INTRODUCTION:

Benzimidazole derivatives have been known in literature for their broad range of biological activities.

During the literature survey, it was found that 2-mercapto benzimidazole derivatives¹ namely, 5(6)-chloro-2-(benzylthio) benzimidazole possesses antifungal activity against Trichophyton interdigitale. Similarly, antibacterial and antifungal activity was also reported for benzimidazole derivatives^2,3. Benomyl⁴ is another antifungal compound having benzimidazole moiety with a carbamido linkage. Also 5-methoxy-2-mercapto benzimidazole has been found as an intermediate for Omeprazole⁵ which is a potent antiulcer agent and used in treatment of Zollinger Ellison syndrome.

Hence, keeping in mind the above mentioned literature, we have reported herewith 16 derivatives of substituted 2-(substituted phenylthiocarbamoyl) benzimidazole, whose structures were confirmed by C,H,N analysis and IR as well as NMR spectra and then were screened for their antimicrobial potency.

MATERIALS AND METHODS:

Melting points were determined in open glass capillaries using Thermonik Precision Melting Point Cum Boiling Point Apparatus Model C-PMB-2 and were uncorrected. Purity of the compounds was checked by precoated TLC plates [E. Merck Kieselgel].

C, H, N analysis was carried out on Thermo Finnegan Flash EA 1112. The IR spectra were recorded using KBr pellets on BOMEM FTIR MB–10452M 5903L spectrophotometer. The ¹H-NMR spectra were recorded on JEOL–AMX–400 (300MHz) spectrophotometer using T.M.S. as an internal standard.

EXPERIMENTAL:

Step I: Preparation of 5-phenoxy-4-chloro-2-nitroaniline (III)

In a 250 ml round bottom flask, a mixture of Phenol (II) (0.949 g, 0.01 mol) and anhydrous potassium carbonate (1.40 g, 0.01 mol) was refluxed in dimethyl formamide at 140°–160°C for 2 hours. To this reaction mixture, 4,5-dichloro-2-nitroaniline (2.07 g, 0.01 mol) (I) dissolved in dimethyl formamide was added drop wise at same temperature. Reaction was further refluxed for 6-8 hours. After completion of reaction, reaction mixture was brought down to room temperature and then was poured into crushed ice. Product (III) separated was filtered and recrystallized from ethanol.

Step II: Preparation of 5-Phenoxy-4-chloro- Ortho Phenylene Diamine (OPD) [IV]

5-phenoxy-4-chloro-2-nitroaniline (0.01 mol) (III) formed using above method, was further reduced at 60° – 80°C with Raney-Nickel and Hydrazine Hydrate in ethanol for 4–6 hours⁶. After completion of reaction, Raney-Nickel was filtered off and decomposed in 10% dil. HCl. Excess of ethanol was removed by vacuum distillation, and the concentrated solution remaining was poured into crushed ice and the product 5-Phenoxy-4-chloro-ortho phenylene diamine (IV) was filtered and dried.

Step III: Preparation of 5-phenoxy-6-chloro-2-mercapto benzimidazole (V).

Potassium hydroxide (0.01 mol) was dissolved in (1:5) mixture of water and ethanol. The mixture was allowed to cool to room temperature and to this mixture carbon disulfide (0.017 mol) was added with constant stirring and this led to the formation of potassium ethylxanthate solution.

4-phenoxy-5-chloro OPD (0.01 mol) (IV) was taken in 15 ml of ethanol and was refluxed. After half an hour, potassium ethylxanthate solution prepared as above was added drop wise and was further refluxed for 8–10 hours.

The reaction mixture was concentrated under vacuum to remove most of the ethanol. The concentrated solution was poured into crushed ice and neutralized with dil. HCl. The solid product 5-phenoxy-6-chloro-2-mercapto benzimidazole (V) separated was filtered, washed with water, dried and recrystallized from ethanol.

Step IV: Synthesis of 5-phenoxy-6-chloro-2-(4-chloro phenyl thiocarbamoyl) benzimidazole (VII-g)

In a 100 ml three necked dry round bottom flask, was placed a mixture of triethyl amine 0.10 g. (0.003 mol) and a solution of 5-phenoxy-6-chloro-2-mercapto benzimidazole (0.002 mol) (V) in 10 ml dry 1,4-dioxane. The reaction mixture was stirred on a mechanical stirrer at 50° – 60°C for about half an hour. To this, was added dropwise, a solution of 4-chloro benzoyl chloride (0.001 mol) (VI) in 10 ml dry 1,4-dioxane at same temperature. After the addition, reaction mass was stirred for 6 – 8 hours. It was kept overnight at room temperature. The solution was poured into crushed ice. The solid separated out was filtered and washed with water and dried. It was then recrystallized from ethanol.

As per the procedures described above, 16 derivatives [VII-a – VII-p] were synthesized and their physical data have been presented in Table-I.

VII-a: Yield: 61%; M.P.: 168°C

IR (KBr): 875, 850, 805 (5-substituted benzimidazole); 1263 (Ar-O-Ar); 1450, 710 (C-S, sulfide linkage), 1550 (C=N stretching), 1691 (C=O stretching)

¹H NMR (δ, ppm): 6.93 – 7.75 [m, 9H (8H, Ar-H), (1H, -NH)]

VII-b: Yield: 60%; M.P.: 182°C

IR (KBr): 881, 856, 812 (5-substituted benzimidazole), 1279 (Ar-O-Ar); 1458, 707 (C-S, sulfide linkage); 1557 (C=N stretching), 1688 (C=O stretching)

¹H NMR (δ, ppm): 6.85 – 8.08 [m, 16H (15H, Ar-H), (1H, -NH)].

VII-d: Yield: 65%; M.P.: 208– 210°C

IR (KBr): 877, 838, 796 (5-substituted benzimidazole), 1259 (Ar-O-Ar), 1443, 716 (C-S, sulfide linkage), 1555 (C=N stretching), 1676 (C=O stretching), 727 (C-Cl Stretching)

¹H NMR (δ, ppm): 6.81-7.89 [m, 15H (14H, Ar-H), (1H, -NH)],

VII-i: Yield: 61%; M.P.: 159°C

IR (KBr): 909, 847, 813 (5-substituted benzimidazole), 1245 (Ar-O-Ar), 1462, 686 (C-S, sulfide linkage), 1580 (C=N stretching), 1695 (C=O stretching), 712, 730 (C-Cl Stretching).

¹H NMR (δ, ppm): 7.03 – 8.18 [m, 12H (11H, Ar-H); (1H, –NH)]

VII-k: Yield: 62%; M.P.: 166 – 168°C

IR (KBr): 930, 855, 824 (5-substituted benzimidazole), 1258 (Ar-O-Ar), 1465, 690 (C-S, sulfide linkage), 1590 (C=N stretching), 1707 (C=O stretching), 739 (C-Cl Stretching).

¹H NMR (δ, ppm): 6.90 – 8.12 [m, 11H (10H, Ar-H); (1H, –NH)]

VII-m: Yield: 62%; M.P.: 170 – 172°C

IR (KBr): 903, 831, 801 (5-substituted benzimidazole), 1270 (Ar-O-Ar), 1440, 718 (C-S, sulfide linkage), 1567 (C=N stretching), 1681 (C=O stretching), 754, 785 (C-Cl stretching).

¹H NMR (δ, ppm): 6.90 – 8.15 [m, 12 H (11H, Ar-H); (1H, –NH)].

VII-p: Yield: 70%; M.P.: 235 – 237°C

IR (KBr): 914, 826, 789 (5-substituted benzimidazole), 1250 (Ar-O-Ar), 1440, 720 (C-S, sulfide linkage), 1585 (C=N stretching), 1701 (C=O stretching), 770 (C-Cl stretching).

¹H NMR (δ, ppm): 6.77 – 8.19 [m, 13 H (12H, Ar-H); (1H, –NH)].

RESULTS AND DISCUSSION:

In vitro antimicrobial evaluation of all synthesized title compounds [VII-a – VII-p] was conducted against Gram Positive bacteria Staphylococcus aureus (ATCC 3750), Gram Negative bacteria Salmonella typhi (NCTC 786) and a fungal strain i.e. Candida albicans (ATCC 10231). The Minimum Inhibitory Concentration (MIC) was defined as the lowest concentration inhibiting the growth of the organism. The test compounds were subjected to in vitro screening by serial tube dilution technique⁷. The detailed results of antimicrobial activity are mentioned in Table-I

Antibacterial activity:

The synthesized title compounds [VII-a – VII-p] were screened for their antimicrobial potency. Muller-Hinton broth was used as the culture medium. It was observed that naphthoxy substituted benzimidazole with dichloro substitution on the phenyl thiocarbamoyl ring was the most potent amongst them. Phenoxy and naphthoxy substitutions on benzimidazole ring with unsubstituted phenyl thiocarbamoyl portion [VII-a – VII-d] was the least effective amongst them. Hence substitution on the phenyl thiocarbamoyl ring is essential for antibacterial activity.

Schematic Representation of the title compounds prepared:

TABLE 1: Physical Properties & Bioactivity of the Title compounds (VII-a – VII-p)

Compound	R	R₁	R₂	M. P. (⁰C)	Yield (%)	Antimicrobial Activity (μg/ml) Minimum Inhibitory Concentration (MIC)
Compound	R	R₁	R₂	M. P. (⁰C)	Yield (%)	S. aureus ATCC 3750	S. typhi NCTC 786	C. Albicans ATCC 10231
VII-a	H	Phenyl	H	168	61	N.A.	N.A.	N.A.
VII-b	H	2-Naphthyl	H	182	60	200	200	200
VII-c	Cl	Phenyl	H	175	63	100	N.A.	N.A.
VII-d	Cl	2-Naphthyl	H	208-210	65	200	200	200
VII-e	H	Phenyl	4-Chloro	153	60	200	100	200
VII-f	H	2-Naphthyl	4-Chloro	172	66	100	100	100
VII-g	Cl	Phenyl	4-Chloro	169	62	100	100	200
VII-h	Cl	2-Naphthyl	4-Chloro	196-198	65	50	100	100
VII-i	H	Phenyl	2,4-Dichloro	159	61	100	200	200
VII-j	H	2-Naphthyl	2,4-Dichloro	170	69	100	50	50
VII-k	Cl	Phenyl	2,4-Dichloro	166-168	62	100	100	100
VII-l	Cl	2-Naphthyl	2,4-Dichloro	187	68	50	100	50
VII-m	H	Phenyl	3,4-Dichloro	170-172	62	100	200	200
VII-n	H	2-Naphthyl	3,4-Dichloro	189-191	68	50	200	50
VII-o	Cl	Phenyl	3,4-Dichloro	215-217	63	50	200	100
VII-p	Cl	2-Naphthyl	3,4-Dichloro	235-237	70	50	50	50

· Elemental Analysis for each compound was within the range of ± 0.4, · N.A. = Not Active upto 200 µg/ml, · Ampicillin (MIC 0.04 μg/ml) used as standard against S. aureus, · Trimethoprim (MIC 0.01 μg/ml) used as standard against S. typhi, · Miconazole (MIC 0.01 μg/ml) as standard against C. albicans.

Antifungal activity:

Similarly, the title compounds were also screened for their antifungal activity against a fungal strain i.e. Candida Albicans. Here, Sabouraud-Dextrose was used as a culture medium. 5-phenoxy benzimidazoles with unsubstituted phenyl thiocarbamoyl moiety [IV-a, IV-c] was inactive against Candida Albicans. Naphthoxy benzimidazole derivatives with dichloro substitution showed better activity i.e. MIC up to 50 µg/ml than the rest of the synthesized compounds.

Overall, naphthoxy derivatives of benzimidazole with dichloro substitution were comparatively more effective against all the three strains, where dichloro substitution at 3,4 position [VII-p] was the most effective.

ACKNOWLEDGEMENT:

The authors wish to thank The Institute of Science, Mumbai for recording spectral analysis and Sophisticated Analytical Instrument Facility (SAIF), IIT Bombay for carrying out the Elemental analysis.

REFERENCES:

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3. Yadav A G, Patil V N et al, Asian Journal of Research in Chemistry, 2(4), 2009, Accepted & In Press.

4. Adams; Dewitt, C; Schlatter; Rudolph; Ger. Offen; 1,956,157 (1970); Cited from, Chem. Abstr. 73, 1970, 3536m.

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Received on 01.12.2009 Modified on 28.01.2010

Asian J. Research Chem. 3(2): April- June 2010; Page 342-345

S. aureus

S. typhi

C. Albicans

ACKNOWLEDGEMENT:

REFERENCES: