m (C@N) are shifted to lower energies. The pyrazine molecule (C4H4N2) has the centrosymmetric and planar structure with D2h symmetry in the vapour and solid phases. The assignments of the infrared spectra of pyrazine in solid [64], liquid [65,66] and vapour [67,68] phases and in solution [64,68,69] have been reported to be in accord with the D2h symmetry. The important infrared frequencies which are observed [68,69] in free pyrazine are 3129, 3100, 1500, 1487, 1179, 1107, 880 and 610. In metal complexes in which both N-atoms of pyrazine are coordinated to two metal atoms, the D2h symmetry is preserved but is removed in unidentate coordination. Several modes of pyrazine in complexes have upward shift in wave number in comparison with those of free pyrazine. This blue shift in pyrazine bands in the complex 2 indicates the coordination through N-atom as in pyridine [70,71]. This blue shift in pyrazine stretching may be due to back bonding from palladium to pyrazine ring through formation of extensive -bonding. Lever et al. [72–75] and Kantara [76] have reported that in complexes where pyrazine is bonded through only one N-atom, the low local symmetry experienced by the pyrazine moiety atoms allows a band to appear in the 950–1000 cm1 region, which may be taken as an evidence for unidentate nature of pyrazine. This band is reported to be absent in the bridged polymers where pyrazine functions as bidentate ligand. The pyrazine complex 2 exhibits a strong band in this region, thereby confirming the unidentate nature of pyrazine in this complex. In order to facilitate assignment of the observed peaks, we have calculated vibrational frequencies and compared them with observed values (Table S1). Moreover, root mean square error (RMSE) value was calculated using Eq. (1)to define which isomer has more adaption with observed spectra. For molecule 1, the RMSE between IR values of observed and trans isomer is 8.59 cm1 and between observed and cis isomer is 34.35 cm1 (trans isomer has more accordance). For molecule 2, the RMSE between IR values of observed and trans isomer is 6.05 cm1 and between observed and cis isomer is 26.12 cm1 (trans isomer has more accordance). These values confirm that in both molecules, the trans isomer is our major product.

sffiffiffiffiffiffiffiffiffiffiffiffiffiffiffiffiffiffiffiffiffiffiffiffiffiffiffiffiffiffiffiffiffiffiffiffiffiffiffiffiffiffiffiffiffi Pn 2 i¼1 ðv exp;i  mcalcu;i Þ RMSE ¼ n

electronic absorption values, it could be easily seen that in both structures, the trans isomers are corresponded to the experimental absorption values. The experimental and theoretical values for 1H and 13C{1H} NMR of both complexes, sodium valproate, imidazole, pyrazine are reported in Tables 2 and 3. The numbering schemes for 1Hand 13C{1H} NMR data of free ligands and complexes are shown in Scheme 1. The 1H NMR spectra of 1 and 2 showed that these compounds represent a molecular structure with an apparent center of inversion, which divided the molecules into two symmetrical parts. These values are in agreement with the structures of square planer proposed for these compounds. Comparison of the 1H NMR spectra of 1, 2 and sodium valproate showed a downfield shift of the former due to the complex formation. This deshielding effect may be due to electron donation of the carboxylate group to the palladium ion. The 1H NMR chemical shifts of the N-donor ligands of 1 and 2 were found in the expected positions of free ligands with almost all protons showing downfield shift, which provide evidence for coordination of ligands to palladium metal through the nitrogen atom of imidazole or pyrazine. The 13C{1H} NMR chemical shift of carbonyl (COO) group depends on the coordination mode and the coordination number of metal ions [77,78]. The carbon atoms of the N-based ligands of complexes 1 and 2 exhibit downfield chemical shift from the free ligand value, supporting complex formation (see Tables 2 and 3) which is explained by the depletion of electron density caused by bond formation with pyrazine or imidazole nitrogen atom. The predicted chemical shift values are in good agreement with the experimental values in trans isomer of complex 1 and 2, its trans isomers for these two complexes might be prepared. Theoretical studies Optimized structures In this part, studies of important aspects of the prepared molecules (complexes 1 and 2) by computational methods have reported. The optimized structures of these molecules are shown in Fig. 1. According to the elemental analysis, the complexes have a ligand (valproate) -to-metal- to ligand (pyrazine or imidazole) ratio of 2:1:2 for Pd(II) ion. Since the structure of tetrahedral for palladium complexes are not usual, the square planar structures were proposed for both complexes. The calculation of the Gibbs free energies showed that the trans isomers are more stable than the cis isomers in both complexes. For complex 1, trans is the

Table 1 Experimental and TD-DFT calculations of electronic spectral transitions for the Pd(II) complexes. Experimental kmax (log e)a

Calculated-trans kmax (f)b

Calculated-cis kmax (f)b

1

282 (3.91) 302 (3.84)

2

275 (3.71) 310 (3.74)

293.0 (0.0083) 306.17 (0.0073) 317.83 (0.0001) 341.40 (0) 377.80 (0) 408.97 (0) 439.61 (0) 265 (0.0011) 303 (0.0009) 368.10 (0.0001) 375.39 (0.0002) 388.90 (0.0002) 398.99 (0.0002) 415.79 (0.0004)

315.16 (0.0003) 325.71 (0.0134) 328.16 (0.0048) 351.17 (0.0081) 375.5 (0.0006) 412.14 (0.0035) 432.98 (0.0023) 329 (0.0003) 415.10 (0.0009) 426.0 (0.0016) 429.57 (0.0003) 446.82 (0.0014) 464.09 (0.0002) 468.88 (0.0003)

ð1Þ

The experimental electronic spectral data for complexes 1 and 2 in water solvent were recorded within 200–600 nm range. In addition, electronic absorption spectra of the complexes 1 and 2 as two trans and cis isomers were calculated using TD-DFT calculations. The experimental and theoretical electronic spectral data are assembled in Table 1. Complexes 1 and 2 have similar spectral features and are very similar to those of the parent ligands with only 1–5 nm shift caused by metal complexation. From the calculated

a b

k in nm (log e), in water solution. The oscillator strength (f).

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Table 2 Experimental and calculated (B3LYP/6-31+G(d,p))1H NMR,

13

C NMR chemical shifts (ppm) of ligands and complex 1.

H, C NO.

1

H NMR Navalp

13

C NMR Navalp

1

H NMR imidazole

13

C NMR imidazole

1

H NMR 1 (Exp.)

1

H NMR 1trans (Cal.)

1

H NMR 1cis (Cal.)

13 C NMR 1 (Exp.)

13

C NMR 1trans (Cal.)

13

1 2 3 4 5 6 7 8 9

– 2.02 1.18 1.05 0.67 – – – –

186.5 48.6 35.0 20.3 13.3 – – – –

– – – – – 7.70 (s) 10.50 (s) 7.03 (s) 7.03 (s)

– – – – – 137.79 – 129.17 120.20

– 2.33 (m) 1.40 (m) 1.29 (m) 0.84 (t) 8.15 (s) 9.53 (s) 7.09(s) 7.37 (s)

– 2.12 1.46 1.32 0.83 8.12 9.50 7.03 7.42

– 1.89 1.68 1.39 0.67 7.18 10.08 6.31 7.12

185.14 47.51 34.67 20.93 14.32 153.45 – 124.13 108.32

185.56 47.14 34.25 20.81 14.56 153.22 – 124.39 108.65

184.06 46.75 33.85 20.31 15.86 155.18 – 123.43 108.08

(tt) (m) (tq) (t)

C NMR 1cis (Cal.)

t: triplet, q: quartet, m: multiplet.

major isomer and it is more stable than cis by 4.00 kcal/mol and 0.33 kcal/mol, respectively in the gas phase and solvent. In addition, for complex 2, trans is the major isomer and it is more stable than cis by 8.94 kcal/mol and 3.22 kcal/mol, respectively in the gas phase and solvent (ethanol).

7 H N

8

NBO and population analyses: frontier orbitals and partial charges Population analyses were employed for all the geometric isomers to extract the energies of the frontier molecular orbitals (FMOs). Graphical presentations of the HOMO and LUMO of all the isomers and their energies (eV) are shown in Fig. 2. Fig. 2 shows noticeable different electron density distributions in the frontier orbitals of the cis and trans isomers of each complex and the electron density in both the LUMO and HOMO orbitals are different from each other. These differences are related to the location of the electron density and its quantity. Therefore, it is obvious that the reactivities of these complexes are different and the energy values of the frontier orbitals confirm these differences. By comparing the energies of the frontier orbitals, the LUMO– HOMO energy gap values in the trans isomers are less than those in the cis isomers in both complexes, which shows the trans isomers are more reactive than cis isomers for these two complexes. NBO calculations are used as a useful method for the determination of many properties, especially for the reproduction of more exact partial atomic charges. The results of these calculations for both isomers of complexes 1 and 2 (Table 4) showed that the nitrogen and oxygen atoms connected to Pd have different charges in the cis and trans isomers and smaller differences can be observed.

6

9

5

4

N

O

1

3

2

O Pd O

O N

3

4 N H

5 1

N

11

12 5

10

13

4

N

O

1

3

2

O Pd O

O

3

N

4 5

N

2

Binding experiments with calf thymus-DNA The mode of interaction of the complexes 1 and 2 with calf thymus DNA (CT-DNA) has been investigated using absorption and emission spectroscopic tools as well as by viscosity measurements.

Table 3 Experimental and calculated (B3LYP/6-31+G(d,p))1H-NMR,

13

Scheme 1. The structural formula of complexes 1 and 2 with numbering ligands for 1 H and 13C{1H} NMR of ligands and the complexes 1 and 2.

C NMR chemical shifts (ppm) of ligands and complex 2.

H, C NO.

1

H NMR pyrazine

13

C NMR pyrazine

H NMR 2 (Exp.)

1

H NMR 2trans (Cal.)

1

H NMR 2cis (Cal.)

13

C NMR 2 (Exp.)

13

C NMR 2trans (Cal.)

13

1 2 3 4 5 10 11 12 13

– – – – – 8.59 8.59 8.59 8.59

– – – – – 145.22 145.22 145.22 145.22

– 2.28 1.42 1.21 0.81 8.87 8.49 8.49 8.87

– 2.18 1.45 1.26 0.79 8.75 8.42 8.42 8.75

– 1.75 1.53 1.17 0.62 8.56 8.29 8.29 8.56

184.91 47.34 33.87 20.83 14.28 147.42 143.42 143.42 147.42

185.16 47.40 34.05 20.65 14.17 147.68 143.87 143.87 147.68

184.56 45.28 33.65 20.19 15.57 145.65 139.40 139.40 145.65

(s) (s) (s) (s)

t: triplet, q: quartet, m: multiplet.

1

(m) (m) (m) (t) (d) (d) (d) (d)

C NMR 2cis (Cal.)

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Fig. 1. Graphical presentation of the optimized structures of the cis (top) and trans (bottom) isomers of complexes 1 (left) and 2 (right).

Absorption spectroscopy Electronic absorption spectroscopy is used as a distinguishing characterization tool for examining the binding mode of metal complexes with DNA [45,79]. In intercalative binding mode, the p⁄ orbital of the intercalated ligand can couple with the p orbital of the base pairs, therefore decreasing the p ? p⁄ transition energy and resulting in bathochromism. However, the coupling p orbital was partially filled by electrons, so decreasing the transition probabilities and concomitantly resulting in hypochromism [80]. The absorption spectra of the free metal complexes and of their compounds with CT-DNA (at a constant concentration of the compounds) are given in Fig. 3 for complexes 1 and 2. The extent of hyperchromism in the absorption band is generally consistent with the strength of intercalative interaction [79–82]. As the concentration of CT-DNA is increased, it was found that the complexes 1 and 2 at 270 nm and 268 nm exhibit hyperchromicity of 3.3/18.90% and 3.5/22.22%, respectively. This feature might be ascribed to the fact that both of the co-complexes could uncoil the helix structure of DNA and made more bases embedding in DNA exposed [83–85]. In order to establish the binding strength of the metal complexes with CT-DNA, the apparent association constant Kb was determined from the spectral titration data using the following equation [86]:

where [DNA] is the concentration of DNA in base pairs, the apparent extinction coefficient ea, ef and eb are respectively the apparent, bound and free extinctions coefficient of each of the compound in respective cases. The Kb, expressed as M1, is derived from the slope of the graph obtained by plotting the [DNA]/(ea  ef) versus [DNA] (Fig. S7). The Kb values for complexes 1 and 2 were estimated to be 0.34  104 M1 (R = 0.99998, n = 5 points) and 0.91  104 M1 (R = 0.99985, n = 5 points), respectively. In order to confirm the binding mode of intercalation of the complexes 1 and 2 with CT-DNA, we employed ethidium bromide (EB) that, interacting with DNA, represents a characteristic indicator of intercalation. The maximal absorption of EB at 477 nm decreased and shifted to 498 nm in presence of DNA (Fig. 4), typically indicating insertion between the base pairs [87]. The observed behavior could be indicative of: (1) being EB strongly bound to complex 1 (or 2), the result is a decrease amount of EB intercalated into DNA; or (2) there exists a competition between the complexes 1 and 2 and EB towards DNA intercalation, so releasing some free EB from DNA–EB complex. On the other hand, here the former account could be irrelevant due to the appearance of a new absorption band.

½DNA ½DNA 1 ¼ þ ðea  ef Þ ðeb  ef Þ K b ðeb  ef Þ

Fluorescence quenching analysis The binding propensity of complexes 1 and 2 to CT-DNA has been analyzed by the steady-state emission quenching experiments

ð2Þ

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Fig. 2. Graphical presentation of the LUMO and HOMO for the optimized structures of the cis and trans isomers of complexes 1 and 2, and their energies (eV).

Table 4 Calculated NBO charges on selected atoms of ligand (valproate, imidazole, pyrazine) and isomers of complex 1 and 2.

Pd N1 N2 O1

1-Cis

1-Trans

2-Cis

2-Trans

0.712 0.543 0.543 0.696

0.716 0.531 0.532 0.715

0.701 0.483 0.483 0.689

0.714 0.476 0.468 0.729

using the emission intensity of ethidium bromide (EB). It is well known that EB can intercalate nonspecifically with DNA, causing a strong fluorescence. Other compounds competing with EB to intercalation in DNA will induce displacement of bound EB and a decrease in the fluorescence intensity. This fluorescence-based competition can provide indirect evidence for the DNA-binding mode. The fluorescence intensity of the EB/DNA system (with excitation wavelength of 522 nm) is reduced by the increasing concentration of the complexes (Fig. 5), and caused by EB migration from a

hydrophobic to an aqueous environment [88]. The quenching of EB bound to DNA by 1 and 2 is in agreement with the linear Stern–Volmer equation [89]:

I0 =I ¼ 1 þ K q ½Q 

ð3Þ

where I0 and I represent the fluorescence intensities in the absence and presence of quencher, respectively. Kq is a linear Stern–Volmer quenching constant, Q is the concentration of the quencher. In the quenching plot (insets of Fig. 5) of I0/I versus [complex], Kq is given by the ratio of the slope to the intercept. The Kq values are 0. 16  104 and 0.14  104 for complexes 1 and 2 respectively implies that both complexes can insert between DNA base pairs. The titration data obtained from the fluorescence experiment can be helpful also to calculate the number of binding sites and the apparent binding constant. In the following equation [90]:

Log ½ðI0  IÞ=I ¼ log K þ n log½Q 

ð4Þ

K and n represent the binding constant and number of binding sites of palladium complex to CT-DNA, respectively. The number of

Fig. 3. Electronic spectral titration of complex 1 (green) and 2 (blue) with CT-DNA at 271 nm in tris–HCl buffer; [complex 1 or 2] = 2.62  105; [DNA]: (a) 0.0, (b) 1.25  106, (c) 2.5  106, (d) 3.75  106, (e) 5.0  106, (f) 6.25  106 mol L1. The arrow (a ? f) denotes the gradual increase of DNA concentration. (For interpretation of the references to colour in this figure legend, the reader is referred to the web version of this article.)

L. Tabrizi et al. / Spectrochimica Acta Part A: Molecular and Biomolecular Spectroscopy 141 (2015) 16–26

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Fig. 4. (a) The electronic spectra of 1.0  105 M EB (A); (A) + 2.5  105 M DNA (B); (B) + 2.5  105 M complex 1 (C) in tris–HCl buffer. (b) The electronic spectra of 1.0  105 M EB (A); (A) + 2.5  105 M DNA (B); (B) + 2.5  105 M complex 2 (C) in tris–HCl buffer.

Fig. 5. Emission spectra of the CT-DNA–EB system in tris–HCl buffer upon titration with complex 1 (green) and complex 2 (blue). kex = 522 nm; [EB] = 9.6  105; [DNA] = 1.25  105; [complex 1 or 2]: (a) 0.0, (b) 1.31  105, (c) 2.62  105, (d) 3.93  105, (e) 5.24  105, (f) 6.55  105 mol L1. The arrow (a ? f) denotes the gradual increase of complex concentration. Inset shows the plot of I0/I vs. [1]; Kq = 0.16  104 (R = 0.99986, n = 5 points) and [2]; Kq = 0.14  104 (R = 0.99943, n = 5 points). (For interpretation of the references to colour in this figure legend, the reader is referred to the web version of this article.)

binding sites n, determined from the intercept of log [(I0  I)/I] versus log [Q] (Fig. S8), are 1.05 and 1.16 for 1 and 2, respectively, indicating the existence of about a single binding site in DNA and a weaker association for the complexes. The K values were calculated to be 0.34  104 and 0.75  104 for 1 and 2, respectively, with a trend similar to the apparent association constant values of the complexes. Viscosity measurements Since optical photophysical probes generally present necessary, but insufficient evidences to further explain the interactions between the complex and DNA, viscosity measurements were performed. Hydrodynamic measurements, sensitive to length change

(i.e. viscosity and sedimentation), are observed as the least ambiguous and the most critical tests of binding in solution in the absence of crystallographic structural data. A classical intercalation model requires that the DNA helix lengthens as base pairs are separated in order to accommodate the binding ligand, causing an increase in DNA viscosity. On the contrary, a partial, non-classical intercalation of compound could bend the DNA helix, reducing its efficient length and along with its viscosity [46]. The results obtained in these viscosity measurement investigations suggest that both the compounds 1 and 2 can intercalate between adjacent DNA base pairs, causing an extension of the helix with a concomitant increase of the DNA viscosity. The effects of both compounds on the viscosity of DNA are shown in Fig. 6. Binding experiments with bovine serum albumin (BSA) Absorption spectral characterization The binding mode of complexes 1 and 2 with BSA were studied by electronic absorption titration with BSA. The absorption spectra of the free metal complexes and of their adducts with BSA are given in Fig. 7 for complex 1 and 2, respectively. The spectra indicate a significant increase in the absorbance of BSA by increasing the concentration of the complex and are indicative of the fact that BSA adsorbs strongly the complex on its surface [91]. From these titration data the apparent association constant (Kapp) of the complexes with BSA has been determined using the following equation [80]:

1=ðAobs  A0 Þ ¼ 1=ðAc  A0 Þ þ 1=K app =ðAc  A0 Þ½comp Fig. 6. Effect of increasing amounts of complexes 1 and 2 on the relative viscosity of CT-DNA at 25 °C.

ð5Þ

where, Aobs is the observed absorbance of the solution containing different concentrations of the complex, A0 and Ac are the

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Fig. 7. Absorption titration spectra of BSA in presence of complex 1 (green) and complex 2 (blue). Concentration range of complex is 0–6.25  106 M1 (a ? f). (For interpretation of the references to colour in this figure legend, the reader is referred to the web version of this article.)

Fig. 8. Fluorescence quenching titration of BSA varying the concentrations of complex 1 (green) and complex 2 (blue), [complex 1 or 2] = 0, 1, 2, 3, 4 and 5  6.35  106 M (a?f). Inset shows the Stern–Volmer plot. (For interpretation of the references to colour in this figure legend, the reader is referred to the web version of this article.)

Table 5 Cytotoxicity of the complexes against selected human tumor cells after 72 h of incubation. Test complex

IC50 (lM) HeLa

Hep-G2

KB

AGZY-83a

1 2 Cisplatin

1.69 ± 0.34 2.52 ± 0.48 0.69 ± 0.14

2.78 ± 0.75 3.22 ± 0.84 1.57 ± 0.51

3.61 ± 1.12 4.45 ± 1.24 1.47 ± 0.23

2.49 ± 0.78 3.76 ± 1.22 2.25 ± 0.51

absorbance of BSA and of the complex at 280 nm, respectively. The enhancement of the absorbance at 280 nm was attributable to the complex absorption at BSA surface. Based on the linear relationship between 1/(Aobs  A0) versus the reciprocal concentration of the complex with a slope of 1/Kapp(Ac  A0) and an intercept equal to 1/(Ac  A0) (Fig. S9), the value of Kapp was determined to be 1.258  105 M1 (R = 0.99987, n = 5 points) and 1.392  105 M1 (R = 0.99981, n = 5 points), for 1 and 2, respectively.

Fluorescence quenching analysis In the fluorescence quenching experiment, the fluorescence emission spectrum of BSA was investigated increasing the concentration of the quencher (Fig. 8). The fluorescence quenching is described by the Stern–Volmer relation, [91] in the same way as described above for CT-DNA binding experiments. From the slope of the regression line in the derived plot of I0/I versus [complex] (insets of Fig. 8) the Kq values for the complexes were determined to be 4.32  104 for 1 (R = 0.99865 for five points) and 4.58  104 for 2 (R = 0.99978 for five points), indicating a strong affinity of both of the complexes to BSA.

Fig. 9. Effect of 3.0 lg/mL of the complexes on breast cancer cells viability after 72 h of incubation. All determinations are expressed as percentage of the control (untreated cells).

Cytotoxicity in vitro study The prepared complexes were tested for their in vitro cytotoxic activity by MTT assay. IC50 values were estimated for four human tumor cell lines, HeLa, Hep-G2, KB and AGZY-83a, which are presented in Table 5. Fig. 9 displays the effect on cell growth after a treatment period of 72 h treatment with 3.0 lg/mL concentration. Among the compounds studied here, both complexes exhibit cytotoxic activity. In contrast, complex 1 reveals more active than

L. Tabrizi et al. / Spectrochimica Acta Part A: Molecular and Biomolecular Spectroscopy 141 (2015) 16–26

complex 2, especially against HeLa cells. It demonstrates a high level of resistance against conventional chemotherapeutic agents. A possibility rate by day 3 to less than 50% of the control values was observed for the complexes. The results coincide with IC50 values reveal. Conclusions We described the synthesis and characterization of two square planar complexes, [Pd(valp)2(imidazole)2] (1), [Pd(valp)2(pyrazine)2] (2) (valp is sodium valproate) by 1H NMR, 13C{1H} NMR, Infrared and UV–Vis spectrometric techniques. The majority of trans isomers were proposed not only by energies, but also by the similarity of its calculated IR frequencies, UV adsorptions and NMR chemical shifts to the experimental values. The complexes have been found to interact with CT-DNA through an intercalative mode, which was investigated by absorption, fluorescence and viscosity measurement tools. The quenching rate constant, binding constant and number of binding sites were calculated according to the relevant fluorescence data. The binding constants indicate that the DNA-binding affinity, as well as the binding trend with BSA, increases from complex 1 to 2, in accordance with the relevant viscosity measurement study. Cytotoxic and antiproliferative studies show that the two complexes exhibit good cytotoxic activity against different cell lines tested in general, especially complex 1 is more effective against HeLa cell lines. Thus the new synthesized palladium(II) complexes may be potential antitumor agent due to its unique interaction mode with DNA. Acknowledgement We are grateful for the financial support from the Department of Chemistry, Isfahan University Of Technology (IUT). Appendix A. Supplementary data Supplementary data associated with this article can be found, in the online version, at http://dx.doi.org/10.1016/j.saa.2015.01.027. References [1] [2] [3] [4] [5] [6] [7] [8] [9] [10] [11] [12] [13] [14] [15] [16] [17] [18] [19] [20] [21] [22] [23]

D. Wang, S.J. Lippard, Nat. Rev. Drug Discovery 4 (2005) 307–320. P.A. Andrews, J.A. Jones, Cancer Commun. 3 (1991) 93–102. M.E. Bianchi, M. Beltrame, G. Paonessa, Science 243 (1989) 1056–1059. D. Bissett, K. McLaughlin, L.R. Kelland, R. Brown, Br. J. Cancer 67 (1993) 742– 748. M.A. Fuertes, J. Castilla, C. Alonso, J.M. Pérez, Curr. Med. Chem. 10 (2003) 257– 266. R.W. Schrier, J. Clin. Invest. 110 (2002) 743–745. J. Reedijk, Curr. Opin. Chem. Biol. 3 (1999) 236–240. F.L. Wimmer, S. Wimmer, P. Castan, S. Cros, N. Johnson, E. Colacio-Rodríguez, Anticancer Res. 9 (1989) 791–793. G. Zhao, H. Sun, H. Lin, S. Zhu, X. Su, Y. Chen, J. Inorg. Biochem. 72 (1998) 173– 177. E. Budzisz, U. Krajewska, M. Rozalski, Pol. J. Pharmacol. 56 (2004) 473–478. A. Divsalar, A.A. Saboury, H. Mansouri-Torshizi, A.A. Moosavi-Movahedi, J. Biomol. Struct. Dyn. 25 (2007) 173–182. L.T. Bozic, M. Juribasic, P. Traldi, V. Scarcia, A. Furlani, Polyhedron 27 (2008) 1317–1328. H. Mansouri-Torshizi, M. Saeidifar, A. Divsalar, A.A. Saboury, S. Shahraki, Bull. Korean Chem. Soc. 31 (2010) 435–441. S. Leininger, B. Olenyuk, P.J. Stang, Chem. Rev. 100 (2000) 853–908. S. Feng, R. Xu, Acc. Chem. Res. 34 (2001) 239–247. M. Yuan, Y. Li, E. Wang, J. Chem. Soc., Dalton Trans. 4 (2002) 2916–2920. Y. Lu, E. Wang, M. Yuan, J. Chem. Soc., Dalton Trans. 15 (2002) 3029–3031. B. Maity, S. Gadadhar, T.K. Goswami, A.A. Karande, A.R. Chakravarty, Eur. J. Med. Chem. 57 (2012) 250–258. R. Loganathan, S. Ramakrishnan, E. Suresh, A. Riyasdeen, M. Abdulkadhar, M.A. Akbarsha, M. Palaniandavar, Inorg. Chem. 51 (2012) 5512–5532. K.E. Erkkila, D.T. Odom, J.K. Barton, Chem. Rev. 99 (1999) 2777–2796. B. Meunier, Chem. Rev. 92 (1992) 1411–1456. G. Pratviel, J. Bernadou, B. Meunier, Angew. Chem., Int. Ed. 34 (1995) 746–769. D.S. Sigman, A. Mazumder, D.M. Perrin, Chem. Rev. 93 (1993) 2295–2316.

25

[24] J. Reedijk, J. Inorg. Biochem. 86 (2001) 89. [25] C.J. Burrows, J.G. Muller, Chem. Rev. 98 (1998) 1109–1152. [26] K.D. Demertzi, M.A. Demertzis, E. Filiou, A.A. Pantazaki, P.N. Yadav, J.R. Miller, Y. Zheng, D.A. Kyriakidis, Biometals 16 (2003) 411–418. [27] S.B.R. Fagundes, Rev. Neurosci. 16 (2008) 130–136. [28] W.H. Feng, S.C. Kenney, Cancer Res. 66 (2006) 8762–8769. [29] R.A. Blaheta, H. Nau, M. Michaelis, J.E. Jr, Curr. Med. Chem. 9 (2002) 1417– 1433. [30] N. Esiobu, N. Hoosein, Antonie Van Leeuwenhoek 83 (2003) 63–68. [31] A.L. Abuhijleh, Polyhedron 15 (1996) 285–293. [32] A.L. Abuhijleh, C. Woods, Inorg. Biochem. 64 (1996) 55–67. [33] A.L. Abuhijleh, C. Woods, Inorg. Chim. Acta 209 (1993) 187–193. [34] A.L. Abuhijleh, J. Inorg. Biochem. 68 (1997) 167–175. [35] M.S. Veitia, F. Dumas, G. Morgant, J.R.J. Sorenson, Y. Frapart, A. Tomas, Biochimie 91 (2009) 1286–1293. [36] P.C. Christidis, P.J. Rentzeperis, M.P. Sigalas, C.C. Hadjikostas, Z. Crystallogr. 176 (1986) 103–112. [37] C.C. Hadjikostas, G.A. Katsoulos, M.P. Sigalas, A. Tsipis, Inorg. Chim. Acta 167 (1990) 165–169. [38] A.L. Abuhijleh, H. Abu Ali, A.H. Emwas, J. Organomet. Chem. 694 (2009) 3590– 3596. [39] M. Darawsheh, H. Abu Ali, A. Latif Abuhijleh, E. Rappocciolo, M. Akkawi, S. Jaber, S. Maloul, Y. Hussein, Eur. J. Med. Chem. 82 (2014) 152–163. [40] H. Abu Ali, M. Darawsheh, E. Rappocciolo, Polyhedron 61 (2013) 235–241. [41] D.M. Griffith, B. Duff, K.H. Suponitsky, K. Kavanagh, M.P. Morgan, D. Egan, C.J. Marmion, J. Inorg. Biochem. 105 (2011) 793–799. [42] S. Satyanarayana, J.C. Dabrowiak, J.B. Chaires, Biochemistry 31 (1992) 9319– 9324. [43] C.V. Kumar, E.H. Asuncion, J. Am. Chem. Soc. 115 (1993) 8541–8553. [44] C.V. Kumar, J.K. Barton, M.J. Turro, J. Am. Chem. Soc. 107 (1985) 5518–5523. [45] J.K. Barton, A.T. Danishefsky, J.M. Golderg, J. Am. Chem. Soc. 106 (1984) 2172– 2176. [46] Y. Xiong, X.F. He, X.H. Zou, J.Z. Wu, X.M. Chen, L.N. Ji, R.H. Li, J.Y. Zhou, R.B. Yu, J. Chem. Soc., Dalton Trans. 1 (1999) 19–24. [47] Ana I. Matesanz, José M. Pérez, Paloma Navarro, José M. Moreno, Enrique Colacio, Pilar Souza, J. Inorg. Biochem. 76 (1999) 29–37. [48] Enjun Gao, Mingchang Zhu, Hongxi Yin, Lei Liu, Wu Qiong, Yaguang Sun, J. Inorg. Biochem. 102 (2008) 1958–1964. [49] 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. 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, D.J. Fox, Gaussian 09, Revision A.02, Gaussian, Inc., Wallingford, CT, 2009. [50] J.L. Calais, Int. J. Quantum Chem. 47 (1993) 101–107. [51] A.D. Becke, J. Chem. Phys. 98 (1993) 5648–5652. [52] C. Lee, W. Yang, R.G. Parr, Phys. Rev. B 37 (1988) 785–789. [53] A.E. Reed, R.B. Weinstock, F. Weinhold, J. Chem. Phys. 83 (1985) 735–746. [54] A.E. Reed, L.A. Curtiss, F. Weinhold, Chem. Rev. 88 (1988) 899–926. [55] R. Dennington II, T. Keith, J. Millam, GaussView, Version 5.0.9, Semichem, Inc., Shawnee Mission, KS, 2008. [56] E. Runge, E.K.U. Gross, Phys. Rev. Lett. 52 (1984) 997–1000. [57] R.E. Stratmann, G.E. Scuseria, M.J. Frisch, J. Chem. Phys. 109 (1998) 8218–8224. [58] M.E. Casida, C. Jamorski, K.C. Casida, D.R. Salahub, J. Chem. Phys. 108 (1998) 4439–4449. [59] M.J. Frisch, J.A. Pople, J.S. Binkley, J. Chem. Phys. 80 (1984) 3265–3269. [60] R. Ditchfield, Mol. Phys. 27 (1974) 789–807. [61] K. Dhara, P. Roy, J. Ratha, M. Manassero, P. Banerjee, Polyhedron 26 (2007) 4509–4517. [62] G.B. Deacon, R.J. Philips, Coord. Chem. Rev. 33 (1980) 227–250. [63] V. Zelenak, Z. Vargova, K. Gyoryova, Spectrochim. Acta Part A Mol. Biomol. Spectrosc. 66 (2007) 262–272. [64] G. Sbrana, V.S. Chettino, R. Righini, J. Chem. Phys. 59 (1973) 2441–2450. [65] R.C. Loard, A.J. Marson, F.A. Miller, Spectrochim. Acta 9 (1957) 113–125. [66] J. Zarembowitch, L. Bokibza-Sebagh, Spectrochim. Acta, A 32 (1976) 605–609. [67] S. Califano, G. Adembri, G. Sbrana, Spectrochim. Acta 20 (1964) 385–396. [68] J.F. Arenas, J.J. Lopez-Navarrate, J.C. Otero, J.I. Macros, A. Cardenate, J. Chem. Soc., Faraday Trans. 81 (1985) 405–416. [69] Z. Kantarci, B. Davarcioglu, C. Bayrak, J. Inclusion Phenom. Macrocyclic Chem. 39 (2001) 115–121. [70] N.N. Greenwood, K. Wade, J. Chem. Soc., A (1960) 1130–1141. [71] J.K. Wilmhurst, H.J. Bernsteim, Can. J. Chem. 35 (1957) 1183–1194. [72] B.P. Lever, J. Lewis, R.S. Nyholm, Nature London 189 (1961) 58–59. [73] B.P. Lever, J. Lewis, R.S. Nyholm, J. Chem. Soc. (1962) 1235–1246 [74] B.P. Lever, J. Lewis, R.S. Nyholm, J. Chem. Soc. (1963) 3156–3158 [75] B.P. Lever, J. Lewis, R.S. Nyholm, J. Chem. Soc. (1963) 5042–5048 [76] H.D. Stidham, J.A. Chandler, J. Inorg. Nucl. Chem. 27 (1964) 397–403. [77] S.J. Lin, T.N. Hong, J.Y. Tung, J.H. Chen, Inorg. Chem. 36 (1997) 3886–3891. [78] D.K. Lavallee, J.D. Doi, Inorg. Chem. 20 (1981) 3345–3349.

26

L. Tabrizi et al. / Spectrochimica Acta Part A: Molecular and Biomolecular Spectroscopy 141 (2015) 16–26

[79] S.A. Tysoe, R.J. Morgan, A.D. Baker, T.C. Strekas, J. Phys. Chem. 97 (1993) 1707– 1711. [80] A.M. Pyle, J.P. Rehmann, R. Meshoyrer, C.V. Kumar, N.J. Turro, J.K. Barton, J. Am. Chem. Soc. 111 (1989) 3051–3058. [81] M. Panda, S. Das, G. Mostafa, A. Castineiras, S. Goswami, Dalton Trans. (2005) 1249–1255 [82] S. Chattopadhyay, C. Sinha, P. Basu, A. Chakravorty, Organometallics 10 (1991) 1135–1139. [83] G. Pratviel, J. Bernadou, B. Meunier, Adv. Inorg. Chem. 45 (1998) 251–312. [84] Q.L. Zhang, J.G. Liu, H. Xu, H. Li, J.Z. Liu, H. Zhou, L.H. Qu, L.N. Ji, Polyhedron 20 (2001) 3049–3055.

[85] Z.S. Yang, Y.L. Wang, G.C. Zhao, Anal. Sci. 20 (2004) 1127–1130. [86] M. Baldini, M. Belicchi-Ferrari, F. Bisceglie, P.P. Dall Aglio, G. Pelosi, S. Pinelli, P. Tarasconi, Inorg. Chem. 43 (2004) 7170–7179. [87] W.D. Wilson, L. Ratmeyer, M. Zhao, L. Strekowski, D. Boykin, Biochemistry 32 (1993) 4098–4104. [88] Y.B. Zeng, N. Yang, W.S. Liu, J. Inorg. Biochem. 97 (2003) 258–264. [89] M.R. Efink, C.A. Ghiron, Anal. Biochem. 114 (1981) 199–227. [90] A. Kathiravan, R. Renganathan, Polyhedron 28 (2009) 1374–1378. [91] C. Chen, X. Qi, B. Zhou, J. Photochem. Photobiol. A: Chem. 109 (1997) 155–158.