CHAPTER-IV An improved process for synthesis of Febuxostat which is an inhibitor of xanthine oxidase.

INTRODUCTION : The clinical manifestations of gout, a spectrum of monoarthritic disorders characterized by crystallization of monosodium urate from supersaturated body fluids into tissues, have been well described for centuries. Although historically associated with royalty and affluent societies, increased longevity and shifts in patterns of diet and lifestyle have led to an increasing prevalence of gout worldwide, including in less-developed countries. 1,2 Attacks of acute gouty arthritis are usually treated with NSAIDs, colchicine, or corticosteroids; however, because hyperuricemia is the primary antecedent biochemical abnormality observed in patients with acute gouty arthritis, urate-lowering agents are the foundation for prevention of further attacks. Colchicine was used originally to treat rheumatic complaints, especially gout. It has toxic side effects which include gastrointestinal upset and neutropenia. 3 Colchicine poisoning has been compared to arsenic poisoning; symptoms start 2 to 5 hours after the toxic dose has been ingested and include burning in the mouth and throat, fever, vomiting, diarrhea, abdominal pain and kidney failure. 4 Hyperuricemia in humans is best defined as a serum uric acid of >6.8 mg/dL, which approaches the limit of solubility for monosodium urate in extracellular fluids. 5 Uric acid is the terminal product of a cascade of metabolic steps produced by xanthine oxidase from xanthine and hypoxanthine, which in turn are produced from purine. Uric acid is more toxic to tissues than either xanthine or hypoxanthine. Uric acid is released in hypoxic conditions. 6 In humans and higher primates, uric acid is the final oxidation (breakdown) product of purine metabolism and is excreted in urine.

Excess serum accumulation of uric acid can lead to a type of arthritis known as gout. 7 This painful condition is the result of needle-like crystals of uric acid precipitating in joints and capillaries. Various factors, such as age, body weight, diet, temperature, and pH, are known to influence both the concentration and solubility of monosodium urate; however, normal

128 physiologic homeostasis is able to maintain serum uric acid levels below the point of supersaturation and subsequent crystal formation. As the total pool of serum uric acid in the body rises, either because of overproduction or underexcretion, the risk of an acute gout attack increases in a continuous manner. The estimated 5-year cumulative risk of gout is <1% in patients with serum uric acid <7 mg/dL, but >25% of those with urate levels >10 mg/dL will likely experience an attack. 8 In chronic gout, polyarticular involvement may be noted. If left untreated, acute gout attacks generally resolve within 2 weeks. Approximately 80% of patients experiencing their first gout attack will have a recurrence within 2 years. 9 If the underlying hyperuricemia is left untreated, intercritical periods become shorter and attacks become more common. Therefore, reducing the total body pool of urate with lifestyle and pharmacologic interventions is an important step in preventing recurrent attacks. The main reasons for high uric acid level are:

• Diet may be a factor in Metabolic Syndrome, fructose and sucrose can cause increased levels of uric acid.

• Eating large amounts of sea salt can cause increased levels of uric acid

• Serum uric acid can be elevated due to reduced excretion by the kidneys.

• Serum uric acid can be elevated due to high intake of dietary purine.

• Fe activates xanthine oxidase (XO) and Cu deactivates it, so that as men accumulate Fe with age and Cu levels decline as testosterone levels drop with age (testosterone increases Cu half life), eventually the high Fe/Cu results in more active XO and higher urate levels.

A xanthine oxidase inhibitor is any substance that inhibits the activity of xanthine oxidase, an enzyme involved in purine metabolism. In humans, inhibition of xanthine oxidase reduces the production of uric acid, and several medications that inhibit xanthine oxidase are indicated for treatment of hyperuricemia and related medical conditions including gout. Xanthine oxidase inhibitors are being investigated for management of reperfusion injury.

129

Xanthine oxidase inhibitors are of two kinds: purine analogues and others. Purine analogues include allopurinol, oxypurinol,10 and tisopurine. Other group of xanthine oxidase inhibitors include febuxostat 11 and inositols. In some cases, for allopurinol, severe life- threatening side effects have been reported. These include a toxicity syndrome dramatized by eosinophilia, vasculitis, rash hepatitis, and progressive renal failure. Therefore, novel non-purine alternatives to allopurinol with potent XO inhibitory activity, but possessing fewer side effects are in great demand. Under efforts to find novel XO inhibitors without purine backbone, 2-phenylthiazoles and 1-phenylpyrazoles had been designed and tested as xanthine oxidase inhibitors. Among them, febuxostat is shown to be promising xanthine oxidase inhibitor. Febuxostat received marketing approval by the European Medicines Agency on April 21, 2008 12 and was approved by the U.S. Food and Drug Administration on February 16, 2009. 13

OMe

OMe

O OMe O N H3C H N NH O OMe N N H Cholchicine Allopurinol

CH3

CH3 O

S O NC

N OH

CH3 Febuxostat

130

PRESENT WORK Chemically febuxostat is 2-(3-cyano-4-isobutoxyphenyl)-4-methyl-1,3-thiazole-5- carboxylic acid which is an inhibitor of xanthine oxidase that is indicated for the use in treatment of hyperuricemia and gout. It is a non-purine selective inhibitor of xanthine oxidase. It works by non-competitively blocking the channel leading to the active site on xanthine oxidase. Xanthine oxidase is needed to successively oxidate both hypoxanthine and xanthine to uric acid. Febuxostat inhibits xanthine oxidase activity, therefore reducing production of uric acid.

CH3

CH3 O

S O NC

N OH

CH3 Febuxostat (1) Process development work was undertaken to make drug available in country. The reported methods for synthesis of febuxostat involve construction of thiazole ring from properly substituted benzene derivative. One of the method to prepare febuxostat ( 1) as per Scheme-114 involves reaction of 4- nitrobezonitrile with KCN in hot DMSO followed by treatment with isobutyl bromide and potassium carbonate to give intermediate 4. Reaction of 4 with thioacetamide in hot DMF gives intermediate 5 which on cyclization using Ethyl-2-chloroacetoacetate followed by alkaline hydrolysis gives febuxostat ( 1).

131

CH 3 CH3

CH 3 CH3 KCN/DMSO, O MeCSNH 2 O2N O Isobutyl bromide/K 2CO 3 DMF 70°C/6.0 h 45°C/ 40 h S CN NC CN NC

4 5 NH2

CH CH3 3

CH CH3 3 Ethyl-2-chloroacetoacetate THF/Ethanol O O Ethanol NaOH O 100°C/ 2 h S 60°C S NC NC COOEt N N OH CH CH3 3 3 Febuxostat (1) Scheme 1

In another synthesis 15 of febuxostat ( 1), 4-hydroxy-3-nitobenzaldehyde is reacted with hydroxylamine and sodium formate in refluxing formic acid to give 4-hydroxy-3- nitobenzbenzonitrile ( 6) which is further treated with thioacetamide in hot DMF to yield corresponding thiobenzamide 7. Cyclization of 7 with ethyl-2-chloroacetoacetate in refluxing ethanol gives intermediate 8, which on o-alkylation with isobutyl bromide in presence of K2CO 3 in hot DMF providing the isobutyl ether 9. The reduction of nitro group

of 9 with H 2/Pd-C gives amino derivative 10 , which on diazotization with NaNO 2/HCl followed by treatment with CuCN and KCN gives 3. The alkaline hydrolysis of 3 gives febuxostat ( 1) (Scheme-2) .

132

MeCSNH Ethyl-2-chloroacetoacetate NH OH 2 2 HO - HO Ethanol HO HCOONa DMF HCl

Formic acid 80°C/ 1 h S reflux/ 5 h O N CN O2N O2N CHO reflux/ 5.0 h 2 NH 6 7 2

CH3 CH3

CH3 Isobutyl bromide CH HO 3 H2,Pd-C DMF/K CO O 2 3 O EtOH, EtOAc S 70°C/18 h RT/24 h S O2N H N COOEt S 2 COOEt O N N 2 COOEt N N CH CH 3 10 3 8 9 CH3

CH3 CH3

CH CH 3 3 THF/Ethanol NaNO 2, HCl O CuCN, KCN O NaOH 60°C S O S NC NC COOEt N N OH

CH3 CH3 Febuxostat (1) 3 Scheme 2

16 In one more synthesis of febuxostat ( 1) (scheme 3), condensation and cyclization of 4-hydroxythiobenzamide with 2-bromoacetoacetic acid ethyl ester in refluxing ethanol provides 11 which is formylated by reaction with hexamethylenetetramine and polyphosphoric acid in hot acetic acid/water to afford 12 . Alkylation of 12 with isobutyl bromide in presence of potassium carbonate and potassium iodide in dimethylformamide gives 13 , which on treatment with formic acid, sodium formate and hydroxylamine hydrochloride yields 3. Finally alkaline hydrolysis of 3 using sodium hydroxide in THF/ethanol gives febuxostat ( 1).

133

Ethyl-2-bromo- OH HO HO acetoacetate HMTA Ethanol S S OHC reflux COOEt PPA/100°C COOEt N N

CH 3 CH3 SNH2

4-Hydroxythiobenzamide 11 12

CH3 CH3

CH3 CH3 Isobutyl bromide O O NH OH.HCl DMF/K 2CO 3 2 S S HCOONa/HCOOH NC OHC COOEt COOEt N N

CH3 CH3 13 3

CH3

CH3 O NaOH

S O THF/Ethanol NC

N OH

CH3 Febuxostat (1) Scheme 3

The first two methods (scheme 1 & 2) have following drawbacks in comparison to scheme 3. These drawbacks include: • Use of hazardous reagents like KCN, CuCN which are industrially unsafe. • Low yields and use of column chromatography for purification process, which makes it industrially unviable.

A key step in the synthesis of febuxostat as per scheme 3 was introduction of formyl group selectively at ortho position to the hydroxyl group in compound 11 . Classical Duff reaction 17 of phenol derivative using HMTA and acetic acid results in ortho formylation.

134

Under similar conditions, the compound 11 failed to give required formyl derivative 12 . However, replacement of acetic acid with polyphosphoric acid gives 12 with 57% yield. In addition to poor yield, polyphosphoric acid has also certain unfortunate physical properties as listed below restricting its use in commercial production. a. It is extremely viscous and is virtually impossible to stir effectively or manipulate conveniently at temperatures below 60°C. b. It is difficult to handle on a large scale, even at elevated temperatures. c. Some organics are only sparingly soluble in PPA, and, in any case, rates of dissolution are low. d. Work up to isolate products from reactions involving PPA is always tedious.

Process development work was initiated starting from easily available 4- hydroxythiobenzamide to find suitable alternative to polyphosphoric acid for introduction of formyl group in compound 11 to get 12 (scheme 4).

135

Ethyl-2-chloro- HO OH acetoacetate HO methanol HMTA S ------> S OHC reflux COOEt COOEt N N CH 3 CH3 SNH2

4-Hydroxythiobenzamide 11 12

CH3 CH3

CH3 CH3 Isobutyl bromide O O NH OH.HCl DMF/K 2CO 3 2 S S HCOONa/HCOOH NC OHC COOEt COOEt N N

CH3 CH3 13 3

CH3

CH3 O Hydrolysis ------> S O NC

N OH

CH3 Febuxostat (1) Scheme 4

Conversion of 4-hydroxythiobenzamide to 11 was studied using cheaply available ethyl-2-chloroacetoacetate in place of ethyl-2-bromoacetoacetate. The reaction was completed in 2 hours by refluxing the mixture of 4-hydroxythiobenzamide and ethyl-2- chloroacetoacetate in methanol. After complete conversion, methanol was distilled under reduced pressure and water was added to the reaction mixture. The pH was adjusted at 6 to 7 using sodium bicarbonate and the reaction mixture was filtered, washed with water and dried. The product was characterized by spectral analysis. The formylation of 11 was studied initially using acetic acid, trifluroacetic acid, methanesulfonic acid (neat) instead of PPA. Using trifluroacetic acid rate of reaction was

136 slow and reaction does not go to completion. Using acetic acid and methanesulfonic acid no reaction was observed. In continuation with the efforts, the formylation of 11 was studied using Eaton’s reagent in presence of HMTA at 80-90°C. To our satisfaction this modification afforded the expected product 12, which was confirmed by mass and NMR spectroscopy. Eaton’s reagent was discovered in year 1973 and was used for preparation of cyclopentenones via intramolecular acylation of olefin acids or their lactones and the preparation of amides via Beckmann rearrangement. Since then, it has been used as better alternative for PPA in many synthetic transformations such as cyclization reaction to form quinolones & quinolone heterocycles 18 , intramolecular cyclization by acylation with carboxylic acid, 19 for quinoline formation 20 and for the formation of Tetrahydroisoquinoline-2-ones. 21 Eaton’s reagent 22 has the following additional advantages over polyphosphoric acid method: a. It is a free flowing colorless liquid that can be poured and stirred without difficulty. b. Organic compounds dissolve readily in this medium. c. Work-up of phosphorus pentoxide-methanesulfonic acid reaction mixture is easy. d. The reagent can be destroyed conveniently with water.

In optimized process, initially Eaton’s reagent was prepared by dissolving P 2O5 in hot

methanesulfonic acid (10% P 2O5 in MsOH) at 90-100°C. After cooling at 40-50°C, hexamethylenetetramine (HMTA) was added slowly below 80°C. Here addition of HMTA to Eaton’s reagent was found to be exothermic. After addition of HMTA, compound 11 was added and the reaction mass was stirred at 85-90°C for 6 hours. The reaction was monitored by HPLC where complete consumption of intermediate 11 was observed. The compound 12 was isolated by quenching the reaction mass in cold water followed by extraction in ethyl acetate. Finally ethyl acetate layer was concentrated under reduced pressure to get solid product. The product was obtained in good yield (> 80%) and high HPLC purity (> 95%).

137

HO HO

S HMTA S OHC COOEt COOEt N P2O5 , MsOH N 85-90°C CH 3 CH3

11 12 Scheme 5

The quantity of P 2O5 in Eaton’s reagent was optimized by carrying out experiments using different molar ratios of P 2O5 with respect to substrate. The best optimized quantity for preparation of Eaton’s reagent was 0.5 equivalents of P 2O5 and 5 times (w/w) methanesulfonic acid on substrate ( 11 ). The reaction temperature was also optimized by carrying out reactions at 25-30°C, 60-65°C and 80-90°C and monitoring the reaction by HPLC. The results are summarized in table-1. Table 1 0 Equivalents of P 2O5 Reaction Temp ( C) Reaction conversion (%) by HPLC after 6 hrs 0.10 80-90 64.45 0.25 80-90 81.54 0.50 80-90 93.62 1.00 80-90 91.14 0.50 25-30 No reaction 0.50 60-65 34.7

The best optimized temperature for this conversion was 80-90°C with period of 6 to 8 hours. Further compound 12 was converted to 13 by o-alkylation with isobutyl bromide. The mixture of 12 , isobutyl bromide and potassium carbonate in DMF was stirred at 75-80°C for 4 to 6 hours. After complete conversion, the reaction mixture was brought at 50-55°C and diluted in cold water for solid procipitation. After stirring the reaction mixture for 1.0 hour, it was filtered, washed with water and dried to give 13 . The product was characterized by spectral analysis.

138

The compound 13 was further converted to intermediate 3 by reaction with hydroxylamine hydrochloride and sodium formate in formic acid. The compound 13 was dissolved in 6.0 volumes of formic acid (98-99% pure). To a clear solution of 13 , hydroxylamine hydrochloride and sodium formate were added and the reaction mixture was stirred at 98-102°C for 4 hours. After complete conversion, the reaction mixture was brought at 70-75°C and diluted in cold water to precipitate solid. After stirring the reaction mixture for 1.0 hour, it was filtered, washed with water and dried to give 3. The product was characterized by spectral analysis. For this conversion, water content in formic acid was found to be critical. When formic acid having water content more than 2% was used, the compound 13 was not get dissolved completely and the reaction mixture became dark brown colored against normal pale yellow color. After addition of hydroxylamine hydrochloride and sodium formate and stirring the reaction mixture at 98-102°C for 4-6 hours, desired product formation was not observed but some other impurities were formed. Another step which attracted our attention was the alkaline hydrolysis of 3 using sodium hydroxide in a mixture of THF and ethanol to give febuxostat ( 1) which needs further purification. To avoid the use of mixture of solvents and further purification of febuxostat so obtained, hydrolysis was studied using aq. sodium hydroxide solution initially in methanol at 50-55°C. To a mixture of compound 3 in 5 volumes of methanol, aq. sodium hydroxide (50% solution ) was added and the reaction mixture was heated at 50-55°C to get clear solution. Reaction was monitored by TLC and was completed in 2 hours. The product was isolated in pure form (HPLC purity > 99.5%) as sodium salt by cooling and filtration of the reaction mixture slurry at 0-10°C temperature. Further sodium salt was converted to febuxostat by acidification using hydrochloric acid in presence of water and ethyl acetate. Eventhough quality of febuxostat obtained by hydrolyzing compound 3 from methanol was much better; the yield obtained was on lower side (50-55%). The lower yield of febuxostat so obtained could be attributed to the higher solubility of its sodium salt in methanol. To overcome this problem and to increase the yield of the final product, hydrolysis was studied in ethanol using sodium hydroxide (1.2 m/m) and 0.5 volumes of water at 70-75°C temperature. The reaction was completed in 2 hours and

139 product was isolated in pure form (HPLC purity > 99.5%) as sodium salt by cooling and filtration of the reaction mixture slurry at 0-10°C temperature. Further sodium salt was converted to febuxostat by acidification using hydrochloric acid in presence of water and ethyl acetate. The yield of final product was observed to be on higher side (65-70%) as compared to that obtained using methanol solvent.

CH3 CH3

Ethanol/Water/NaOH CH3 CH3 70-75°C O O Water/HCl S O S Ethyl acetate NC NC COOEt N N OH

CH3 CH3 Febuxostat (1) 3 Scheme 6 In conclusion, we have successfully developed a simple, cost effective and industrially scalable process for synthesis of febuxostat using Eaton’s reagent, a good substitute to polyphosphoric acid.

140

EXPERIMENTAL SECTION The 1H NMR spectra were recorded in DMSO-d6 on Varian 400 MHz; the chemical shifts were reported in δ ppm relative to TMS. The IR spectra were recorded in the solid state KBr dispersion using Perkins Elmer FT–IR spectrophotometer. The mass spectra were recorded on Applied Biosystems spectrometer. The melting points were determined by using Buchi apparatus .The solvents and reagents were used without purification. The purity by HPLC of all the compounds was checked on Waters make HPLC system using reverse phase C-18 column. 2-(4-hydroxyphenyl)-4-methyl-5-thiazole carboxylic acid ethyl ester (11): HO

S COOEt N

CH3 4-Hydroxybenzothiamide (1500 g, 9.79 mol), ethyl-2-chloroacetoacetate (1610 g, 9.79 mol) and methanol (3.0 L) were heated to reflux (65-70°C) for 4 h. Methanol (1.5 L) was distilled out from the reaction mixture at 65-70 °C and water (22.5 L) was added slowly. The reaction mixture was brought at 20 to 25°C, filtered and the residue was washed with 15.0 L of water. The wet cake was dried at 55 to 60 °C to give 11 with > 98.0 % purity by HPLC

[λmax : 225 nm; Mobile phase- Acetonitrile: Buffer (pH 2.5) in 20:80 ratio]. Yield : 2250 g, (87% yield)

Molecular formula : C13 H13 O3NS Melting point : 182-184°C IR (KBr) : 3097, 2995, 2984, 2908, 2802, 2677, 2596, 2473, 1898, 1723, 1606, 1536, 1521, 1426, 1407, 1375, 1316, 1285, 1268, 1172, 1095 cm -1 MS( m/z) : 264 1H NMR, : δ 1.29(t, J=7.1 Hz, 3 H), 2.65 (s, 3 H), 4.27 (q, J=7.1 Hz, 2 H), 6.87

(DMSO-d6, 400 MHz) (d, J=8.4 Hz, 2 H), 7.83 (d, J=8.3 Hz, 2 H), 10.21 (s, 1H) 13 C NMR : 14.1, 17.2, 60.8, 115.9, 119.6, 123.5, 128.3, 160.1, 160.6, 161.5,

(DMSO-d6, 100 MHz) 169.4. Elemental analysis : Calcd.: C, 59.31; H, 4.94; N, 5.32; S, 12.16. Found: C, 59.34;

141

H, 4.87; N, 5.36; S, 12.14%. 2-(3-formyl-4-hydroxyphenyl)-4-methyl-5-thiazole carboxylic acid ethyl ester (12): HO

S OHC COOEt N

CH3 A mixture of methanesulfonic acid (10.0 Kg) and phosphorus pentoxide (540 g, 3.80 mol) was heated at 85 to 90°C for 1.0 h to get clear solution. After cooling at 60-65 °C, hexamethylenetetramine (1278 g, 9.23mol) was added portion wise by maintaining the reaction mass temperature below 90ºC. Compound 11 (2000 g, 7.6 mol) was added and the reaction mass was heated at 85 to 90 °C for 2 to 4 h. The reaction mixture was brought at 65 to 70 °C and quenched slowly in chilled water (20.0 L) at 10 to 30 °C. Ethyl acetate (14.0 L) was added into the reaction mass and stirred the reaction mass at 30 to 35 °C for 2 h. Lower aqueous layer was separated and water (20.0 L) was added to the organic layer. After adjusting the reaction mass pH at 7.0 to 7.5 with sodium bicarbonate, ethyl acetate was distilled completely at 60 to 65 °C under vacuum. The reaction mixture was brought at 25 to 30 °C, filtered and the residue was washed with water (20.0 L). The wet cake was dried at

55 to 60°C to give 12 with 96.0% purity by HPLC [ λmax : 225 nm; Mobile phase- Acetonitrile: Buffer (pH 3.5) in 30:70 ratio]. Yield : 1792 g, (81% yield)

Molecular formula : C14 H13 O4NS Melting point : 112-114°C IR (KBr) : 2999, 2989, 2869, 1705, 1664, 1652, 1617, 1592, 1524, 1497, 1426, 1376, 1317, 1288, 1268, 1210, 1172, 1094, 1006, 827, 738 cm -1 MS( m/z) : 292 1H NMR : δ 1.30 (t, J=7.1 Hz, 3 H), 2.65 (s, 3 H), 4.28 (q, J=7.1 Hz, 2 H),

(DMSO-d6, 400 MHz) 7.10 (d, J=8.7 Hz, 1 H), 8.08 (dd, 2J=6.3 Hz & 2.4 Hz, 1 H), 8.21 (d, J=2.4 Hz, 1 H), 10.31 (s, 1H) 13 C NMR : 14.4, 17.4, 61.5, 118.5, 120.5, 122.6, 123.7, 127.3, 134.0, 160.4,

(DMSO-d6, 100 MHz) 161.7, 163.4, 168.2, 191.0.

142

Elemental analysis : Calcd.: C, 57.73; H, 4.46; N, 4.81; S, 11.0. Found: C, 57.61; H, 4.42; N, 4.85; S, 11.16%.

2-(3-formyl-4-isobutyloxy phenyl)-4-methyl-5-thiazole carboxylic acid ethyl ester (13):

CH3

CH3 O

S OHC COOEt N

CH3 A mixture of compound 12 (1750 g, 6.01 mol), DMF (10.5 L), isobutyl bromide (2055 g, 15.0 mol) and potassium carbonate (2070 g, 15.0 mol) was heated at 75 to 80 °C for 4 to 6 h. Reaction mass was brought at 50 to 55 °C and quenched slowly in chilled water (17.5 L) at 25 to 30 °C. After stirring at 25 to 30°C for 1 h, the reaction mixture was filtered and the residue was washed with water (17.5 L). The wet cake was dried at 55 to 60 °C to give 13 with > 97.0 % purity by HPLC [ λmax : 225 nm; Mobile phase- Acetonitrile: Buffer (pH 2.0) in 50:50 ratio]. Yield : 2003 g, (96% yield)

Molecular formula : C18 H21 O4NS Melting point : 160-161°C IR (KBr) : 2969, 2958, 2873, 1710, 1685, 1608, 1505, 1470, 1430, 1391, 1368, 1321, 1285, 1259, 1175, 1104, 996, 962, 823 cm -1 MS( m/z) : 348 1 H NMR : δ 1.03 (d, J=6.7 Hz, 6 H), 1.30 (t, J=7.1 Hz, 3 H), 2.12 (m, 1

(DMSO-d6, 400 MHz) H), 2.68 (s, 3 H), 4.01 (d, J=6.4 Hz, 2 H), 4.27-4.32 (m, 2 H), 7.37 (d, J=8.7 Hz, 1H), 8.22-8.26 (m, 2 H), 10.41 (s, 1H) 13 C NMR : 14.1, 17.1, 18.8, 27.6, 61.1, 74.8, 114.5, 120.8, 124.4, 124.6,

(DMSO-d6, 100 MHz) 125.9, 134.1, 160.2, 161.3, 162.8, 167.7, 188.6. Elemental analysis : Calcd.: C, 62.24; H, 6.05; N, 4.03; S, 9.22. Found: C, 62.22; H, 6.03; N, 4.07; S, 9.25%.

143

2-(3-cyano-4-isobutyloxy phenyl)-4-methyl-5-thiazole carboxylic acid ethyl ester (3):

CH3

CH3 O

S NC COOEt N

CH3 A mixture of compound 13 (2000 g, 5.76 mol) and formic acid (12.0 L) was heated at 60 to 65°C for 1.0 hour to get clear solution. After cooling at 25-30 °C, hydroxylamine hydrochloride (580 g, 8.35 mol) and sodium formate (745 g, 10.95 mol) were added and the reaction mass was heated at 100 to 102°C for 4 to 6 h. The reaction mixture was brought at 70 to 75 °C and quenched slowly in chilled water (20.0 L) at 10 to 30 °C. After stirring at 25 to 30°C for 1 h, the reaction mixture was filtered and the residue was washed with water (30.0 L). The wet cake was dried at 55 to 60 °C to give 3 with > 95.0% purity by HPLC

[λmax : 225 nm; Mobile phase- Acetonitrile: Buffer (pH 3.5) in 30:70 ratio]. Yield : 1842 g, (93% yield)

Molecular formula : C18 H20 O3N2S Melting point : 174-176°C IR (KBr) : 2973, 2933, 2875, 2360, 2227, 1712, 1606, 1511, 1471, 1429, 1369, 1301, 1262, 1171, 1128, 1100, 1014, 825 cm -1 MS( m/z) : 345 1 H NMR : δ 1.02 (d, J=6.7 Hz, 6 H), 1.31 (t, J=7.1 Hz, 3 H), 2.09 (m, 1 H),

(DMSO-d6, 400 MHz) 2.68 (s, 3 H), 4.00 (d, J=6.5 Hz, 2 H), 4.30 (q, J=7.1 Hz, 2 H), 7.37 (d, J=9.0 Hz, 1H), 8.24 (dd, 2J=6.5 Hz & 2.3 Hz, 1 H), 8.30 (d, J=2.3 Hz, 1 H) 13 C NMR : 14.1, 17.1, 18.6, 27.5, 61.2, 75.1, 101.5, 113.9, 115.3, 121.3,

(DMSO-d6, 100 MHz) 125.1, 131.6, 133.1, 160.1, 161.3, 162.1, 166.7. Elemental analysis : Calcd.: C, 62.79; H, 5.81; N, 8.14; S, 9.30. Found: C, 62.84; H, 5.85; N, 7.85; S, 9.36%.

144

Febuxostat (1):

CH3

CH3 O

S O NC

N OH

CH3 To a solution of compound 3 (1800 g, 5.23 mol) in ethanol (7.2 L), sodium hydroxide solution (251 g, 6.27 mol dissolved in 900 mL water) was added slowly over 1.0 h and the reaction mixture was heated at 70 to 75°C for 4 to 6 h. Reaction mixture was cooled at 0 to 5°C, filtered and the residue was washed with chilled ethanol (3.6 L). The wet cake was dried at 55 to 60 °C to give sodium salt of febuxostat (1485 g, 84.0% yield with 99.5% purity by HPLC). Sodium salt of febuxostat (1400g, 4.14 mol) was admixed with water (14.0 L) and ethyl acetate (9.8 L) and the reaction mass pH was adjusted at 4.0 to 5.0 by drop wise addition of conc. HCl (420 mL). The reaction mixture was heated at 65 to 70°C to get clear solution of two layers. After separation of aq. Layer from bottom, org. layer was further washed with water (4.2 L) at 65 to 70°C. To org. layer water (14.0 L) was added and ethyl acetate was distilled at 65 to 70°C under vacuum. The reaction mixture was brought at 25 to 30°C, filtered and the residue was washed with 5.6 L of water. The wet cake was dried at 55 to

60°C to give 1 with 99.5% purity by HPLC [λmax : 225 nm; Mobile phase- Acetonitrile: Buffer (pH 2.5) in 45:55 ratio]. Yield : 1125 g, (86% yield)

Molecular formula : C16 H16 O3N2S Melting point : 201-202°C IR (KBr) : 3538, 3463, 2962, 2876, 2546, 2361, 2229, 1702, 1684, 1605, 1511, 1427, 1370, 1298, 1284, 1170, 1129, 1117, 1012, 956, cm -1 MS( m/z) : 317 1 H NMR : δ 1.01 (d, J=6.7 Hz, 6 H), 2.09 (m, 1 H), 2.65 (s, 3 H), 3.99

(DMSO-d6, 400 MHz) (d, J=6.5 Hz, 2 H), 7.35 (d, J=9.0 Hz, 1H), 8.20 (dd, 2J=6.6 Hz & 2.3 Hz, 1 H), 8.26 (d, J=2.3 Hz, 1 H), 13.41 (brs, 1 H)

145

13 C NMR : 17.2, 18.9, 27.8, 75.3, 101.6, 113.9, 115.6, 122.9, 125.4, 131.5,

(DMSO-d6, 100 MHz) 133.2, 159.8, 162.2, 162.9, 166.4 Elemental analysis : Calcd.: C, 60.75; H, 5.06; N, 8.86; S, 10.12. Found: C, 60.72; H, 4.94; N, 9.04; S, 10.32%.

146

1H NMR Spectra of compound 11

13 CNMR Spectra of compound 11

147

IR Spectra of compound 11

Mass Spectra of compound 11

148

HPLC Chromatogram of compound 11

149

1H NMR Spectra of compound 12

13C NMR Spectra of compound 12

150

IR Spectra of compound 12

Mass Spectra of compound 12

151

HPLC Chromatogram of compound 12

152

1H NMR Spectra of compound 13

13C NMR Spectra of compound 13

153

IR Spectra of compound 13

Mass Spectra of compound 13

154

HPLC Chromatogram of compound 13

155

1H NMR Spectra of compound 3

13C NMR Spectra of compound 3

156

IR Spectra of compound 3

Mass Spectra of compound 3

157

HPLC Chromatogram of Compound 3

158

1H NMR Spectra of compound 1

13 C NMR Spectra of compound 1

159

IR Spectra of compound 1

Mass Spectra of compound 1

160

HPLC Chromatogram of Compound 1

161

1H NMR Spectra of compound 14

13C NMR Spectra of compound 14

162

Mass Spectra of compound 14

163

REFERENCES 1. Saag K. G, Choi H. Epidemiology, risk factors, and lifestyle modifications for gout. Arthritis Res Ther. 2006;8 (Suppl 1). 2. Annemans L, Spaepen E, Gaskin M, et al. Gout in the UK and Germany: Prevalence, comorbidities and management in general practice 2000–2005. Ann Rheum Dis . 2008;67:960–966. 3. Colchicine for acute gout: updated information about dosing and drug interactions". National Prescribing Service . 14 May 2010. 4. Colchicine. National Institute for Occupational Safety and Health. Emergency Response Safety and Health Database, August 22, 2008. 5. Wortmann RL. Chapter 87: Gout and hyperuricemia. In: Firestein GS, Budd RC, Harris ED Jr, et al, eds. Kelley’s Textbook of Rheumatology . 8th ed. Philadelphia, Pa: Saunders/Elsevier; 2009. 6. Baillie, J.K.; M.G. Bates, A.A. Thompson, W.S. Waring, R.W. Partridge, M.F. Schnopp, A. Simpson, F. Gulliver-Sloan, S.R. Maxwell, D.J. Webb (2007-05). "Endogenous urate production augments plasma antioxidant capacity in healthy lowland subjects exposed to high altitude". Chest 131 (5): 1473–1478.] 7. Heinig M, Johnson RJ (December 2006). "Role of uric acid in hypertension, renal disease, and metabolic syndrome". Cleveland Clinic Journal of Medicine 73 (12): 1059–64.] 8. Campion EW, Glynn RJ, DeLabry LO. Asymptomatic hyperuricemia. Risks and consequences in the Normative Aging Study. Am J Med . 1987;82:421–426. 9. Yu TF, Gutman AB. Efficacy of colchicine prophylaxis in gout. Prevention of recurrent gouty arthritis over a mean period of five years in 208 gouty subjects. Ann Intern Med . 1961;55:179–192. 10. Iwanaga T, Kobayashi D, Hirayama M, Maeda T, Tamai I (December 2005). "Involvement of uric acid transporter in increased renal clearance of the xanthine oxidase inhibitor oxypurinol induced by a uricosuric agent, benzbromarone". Drug metabolism and disposition: the biological fate of chemicals 33 (12): 1791–5. 11. Becker MA, Schumacher HR, Wortmann RL, et al. (March 2005). "Febuxostat, a novel nonpurine selective inhibitor of xanthine oxidase: a twenty-eight-day, multicenter, phase II,

164

randomized, double-blind, placebo-controlled, dose-response clinical trial examining safety and efficacy in patients with gout". Arthritis and rheumatism 52 (3): 916–23. 12. “Adenuric (febuxostat) receives marketing authorisation in the European Union". http://www.ipsen.com/articles/mediacentre/pressreleases/20080505___autorisation_adenuric _eu_10.pdf. 13. "Uloric Approved for Gout". U.S. News and World Report. http://health.usnews.com/articles/health/healthday/2009/02/16/uloric-approved-for- gout.html. 14. Hasegawa, M. Heterocycles, Vol. 47, No.2, 1998 (p.857-864), 1998. 15. Kondo, S.; Fukushima, H.; Hasegawa, M.; et al. (Teijin Ltd.) US 5614520. 16. Watanabe Kanezo; Yarino, Tatsuo; Hiramatsu, Toshiyuki. (Teijin Ltd.) JP 1998045733. 17. J. C. Duff, J. Chem. Soc., 547, 1941. 18. Daniel Zewge, Cheng-yi Chen, Curtis Deer, Peter G. Dormer, Dave L. Hughes, J. Org. Chem . 2007 , 72, 4276-79. 19. R. L. Dorow, Paul M. Herrinton, Richard A. Hohler, Mark T. Maloney, Michael A. Mauragis, William E. McGhee, Jeffery A. Moeslein, Joseph W. Strohbach, Michael F. Veley. OPRD, 2006 , 10, 493-499. 20. Muthusamy Jayaraman, Phillip E. Fanwick, Mark Cushman, J. Org. Chem . 1998 , 63, 5736- 5737. 21. Luckner G. Ulysse, Qiang Yang, Mark D. McLaws, Daniel K. Keefe, Peter R. Guzzo, and Brian P. Haney. OPRD, 2010 , 14, 225-228 22. Eaton, P. E.; Carlson, G. R.; Lee, J. T.; J. Org. Chem. 1973 , 38, 4071.

165