Substituted benzimidazoles: A novel class of anti-tubercular agents

Reaction of O-phenylenediamine with phthalic anhydride gives 2H-benzimidazole -1-yl-3, 8 benzpyrolidine 2-one (I) This on condensation with anthranilic acid and 4-aminoacetophenone gives II. II on reaction with different aldehydes in presence of sodium hydroxide and ethanol stirred at room temperature for 24 h gives N-[4-(3-Aryl-acryloyl)–phenyl]–2-(1H– benzimidazol–2–yl)–benzamide (III-X). All the compounds have been characterized by elemental analysis, IR and NMR and Mass spectral analysis. All the compounds have been evaluated for their anti-tubercular activity against different stain of mycobacterium species.


Introduction
Benzimidazole nucleus has been of great interest to synthetic and medicinal chemists for a long time due to their unique chemical & biological properties. Benzimidazoles & its derivatives have been found to possess biological activity such as anti-viral, 1 anti-baterial, 2 anti-tungal, 3 antiinflamatory 4 and anti-cancer 5 activity etc. Recently including tuberculastatic activity [6][7][8][9] of benzimidazole derivatives was reported. These reports prompted us to synthesize different Benzimidazole derivatives (III-X). Continuous increase in bacterial resistance to existing drugs has been resulted due to wide spread use of antibiotics leading to research on new substances possessing antimicrobial activity [10][11][12][13][14][15][16][17] In the present study the N-[4-(3 -Aryl-acryloyl)phenyl] -2-(1 Hbenzimidazol -2yl)benzamide (III-X) were prepared by the reaction of O-Phenylenediamine with phthalic anhydride and heated under reflux at 110° C yields compound (I), this on condensing with anthranilic acid and 4-aminoacetophenone in dimethyl formamide heated under reflux at 110° C gives II. The compound II is on treating with different aldehydes in presence of 40% sodium hydroxide and ethanol stirred at room temperature for 24 h gives III-X. The resulted compounds were characterized on the basis of elemental and spectral analysis.

Experimental
The purity of the synthesized compounds were ascertained by thin layer chromatography on silica gel G in various solvent systems using iodine vapours as detecting agent. Melting points were determined in open capillary tubes and are uncorrected. Infrared spectra were recorded on perkin -Elmer FT-IR spectrometer (Spectrum RX1) as KBr disks. Proton NMR spectra were recorded on varian AMX 400 instrument (300 MHz) using tetra methyl silane as internal standard.

Results and Discussion
Antimicrobial Activity All the compounds were screened for their antibacterial activity against gram positive strains (Staphylococcus aures, proteus vulgaris and streptococcus pyogens) and gram negative strains (Escherichia coli, pseudomonas aeroginosa and klebsiella pneumonia) using ciprofloxacin as a standard.
The antifungal activity was performed against candida albicans and Cryptococcus informans using griseofulvin as a standard. The MIC of the compounds tested in this study was determined according to the method of Goto et al 18 by a serial dilution technique. The inoculum size was approximately 10 6 colony forming units (cFu/m).
The reference standard ciprofloxacin inhibits the gram negative bacteria viz., E.coli, pseudomonas and klebsiella pneumonia at a MIC of 0.15g/ml and 0.2g/ml respectively whereas against gram positive bacteria viz., S. aures, P. Vulgaris and St.Pyogens at a MIC of 10g/ml and 15g/ml respectively. The screening results revealed that the compound III, VI, VII, IX and X showed good activity against gram positive strains (S. aures) and gram negative strain (E.Coli) than other compound. The screening results are given in Table III. The majority of the active compounds possessed phenyl substituted derivatives.

Scheme of Synthesized Compounds
The physical data of the synthesized compounds recorded in Table I.

Tuberculostatic Activity
The compounds obtained were tested towards the standard Mycobacterium tuberculosis H37Rv strain as well as two strains isolated from tuberculosis patients: one resistant to the isonicotinic acid (INH), ethambutal (ETB) and rifampicin (RFP), the other fully susceptible to the tuberculostatics administered. Tuberculostatic activity was determined in vitro by classical test tube method with Youman's liquid medium containing 10% of bovine serum. On the ground of the minimum inhibiting concentration (MIC) values obtained one may conclude that some of the compounds tested exhibited a high tuberculostatic activity. The most active compounds were III, VI, VII, VIII, IX and X as their MIC values were within limits of 6-50g/ml ( Table 2).The majority of the compounds possessed amide substituted phenyl substitutents of benzimidazole system. The presence of benzimidazole alone did not show the activity.