Synthesis , Characterization and Antimicrobial Activity of Imidazole Derivatives Based on 2-chloro-7-methyl-3-formylquinoline

A series of oxazole and thereof imidazole derivatives were prepared from 2-chloro-7-methyl-3-formyl quinoline. The structures of all synthesized compounds were elucidated by elemental, IR, HNMR, CNMR spectra. Supplementary to these, they were assayed in vitro for their antimicrobial activity; it was revealed that some synthesized derivatives were exhibiting competent biological activity against both gram negative & gram positive bacterial species and fungal microorganisms.


Introduction
Heterocyclic compounds of nitrogen containing five membered ring systems have been described for their biological activity against various micro organisms. 1,2Besides this, the chemistry of quinoline and imidazoles have also been reviewed in literature.2][3][4][5] Considerable interest has been created in the chemistry of quinoline derivatives due to their versatile therapeutic activities like bactericidal, antihistaminic, antimalarial, antidepressant, analgesic, anti-ulcer, antiviral, herbicidal, antitumor, anti-allergic, anticonvulsant, anti-inflammatory etc. [6][7][8][9] Almost every class of imidazole derivatives has been used for different reactions to produce enormous number of heterocycles.1][12][13][14][15] The emergence of powerful and elegant imidazole has stimulated major advances in chemotherapeutic agents of remarkable significance in medicine, biology and pharmacy.Besides this, it is also reported [15][16] that imidazole compounds are one of the effective antifungal agents.Considering the importance of both moieties Quinoline and Imidazole, extending our previous work 17,18 we planned to synthesize imidazole derivatives from 2-chloro-7-methyl-3-formylquinoline.The whole synthesis route is shown in scheme 1.

Material and Methods
Acetanilide and their derivatives were purified by crystallization in R-spirit.DMF and phosphorous oxychloride used were of analytical reagent grade.All of the organic solvents and Hippuric acid, acetic anhydride, sodium acetate used were of analytical reagent grade.Eight diamines were used after recrystallization.The 2-chloro-7-methyl-3-formyl quinoline was synthesized by Vilsmeier-Haack reaction by the procedure reported in the literature. 16,19elting points were measured in an open capillary tube and are uncorrected.Elemental analysis was obtained using Perkin Elmer (USA) 2400, series II CHN-analyser.In addition to this, the nitrogen content in all the imidazoles was estimated by Kjeldhal's method. 20IR spectra were recorded on a NICOLET-400 D spectrophotometer, 1 H NMR spectra in CDCl 3 /DMSO-d 6 at 400 MHz on a FT-NMR, R-1500 spectrometer (chemical shift in δ ppm) relative to TMS as an internal standard.Reactions were monitored by TLC, using silica gel as an adsorbent and ethyl acetate-hexane in different ratios as eluent.
Dimethyl formamide (9.6 ml, 0.125 M) at 0 0 C was taken in a three necked flask equipped with a drying tube and phosphoryl oxychloride (32.2 ml, 0.35 M) was added drop wise under continuous stirring.To this solution, 3-methyl acetanilide (0.05 M) was slowly added with continuous stirring.After five minutes, solution was heated under reflux for 1 hour at 80-90 0 C. The reaction mixture was poured into ice water (300 ml) and stirred for further 30 mins at 0-10 0 C. The 2-chloro-7-methyl-3-formylquinoline so obtained, was filtered and washed with water.It was crystallized using R-spirit.The yield was 82%.
Hippuric acid 2-chloro-7-methyl-3-formyl quinoline IMDT Antimicrobial assay: The antimicrobial activities were determined using agar-cup method by measuring the zone of inhibition in mm.All newly synthesized compounds were screened in vitro for their antibacterial activity against Gram positive species (Bacillus subtilis, Bacillus megaterium) and Gram negative species (Escherichia coli, Pseudomonas aeruginosa), while antifungal activity was tested against Aspergillus niger and C. albicans at concentration of 75 μg/ml.Streptomycin was used as a standard drug for antibacterial screening, while Imidil was used as a standard drug for antifungal screening and solvent DMSO was used as a control.Each experiment was made in triplicate and the average reading was taken.The results are summarized in Table-3.

Synthesis, characterization and antimicrobial activity of imidazole derivatives based on 2-chloro-7-methyl-3-formylquinoline
The test was performed by using the agar cup borer method, with some modifications using Streptomycin and Imidil as reference for bacterial and fungal culture respectively. 27A test tube containing sterile melted top agar (1.5 %) previously cooled at room temperature with 0.2 ml suspension of the test culture, mixed methodically and poured in the petri dish containing sterile base agar medium (autoclaved at 121°C for 15 min.)then allowed it to solidify.The cup borer was sterilized by dipping into absolute ethanol and flaming it then allowed to cool it.With the help of sterile cup-borer, three cups in the agar-plate were marked and were injected with 0.1 ml of test solution, 0.1 ml of standard solution and 0.1 ml of DMSO solvent respectively.Then the plates were allowed to diffuse for 20 min. in refrigerator at 4-5°C.The plates were then incubated in upright position at 37°C for 24 hrs.After incubation, the relative susceptibility of the micro-organisms to the potential antimicrobial agent is demonstrated by a clear zone of growth inhibition around the cup.The inhibition zone caused by the various compounds on the micro-organisms was measured and the activity was rated on the basis of the size of the inhibition zone.

Results and Discussion
The elemental analysis of the prepared compounds is given in Table-1  In all the imidazole derivatives vinylic proton is seen around 6 ppm (δ).The aromatic protons are assigned to resonances in the range of (δ) 7.00 to 8.2 ppm.The resonance due to -NH 2 moiety is attributed to the peak in the range of 6.5 to 6.8 ppm.The resonance due to -CH 3 is observed at 2-2.2 ppm.The compounds containing 4,4'-diamino diphenyl methane has a >CH 2 moiety attached to benzene ring and this >CH 2 is highly deshielded.This is reflected in the proton NMR signal of >CH 2 group seen at 3.69 ppm.The 13 C NMR peaks are quite interesting in all these imidazoles derivatives where the peak around 165 ppm is attributed to C of >C=O (Table -2).In all the compounds the peak at 158 ppm is assigned to Cl-C=N moiety.The peak at 148 ppm is likely due to >C=N moiety.The peaks in the region 110-130 ppm are attributed to aromatic ring.The compounds containing 4,4'-diamino diphenyl methane shows a peak at 40 ppm which is due to =CH 2 group attached to both the rings.Practically in all the compounds -NH 2 asymmetric stretching vibration is assigned to a peak around 3400 cm -1 , while a peak around 3250 cm -1 is attributed to -NH 2 symmetric stretching vibration.The =CH stretching vibration in the vinyl moiety is attributed to the absorption at ~3040 cm -1 .The aromatic C-H stretching frequency, as expected is observed at around ~3010 cm -1 .The strong absorption at ~1700 cm -1 is found to be present in majority of the compounds.The absorption will have contributions from stretching of >C=O and >C=N.The strong absorption at 1650 cm -1 have contributions from υ C=N, υ C=C and bending of -NH 2 .In most of the compounds the C-C stretching of the aromatic ring is around 1540 cm -1 .
A fairly strong absorption at ~1300 cm -1 is assigned to C-N stretching.The strong absorption in the region 810-840 cm -1 is due to C-H out of plane bending in aromatic ring.The C-Cl stretching is attributed to the strong absorption in the region 740-720 cm -1 .Compounds containing O=S=O moiety show strong absorption in the region of 1050-1200 cm -1 is due to O=S=O stretching.The C-H bending in the vinyl moiety is seen as a strong band around 800 cm -1 in all the compounds.The compounds containing -CH 3 group shows peaks due to asymmetric and symmetric bending of -CH 3 group at 1475 and 1375 cm -1 respectively and absorption at ~550 cm -1 in the bromo compounds is assigned to C-Br stretching.
The synthesized compounds were screened 'in vitro' for antimicrobial activity.From the data presented in Table-3, it is clear that out of 8 imidazole compounds IMMD, IMBD, IMDM exhibited moderate inhibition against gram negative bacterial species and especially against Escherichia coli while IMBD, IMDM and IMOTD showed maximum activity against most gram negative organisms.Against gram positive organisms almost all compound of the series exhibited maximum inhibition, especially IMPD and IMBD showed highest inhibition against B. megaterium, while IMMD and IMDT showed good inhibition against fungal organism especially C. albicans.The other compounds exhibited moderate to less inhibition against fungal species, but IMMD showed good inhibition.