Synthesis, spectroscopic characterization, DFT studies and antibacterial assays of a novel silver(I) complex with the anti-inflammatory nimesulide

Polyhedron 36 (2012) 112–119

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Synthesis, spectroscopic characterization, DFT studies and antibacterial assays of a novel silver(I) complex with the anti-inflammatory nimesulide Raphael E.F. de Paiva a, Camilla Abbehausen a, Alexandre F. Gomes b, Fábio C. Gozzo b, Wilton R. Lustri c, André L.B. Formiga a, Pedro P. Corbi a,⇑ a b c

Inorganic Chemistry Department, Institute of Chemistry, University of Campinas – UNICAMP, P.O. Box 6154, 13083-970 Campinas, SP, Brazil Organic Chemistry Department, Institute of Chemistry, University of Campinas – UNICAMP, P.O. Box 6154, 13083-970 Campinas, SP, Brazil Biological and Health Sciences Department – UNIARA, 14801-320 Araraquara, SP, Brazil

a r t i c l e

i n f o

Article history: Received 16 December 2011 Accepted 3 February 2012 Available online 9 February 2012 Keywords: Silver(I) Nimesulide Metal complex Density functional theory ESI-QTOF-MS Antibacterial activity

a b s t r a c t A novel silver(I) complex with nimesulide (NMS) of composition AgC13H11N2O5S was synthesized and characterized by chemical and spectroscopic measurements, density functional theory (DFT) studies and biological assays. Infrared (IR), ESI-QTOF-mass spectrometric (MS) analyses and 1H, 13C and [1H–15N] nuclear magnetic resonance (NMR) studies indicate that NMS acts as a bidentate ligand, being coordinated to Ag(I) through the nitrogen and one of the oxygen atoms of the sulphonamide group. The proposed structure based on the experimental data was confirmed as a minimum of the potential energy surface (PES) with the calculation of the hessians, showing no imaginary frequencies. Also, the theoretical IR spectra of the free ligand and of the silver(I) complex (Ag–NMS) are in a good agreement with the experimental data. Theoretical Time-Dependent DFT (TD-DFT) studies confirmed that the observed transitions in the UV–Vis spectra of the NMS and Ag–NMS are due to p–p⁄ transitions. The antibacterial activity of Ag–NMS was evaluated by an antibiogram assay, using the disk diffusion method. The complex showed an effective antibacterial activity against Staphylococcus aureus (Gram-positive), and Escherichia coli and Pseudomonas aeruginosa (Gram-negative) pathogenic bacterial cells. Ó 2012 Elsevier Ltd. All rights reserved.

1. Introduction Biological and pharmacological activities of silver(I)-based compounds have been explored for a long time [1,2]. The first report concerning the use of silver in wound treatment is dated to the 18th century, when silver nitrate was used in the treatment of ulcers [3]. In the 20th century, the biological activities of silver compounds were shown to be associated primarily to presence of silver(I) ions [4]. In the 1940s, however, with the advances in the isolation of the fungus Penicillium notatum and purification of penicillin, penicillin-derived antibiotics started to be used worldwide as the main pharmaceutical agents against bacterial infections,

Abbreviations: NMS, nimesulide; Ag–NMS, silver(I) complex with nimesulide; NMR, nuclear magnetic resonance; IR, infrared; ESI-QTOF-MS, electrospray ionization quadrupole time-of-flight mass spectrometry; TMS, tetramethylsilane; DMSO, dimethyl sulfoxide; LANL2DZ, Los Alamos National Laboratory-2nd version on double zeta function; B3LYP, Becke3–Lee Young Parr functional; ATCC, American type collection cell; MH, Mueller–Hinton agar; BHI, Brain–heart infusion medium; CFU, colony forming unit; PCM, polarizable continuum model; PES, potential energy surface. ⇑ Corresponding author. Tel.: +55 19 35213130; fax: +55 19 35213023. E-mail address: [email protected] (P.P. Corbi). 0277-5387/$ - see front matter Ó 2012 Elsevier Ltd. All rights reserved. doi:10.1016/j.poly.2012.02.002

drastically reducing the use of silver compounds [5,6]. Only in the 1960s, when Moyer et al. [7] introduced the use of a 0.5% silver nitrate solution for the treatment of burns, did silver-based compounds start to be considered again. Moyer proposed that at this low concentration the silver nitrate does not interfere with epidermal proliferation and possess antibacterial properties against Staphylococcus aureus, Pseudomonas aeruginosa and Escherichia coli [8]. In 1968, silver-sulfadiazine, a silver(I)–sulphonamide complex, was introduced clinically for treatment of burns and skin wounds. Due to its efficacy, silver-sulfadiazine has been used in medicine since the 1970s as an antibacterial agent. At the present time, most of the silver(I)-based compounds are commercially available in the form of wound dressings, such as ActicoatÒ (Smith and Nephew) and ActisorbÒ (Johnson and Johnson) [9], which have some intrinsic benefits like stimulation of healing in indolent wounds, prophylactic applications and treatment of critically colonized wounds [10]. Silver(I)-based antiseptics have been shown to possess lower propensity to induce antimicrobial resistance than other antibiotics [11], and simultaneously have a remarkably low human toxicity when compared to other heavy metal ions [12], which are desirable properties for antimicrobial agents. According to the literature, there are three possible mechanisms for inhibition of bacteria growth by

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silver(I) ions: (a) interference in the electron transport system, (b) DNA-binding and (c) interaction with the cell membrane [13]. One approach to obtain new silver(I)-based compounds is to combine some specific classes of biologically active molecules with silver ions [14]. In some cases, silver complexes with antibiotics have been shown to be more effective than the free antibiotics [15]. Following this approach, new metal–organic complexes with antibacterial activities have been synthesized and described in the literature. Kazachenko et al. [16] investigated the synthesis and antimicrobial activities of silver complexes with the amino acids histidine and tryptophan. Both compounds showed a good antibacterial activity against Gram-negative and Gram-positive bacterial strains, with low toxicity. Ruan et al. [17] synthesized silver(I) complexes with carbazole carboxylate ligands exhibiting an antibacterial activity superior to penicillin against strains of Bacillus subtilis. Isab and Wazeer [18] reported the synthesis and characterization of a monodentate silver(I) complex with captopril, which exhibits a high antimicrobial activity against P. aeruginosa when compared to the free ligand. Poyraz et al. [19] reported the synthesis of silver(I) complexes with triphenylphosphine and aspirin or with salicylic acid. The complexes exhibited higher cytotoxic activity than cisplatin against leiomyosarcoma cells and human breast adenocarcinoma cells. Furthermore, a recently published silver(I) complex with acesulfame was also shown to possess inhibitory activity against Mycobacterium tuberculosis with a minimal inhibitory concentration (MIC) value of 11.6 lmol L1, and also against E. coli, P. aeruginosa and Enterococcus faecalis [20]. The sulphonamides represent an attractive class of ligands with antibiotic properties. Such compounds are characterized by the presence of a sulphonamide group (O2SNH). Nimesulide and sulfadiazine are two well known examples of sulphonamides. Specifically, nimesulide (C13H12N2O5S, Fig. 1, NMS) is a nonsteroidal anti-inflammatory drug (NSAID), used in the treatment of acute or chronic inflammatory processes of the respiratory tract and the oral cavity, tendinitis and rheumatoid arthritis [21]. The activity of NMS is associated with its antioxidant capability, since NMS can block the formation of the radical anion superoxide (O2) and other reactive oxygen species (ROS), which are related to the inflammatory process and pain. In addition, NMS selectively inhibits action of cyclooxygenase-2 (COX-2) and the inflammatory process. Due to the presence of the sulphonamide group, nimesulide can be considered a suitable ligand to coordinate both hard and soft acids. The present work describes the synthesis, spectroscopic characterization and antibacterial assays of a novel silver(I) complex with nimesulide.

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2. Experimental 2.1. Materials and equipment Nimesulide (>99%) and analytical grade silver(I) nitrate (AgNO3) were purchased from Sigma–Aldrich laboratories. Potassium hydroxide (>85%) was purchased from Fluka. Elemental analyses for carbon, hydrogen and nitrogen were performed using a Perkin Elmer 2400 CHNS/O Analyzer. Infrared (IR) spectra were measured in KBr pellets using a Bomem MB-Series Model B100 FT-IR spectrophotometer in the range 4000–400 cm–1 with resolution of 4 cm–1. Solution-state 1H, 13C and the [1H–15N] correlation nuclear magnetic resonance spectra were recorded on a BrukerAvance III 400 MHz (9.395T). The 1H NMR spectra were obtained at 400.1 MHz while the 13C–{1H} spectra were obtained at 125.7 MHz. Samples were analyzed in deuterated dimethylsulfoxide-d6 solutions and the chemical shifts were given relative to tetramethylsilane (TMS). The UV–Vis spectra were recorded in 1.67  10–6 mol L1 water solutions using 10.00 mm quartz cuvettes in a Hewlett–Packard 8453A diode array spectrophotometer. Electrospray ionization quadrupole time-of-flight mass spectrometry (ESI-QTOF-MS) measurements were carried out in a Waters Synapt HDMS instrument (Manchester, UK). The complex was dissolved in methanol and the resulting solution was directly infused into the instrument’s ESI source at a flow rate of 15 lL min1. Typical acquisition conditions were capillary voltage 3 kV, sampling cone voltage 20 V, source temperature 100 °C, desolvation temperature 200 °C, cone gas flow 30 L h1, desolvation gas flow 900 L h1, Trap and Transfer collision energies at 6 and 4 eV, respectively. The ESI(+) mass spectra (full scans) and fragment ion spectra for quadrupole-isolated ions (QTOF-MS/MS) were acquired in reflectron V-mode at a scan rate of 1 Hz. For fragment ion experiments, the desired ion was isolated in the mass-resolving quadrupole, and the collision energy of the Trap cell was increased until the sufficient fragmentation was observed. Prior to the analyses, the instrument was calibrated with phosphoric acid oligomers (H3PO4 0.05% in H2O/MeCN 50:50). 2.2. Synthesis of the complex The silver(I) complex with nimesulide (Ag–NMS) was synthesized by the reaction of 10.0 mL of an aqueous solution containing 8.0  104 mol (0.2466 g) NMS and 8.8  104 mol (0.0494 g) KOH with 2.0 mL of an aqueous solution containing 8.8  104 mol (0195 g) AgNO3. The synthesis was carried out with stirring and at room temperature. After 1 h of constant stirring, the yellowish solid obtained was vacuum filtered, washed with cold water, and dried in a desiccator over P4O10. Anal. Calc. for AgC13H11N2O5S: C, 37.6; H, 2.7; N, 6.7. Found: C, 37.8; H, 2.4; N, 6.7%. The yield of the synthesis was 89%. The Ag–NMS complex is slightly soluble in water, and it is soluble in methanol, ethanol, acetonitrile and dimethylsulfoxide. No single crystals were obtained to provide an X-ray crystallographic study. 2.3. Molecular modeling

Fig. 1. Schematic structure of NMS with carbon atom numbering.

Geometric optimizations were carried out using GAMESS software [22] with a convergence criterion of 10–4 a.u. in a conjugate gradient algorithm. The LANL2DZ [23] effective core potential was used for silver and the atomic 6-31G (d, p) basis set [24–27] for all other atoms. Density functional theory (DFT) calculations were performed by using the B3LYP [28,29] gradient-corrected hybrid to solve the Kohn–Sham equations with a 10–5 a.u. convergence criterion for the density change. The final geometries were confirmed as minima of the potential energy surface (PES) with calculation of the hessian.

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The harmonic vibrational frequencies and intensities were calculated at the same level of theory with the analytical evaluation of second derivatives of energy as a function of atomic coordinates, and the calculated intensities were used to generate the theoretical spectra. Frequencies were scaled by a factor of 0.9614, as recommended by Scott and Radom [30]. Simulated vibrational spectra were obtained from the sum of Lorentzian functions with 20 cm1 half-bandwidths using the software MOLDEN 4.7 [31]. TD-DFT was employed to calculate theoretical UV–Vis spectra and, in these calculations, the Polarizable Continuum Model (PCM) [32] was used to simulate the effect of water in the electronic transitions. 2.4. Biological assays Three referenced bacteria (E. coli – ATCC 25922, P. aeruginosa – ATCC 27853, and S. aureus – ATCC 25923) were used in this study. The antibiogram assay was performed by the disc diffusion method [33,34]. The Ag–NMS complex was tested in Mueller–Hinton (MH) agar plates. The microorganisms (E. coli, P. aeruginosa and S. aureus) were transferred to separate test tubes containing 5.0 mL of sterile brain heart infusion (BHI) medium and incubated for 18 h at 35– 37 °C. Sufficient inocula were added in new tubes until the turbidity equaled 0.5 McFarland (1.5  108 CFU mL1). The bacterial inocula diluted with BHI (McFarland standard) were uniformly spread using sterile cotton swabs on sterile Petri dishes of MH agar. Sterile filter paper discs of 10 mm in diameter were aseptically impregnated with 800 lg of Ag–NMS according to the following procedure: 20 mg of Ag–NMS complex were diluted in 1.0 mL of absolute ethanol and homogenized. Then, 40 lL of the solution were taken with a micropipette and transferred to the paper discs. Sterile discs impregnated with 800 lg of pure NMS, used as a negative control, were prepared as follows: 20 mg of NMS were dissolved in 1.0 mL of water and 40 lL of the homogenized solution were transferred to the paper discs. Three positive controls were used: ceftriaxone, gentamicine and AgNO3. Discs impregnated with Ag–NMS, pure NMS and the positive controls were dried and sterilized in a vertical laminar flow under UV radiation for 45 min before the experiment. All impregnated discs were placed on the surface of the solid agar. The plates were incubated for 18 h at 35–37 °C and examined thereafter. Clear zones of inhibition formed around the discs were measured and the complex sensitivity was assayed from the diameter of the clear inhibition zones (in millimeters). All experiments were performed in duplicate. 3. Results and discussion 3.1. Nuclear magnetic resonance spectroscopy Solution state 1H, 13C and [1H–15N] heteronuclear multiple bond coherence (HMBC) NMR spectra were obtained in order to identify the coordination sites of NMS to Ag(I). The spectra of the Ag–NMS complex were analyzed by comparison to the NMR spectra of free NMS. The structure of NMS with carbon assignments is shown in Fig. 1. Nitrogen coordination of the sulphonamide group of NMS to Ag(I) was primarily evaluated by [1H–15N] multiple bond coherence NMR spectroscopy [35,36]. The 15N spectra are provided in Fig. 2. In the NMS spectrum, the 15N chemical shift of the nitrogen atom of the sulphonamide group is observed at 115.1 ppm, while in the Ag–NMS spectrum, this signal is observed at 144.3 ppm. The observed Dd (d complex  d ligand) of 29.2 ppm indicates coordination of NMS to Ag(I) through the nitrogen atom of the sulphonamide group. On the other hand, the nitrogen atom of the NO2

Fig. 2. Heteronuclear [1H–15N] multiple bond coherence spectra of (a) NMS and (b) Ag–NMS.

group appears at 369.7 ppm in the NMS spectrum, while for the complex, it appears at 371.0 ppm. The minor chemical shift of 1.3 ppm, when compared to the 15N shift of the nitrogen of the sulphonamide group, suggests that NMS is not coordinated to Ag(I) through the NO2 group. Solution state 1H NMR spectroscopy was applied to obtain further details about the coordination sites of NMS to Ag(I). The obtained spectra for NMS and Ag–NMS are presented in Fig. 3, while the corresponding assignments and chemical shifts are presented in the Table 1. First, the hydrogen which appears as a singlet at 10.2 ppm in the NMS 1H NMR spectrum is no longer observed in the spectrum of the Ag–NMS complex. This signal is attributed to the hydrogen atom bonded to the nitrogen of the sulphonamide group of NMS. This data leads us to consider the loss of the hydrogen atom of the (O2SNH) group of NMS and nitrogen coordination to the metal center. This evidence, in addition to the expressive 15N chemical shift of the (N–H) group when the ligand and the complex data are compared, confirms nitrogen coordination of the sulphonamide group to Ag(I). Observing the Dd values presented in the Table 2, it is also possible to note that all hydrogens are shifted upfield upon coordination. The most pronounced shift (0.42 ppm) occurs for the hydrogen atoms of the methyl group, which is bonded to the sulfur atom of the sulphonamide group. Solution state 13C NMR spectroscopy was employed to evaluate the electronic density changes in the NMS molecule upon coordination. Table 2 summarizes the carbon chemical shifts for NMS and the Ag–NMS complex. The Table 2 shows that changes in the electronic density of the carbon atoms do not strictly follow the same standard as the changes of the hydrogen atoms upon coordination. It is also evident that the methyl carbon presents a pronounced upfield Dd

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Fig. 3. 1H NMR spectra of (a) NMS and (b) Ag–NMS.

Table 1 The 1H NMR assignments, chemical shifts and multiplicity for NMS and Ag–NMS – Dd represents the difference between the chemical shifts of the complex and the free ligand. Assignment

NMS d (ppm)

Ag–NMS d (ppm)

Dd (ppm)

Multiplicity

CH3 20 ,60 40 30 ,50 3 6 5 NH

3.20 7.16–7.18 7.25–7.29 7.47–7.51 7.53–7.54 7.73–7.75 8.01–8.04 10.2

2.78 6.93–6.95 7.07–7.11 7.27–7.31 7.41–7.42 7.58–7.60 7.90–7.93 –

0.42 0.23 0.18 0.20 0.12 0.15 0.11 –

s d t t d d dd s

s = singlet, d = doublet, dd = double doublet and t = triplet.

Table 2 13 C NMR assignments and chemical shifts for NMS and the Ag–NMS complex. Assignment

NMS d (ppm)

Ag–NMS d (ppm)

Dd (ppm)

CH3 3 6 5 20 ,60 40 30 ,50 1 10 2 4

40.89 112.4 119.3 119.5 121.1 124.9 130.3 135.6 143.1 147.5 155.0

38.52 113.4 117.9 118.4 120.4 123.7 129.9 137.2 145.6 140.0 156.2

2.37 1.0 1.4 1.1 0.7 1.2 0.4 1.6 2.5 0.5 1.2

upon coordination, just like the hydrogen atoms of this group. This evidence leads us to consider that the sulphonyl group (O@S@O) is also participating in the ligand coordination to Ag(I) since the methyl group exhibits remarkable chemical shifts on 1H and 13C NMR spectra when the ligand and the complex are compared. 3.2. Infrared vibrational spectroscopy The infrared (IR) vibrational spectra of different sulphonamides have been studied and reported in the literature [37,38]. For S-methyl sulphonamides, the m(N–H) vibration band is observed in the range 3320–3250 cm1. Moreover, four characteristic bands of the sulphonyl group are observed: the mas(O@S@O) around 1350 cm1, the ms(O@S@O) around 1160 cm1, and the scissoring

Fig. 4. Infrared vibrational spectra of (a) NMS and (b) Ag–NMS.

and wagging deformation modes in the ranges 568–520 and 529– 487 cm1, respectively. Lastly, the m(C–S) vibration band is expected in the range 773–754 cm1. The IR spectra of NMS and the Ag–NMS complex are shown in Fig. 4. The IR spectrum of NMS exhibits all the characteristic sulphonamide bands. The (N–H) stretching appears as a sharp and intense signal at 3284 cm1. The four signals of the sulphonyl group also appear: the mas(O@S@O) at 1342 cm1, ms(O@S@O) at 1153 cm1, the scissoring deformation at 553 cm1 and the wagging deformation at 516 cm1. The (C–S) stretching at 754 cm1and the two characteristic bands of the nitro (NO2) group mas(NO2) at 1522 cm1 and ms(NO2) at 1317 cm1 are also observed. The Ag–NMS IR spectrum supplies remarkable evidence about coordination of NMS to Ag(I). The first, and most pronounced evidence, is the disappearance of the m(N–H) band, due to the loss of the hydrogen atom of the sulphonamide group upon coordination. Furthermore, the mas(O@S@O) and ms(O@S@O) vibration bands are shifted by 48 cm1 to lower energy values. This suggests the participation of the sulphonyl group in the coordination of the NMS to Ag(I). It is interesting to note that the shift of the mas(O@S@O) and ms(O@S@O) bands to lower energies in the IR spectrum of Ag–NMS is related to the weakening of the S@O bond. Nakamoto [39] has shown that this effect is observed when the coordination occurs through the oxygen atom of the sulphonyl group. Based on the observed data, coordination of NMS to Ag(I) seems to occur in a bidentate form through the nitrogen atom and one of the oxygen atoms of the sulphonamide group. 3.3. Electronic absorption spectroscopy The UV–Vis electronic absorption spectra of NMS and the Ag– NMS complex are shown in Fig. 5. The NMS spectrum is dominated by p–p⁄ transitions and exhibits an absorption band in the visible region at 389 nm. This transition is centered in the nitro-aryl ring and it is low-lying due to the electron-withdrawing properties of the NO2 substituent. There are at least three less intense absorption bands in the UV region at 195, 225, and 270 nm. Coordination to Ag(I) does not change the spectrum profile but the bands become stronger, especially the one in the visible at 389–391 nm, which is responsible for the Ag–NMS complex intense yellow colour, is enhanced by a factor of 22. Molar extinction coefficient varies from 570 mol1 cm1 in NMS to 12 655 mol1 cm1 in Ag–NMS.

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Fig. 5. Experimental (solid) and theoretical (vertical dash lines) UV–Vis electronic absorption transitions of (a) Ag–NMS and (b) NMS. Top and right axis correspond to the parameters of the theoretical spectra.

Fig. 6. Fragment mass spectrum for the Ag–NMS monoprotonated ion, [AgNMS+H]+, of m/z 414.95.

This is strong evidence that one of the coordination sites is the nitrogen atom of the sulphonamide group, which is bonded to the nitro-aryl ring. 3.4. Mass spectrometric measurements The ESI-QTOF-MS analyses of Ag–NMS demonstrate the presence of the proposed monomeric form of the complex in solution (Supplementary data #1) as single charged monoprotonated ions ([AgNMS+H]+, m/z 414.95), as well as minor sodium and potassium adducts, [AgNMS+Na]+ and [AgNMS+K]+ at m/z 436.93 and 444.24, respectively. There were also observed, even in low abundance, the dimeric complex ([Ag2NMS2+H]+, m/z 828.91), as well as the respective sodium and potassium adducts. Trimeric ions were only observed as sodium and potassium adducts ([Ag3NMS3+Na]+ and [Ag3NMS3+K]+) at m/z 1264.82 and 1280.80, respectively. The free NMS ligand was also observed, mainly as sodium and potassium adducts of m/z 331.04 and 347.01, respectively. The presence of

Fig. 7. Optimized structure for the Ag–NMS complex.

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Fig. 8. Kohn–Sham orbitals for (a) NMS and (b) Ag–NMS complex. Silver (gray), carbon (dark gray), nitrogen (blue), oxygen (red) and hydrogen (white).

Table 3 Antibiotic sensitive profile of bacterial strains against NMS, Ag–NMS, AgNO3, gentamicine and ceftriaxone. Compounds

NMS AgNO3 Ag–NMS Ceftriaxone Gentamicine

Results P. aeruginosa

S. aureus

Inhibition zone diameter (mm)

Inhibition zone diameter (mm)

Inhibition zone diameter (mm)

0.0 20.0 12.0 34.0 24.0

0.0 25.0 (±0.1) 18.0 (±0.1) >42.0 (±0.1) 24.0 (±0.1)

0.0 12.0 18.0 30.0 20.0

(±0.1) (±0.1) (±0.1) (±0.1)

sodium and potassium ions is most probably due to the use of potassium hydroxide during the synthesis of the complex. An isotope pattern comparison for the monoprotonated [AgNMS+H]+ ion (see Supplementary data #1) shows a good agreement with the theoretical predictions. The mass error was 0.5 ppm for the monoprotonated ion C13H11N2O5SAg+ (Calc. m/z 414.9518, exp. m/z 414.9516). To further confirm the proposed composition of the [AgNMS+H]+ species, fragment ion spectra were acquired for the monoprotonated ion (Fig. 6). The fragment ion spectrum of [AgNMS+H]+shows the loss of CH3SO2 (79 Da), which corresponds to the methyl-sulphone group of the NMS ligand, followed by the loss of Ag (107 Da), as well a direct loss of the NMS ligand (308 Da) from the precursor. The obtained data confirmed the previously proposed composition.

E. coli

(±0.1) (±0.1) (±0.1) (±0.1)

3.5. Molecular modeling Geometries of NMS and Ag–NMS were analyzed by theoretical calculations using density functional theory (DFT) studies. The equilibrium geometries were confirmed by the vibrational analysis, showing no imaginary frequencies. The calculated geometry for NMS is in a good agreement with the previously reported crystallographic data [40]. According to the IR and NMR spectroscopic data, NMS is coordinated to the silver atom through the nitrogen of the sulphonamide group, and the oxygen of sulphonyl group could also be participating in ligand coordination. So, in order to confirm if coordination of NMS to Ag(I) occurs in a monodentate form through the N atom or

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in a bidentate form through the N and O atoms of the (O2SNH) group, calculations were performed for the mono and bidentate coordinations. Attempts to obtain coordination of NMS to the metal solely through nitrogen atom were unsuccessful since the structure converged to coordination through the nitrogen atom of sulphonamide and also through the oxygen of the sulphonyl group. Calculations confirmed that the bidentate structure is a minimum in the potential energy surface (PES), with the silver atom coordinating through both the nitrogen and the oxygen atoms of sulphonamide group. The optimized structure for the Ag–NMS complex is presented in the Fig. 7. The calculated Ag–N1 and Ag– O1 bond distances are 1.952 and 2.572 Å. The calculated angles N1–Ag–O1 and Ag–N1–S are 66.2° and 83.2°. The detailed bond distances and angles are reported in Supplementary data #2. The simulated IR spectra of NMS and Ag–NMS are in good agreement with the experimental spectra, confirming all the assignments. The simulated N–H stretching in the NMS spectrum appears at 3421 cm1 and the experimental data shows it at 3284 cm1. Experimental absorptions in the IR spectrum of NMS at 1342 and 1153 cm1 were confirmed by the simulated spectra as the (O@S@O) asymmetrical and symmetrical stretching modes, respectively. In this case, the calculated frequencies were1378 and 1134 cm1, respectively. The experimental spectrum of Ag–NMS shows a shift to low energies of both O@S@O stretching modes of 48 cm1 after coordination of the ligand to Ag(I). The simulated spectra show that the asymmetrical stretching mode is expected to be shifted to lower energies by 86 cm1, which corroborates to the proposition of the participation of one of the oxygen atoms of the sulphonyl group in the coordination of NMS to Ag(I). The Time Dependent (TD-DFT) calculations permitted us to explain the nature of the transitions observed in the UV–Vis spectra. The predicted absorptions were showed with the experimental electronic absorption spectra for both NMS and Ag–NMS in Fig. 5. All the observed bands are p–p⁄ transitions and the HOMO–LUMO is the low-lying one. As observed in Fig. 8, these Kohn–Sham orbitals are centered in the aryl groups and the LUMO has a significant influence from the NO2 group, explaining why this transition is low-lying. Essentially, all other transitions are of the same kind since all of them have origin in p orbitals. As seen in Fig. 8, the frontier Kohn–Sham orbitals for the complex are very similar to the NMS ones. Apart from small shifts in the energies, calculations confirm that coordination does not change the nature of the transitions. Since the calculated oscillator strength is higher for Ag–NMS when compared to NMS, the p–p⁄ transitions bands are more intense in the complex (data available in Supplementary data #3). 3.6. Biological assays The Ag–NMS complex was evaluated concerning its antibacterial activities. Table 3 summarizes the results obtained for NMS and the Ag–NMS complex against Gram-negative (E. coli and P. aeruginosa) and Gram-positive (S. aureus) microorganisms. Silver nitrate and the comercial antibiotics ceftriaxone and gentamicine were used as positive controls. It was observed that impregnated paper discs with Ag–NMS exhibited inhibition zones for P. aeruginosa, S. aureus and E. coli, of 12.0 ± 0.1, 18.0 ± 0.1 and 18.0 ± 0.1 mm, respectively. These results confirm the sensitivity of the tested bacterial cells to the Ag–NMS complex. The observed data for the complex are similar to silver nitrate and gentamicine for the same tested strains, and also to the Ag(I)–acesulfame complex early reported [20]. Nimesulide alone did not show inhibitory activity against the considered bacterial strains under the same experimental conditions. As reported earlier, silver ions are considered responsible for the antibiotic activity of silver complexes due to their high reactivity,

binding capacity to tissue proteins and capacity to lead to structural changes in the bacterial cell wall. Silver also binds to bacterial DNA and RNA, leading to inhibition of bacterial replication [41,42]. The results obtained show the potential application of the Ag–NMS complex, for example, in topic formulations for the treatment of wounds and burns. 4. Conclusions The molar composition of the Ag–NMS complex was found to be 1:1 M/L. Infrared, ESI-QTOF-MS, UV–Vis and 1H, 13C and 15N NMR spectroscopic data permitted proposing coordination of the ligand to Ag(I) through the nitrogen and oxygen atoms of sulphonamide group. Biological assays showed antibacterial activity of the complex against Gram-positive and -negative bacterial strains. Acknowledgments This study was supported by grants from the Brazilian Agencies FAPESP (São Paulo State Research Foundation, proc. 2006/55367-2 and2008/57805-2), and CNPq (Conselho Nacional de Desenvolvimento Científico e Tecnológico, proc. 472468/2010-3 and 573672/2008-3). Professor Corbi is also grateful to Professor Carol H. Collins for English revision of the manuscript. Appendix A. Supplementary data Supplementary data associated with this article can be found, in the online version, at doi:10.1016/j.poly.2012.02.002. References [1] [2] [3] [4] [5] [6] [7] [8] [9] [10] [11] [12] [13] [14]

[15] [16] [17] [18] [19]

[20]

[21] [22]

[23] [24] [25] [26] [27] [28]

I. Tsyba, B.B.-K. Mui, R. Naguchi, K. Nomiya, Inorg. Chem. 42 (2003) 8028. K. Nomiya, S. Takahashi, R. Noguchi, J. Chem. Soc., Dalton Trans. (2000) 2091. H.J. Klasen, Burns 26 (2000) 117. R.H. Demling, L. DeSanti, Wounds 13 (Suppl A) (2001) 4. I. Chopra, J. Antimicrob. Chemother. 59 (2007) 587. R.H. Demling, M.D.L. DeSanti, Burns 28 (2002) 264. C.A. Moyer, L. Brentano, D.L. Gravens, H.W. Margraf, W.W. Monafo, Arch. Surg. 90 (1965) 812. C.G. Bellinger, H. Conway, Plast. Reconstr. Surg. 45 (1970) 582. S. Silver, L.T. Phung, G. Silver, J. Ind. Microbiol. Biotechnol. 33 (2006) 627. R.G. Sibbald, H. Orsted, G.S. Schultz, P. Coutts, D. Keast, Ost. Wound Man. 49 (2003) 23. S.A. Jones, P.G. Bowler, M. Walker, D. Parsons, Wound Repair Regen. 12 (2004) 288. Md.S.A.S. Shah, M. Nag, T. Kalagara, S. Singh, S.V. Manorama, Chem. Mater. 20 (2008) 2455. K. Nomiya, H. Yokoyama, J. Chem. Soc., Dalton Trans. (2002) 2483. A. Kascatan-Nebioglu, A. Melaiye, K. Hindi, S. Durmus, M.J. Panzner, L.A. Hogue, R.J. Mallett, C.E. Hovis, M. Coughenour, S.D. Crosby, A. Milsted, D.L. Ely, C.A. Tessier, C.L. Cannon, W.J. Youngs, J. Med. Chem. 49 (2006) 6811. D.P. Rocha, G.F. Pinto, R. Ruggiero, C.A. de Oliveira, W. Guerra, A.P.S. Fontes, T.T. Tavares, I.M. Marzano, E.C. Pereira-Maia, Quim. Nova 34 (2011) 111. A.S. Kazachenko, A.V. Legler, O.V. Per’yanova, Y.A. Vstavskaya, Pharm. Chem. J. 34 (2000) 257. B. Ruan, Y. Tian, H. Zhou, J. Wu, Z. Liu, C. Zhu, J. Yang, H. Zhu, J. Organomet. Chem. 694 (2009) 2883. A.A. Isab, M.I.M. Wazeer, Spectrochim. Acta A 65 (2006) 191. M. Poyraz, C.N. Banti, N. Kourkoumelis, V. Dokorou, M.J. Manos, M. Simcic, S. Golic-Grdadolnik, T. Mavromoustakos, A.D. Giannoulis, I.I. Verginadis, K. Charalabopoulos, S.K. Hadjikakou, Inorg. Chim. Acta 375 (2011) 114. M. Cavicchioli, A.C. Massabni, T.A. Heinrich, C.M. Costa-Neto, E.P. Abrão, B.A.L. Fonseca, E.E. Catellano, P.P. Corbi, W.R. Lustri, C.Q.F. Leite, J. Inorg. Biochem. 104 (2010) 533. B.J.R. Whittle, Fundam. Clin. Pharmacol. 17 (2003) 275. M.W. Schmidt, K.K. Baldridge, J.A. Boatz, S.T. Elbert, M.S. Gordon, J.H. Jensen, S.K.N. Matsunaga, K. Nguyen, S. Su, T.L. Windus, M. Dupuis, J.A. Montgomery Jr., J. Comput. Chem. 14 (1993) 1347. P.J. Hay, W.R. Wadt, J. Chem. Phys. 82 (1985) 299. R. Ditchfie, W.J. Hehre, J.A. Pople, J. Chem. Phys. 54 (1971) 724. W.J. Hehre, R. Ditchfie, J.A. Pople, J. Chem. Phys. 56 (1972) 2257. M.M. Francl, W.J. Pietro, W.J. Hehre, J.S. Binkley, M.S. Gordon, D.J. DeFrees, J.A. Pople, J. Chem. Phys. 77 (1982) 3654. P.C. Harihara, J.A. Pople, Theor. Chim. Acta 28 (1973) 213. A.D. Becke, J. Chem. Phys. 98 (1993) 5648.

R.E.F. de Paiva et al. / Polyhedron 36 (2012) 112–119 [29] [30] [31] [32] [33] [34]

C.T. Lee, W.T. Yang, R.G. Parr, Phys. Rev. B37 (1988) 785. A.P. Scott, L. Radom, J. Phys. Chem. 100 (1996) 16502. G. Schaftenaar, J.H. Noordik, J. Comput.-Aided Mol. Des. 14 (2000) 123. S. Miertus, E. Scrocco, J. Tomasi, Chem. Phys. 55 (1981) 117. A.W. Bauer, W.M. Kirby, J.C. Sheris, M. Turck, Am. J. Clin. Pathol. 45 (1996) 493. Clinical and Laboratory Standards Institute, Performance standards for antimicrobial susceptibility testing, seventeenth informational supplement, Wayne, PA, USA, 2007. [35] (a) N. Juranic, S. Macura, Inorg. Chim. Acta 217 (1994) 213; (b) P.P. Corbi, A.C. Massabni, T.A. Heinrich, C.M. Costa-Neto, J. Coord. Chem. 61 (2008) 2470; (c) P.P. Corbi, A.C. Massabni, Spectrochim. Acta A 64 (2006) 418.

119

[36] C. Abbehausen, J.F. Castro, M.B.M. Spera, T.A. Heinrich, C.M. Costa-Neto, W.R. Lustri, A.L.B. Formiga, P.P. Corbi, Polyhedron 30 (2011) 2354. [37] J.N. Baxter, J. Cymerman-Craig, J.B. Willis, J. Chem. Soc. (1955) 669. [38] M. Goldstein, M.A. Russell, H.A. Willis, Spectrochim. Acta A 25 (1969) 1275. [39] K. Nakamoto, Infrared and Raman Spectra of Inorganic and Coordination Compounds – Part B, 5th ed., John Wiley & Sons, New York, USA, 1997. pp. 59– 104. [40] L. Dupont, B. Pirotte, B. Masereel, J. Delarge, J. Geczy, Acta Crystallogr., Sect. 51 (1995) 507. [41] M. Rai, A. Yadav, A. Gade, Biotechnol. Adv. 27 (2009) 76. [42] J.J. Castellano, S.M. Shafii, F. Ko, G. Donate, T.E. Wright, R.J. Mannari, Int. Wound J. 4 (2007) 114.