Synthesis, characterization and biological evaluation of novel tetrasubsituted Imidazole compounds

: New classes of tetrasubsituted imidazole based compounds were synthesized using multicomponent one pot synthesis scheme through cyclocondensation reaction of benzil, aromatic primary amines, aldehydes and ammonium acetate in glacial acetic acid. The synthesized compounds have been analyzed and characterized by melting point, color, conductivity method, CHN analysis, FT-IR and UV-Visible. The reaction proceeding was examined by TLC after regular intervals of period. To test biological activity, the synthesized compounds have been examined against various bacterial strains. From the analysis of the antibacterial activity of these synthesized compounds demonstrated that all three imidazole compounds have considerable to significant activity against the strains, and compound K2 was found potent comparatively. not-for-profit


Color / Melting point / Physical appearance / yield:
The newly prepared tetrasubsituted imidazole derivatives (K1, K2 and K3) inert against climate and humidity at room temperature.. They exist in crystalline form and have color differentiates. Color, physical appearance and melting points of synthesized compounds are shown in table 1.

Conductance Values:
The conductivity of synthesized compounds was determined at room temperature. About 1 M solution of synthesized compounds was prepared using DMSO as a solvent to check their conductance. The conductance values of synthesized compounds were low as they are organic compounds of covalent nature and nonelectrolyte having conductivity range from 12 to 15 Ω -1 cm -2 mol -1 as shown in table 2.

UV-Visible study
The λmax of all synthesized compounds was determined experimentally in the solvent phase. The experimentally determined values are tabulated in table 3. It was observed that compound K3 has highest λmax while K1 possess lowest one.

IR Spectra of 5-methyl-2-(2-methyl-4,5-diphenyl-1Himidazol-1-yl)phenol (K1)
Selected IR values of K1 are given in Table 4. The IR spectra of K1 exhibited that a new peak is revealed at 1650 cm -1 which may be due to presence of stretching frequency of C=N . This indicates that the probable imino bond (C=N) might be shaped due to cyclocondensation of benzil, primary amine, aldehyde and ammonium acetate. The C-N peak occurred at 1440 cm-1, which also suggests the development of the required compound. Peak appeared at 3345 cm -1 due to O-H stretching frequency. New peaks observed at 3050 cm -1 and 2912 cm -1 due to sp 2 (C-H) and sp 3 (C-H) stretching frequencies. Peak at 1680 cm -1 was vanished that directs the absence of benzil. However, the remaining peaks of different groups did not undergo significant change.

IR spectra of 2-[2-(furan-2-yl)-4,5-diphenyl-1Himidazol-1-yl]-5-methylphenol (k2)
The few selected IR values of K2 are given in Table 4. Data revealed that the bands due to the azomethine (C=N) linkage observe at 1658 cm -1 . The peak of C-N occurred in the range of 1419 cm-1. The peak due to formation of C=N bond showed that our required product might be formed by condensation of benzil with p-cresolamine and furfural. Peak at 1575 cm -1 might be due to (C=C) of aromatic imidazole ring. Peak due to sp 2 (C-H) also appeared at (3058 cm -1 ). The band due to OH stretching frequency appeared at 3315 cm -1 . Peak due to C-O stretching frequency also observed at 1325 cm -1 . However, the remaining peaks of different groups did not change any more.

IR spectra of 5-methyl-2-(2,4,5-triphenyl-1H-imidazole-1-yl)phenol (K3)
Some selected IR values of K3 are given in Table 4. Peak at 1656 cm -1 indicates the formation of C=N linkage present in imidazole ring. This shows that imino bond which is required to establish might be formed by condensation of benzil with pcresolamine and benzaldehyde. Peak at 1522 cm -1 shows the presence of C=C which may be of aromatic imidazole ring. Peak of sp 2 (C-H) appeared at 3065cm -1 . Another new peak also appeared at 1449 cm -1 which may be due to C-N stretching frequency. This also indicates the formation of required product. Peaks at 3305 cm -1 due to OH stretching frequency and at 2908 cm -1 due to sp 3 (C-H) are the indications of desired product development. Peak at 1680 cm -1 due to benzil carbonyl was missing that shows the complete condensation reaction of benzil with other reagents. All other peaks were not changed significantly.

Antibacterial Activity (in-vitro)
The biological activity of all three synthesized compounds (K1, K2, K3) was checked against one Gram negative (Escherichia coli) and two Gram positive (Bacillus subtilis, Staphylococcus aureus) following published literature [31]. The results obtained by compounds were compared with a standard drug (Ciprofloxacin). Synthesized compounds had major action toward bacterial species. Compound K2 has been found to have the greatest efficacy against certain strains of bacteria. On the other hand, lowest activity was shown by compound K1. The compounds K2 and K3 presented promising results (>20mm) against all bacterial strains. The compound K2 demonstrated maximum activity among all three compounds (23mm) against Bacillus subtilis (Table 5).

III. CONCLUSION
In present work, we synthesized three novel tetrasubsituted imidazole compounds using multicomponent one pot synthesis scheme. To elucidate the structures of synthesized compounds, characterization was performed by spectroscopic techniques (FT-IR and UV-Visible). Chemically modified compounds have been checked against different bacterial strains to verify biological activity (Staphylococcus aureus, Bacillus subtilis, Escherichia coli). The present study demonstrated that the novel synthesized compounds possess reasonable activity against bacterial strains, worth considering for further studies.

MATERIALS & METHODS
All chemicals used in this work were of laboratory quality (analytical) obtained from Sigma Aldrich by chemical suppliers. The purchased chemicals were used in the synthesis without any further refinement and action. The solvents used in the research were also purchased from chemicals suppliers and before use they were distilled. The reactions were monitored using thin layer chromatography proceedings and chromatographic plates were irradiated in U.V and then assessed in iodine vapors. In order to check the percentage composition of newly synthesized compounds, elemental analysis by EL III CHNOS elemental analyzer (Elementar, Hanau, Germany) was accomplished.

Instrumentation
For the purpose of both heating and stirring the reaction materials, magnetic stirrer hot plate with stirring range 50 to 1200rpm and heating range (60-200 °C) was applied. To weigh the materials, AX200, Shimadzu, Japan modal was used. The melting point of newly synthesized compounds was determined by using melting point apparatus (Gallen Kamp). Agilent technology (Cary-620) FTRIR spectrophotometer was used to check the presence of desired functional groups of synthesized compounds by taking IR Spectra. SHIMADZU UV 240 spectrophotometer was used to obtain the spectrum of U.V/Visible technique. To measure the conductivity of synthesized compounds, SDT-600 conductivity meter was used. Elemental analysis was performed by using EL III CHNOS elemental analyzer (Elementar, Hanau, Germany) to check the percentage composition of elements in synthesized compounds.

General method for the synthesis of compounds; K1, K2 and K3
Tetrasubsituted imidazole derivatives (K1, K2 and K3) were synthesized according to reported protocol with small variations.

Synthesis of 5-methyl-2-(2-methyl-4,5-diphenyl-1Himidazol-1-yl)phenol (K1)
Benzil (1.05 g, 0.005 mmol) and acetaldehyde (0.22 g, 0.005 mmol) were dissolved in glacial acetic acid at room temperature. After that, p-cresolamine (0.62 g, 0.005 mmol) and ammonium acetate (0.38 g, 0.005mmol) were added to reaction mixture. The reaction was with a reflux the mixture at 110 °C for 12 hours. TLC was used to monitor the reaction progress. After 12 hours, the volume of the mixture was reduced to half by heating. For slow evaporation, the mixture was kept in a beaker. Crystals of K1 were obtained within a week ( Figure 5).

Synthesis of 2-[2-(furan-2-yl)-4,5-diphenyl-1Himidazol-1-yl]-5-methylphenol (K2)
Benzil (1.05 g, 0.005 mmol) and furfural (0.36ml, 0.005 mmol) were dissolved in glacial acetic acid at room temperature. After that, p-cresolamine (0.62 g, 0.005 mmol) and ammonium acetate (0.38 g, 0.005mmol) were added to reaction mixture. The reaction was by the refluxing of the mixture at 110 °C for 12 hours. TLC was used to monitor the reaction progress. After 12 hours, the volume of the mixture was reduced to half by heating. For slow evaporation, the mixture was kept in a beaker. Crystals of K2 were obtained within a week ( Figure 6). Crystals purification was performed by first washing with ethyl acetate and then with ethyl alcohol.

Synthesis of 5-methyl-2-(2, 4, 5-triphenyl-1H-imidazole-1-yl) phenol (K3)
Benzil (1.05 g, 0.005 mmol) and benzaldehyde (0.53 g, 0.005 mmol) were dissolved in glacial acetic acid at room temperature. After that, p-cresolamine (0.62 g, 0.005 mmol) and ammonium acetate (0.38 g, 0.005mmol) were added to reaction mixture. The reaction was done by refluxing the mixture. at 110 °C for 12 hours. TLC was used to monitor the reaction progress. After 12 hours, the volume of the mixture was reduced to half by heating. For slow evaporation, the mixture was kept in a beaker. Crystals of K3 were obtained within a week (Figure 7). Crystals purification was performed by first washing with ethyl acetate and then with ethyl alcohol. The yield of desired compound was about 70 % and that of melting point was 206-208 °C. Elemental analysis: Calculated for C 28