Synthesis and Biological Evaluation of Naphthylmethyl-1, 3, 4- Oxadiazoles for Antioxidant and Antibacterial Activities

 

Soumya J and Rajitha G*

Institute of Pharmaceutical Technology,

Sri Padmavathi Mahila Viswavidyalayam (Women’s University), Tirupati-517502, Andhra Pradesh, India

*Corresponding Author E-mail: rajitha_galla@yahoo.com

A Series of 2-((naphthalen-1-yl) methyl)-5-substituted-1,3,4-oxadiazole derivatives were synthesized by refluxing naphthyl acetic acid hydrazide with substituted aromatic acids in presence of phosphorous oxychloride. The intermediate naphthyl acetic acid hydrazide (2) was prepared by treatment of naphthyl acetic acid ester (1) with hydrazine hydrate. The chemical structures of all the twelve synthesized compounds were confirmed by means of IR, ¹H NMR and Mass spectral data. All the title compounds were screened for antioxidant activity by Nitric oxide scavenging activity, reduction of DPPH and antibacterial activity using Cup plate method. Among the series of the compounds  4-hydroxy-3,5-dimethoxy (3k), 3-pyridyl (3h), 4-pyridyl (3i) derivatives showed  good antioxidant activity in two invitro models. 4-chloro (3b)  4-hydroxy-3,5-dimethoxyderivative(3k), 4-pyridyl (3i) and 3-Pyridyl (3h) derivatives showed good antibacterial activity.

 

 

 

INTRODUCTION:

Substituted 1,3,4-Oxadiazoles were reported to possess several interesting biological activities such as antimicrobial¹, anticancer², antioxidant activity³, antiinflammatory⁴, anti-fungal⁵, antimicrobial⁶ and anti-tubercular⁷ activities etc. These findings prompted us to synthesize novel 1,3,4-oxadiazoles with the hope of obtaining compounds with potent biological activity.

 

 

 

MATERIALS:

Carboxylic acids were procured from Merck, SD fine chemicals, Aldrich and Sigma. All other chemicals are of AR grade. All Melting points were determined using open capillary tube method and were uncorrected. The purity of the compounds was monitored by ascending thin layer chromatography (TLC) on silica gel G (Merck Kieselgel 60 F₂₅₄) and visualized by iodine vapour. Developing solvents were chloroform: methanol (9:1). IR spectra were recorded using KBr on FTIR spectrometer and proton nuclear magnetic resonance (¹H NMR) spectra were recorded on Bruker (400 MHz) Avance in DMSO-d6. Chemical shifts are reported in parts per million (ppm) using tetramethylsilane (TMS) as an internal standard. Mass spectra were performed on Agilent 6310 Ion Trap spectrometer under EI. The agar medium was purchased from HI Media Laboratories Limited, Mumbai, India.

 

METHODS:

Synthesis of naphthyl acetic acid ester (1):

A mixture of naphthyl acetic acid (0.246mol), absolute ethanol (2.5mol) and conc. H₂SO₄ (2.7ml) was refluxed for about 4h. The excess solvent was distilled off and allowed to cool and poured into crushed ice. The solution was extracted with carbon tetra chloride. CCl₄ layer was collected and sodium bicarbonate was added to remove excess acid, then filtered. CCl₄ layer was evaporated to get thick concentrated ester. Yield 86%;  bp 210⁰C.

Synthesis of naphthyl acetic acid hydrazide (2):

The ester (0.01mol) obtained in the earlier step and hydrazine hydrate (0.01mol) were dissolved in ethanol (20ml) and refluxed for about 3h.  The solid was separated by filtration and recrystallized from ethanol to afford naphthyl acetic acid hydrazide. Yield 70% mp 140⁰C.

 

General method of synthesis of naphthyl acetic acid oxadiazole (3a-3l):

A mixture of naphthyl acetic acid hydrazide (0.01mol)and appropriate aromatic acid (0.01 mol) were dissolved in  phosphorus oxy chloride and refluxed for 5 h .The mixture was cooled and poured into ice cold water with stirring. The mass obtained was filtered and washed with water and recrystallized from ethanol.

 

All the compounds (3a-3l) were prepared by similar procedure (Scheme). The physical data of these compounds such as melting point, recrystallization solvent and yield are given in Table 1.

 

 

Scheme

 

Table 1: Physical data of synthesized compounds of oxadiazole derivatives 3a-3l:

Compound	Ar	Formula	M.P(0C)	Yield (%)
3a		C19h13 n2o	144-146	70
3b		C19h13 n2ocl	148-150	65
3c		C19h13 n2o3	208-210	60
3d		C20h16 n2o2	142-144	70
3e		C21h18 n2o3	198-200	70
3f		C19h15n3o	198-200	65
3g		C19h15 n3o2	196-198	60
3h		C18h13 n3o	186-188	58
3i		C18h13 n3o	184-186	56
3j		C19h12 n2o5	214-216	60
3k		C21h18 n2o4	212-214	65
3l		C19h14 n2o2	180-182	60

Recrystallization solvent: Ethanol.

 

 

 

BIOLOGICAL EVALUATION:

Antioxidant Activity

1. Assay of Nitric Oxide (NO) scavenging activity⁸:

Sodium nitroprusside (10mM) in phosphate buffer pH 7.4; was incubated with 100 mM concentrations of drug dissolved in a suitable solvent (dioxane/methanol) and tubes were incubated at 25^oC for 120 minutes.  2 mL of incubation solution was removed and diluted with 2 mL of Griess reagent. The absorbance of the chromophore formed during diazotization of nitrite with sulphanilamide and on subsequent coupling with N-napthylethylene diamine was read at 546 nm. Control experiments without test compound were conducted in an identical manner.

 

2. Interaction with stable free radical DPPH⁹:

DPPH assay was performed as described. Solutions of various drugs at 100 mM concentration were added to 100 mM DPPH in 95% ethanol and tubes were kept at an ambient temperature for 20 minutes and absorbance was measured at 517 nm.  Ethanol was used as a blank solution and DPPH solution in ethanol served as the control. The results of antioxidant studies of (3a-3l) are given in Table 2.

Antibacterial Activity

The antibacterial activity of title compounds has been assayed by cup-plate method¹⁰ by measuring zone of inhibition against four different strains of bacteria. All the test compounds were screened for antibacterial activity against two gram-positive bacteria Bacillus subtilis, Staphylococcus aureus and two gram-negative bacteria Escherichia coli and Klebsiella pneumoniae at a concentration of 10mg/10ml. Streptomycin was used as standard drug at a concentration of 1mg/ml. Stock solutions of the synthesized compounds were prepared in concentrations of 100µg/ml using DMSO as solvent for anti bacterial activity. The zone of inhibition was measured using mm scale and the values of antibacterial activity of     (3a-3l) are given in Table 3.

 

Cup plate method: The nutrient agar medium was sterilized by autoclaving at 121ºC (15lb/sq inches) for 15min. The petriplates and other required glassware was sterilized in hot air oven at 160ºC, for an hour. Into each sterilized petriplate (10cm diameter), about 20ml of molten nutrient agar medium was poured and inoculated with the respective strain of bacteria (6ml of inoculum to 300mlof nutrient agar medium) was transferred aseptically.

 

DMSO was used as a solvent control. Laminar airflow bench was swapped with 70% alcohol and UV lamp was switched on. After 30min, the UV lamp was switched off. All the reagents, media, inoculums and glassware were placed in laminar airflow bench observing all aseptic conditions. The plates were incubated within minutes of preparation of suspension, so that density does not change. A sterile cotton swab over was dipped into the suspension and the medium was inoculated by even streaking of the swab over the entire surface of the plate in three directions. After inoculums had dried, cups of diameter 6mm were made in the agar plate with a sterile cork borer. The test samples were added to these cups with a micro pipette and plates were then incubated at 37ºC for 24hrs.

 

 

Table 2: Antioxidant activity of 2-((naphthalen-1-yl) methyl)-5-substituted-1,3,4 oxadiazoles:

Compound	Ar	Nitric oxide scavenging activity at 100µM	Reduction of DPPH  at 100µM
3a		42	45
3b		48	45
3c		50	46
3d		50	50
3e		48	48
3f		48	38
3g		50	45
3h		63	60
3i		62	58
3j		40	50
3k		65	62
3l		60	55
		80	78

       

                                              

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

Table 3: Antibacterial activity of 2-((naphthalen-1-yl) methyl)-5-substituted-1,3,4 oxadiazoles (3a-3l):

Zone of inhibition of test compounds at a concentration of 100µg/mL was measured

B. subtilis: Bacillus subtilis   E. coli: Escherichia coli; K. pneumoniae: Klebsiella pneumonia    S. aureus: Staphylococcus aureus.

 

 

 

RESULTS AND DISCUSSION:

All the derivatives 3a-3l were synthesized as per scheme. The final products were purified by the recrystallization techniques with ethanol. The structures of these compounds were established by IR, ¹H NMR and Mass spectral data. The spectral studies of the synthesized compounds were given below.

 

Antioxidant activity:

The newly synthesized compounds (3a-3l) were evaluated for in vitro antioxidant activity by Nitric oxide scavenging activity and reduction of DPPH. Among all the compounds  4-hydroxy-3,5-dimethoxy (3k), 3-pyridyl (3h), 4-pyridyl (3i) derivatives showed  good antioxidant activity in both the two invitro models.

 

Antibacterial activity:

The compounds 4-chloro (3b), 4-hydroxy-3,5-dimethoxyderivative (3k), 3-pyridyl (3h) and 4-pyridyl (3i) derivatives showed good activity against Bacillus subtilis, Escherichia coli and Klebsiella pneumonia and compounds with 4-nitro (3c), 4-hydroxy (3l), derivatives showed moderate activity against Bacillus subtilis, Escherichia coli and Klebsiella  pneumoniae.

 

SPECTRAL DATA:

Compound 3a: 2-((naphthalen-1-yl) methyl)-5-phenyl-1,3,4-oxadiazole

IR (KBr cm^-1): 2928 (Ar C-H), 1697 (C=N), 1167 (C-O-C); ¹H NMR (d ppm): 4.01 (s, CH₂, 2H), 7.41- 7.92 (m, ArH, 12H); MASS: m/z: 286(M⁺).

 

Compound 3b: 2-((naphthalen-1-yl) methyl)-5-(4-chlorophenyl)-1,3,4-oxadiazole 

IR (KBr cm^-1): 2969 (Ar C-H), 1643 (C=N), 1122 (C-O-C), 673 (C-Cl); ¹H  NMR (d ppm): 4.04 (s, CH₂, 2H), 7.46- 7.96 (m, ArH, 12H); MASS: m/z: 286(M⁺).

 

Compound 3c: 2-((naphthalen-1-yl) methyl)-5-(4-nitrophenyl)-1,3,4-oxadiazole

IR (KBr cm^-1): 2853(ArC-H), 1664(C=N), 1083(C-O-C), 1521(N-O); ¹H NMR (d ppm): 4.06 (s, CH₂, 2H), 7.48- 7.98 (m, ArH, 12H); MASS: m/z: 286(M⁺).

 

Compound 3d: 2-((naphthalen-1-yl) methyl)-5-(4-methoxyphenyl)-1,3,4-oxadiazole

IR (KBr cm^-1): 2928 (ArC-H), 1697 (C=N ), 1167 (C-O-C).

 

Compound 3e: 2-((naphthalen-1-yl) methyl)-5-(3,4-dimethoxyphenyl)-1,3,4-oxadiazole

IR(KBr cm^-1): 2924 (ArC-H ), 1177 (C-O-C).

 

Compound 3f: 2-((naphthalen-4-yl) methyl)-5-(4-amino phenyl)-1,3,4-oxadiazole

                              IR (KBr cm^-1): 3331 (NH), 1170 (C-O-C); ¹H NMR (d ppm) 4.14 (s, -CH₂, 2H), 7.03-7.93 (m, ArH, 11H), 4.04 (s, - NH_2, 2H); MASS: m/z: 302(M⁺).

 

Compound 3g: 2-((naphthalen-1-yl) methyl)5-(2-hydroxy-5-aminophenyl)-1,3,4-oxadiazole

                              IR (KBr cm^-1): 3252(phenolic-OH), 1165(C-O-C); ¹H NMR (d ppm): 4.13(s,-CH₂, 2H), 7.15-7.96 (m, ArH, 10H), 4.04(s, - NH₂, 2H); MASS: m/z: 318(M⁺).

 

Compound 3h: 2-((naphthalen-1-yl) methyl)-5-(pyridin-3-yl)-1,3,4-oxadiazole

                           IR (KBr cm^-1): 2942(ArC-H), 1168(C-O-C); ¹H NMR (DMSO-d6, 400 MHz) d ppm: 4.02(s, -CH₂, 2H), 6.9-8.4 (m, ArH, 11H); MASS: m/z: 288(M⁺).

 

Compound 3i: 2-((naphthalen-1-yl) methyl)-5-(pyridin-4-yl)-1,3,4-oxadiazole

IR (KBr cm^-1): 2926(ArC-H ), 1641(C=N), 1155(C-O-C).

 

 

Compound 3j: 2-((naphthalen-1-yl) methyl)-5-(3,5-dinitrophenyl)-1,3,4-oxadiazole

IR(KBr cm^-1): 2885(ArC-H), 1627(C=N), 1174(C-O-C), 1545(N-O).

Compound 3k: 2-((naphthalen-1-yl) methyl)-5-(3,5-dimethoxy-4-hydroxyphenyl)- 1,3,4-oxadiazole

IR(KBr cm^-1): 3452(phenolic-OH ),2942 (ArC-H ), 1130(C-O-C).

Compound 3l: 2-((naphthalen-1-yl) methyl)-5-(4-hydroxy phenyl)- 1,3,4-oxadiazole

IR(KBr cm^-1): 3482 (phenolic-OH ), 1648 (C=N), 1121(C-O-C).

 

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Received on 23.12.2014         Modified on 10.01.2015

Accepted on 20.01.2015         © AJRC All right reserved