Synthesis and Evaluation of Some New Benzimidazole Derivatives for their Anti-Microbial and Anti-Inflammatory Activities

 

Santosh Dighe*, Nachiket Dighe, Pankaj S. Shinde, Ravi Lawre and Sunil Nirmal

PRES’s Pravara Rural College of Pharmacy, Loni, MS, India-413736

*Corresponding Author E-mail:

 

The synthesis, structure and biological activity of Benzimidazole derivatives have long been the focus of research interests in the field of Medicinal Chemistry. A number of Benzimidazole derivatives have been reported to possess interesting biological activities such as antimicrobial activity, 5-HT4 Receptor Antagonists activity, anticancer activity.

 

All synthesized compound were characterized by IR, H­¹-NMR and elemental Analysis. All the compounds were evaluated for Antibacterial activity at the concentration of 200 µcg/mL by using cup-plate agar diffusion method. The activity was carried out on different micro-organisms (E.coli, S. aureus, A.niger, C. albicans) measured in terms of zone of inhibition and compared the standard drug Levofloxacin and Amphotericin B for antimicrobial activity. All the newly synthesized derivatives were screened for Anti-inflammatory activity by an in-vitro method of Inhibition of protein denaturation using Ibuprofen as a standard. These compounds with the suitable molecular modification may prove as a drug of choice in the treatment of microbial infectious disease in future.

 

 

INTRODUCTION

The need of new anti-microbial agents is justified because more microorganisms are being resistance to the present drugs available in the market. Word wide researchers are trying to synthesize new drugs with better pharmacokinetic and dynamic properties with less adverse effects. The literature survey suggests that the Benzimidazole have proved to be good bioactive molecules. They have shown diverse biological activities like anti-bacterial¹, anti-fungal², anti-inflammatory³, 5-HT4 Receptor Antagonists activity⁴, angiotensin-II receptor antagonists⁵, monoamine oxidase inhibitors (MAOIs)⁶, potent AMP-activated protein kinase activators⁷, and anticancer activity⁸ etc. Therefore in view of above facts it was thought of interest to synthesize some Benzimidazole Derivatives. IR, 1H-NMR Spectra and CHN analysis confirmed the structures of the final compounds. The proposed compounds were screened for their antibacterial and anti-inflammatory activities with the standard drugs in the well-equipped microbiology and pharmacology lab by using standard methods.

 

MARERIALS AND METHODS:

EXPERIMENTAL:

Melting points were determined in open capillary method and are uncorrected. Purity of the compound was checked on Silica gel TLC plates. IR spectra were recorded on Jasco FT/IR-4100 spectrophotometer using KBr disc method. 1HNMR spectra were recorded on Bruker Advance –II 400, DMSO as internal standard. Combustion analyses were found to be within the limits of permissible errors.

 

ANTIBACTERIAL ACTIVITY:

The newly synthesized compounds were screened for their antibacterial activity against

 

Escherichia coli (MTCC 443), Bacilus subtilis (ATCC12228) and Staphylococcus aureus (ATCC25923) bacterial strains by disc diffusion method. In all the determinations tests were performed in triplicate and the results were taken as a mean of three determinations. Levofloxacin was used as a standard drug ⁹.

 

ANTI-INFLAMMATORY ACTIVITY:

In-vitro anti-inflammatory activity

Inhibition of protein denaturation

The standard drug and synthesized compounds were dissolved in minimum quantity of dimethyl formamide (DMF) and diluted with phosphate buffer (0.2 M, pH 7.4). Final concentration of DMF in all solution was less than 2.5%. Test solution (1mL) containing different concentrations of drug was mixed with 1 mL of 1mM albumin solution in phosphate buffer and incubated at 27° + 1°C in BOD incubator for 15 min. Denaturation was induced by keeping the reaction mixture at 60° + 1° C in water bath for 10 min. After cooling, the turbidity was measured at 660 nm (UV-Visible Spectrophotometer). Percentage of inhibition of denaturation was calculated from control where no drug was added. Each experiment was done in triplicate and average is taken. The Ibuprofen was use as standard drug. The percentage inhibition of denaturation was calculated by using following formula.

 

% of Inhibition = 100 X [1- Vt / Vc]

 

Where,

Vt = Mean absorbance of test sample.

Vc = Mean absorbance of control^10-12.

 

PROCEDURE FOR SCHEME:

Synthesis of 2-substituted Benzimidazole [A₁ to A₁₁]

0.01 mole of O-phenyldiamine was refluxed with 0.01 mole of CS₂ in presence of 10ml of KOH (10%) for 1 hour. After which the resulting reaction mixture was made to react with substituted aromatic amine to get substituted benzimidazole-2-thiol derivatives.

 

Synthesis of 2-substituted Benzimidazole [B₁ to B₁₁]

0.01 mole of chloro substituted O-phenyldiamine was refluxed with 0.01 mole of CS₂ in presence of 10ml of KOH (10%) for 1 hour. After which the resulting reaction mixture was made to react with substituted aromatic amine to get substituted benzimidazole-2-thiol derivatives.

SCHEME I:

 

Comp. Code	Ar	Comp. Code	Ar	Comp. Code	Ar
A1		A5		A9
A2		A6		A10
A3		A7		A11
A4		A8

SCHEME-II

 

 

Comp. Code	Ar	Comp. code	Ar
B1		B7
B2		B8
B3		B9
B4		B10
B5		B11
B6

 

 

SPECTRAL DATA:

A₁: IR (KBr) cm^-1: 3213.45 (-NH str.), 3063.06 (Ar-CH str.), 2885.60 (-NH str.), 1525.32 (-C=N str), 1245.36 (-C-N str). ¹H NMR: (δ ppm): 6.63-8.36 (8 H phenyl), 12.5 (1H NH), 4.0 (1H NH aliphatic).

 

A₂: IR (KBr) cm^-1: 1510 (- N –O str ), 3010.23 (Ar-CH str.), 1525.32 (-C=N str), 1245.36 (-C-N str). ¹H NMR: (δ ppm): 6.63-8.36 (7 H phenyl), 12.5 (1H NH), 4.0 (1H NH aliphatic).

 

A₃: IR (KBr) cm^-1: 1510 (- N –O str ), 3213.45 (-NH str.), 3010.23 (Ar-CH str.), 1525.32 (-C=N str) 1245.36 (-C-N str). ¹H NMR: (δ ppm): 6.63-8.36 (7 H phenyl), 12.5 (1H NH), 4.0 (1H NH aliphatic).

 

A₄: IR (KBr) cm^-1: 740 (-C –Cl str), 3213.45 (-NH str.), 3010.23 (Ar-CH str.), 1525.32 (-C=N str), 1245.36 (-C-N str). ¹H NMR: (δ ppm): 6.63-8.36 (7 H phenyl), 12.5 (1H NH), 4.0 (1H NH aliphatic).

 

A₅: IR (KBr) cm^-1: 740 (-C –Cl str), 3010.23 (Ar-CH str.), 1525.32 (-C=N str), 1245.36 (-C-N str). ¹H NMR: (δ ppm): 6.63-8.36 (7 H phenyl), 12.5 (1H NH), 4.0 (1H NH aliphatic).

 

A₆: IR (KBr) cm^-1: 3010.23 (Ar-CH str.), 1510 (- N –O str), 1650 (-C=O str), 3200 (-O-H str.),1525.32 (-C=N str), 1245.36 (-C-N str). ¹H NMR: (δ ppm): 6.63-8.36 (7 H phenyl), 12.5 (1H NH), 4.0 (1H NH aliphatic), 12.74 (1H COOH).

 

A₇: IR (KBr) cm^-1: 3213.45 (-NH str.), 1650 (-C=O str),3010.23 (Ar-CH str.), 1525.32 (-C=N str), 1245.36 (-C-N str). ¹H NMR: (δ ppm): 6.63-8.36 (8 H phenyl), 12.5 (1H NH), 4.0 (1H NH aliphatic).

 

A₈: IR (KBr) cm^-1: 3010.23 (Ar-CH str.), 1650 (-C=O str), 1525.32 (-C=N str), 1245.36 (-C-N str). ¹H NMR: (δ ppm): 6.63-8.36 (8 H phenyl), 12.5 (1H NH), 2.0 (1H NH aliphatic), 8.0 (1H NH C=O).

 

A₉: IR (KBr) cm^-1: 740 (-C –Cl str), 3213.45 (-NH str.), 3010.23 (Ar-CH str.), 1525.32 (-C=N str), 1245.36 (-C-N str). ¹H NMR: (δ ppm): 6.63-8.36 (8 H phenyl), 12.5 (1H NH), 4.0 (1H NH aliphatic).

 

A₁₀: IR (KBr) cm^-1: 3213.45 (-NH str.), 1650 (-C=O str), 3063.06 (Ar-CH str.), 2885.60 (-NH str.), 1525.32 (-C=N str), 1245.36 (-C-N str). ¹H NMR: (δ ppm): 6.63-8.36 (8 H phenyl), 12.5 (1H NH), 8.0 (1H NH aliphatic), 2.04 (3H CH₃).

 

A₁₁: IR (KBr) cm^-1: 3213.45 (-NH str.), 3063.06 (Ar-CH str.), 2885.60 (-NH str.), 1525.32 (-C=N str), 1245.36 (-C-N str). ¹H NMR: (δ ppm): 6.63-8.36 (8 H phenyl), 12.5 (1H NH), 4.0 (1H NH aliphatic) 7.40 (2H NH₂).

 

B₁: IR (KBr) cm^-1: 3213.45 (-NH str.), 740 (-C –Cl str), 3063.06 (Ar-CH str.), 2885.60 (-NH str.), 1525.32 (-C=N str), 1245.36 (-C-N str). ¹H NMR: (δ ppm): 6.63-8.36 (8 H phenyl), 12.5 (1H NH), 4.0 (1H NH aliphatic).

 

B₂: IR (KBr) cm^-1:1510 (- N –O str ), 3010.23 (Ar-CH str.), 740 (-C –Cl str), 1525.32 (-C=N str), 1245.36 (-C-N str). ¹H NMR: (δ ppm): 6.63-8.36 (7 H phenyl), 12.5 (1H NH), 4.0 (1H NH aliphatic).

 

B₃: IR (KBr) cm^-1:1510 (- N –O str ), 3213.45 (-NH str.), 3010.23 (Ar-CH str.), 740 (-C –Cl str), 1525.32 (-C=N str) 1245.36 (-C-N str). ¹H NMR: (δ ppm): 6.63-8.36 (7 H phenyl), 12.5 (1H NH), 4.0 (1H NH aliphatic).

 

B₄: IR (KBr) cm^-1:740 (-C –Cl str), 3213.45 (-NH str.), 3010.23 (Ar-CH str.), 1525.32 (-C=N str), 1245.36 (-C-N str). ¹H NMR: (δ ppm): 6.63-8.36 (7 H phenyl), 12.5 (1H NH), 4.0 (1H NH aliphatic).

 

B₅: IR (KBr) cm^-1:740 (-C –Cl str), 3010.23 (Ar-CH str.), 1525.32 (-C=N str), 1245.36 (-C-N str). ¹H NMR: (δ ppm): 6.63-8.36 (7 H phenyl), 12.5 (1H NH), 4.0 (1H NH aliphatic).

 

B₆: IR (KBr) cm^-1:3010.23 (Ar-CH str.), 1510 (- N –O str), 740 (-C –Cl str), 1650 (-C=O str), 3200 (-O-H str.), 1525.32 (-C=N str), 1245.36 (-C-N str). ¹H NMR: (δ ppm): 6.63-8.36 (7 H phenyl), 12.5 (1H NH), 4.0 (1H NH aliphatic), 12.74 (1H COOH).

 

B₇: IR (KBr) cm^-1:3213.45 (-NH str.), 1650 (-C=O str), 3010.23 (Ar-CH str.), 1525.32 (-C=N str), 740 (-C –Cl str), 1245.36 (-C-N str). ¹H NMR: (δ ppm): 6.63-8.36 (8 H phenyl), 12.5 (1H NH), 4.0 (1H NH aliphatic).

 

B₈: IR (KBr) cm^-1:3010.23 (Ar-CH str.), 1510 (- N –O str), 1525.32 (-C=N str), 740 (-C –Cl str), 1245.36 (-C-N str). ¹H NMR: (δ ppm): 6.63-8.36 (8 H phenyl), 12.5 (1H NH), 4.0 (1H NH aliphatic).

 

B₉: IR (KBr) cm^-1:740 (-C –Cl str), 3213.45 (-NH str.), 3010.23 (Ar-CH str.), 1525.32 (-C=N str), 1245.36 (-C-N str). ¹H NMR: (δ ppm): 6.63-8.36 (8 H phenyl), 12.5 (1H NH), 4.0 (1H NH aliphatic).

 

B₁₀: IR (KBr) cm^-1:3213.45 (-NH str.), 740 (-C –Cl str), 1650 (-C=O str), 3063.06 (Ar-CH str.), 2885.60 (-NH str.), 1525.32 (-C=N str), 1245.36 (-C-N str). ¹H NMR: (δ ppm): 6.63-8.36 (8 H phenyl), 12.5 (1H NH), 8.0 (1H NH aliphatic), 2.04 (3H CH₃).

 

B₁₁: IR (KBr) cm^-1:3213.45 (-NH str.), 740 (-C –Cl str), 3063.06 (Ar-CH str.), 2885.60 (-NH str.), 1525.32 (-C=N str), 1245.36 (-C-N str). ¹H NMR: (δ ppm): 6.63-8.36 (8 H phenyl), 12.5 (1H NH), 4.0 (1H NH aliphatic) 7.40 (2H NH₂).

 

 

 

Table no. 1: Analytical and Physicochemical data of the synthesized compounds (A₁-A₁₁)

 

Table no. 2: Analytical and Physicochemical data of the synthesized compounds (B₁-B₁₁)

 

 

Table no: 3 Antibacterial activity of synthesized compounds (A₁-A₁₁) (Scheme-I)

 

Table no: 4 Antibacterial activities of synthesized compounds (B₁-B₁₁) (Scheme-II)

Table no. 5: Anti-inflammatory activity of synthesized compounds (A₁-A₁₁) (Scheme-I)

% inhib. =% inhibition

 

Table no. 6: Anti-inflammatory activity of synthesized compounds (B₁-B₁₁) (Scheme-II)

% inhib. =% inhibition

 

 

Result and DISCUSSION:

Antibacterial activity:

The compounds A₁, A₂, A₃, A₅, A₈, B₄, B₅ ,B_6, B₇, B₈, B₉ has  excellent Antibacterial activity against S. aureus, the compounds A₁, B₄, B₅ have shown Antibacterial activity against B. subtilis, while A₄, B₂, B₃, B₆, B­₇, B₈, B₉ shows Antibacterial activity against E.coli., when compared with standard Levofloxacin

 

Fig. no. 1: Anti-bacterial activity of synthesized compounds (scheme-I and II)

Anti-Inflammatory Activity:

All the compounds were evaluated for Anti-inflammatory activity by Carrageenan Induced Rat hind Paw method. The synthesized compounds A₁, A₃, A₅, A₆, A₈, B₁,B₃, B₆, and B₈ showed better anti-inflammatory activity with 69.66 % inhibition and it was found comparable with standard drug ibuprofen (70.78% inhibition)  at the same dose (100 µg/kg).

 

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Received on 12.11.2014         Modified on 12.12.2014

Accepted on 15.12.2014         © AJRC All right reserved