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http://informahealthcare.com/enz ISSN: 1475-6366 (print), 1475-6374 (electronic) J Enzyme Inhib Med Chem, Early Online: 1–10 ! 2013 Informa UK Ltd. DOI: 10.3109/14756366.2013.815178

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ORIGINAL ARTICLE

Metal-based carboxamide-derived compounds endowed with antibacterial and antifungal activity Muhammad Hanif1, Zahid H. Chohan1, Jean-Yves Winum2, and Javeed Akhtar3 1

Institute of Chemical Sciences, Bahauddin Zakariya University, Multan, Pakistan, 2Institut des Biomole´cules Max Mousseron (IBMM), UMR 5247, CNRS-UM1-UM2, Baˆtiment de Recherche Max Mousseron, Ecole Nationale Supe´rieure de Chimie de Montpellier, Montpellier Cedex, France, and 3 Nanoscience & Materials Synthesis Group, Department of Physics, COMSATS CIIT, Islamabad, Pakistan Abstract

Keywords

A series of three bioactive thiourea (carboxamide) derivatives, N-(dipropylcarbamothioyl)thiophene-2-carboxamide (L1), N-(dipropylcarbamothioyl)-5-methylthiophene-2-carboxamide (L2) and 5-bromo-N-(dipropylcarbamothioyl)furan-2-carboxamide (L3) and their cobalt(II), copper(II), nickel(II) and zinc(II) complexes (1)–(12) have been synthesized and characterized by their IR,1H-NMR spectroscopy, mass spectrometry and elemental analysis data. The Crystal structure of one of the ligand, N-(dipropylcarbamothioyl)thiophene-2-carboxamide (L1) and its nickel(II) and copper(II) complexes were determined from single crystal X-ray diffraction data. All the ligands and metal(II) complexes have been subjected to in vitro antibacterial and antifungal activity against six bacterial species (Escherichia coli, Shigella flexneri, Pseudomonas aeruginosa, Salmonella typhi, Staphylococcus aureus and Bacillus subtilis) and for antifungal activity against six fungal strains (Trichophyton longifusus, Candida albicans, Aspergillus flavus, Microsporum canis, Fusarium solani and Candida glabrata). The in vitro antibacterial and antifungal bioactivity data showed the metal(II) complexes to be more potent than the parent ligands against one or more bacterial and fungal strains.

Antibacterial, antifungal, carboxamides, crystal structures, metal(II) complexes

Introduction Despite the availability of a large number of antimicrobials, the emergence of antibiotic resistance at the same time has created a substantial therapeutic need for designing and synthesis of new class of compounds endowed with antimicrobial activity, possibly acting through mechanism of action, distinct from known antimicrobial agents. Some metals have been used as drugs to treat various diseases and conditions. Platinum compounds, cisplatin (cis-[Pt(NH3)2Cl2]), carboplatin and oxaliplatin are among the most widely used cancer therapeutic agents. Gold drugs, myocrisin and auranofin are used for the treatment of rheumatoid arthritis. Thiourea (carboxamide) and its derivatives display a wide range of bioactivity1–4. Studies have indicated that thioureas show even a strong antifungal activity comparable to the commonly used antifungal, ketoconazole5. Antimicrobial and insecticidal properties of thioureas have been well-documented6,7 and used not only in the control of plant pathogenic fungi8 but also have been shown to possess antitubercular, antithyroid, anthelmintic, antibacterial, insecticidal and rodenticidal properties9,10. The process of chelation/coordination plays an important role in biological systems11. It has been suggested that biological activity of many compounds is enhanced12–14 upon chelation with the metal(II) ions. The oxygen and sulfur atoms of carbonyl (C ¼ O) and thiocarbonyl (C ¼ S) groups in thiourea potentially Address for correspondence: Zahid H. Chohan, Institute of Chemical Sciences, Bahauddin Zakariya University, Multan, 60800, Pakistan. Tel: +92 619210142. E-mail: [email protected]

History Received 3 May 2013 Revised 27 May 2013 Accepted 7 June 2013 Published online 31 July 2013

act as donor atoms for coordination with the metal(II) ions. The complexation capability of thioureas is well established15, and the bioactivities of different metal complexes of thiourea have been studied for antifungal activity against phytopathogenic fungi and yeast16–18. In view of the significant structural and biological behavior, we wish to report herein the synthesis of three carboxamide derivatives: N-(dipropylcarbamothioyl)thiophene2-carboxamide (L1), N-(dipropylcarbamothioyl)-5-methylthiophene-2-carboxamide (L2) and 5-bromo-N-(dipropylcarbamothioyl)furan-2-carboxamide (L3) and their cobalt(II), nickel(II), copper(II) and zinc(II) complexes (1)–(12). These compounds have been further investigated for their antimicrobial profile against four Gram-negative (Escherichia coli, Shigella sonnei, Pseudomonas aeruginosa, Salmonella typhi) and two Gram-positive (Staphylococcus aureus, Bacillus subtilis) bacterial strains and, for antifungal activity against six fungal strains, Trichophyton longifusus, Candida albicans, Aspergillus flavus, Microsporum canis, Fusarium solani and Candida glaberata. In this study, we also wish to report the crystal structures of the ligand N-(dipropylcarbamothioyl)thiophene-2-carboxamide (L1) and two of its nickel(II) and copper(II) complexes (2) and (3).

Experimental All chemicals used were of reagents grade. All metal salts were used as acetate. Melting points were recorded on Fisher Johns melting point apparatus. Infrared spectra were recorded on Shimadzu FT-IR spectrometer. The C, H and N analyses was carried out using a Perkin Elmer (Waltham, MA) model. The 1H

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M. Hanif et al.

and 13C-NMR spectra were recorded in DMSO-d6 using TMS as internal standard on a Bruker Spectrospin Avance DPX-500 spectrometer. Electron impact mass spectra (EIMS) were recorded on JEOL MS Route Instrument. In vitro antibacterial and antifungal properties were studied at HEJ Research Institute of Chemistry, International Centre for Chemical Sciences, University of Karachi, Pakistan, and Department of Chemistry, The Islamia University, Bahawalpur, Pakistan.


Synthesis of N-(dipropylcarbamothioyl)thiophene-2-carboxamide (L1) The ligand, N-(dipropylcarbamothioyl)thiophene-2-carboxamide (L1) was synthesized by the reported method19, in which a solution of thiophenyl carbonyl chloride (20 mmol, 2.14 mL) in dry acetone (30 mL) was added dropwise to a solution of KSCN (20 mmol, 1.94 g) in acetone (30 mL). The reaction mixture was refluxed for 1 h and then cooled to room temperature. A solution of dipropylamine (20 mmol, 2.2 mL) in acetone (20 mL) was added dropwise to the mixture during 20 min and refluxing continued for 2 h. Then, KCl was removed by filtration and light yellow filtrate was placed into water (250 mL) and few drops of conc. HCl were added under constant agitation. A wax-like material was formed which was separated by filtration, washed several times with cold water and then with diethyl ether. The crude product was purified by recrystallization from a solution mixture of ethanol-dichloromethane (1:1). As a result of recrystallization, fine shiny colorless crystals of (L1) were obtained, which were suitable for X-rays analysis. The same procedure was used for the preparation of other new ligands (L2) and (L3). N-(Dipropylcarbamothioyl)thiophene-2-carboxamide (L1) Yield: 82%. Color (white crystalline solid). M.p. 134 C. IR (KBr, cm1): 3265 (NH), 3075–3025 (CH), 1675 (C ¼ O), 1580 (aromatic C C), 1240 (C ¼ S), 890 (C–S). 1H NMR (DMSO–d6, , ppm): 11.51 (s, 1H, CONH), 8.18 (d, 1H, J ¼ 5.5 Hz, thienyl C4–H), 8.0 (d, 1H, J ¼ 4.2 Hz, thienyl C2–H), 7.35 (dd, 1H, J ¼ 5.5, 4.2 Hz, thienyl C3–H), 2.50 (t, 4H, J ¼ 6.84 Hz, C7,10–H), 1.25 (sextet, 4H, J ¼ 6.84, 6.86 Hz, C8,11–H), 0.95 (t, 6H, J ¼ 6.86 Hz, C9,12–H). 13C NMR (DMSO–d6, , ppm): 178.75 (C ¼ S), 171.0 (C ¼ O), 137.73 (C1), 135.13 (C2), 134.53 (C3), 132.78 (C4), 49.5 (C7, C10), 37 (C8, C11), 20.95 (C9, C12). EIMS (70 eV) m/z: 270.42. Anal. Calcd. for C12H18N2S2O (270.42): C: 53.30; H: 6.71; N: 10.36; S: 23.72; Found: C: 53.26; H: 6.69; N: 10.38; S: 23.70%. N-(Dipropylcarbamothioyl)-5-methylthiophene-2-carboxamide (L2) Yield: 79%. Color (white crystalline solid). M.p. 127 C. IR (KBr, cm1): 3270 (NH), 3065–3030 (CH), 1670 (C ¼ O), 1577 (aromatic C ¼ C), 1250 (C ¼ S), 885 (C–S). 1H NMR (DMSO– d6, , ppm): 11.56 (s, 1H, CONH), 7.75 (d, 1H, J ¼ 4.1 Hz, thienyl C2–H), 7.15 (d, 1H, J ¼ 4.1 Hz, thienyl C3–H), 2.55 (t, 4H, J ¼ 6.84 Hz, C7,10–H), 2.50 (s, 3H, thienyl –CH3), 1.27 (sextet, 4H, J ¼ 6.84, 6.85 Hz, C8,11–H), 0.92 (t, 6H, J ¼ 6.85 Hz, C9,12– H). 13C NMR (DMSO–d6, , ppm): 178.92 (C ¼ S), 171.32 (C ¼ O), 139.73 (C1), 137.13 (C4), 135.53 (C2), 130.18 (C3), 50.5 (C7, C10), 37.2 (C8, C11), 20.75 (C9, C12), 15.5 (thienyl –CH3). EIMS (70 eV) m/z: 284.44. Anal. Calcd. for C13H20N2S2O (284.44): C: 54.89; H: 7.09; N: 9.85; S: 22.55; Found: C: 54.85; H: 7.11; N: 9.90; S: 22.59%. 5-Bromo-N-(dipropylcarbamothioyl)furan-2-carboxamide (L3) Yield: 80%. Color (off white crystalline solid). M.p. 119 C. IR (KBr, cm1): 3275 (NH), 3070–3030 (CH), 1680 (C ¼ O), 1575

J Enzyme Inhib Med Chem, Early Online: 1–10

(aromatic C ¼ C), 1245 (C ¼ S), 1150 (C–O), 685 (C–Br). 1H NMR (DMSO–d6, , ppm): 11.60 (s, 1H, CONH), 7.62 (d, 1H, J ¼ 3.5 Hz, furanyl C2–H), 7.45 (d, 1H, J ¼ 3.5 Hz, furanyl C3–H), 2.60 (t, 4H, J ¼ 6.84 Hz, C7,10–H), 1.22 (sextet, 4H, J ¼ 6.86 Hz, C8,11–H), 0.90 (t, 6H, J ¼ 6.84, 6.86 Hz, C9,12–H). 13C NMR (DMSO–d6, , ppm): 178.90 (C ¼ S), 171.5 (C ¼ O), 139.73 (C1), 135.13 (C4), 130.53 (C2), 127.78 (C3), 50 (C7, C10), 38 (C8, C11), 21.10 (C9, C12). EIMS (70 eV) m/z: 333.24. Anal. Calcd. for C12H17BrN2S2O (333.24): C: 43.25; H: 5.14; N: 8.41; S: 9.62; Found: C: 43.30; H: 5.17; N: 8.38; S: 9.65%. General procedure for the synthesis of novel metal(II) complexes (1)–(12) To a stirred solution of the respective ligands (1 mmol) in methanol (20 mL) was added dropwise a solution of appropriate metal(II) acetate. nH2O (n ¼ 0, 1 or 4) (0.5 mmol) in methanol (15 mL). The reaction mixture was refluxed for 2 h. Colored precipitates were formed during refluxing. The precipitated product thus formed was filtered, washed with methanol and dried under vacuum. The precipitates were dissolved in a mixture of ethanol and dichloromethane (1:1) and a clear solution was kept in a refrigerator for one week. Suitable pink and green crystals for X-ray studies were obtained for complexes (2) and (3). The same method was used for the preparation of all other complexes. Physical, analytical and spectral data are given in Tables 1 and 2. NMR data of the Zn(II) complexes [Zn (L1-H)2] (4) 1

H NMR (DMSO–d6, , ppm): 8.26 (d, 2H, J ¼ 5.5 Hz, thienyl C4–H), 8.06 (d, 2H, J ¼ 4.2 Hz, thienyl C2–H), 7.36 (dd, 2H, J ¼ 5.5, 4.2 Hz, thienyl C3–H), 2.65 (t, 8H, J ¼ 6.84 Hz, C7,10–H), 1.29 (sextet, 8H, J ¼ 6.84, 6.86 Hz, C8,11–H), 1.0 (t, 12H, J ¼ 6.86 Hz, C9,12–H). 13C NMR (DMSO–d6, , ppm): 180.0 (C ¼ S), 172.4 (C ¼ O), 137.98 (C1), 135.47 (C2), 134.83 (C3), 133.18 (C4), 49.72 (C7, C10), 37.20 (C8, C11), 21.10 (C9, C12). [Zn (L2-H)2] (8) 1

H NMR (DMSO–d6, , ppm): 7.83 (d, 2H, J ¼ 4.1 Hz, thienyl C2–H), 7.21 (d, 2H, J ¼ 4.1 Hz, thienyl C3–H), 2.71 (t, 8H, J ¼ 6.84 Hz, C7,10–H), 2.55 (s,6H, thienyl –CH3) 1.33 (sextet, 8H, J ¼ 6.84, 6.85 Hz, C8,11–H), 0.96 (t, 12H, J ¼ 6.85 Hz, C9,12–H). 13 C NMR (DMSO–d6, , ppm): 180.5 (C ¼ S), 172.65 (C ¼ O), 139.99 (C1), 137.30 (C4), 135.77 (C2), 130.34 (C3), 50.66 (C7, C10), 37.35 (C8, C11), 20.90 (C9, C12), 15.64 (thienyl -CH3). [Zn (L3-H)2] (12) 1 H NMR (DMSO–d6, , ppm): 7.72 (d, 2H, J ¼ 3.5 Hz, furanyl C2–H), 7.52 (d, 2H, J ¼ 3.5 Hz, furanyl C3–H), 2.77 (q, 8H, J ¼ 6.84 Hz, C7,10–H), 1.28 (m, 8H, J ¼ 6.86 Hz, C8,11–H), 0.95 (t, 12H, J ¼ 6.84, 6.86 Hz, C9,12–H). 13C NMR (DMSO–d6, , ppm): 180.6 (C ¼ S), 172.95 (C ¼ O), 140.0 (C1), 135.35 (C4), 130.75 (C2), 127.95 (C3), 50.25 (C7, C10), 38.21 (C8, C11), 21.25 (C9, C12).

Pharmacology Antibacterial Studies The synthesized ligands, (L1)–(L3), and their respective metal(II) complexes (1)–(12) were tested against four Gram-negative (E. coli, S. sonnei, P. aeruginosa, S. typhi) and two Gram-positive (S. aureus, B. subtilis) bacterial strains by the disk diffusion method20,21. The test compounds (ligand/complex) were dissolved

Metal-based carboxamide-derived compounds endowed with antibacterial and antifungal activity

DOI: 10.3109/14756366.2013.815178

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Table 1. Physical measurements and analytical data of metal(II) complexes (1)–(12). Elemental analysis Calc (found) %


Compd. Structure

Yield (%)

MW/Formula

M.P( C)

C

H

N

M

[597.75] C24H34N4S4O2Co [1195.02] C48H68N8S8O4Ni2 [602.36] C24H34N4S4O2Cu [604.22] C24H34N4S4O2Zn [625.80] C26H38N4S4O2Co [1251.12] C52H76N8S8O4Ni2 [630.41] C26H38N4S4O2Cu [632.25] C26H38N4S4O2Zn [723.41] C24H32Br2N4S2O4Co [1446.34] C48H64Br4N8S4O8Ni2 [728.02] C24H32Br2N4S2O4Cu [729.85] C24H32Br2N4S2O4Zn

161–162

48.22 48.20 48.24 48.21 47.85 47.73 47.65 47.62 49.90 49.98 49.91 50.11 49.53 49.59 49.39 49.44 39.84 39.78 39.86 39.90 39.59 39.63 39.49 39.59

5.73 5.72 5.74 5.75 5.69 5.70 5.62 5.60 6.12 6.10 6.12 6.09 6.07 6.10 6.05 6.09 4.45 4.47 4.46 4.47 4.43 4.40 4.41 4.44

9.37 9.34 9.38 9.34 9.30 9.31 9.27 9.29 8.95 8.88 8.95 9.02 8.88 8.90 8.86 8.91 7.74 7.73 7.74 7.76 7.69 7.71 7.67 7.66

9.86 9.82 9.82 9.85 10.50 10.50 10.80 10.50 9.41 9.34 9.39 9.36 10.10 10.00 10.30 10.30 8.14 8.15 8.11 8.09 8.73 8.74 8.95 8.99

1

(1)

[Co(L -H)2]

80

(2)

[Ni(L1-H)2]2

79

(3)

[Cu(L1-H)2]

80

(4)

[Zn(L1-H)2]

77

(5)

[Co(L2-H)2]

78

(6)

[Ni(L2-H)2]2

79

(7)

[Cu(L2-H)2]

77

(8)

[Zn(L2-H)2]

75

(9)

[Co(L3-H)2]

78

(10)

[Ni(L3-H)2]2

81

(11)

[Cu(L3-H)2]

78

(12)

[Zn(L3-H)2]

76

164–165 157–158 168–170 149–150 159–160 153–154 163–164 139–140 149–150 147–149 136–137

Table 2. Conductivity, magnetic and spectral data of metal(II) complexes (1)–(12).

No.

M (1 cm2 mol1)

B.M meff

(1)

12.4

3.70

18 440

(2)

11.4

Dia

13 350, 19 100

(3)

14.7

1.80

15 255, 21 400

(4)

15.5

Dia

27 100

(5)

13.4

3.85

18 290

(6)

16.3

Dia

13 475, 18 890

(7)

11.9

1.88

15 370, 21 650

(8)

17.2

Dia

26 990

(9)

16.1

3.97

17 980

(10)

10.4

Dia

13 560, 19 210

(11)

11.9

1.86

15 460, 21 050

(12)

13.1

Dia

27 355

max (cm1)

(10 mg/mL) in DMSO. A known volume (10 mL) of the solution was applied with the help of a micropipette onto the sterilized filter paper discs. The discs were dried at room temperature overnight and stored in sterilized dry containers. Disks soaked with 10 mL of DMSO and dried in air at room temperature were

IR (cm1) 3075–3025 (CH), 1644 (C ¼ O), 1210 (C ¼ S), 890 (C–S), 535 (M–O), 460 (M–S) 3075–3025 (CH), 1645 (C ¼ O), 1207 (C ¼ S), 890 (C–S), 538 (M–O), 468 (M–S) 3075–3025 (CH), 1640 (C ¼ O), 1212 (C ¼ S), 890 (C–S), 542 (M–O), 467(M–S) 3075–3025 (CH), 1642 (C ¼ O), 1215 (C ¼ S), 890 (C–S), 545 (M–O), 464 (M–S) 3065–3030 (CH), 1635 (C ¼ O), 1215 (C ¼ S), 885 (C–S), 539 (M–O), 464 (M–S) 3065–3030 (CH), 1630 (C ¼ O), 1212 (C ¼ S), 885 (C–S), 548 (M–O), 462 (M–S) 3065–3030 (CH), 1636 (C ¼ O), 1215 (C ¼ S), 885 (C–S), 550 (M–O), 468 (M–S) 3065–3030 (CH), 1630 (C ¼ O), 1210 (C ¼ S), 885 (C–S), 545 (M–O), 467 (M–S) 3070–3030 (CH), 1640 (C ¼ O), 1215 (C ¼ S), 1150 (C–O), 680 (C–Br), 540 (M–O), 470 (M–S) 3070–3030 (CH), 1645 (C ¼ O), 1213 (C ¼ S), 1150 (C–O), 680 (C–Br), 549 (M–O), 466 (M–S) 3070–3030 (CH), 1635 (C ¼ O), 1215 (C ¼ S), 1150 (C–O), 680 (C–Br), 544 (M–O), 464 (M–S) 3070–3030 (CH), 1645 (C ¼ O), 1210 (C ¼ S), 1150 (C–O), 680 (C–Br), 546 (M–O), 465 (M–S)

used as the negative control. The standard antibiotic discs used as positive control were prepared as mention above in the laboratory by applying a known concentration of the standard antibiotic solution. Ampicillin was used as standard antibiotic. Bacterial culture was grown in nutrient broth medium at 37 C overnight

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M. Hanif et al.


and spread on to solidified nutrient agar medium in Petri plates using sterilized cotton swabs. Test and control disks were then applied to the medium surface with the help of sterilized forceps. The plates were incubated at 37 C for 24–48 h. The results were recorded by measuring the zone of inhibition in millimeter against each compound. The experiments were carried out in triplicate, and the values obtained were statistically analyzed.

The ligands showed different inhibitory effect on one or more bacterial and fungal strains which generally increased upon chelation/coordination with the metal(II) ions. IR spectra The characteristic IR spectral bands of ligands (L1)–(L3) and their metal(II) complexes (1)–(12) are reported in experimental section in Table 2. The IR spectra of all the synthesized ligands displayed24 strong absorption band at 3265–3275 cm1 due to N–H vibrations and medium absorption bands at 1670–1680 and 1240–1250 cm1, respectively, due to carbonyl (C ¼ O) and thiocarbonyl (C ¼ S) vibrations, strongly supporting the preparation of desired new compounds. The ligands (L1) and (L2) showed bands at 885–890 cm1 assigned to thienyl (C–S) group. The spectrum of ligand (L3) showed peak at 1153 cm1 assigned to furanyl (C–O) group. The same ligand, (L3) also showed another band at 685 cm1 giving a clue of the presence of C–Br. The IR spectra of all the metal(II) complexes showed significant changes on comparison to the spectra of the corresponding ligands. The most striking change observed was the presence of strong absorption band at 3265–3275 cm1 due to N–H vibrations in the spectra of free ligands which disappeared in spectra of the metal(II) complexes. The carbonyl (C ¼ O) and thiocarbonyl (C ¼ S) bands, originally appearing at 1670–1680 and 1240– 1250 cm1 in the spectra of the ligand, shifted to lower frequency by 25–40 cm1 at 1630–1545 and 1207–1215 cm1, respectively, in the spectra of their metal(II) complexes, indicating involvement of in coordination with the metal(II) ions. The decrease in frequency is suggested due to delocalization of electrons25. Coordination of carbonyl-O and thiocarbonyl-S is further justified by the appearance of new bands at 460–470 and 535–550 cm1 due to M–S and M–O linkages, respectively. This linkage is also supported by X-ray crystallographic structures of the Ni(II) and Cu(II) complexes (2) and (3), respectively.


In vitro antifungal activity Antifungal activities of all the compounds was studied against six fungal strains (T. longifusus, C. albicans, A. flavus, M. canis, F. solani and C. glabrata) according to literature protocol22,23. Sabouraud dextrose agar (Oxoid, Hampshire, England) was seeded with 105 (cfu) mL1 fungal spore suspensions and transferred to petri dishes. Disks soaked in 20 ml (200 mg/mL in DMSO) of test compounds were placed at different positions on the agar surface. The plates were incubated at 32 C for 7 d. The results were recorded as percentage of inhibition and compared with the standard drugs miconazole and amphotericin B.

Results and discussion The carboxamide derivatives (L1)–(L3) were synthesized by the reaction of potassium thiocyanate with thiophene-2-carbonyl chloride/5-methyl-thiophene-2-carbonyl chloride/5-bromo-furan2-carbonyl chloride in dry acetone followed by the condensation of the resulting product with dipropylamine (Scheme 1). These ligands were soluble in ethanol, ethyl acetate, DMF and DMSO, slightly soluble in tetrahydrofuran, diethyl ether and insoluble in aliphatic and aromatic hydrocarbons. The metal(II) complexes (1)–(12) were obtained by the stoichiometric reaction of the corresponding ligands with metals [Co(II), Ni(II), Cu(II) or Zn(II)] as acetate in a molar ratio M:L (1:2). All metal(II) complexes were air and moisture stable. They were soluble in mixture of ethanol and dichloromethane (1:1), DMF and DMSO. The structure of ligand, N-(dipropylcarbamothioyl)thiophene-2carboxamide (L1), and its nickel(II) (2) and copper(II) (3) complexes were determined from single crystal X-ray diffraction data. Physical measurements and analytical data of the metal(II) complexes (1)–(12) are given in Tables 1 and 2. These compounds have been investigated for in vitro antibacterial activity against four Gram-negative (E. coli, S. sonnei, P. aeruginosa, S. typhi) and two Gram-positive (S. aureus, B. subtilis) bacterial strains, and antifungal activity against six fungal strains (T. longifusus, C. albicans, A. flavus, M. canis, F. solani and C. glabrata).

1

H NMR spectra

1

H NMR spectral data of the ligands (L1)–(L3) and their diamagnetic Zn(II) complexes are recorded in the experimental part. The exhibited signals of all the protons due to heteroaromatic/aromatic groups were found26 in their expected region. The spectra of all the ligands (L1)–(L3) showed a broad signal at 11.50 ppm due to N–H proton. The ligand (L1) displayed C2–H and C4–H protons as a doublet and C3–H proton as double of the doublet at 8.0–8.18 and 7.35 ppm, respectively. The ligands (L2)

Scheme 1. Preparation of ligands (L1)–(L3) and their metal(II) complexes (1)–(12). O R

O

R

HN(CH2CH2CH3)2

KSCN N

Cl

C

M(CH3COO)2

1

4

L1: R =

S

2

3

2

3 H3C

N

M S

S 2 1

4 Br

N

2S 3

L 3: R =

O

O

1

4

R

R

2S L2: R =

9 CH3 1S 1O 8 7 2 5 1 6 N N R 10 11 12 H CH3 (L1)-(L3)

N

N

2O H3C

M = Co(II), Ni(II), Cu(II) and Zn(II)

CH3

H3C

(1)-(12)

CH3

DOI: 10.3109/14756366.2013.815178



and (L3) also showed C2–H and C3–H protons as a doublet at 7.62–7.75 and 7.15–7.45 ppm, respectively. However, the ligand (L2) displayed three protons of methyl (C4–CH3) as a singlet at 2.50 ppm. Furthermore, the spectra of all the ligands displayed protons of propyl group (C7,10–H, C8,11–H and C9,12–H) as triplet to sextet at 0.90–2.60 ppm. The high value of N–H proton is due to deshielding effect of carbonyl (C ¼ O) and thiocarbonyl (C ¼ S) groups. On comparison with the spectra of Zn(II) complexes, the N–H proton disappeared in the spectra of the Zn(II) complexes which is also supported by the IR spectra. All other protons underwent downfield shift by 0.05–0.15 ppm due to increased conjugation27 in the spectra of the Zn(II) complexes. 13

C NMR spectra

The 13C NMR spectral data are reported along with their possible assignments in the experimental section. The conclusion obtained from these studies provides further support to the mode of bonding explained in their IR and 1H NMR spectral data. The 13C NMR spectra of all the ligands showed peaks at 171.0–171.5 and 178.75–178.90 ppm for carbonyl-C and thiocarbonyl-C, respectively. All ligands displayed thiophene and furane carbons in the region at 130.18–139.73 and 127.78–139.73 ppm, respectively. The spectra of Zn(II) complexes showed downfield shifting of carbonyl–C and thiocarbonyl–C from 171.0–171.5 and 178.75– 178.90 ppm in free ligands to 172.4–172.95 and 180.0–180.6 ppm in the zinc(II) complexes, respectively, revealing the coordination of carbonyl-CO and thiocarbonyl-CS with the Zn(II) metal ion. Furthermore, all other carbons in the spectra of the Zn(II) complexes underwent downfield shifting by 0.15–0.4 ppm due to increased conjugation and coordination27. Molar conductivity and magnetic properties of the metal(II) complexes (1)–(12) The molar conductance values of all the metal complexes (1)–(12) in DMF (Table 2) fall in the range 11.4–15.5 1cm2mol1 showing their non-electrolytic nature28. The room temperature magnetic moment values of the Co(II), Ni(II), Cu(II) and Zn(II) complexes are given in Table 2. The values for Co(II) complexes are in the range of 3.70–3.97 B.M expected for three unpaired electrons favorably away from higher values usually in the range of 4.3–4.8 B.M and are in agreement with their tetrahedral geometry29. The Cu(II) and Ni(II) complexes showed meff values compatible for their square-planer geometry. The Zn(II) complexes exhibited diamagnetic nature. Figure 1. Molecular structure of ligand (L1).

5

Electronic spectra The electronic spectral values of Co(II), Ni(II), Cu(II) and Zn(II) complexes are recorded in Table 2. The electronic spectra of Co(II) complexes showed only one absorption band in the visible region at 17 980–18 440 cm1 assigned30 to the transition 4A2 (F)!4T1(F). This in turn suggests tetrahedral geometry for the Co(II) complexes. This is also supported by magnetic moment values (3.70–3.97 B.M) of Co(II) complexes. The Ni(II) complexes showed two bands at 13 350–13 560 cm1 and at 18 990–19 500 cm1 assigned to the transitions 1A1g ! 1B2g and 1 A1g ! 1A2g, respectively, consistent with their well-defined square-planar geometry. The spectra of Cu(II) complexes exhibited low-energy absorption bands at 15255–15460 cm1 assigned to the transitions, 2B1g ! 2E1g. The high-energy bands at 21 050–21 650 cm1 was assigned to the transition, 2B1g ! 2A1g. These transitions as well as the measured magnetic moment values (1.80–188 B.M) suggest31 a square-planar geometry for Cu(II) complexes. The diamagnetic Zn(II) complexes did not show any d–d transitions and their spectra were dominated only by the charge transfer band at 26 990–27 355 cm1, proposing a tetrahedral geometry for the Zn(II) complexes.

X-ray crystallographic studies Single crystal X-ray structure of ligand (L1) The ligand L1 is already reported19 as mentioned in the experiment, but its crystal structure has not been reported yet. The molecular structure of N-(dipropylcarbamothioyl)thiophene2-carboxamide (L1) (Figures 1 and 2) shows the expected bond lengths and bond angles32,33. The crystal data is given in Table 3. The C6–S1 and C5–O1 bonds show a double bond character with ˚ and 1.229(3) A ˚ , respectively. All of the bond lengths of 1.677(2) A ˚ and C10–N2 1.472(3) A ˚ represent CN bonds, C7–N2 1.467(3) A ˚ , C6–N1 1.421(3) A ˚ and C5– single bond and C6–N2 1.326(3) A ˚ N1 1.359(3) A indicate a partial double bond character (Tables 4 and 5). In the title compound, C12H18N2S2O, the carbonyl (C ¼ O) group is nearly coplanar with the thiophene ring [C2–C1–C5– O1 ¼ 176.2 (2)]. The propyl group is coplanar with S atom of thiocarbonyl (S ¼ O) group [C10–N2–C6–S1 ¼ 179.94 (16)] (Table 6). Molecular structure of nickel(II) (2) and copper(II) (3) complexes The crystal structures of compounds (2) and (3) were obtained by single crystal X-ray studies and are shown as Figures 3–5.

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M. Hanif et al.


˚ ) of ligand (L1). Table 4. Selected bond lengths (A ˚) Bond length (A

Atoms


S1–C6 S2–C1 N1–C5 N1–C6 N2–C6 C10–C11 C1–C2 C11–C12 C3–C4

1.677 1.727 1.359 1.421 1.326 1.522 1.367 1.515 1.340

(2) (2) (3) (3) (3) (3) (3) (3) (4)

Atoms

˚) Bond length (A

S2–C4 O1–C5 C8–C9 N1–H1N N2–C7 N2–C10 C1–C5 C2–C3 C7–C8

1.700 (3) 1.229 (3) 1.525 (3) 0.81 (2) 1.467 (3) 1.472 (3) 1.466 (3) 1.412 (3) 1.522 (3)

Table 5. Selected bond angles (deg) of ligand (L1).

Figure 2. The unit cell packing in ligand (L1). Table 3. Crystal data and structural refinement for ligand (L1). Empirical formula Formula weight Temperature Color Crystal system, space group A B C V Z, Calculated density Absorption coefficient (m) Crystal size F(000) Theta range for data collection () Parameters Refinement method max, min Rint Measured reflections Unique reflections R[F242(F2)] w wR(F2) (D/)max Dmax, Dmin CCDC number CIF file

C12H18N2OS2 270.40 100(2) K Colorless Orthorhombic, Fdd2 ˚ 37.384 (6) A ˚ 8.1949 (13) A ˚ 18.735 (3) A ˚3 5739.5 (15) A 16, 1.252 Mg m–3 0.36 mm–1 0.30 0.30 0.30 mm 2304 2.8–23.6 159 Refinement on F2 26.4 , 2.2 0.039 8028 2370 0.031 1/[2(Fo2) þ (0.0318P)2] where P ¼ (Fo2 þ 2Fc2)/3 0.065 0.001 ˚ 3, 0.17 e A ˚ 3 0.39 e A 884603 s3355m

Crystal refinement parameters are given in Table 7, and selected bond lengths and bond angles are given in Tables 8–12. The bond lengths and angles are all present in the expected34 range. The molecular structure of complex (2) shows that it exists as dimer form in asymmetric unit (Figure 3). Data collection and refinement of complex (2) are listed in Table 7 (supplementary material). The two carbonyl groups (C5 ¼ O1 and C17 ¼ O2) and (C29 ¼ O3 and C41 ¼ O4) adopt a cis-configuration around the nickel atoms Ni1 and Ni2, respectively (Table 8). The four

Atoms

Bond angle (degree)

Atoms

Bond angle (degree)

C7–C8–C9 C6–N2–C7 C7–N2–C10 C2–C1–S2 N2–C10–C11 O1–C5–N1 N1–C5–C1 N2–C6–S1 C5–N1–C6 C6–N2–C10 O1–C5–C1 N2–C7–C8 C10–N2–C6–S1 C10–N2–C6–S1 C5–N1–C6–S1 C6–N1–C5–O1 C6–N1–C5–C1

110.4 (2) 120.11 (16) 115.41 (17) 111.40 (17) 112.44 (18) 121.5 (2) 116.19 (19) 125.23 (16) 120.54 124.45 (18) 122.3 (2) 111.8 (2) –179.94 (16) –179.94 (16) 98.2 (2) 12.3 (3) –169.20 (18)

C5–N1–C6 C6–N2–C10 C2–C1–C5 C5–C1–S2 C12–C11–C10 O1–C5–C1 N2–C6–N1 N1–C6–S1 C6–N2–C7 N2–C10–C11 N1–C5–C1 C7–N2–C6–N1 C10–N2–C6–N1 C5–N1–C6–N2 C6–N2–C7–C8 C10–N2–C7–C8 N2–C7–C8–C9

120.54 (17) 124.45 (18) 131.5 (2) 117.06 (16) 112.9 (2) 122.3 (2) 116.25 (18) 118.52 (15) 120.11 (16) 112.44 (18) 116.19 (19) 178.83 (18) 0.5 (3) 82.5 (2) 86.4 (2) 92.1 (2) 176.64 (19)

˚ ) of ligand (L1). Table 6. Hydrogen-bond geometry (A D–H. . .A i

N1–H1N. . .S1

D–H

H...A

D...A

D–H...A

0.81(2)

2.62(3)

3.3762(19)

157(2)

Symmetry code: (i) x þ 1/2, y þ 1/2, z.

bidentate S,O-ligands (L1) adopt a meridional conformation with oxygen atoms in cisoidal coordinative geometry. The four sulfur hetero atoms are also in cisoidal coordinative geometry (Table 9). In this complex, oxygen(O) and sulfur(S) atoms from each ligands coordinated with the central metal atom Ni(II), in a cis-fashion, with slightly distorted square planar geometry33. In the structure, there are hydrogen bonds O2...S4B. . .S4 and N1. . .N2B. . .S1, these interactions are intermolecular interactions (Table 10). The labeling of atom and packing in unit cell are given in Figures 3 (supplementary material) and 4. The molecular structure of the copper(II) complex (3) is based on a monomer unit in which each copper atom is surrounded by four (two oxygen and two sulfur) atoms, two (one oxygen and one sulfur) atoms from each chelating ligands (Figure 5). In this complex, oxygen(O) and sulfur(S) atoms from each ligands coordinated with the central metal atom Cu(II) in a cis-fashion, with slightly distorted square planar geometry. All bond lengths are in the expected ranges33. In both chelate rings, the distances of N1–C6 (1.333(13) A ) and N3–C18 (1.330(14) A ) in the thiourea fragment are nearly same, but the distances of N1–C5 (1.314(14) A ) and N3–C17 (1.336 (14) A ) (Table 11) are slightly different, which support the slightly distorted square planar geometry of copper(II) complex

DOI: 10.3109/14756366.2013.815178


7


Figure 3. Molecular structure of Ni(II) complex (2).

Antibacterial bioassay

Figure 4. The unit cell packing in Ni(II) complex (2).

˚ ; O2– (3). The bond lengths of the carbonyl O1–C5 1.217(14) A ˚ and thiocarbonyl S1–C6 1.705(14) A ˚ ; S3–C18 C17 1.191(14) A ˚ groups lie between those for double and single 1.689(15) A bonds (Table 11). The same behavior is observed for C–N bond lengths, C–N bond lengths for the investigated complex is ˚, shorter than the average single C–N bond length of 1.48 A and greater than average double C ¼ N bond length ˚ , being C5–N1 ¼ 1.314(14) A ˚ , C6–N1 ¼ 1.333(13) A ˚, 1.16 A ˚, ˚, C18–N3 ¼ 1.330(14) A C17–N3 ¼ 1.336(14) A C18–N4 ¼ ˚ , C6–N2 ¼ 1.352(14) A ˚ , thus showing varying 1.340(15) A degrees of double and single bond character. The molecular structure data Cu(II) complex (3) with the atom-numbering scheme are given in Tables 7, 11 and 12.

Antibacterial activity of the synthesized ligands (L1)–(L3) and their metal(II) complexes (1)–(12) was determined against four Gram-negative (E. coli, S. sonnei, P. aeruginosa, S. typhi) and two Gram-positive (S. aureus, B. subtilis) bacterial strains and the obtained data are recorded in Table 13. The antibacterial activity was compared with the activity of standard drug (ampicillin) considering its activity as 100%. The synthesized compounds showed varying degree of inhibitory effects: low (up to 33%), moderate (up to 53%) and significant (above 53%). The obtained results indicated that the ligand (L1) exhibited moderate activity (42–52%) against all tested bacterial strains. The ligand (L2) possessed moderate activity (38–45%) against (a)–(d) and (f) and weak activity (33%) against strain (e). Similarly, the ligand (L3) displayed moderate activity (37–50%) against (c)–(f) and weak activity (30–33%) against (a) and (b) bacterial strains. However, the metal(II) complex (1) showed significant activity (62%) against (c) and (f) and moderate activity (43–50%) against (a), (b), (d) and (e) bacterial strains. Similarly, the complex (2) displayed significant activity (56–63%) against (c), (e) and (f) and moderate activity (42–50%) against (a), (b) and (d) bacterial strains. The complex (3) possessed significant activity (56–66%) against (b)–(d) and (f) and moderate activity (48–50%) against (a) and (e) bacterial strains. The complexes, (4), (7) and (8) also showed significant activity (56–65%) against (c) and (f) and moderate activity (38–52%) against (a), (b), (d) and (e) bacterial strains. In the same way, the metal(II) complexes, (6), (9) and (10) displayed significant activity (55–57%) against (c), (d) and (f) bacterial strains, respectively, and the complexes (11) and (12) also possessed significant activity (55–62%) against (d) and (f) bacterial strains. Moreover, the complexes (6), (9), (10), (11) and (12) showed moderate activity (38–52%) against other tested bacterial strains. However the complex (5) possessed moderate activity (46–52%) against (a) and (c)–(f) bacterial strains and weak activity (33%) against strain (b). Conclusively, the ligands (L1)–(L3) possessed smaller average activity value (11.8 mm, 42.59%) than the average activity value (14.0 mm, 50.54%) of the metal(II) complexes (1)–(12), which showed that the activity is enhanced34 upon coordination. In vitro antifungal bioassay The ligands (L1)–(L3) and their metal(II) complexes (1)–(12) were subjected to screening for their antifungal activity against, T. longifusus, C. albicans, A. flavus, M. canis, F. solani and

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Figure 5. Molecular structure of Cu(II) complex (3).

Table 7. Crystal data and details structure determinations of nickel(II) (2) and copper(II) (3) complexes.

˚ ) of nickel(II) complex (2). Table 8. Selected bond lengths (A Atoms

Data Empirical formula Formula weight Temperature Color Crystal system, space group a b c a b g V Z, Calculated density Absorption coefficient (m) Crystal size F(000) Theta range for data collection () Parameters max, min Rint Measured reflections Unique reflections Observed reflections I42(I) R[F242(F2)] wR(F2) (D/)max Dmax, Dmin CCDC number CIF file

(2)

(3)

C24H34N4NiO2S4 597.50 100 K Dark brown Triclinic, P-1

C24H34CuN4O2S4 602.33 100 K Green Monoclinic, C2/c

˚ 8.7115 (16) A ˚ 13.129 (2) A ˚ 25.114 (2) A 96.240 (4) 90.575 (4) 96.782 (4) ˚3 2834.6 (9) A 4, 1.400 Mg m3 1.01 mm1 0.40 0.20 0.1 mm 1256 2.5–17.8

˚ 30.86 (1) A ˚ 9.108 (3) A ˚ 24.803 (8) A

˚3 5632 (3) A 8, 1.421 Mg m3 1.10 mm1 0.60 0.10 0.05 mm 2520 2.4–13.2

934 25.0 , 1.9 0.100 14 728 9852 3882

239 23.3 , 1.6 0.267 12 107 4040 1027

0.060 0.116 50.001 ˚ 3, 0.55 e A ˚ 3 0.46 e A 774653 s3363 m

0.071 0.163 0.002 ˚ 3, 0.47 e A ˚ 3 0.46 e A 888415 s3365 m

126.115 (7)

C. glabrata fungal strains and results are recorded in Table 14. The obtained data were compared with the standard drugs miconazole and amphotericin B. The antifungal data showed that the ligand (L1) has moderate (40–50%) activity against (a), (b) and (d)–(f) and weak activity (32%) against (c) fungal strains. The ligand (L2) possessed significant activity (63%) against (d) and moderate activity (37–42%) against (b), (c) and (f) and weak

˚) Bond length (A

Ni1–O1 Ni1–O2 Ni1–S1 Ni1–S3 O1–C5 O2–C17 S1–C6 S3–C18

1.857 1.871 2.149 2.133 1.256 1.268 1.689 1.729

˚) Bond length (A

Atoms

(5) (5) (2) (2) (9) (9) (8) (8)

Ni2–O4 Ni2–O3 Ni2–S7 Ni2–S5 O3–C29 O4–C41 S5–C30 S7–C42

1.863 1.871 2.140 2.143 1.259 1.269 1.725 1.719

(4) (4) (2) (18) (7) (7) (7) (7)

Table 9. Selected bond angles (deg) of nickel(II) complex (2). Atoms

Bond angle (degree)

Atoms

Bond angle (degree)

84.6 (2) 95.1 (2) 95.1 (1) 85.50 (8) 131.2 (4) 132.6 (4) 108.5 (3) 109.5 (2) 4 (1) 8 (1) 2.4 (7)

O4 Ni2 O3 O4 Ni2 S7 O3 Ni2 S7 C29O3Ni2 C30N6C34 C30N6C31 C41N7C42 C2 C3 C4 O1 Ni1 S1 C6 O1 C5 C1 S2 O2 C17 C13 S4

85.30 (19) 95.07 (15) 177.23 (15) 130.7 (4) 122.8 (6) 120.7 (5) 122.6 (6) 112.2 (6) 10.1 (3) –174.9 (5) –176.5 (6)

O1 Ni1 O2 O1 Ni1 S1 O2 Ni1 S3 S1 Ni1 S3 Ni1 O1 C5 Ni1 O2 C17 Ni1 S1 C6 Ni1 S3 C18 Ni1 O1 C5 N1 Ni1 O2 C17 N3 Ni1 S3 C18 N3

˚ ) of nickel(II) Table 10. Van der Waals-virtual bond parameters (A complex (2). A. . .B. . .C N1. . ..N2B. . .S1 O2–S4B. . .C14B

A. . .B

B. . .C

A...C

A. . .B...C

2.413(5) 2.921(5)

2.581(5) 2.486(5)

2.755(5) 3.626(5)

66.85(3) 83.87(4)

activity (19–30%) against (a) and (e) fungal strains. Similarly, the ligand (L3) displayed moderate activity (39–52%) against (a) and (c)–(f) and weak activity (25%) against (b) fungal strains. The compounds (1) and (3) possessed significant activity (54–56%) against (b) and moderate activity (35–50%) against (a) and (c)–(f) fungal strains. The activity of compounds (2) and (4) was found to

DOI: 10.3109/14756366.2013.815178


˚ ) of copper(II) complex (3). Table 11. Selected bond lengths (A Atoms

˚) Bond length (A

S1–C6 S3–C18 N1–C5 N1–C6 Cu1–S1 Cu1–S3

1.705 1.689 1.314 1.333 2.221 2.196

Table 14. Antifungal bioassay (concentration used 200 mg/mL) of ligands (L1)–(L3) and metal(II) complexes (1)–(12).

˚) Bond length (A

Atoms

(14) (15) (14) (13) (4) (4)

O1–C5 O2–C17 N4–C18 N2–C6 Cu1–O1 Cu1–O2

1.217 1.191 1.340 1.352 1.916 1.929

(14) (14) (15) (14) (8) (8)

Table 12. Selected bond angles (deg) of copper(II) complex (3). Journal of Enzyme Inhibition and Medicinal Chemistry Downloaded from informahealthcare.com by East Carolina University on 08/10/13 For personal use only.

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Atoms

Bond angle (degree)

Atoms

O1–Cu1–O2 O1–Cu1–S3 O2–Cu1–S3 O1–Cu1–S1 O2–Cu1–S1 S3–Cu1–S1 C6– S1–Cu1 S1–Cu1–S3–C18 O2–Cu1–O1–C5 Cu1–O1–C5–N1 Cu1–S1–C6–N1

85.7 (3) 164.6 (3) 92.7 (3) 92.0 (3) 169.4 (3) 92.15 (15) 108.4 (5) 151.1 (5) 162.5 (14) 9 (2) 3.2 (14)

C18–S3–Cu1 C5–O1–Cu1 C17–O2–Cu1 C14–C13–S4 C14–C13–C17 C5–N1–C6 C6–N2–C7 O2–Cu1–S1–C6 S3–Cu1–S1–C6 C6–N1–C5–C1 Cu1–S1–C6–N2

Bond angle (degree) 106.4 135.9 129.9 115.6 127.1 124.9 120.7 –76.4 166.2 178.8 –178.6

(5) (9) (9) (10) (12) (13) (11) (16) (5) (11) (9)

Table. 13 Antibacterial bioassay (concentration used 1 mg/mL of DMSO) of ligands (L1)–(L3) and metal(II) complexes (1)–(12).

[% Inhibition] Compounds 1

(L ) (L2) (L3) (1) (2) (3) (4) (5) (6) (7) (8) (9) (10) (11) (12) SD

(a)

(b)

(c)

(d)

(e)

(f)

SA

Average

39 19 39 38 42 44 41 19 23 20 25 40 37 39 44 A

50 39 25 54 52 56 50 43 43 38 37 29 25 20 27 B

32 37 52 35 38 36 35 38 36 43 41 55 53 58 53 C

43 63 43 46 44 46 48 65 66 68 62 44 48 47 44 D

34 30 44 30 35 39 33 34 32 35 37 47 46 46 48 E

47 42 47 48 52 50 49 50 46 45 44 50 49 51 49 F

7.14 14.6 9.24 8.99 7.05 7.28 7.45 15.5 14.7 15.7 12.1 9.86 10.3 13.1 9.06 –

40.8 38.3 41.7 41.8 43.8 45.1 42.7 41.5 41.0 41.5 41.0 44.1 43.0 43.5 44.1 –

Activity of ligands (L1)–(L3) ¼ 40.2%; Average activity of complexes (1)–(12) ¼ 42.7%; (a) ¼ T. longifusus (b)¼C. albicans (c) ¼ A. flavus (d) ¼ M. canis (e) ¼ F. Solani (f) ¼ C. glabrata, SD ¼ Standard Drugs MIC mg/mL; A ¼ Miconazole (70 mg/ mL:1.6822 107 M/mL), B ¼ Miconazole (110.8 mg/mL:2.6626 10– 7 M/mL), C ¼ Amphotericin B (20 mg/mL:2.1642 108 M/mL), D ¼ Miconazole (98.4 mg/mL:2.3647 107 M/mL), E ¼ Miconazole (73.25 mg/mL: 1.7603 107 M/mL), F ¼ Miconazole (110.8 mg/mL: 2.66266 107 M/mL). SA ¼ Statistical Analysis.

[Zone of Inhibition (mm)] Gram negative

Conclusion

Gram positive

Compounds

(a)

(b)

(c)

(d)

(e)

(f)

(SA)

Average

(L1) (L2) (L3) (1) (2) (3) (4) (5) (6) (7) (8) (9) (10) (11) (12) SD

11 10 08 13 11 13 13 12 13 10 10 12 11 13 10 26

12 10 08 11 12 16 12 08 11 12 12 10 11 12 10 24

15 13 12 20 18 18 19 15 18 19 18 14 15 13 15 32

13 11 14 12 14 17 14 14 13 12 13 16 13 17 16 28

13 09 11 13 17 13 14 13 13 12 12 14 12 13 13 27

15 13 1 14 2 18 17 18 18 15 14 19 17 13 16 16 18 29

1.60 67 50 3.61 2.32 2.10 2.45 2.28 2.06 3.74 2.66 2.04 2.10 2.00 3.14 2.73

13.1 11.0 11.3 14.5 14.8 16.0 15.0 13.0 13.3 14.0 13.3 13.1 13.0 14.0 13.7 27.7

Activity of Ligand (L1)–(L3) ¼ 11.8 mm; average activity of Complexes (1)–(12) ¼ 14.0 mm; (a)¼E. coli (b)¼S. sonnei (c)¼P. aeruginosa (d)¼S. typhi (e)¼S. aureus (f)¼B. subtilis, Activity 510 ¼ weak; 410 ¼ moderate; 416 ¼ significant: SD ¼ Standard Drug (Ampicillin). SA ¼ Statistical Analysis.

be moderate (35–52%) against (a)–(d) and (f) and weak (30–33%) against (e) fungal strains. The compounds, (5)–(8) showed significant activity (62–68%) against (d), moderate activity (34– 50%) against (b), (c), (e), (f) and weak activity (19–25%) against (a). Similarly, the compounds (9)–(12) possessed significant activity (54–58%) against (c), moderate activity (37–51%) against (a), (d)–(f) and weak activity (20–29%) against (b) fungal strains. The average activity data comparison showed that the metal(II) complexes exhibited greater average activity value (42.7%) than the average activity value of the ligands (40.2%). These conclusions indicated that antifungal activity of the ligands increased upon chelation/coordination with the metal(II) ions.

The ligands (L1)–(L3) act as bidentate and coordinate through carbonyl-O and thiocarbonyl-S to the metal(II) ion. The bonding of ligands to the metal(II) ion is supported by their analytical, IR, 1 H NMR, 13C NMR, electronic and magnetic studies which have showed the Ni(II) and Cu(II) complexes to observe a square planar and Co(II) and Zn(II) complexes a tetrahedral geometry. These observations are further supported by the X-ray crystallographic data of Ni(II) and Cu(II) complexes. The X-ray crystallographic data of Ni(II) complex showed that the complex (2) exist in dimer form as asymmetric unit. In vitro antibacterial and antifungal studies of the ligands and their metal complexes against representative bacterial and fungal strains revealed that the ligands, (L1)–(L3) and their Co(II), Ni(II), Cu(II) and Zn(II) complexes were found to possess moderate-to-significant activity. This activity is enhanced upon chelation, and the metal complexes thus become more potent than the parent ligands against one or more bacterial and fungal species.

Acknowledgements We are thankful to HEJ research Institute of Chemistry, University of Karachi, Pakistan, for providing their help in taking NMR, mass spectral and antibacterial/antifungal data.

Declaration of interest This work was supported by Collaboration between University of Manchester, U.K and Bahauddin Zakariya University, Multan (Pakistan) on Indigenous Fellowship program sponsored by the Higher Education Commission, Government of Pakistan to one of us (MH). The authors report no conflicts of interest. The authors alone are responsible for the content and writing of this article.

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