научная статья по теме SYNTHESIS AND BIOLOGICAL EVALUATION OF NOVEL ISOXAZOLO[4,3-E]INDOLES AS ANTIBACTERIAL AGENTS Химия

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EHOOPrAHH^ECKÄM XHMH3, 2013, moM 39, № 2, c. 240-243

SYNTHESIS AND BIOLOGICAL EVALUATION OF NOVEL ISOXAZOLO[4,3-e]INDOLES AS ANTIBACTERIAL AGENTS

© 2013 Mehdi Pordel#, Amin Abdollahi, Bahareh Razavi

Department of Chemistry, Mashhad Branch, Islamic Azad University, Mashhad, Iran Received July 30, 2012; in final form August 6, 2012

Abstract—The synthesis of a new series of 8-bromo-6-alkyl-1-aryl-6H-isoxazolo[4,3-e]indole derivatives is described. All the newly synthesized compounds were screened for their antibacterial activity against Escherichia coli HB101, Staphylococcuse aureus pathogens (methicillin resistant S. aureus and methicillin susceptible S. aureus), Pseudomonas aeruginosa, and Bacillus subtilis; also MIC values of these compounds were determined.

Keywords: isoxazolo[4,3-e]indole, antibacterial agents, Staphylococcus aureus, MIC DOI: 10.7868/S0132342313020103

INTRODUCTION

Indoles are a pervasive class of compounds found in abundance in biologically active compounds such as pharmaceuticals [1, 2], agrochemicals [3] and alkaloids [4]. Indole myriad derivatives have, therefore, captured the attention of organic synthetic chemists. On the other hand, isoxazoles have found continuing application in medicinal chemistry, several examples of which have advanced to general medical practice [5]. As potential new chemical entities advance from in vitro screening through in vivo study towards clinical trials, pharmacokinetic properties collectively referred to as ADMET for Absorption, Distribution, Metabolism, Excretion and Toxicity [6, 7] become important considerations: half of drug attrition can be attributed to poor ADMET properties. An interesting facet of the biology of isoxazoles is that one significant route of metabolism and excretion is C-5 methyl hydroxyla-tion [8, 9], mediated by the cytochrome P450 isoform 3A4 [10], often followed by conjugation to the glucu-ronide [11]. The existence of a safe route for drug metabolism is an important design feature for potential investigational new drugs containing the isoxazole. Combination of the isoxazole moiety with the indole nucleus may enhance these activities [12, 13].

# Corresponding author: tel.: 0098-0511-8414182; fax: 00980511-8424020; e-mail: mehdipordel58@yahoo.com.

Based on these facts and in continuation of research work on the synthesis of bioactive heterocycles [14—17] and evaluation of their biological activities, we decided to examine the transformation of 3-bro-mo-1-alkyl-5-nitro-1H-indoles to new heterocyclic system isoxazolo[4,3-e]indole by the reaction of nu-cleophilic hydrogen substitution, which may result in interesting biological activities.

RESULTS AND DISCUSSION

The target compounds 8-bromo-6-alkyl-1-aryl-6H-isoxazolo[4,3-e]indole derivatives (3a—h) were obtained as described in Scheme. The required starting materials 3-bromo-1-alkyl-5-nitro-1H-indoles (1a—d) were prepared by reaction of 5-nitro-1H-in-dole with bromine [18] and then different alkyl halides in DMF and KOH [19] using literature methods. The reaction of (1a—d) with arylacetonitriles (2a,b) led to the formation of the new 8-bromo-6-alkyl-1-aryl-6H-isoxazolo[4,3-e]indole (3a—h) via the nucleophilic substitution of hydrogen in basic EtOH solution in moderate yields ( cf. [20—23]) (Scheme). The yield of the reaction was very low when instead of 3-bromo-1-alkyl-5-nitro-1H-indoles (1a—d), 1-alkyl-5-nitro-1H-indoles was used in this reaction. This can be due to the presence of bromine in the indole ring of compounds (1a—d) which are more electron-deficient than in 1-alkyl-5-nitro-1H-indoles.

SYNTHESIS AND BIOLOGICAL EVALUATION

241

Antibacterial activity (MIC, mL 1) of compounds 3a—h against methicillin-resistant (MRSA) and methicillin-suscep-tible Staphylococcus aureus (MSSA)

Compd. MRSA MSSA Compd. MRSA MSSA

(3a) 22.0 15.2 (3f) 10.0 10.0

(3b) 15.7 10.0 (3g) 4.3 4.3

(3c) 10.0 10.0 (3h) 5.0 5.0

(3d) 17.5 10.6 Erythromycin 32.0 32.0

(3e) 15.3 15.3 Cephalexin 72.0 4.6

3a: R = Et, R' = Cl (49%), 3c: R = Bu, R' = Cl (43%), 3e: R = Et, R' = Me (51%), 3g: R = Bu, R' = Me (55%), 3b: R = Et, R' = Cl (45%), 3d: R = iso-Bu, R' = Cl (55%), 3f: R = Pr, R' = Me (50%), 3h: R = iso-Bu, R' = Me (60%)

Scheme

The structural assignments of compounds (3a—h) were based on the analytical and spectral data. For example, in the XH NMR spectrum of (3a), there are the doublet signals at 8 7.40 (J = 9.6 Hz, 1 H), 8 7.47 (J= = 9.6 Hz, 1 H), 8 7.57 (J = 8.5 Hz, 2 H), 8 8.85 (J = = 8.5 Hz, 2 H) ppm and singlet signal at 8 7.85 ppm attributed to seven protons of aromatic rings. Moreover, the 13C NMR spectrum, molecular ion peak at m/z 379 (M+ + 4) and microanalytical data strongly support the tricyclic structure of compound (3a).

ANTIMICROBIAL ACTIVITY

The test compounds listed in table were screened for the antibacterial activity against Escherichia coli HB101 (BA-7601C), Staphylococcuse aureus pathogens [methicillin resistant S. aureus (MRSA) and me-thicillin susceptible S. aureus (MSSA - ATCC 1112)], Pseudomonas aeruginosa (PTCC 1431), and Bacillus subtilis (PTCC 1365).

The Minimum inhibitory concentrations (MIC) of (3a—h) were determined in dilution test tube method, which had been introduced by NCCLS (National Committee for Clinical Laboratory Standards) [24]. For broth dilution methods, in which decreasing concentrations of the antimicrobial agents must be tested, usually prepared in serial two fold dilution of a broth medium is placed in tubes which will support the growth of the test microorganism (104 CFU mL-1).

After sufficient incubation (18 h), the tubes are examined for turbidity, indicating growth of the microorganism. The organism will grow in the tube that does not contain enough antimicrobial agents to inhibit growth. For further confidence, the samples were cultured onto Petri dishes containing Muller Hinton agar (18 h at 37oC). The lowest concentration of the antibacterial agent that prevents growth of the test organism, as detected by lack of visual turbidity (matching the negative growth control), is designated the minimum inhibitory concentration (MIC). A serial dilution of tested compounds (final concentration of 400 to 0.4 ^g mL-1), were added to the test bacteria in Mueller-Hinton broth and were incubated at 37oC for 18 h. Growth was presented in the medium control and was absent from the inoculum control [25].

The result indicates that these compounds are only effective against gram positive bacteria. It has been observed from the table, all compounds were found to exhibit considerable antibacterial activities against the mentioned organisms. These results are compared with MIC values of Cephalexin (72 and 4.6 ^g mL-1 and Erythromycin (32 and 32 ^g mL-1). As the data in table display, compound 3g shows the best inhibitory activity against methicillin resistant S. aureus (MRSA) and methicillin susceptible S. aureus (MSSA-ATCC 1112).

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CONCLUSIONS

To summarize, we have synthesized some novel derivatives of isoxazolo[4,3-e]indole and shown them to be very effective S. aureus growth inhibitors. Such compounds would appear to offer a suitable template for the design of more powerful antibacterial agents and further studies are under way to this end in our laboratory.

EXPERIMENTAL

Melting points were measured on an Electrother-maltype-9100 melting-point apparatus. XH and 13C NMR spectra (8, ppm, J, Hz) were recorded on a Bruk-er Avance DRX-400 FT spectrometer at 400 MHz for 1H and 100 MHz for 13C using CDCl3 as solvent. The XH and 13C NMR chemical shifts were referenced to tetramethylsilane (TMS) as internal standard. The mass spectra were recorded on a Varian Mat, CH-7 at 70 eV. Elemental analysis was performed on a Thermo Finnigan Flash EA microanalyzer.

The microorganisms S. aureus ATCC 1112 were purchased from Pasteur Institute of Iran and S. aureus (methicillin resistant) was isolated from different specimens which were referred to the Microbiological Laboratory of Ghaem Hospital of Medical University of Mashhad-Iran and its methicillin resistance was tested according to the NCCLS guidelines [24].

General procedure for the synthesis of (3a—h) from (1a—d) and (2a,b).

Compounds (1a—d) (10 mmol) and (2a,b) (12 mmol) were added with stirring to a solution of KOH (20 g, 357 mmol) in ethanol (80 mL). The mixture was re-fluxed with stirring for 4 h, and then poured into water. The precipitate was collected by filtration, washed with water and air-dried to give crude (3a—h). More purification was achieved by crystallization from suitable solvent such as n-hexane-ethyl-acetate, MeOH or EtOH.

8-Bromo-1-(4-chlorophenyl)-6-ethyl-6H-isoxazo-lo[4,3-e]indole (3a) was obtained as pale yellow crystals (EtOH), yield (49%), M.p.: 151-153°C; 1H NMR (CDCl3) 8 1.57 (3 H, t, J = 7.3), 4.27 (2 H, q, J = 7.3 ), 7.40 (1 H, d, J = 9.6 ), .47 (1 H, d, J = 9.6 ), 7.57 (2 H, d, J = 8.5), 7.85 (1 H, s), 8.85 (2 H, d, J = = 8.5); 13C NMR (CDCl3): 8 15.25, 43.56, 93.89, 112.24, 124.33, 127.45, 131.02, 132.87, 132.95, 133.23, 145.87, 148.55, 158.83, 160.19, 164.17; MS (m/z) 379 (M+4); Anal. Calcd. for C17H12BrClN2O (375.6): C, 54.36; H, 3.22; N, 7.46. Found: C, 54.21; H, 3.18; N, 7.58.

8-Bromo-1-(4-chlorophenyl)-6-propyl-6H-isoxa-zolo[4,3-e]indole (3b) was obtained as pale yellow needles (MeOH), yield (45%), M.p.: 141-143°C; 1H NMR (CDCl3) 8 0.98 (3 H, t, J= 7.3), 1.95-1.87 (2 H, m,), 4.21 (2 H, t, J = 7.3), 7.41 (1 H, d, J = 9.6), 7.46 (1 H, d, J = 9.6), 7.56 (2 H, d, J = 8.5), 7.85 (1 H,

s), 8.85 (2 H, d, J = 8.5) ppm; 13C NMR (CDCl3): 8 11.23, 27.09, 44.59, 93.75, 112.23, 124.56, 127.56, 131.08, 132.90, 132.96, 133.23, 145.89, 148.63, 158.91, 160.23, 164.15; MS (m/z) 393 (M+4); Anal. Calcd for C18H14BrClN2O (389.7): C, 55.48; H, 3.62; N, 7.19. Found: C, 55.65; H, 3.69; N, 7.02.

8-Bromo-6-butyl-1-(4-chlorophenyl)-6H-isoxazo-lo[4,3-e]indole (3c) was obtained as yellow needles (MeOH), yield (43%), M.p.: 154-1

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