Synthesis, Characterization of some Novel 6h-Indolo (2, 3-b) Quinoxaline Fused Azetidinones as Potential Bioactive Molecules

Sai Padmini. D ¹, Narayana Babu. M ², Madhavan. V ²

¹PRRM College of Pharmacy, Kadapa , Andhra Pradesh-516003

²M S Ramaiah College of Pharmacy, MSR Nagar, Bangalore

*Corresponding Author E-mail: dspadmini.pharma@gmail.com

ABSTRACT:

Synthetic analogs of indoles and azetidinones are reported to possess various pharmacological activities such as anti-inflammatory, antidepressant, neuroleptic, cytotoxic, antitubercular, antihypercholesterolemic, antioxidant activities along with wide range of antimicrobial activity. Isatin (indole-2,3-dione) on treatment with o-phenelynediamine in presence of ethanol and glacial acetic acid gave 6H-indolo(2,3 b)quinoxaline [M1] which on treatment with ethylchloroacetate gave Ethyl [6H-indolo(2,3 b)quinoxaline)-1-acetate [M2] which on amination with hydrazine hydrate afforded 6H-indolo(2,3 b)quinoxaline-1- acetic acid hydrazide[M3].New series of N-(3-chloro-2 oxo-4 aryl azetidine-1-yl)-2-[6H-indolo (2, 3 b) quinoxaline-6-yl] acetamides[M4] were synthesized by condensation of 6H-indolo (2,3b) quinoxaline-1-acetic acid with different aromatic aldehydes to produce a series of schiff’s bases. β-lactam moiety was incorporated on Schiff’s base through cycloaddition reaction using chloroacetyl chloride in presence of triethyl amine. The synthesized compounds were charecterised by IR, ¹H NMR and Massspectral studies. All the newly synthesized compounds have been evaluated for their antimicrobial activity against gram-positive and gram-negative bacteria and fungi The synthesized compounds were active against gram positive, gram negative bacterial and fungal species. Para nitro substituted derivatives are good antimicrobial agents among all the derivatives.

KEYWORDS: Indoloquinoxalines, Schiff base, azetidinone, anti-bacterial, antifungal activity.

INTRODUCTION:

Azetidinones which are part of the antibiotic structure are known to exhibit interesting biological activities. A large number of 3-chloro monocyclic β- lactams possess powerful antibacterial^1-5, antifungal^6,7, anti-tubercular⁸, anticonvulsant^9,10, anti-inflammatory^11-15, enzyme inhibition activities. They are also effective on central nervous system. Many derivatives of quinoxaline show a wide variety of biological activities¹⁶ which made them privileged structure in combinatorial drug discovery libraries.

Quinoxaline derivatives constitute the basis of many insecticides, fungicides, herbicides as well as being important in human health as receptor antagonists. Synthetic quinoxaline moiety is a part of number of antibiotics such as levomycin, actinomycin etc. In view of these observations, we have synthesized some novel heterocyclic derivatives obtained by tethering indoloquinoxaline with azetidinone and studied their biological activity.

MATERIALS AND METHODS:

The chemicals and reagents used in the present work were of AR and LR grade, procured from Aldrich, Hi-media, Merck, S. d– Fine Chem Ltd and Sigma. The melting points of the synthesized compounds were determined by open capillary tube method and all the compounds gave sharp melting points and were uncorrected. The purity of the compounds was ascertained by thin layer chromatography using silica gel-G as stationary phase. The IR spectra of the synthesized compounds were recorded on a Fourier Transform IR spectrophotometer (model Shimadzu 8400S) in the range of 400-4000 using diffuse reflectance system and values of ν_max are reported in cm^-1. ¹H NMR spectra were recorded on amx-400 NMR spectrometer and chemical shifts (δ) are reported in parts per million downfield from internal reference Tetramethylsilane (TMS). Mass spectra were recorded on Shimadzu LC-MS model 2010A. Elemental analysis of compounds was performed on FLASH EA 1112 SERIES.

Preparation of 6H-Indolo (2, 3 b) quinoxaline(M1):

1.08 g of orthophenylenediamine (0.01mol) and solution of 0.147g indole-2, 3-dione (Isatin) (0.01mol) were dissolved in ethanol and refluxed at 65°C for one hour and cooled to room temperature. A dark yellow crystalline precipitate was formed. The precipitate was recrystalised from ethanol.Yield81%.Rf: 0.57, m.p 265-267°C; IR(KBr)(cm^-1):1610 cm^-1 (C-N str), 3390 cm^-1(N-H str), 3070(C-H Straomatic).

Preparation of ethyl 2-(6H-indolo [3, 2-b] quinoxalin-6-yl) acetate(M2):

0.01 mol of 6H-Indolo (2, 3 b) quinoxaline was dissolved in dry acetone (50 ml) and a solution of ethylchloroacetate( 0.01 mol) was added in presence of anhydrous potassium carbonate. The resulting mixture was refluxed on for 13 hrs. The solution was evaporated to dryness and the residue was washed with ethanol (50ml), filtered and recrystallized from methanol to give the desired yellow crystalline product. Yield 68%, Rf:0.61,m.p 210-212°C; IR(KBr)(cm^-1):1610 cm^-1 ( C-N str), 1217 cm^-1(C-O str), 3058(C-H Straomatic), 1739 cm^-1(C-O str ester).

Synthesis of 2-(6H-indolo [3, 2-b] quinoxalin-6-yl) acetohydrazide(M3):

Ethyl 2-(6H-indolo [3, 2-b] quinoxalin-6-yl) acetate (0.01 mol, 3 g) and hydrazine hydrate (0.01 mol, 0.9 ml) were taken in solvent 1, 4-dioxan (50 ml) and refluxed for 5 hrs at 60-70°C until Pale yellow solid mass had appeared which was filtered, dried and recrystallized with ethanol.yield:61%, Rf: 0.54, m.p 282-284°C; IR(KBr)(cm^-1):3274 cm^-1 (N-H str), 1693 cm^-1(C-O str), 3047(C-H Straomatic), 2921 cm^-1(C-H str aliphatic).

Synthesisof (e)-n'-arylidene2-(6h-indolo [3, 2-b] quinoxalin-6-yl) acetohydrazide(M4 a-h):

2-(6H-indolo [3, 2-b] quinoxalin-6-yl) acetohydrazide (0.01 mol, 2.91 g) (03) and the aromatic aldehyde were dissolved in 30 ml methanol with vigorous stirring at 40-50˚C. Glacial acetic acid was added and allowed to reflux for further 3 hrs at 40°C. The mixture was poured into ice-cold water and stirred. The precipitate obtained was filtered, dried and recrystallized from chloroform.

Representative physical and Spectral data^17,18:

4a: m.p 328-330, yield 54%, IR (KBr) (cm^-1): 1610 cm^-1(C=O str), 1693 cm^-1, 1436. Cm-1(C=N str), 2979 cm^-1, 2927 cm^-1 (Aliphatic CH str). 3089 cm^-1, 3143 cm^-1 (Ar CH str). 3018 cm^-1(N-H str), 1103 cm^-1 (Ar C-Cl str).

4b:m.p 298-300, yield 55%,IR (KBr) (cm^-1) :3236 cm^-1 (N-H str),1739 cm^-1(C=O str), 3161cm^-1(Ar CH str),1583 cm^-1 (C=N) 1427 cm^-1(imine),3018 cm^-1, 2977 cm^-1(Aliphatic CH str),1427 cm^-1, 1407 cm^-1(Aliphatic CH str). 4c:m.p 326-328, yield 53%, IR (KBr) (cm^-1)1407 cm^-1( C=N str Imine) 1521 cm^-1,1344 cm^-1 (Aliphatic CH str),3020 cm^-1, 3078 cm^-1 (Ar CH str), 3145 cm^-1 (N-H str),1134 cm^-1(C-Cl str Ar).

4d:m.p 245-247, yield 59%, IR (KBr) (cm^-1) : 3399 cm^-1(N-H str),2979 cm^-1(Ar CH str),1679 cm^-1(C=O str),1514 cm^-1 (C=N Imine) 2931 cm^-1(Aliphatic CH str),1436 cm^-1(Aliphatic CH str)1577 cm^-1,1234 cm^-1(O=N=O str). 4e:m.p 267-269, yield 54%, IR (KBr) (cm^-1):3137 cm^-1 (N-H str),3072 cm^-1(Ar CH str),1677 cm^-1 (C=O str),1550 cm^-1 (C=N str Imine),3041 cm^-1, 2991 cm^-1(Aliphatic CH str), 1487 cm^-1, 1598 cm^-1,1232cm^-1(O=N=O str).

4f:m.p 216-218, yield 65%, IR (KBr) (cm^-1): 2900 cm^-1(Ar CH str),1683 cm^-1(C=O str),1406 cm^-1(C=N str Imine), 3003 cm^-1(Aliphatic CH str),1488 cm^-1, 1330 cm^-1 (Aliphatic CH str),3096 cm^-1 (O-H str). 4g:m.p 196-197, yield 57%, IR (KBr) (cm^-1):2995 cm^-1 (Ar CH str),1618 cm^-1(C=O str),1407 cm^-1 (C=N str),1514 cm^-1(C=N str Imine),1211 cm^-1(C-O-C Asym)1180 cm^-1(C-O-C Sym)3080cm^-1 (O-H str). 4h:m.p 228-230, yield 64%, IR (KBr) (cm^-1):3143 cm^-1 (N-H str), 2344 cm^-1 (Ar CH str), 1591 cm^-1 (C=N str),1519 cm^-1 (C=N str Imine), 2937 cm^-1 ( C-H str)2937 cm^-1 ( C-H str)1366 cm^-1 (Aliphatic CH str)1366 cm^-1 (Aliphatic CH str)1519 cm^-1 (C=N str Imine).

Synthesis of n-(3-chloro-2-oxo-4-arylazetidin-1-yl)-2-(6h-indolo[3,2-b] quinoxalin-6-yl) acetamide. (M5 a-h).

A solution of Chloro-acetyl chloride (1.12ml 0.01M) in 1,4-Dioxan was added drop wise to a well stirred solution of (E)-N'-Arylidene-2-(6H-indolo [3, 2-b] quinoxalin-6-yl) acetohydrazide (0.01M) and Triethylamine (0.02M) in 1,4-Dioxan. After the addition had been completed, the solution was stirred for 24 hrs. The reaction mixture was added to ice cold water. The separated solid was filtered and purified from 1, 4-Dioxan: water (80:20). Physical data included in table-1 and spectral data included in table-2.

Figure 1: Synthetic scheme

Table-1physical data of compounds (5a-h)

compound	R	Molecular formula	Mol. weight	M.P(^oc)	%yield	Rf value
5a	2,4 -di chloro	C₂₅H₁₆Cl₃N₅O₂	524	323	59	0.66
5b	H	C₂₅H₁₈ClN₅O₂	456	289	50	0.54
5c	2,6- di chloro	C₂₅H₁₆Cl₃N₅O₂	524	329	55	0.62
5d	2- nitro	C₂₅H₁₇ClN₆O₄	500.9	241	56	0.60
5e	4-nitro	C₂₅H₁₇ClN₆O₄	500.9	242	64	0.59
5f	2-hydroxy	C₂₅H₁₈ClN₅O₃	471	220	59	0.64
5g	3-ethoxy,4-hydroxy	C₂₇H₂₂N₅O₄	515.5	193	62	0.65
5h	4-dimethyl amino	C₂₇H₂₃ClN₆O₂	499	224	66	0.63

Table-2 : Spectral data of compounds 5a-h

Compound	IR spectral data(cm^-1)	¹H NMR data d (ppm)	Mass Spectral data
5a	1602, 1714 (C=O str),1627( C=N str), 3016,3056 (Ali CH str) 1508, 1456(Ali CHstr),3132,3167 (Ar CH str),3267 (N-H str)796(C-Cl str)	4.1 (2H, CH₂),5.3 (1H, CH),5.6 (1H, CH),7.3-7.9 (7H, Ar),8.1-8.7 (4H, Ar) 10.5 (1H, NH)	525(M⁺)
5b	1772,1716 (C=O str),1583( C=N str),2836,3062(AliCHstr),1496,1394(Ali CH str)2979(Ar CH str)3062(N-H str)	4.0 (2H, CH₂),5.3(2H,CH) 7.2-7.9 (7H, Ar),8.1-8.4 (6H, Ar) 10.3 (1H, NH)	455(M⁺)
5c	1741,1716.53 (C=Ostr),1568,1618(C=N str)2918,2956 (Ali CH str), 1523, 1344(Ali CH str),3076,3215(Ar CH str), 3280(N-H str),748(C-Cl str)	4.3 (2H, CH₂),5.3 (2H, CH)7.1-7.8 (7H, Ar)8.0-8.4 (4H, Ar) 10.4 (1H, NH)	524(M-1)
5d	3134(N-H str),3016 (Ar CH str) 1710,1766(C=O str),1594 (C=N),2977 (Ali CH str),1485,(O=N=O str)792(C-Cl str	4.2 (2H, CH₂),5.6 (1H, CH)5.8 (1H, CH)7.4-7.8 (8H, Ar)8.1-8.4 (4H, Ar) 10.9 (1H, NH)	501(M⁺)
5e	3134 (N-H str),3068(Ar CH str)1712,1654(C=O str)1519 (C=N str) 30683, 1519 (Aliphatic CH str)1602,1238 (O=N=O str)	4.3 (2H, CH₂)5.6 (1H, CH)5.7 (1H, CH)7.3-7.9 (8H, Ar)8.0-8.3 (4H, Ar) 10.6 (1H, NH)	500 (M-1)
5f	3010 (N-H str),2936 (Ar CH str) 1685, 1737.74 (C=O str),1596 (C=N str) 3096, 1407 (Ali CH str),3120(O-H str)	4.2 (2H, CH₂)5.3 (1H, CH)5.6 (1H, CH)6.8 (2H, Ar)7.0-7.9 (9H, Ar)8.1 (1H, Ar) 8.9 (1H, OH) 10.7 (1H, NH)	472(M⁺)
5g	2985 (Ar CH str),1614,1710(C=O str),1410 (C=N str),1216(C-O-C Asym),3141 (O-H str) 970(C-Cl)	1.6 (3H, CH₃)4.2 (2H, CH₂)4.6 (2H, CH₂)5.4 (1H, CH)5.5 (1H, CH) 7.1-7.7 (8H, Ar)8.0-8.5 (3H, Ar) 9.3 (1H, OH) 10.5 (1H, NH)	516 (M⁺)
5h	3062 (N-H str),2995 (Ar CH str) 1616,1706 (C=O str),1595 (C=N str) 2879,2974(AliCH str),1458,1488(Ali CH str)	2.8(6H, CH₃),4.7 (2H, CH₂),5.2 (1H, CH),5.6 (1H, CH)6.8 (1H, Ar)7.0-7.8 (10H, Ar)10.5 (1H, NH)	498(M⁺)

Table 3: Antimicrobial activity of compounds (5A-5H)

S. No.	Compound	Antibacterial activity zone of inhibition in (mm)				Antifungal activity zone of inhibition in (mm)
S. No.	Compound	S.aureus (Gram +ve)	B.subtilis (Gram +ve)	Klebsiella (Gram -ve)	P. vulgaris (Gram -ve)	Aspergillusniger	Candida albicans
1	5A	16	18	15	16	11	10
2	5B	13	11	12	13	06	06
3	5C	15	17	15	17	09	10
4	5D	13	15	14	16	08	08
5	5E	15	15	16	14	06	09
6	5F	15	13	13	13	14	12
7	5G	18	20	13	11	09	10
8	5H	14	15	13	13	10	13
9	Amoxicillin	36	30	32	30	-	-
10	Ciprofloxacin	40	37	35	35	-	-
11	Fluconazole	-	-	-	-	32	32
12	Amphotericin B	-	-	-	-	29	25
12	Control (DMSO)	NI	NI	NI	NI	NI	NI

Note: All the values are mean of triplicates NI: no inhibition - : not tested

The standards used are Ciprofloxacin and Amoxicillin. All compounds have shown antibacterial activity. Compound 5G has highest activity among all the derivatives against gram-positive bacteria. Compound 5c has highest activity among all the derivatives against gram-negative bacteria.

(b) Antifungal activity:

The antifungal activity of newly synthesized derivatives has been evaluated against Aspergillusniger and C. albicans. The standards used are Fluconazole and Amphoterericin B. All compounds have shown antifungal activity.

CONCLUSION:

In conclusion, several substituted N-(3-chloro-2-oxo-4 aryl azetidine-1-yl)-2-[6 H-indolo(2,3 b) quinoxalin-6-yl] acetamides [5A-H] were synthesized. The biological evaluation was undertaken to evaluate the effect of substitutions on antimicrobial activity. Para nitro and para hydroxyl derivatives are good antimicrobial agents. The derivatives of Azetidinones tethered with indolo (2, 3-b) quinoxaline moiety show reasonable antimicrobial activity. With these encouraging results, all the synthesized compounds can be further explored for structural modification and detailed microbiological investigations to arrive at possibly newer potent antimicrobials.

ACKNOWLEDGEMENT:

The authors are thankful to Gokula Education Foundation and M.S. Ramaiah College of Pharmacy, Bangalore for providing necessary facilities.

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Received on 06.11.2015 Modified on 23.11.2015

Asian J. Research Chem. 8(12): December 2015; Page 716-720

DOI: 10.5958/0974-4150.2015.00115.7