Transition Metal Complexes of Isonicotinoyl-- hydrazone-4-diphenylaminobenzaldehyde : Synthesis , Characterization and Antimicrobial Studies

A series of complexes of Cu(II), Ni(II), Co(II), Mn(II) and Cd(II) with isonicotinoylhydrazone-4-diphenylaminobenzaldehyde (INHDAB) has been reported. The complexes have been characterized by analytical data, IR, UV-Vis, NMR spectra, magnetic susceptibility values, thermal analysis and for the Cu(II) complex the ESR spectrum has been registered. The biological activity of these complexes were investigated against Staphylococcus aureus, Escherichia coli, Pseudomonas aeruginosa, Salmonella enteritidis, Shigella flexneri bacteria. The INHDAB ligand is coordinate at the metallic ions by oxygen amide (O=C) and the azomethine nitrogen.


Introduction
The heterocyclic chemistry offers the results of the researches show a lot of bioactive substances.The parameters of the chemical structure, the physical and electronical characteristics of the molecule are determined factors in the manifestation of the bioactivity [1][2][3] .Hydrazones and Schiff bases were the subject of many interesting studies due to their important applications in much synthetic areas especially in indicators-chemistry. Hydrazones are being used extensively in detection and quantitative determination of several metals, for the preparation of compounds having diverse structures, analytical chemistry for the identification and isolation of carbonyl compounds 4 .However, the most valuable property of hydrazones is perhaps their great physiological activity.It is well known that the hydrazone group provides a wide range of application in biological and pharmaceutical fields.Therefore, a number of hydrazide-hydrazone derivatives have been claimed to possess interesting antibacterial-antifungal, anticonvulsant, antiinflammatory, antimalarial and antituberculosis activities [5][6][7][8][9][10][11][12] .In the continuation of our earlier studies on the complexes with the ligands from the aroylhydrazone class 13,14 we report here the synthesis and characterization of the complexes of Cu(II), Ni(II), Co(II), Mn(II), Cd(II) with the isonicotinoylhydrazone-4-diphenylaminobenzaldehyde ligand.

Experimental
All reagents and solvents used are of the type AR and were used without further purification.The metal content and chlorine was obtained by the literature methods 15 and carbon, hydrogen and nitrogen were determined with an analyzer CHN-Hewlett Packard 185.The IR spectra were recorded between 4000-400 cm -1 on a BIORAD-FT-IR 135 FTS spectrophotometer in a disc of anhydrous KBr.The electronic spectra in reflection (300-1100 nm) were obtained on a VSU-2P Zeiss-Jena spectrophotometer using MgO as a standard.The ESR spectrum for the Cu(II) complex were registered at room temperature (293 K) on a microcrystalline powder with an ART 5 spectrophotometer.The magnetic moments were determined by the Faraday method at the room temperature. 1H and 13 C NMR spectra were recorded on a device Varian Gemini 300BB in DMSO-d 6 .The molar conductivity of the complexes was measured with a HACH-sens ion 5-conductivitymeter to the solutions in DMF 10 -3 M. The thermic analysis was realized with an MOM-Q-1500 D derivatograph in air with a heating rate of 5 o C/min.The biological activity of the complexes was studied by the diffusion technic in agar plates using DMF as solvent at the concentration of 150 µg/mL.

Synthesis of ligand (INHDAB)
A solution of 0.001 mol of isonicotinoylhydrazine and 0.001 mol of 4-diphenylaminobenzaldehyde in 75 mL (MeOH + C 6 H 6 ) was refluxed for 4 h on a water-bath.After the solution was concentrated the precipitate was filtered and recrystallized from ethanol (Figure 1).

Synthesis of complexes
A methanolic solution of metal chloride MCl2 (0.002 mol/25 mL MeOH) was added to a mixture of isonicotinoylhydrazine (0.002 mol INH/40 mL MeOH) and 4-diphenylaminobenzaldehyde (0.002 mol DAB/60 mL MeOH + C 6 H 6 ).The reaction mixture was refluxed on a water-bath for 4 h after which a part of the solvent was removed by distillation.The precipitated complexes were filtered, washed with methyl alcohol and then with ether and finally dried in vacuum on anhydrous CaCl 2 .

Results and Discussion
The obtained complexes were coloured powders, stable for a long time in the open atmosphere, insoluble in methanol, ethanol, chloroform and acetone; the complex combinations of Ni(II), Co(II), Mn(II) are soluble in DMF but the complexes of Cu(II), Cd(II) are partly soluble.The analytical data suggest a ratio of 1:1 (metal : ligand) for the complexes of Cu(II), Ni(II), Cd(II) and of 1:2 for the complexes of Co(II), Mn(II) (Table 1).The presence of lattice water was confirmed by TG analysis.The molar conductivity corresponding for the Ni(II), Co(II), Mn(II) complexes present low values and in this way it can be assigned a structural formula of non-electrolyte for these.

Infrared spectra
The IR spectra analysis gave information about the mode of coordination of the INHDAB ligand to the metallic ions.The characteristic bands are presented in Table 2.In the ligand spectrum νNH vibration from the imide group is located at 3053 cm -1 .The amide band I is situated at 1672 cm -1 and the band at 1590 cm -1 is assigned to the vibration corresponding to the azomethine group 16,17 .The amide II (δ NH) and amide III (γ NH) vibrations give bands in spectrum situated at 1554 cm -1 and 1331 cm -1 respectively.The weak band from 604 cm -1 shows the "β" deformation in plane from the pyridinic ring.IR spectra of the complexes of Ni(II), Co(II) contain a broad band situated in the range 3400 -3350 cm -1 ; this band is associated to the lattice water from the composition of these complexes 18 .The imido νNH frequency appears in the region 3091-3056 cm -1 and as cosequence the imide group does not changed its structure during the coordination.The amide I vibration shift towards lower values and as a result the hydrazone ligand is coordinated to the metal ions by mean of the amide oxygen (O=C) 19 .The azomethine frequency band νN=C is found to the lower values suggests the azomethine nitrogen coordination in all the complexes 20,21 .The deformation in plane for the pyridinic ring is found at close values from that of free ligand and thus suggests that the nitrogen from the pyridinic ring is not involved in the coordination 22 .The coordination of the azomethine nitrogen of the INHDAB ligand to the Cu(II), Ni(II), Co(II), Mn(II), Cd(II) ions was also proved by the νM-N vibrations appearing in the range 443-412 cm -1 , 23 which were absent in the spectrum of the ligand.

NMR spectra
The INHDAB ligand formation is confirmed by the singlet from δ 8.40 ppm assigned to the azomethine proton (-N=CH-) in 1 H NMR spectrum but also by the peak from δ 140.95 ppm associated to the azomethine carbon in the 13 C NMR spectrum.Beside these values in the 1 H NMR spectrum of the ligand show signal to δ 11.97; δ 8.78 ppm assigned to the protons amide -NH-8; H-2,6 pyridine and also peaks in the 13 C NMR spectrum to δ 161.19; δ 149.25 ppm corresponding to the atoms of carbon amide-7; C-2,6 pyridine.In 1 H NMR spectrum of the complex of Cd(II) the signals of the protons -NH-8 amide and azomethine shift with 0.34 ppm and respectively 0.41 ppm result which sustain the coordination of the ligand by the amide oxygen O=C and the azomethine nitrogen.The signals of the atoms of carbon C-7 and C-10 in the 13 C NMR spectrum present shifts of 2.76 ppm respectively 2.48 ppm and pointed clearly the involvement of the atoms of amide oxygen and azomethine nitrogen in coordination.The atoms of carbon C-2,6 have signals in the same region of the spectrum and the shift is very small and in this way is confirmed that the pyridinic nitrogen is not involved in the coordination.

Electronic spectra, ESR and magnetic determinations
The electronic spectra (Table 3) and the magnetic moments support the stereochemistry of the complexes.The electronic spectrum of the INHDAB ligand presents a maximum of absorption at 28169 cm -1 assigned to the n → π * transition from the C = O and C = N chromophore groups.This transition could be found again in the complex combinations spectra but shifts towards lower frequencies, result which indicates the coordination of the ligand at the metallic ions 24 .
Table 3  The Cu(II) complex displays electronic spectral bands in the region 19607 cm -1 and 14492 cm -1 , which may be assigned to 2 B 1g → 2 E g and 2 B 1g → 2 A 1g transitions in a square-planar stereochemistry 25 .The experimental value of the magnetic moment is of 1.83 BM and suggests a monomeric square-planar geometry.The ESR spectrum of a polycrystalline sample of the complex measured at room temperature gave g ║ and g ┴ values in the range of 2.164 and 2.073, respectively.The high intensity of the signal confirms the monomeric molecular formula.The g ║ and g ┴ values were > 2, corresponding to an axial symmetry with all main axes disposed parallelly.The fact that g ║ > g ┴ > 2.0023 supports a ground state of the Cu(II) ion with the unpaired electron in the d x

2
-y 2 orbital and from the ESR spectrum there results an square-planar geometry 26 .The G parameter determined as G = [(g ║ -2) / (g ┴ -2)] was found to be much less than 4, suggesting considerable interaction in the solid state.For the Ni(II) complex, two absorption bands appeared at 17605 cm -1 and 21834 cm -1 .These bands could be assigned to the transitions 1 A 1g → 1 A 2g and 1 A 1g → 1 B 1g which characterize the Ni(II) ion in the square-planar stereochemistry 27 .The diamagnetism of the complex confirms this symmetry.In the spectrum of Co(II) complex, the bands at 10224 cm -1 , 15137 cm -1 and 19920 cm -1 are associated to the transitions 4 T 1g (F) → 4 T 2g (F) (ν 1 ), 4 T 1g (F) → 4 A 2g (F) (ν 2 ) respectively 4 T 1g (F) → 4 T 1g (P) (ν 3 ).These transitions are specified to the Co(II) ion in the field of octahedral symmetry and the magnetic moment of 4.92 BM corresponds to this geometry 19 .The ligand field splitting energy (10 Dq), interelectronic repulsion parameter (B) and nephelauxetic ratio (β) for the Co(II) complex were calculated using the secular equations given by König 28 and these correspond to the 10 Dq = 11388 cm -1 , B = 724 cm -1 , β = 0.74 values.
10 Dq = 2ν 1 - The value of the β parameter indicates a moderate covalent character for the metalligands bonds.In the case of the Mn(II) complex, the ,,d-d" transitions doubly forbidden from the fundamental term 6 A 1g towards the quartet terms 4 A 1g (G), 4 E g (G), 4 T 2g (G), 4 T 1g (G) exhibit a very small intensity and are concealed by the intraligand transitions 29 .The absorption band at 24096 cm -1 can be attributed to the charge transfer of ligand → metal.The magnetic moment of 5.86 BM supports an octahedral configuration of the type high spin around the Mn(II) ion.The Cd(II) complex presents an intense band situated at 25188 cm -1 which can be assigned to the charge transfer transition L → M 30 .The electronic spectroscopy doesn ' t permit the establish of a clear stereochemistry for the Cd(II) ion but taking into consideration the bidentate behaviour of the INHDAB (HL) ligand as well as the tendency of the Cd(II) ion for the tetrahedral geometry in the tetracoordination complexes, we propose this type of stereochemistry.

Thermal analysis
Thermal analysis by the TG and DTG techniques has proved to be very useful in determining the crystal water content in complexes and their thermal stability and decomposition mode under a controlled heating rate.The determined temperature ranges and the corresponding percent mass losses are given in Table 4.For the complexes of Ni(II), Co(II) the lattice water is eliminated in the domain 85 -155 o C. At high temperatures (t > 240 o C) the complexes were decomposed and the INHDAB ligand was eliminated in two steps by oxidative degradations; in the final step (≈ 700 o C) results the metallic oxides as stable residue: CuO, NiO, Co 3 O 4 , Mn 2 O 3 and CdO.Data from the thermal analysis sustain the molecular formulas assigned for these complexes.The structural formulas assigned to the complexes are presented in Figure 2.  Table 5 shows the obtained results expressed by the diameter of the inhibition zone of the growth bacteria by the tested compounds.The results show that INHDAB exhibited weak activity.The complexes of Cu(II), Co(II) show a weak action compared to all the germs and the complexes of Ni(II), Mn(II) are a little more active compared to Staphylococcus aureus, Pseudomonas aeruginosa and Shigella flexneri.The Cd(II) complex manifest maximum activity to Staphylococcus aureus, Escherichia coli, Pseudomonas aeruginosa and this effect is assigned to the donor atoms which are not involved in the coordination but become more active by the formation of the complex combination 20 .The increased activity of the chelates can be explained based on the overtone concept and the Tweedy chelation theory 31 .According to the overtone concept of cell permeability, the lipid membrane surrounding the cell favours the passage of only lipid-soluble materials, which means that liposolubility is an important factor controlling antimicrobial activity.On chelation, the polarity of metal ion is reduced to a greater extent due to overlap of the ligand orbital and partial sharing of its positive charge with the donor groups.In addition, it is also due to delocalization of the π-electrons over whole chelate ring, enhancing the penetration of the complexes into the lipid membranes and the blocking of the metal binding sites of the enzymes of the microorganisms.

Table 1 .
Analytical and physical data of the complexes.

Table 2 .
Characteristic IR bands (cm -1 ) of the INHDAB ligand and its complexes.

.
Reflection spectra for the ligand and complexes.

Table 4 .
Thermal analysis for the complex combinations.

Table 5 .
Antimicrobial activity of INHDAB and its metal complexes.