Synthesis Characterization and Antimicrobial Activity Studies of Some Transition Metal Complexes Derived from 3-Chloro-N′-[(1E)-(2-hydroxy phenyl)methylene]-6-methoxy-1-benzothiophene-2-carbohydrazide

A series of new coordination complexes of Cu(II), Co(II), Ni(II), Zn(II), Hg(II), Mn(II), and Fe(III) with the Schiff base 3-chloro-N′-[(1E)-(2-hydroxy phenyl)methylene]-6-methoxy-1-benzothiophene-2-carbohydrazide (HL) have been synthesized and characterized by elemental analysis, electrical conductivity measurements, IR spectra, 1H NMR, mass spectral data, electronic spectra, magnetic susceptibility, ESR spectra, TGA, and Powder XRD data. The Schiff base behaves as tridentate ONO donor ligand and forms the complexes of the type ML2 (metal-ligand) stoichiometry for Cu(II), Co(II), Ni(II), and Mn(II) complexes and ML stoichiometry for Zn(II), Hg(II), and Fe(III) complexes. All the complexes are colored and nonelectrolytes. It is found that Cu(II), Co(II), Ni(II), Mn(II) and Fe(III) complexes have exhibited octahedral geometry whereas Zn(II) and Hg(II) complexes exhibited tetrahedral geometry. The ligand and its metal complexes have been screened for their antibacterial activity against E. coli and S. aureus and antifungal activity against A. niger and A. flavus.


Introduction
Metal complexes with potentially tridentate and tetradentate ligands have evoked much interest in coordination chemistry [1]. Schiff base complexes of transition metals have played prominent role in the development of coordination chemistry [2]. Several Schiff base metal complexes have been studied because of their industrial and biological applications [3][4][5]. Schiff bases containing polyfunctional groups offer many practical advantages and unique structural environment for complexation [6]. Chen et al. [7] have reported Cu(II) complexes of thiophene-2,5-dicarboxylic acid with a view of constructing diverse low dimensional coordination polymers and the model coordination compounds. Literature survey reveals that many benzothiophene [8][9][10] derivatives are known to possess good biological activities like antimicrobial, anti-inflammatory, analgesic, diuretic, and antiviral activities.

Material and Method.
All the chemicals are of reagent grade. Solvents were dried and distilled before use according to standard procedure [17]. The precursor 3-chloro-6methoxy benzothiophene-2-carbohydrazide was prepared by literature method [18]. The absorption of metal and chloride containt of the complexes were carried out by the standard procedure. [19]. The metal chlorides used were in their hydrated form.

Synthesis of the Ligand HL.
An equimolar mixture of 3-chloro-6-methoxy-benzothiophene-2-carbohydrazide (0.001 mol) and salicylaldehyde (0.001 mol) in ethanol (30 mL) was refluxed in presence of catalytic amount of glacial acetic acid (1-2 drops) for about 6 h on water bath. The reaction mixture was cooled to room temperature, and the separated Schiff base (HL) (Figure 9) was collected by filtration, washed with ethanol, dried and recrystallized from absolute ethanol (Scheme 1).

Preparation of Cu(II), Co(II), Ni(II), Zn(II), Hg(II), Mn(II), and Fe(III) Complexes of Ligand HL.
To the hot solution of 3-chloro-N -[(IE)-(2-hydroxy phenyl)methylene]-6-methoxy-1-benzothiophene-2-carbohydrazide (HL) ( Figure 10) (0.002 mol) in ethanol (30 mL) was added a hot ethanolic solution (10 mL) of respective metal chloride (0.002 mol); the reaction mixture was refluxed on a steam bath for 4 h, then sodium acetate (0.5 g) was added to it and refluxed for further 2 h. It was then poured into distilled water. The resulting solid complexes were collected by filtration, washed with sufficient quantity of distilled water, then with hot ethanol to apparent dryness, and dried in a vacuum over anhydrous calcium chloride in a desiccator ( The bacterial and fungal cultures were inoculated in nutrient broth (inoculation medium) and incubated overnight at 37 ∘ C. Inoculated medium containing 24 h grown culture was added aseptically to the nutrient medium and mixed thoroughly to get a uniform distribution. This solution was poured (25 mL in each dish) into Petri dishes and then allowed to attain room temperature. Wells (6 mm in diameter) were punched carefully using a sterile cork borer and were filled with test solution 25 L. The plates were allowed to stand for an hour in order to facilitate the diffusion of the drug solutions, then the plates were incubated at 37 ∘ C for 24 h for bacteria and 48 h for fungi and the diameter of the zone of inhibition was measured [21]. The results were compared with those of standard drug streptomycin for bacterial and fluconazole for fungal activity of the same concentration as that of the test compounds under identical conditions.

Antioxidant Activity by DPPH Radical
Scavenging Activity. The free radical-scavenging activity of the compounds was measured in terms of hydrogen donating or radical scavenging ability using the stable radical DPPH described by Blois method [22][23][24]. Briefly, stock solution of the sample (0.001 g/mL) was prepared by dissolving it in DMSO. 3 mL solutions of each with varying concentration (25-100 g) were prepared from the stock solution in methanol. Solution of DPPH (0.01 mM) in methanol was prepared and 1 mL of this solution was added to the above test solutions. The mixture was shaken vigorously and incubated for 30 min, and then the absorbance was measured at 517 nm. All the tests were run in triplicate and expressed as the mean ± standard deviation (S.D). Vit-C and Vit-E were used as standard or positive control, parallel to the test compound and in the absence of the test compound/standard used as the negative control. The capability to scavenge the DPPH radical was calculated using the fallowing equation: Percentage of scavenging activity = − × 100, (1) where corresponds to the absorbance of the negative control that is without sample; is the absorbance of sample with complex or ligand.

Results and Discussion
The physical and analytical data of the synthesized ligand HL and its Cu(II), Co(II), Ni(II), Zn(II), Hg(II), Mn(II), The Scientific World Journal 3  [26] is evidenced by the appearance of a broad band at 2901 cm −1 . Vibrations because C-O was observed at 1235 cm −1 . The band due to C-S-C of thiophene ring was found to appear at 1518 cm −1 ( Table 2).

IR Spectra of Cu(II), Co(II), Ni(II), Zn(II), Hg(II), Mn(II), and Fe(III) Complexes of Ligand HL.
Appearance of band in the region 3112-3334 cm −1 in the IR spectra of Cu(II), Co(II), Ni(II), Mn(II), and Fe(III) complexes of ligand HL is assigned to ] NH of amide function [27]; further appearance of strong intensity bands in the region 1585-1635 cm −1 with a shift in the lower frequency side in these complexes by 16-66 cm −1 when compared to C=O of ligand which appear at 1651 cm −1 indicates the involvement of C=O oxygen atom in bonding with metal ions as such without undergoing enolization. But in case of Zn(II) and Hg(II) complexes of ligand HL, disappearance of the band due to ] NH of amide function [28] and carbonyl function which were found to appear at 3351 and 1651 cm −1 , respectively, in case of ligand clearly indicates the enolization of carbonyl function and subsequently coordination of enolized carbonyl oxygen to Zn(II) and Hg(II) ion via deprotonation [27]. The fact of enolization of amide function was further conformed by the appearance of a new band (enhanced intensity) in the region 1601 cm −1 in the case of these Zn(II) and Hg(II) complexes. Medium intensity band which appeared at 1618 cm −1 in case of ligand HL was found to be shifted to lower frequency side by 58-83 cm −1 and appeared in the region 1535-1560 cm −1 in all these complexes of ligand HL suggesting the coordination of the nitrogen atom of the azomethine function of ligand with metal atoms. In the IR spectra of all these complexes, the absence of band due to intramolecular hydrogen bonded phenolic OH vibration which appeared at 2901 cm −1 in ligand clearly indicates the deprotonation of hydrogen bonded phenolic OH group during complexation with metal ions. The band due to phenolic C-O function observed at 1235 cm −1 in case of ligand HL has been found to be shifted to higher frequency side by 83-67 cm −1 in the complexes of the ligand HL under present study and appeared in the region 1302-1318 cm −1 , confirms the coordination of phenolic oxygen with the metal ions via deprotonation.
The thiophene ring vibration of the ligand which appeared at 1518 cm −1 in case of ligand remained unaffected in all the complexes and appeared at about the same region  Table 2.

Mass Spectrum of the Ligand HL.
In the mass spectrum of the ligand HL (Figure 2), the molecular ion peak M + • was observed at m/z 360, 362 (96%, 33%). The molecular ion underwent fragmentation to give fragment ions at m/z 241, 243 (92%, 32%) due to the expulsion of C 7 H 5 NO radical from the molecular ion. This fragment ion on further simultaneous loss of CS and HCNO species gave another fragment ion recorded at m/z 154, 156 (100%, 31%), which is also a base peak. This fragment ion further loses a chloride radical and gave a fragment ion recorded at m/z 119 (3%), (Scheme 2).

Electronic Spectra of Cu(II), Co(II), Ni(II), Mn(II), and Fe(III) Complexes of the Ligand HL.
Electronic spectral data of the Cu(II), Co(II), Ni(II), Mn(II), and Fe(III) complexes of the ligand HL are given in Table 3. Electronic spectral studies of all these complexes were carried out in DMF at 10 −3 M concentration.

Ni(II) Complex.
The ground state of Ni(II) in octahedral coordination is 3 A 2g (t 2g 6 eg 2 ). The Ni(II) complex shows three transitions in an octahedral field, namely, The bands in the region ∼10000, ∼12000, and ∼ 25000 cm for the transitions mentioned above are characteristics of octahedral geometry.
The electronic spectrum of Ni(II) complex of the ligand HL under present investigation exhibited three bands in the region 10590 cm −1 , 16598 cm −1 , and 25617 cm −1 which are assigned to 3 , and 3 A 2g (F) → 3 T 1g (P) (] 3 ) transition. All these observations favor the octahedral geometry for Ni(II) complex of the present study.

Mn(II) Complex.
The ground term of Mn(II) ion is in the sextet. The only sextet term of the d 5 configuration in octahedral stereochemistry is the 6 A 1g . The transitions of the spectrum are assigned as from the 6 A 1g ground term to the quartet excited term. Electronic absorption spectra of Mn(II) octahedral complex are expected to show four spin allowed transitions. The four narrow absorption bands approximately around 18000 cm −1 , 24750 cm −1 , 29500 cm −1 , and 31900 cm −1 were assigned to 6     Ni(II) complex the observed magnetic moment value is 2.90 BM which is well within the expected range for Ni(II) complex with octahedral stereochemistry 2.83-4.00 BM [36,37]. For Mn(II) and Fe(III) complexes the observed magnetic moment value is 5.68 BM and 5.91 BM, respectively, which are characteristic of octahedral geometry (Table 1).

ESR Spectral Studies of the Cu(II) Complex of the Ligand HL.
The X-Band ESR spectrum of the powder Cu(II) complex was recorded at room temperature using DPPH as reference standard. One unpaired electron in Cu(II) complex with 2 B 1g as ground state lies in dx 2 -y 2 orbital and follows the trend ‖ > ⊥ > ( = 2.0036 free ion value). The observed ‖ = 2.36, ⊥ = 2.07 values of the Cu(II) complex under the present study followed the same trend ‖ > ⊥ > which suggest the presence of unpaired electron in dx 2 -y 2 orbital giving octahedral geometry [35].
The observed = 5.14 for the complexes under present study evidenced the monomeric nature of the complexes; this fact is further supported by the absence of a band corresponding to ΔMS = ±2 transition in the observed ESR spectrum which is characteristic of monomeric complex (Table 3). Consider 3.8. Thermogravimetric Analysis. Thermogravimetry (TG) is a technique in which the change in mass of the sample is determined as a function of temperature and/or time. Among all the methods, TG is the most widely used one. From TG curve, information related to the thermal stabilities, composition of the initial sample, intermediate compounds that are formed, and the final residue could be obtained ( Figure 3). The TGA study on [Cu(HL) 2 ] and [Co(HL) 2 ] was carried out in the temperature range 38.69 ∘ C to 800 ∘ C.

Cu(II) Complex. The decomposition studies of the compound Cu(II) complex [Cu(HL) 2 ] have been carried out.
In the thermogram of the [Cu(HL) 2 ], ( Figure 4) the first stage of the decomposition represents the weight loss of N 2 H 2 at 97.52 ∘ C, with weight loss of 4.14%. The theoretical weight loss for this decomposition was 3.83% agreeing with observed value of 4.14%. The complex underwent further degradation and gave break at 218.27 ∘ C with a weight loss of 26.22%, which corresponds to the simultaneous decomposition of the C 9 H 6 OSCl and H 2 species. This practical weight loss 26.22% is in accordance with theoretical weight loss of 25.47%. The third stage of decomposition occurs at 290.97 ∘ C, with weight loss of 25.53% which corresponds to the decomposition of C 9 H 6 OSCl species. This practical weight loss 25.53% is in accordance with theoretical weight loss of 25.80%. The fourth stage degradation at 328 ∘ C with weight loss of 16.73% corresponds to the decomposition of C 7 H 6 N 2 O species. This practical weight loss 16.73% is in accordance with theoretical weight loss 16.84%. Thereafter compound showed a gradual decomposition up to 800 ∘ C and onwards. The weight of the residue corresponds to copper oxide. The thermal decomposition of [Cu(HL) 2 ] with probable assignments is given in Table 4.  Loss of mass (%) Probable assignments

Powder X-Ray Diffraction
3.9.1. Cu(II) Complex. Powder X-ray diffraction pattern for Cu(II) complex of ligand HL has been depicted in Figure 6.
The Copper complex of [Cu(HL) 2 ] showed nine reflections in the range of 5-80 ∘ (2 ) arising from diffraction of X-ray by planes of complex. The interplanar spacing (d) has been calculated from the position of intense peak using Bragg's equation where = wavelength of X-ray used (Cu K = 1.54Å).
The Scientific World Journal 9  The calculated spacing together with relative intensities with respect to most intense peak has been recorded in Table 6.
The 2 value with maximum intensity of the peak for the compound was found to be 25.457 (2 )   been listed in Table 6. The entire important peaks have been indexed and unit cell calculations have been made for cubic symmetry of the complex. The observed values of interplanar distance have been compared with the calculated ones. It was observed that there is good agreement between the calculated and observed values. The experimental values of sin 2 /common factor are recorded for each peak in Figure 6. The (h 2 + k 2 + l 2 ) values are 1, 2, 3, 4, 7, 10, 12, 15, and 20. The presence of forbidden no 7 and 15 indicates that the Cu(II) complex may belong to hexagonal or tetragonal system.  3.9.2. Co(II) Complex. Powder X-ray diffraction pattern for Co(II) complex of ligand HL has been depicted in Figure 7. The copper complex of [Co(HL) 2 ] showed eleven reflections in the range of 5-80 ∘ (2 ) arising from diffraction of X-ray by planes of complex. The interplanar spacing (d) has been calculated from the position of intense peak using Bragg's equation The calculated spacing together with relative intensities with respect to most intense peak has been recorded in Table 7.
The 2 value with maximum intensity of the peak for the compound was found to be 26.5511 (2 ) which corresponds to = 3.35453Å. The 2 values for the prominent peaks have been listed in Table 7. The entire important peaks have been indexed and unit cell calculations have been made for cubic symmetry of the complex. The observed values of interplanar distance have been compared with the calculated ones. It was observed that there is good agreement between the calculated and observed values. The experimental values of sin 2 /common factor are recorded for each peak in Figure 7. The (h 2 + k 2 + l 2 ) are 1, 2, 3,5,9,12,17,20,30,43, and 54. The absence of forbidden no 7, 15, and 23 conforms the cubic symmetry. The experimental values are in good agreement with (h 2 + k 2 + l 2 ) values of primitive type cubic cell, with lattice parameter equal to = = = 9.9518.

Antimicrobial Activity.
Antimicrobial activity was carried out by the cup-plate method [33]. The ligand HL and its Cu(II), Co(II), Ni(II), Zn(II), Hg(II), Mn(II), and Fe(III) complexes have been tested for their antibacterial and antifungal activity at 1 mg/mL concentration. The results of the antimicrobial activity have been presented in Table 8. The results of the antibacterial activity testing showed that the complexes of Hg(II) and Mn(II) showed good activity, complexes of Cu(II), Co(II), Ni(II), and Fe(III) exhibited moderate activity, and ligand HLand complex of Zn(II) showed less activity against E. coli when compared with that of standard drug Streptomycin. The complexes of Hg(II) and Mn(II) showed good activity, and complexes of Cu(II), Co(II), Ni(II), and Fe(III) exhibited moderate activity against S. aureus when compared with that of standard drug Streptomycin.
The results of the antifungal activity testing of the ligand HL and its Cu(II), Co(II), Ni(II), Zn(II), Hg(II), Mn(II), and M = Hg and Zn

Antioxidant Activity by DPPH Radical Scavenging
Activity. DPPH scavenging activity of ligand and its complexes against vitamin-E as standard were analyzed at 50-100 g in DMF solution at = 30 min. This investigation indicates that there is a great possibility of finding potent antioxidants. The Cu(II), Ni(II) Zn(II), and Fe(III) complexes have exhibited very good free radical scavenging activity, and Co(II) showed moderate activity. Ligand HL and its Hg(II) and Mn(II) complexes showed less activity compared with Vit-E. The bar graph representation of percentage of free radical scavenging activities is shown in Figure 8.

Conclusion
In the light of above discussion we have proposed that Cu(II), Co(II), Ni(II), Mn(II), and Fe(III) complexes have exhibited octahedral geometry whereas Zn(II) and Hg(II) complexes exhibited tetrahedral geometry. The ligand behaves as ONO tridentate chelating agent coordinating through the deprotonation of hydroxyl group, carbonyl group, and azomethine nitrogen. The elemental analysis, electrical conductivity measurements, IR spectra, 1 H NMR, mass spectral data, electronic spectra, magnetic susceptibility, ESR spectra, and TGA revealed mononuclear nature of the complexes. The ligand HL was found to be less active against the bacteria as well as fungi whereas its complexes were found to be highly active against the bacteria as well as fungi and some of the complexes showed good antioxidant activity. On the basis of spectral evidence following structures have been assigned for synthesized complexes.