Zahra Kazemi

Amiri Hadi

Valiollah Rudbari

Mehdi Mirkhani

Majid Sahihi

Sharam Moghadam

Iraj Tangestaninejad

Mohammadpoor-Baltork

Tangestaninejad

Raj Mohammadpoor-Baltork

Synthesis, characterization, crystal structure, DNA-and HSA-binding studies of a dinuclear Schiff base Zn(II) complex derived from 2-hydroxynaphtaldehyde and 2-picolylamine

Keywords:

niveau recherche, publiés ou non, émanant des établissements d'enseignement et de recherche français ou étrangers, des laboratoires publics ou privés.

Introduction

Nowadays, cancer is known as an important agent of worldwide death which can be cured by damage to DNA of cancer cells. An anti-cancer drug binds to DNA of cancer cells and inhibits DNA replication and transcription [1], [2]. Great investigations have been performed on metalbased pharmaceutical molecules since use of cisplatin as an anti-cancer drug [3]. The metal complexes have high affinity to DNA and can be introduced as anti-cancer drug candidates [4].

Moreover, literatures reported that the coordination of metal to ligand increases the biological activity of compounds [5], [6]. Schiff bases have attracted a great attention in bioinorganic field among other ligands due to their stability, biocompatibility and structural variability [4], [7].

Zinc(II) is the most abundant essential transition metal after Fe 3+ in human body and many enzymes such as carboxypeptidase and carbonic anhydrase use Zinc(II) in their active sites [8], [9].

Hence it has attracted a great attention in medicine field [10] and many studies have been carried out on Zn(II) complex and its biological properties such as antimicrobial [11] and antitumor [12], [13], [14] activity. Some studies reported the bigger DNA binding constant for Zn in comparison to the other metals such as Cu and Ni [15]. Moreover, researches revealed that bimetallic complexes have higher affinity with DNA in contrast with their mononuclear complex analogue [START_REF] Smith | Noncovalent DNA binding of metal complexes[END_REF].

DNA is known as a major target for anticancer drugs. Thus, the study of interaction between Schiff base complexes and DNA is critical step to design and synthesis of new pharmaceutical molecules which is an active area of research [START_REF] Shahabadi | [END_REF], [18]. The small molecules including drugs can interact with DNA through three models: (I) electrostatic binding between cation species and negative charged DNA phosphate residues (II) interaction with grooves of DNA by hydrogen bond or van der Waals interactions, and (III) intercalative binding [19], [20], [21].

Human serum albumin (HSA) is the most abundant plasma carrier protein, which is involved in binding and transport drugs in the blood. Therefore, the first step to know the mechanism of anti-cancer drug in body is the investigation its HSA binding. The Schiff base complexes as candidate for anti-cancer drugs have high affinity for binding to HSA [22], [23].

In this work, we have synthesized a new tridentate Schiff base ligand (HL) and its dinuclear Zinc(II) complex Zn2L2(NO3)2 (see Scheme 1, Scheme 2). The schiff base ligand and its Zn(II) complex were characterized by FT-IR, 1 H NMR, 13 C NMR and elemental analysis. In addition, the molecular structure of complex was determined by single crystal X-ray diffraction technique.

Furthermore, HSA and DNA binding of dinuclear Zn(II) complex was investigated using fluorescence quenching, UV-Vis spectroscopy, molecular dynamics simulation and molecular docking methods. Scheme 1. Synthesis of Ligand (HL). Scheme 2. Synthetic route for preparation of Zn2L2(NO3)2 complex.

Experimental

Materials

Human serum albumin (HSA), fish sperm DNA, Tris(hydroxymethyl)-aminomethane (Tris) and ethidium bromide (3,8-diamino-5-ethyl-6-phenylphenanthridinium bromide, EtBr) were purchased from Sigma-Aldrich. 2-Hydroxy-1-naphtaldehhyde and 2-picolylamine were obtained from Merck chemical Co. and used without further purification. All salts used for buffer preparation were analytical grade and dissolved in double distilled water. All of the solutions were used freshly after preparation.

Apparatus and software

The FT-IR spectrum was recorded on a JASCO, FT/IR-6300 spectrometer (4000-400 cm -1 ) in KBr pellets. 1 H and 13 C NMR spectra were recorded on a Bruker Avance 400 spectrometer using CDCl3 as solvent for the ligand and DMSO for the Zn(II) complex. The UV-Vis spectra were recorded on a JASCO V-570 spectrophotometer. Fluorescence measurements were carried out on Shimadzu RF-5000 spectrofluorometer at room temperature.

Synthesis of Schiff base ligand

A methanolic solution (10 ml) of 2-Picolylamine (10 mmol) was slowly added to a solution of 2hydroxy-1-naphtaldehyde (10 mmol) in absolute methanol (10 ml). Then the mixture was stirred at room temperature for 2 h. The reaction mixture was allowed to stand for slow evaporation of the solvent until the formation of yellow oil. After that the resulting yellow oil re-dissolved in a minimum amount of methanol and light yellow precipitate was extracted by diethyl ether. Yield

Synthesis of Zn(II) complex

Methanolic solution (10 ml) of 2-hydroxy-1-naphtaldehyde (5 mmol) was added dropwise to a methanolic solution (10 ml) of 2-picolylamin (5 mmol). The mixture was stirred for 2 h at room temperature. Then a solution of triethylamine (7 mmol) in absolute methanol (5 ml) was added to the mixture. The mixture stirred for 10 min, and then a solution of Zn(NO3)2⋅6H2O (10 mmol) in absolute MeOH (10 ml) was added dropwise. Upon addition, immediate precipitation of Zn2L2(NO3)2 complex occurs. The resulting solution was stirred for 2 h in ambient temperature.

The yellow powder was isolated from the solution and purified by washing with cold methanol.

Appropriate single crystals for X-ray crystallography were obtained in methanol by

Single crystal diffraction method

The X-ray diffraction measurement for Zn(II) complex, Zn2L2(NO3)2, was made on a STOE IPDS-II diffractometer with graphite monochromated Mo Kα radiation. A yellow small crystal was mounted on a glass fiber and used for data collection. Data were collected at a temperature of 298 K and in a series of ω scans in 1° oscillations and integrated using the Stoe X-AREA software package [START_REF] Stoe | Program for the Acquisition and Analysis of Data XeAREA[END_REF]. A numerical absorption correction was applied using X-RED [START_REF] Stoe | Program for Data Reduction and Absorption Correction Xe[END_REF] and X-SHAPE [START_REF] Stoe | Program for Crystal Optimization for Numerical Absorption Correction Xe[END_REF] softwares. The data was corrected for Lorentz and Polarizing effects. Structure was solved by direct methods using SIR2004 [START_REF] Burla | [END_REF]. The non-hydrogen atoms were refined anisotropically by the full matrix least squares method on F 2 using SHELXL [28]. All the hydrogen (H) atoms were placed at the calculated positions and constrained to ride on their parent atoms. Crystallographic data are listed in Table 1. Selected bond distances and angles are summarized in Table S1.

DNA binding studies

The stock solution of FS-DNA was prepared in 50 mM Tris buffer at pH 7.5 using double-distilled deionized water and stored at 4 °C. The FS-DNA concentration was determined by the absorption intensity at 260 nm and the molar absorption coefficient 6600 M -1 cm -1 . Purity of FS-DNA solution was confirmed by ratio of UV absorbance at 260 and 280 nm (A260/A280 = 1.9), indicating that FS-DNA is free from protein impurity [29], [30]. A stock solution of the Zn(II) complex was prepared by dissolving the complex in an aqueous solution of DMF as the co-solvent and then diluted suitably with the corresponding buffer to the required concentrations. The final DMF concentration never exceeded 0.3% v/v. The UV-Vis spectral features of dinuclear Zn(II) complex did not change on keeping its buffered or DMF solution for 24 h and no precipitation or turbidity was observed even after long storage at room temperature (at least 4 weeks after preparation), which indicates the stability of the Zn(II) complex in different media. UV-Vis absorption spectra of Zn(II) complex (2 × 10 -5 M) in absence and presence of various amounts of DNA (0-67 μM)

were recorded. All complex-DNA solutions were allowed to incubate for 5 min before recording the absorption spectra and then the absorptions were recorded to calculate the intrinsic binding constant, Kb of the dinuclear Zn(II) complex to FS-DNA.

Furthermore, fluorescence quenching experiments carried out using quartz cuvette with 1 cm optical path length and the excitation and emission slits set at 5 and 10 nm, respectively. To this aim, the FS-DNA solution was stirred with EtBr with molar ratio of DNA:EtBr 10:1 for 1 h at 4 °C. Then, various amounts of Zn(II) complex (0-45 μM) were added to this mixture. The fluorescence spectra were measured in the range of 550-650 nm with exciting wavelength at 520 nm. In each measurement after addition of FS-DNA, the mixture was allowed to stand for 5 min.

HSA Binding experiments

A stock solution of HSA was prepared by dissolving the desired amount of HSA in 50 mM phosphate buffer (pH = 7). The HSA stock solution was stored at 4 °C in the dark and used within 2 h. HSA concentration was determined by UV-Vis spectrophotometry using the molar absorption coefficient 35700 M -1 cm -1 at 278 nm [31]. UV-Vis spectra of dinuclear Zn(II) complex were recorded with addition of successive amount of HSA with molar ratios of the [HSA]/[Complex] = 0-4.5 at room temperature. Fluorescence quenching experiment was also done to more investigation of interaction between HSA and Zn(II) complex. In this experiment 2 mL of HSA solution (4 μM) was placed into the cell and various amounts of 100 μM dinuclear Zn(II) complex were added to the cell. The fluorescence intensity was measured with excitation wavelength at 295 nm and emission wavelength rang of 300-370 nm. As mentioned above, the mixture was allowed to incubate for 5 min after addition of HSA or complex.

Molecular dynamics simulation

The known crystal structures of DNA (PDB ID: 423D) with sequence d(ACCGACGTCGGT)2 and HSA (PDB ID: 1BM0) were taken from the Brookhaven Protein Data Bank (www.rcsb.org). A 10 ns MD study was carried out using GROMACS4.5.6 package [32], [33] and GROMOS96 43a1

force field [START_REF] Van Gunsteren | Biomolecular Simulation: The GROMOS96 Manual and User Guide[END_REF], [START_REF] Van Gunsteren | Encyclopedia of Computational Chemistry[END_REF] for DNA and HSA. The topology parameters of DNA and HSA were generated using gromacs. The macromolecules were located in a cubic box with the periodic boundary conditions. The SPC water model [START_REF] Berendsen | Intermolecular Forces[END_REF] was employed for the water molecules. The sufficient amounts of sodium ions were added to balance the charge of system. The system was energy-minimized using the steepest descent method for the 4000 time-step [START_REF] Hess | [END_REF]. Next, position restraint procedure was performed in NVT and NPT ensembles with time duration of 200 ps. The system was equilibrated at constant temperature of 300 K and constant pressure of 1.0 bar. Finally, a 10 ns MD simulation was carried out at 1 bar and 300 K. The Particle-Mesh Ewald (PME) method [38], [39] and the cut-off method were used for the long-range electrostatic interactions and the van der Waals interactions, respectively. A 7 Å cutoff for the van der Waals interactions was used. The equation of motion was integrated by the leap-frog algorithm with the 2 fs timestep and the atomic coordinates were recorded to the trajectory file every 0.5 ps for further analysis.

The stability of simulated systems was scanned using the root-mean-square deviations (RMSDs)

of HSA and DNA with respect to their initial structures. The average RMSD values of the HSA and DNA backbones were 8.21 nm and 5.39 nm, respectively. The equilibrated conformation of the HSA and DNA were used for docking.

Docking procedure

Docking study was carried out to indicate the active binding site for dinuclear Schiff base Zn(II) complex. The 3D structure of the metal complex was generated using the CIF file of its X-ray crystal structure. The CIF file was converted to the PDB format by using the Mercury software.

The equilibrated conformations of the HSA and DNA in MD simulation were used for docking and missing hydrogen atoms were added at appropriate geometry. Flexible-ligand docking was performed by AutoDock 4.2 molecular-docking program using the implemented empirical free energy function and the Lamarckian Genetic Algorithm [40]. The Gasteiger charges were added to prepare the macromolecule input file for docking and the Auto Grid was used to calculate Grids.

A grid map with 90 × 90 × 90 points and a grid-point spacing of 0.375 Å was applied in all dockings. 200 docking runs with 25,000,000 energy evaluations for each run were performed.

Result and discussion

IR spectra

The comparison of IR spectrum of ligand with complex is an evidence for the coordination of metal to ligand. The sharp band assigned to imine group is observed at 1620 cm -1 for free ligand and at 1623 cm -1 for binuclear complex.

The absorption band at 1367 cm -1 and 1472 cm -1 in the spectrum of free ligand can be assigned to C O and C N stretching of the ligand respectively. However, after coordination of the metal ion to the ligand via oxygen and nitrogen, these bands shifted to 1336 cm -1 and 1455 cm -1 .

Furthermore, elimination of the stretching frequency of O H group after complexation is result of deprotonation of the ligand [41].

NMR study

1 H NMR and 13 C NMR spectra of schiff base ligand and Zn2L2(NO3)2 complex were measured in CDCl3 and DMSO, respectively. The structure of Zn2L2(NO3)2 is symmetric, therefore 1 H NMR and 13 C NMR of two coordinated ligands is identical.

The singlet peak due to the imine proton in the ligand was observed at 8.9 ppm which shifted to 9.5 ppm for Zn2L2(NO3)2 complex. The resonance signal observed at 13.13 ppm assign to OH group that is not observed in the 1 H NMR spectrum of complex. It confirms that the hydroxyl groups are fully deprotonated and the oxygen acts as a coordination atom. In 13 C NMR of schiff base ligand, the peak at 175.1 ppm assignable to the imine carbon atoms, confirms the presence of the Schiff base ligand in the complexes [41]. The 1 H NMR spectrum of complex shows a peak at 5.3 ppm assigned to H5 (Fig. S1). Also, aromatic protons of Zn(II) complex appear in the region of 6.9-8.5 ppm (see Fig. 1).

Fig. 1. Molecular structure of HL showing the lettering scheme for NMR assignments.

X-ray structure

The crystal structure of Zn2L2(NO3)2 consists of a discrete centro-symmetric dimeric unit (Fig. 2).

Selected bond length sand angles are summarized in Table S1 in complex involving bidentate nitrate [42].

As we know, non-covalent interactions are at the core of most chemical and biological processes and hence knowledge of their nature, strength, occurrence and consequences is of paramount importance [43]. The weak non-covalent interactions such as C H⋯X (X = O, N, S, halogen) and π-π stacking, while weaker than the classical H-bonds, also play notable roles in conformation, crystal packing, supramolecular assembly and physicochemical properties and thus have implications in drug design, material design and supramolecular synthesis [43], [44], [45], [46], [47], [48], [49].

Although there are not classical hydrogen bonds in the structure, but both uncoordinated oxygen atoms of the nitrate ions are involved in C H⋯O non-classical hydrogen bonds (Fig. 3). Nonclassical intermolecular hydrogen bond occurs between C(6) H( 6)⋯O(4) [2.58 Å] atoms and C( 16) H( 16)⋯O(2) [2.49 Å] atoms (Fig. 3).

Fig. 3. Intramolecular hydrogen bond geometries (Å, °).

Another interesting feature of the Zn2L2(NO3)2 complex is the presence of C H⋯π and π⋯π interactions (Fig. 4, Fig. 5). Weak C H⋯π edge-to-face interactions are present between CH of Schiff base group with H⋯π distance of 2.67 Å for C(12) H(12B)⋯Cg (9) [Cg( 9) is centroid for C(4)/C(9) ring] (Fig. 4). Also, in the crystal packing π⋯π interactions are exist between the naphthalene ring and pyridine ring of neighbor molecule, Cg(7)⋯Cg( 9), with centroid-centroid distance of 3.5498( 14), 3.5082(12) Å, in which Cg (7) and Cg( 9) are centers of N(2) C( 13) C( 14) C( 15) C( 16) C( 17) and C(4) C( 5) C( 6) C( 7) C( 8) C( 9) rings, respectively (Fig. 5, Scheme 3).

DNA binding studies

DNA interactions play an important role in the field of medicinal chemistry, since it is the primary intracellular target of anticancer drugs. The interaction between small molecules and DNA can cause DNA damage in cancer cells, blocking the division of cancer cells, and resulting in cell death. It includes conformational and structural changes of DNA molecules which play an important role in the biological systems. Thus, to further demonstrate the mechanism of anticancer action of the dinuclear Zn(II) complex, its interaction with FS-DNA was analyzed by spectroscopic techniques.

UV-Vis absorption

Recording UV-Visible spectrum of metal complex upon addition of various amount of DNA is a powerful technique to investigate DNA binding of complex. Electronic absorption spectra can give the information regarding the electronic perturbations in the complex made by interaction with DNA [50]. Generally, hyperchromic or hypochromic effect and red or blue shift are observed in the UV-Vis spectra of drugs upon binding of drugs to DNA. Hypochromic and red shift are indicative of intercalation mode [51]. Intercalation mode is interaction between π * -orbital of complex with π-orbital of DNA base pairs. Therefore, energy level of π * -orbital of complex decreases which causes red shift in UV-Vis spectrum of complex. Furthermore, the coupled π *orbital is filled and so the probability of electron transition decreased and hypochromic is observed [51], [52]. On the other hand, hyperchromic effect in spectrum of drug is due to damage of the DNA double-helix structure [53]. This change in spectrum can be attributed to groove binding or electrostatic interaction. The minor or major groove binding causes damage to DNA at the exterior phosphate backbone [53], [54].

The absorption spectrum of the synthesized Zn(II) complex indicates two absorption bands attributed to π → π * and n → π * at 305 and 380 nm. The n → π * absorption band was chosen to determine the interaction of this complex with DNA. Through addition of various amount of FS-DNA (1.5 × 10 -4 M) with molar ratio of [DNA]/[Complex], 0-6, a hypochromic effect accompanied by red shift in spectrum of complex is observed (Fig. 6).

Fig. 6. Electronic absorption spectra of Zn(II) complex upon addition of various amounts of DNA.

[Complex] = 2 × 10 -5 M, [DNA] = 1.5 × 10 -4 M.

Although, observation a hypochromic effect in UV-Vis spectrum with increase amount of DNA reveals an intercalative mode, some studies reported that this is not a correct conclusion necessarily. The hypochromic effect can be observed as a result of other binding mode such as groove binding due to the changes in the solvent distribution and the orientation effect in moving into the more hydrophobic environment of groove [55], [56]. The crystal structure of the complex (Fig. 2) shows that this complex is not planner. Hence, probability of intercalation for this complex is very low. This spectral characteristic suggests that this complex interacts with FS-DNA through the partial insertion of the ligand aromatic rings to the FS-DNA groove. Moreover, the small redshift may be attributed to the groove binding affinity of the complex which is due to the presence of a phenyl ring in the coordinated ligand [57].

To assess the binding ability of the dinuclear Zn(II) complex with FS-DNA, the intrinsic binding constant (Kb) was determined using following equation [21]:

where The obtained data are lower than the mentioned Kb for classical intercalators (ethidium bromide and [Ru(phen)DPPZ]) that their binding constants are in the order of 10 6 -10 7 M -1 [59]. Therefore, the synthesized Schiff base complex can be bind to FS-DNA via groove binding.

Fluorescence quenching experiment

EtBr emits only a weak fluorescence emission in solution due to hydrogen transfer from one of amino group of EtBr into solution which causes nonradiative decay, while a significant increase of fluorescence intensity of EtBr is observed with the addition of FS-DNA. Since the EtBr molecules can be intercalated into the double helix of DNA [60], [61], [62], [63]. In the present study, quenching fluorescence of DNA-EtBr by addition of Zn(II) Schiff base complex is measured (Fig. 7). The absorptions of the dinuclear Zn(II) complex at the excitation and emission wavelengths were approximately zero in all concentrations. Hence, a reduction in the emission intensity is independent of the inner filter effect. However, this complex cannot intercalate to the DNA duplex, but it can displace EtBr in a non-competitive manner by changing the DNA conformation. Consequently, the DNA-bound EtBr molecules are converted to their free form in solution. In order to determine the binding ability between the complex and FS-DNA, the Stern-Volmer quenching plot was obtained by monitoring the fluorescence quenching of the EtBr-DNA with increasing the concentration of the Zn(II) complex according to the Stern-Volmer equation [64]:

where F0 and F are the fluorescence intensity of FS-DNA in absence and presence of Zn(II) complex, and Ksv is the Stern-Volmer quenching constant, Kq is the quenching rate constant of DNA and τ is the average lifetime of DNA without quencher which is typically equal to 10 -8 s for biomacromolecules [65]. Ksv is determined from the plot of F0/F vs [Q]. In the present study, Ksv was obtained 1.9 × 10 4 M -1 (Fig. 8). The value of Ksv shows that this complex has high quenching efficiency and a significant degree of binding to FS-DNA. Fluorescence quenching is classified to two mechanisms: static quenching and dynamic quenching. In the static mechanism, the fluorophore and the quencher collide together in the ground state while fluorophore and quencher collide together in the excited state in dynamic mechanism [65]. Linearity of the Stern-Volmer plot indicates that quenching fluorescence has only one mechanism, dynamic or static [66].

In this study, the value of Kq was obtained 1.9 × 10 12 M -1 S -1 that is than limiting diffusion rate constant of the diffusional quenching for biopolymers (2 × 10 10 M -1 S -1 ). This observation supports that quenching fluorescence of DNA occurs by static mechanism [65]. Moreover, binding constant (Kb) of the Zn(II) complex and FS-DNA can be calculated by this experiment using following equation [65]:

where F0 and F are the fluorescence intensity of FS-DNA in absence and presence of Schiff base complex, respectively. [Q] is the concentration of quencher that quencher is Zn(II) Schiff base complex here. "Kb" is obtained from the plot of Ln [Q] vs Ln�0-�� as a slope. Furthermore, "n" which is the number of binding site per nucleic acid is y-intercept of the plot. The intrinsic binding constant (Kb) of Zn(II) Schiff base complex and FS-DNA is 3.9 × 10 4 M -1 and (n) is 0.72 (Fig. S2). This result is in good agreement with the UV-Vis spectroscopy result. Also, Eq. ( 4) can be used to investigate the apparent binding constant (Kapp).

where [complex]50 is the concentration of complex that makes a 50% reduction of the initial fluorescence intensity of the EtBr-DNA system, �EthBr = 1 × 10 7 M -1 is the binding constant of EtBr with DNA and [EtBr] = 1 × 10 -5 M [67]. The apparent binding constant was obtained as 2.2 × 10 6 M -1 which is lower than the binding constant of classical intercalators and metallointercalators [68].

HSA binding studies

The drug-HSA interactions in the blood stream can influence the chemical stability, toxicity, and distribution of the drug. These interactions significantly influence the pharmacokinetics and pharmacodynamics of drugs. Consequently, it is important to know the binding affinity of HSA for drugs. Hence, the interaction of the dinuclear Zn(II) complex with HSA was investigated using fluorescence quenching and UV-Vis absorption spectroscopy.

Electronic absorption spectroscopy

UV-Vis absorption spectroscopy is an effective technique to study of HSA-drug binding. The titration of complex with increasing amount of HSA with the molar ratio of [HAS]/[Complex] = 0-4.5 was carried out and its absorption spectra were measured (Fig. 8). Eq. ( 1) was applied to obtain constant binding of HSA and dinuclear Zn(II) complex. All of calculations and definitions are similar to DNA binding study. Constant binding was calculated as 1 × 10 4 M -1 .

Fluorescence spectroscopy

Fluorescence of HSA protein is originated from three amino acids, i.e. tryptophan (Trp), tyrosine (Tyr), and phenylalanine (Phe) residues. Fluorescence is a useful approach for investigating intermolecular interactions because of the photophysical character of the fluorophoreis sensitive to the polarity of its surrounding environment [69].

Decrease of fluorescence intensity through the addition of complex can confirm the binding of dinuclear Zn(II) complex to HSA [70]. [HSA] = 4 × 10 -6 M, λex = 295 nm.

The binding constant (Kb) of the metal complex and HSA can be calculated using Eq. (3).

The Kb value is calculated as 1.7 × 10 4 M -1 and n is determined as 1.1 (Fig. S3).

Energy transfer between dinuclear Zn(II) complex and HSA

Fluorescence resonance energy transfer (FRET) also known as Förster's resonance energy transfer is an interaction between the excited molecule and its adjacent molecule. Upon this interaction, non-radiative energy transfer take places from a donor molecule in excited state to an acceptor in ground state [71], [72]. According to this theory, Three conditions are required to energy transfer:

(1) the donor should have fluorescence, (2) the fluorescence emission spectrum of the donor and the UV-Vis spectrum of the acceptor should have sufficient overlap and (3) the small distance between donor and acceptor (<8 nm) [72]. The distance and efficiency of energy transfer (E) between tryptophan residue of protein (HSA) and drug (complex) has been calculated using this theory through the following equation:

where F0 and F are fluorescence intensities of HSA in the absence and presence of complex, respectively and R0 is the critical distance when the transfer efficiency is 50%; r is the distance between donor and acceptor. R0 can be calculated by Eq. ( 6) [73]:

In the above equation, the term K 2 is the orientation factor of the dipoles; is the refracted index of medium, J is the overlap integral of the fluorescence spectrum of the donor with absorption spectrum of the acceptor and φ is the fluorescence quantum yield of the donor. The value of J can be calculated by the following expression:

where F(λ) is the fluorescence intensity of the donor in the absence of the acceptor at wavelength λ and ɛ is the molar absorption coefficient of the acceptor at λ. In the present case, K 2 = 2/3, N = 1.336 and φ = 0.15 for HSA. Therefore, according to Eqs. ( 5), ( 6), (7) and Fig. S4, J = 3.6 × 10 -14 cm 3 L mol -1 , E = 0.36, R0 = 3.17 nm and r = 3.6 nm. The value of r is less than 8 nm and 0.5R0 < r < 1.5R0, suggesting energy transfer from HSA to dinuclear Zn(II) complex occurs with high probability. Furthermore, the value of r is higher than R0 and it indicates the presence of a static quenching mechanism in the interaction of HSA and complex [74].

Molecular docking of the Zn complex with DNA

The Zn(II) complex was docked to the best conformer of DNA obtained from the MD simulation.

The results revealed that the Zn complex fitted into the DNA major groove with the binding energy of -7.7 kcal mol -1 , indicating a high binding affinity between DNA and the complex (Fig. 10).

This result is in consistent with UV-Vis spectroscopy and fluorescence quenching results. There are four categories of hydrophobic contacts between the complex atoms and bases of DNA, including, DC 9, DG 10, DG 11 and DC 15 (Fig. 11). Furthermore, complex forms a hydrogen bond with DC 14, thus this nucleotide plays a significant role in stabilizing of Zn(II) complex interactions.

Molecular docking of the Zn complex with HSA

HSA is the most abundant protein in human blood plasma and consists of 585 amino acids, with three α-helical domains (I, II and III), each containing two subdomains (A and B) [75]. HSA transports drugs, hormones, fatty acids, and other compounds, and maintains osmotic pressure, among other functions [76], [77]. The previous crystal structure studies showed that most ligands are binded to one of the two binding sites in subdomains IIA and IIIA [78], [79], [80], [81], [82].

Also, D-shaped cavity in subdomain IB is the binding site of some compounds [83], [84], [85].

The Zn(II) complex was docked to the best conformer of HSA obtained from the MD simulation.

The best energy-ranked result of the interaction between complex and protein in all runs of docking procedure is shown in Fig. 11. As seen Zn(II) complex was situated within subdomain IIA in site 1 with the binding energy of -11.14 kcal mol -1 . Furthermore, this compound was adjacent to some hydrophobic residues in subdomain IIA of HSA (Ala 210, Ala 213, Val 482 and Leu 481). On the other hand, complex was able to form a π-cation interaction with Lys 286, thus this residue plays a significant role in stabilizing of Zn complex interactions (Fig. S5).

Conclusion

In this study, a dinuclear Zn(II) Schiff base complex, Zn2L2(NO3)2, was synthesized from reaction of Zn(NO3)2 and a tridentate ligand, HL, at ambient temperature. The ligand and complex were characterized by FT-IR, 1 H NMR, 13 C NMR and elemental analysis (CHN). Furthermore, structure of Zn(II) Schiff base complex was determined by single-crystal X-ray analysis. The complex, Zn2L2(NO3)2, is centrosymmetric dimer in which deprotonated phenolates bridge the two Zn(II) atoms and link the two halves of the dimer. In structure, the Zinc(II) ions have a highly distorted six-coordinate structure, bonded to two oxygen atoms of a bidentate nitrate group, the nitrogen atom of pyridine, the nitrogen atom of imine group and the two oxygen atoms of two phenolate groups. Moreover, DNA and HSA binding of complex were investigated using fluorescence quenching, UV-Vis spectroscopy, molecular docking and molecular dynamics simulation methods. Fluorescence quenching and UV-Vis spectroscopy results represent that dinuclear Zn(II) complex binds to DNA and HSA with binding constants of (3.60 ± 0.18) × 10 4 M -1 and (1.35 ± 0.24) × 10 4 M -1 , respectively. Molecular docking studies revealed the binding of dinuclear Zn(II) complex to the major groove of DNA and IIA site of HSA by formation of hydrogen bond, π-cation and hydrophobic interactions. These results reflect the strong interaction with DNA and HSA which is in consistent with the fluorescence quenching UV-Vis spectroscopy.

  supplementary. The dinuclear unit is formed by two Zn(II) atoms labeled Zn(1) and Zn(1′) (′ = -x, -y + 1, -z), bridged by the two phenoxo oxygen atoms O(1) and O(1′) of the Schiff-base ligands, L. Each of the two equivalent zinc atoms presents a distorted octahedral environment formed by the deprotonated chelate Schiff base ligand, a bidentate nitrate anion and a phenoxo oxygen atom from the symmetry related Schiff base ligand (Fig.2). The Schiff base ligand coordinates the Zn(II) ion in facial (fac) configuration through the imine nitrogen N(1), the nitrogen atom of pyridine N(2), and the phenoxo oxygen atom O(1) with usual phenoxo-bridged bond distances (TableS1in supplementary)[42]. Note that the double oxo bridge connecting both Zn(II) ions is slightly asymmetric because each Zn(II) ion is closer to its own phenoxo oxygen atom (Zn(1) O(1) = 2.0188(18) Å) than to the phenoxo oxygen atom of the symmetry related Schiff base (Zn(1) O(1′) = 2.0611(16) Å). The two Zn atoms are separated by 3.1428(7) Å and the Zn O(1)Zn′ angle is 100.76(7)°.

Fig. 2 .

 2 Fig. 2. ORTEP representation of Zn2L2(NO3)2. Displacement ellipsoids are drawn at the 50% probability level and H atoms are shown as small spheres of arbitrary radii.

Fig. 4 .

 4 Fig. 4. Analysis of X H⋯Cg (Pi-Ring) Interactions (H⋯Cg < 4.0 Å).

Fig. 5 .

 5 Fig. 5. Aromatic interaction parameters (Å and °) for description of π-π interactions.

  [DNA] is the concentration of FS-DNA; ��, �� and �� are the apparent extinction coefficient, the extinction coefficient for free metal and the extinction coefficient for the complex in fully bound form, respectively. �� was determined by calibration curve and �� is the ratio of Aobs to[Complex]. A plot of [DNA] vs [DNA]��-�� gives Kb as ratio of slop to y-intercept.The binding constant of FS-DNA and dinuclear Zn(II) complex is 3.3 × 10 4 M -1 which is comparable to obtained data for the other dinuclear Zn(II) complexes.Patel et al. reported the Kb value for synthesized homo dinuclear Zinc complexes and DNA in the range of (1-3.5) × 10 4 M -1[58].

Fig. 7 .Fig. 8 .

 78 Fig. 7. Emission spectra of the EtBr-DNA system in the absence and presence of the various concentrations of Zn(II) complex. [DNA] = 10 -4 M, [EtBr] = 10 -5 M,

Fig. 9 Fig. 9 .

 99 Fig. 9. Changes in the fluorescence spectra of HSA in the presence of Zn(II) complex.

Fig. 10 .

 10 Fig. 10. The docking pose of Zn complex-DNA (the small green spheres show H-bond interaction).

Fig. 11 .

 11 Fig. 11. (Left): Presentation of the interaction of Zn(II) complex with the major groove side of DNA. (Right): The docking pose of Zn(II) complex-HSA (π-cation interaction showed as yellow cone).

  88%, Anal. Calc. for C17H14N2O: C, 77.84; H, 5.38; N, 10.68. Found: C, 77.51; H, 5.71; N, 10.23.

	FT-IR (KBr, cm -1 ): (O	H) 3365 (br, vs); (C	N) 1620 (vs). 1 H NMR (CDCl3, 400 MHz,
	298 K) δ/ppm:13.13 (1H, s, OH),8.9 (s, 1H, N	CH),8.5 (d, 1H, Ar), 8.2 (d, 1H, Ar), 7.9 (t, 1H,
	Ar), 7.7 (d, 1H, Ar), 7.6 (d, 1H, Ar), 7.5 (t, 1H, Ar), 7.4 (t, 1H, Ar), 7.2 (t, 1H, Ar), 7.0 (d, 1H, Ar),

6.9 (10H, Ar), 4.8 (s, 2H, CHH).

13

C NMR (CDCl3, 400 MHz, 298 K) δ/ppm: 175.1, 164.8, 158.0, 149.4, 139.1, 137.1, 129.4, 129.1, 128.0, 124.4, 124.0, 122.9, 121.9, 119.0, 118.4, 118.1, 58.3.

Table 1 .

 1 Crystal data and structure refinement for

Zn2L2(NO3)2.

	Empirical formula	C34H26N6O8Zn2 0 ⩽ l ⩽ 18
	Formula weight Reflections collected	777.35 10,844
	Temperature Independent reflections	298(2) K 2895 [R(int) = 0.0327]
	Wavelength Data completeness	0.71073 Å 100.0%
	Crystal system Refinement method	Orthorhombic Full-matrix least-squares on F 2
	Space group Data/restraints/parameters	Pbca 2895/0/226
	Unit cell dimensions Goodness-of-fit on F 2	a = 14.517(3) Å 0.805
	Final R indices [I > 2σ (I)]	b = 14.626(3) Å R1 = 0.0280
		c = 15.449(3) Å wR2 = 0.0471
	Volume R indices (all data)	3280.1(11) Å 3 R1 = 0.0675
	Z	4 wR2 = 0.0520
	Density (calculated) Largest diff. peak and hole	1.574 Mg/m 3 0.136 and -0.255 e Å -3
	Absorption coefficient	1.525 mm -1
	F(0 0 0)	1584
	Theta range for data collection	2.38-25.00°
	Index ranges	-17 ⩽ h ⩽ 17
		-17 ⩽ k ⩽ 17

Acknowledgement

The authors are grateful to the Research Council of the University of Isfahan for financial support of this work.

Appendix A. Supplementary material CCDC 1042425 for Zn2L2(NO3)2 contains the supplementary crystallographic data for this complex. Copies of the data can be obtained, free of charge, on application to CCDC, 12 Union Road, Cambridge CB21EZ, UK (Fax: +44 1223 336033 or e-mail: deposit@ ccdc.cam.ac.uk). Supplementary data associated with this article can be found, in the online version, at http://dx.doi.org/10.1016/ j.molstruc.2015.04.033.


Mahsa Khorshidifard

Amiri Hadi

Banafshe Rudbari

Mehdi Askari

Mostafa Riahi Sahihi

Fariba Farsani

Giuseppe Jalilian

Bruno

Hadi Amiri Rudbari

Banafshe Askari

Mehdi Sahihi

Riahi Farsani

Fariba Jalilian

Giuseppe Bruno

Cobalt(II), copper(II), zinc(II) and palladium(II) Schiff base complexes: Synthesis, characterization and catalytic performance in selective oxidation of sulfides using hydrogen peroxide under solvent-free conditions

Keywords:

ou non, émanant des établissements d'enseignement et de recherche français ou étrangers, des laboratoires publics ou privés.

Introduction

The selective oxidation of organosulfur compounds has attracted much attention from the viewpoint of organizing chemical processes for the preparation of synthetically useful sulfoxides, and sulfones, which are important and versatile intermediates in the synthesis of natural products and biologically significant molecules [1], [2], ligands in asymmetric catalysis [3] and oxo-transfer reagents [4], [5]. Among a lot of different oxidants, aqueous hydrogen peroxide has been one of the most noteworthy "green oxidants", offering the advantages of safety in storage and operation, being cheap, readily available, of highly effective oxygen content and environmentally benign, with the formation of water as the only by-product [6]. Different catalysts, such as polyoxometallates and transition-metal Schiff-base complexes, have been used for H2O2-based oxidation of sulfides [7], [8], [START_REF] Sheldon | Metal Catalyzed Oxidations of Organic Compounds[END_REF], [START_REF] Afrasiabi | [END_REF], [11], [12], [13], [14], [15], [16], [17], [START_REF]The Porphyrin Handbook[END_REF], [19], [20]. However, the oxidation of sulfides to sulfones has been much less investigated as compared to the oxidation of sulfides to sulfoxides. Also, many of these systems suffer from one or more limitations, such as high cost, the requirement of a promoter or a co-catalyst, high temperature, long reaction time, chlorohydrocarbon solvents and excessive H2O2. These defects are becoming more apparent in view of the developing environmental concerns in recent years [7], [START_REF] Afrasiabi | [END_REF], [11]. In addition, most of the catalysts are required to use a solvent in the reaction, while it is highly favorable to expand a process without a solvent from the viewpoint of green chemistry.

Schiff base ligands have played an important role in the development of coordination chemistry, especially their metal complexes exhibit wide applications in biological and industrial systems [START_REF] Holm | [END_REF], [22], [23], [24], [25], [26]. Furthermore, Schiff bases are very important tools for inorganic chemists, as they are widely used to design molecular ferromagnets, in catalysis, in biological modeling applications, as liquid crystals and as heterogeneous catalysts [27], [28], [29]. Unfortunately, application of Schiff base complexes as a catalyst for the solvent-free oxidation of sulfides with hydrogen peroxide is very rare [30], [31], [32], [33].

So current demand for environmentally friendly processes requires the synthesis of new catalysts and the development of green oxidation methods that use clean oxidants, such as hydrogen peroxide, without any solvent. As part of our research program on the synthesis, characterization and catalytic activity of Schiff base ligands and complexes [34], [35], [36], [37], in this paper, we synthesized a bidentate Schiff base ligand, HL, derived from salicylaldehyde and tertbutylamine (Scheme 1). In order to investigate the coordination modes of this ligand, cobalt(II), copper(II), zinc(II) and palladium(II) complexes were synthesized (Scheme 2). After synthesis and characterization of the complexes, we used them in the no-solvent oxidation of sulfides with hydrogen peroxide as a green oxidant. This process is clean, safe, selective, simple and costeffective for the oxidation of sulfides to the corresponding sulfones. Scheme 1. Synthesis of the ligand (HL). Scheme 2. Synthetic routes for the preparation of the complexes.

Experimental section

Chemicals and instrumentation

All the chemicals were purchased from Merck Co. and used without further purification. The FT-IR spectra were recorded on a JASCO, FT/IR-6300 spectrometer (4000-400 cm -1 ) in KBr pellets. 1 H and 13 C NMR spectra were recorded on a Bruker Avance 400 spectrometer using CDCl3 (for the palladium(II) complex) and DMSO-d6 (for the ligand and zinc(II) complex) as solvents. The elemental analysis was performed on Leco, CHNS-932 and Perkin-Elmer 7300 DV elemental analyzers. UV-Vis spectra were recorded on a JASCO V-670 UV-Vis spectrophotometer (200-700 nm). The oxidation products were quantitatively analyzed by gas chromatography (GC) on a Shimadzu GC-16A instrument using a 2 m column packed with silicon DC-200 and an FID detector.

Single crystal diffraction studies

X-ray data for complexes, CoL2, CuL2, ZnL2 and PdL2, were collected on a STOE IPDS-II diffractometer with graphite monochromated Mo Kα radiation.

For CuL2, CoL2, ZnL2 and PdL2 the crystals were dark green, red, yellow and light red, respectively. A high-quality piece of crystal was chosen in each case using a polarizing microscope and they were mounted on a glass fiber, then used for data collection. Cell constants and an orientation matrix for data collection were obtained from the least-squares refinement of diffraction data from 5922 for CuL2, 3108 for CoL2, 3501 for ZnL2 and 2450 for PdL2 unique reflections. Data were collected at a temperature of 298(2) K in a series of ω scans in 1° oscillations and integrated using the Stöe X-AREA software package [START_REF]Version 1.30, Program for the Acquisition and Analysis of Data[END_REF]. A numerical absorption correction was applied using the X-RED [39] and X-SHAPE [39] software packages. The data were corrected for Lorentz and Polarizing effects. The structures were solved by direct methods using SIR2004 [START_REF] Burla | [END_REF]. The non-hydrogen atoms were refined anisotropically by the full matrix least squares method on F 2 using SHELXL [41]. All hydrogen atoms were added at ideal positions and constrained to ride on their parent atoms.

In the crystal structures of CoL2 and ZnL2, the C9, C10 and C11 carbon atoms of the tert-butyl group were disordered over two sites and refined with site occupancy factors 0.67:0. -O). 1 H and 13 C NMR (DMSO-d6, 400 MHz, 298 K): Table 3.

Preparation of the complexes

General procedures for the preparation of CoL2, CuL2, ZnL2 and PdL2.

A MeOH solution (20 cm 3 ) of tert-butylamine (2 mmol) was added dropwise to a MeOH solution (20 cm 3 ) of the salicylaldehyde (2 mmol). The yellow solution was stirred for 2 h at ambient temperature, then a solution of triethylamine (3 mmol) in absolute MeOH (5 cm 3 ) was added to the solution. The solution turned dark yellow and was stirred for 10 min, then a solution of the appropriate metal salt (1 mmol) in absolute MeOH (20 cm 3 ) was added dropwise. The resulting solution was stirred for 12 h at ambient temperature for the preparation of the CoL2, CuL2 and ZnL2 complexes, while for the palladium(II) complex, PdL2, the solution was refluxed overnight.

Cobalt(II) complex (CoL2)

The red solution was slowly evaporated to dryness at room temperature to yield a red solid which was purified by washing with 50 ml of diethyl ether several times and the obtained green precipitate was dried in air. Appropriate single crystals for X-ray crystallography were obtained directly from the reaction mixture. The typical yield was 62%. Anal. Calc. for C22H28CoN2O2: C, 64.23; H, 6.86; N, 6.81. Found: C, 64.18; H, 7.00; N: 6.89%. Selected IR data (KBr, cm -1 ):

3056-2900 (w, C-H aliphatic and aromatic), 1601 (νC N), 1174 and 1145 (νC-O).

Copper(II) complex (CuL2)

The resultant red solution was allowed to stand overnight. After concentration at room temperature, a red precipitate was collected by filtration. Appropriate single crystals for X-ray crystallography were obtained directly from the reaction mixture.

Zinc(II) complex (ZnL2)

The yellow solution was slowly evaporated to dryness at room temperature to yield a yellow solid which was purified by washing with 50 ml of diethyl ether about five times and the obtained yellow precipitate was dried in air. Appropriate single crystals for X-ray crystallography were obtained directly from the reaction mixture. 1174 and 1148 (νC-O). 1 H and 13 C NMR (DMSO-d6, 400 MHz, 298 K): Table 3.

Palladium(II) complex (PdL2)

A green precipitate was collected by filtration and purified by washing with 50 ml of diethyl ether several times and then dried in air. 1 H and 13 C NMR (CDCl3, 400 MHz, 298 K): Table 3.

Catalytic activity and optimization of the reaction conditions

Oxidation of thioanisole with H2O2 catalyzed by CoL2, CuL2, ZnL2 and PdL2 Schiff base complexes

To find the optimized conditions, we investigated the oxidation of thioanisole as a model substrate using hydrogen peroxide (Scheme 3). The effect of different reaction parameters (H2O2 amount, temperature and mmol of catalyst) were studied for the thioanisole oxidation.

General procedure for the catalytic oxidation of sulfides under solvent free conditions

To a mixture of the sulfide (1 mmol) and 30% H2O2 (2 mmol), the catalyst (1 mmol) was added and the mixture was stirred at 50 °C for a specified time. The progress of the reaction was monitored by TLC (petroleum ether/ethylacetate 8:3) and GC. After completion of the reaction, the product was extracted with ethyl acetate and the catalyst was separated by filtration. The combined organics were washed with brine (5 ml) and dried over anhydrous Na2SO4. Further purification was achieved by short-column chromatography on silica gel with EtOAc/n-hexane as the eluent.

Results and discussion

The Schiff base ligand HL, obtained by the self-condensation reaction between tert-butylamine and salicylaldehyde in absolute methanol as solvent at ambient temperature and reported in Scheme 1, is a yellow oil, stable in air and moderately soluble in the most common organic solvents. Cobalt(II), copper(II), zinc(II) and palladium(II) Schiff-base complexes were obtained by treating CoCl2•6H2O, CuCl2•2H2O, Zn(NO3)2•6H2O and PdCl2 with two molar equivalents of the ligand in the presence of triethylamine, respectively (Scheme 2). The ligand and all the complexes were characterized by elemental analysis (CHN) and FT-IR spectroscopy. Also, the Schiff base ligand and its zinc(II) and palladium(II) complexes were investigated by NMR techniques. The molecular structures of all the complexes were determined by the single crystal X-ray diffraction technique.

IR spectra

In the FT-IR spectrum of the ligand, a sharp band appeared at 1628 cm -1 , which is attributed to the C N vibrations of the imine group [42], [43], [44]. In the FT-IR spectra of the Schiff base complexes this band was shifted to a lower wavenumber and appeared at 1601, 1615, 1613 and 1607 cm -1 for the cobalt(II), copper(II), zinc(II) and palladium(II) complexes, respectively. This shift could be attributed to a weakening of the M-N bonds on adduct formation and this can be explained by the donation of electrons from the nitrogen atom to the empty d-orbitals of the metal atom [37], [45], [46], [47].

The spectrum of the free ligand shows a strong band at 1196 cm -1 , assigned to the phenolic C-O stretching. However, after complexation of the C-O group via the oxygen atom to the metal ion, these bands were observed at 1174 and 1145 cm -1 for cobalt(II), 1181 and 1146 cm -1 for copper(II), 1174 and 1148 cm -1 for zinc(II) and 1180 and 1148 cm -1 for the palladium(II) complex. In these complexes, the absence of the phenolic O-H vibration indicates that ligand is deprotonated.

NMR spectra

The Schiff-base ligand, HL, and diamagnetic zinc(II) and palladium(II) complexes, ZnL2 and PdL2, were studied by 1 H and 13 C NMR experiments (Table 3). The 1 H and 13 C NMR spectra of the ligand and the complexes were run immediately and gave the expected simple spectra, indicating the integrity of the ligand and the complexes. The spectra of the complexes obtained after 12, 24 and 120 h were similar to the initial spectra, indicating that the complexes are stable in solution. The 1 H NMR spectra of the complexes are similar to that of the ligand, but with slight shifts to lower fields. The signal for the imine proton in the zinc(II)

and palladium(II) complexes appears at 8.52 and 7.32 ppm, respectively and this downfield shift with respect to the corresponding signal in the free ligand, indicates that the metal-nitrogen bond is retained in solution. Observing no signal corresponding to hydroxyl protons at 14.3 ppm, suggests that the hydroxyl groups are fully deprotonated and the oxygen atoms are most likely coordinated to the metal ions. The 13 C NMR spectra show 9 signals for the ZnL2 and PdL2 complexes. The peak at 168.7 ppm for ZnL2 and 159.7 ppm for PdL2, assignable to the imine carbon atoms, confirms the presence of the Schiff base ligand in the complexes [48].

Description of the crystal structures

Structural description of PdL2

In the solid state, PdL2 has a crystallographic center of symmetry that is right in the middle point of the palladium(II) ion in a planar-transoid conformation (Fig. 1). The molecular unit is centrosymmetric and is made up of equivalent halves. The tert-butyl groups show a fixed conformation in the crystal packing. The molecular structure of PdL2 is shown in Fig. 1.

Selected bond distances and angles are listed in Table 2. The crystallographic data reveal that the Pd(II) complex has distorted planar configuration and is four-coordinated by two phenolate oxygen and two imine nitrogen atoms of two Schiff base ligands. The ligands coordinated to the Pd(II) center in a trans geometry with respect to each other.

The Pd-O distance of 1.987(3) Å and the Pd-N bond length of 2.066(3) Å are similar to those seen in related complexes [49], [50], [51]. For instance, distances of 1.979(1) and 1.982(1) Å for the Pd-O bonds and 2.013(1) and 2.015(1) Å for the Pd-N bonds are found in a related di-(2allyliminomethylphenolato)palladium(II) complex derived from allylamine and salicylaldehyde [51]. The C N bond distance is 1.285( 5) Å (N1 C7), which is consistent with a slight elongation of the C-N double bond when it coordinates to a metal center [50]. Refs. [53], [54], [55], [56], [57], [58], [59]), step configuration (with the distance between the mean planes of the aromatic rings greater than d = 0.4 Å, see for example

Refs. [60], [61], [62], [63], [64], [65], [66], [67]) and bowl configuration (see for example

Refs. [68], [69], [70], [71], [72], [73]). Scheme 4. Planar, step and bowl configurations in bis(salicylidenimine)palladium(II) complexes.

-square of Pd, O, and N atoms;chelate rings; arene rings.

In the Pd(N,O)2 complexes with a planar configuration the Pd atom lies in the plane of the chelate rings (see for example Refs. [53], [54], [55], [56], [57], [58], [59]). The two sixmembered chelate rings and their annulated phenyl rings are coplanar (Scheme 4a). The complexes have an inversion symmetry. Frequently, however, a characteristic feature of Pd(N,O)2 complexes is an envelope configuration of the six-membered chelate rings, with the Pd atom deviating from the plane of the other 5 atoms of the chelate rings. A direct consequence is the alternative step or a bowl configuration (Scheme 4b and c). In the step configuration the planes between the aromatic rings are parallel to each other (see for example

Refs. [60], [61], [62], [63], [64], [65], [66], [67]), though complexes still have Ci symmetry.

Some Pd(II) complexes adopt a bowl configuration (see for example

Refs. [68], [69], [70], [71], [72], [73]). At best they have C2 symmetry. In the PdL2 complex, the chelate rings have the envelope form and the structure is the step configuration.

Structural description of CoL2, CuL2 and ZnL2

An ORTEP view of CoL2, CuL2 and ZnL2 with the atom-numbering scheme is presented in Fig. 2, Fig. 3, Fig. 4 and the crystallographic data and selected bond lengths and angles are collected in Table 1, Table 2. The crystallographic data reveal that the metal center is four-coordinated by two phenolate oxygen and two imine nitrogen atoms of two Schiff base ligands. The geometry around the metal center is distorted tetrahedral, with Pca2(1), P2( 1)2( 1)2( 1) and P2(1) space groups for CoL2, CuL2 and ZnL2, respectively. The ligands coordinate to the metal center in a cis geometry with respect to each other. In the crystal structures of all the complexes, the atoms of the phenyl ring plane [C(1)C( 2)C( 3)C( 4)C( 5)C( 6) and C(12)C( 13)C( 14)C( 15)C( 16)C( 17 As depicted in Fig. 2, Fig. 3, Fig. 4, all of the complexes have a distorted tetrahedral geometry, with bidentate coordinated ligands which have a cis geometry with respect to each other. The N(1)-M-N(2) angle in CuL2 is larger than that for the CoL2 and ZnL2 complexes (Fig. 5). With such an angle in CuL2, the distance and steric hindrance between the two butyl groups in CuL2 are greater and lower respectively than in the CoL2 and ZnL2 complexes. naphthaldi-minato]copper(II) has a bulky c-octyl substituent (c stands for cyclo) that determines a flat-tetrahedral coordination sphere [76]. Thus, steric hindrance of the N substituents is not the only factor influencing the coordination sphere in the solid state.

In our case, CuL2 is more distorted than bis(N-tert-butyl-2-oxy-1-naphthaldiminato)copper(II), according to their Dih values of 53.6° and 45.4°, respectively [75]. In CoL2 and ZnL2, the dihedral angle between the planes N(1)-M(1)-O(1) and N(2)-M(1)-O(2) are 87.1° and 83.1°, respectively, and the two bidentate ligands in these complexes are almost perpendicular to each other.

Electronic spectra

Electronic spectra of the free Schiff base ligand and the complexes were recorded in a CHCl3 solution. The absorption spectra for the Schiff base ligand (H2L) and all the complexes are shown in Fig. 6. The electronic spectrum of the Schiff base ligand exhibits three main peaks at 255, 316 and 409 nm with a shoulder at 240 nm (Fig. 6). The first and second peaks (240 and 255 nm) are attributed to benzene π → π * transitions.

The azomethine π → π * and n → π * transitions are viewed at 316 and at 409 nm as broad peaks with low intensity, respectively [77]. In the complexes CoL2, CuL2, ZnL2 and PdL2, the broad band that appears in the range 350-450 nm with more intensity than free Schiff base ligand may be due to MLCT (comprised of a transition between the metal centered HOMO to the ligand centered LUMO (C N(π * )), LMCT, d-d and azomethine n → π * transitions. This band is in the range usually observed for azomethine n → π * , d-d and LMCT transitions [36], [78]. According to the d 10 electronic configuration (+2 formal oxidation state) of zinc, the Zn(II) complex shows no d-d transition.

Catalytic activity of the different complexes in the solvent free oxidation of sulfides

Influence of different amounts of catalyst

We first studied the oxidation of thioanisole as a model substrate using 30% aqueous hydrogen peroxide as the oxidant under solvent-free conditions. The amount of catalyst had a considerable effect on the conversion of thioanisole to the corresponding sulfone. The results are given in Table 4 and show that the reaction is sensitive to the catalyst amounts for each case. These results showed that on increasing the catalyst amount, the reaction conversion enhances without significant changing the selectivity for all the catalysts. The role of the catalyst is very important in the reaction because this reaction has low conversion without any catalyst (Table 4, entry 17).

Also, among the four catalysts, ZnL2 showed a better catalytic performance compared to the others.

Influence of different amounts of oxidant

After optimization of the catalyst amount, we demonstrated the effect of the amount of oxidant on the oxidation of thioanisole in the presence of each of the four catalysts. With 0.5 mmol of H2O2 all catalysts showed low conversions (entries 1, 5, 9 and 13, Table 5); in contrast, on raising the amount of oxidant, higher conversions were obtained; among them all, ZnL2 provided the highest conversion of 99% and selectivity of 99%. Furthermore, it can be seen in Table 5 that with an increase in the amount of the H2O2 with 0.01 mmol catalyst at 50 °C, the conversion of thioanisole increased, but selectivity for sulfoxide decreased quickly.

Influence of reaction temperature

In the present work for optimization of the reaction conditions, we also studied the effect of temperature on the catalytic performance of CoL2, CuL2, ZnL2 and PdL2 (Table 6). Obviously, a substantially improved conversion of thioanisole to the corresponding sulfone and sulfoxide was observed with an increase in the temperature, but in all cases a decrease in selectivity to sulfoxide was observed; for example at 85 °C, an increase in the oxidation activity was observed, which caused the increased formation of the methyl phenyl sulfone, but selectivity for sulfoxide decreased for all catalysts.

Influence of the geometrical structure of the catalyst

In this work, the catalytic activity of four kinds of metal complexes were examined (Fig. 7, Fig. 10). Among them, ZnL2 showed the best catalytic activity for the oxidation of sulfide and CoL2 showed the lowest catalytic performance under these reaction conditions. In general, there are some different parameters affecting the performance of a catalyst. Here, we studied the influence of the geometrical structure of the metal complex. As previously reported [79], [80], [81], formation of a metal-peroxo intermediate occurs in some oxidation reactions that shows the geometrical structure of the complex plays an important role in its catalytic activity. In the present study, according to Fig. 5, the CoL2 complex has more steric hindrance around the metal center due to the geometry of the ligands. So the bulkiness of the ligands for CoL2 probably lead to a further lowering of the accessibility of the metal center in the catalyst and so reduce the catalytic efficiency of the complex (the slope of the conversion curve for this complex is lower than for the other ones) (Fig. 7, Fig. 10).

Oxidation of different sulfides

The oxidation of thioanisole to the corresponding sulfone, catalyzed by CoL2, CuL2, ZnL2 and PdL2 with 2 mmol of H2O2 at 50 °C, was determined as a function of the reaction time (Fig. 7, Fig. 10). The results indicate that at the beginning of the reaction the sulfoxide is formed selectively, but the amount of sulfone increases slightly over time. It is remarkable that sulfone can be obtained with high selectivity with 2 equivalents of 30% H2O2 in the presence of all the complexes in 3 h under solvent free conditions.

In an effort to extend the scope of the reaction under the optimized conditions, the CuL2, ZnL2 and PdL2 catalysts were applied to the oxidation of various types of aliphatic and aromatic sulfides and the results are summarized in Table 7. CoL2 did not show good catalytic activity, for this reason we did not use it for oxidation of different sulfides. The different sulfides were selectively oxidized to the corresponding sulfones with 2 mmol H2O2 under solvent free conditions. The oxidation of allylsulfides and hydroxysulfides proceeded chemoselectively to the corresponding sulfones, without the epoxidation of the double bonds and dehydrogenation of the hydroxyl groups (entries 6, 7 and 8). Thioanisole was oxidized under solvent-free conditions to the corresponding sulfone in 93% yield with 2 mmol H2O2 and ZnL2 as a catalyst (Table 7, entry 2). The CuL2 catalyst gave sulfone in 80% conversion, while the PdL2 catalyst gave methyl phenyl sulfone in 74% conversion (Table 7, entry 2). Dibutyl sulfide, thiophene and all the aromatic sulfides, such as methylphenyl sulfide, diphenyl sulfide and the substituted diphenyl sulfide (4-nitro), were oxidized to the corresponding sulfones with a good conversion in the presence of each of the three catalysts (Table 7, entries 1-5). Unfortunately, with this protocol DBT was oxidized into sulfone (Table 7, entry 9) with a low conversion. Although DBT was oxidized into sulfone with low conversion under the reaction conditions, these results show the good capability of this method for the oxidation of several types of sulfides to the corresponding sulfones.

Scheme 3 .

 3 Scheme 3. Oxidation of thioanisole with hydrogen peroxide in the presence of CoL2, CuL2, ZnL2 and PdL2 Schiff base complexes as catalysts under solvent-free conditions.

Fig. 1 .

 1 Fig. 1. Perspective view of PdL2 constituted by the asymmetric unit (filled drawings) showing the numbering scheme and the centrosymmetric half part (empty drawings). Thermal ellipsoids are drawn at the 50% probability level, while the hydrogen size is arbitrary. The suffix A corresponds to the symmetry code -x, -y, -z.

  The O-Pd-N bond angles [O(1)-Pd(1)-N(1) = 88.45(12)° and O(1)-Pd(1)-N(1A) = 91.55(12)°] are close to 90°. Moreover the coordination of the two NO bidentate chelate ligands to the Pd(II) ion results in the formation of two six-membered rings (Pd(1)/O(1)/C(1)/C(6)/C(7)/N(1) and Pd(1)/O(1A)/C(1A)/C(6A)/C(7A)/N(1A)). A search for salicylidenimine palladium(II) complexes in the Cambridge Structural Database System 2014 showed that except in one case [52] the N,O-ligand atoms in bis(salicyli-denimine) palladium(II) complexes, Pd(N,O)2, adopt a trans arrangement in the square planar Pd(II) configuration. For salen-type complexes, Pd(ONNO), the N,O-ligand atoms are cis to each other, and they are excluded from the following analysis. The trans-Pd(N,O)2 complexes can subdivide into three classes (Scheme 4): planar configuration (with the distance between the mean planes of the aromatic rings being less than d = 0.4 Å, see for example

Fig. 2 .Fig. 3 .Fig. 4 .

 234 Fig. 2. View of CoL2 in the asymmetric unit showing the numbering scheme. Thermal ellipsoids are drawn at the 50% probability level, while the hydrogen size is arbitrary. The C9, C10 and C11 carbon atoms of the tert-butyl group are disordered over two sites and refined with site occupancy factors 0.67:0.33. Only the major component of the disordered tert-butyl group is shown.

  )] and the chelate ring formed by the same ligand plane [O(1)/M(1)/N(1)/C(7)/C(6)/C(1) and O(2)/M(1)/N(2)/C(18)/C(17)/C(12)] are nearly coplanar with dihedral angles of 3.82(16)° and 2.17

  (19)° for CoL2, 2.53 (12)° and 11.13 (11)° for CuL2 and 5.48 (13)° and 5.00 (11)° for ZnL2, respectively. Examination of the main metal-ligand distances shows that the Co⋯N, Cu⋯N and Zn⋯N distances are longer than the Co⋯O, Cu⋯O and Zn⋯O distances. The M⋯N and M⋯O distances are 2.003(3) and 1.883(3) Å for CoL2, 1.995(2) and 1.8957(17) Å for CuL2 and 2.0200(19) and 1.911(2) Å for ZnL2.

Fig. 5 .

 5 Fig. 5. Distances (Å) and angles (°) between the two butyl groups in the CoL2, CuL2 and ZnL2 complexes.

Fig. 6 .

 6 Fig. 6. Electronic spectra of the Schiff base ligand and the complexes (A: H2L, B: CuL2, C: CoL2, D: PdL2, E: ZnL2).

Fig. 7 .Fig. 8 .

 78 Fig. 7. Reaction profile of the oxidation of thioanisole catalyzed by CoL2 with 2 mmol of H2O2 at 50 °C.

Fig. 9 .

 9 Fig. 9. Reaction profile of the oxidation of thioanisole catalyzed by ZnL2 with 2 mmol of H2O2 at 50 °C.

Fig. 10 .

 10 Fig. 10. Reaction profile of the oxidation of thioanisole catalyzed by PdL2 with 2 mmol of H2O2 at 50 °C.

Table 1 .

 1 33 and 0.65:0.35 for CoL2 and ZnL2, respectively (details are available in the archived CIF). Crystallographic data for the complexes are listed in Table 1. Selected bond distances and angles are summarized in Table 2. Crystal data and structure refinement.

	Empty Cell	CuL2	CoL2	PdL2	ZnL2
	Empirical formula	C22H28N2O2Cu C22H28N2O2Co C22H28N2O2Pd C22H28N2O2Zn
	Formula weight	416.00	411.39	458.86	417.83
	T (K)	298(2)	298(2)	298(2)	298(2)
	λ (Å)	0.71073	0.71073	0.71073	0.71073
	Crystal system	orthorhombic	orthorhombic	monoclinic	monoclinic
	Space group	P2(1)2(1)2(1)	Pca2(1)	C2/c	P2(1)
	Unit cell dimensions
	a (Å)	9.1377(18)	20.1258(14)	19.308(2)	10.625(2)
	b (Å)	11.192(2)	10.9506(8)	7.7468(7)	9.982(2)
	c (Å)	21.367(4)	9.8769(9)	14.6967(16)	10.821(2)
	β (°)			108.753(9)	110.81(3)
	V (Å 3 )	2185.3(8)	2176.8(3)	2081.6(4)	1072.8(4)
	Z	4	4	4	2
	Dcalc (Mg m -3 )	1.264	1.255	1.464	1.294

Table 2 .

 2 Selected bond distances (Å) and angles (°) for

CuL2, CoL2, ZnL2 and PdL2.

	Empty Cell	CuL2	CoL2	ZnL2	Empty Cell	PdL2
	M(1)-O(1)	1.8957(17) 1.883(3)	1.911(2)	Pd(1)-O(1)	1.987(3)
	M(1)-N(1)	1.995(2)	2.003(3)	2.0200(19) Pd(1)-N(1)	2.066(3)
	N(1)-C(7)	1.285(3)	1.272(4)	1.275(4)	N(1)-C(7)	1.285(5)
	N(2)-C(18)	1.286(3)	1.281(4)	1.272(3)
	O(1)-M(1)-O(2)	137.28(9)	113.66(12) 114.19(9)	O(1)-Pd(1)-O(1A)	180.00(17)
	O(1)-M(1)-N(1)	95.88(8)	97.94(12)	97.58(11)	O(1)-Pd(1)-N(1)	88.45(12)
	O(2)-M(1)-N(1)	100.18(7)	113.45(11) 96.65(9)	O(1)-Pd(1)-N(1A)	91.55(12)
	N(1)-M(1)-N(2)	145.58(8)	122.03(12) 126.95(13) N(1)-Pd(1)-N(1A)	180.0
	O(2)-M(1)-N(2)	93.22(8)	96.71(11)	96.65(9)

2.3. Synthesis of the Schiff-base ligand (HL: 2-tert-butyliminomethyl-phenol)

	Tert-butylamine (10 mmol) was dissolved in 30 ml of absolute methanol or chloroform and was
	added slowly to a stirring solution of salicylaldehyde (10 mmol) in 30 ml of absolute methanol or
	chloroform at ambient temperature. The color immediately changed to yellow. The mixture was
	then stirred for 2 h at ambient temperature before removal of the solvent under vacuum. The
	result was a yellow oil. Yield 89%, Anal. Calc. for C11H15ON: C, 74.54; H, 8.53; N, 7.90. Found:

C, 74.49; H, 8.56; N, 7.88%. FT-IR (KBr, cm -1 ): 3480 (νO-H, br, vs), 3060-2871 (C-H aliphatic and aromatic), 1628 (νC N), 1196 (νC

Table 3 .

 3

	Empty Cell HL	ZnL2	PdL2	Empty Cell	HL	ZnL2	PdL2
	H7	8.48(s)	8.52(s)	7.32(s)	C1	161.4	169.3	165.0
	H5	7.44(d of d) 7.40(d)	7.14(d of t) C7	160.9	168.7	159.7
	H3	7.29(d of t) 7.25(t)	7.17(d of t) C3	131.9	137.1	134.2
	H2	6.87(d)	6.63(d)	6.80(d)	C5	131.8	132.1	133.5
	H4	6.84(t)	6.56(t)	6.58(d of t) C4	117.9	122.2	119.8

1

H and

13

C NMR data for the Schiff base ligand (HL), ZnL2 and PdL2 complexes. 1. Download : Download full-size image Empty Cell 1 H NMR Empty Cell Empty Cell 13 C NMR Empty Cell s, singlet; d, doublet; t, triplet; m, multiplet.

Table 4 .

 4 Oxidation of thioanisole with different amounts of catalyst in the presence H2O2. a

	Entry	Catalyst	mmol of catalyst	Conversion (%) b	Selectivity (%) c
	1	Co	0.001	20	97
	2	Co	0.005	23	99
	3	Co	0.01	29	99
	4	Co	0.05	30	99
	5	Cu	0.001	48	94
	6	Cu	0.005	61	99
	7	Cu	0.01	80	99
	8	Cu	0.05	83	99
	9	Zn	0.001	63	98

Table 5 .

 5 Oxidation of thioanisole with various amounts of H2O2 in the presence of Co, Ni, Zn and Pd Schiff base a complexes.

	Entry	Catalyst	mmol H2O2	Conversion (%) b	Selectivity (%) c
	1	Co	0.5	17	66
	2	Co	1	20	81
	3	Co	2	29	99
	4	Co	4	35	99
	5	Cu	0.5	33	58
	6	Cu	1	51	77
	7	Cu	2	80	99
	8	Cu	4	86	99
	9	Zn	0.5	37	63
	10	Zn	1	59	72
	11	Zn	2	93	99
	12	Zn	4	96	99
	13	Pd	0.5	30	69
	14	Pd	1	50	80

c Selectivity for sulfone.

Table 6 .

 6 Oxidation of thioanisole with different catalysts at various temperatures. a

	Entry	Catalyst	Temperature (°C)	Conversion (%) b	Selectivity (%) c
	1	Co	25	16	45
	2	Co	50	29	99
	3	Co	60	31	99

Table 7

 7

	CuL2	ZnL2	PdL2

. The oxidation of various sulfides with CoL2, CuL2, ZnL2, and PdL2 Schiff-base complexes as catalyst using 30% aqueous H2O2. a Entry Sulfide Time (h) Conversion (%) b Selectivity(%) c

Acknowledgement

Support for this research by the University of Isfahan is acknowledged.

Conclusion

In We have demonstrated the effectiveness of these complexes as catalysts for the green oxidation of sulfides to the corresponding sulfones with hydrogen peroxide. In this system the reactions can be carried out under solvent-free conditions as a green sustainable method using all the catalysts in the presence of H2O2. Conversions of 20-96%, and selectivities of 98-100% for sulfone are observed with the four compounds as catalysts. Also, we found that ZnL2 shows a better catalytic activity for the oxidation of sulfide and CoL2 shows a lower catalytic performance with these reaction conditions. Overall, we recommend this green, simple, clean, and economical procedure for the oxidation of different sulfides to the corresponding sulfones.

Appendix