Synthesis, Characterization and Screening of Some Schiff Bases as Potential Antimicrobial Agents

Schiff bases derived from 4-amino antipyrene were prepared, and IR and NMR spectral analysis characterized their structure. The Schiff bases produced were (1) 4-((4-chlorobenzylidine)amino)-1,5-dimethyl-1H-pyrazole-3(2H)-one. structure of the compound, solvent used and the bacterial strain under consideration. However, from the present results, it appears that cinnamaldehyde as side chain with 4-amino antipyrene as central ligand and DMF as a solvent to be best in inhibiting the studied bacterial strains.


INTRODUCTION
The molecular manipulation of promising lead compounds is still a major line of approach to new drugs. Molecular manipulations involves the efforts to combine the separate groups of similar activity into one compound, thus making structural changes into the compound leading to changes in the biological activity.
A variety of Schiff bases have been synthesized from various compounds, the main aim being in search of new lead molecules. Savich et al. [17] synthesized Schiff bases from 2-hydroxy-1naphthaldehyde with arylamides. Fioravanti et al.
[18] observed antibacterial and antifungal activity in few Schiff bases derived from N-heteroaryl benzylamines. Shetye et al. [19] have reported the synthesis of Schiff bases derived from 3phenyl salicyaldehyde and studied their biological properties. Yang et al. [20] prepared Schiff bases by using salicylaldehyde and aryl amines. Saghatforoush et al. [21] have prepared tetradentate Schiff bases derived from amino thio ether pyridine and salicylaldehyde derivatives and studied their metal complexes. Schiff bases from 4-amino-N-carbamimidoyl benzene sulfonamide have been synthesized by Singh et al. [22] and their antimicrobial activities have also been studied. Amanullah et al. [23] synthesized some novel Schiff bases and studied their cytotoxic, antibacterial activity and physicochemical properties. Deshpande et al. [24] have synthesized some heterocyclic Schiff bases.
Considering the wide range of biological activities of Schiff bases, in the present work, some new Schiff bases have been synthesized from 4-amino antipyrene with different side chains and to evaluate their potency as anti bacterial agents.

EXPERIMENTAL
The following Schiff bases have been synthesized:

Experimental Procedure
Equimolar mixture of 4-amino antipyrine and different aldehydes were dissolved in methanol and the mixture was refluxed for 10-15 hours at 65-70ºC. The mixture was then poured on crushed ice, filtered and dried.

Antibacterial Activity
The in vitro antibacterial activity of the synthesized compounds was evaluated against two Gram positive bacteria (S. subfava NCIM 2178 and B. megaterium ATCC 9885) and four Gram negative bacteria (P. pseudoalcaligenes ATCC 17440, P. vulgaris NCTC 8313, C. freundii ATCC 10787 and E. aerogenes ATCC 13048).
The antibacterial activity was done by the Agar ditch method using Mueller Hinton Agar No.2 as the nutrient medium [27]. Stock solutions (10 mg/ml) were prepared by dissolving the compound in DMSO or DMF. The bacterial strains were activated by inoculating a loop full of test strain in 25 ml of N-broth and the same was incubated for 24 h in an incubator at 37ºC. 0.2 ml of the activated strain was inoculated in Mueller Hinton Agar. Mueller Hinton Agar kept at 45ºC was then poured in the Petri dishes and allowed to solidify. After solidification of the media, 0.85 cm ditch was made in the plates using a sterile cork borer and these were completely filled with the test solution. The plates were incubated for 24 h at 37ºC. The mean value obtained for the three wells was used to calculate the zone of growth inhibition of each sample. The controls were maintained for each bacterial strain and each solvent, where pure solvent was inoculated into the well. The inhibition zone formed by these compounds against the particular test bacterial strain determined the antibacterial activities of the synthetic compounds.

RESULTS AND CONCLUSION
Seven compounds were synthesized and IR and NMR spectral data analysis confirmed their molecular structure. Their data are given below: The molecular formula, molecular weight, melting point, % yield and R f values along with the solvent systems of 7 Schiff bases are given in Table 1.
The antibacterial activity of seven synthetic compounds derived from 4-amino antipyrene in DMSO and DMF against B. megaterium and S. subfava are shown in Fig. 1. Maximum antibacterial activity was shown by SBS3 in both the solvents against B. megaterium; SBS3 showed more activity in DMF than in DMSO. SBS1 and SBS2 showed slight activity only in DMF while SBS5 and SBS6 showed activity only in DMSO against this bacterial strain. SBS4 showed some activity in DMSO and very negligible activity in DMF. SBS7 did not show any activity in either of the solvent. Against S. subfava, all the compounds show quite less activity in both the solvents as compared to B. megaterium. SBS3 showed maximum activity in both the solvents against this bacterial strain also. SBS1, SBS2 and SBS7 showed slight activity only in DMF and rest of the compounds did not show any activity in both the solvents against this bacterial strain. The differential response of the synthesized compounds is because of the difference in their structure and also the solvent in which they are dissolved. All the seven compounds have 4-amino antipyrene as their central ligand and the side chains are different. In SBS1 it is chlorobenzaldehyde, in SBS2 it is hydroxy benzaldehyde, in SBS3 it is cinnamaldehyde, in SBS4 it is vanillin, in SBS5 it is benzaldehyde, in SBS6 it is furfuraldehyde and in SBS7 it is p-anisaldehyde.
The antibacterial activity against the Gram negative bacteria C. freundii and P. vulgaris in both polar solvents DMSO and DMF are shown in Fig. 2. Here an entirely different trend was observed. Only three compounds i.e. SBS1, SBS2 and SBS3 in DMF showed antibacterial activity against C. freundii and maximum activity was shown by SBS3. All other compounds in both the solvents did not show any activity against C. freundii. All the seven compounds showed activity against P. vulgaris extracted in DMSO while none of the compounds exhibited antibacterial activity in DMF. Maximum activity was shown by SBS1, closely followed by SBS2 and SBS3 while minimum activity was shown by SBS6 and SBS7. This again firmly proves and confirms our earlier conclusion that there is a direct and strong relation between structures of the compound, solvent in which it is extracted and the bacterial strain against which it is being studied.
The antibacterial activity against P. pseudoalkaligens and E. aerogenes are shown in Fig. 3. All the seven compounds synthesized from 4-amino antipyrene extracted in DMSO could not inhibit the growth of either of these two Gram negative bacteria.
The compound extracted in DMF inhibited both these bacteria but a differential effect was envisaged. All the compounds extracted in DMF could inhibit P. pseudoalkaligens but each compound had a different level of inhibition. Maximum inhibitory activity was shown by SBS4 and SBS5 followed by SBS3, SBS6 and SBS7 respectively. Minimum activity was by SBS1 and SBS2. In E. aerogenes, only SBS3 showed some inhibition while all the other compounds showed no activity.

. Antibacterial activity of synthesized compounds against gram negative Bacteria
In the present work, among the seven synthesized compounds and the two solvents used, the best activity was shown by SBS3 in DMF solvent. Hence, it can be concluded that cinnamaldehyde as side chain with 4-amino antipyrene as central ligand and DMF as solvent are the best in inhibiting the studied bacterial strains. This once again poves the earlier conclusion that antibacterial activity is dependent on molecular structure of the compound, solvent used and the bacterial strain under consideration [25,26]. Such screening of various organic compounds and identifying active agents is the need of the hour; because successful prediction of lead molecule and drug like properties at the onset of drug discovery will pay off later in drug development.