Synthesis and Biological Evaluation of New Pyridine Derivatives

 

Yogesh Jadhav, Kushal Dawane, Vrushali Patil, A.S. Bobade*, S.V. Athlekar and Abhay Chowdhary.

Department of Chemotherapy, Haffkine Institute for Training, Research and Testing, Acharya Donde Marg, Parel, Mumbai-400012.

*Corresponding Author E-mail: j.yogi@yahoo.com

ABSTRACT:

The 3-Hydroxy pyridine on reaction with ethylchloro acetate in presence of K₂CO₃ followed by the reaction with hydrazine hydrate resulted in the formation of 3-pyridoxy acetyl hydrazide (III), which on further the reaction with CS₂ and KOH cyclized to give (IV) which on reaction with different acid chloride form 2-(3-pyridoxymethyl)-5-(4-substitutedphenyl)-1,3,4-triazole. These structures are determined by the elemental analysis and spectral data (IR, 1H-NMR). These new derivatives are evaluated for in vitro antimicrobial activity against Staphylococcus aureus (ATCC 3750), Salmonella typhi (NCTC 786), Candida albicans (ATCC 10231) and Aspergillus niger (ATCC 16404).

 

 

 

INTRODUCTION:

Large number of pyridine derivatives exhibit interesting biological activities. Especially 3-Pyridine derivatives such as Isoniazides and Nicotinamide are commonly used as drug for the treatment of tuberculosis¹. The presence of Oxygen and nitrogen in the heterocyclic system has attracted the attention of medicinal chemist because the diverse biological activities and profound efficacy. 1, 3, 4-Oxadiazole nucleus, constitute a unique class of nitrogen and oxygen containing five membered heterocycles associated with various biological activities such as anti-bacterial, anti-inflammatory, pesticidal, anti-hypertensive^2-5. The recent review of literature has highlighted the significance of medicinal importance of oxadiazole and their derivatives⁶ Also it has been reported that 2,5-disubstituted-1,3,4-oxadiazoles shows good anti-bacterial and significant pesticidal activity⁷.

 

MATERIALS AND METHODS:

All raw materials used in the synthesis have been obtained from Fluka AG (Buchs, Switzerland) and Aldrich Chemical and Co. Inc. (Milwaukee, WI, USA). All the commercial solvents used in the experimental work were redistilled and dried before use.

 

Melting points of the synthesized compounds was recorded on Thermonik Melting Point Apparatus (Campbell Electronics, Mumbai, India) and are uncorrected. TLC was performed on silica gel coated plates using iodine vapour as visualizing agent. IR spectra were recorded on Hartmann-Braun, MB series (Bomem, Quebec, Canada), ¹H NMR spectra were recorded on 300 MHz instrument (Jeol Ltd., Tokyo, Japan), using deuterated solvent DMSO, at Institute of Science, Mumbai. Elemental analyses were carried out on at SAIF, IIT Powai, Mumbai. All compounds were subjected to antibacterial screening against Staphylococcus aureus (ATCC 3750), Salmonella typhi (NCTC 786) using tube dilution technique⁸. Ampicillin and Trimethoprim were used as the standard drugs against S. aureus and S. typhi, respectively. The SAR of antifungal agents has revealed that the basic nitrogen-carbon linkage is an essential feature for the activity⁹. All the synthesized compounds were screened for antifungal activity against standard strain of Candida albicans (ATCC 10231) and Aspergillus niger (ATCC 16404) on Sabouraud-Dextrose broth by tube dilution method using Miconazole as standard drug.

 

EXPERIMENTAL:

Step I : Synthesis of 3-pyridoxy acetyl hydrazide (III)

A mixture of 3-hydroxy pyridine 9.5g (0.1mole) and potassium carbonate 13.8g (0.1mole) was stirred and refluxed for 6 hours. Cooled to room temperature and 11.1ml (0.1mole) of ethyl chloroacetate was added to it drop wise with stirring. After complete addition stirred for one hours and whole reaction extracted with ether.

 

Table No. 1 :- Physical properties and elemental composition of 2-(Substituted thiocarbamoyl)-5-(3-pyridoxymethylene)-1,3,4-oxadiazole derivatives.

 

 

Comp. No	R	Yield (%)	M.P. (0C)	Colour and Nature	Elemental Analysis
					%	C	H	N
1.	4-Chloro	55	156	Brown Amorphous Solid	Cal.	51.79	2.87	12.08
					Found	51.63	2.65	11.82
2.	4-Bromo	58	160	Brown Amorphous Solid	Cal.	45.91	2.55	10.71
					Found	46.12	2.73	10.56
3.	4-Fluro	61	152	Yellow Amorphous Solid	Cal.	54.38	3.02	12.68
					Found	54.11	3.34	12.91
4.	2-Bromo	52	168	Gray Amorphous Solid	Cal.	45.91	2.55	10.71
					Found	45.68	2.36	10.88
5.	4-Methoxy	47	173	Off White Amorphous Solid	Cal.	55.97	3.79	12.24
					Found	55.61	3.58	12.42

 

Excess of solvent was evaporated in vacuum. Residual liquid was added drop wise to a stirred solution of 9.7ml (0.2mole) of hydrazine hydrate in absolute alcohol at room temperature. Mixed well and refluxed for 3 hours.

 

Excess of solvent was distilled off. After cooling white amorphous solid was obtained. Crystallized from ethanol.

Yield = 48%, M.P.=148°C

 

Elemental Analysis (C₇H₉N₃O₂) :-  C% = 50.40, H% = 5.09, N% = 24.86

IR (KBr) cm^-1:- 3000-3100 (C-H str. Aromatic), 1600 (C=C str. Aromatic), 1325 (C-O-C str Ether linkage), 825 (C-H def Aromatic).

 

Step II: Synthesis of 2-(3-pyridoxymethyl)-5-mercapto-1,3,4-oxadiazole (IV)

To a stirred ethanolic solution of 4g of KOH and 1.67g (0.01mole) of 3-pyridoxy acetyl hydrazide (III), 1.8ml (0.03mole) of CS₂ were added at room temperature. Then the reaction mixture was refluxed till evolution of H₂S ceased. Thereafter it was cooled, diluted with cold water and acidified with glacial acetic acid. The white cotton wooly solid separated was washed with water and crystallized from ethanol.

Yield =  66%, M. P. =  125°C

 

Elemental Analysis (C₉H₁₁N₃O₂S) :-  C% = 49.53, H% = 4.99, N% = 18.48

 

IR (KBr) cm^-1:- 3000-3150 (C-H str. Aromatic), 1575 (C=C str. Aromatic), 1317 (C-O-C str Ether linkage), 870 (C-H def Aromatic).

 

Step II : Synthesis of 2-(4-Bromophenyl thiocarbamoyl)-5-(3-pyridoxymethylene)-1,3,4-oxadiazole  (Compound No. 2, Table No. 1)

Solution of 2.09g (0.01mole) of 2-(3-pyridoxymethyl)-5-mercapto-1,3,4-oxadiazole (IV) and 1.4ml (0.001mole) of 4-bromobenzoyl chloride in dry pyridine was refluxed for 2 hours. Cooled and poured into crushed ice. Solid thus separated was filtered, washed with 20ml of water and crystallized from ethanol.

Yield = 58%, M. P. = 160°C

 

Elemental Analysis (C₉H₁₁N₃O₂S) :-  C% = 46.12, H% = 2.73, N% = 10.56

IR (KBr) cm^-1 -: 2997-3300 (C-H str. Aromatic), 1690 (C=O Str.), 1590 (C=C str. Aromatic), 1310 (C-O-C str Ether linkage), 907 (C-H def Aromatic), 556 (C-Br str).

¹H NMR (CDCl₃) δ ppm -: 3.16 (s, 2H,-OCH₂), 7.0-7.92 (m, 8H, Ar-H).

All other analogs were prepared using similar method, physical properties and elemental analysis results of which are recorded in Table No. 1.

 

DISCUSSION:

In present work, the synthesis of 2-(Substituted thiocarbamoyl)-5-(3-pyridoxymethylene)-1,3,4-oxadiazole derivatives have been described. The following points are to be noted related to Synthesis and Biological screening:

 

1.           The key step in the synthesis of 3-pyridoxyacetyl hydrazide (III) is the formation of potassium salt of 3-hydroxy pyridine. For this anhydrous K₂CO₃ stirred with 3-hydroxy pyridine for about three to five hours at 50⁰C to provide complete salt formation.

 

2.           It was found that 3- pyridoxyacetate, as containing ester functional group tends to de-esterify in presence of water (humidity) and acid (liberated HCl). To overcome this, excess of K₂CO₃ (2.5 equivalent) in anhydrous condition was used. Also separation of 3- pyridoxyacetate from reaction mixture found very difficult in our laboratory conditions. Hence we use ethernal extract of reaction mixture to prepare 3-pyridoxyacetyl hydrazide (III).

 

3.           2-(3-pyridoxymethyl)-5-mercapto-1,3,4-oxadiazole (IV) was found highly hygroscopic hence must be used immediately when prepared.

 

 

Table No. 2 :- Biological activity of 2-(Substituted thiocarbamoyl)-5-(3-pyridoxymethylene)-1,3,4-oxadiazole derivatives.

 

Comp. No.	Substituents R	Anti Microbial Activity (µ/ml) [MIC]
		Anti bacterial		Anti Fungal
		S. aureus	S. typhi	C. abicans	A. niger
1.	4-Chloro	50	100	50	100
2.	4-Bromo	200	200	200	200
3.	4-Fluro	50	50	100	50
4.	2-Bromo	N.A.	200	N.A.	N.A.
5.	4-Methoxy	50	100	100	100

·                  Ampicillin (MIC-0.04 mg/ml) used as standard against S. aureus.

·                  Trimethoprim (MIC 0.01 mg/ml) used as standard against S. typhi.

·                  Miconazole (MIC 6.25 mg/ml) as standard against C. albicans and A. niger.

·                  N.A. -: Not active at 200 mg/ml.

 

 

Schematic Representation :-

4.           Among all the five synthesized titled compounds, (Refer Table No. 2) 2-bromo substituent (Compound No. 4) is found not active, 4-bromo substituent (Compound No. 2) exhibited activity upto 200mg/ml and rest of the compounds shows good activity against S.aureus.

 

5.           The compounds with 2-bromo substituent (Compound No. 4) and 4-bromo substituent (Compound No. 2) exhibited activity upto 200mg/ml whereas rest of the compounds exhibited moderate to good activity upto 100mg/ml against S.typhi. 4-Fluro substitution (Compound No. 3) shows encouraging activity at 50mg/ml against S.typhi.

 

6.           2-Bromo substituent (Compound No. 4) is found not active against both fungal strains while 4-bromo substituent (Compound No. 2) exhibited activity upto 200mg/ml. All other substitutions show moderate to good activity against both fungal strains.

 

ACKNOWLEDGEMENT:

The authors are thankful to SAIF (Sophisticated Analytical Instrument Facility), IIT (Indian Institute of Technology), Powai, Mumbai for carrying out the elemental analysis (CHN) and also thankful to Institute of Science, Fort, Mumbai for recording the NMR spectra.

 

REFERENCES:

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3.       Ramalingam R. T., Deshmukh A. A. et al; J. Indian Chem. Soc.; (1981), 58, 269.

4.       Hiroshi K., Isao H. and Shigeki O.; Zassokenkyn; (1969), 8, 46. cited Chem. Abstr.; (1970), 73, 108544.

5.       Ponticello G. S., Englehardt E. L. and Baldwin J. J.; J. Heterocyclic Chem.; (1980), 17, 425.

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7.       Hosam Saad et al; Indian J. Chem.; (1996), 35B, 980.

8.       E. Joan Stokes, Clinical Bacteriology, 4^th Edition (1995), 226.

9.       Wilson Giswold’s “Textbook of Organic Medicinal Chemistry and Pharmaceutical Chemistry”, 11^th Edition, 455.

 

 

Received on 19.04.2010        Modified on 02.05.2010

Accepted on 22.05.2010        © AJRC All right reserved