Syntheses of aromatic aldehyde imine derivatives of 2-thiobenzyl-1,3,4-thiadiazole and evaluation of their anticonvulsant activity

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1 Indian Journal of Chemistry Vol. 49B, February 2010, pp ote yntheses of aromatic aldehyde imine derivatives of 2-thiobenzyl-1,3,4-thiadiazole and evaluation of their anticonvulsant activity Bahar Ahmed* & Md. Yusuf Department of Pharmaceutical Chemistry, Faculty of Pharmacy, Jamia Hamdard, ew Delhi , India eceived 27 January 2009; accepted (revised) 28 May 2009 Benzyl and chlorobenzyl substituted 1, 3, 4-thiadiazole imine derivatives have been prepared by refluxing aromatic aldehyde imine derivatives and benzyl chloride/ 4-chloro benzyl chloride in ethanolic potassium hydroxide. The structures of the compounds have been determined by spectral and chemical methods. The anticonvulsant activity has been carried out using Karl et al. method. All the reported compounds show good anticonvulsant activity wherein chlorobenzyl substituted compounds show potent anticonvulsant activity against phenytoin used as the standard reference drug. eurotoxicity has been screened through the ota rod and ethanol potentiation test. The compounds did not show any neurotoxicity. Keywords: 1,3,4-Thiadioazole, maximum electroshock seizure method, rotarod method, ethanol potentiation test During the recent years there has been intense investigation of different classes of thiadiazole compounds, many of which were found to be pharmacologically active. Among them the anticonvulsant effect has been of significance importance 1-3. Dogan et al. 4 and rivastava et al. 5 synthesized several disubstituted-1, 3, 4-thiadiazoles and evaluated their anticonvulsant activity. The above compounds have poor lipophilicity and thus do not exhibit good results. We have prepared some new thiosubstituted-1,3,4- thiadiazole derivatives, which showed better anticonvulsant activity due to their carbonic anhydrase inhibitory activity. The synthesized aromatic aldehyde imines derivatives of 2-thiobenzyl-1,3,4-thiadiazole have good lipophilic activity due to sulphur and imine moieties and thus had shown comparatively better anticonvulsant activity which could be due to carbonic anhydrase inhibitory activity. Experimental ection Infrared spectra were scanned in KBr on Bio-ed FTI-spectrophotometer. uclear magnetic resonance spectra were scanned in Bruker model D-400 MHz M instrument in DMO-d 6 and CD 3 using tetramethylsilane (TM) as the internal reference. Mass spectrum was recorded on a JOEL 102\ DA mass spectrometer using Argon\ enon as FAB (Fast Atom Bombardment) gas in matrix of m-nitro benzyl alcohol with a accelerating voltage of 10 KV. All chemicals used were of analytical reagent grade. Electroconvulsometer was used to evaluate the anticonvulsant activity. otating rod with 24 rpm and ethanol were used for evaluating neurotoxicity. ynthesis of 5-amino-1, 3, 4-thiadiazole-2-thiol 6 Thiosemicarbazide 45.5 g, (0.25 mole) was suspended in absolute ethanol, and then anhydrous sodium carbonate (24 g) in carbon disulphide (0.25 mole, 46 g) was added. The mixture was heated with stirring under reflux for 1 hr and then heated on steam bath for 4 hr. The completion of the reaction was indicated by TLC ( Thin Layer Chromatography) using silica gel as stationary phase and TEF (Toluene: Ethyl acetate: Formic acid) 5:4:1 as mobile phase, yield 84%; m.p. 232ºC; 1 H M (400 MHz, DMO-d 6, TM): δ 7.99 (2H, s, H 2 ), 2.93 (H, s, H); FTI (KBr): 2553 cm -1 (-H str, thiol). yntheses of aromatic aldehyde imine derivatives of 5-amino-1,3,4-thiadiazole-2-thiol (1a-e). Amino-1,3,4-thiadiazole-2-thiol (0.02 mole) was added in benzaldehyde (0.02 mole) in 25 ml absolute ethanol. The reaction mixture was refluxed till the completion of reaction (as indicated by TLC using silica gel as stationary phase and TEF 5:4:1 as mobile phase), the reaction mixture was concentrated and kept over night below 20 o C for crystallization. The solid separated was filtered under vacuum and recrystallized from 95% ethanol (Figure 1). 5-[(2-Chlorophenyl) methylene] amino-1,3,4- thiadiazole-2-thiol 1a: Yield 55%; m.p C; 1 H M (400 MHz, DMO-d 6, TM): δ (4H, m, Ar Bz), 9.15 (H, s, CH=), 2.53 (H, s, H); FTI (KBr): (Ar C-H str, Bz), 2553 (-H str, thiol), 1610 (Ar C=C str, Bz), 1540 (C= str, imine), 720 (Ar C-H def, o-disubs-bz), 680 cm -1 (Ar C- str, Bz); FABM: m/z [M + ] 256, 220, 165, 138, 112; Anal. Calcd. for C 9 H : C, 42.27; H, 2.36;, Found: C, 42.24; H, 2.35;, 12.89%.

2 242 IDIA J. CHEM., EC B, FEBUAY 2010 H H 2 abs. ethanol H 2 + Carbon a 2 CO 3 Thiosemicarbazide disulphide O H 2 H 5-amino-1,3,4- refluxed in methanol 1a-1e thiadiazole-2-thiol H Compounds 1a, 2ia, 2iia 2-1b, 2ib, 2iib 4-1c, 2ic, 2iic 2-O 2 1d, 2id, 2iid 4-O 2 1e, 2ie, 2iie 4-F reflux in alc. KOH 2ia-2ie 2iia-2iie Figure 1 yntheses of aromatic aldehyde imines derivatives of 2-thiobenzyl-1,3,4-thiadiazole 5-[(4-Chlorophenyl)methylene]amino-1,3,4-thiadiazole-2-thiol 1b: Yield 70%; m.p C; 1 H M (400 MHz, DMO-d 6, TM): δ (4H, m, Ar Bz), 9.72 (H, s, CH=), 2.41 (H, s, H); FTI (KBr): (Ar C-H str, Bz), 2546 (-H str, thiol), 1612 (Ar C=C str, Bz), 1541 (C= str, imine), 800 (Ar C-H def, p-disubs-bz), 685 cm -1 (Ar C- str, Bz); FABM: m/z [M + ] 256, 219,165, 138, 111, 92; Anal. Calcd. for C 9 H : C, 42.27; H, 2.36;, Found: C, 42.26; H, 2.34;, 12.87%. 5-[(2-itrophenyl)methylene]amino-1,3,4-thiadiazole-2-thiol 1c: Yield 40%; m.p C; 1 H M (400 MHz, DMO-d 6, TM): δ (4H, m, Ar Bz), 9.12 (H, s, CH=), 2.54 (H, s, H); FTI (KBr): (Ar C-H str, Bz), 2556 (-H str, thiol), 1607 (Ar C=C str, Bz), 1566 (C= str, imine), 715 cm -1 (Ar C-H def, o-disubs-bz); FABM: m/z [M + +1] 268, 176, 151, 145, 92; Anal. Calcd. for C 9 H 6 4 O 2 2 : C, 40.59; H, 2.27;, Found: C, 40.56; H, 2.24;, 21.07%. 5-[(4-itrophenyl)methylene]amino-1,3,4-thiadiazole-2-thiol 1d: Yield 68%; m.p C; 1 H M (400 MHz, DMO-d 6, TM): δ (4H, m, Ar Bz), 9.74 (H, s, CH=), 2.47 (H, s, H); FTI (KBr): (Ar C-H str, Bz), 2548 (-H str, thiol), 1619 (Ar C=C str, Bz), 1548 (C= str, imine), 805 cm -1 (Ar C-H def, p-disubs-bz); FABM: m/z [M + ] 267, 176, 151, 145, 92; Anal. Calcd. for C 9 H 6 4 O 2 2 : C, 40.59; H, 2.27;, Found: C, 40.55; H, 2.23;, 21.04%. 5-[(2-Florophenyl) methylene] amino-1,3,4-thiadiazole-2-thiol 1e: Yield 52%; m.p C; 1 H M (400 MHz, DMO-d 6, TM): δ (4H, m, Ar Bz), 9.23 (H, s, CH=), 2.73 (H, s, H); FTI (KBr): (Ar C-H str, Bz), 2547 (-H str, thiol), 1611 (Ar C=C str, Bz), 1539 (C= str, imine), 719 cm -1 (Ar C-H def, o-disubs-bz); FABM: m/z [M + +1] 241, 167, 145, 121, 93; Anal. Calcd. for C 9 H 6 F 3 2 : C, 45.17; H, 2.53;, Found: C, 45.13; H, 2.53;, 17.54%. Aromatic aldehyde imine derivatives 2a-e A mixture of (0.005 mole) of aromatic aldehyde imine derivatives and mole of benzyl chloride/ 4-chloro benzyl chloride in ethanolic alkali (0.08 g KOH in 20 ml EtOH) was refluxed for specified hr. After the completion of reaction, the reaction mixture was poured into crushed ice and stirred by a glass rod. A crude precipitate was obtained, which was filtered and recrystallized from acetone (Figure 1). 5-(Benzylthio)--[(2-chlorophenyl) methylene]- 1,3,4-thiadiazol-2-amine 2ia: Yield 76%; m.p C; 1 H M (400 MHz, DMO-d 6, TM): δ (9H, m, Ar Bz), 10.1 (H, s, CH=), 4.36 (2H, s, CH 2 ); FTI (KBr): (Ar C-H str, Bz), 1608 (Ar C=C str, Bz), 1534 (C= str, imine), 718 (Ar C-H def, o-disubs-bz), 677 cm -1 (Ar C- str, Bz); FABM: m/z [M + ] 345, 221, 138; Anal. Calcd. for C 16 H : C, 55.56; H, 3.50;, Found: C, 55.53; H, 3.45;, 12.09%.

3 OTE (Benzylthio)--[(4-chlorophenyl) methylene]- 1,3,4-thiadiazol-2-amine 2ib: Yield 62%; m.p C; 1 H M (400 MHz, DMO-d 6, TM): δ (9H, m, Ar Bz), 9.98 (H, s, CH=), 4.35 (2H, s, CH 2 ); FTI (KBr): (Ar C-H str, Bz), 1610 (Ar C=C str, Bz), 1539 (C= str, imine), 780 (Ar C-H def, p-disubs-bz), 682 cm -1 (Ar C- str, Bz); FABM: m/z [M + ] 346, 222, 138; Anal. Calcd. for C 16 H : C, 55.56; H, 3.50;, Found: C, 55.53; H, 3.45;, 12.09%. 5-(Benzylthio)--[(2-nitrophenyl) methylene]- 1,3,4-thiadiazol-2-amine 2ic: Yield 70%; m.p C; 1 H M (400 MHz, DMO-d 6, TM): δ (9H, m, Ar Bz), (H, s, CH=), 4.42 (2H, s, CH 2 ); FTI (KBr): (Ar C-H str, Bz), 1607 (Ar C=C str, Bz), 1593 (C= str, imine), 719 cm -1 (Ar C-H def, o-disubs-bz),; FABM: m/z [M + ] 356, 221, 149; Anal. Calcd. for C 16 H 12 4 O 2 2 : C, 53.92; H, 3. 39;, Found: C, 53.93; H, 3.34;, 17.99%. 5-(Benzylthio)--[(4-nitrophenyl) methylene]- 1,3,4-thiadiazol-2-amine 2id: Yield 68%; m.p ºC. 1 H M (400 MHz, DMO-d 6, TM): δ (9H, m, Ar Bz), (H, s, CH=), 4.52 (2H, s, CH 2 ); FTI (KBr): (Ar C-H str, Bz), 1611 (Ar C=C str, Bz), 1598 (C= str, imine), 777 cm -1 (Ar C-H def, p-disubs-bz); FABM: m/z [M + +1] 357, 222, 148; Anal. Calcd. for C 16 H 12 4 O 2 2 : C, 53.92; H, 3. 39;, Found: C, 53.91; H, 3.32;, 17.95%. 5-(Benzylthio)--[(4-fluorophenyl) methylene]- 1,3,4-thiadiazol-2-amine 2ie: Yield 71%; m.p C; 1 H M (400 MHz, DMO-d 6, TM): δ (9H, m, Ar Bz), (H, s, CH=), 4.40 (2H, s, CH 2 ); FTI (KBr): (Ar C-H str, Bz), 1603 (Ar C=C str, Bz), 1536 (C= str, imine), 783 cm -1 (Ar C-H def, p-disubs-bz); FABM: m/z [M + +1] 331, 222, 123; Anal. Calcd. for C 16 H : C, 55.56; H, 3.50;, Found: C, 55.53; H, 3.45;, 12.09%. 5-(4-Chlorobenzylthio)--[(2-chlorophenyl) methylene]-1,3,4-thiadiazol-2-amine 2iia: Yield 40%; m.p C; 1 H M (400 MHz, DMO-d 6, TM): δ (8H, m, Ar Bz), 10.2 (H, s, CH=), 4.41 (2H, s, CH 2 ); FTI (KBr): (Ar C-H str, Bz), 1610 (Ar C=C str, Bz), 1528 (C= str, imine), 778 (Ar C-H def, p-disubs-bz), 702 (Ar C-H def, o-disubs-bz), 675 cm -1 (Ar C- str, Bz); FABM: m/z [M + +1] 381, 138, 125; Anal. Calcd. for C 16 H : C, 50.53; H, 2.92;, Found: C, 50.53; H, 2.91;, 11.09%. 5-(4-Chlorobenzylthio)--[(4-chlorophenyl) methylene]-1,3,4-thiadiazol-2-amine 2iib: Yield 61%; m.p C; 1 H M (400 MHz, DMO-d 6, TM): δ (8H, m, Ar Bz), (H, s, CH=), 4.40 (2H, s, CH 2 ); FTI (KBr): (Ar C-H str, Bz), 1610 (Ar C=C str, Bz), 1528 (C= str, imine), 778 (Ar C-H def, p-disubs-bz), 673 cm -1 (Ar C- str, Bz); FABM: m/z [M + ] 380, 138, 125; Anal. Calcd. for C 16 H : C, 50.53; H, 2.92;, Found: C, 50.51; H, 2.89;, 11.04%. 5-(4-Chlorobenzylthio)--[(2-nitrophenyl) methylene]-1,3,4-thiadiazol-2-amine 2iic: Yield 60%; m.p C; 1 H M (400 MHz, DMO-d 6, TM): δ (8H, m, Ar Bz), 10.34(H, s, CH=), 4.43 (2H, s, CH 2 ); FTI (KBr): (Ar C-H str, Bz), 1609 (Ar C=C str, Bz), 1530 (C= str, imine), 780 (Ar C-H def, p-disubs-bz), 702 cm -1 (Ar C- H def, o-disubs-bz),; FABM: m/z [M + +1] 391, 150, 125; Anal. Calcd. for C 16 H 11 4 O 2 2 : C, 49.17; H, 2.84;, Found: C, 49.13; H, 2.83;, 14.33%. 5-(4-Chlorobenzylthio)--[(4-nitrophenyl) methylene]-1,3,4-thiadiazol-2-amine 2iid: Yield 53%; m.p C; 1 H M (400 MHz, DMO-d 6, TM): δ (8H, m, Ar Bz), 10.39(H, s, CH=), 4.51 (2H, s, CH 2 ); FTI (KBr): (Ar C-H str, Bz), 1603 (Ar C=C str, Bz), 1526 (C= str, imine), 772 cm -1 (Ar C-H def, p-disubs-bz); FABM: m/z [M + ] 390, 150, 126; Anal. Calcd. for C 16 H 11 4 O 2 2 : C, 49.17; H, 2.84;, Found: C, 49.14; H, 2.85;, 14.35%. 5-(4-Chlorobenzylthio)--[(4-fluorophenyl) methylene]-1,3,4-thiadiazol-2-amine 2iie: Yield 56%; m.p C; 1 H M (400 MHz, DMO-d 6, TM): δ (8H, m, Ar Bz), 9.23 (H, s, CH=), 2.73 (H, s, H); FTI (KBr): (Ar C-H str, Bz), 2547 (-H str, thiol), 1611 (Ar C=C str, Bz), 1539 cm -1 (C= str, imine); FABM: m/z [M + +1] 365, 122, 125; Anal. Calcd. for C C 16 H 11 F 3 2 : C, 52.81; H, 3.50;, Found: C, 52.83; H, 3.51;, 11.40%. The practical yields of the following prepared compounds were in the following decreasing order 2i (a) > 2i (e) > 2i (c) > 2i (d) > 2i (b) and 2ii (b) >2ii (c) >2ii (e) >2ii (d) > 2ii (a), and the melting points were found as 2i (e) < 2i (d) < 2i (b) < 2i (a) < 2i (c) and 2ii (a) < 2ii (b) < 2ii (e) < 2ii (a) < 2ii (c). These data showed that p-chlorobenzyl derivatives had low yields and melting points as compared to benzyl derivatives. Anticonvulsant activity The investigations were conducted on albino mice of either sex (25-30 g). The albino mice were kept under standard conditions at an ambient temperature

4 244 IDIA J. CHEM., EC B, FEBUAY 2010 Table I Anticonvulsant and neurotoxicity activity of the synthesized compounds by ME test in albino mice Compd ME-induced hind limb extension (% Protection) * ecovery otarod test # esult of ethanol Potentiation test $ 2ia 83.3 oon (+) (+) 2ib 83.3 oon (+) (+) 2ic 83.3 oon (+) (+) 2id 83.3 oon (+) (+) 2ie 100 oon (+) (+) 2iia 100 Very soon (+) (+) 2iib 100 Very soon (+) (+) 2iic 100 Very soon (+) (+) 2iid 100 Very soon (+) (+) 2iie 100 Very soon (+) (+) 5-Amino-1,3,4-50 oon ( ) ( ) thiadiazole-2-thiol Phenytoin 100 Very soon (+) (+) *Test compounds and Phenytoin (reference) dissolved in propylene glycol were administered i.p 20 mg/ kg in a group of 6 mice each, 30 min before the test. # Dose was administered i.p and neurotoxicity was measured after 30 min. $ Mice were treated with the test compounds and 1 hour later with ethanol 2.5 g/kg i.p neurotoxicity was measured after 30 min. The signs indicates 50% or more passed (+) or fail ( ) the neurotoxicity testing. of 25±2 C; Food and water were withdrawn prior to the experiment. The standard drug phenytoin and the test drugs were given ip. at the dose of 20 mg/ kg in propylene glycol. The anticonvulsant activity was evaluated by ME test method (Maximal electroshock seizure method) 7. upra maximal electroshock of current intensity of 54 ma, 60 Hz was given to mice for 0.2 sec. The abolition of the hind limb tonic extensor spasm was recorded as an increased anticonvulsant activity (Table I). eurotoxicity Test otarod Test 8 : The disruptive effects on motor coordination were assessed using the otarod thread mill mouse test. The animals were placed on a rotating rod (24 rpm) and then observed for 5 min. The skeletal muscle relaxation induced by a test compound could be evaluated by testing the ability of mice or rats to remain on a revolving rod. This forced motor activity has subsequently been used by many investigators. The dose, which impairs the ability of 50% of the mice to remain on the revolving rod is considered the endpoint. Dose was administered intraperitonially and neurotoxicity was measured after 30 min. 50% or more passed the neurotoxicity testing, (-) shows 50% or more failed the neurotoxicity testing. Ethanol Potentiation Test 9 : Mice were treated with the test compound and 1 hr later with ethanol (2.5 g/kg i.p). This dose of ethanol did not induce lateral position in the control animals. The number of mice that were in the lateral position after receiving ethanol in each group was determined (Table I). esults and Discussion The parent compound 5-amino-1,3,4-thiadiazole-2- thiol showed 50% protection towards ME-induced hind limb extension and showed soon recovery but it did not pass the neurotoxicity tests perfectly. After the formation of their imines-thiobenzyl derivatives (2ii series) it was inferenced that chlorobenzylated imines have shown 100% protection towards ME-induced hind limb extension with fast recovery and haven t shown any neurotoxicity. On the other hand benzylated imines also showed 83.3% protection towards ME-induced hind limb extension with slow recovery and have not shown any neurotoxicity.

5 OTE 245 C 7 H 5 C 7 H 6 C 7 H 7 C 16 H [M] + C 9 H C 2 m\z 89 C 2 2 m\z: 107 C 16 H m\z: 307 Figure 2 Mass fragmentation pattern of the series compounds The I spectra of compound exhibited absorption bands in the range cm -1 due to aromatic CH bonds stretching, cm -1 due to H, cm -1 due to aromatic C-H groups, cm - 1 due to imine C= stretching and cm -1 due to C-H. Other peaks in the I spectra of the compounds were found quite informative and characteristic. The M spectra also displayed the characteristic peaks for different chemical compounds. The peaks in the range were due to different aromatic protons. The peaks in the range 8-10 were characteristic to imino-proton; peak in the range was assigned to thiol-proton. The peaks in the range were also found due to methylene group. Other peaks in the M spectra of different compounds were also characteristic to the particular protons in the compounds. The structure of the compounds was substantiated with the help of mass spectral fragmentation pattern of the selected compounds (Figure 2). Chlorobenzyl substituted compounds have showed activity same as like as the standard reference Phenytoin, while the normal benzyl derivatives have also showed good activity, but not much as the chlorobenzyl substituted compounds, which indicated that chlorine or other electronegative groups have increased the anticonvulsant activity. All the compounds have passed the neurotoxicity test. Acknowledgements The authors are thankful to the M ection Jamia Hamdard, ew Delhi for scanning the M spectra, and Arbro Pharmaceuticals for scanning I spectra.

6 246 IDIA J. CHEM., EC B, FEBUAY 2010 One of the authors (MY) is also thankful to UGC, ew Delhi for awarding the GATE scholarship. eferences 1 Chapleo C B, Myers P L, aville J F, mith A C B, tillings M, Tulloch I F, Walter D &Welbourn A P, J Med Chem, 29, 1986, tillings M, Walter D & Welbourn A P, J Med Chem, 29, 1986, Chapleo C B, Myers P L, aville J F, mith A C B, Tulloch I F, Walter D & Welbourn A P, J Med Chem, 30, 1987, Dogan H, Duran A, ollas, ener G, Uysal K M & Gulen D, Bioorg Med Chem 10, 2002, rivastava K, rivastava & rivastava D, Indian J Chem, 38B, 1999, Petrow V, tephenson O, Thomas A J & Wild, J Amer Chem oc, 2, 1952, Kral A C, Penny J K, While B G, Kupferlerg H G & wingard E A, Epilepsia, 19, 1978, Vogel W H, chcllens B A, andow J, Muller G & Vogel W F, Drug Discovery and evaluation Pharmacological assay, (pringer-verlag, Berlin Heidelberg) erci F, Pocar D, Guido M, Loche A & Perlini V, J Med Chem, 44, 2001, 931.