Microwave Synthesis, Spectral, Thermal, and Pharmacological Evaluation of Some Metal Complexes Containing Benzene Sulfonyl Thiourea Moiety

Hassan AM¹, Abd El-Wahab ZH¹, Salman A A¹, Abd El-Rahman NM², Salama TAM³, El-Tantawy ASM⁴ and Khalid MW^5*
*Correspondence: Khalid MW, Department of Pharmaceutical Chemistry, National Organization for Drug Control and Research, Giza, Egypt, Tel: +201007068070, Email:

Keywords

Thermal analysis • FT-IR • ¹HNMR

Introduction

Metal complexes are being nowadays widely used as analytical and antimicrobial reagents owing to their versatile activities. Also, metal complexes have been found to possess important biological and catalytic activity [1]. The uprising need to understand bio-coordination chemistry helps in improving the design of metal complexes and interpreting their kinetics and thermodynamic actions under biologically relevant conditions [2]. Metal complexes have become of great influence in the medical practice [3]. In some cases, they were used as antibacterial [4], antifungal [4], antioxidant [5], anti-inflammatory [6], and anticancer [7] agents.

Sulfonamides are synthetic antibiotics that inhibit bacterial multiplication. They occupy a unique position in the drug industry and exhibit a wide spectrum of biological activities. A sulfonyl group plays an important role as a key constituent of number of biologically active molecules. Several drugs containing sulfonamide functionality are in clinical uses which include antibacterial and antifungal drug, carbonic anhydrase inhibitors and antiinflammatory agents [8]. They are also found to have extensive applications in cancer chemotherapy. A general method for the synthesis of sulfonamides involves the coupling of sulfonyl chloride with primary or secondary amine or a substituted amine [9]. The presence of several potential donor sites, e.g. the sulfonamidic nitrogen, one amino nitrogen, and two sulfonyl oxygen atoms make them versatile complexing agents with metal ions.

Thiourea derivatives (TUD) are important class of molecules having chemical and biological applications. They are of marked significance industrially and physiologically. They, also, have been incorporated in various therapeutics as anti-inflammatory [10], antioxidant [11], antibacterial [12], antifungal [13] and anticancer [14] agents. Based on the just mentioned properties of both sulfonyl and thiourea groups, we are taking the advantage of incorporating both in the synthesis and characterization of organic ligand (H₂L¹) N-(2-chloro phenyl)-N’ benzenesulfonyl thiourea and its incorporation in metal complex formation to be tested for its biological activity.

In the background of green chemistry, microwave-assisted synthesis provide a clean, fast, low cost, eco-friendly, solvent free and higher yield rather than conventional heating processes, aiming to introduce energy into the system [15]. In this work, we present the synthesis of Pt (II), Pd (II) and Au (III) metal complexes and their coordination behavior towards the unsymmetrical organic ligand N-(2-chloro phenyl)-N ’ benzenesulfonyl thiourea using microwave-assisted technique. The prepared metal complexes have been characterized by elemental analysis, FT-IR, NMR, UVVis, thermal analysis and magnetic susceptibility. They were tested for their antibacterial activity against Gram positive, Gram-negative bacteria as well as fungi. Also anti-cancer assay was applied against two cell lines; breast cancer cells (MCF-7) and liver cancer cells (HepG-2). The molecular compositions of the complexes were proposed on the basis of chemical analyses and spectrophotometric evidences. The metal complexes obtained can be used as precursors for the formulation of new compounds with significant biological activities.

Experimental Procedure

Chemicals and instrumentation

All chemicals used in the present work were of analytical AR grade. All the reagents were obtained from Sigma Aldrich. Metals were purchased from Loba Chemie. The solid reflectance spectra of the ligand and their metal complexes were recorded in the solid state on a Shimadzu 3101 PC spectrophotometer in the UV/Vis range of 200-800 nm. The TG/DTG were recorded from ambient to 1000°C with a heating rate of 10°C min^-1 under a nitrogen atmosphere (20 mL min^-1) using Shimadzu TG-50H and DTA-60H thermal analyzer. Elemental analysis of the constituting elements was carried out on Perkin Elmer 2400 model. The metal cations were determined using Atomic absorption instrument Perkin Elmer (AAS) for the quantitative determination of chemical elements using the absorption of optical radiation (light) by free atoms in gaseous state. The IR spectra were recorded using KBr pellets in the region 4000-400 cm^-1 on FT-IR spectrophotometer Shimadzu 8201. The ¹H-NMR spectra were recorded on a varian F.T-300 MHz spectrometer using TMS as internal standard and DMSO d6 as solvent. The molar conductivities were performed in DMF solution (1 × 10^-3 M), by using Jenway 4010 conductivity meter. The Magnetic susceptibility of the metal complexes in the solid state was recorded at room temperature on a Sherwood magnetic susceptibility balance. Mass spectra of the solid ligand and its metal complexes were measured at 70 eV on a JEOL JMS-AX 500 spectrometer.

Ligand and their metal complexes were subjected to be tested for in-vitro antibacterial activity against two species of Gram positive bacteria and two species of Gram negative bacteria as well as two species of fungi. Also anticancer assay was tested against two cell lines; breast cancer cells (MCF-7) and liver cancer cells (HepG-2).

Ligand (H₂L¹) synthesis

The ligand under study involves a two- step process for its preparation. The first step includes 1 h refluxing of equimolar amounts of benzenesulfonyl chlorine (0.01 mol) and ammonium thiocyanate (0.01 mol) dissolvedin acetone (50 mL) to form benzene sulfonyl isocyanate. Secondly, 4 h refluxing of equimolar amounts of benzenesulfonyl isothiocyanate (0.01 mol) with 2- chloro aniline (0.01 mol) in (50 mL) acetone. The proposed chemical formula of the formed H₂L¹ is C₁₃H₁₁N₂O₂S₂Cl (H₂L¹) and the chemical structure is as shown in Figure 1.

The obtained solid was filtered, recrystallized using ethanol and dried over anhydrous CaCl₂ in a dessicator.

Preparation of Pd (ll), Pt (ll) and Au (lll) complexes using microwave-assisted irradiation

The lN-(2-chloro phenyl)-N’ benzenesulfonyl thiourea ligand and the metal salts (Au (III) chloride trihydrate, Pt (II) chloride anhydrous and Pd (II) chloride anhydrous) were mixed in equimolar ratio (1:1) (metal: ligand) in a grinder.

Then the reaction mixture of each was irradiated in 1-3 mL of dry ethanol. The reaction was completed in a short time (3-10 min) with a high % yield.

The obtained product was washed several times with ethanol and ether and finally dried in a desiccator under reduced pressure over anhydrous CaCl₂. The progress of the reaction and purity of the product was monitored by TLC using silica gel G (yield: 89-90%). The results are tabulated in the Table 1 and Table 2 and from the results we can conclude that the future scope of the investigated compound.

Pharmacological activity

The obtained metal complexes were screened for their activities as antibacterial, against Gram-positive of Streptococcus pneumonia (RCMB 010010) and Bacillus subtilis (RCMB 010067); Gram-negative species of Pseudomonas aeruginosa (RCMB 010043) and Escherichia coli (RCMB 010052) as well as anti-fungal of Aspergillus fumigatus (RCMB 02568) and Candida albicans (RCMB 05036). Antimicrobial activity was tested by the disc diffusion method [16]. The Amphotricine B, Ampicillin and Gentamicine were used as standard references for Gram-positive, Gram-negative bacteria and fungi, respectively, serving as positive controls. Nutrient agar was prepared then autoclaved at 121°C for 15 min, cooled and finally poured in Petri dishes. Water/methanol (99:1 v/v) was used as solvent. The filter paper discs were soaked in different dilutions of the ligand and its metal complexes (50 to 1000 ppm) to find out the minimum concentration required for inhibition of microbial growth. The discs were then dried and placed in Petri dishes previously spotted with the just mentioned antibacterial and antifungal species. After 24 h incubation at 37°C, the inhibition zone around each disc was measured in mm by the help of a Zone Reader. The average value was determined in triplicate readings for each zone of inhibition in millimeter. The highest antibacterial action was for H₂L¹ complex against species. It is obvious that the incorporation of the ligand in metal complexes exhibited higher biological activity. This may be attributed to the donated coordination bonds to the central metal ions in metal chelates, thus delocalizing the - electron over the chelate ring. Accordingly, increasing the lipophilicity of the metal ion, which favors the metal complex permeation through the lipoid layers of the bacterial and fungal membranes [17,18].

Cytotoxic activity

Thiourea derivatives play an important role as anticancer agents [19], due to their good inhibitory activity against protein tyrosine kinases (PTKs) [20]. H₂L¹ ligand, also a thiourea derivative, and its metal complexes, were tested for their inhibitory activity towards tumor cell lines of breast (MCF-7) and liver cancer cells (HepG-2). An MTT colorimetric assay was used to plot a dose response curve required to kill 50% of cell population (IC₅₀). The results are shown in Tables 3 and 4 and Figure 2. The most active against the liver carcinoma cell line (HepG2), showing an IC₅₀ value of 4.8 μg mL^-1 compared to 4.9 and 18.8 μg mL-1 for the known anticancer drugs, 5- flourouracil and cisplatin, respectively. A lower tendency was observed towards breast cancer cells inhibition than those observed for liver carcinoma. The breast cancer cell line (MCF-7) inhibition for the metal complexes was 4a, 3c, 4b, and 4c, in descending order.

Applications

Anti-microbial activity

The spectrum, Figure 15 and Table 3; reveals a noticeable growth inhibition of Gram-positive, Gram-negative bacteria and fungi examined at concentrations as low as 5 μg/mL.

Anticancer activity

The free ligand H₂L¹, H₃L₂ may have a promising potential as an anticancer agent, attributed to the ligands possible alternative modes of cytotoxic action. The cytotoxicity activities of Au- H₂L¹ was tested against (MCF-7) and (HepG-2) human tumor cell lines. The reported results in terms of IC₅₀ value was recorded in Table 4, Figures 16 and 17 for comparison, the cytotoxicity of Cisplatin, as standard antitumor drug was evaluated and produced (IC₅₀ mL) under the same conditions, Au- H₂L¹> H₂L¹ against breast carcinoma cells; H₂L¹ > Au- H₂L¹ against Hepatocellular carcinoma cells.

From the obtained results;

Au- H₂L¹ has more efficiency than H₂L¹on the breast carcinoma cells.

Au- H₂L¹ has less efficiency than H₂L¹ on the Hepatocellular carcinoma cell.

Conclusion

The prepared metal complexes have been characterized by elemental analysis, FT-IR, 1HNMR, UV-Vis, thermal analysis and magnetic susceptibility. They were tested for their antibacterial activity against Gram positive, Gram-negative bacteria as well as fungi. Also anti-cancer assay was applied against two cell lines; breast cancer cells (MCF-7) and liver cancer cells (HepG-2). The IR spectral data indicate that the ligand H₂L¹ coordinates to metal ions through sulfonyl and thionyl groups in a tetradentate square planer manner for PdII and PtII ions while in case of AuIII it takes an octahedral shape. Based on spectrophotometric investigations, it is suggested that Pd(II), Pt(II) and Au (III) form 1:1 complexes. The molecular compositions of the complexes were proposed on the basis of chemical analyses and spectrophotometric evidences.

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