Antibacterial activity of the ligand and complexes were determined by the agar well diffusion method [12] . E. coli, P. aeruginosa and S. aureus. Strains of three bacterial species which included Gram-positive bacteria, namely Staphylococcus aureus and Gram-negative bacteria namely Pseudomonas aeruginosa and Escherichia coli were investigated. Ciprofloxacin was used as the standard antibacterial agents. The bacteria isolates were subcultured on nutrient agar plates and incubated at 37˚C for 24 h. A loop full of bacteria cells from the nutrient agar plates was incubated into a nutrient broth (50 ml) at 37˚C for 18 h with vigorous shaking. Using a sterile glass spreader, 18 h bacterial cultures (100 µl) were used to spread a bacterial lawn on nutrient agar [13] . The bacterial strains were grown at 37˚C overnight and maintained on nutrient agar. A stock solution of the compounds were prepared in DMF at 50˚C to give a final concentrations; after pouring into plates and allow the agar to set, plates were inoculated with standardized inocula of the test bacteria, and further incubated at 37˚C for 24 h under aseptic conditions.

The NMR show doublet-doublet for methylene groups (CH₂; 2H) at 1.82 ppm and multiple at 2.35 - 2.40 for (CH₂; 2H), 4.27 ppm, doublet-doublet for protons CH. While the protons at 7.31 - 7.67 ppm were for aromatic ring while amino group located at 8.02 ppm.

Scheme 1. Synthetic procedure used for preparation Schiff base derivative.

The infrared spectrum of the ligand, showed four characteristic bands at 3316 cm⁻¹, 3072 cm⁻¹, 1661 cm⁻¹ and 1544 cm⁻¹, Table 2, which are attributed to amine band (N-H), aromatic band (C-H aromatic), carbonyl band (C=O) and azomethane band (C=N) respectively. The presence of these four bands in the FT-IR spectrum of this ligand indicates the reaction of isatine and 1,2-diamino- cyclohexane and formation of azomethane group in the ligand. The primary (NH₂) group of the reactant isatine has two IR bands and upon reaction with 1,2-diaminocyclohexane, these two bands are disappeared and a band at 3316 cm⁻¹, Table 2, is appeared which belongs to υ (N-H) of the secondary amine (N-H) of the ligand. This is an indication of the reaction of the isatine with 1,2-diaminocyclohexane and formation of the ligand.

Treatment of the ligand with metal chloride of (Co(II), Ni(II) and Cu(II)) in a solvent gave complexes of the types [M(Ligand)Cl₂], where M = Co(II) and Cu(II), [M(Ligand)], where M = Ni(II). The prepared metal complexes are characterized as follow: Elemental analysis data for the synthesized ligand and its metal complexes are shown in Table 1. The metal complexes are consistent with the suggested stoichiometries.

Molar conductivities of all synthesized complexes are measured for (1 × 10⁻³ M) solution in DMSO at room temperature, Table 1 and indicates that they are nonelectrolyte and consistent with the proposed formula for them. The infrared spectra of the prepared ligand and its metal complexes are measured in the range (200 - 4000) cm⁻¹ using cesium iodide (CsI) disc. The characteristic band frequencies are arranged in Table 2.

The infrared spectral data were discussed according to the functional groups present in the prepared complexes as follow: IR data given in Table 2 show broadening of ?N=C and carbonyl peaks, which suggest coordination through these sites. While the peak of −NH remains unaltered, and the peak of non- coordinated carbonyl diminishes due to tautomerism, it can be proposed that the carbonyl peak diminishes due to hydrogen bond formation with free noncoordinated secondary amine group. Therefore, the proposed structure of the coordination complexes produced is given as shown in Figure 1.

Conductance of the complexes are given in Table 1, conductance data show that the metal complexes are non electrolyte indicating the chloride are located inside

the coordination sphere and are directly involved in coordination with the metal center except Ni(II).

The magnetic susceptibility of synthesized complexes is shown in Table 1. The magnetic susceptibility of Co(II) complex is 4.3 B.M [14] , which denotes to an octahedral geometry. The magnetic susceptibility of Ni(II) complex, 3, is 0.06 B.M, which indicates a square-planar geometry [15] . The magnetic susceptibility of Cu(II) complex, is 3.67 B.M, which suggests an octahedral geometry arrangement around Cu(II) [16] .

The treatment of infectious diseases still remains an important and challenging problem because of a combination of factors including emerging infectious diseases and the increasing number of multi-drug resistant microbial pathogens. In spite of a large number of antibiotics and chemotherapeutics available for medical use, at the same time the emergence of old and new antibiotic resistance created in the last decades revealed a substantial medical need for new classes of antimicrobial agents. There is a real perceived need for the discovery of new compounds endowed with antimicrobial activity, possibly acting through

mechanism of action, which is distinct from those of well-known classes of antimicrobial agents to which many clinically relevant pathogens are now resistant [17] [18] [19] . The field of bioinorganic chemistry, which deals with the study of role of metal complexes in biological systems, has opened a new horizon for scientific research in coordination compounds. A large number of compounds are important from the biological point of view [20] [21] . The results of in vitro antibacterial activity of the ligand and its Co(II), Ni(II) and Cu(II) complexes are presented in Figure 5. Amoxicillin was used as positive standards and DMSO was used as negative control for these antibacterial activities. In general, metal (II) complexes have been shown to be more effective than the free ligands and the same was observed in this study, i.e., that the complexes are more active than the parent ligand. The Cu(II) and Co-complexes were found to show bactericidal activity against selected types of bacteria. A fairly moderate effectiveness is exhibited by Ni(II) complex towards E. coli. The ligand show less activity towards E. coli and P. aeruginosa respectively, but had no activity against the organisms E. coli within the range of concentrations considered. The variation in the activity

of different complexes against different organisms depends either on the impermeability of cells of the microbes or differences in the ribosomes of microbial cells [22] . The higher antibacterial activity of M-complexes than the free Schiff base ligands can be explained by chelation of the ligand with metal ions [23] as metal chelates display both polar and nonpolar properties; this makes them suitable for permeation into cells and tissues. The polarity of the metal ion will be reduced to a greater extent due to the overlap of the ligand orbital upon chelation, and partial sharing of the positive charge of the metal ion with donor groups. Chelation increases the delocalization of π-electrons over the entire chelate ring and enhances the penetration of the complexes into lipid membranes [23] [24] . It also increases the hydrophilic and lipophilic nature of the central metalions, probably leading to lipo-solubility and permeability through the lipid layer of cell membranes. Further, lipophilicity, which controls the rate of entry of molecules into the cell, is modified by coordination, so the metal complex can become more active than the free ligand [25] .