Synthesis and Biological Screening of some Novel 3-Subsituted Indole Derivatives

 

Raju Kundavaram¹*, M. Gayathri Devi¹, B. Siva Kumar², T. Archana³, M. Gowri Manoja⁴,

Dr. J. Naga Sai Kumar⁵

¹Assistant Professor, Pharmaceutical Chemistry, Karnataka College of Pharmacy, Bangalore.

²Creative Educational Society College of Pharmacy, Chinna Tekuru, Andhra Pradesh.

³Assistant Professor, Pharmaceutics, Karnataka College of Pharmacy, Bangalore.

⁴Assistant Professor, Pharmaceutical Analysis and Quality Assurance, Anurag Group of Institutions, Hyderabad.

⁵Assistant Professor, Pharmacy Practice, Karnataka College of Pharmacy, Bangalore.

 

ABSTRACT:

In the present work Indole and substituted glycolic acid have been chosen as starting materials. These are broad class of chemical compounds, with many important pharmacological properties. Because of ease of reaction with electrophiles, indole is widely used in various pharmaceutical preparations. The present investigation was concerned with synthesis of some novel 3-substituted indole derivatives with objective of discovery novel and potent analgesic, anti-inflammatory and anti-oxidant agents. These compounds were synthesized by cyclisation of indole-3-acetic acid with different aromatic amines. The structures of new compounds were confirmed by FT IR, 1HNMR and Mass. These compounds were tested in-vivo for analgesic, anti-inflammatory activates and biochemical studies antioxidant activity. The statistical analysis was carried out by ANOVA. All the compounds tested (NHR-I-NHR-VII) showed analgesic, anti-inflammatory and antioxidant activities. Among seven compounds three compounds (NHR-I, NHR-II, NHR-IV) showed significant analgesic, anti-inflammatory and antioxidant activities. Compounds NHR-II and NHR-V showed 74.38% and 70.68% inhibition of paw oedema in anti-inflammatory activity compared to 75.28% inhibition of standard Indomethacin. Compounds NHR-I and NHR-II showed 68.84% and 73.30 % inhibition against acetic acid induced writings’ in analgesic activity compared to 75.23% inhibition of standard Diclofenac. Compound NHR-II showed 71.6% inhibition of DPPH radical in Anti-oxidant activity compared to 84% inhibition of standard Ascorbic acid. The results showed that incorporation of appropriately substituted aromatic amines at third position of Indole nucleus can afford good analgesic, anti-inflammatory and antioxidant with reduced side effects of compounds.

 

 

 

INTRODUCTION:

INDOLES

Indole is an aromatic, heterocyclic, organic compound. It has a bicyclic structure, consisting of a six-membered benzene ring fused to a five-membered pyrrole ring at 2,3 – position. The name indole is portmanteau of the words indigo and oleum, since indole was first isolated by treatment of the indigo dye with oleum.

 

Indole chemistry began to develop with the study of the dye indigo found in many natural products such as the indole alkaloids, fungal metabolites and marine natural products which is benzopyrrole in which the benzene and pyrrole rings are fused through the 2- and 3-positions of the pyrrole nucleus. The IUPAC name of the indole is 1H-benzo (b) pyrrole, it is being the b-face benzo-fused isomer.

 

Indole is a solid at room temperature, can be produced by bacteria as a degradation product of the amino acid tryptophan. It also occurs in coal tar. Indoles are basic in nature and form salts with acids (Morrin Acheson et al., 2009)^1,2. Pandeya et al. synthesized a series of p-nitro phenyl substituted semicarbazones. The newly synthesized derivatives were screened for their anticonvulsant activities against maximal electroshock (MES), subcutaneous pentylenetetrazole (scPTZ) and subcutaneous strychnine (scSTY) tests. (Pandeya et al., 1999)³. Gerard A. Pinna et al. investigated various new derivatives and structural analogues of N-(1-ethyl-2-pyrrolidinylmethyl)-4,5-dihydro-1H-benzo [g] indole-3-carboxamide, a representative term of a series of 2-aminomethyl pyrrolidinyl derived 4,5-dihydrobenzo[g] indole carboxamides with good D₂-like affinity, and evaluated for their ability to bind to dopamine D₂-like receptors in vitro. From these studies, compound (2-chloro-N-(1-ethyl-2-pyrrolidinylmethyl)-5, 6-dihydro-4H benzo [6, 7] cyclohepta [b] pyrrole-3-carboxamide), was found to possess a potent affinity for D₂-like receptors. (Gerard A. Pinna et al., 2002)⁴. Istvan Borza et al., synthesized a novel series of indole-2-carboxamide derivatives and identified as NR2B selective NMDA receptor antagonists. The influence of the number and position of OH groups on the indole skeleton as well as the substitution of the piperidine ring on the biological activity of the compounds was studied. From the results the compound was found to be more potent among the all derivatives. (Istvan Borza et al., 2003.)⁵.

 

Able, E et al., synthesized some istatin and indole oximes and evaluated for their anti-hypertensive activity. (Able, E et al., 2003)⁶, VitoBoido et al., reported A set of ten 2-phenyl-3-(quinolizidin-1-yl)-5-substituted indoles was prepared through the Fischer cyclization of lupin and epilupinylphenylketone4-substituted phenyl hydrazones. Compounds were tested for antiaggregating activity on human platelets activated by adenosine diphosphate (ADP), collagen and adrenaline. (VitoBoido et al.,2004)⁷. Girolamo Cirrincione et al., reported the Eight derivatives of the new ring system [1,2,3,5] tetrazino [5,4-a] indole-4-one 7, were synthesised in good yields by reaction of 2-diazoindoles with alkyl or aryl isocyanates. Compounds 7 were screened for their activity against human tumour cell lines. (Girolamo Cirrincione et al., 2005)⁸.

 

Importance of the proposed investigation:

Indoles found to have analgesic, anti-inflammatory, antioxidant activities which are potent molecules and hence indole derivatives can effectively treat these diseases. The starting materials in the present work are Indole and substituted glycolic acid which are broad class of chemical compounds, with many important pharmacological properties. It is widely used in various pharmaceutical preparations. Therefore, various efforts have been made to produce new compounds with different properties and least possible unwanted effects. The present investigation mainly concerned with synthesis and characterization of some novel 3-substituted Indole derivatives and evaluated for their potent analgesic, anti-inflammatory and anti-oxidant activities with least possible unwanted effects.

 

MATERIAL AND METHODS:

Research Design:

To start the work, initially literature survey has been done. After literature survey, a general scheme was prepared as follows.

 

 

Indole and substituted glycolic acid have been chosen as starting materials. The formations of the final products were monitored by TLC. The completed products showed significant color under UV chamber. All the compounds prepared were purified by recrystallization with suitable solvents. Structures of the synthesized compounds have been characterized and confirmed by IR, NMR and Mass techniques.

 

Chemicals and Instruments:

The chemicals used in this project was obtained from SD Fine chemicals, the instruments such as FT-IR was of Shimadzu make, NMR of Joel D-3000, and UV Spectrophotometer of Labo Med Inc 2602.

Table No. 1: Summary of the synthesized compounds

S. No.	Compound code	Molecular formula	Molecular weight	Colour	Solubility	Melting point	% of Yield
1	NHR-I	C16H15N2O	250	Light buff	Chloroform, DMSO	126-128°C	78
2	NHR-II	C15H13N2OCl	284.5	Buff	Chloroform, Ethanol	80-82°C	80
3	NHR-III	C17H16N2o2	280	Dark brown	Chloroform, DMSO		79
4	NHR-IV	C17H16N2O	264	Brick red	Chloroform, acetone	166-167°C	83
5	NHR-V	C22H18N4O	354	Dark yellow	Chloroform, acetone. DMSO	130-132°C	80

 

RESULTS:

Synthetic methodology (table-1).

 

Spectral results: The synthesized compounds were characterized by IR, NMR and Mass spectral studies IR spectra were recorded on SHIMADZU as KBr disc and Mull technique (γ cm^-1).¹H-NMR spectra were recorded on Bruker AC 80 MHz Spectrophotometer using CDCl₃ as an internal standard (chemical shift in δ, ppm).Mass spectra were recorded on an Electron impact Mass spectrometer at 70 eV using direct insertion probe. The molecular weight was calculated in positive mode by standard addition method.

 

1.     Compound: 2-(1H-indol-3-yl)-N-phenylacetamide

Code: NHR-I

 

Fig. 1: FT-IR, ¹H NMR, Mass Spectra of Compound NHR-I

 

2.     Compound: N-(3-chlorophenyl)-2-(1H indole-3-yl) acetamide

Code: NHR-II

 

 

Fig. 2: FT-IR, ¹H NMR, Mass Spectra of Compound NHR-I

 

3.     Compound: 2-(1H-indole-3-yl)-N-(3-methoxyphenyl) acetamide

Code: NHR-III

 

Fig. 3: FT-IR ¹H NMR Spectra of Compound NHR-III

 

Table No. 2: IR and ¹H-NMR and Mass spectral data of compound NHR-I, II, III

Data of Compound NHR-I
S. No.	FTIR		1H-NMR		Mass
	Vibrational moiety	Wave number, γ (cm-1)	Chemical shift (δ, ppm)	Nature of proton	Data of positive addition method	Molecular weight
1.	Indole N-H stretch	3432	3.928	2H, s, CH2	(M + BP) = 250	250
2.	N-H stretch of RCONHR1	3276	7.0-7.6	10H, s, of Aromatic ring
3.	C-H stretch in aromatic ring	2854	8.2	1H, br-s, NH
4.	C-H stretch in aliphatic ring	2930	8.4	1H, br-s, NH
5.	C=O stretch in RCONHR1	1654
Data of Compound NHR-II
S. No.	FTIR		1H-NMR		Mass
	Vibrational moiety	Wave number, γ (cm-1)	Chemical shift (δ, ppm)	Nature of proton	Data of positive addition method	Molecular weight
1.	Indole N-H stretch	3372	3.928	2H, s, CH2 Aliphatic ring	(M + BP) = 250	250
2.	Br N-H stretch of RCONHR1	3314	7.0-7.6	9H, m, of Aromatic ring
3.	C-H stretch in aromatic ring	2854	8.1	1H, br-s, NH
4.	C-H stretch in aliphatic ring	2930	8.3	1H, br-s, NH
5.	C=O stretch in RCONHR1	1654
Data of Compound NHR-III
S. No.	FTIR		1H-NMR		Mass
	Vibrational moiety	Wave number, γ (cm-1)	Chemical shift (δ, ppm)	Nature of proton	Data of positive addition method	Molecular weight
1.	Indole N-H stretch	3413	3.89	3H, s, CH2 Aliphatic ring	(M +) = 281 Base peak	281 277
2.	Br N-H stretch of RCONHR1	3294	3.75	3H, s, of OCH3
3.	C-H stretch in aromatic ring	2854	8.1	1H, br-s, NH
4.	C-H stretch in aliphatic ring	2929	8.3	1H, br-s, NH
5.	C=O stretch in RCONHR1	1666	6.60	1H,m of Aromatic ring

 

Pharmacological Studies:

Analgesic and anti-inflammatory activities of synthesised compounds were evaluated by in vivo methods.

 

Analgesic Activity:

The analgesic activity of the compounds was evaluated by acetic acid induced writhing model and Eddy’s hot plate methods.

 

Results of Analgesic Activity using acetic acid induced writhing in Mice:

Analgesic effect of synthesized derivatives was measured in acetic acid induced writhing test. Data are expressed as the Mean ± SEM (n=5), one-way analysis of variance (ANOVA) followed by multiple comparison method. It is represented in the Fig. 4

 

Fig. 4: Bar diagram showing Analgesic activity results

 

Results of Analgesic Activity using Eddy’s Hot-Plate Test in Mice:

 

Table No. 3: Eddy’s Hot-Plate Test

S. No.	Compound Group	Response in minutes
		30min	60min	90min
1	Control	2±0.29	3.83±0.8	3±0.52
2	Standard (Diclofenac)	6±0.45ns	7±1.16**	9.16±0.76**
3	NHR-I	3.33±0.5ns	6.6±0.25*	5.83±0.7**
4	NHR –II	3.33±0.4**	6±0.2**	6.83±0.3**
5	NHR –III	4.66±0.5***	6.1±0.4**	8.33±0.33**

 

Effects of the all compounds on the latency time of mice exposed to the hot plate test were studied. All values are expressed as Mean ± Sem n=5, one way analysis of variance (ANOVA) followed by multiple comparison Dunnett’s test ^*P<0.005, ^**P<0.01, and^***P<0.001 as compared to control group. Some of the values are said to be NS (no significant)

 

Anti-Inflammatory Activity:

The anti-inflammatory activity of the compounds was evaluated in Carrageenan-induced paw method.

 

Results of Anti-inflammatory Activity in Mice using Carrageenan-induced paw oedema model in mice:

 

Table No. 4: Anti-inflammatory activity (Percentage inhibition of paw volume)

S. No.	Compound Group	% Inhibition of Paw oedema Time in minutes
		30min	60min	90min	180 min
1	Standard	77	79	83	75.28
2	NHR-I	56.74	55.61	48.03	62.99
3	NHR –II	74.38	72.70	73.51	74.38
4	NHR –III	41.97	26.03	21.05	62.73

 

Fig. 5: Graph showing Anti-inflammatory activity results

 

DISCUSSION:

The wide literature survey kept in view for the present research work, and it has been carried out to synthesize the 3-substituted indole acetamide and their evaluation for the analgesic, anti-inflammatory anti-oxidant activities.

“3-substituted some novel indole derivatives” were synthesized in two steps facile procedure. The reaction involves interaction of indole, Glycolic acid and substituted anilines. Seven derivatives were synthesized, characterized and evaluated for analgesic, anti-inflammatory and antioxidant activities. The purity of the title compounds were tested in the precoated TLC plates, structural characterization was performed by IR, 1H NMR and Mass. All the three derivatives were screened for their analgesic, anti-inflammatory and antioxidant activities. Synthesized derivatives were evaluated for analgesic activity in this activity, Diclofenac was used as standard drug.

 

ANALAGESIC ACTIVITY:

Acetic acid induced writhing model:

From the results the synthesized derivatives screened at 20 mg/kg body weight NHR-I has shown 68.8% of inhibition which is more potent when compare to control and less potent than standard (72.5%) due to unsubstituted aniline. The result of NHR-II revealed that 73.30% of inhibition which is less potent than standard Diclofenac may be presence of Chlorine group at Meta position might be favoured more potent analgesic activity.

 

NHR-III has given 61.6% inhibition which is less potent than all other derivatives and standard due to presence of electron releasing group like Methoxy at Meta position.

 

Eddy’s hot plate method:

All derivatives which are screen by Acetic acid induced model where also screened for their Analgesic activity using Eddy’s hot plate method at different time intervals (30, 60, and 90). The results at various reaction time of all the derivatives. As the reaction time increase the time taken for the jumping also increases. Among all the novel 3-substituted indole derivatives NHR-III have shown potent more activity when compared to control and almost equipotent to that of standard. This may due to presence of electron donating group Methoxy of NHR-III at Meta position.

 

ANTI-INFLAMMATORYACTIVITY:

Carrageenan-induced paw oedema model:

From the results of synthesized derivatives screened at 20 mg/kg body weight, NHR-I shown percentage of inhibition 62.9 which is shown to be more potent than control and less potent than standard that of 75.28% may be unsubstituted aniline. NHR-II revealed 74.38% of inhibition which shows that it is equipotent to that of standard Indomethicin (75.28%) may be presence of Chlorine group at meta position. NHR-III has given 62.73% of inhibition which is more potent.

 

SUMMARY:

Indole is a common component of fragrances and the precursor to many pharmaceuticals. The biological importance of Indole heterocyclic directly associated with their pharmacological and medical potential, Indole extremely attractive and rewarding research targets and has motivated countless researchers to study their synthesis and pharmacological properties. Indole posses’ wide range pharmacological activities like analgesic, antibacterial, antifungal anticancer anti-diabetic anti-inflammatory antioxidant etc, due to its pharmacological activities inspired us to synthesize novel 3-substituted Indole derivatives and screen for their anti-inflammatory analgesic and antioxidant activities respectively. The structures of novel compounds were confirmed by FT-IR, ¹HNMR and Mass. These compounds (NHR-I-NHR-III) were tested in-vivo for analgesic, anti-inflammatory activates and biochemical studies antioxidant activity. Compounds NHR-I and NHR-II showed 68.84% and 73.30 % inhibition against acetic acid induced writings’ in analgesic activity compared to 75.23% inhibition of standard Diclofenac. Compound NHR-II has showed 71.6% inhibition of DPPH radical in Anti-oxidant activity compared to 84% inhibition of standard Ascorbic acid. The results showed that incorporation of appropriately substituted aromatic amines at third position of Indole nucleus can afford good analgesic, anti-inflammatory and antioxidant with reduced side effects of compounds.

 

CONCLUSION AND FURTHER SCOPE OF RESEARCH:

All the title compounds (NHR-I to NHR-III) were synthesized and characterized by analytical data and screened for their analgesic, anti-inflammatory and antioxidant activates. Among all the derivatives, the compound NHR-II N-(3-chlorophenyl)-2-(1H indole-3-yl) acetamide) was found to be more potent analgesic when compared with all other derivatives and less potent than standard in acetic acid induced writhing model. The compounds NHR-III and has shown more potent analgesic activity when compare to other derivatives and less potent than standard in Eddy’s hot plate method. The derivatives NHR-II N-(3-chlorophenyl)-2-(1H indole-3-yl) acetamide) was found to be more potent when compared with all other derivatives and less potent than standard in the Carrageenan-induced paw oedema model.

 

ACKNOWLEDGEMENTS:

I am very grateful to my parents who have struggled their present days for the wealth of my tomorrow, who shows their blessings, love and encouragement in every activity of me which gave strength and power to achieve my goals, even in hard times.

 

With great pleasure I acknowledge my sincere thanks to honorable chairman, honorable secretary and founder of Karnataka Educational Trust Prof. Basawaraj Ramnal and our Director Dr K. Ramesh for granting us the permission to carry out the research work. I express my heartfelt thanks to Miss. Julisa Suwail, and Mrs. M. Gayathri Devi for their support to complete this work.

 

REFERENCES:

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Received on 24.12.2019           Modified on 05.03.2020

Accepted on 06.05.2020         © RJPT All right reserved