Title of Invention	NOVEL COMPOUNDS FOR THE MANAGEMENT OF AGING-RELATED AND DIABETIC VASCULAR COMPLICATIONS, PROCESS FOR THEIR PREPARATION AND COMPOSITION COMPRISING THEM.
Abstract	TITLE: NOVEL COMPOUNDS FOR THE MANAGEMENT OF AGING-RELATED AND DIABETIC VASCULAR COMPLICATIONS, PROCESS FOR THEIR PREPARATION AND COMPOSITION COMPRISING THEM. The invention disclose novel compounds of the pyridinium series useful for the managment of diabetes and aging-related vascular complilcations, including kidney disease, nerve damage, atherosclerosis, retinopathy, dermatological disprders and discoloration of teeth by breakeing performed AGE. The invention also discloses, method for preparation of the novel compounds of the series and pharmaceutical composition having one or more compounds as defined above as active ingredients.

Title of Invention

NOVEL COMPOUNDS FOR THE MANAGEMENT OF AGING-RELATED AND DIABETIC VASCULAR COMPLICATIONS, PROCESS FOR THEIR PREPARATION AND COMPOSITION COMPRISING THEM.

Abstract

TITLE: NOVEL COMPOUNDS FOR THE MANAGEMENT OF AGING-RELATED AND DIABETIC VASCULAR COMPLICATIONS, PROCESS FOR THEIR PREPARATION AND COMPOSITION COMPRISING THEM. The invention disclose novel compounds of the pyridinium series useful for the managment of diabetes and aging-related vascular complilcations, including kidney disease, nerve damage, atherosclerosis, retinopathy, dermatological disprders and discoloration of teeth by breakeing performed AGE. The invention also discloses, method for preparation of the novel compounds of the series and pharmaceutical composition having one or more compounds as defined above as active ingredients.

Full Text	FIELD OF THE INVENTION The present invention relates to a new class of compounds of pyridinium series and to their use in treatment of diabetes and related illnesses. More particularly the invention relates to compounds of this series, methods for their preparation, pharmaceutical composition containing these compounds and their use in the treatment of complications of diabetes mellitus. The compounds of this series exhibit AGE breaking activity, which is essential for the treatment of diabetic and aging-related complications including kidney disease, nerve damage, atherosclerosis, retinopathy and dennatological conditions. The invention also extends to the method of reversing the discoloration of teeth resulting from nonenzymatic browning in the oral cavity which comprises administration of an amount effective to reverse pre-formed advanced glycosylation crosslinks. BACKGROUND OF THE INVENTION Maillard in 1912 found that reducing sugars, such as glucose and ribose react with proteins to form brown pigments. Further studies have shown that this is an irreversible non-enzymatic reaction, which occurs in several natural systems including stored foodstuff. Maillard reaction occurs in two stages, early and advanced. Initially, proteins react with glucose to form stable Amadori products, which subsequently cross-links to form advanced glycation end products (AGE). In most cases, the formation of AGE also accompanies browning of the proteins and increase in the fluorescence. In diabetes, where blood glucose level is significantly higher than normal, the reaction of glucose with several proteins such as haemoglobin, lens crystallin and collagen, gives rise to the formation of AGE, which in turn, is responsible for the complications associated with diabetes, such as nephropathy, microangiopathy, endothelial dysfunction and other organ dysfunctions. In addition, the activity of several growth factors, such as basic fibroblast growth factor, is also impaired. AGE products, unlike normal proteins in tissue, have a slower rate of turnover and replenishment. It has been reported that AGE products may in fact elicit a complex immunological reaction involving RAGE (Receptor for Advanced Glycation End Products) receptors and activation of several incompletely defined immunological processes. It has been documented that diabetes with evidence of microangiopathy and macroangiopathy also show evidence of oxidative stress, the mechanism of which has not been elucidated. In vitro AGE formation can be studied in the laboratory by incubating reducing sugars, such as ribose or glucose with bovine serum albumin. AGE formation can be detected by increase in the fluorescence or increased cross reactivity with anti-AGE antibodies. The increase in fluorescence seems to precede formation of AGE specific antigenic epitopes. This increase in fluorescence is used to monitor the increased AGE formation in vitro (Brownlee M et al, Science 1986; 232:1629-1632). In addition to the increase in the fluorescence, one of the most important features of in vitro AGE formation is the formation of antigenic epitopes that are specific to AGE and not to the native proteins. Therefore, it is possible to raise antibodies against advanced glycation end products of one protein and use them to detect AGE formation in other proteins. This has served as an important analytical tool in AGE research. Due to the clinical significance of AGE formation, many approaches are being used to diagnose, prevent, or revert AGE formation in the body. The formation of AGE could be inhibited by reacting with an early glycosylation product that results from the original reaction between the target protein and glucose. The inhibition was believed to take place as the reaction between the inhibitor and the early glycosylation product appeared to interrupt the subsequent reaction of the glycosylated protein with additional protein material to form the cross linked late stage product. Compounds like aminoguanidine act to inhibit AGE formation by such mechanism. The formation of AGE on long-lived proteins is also associated with cross- linking of these proteins. The AGE derived protein cross-links have been shown to be cleaved by compounds like N- phenacyl thiazolium bromide (PTB), which reacts with and cleaves covalent, AGE derived protein cross links (Vasan et al. Nature 1996; 382: 275-278 ; US 5,853,703, Date of Patent : Dec. 29, 1998). The mechanism of reducing the AGE content in tissues is expected to take place relatively rapidly, in contrast to aminoguanidine, which acts slowly by its very nature of mechanism of action. This current specification is related to compounds of pyridinium class, which break pre-formed AGE, like PTB, and in some cases even more effectively by than PTB. SUMMARY OF THE INVENTION The main objective of the present invention is to provide a new class of compounds of the pyridinium series which are useful for the management of diabetes and aging related vascular complications and particularly in the treatment of complications of diabetes mellitus and other aging related conditions including kidney disease, nerve damage, atherosclerosis, retinopathy and dermatological conditions. The invention also extends the method to reverse the discoloration of teeth resulting from nonenzymatic browning in the oral cavity which comprises administration of an amount effective to reverse the pre-formed advanced glycosylation crosslinks, etc. Another object of the present invention is to provide compounds of the pyridinium series, which exhibit AGE breaking activities. Yet another object of the present invention is to provide a method of preparation of compounds of the pyridinium series which exhibit AGE breaking activities. Still another object of the invention is to provide pharmaceutical compositions with a new class of compounds of the pyridinium series according to the invention and their pharmaceutically acceptable salts in combination with suitable carriers, solvents, excepients, diluents and other media normally employed in preparing such compositions. Still another object of the invention is to provide a method of treatment of a diabetic patient by administration of the compounds of the invention, either singly or in combination with drugs for anti-diabetic therapy, or pharmaceutically acceptable salts thereof in required dosage in admixture with pharmaceutically acceptable diluent, solvent, excepients, carriers or other media as may be appropriate for the purpose. DETAILED DESCRIPTION OF THE INVENTION The present invention provides for a new class of AGE breakers, of general formula I, wherein R1 is -R4-R5 or -N(R7) N (R7) R9; R4 is selected from the group -N(R7)R6O, -N(R7)R6N(R7)-, OR6O, -OR6N(R7)-, where Re is alkyl; R5 is selected from the group alkyl, aryl, including heteroaryl, -COR7, SO2R7, where R7 is selected from the group H, alkyl or aryl, including heteroaryl; R2 is selected from the group F, Cl, Br, I, OR7, NO2, alkyl, aryl including heteroaryl, formyl, acyl, C(O)NR7R10, C(O)OR7, NR7R10, N=C(R7)(R10), SR7, SO2NH2, SO2 alkyl and SO2aryl, and m is 0, 1 or 2 R3 is selected from the group R7, OR7, N(R7) (R10), N=C(R7) (Rio), N(R7) N(R7) (R10), N(R7) N=C(R7) (R,o) and CH(R7)C(O)R8 where Rs is selected from the group R7, OR7 and NR7R10; R9 is selected from the group consisting of hydrogen, alkyl, aryl, including heteroaryl, C(O)R10, -SO2R10, -C(S)NHR10, -C(NH) NH (R10), -C(O) NHR10, Rio is selected for the group H, alkyl or aryl, including heteroaryl and in each case optionally different from substituent R7 X is selected from group consisting of a halide ion, acetate ion, perchlorate ion, sulfonate ion, oxalate ion, citrate ion, tosylate ion, maleate ion, mesylate ion, carbonate ion, sulfite ion, phosphoric hydrogen ion, phosphonate ion, phosphate ion, BF4-, PF6-, etc. with proviso that (i) when two alkyl groups are present on the same carbon or nitrogen, they are optionally linked together to form a cyclic structure and (ii) the nitrogen of heteroaryl ring of R10, when present, is optionally quaternized with compound such as X-CH2C(O)-R.3 As used herein, "alkyl" refers to an optionally substituted hydrocarbon group joined by single carbon-carbon bonds and having 1 to 8 carbon atoms joined together. The alkyl hydrocarbon group may be linear, branched or cyclic, saturated or unsaturated. The substituents are selected from F, Cl, Br, I, N, S, 0 and aryl. Preferably, no more than three substituents are present. As used herein "aryl" refers to an optionally substituted aromatic group with atleast one ring having a conjugated pi- electron system, containing upto two conjugated or fused ring systems. Aryl includes carbocyclic aryl, heterocyclic aryl and biaryl groups, all of which may be optionally substituted. The substituents are selected from F, Cl, Br, I, N, S. O and straight chain or branched C1-C6 hydrocarbon. The novel compounds of the invention of general formula I having m as 0 and - COR1 at position 3 are listed in Table 1A and the novel compounds of the invention of general formula I having m as 0 and - COR1 at position 4 are listed in Table 1B. The following compounds suggested are by way of example alone of the representative compounds of the general formula I as defined above and in no way restrict the invention. l-[1-(2-thien-2-yl-2-oxoethyl)-6-methyl-3-carbonylpyridinium]-2-[l- (2-Thien-2"-yl-2-oxoethyl )-3-carbonyl pyridinium ] hydrazine dichloride(compound no: 1). 1 -(2-thien-2"-yl-2-oxoethyl)-3-(isopropylsulfonyl hydrazino carbonyl) pyridiniiun broraide(compound no: 2.). 1-(2-(4-benzyl piperidin-1 -yl)-2-oxoethyl)-3-(methanesulfonyl hydrazino carbonyl) pyridinium chloride(compound no: 3 ). 1-(2-(2-ethoxy carbonyl pyrrolidin-l-yl)-2-oxoethyl)-3-(methanesulfonyl hydrazino carbonyl) pyridinium chloride, (compound no: 40- 1-(2-thien-2"-yl-2-oxoethyl)-3-(raethanesulfonyl hydrazino carbonyl )-5- bromo pyridinium bromide, (compound no: 5"). 1-(2-thien-2"-yl-2-oxoethyl)-3-(ethoxycarbonyl hydrazino carbonyl) pyridinium bromide, (compound no: 6 ). 1-(2-(5-chlorothien-2-yl)-2-oxoethyl)-3-(methanesulfonyl hydrazino carbonyl) pyridinium bromide (compound no: 1 ). N,N"-bis[3-carbonyl-1-(2-(4-nitrothien-2-yl)-2-oxoetJiyl)pyridinium] hydrazine dichloride. (compound no: 8). 1-(2-thien-2"-yl-2-oxoethyl)-3-(methanesulfonyl hydrazino carbonyl) -6-methyl pyridinium bromide, (compound no: 9 ). N,N"-bis[3-carbonyl-1-(2-(5-methylthien-2-yl)-2-oxoethyl) pyridinium] hydrazine dichloride. (compound no: 10). N,N"-bis[3-carbonyl-1-(2-(2-ethoxycarbonyl pyrrolidin-l-yl)-2- oxoethyl) pyridinium] hydrazine dichloride. (compound no:n ). 1-[1-(2-thien-2"-yl-2-oxoethyl)-5-aminocarbonyl-3-carbonyl pyridinium]-2-[1-(2-Thien-2"-yl-2-oxoethyl )-3-carbonyl pyridinium] hydrazine dichloride (compound no: 12). 1-(2-(4-carbethoxy-thiazolidin-3-yl)-2-oxoethyl)-3-(methanesulfonyl hydrazino carbonyl) pyridinium chloride (compound no: 13 ). N,N"-bis[3-carbonyl-1-(2-(5-chlorothien-2-yl)-2-oxoethyl)pyridinium] hydrazine dichloride. (compound no: 14). 1-(2-(5-methylthien-2-yl)-2-oxoethyl)-3-(methanesulfonyl hydrazino carbonyl) pyridinium chloride (compound no:15). 1 -(2-(4-nitrothien-2-yl)-2-oxoethyl)-3 -(methane sulfonyl hydrazino carbonyl) pyridinium bromide, (compound no;16"). 1-(2-phenylamino-2-oxoethyl)-3-(phenyl hydrazino carbonyl) pyridinium chloride (compound no: 11). 1-(2-phenylamino-2-oxoethyl)-4 -[ 2-(benzoyloxy) ethylamino carbonyl ] pyridinium chloride (compound no: 18"). 1 -2-(5-nitro-thien-2-yl)-2-oxoethyl)-3-(methanesulfonyl hydrazino carbonyl) pyridinium chloride (compound no: .19). 1 -(2-thien-2"-yl-2-oxoethyl)-3-(trifluoromethanesulfonyl hydrazino carbonyl) - pyridinium bromide, (compound no. 20,). 1-(2-thien-2"-yl-2-oxoethyl)-3-(phenyl hydrazino carbonyl) pyridinium bromide (compound no.21). 1-(2-thien-2"-yl-2-oxocthyl)-3-(p-methoxy phenyl sulfonyl hydrazino carbonyl) pyridinium bromide (compound no. 22). 1-(2-ethoxy-2-oxoethyl)-3-(phenyl aminocarbonyl hydrazino carbonyl) pyridinium bromide (compound no. 23). 1-(2-ethoxy-2-oxoemyl)-3-(p-toluene sulfonyl hydrazino carbonyl) pyridinium bromide (compound no. 24). 1-(2-phenyl-2-oxoethyl)-3-(phenylamino carbonyl hydrazino carbonyl) pyridinium bromide (compound no. 25). 1 -(2-phenylamino-2-oxoethyl)-3-(benzyl sulfonyl hydrazino carbonyl) pyridinrumchloride. (compound no. 26 ). 1-(2-phenyl-2-oxoethyl)-4-(methanesulfonyl hydrazino carbonyl) pyridinium bromide (compound no. 27). 1-(2-phenyl-2-oxoethyl)-3-(phenyl hydrazino carbonyl) pyridinium bromide, (compound no. 28). 1-(2-ethoxy-2-oxoethyl)-4-[2-(benzoyloxy) ethyl amino carbonyl ] pyridinium bromide (compound no. 29 ). 1-(2-ethoxy-2-oxoethyl)-3-(phenyl hydrazino carbonyl) pyridinium bromide.(compound no.3o ). 1-(2-phenyl-2-oxoethyl)-3-(p-methoxyphenyl sulfonyl hydrazino carbonyl) pyridinium bromide, (compound no. 31). 1-(2-phenyl-2-oxoethyl)- 4-[2-(benzoyloxy) ethyl amino carbonyl ] pyridinium bromide, (compound no.32.). 1-(2-ethoxy-2-oxoethyl)- 4-(p-methanesulfonyl hydrazino carbonyl ) pyridinium bromide, (compound no. 33). In place of specific halide salts of the compounds listed above, these compounds may also be in the form of other pharmaceutically acceptable salts falling within the definition of X as given above. The words "and pharmaceutically acceptable salts thereof as used herein following the names of specific compounds of general formula (I) of the invention means that such compounds encompass other pharmaceutically acceptable salts falling within the definition of X. According to the embodiment of the present invention, the present compounds are used for the treatment of diabetic complications, and aging related vascular and neurovascular complications including kidney disease, nerve damage, atherosclerosis, retinopathy, inflammatory discorders, immunological disorders, oxidative stress, dermatological conditions, and cosmetic conditions including colouration of teeth occurring due to the higher levels of preformed AGE. The increased levels of preformed AGE can be brought under control by breaking the AGE products using compounds mentioned in the invention. The invention also provides a process for the preparation of novel compounds of the pyridinium series. The novel compounds of general formula I, are prepared from properly substituted pyridine derivatives followed by quarternization with appropriate reagent by refluxing in alcoholic solvents like, methanol, ethanol, propanol, etc and high boiling solvents like toluene or xylene etc, for 6-48 hrs. to give the desired compounds. The examples of substituted pyridine derivatives which can be used for preparation of specific compounds of the invention are given below: 1. N,N"-bis(nicotinyl)hydrazine 2. 3-[(2-pyridyl)hydrazinocarbonyl]pyridine 3. 3-[2-methanesulfonyl)hydrazinocarbonyl]pyridine 4. 3-[(2-benzoyloxy)ethylamiiiocarbonyl]pyridine 5. 3-[(2-phenylsulfonyl)hydrazinocarbonyl] pyridine 6. 3-[(2-acetoxy)ethyloxycarbonyl]pyridine 7. 3-[(2-benzoyloxy)ethyloxycarbonyl]pyridine 8. 3-[(2-methoxy)ethyloxycarbonyl]pyridine 9. 3-[(2-phenylaminocarbonyl)hydrazinocarbonyl]pyridine 10. 3-[(2-acetoxy)ethylaminocarbonyl]pyridine 11. 3-[(2-(4-methylphenyl sulfonylhydrazinocarbonyl))]pyridine 12. 3-[(2-benzoyl)- hydrazino carbonyl]pyridine 13. 3-[(2-phenyhnethane sulfonyl) hydrazino carbonyl] pyridine 14. 3-[(2-(3-cyclohexylpropanoyl) hydrazino carbonyl]pyridine 15. 3-[(2-methoxy)ethylaniinocarbonyllpyridine 16. 3-[l-oxo-1 -(2-methoxycarbonyl)pyridyl Jhydrazine pyridine The examples of quaternizing agents which may be used in the reaction are given below: 1. 2-bromoacetyl thiophene 2. 2-chloroacetyl thiopene 3. phenacylbromide 4. phenacyl chloride 5. 2,4-dichllrophenacylbromide 6. N-phenyl chloroacetamide 7. N-cyclopropyl chloroacetamide 8. ethylbromoacetate 9. bromo acetylfuran 10. N-isopropyl chloroacetamide 11. N-chloroacetyl-2-pyrrolidinone 12. chloroacetic acid In-vitro screening for AGE-breaking Activity The in vitro AGE formation, studied in the laboratory, by incubating reducing sugar glucose, with protein bovine serum albumin, resulted in browning of solution and increase in the fluorescence. Fluorescence was used as the criteria to monitor the increased AGE formation. Example 1 AGE breaker activity has been confirmed by the screening procedure as mentioned below: Materials: Bovine serum albumin (fraction V) (BSA) Glucose, analytical grade Phosphate buffered saline (PBS) Equipment: Microplate ELISA Reader - Spectramax Plus (Molecular Devices, USA) Microplate washer, (Bio -Tec Instruments, USA) pH meter Methods of experiment: Elisa (Enzyme Linked Immunosorbent Assay) 160 mg/ml of protein, bovine serum albumin, BSA and 1.6M glucose sugar were dissolved in phosphate buffered saline, PBS. Sodium azide was added at 0.02% concentration as a preservative. The solution was filtered asceptically through a 0.22 µM filter and kept for aging at 37°C for 16 weeks. After 16 weeks the solution was dialyzed against PBS, aliquoted and stored at - 20°C. To determine the AGE breaking activity, 10µg/ml of the 16 weeks AGE-BSA was incubated with different concentrations of the test compounds at 37°C for 24 hours and AGE breaking activity of the test compounds by ELISA was determined. ELISA was performed as follows: 1. Different concentrations of 16 weeks AGE-BSA were coated on a microtitre plate as standard. Each concentration is coated in triplicates. 2. The test samples were coated on microtitre plate at a concentration of 5 ng. to 20 ng per well in triplicates. 3. The plate was incubated at 37°C for one hour. 4. After incubation the plate was washed with PBST (PBS with 0.05% Tween 20). 5. Blocking with 5% skimmed milk in PBS at 37°C for one hour was done. 6. The plate was washed with PBST. 7. Primary antibody against AGE-BSA was added and the plate is incubated at 37°C for one hour. 8. The plate was washed with PBST 9. Secondary antibody anti rabbit HRPO (Horse-Radish Per Oxidase) conjugate was added and the plate is incubated at 37°C for one hour. 10. The plate was washed with PBST. 11. Colour development with OPD (orthophenylenediamine dihydrochloride) and hydrogen peroxide was done. 12. OD (optical density) at (450nm reading - 620nm reading) was measured after incubation at 37°C for 15 minutes with Microplate ELISA Reader. The breaker activity of the compounds were determined by the following formula: OD450-620Control= Absorbance of 20ng AGE-BSA after incubation at 37°C for 24 hours without test compound OD450-620Test= Absorbance of 20ng AGE-BSA after incubation at 37°C for 24 hours with required concentration of test compound Using specific examples, the % AGE breaking activity was calculated and recorded in Table 2. Hence compounds -1,2,5-10,12,14,16,17,19 and 20 have superior AGE breaking activity compared to PTB, of which the potency of compounds .1,8-10,12,14,16,17,19 and 20 are significantly much higher. The following examples give method of preparation of the specific novel compounds of the invention as given in Table 1. The following compounds suggested are by way of example alone and in no way restrict the invention. Example 2. 1-[1-(2-Thien-2"-yl-2-oxoethvI)-6methyl-3-carbonvl pvridiniumj-2- [1-(2-Thien-2"-yl-2-oxoethvl )-3--carbonvl pyridinium ] hvdrazine dichloridefcompound no: 1 ), Yield " : 40% M.P. : 76-80 °C (dec) IR(KBr,cm-1) : 1637,1513 1HNMR (DMSO d6, 400 MHz) 5 : 11.69(2H,s), 9.59- 9.53(2H,d), 9.19(2H,m), 9.05(lH,d), 8.46-8.43(1H,t) ,8.34(lH,d), 8.27- 8.23(4H,m), 7.45-7.41 (2H,ra) ,6.56(2H,s) ,6.48(2H,s),2.81(3H,s). Mass(m/z) : 505,506,507. Example 3 1-(2-Thien-2"-vl-2-oxoethvl)-3-(isopropvlsulfonvI hvdrazino carbonvl) pyridinium bromide(compound no: 2.), Yield . : 70% M.P : 90-95°C (dec) IR(KBr.cm-1) 1638,1589 1HNMR (DMSO d6, 400 MHz) 5 : 11 .27(1H,s) ,9.91(1H,s), 9.60(1H,s) ,9.19-9.15(2H5m), 8.42-8.36(1 H,m) ,8.25-8.21(2H,m) ,7.43- 7.41(1H,t) ,6.45(2H,s), 1.35-1.34(6H,d). Mass(m/z) : 368,369,370 Example 4- 1-(2-(4-Benzvl piperidin-l-vl)-2-oxoethvl)-3-(methanesulfonvl hvdrazino carbonvl) pyridinium chloride(compound no: 3.), Yield : 17% M.P : 76-78°C IR (KBr,cm-1) : 1684,1650,1556,1540. 1HNMR (DMSO d6, 400 MHz) 5 : 11.46(1H,s) ,9.55(1H,s), 9.46(1H,s) ,9.09-9.03(2H,m), 8.36-8.32(1H,t), 7.33-7.29(2H,m), 7.23- 7.19(3H,m), 5.88-5.79(2H,m) ,4.30-4.27(1H,d) ,3.76-3.73(1H,d), 3.10(4H,m) ,2.64(1H,t) ,2.57-2.55(2H,d), 1.85(1H,bs) ,1.72- 1.63(2H,t),1.36-1.28(1H,q) ,1.13-1.03(1H,m) Mass(m/z) : 431,432,433 Example S 1-(2-(2-Ethoxv carbonvl pvrrolidin-l-vl)-2-oxoethvl)-3- (methanesulfonvl hydrazino carbonvl) pvridinium chloride, (compound no: 4 ), Yield : 14% M.P : 88-91°C IR(KBr,cm1) : 1735,1665,1539 1HNMR (DMSO d6, 400 MHz) 5 : 11.48(1H,s) ,9.96(1H,s) ,9.46(1H,s) ,9.09-9.05(2H,m) ,8.38-8.34(1H,t), 5.94-5.80(2H,q) ,4.37- 4.36(1H,d), 4.08-4.06(2H,d), 3.68-3.65(2H,m), 3.09(4H,m) ,2.23- 2.18(2H,m), 2.04 -1.93(3H,m) ,1.18-1.09(3H,t) Mass(m/z) : 399,400,401 Example 6 1-(2-Thien-2"-vl~2-oxoethvI)-3-(methanesulfonyl hydrazino carbonyl )-5-bromo pvridinium bromide, (compound no: 5.), Yield : 54% M.P : Above 190-195 °C(dec) IR(KB,cm-1) : 1682,1557,1540,1520 1HNMR (DMSO d6, 400 MHz) 8 : 11.35(1H,s) ,10.01(1H,s) ,9.57-9.54(2H,d), 9.32(1H,s), 8.26-8.22(2H,m),7.42(1H,s),6.39(2H,s), 3.08(3H,s) Mass(m/z) : 418,419,420 Example .7- 1-(2-Thien-2"-yl-2-oxoethyl)-3-(ethoxvcarbonvI hvdrazino carbonvl) pvridinium bromide, (compound no: 6 ), Yield : 69% M.P : 155-157°C IR(KBr,cm-1) : 1731,1665,1637 1HNMR (DMSO d6, 400 MHz) 5 : 11.04(1H,s), 9.59(1H,s) , 9.53(1H,s), 9.18(1H,s), 9.05-9.04(1H,d), 8.42(1H,s), 8.25- 8.23(2H,m), 7.43(1H,s), 6.46(2H,s), 4.12-4.11(2H,s), 1.23(3H,s) Mass (m/z) : 334,335,336 Example .8 1-(2-(5-chloro-thien-2-yI)-2-oxoethyl)-3-(methanesulfonyl hydrazino carbonyl) pyridinium bromide (compound no: 7-) Yield : 87% M.P : 228-230°C IR (KBr,cm-1) : 1708,1664,1631,1550 "HNMR (DMSO d6, 400 MHz) 5 : 11.40(1H,s), 9.98(1H,s), 9.50(1H,s), 9.15(1H,d) , 9.061H,d), 8.43-8.39(1H,t), 8.16-8.15(1H,d), 7.51-7.50(1 H,d), 6.41 (2H,s), 3.09 (3H,s) Mass (m/z) : 374,375,376,377 Example .9 N-N"-Bis[3-carbonyl-1-(2-(4-nitro-thien-2-yl)-2-oxoethyl)pvridinium] hvdrazine dichloride. (compound no: .8 ), Yield : 27% M.P : 204-207°C IR (KBr,cm-1) : 1681,1539,1514 1HNMR (DMSO d6, 400 MHz) 5 : 11.90(2H,s), 9.63(2H,s), 9.31-9.30(4H,m), 9.24 -9.22(2H,m), 8.87(2H,s), 8.49-8.46(2H,t), 6.56 (4H,s) Mass (m/z) : 581,582,583 Example 10 1-(2-Thien-2"-yl-2-oxoethyl-3-(methanesulfonyl hydrazino carbonyl) -6-methyl pyridinium bromide, (compound no:9 ), Yield : 14% M.P : 90-95°C(dec) IR (KBr,crn1) : 1677,1575 1HNMR (DMSO d6, 400 MHz) 5 : 11.32(1H,s), 9.97(1H,s) 9.52(1H,s), 8.94-8.92(1H,d) , 8.32-8.24(3H,m), 7.44(1H,t), 6.54(2H,s), 3.08(3H,s), 2.79(3H,s) Mass (m/z) : 354,355,356 Example 11 N-N"-Bis[3-carbonyl-1-(2-(5-methvl-thien-2-vl)-2-oxoethvn pvridiniuml hydrazine dichloride. (compound no: 10), Yield : 37% M.P : Above 166-168°C(dec) IR(KBr,cm-1) : 1666,1500 1HNMR (DMSO 4, 400 MHz) 5 : 11.73(2H,s), 9.59(2H,s), 9.19-9.15(4H,d) 8.45-8.42(2H,t), 8.06-8.05(2H,d) ,7.15-7.14(2H,d), 6.43 (4H,s), 2.59(6H,s) Mass(m/z) : 519,520,521,522 Example 12 N-N"-Bis [3-carbonvl-1 -(2-(2-ethoxy carbonvl pyrrolidin-1 -yl)-2- oxoethvl) pvridiniuml hvdrazine dichloride. (compound no: 11), Yield : 28% M.P : 118-120°C IR (KBr,cm-1) : 1660,1510 1HNMR (DMSO d6, 400 MHz) 5 : 11.75(2H,s), 9.51(2H,s), 9.20-9.10(4H,m) 8.43-8.40(2H,t), 5.97-5.83(4H,m) ,4.39-4.36(2H,m), 4.27-4.22(1H,q), 4.12-4.05(4H,m), 3.71-3.63(4H,m), 3.48-3.40(1H,m), 2.26-2.19(2H,m),2.05-1.91(5H,m), 1.30-1.27(1H,t), 1.19-1.15(5H,t) Mass(m/z) : 609,610,611 Example 13 l-[1-(2-Thien-2"-vl-2-oxoethvl)-5-aminocarbonyl-3-carbonvl pyridiniuml-2-f1-(2-Thien-2"-vl-2-oxoethyl )-3-carbonvl pvridiniuml hvdrazine dichloride (compound no: 12) Yield : 54% M.P : Above 127-129°C(dec) IR (KBr,cm-1) : 1678,1513 1HNMR (DMSO d6, 400 MHz) 5 : 11.86(2H,s), 9.83- 9.64(4H,t), 9.24 -9.23(2H,s), 8.82(1H,s), 8.48-8.45(1 H,t), 8.34(1H,s) 8.26-8.24(4H,m), 7.44 -7.42(2H,d), 6.52-6.46(4H,d) Mass (m/z) : 534,535,536 Example 14 1-(2-(4-carbethoxv-thiazolidin-3vl)-2--oxoethvl)-3-(methanesulfonyl hvdrazino carbonvl) pyridinium chloride (compound no: 13), Yield : 29% M.P : 190-192°C IR(KBr,cm-1) : 1673,1541 1HNMR (DMSO d6, 400 MHz) 5 : 11.50(1H,s), 9.55(1H,s), 9.48(1H,s), 9.12-9.08(2H,m), 8.39-8.34(1H,t), 6.04 - 5.99(2H,m) , 4.94 -4.91(1H,m), 4.87-4.84(1H,d), 4.73-4.71(1H,d), 4.28-4.23(1H,q), 4.14 -4.09(1H,q), 3.43-3.38(1H,m),3.27-3.22(1H,m), 3.10(3H,s) ,1.30- 1.27(1H,t), 1.20-1.17(2H,m) Mass(m/z) : " 439,440,441 Example 15 N-N"-Bis[3-carbonvl-1-(2-(S-chIoro-thien-2-vn-2-oxoethvl) pyridinium] hydrazine dichloride. (compound no: 14:), Yield : 35% M.P : Above 200-205°C (dec) IR(KBr,cm-1) : 1674,1590,1500 1HNMR (DMSO d6, 400 MHz) 5 : 11.90(2H,s), 9.64 - 9.61(2H,d), 9.29-9.20(4H,m) , 8.47-8.44(2H,t), 8.18-8.17(2H,d),7.51- 7.50(2H,d),6.49-6.48(4H,s) Mass(m/z) : 559,560,561,562,563,564 Example 16 1-(2-(5-Methyl-thien-2-vn-2-oxoethvl)-3-(methanesulfonyl hydrazino carbonvl) pyridinium chloride (compound no: 15), Yield : 22% M.P : 196-198°C IR(KBr,cm-1) : 1689,1657 1HNMR (DMSO d6, 400 MHz) 5 : 11.47(1H,s) ,9.98(1H,s) ,9.53(1H,s) ,9.17-9.16(1H,d), 9.09-9.07(1H,d), 8.42-8.38(1H,t), 8.06- 8.05(1H,d), 7.15-7.14(1H,d), 6.41 (2H,s), 3.09(3H,s), 2.59(3H,s) Mass(m/z) : 354,355,356,357 Example 17 1-(2-(4-Nitro-thien-2-yl)-2-oxoethyl)-3-(methanesulfonvl hydrazino carbonvl) pyridinium bromide, (compound no: 16). Yield : 52% M.P : Above 200-205°C(dec) IR(KBr,cm-1) : 1688,1631,1541 1HNMR (DMSO d6, 400 MHz) 5 : 11.41(1H,s), 9.50(1H,s) 9.309-9.306(1H,d), 9.17-9.15(1H,d) ,9.09-9.07(1H,d), 8.866- 8.862(1H,d) ,8.45-8.41(1H,t), 6.50(2H,s), 3.09(3H,s) Mass(m/z) : 385,386,387 Example 18 1-(2-Phenylamino-2-oxoethyI)-3-(Phenyl hydrazino carbonyl) pyridinium chloride (compound no: 17), Yield : 45% M.P : 165-167°C IR(KBr,cm-1) : 1679,1626,1600,1497 1HNMR (DMSO d6, 400 MHz) 8 : 11.18(1H,s), 11.10(1H,s) , 9.62(1H,s) , 9.24-9.22(1H,d), 9.17-9.15(1H,d), 8.40-8.36(1H,t), 8.19(1H,s), 7.63-7.61(2H,d) , 7.37-7.33(2H,t), 7.20-7.16(2H,t), 7.12- 7.09(1H,t), 6.88-6.86(2H,d), 6.78-6.74(1H,t) , 5.78(2H,s) Mass(m/z) : 347,348,349 Example 19 1-(2-Phenvlamino-2-oxoethyl)-4 -I 2-(benzovIoxy) ethylamino carbonvl] pyridinium chloride (compound no: 18), Yield : 40% M.P : 178-180°C IR (KBr,cm-1) : 1700,1666,1559 1HNMR (DMSO d6, 400 MHz) 5 : 11.13(1H,s), 9.74 - 9.71(1H,t), 9.23-9.22(2H,d), 8.52-8.50(2H,d), 8.01-7.99(2H,d), 7.68-7.60(3H,m) , 7.54-7.5l(2H,t), 7.36-7.32(2H,t), 7.12-7.08 (1H,t), 5.75 (2H,s), 4.47-4.45(2H,t), 3.77-3.72(2H,q). Mass (m/z) : 404,405,406 Example 20 l-2-(5-Nitro-thien-2-yl-2-oxoethvI)-3-(methanesulfonyl hydrazino carbonvl) pyridinium chloride (compound no: 19), Yield : 10% M.P : Above 105-110°C(dec) IR(KBr,cm-1) : 1680,1620 "HNMR (DMSO d6, 400 MHz) 5 : 11.48(1H,s), 9.98(l.H,s), 9.52(1H,s), 9.16-9.10(2H,m) ,8.45-8.41(1H,t) , 8.35-8.34(1H,d) ,8.25- 8.24(1H,d), 6.50(2H,s), 3.09(3H,s). Mass (m/z) : 385,386,387 Example 31 1-(2-Thien-2"-vl-2-oxoethvI)3-(TrifIuromethanesulfonvl hydrazino carbonvl)-pyridinium bromide (compound no: 20), Yield : 22% M.P : 77-79°C IR (KBr,cm-1) : 2960, 1690, 1673, 1591 1HNMR (DMSO d6, 400 MHz) 5 : 11.76(1H,s), 11.27(1H,s), 9.61(1H,s), 9.20-9.19(1H,d) ,9.07-9.05(1H,d), 8.44- 8.41(1H,t) ,8.25-8.22(2H,m), 7.34 - 7.41 (1H,m), 6.46 (2H,s). Mass (m/z) : 394,395,396 Example 22 1-(2-Thien-2"-vl-2-oxoethvl)-3-(phenvl hvdrazino carbonvl) pvridinium bromide (compound no. 21), Yield : 10% M.P : 192-194°C IR(KBr,cm-1) : 1669,1663,1603, 1HNMR (DMSO d6, 400 MHz) 5 : 10.99(1H,s), 9.54(1 H,s), 9.17-9.14(2H,t), 8.44-8.41(1H,t), 8.25-8.22(3H,m), 7.43-7.41(1H,t), 7.20-7.16(2H,t), 6.87-6.85(2H,d) , 6.79-6.75(1H,t), 6.46(2H,s) Mass(m/z) : 338,339,340 Example 23 1-(2-Thien-2"-vl-2-oxoethvl)-3-(p-methoxy phcnvl sulfonvl hvdrazino carbonvl) pvridiniuin bromide (compound no. 22.), Yield : 28% M.P : 126-]28°C IR(KBr.cm-1) : 1672,1653,1596 1HNMR (DMSO 4, 400 MHz) 5 : 11.34 -11.33(1H,d), 10.27-10.26(1H,d), 9.34(1H,s), 9.13-9.12(1H,d), 8.94-8.92(1H,d), 8.38-8.34(1H,t) ,8.24-8.19(2H,m),7.82-7.75(2H,m) ,7.42-7.40(1H,t), 7.07-7.04(2H,d) ,6.40(2H,s), 3.81(3H,s). Mass(m/z) : 432,433,434 Example 24 1-(2-Ethoxv-2-oxoethv))-3-(phenyl aminocarbonvl hvdrazino carbonvl) pyridinium bromide, (compound no. 23), Yield : 25% M.P : 183-185°C IR(KBr,cm-1) : 1746,1717,1682 1HNMR (DMSO d6, 400 MHz) 5 : 11.02(1H,s), 9.57(1H,s), 9.22-9.21(1H,d), 9.11-9.09(1H,d), 9.00(1H,s), 8.57(1H,s) ,8.44- 8.41(1H,m),7.47-7.45(2H,d) ,7.29-7.25(2H,t), 7.00-6.96(1H,t), 5.74(2H,s), 4.28-4.23(2H,q), 1.28-1.25(3H,t). Mass(m/z) : 343,344,345,346 Example 25 1-(2-Ethoxv-2-oxoethyl)-3-(p-toluene sulfonvl hvdrazino carbonvl) pvridinium bromide, (compound no. 24-), Yield : 54% M.P : 174-176°C IR(KBr,cm-1) : 1746,1712,1634 1HNMR (DMSO d6, 400 MHz) 5 : 11.33(1H,s), 10.36(1H,s), 9.37(1H,s), 9.18-9.16(1H,d), 8.93-8.91(1H,d), 8.37- 8.33(1H,t), 7.78-7.76(2H,d), 7.37-7.35(2H,d), 5.68 (2H,s), 4.26- 4.20(2H,q),2.37(3H,s),1.27-1.23(3H,t). Mass(m/z) : 378,379,380,381 Example 36 1-(2-Phenvl-2-oxoethvl)-3-(Phenylamino carbonyl hydrazino carbonvl) pyridinium bromide, (compound no.25), Yield : 70% M.P : 206-208°C IR(KBr,cm-1) : 1713,1684,1634 1HNMR (DMSO d6, 400 MHz) 5 : 11.05(1H,s), 9.55(1H,s), 9.18-9.13(2H,m), 9.02(1H,s), 8.59(1H,s), 8.49-8.45(1H,m) ,8.09- 8.07(2H,d), 7.84-7.80(1H,t) , 7.71-7.67(2H,t), 7.49-7.47(2H,d), 7.30- 7.26(2H,t), 7.01-6.97 (1H,t) ,6.56(2H,s). Mass (m/z) : 375,376,377 Example 27 1-(2-Phenylamino-2-oxoethvI)-3-(benzvI sulfonyl hydrazino carbonvl) pyridiniumchloride. (compound no.26), Yield : 48% M.P : 208-210°C IR(KBr,cm-1) : 1712,1681,1632 1HNMR (DMSO d6, 400 MHz) 8 : 11.46(1H,s) ,10.80(1H,s), 9.59(1H,s), 9.22-9.20(1H,d) ,9.08-9.06(1H,d), 8.38-8.36(1H,t),7.60- 7.58(2H,d) ,7.49(2Hsm), 7.39-7.34(5H,m),7.13-7.10(1 H,t), 5.74(2H,s), 4.52(2H,s). Mass(m/z) : 425,426,427,428 Example 28 1-(2-Phenyl-2-oxoethyl)-4-(methanesulfonvl hvdrazino carbonyl) pyridinium bromide (compound no. 27), Yield : 10% M.P : 190-192°C IR(KBr,cm-1) : 1679,1630,1650 1HNMR (DMSO d6, 400 MHz) 5 : 11.54(1H,s), 10.03(1H,s), 9.20-9.18(2H,d), 8.59-8.57(2H,d), 8.10-8.08(2H,d), 7.84 -7.80(1H,t), 7.71-7.67(2H,t), 6.56(2H,s), 3.08(3H,s). Mass(m/z) : 334,335,336 Example 2S 1-(2-PhenvI-2-oxoethyl)-3-(phenyl hvdrazino carbonvl) pvridinium bromide (compound no."28"), Yield : 36% M.P : 204-206°C IR(KBr,cm-1) : 1686,1653,1630 1HNMR (DMSO d6, 400 MHz) 6 : 11.01(1H,s) ,9.53(1H,s) ,9.17-9.16(2H,m), 8.46-8.42(1H,t), 8.09-8.07(2H,d), 7.82-7.78(1 H,t), 7.69-7.65(2H,t), 7.20-7.16(2H,t), 6.88-6.86(2H,d), 6.79-6.75(1H,t), 6.56(2H,s) Mass(m/z) : 332,333 Example 30 1-(2-Ethoxv-2-oxoethyl)-4-[2-(benzovIoxv) ethyl amino carbonvl ] pvridinium bromide (compound no.29), Yield : 82% M.P : 154-156°C IR(KBr,cm-1) : 1742,1719,1707,1675 1HNMR (DMSO d6, 400 MHz) 5 : 9.57-9.54(1H,t), 9.22- 9.20(2H,d), 8.51-8.49(2H,d), 8.00-7.98(2H,d), 7.68-7.64(1H,t), 7.54 -7.51(2H,t), 5.72(2H,s), 4.47-4.44(2H,t), 4.27-4.21(2H,q), 3.76- 3.72(2H,q), 1.27-1.24. (3H,t) Mass(m/z) : 357,358,359. Example 31 1-(2-Ethoxv-2-oxoethyl)-3-(phenvl hvdrazino carbonvl) pvridinium bromide.(compound no. 30), Yield : 37% M.P : 185-187°C IR (KBr,cm-1) : 1740,1690,1630. 1HNMR (DMSO d6, 400 MHz) 5 : 11.01(1H,s), 9.58(1H,s), 9.23-9.14(2H,m), 8.42-8.39(1H,t), 8.19(1H,s), 7.20-7.16(2H,t), 6.87- 6.85(2H,d), 6.78-6.75(1H,t), 5.75(2H,s), 4.28-4.22(2H,q), 1.28-1.24(3H,t) Mass(m/z) : 300,301,302. Example 32. 1-(2-PhenvI-2-oxoethvl)-3-(p-methoxvphenvl sulfonvl hydrazino carbonvl) pvridinium bromide (compound no. 31), Yield : 59% M.P : 188-190°C IR(KBr,cm-1) : 1671,1634,1580. 1HNMR (DMSO d6, 400 MHz) 5 : 11.26-11.25(1H,d), 10.17- 10.16(1H,d), 9.24(1H,s), 9.03-9.01(1H,d), 8.87-8.85(1H,d) , 8.31-8.27(1H,t), 7.97-7.96(2H,d), 7.74 -7.69(3H,m), 7.60-7.56(2H,t) , 6.99-6.97(2H,d), 6.40(2H,s), 3.73(3H,s). Mass(m/z) : 426,427,428,429 Example 33 1-(2-Phenvl-2-oxoethvl)- 4-[2-(benzovloxv) ethyl amino carbonvl ] pvridinium bromide (compound no.32), Yield : 92% M.P : 202-204°C IR(KBr,cm-1) : 1715,1692,1650 1HNMR (DMSO d6, 400 MHz) 5 : 9.55(1H,s), 9.14- 9.13(2H,d), 8.52-8.51(2H,d), 8.07-7.99(4H,m), 7.80-7.51(6H,m), 6.52(2H,s), 4.46(2H,s), 3.76-3.75(2H,s). Mass(m/z) : 389,390,391,392 Example 34- 1-(2-Ethoxy-2-oxoethyl)- 4-(p-methanesulfonyl hvdrazino carbonvl) pvridinium bromide, (compound no. 33), Yield . : 45% M.P : 94-96°C IR(KBr,cm-1) : 1726,1681,1643 1HNMR (DMSO d6, 400 MHz) 5 : 11.49(1H,s) ,9.98(1 H,s) ,9.23-9.21(2H,d), 8.54-8.52(2H,d), 5.73(2H,s), 4.28-4.22(2H,q), 3.09(3H,s), 1.28-1.25(3H,t). Mass(nVz) : 302,303,304,305. Pharmaceutical Compositions Pharmaceutical compositions may be prepared with a pharmaceutically effective quantity of compounds of general formula I, individually or in combination. The following pharmaceutical formulations suggested are by way of example alone and in no way restrict the forms in which they can be used. Oral formulations Oral formulations may be administered as solid dosage forms for example pellets, powders, sachets or discreet units such as tablets or capsules and like. Other orally administered pharmaceutical preparations include monophasic and biphasic liquid dosage forms either in ready to use form or forms suitable for reconstirution such as mixtures, syrups, suspensions or emulsions. The preparations in addition may contain diluents, dispersing agents, buffers, stabilizers, solubilizers, surfactants, preservatives, chelating agents and/ or other pharmaceutical additives as are used. Aqueous or non aqueous vehicle or their combination may be used and if desired may contain suitable sweetener, flavoring agent or similar substances. In case of suspension or emulsion a suitable thickening agent or suspending agent or emulsifying agent may be present in addition. Alternatively, the compounds may be administered as such in their pure form unassociated with other additives for example as capsules or sachets. It may also be administered with a vehicle. Pharmaceutical preparations can have a slow, delayed or controlled release of active ingredients as is provided by a matrix or diffusion controlled system. When the present invention or its salts or suitable complexes is presented as a discreet unit dosage form like tablet, it may contain in addition medically inert excipients as are used in the art. Diluents such as starch, lactose, dicalcium phosphate, talc, magnesium stearate, polymeric substances like methyl cellulose, fatty acids and derivatives, sodium starch glycollate, etc. may also be used. Example 35 Preparation of oral dosage form: A typical tablet has the following composition: Active ingredient of formula I as given above Lactose 135 mg Starch 76 mg Polyvinyl pyrolidone (K-30) 2 mg Talc 1-5 mg Magnesium Stearate 1.0 mg Parenteral Formulations For parenteral administration, the compounds or their salts or suitable complexes thereof may be present in a sterile vehicle which may be an aqueous or non aqueous vehicle or a combination thereof. The examples of vehicles are water, ethyl oleate, oils and derivatives of polyols, glycols and their derivatives. It may contain additives common in injectable preparations like stabilizers, solubilizers, pH modifiers, buffers, antioxidants, cosolvents, complexing agents, tonicity modifiers, etc. Some suitable additives are for example tartrate, citrate or similar buffers, alcohol, sodium chloride, dextrose and high molecular weight polymers. Another alternative is sterile powder reconstitution. The compound may be administered in the form of injection for more than once daily administration, or intravenous infusion/ drip or suitable depot preparation. Example 36 Preparation suitable for parenteral administration has the following composition: Active ingredient of formula I as given above Polyethylene glycol (400) 0.75 ml Sodium metabisulphite 0.01 % Isotonic saline/ WFI q.s. Other Formulations. For the dermatological application and for the discoloration of teeth, the recommended formulations are lotions, oral rinse and toothpaste containing appropriate amount of the compounds of the general formula I. The above examples are presented by way of illustration alone and in no way limit the scope of the invention. WE CLAIM: 1. A Compound " which is selected from the group consisting of the following compounds: (^)l-[1-(2-thien-2"-yl-2-oxoethyl)-5-aminocarbonyl-3-carbonyl pyridinium]-2-[1-(2-thien-2"-yl-2-oxoethyl )-3-carbonyl pyridinium] hydrazine dichloride and pharmaceutically acceptable salts thereof, ( b) 1 -(2-thien-2"-yl-2-oxoethyl)-3-(trifluoromethanesulfonyl hydrazino carbonyl) - pyridinium bromide and pharmaceutically acceptable salts thereof, (c)l-[1-(2-thien-2"-yl-2-oxoethyl)-6-methyl-3-carbonyl pyridinium]-2- [1-(2-thien-2"-yl-2-oxoethyl )-3-carbonyl pyridinium ] hydrazine dichloride and pharmaceutically acceptable salts thereof, (dd) N,N"-bis[3-carbonyl-1-(2-(5-methyl-thien-2-yl)-2-oxoethyl) pyridinium] hydrazine dichloride and pharmaceutically acceptable salts thereof, (e) N,N"-bis[3-carbonyl-1-(2-(5-chloro-thien-2-yl)-2-oxoethyl) pyridinium] hydrazine dichloride and pharmaceutically acceptable salts thereof, (f ) 1-(2-tm"en-2"-yl-2-oxoethyl)-3-(methanesulfonyl hydrazino carbonyl )-6-methyl pyridinium bromide and pharmaceutically acceptable salts thereof, (9)N,N"-bis[3-carbonyl-1-(2-(4-nitro-thien-2-yl)-2-oxoethyl)pyridinium] hydrazine dichloride and pharmaceutically acceptable salts thereof, (h ) 1-(2-phenylamino-2-oxoethyl)-3-(phenyl hydrazino carbonyl) pyridinium chloride and pharmaceutically acceptable salts thereof, (i ) 1-(2-(4-nitro-thien-2-yl)-2-oxoethyl)-3-(methanesulfonyl hydrazino carbonyl) pyridinium bromide and pharmaceutically acceptable salts thereof, (j ) 1-(2-(5-nitro-thien-2-yl)-2-oxoethyl)-3-(methanesulfonyl hydrazino carbonyl) pyridinium chloride and pharmaceutically acceptable salts thereof, (k.) 1 -(2-(5-chloro-thien-2-yl)-2-oxoethyl)-3-(methanesulfonyl hydrazino carbonyl) pyridinium bromide and pharmaceutically acceptable salts thereof, (l) 1-(2-thien-2"-yl-2-oxoethyl)-3-(ethoxycarbonyl hydrazino carbonyl) pyridinium bromide and pharmaceutically acceptable salts thereof, (m) 1-(2-thien-2"-yl-2-oxoethyl)-3-(isopropylsulfonyl hydrazino carbonyl) pyridinium bromide and pharmaceutically acceptable salts thereof, (ft..) 1-(2-tliien-2"-yl-2-oxoethyl)-3-(metlianesulfonyl hydrazino carbonyl) -5-bromo pyridinium bromide and pharmaceutically acceptable salts thereof, (o ) 1-(2-(2-ethoxy carbonyl pyrrolidin-l-yl)-2-oxoethyl)-3- (methanesulfonyl hydrazino carbonyl) pyridinium chloride and pharmaceutically acceptable salts thereof, (p ) 1-(2-(5-methyl-thien-2-yl)-2-oxoethyl)-3-(methanesulfonyl hydrazino carbonyl) pyridinium chloride and pharmaceutically acceptable salts thereof, (q) 1-(2-(4-carbethoxy-thiazolidin-3-yl)-2-oxoethyl) -3- (methanesulfonyl hydrazino carbonyl) pyridinium chloride and pharmaceutically acceptable salts thereof, (r ) 1-(2-(4-benzyl piperidin-l-yl)-2-oxoethyl)-3-(methanesulfonyl hydrazino carbonyl) pyridinium chloride and pharmaceutically acceptable salts thereof, (s) N,N"-bis[3-carbonyl-1 -(2-(2-ethoxycarbonyl pyrrolidin-1 -yl)-2- oxoethyl) pyridinium] hydrazine dichloride and pharmaceutically acceptable salts thereof; (t ) 1-(2-phenylamino-2-oxoethyl)-4 -[ 2-(benzoyloxy) ethylamino carbonyl ] pyridinium chloride and pharmaceutically acceptable salts thereof, ( u) 1-(2-thien-2"-yl-2-oxoethyl)-3-(phenyl hydrazi.no carbonyl) pyridinium bromide and pharmaceutically acceptable salts thereof, (v) 1-(2-thien-2"-yl-2-oxoethyl)-3-(p-methoxy phenyl sulfonyl hydrazino carbonyl) pyridinium bromide and phannaceutically acceptable salts thereof, (W) 1-(2-ethoxy-2-oxoethyl)-3-(phenyl aminocarbonyl hydrazino carbonyl) pyridinium bromide and phannaceutically acceptable salts thereof, (X .) 1-(2-ethoxy-2-oxoethyl)-3-(p-toluene sulfonyl hydrazino carbonyl) pyridinium bromide and phannaceutically acceptable salts thereof, (y) 1-(2-phenyl-2-oxoethyl)-3-(phenylamino carbonyl hydrazino carbonyl) pyridinium bromide and pharmaceutically acceptable salts thereof, (z ) 1-(2-phenylamino-2-oxoethyl)-3-(benzyl sulfonyl hydrazino carbonyl) pyridiniumchloride and pharmaceutically acceptable salts thereof, (za.) 1-(2-phenyl-2-oxoethyl)-4-(methanesulfonyl hydrazino carbonyl) pyridinium bromide and phannaceutically acceptable salts thereof, (zb) 1-(2-phenyl-2-oxoethyl)-3-(phenyl hydrazino carbonyl) pyridinium bromide and pharmaceutically acceptable salts thereof, (zc) 1 -(2-ethoxy-2-oxoethyl)-4-[2-(benzoyloxy) ethyl ammo carbonyl ] pyridinium bromide and pharmaceutically acceptable salts thereof, (Zd) 1-(2-ethoxy-2-oxoethyl)-3-(phenyl hydrazino carbonyl) pyridinium bromide and pharmaceutically acceptable salts thereof, (ze) 1-(2-phenyl-2-oxoethyl)-3-(p-methoxyphcnyl sulfonyl hydrazinc carbonyl) pyridinium bromide and pharmaceulically acceptable salts thereof, (zf) 1-(2-phenyl-2-o.\oelhyl)-4-[2-(bcnzoyloxy) ethyl amino carbonyl] pyridinium bromide and pharmaceutically acceptable salts thereof, and (zg) 1-(2-elhoxy-2-oxocthyl)-4-(p-methancsulfonyl hydra/ine carbonyl) pyridinium bromide and pharmaceutically acceptable salts thereof. 2. A process for the preparation of the compound as claimed in claim 1, which comprises : i) refluxing pyridinc derivative selected from : 1. N,N" -bis(nicotinyl)hydrazinc 2. 3-[(2-pyridyl)hydrazinocarbonyl]pyridine 3. 3-\|(2-methanesulfonyl)hydrazinocarbonyl]pyridine 4. 3-[(2-benzoyloxy)cthylaminocarbonyl]pyridine 5. 3-\|(2-phyenylsuironyl)hydrazinocarbonyl]pyridine 6. 3-\|(2-acctoxy)ethyloxycarbonyl]pyridinc 7. 3-\|(2-ben/.oyloxy)ethyloxycarbonyl\|pyridinc 8. 3-[(2-methoxy)cthyloxycarbonyl]pyridine 9. 3-[(2-phycnylaminocarbonyl)hydra/inocarbonyl\|pyridinc 10. 3-[(2-acetoxy)ethylaminocarbonyl]pyridine 11. 3-((2-(4-methylphenyl sulfonyl)hydra/.inocarbonyl)]pyridine 12. 3-[(2-benzoyl)- hydrazino carbonyl ]pyridinc 13. 3-\|(2-phenylmcthanc sulfonyl) hydrazino carbonyl ]pyridinc 14. 3-[(2-(3- cyclohexylpropanoyl) hydrazino carbonyl]pyridine 15. 3-[(2-methoxy)ethylaminocarbonyl ]pyridinc 16. 3-\| l-oxo-1-(2-methoxycarbonyl)pyridyl\|hydrazino pyridinc with a quaternizing agent selected from the group comprising of : 1. 2-bromoacetyl thiophene 2. 2-chloroacetyl thiopene 3. phenacylbromide 4. phenacylchloride 5. 2,4-dichlorophenacylbromide 6. N- phenyl chloroacetamide 7. N- cyclopropyl chloroacetamide 8. cthylbromoacctatc 9. bromo acetylfuran 10. N- isopropylchloroacetamide 11. N- chloroacetyl-2-pyrrolidinone 12. chloroacetic acid in a solvent selected from the group comprising of methanol, ethanol, propanol, toluene or xylene under reflux for 6 to 48 hours. ii) isolating the compound from step (i) 3. A pharmaceutical composition for treatment of diabetic complictions and aging related diseases which comprises a pharmaceutically effective amount of one or more compounds as claimed in claim 1 or pharmaceutically acceptable salt(s) thereof in admixture with a pharmaceutically acceptable carrier, diluent, solvent or excepient. 4. The pharmaceutical composition as claimed in claim 3, in the form of an oral formulation. 5. The pharmaceutical composition as claimed in claim 4, wherein said pharmaceutically acceptable carrier is selected from group consisting of starch, lactose, polyvinyl pyrolidone (K- 30), talc and magnesium stearatc. 6. The Pharmaceutical composition as claimed in claim 3 in the form of a parenteral formulation. 7. A method for the preparation of a parenteral formulation as claimed in claim 6. which comprises dissolving one or more compounds as defined in claim 1, in polyethylene glycol 400 and diluting the solution so obtained, with an isotonic solution or water to a desired concentration. 8. Pharmaceutical composition as claimed in claim 5, in the form of a lotion, oral rinse and toothpaste. 9. A process for preparation of pyridinium compounds substantially as herein described particularly with reference to the examples. 10. A pharmaceutical composition substantially as herein described particularly with reference to examples. The invention discloses novel compounds of the pyridinium series useful for the management of diabetes and aging-related vascular complications, including kidney disease, nerve damage, atherosclerosis, retinopathy, dermatological disorders and discoloration of teeth, by breaking performed AGE. The invention also discloses, method for preparation of the novel compounds of the series and pharmaceutical composition having one or more compounds as defined above as active ingredients.

Full Text

FIELD OF THE INVENTION
The present invention relates to a new class of compounds of pyridinium
series and to their use in treatment of diabetes and related illnesses. More
particularly the invention relates to compounds of this series, methods for their
preparation, pharmaceutical composition containing these compounds and their
use in the treatment of complications of diabetes mellitus. The compounds of this
series exhibit AGE breaking activity, which is essential for the treatment of
diabetic and aging-related complications including kidney disease, nerve damage,
atherosclerosis, retinopathy and dennatological conditions. The invention also
extends to the method of reversing the discoloration of teeth resulting from
nonenzymatic browning in the oral cavity which comprises administration of an
amount effective to reverse pre-formed advanced glycosylation crosslinks.
BACKGROUND OF THE INVENTION
Maillard in 1912 found that reducing sugars, such as glucose and ribose
react with proteins to form brown pigments. Further studies have shown that this
is an irreversible non-enzymatic reaction, which occurs in several natural systems
including stored foodstuff. Maillard reaction occurs in two stages, early and
advanced. Initially, proteins react with glucose to form stable Amadori products,
which subsequently cross-links to form advanced glycation end products (AGE).
In most cases, the formation of AGE also accompanies browning of the proteins
and increase in the fluorescence.
In diabetes, where blood glucose level is significantly higher than normal,
the reaction of glucose with several proteins such as haemoglobin, lens crystallin
and collagen, gives rise to the formation of AGE, which in turn, is responsible for
the complications associated with diabetes, such as nephropathy,
microangiopathy, endothelial dysfunction and other organ dysfunctions. In
addition, the activity of several growth factors, such as basic fibroblast growth
factor, is also impaired. AGE products, unlike normal proteins in tissue, have a
slower rate of turnover and replenishment. It has been reported that AGE
products may in fact elicit a complex immunological reaction involving RAGE
(Receptor for Advanced Glycation End Products) receptors and activation of
several incompletely defined immunological processes. It has been documented
that diabetes with evidence of microangiopathy and macroangiopathy also show
evidence of oxidative stress, the mechanism of which has not been elucidated.
In vitro AGE formation can be studied in the laboratory by incubating
reducing sugars, such as ribose or glucose with bovine serum albumin. AGE
formation can be detected by increase in the fluorescence or increased cross
reactivity with anti-AGE antibodies. The increase in fluorescence seems to
precede formation of AGE specific antigenic epitopes. This increase in
fluorescence is used to monitor the increased AGE formation in vitro (Brownlee
M et al, Science 1986; 232:1629-1632). In addition to the increase in the
fluorescence, one of the most important features of in vitro AGE formation is the
formation of antigenic epitopes that are specific to AGE and not to the native
proteins. Therefore, it is possible to raise antibodies against advanced glycation
end products of one protein and use them to detect AGE formation in other
proteins. This has served as an important analytical tool in AGE research.
Due to the clinical significance of AGE formation, many approaches are
being used to diagnose, prevent, or revert AGE formation in the body. The
formation of AGE could be inhibited by reacting with an early glycosylation
product that results from the original reaction between the target protein and
glucose. The inhibition was believed to take place as the reaction between the
inhibitor and the early glycosylation product appeared to interrupt the subsequent
reaction of the glycosylated protein with additional protein material to form the
cross linked late stage product. Compounds like aminoguanidine act to inhibit
AGE formation by such mechanism.
The formation of AGE on long-lived proteins is also associated with cross-
linking of these proteins. The AGE derived protein cross-links have been shown
to be cleaved by compounds like N- phenacyl thiazolium bromide (PTB), which
reacts with and cleaves covalent, AGE derived protein cross links (Vasan et al.
Nature 1996; 382: 275-278 ; US 5,853,703, Date of Patent : Dec. 29, 1998). The
mechanism of reducing the AGE content in tissues is expected to take place
relatively rapidly, in contrast to aminoguanidine, which acts slowly by its very
nature of mechanism of action. This current specification is related to compounds
of pyridinium class, which break pre-formed AGE, like PTB, and in some cases
even more effectively by than PTB.
SUMMARY OF THE INVENTION
The main objective of the present invention is to provide a new class of
compounds of the pyridinium series which are useful for the management of
diabetes and aging related vascular complications and particularly in the treatment
of complications of diabetes mellitus and other aging related conditions including
kidney disease, nerve damage, atherosclerosis, retinopathy and dermatological
conditions. The invention also extends the method to reverse the discoloration of
teeth resulting from nonenzymatic browning in the oral cavity which comprises
administration of an amount effective to reverse the pre-formed advanced
glycosylation crosslinks, etc.
Another object of the present invention is to provide compounds of the
pyridinium series, which exhibit AGE breaking activities.
Yet another object of the present invention is to provide a method of
preparation of compounds of the pyridinium series which exhibit AGE breaking
activities.
Still another object of the invention is to provide pharmaceutical
compositions with a new class of compounds of the pyridinium series according
to the invention and their pharmaceutically acceptable salts in combination with
suitable carriers, solvents, excepients, diluents and other media normally
employed in preparing such compositions.
Still another object of the invention is to provide a method of treatment of a
diabetic patient by administration of the compounds of the invention, either singly
or in combination with drugs for anti-diabetic therapy, or pharmaceutically
acceptable salts thereof in required dosage in admixture with pharmaceutically
acceptable diluent, solvent, excepients, carriers or other media as may be
appropriate for the purpose.
DETAILED DESCRIPTION OF THE INVENTION
The present invention provides for a new class of AGE breakers, of general
formula I,
wherein
R1 is -R4-R5 or -N(R7) N (R7) R9;
R4 is selected from the group -N(R7)R6O, -N(R7)R6N(R7)-, OR6O, -OR6N(R7)-,
where Re is alkyl;
R5 is selected from the group alkyl, aryl, including heteroaryl, -COR7, SO2R7,

where R7 is selected from the group H, alkyl or aryl, including heteroaryl;
R2 is selected from the group F, Cl, Br, I, OR7, NO2, alkyl, aryl including
heteroaryl, formyl, acyl, C(O)NR7R10, C(O)OR7, NR7R10, N=C(R7)(R10), SR7,
SO2NH2, SO2 alkyl and SO2aryl,
and m is 0, 1 or 2
R3 is selected from the group R7, OR7, N(R7) (R10), N=C(R7) (Rio), N(R7) N(R7)
(R10), N(R7) N=C(R7) (R,o) and CH(R7)C(O)R8
where Rs is selected from the group R7, OR7 and NR7R10;
R9 is selected from the group consisting of hydrogen, alkyl, aryl, including
heteroaryl, C(O)R10, -SO2R10, -C(S)NHR10, -C(NH) NH (R10), -C(O) NHR10,
Rio is selected for the group H, alkyl or aryl, including heteroaryl and in each
case optionally different from substituent R7
X is selected from group consisting of a halide ion, acetate ion, perchlorate ion,
sulfonate ion, oxalate ion, citrate ion, tosylate ion, maleate ion, mesylate ion,
carbonate ion, sulfite ion, phosphoric hydrogen ion, phosphonate ion, phosphate
ion, BF4-, PF6-, etc.
with proviso that
(i) when two alkyl groups are present on the same carbon or nitrogen, they are
optionally linked together to form a cyclic structure and
(ii) the nitrogen of heteroaryl ring of R10, when present, is optionally
quaternized with compound such as X-CH2C(O)-R.3
As used herein, "alkyl" refers to an optionally substituted hydrocarbon
group joined by single carbon-carbon bonds and having 1 to 8 carbon atoms
joined together. The alkyl hydrocarbon group may be linear, branched or cyclic,
saturated or unsaturated. The substituents are selected from F, Cl, Br, I, N, S, 0
and aryl. Preferably, no more than three substituents are present.
As used herein "aryl" refers to an optionally substituted aromatic group
with atleast one ring having a conjugated pi- electron system, containing upto two
conjugated or fused ring systems. Aryl includes carbocyclic aryl, heterocyclic aryl
and biaryl groups, all of which may be optionally substituted. The substituents are
selected from F, Cl, Br, I, N, S. O and straight chain or branched C1-C6
hydrocarbon.
The novel compounds of the invention of general formula I having m as 0
and - COR1 at position 3 are listed in Table 1A and the novel compounds of the
invention of general formula I having m as 0 and - COR1 at position 4 are listed in
Table 1B. The following compounds suggested are by way of example alone of
the representative compounds of the general formula I as defined above and in no
way restrict the invention.
l-[1-(2-thien-2-yl-2-oxoethyl)-6-methyl-3-carbonylpyridinium]-2-[l-
(2-Thien-2"-yl-2-oxoethyl )-3-carbonyl pyridinium ] hydrazine
dichloride(compound no: 1).
1 -(2-thien-2"-yl-2-oxoethyl)-3-(isopropylsulfonyl hydrazino carbonyl)
pyridiniiun broraide(compound no: 2.).
1-(2-(4-benzyl piperidin-1 -yl)-2-oxoethyl)-3-(methanesulfonyl
hydrazino carbonyl) pyridinium chloride(compound no: 3 ).
1-(2-(2-ethoxy carbonyl pyrrolidin-l-yl)-2-oxoethyl)-3-(methanesulfonyl
hydrazino carbonyl) pyridinium chloride, (compound no: 40-
1-(2-thien-2"-yl-2-oxoethyl)-3-(raethanesulfonyl hydrazino carbonyl )-5-
bromo pyridinium bromide, (compound no: 5").
1-(2-thien-2"-yl-2-oxoethyl)-3-(ethoxycarbonyl hydrazino carbonyl)
pyridinium bromide, (compound no: 6 ).
1-(2-(5-chlorothien-2-yl)-2-oxoethyl)-3-(methanesulfonyl hydrazino
carbonyl) pyridinium bromide (compound no: 1 ).
N,N"-bis[3-carbonyl-1-(2-(4-nitrothien-2-yl)-2-oxoetJiyl)pyridinium]
hydrazine dichloride. (compound no: 8).
1-(2-thien-2"-yl-2-oxoethyl)-3-(methanesulfonyl hydrazino carbonyl)
-6-methyl pyridinium bromide, (compound no: 9 ).
N,N"-bis[3-carbonyl-1-(2-(5-methylthien-2-yl)-2-oxoethyl) pyridinium]
hydrazine dichloride. (compound no: 10).
N,N"-bis[3-carbonyl-1-(2-(2-ethoxycarbonyl pyrrolidin-l-yl)-2-
oxoethyl) pyridinium] hydrazine dichloride. (compound no:n ).
1-[1-(2-thien-2"-yl-2-oxoethyl)-5-aminocarbonyl-3-carbonyl
pyridinium]-2-[1-(2-Thien-2"-yl-2-oxoethyl )-3-carbonyl pyridinium]
hydrazine dichloride (compound no: 12).
1-(2-(4-carbethoxy-thiazolidin-3-yl)-2-oxoethyl)-3-(methanesulfonyl
hydrazino carbonyl) pyridinium chloride (compound no: 13 ).
N,N"-bis[3-carbonyl-1-(2-(5-chlorothien-2-yl)-2-oxoethyl)pyridinium]
hydrazine dichloride. (compound no: 14).
1-(2-(5-methylthien-2-yl)-2-oxoethyl)-3-(methanesulfonyl hydrazino
carbonyl) pyridinium chloride (compound no:15).
1 -(2-(4-nitrothien-2-yl)-2-oxoethyl)-3 -(methane sulfonyl hydrazino
carbonyl) pyridinium bromide, (compound no;16").
1-(2-phenylamino-2-oxoethyl)-3-(phenyl hydrazino carbonyl)
pyridinium chloride (compound no: 11).
1-(2-phenylamino-2-oxoethyl)-4 -[ 2-(benzoyloxy) ethylamino
carbonyl ] pyridinium chloride (compound no: 18").
1 -2-(5-nitro-thien-2-yl)-2-oxoethyl)-3-(methanesulfonyl hydrazino
carbonyl) pyridinium chloride (compound no: .19).
1 -(2-thien-2"-yl-2-oxoethyl)-3-(trifluoromethanesulfonyl hydrazino
carbonyl) - pyridinium bromide, (compound no. 20,).
1-(2-thien-2"-yl-2-oxoethyl)-3-(phenyl hydrazino carbonyl) pyridinium
bromide (compound no.21).
1-(2-thien-2"-yl-2-oxocthyl)-3-(p-methoxy phenyl sulfonyl hydrazino
carbonyl) pyridinium bromide (compound no. 22).
1-(2-ethoxy-2-oxoethyl)-3-(phenyl aminocarbonyl hydrazino carbonyl)
pyridinium bromide (compound no. 23).
1-(2-ethoxy-2-oxoemyl)-3-(p-toluene sulfonyl hydrazino carbonyl)
pyridinium bromide (compound no. 24).
1-(2-phenyl-2-oxoethyl)-3-(phenylamino carbonyl hydrazino carbonyl)
pyridinium bromide (compound no. 25).
1 -(2-phenylamino-2-oxoethyl)-3-(benzyl sulfonyl hydrazino carbonyl)
pyridinrumchloride. (compound no. 26 ).
1-(2-phenyl-2-oxoethyl)-4-(methanesulfonyl hydrazino carbonyl)
pyridinium bromide (compound no. 27).
1-(2-phenyl-2-oxoethyl)-3-(phenyl hydrazino carbonyl) pyridinium
bromide, (compound no. 28).
1-(2-ethoxy-2-oxoethyl)-4-[2-(benzoyloxy) ethyl amino carbonyl ]
pyridinium bromide (compound no. 29 ).
1-(2-ethoxy-2-oxoethyl)-3-(phenyl hydrazino carbonyl) pyridinium
bromide.(compound no.3o ).
1-(2-phenyl-2-oxoethyl)-3-(p-methoxyphenyl sulfonyl hydrazino
carbonyl) pyridinium bromide, (compound no. 31).
1-(2-phenyl-2-oxoethyl)- 4-[2-(benzoyloxy) ethyl amino carbonyl ]
pyridinium bromide, (compound no.32.).
1-(2-ethoxy-2-oxoethyl)- 4-(p-methanesulfonyl hydrazino carbonyl )
pyridinium bromide, (compound no. 33).
In place of specific halide salts of the compounds listed above,
these compounds may also be in the form of other pharmaceutically
acceptable salts falling within the definition of X as given above. The
words "and pharmaceutically acceptable salts thereof as used herein
following the names of specific compounds of general formula (I) of the
invention means that such compounds encompass other
pharmaceutically acceptable salts falling within the definition of X.
According to the embodiment of the present invention, the
present compounds are used for the treatment of diabetic
complications, and aging related vascular and neurovascular
complications including kidney disease, nerve damage, atherosclerosis,
retinopathy, inflammatory discorders, immunological disorders,
oxidative stress, dermatological conditions, and cosmetic conditions
including colouration of teeth occurring due to the higher levels of
preformed AGE. The increased levels of preformed AGE can be
brought under control by breaking the AGE products using compounds
mentioned in the invention.
The invention also provides a process for the preparation of
novel compounds of the pyridinium series.
The novel compounds of general formula I, are
prepared from properly substituted pyridine derivatives followed by
quarternization with appropriate reagent by refluxing in alcoholic
solvents like, methanol, ethanol, propanol, etc and high boiling solvents
like toluene or xylene etc, for 6-48 hrs. to give the desired
compounds.
The examples of substituted pyridine derivatives which can be
used for preparation of specific compounds of the invention are given
below:
1. N,N"-bis(nicotinyl)hydrazine
2. 3-[(2-pyridyl)hydrazinocarbonyl]pyridine
3. 3-[2-methanesulfonyl)hydrazinocarbonyl]pyridine
4. 3-[(2-benzoyloxy)ethylamiiiocarbonyl]pyridine
5. 3-[(2-phenylsulfonyl)hydrazinocarbonyl] pyridine
6. 3-[(2-acetoxy)ethyloxycarbonyl]pyridine
7. 3-[(2-benzoyloxy)ethyloxycarbonyl]pyridine
8. 3-[(2-methoxy)ethyloxycarbonyl]pyridine
9. 3-[(2-phenylaminocarbonyl)hydrazinocarbonyl]pyridine
10. 3-[(2-acetoxy)ethylaminocarbonyl]pyridine
11. 3-[(2-(4-methylphenyl sulfonylhydrazinocarbonyl))]pyridine
12. 3-[(2-benzoyl)- hydrazino carbonyl]pyridine
13. 3-[(2-phenyhnethane sulfonyl) hydrazino carbonyl] pyridine
14. 3-[(2-(3-cyclohexylpropanoyl) hydrazino carbonyl]pyridine
15. 3-[(2-methoxy)ethylaniinocarbonyllpyridine
16. 3-[l-oxo-1 -(2-methoxycarbonyl)pyridyl Jhydrazine pyridine
The examples of quaternizing agents which may be used in the reaction are given below:
1. 2-bromoacetyl thiophene
2. 2-chloroacetyl thiopene
3. phenacylbromide
4. phenacyl chloride
5. 2,4-dichllrophenacylbromide
6. N-phenyl chloroacetamide
7. N-cyclopropyl chloroacetamide
8. ethylbromoacetate
9. bromo acetylfuran
10. N-isopropyl chloroacetamide
11. N-chloroacetyl-2-pyrrolidinone
12. chloroacetic acid
In-vitro screening for AGE-breaking Activity
The in vitro AGE formation, studied in the laboratory, by incubating reducing
sugar glucose, with protein bovine serum albumin, resulted in browning of solution and
increase in the fluorescence. Fluorescence was used as the criteria to monitor the
increased AGE formation.
Example 1
AGE breaker activity has been confirmed by the screening
procedure as mentioned below:
Materials:
Bovine serum albumin (fraction V) (BSA)
Glucose, analytical grade
Phosphate buffered saline (PBS)
Equipment:
Microplate ELISA Reader - Spectramax Plus (Molecular Devices,
USA)
Microplate washer, (Bio -Tec Instruments, USA)
pH meter
Methods of experiment: Elisa (Enzyme Linked Immunosorbent Assay)
160 mg/ml of protein, bovine serum albumin, BSA and 1.6M
glucose sugar were dissolved in phosphate buffered saline, PBS.
Sodium azide was added at 0.02% concentration as a preservative. The
solution was filtered asceptically through a 0.22 µM filter and kept for
aging at 37°C for 16 weeks. After 16 weeks the solution was dialyzed
against PBS, aliquoted and stored at - 20°C.
To determine the AGE breaking activity, 10µg/ml of the 16
weeks AGE-BSA was incubated with different concentrations of the
test compounds at 37°C for 24 hours and AGE breaking activity of the
test compounds by ELISA was determined.
ELISA was performed as follows:
1. Different concentrations of 16 weeks AGE-BSA were coated on a
microtitre plate as standard. Each concentration is coated in triplicates.
2. The test samples were coated on microtitre plate at a concentration
of 5 ng. to 20 ng per well in triplicates.
3. The plate was incubated at 37°C for one hour.
4. After incubation the plate was washed with PBST (PBS with 0.05%
Tween 20).
5. Blocking with 5% skimmed milk in PBS at 37°C for one hour was
done.
6. The plate was washed with PBST.
7. Primary antibody against AGE-BSA was added and the plate is
incubated at 37°C for one hour.
8. The plate was washed with PBST
9. Secondary antibody anti rabbit HRPO (Horse-Radish Per Oxidase)
conjugate was added and the plate is incubated at 37°C for one hour.
10. The plate was washed with PBST.
11. Colour development with OPD (orthophenylenediamine
dihydrochloride) and hydrogen peroxide was done.
12. OD (optical density) at (450nm reading - 620nm reading) was
measured after incubation at 37°C for 15 minutes with Microplate
ELISA Reader.
The breaker activity of the compounds were determined by the
following formula:

OD450-620Control= Absorbance of 20ng AGE-BSA after incubation at
37°C for 24 hours without test compound
OD450-620Test= Absorbance of 20ng AGE-BSA after incubation at 37°C
for 24 hours with required concentration of test compound
Using specific examples, the % AGE breaking activity was calculated
and recorded in Table 2.
Hence compounds -1,2,5-10,12,14,16,17,19
and 20 have superior AGE breaking activity compared to PTB, of
which the potency of compounds .1,8-10,12,14,16,17,19 and 20
are significantly much higher.
The following examples give method of preparation of the
specific novel compounds of the invention as given in Table 1. The
following compounds suggested are by way of example alone and in no
way restrict the invention.
Example 2.
1-[1-(2-Thien-2"-yl-2-oxoethvI)-6methyl-3-carbonvl pvridiniumj-2-
[1-(2-Thien-2"-yl-2-oxoethvl )-3--carbonvl pyridinium ] hvdrazine
dichloridefcompound no: 1 ),
Yield " : 40%
M.P. : 76-80 °C (dec)
IR(KBr,cm-1) : 1637,1513
1HNMR (DMSO d6, 400 MHz) 5 : 11.69(2H,s), 9.59-
9.53(2H,d), 9.19(2H,m), 9.05(lH,d), 8.46-8.43(1H,t) ,8.34(lH,d), 8.27-
8.23(4H,m), 7.45-7.41 (2H,ra) ,6.56(2H,s) ,6.48(2H,s),2.81(3H,s).
Mass(m/z) : 505,506,507.
Example 3
1-(2-Thien-2"-vl-2-oxoethvl)-3-(isopropvlsulfonvI hvdrazino
carbonvl) pyridinium bromide(compound no: 2.),
Yield . : 70%
M.P : 90-95°C (dec)
IR(KBr.cm-1) 1638,1589
1HNMR (DMSO d6, 400 MHz) 5 : 11 .27(1H,s) ,9.91(1H,s),
9.60(1H,s) ,9.19-9.15(2H5m), 8.42-8.36(1 H,m) ,8.25-8.21(2H,m) ,7.43-
7.41(1H,t) ,6.45(2H,s), 1.35-1.34(6H,d).
Mass(m/z) : 368,369,370
Example 4-
1-(2-(4-Benzvl piperidin-l-vl)-2-oxoethvl)-3-(methanesulfonvl
hvdrazino carbonvl) pyridinium chloride(compound no: 3.),
Yield : 17%
M.P : 76-78°C
IR (KBr,cm-1) : 1684,1650,1556,1540.
1HNMR (DMSO d6, 400 MHz) 5 : 11.46(1H,s) ,9.55(1H,s),
9.46(1H,s) ,9.09-9.03(2H,m), 8.36-8.32(1H,t), 7.33-7.29(2H,m), 7.23-
7.19(3H,m), 5.88-5.79(2H,m) ,4.30-4.27(1H,d) ,3.76-3.73(1H,d),
3.10(4H,m) ,2.64(1H,t) ,2.57-2.55(2H,d), 1.85(1H,bs) ,1.72-
1.63(2H,t),1.36-1.28(1H,q) ,1.13-1.03(1H,m)
Mass(m/z) : 431,432,433
Example S
1-(2-(2-Ethoxv carbonvl pvrrolidin-l-vl)-2-oxoethvl)-3-
(methanesulfonvl hydrazino carbonvl) pvridinium chloride,
(compound no: 4 ),
Yield : 14%
M.P : 88-91°C
IR(KBr,cm1) : 1735,1665,1539
1HNMR (DMSO d6, 400 MHz) 5 : 11.48(1H,s) ,9.96(1H,s)
,9.46(1H,s) ,9.09-9.05(2H,m) ,8.38-8.34(1H,t), 5.94-5.80(2H,q) ,4.37-
4.36(1H,d), 4.08-4.06(2H,d), 3.68-3.65(2H,m), 3.09(4H,m) ,2.23-
2.18(2H,m), 2.04 -1.93(3H,m) ,1.18-1.09(3H,t)
Mass(m/z) : 399,400,401
Example 6
1-(2-Thien-2"-vl~2-oxoethvI)-3-(methanesulfonyl hydrazino
carbonyl )-5-bromo pvridinium bromide, (compound no: 5.),
Yield : 54%
M.P : Above 190-195 °C(dec)
IR(KB,cm-1) : 1682,1557,1540,1520
1HNMR (DMSO d6, 400 MHz) 8 : 11.35(1H,s) ,10.01(1H,s)
,9.57-9.54(2H,d), 9.32(1H,s), 8.26-8.22(2H,m),7.42(1H,s),6.39(2H,s),
3.08(3H,s)
Mass(m/z) : 418,419,420
Example .7-
1-(2-Thien-2"-yl-2-oxoethyl)-3-(ethoxvcarbonvI hvdrazino carbonvl)
pvridinium bromide, (compound no: 6 ),
Yield : 69%
M.P : 155-157°C
IR(KBr,cm-1) : 1731,1665,1637
1HNMR (DMSO d6, 400 MHz) 5 : 11.04(1H,s), 9.59(1H,s) ,
9.53(1H,s), 9.18(1H,s), 9.05-9.04(1H,d), 8.42(1H,s), 8.25-
8.23(2H,m), 7.43(1H,s), 6.46(2H,s), 4.12-4.11(2H,s), 1.23(3H,s)
Mass (m/z) : 334,335,336
Example .8
1-(2-(5-chloro-thien-2-yI)-2-oxoethyl)-3-(methanesulfonyl hydrazino
carbonyl) pyridinium bromide (compound no: 7-)
Yield : 87%
M.P : 228-230°C
IR (KBr,cm-1) : 1708,1664,1631,1550
"HNMR (DMSO d6, 400 MHz) 5 : 11.40(1H,s), 9.98(1H,s),
9.50(1H,s), 9.15(1H,d) , 9.061H,d), 8.43-8.39(1H,t), 8.16-8.15(1H,d),
7.51-7.50(1 H,d), 6.41 (2H,s), 3.09 (3H,s)
Mass (m/z) : 374,375,376,377
Example .9
N-N"-Bis[3-carbonyl-1-(2-(4-nitro-thien-2-yl)-2-oxoethyl)pvridinium]
hvdrazine dichloride. (compound no: .8 ),
Yield : 27%
M.P : 204-207°C
IR (KBr,cm-1) : 1681,1539,1514
1HNMR (DMSO d6, 400 MHz) 5 : 11.90(2H,s), 9.63(2H,s),
9.31-9.30(4H,m), 9.24 -9.22(2H,m), 8.87(2H,s), 8.49-8.46(2H,t), 6.56
(4H,s)
Mass (m/z) : 581,582,583
Example 10
1-(2-Thien-2"-yl-2-oxoethyl-3-(methanesulfonyl hydrazino carbonyl)
-6-methyl pyridinium bromide, (compound no:9 ),
Yield : 14%
M.P : 90-95°C(dec)
IR (KBr,crn1) : 1677,1575
1HNMR (DMSO d6, 400 MHz) 5 : 11.32(1H,s),
9.97(1H,s) 9.52(1H,s), 8.94-8.92(1H,d) , 8.32-8.24(3H,m),
7.44(1H,t), 6.54(2H,s), 3.08(3H,s), 2.79(3H,s)
Mass (m/z) : 354,355,356
Example 11
N-N"-Bis[3-carbonyl-1-(2-(5-methvl-thien-2-vl)-2-oxoethvn
pvridiniuml hydrazine dichloride. (compound no: 10),
Yield : 37%
M.P : Above 166-168°C(dec)
IR(KBr,cm-1) : 1666,1500
1HNMR (DMSO 4, 400 MHz) 5 : 11.73(2H,s), 9.59(2H,s),
9.19-9.15(4H,d) 8.45-8.42(2H,t), 8.06-8.05(2H,d) ,7.15-7.14(2H,d),
6.43 (4H,s), 2.59(6H,s)
Mass(m/z) : 519,520,521,522
Example 12
N-N"-Bis [3-carbonvl-1 -(2-(2-ethoxy carbonvl pyrrolidin-1 -yl)-2-
oxoethvl) pvridiniuml hvdrazine dichloride. (compound no: 11),
Yield : 28%
M.P : 118-120°C
IR (KBr,cm-1) : 1660,1510
1HNMR (DMSO d6, 400 MHz) 5 : 11.75(2H,s), 9.51(2H,s),
9.20-9.10(4H,m) 8.43-8.40(2H,t), 5.97-5.83(4H,m) ,4.39-4.36(2H,m),
4.27-4.22(1H,q), 4.12-4.05(4H,m), 3.71-3.63(4H,m), 3.48-3.40(1H,m),
2.26-2.19(2H,m),2.05-1.91(5H,m), 1.30-1.27(1H,t), 1.19-1.15(5H,t)
Mass(m/z) : 609,610,611
Example 13
l-[1-(2-Thien-2"-vl-2-oxoethvl)-5-aminocarbonyl-3-carbonvl
pyridiniuml-2-f1-(2-Thien-2"-vl-2-oxoethyl )-3-carbonvl pvridiniuml
hvdrazine dichloride (compound no: 12)
Yield : 54%
M.P : Above 127-129°C(dec)
IR (KBr,cm-1) : 1678,1513
1HNMR (DMSO d6, 400 MHz) 5 : 11.86(2H,s), 9.83-
9.64(4H,t), 9.24 -9.23(2H,s), 8.82(1H,s), 8.48-8.45(1 H,t), 8.34(1H,s)
8.26-8.24(4H,m), 7.44 -7.42(2H,d), 6.52-6.46(4H,d)
Mass (m/z) : 534,535,536
Example 14
1-(2-(4-carbethoxv-thiazolidin-3vl)-2--oxoethvl)-3-(methanesulfonyl
hvdrazino carbonvl) pyridinium chloride (compound no: 13),
Yield : 29%
M.P : 190-192°C
IR(KBr,cm-1) : 1673,1541
1HNMR (DMSO d6, 400 MHz) 5 : 11.50(1H,s), 9.55(1H,s),
9.48(1H,s), 9.12-9.08(2H,m), 8.39-8.34(1H,t), 6.04 - 5.99(2H,m) ,
4.94 -4.91(1H,m), 4.87-4.84(1H,d), 4.73-4.71(1H,d), 4.28-4.23(1H,q),
4.14 -4.09(1H,q), 3.43-3.38(1H,m),3.27-3.22(1H,m), 3.10(3H,s) ,1.30-
1.27(1H,t), 1.20-1.17(2H,m)
Mass(m/z) : " 439,440,441
Example 15
N-N"-Bis[3-carbonvl-1-(2-(S-chIoro-thien-2-vn-2-oxoethvl)
pyridinium] hydrazine dichloride. (compound no: 14:),
Yield : 35%
M.P : Above 200-205°C (dec)
IR(KBr,cm-1) : 1674,1590,1500
1HNMR (DMSO d6, 400 MHz) 5 : 11.90(2H,s), 9.64 -
9.61(2H,d), 9.29-9.20(4H,m) , 8.47-8.44(2H,t), 8.18-8.17(2H,d),7.51-
7.50(2H,d),6.49-6.48(4H,s)
Mass(m/z) : 559,560,561,562,563,564
Example 16
1-(2-(5-Methyl-thien-2-vn-2-oxoethvl)-3-(methanesulfonyl
hydrazino carbonvl) pyridinium chloride (compound no: 15),
Yield : 22%
M.P : 196-198°C
IR(KBr,cm-1) : 1689,1657
1HNMR (DMSO d6, 400 MHz) 5 : 11.47(1H,s) ,9.98(1H,s)
,9.53(1H,s) ,9.17-9.16(1H,d), 9.09-9.07(1H,d), 8.42-8.38(1H,t), 8.06-
8.05(1H,d), 7.15-7.14(1H,d), 6.41 (2H,s), 3.09(3H,s), 2.59(3H,s)
Mass(m/z) : 354,355,356,357
Example 17
1-(2-(4-Nitro-thien-2-yl)-2-oxoethyl)-3-(methanesulfonvl hydrazino
carbonvl) pyridinium bromide, (compound no: 16).
Yield : 52%
M.P : Above 200-205°C(dec)
IR(KBr,cm-1) : 1688,1631,1541
1HNMR (DMSO d6, 400 MHz) 5 : 11.41(1H,s), 9.50(1H,s)
9.309-9.306(1H,d), 9.17-9.15(1H,d) ,9.09-9.07(1H,d), 8.866-
8.862(1H,d) ,8.45-8.41(1H,t), 6.50(2H,s), 3.09(3H,s)
Mass(m/z) : 385,386,387
Example 18
1-(2-Phenylamino-2-oxoethyI)-3-(Phenyl hydrazino carbonyl)
pyridinium chloride (compound no: 17),
Yield : 45%
M.P : 165-167°C
IR(KBr,cm-1) : 1679,1626,1600,1497
1HNMR (DMSO d6, 400 MHz) 8 : 11.18(1H,s), 11.10(1H,s)
, 9.62(1H,s) , 9.24-9.22(1H,d), 9.17-9.15(1H,d), 8.40-8.36(1H,t),
8.19(1H,s), 7.63-7.61(2H,d) , 7.37-7.33(2H,t), 7.20-7.16(2H,t), 7.12-
7.09(1H,t), 6.88-6.86(2H,d), 6.78-6.74(1H,t) , 5.78(2H,s)
Mass(m/z) : 347,348,349
Example 19
1-(2-Phenvlamino-2-oxoethyl)-4 -I 2-(benzovIoxy) ethylamino
carbonvl] pyridinium chloride (compound no: 18),
Yield : 40%
M.P : 178-180°C
IR (KBr,cm-1) : 1700,1666,1559
1HNMR (DMSO d6, 400 MHz) 5 : 11.13(1H,s), 9.74 -
9.71(1H,t), 9.23-9.22(2H,d), 8.52-8.50(2H,d), 8.01-7.99(2H,d),
7.68-7.60(3H,m) , 7.54-7.5l(2H,t), 7.36-7.32(2H,t), 7.12-7.08 (1H,t),
5.75 (2H,s), 4.47-4.45(2H,t), 3.77-3.72(2H,q).
Mass (m/z) : 404,405,406
Example 20
l-2-(5-Nitro-thien-2-yl-2-oxoethvI)-3-(methanesulfonyl hydrazino
carbonvl) pyridinium chloride (compound no: 19),
Yield : 10%
M.P : Above 105-110°C(dec)
IR(KBr,cm-1) : 1680,1620
"HNMR (DMSO d6, 400 MHz) 5 : 11.48(1H,s), 9.98(l.H,s),
9.52(1H,s), 9.16-9.10(2H,m) ,8.45-8.41(1H,t) , 8.35-8.34(1H,d) ,8.25-
8.24(1H,d), 6.50(2H,s), 3.09(3H,s).
Mass (m/z) : 385,386,387
Example 31
1-(2-Thien-2"-vl-2-oxoethvI)3-(TrifIuromethanesulfonvl hydrazino
carbonvl)-pyridinium bromide (compound no: 20),
Yield : 22%
M.P : 77-79°C
IR (KBr,cm-1) : 2960, 1690, 1673, 1591
1HNMR (DMSO d6, 400 MHz) 5 : 11.76(1H,s),
11.27(1H,s), 9.61(1H,s), 9.20-9.19(1H,d) ,9.07-9.05(1H,d), 8.44-
8.41(1H,t) ,8.25-8.22(2H,m), 7.34 - 7.41 (1H,m), 6.46 (2H,s).
Mass (m/z) : 394,395,396
Example 22
1-(2-Thien-2"-vl-2-oxoethvl)-3-(phenvl hvdrazino carbonvl)
pvridinium bromide (compound no. 21),
Yield : 10%
M.P : 192-194°C
IR(KBr,cm-1) : 1669,1663,1603,
1HNMR (DMSO d6, 400 MHz) 5 : 10.99(1H,s), 9.54(1 H,s),
9.17-9.14(2H,t), 8.44-8.41(1H,t), 8.25-8.22(3H,m), 7.43-7.41(1H,t),
7.20-7.16(2H,t), 6.87-6.85(2H,d) , 6.79-6.75(1H,t), 6.46(2H,s)
Mass(m/z) : 338,339,340
Example 23
1-(2-Thien-2"-vl-2-oxoethvl)-3-(p-methoxy phcnvl sulfonvl
hvdrazino carbonvl) pvridiniuin bromide (compound no. 22.),
Yield : 28%
M.P : 126-]28°C
IR(KBr.cm-1) : 1672,1653,1596
1HNMR (DMSO 4, 400 MHz) 5 : 11.34 -11.33(1H,d),
10.27-10.26(1H,d), 9.34(1H,s), 9.13-9.12(1H,d), 8.94-8.92(1H,d),
8.38-8.34(1H,t) ,8.24-8.19(2H,m),7.82-7.75(2H,m) ,7.42-7.40(1H,t),
7.07-7.04(2H,d) ,6.40(2H,s), 3.81(3H,s).
Mass(m/z) : 432,433,434
Example 24
1-(2-Ethoxv-2-oxoethv))-3-(phenyl aminocarbonvl hvdrazino
carbonvl) pyridinium bromide, (compound no. 23),
Yield : 25%
M.P : 183-185°C
IR(KBr,cm-1) : 1746,1717,1682
1HNMR (DMSO d6, 400 MHz) 5 : 11.02(1H,s), 9.57(1H,s),
9.22-9.21(1H,d), 9.11-9.09(1H,d), 9.00(1H,s), 8.57(1H,s) ,8.44-
8.41(1H,m),7.47-7.45(2H,d) ,7.29-7.25(2H,t), 7.00-6.96(1H,t),
5.74(2H,s), 4.28-4.23(2H,q), 1.28-1.25(3H,t).
Mass(m/z) : 343,344,345,346
Example 25
1-(2-Ethoxv-2-oxoethyl)-3-(p-toluene sulfonvl hvdrazino carbonvl)
pvridinium bromide, (compound no. 24-),
Yield : 54%
M.P : 174-176°C
IR(KBr,cm-1) : 1746,1712,1634
1HNMR (DMSO d6, 400 MHz) 5 : 11.33(1H,s),
10.36(1H,s), 9.37(1H,s), 9.18-9.16(1H,d), 8.93-8.91(1H,d), 8.37-
8.33(1H,t), 7.78-7.76(2H,d), 7.37-7.35(2H,d), 5.68 (2H,s), 4.26-
4.20(2H,q),2.37(3H,s),1.27-1.23(3H,t).
Mass(m/z) : 378,379,380,381
Example 36
1-(2-Phenvl-2-oxoethvl)-3-(Phenylamino carbonyl hydrazino
carbonvl) pyridinium bromide, (compound no.25),
Yield : 70%
M.P : 206-208°C
IR(KBr,cm-1) : 1713,1684,1634
1HNMR (DMSO d6, 400 MHz) 5 : 11.05(1H,s), 9.55(1H,s),
9.18-9.13(2H,m), 9.02(1H,s), 8.59(1H,s), 8.49-8.45(1H,m) ,8.09-
8.07(2H,d), 7.84-7.80(1H,t) , 7.71-7.67(2H,t), 7.49-7.47(2H,d), 7.30-
7.26(2H,t), 7.01-6.97 (1H,t) ,6.56(2H,s).
Mass (m/z) : 375,376,377
Example 27
1-(2-Phenylamino-2-oxoethvI)-3-(benzvI sulfonyl hydrazino
carbonvl) pyridiniumchloride. (compound no.26),
Yield : 48%
M.P : 208-210°C
IR(KBr,cm-1) : 1712,1681,1632
1HNMR (DMSO d6, 400 MHz) 8 : 11.46(1H,s) ,10.80(1H,s),
9.59(1H,s), 9.22-9.20(1H,d) ,9.08-9.06(1H,d), 8.38-8.36(1H,t),7.60-
7.58(2H,d) ,7.49(2Hsm), 7.39-7.34(5H,m),7.13-7.10(1 H,t),
5.74(2H,s), 4.52(2H,s).
Mass(m/z) : 425,426,427,428
Example 28
1-(2-Phenyl-2-oxoethyl)-4-(methanesulfonvl hvdrazino carbonyl)
pyridinium bromide (compound no. 27),
Yield : 10%
M.P : 190-192°C
IR(KBr,cm-1) : 1679,1630,1650
1HNMR (DMSO d6, 400 MHz) 5 : 11.54(1H,s), 10.03(1H,s),
9.20-9.18(2H,d), 8.59-8.57(2H,d), 8.10-8.08(2H,d), 7.84 -7.80(1H,t),
7.71-7.67(2H,t), 6.56(2H,s), 3.08(3H,s).
Mass(m/z) : 334,335,336
Example 2S
1-(2-PhenvI-2-oxoethyl)-3-(phenyl hvdrazino carbonvl) pvridinium
bromide (compound no."28"),
Yield : 36%
M.P : 204-206°C
IR(KBr,cm-1) : 1686,1653,1630
1HNMR (DMSO d6, 400 MHz) 6 : 11.01(1H,s) ,9.53(1H,s)
,9.17-9.16(2H,m), 8.46-8.42(1H,t), 8.09-8.07(2H,d), 7.82-7.78(1 H,t),
7.69-7.65(2H,t), 7.20-7.16(2H,t), 6.88-6.86(2H,d), 6.79-6.75(1H,t),
6.56(2H,s)
Mass(m/z) : 332,333
Example 30
1-(2-Ethoxv-2-oxoethyl)-4-[2-(benzovIoxv) ethyl amino carbonvl ]
pvridinium bromide (compound no.29),
Yield : 82%
M.P : 154-156°C
IR(KBr,cm-1) : 1742,1719,1707,1675
1HNMR (DMSO d6, 400 MHz) 5 : 9.57-9.54(1H,t), 9.22-
9.20(2H,d), 8.51-8.49(2H,d), 8.00-7.98(2H,d), 7.68-7.64(1H,t), 7.54
-7.51(2H,t), 5.72(2H,s), 4.47-4.44(2H,t), 4.27-4.21(2H,q), 3.76-
3.72(2H,q), 1.27-1.24. (3H,t)
Mass(m/z) : 357,358,359.
Example 31
1-(2-Ethoxv-2-oxoethyl)-3-(phenvl hvdrazino carbonvl) pvridinium
bromide.(compound no. 30),
Yield : 37%
M.P : 185-187°C
IR (KBr,cm-1) : 1740,1690,1630.
1HNMR (DMSO d6, 400 MHz) 5 : 11.01(1H,s), 9.58(1H,s),
9.23-9.14(2H,m), 8.42-8.39(1H,t), 8.19(1H,s), 7.20-7.16(2H,t), 6.87-
6.85(2H,d), 6.78-6.75(1H,t), 5.75(2H,s), 4.28-4.22(2H,q), 1.28-1.24(3H,t)
Mass(m/z) : 300,301,302.
Example 32.
1-(2-PhenvI-2-oxoethvl)-3-(p-methoxvphenvl sulfonvl hydrazino
carbonvl) pvridinium bromide (compound no. 31),
Yield : 59%
M.P : 188-190°C
IR(KBr,cm-1) : 1671,1634,1580.
1HNMR (DMSO d6, 400 MHz) 5 : 11.26-11.25(1H,d),
10.17- 10.16(1H,d), 9.24(1H,s), 9.03-9.01(1H,d), 8.87-8.85(1H,d) ,
8.31-8.27(1H,t), 7.97-7.96(2H,d), 7.74 -7.69(3H,m), 7.60-7.56(2H,t)
, 6.99-6.97(2H,d), 6.40(2H,s), 3.73(3H,s).
Mass(m/z) : 426,427,428,429
Example 33
1-(2-Phenvl-2-oxoethvl)- 4-[2-(benzovloxv) ethyl amino carbonvl ]
pvridinium bromide (compound no.32),
Yield : 92%
M.P : 202-204°C
IR(KBr,cm-1) : 1715,1692,1650
1HNMR (DMSO d6, 400 MHz) 5 : 9.55(1H,s), 9.14-
9.13(2H,d), 8.52-8.51(2H,d), 8.07-7.99(4H,m), 7.80-7.51(6H,m),
6.52(2H,s), 4.46(2H,s), 3.76-3.75(2H,s).
Mass(m/z) : 389,390,391,392
Example 34-
1-(2-Ethoxy-2-oxoethyl)- 4-(p-methanesulfonyl hvdrazino carbonvl)
pvridinium bromide, (compound no. 33),
Yield . : 45%
M.P : 94-96°C
IR(KBr,cm-1) : 1726,1681,1643
1HNMR (DMSO d6, 400 MHz) 5 : 11.49(1H,s) ,9.98(1 H,s)
,9.23-9.21(2H,d), 8.54-8.52(2H,d), 5.73(2H,s), 4.28-4.22(2H,q),
3.09(3H,s), 1.28-1.25(3H,t).
Mass(nVz) : 302,303,304,305.
Pharmaceutical Compositions
Pharmaceutical compositions may be prepared with a
pharmaceutically effective quantity of compounds of general formula I,
individually or in combination. The following pharmaceutical
formulations suggested are by way of example alone and in no way
restrict the forms in which they can be used.
Oral formulations
Oral formulations may be administered as solid dosage forms for
example pellets, powders, sachets or discreet units such as tablets or
capsules and like. Other orally administered pharmaceutical
preparations include monophasic and biphasic liquid dosage forms
either in ready to use form or forms suitable for reconstirution such as
mixtures, syrups, suspensions or emulsions. The preparations in
addition may contain diluents, dispersing agents, buffers, stabilizers,
solubilizers, surfactants, preservatives, chelating agents and/ or other
pharmaceutical additives as are used. Aqueous or non aqueous vehicle
or their combination may be used and if desired may contain suitable
sweetener, flavoring agent or similar substances. In case of suspension
or emulsion a suitable thickening agent or suspending agent or
emulsifying agent may be present in addition. Alternatively, the
compounds may be administered as such in their pure form
unassociated with other additives for example as capsules or sachets. It
may also be administered with a vehicle. Pharmaceutical preparations
can have a slow, delayed or controlled release of active ingredients as
is provided by a matrix or diffusion controlled system.
When the present invention or its salts or suitable complexes is
presented as a discreet unit dosage form like tablet, it may contain in
addition medically inert excipients as are used in the art. Diluents such
as starch, lactose, dicalcium phosphate, talc, magnesium stearate,
polymeric substances like methyl cellulose, fatty acids and derivatives,
sodium starch glycollate, etc. may also be used.
Example 35
Preparation of oral dosage form:
A typical tablet has the following composition:
Active ingredient of formula I as given above
Lactose 135 mg
Starch 76 mg
Polyvinyl pyrolidone (K-30) 2 mg
Talc 1-5 mg
Magnesium Stearate 1.0 mg
Parenteral Formulations
For parenteral administration, the compounds or their salts or
suitable complexes thereof may be present in a sterile vehicle which
may be an aqueous or non aqueous vehicle or a combination thereof.
The examples of vehicles are water, ethyl oleate, oils and derivatives of
polyols, glycols and their derivatives. It may contain additives common
in injectable preparations like stabilizers, solubilizers, pH modifiers,
buffers, antioxidants, cosolvents, complexing agents, tonicity modifiers,
etc.
Some suitable additives are for example tartrate, citrate or
similar buffers, alcohol, sodium chloride, dextrose and high molecular
weight polymers. Another alternative is sterile powder reconstitution.
The compound may be administered in the form of injection for more
than once daily administration, or intravenous infusion/ drip or suitable
depot preparation.
Example 36
Preparation suitable for parenteral administration has the
following composition:
Active ingredient of formula I as given above
Polyethylene glycol (400) 0.75 ml
Sodium metabisulphite 0.01 %
Isotonic saline/ WFI q.s.
Other Formulations.
For the dermatological application and for the discoloration of
teeth, the recommended formulations are lotions, oral rinse and
toothpaste containing appropriate amount of the compounds of the
general formula I.
The above examples are presented by way of illustration alone
and in no way limit the scope of the invention.
WE CLAIM:
1. A Compound " which is selected from the
group consisting of the following compounds:
(^)l-[1-(2-thien-2"-yl-2-oxoethyl)-5-aminocarbonyl-3-carbonyl
pyridinium]-2-[1-(2-thien-2"-yl-2-oxoethyl )-3-carbonyl pyridinium]
hydrazine dichloride and pharmaceutically acceptable salts thereof,
( b) 1 -(2-thien-2"-yl-2-oxoethyl)-3-(trifluoromethanesulfonyl hydrazino
carbonyl) - pyridinium bromide and pharmaceutically acceptable salts
thereof,
(c)l-[1-(2-thien-2"-yl-2-oxoethyl)-6-methyl-3-carbonyl pyridinium]-2-
[1-(2-thien-2"-yl-2-oxoethyl )-3-carbonyl pyridinium ] hydrazine
dichloride and pharmaceutically acceptable salts thereof,
(dd) N,N"-bis[3-carbonyl-1-(2-(5-methyl-thien-2-yl)-2-oxoethyl)
pyridinium] hydrazine dichloride and pharmaceutically acceptable salts
thereof,
(e) N,N"-bis[3-carbonyl-1-(2-(5-chloro-thien-2-yl)-2-oxoethyl)
pyridinium] hydrazine dichloride and pharmaceutically acceptable salts
thereof,
(f ) 1-(2-tm"en-2"-yl-2-oxoethyl)-3-(methanesulfonyl hydrazino
carbonyl )-6-methyl pyridinium bromide and pharmaceutically
acceptable salts thereof,
(9)N,N"-bis[3-carbonyl-1-(2-(4-nitro-thien-2-yl)-2-oxoethyl)pyridinium]
hydrazine dichloride and pharmaceutically acceptable salts thereof,
(h ) 1-(2-phenylamino-2-oxoethyl)-3-(phenyl hydrazino carbonyl)
pyridinium chloride and pharmaceutically acceptable salts thereof,
(i ) 1-(2-(4-nitro-thien-2-yl)-2-oxoethyl)-3-(methanesulfonyl hydrazino
carbonyl) pyridinium bromide and pharmaceutically acceptable salts
thereof,
(j ) 1-(2-(5-nitro-thien-2-yl)-2-oxoethyl)-3-(methanesulfonyl
hydrazino carbonyl) pyridinium chloride and pharmaceutically
acceptable salts thereof,
(k.) 1 -(2-(5-chloro-thien-2-yl)-2-oxoethyl)-3-(methanesulfonyl
hydrazino carbonyl) pyridinium bromide and pharmaceutically
acceptable salts thereof,
(l) 1-(2-thien-2"-yl-2-oxoethyl)-3-(ethoxycarbonyl hydrazino carbonyl)
pyridinium bromide and pharmaceutically acceptable salts thereof,
(m) 1-(2-thien-2"-yl-2-oxoethyl)-3-(isopropylsulfonyl hydrazino
carbonyl) pyridinium bromide and pharmaceutically acceptable salts
thereof,
(ft..) 1-(2-tliien-2"-yl-2-oxoethyl)-3-(metlianesulfonyl hydrazino carbonyl)
-5-bromo pyridinium bromide and pharmaceutically acceptable salts
thereof,
(o ) 1-(2-(2-ethoxy carbonyl pyrrolidin-l-yl)-2-oxoethyl)-3-
(methanesulfonyl hydrazino carbonyl) pyridinium chloride and
pharmaceutically acceptable salts thereof,
(p ) 1-(2-(5-methyl-thien-2-yl)-2-oxoethyl)-3-(methanesulfonyl hydrazino
carbonyl) pyridinium chloride and pharmaceutically acceptable salts
thereof,
(q) 1-(2-(4-carbethoxy-thiazolidin-3-yl)-2-oxoethyl) -3-
(methanesulfonyl hydrazino carbonyl) pyridinium chloride and
pharmaceutically acceptable salts thereof,
(r ) 1-(2-(4-benzyl piperidin-l-yl)-2-oxoethyl)-3-(methanesulfonyl
hydrazino carbonyl) pyridinium chloride and pharmaceutically
acceptable salts thereof,
(s) N,N"-bis[3-carbonyl-1 -(2-(2-ethoxycarbonyl pyrrolidin-1 -yl)-2-
oxoethyl) pyridinium] hydrazine dichloride and pharmaceutically
acceptable salts thereof;
(t ) 1-(2-phenylamino-2-oxoethyl)-4 -[ 2-(benzoyloxy) ethylamino
carbonyl ] pyridinium chloride and pharmaceutically acceptable salts
thereof,
( u) 1-(2-thien-2"-yl-2-oxoethyl)-3-(phenyl hydrazi.no carbonyl) pyridinium
bromide and pharmaceutically acceptable salts thereof,
(v) 1-(2-thien-2"-yl-2-oxoethyl)-3-(p-methoxy phenyl sulfonyl
hydrazino carbonyl) pyridinium bromide and phannaceutically
acceptable salts thereof,
(W) 1-(2-ethoxy-2-oxoethyl)-3-(phenyl aminocarbonyl hydrazino
carbonyl) pyridinium bromide and phannaceutically acceptable salts
thereof,
(X .) 1-(2-ethoxy-2-oxoethyl)-3-(p-toluene sulfonyl hydrazino carbonyl)
pyridinium bromide and phannaceutically acceptable salts thereof,
(y) 1-(2-phenyl-2-oxoethyl)-3-(phenylamino carbonyl hydrazino
carbonyl) pyridinium bromide and pharmaceutically acceptable salts
thereof,
(z ) 1-(2-phenylamino-2-oxoethyl)-3-(benzyl sulfonyl hydrazino
carbonyl) pyridiniumchloride and pharmaceutically acceptable salts
thereof,
(za.) 1-(2-phenyl-2-oxoethyl)-4-(methanesulfonyl hydrazino carbonyl)
pyridinium bromide and phannaceutically acceptable salts thereof,
(zb) 1-(2-phenyl-2-oxoethyl)-3-(phenyl hydrazino carbonyl)
pyridinium bromide and pharmaceutically acceptable salts thereof,
(zc) 1 -(2-ethoxy-2-oxoethyl)-4-[2-(benzoyloxy) ethyl ammo carbonyl ]
pyridinium bromide and pharmaceutically acceptable salts thereof,
(Zd) 1-(2-ethoxy-2-oxoethyl)-3-(phenyl hydrazino carbonyl)
pyridinium bromide and pharmaceutically acceptable salts thereof,
(ze) 1-(2-phenyl-2-oxoethyl)-3-(p-methoxyphcnyl sulfonyl hydrazinc carbonyl)
pyridinium bromide and pharmaceulically acceptable salts thereof,
(zf) 1-(2-phenyl-2-o.\oelhyl)-4-[2-(bcnzoyloxy) ethyl amino carbonyl] pyridinium
bromide and pharmaceutically acceptable salts thereof, and
(zg) 1-(2-elhoxy-2-oxocthyl)-4-(p-methancsulfonyl hydra/ine carbonyl) pyridinium
bromide and pharmaceutically acceptable salts thereof.
2. A process for the preparation of the compound as claimed in claim 1, which comprises :
i) refluxing pyridinc derivative selected from :
1. N,N" -bis(nicotinyl)hydrazinc
2. 3-[(2-pyridyl)hydrazinocarbonyl]pyridine
3. 3-|(2-methanesulfonyl)hydrazinocarbonyl]pyridine
4. 3-[(2-benzoyloxy)cthylaminocarbonyl]pyridine
5. 3-|(2-phyenylsuironyl)hydrazinocarbonyl]pyridine
6. 3-|(2-acctoxy)ethyloxycarbonyl]pyridinc
7. 3-|(2-ben/.oyloxy)ethyloxycarbonyl|pyridinc
8. 3-[(2-methoxy)cthyloxycarbonyl]pyridine
9. 3-[(2-phycnylaminocarbonyl)hydra/inocarbonyl|pyridinc
10. 3-[(2-acetoxy)ethylaminocarbonyl]pyridine
11. 3-((2-(4-methylphenyl sulfonyl)hydra/.inocarbonyl)]pyridine
12. 3-[(2-benzoyl)- hydrazino carbonyl ]pyridinc
13. 3-|(2-phenylmcthanc sulfonyl) hydrazino carbonyl ]pyridinc
14. 3-[(2-(3- cyclohexylpropanoyl) hydrazino carbonyl]pyridine
15. 3-[(2-methoxy)ethylaminocarbonyl ]pyridinc
16. 3-| l-oxo-1-(2-methoxycarbonyl)pyridyl|hydrazino pyridinc
with a quaternizing agent selected from the group comprising of :
1. 2-bromoacetyl thiophene
2. 2-chloroacetyl thiopene
3. phenacylbromide
4. phenacylchloride
5. 2,4-dichlorophenacylbromide
6. N- phenyl chloroacetamide
7. N- cyclopropyl chloroacetamide
8. cthylbromoacctatc
9. bromo acetylfuran
10. N- isopropylchloroacetamide
11. N- chloroacetyl-2-pyrrolidinone
12. chloroacetic acid
in a solvent selected from the group comprising of methanol, ethanol, propanol, toluene or
xylene under reflux for 6 to 48 hours.
ii) isolating the compound from step (i)
3. A pharmaceutical composition for treatment of diabetic complictions and aging related
diseases which comprises a pharmaceutically effective amount of one or more compounds as
claimed in claim 1 or pharmaceutically acceptable salt(s) thereof in admixture with a
pharmaceutically acceptable carrier, diluent, solvent or excepient.
4. The pharmaceutical composition as claimed in claim 3, in the form of an oral
formulation.
5. The pharmaceutical composition as claimed in claim 4, wherein said pharmaceutically
acceptable carrier is selected from group consisting of starch, lactose, polyvinyl pyrolidone (K-
30), talc and magnesium stearatc.
6. The Pharmaceutical composition as claimed in claim 3 in the form of a parenteral
formulation.
7. A method for the preparation of a parenteral formulation as claimed in claim 6. which
comprises dissolving one or more compounds as defined in claim 1, in polyethylene glycol 400
and diluting the solution so obtained, with an isotonic solution or water to a desired
concentration.
8. Pharmaceutical composition as claimed in claim 5, in the form of a lotion, oral rinse and
toothpaste.
9. A process for preparation of pyridinium compounds substantially as herein described
particularly with reference to the examples.
10. A pharmaceutical composition substantially as herein described particularly with
reference to examples.
The invention discloses novel compounds of the pyridinium series useful for the
management of diabetes and aging-related vascular complications, including kidney disease,
nerve damage, atherosclerosis, retinopathy, dermatological disorders and discoloration of teeth,
by breaking performed AGE.
The invention also discloses, method for preparation of the novel compounds of the series
and pharmaceutical composition having one or more compounds as defined above as active
ingredients.

Documents:

604-CAL-2001-CORRESPONDENCE.pdf

604-CAL-2001-FORM 27.pdf

604-CAL-2001-FORM-27.pdf

604-cal-2001-granted-abstract.pdf

604-cal-2001-granted-assignment.pdf

604-cal-2001-granted-claims.pdf

604-cal-2001-granted-correspondence.pdf

604-cal-2001-granted-description (complete).pdf

604-cal-2001-granted-examination report.pdf

604-cal-2001-granted-form 1.pdf

604-cal-2001-granted-form 18.pdf

604-cal-2001-granted-form 2.pdf

604-cal-2001-granted-form 3.pdf

604-cal-2001-granted-form 5.pdf

604-cal-2001-granted-gpa.pdf

604-cal-2001-granted-letter patent.pdf

604-cal-2001-granted-reply to examination report.pdf

604-cal-2001-granted-specification.pdf

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Patent Number

214276

Indian Patent Application Number

604/CAL/2001

PG Journal Number

06/2008

Publication Date

08-Feb-2008

Grant Date

07-Feb-2008

Date of Filing

19-Oct-2001

Name of Patentee

TORRENT PHARMACEUTICALS LTD,

Applicant Address

CENTRAL PLAZA, 1ST FLOOR, ROOM #-106, 2/6, SARAT BOSE ROAD, CALCUTTA-700020, WEST BENGAL, INDIA.

Inventors:

#	Inventor's Name	Inventor's Address
1	SANKARANARAYANAN ALANGUDI	B-7, SWASTIK PARK PREMCHANDNAGAR ROAD OPP. JUDGES'BUNGLOWS AHMEDABAD-380 015, INDIA.

PCT International Classification Number

A61K31/444,A61P9/10

PCT International Application Number

N/A

PCT International Filing date

PCT Conventions:

#	PCT Application Number	Date of Convention	Priority Country
1	09/939,702	2001-08-28	U.S.A.