RJPS Vol No: 14 Issue No: 1 eISSN: pISSN:2249-2208
BP Nandeshwarappa 1, SK Manjunatha*2, DK Ramesh 2, M Suchitra2, SO Sadashiv3
1Department of studies in Chemistry, Davangere University, Davangere
2Department of Pharmaceutical Chemistry, Bapuji Pharmacy College, Davangere
3Department of Food Technology, Davangere University, Davangere
Corresponding author:
Dr. Manjunatha S. Katagi, Department of Pharmaceutical Chemistry, Bapuji Pharmacy College Davangere-577004, Karnataka, India, E-mail: manju_mpharm@rediffmail.com
Received Date: 08/11/2020 Accepted Date : 09/12/2020
Abstract
Background: The design of new agents, active against resistant organism is of critical importance. The current antimicrobial chemotherapy still suffers from two major limitations. The first is the lack of selectivity of conventional antimicrobial agents, which in turn brings about unwanted side effects. The second is acquisitions by the microorganism of multi drug resistance.
Objective: To synthesize a series of quinoline-2-one linked thiadiazole derivatives and to asses in vitro antimicrobial activity.
Method: The derivatives of quinoline-2-one linked thiadiazole were synthesized, purified and characterized on the basis of physiochemical and spectral studies. The synthesized compounds were subjected for in vitro antimicrobial activity by cup-plate agar diffusion.
Results: Among the eight synthesized novel compounds, three compounds 4b, 4c and 4f shows promising antibacterial activity as compared to Ciprofloxacin (100 µg/ml), however most of them 4b, 4c, 4e and 4f showed potent antifungal activity as compared to Fluconazole (100 µg/ml).
Conclusion: Despite continued efforts to discover improved antimicrobial agents, there has been little success towards discover of antimicrobial agents. The compounds having fluoro substitution at 4th & 2,4-dichloro positions showed satisfactory antibacterial and antifungal activity.
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Introduction
The knowledge in the synthesis of higher nitrogen containing heterocyclic system has evolved greater interest just because of their enormous usage in chemotherapy1. Although considerable advances have been achieved over recent decades in the field of research and development of novel structural prototypes as effective antimicrobials, current antimicrobial chemotherapy still suffers from two major limitations. The first is the lack of selectivity of conventional antimicrobial agents, which in turn brings about unwanted side effects. The second is acquisitions by the microorganism of multi drug resistance. The design of new agents, active against resistant organism is of critical importance. In the field of quinolone antimicrobial agents, the new generation of quinolone has achieved significant improvements in terms of potency, spectrum and pharmacokinetic properties. But these agents faced a rapid increase of resistance from gram-positive organisms. Therefore, enhancing the potency of quinolone especially against gram-positive organism has become most urgent2.
As there is increased development of resistance to antimicrobial drugs, there is an urgent need of finding out a new and broad spectrum antimicrobial agents worldwide. Heterocyclic compounds containing nitrogen, is an indispensable structural unit for both the chemist and the biochemist. Among the antimicrobial agents discovered in recent years the various 2-quinolone as antimicrobial agent has stimulated remarkable interest in the synthesis of 2-quinolones bearing heterocycles3. Numerous biological activities of 2-quinolone have been described; antimicrobial4-6, antioxidant and anti-inflammatory7, antitumor8, Fornesyl transferase inhibitor9, antiangiogenic10, acetylcholinesterase reactivators11-14, and anti-tuberculosis15-17.
In spite of such stimulating properties it was contemplated to synthesize some newer congeners of 2-quinolone linked thiadiazole with a need to explore their potency as better antibacterial and antifungal activity. The 4-hydroxy-6-methyl/ phenyl-2H-pyrano [3,2-c]quinoline-2,5(6H)-diones were synthesized according to literature procedure and subjected to hydrolysis to yield 1a and 1b6, further 1a and 1b subjected to bromination to yield 3-bromoacetyl-4-hydroxy-1-methyl/phenyl quinolin-2(1H)-one 2a and 2b18. The substituted thiadiazole (3a-3d) were synthesized according to literature procedure19. The synthesized compounds 4a-4d and 4e-4h were accomplished by condensing with different substituted thiadiazole. The compounds thus obtained were characterized by physiochemical and spectral data. The physicochemical properties of title compounds were reported in Table 1.
Experimental Procedure
Merck, S.D Fine-Chem. Limited, Mumbai supplied all the chemicals and solvents . All the obtained solvents and chemicals were purified by either distillation or recrystallization before use. Veego (VMP-MP) melting point apparatus were used to determine the melting point and are uncorrected. The IR spectra of the newly synthesized compounds were recorded using KBr on Jasco FTIR spectrometer (model-4100). The 1H NMR of the compounds synthesized was recorded on Bruker avance II 400 NMR spectrometer (with TMS as internal references) at Sophisticated Analytical and Instrumentation Facility (SAIF), Punjab University (Chandigarh). The mass spectra (LCMS) were recorded on a MAT 120 in SAIF, Punjab University, Chandigarh. (India)
General procedure for synthesis of 3-(bromoacetyl) -4-hydroxy-1-methyl (2a)/phenyl (2b) quinolin-2 (1H)-one.
In 50ml glacial acetic acid and gently heated at 80°C with continuous stirring for 30 min. The equimolar quantity of bromine (0.065 mol) in glacial acetic acid (10 ml) was slowly added in a drop wise manner for a period of 1 h and heating of the solution was continued until a slight change in the color of the solution was observed. The obtained mixture was cooled to room temperature. The products (2a and 2b) were obtained as yellow crystalline solid and recrystallized from ethanol.
3-(Bromoacetyl)-4-hydroxy-1-methylquinolin-2 (1H)-one (2a).
The compound 2a was prepared and purified as per the above mentioned procedure: yield 10.26 g (73%), mp 174-176 0C, IR (KBr, v, cm-1): 756.14 cm-1 and 1190.18 (CH2 -Br), 1615.47 cm-1 (-C=O amide), 3046.24cm-1 (aromatic -C-H stre). 1 H NMR (400MHz, CDCl3 ) δ (ppm): δ 3.78 (s, 3H, -NCH3 ), 4.98 (s, 2H, -CH2 -Br), 7.72 – 8.35 (m, 4H, Ar-H), 15.82 (s, 1H, -OH).
3-(Bromoacetyl)-4-hydroxy-1-phenylquinolin-2 (1H)-one (2b)
The compound 2b was prepared and purified as per the above mentioned procedure: yield 12.24 g (67.43%), mp 192-194 0, IR (KBr, v, cm-1): 786.11 cm-1 and 1210.06 (CH2-Br), 1620.31 cm-1 (-C=O amide), 3058.11cm-1 (aromatic -C-H stre). 1H NMR (400MHz, CDCl3) δ (ppm): δ 5.23 (s, 2H, CH2-Br), 7.65 – 8.85 (m, 9H, Ar-H), 15.91 (s, 1H, -OH).
General procedure for synthesis of 3-(2-(substituted 1,3,4-thiadiazol-2-ylamino)acetyl)-4-hydroxy-1- methyl (4a-4d)/phenyl(4e-4h)quinolin-2(1H)-ones.
A mixture of an appropriately substituted thiadiazole (one of 3a-3d, 0.004 mol) and 3-(2-bromoacetyl)-4-hydroxy-1-methylphenylquin olin-2(1H)-one (2a/2b, 0.004 mol) was dissolved in 45 mL of glacial acetic acid with constant stirring. The obtained mixture was then heated at reflux temperature for about 6 to13 h. The progress of the reaction was monitored by the TLC. The obtained mixture was allowed to attain the room temperature. The solvent was removed under vacuum. The ethyl acetate was used to dissolve the obtained residue, washed twice with water, and dried over Na2SO4. Further, the final compounds were purified by removal of the solvent followed by column chromatography using 200-400 mesh silica gel eluting with methanol (12%) in dichloromethane.
3-(2-(5-Phenyl-1,3,4-thiadiazol-2-ylamino)acetyl) -4-hydroxy-1-methylquinolin-2(1H)-one (4a).
IR (KBr, v, cm-1): 1620.9 cm-1 (-COCH2 stre), 1740.5 cm-1 (-C=O amide), 2968.4 cm-1 (Ar-CH stre), 3121.8 cm-1 (-NH), 3393.8 cm-1 (-OH); 1 H NMR (400MHz, DMSO-d6) δ (ppm): 2.21 (s, 3H, N-CH3), 6.28 (s, 1H, -NH), 6.54 (s, 2H, COCH2), 6.80–8.65 (m, 9H, Ar-H), 12.84 (s, 1H, -OH); LCMS: C20H16N4O3S (M+) m/z 392.24; calcd. 392.09.
3-(2-(5-(4-Fluorophenyl)-1,3,4-thiadiazol-2-ylamino) acetyl)-4-hydroxy-1-methylquinolin-2(1H)-one (4b).
IR (KBr, v, cm-1): 1627.2 cm-1 (-COCH2 stre), 1738.9 cm-1 (-C=O amide), 2943.7 cm-1 (Ar-CH stre), 3101.0 cm-1 (-NH), 3363.0 cm-1 (-OH); 1H NMR (400MHz, DMSO-d6) δ (ppm): 2.15 (s, 3H, N- CH3), 6.21 (s, 1H, NH), 6.38 (s, 2H, COCH2), 6.90– 8.51 (m, 8H, Ar-H), 12.31 (s, 1H, -OH); LCMS: C20H15FN4O3S (M+) m/z 410.42; calcd. 410.08.
3-(2-(5-(2,4-Dichlorophenyl)-1,3,4-thiadiazol-2-ylamino) acetyl)-4-hydroxy-1-methylquinolin-2(1H)-one (4c).
IR (KBr, v, cm-1): 1640.4 cm-1 (-COCH2 stre), 1704.2 cm-1 (-C=O amide), 2988.7 cm-1 (Ar-CH stre), 3124.0 cm-1 (-NH), 3394.5 cm-1 (-OH); 1H NMR (400MHz, DMSO-d6) δ (ppm): 2.48 (s, 3H, N-CH3), 5.98 (s, 1H, -NH), 6.18 (s, 2H, COCH2), 7.21–8.64 (m, 7H, Ar-H), 12.85 (s, 1H, -OH); LCMS: C20H14Cl2N4O3S (M+) m/z 460.54; calcd. 460.02.
3-(2-(5-Phenyl-1,3,4-thiadiazol-2-ylamino)acetyl) -4-hydroxy-1-phenylquinolin-2(1H)-one (4e).
IR (KBr, v, cm-1): 1628.2 cm-1 (-COCH2 stre), 1680.2 cm-1 (-C=O amide), 3012.7 cm-1 (Ar-CH stre), 3145.5 cm-1 (-NH), 3376.2 cm-1 (-OH); 1H NMR (400MHz, DMSO-d6) δ (ppm): 6.02 (s, 1H, -NH), 6.32 (s, 2H, COCH2), 6.92–8.74 (m, 14H, Ar-H), 13.15 (s, 1H, -OH); LCMS: C25H18N4O3S (M+) m/z 454.54; calcd. 454.11.
3-(2-(5-(4-Fluorophenyl)-1,3,4-thiadiazol-2-ylamino) acetyl)-4-hydroxy-1-phenylquinolin-2 (1H)-one (4f).
IR (KBr, v, cm-1): 1634.4 cm-1 (-COCH2 stre), 1694.4 cm-1 (-C=O amide), 3022.4 cm-1 (Ar-CH stre), 3112.5 cm-1 (-NH), 3354.8 cm-1 (-OH); 1 H NMR (400MHz, DMSO-d6) δ (ppm): 5.78 (s, 1H, -NH), 6.12 (s, 2H, COCH2), 6.88–8.59 (m, 13H, Ar-H), 13.24 (s, 1H, OH); LCMS: C25H17FN4O3S (M+) m/z; 472.48 calcd. 472.10.
3-(2-(5-(2,4-dichlorophenyl)-1,3,4-thiadiazol-2-ylamino) acetyl)-4-hydroxy-1-phenylquinolin-2(1H)-one (4g).
IR (KBr, v, cm-1): 1645.4 cm-1 (-COCH2 stre), 1692.4 cm-1 (-C=O amide), 3029.2 cm-1 (Ar-CH stre), 3124.5 cm-1 (-NH), 3388.9 cm-1 (-OH); 1 H NMR (400MHz, DMSO-d6) δ (ppm): 5.84 (s, 1H, -NH), 6.02 (s, 2H, COCH2), 6.88–8.59 (m, 12H, Ar-H), 13.33 (s, 1H, OH).
Antimicrobial Screening20-23
All the synthesized compounds 4a-4h have been screened in vitro for their antibacterial activity against gram-negative bacteria Escherichia coli, Pseudomonas aeruginosa and gram-positive bacteria Staphylococcus aureus, Bacillus substilis, while antifungal activity against Candida albicans and Asparagillus niger at 100 µg/ml concentration by cup-plate agar diffusion method using dimethylsulfoxide as solvent. After 24h and 48h of incubation at 370 ±1, the antibacterial and antifungal activity respectively was determined by measuring the zones of inhibition in mm. Standard antibacterial ciprofloxacin and fungicide fluconazole were used under similar condition for comparison. Control test with solvent were performed for every assay but showed no inhibition of microbial growth. The values of antibacterial and antifungal activity were depicted in Table 2.
Results
4-Hydroxy-6-methyl/phenyl-2H-pyrano[3,2-c]quin oline-2,5(6H)-dione, were synthesized according to literature procedure6 and subjected to hydrolysis to yield 1a and 1b, further 1a and 1b subjected to bromination to yield 3-bromoacetyl-4- hydroxy-1-methyl/phenyl quinolin-2(1H)-one 2a and 2b18. The synthesis of title compounds 4a-4h was accomplished by condensing with different substituted thiadiazole. The newly synthesized compounds structures were confirmed by their spectral data. The spectral data of 4b exhibit IR band at 1627.2 cm-1 and 1738.9 cm-1 was due to amide carbonyl and acetyl groups. Another peak at 2943.7 cm-1 mainly because of aromatic –C-H stretching. Further, a peak at 3101.0 cm-1 mainly because of –N-H stretching, and 3363.0 cm-1 of hydroxyl group indicates the completion of the reaction. Their structure was further confirmed by their 1 H NMR spectral data that exhibited three protons of N-methyl signal found at δ value 2.15 ppm, whereas aromatic protons signal showed the δ value 6.90-8.51 ppm. The appearance of a signal at δ value 12.31 ppm indicates one proton of hydroxyl (-OH) group and δ value 6.21 ppm indicates one proton of amine (-NH) group. The mass spectra of the compound 4b (m+) found 410.42, which was one more proof for the confirmation of the title compound. The physicochemical data of the synthesized compounds are listed in Table 1. Some of these compounds have shown good antibacterial and potent antifungal agents.
Sa=Staphylococcus aureus, Bs= Bacillus substilis, Ec= Escherichia coli, Pa= Pseudomonas aeruginosa, Ca= Candida albicans, An= Asparagillus niger, NA= No activity, ND= Not determined.
Discussion
The synthesized compounds of 3-(2-(substituted 1,3,4-thiadiazol-2-ylamino)acetyl)-4-hydroxy-1- methyl (4a-4d)/phenyl(4e-4h)quinolin-2(1H)-ones were screened for their in vitro antibacterial activity against gram positive bacteria (Staphylococcus aureus, Bacillus substilis) and gram negative bacteria (Escherichia coli, Pseudomonas aeruginosa) and for antifungal activity (Candida albicans, Asparagus niger) by measuring the zone of inhibition at concentration 100 µg/mL (mm). Compounds 4b (20, 23, 25, 27mm), 4c (21, 25, 32, 30mm) and 4f (20, 23, 25, 27mm) showed promising antibacterial activity as compared with standard Ciprofloxacin (23, 26, 34, 31mm) and compounds 4b (31, 34mm), 4c (27, 29mm), 4e (28, 33mm), and 4f (31, 34mm) showed good antifungal activity against standard Fluconazole (33, 36mm).
Conclusion
4-Hydroxy-1-methyl/phenyl-3-(substituted thiadiazole)quinolin-2(1H)-one compounds (4a-4h) were successfully synthesized and characterized on the basis of physicochemical and spectral studies. Further evaluated for their antimicrobial activity against Staphylococcus aureus, Bacillus substilis, Escherichia coli, Pseudomonas aeruginosa, Candida albicans and Asparagillus niger. Among synthesized novel compounds 4b, 4c, 4f shows promising antibacterial activity as compared to Ciprofloxacin (100 µg/ml), however most of them 4b, 4c, 4e and 4f showed potent antifungal activity as compared to Fluconazole (100 µg/ml).
Suitable molecular modification of these compounds may generate potent antimicrobial agents in future.
Acknowledgement
The authors are thankful to Principal, Bapuji Pharmacy College, Davangere, for providing necessary facilities to carry out this research work. We are also thankful to Shree Dhanvantary Pharmaceutical Analysis and Research Centre (SDPARC), Kim (Surat), and Director, RSIC, Punjab University, Chandigarh, India, for providing analytical data.
Supporting Files
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