1. INTRODUCTION:

The versatility isatin based Schiff base compounds having, acyl, aroyl and heteroacroyl Schiff bases have addi- tional donor sites >C=O, >C=N-, etc. has made them good chelating agents that can form a variety of complexes with various transition and inner transition metals and has attracted the attention of many researchers [1]. Moreover, Schiff bases are regarded as privileged ligands [2]. Due to their capability to form complexes with different transition metals can act as catalysts for many different reactions [3, 4]. Recently, complexes of type, [ML]Cl(2) [M=Co(II), Ni(II), Cu(II) and Zn(II)] were reported [5] where, the Schiff base ligand L= bis(indoline-2-one)triethylenetetramine obtained from condensation of triethylenetetramine and isatin.

Transition metals, in combination with a variety of ligands, has been shown to exhibit cytotoxic or antibiotic activity. The transition metals, primarily Cd(II), Ni(II), Co(II) and Zr(IV) has been linked with a variety of ligands with various degrees of cytotoxic and/or antibiotic activity [6]. The emergence of strains resistant to antimicrobial agents may be bringing to an end the so called “antibiotic era” [7].

The widespread and increasing resistance of bacterial and fungal pathogens to commonly used antibiotics and chemotherapeutics has provided the necessary impetus to find alternative drugs and/or therapies to which microorganisms will not easily develop resistance. One example of these relatively novel strategies (therapies) is antimicrobial metal complexes, which is expected to be useful in the treatment of several severe infections[8]. To date, there has been no signiﬁcant systematic study designed to elucidate the role of the transition metal complexes [e.g., Metal complexes of Cd (II), Ni (II), Co (II) and Zr (IV)] or ligand in antibiotic activity.

Earlier, we studied electronic properties of N₂O₄schiff base ligand containing metal complexes of Cd(II), Pd(II), Hg(II) and Zr(IV) [9-11]. In this study, we are motivated to undertake a systematic study of preparation, characterization, the MICs, minimal bactericidal concentrations (MBCs), and antimicrobial activities of a group of newly prepared [M(L)(Cl)]Cl type transition metalcomplexes [where, M = Cd(II), Pd(II), Hg(II) and Zr(IV) and L = bis(indoline-2-one)diethylenetriamine].

2. EXPERIMENTAL:

2.1 Measurements and materials:

Electronic spectra were recorded on a Thermoelectron Nicolet evolution 300 UV-Vis spectrophotometer. All chemicals were commercial products and were used as supplied.

2.2 Synthesis of bis(indoline-2-one)diethylenetriamine (L):

To a stirring solution of isatin (0.2942g, 2 mmol) dissolved in 25ml of ethanol, a solution of diethylenetriamine (0.1030mL,1mmol) in 10mL ethanol was added drop wise. This has resulted a dark orange solution, which was refluxed for 6h.The reaction mixture was cooled and kept for evaporation at room temperature leading to isolation of solid product. The product thus formed was filtered washed several times with ethanol and finally with diethyl ether and dried in vacuum over anhydrous CaCl₂. The product was found to be soluble in DMF and DMSO [Scheme 1].

Scheme 1:Synthesis of Bis(indoline-2-one)diethylenetriamine (L).

2.3 Synthesis of Metal- schiff base [bis(indoline-2-one)diethylenetriamine] complexes:

1 mmol (0.2013gm) solution of CdCl₂ dissolved in ethanol (15 mL) was taken in a two necked round bottom flask and kept on magnetic stirring. To this solutions of isatin (0.2942 gm, 2 mmol) dissolved in 25 mLof ethanol and diethylenetriamine (DETA) (0.1030 mL, 1 mmol) in ethanol (10 mL) were simultaneously added Drop wise followed by refluxing for about 8h leading to isolation of precipitated product. The complexes thus formed were filtered and washed several times with ethanol to remove any traces of unreacted starting materials and were further washed with diethyl ether and dried in vacuum over anhydrous CaCl₂. The complexes were soluble in DMF and DMSO[Scheme 2].

Scheme 2: Synthesis of Cd(II) Schiff base complex. Where, M = Cd(II), Pd(II), Hg(II) and Zr(IV)

3. RESULTS AND DISCUSSION:

3.1 Physical properties of the complexes:

Some physical properties of the complexes are shown in the (Table 1). The molar conductance values are in the region range 0.2 to 1.4 Ω^‑1_cm²_mol^-1_.The observed values of effective moment (µ_eff) of the complexes at room temperature shown that all the complexes are paramagnetic except [Cd(L)(Cl)]Cl which is diamagnetic in nature. The melting point ranges from 225-255.

3.2 Infrared spectra:

IR spectral data are shown in (Table 2). The strong bands obtained at 3400, 1715 and 1620 cm^-1 due to ν(-NH), ν(C=O) and ν(C=N) respectively. The presence of metal ligand bonding is evident from the appearance of ν(M-O), ν(M-N) and ν(M-N)(-NH) at around 750, 400 and 500 cm^-1 respectively in the spectra of the complexes.

3.3 Electronic spectral studies:

The observed d-d transitions of the complexes are given in Table-2. In addition to d→d transitions, the complexes obtained charge transfer band. The results are consistent with the result of Al-Resayeset. al.[5]. The electronic spectra of Schiff base ligand bis(indoline-2-one)diethylenetriamine and its Cd(II) complex is shown in Figure-1(A and B) respectively.

Table 1:Physical properties of complexes

Compounds	Colour	Melting point (±5⁰C)	Molar conductance Ohm ^-1cm²mol^-1	µ_eff(B.M)
[Cd(L)(Cl)]Cl	Red	250	1.065	Dia.
[Ni(L)](Cl)]Cl	Red	255	0.210	0.84
[Co(L)(Cl)]Cl	Ash	225	1.385	5.41
[Zr(L)(NO₃)]NO₃	Red	225	0.210	0.79

Where, L= H₂₀C₁₉N₅O₂

Table 2:Major IR spectral data (cm^-1) with their assignment and electronic spectral data of the Schiff base complexes

Compounds	Υ (N-H)	υ(C=O)	υ(C=N)	υ(M-O)	υ (M-N)	υ(MN)(NH)	υ(M-Cl/ NO₃)	d→d (λ/nm)
[Cd(L)(Cl)]Cl	3455	1715	1626	754	397	500	350	530
[Ni(L)(Cl)]Cl	3302	1719	1616	754	350	671	328	445
[Co(L)(Cl)]Cl	3397	1717	1620	756	395	440	351	520
[Zr(L)(NO₃)]NO₃	3410	1710	1622	756	498	665	345	535

Where, L= H₂₀C₁₉N₅O₂

Figure1:Electronic spectra of Schiff ligand bis(indoline-2-one)diethylenetriamine (A) and its Cd(II) complex (B).

4. Antibacterial Screening:

The objective of the study is to evaluate the antibacterial activity of transition metal complexes. Antibacterial activities of complexes A = [Cd(L)(Cl)]Cl, B = [Ni(L)(Cl)]Cl, C = [Co(L)(Cl)]Cl, D = [Zr(L)(NO₃)]NO₃were observed by disc diffusion assay [7,8]. A total of six Gram positive and Gram negative bacteria were used in this antimicrobial screening. Complexes A-D (30mg/disc) were prepared by dissolving with dimethyl sulfoxide (DMSO). To compare the antibacterial activity, kanamycin (30mg/disc) was used as standard antibiotic. As a negative control, a blank disc impregnated with solvent, DMS followed by drying off was used.

4.1 Minimum inhibitory concentration (MIC)assay:

The minimum inhibitory concentrations (MIC) were determined by serial dilution technique [Table 3] (8,9) in the presence of standard Kanamycin. bacterial inocula were prepared at 5×10⁶ - 5×10⁷cfu/ml. Final adjustment were made using optical density measurement for bacteria (absorbance 0.05 at a wavelength of 660 nm).

Table 3: Antimicrobial activity of complexes and standard Kanamycin

Bacterial strains	Zone of inhibition, diameter in mm (30µg/disc)
	A		B		C		D		K
Gram positive
Bacillus caerius	21	12		08		12		30
Staphylococcus aureus	24	09		12		09		31
Bacillus subtilis	22	12		12		13		29
Gram negative
Escherichia coli	12	08		10		08		32
Shigelladysenteriae	24	12		12		10		32
ShigellaSonnei	22	16		16		13		33

Complexes A = [Cd(L)(Cl)]Cl, B = [Ni(L)(Cl)]Cl, C = [Co(L)(Cl)]Cl,and D = [Zr(L)(NO₃)](NO₃),L= H₂₀C₁₉N₅O₂and K = Kanamycin.

It was observed that A, [Cd(L)]Cl₂complex showed great antibacterial activities against Gram positive Bacillus caerius, Staphylococcus aureus and Bacillus subtilis and Gram negative Shigella Sonnei Shigellady senteriae but less active in Escherichia coli. However, complexes B, C and D showed less antibacterial activity against six microorganisms.

Table 4:minimum inhibitory concentrations (MIC) of complexes and standard Kanamycin

Bacterial strains	MIC (µg/disc)
Bacterial strains	A	B	C	D	K
Gram positive
Bacillus caerius	2	128	256	256	2
Staphylococcus aureus	8	256	128	256	2
Bacillus subtilis	4	128	128	128	4
Gram negative
Escherichia coli	16	128	128	64	8
Shigelladysenteriae	4	128	256	128	4
ShigellaSonnei	2	64	64	128	2

The minimum inhibitory concentrations (MIC) of complex A against Bacillus caerius, Staphylococcus aureus, Bacillus subtilis, Escherichia coli, Shigellady senteriae and Shigella Sonnei were found to be 2, 8, 4, 16, 4 and 2 mg/ml. respectively; that of 128, 256, 128, 128,128 and 64mg/ml, respectively for complex B; that of 256, 128, 128,128, 256 and 64mg/ml, respectively for complex C. The MIC of the complex D against all of the above pathogenic bacteria is observed as 256, 256, 128, 64,128 and 128 mg/ml, respectively [Table 4].

It is concluded that among the tested complexes, the Cd(II) complex possesses very high antimicrobial activity with a minimum inhibitory concentration. Further investigations are required to explore the exact mechanism of their cytotoxic properties which may be helpful for exploring new type of potent cytotoxic agent(s) with the hope of adding novel and alternative chemotherapeutic agent(s) in clinical implications.

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Received on 27.05.2014 Modified on 22.06.2014

Asian J. Research Chem. 7(7): July 2014; Page 619-621