INTRODUCTION:

Schiff base plays an important role in inorganic chemistry as they easily form stable complexes with most transition metal ions. Studies on synthesis, structural investigation and reaction of transition metal Schiff base complexes have received a renewed attention in recent years, because of their biological activities. Hydrazones derivative Schiff bases are widely used in analytical chemistry as a selective metal extracting agents as well as spectroscopic determination of certain transition metal ions^1,2, e.g., thiosemicarbazones^3,4. Most of the thiosemicarbazones contain NNS donor atoms; however, very little information is available for hydrazones containing ONO or NNO donor atoms. Copper (II) complex of salicylaldehyde benzoylhydrazone has shown to be a potent inhibitor of DNA synthesis and cell growth. This hydrazone has mild bacteriostatic activity and a range of analogues compounds has been investigated as potential for oral iron chelating drugs for genetic disorders such as thalassemia⁵. Another interesting feature of aroylhydrazones is the possibility of keto-enol tautomeric interconversion⁶. As a consequence, aroylhydrazones can act as neutral, monoanionic or dianionic ONO tridentate donor ligands⁷.

The coordination compounds of aroylhydrazones have been reported to act as enzymes inhibitors and also useful for their pharmacological applications. Isonicotinic acid hydrazide [INH] is a drug of proven therapeutic importance and is used as bacterial ailments e.g., tuberculosis ⁸. Hydrazones derived from condensation of isonicotinic acid hydrazide with pyridine aldehyde have been found to show better antitubercular activity than INH ⁹. In view of the versatile importance of hydrazones, we herein described the synthesis, characterization and antimicrobial study of isonicotinic acid hydrazide derivative Schiff base transition metal ion complexes.

MATERIALS AND METHODS:

All the chemicals were analytical grade and used as received without further purification.

Synthesis of complexes

sonicotinic acid hydrazide (0.274 g, 2 mmol) was dissolved in 20 ml of methanol; to it pyridine-2-carbaxaldehyde (0.214 g, 2 mmol) in 10 ml of methanol was added. The resulting solution was stirred for an hour at 60^oC. The yellow coloured solution was then refluxed to 3 hours and solution changes into red in colour. To the red coloured solution, manganese acetate (1 mmol, 0.245 g) in 15 ml methanol was added drop-wise with constant stirring. The resulting solution was stirred magnetically at 60^oC for two hours. The solution was then cooled at room temperature; crystalline precipitate was formed and collected by filtration, dried in vacuum desiccators over anhydrous CaCl₂. By following the same procedure other complexes were also synthesised. However, cobalt, nickel and copper nitrates were used for complex 2, 3 and 4 respectively.

Physical measurements

Elemental analyses (carbon, hydrogen and nitrogen) were performed on a Perkin - Elmer 2400 – II elemental analyzer (Table 1). Infrared spectra were recorded on a Perkin-Elmer FTIR spectrometer in the range of 4000–400 cm^-1 using KBr pellets. The absorption spectroscopy measurements were performed using a SHIMADZU UV-VISIBLE 2450 Spectrophotometer at 298 K. Magnetic susceptibility measurements were performed using a Sherwood magnetic susceptibility balance (M.S.D) using copper sulphate pentahydrate as standard substance at room temperature and diamagnetic corrections were made using Pascal’s constants.

Antimicrobial activity

The antimicrobial activity of all the synthesized complexes (1-4) were examined against different gram-positive (EF= Enterococcus faecalis, SA= Staphylococcus aureus); gram-negative (PA= Pseudomonas aeruginosa, EC= Escherichia coli, ST= Salmonella typhi) by using disc diffusion method ¹⁰. Petriplates (100 mm) was prepared with 20 ml of sterile nutrient agar (NA) for testing the bacterial. The test cultures were swabbed on the top of the solidified media and allowed to dry for 10 min. Stock solutions of each compounds were diluted in DMSO to produce 1000 μg/ml. The dilutions of the compounds were deposited on 4 mm diameter sterile filters paper discs (10μl/disc) which were subsequently placed on the inoculated petriplates and left for 10 min at room temperature for compound diffusion. Negative control was prepared using DMSO. Ciprofloxacin (5μg/disc) for bacteria were served as positive control. The plates were incubated at 37°C for 24 hrs. The experiment was repeated thrice and the average results were recorded. MIC was defined as the lowest concentration of extract that inhibited visible growth on agar.

RESULTS AND DISCUSSION:

Infrared Spectroscopy

The selected infrared vibration frequencies of the complexes 1-4 are listed in Table 2. The FTIR spectra of the synthesized complexes were recorded within 4000-400 cm^-1. The strong absorption band at 1593-1603 cm^-1are attributed to the C=O stretching frequencies. These bands are assigned to the ν(C=O) of amide functionality. After complexation, these bands have been shifted to lower wave numbers which supports the involvement of C=O in coordination to metals ions ^11,12. The very strong and sharp bands were located at 1500-1521 cm^-1 are assigned to the ν(C=N) stretching vibration of azomethine and these bands are shifted to lower wave numbers supporting the participation of the azomethine group of ligand in binding to the metal ions^13,14. The sharp absorption bands at 1151-1157 cm^-1 may be assigned to the stretching absorption for ν(N-N) ^15,16. New absorption bands with medium to weak intensities appears in the regions 419-667 cm^-1 in the complexes are assigned to ν(M-O) and ν(M-N) ¹⁷.

Table 1: Analytical data for the complexes (1- 4)

Empirical formula (Formula weight)	Found (calculated)
Empirical formula (Formula weight)	C(%)	H(%)	N(%)	µeff(B.M.)
1. C₂₄H₂₀MnN₈O₂ (507.1)	56.81(56.77)	3.97(3.99)	22.08(22.04)	5.7
2.C₂₄H₂₀CoN₈O₈ (511.1)	56.37(56.39)	3.95(3.91)	21.91(21.89)	4.6
3.C₂₄H₂₀N₈NiO₂ (510.1)	56.39(56.35)	3.94(3.90)	21.92(21.76)	2.7
4. C ₄H₂₀CuN₈O₂ (515.1)	55.86(55.89)	3.91(3.87)	21.72(21.76)	1.7

Table 2: Characteristic IR absorption bands (cm^-1) of complexes (1-4).

Complex	ν(C=O)	ν(C=N)	ν(N-N)	ν(M-O)	ν(M-N)
1. Mn(II)complex	1606	1510	1151	533	452
2. Co(II)complex	1593	1500	1154	623	419
3. Ni(II)complex	1598	1504	1154	524	426
4. Cu(II)complex	1603	1521	1157	667	488

Table 3: Thermal analyses data of complexes (1-4)

Complexes (F.Wt)	Dissociation Stage	Temp. range in TG^oC	Weight loss found (calculated) %	Decomposition assignment
C₂₄H₂₀MnN₈O₂ (507.1)	Stage I StageII Stage III	342-387 388-634 655-815	44.4(44.5) 43.7(44.5) 14.3(13.9)	Loss of one mole of ligand Loss of one mole of ligand Formation of MnO
C₂₄H₂₀CoN₈O₂ (511.1)	StageI StageII StageIII	213-495 496-800 801-890	44.9(44.2) 44.8(44.2) 15.1(14.6)	Loss of one mole of ligand Loss of one mole of ligand Formation of CoO
C ₂₄H₂₀NiN₈O₂ (510.1)	Stage I	278-502	44.1(44.3)	Loss of one mole of ligand
	Stage II	503-719	44.1(44.3)	Loss of one mole of ligand
	Stage III	720-888	15..1(14.4)	Formation of NiO
C₂₄H₂₀CuN₈O₂ (515.1)	Stage I	251-296	44.1(43.8)	Loss of one mole of ligand
	Stage II	297-480	44.8(43.8)	Loss of one mole of ligand
	Stage III	481-874	14.8(15.1)	Formation of CuO

UV-Vis. spectra

Mn(II) ion in the complex is d⁵ system and its transitions are both Laporte and spin-forbidden. However, due to instantaneous distortion of the octahedral structure around the metal cation, complex shows an absorption band at 22,809 cm^-1 ^{18, 8}. The UV-Vis. spectra of Co(II) complex shows an absorption peak at 22,779 cm^-1 which may be assigned for ⁴T_1g→⁴T_2g transition ^18,19 , generally observed in Co(II) octahedral complexes.

The electronic spectrum of the Ni(II) complex, showed absorption band at 22,696 cm^-1 which may be assigned to ³A_1g(F)→³T_2g (F), while the absorption due to ³A_2g(F)→³T_2g(P) is overlapped with the ligand absorption bands. This indicates that the Ni(II) ion coordinated to NNO site in an octahedral geometry ^18,20. The spectra of the copper(II) complex showed broad absorption bands at 14,144 cm^-1 which could be attributed to the ²B_1g→²A_1g transitions characterized Cu(II) ion in a octahedral geometry ^18,21.

Magnetic susceptibility measurements

The magnetic susceptibility measurement of the complexes at room temperature was found to be (1) 5.3 B.M., (2) 4.9 B.M., (3) 2.7 B.M. and (4) 1.7 B.M. which is closed to theoretical value of metal (II) octahedral complexes (Table 1) ⁸.

Thermal analyses

The TG-DTA results of the solid Mn(II), Co(II), Ni(II) and Cu(II) complexes are listed in Figure 1, Table 3. The result shows that all complexes have similar decomposition pattern which is in good agreement with the formulae suggested from the analytical data. All the complexes lost one mole of the ligand at first stage and the second mole of the ligand was lost at the second stage accompanied with the formation of the metal oxides ^22,23,24.

Figure: 1 TG plots of the complexes 1-4.

Based on the experimental results obtained from TGA, magnetic moment, elemental analyses, IR spectra, UV spectra, the bonding between Schiff base hydrazone and metal(II) ion can be formulated as in figure 2.

Figure: 2 Molecular structures of the metal (II) complexes

Antimicrobial activity and minimum inhibitory concentration, MIC

The tests were carried for concentrations of 125, 250, 5×10², 1×10³ µg/ml DMSO solutions of the complexes. The inhibition zones caused by the various complexes on the different microorganisms were examined. Activity data of the complexes were observed and compared with common standard Ciprofloxacin (Cip.). The results of the preliminary screening test are listed in Table 4. The antibacterial result shown good activity against some bacteria (E. faecalis, S. aureus, P. aeruginosa, E. coli, S. typhi) in comparison with the standard drug ciprofloxacin (Cip.) except Mn(II) complex. The Co(II) complex showed very good activity against EF, SA, PA, EC and ST at a concentration of 5×10² and 1×10³and MIC (125µg and 250µg). The Ni(II) complex resulted very good activity against EF, SA , ST and PA with the concentration of 5×10² and 1×10³ and MIC found between 62.5µg and 500µg. A similar type of activity was shown by Cu(II) complex also very good activity against EF, SA and ST at 5×10² and 1×10³ concentrations and MIC found to be between 125µg and 500µg Figure 3 (a) and (b).

(a)

(b)

Figure: 3 Zone of inhibition (mm) at 5x10²(a) and 1x10³ (b)

concentrations of complexes (2-4)

CONCLUSION

In the present work we have synthesised four new complexes of Mn(II), Co(II), Ni(II) and Cu(II) with N'-(pyridin-2-yl) methylene) nicotino hydrazide and characterized by different analytical techniques and found that the complexes are inconsistence with our expected structures and revealed octahedral geometry around metal ions complexes; where the ligand act as tridentate chelate with NNO donor sites forming five- membered chelate rings. Antimicrobial activity of the synthesised complexes was done in comparison with Ciprofloxacin as standard to reveal the potency of the synthesized complexes. In all the five selected strains E. faecalis, S. aureus, P. aeruginosa, E. coli, and S. typhi showed sensitivity to all complexes except manganese complex at higher concentrations (1000 μg/ml) and no sensitivity at lower concentrations.

Table 4: Antibacterial activity (inhibition zone) of complexes 1- 4

Name of the compound	Name of microorganism	Inhibition (mm) (Different conc. of each complex. (µg/disc)
Name of the compound	Name of microorganism	1x10³	5x10²	250	125	Cip.	MIC(µg/ml)
	EF	-	-	-	-	14	>1x10³
Mn(II)complex	SA	-	-	-	-	16	>1x10³
	ST	-	-	-	-	14	>1x10³
	PA	-	-	-	-	12	>1x10³
	EC	-	-	-	-	12	>1x10³
Co(II)complex	EF	9	7	-	-	14	250
	SA	10	8	-	-	16	125
	ST	10	7	-	-	14	125
	PA	11	8	-	-	12	125
	EC	8	7	-	-	12	250
Ni(II)complex	EF	10	8	-	-	14	250
	SA	13	10	8	-	16	62.5
	ST	8	-	-	-	14	250
	PA	7	-	-	-	12	250
	EC	7	-	-	-	12	500
Cu(II)complex	EF	10	8	-	-	14	250
	SA	14	12	8	-	16	125
	ST	8	-	-	-	14	250
	PA	-	-	-	-	12	>1x10³
	EC	-	-	-	-	12	>1x10³

Gram-positive = EF - Enterococcus faecalis; SA- Staphylococcus aureus. Gram-negative = PA - Pseudomonas aeruginosa; EC - Escherichia coli; ST – Salmonella typhi. Standard = Cip. – Ciprofloxacin

ACKNOWLEDGEMENT

One of the authors (U.I.S.) would like to acknowledge UGC for financial support of this work, under the Rajiv Gandhi National Fellowship scheme in Science 2009-2010 (vide order No.F.14-2(SC)/2009(SA-III).

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