Synthesis and Antibacterial Evaluation of Some Oxadiazole Derivatives

ANTIMICROBIAL ACTIVITY OF SOME NEW OXADIAZOLE

DERIVATIVES

Zuhair Muhi-eldeen1 , Ghada Juma’a 2, Elham Al-kaissi 1, Lina Nouri 3,

1College of Pharmacy, University of Petra, Amman – Jordan

2College of Pharmacy, University of Kufa, Kufa, Iraq

3College of Pharmacy, University of Baghdad, Baghdad, Iraq

• Department of Pharmaceutics, School of Pharmacy, Petra University, P.O. Box 961343 Amman, Jordan, Fax 00962-6-5715551, Tel. 00962-6-05715546

E. mail:

ABSTRACT

Alkyl, alkenyl, sulfonyl, thiocarbamates and Mannich derivatives were synthesized and characterized through IR, NMR, and Elemental analysis. It is of interest to report the isomerization rearrangement of propynyl to allene group in Mannich reaction under basic condition.

The most promising compound as antibacterial agent was 5-(pyridyl)-1, 3, 4-oxadiazole-2-benzylthiocarbamates.

Keywords: Mannich derivative, Antibacterial, Isomerization, Propynyl, Allene

INTRODUCTION

A large number of oxadiazole derivatives have been prepared and many of these compounds have shown a wide spectrum of antimicrobial activity (1-4). The observation that some oxadiazoles with different substituents at different location on the heterocyclic ring resulted in fungicidal (5, 6) and antibacterial agents (7- 10) of various potencies. Despite a number of antimicrobial agents available for treatment of microbial infections, emergence of multi-drug resistant organism has posed a great challenge to the scientists (11-17) for continuous looking for new antimicrobial agents. This promoted our interest to synthesize a new series of substituted 2- mercapto – 5 – phenyl or 5 – (4 – pyridyl) – 1,3,4 – oxadiazole derivatives in which the mercapto group was converted to alkyl, alkenyl , alkynyl, sulfon, thiocarbamate and mannich derivatives as a new compounds of possible antimicrobial activity and to investigate the influence of unsaturation on their activity profile.

MATERIALS AND METHODS

Experimental

Melting point was determined by using a calibrated Thomas-Hoover melting apparatus. IR spectra were recorded using a Perkin – Elmer 257 spectrophotometer; NMR spectra were carried out on Varian EM-390, 90MHZ spectrometer using tetramethylsilane as the internal reference.

Microanalyses were performed in the Laboratories of Dr. BERNHARDT, Mulheim, West Germany and in the Laboratories of the Oil Exploration Company; Iraq.

The analyses are shown in (Table 4, 5, 6) or included in the experimental part and indicated only by symbols of the elements analyzed, the result obtained had a maximum deviation of ± 0.4% from the theoretical value.

Starting material

Ethylisonicotinate and isonicotinic hydrazides were prepared as previously described by Burrus and British pharmaceutical codex. Benzoic acid hydrazide was synthesized according to method reported in Vogel (18).

5-Subtituted-2-mercapto-1, 3, 4-oxadiazoles (1, 2)

To a solution containing 400ml of 95% ethanol and (0.1 mole,5.6g) of potassium hydroxide (dissolved in 15ml of water), (0.1 mole) of the appropriate hydrazide was added. After the solution occurred (6.6ml, 0.11 moles) of carbon disulfide was added and the mixture was refluxed for 3 hrs. The solution then was concentrated to a small volume and the residue was dissolved in water. A precipitate was obtained by adding the solution to ice containing hydrochloric acid. The solid was filtered off, dried and recrystallized from ethanol.

The IR spectra showed a weak SH stretching absorption at 2600 cm-1.

Melting point for 5-phenyl and 5-(4-pyridyl)-2-mercapto-1, 3, 4-oxadiazoles 1 and 2 were 270 – 271 oC and 218 – 220 oC respectively and are consistent with their previously reported melting points (19).

2- Alkyl/alkenyl/aralkyl-thio-5-substituted-1, 3, 4-oxadiazoles (3-9)

To a stirred solution of 1 or 2 (0.01 mole) in 20ml absolute ethanol, was added (0.01mole, 0.56g) potassium hydroxide, the solution was refluxed for 30 minutes. Then the appropriate alkyl or allyl halide (0.01 mole) was added dropwise to the stirred reaction mixture, which was then refluxed for two hrs. After cooling, the mixture was filtered and the filtrate poured into ice- cold water.

The crude product was collected and recrystallized from ethanol-water. The oily alklated products, were separated as oil drops after pouring the mixture on ice-cold water.

The aqueous mixture was then extracted with ether, the ether layer dried over anhydrous sodium sulfate, the ether was evaporated and the product was purified from acetone. The physical data, the prepared compounds and their IR spectral analyses were reported in (Table 1 and 4) respectively.

2- Alkyl/aralkyl sulfonyl-5-substituted-1, 3, 4-oxadiazoles (10-14)

A solution of 0.7g potassium permanganate in 8ml water was added to a stirred ice-cold suspension of 0.002 mole of the appropriate 2- alkyl thio -5- substituted -1,3,4-oxadiazoles 3-9 in 5ml glacial acetic acid. Cooling and stirring was continued for four hrs, then the reaction temperature was brought to 0 oC and saturated sodium sulfite solution was added gradually with stirring until the color of the permanganate disappeared. The reaction mixture was then filtered and the precipitate washed with water and crystallized from ethanol-water. The physical data of the prepared compounds and their IR spectral analyses were reported in (Table 2 and 5) respectively.

5-(4-pyridyl)-1, 3, 4-oxadiazolyl-2-thiocarbanates (15, 16)

A mixture of 5-(4-pyridyl)-2-mercapto-1,3,4-oxadiazole (0.005 mole) absolute ethanol (20ml), potassium hydroxide (0.005 mole, 0.28g), and anhydrous sodium carbonate (0.005 mole, 0.42g) was refluxed for 30 minutes. The mixture was then cooled to room temperature, and the appropriate chloroformate (0.005 mole) was added dropwise with continuous stirring, the mixture was heated at (30-35 oC) for one hr, cooled to room temperature and poured into ice-water. The precipitated product was filtered, washed with water, and recrystallized from ethanol-water. Yields melting point and IR spectral analyses were reported in (Table 3 and 6).

5-phenyl-1, 3, 4-oxadiazolyl-2-thiocarbamates (17)

To a solution of 0.01 mole of 5-phenyl-2-mercapto-1, 3, 4-oxadiazole in 20 ml absolute ethanol was added (0.84 mole, 0.01g) of anhydrous sodium carbonate, then (0.01 mole) of the appropriate chloroformate was added dropwise. The reaction mixture was refluxed for 1-2 hrs; cooled to room temperature and poured on 100 ml ice water. The precipitated carbonate was filtered off, washed with water and crystallized from ethanol-water. Physical data of the prepared compounds, and the IR spectral analyses were shown in (Table 3 and 6).

2-Allenyl thio-5-substituted-1, 3, 4-oxadiazole (18)

To a stirred solution of 1 (0.01 mole) in 20 ml absolute ethanol, (0.01 mole, 0.56g) of potassium hydroxide was added, the solution was refluxed for 30 minutes, then propargylbromide (0.011 mole) was added dropwise and refluxed for 2 hrs.

After cooling to room temperature, the mixture was filtered and the filtrate poured into ice-cold water. The precipitate was filtered off and recrystallized from ethanol-water. The physical data were shown in Table 1.

2-(2-propynyl)-thio-5-phenyl-1,3,4-oxadiazole (18)

To a solution of 2-mercapto-5-phenyl-1, 3, 4-oxadiazole (0.01 mole) in ethanol (50 ml), propargyl bromide (0.012 mole, 2 ml) was added dropwise.

When the addition was completed, the reaction mixture was stirred for 3 hrs at room temperature. Evaporation of the ethanol under reduced pressure yielded a crude product which was extracted with ethyl acetate and dried over anhydrous sodium sulphate. The ethanol was removed under reduced pressure. The crude solid was crystallized from ethyl acetate-ethanol mixture as a white solid in 65% yield.

The IR spectra showed the following absorption bands (KBr, cm-1) 3270 (=CH), 3020 (ArH), 2120 (C=C), the NMR spectra showed the following characteristic chemical shifts (CDCI3, δ) 3.5 (doublet, 2H, CH2-C=, Jz =2,2) 205 (triplet, IH, =CH, Jz = 2,2). Other signals in the spectra are consistent with various protons in the aromatic ring.

2-(4-pyrrolidino-2-butynyl) thio-5-phenyl-1, 3, 4-oxadiazole (19)

A mixture of 5-phenyl-(2-propynyl)-1,3,4- oxadiazole (0.003 mole), paraformaldehyde (0.0033 mole), pyrrolidine (0.003 mole) and a catalytic amount of cuprous chloride in 10 ml per oxide-free dioxone. The mixture was stirred at room temperature for 10 minutes then was heated at 70 – 75 oC for three hours. Concentration of the reaction mixture under reduced pressure gave brown syrup, which was suspended in water and neutralized with sodium carbonate solution.

Extraction of the syrup with ethyl acetate, dried over anhydrous sodium sulfate. The chromatography on silica gel resulted in the desired Mannich product. The IR spectra showed the following characteristic absorption bands (KBr, cm-1) 3050 (CH, ArH), 2100 (very weak, C=C); The NMR spectra showed the following characteristic chemical shifts (DMSO, δ).

4.5 (triplet, 2H, S-CH2-C≡, J= 2.2Hz) 3.4 (triplet, 2H, ≡C-CH2-N, J=2.2Hz).

Other signals in the NMR spectra were consistent with the protons in aromatic ring and the pyrrolidine.

ANTIMICROBIAL ACTIVITY

All tested compounds were assayed for their antimicrobial activity according to the macrodilution method of the National Committee for Clinical Laboratory Standards

(NCCLS)(20-21) recommendations, against three standard bacterial strains, Staphylococcus aureus( ATCC 29213), Escherichia coli (ATCC 25922) and Pseudomonas aeruginosa (ATCC 27953). The compounds were evaluated against clinical isolated Candida albicans for their antifungal activity. The evaluation was done using dilution method as shown in (Table 7). Increasing concentrations of the compounds were incorporated into Muller Hinton broth (BBL Microbiology Systems) for bacteria and Sabouraud’s broth for Candida.

MIC breakpoints for defining susceptibility were in accordance with the description by National Committee for Clinical laboratory Standards (20).

RESULTS AND DISCUSSION

The new compounds (Table 1, 2, 3) and compounds (18, 19, 20) were prepared as depicted in Scheme 1. 2 – mercapto – 5 – substituted – 1, 3, 4 – oxadiazoles 1, 2 were prepared as previously described by Young and Wood (19)

The potassium salts of 1, 2 were converted to their corresponding thioethers 3-9 through reaction with the appropriate alkyl halides (Table 1). Oxidation of the thioethers upon treatment with potassium permanganate in glacial acetic acid yielded the corresponding sulfonyl derivatives 10-14 (Table 2).

Treatment of 1, 2 with chloroformate in the presence of sodium carbonate afforded the corresponding thiocarbamates 15-17 (Table 3). The physical constant, IR, and elemental analyses were consistent with assigned structures (Table 4, 5, 6). Treatment of 1 with alcoholic sodium or potassium hydroxide afforded 2 – allenyl thio – 5 – phenyl – 1, 3, 4 – oxadiazole 8.

The formation of the allene may result from the isomerization of the kinetically controlled product 18 into the thermodynamically controlled product 8 as illustrated in scheme 2.

The ir and elemental analysis were in agreement with the assigned structures, whoever alkylation of 1 with propargyl bromide in absolute ethanol afforded the 2 – (2-propynyl) – thio – 5 – phenyl – 1,3,4 – oxadiazole 19 .

Compound were subjected to mannich reaction utilizing paraformaldehyde, pyrrolidine as secondary amine and a catalytic quantity of cuprous chloride in peroxide – free dioxane yielded the N – 2 – (4 – pyrrolidine – 2 – butynyl) thio – 5 – phenyl – 1,3,4 – oxadiazole 19.

The IR, NMR and elemental analysis were consistent with prepared compound. It is of interest to report here that in our previous publications (22, 23).

The alkylation of 2 – mercapto – 5 – substituted - 1, 3, 4 – thiadiazoles or

3 – mercapto – 4,5 – disubtituted – 1,2,4 – triazoles in basic ethanolic solution yielded the 2 – propynyl thio rather than the allenyl thio – derivatives.

These results may suggest that the oxygen atom in the heterocyclic ring exert a greater electronic polarization on the neighboring mercapto group than sulfur or nitrogen within the heterocyclic ring and that resulted in the isomerization of 2 – propynyl to allene group.

ANTIMICROBIAL ACTIVITY

The lowest concentration which inhibited growth was considered as the MIC. The synthesized compounds showed low antibacterial activity against P. aeruginosa but good activity against S. aureus and E. coli. Only thiocarbamate 16 and 17 were effective against Gram positive and Gram negative bacteria with less activity against fungi. These results promote our interest to investigate further the thiocarbamate series.

Table 1: Physical data of the 2-alkyl, alkenyl thio-5-substituted-1, 3, 4-oxadiazoles

* These compounds contain ½ mole of H2O

§ These compounds contain 1 mole of H2O

Table 2: Physical data of the 2-alkylsulfonyl -5-substituted-1, 3, 4-oxadiazoles

Table 3: Physical data of 5-substituted-1, 3, 4-oadiazolyl -2-thiocarbamates

Table 4: Selected infrared data of 5-substituted-2-alkyl/alkenyl/aralkyl thio-1, 3, 4-oxadiazoles

* Abbreviations: (s) Strong, (m) Medium, (w) Weak.