Asian J. Research Chem. 2(2): April.

-June, 2009 ,

ISSN 0974-4169 RESEARCH ARTICLE

www.ajrconline.org

Synthetic and Pharmacological Evaluation of Some Pyridine Containing Thiazolidinones

1

KYDSCTs College of Pharmacy, Sakegaon, Tal. Bhusawal, Dist. Jalgaon, (M.S), India. 2 TVESs College of Pharmacy, Faizpur, Tal. Yawal, Dist. Jalgaon, (M.S), India. *Corresponding Author E-mail: sandipfirke@rediffmail.com

Firke SD*1, Firake BM1, Chaudhari RY2 and Patil VR2

ABSTRACT

A series of N-[3-(aryl/alkyl substituted)-4-oxo-1, 3-thiazolidin-2-ylidene]-2-(pyridine-2-yloxy) acetohydrazides were synthesized using appropriate synthetic route. These compounds were synthesized by their analytical and spectral data. All the newly synthesized compounds were examined for their antidiabetic activity using GOD-POD method on Wistar strain rats. The acute toxicity study (LD50) values of these compounds were determined. The test compounds showed significant antidiabetic activity on evaluation.

KEY WORDS: Thiazolidinone, Pyridine, Antidiabetic activity.

A number of thiazolidinone derivatives have been reported to possess diversified activities including hypoglycemic action.1 Thiazolidinone ring is a main pharamacophoric group responsible for antidiabetic activity. Therefore, it was planned to choose thiazolidinone as a lead molecule for molecular modification to enhance the specificity and potency of action and to reduce the toxicity. Compounds carrying the thiazolidinone ring have been reported to demonstrate a wide range of pharmacological activities which include anticonvulsant2, antimicrobial3, 4 5 antiinflammatory , antihistaminic , anti-hypertensive6, and hypnotic7, antidiabetic8, 9 activities. Heterocyclic ring like pyridine ring also plays important role in antidiabetic activity of some drugs (pioglitazone, rosiglitazone). In general, pyridine ring and substituted thiazolidinone ring are essential for antidiabetic activity. The proposed work involves syntheses of some novel N-[3-(aryl/alkyl substituted)-4-oxo-1, 3-thiazolidin-2-ylidene]- -(pyridine-2yloxy) acetohydrazides with the aim of obtaining the new antidiabetic agents.

INTRODUCTION:

boiling ethanol containing fused sodium acetate gives the corresponding N-[3-(aryl/alkyl substituted)-4-oxo-1, 3-thiazolidin-2-ylidene]-2-(pyridine-2-yloxy) acetohydrazides (9-13). The synthetic route is depicted in Scheme 1. Thus in the present investigation, five different derivatives of N-[3-(aryl/alkyl substituted)-4-oxo-1, 3thiazolidin-2-ylidene]-2-(pyridine-2-yloxy) acetohydrazides were synthesized and evaluated for their antidiabetic activity.

EXPERIMENTAL:

Ethyl (pyridine-2-yloxy) acetate (compound 2) was synthesized in an excellent yield by electrophillic substitution on 2-hydroxy pyridine using ethyl chloroacetate under the reflux condition. Compound 2 on amination with hydrazine hydride yield 2-(pyridine2-yloxy) acetohydrazide 3. Reaction of 3 with alkyl/aryl isothiocynate in ethanol gives compounds 4-8. The Synthesis of 2: To a mixture of triethylamine (5.32 g, cyclization reaction of 4-8 with chloroacetic acid in 0.0525 mol) and 2-hydroxy pyridine (5 g, 0.0525 mol), a solution of ethyl chloroacetate (5.6ml, 0.0525 mol) in 1:4 Dioxane (50 ml) was added drop wise. The Received on 06.04.2009 Modified on 21.05.2009 temperature was maintained at 90C for 1 hr and then Accepted on 15.06.2009 AJRC All right reserved the reaction mixture was stirred for 7-8 hrs. The excess Asian J. Research Chem. 2(2): April.-June, 2009 page 157-161 solvent was removed under reduced pressure. Then the reaction mixture was poured in ice-cold water and was

Melting points and boiling points were determined in open capillaries and were uncorrected. Purity of the compounds was ascertained by TLC plates using silica gel G coated glass plates using chloroform-methanol as irritant and iodine vapour as detecting agent. IR spectra were recorded using KBr pellets on FTIR 8101, Shimadzu, Japan, 1H NMR spectra and Mass spectra (FAB-MS) were recorded on Varian 300 MHz instrument and 70eV on Jeol D-300 spectrometer (Jeol Ltd, Tokyo, Japan). All the solvents and chemicals used for the synthesis were of S. d. Finechemicals Limited, Mumbai. The starting materials were obtained from Lancaster Limited and Alkali Metals Limited, Hyderabad.

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Table 1: Physical data of N-(substituted aryl/alkyl)-2-[(pyridine-2- yloxy) acetyl] carbothioamides (4-11). Compound No. Name of aliphatic / aromatic isothiocynate Yield Rf value m.p.(0c) (Uncorrected) 4 Phenyl 95% 0.68 262-264 5 Ethyl 93% 0.65 116-118 6 p-chlorophenyl 89% 0.59 226-228 7 2, 4- dichlorophenyl 88% 0.60 215-216 8 Methyl 90% 0.64 136-137 Table 2: Physical and analytical data of compounds. Compound No. R Molecular formula m.p.(0C) uncorrected Yielda (%) Mass[M+2] 9 Phenyl C16H14N4O3S 84-86 65 342b 10 Ethyl C12H14N4O3S 116-118 62 294 11 p-chlorophenyl C16H13N4O3S 130-132 55 376b 12 2,4-dichlorophenyl C16H12N4O3S 116-118 62 410b 13 Methyl C11H12N4O3S 118-120 59 279 a All the compounds were recrystalized from ethanol. b Values represent [M+2] due to appearance of an isotopic peak. Table 3: Intraday Effect of Different Aryl/ Alkyl Substituted Thiazolidinone Derivatives on Serum Glucose at 1st day. Average Serum Glucose Level (mg/dL) at (1st day) Compound No. 0 hr 1 hr 3 hr 5 hr 7 hr Control 275.151.87 276.120.78 269.212.22 266.023.10 262.012.43 Alloxan 270.62.80 275.231.87 289.401.01 296.083.12 302.612.09 Standard 282.002.80 242.271.63 201.424.21 159.110.99 122.155.63 9 276.023.36 245.422.50 212.123.10 170.211.56 126.061.23 10 294.315.32 274.103.21 247.021.02 191.380.19 138.150.96 11 298.324.23 290.430.49 261.233.10 220.315.20 181.262.14 12 294.431.93 288.112.63 276.402.90 253.240.62 245.221.83 13 283.111.23 276.102.01 243.171.94 196.161.03 148.190.96 The values are presented as mean S. E. M. of six determinations p<0.01, Students t- test compared with diabetic control. Table 4: Effect of Different Aryl/ Alkyl Substituted Thiazolidinone Derivatives on Serum Glucose at 1st, 3rd, 7th day. Average Serum Glucose Level (mg/dL) Compound No. 1st day 3rd day 7th day Control 237.023.10 229.211.23 198.133.21 Alloxan 270.62.80 336.020.98 302.311.34 Standard 159.110.99 124.152.63 105.215.32 9 170.211.56 140.085.20 118.161.10 10 191.380.19 164.321.03 124.430.69 11 290.315.20 261.113.36 221.122.12 12 253.240.62 220.430.36 191.032.26 13 196.161.03 176.214.20 159.211.21 The values are presented as mean S. E. M. of six determinations p<0.01, Students t- test compared with diabetic control.

extracted with chloroform. The chloroform layer was separated and the chloroform was removed under vacuum. The liquid product obtained was recrystallized from chloroform. Rf: 0.56 (Chloroform: methanol, 8:2); b.p. 68-69C IR: 1690cm (C=O), 2985 cm (C-H stretching); H NMR (CDCl3): 1.20 (t, 3H, CH3), 4.83-4.96 (q,2H, CH2), 4.70 (s, 2H, OCH2), 6.38 (t, 1H, pyridine C-3), 7.75 (q, 1H, pyridine C-4), 6.68 (q, 1H, pyridine C-5), 7.79 (t, 1H, pyridine C-6). Synthesis of 3: Compound 2 (3.75 ml, 0.025 mol) was added drop wise to hydrazine hydrate (1.3 ml, 0.025 mol) in a round bottom flask. The mixture was heated gently under reflux for 15 minutes and enough absolute ethanol was added through the condenser to produce a clear solution. Then the reaction mixture was refluxed
-1 -1 1

for further 2-3hrs. The ethanol was distilled off. The solid crystals were filtered and recrystallized from ethanol. Yield: 62%, Rf: 0.62 (Chloroform: methanol, 8:2), m.p.: 105-107C IR: 3382, 3355 cm-1(NHNH2), 1720 cm-1(C=O); 1H NMR (CDCl3): 4.40(s, 2H, NH2), 4.68 (s, 2H, OCH2), 7.75 (s, 1H, CONH), 6.35 (t, 1H, pyridine C-3), 7.78 (q, 1H, pyridine C-4), 6.70 (q, 1H, pyridine C-5), 7.79 (t, 1H, pyridine C-6). Syntheses of 4-8: To a solution of 3 (1.68g, 0.01 mol) in ethanol (50 ml), various aliphatic/aromatic isothiocynates (0.01 mol) were added and the reaction mixture was refluxed for 12 hrs. Excess solvent was removed under vacuum. The residue obtained was washed with diethyl ether and was recrystallized from methanol.

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Scheme 1: General scheme for syntheses of compounds 9-13 (R= phenyl, ethyl, p-chlorophenyl, 2, 4- dichlorophenyl, methyl)
ClC H
2

Asian J. Research Chem. 2(2): April.-June, 2009 ,

O N O C H O C
2 2

C O O C o

N H

N H

O H

90

c 1h

O N O C H
2

2 - h y d r o x y p y r id in e (1 )

TE A

E t h y l( p y r id in e - 2 - y lo x y ) a c e ta te (2 )

R e f lu x 2 -3 h

N H

N H

2 - ( p y r id in e - 2 - y lo x y ) a c e t o h y d r a z id e ( 3 ) R N C S R e f lu x1 2 h

O N O C H
2

N H R

H N S N H

N - ( s u b s t it u t e d a r y l / a lk y l) - 2 - [ ( p y r id in e - 2 - y lo x y ) a c e t y l] a c e t o h y d r a z id e (4 -8 ) ClC H2C O O H R e f lu x 1 0 h s o d iu m a c e t a t e

O N O C H
2

N H R O N

N S

N '[ 3 - ( a lk y l / a r y l s u b s t it u t e d ) - 4 - o x o - 1 , 3 - t h ia z o lid in - 2 - y lid in e ] - 2 - ( p y r id in e - 2 - y lo x y ) a c e t o h y d r a z id e . (9 -1 3 )

The yield and physical data are summarized in Table 1. 4: IR: 3382, 3355 cm-1(NHNH2), 1720 cm-1(C=O), 1360 cm-1(C=S); 1H NMR (CDCl3): 7.58 (m, 4H, Ar.), 4.82 (s, 2H, OCH2), 7.78 (s, 1H, CONH), 6.34 (t, 1H, pyridine C-3), 7.72 (q, 1H, pyridine C-4), 6.79 (q, 1H, pyridine C-5), 7.76 (t, 1H, pyridine C-6).

5: IR: 3212, 3225 cm-1(NH), 1730 cm-1(C=O), 1365 cm1 (C=S); 1H NMR (CDCl3): 1.28 (m, 3H, CH3), 4.72 (s, 2H, OCH2), 7.75 (s, 1H, CONH), 6.44 (t, 1H, pyridine C-3), 7.76 (q, 1H, pyridine C-4), 6.69 (q, 1H, pyridine 9. IR: 3220 cm-1 (N-H), 1720 cm-1 (C=O), 1585 cm-1 C-5), 7.66 (t, 1H, pyridine C-6). (C=N), 3010 cm-1 (C-H); 1H NMR :( CDCl3) 6.38, 7.75, 6.68, 7.79 (C-H, 2-pyridine), 3.76, 4.83, 3.24 6: IR: 3217, 3232 cm-1(NH), 1736 cm-1(C=O), 1315 cm- (CH2), 7.0 (-NH-), 1.20(CH3); FAB-MS: (m/z, 1 (C=S); 1H NMR (CDCl3): 7.39-7.65 (m, 4H, ArH), 100%): 342 ([M+], 100%) 4.72 (s, 2H, OCH2), 7.75 (s, 1H, CONH), 6.34 (t, 1H, pyridine C-3), 7.73 (q, 1H, pyridine C-4), 6.62 (q, 1H, 10. IR: 3325 cm-1 (N-H), 1722 cm-1 (C=O), 1583 cm-1 pyridine C-5), 7.76 (t, 1H, pyridine C-6). (C=N), 3015 cm-1 (C-H); 1H NMR :( CDCl3) 6.38, 7.69, 6.68, 7.75 (C-H, 2-pyridine), 3.76, 4.83 (CH2), 7: IR: 3219, 3222 cm-1(NH), 1734 cm-1(C=O), 1315 cm- 7.0 (-NH-), 7.14, 7.06, 7.06, 7.14, 7.07 (Phenyl ring); 1 (C=S); 1H NMR (CDCl3): 7.29-7.65 (m, 3H, ArH), FAB-MS: (m/z, 100%): 294 ([M+], 100%) 4.73 (s, 2H, OCH2), 7.75 (s, 1H, CONH), 6.44 (t, 1H, pyridine C-3), 7.73 (q, 1H, pyridine C-4), 6.65 (q, 1H, 11. IR: 3400 cm-1 (C-H str. pyridine), 1552 cm-1 (N-H pyridine C-5), 7.73 (t, 1H, pyridine C-6). str.), 1730 cm-1, 1650 cm-1 (C=O), 1525 cm-1 (C=N),

Syntheses of 9-13: A mixture of the N-(substituted aryl/alkyl)-N(-2-pyridine-2-yloxy) acetyl thiosemicarbazides (3.03g, 0.01 mol), chloroacetic acid (0.93g, 0.01 mol) and sodium acetate (0.81g, 0.01 mol) in ethanol (60 ml) was refluxed for 10 hrs. The mixture was cooled and diluted with enough water to develop turbidity and left overnight for complete separation of the product. Then the compounds were filtered and recrystallized from ethanol. The yield and physical data are summarized in Table 2.

3035 cm-1 (C-H, Aromatic str.), 1208 cm-1 (C-O-C str.); 8: IR: 3216, 3215 cm-1(NH), 1735 cm-1(C=O), 1362 cm- 1 H NMR (CDCl3) 6.38, 7.69, 6.68, 7.75 (C-H, 21 (C=S); 1H NMR (CDCl3): 1.18(m, 2H, CH2), 4.70 (s, pyridine), 3.81, 4.83 (CH2), 7.0 (-NH-), 7.58, 7.25, 2H, OCH2), 7.73 (s, 1H, CONH), 6.54 (t, 1H, pyridine 7.25, 7.58, (Phenyl ring); FAB-MS: (m/z, 100%): 378 C-3), 7.72 (q, 1H, pyridine C-4), 6.72 (q, 1H, pyridine ([M++2], 100%)). C-5), 7.76 (t, 1H, pyridine C-6).

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In the present investigation, different derivatives of N[3-(aryl/alkyl substituted)-4-oxo-1, 3-thiazolidin-2ylidene]-2-(pyridine-2-yloxy) acetohydrazides (9-13) were synthesized and evaluated for their physical, Cut-Off Lethal Dose (LD50): All the compounds synthesized were tested for acute analytical and spectral data (Table 2). toxicity test. No toxicity was observed at the doses of 300, 1000, 2000 mg/kg of body weight but it was The structures of compounds 9-13 were confirmed on observed that more than 50% of animals were died at the the basis of spectral data. IR spectrum showed -1 -1 dose of 2000 mg/kg of body weight. Thus for the absorption peaks at 1552 cm and 1650 cm for the Nscreening of antidiabetic activity, the dose selected was H stretching and C=O stretching of amide groups 1 200 mg/kg of body weight (i.e., 1\10 of the 2000 mg/kg respectively. The H-NMR spectrum exhibited signals 10 attributed to the proton at 6.38, 7.69, 6.68, 7.75 of body weight) as per the OECD guidelines. indicating the presence of pyridine ring, while the signals at 7.0 indicated the presence of acetohydrazide Drugs Used: Metformin was given to rats at a dose of 5 mg/kg body linkage. weight, as a reference standard. The results of antidiabetic activity of test compounds were given in Table 3, 4. Compounds No. 9 and 10 were Induction of Diabetes: A single dose (150 mg/kg, body weight) of Alloxan found to be most efficient i.e. 48% and 52% reduction of monohydrate (5%w/v in sterile water) was dissolved in serum glucose level respectively at 200 mg/kg dose. normal saline used for the induction of diabetes and injected intraperitoneally to Wistar albino rats weighing CONCLUSION: 150-200 g. The induction of diabetes was confirmed by A series of N-[3-(4-alkyl/aryl substituted)-4-oxo-1, 3estimation of elevated fasting blood glucose level. The thiazolidin-2 ylidene]-2-(pyridine-2-yloxy) rats having blood glucose level above 200 mg/dl of acetohydrazides were synthesized using appropriate blood were selected for the study. synthetic route and screened for antidiabetic activity. It can be concluded that, the number of compounds Groups Design: showed antidiabetic activity, out of which 9 and 10 These rats were divided into various groups with 6 rats showed appreciable antidiabetic activity. Thus research each. The rats in group I (control) were administered work was undertaken for substitution at 3 position of distilled water orally. Group II was treated as the thiazolidinone ring. The encouraging results showed diabetic control (Alloxan 150 mg/kg, i.p.). Group III may lead to the development of novel antidiabetic drugs was treated with metformin (5mg/kg, orally), while if explored further. groups IV, V, VI, VII, VIII were treated with test compounds. Treatment with compounds was started on

Animals: Wistar albino rats of either sex weighing between 150 200 g were used for the study. The animals were housed in standard environmental conditions of temperature (2520C), humidity (5510%) and light (12:12 hr light: dark cycle). Rats were supplied with standard laboratory diet and water ad libitum. Animals were deprived of food for at least 18 hrs but were allowed free access to drinking water.

the 6th day of Alloxan treatment (i.e. Day 1) and was continued for 8th day (i.e. Day 3), 12th day (i.e. Day 7) of Alloxan treatment. Before this treatment, intraday serum glucose estimation was also carried out (i.e. after 0hr, 1hr, 3hr, 5hr, and 7hr on the 6th day of Alloxan treatment). All the drugs were given orally as a single dose. All the groups were subjected to serum glucose estimation by withdrawing 0.5 ml of blood from the -1 -1 -1 retro orbital plexus under light ether anesthesia. The 13. IR: 3326 cm (N-H), 1729 cm (C=O), 1583 cm (C=N), 3025 cm-1 (C-H); 1H NMR :( CDCl3) 6.38, blood glucose concentration was estimated in 7.69, 6.68, 7.75 (C-H, 2-pyridine), 3.76, 4.83 (CH2), spectrophotometer at 505 nm. 7.0 (-NH-), 7.14, 7.06, 7.06, 7.14, 7.07 (Phenyl ring); Sample Collection: FAB-MS: (m/z, 100%): 294 ([M+], 100%) Blood was collected from retro orbital plexus of the eye under light ether anesthesia using capillary tube. Blood was collected in fresh vials containing sodium fluoride PHARMACOLOGICAL EVALUATION: and sodium oxalate as anti coagulant. Antidiabetic activity: All the compounds synthesized were tested for antidiabetic activity, the fasting serum glucose levels were determined according to GOD-POD method.11

12: IR: 3410 cm-1 (C-H str. pyridine), 1552 cm-1 (N-H str.), 1710 cm-1, 1640 cm-1 (C=O), 1580 cm-1 (C=N), 3020 cm-1 (C-H, Aromatic str.), 1260 cm-1 (C-O-C str.); 1 H NMR (CDCl3) 6.38, 7.69, 6.68, 7.75 (C-H, 2pyridine), 3.81, 4.83 (CH2), 7.0 (-NH-), 7.26, 7.13, 7.52 (Phenyl ring); FAB-MS: (m/z, 100%): 412 ([M+ +2], 100%.

Asian J. Research Chem. 2(2): April.-June, 2009 ,

RESULTS AND DISCUSSION:

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Joy JM, et al. Evaluation of hypoglycemic effects of 4thiazolidinones. Indian Drugs. 2005; 42(1): 47. 2. Ragab FA, et al. Egypt J. Pharm. Sci. 1993; 34: 387. 3. Hassan HY, et al. Synthesis and antimicrobial activity of Pyridines bearing thiazoline and thiazolidinones moieties. Chem. Pharm. Bull. 1998; 46(5): 863. 4. Patel PB, Trivedi JJ. Synthesis of 2-aryl-3 -aryloxyethyl4-thazolidinones and their 1, 1-dioxides. J. Ind. Chem. Soc. 1977; 54:765. 5. Vittoria DM, et al. Synthesis and antihistaminic activity of some thazolidin-4-ones. J. Med. Chem. 1992; 35: 2910. 6. Omar AM, Eshba NH. J. Pharm. Sci. 1984; 73: 1166. 7. Chaudhary M, et al. CNS depressant activity of pyrimidyltiazolidones and their selective inhibition of NAD- depressant pyruvate oxidation. J. Pharm. Sci. 1976; 65: 443. 8. Bue-Vallesky, et al. United States Patent. 1996; US5:523:314. /ChemAbst, 1996; 123: 13816. 9. Panetta JA, et al. United States Patent. 1997; US5:661:168. / Chem Abst, 1997; 125: 117581. 10. OECD (2000), Guidance Documents on Acute Oral Toxicity, Environmental Health and safety Monograph Series on Testing and Assessment No 24. 11. Henry JB. Clinical and diagnosis management by laboratory methods. W. B. Saunders, H. B. J. International, 1991.

REFERENCES:
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