Crystal Growth, Spectral, Magnetic, Antibacterial and Antifungal Studies of Co(II) and Ni(II) Complexes of 4-Nitrobenzoic Acid

Abstract

Single crystals of cobalt(II) and nickel(II) complexes of 4-nitrobenzoic acid have been successfully grown by gel diffusion technique. The grown crystals were characterized by elemental analysis, FT-IR and UV-Visible spectroscopy. The structure of the grown crystals was elucidated using single crystal X-ray diffraction studies. The title compounds $[Co(Pnba)_2(H_2O)_4]{\cdot}2H_2O$ 1 and $[Ni(Pnba)_2(H_2O)_4]{\cdot}2H_2O$ 2 where PnbaH=4-nitrobenzoic acid, crystallize in centrosymmetric triclinic space group P-1. Magnetic susceptibility measurements reveal that the compounds are paramagnetic in nature. The mechanical strength of the grown crystals was determined by Vicker's microhardness studies. The ligand (4-nitrobenzoic acid) and the complexes have been screened for their biological activity against various bacteria and fungi. The activity data show that the biological activity of the complexes is higher than that of the ligand.

INTRODUCTION

Metal carboxylates play an important role in inorganic and bioinorganic chemistry. The fascinating structural features of metal carboxylates can be attributed to the versatile ligational behaviour of the carboxylate group which can function as a monodentate or bidentate or bridging type of ligand.1−3 The study of coordination complexes have gained importance during the recent years especially due to the slow release or long acting drugs in nutrition and in the study of metabolism.4 Transition metal aromatic carboxylates have shown promising results when screened against several pathogenic bacteria and fungi.5−7 Therefore the transition metal complexes were studied for their antibacterial and antifungal activity. Moreover cobalt and copper ions have been used to inhibit the growth of harmful microbes for centuries. 4-Nitrobenzoic acid is used as a raw material for the synthesis of benzocaine, folic acid and benzoyl glutamic acid. Eventhough the crystal structure of the title compounds have been reported earlier,8−10 we have grown the crystals by a different method. The compounds crystallize in a triclinic space group P-1. The grown crystals are characterized by elemental analysis, FT-IR spectroscopy and UV-Visible spectroscopy. Powder X-ray diffraction study was conducted to confirm the crystalline nature of the compound. Magnetic susceptibility measurement of the complexes was also carried out. The mechanical strength of the compounds was determined by Vicker’s microhardness studies. The antibacterial and antifungal activity of the compounds was studied using agar-well diffusion method.

 

EXPERIMENTAL PROCEDURE

Growth Procedure

All commercial reagents were of analytical grade and used without further purification. Single crystals of the title compounds were grown by single gel diffusion technique. Sodium metasilicate (Na2SiO3·9H2O) was used to prepare the gel. The apparatus for crystallization consists of borosilicate glass tubes of length 20 cm and diameter 2.5 cm. The gel was prepared by mixing 4-nitrobenzoic acid in sodium metasilicate solution of varying density (1.03−1.05 gcm−3) with continuous stirring to avoid local ion concentration which may cause premature local gelling and make the final solution inhomogeneous.11 The pH of the gel was adjusted using glacial acetic acid. The gel solutions of different pH ranging from 5.5 to 7 were transferred to several glass tubes. These glass tubes were well sealed to eliminate surface contamination. Sufficient time was given to the gel to set. Once gelled, an aqueous solution of metal solution was poured along the walls of the tubes without tampering the meniscus of the gel, so that the ligand to metal stochiometry is 2:1. The open end of the test tubes was again covered to avoid evaporation of the solution and contamination.

Characterization

The carbon, nitrogen and hydrogen contents in the obtained crystals were determined using Elementar Vario-EL 111 CHNS analyzer. Single crystal XRD analysis of the crystals was carried out using a Bruker AXS Kappa Apex2 CCD diffractometer, with graphite monochromated Mo Kα (λ=0.71073Å) radiation. The powder X-ray diffraction experiment was carried out in a Bruker AXS D8 advance X-ray powder diffractometer using Cu Kα radiation (λ=1.54056Å). The FT-IR spectra were recorded in the range 4000−400 cm−1 using Thermo Nicolet, Avatar 370 spectrometer in KBr pellets. UV-Visible absorption spectrum was recorded using Varian Cary 5000 UV-Vis-NIR spectrometer in the range 200−1200 nm. Magnetic measurements were carried out on a Sherwood Scientific MK1 magnetic balance using Gouy method. Vicker’s microhardness measurements were taken using Shimadzu HMV-2TAW microhardness tester, fitted with a diamond indenter.

Antimicrobial Activity

The ligand and the title compounds were screened against three Gram positive (Staphylococcus aureus, Bacillus subtilis and Klebsiella pneumonia) and two Gram negative (Escherichia coli and Streptococcus mutans) bacteria using agar-well diffusion method. The activity of the compounds was found out by measuring the diameter of the zone of inhibition in millimeters. In vitro antibacterial activity of the ligands and compounds has been evaluated and compared with the standard antibiotic, Gentamycin. The Muller Hinton agar and the nutrient broth were prepared and poured in the petriplates and allowed to cool. These petriplates were seeded with the tested bacterial strains such as Staphylococcus aureus, Bacillus subtilis, Streptococcus mutans, Klebsiella pneumoniae and Escherichia coli. After seeding, wells of approximately 10 mm was bored using a well cutter and different concentrations (25, 50,100 μg/ml) of the investigated compounds were added. The plates were incubated at 37 °C for 24 hours. The diameter of the zone of inhibition was measured in millimeters after overnight incubation.

Antifungal Activity

The antifungal activity of the ligand and the complexes has been investigated by agar-well diffusion method. Aspergillus niger and Candida albicans were employed for testing the antifungal activity of both ligand and the complexes. Muller Hinton agar was prepared and overnight grown fungi were swabbed. Wells of approximately 10 mm was bored using a well cutter and samples of different concentrations (50, 100 μg/ml) was added. After the completion of overnight incubation, the diameter of the zone of inhibition was measured in millimeters and compared with the standard antibiotic, Clotrimazole.

 

RESULTS AND DISCUSSION

Crystal Growth

Block shaped transparent crystals of the title compounds were grown by single gel diffusion technique as described above. Crystals suitable for single crystal X-ray diffraction appeared at the gel interface within two weeks, under the conditions of gel density 1.05 gcm−3 and pH 6.5. This was the optimum condition for growth of the crystals. The photographs of the as grown crystals are shown in Figs. 1a and 1b.

Elemental Analysis

The chemical composition of the compounds was determined by elemental analysis. In compound 1, the experimental and calculated percentages of carbon, nitrogen and hydrogen are %C=34.77 (calc.=34.79), %N=3.89 (calc.=3.72) and %H=5.65 (calc. 5.79) and in compound 2, they are %C=33.65 (calc.=33.55), %H=4.72 (calc.=3.99) and %N=5.64 (calc.=5.59). The experimental and calculated values are in good agreement with each other within the acceptable limits. The calculated values agree with the formulae [M(Pnba)2(H2O)4]·2H2O where M=Co(II) and Ni(II).

Powder X-ray Diffraction Studies

To confirm the crystalline nature of the title compounds, powder X-ray diffraction studies were carried out and the crystalline nature of the grown crystals has been revealed from the well defined Bragg’s peaks at the specific 2θ angles. The experimental powder X-ray diffraction patterns was compared with the X-ray diffraction patterns obtained from the single crystal XRD data using Mercury software in order to check the phase purity of the compounds. The peaks observed at specific 2θ angles in the experimental powder X-ray diffractograms are in good agreement with the simulated XRD patterns obtained from single crystal XRD data which indicates that the compounds possess bulk phase purity.12 The simulated and experimental powder X-ray diffraction patterns of compounds 1 and 2 are shown in Figs. 2a and 2b.

Figure 1.(a) Photograph of the grown crystal of compound 1. (b) Photograph of the grown crystal of compound 2.

Figure 2.(a) Powder X-ray diffraction patterns (experimental and simulated) of 1. (b) Powder X-ray diffraction patterns (experimental and simulated) of 2.

Table 1.Crystal Growth, Spectral, Magnetic, Antibacterial and Antifungal Studies of Co(II) and Ni(II) Complexes of 4-Nitrobenzoic Acid

X-ray Crystallography

The single crystal X-ray diffraction data of the compounds were collected using a Bruker AXS Kappa Apex2 CCD diffractometer, with graphite monochromated Mo Kα (λ=0.71073Å) radiation. The unit cell dimensions and intensity data were recorded at 296 K. The structures was solved with direct methods using SIR9213 and refinement was carried out by full-matrix least squares on F2 using SHELXL-97.14 All non-hydrogen atoms were refined anisotropically. The hydrogen atoms were located from the difference Fourier maps and refined isotropically. Molecular graphics tool used was DIAMOND Version 3.1f15 and MERCURY.16 The technical details of data collection and some selected refinement parameters of compounds 1 and 2 are given in Table 1. Selected bond lengths and bond angles of compounds 1 and 2 are summarized in Tables 2 and 3 respectively.

Table 2.Symmetry transformations used to generate equivalent atoms: #1−x+1, −y+1, −z+2

Table 3.Symmetry transformations used to generate equivalent atoms: #1–x+1, –y+2, –z+2

Crystal structures of [Co(Pnba)2(H2O)4]·2H2O (1) and [Ni(Pnba)2(H2O)4]·2H2O (2)

To the best of our knowledge, this is first report of the crystal growth of the compounds 1 and 2 by single gel diffusion technique. The compounds 1 and 2 crystallize in a triclinic space group P-1 and are having a centrosymmetric crystal structure. In compound 1, the monomeric molecule consists of a Co(II) ion at the center which is six coordinated. The carboxylate group (O3) of each PnbaH ligand binds the Co(II) ion in a monodendate manner satisfying two coordination position. The other four coordination positions are satisfied by oxygen atoms (O2w & O3w) of each of the four water molecules. The Co–O bond distances are found to be Co1–O3 (2.1296(11) Å), Co1–O2w (2.0803(13) Å) and Co1–O3w (2.0891(14) Å). The lattice site also contains two water molecules. The molecular structure with atom numbering scheme is shown in Fig. 3. In the molecule, the coordinated as well as the lattice water molecules are involved in hydrogen bonding linking the complex into a chain. The oxygen atoms, O(1) and O(2) connected to the nitrogen atom are hydrogen bonded to the lattice and coordinated water molecules of the adjacent molecule. The coordinated water is also involved in intramolecular hydrogen bonding. The intermolecular and intramolecular hydrogen bonding contributes to the stability of the compound thereby forming a supramolecular network. The hydrogen bonding interactions are shown in Table 4. The supramolecular arrangement of the molecule is shown in Fig. 4.

Figure 3.Molecular structure of 1 with atom numbering scheme. Hydrogen atoms of water molecules are omitted for clarity.

Table 4.Symmetry transformations used to generate equivalent atoms: #1 −x+1, −y+2, −z+2 #2 x+1, y, z #3 −x+2, −y+1, −z+2 #4 –x+1, −y, −z+2 #5 x+1, y, z

The compound 2 comprises of a Ni(II) ion at the center coordinated to six oxygen atoms, two oxygen atoms each from the carboxylic group of PnbaH ligand and four oxygen atoms (O2w & O3w) each from four water molecules and the bond lengths are Ni1–O2w (2.055(3) Å) and Ni1–O3w (2.064(3) Å). The carboxylic oxygen atoms (O4) coordinate to the Ni(II) ion in a monodentate fashion and the Ni1–O4 bond length is (2.078(3) Å). Two water molecules are also present in the lattice site of the compound. The asymmetric unit with atom numbering scheme is shown in Fig. 5.

Figure 4.Supramolecular arrangement of the 1 viewed along a-axis.

Figure 5.Molecular structure of 2 with atom numbering scheme. Hydrogen atoms of water molecule are omitted for clarity.

The compound 2 undergoes extensive hydrogen bonding due to the presence of coordinated and lattice water molecules. The hydrogen bonding interactions are shown in Table 5. The compound forms both intermolecular and intramolecular hydrogen bonds thereby generating a supramolecular network. The supramolecular network with well defined channels can be qualitatively viewed along a-axis (Fig. 6).

FT-IR Spectroscopy

FT-IR spectroscopic method is used to determine the molecular structure and to identify the functional groups in the grown crystals. The IR spectra of the compounds exhibit several sharp bands in the mid-infrared region, many of which are due to the vibrations of organic moiety.17,18 The free acids do not exhibit any significant signals above 3100 cm−1 in the FT-IR spectra indicating that the –OH group of the carboxylic acid is strongly H-bonded to the nitro substituent in the solid state.18 The vibrational bands observed between 3200 and 3600 cm−1 in the FT-IR spectra of 1 and 2 indicate the presence of water in it. The carbonyl absorption, observed at 1692 cm−1 in the FT-IR spectrum of PnbaH is absent in the complexes indicating that the ligand coordinates to the metal ion through the carboxylate group after deprotonation.19,20 The asymmetric and symmetric stretching vibrations of the carboxylate groups observed at 1514 and 1378 cm−1 in 1 and 1518 and 1347 cm−1 in 2 support this observation. The asymmetric and symmetric stretching vibrations of the nitro group in the aromatic ring observed at 1543 and 1350 cm−1 in FT-IR spectrum of the ligand does not show appreciable change in the complexes indicating that the nitro group is not involved in bonding. The FT-IR spectra of 1 and 2 are shown in Figs. S1 and S2 in the supplementary information.

Table 5.Symmetry transformations used to generate equivalent atoms: #1 −x+1, −y+2, −z+2 #2 x+1, y, z #3 x+1, y, z+1 #4 x, y+1, z #5 −x, −y+2, −z+1

Figure 6.Supramolecular arrangement of the 2 viewed along a-axis.

UV-Vis Spectral Studies

The absorbance spectra of the compounds were recorded in solid state, due to their low solubility in common solvents. The absorption band observed between 250 and 300 nm, in the compounds 1 and 2 can be attributed to intraligand transitions of the PnbaH ligand.21 A broad band observed at 503 nm in compound 1 can be attributed to 4T1g(F)→4A2g transitions which suggests the octahedral geometry of the Co(II) complexes.22,23 The absorbance spectrum of complex 1 and 2 is shown in Figs. S3 and S4 (supplementary information). The absorption spectrum of compound 2 shows a broad band at 707 nm corresponding to the 3A2g (F)→3T1g (F) indicating that the compound possess a six coordinate octahedral geometry.24

Micro Hardness Studies

Hardness can be defined as a measure of the resistance to deformation. Vicker’s micro hardness measurements were done on the grown crystals using a Shimadzu HMV-2TAW micro hardness tester fitted with a diamond pyramidal indenter. The diamond pyramidal indenter was used with a constant indentation time of 14 seconds. The load applied was 25 g. Hardness value is calculated from the relation:

Hv = 1.8544 P/d2 Kg/mm2

where P is the applied load in Kg and d is the average diagonal length of the indented impression in millimeter. The micro hardness value was taken as the average of several indentations made. The value of hardness was found to be 39.49 Kg/mm2 for compound 1 and 42.16 Kg/mm2 for compound 2. The hardness may be attributed to the stress required for the homogeneous nucleation in the small dislocation free region indented.25

Magnetic Susceptibility

Magnetic susceptibility of the powdered compounds was determined using a Sherwood scientific MK1 balance at room temperature. The magnetic susceptibility measurements showed that the compound was paramagnetic in nature. After applying suitable diamagnetic corrections, the effective magnetic moment of the compound, μeff was calculated using the relation:

μeff = 2.828(χmcorr T)1/2

where χmcorr is the molar magnetic susceptibility corrected using Pascal’s constants for diamagnetism of all atoms in the compound and T is the absolute temperature. The effective magnetic moment of the compound 1 was found to be 4.61 BM which suggests a high spin octahedral geometry for the complex.26,27 Compound 2 has an effective magnetic moment of 2.77 BM indicating that the compound possesses an octahedral geometry.28

Antibacterial Studies

The antibacterial activity of the ligand, PnbaH and the complexes were determined by agar-well diffusion method as described above. Gentamycin was used as the standard. The growth inhibition of Gentamycin for Staphylococcus aureus, Bacillus subtilis, Streptococcus mutans, Klebsiella pneumoniae and Escherichia coli was found to be 22, 35, 35, 30 and 38 mm respectively. The diameter of the zone of inhibition formed around the wells by the ligand and the complexes was calculated for three different concentrations and the photograph showing the antibacterial activity of the compounds 1 and 2 are shown in Fig. 7 and 8 respectively. It was found that the complexes are more active compared to the free ligand, but less active when compared with the standard Gentamycin in the case of all bacterias except Staphylococcus aureus. The diameter of zone of inhibition in millimeters is shown in Table 6. From the table, it was found that the compound 1 is more active compared to the ligand and the compound 2. The inhibition zones of the ligand as well as the complexes were found to increase with increase in concentration. The increased antimicrobial activity of the metal complexes over ligands is favored by five factors namely a) the chelate effect, b) the nature of coordinated ligands, c) the total charge of the complex, d) the nature of the ion neutralizing the ionic complex and d) the nuclearity of the metal center in the complex. 29,30 On chelation the polarity of the metal ion will be reduced to a greater extent due to overlap of ligand orbital and partial sharing of the positive charge of the metal ion with donor groups. The delocalization of the π-electrons over the whole chelate ring also increases which enhances the lipophilicity of the complexes.31 The increased lipophilicity enhances the penetration of the complexes into lipid membranes and blocking of the metal binding sites in the enzymes of microorganisms. The respiration process of the cell is also disturbed by the complexes thereby blocking the synthesis of the proteins that restricts further growth of the organism.32

Figure 7.Antibacterial activity of 1 against Streptococcus mutans, Klebsiellla pneumoniae, E. coli, Bacillus subtilis, Staphylococcus aureus.

Figure 8.Antibacterial activity of 2 against Klebsiellla pneumoniae, E. coli, Streptococcus mutans, Bacillus subtilis, Staphylococcus aureus.

Antifungal Activity

The antifungal activity of the ligand and the complexes are determined using agar-well diffusion method as described above. The standard used for the antifungal study was Clotrimazole. The growth inhibition of Clotrimazole for Candida albicans and Aspergillus niger was found to be 17 and 20 mm. The ligand and the complexes were found to be more active compared to the standard. The activity of the investigated samples was found to increase with the concentrations of the samples. The diameter of zone of inhibition in millimeters for the ligand and the complexes is shown in Table 7 and the photograph of antifungal activity of the compounds 1 and 2 are shown in Figs. 9 and 10 respectively.

Table 6.Antibacterial activity of the ligand (PnbaH) and the complexes 1 and 2 at different concentrations (25, 50, 100 μg/ml)

Figure 9.Antifungal activity of 1 towards Aspergillus niger and Candida albicans.

Figure 10.Antifungal activity of 2 towards Aspergillus niger and Candida albicans.

Table 7.(−): no activity observed against the fungal species

 

CONCLUSION

X-ray quality single crystals of Co(II) and Ni(II) complexes of 4-nitrobenzoic acid were successfully grown by gel diffusion technique. The composition of the complexes was obtained from elemental analysis and was found to be [Co(Pnba)2(H2O)4]·2H2O and [Ni(Pnba)2(H2O)4]·2H2O. The presence of water in the compounds 1 and 2 was confirmed from FT-IR spectral studies. Powder X-ray diffraction studies confirmed the crystalline nature of the complexes and on comparison with the simulated X-ray diffraction patterns from single crystal X-ray diffraction data, the bulk purity of the crystals were confirmed. Single crystal X-ray diffraction studies showed that the monomeric compounds 1 and 2 crystallized in centrosymmetric triclinic space group P-1. The UV-visible spectral data and the magnetic moment values confirmed the octahedral geometry of the complexes. The mechanical strength of the grown crystals was confirmed by Vicker’s microhardness studies. The complexes show good antifungal and antibacterial activity compared to the free ligand.