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Design, synthesis and biological evaluation of some novel isoniazid cyclocondensed azetidinones
d r u g i n v e n t i o n t o d a y 5 ( 2 0 1 3 ) 1 0 0 e1 0 4
Available online at www.sciencedirect.com
journal homepage: www.elsevier.com/locate/dit
Original Article
Design, synthesis and biological evaluation of some novel isoniazid cyclocondensed azetidinones Karthikeyan Elumalai a,c,*, Mohammed Ashraf Ali a, Manogaran Elumalai b, Kalpana Eluri b, Sivaneswari Srinivasan d, Sujit Kumar Mohanti c, Anil Thota c a
New Drug Discovery Research, Department of Medicinal Chemistry, Sunrise University, Alwar, Rajasthan 301030, India b Faculty of Pharmaceutical Sciences, UCSI University, Cheras, Kuala Lumpur 56000, Malaysia c Department of Pharmaceutical Chemistry, Jayamukhi Institute of Pharmaceutical Sciences, Warangal 506 332, India d Department of Pharmaceutics, Jayamukhi College of Pharmacy, Warangal 506 332, India
article info
abstract
Article history:
Background: In the present study, a series of novel azetidinone derivatives synthesized
Received 20 May 2013
because of its potent antimicrobial and antimycobacterial activity.
Accepted 25 May 2013
Method: Compounds (10ae10j) were synthesized by reacting Schiff’s base of isoniazid reacted with, chloro acetyl chloride in presence of triethyl amine and 1, 4-dioxane as an efficient catalyst, analyzed for their structures. in vitro antimicrobial and antimycobacterial
Keywords:
activity were carried out.
Azetidinones
Results: Among the synthesized compounds, compound 10b and 10i was found to be the
Antimycobacterial
most potent against gram-positive bacteria Bacillus subtilis, gram-negative bacteria Escher-
Isoniazid cyclocondensed
ichia coli, Mycobacterium tuberculosis CIP and M. tuberculosis H37Rv.
Mycobacterium tuberculosis CIP and
Conclusion: A series of novel azetidinone derivatives of biological interest were synthesized
H37RV strain
and analyzed, suggests that it an interesting compound compared to the current therapeutic agents and are considered to investigate further for the same. Copyright ª 2013, JPR Solutions; Published by Reed Elsevier India Pvt. Ltd. All rights reserved.
1.
Introduction
The 2-azetidinone (b-lactam) ring system is the common structural feature of a number of broad spectrum b-lactam antibiotics, including penicillin, cephalosporin, carbapenems, nocardicins, monobactams, clavulanic acid, sulbactams and tazobactams, which have been widely used as
chemotherapeutic agents to treat bacterial infections and microbial diseases. Most of the researches up to early 90s focused on synthesis of 2-azetidinones and study of their antibacterial property. In recent years, renewed interest has been focused on the synthesis and modification of b-lactam ring to obtain compounds with diverse pharmacological activities like cholesterol absorption inhibitory activity, human tryptase,
* Corresponding author. New Drug Discovery Research, Department of Medicinal Chemistry, Sunrise University, Alwar, Rajasthan 301030, India. Tel.: þ91 9573396024 (mobile). E-mail address: [email protected] (K. Elumalai). 0975-7619/$ e see front matter Copyright ª 2013, JPR Solutions; Published by Reed Elsevier India Pvt. Ltd. All rights reserved. http://dx.doi.org/10.1016/j.dit.2013.05.007
d r u g i n v e n t i o n t o d a y 5 ( 2 0 1 3 ) 1 0 0 e1 0 4
thrombin and chymase inhibitory activity, antidiabetic, antiinflammatory, anti-parkinsonian and anti-HIV activity. Azetidin-2-one, a four-membered cyclic lactam (b-lactam) skeleton has been recognized as a useful building block for the synthesis of a large number of organic molecules by exploiting the strain energy associated with it. The Staudinger reaction ([2 þ 2] ketene-imine cycloaddition reaction) is regarded as one of the most fundamental and versatile methods for the synthesis of structurally diverse 2-azetidinone derivatives, although many synthetic methods have been developed to date.1,2 Azetidin-2ones can also be synthesized by enolate-imine condensations and cyclization reactions. Efforts have been made in exploring such new aspects of b-lactam chemistry versatile intermediates for the synthesis of aromatic b-amino acids and their derivatives, peptides, polyamines, polyamino alcohols, amino sugars and polyamino ethers the cyclic 2-azetidinone skeleton has been extensively used as a template to build the heterocyclic structure fused to the four-membered ring. This provides an access to diverse structural type of synthetic target molecules lacking b-lactam ring structure.3 Azetidinones are of great biological interest, especially as anti-tubercular, antibacterial, anticonvulsant, anti-tumour, antimalarial, anti-emetic, antihistaminic, antipsychotic and as anti-inflammatory agents.4e12 The chemical structure of isoniazid provides a most valuable molecular template for the development of agents able to interact with a wide variety of biological activities. The synthesis of isoniazid condensed with azetidinones are not reported so for. Hence, it was thought worthwhile to synthesize new congeners by incorporating isoniazid and azetidinones moieties in a single molecular frame work and to evaluate their antimicrobial and antimycobacterial activity.
2.
Materials and methods
The entire chemicals were supplied by E. Merck (Germany) and S.D fine chemicals (India). Melting points were determined by open tube capillary method and are uncorrected. Purity of the compounds was checked on thin layer chromatography (TLC) plates (silica gel G) in the solvent system chloroform, benzene and ethyl acetate as mobile phase (60:30:10) the spots were located under iodine vapours or UV light. IR spectrums were obtained on a PerkineElmer 1720 FTIR spectrometer (KBr Pellets). 1H NMR spectra were recorded or a Bruker AC 300 MHz spectrometer using TMS as internal standard in DMSO/CDCl3. Mass spectra were obtained using Shimadzu LCMS 2010A under ESI ionization technique.
2.1.
Synthesis of Schiff’s bases (5aej)
A mixture of equimolar quantities of compound isoniazid (0.01 mol) and appropriate aryl and heteroaryl aldehyde (0.01 mol) were dissolved in ethanol (95%). The contents were refluxed for a period of 3.5 h on a steam bath. The solid obtained was separated out and recrystallized from ethanol. The compounds with 58e82 percent yield and the compounds melted between 164 and 210 C. The IR spectra of compounds 5aej showed strong absorption bands for carbonyl group (1714 cm1), aromatic CeH stretching (3152 cm1) and aromatic C]C stretching (1640 and 1510 cm1). 1H NMR spectrum of
101
compounds 5aej showed a quartet for methine protons at 7.4 (1H, N]CHeR), multiplets at 6.6e8.1 (AreH). Compounds 5c and 5e showed a singlet at 4.64 due to the signals of third CH group.
2.2.
Synthesis of azetidinones (10aej)
Triethyl amine (0.01 mol) in 1, 4-dioxane, chloroacetyl chloride (0.01 mol) was added drop wise to a solution of the compounds 5aej (0.005 mol) and at room temperature. The reaction mixture was stirred for 15 min. The mixture was then refluxed for 4 h on a water bath. The solid obtained after removal of 1, 4-dioxane was recrystallized from ethanol. The yield, melting point and spectral characterization of the compounds were reported in analytical data.
2.3.
Analytical data
2.3.1. N-(3-chloro-2-oxo-4-phenylazetidin-1-yl) pyridine-4carboxamide (10a) Colourless amorphous solid, M.P: 236e238 C, Yield-52%, IR (KBr, cm1): 3290 (NeH), 3074 (ArCeH), 1652 (C]O, amide), 1490 (ArC]C), 1332 (CeN), 703 (CeCl), 1H NMR (DMSO-d6) d: 1.87 (d, 1H, NeCHeC), 2.56 (d, 1H, CeCHeCl), 7.10 (m, 4H, ArH), 7.45 (m,4H, ArH), 10.12 (s, 1H, CONH). MS (m/z): Mþ calculated 301.06, found 300.94.
2.3.2. N-[3-chloro-2-oxo-4-(pyridin-4-yl) azetidin-1-yl] pyridine-4-carboxamide (10b) Colourless amorphous solid, M.P: 272e274 C, Yield-64%, IR (KBr, cm1): 3274 (NeH), 3086 (ArCeH), 1664 (C]O, amide), 1560 (ArC] C), 1346 (CeN), 708 (CeCl), 1H NMR (DMSO-d6) d: 1.87 (d, 1H, NeCHeC), 2.56 (d, 1H, CeCHeCl), 7.10 (m, 4H, ArH), 7.45 (m, 4H, ArH), 10.12 (s, 1H, CONH). MS (m/z): Mþ calculated 302.05, found 301.84.
2.3.3. N-[3-chloro-2-oxo-4-(pyridin-3-yl) azetidin-1-yl] pyridine-4-carboxamide (10c) Colourless amorphous solid, M.P: 256e258 C, Yield-58%, IR (KBr, cm1): 3238 (NeH), 3066 (ArCeH), 1710 (C]O, amide), 1520 (ArC]C), 1368 (CeN), 709 (CeCl), 1H NMR (DMSO-d6) d: 1.87 (d, 1H, NeCHeC), 2.56 (d, 1H, CeCHeCl), 7.10 (m, 4H, ArH), 7.45 (m, 4H, ArH), 10.12 (s, 1H, CONH). MS (m/z): Mþ calculated 302.05, found 301.90.
2.3.4. N-[3-chloro-2-(3-chlorophenyl)-4-oxoazetidin-1-yl] pyridine-4-carboxamide (10d) Colourless amorphous solid, M.P: 212e214 C, Yield-59%, IR (KBr, cm1): 3244 (NeH), 3079 (ArCeH), 1694 (C]O, amide), 1478 (ArC]C), 1384 (CeN), 705 (CeCl), 1H NMR (DMSO-d6) d: 1.87 (d, 1H, NeCHeC), 2.56 (d, 1H, CeCHeCl), 7.10 (m, 4H, ArH), 7.45 (m, 4H, ArH), 10.12 (s, 1H, CONH). MS (m/z): Mþ calculated 335.02, found 334.88.
2.3.5. N-[3-chloro-2-(3-nitrophenyl)-4-oxoazetidin-1-yl] pyridine-4-carboxamide (10e) Colourless amorphous solid, M.P: 220e223 C, Yield-43%, IR (KBr, cm1): 3247 (NeH), 3064 (ArCeH), 1721 (C]O, amide), 1512 (ArC] C), 1386 (CeN), 707 (CeCl), 1H NMR (DMSO-d6) d: 1.87 (d, 1H, NeCHeC), 2.56 (d, 1H, CeCHeCl), 7.10 (m, 4H, ArH), 7.45 (m, 4H,
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ArH), 10.12 (s, 1H, CONH). MS (m/z): Mþ calculated 346.04, found 345.86.
2.3.6. N-[3-chloro-2-(furan-2-yl)-4-oxoazetidin-1-yl] benzamide (10f) Colourless amorphous solid, M.P: 196e198 C, Yield-44%, IR (KBr, cm1): 3310 (NeH), 3093 (ArCeH), 1652 (C]O, amide), 1468 (ArC]C), 1372 (CeN), 707 (CeCl), 1H NMR (DMSO-d6) d: 1.87 (d, 1H, NeCHeC), 2.56 (d, 1H, CeCHeCl), 7.10 (m, 4H, ArH), 7.45 (m, 4H, ArH), 10.12 (s, 1H, CONH). MS (m/z): Mþ calculated 290.04, found 289.95.
2.3.7. N-{3-chloro-2-oxo-4-[(Z )-2-phenylethenyl] azetidin-1yl} benzamide (10g) Colourless amorphous solid, M.P: 212e215 C, Yield-39%, IR (KBr, cm1): 3248 (NeH), 3058 (ArCeH), 1712 (C]O, amide), 1464 (ArC]C), 1359 (CeN), 704 (CeCl), 1H NMR (DMSO-d6) d: 1.87 (d, 1H, NeCHeC), 2.56 (d, 1H, CeCHeCl), 7.10 (m, 4H, ArH), 7.45 (m, 4H, ArH), 10.12 (s, 1H, CONH). MS (m/z): Mþ calculated 326.08, found 325.75.
2.3.8. N-[3-chloro-2-(4-chlorophenyl)-4-oxoazetidin-1-yl] pyridine-4-carboxamide (10h) Colourless amorphous solid, M.P: 276e279 C, Yield-57%, IR (KBr, cm1): 3274 (NeH), 3048 (ArCeH), 1710 (C]O, amide), 1463 (ArC]C), 1332 (CeN), 705 (CeCl), 1H NMR (DMSO-d6) d: 1.87 (d, 1H, NeCHeC), 2.56 (d, 1H, CeCHeCl), 7.10 (m, 4H, ArH), 7.45 (m, 4H, ArH), 10.12 (s, 1H, CONH). MS (m/z): Mþ calculated 335.02, found 355.48.
2.3.9. N-[3-chloro-2-(4-fluorophenyl)-4-oxoazetidin-1-yl] pyridine-4-carboxamide (10i) Colourless amorphous solid, M.P: 282e284 C, Yield-65%, IR (KBr, cm1): 3246 (NeH), 3085 (ArCeH), 1652 (C]O, amide), 1438 (ArC]C), 1337 (CeN), 707 (CeCl), 1H NMR (DMSO-d6) d: 1.87 (d, 1H, NeCHeC), 2.56 (d, 1H, CeCHeCl), 7.10 (m, 4H, ArH), 7.45 (m, 4H, ArH), 10.12 (s, 1H, CONH). MS (m/z): Mþ calculated 319.05, found 318.94.
NH
O
Colourless amorphous solid, M.P: 226e228 C, Yield-53%, IR (KBr, cm1): 3276 (NeH), 3092 (ArCeH), 1708 (C]O, amide), 1474 (ArC]C), 1384 (CeN), 706 (CeCl), 1H NMR (DMSO-d6) d: 1.87 (d, 1H, NeCHeC), 2.56 (d, 1H, CeCHeCl), 7.10 (m, 4H, ArH), 7.45 (m, 4H, ArH), 10.12 (s, 1H, CONH). MS (m/z): Mþ calculated 335.02, found 335.26.
2.4.
In vitro antimicrobial activity
The in vitro antibacterial activities were tested against grampositive bacteria Bacillus subtilis and gram-negative bacteria Escherichia coli by standard serial dilution method using a stock solution of 100 mg/ml concentrations.13,14 Double strength nutrient broth was used as culture media and dimethyl sulphoxide (DMSO) was used as solvent control. The stock solutions of the test compounds were serially diluted in test tubes containing 1 ml of sterile medium to get the concentration of 50e3.12 mg/ml and then inoculated with 100 mL of suspension of respective microorganism in sterile saline. Norfloxacin was used as standard drug. The inoculated test tubes were incubated at 37 1 C for 24 h.
2.5.
In vitro antimycobacterial activity
Antimycobacterial activity was performed following a protocol previously reported.15 Synthesized compounds were preliminarily assayed against to freshly isolate clinical strains, Mycobacterium fortuitum CA10 and Mycobacterium tuberculosis B814, according to the dilution method in agar. Growth media were MuellereHilton (Difco) containing 10% of OADC (oleic acid, albumin and dextrose complex) for M. fortuitum and Middle brook 7H11 agar (Difco) with 10% of OADC for M. tuberculosis. Substances were tested at single dose of 100 mg/ mL. The active compounds were then assayed for inhibitory activity against a panel of mycobacterial (M. tuberculosis CIP, M. tuberculosis H37Rv) in middle brook 7H11 agar by a standard
NH
O
NH2 +
N
2.3.10. N-[3-chloro-2-(2-chlorophenyl)-4-oxoazetidin-1-yl] pyridine-4-carboxamide (10j)
N
R Cl
a
O
+
RCHO
Cl
N
b
NH
O
O N
R
Cl
N
Scheme 1 e Reagents and conditions: (a) 95% of ethanol, reflux 3.5 h (b) triethyl amine, 1, 4-dioxane, stir-20 min, reflux 4 h.
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d r u g i n v e n t i o n t o d a y 5 ( 2 0 1 3 ) 1 0 0 e1 0 4
Table 1 e In vitro antimicrobial and antimycobacterial activity of synthesized azetidinones. S. no
1 2 3 4 5 6 7 8 9 10 11 12
Compound no.
R
Bacillus subtilis MIC (mmol/mL)
Escherichia coli MIC (mmol/mL)
Mycobacterium tuberculosis CIP MIC (mg/ml)
Mycobacterium tuberculosis H37Rv MIC (mg/ml)
10a 10b 10c 10d 10e 10f 10g 10h 10i 10j Norfloxacin Pyrazinamide
Phenyl 4-Pyridyl 3-Pyridyl 3-Chlorophenyl 3-Nitrophenyl 2-Furyl Cinnamyl 4-Chlorophenyl 4-Flurophenyl 2-Chlorophenyl Standard Standard
0.0435 0.0114 0.0288 0.0209 0.0398 0.0218 0.0296 0.0110 0.0104 0.0119 0.0116 e
0.0356 0.0112 0.0636 0.0538 0.0584 0.0664 0.0648 0.0526 0.0111 0.0584 0.0142 e
3.08 1.14 >2.48 >2.26 1.84 2.24 >3.35 2.62 1.08 2.18 e 1.10
2.98 1.02 2.45 3.26 1.65 2.65 3.45 2.65 1.04 2.21 e 1.13
twofold dilution method. Plates were incubated at 37 C for 3 or 28 days. Pyrazinamide was used as reference compound. After cultivation, MICs were read as minimal concentrations of drugs completely inhibiting visible of mycobacterial growth.
3.
Results and discussion
A series of ten novel azetidinone derivatives were synthesized by cyclocondensation reaction using triethyl amine as an efficient catalyst. All the titled compounds were purified and analyzed by IR, 1H NMR and MS for their structure and all the compounds were evaluated for antimicrobial and antimycobacterial activity.
3.1.
Chemistry
Synthesis of azetidinone derivatives by adopting condensation reaction was performed by following step as outlined in Scheme 1. In the synthesis Schiff’s base of isoniazid reacted with, chloro acetyl chloride in presence of triethyl amine and 1, 4-dioxane under neat conditions resulting in the formation of the product. The reaction times were found to be 4 h. Totally, ten compounds (10ae10j), various substituted azetidinone derivatives, were synthesized with the yield ranging from 39 to 65 percent. The present protocol best describes the synthesis of azetidinone derivative compounds, were found to be novel and not reported elsewhere.
3.2. Antimicrobial activity and antimycobacterial activity Analyzing the activities of the synthesized compounds, the following structure activity relationships (SARs) were obtained. The N1 position of azetidinone derivatives contain isoniazid groups contributed towardsactivity against gram-positive B. subtilis, gram-negative E. coli, M. tuberculosis CIP and H37RV strain. The second position of the azetidinone it contain heteroaryl and substituted aryl group showed more potent antimicrobial and antimycobacterial action (Table 1). Among the reported compounds, compound (10i) contain fluro substituted phenyl group at 2nd position of azetidinone showed potent
antimicrobial and antimycobacterial activity when compared with chloro, nitro and heteroaryl group substituted azetidinones due to strong electron withdrawing nature of the ring. The order of reactivity of the titled compound contain substituted phenyl ring at 2nd position of azetidinones F > NO2 > Cl. Our present study makes it an interesting compound when compared to the current therapeutic agents and are considered the candidates to investigate further for the same.
4.
Conclusion
A series of novel isoniazid cyclocondensed azetidinone of biological interest were synthesized and analyzed for their structures. The azetidinone compounds were prepared by using triethyl amine in 1, 4-dioxane as an efficient catalyst. The importance of substitutions at the second positions of azetidinones was studied towards the antimicrobial and antimycobacterial activity. Almost all of the titled compounds exhibited weak, moderate, or high antimicrobial and antimycobacterial activity. Compounds, such as 10b and 10i, exhibited potential antimicrobial and antimycobacterial activity. Some of new derivatives showed an in vitro activity against gram-positive, gram-negative and M. tuberculosis better than that of antimicrobial drug norfloxacin and antitubercular drug pyrazinamide. Among the compounds reported here in, compound (10i) is arguably the most potent and our present study makes it an interesting compound when compared to the current therapeutic agents and are considered the candidates to investigate further for the same.
Conflicts of interest All authors have none to declare.
Acknowledgements The authors wish to thank Sunrise University for research support and Tuberculosis Research Center, Chennai, India. Also wish to thank JPR Solutions for publication support.
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references
1. Palomo C, Aizpurua JM, Ganboa I, Oiarbide M. Asymmetric synthesis of beta-lactams by Staudinger ketene-imine cycloaddition reaction. Eur J Org Chem. 1999;12:3223e3235. 2. Xu Jiaxi. Stereo selectivity in the synthesis of 2-azetidinones from ketenes and amines via Staudinger reaction. Arkivoc. 2009;4:21e44. 3. Benito A, Alberto RV. New intramolecular cyclization and rearrangement processes based on the radical aryl-aryl coupling of arylsubstituted 2-azetidinones. Tetrahedron Lett. 1998;36:6589e6592. 4. Brickner SJ, Gaikema JJ, Zurenko GE, Greenfield IJ, Manninen PR, Ulanowicz DA. N-acyl 3-alkylidenyl- and 3alkyl azetidin-2-ones: a new class of monocyclic beta-lactam antibacterial agents. 1. Structure-activity relationships of 3isopropylidene and 3-isopropyl analogs. J Antibiot (Tokyo). 1992;45:213e226. 5. Kagthara P, Teja S, Rajeev D, Parekh HH. Synthesis of some 2azetidinones as potential antitubercular agents. Indian J Heterocycl Chem. 2000;10:9e12. 6. Bhanvesh DN, Desai KR. Synthesis of some heterocyclic Schiff base and azetidinones compounds and their anti-bacterial activity. Asian J Chem. 2004;16:1449e1452. 7. Gildasio AS, Luciana MMC, Fernanda CFB, Ana LPM, Eliezer JB, Carlos AMF. New class of potent anti-nociceptive and antiplatelet 10H-phenothiazine-1-acylhydrazone derivatives. Bioorg Med Chem. 2004;12:3149e3158. 8. Mia LL, Ralph RR, Jesse MN, et al. Synthesis, characterization and anticonvulsant activity of enaminones. Part 5:
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investigations on 3-carboalkoxy-2-methyl-2, 3-dihydro-1Hphenothiazin-4[10H]-one derivatives. Bioorg Med Chem. 1998;6:2289e2299. Gdupi RH, Jeeson M, Bhat AR. Synthesis of 2(20 -carboxy-50 nitro phenyl)-3, 4-substitute azetidin-2-ones as antiinflammatory and antimicrobial agents. Indian J Heterocycl Chem. 1996;6:189e196. Choudhari BP, Mulwad VV. Synthesis and antimicrobial screening of N-[60 -coumarinyl amino-3-chloro-4-aryl azetidine-2-ones. Indian J Heterocycl Chem. 2003;2:197e200. Pandey VK, Gupta VD, Upadhyay M, Singh VK, Tandon M. Synthesis, characterization and biological activity of 1, 3, 4substituted 2-azetidinones. Indian J Chem. 2005;44:158e162. Oza HB, Datta NJ, Joshi DG, Parekh HH. Synthesis of some new-2-azetidinones as potential antimicrobial agents. Indian J Heterocycl Chem. 2003;12:275e276. Zhu Bin, Marinelli Brett A, Goldschmidt Raul, et al. Synthesis and antibacterial activity of 7-(1, 2, 3, 4-tetrahydropyrrolo [1, 2-a]-pyrazin-7-yl) quinolones. Bioorg Med Chem Lett. 2009;19: 4933e4936. Avupati Vasudeva Rao, Yejella Rajendra Prasad, Guntuku Girijasankar, Gunta Pradeepsagar. Synthesis, characterization and in vitro biological evaluation of some novel diarylsulfonylureas as potential cytotoxic and antimicrobial agents. Bioorg Med Chem Lett. 2012;22: 1031e1035. Biava M, Porretta GC, Poce G. Identification of a novel pyrole derivative endowed with antimycobacterial activity and protection index comparable to that of the current antitubercular drugs streptomycin and rifampicin. Bioorg Med Chem. 2010;18:8076e8084.
Available online at www.sciencedirect.com
journal homepage: www.elsevier.com/locate/dit
Original Article
Design, synthesis and biological evaluation of some novel isoniazid cyclocondensed azetidinones Karthikeyan Elumalai a,c,*, Mohammed Ashraf Ali a, Manogaran Elumalai b, Kalpana Eluri b, Sivaneswari Srinivasan d, Sujit Kumar Mohanti c, Anil Thota c a
New Drug Discovery Research, Department of Medicinal Chemistry, Sunrise University, Alwar, Rajasthan 301030, India b Faculty of Pharmaceutical Sciences, UCSI University, Cheras, Kuala Lumpur 56000, Malaysia c Department of Pharmaceutical Chemistry, Jayamukhi Institute of Pharmaceutical Sciences, Warangal 506 332, India d Department of Pharmaceutics, Jayamukhi College of Pharmacy, Warangal 506 332, India
article info
abstract
Article history:
Background: In the present study, a series of novel azetidinone derivatives synthesized
Received 20 May 2013
because of its potent antimicrobial and antimycobacterial activity.
Accepted 25 May 2013
Method: Compounds (10ae10j) were synthesized by reacting Schiff’s base of isoniazid reacted with, chloro acetyl chloride in presence of triethyl amine and 1, 4-dioxane as an efficient catalyst, analyzed for their structures. in vitro antimicrobial and antimycobacterial
Keywords:
activity were carried out.
Azetidinones
Results: Among the synthesized compounds, compound 10b and 10i was found to be the
Antimycobacterial
most potent against gram-positive bacteria Bacillus subtilis, gram-negative bacteria Escher-
Isoniazid cyclocondensed
ichia coli, Mycobacterium tuberculosis CIP and M. tuberculosis H37Rv.
Mycobacterium tuberculosis CIP and
Conclusion: A series of novel azetidinone derivatives of biological interest were synthesized
H37RV strain
and analyzed, suggests that it an interesting compound compared to the current therapeutic agents and are considered to investigate further for the same. Copyright ª 2013, JPR Solutions; Published by Reed Elsevier India Pvt. Ltd. All rights reserved.
1.
Introduction
The 2-azetidinone (b-lactam) ring system is the common structural feature of a number of broad spectrum b-lactam antibiotics, including penicillin, cephalosporin, carbapenems, nocardicins, monobactams, clavulanic acid, sulbactams and tazobactams, which have been widely used as
chemotherapeutic agents to treat bacterial infections and microbial diseases. Most of the researches up to early 90s focused on synthesis of 2-azetidinones and study of their antibacterial property. In recent years, renewed interest has been focused on the synthesis and modification of b-lactam ring to obtain compounds with diverse pharmacological activities like cholesterol absorption inhibitory activity, human tryptase,
* Corresponding author. New Drug Discovery Research, Department of Medicinal Chemistry, Sunrise University, Alwar, Rajasthan 301030, India. Tel.: þ91 9573396024 (mobile). E-mail address: [email protected] (K. Elumalai). 0975-7619/$ e see front matter Copyright ª 2013, JPR Solutions; Published by Reed Elsevier India Pvt. Ltd. All rights reserved. http://dx.doi.org/10.1016/j.dit.2013.05.007
d r u g i n v e n t i o n t o d a y 5 ( 2 0 1 3 ) 1 0 0 e1 0 4
thrombin and chymase inhibitory activity, antidiabetic, antiinflammatory, anti-parkinsonian and anti-HIV activity. Azetidin-2-one, a four-membered cyclic lactam (b-lactam) skeleton has been recognized as a useful building block for the synthesis of a large number of organic molecules by exploiting the strain energy associated with it. The Staudinger reaction ([2 þ 2] ketene-imine cycloaddition reaction) is regarded as one of the most fundamental and versatile methods for the synthesis of structurally diverse 2-azetidinone derivatives, although many synthetic methods have been developed to date.1,2 Azetidin-2ones can also be synthesized by enolate-imine condensations and cyclization reactions. Efforts have been made in exploring such new aspects of b-lactam chemistry versatile intermediates for the synthesis of aromatic b-amino acids and their derivatives, peptides, polyamines, polyamino alcohols, amino sugars and polyamino ethers the cyclic 2-azetidinone skeleton has been extensively used as a template to build the heterocyclic structure fused to the four-membered ring. This provides an access to diverse structural type of synthetic target molecules lacking b-lactam ring structure.3 Azetidinones are of great biological interest, especially as anti-tubercular, antibacterial, anticonvulsant, anti-tumour, antimalarial, anti-emetic, antihistaminic, antipsychotic and as anti-inflammatory agents.4e12 The chemical structure of isoniazid provides a most valuable molecular template for the development of agents able to interact with a wide variety of biological activities. The synthesis of isoniazid condensed with azetidinones are not reported so for. Hence, it was thought worthwhile to synthesize new congeners by incorporating isoniazid and azetidinones moieties in a single molecular frame work and to evaluate their antimicrobial and antimycobacterial activity.
2.
Materials and methods
The entire chemicals were supplied by E. Merck (Germany) and S.D fine chemicals (India). Melting points were determined by open tube capillary method and are uncorrected. Purity of the compounds was checked on thin layer chromatography (TLC) plates (silica gel G) in the solvent system chloroform, benzene and ethyl acetate as mobile phase (60:30:10) the spots were located under iodine vapours or UV light. IR spectrums were obtained on a PerkineElmer 1720 FTIR spectrometer (KBr Pellets). 1H NMR spectra were recorded or a Bruker AC 300 MHz spectrometer using TMS as internal standard in DMSO/CDCl3. Mass spectra were obtained using Shimadzu LCMS 2010A under ESI ionization technique.
2.1.
Synthesis of Schiff’s bases (5aej)
A mixture of equimolar quantities of compound isoniazid (0.01 mol) and appropriate aryl and heteroaryl aldehyde (0.01 mol) were dissolved in ethanol (95%). The contents were refluxed for a period of 3.5 h on a steam bath. The solid obtained was separated out and recrystallized from ethanol. The compounds with 58e82 percent yield and the compounds melted between 164 and 210 C. The IR spectra of compounds 5aej showed strong absorption bands for carbonyl group (1714 cm1), aromatic CeH stretching (3152 cm1) and aromatic C]C stretching (1640 and 1510 cm1). 1H NMR spectrum of
101
compounds 5aej showed a quartet for methine protons at 7.4 (1H, N]CHeR), multiplets at 6.6e8.1 (AreH). Compounds 5c and 5e showed a singlet at 4.64 due to the signals of third CH group.
2.2.
Synthesis of azetidinones (10aej)
Triethyl amine (0.01 mol) in 1, 4-dioxane, chloroacetyl chloride (0.01 mol) was added drop wise to a solution of the compounds 5aej (0.005 mol) and at room temperature. The reaction mixture was stirred for 15 min. The mixture was then refluxed for 4 h on a water bath. The solid obtained after removal of 1, 4-dioxane was recrystallized from ethanol. The yield, melting point and spectral characterization of the compounds were reported in analytical data.
2.3.
Analytical data
2.3.1. N-(3-chloro-2-oxo-4-phenylazetidin-1-yl) pyridine-4carboxamide (10a) Colourless amorphous solid, M.P: 236e238 C, Yield-52%, IR (KBr, cm1): 3290 (NeH), 3074 (ArCeH), 1652 (C]O, amide), 1490 (ArC]C), 1332 (CeN), 703 (CeCl), 1H NMR (DMSO-d6) d: 1.87 (d, 1H, NeCHeC), 2.56 (d, 1H, CeCHeCl), 7.10 (m, 4H, ArH), 7.45 (m,4H, ArH), 10.12 (s, 1H, CONH). MS (m/z): Mþ calculated 301.06, found 300.94.
2.3.2. N-[3-chloro-2-oxo-4-(pyridin-4-yl) azetidin-1-yl] pyridine-4-carboxamide (10b) Colourless amorphous solid, M.P: 272e274 C, Yield-64%, IR (KBr, cm1): 3274 (NeH), 3086 (ArCeH), 1664 (C]O, amide), 1560 (ArC] C), 1346 (CeN), 708 (CeCl), 1H NMR (DMSO-d6) d: 1.87 (d, 1H, NeCHeC), 2.56 (d, 1H, CeCHeCl), 7.10 (m, 4H, ArH), 7.45 (m, 4H, ArH), 10.12 (s, 1H, CONH). MS (m/z): Mþ calculated 302.05, found 301.84.
2.3.3. N-[3-chloro-2-oxo-4-(pyridin-3-yl) azetidin-1-yl] pyridine-4-carboxamide (10c) Colourless amorphous solid, M.P: 256e258 C, Yield-58%, IR (KBr, cm1): 3238 (NeH), 3066 (ArCeH), 1710 (C]O, amide), 1520 (ArC]C), 1368 (CeN), 709 (CeCl), 1H NMR (DMSO-d6) d: 1.87 (d, 1H, NeCHeC), 2.56 (d, 1H, CeCHeCl), 7.10 (m, 4H, ArH), 7.45 (m, 4H, ArH), 10.12 (s, 1H, CONH). MS (m/z): Mþ calculated 302.05, found 301.90.
2.3.4. N-[3-chloro-2-(3-chlorophenyl)-4-oxoazetidin-1-yl] pyridine-4-carboxamide (10d) Colourless amorphous solid, M.P: 212e214 C, Yield-59%, IR (KBr, cm1): 3244 (NeH), 3079 (ArCeH), 1694 (C]O, amide), 1478 (ArC]C), 1384 (CeN), 705 (CeCl), 1H NMR (DMSO-d6) d: 1.87 (d, 1H, NeCHeC), 2.56 (d, 1H, CeCHeCl), 7.10 (m, 4H, ArH), 7.45 (m, 4H, ArH), 10.12 (s, 1H, CONH). MS (m/z): Mþ calculated 335.02, found 334.88.
2.3.5. N-[3-chloro-2-(3-nitrophenyl)-4-oxoazetidin-1-yl] pyridine-4-carboxamide (10e) Colourless amorphous solid, M.P: 220e223 C, Yield-43%, IR (KBr, cm1): 3247 (NeH), 3064 (ArCeH), 1721 (C]O, amide), 1512 (ArC] C), 1386 (CeN), 707 (CeCl), 1H NMR (DMSO-d6) d: 1.87 (d, 1H, NeCHeC), 2.56 (d, 1H, CeCHeCl), 7.10 (m, 4H, ArH), 7.45 (m, 4H,
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ArH), 10.12 (s, 1H, CONH). MS (m/z): Mþ calculated 346.04, found 345.86.
2.3.6. N-[3-chloro-2-(furan-2-yl)-4-oxoazetidin-1-yl] benzamide (10f) Colourless amorphous solid, M.P: 196e198 C, Yield-44%, IR (KBr, cm1): 3310 (NeH), 3093 (ArCeH), 1652 (C]O, amide), 1468 (ArC]C), 1372 (CeN), 707 (CeCl), 1H NMR (DMSO-d6) d: 1.87 (d, 1H, NeCHeC), 2.56 (d, 1H, CeCHeCl), 7.10 (m, 4H, ArH), 7.45 (m, 4H, ArH), 10.12 (s, 1H, CONH). MS (m/z): Mþ calculated 290.04, found 289.95.
2.3.7. N-{3-chloro-2-oxo-4-[(Z )-2-phenylethenyl] azetidin-1yl} benzamide (10g) Colourless amorphous solid, M.P: 212e215 C, Yield-39%, IR (KBr, cm1): 3248 (NeH), 3058 (ArCeH), 1712 (C]O, amide), 1464 (ArC]C), 1359 (CeN), 704 (CeCl), 1H NMR (DMSO-d6) d: 1.87 (d, 1H, NeCHeC), 2.56 (d, 1H, CeCHeCl), 7.10 (m, 4H, ArH), 7.45 (m, 4H, ArH), 10.12 (s, 1H, CONH). MS (m/z): Mþ calculated 326.08, found 325.75.
2.3.8. N-[3-chloro-2-(4-chlorophenyl)-4-oxoazetidin-1-yl] pyridine-4-carboxamide (10h) Colourless amorphous solid, M.P: 276e279 C, Yield-57%, IR (KBr, cm1): 3274 (NeH), 3048 (ArCeH), 1710 (C]O, amide), 1463 (ArC]C), 1332 (CeN), 705 (CeCl), 1H NMR (DMSO-d6) d: 1.87 (d, 1H, NeCHeC), 2.56 (d, 1H, CeCHeCl), 7.10 (m, 4H, ArH), 7.45 (m, 4H, ArH), 10.12 (s, 1H, CONH). MS (m/z): Mþ calculated 335.02, found 355.48.
2.3.9. N-[3-chloro-2-(4-fluorophenyl)-4-oxoazetidin-1-yl] pyridine-4-carboxamide (10i) Colourless amorphous solid, M.P: 282e284 C, Yield-65%, IR (KBr, cm1): 3246 (NeH), 3085 (ArCeH), 1652 (C]O, amide), 1438 (ArC]C), 1337 (CeN), 707 (CeCl), 1H NMR (DMSO-d6) d: 1.87 (d, 1H, NeCHeC), 2.56 (d, 1H, CeCHeCl), 7.10 (m, 4H, ArH), 7.45 (m, 4H, ArH), 10.12 (s, 1H, CONH). MS (m/z): Mþ calculated 319.05, found 318.94.
NH
O
Colourless amorphous solid, M.P: 226e228 C, Yield-53%, IR (KBr, cm1): 3276 (NeH), 3092 (ArCeH), 1708 (C]O, amide), 1474 (ArC]C), 1384 (CeN), 706 (CeCl), 1H NMR (DMSO-d6) d: 1.87 (d, 1H, NeCHeC), 2.56 (d, 1H, CeCHeCl), 7.10 (m, 4H, ArH), 7.45 (m, 4H, ArH), 10.12 (s, 1H, CONH). MS (m/z): Mþ calculated 335.02, found 335.26.
2.4.
In vitro antimicrobial activity
The in vitro antibacterial activities were tested against grampositive bacteria Bacillus subtilis and gram-negative bacteria Escherichia coli by standard serial dilution method using a stock solution of 100 mg/ml concentrations.13,14 Double strength nutrient broth was used as culture media and dimethyl sulphoxide (DMSO) was used as solvent control. The stock solutions of the test compounds were serially diluted in test tubes containing 1 ml of sterile medium to get the concentration of 50e3.12 mg/ml and then inoculated with 100 mL of suspension of respective microorganism in sterile saline. Norfloxacin was used as standard drug. The inoculated test tubes were incubated at 37 1 C for 24 h.
2.5.
In vitro antimycobacterial activity
Antimycobacterial activity was performed following a protocol previously reported.15 Synthesized compounds were preliminarily assayed against to freshly isolate clinical strains, Mycobacterium fortuitum CA10 and Mycobacterium tuberculosis B814, according to the dilution method in agar. Growth media were MuellereHilton (Difco) containing 10% of OADC (oleic acid, albumin and dextrose complex) for M. fortuitum and Middle brook 7H11 agar (Difco) with 10% of OADC for M. tuberculosis. Substances were tested at single dose of 100 mg/ mL. The active compounds were then assayed for inhibitory activity against a panel of mycobacterial (M. tuberculosis CIP, M. tuberculosis H37Rv) in middle brook 7H11 agar by a standard
NH
O
NH2 +
N
2.3.10. N-[3-chloro-2-(2-chlorophenyl)-4-oxoazetidin-1-yl] pyridine-4-carboxamide (10j)
N
R Cl
a
O
+
RCHO
Cl
N
b
NH
O
O N
R
Cl
N
Scheme 1 e Reagents and conditions: (a) 95% of ethanol, reflux 3.5 h (b) triethyl amine, 1, 4-dioxane, stir-20 min, reflux 4 h.
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Table 1 e In vitro antimicrobial and antimycobacterial activity of synthesized azetidinones. S. no
1 2 3 4 5 6 7 8 9 10 11 12
Compound no.
R
Bacillus subtilis MIC (mmol/mL)
Escherichia coli MIC (mmol/mL)
Mycobacterium tuberculosis CIP MIC (mg/ml)
Mycobacterium tuberculosis H37Rv MIC (mg/ml)
10a 10b 10c 10d 10e 10f 10g 10h 10i 10j Norfloxacin Pyrazinamide
Phenyl 4-Pyridyl 3-Pyridyl 3-Chlorophenyl 3-Nitrophenyl 2-Furyl Cinnamyl 4-Chlorophenyl 4-Flurophenyl 2-Chlorophenyl Standard Standard
0.0435 0.0114 0.0288 0.0209 0.0398 0.0218 0.0296 0.0110 0.0104 0.0119 0.0116 e
0.0356 0.0112 0.0636 0.0538 0.0584 0.0664 0.0648 0.0526 0.0111 0.0584 0.0142 e
3.08 1.14 >2.48 >2.26 1.84 2.24 >3.35 2.62 1.08 2.18 e 1.10
2.98 1.02 2.45 3.26 1.65 2.65 3.45 2.65 1.04 2.21 e 1.13
twofold dilution method. Plates were incubated at 37 C for 3 or 28 days. Pyrazinamide was used as reference compound. After cultivation, MICs were read as minimal concentrations of drugs completely inhibiting visible of mycobacterial growth.
3.
Results and discussion
A series of ten novel azetidinone derivatives were synthesized by cyclocondensation reaction using triethyl amine as an efficient catalyst. All the titled compounds were purified and analyzed by IR, 1H NMR and MS for their structure and all the compounds were evaluated for antimicrobial and antimycobacterial activity.
3.1.
Chemistry
Synthesis of azetidinone derivatives by adopting condensation reaction was performed by following step as outlined in Scheme 1. In the synthesis Schiff’s base of isoniazid reacted with, chloro acetyl chloride in presence of triethyl amine and 1, 4-dioxane under neat conditions resulting in the formation of the product. The reaction times were found to be 4 h. Totally, ten compounds (10ae10j), various substituted azetidinone derivatives, were synthesized with the yield ranging from 39 to 65 percent. The present protocol best describes the synthesis of azetidinone derivative compounds, were found to be novel and not reported elsewhere.
3.2. Antimicrobial activity and antimycobacterial activity Analyzing the activities of the synthesized compounds, the following structure activity relationships (SARs) were obtained. The N1 position of azetidinone derivatives contain isoniazid groups contributed towardsactivity against gram-positive B. subtilis, gram-negative E. coli, M. tuberculosis CIP and H37RV strain. The second position of the azetidinone it contain heteroaryl and substituted aryl group showed more potent antimicrobial and antimycobacterial action (Table 1). Among the reported compounds, compound (10i) contain fluro substituted phenyl group at 2nd position of azetidinone showed potent
antimicrobial and antimycobacterial activity when compared with chloro, nitro and heteroaryl group substituted azetidinones due to strong electron withdrawing nature of the ring. The order of reactivity of the titled compound contain substituted phenyl ring at 2nd position of azetidinones F > NO2 > Cl. Our present study makes it an interesting compound when compared to the current therapeutic agents and are considered the candidates to investigate further for the same.
4.
Conclusion
A series of novel isoniazid cyclocondensed azetidinone of biological interest were synthesized and analyzed for their structures. The azetidinone compounds were prepared by using triethyl amine in 1, 4-dioxane as an efficient catalyst. The importance of substitutions at the second positions of azetidinones was studied towards the antimicrobial and antimycobacterial activity. Almost all of the titled compounds exhibited weak, moderate, or high antimicrobial and antimycobacterial activity. Compounds, such as 10b and 10i, exhibited potential antimicrobial and antimycobacterial activity. Some of new derivatives showed an in vitro activity against gram-positive, gram-negative and M. tuberculosis better than that of antimicrobial drug norfloxacin and antitubercular drug pyrazinamide. Among the compounds reported here in, compound (10i) is arguably the most potent and our present study makes it an interesting compound when compared to the current therapeutic agents and are considered the candidates to investigate further for the same.
Conflicts of interest All authors have none to declare.
Acknowledgements The authors wish to thank Sunrise University for research support and Tuberculosis Research Center, Chennai, India. Also wish to thank JPR Solutions for publication support.
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