FORM 2
THE PATENTS ACT 1970
(39 OF 1970)
&
The patent Rules, 2003
COMPLETE SPECIFICATION
A Process for the Preparation of Ursodeoxycholic acid
SeQuent Scientific Limited
A Company Incorporated Under The Companies Act, 1956
Having Registered Office at 301, 'Dosti Pinnacle', 3rd Floor,
Plot No.E7, Road No.22, Wagle Industrial Area,
Thane (W)-400 604
The following specification particularly describes the invention and the manner in which it is, to be performed:

Field of Invention
The present invention relates to a novel, cost-effective process for the preparation of a cholelitholytic agent, which is commonly known as ursodeoxycholic acid.
Background of the Invention
Ursodeoxycholic acid is a bile acid, an anticholelithogenic, which suppresses hepatic synthesis and secretion of cholesterol and also inhibits intestinal absorption of cholesterol. Due to its many functions, ursodeoxycholic acid is an active ingredient of considerable interest in human therapy; for example, it promotes the dissolution of gallstones, lowers the percentage of cholesterol in the blood and is also used as a diuretic and as fat metabolism accelerator.
Ursodeoxycholic acid is marketed under the trade name "Actigall". It has been approved for the treatment of prevention of gallstone formation and primary biliary cirrhosis. Ursodeoxycholic acid is chemically known as 3α,7β-dihydroxy-5p-cholan-24-oic acid which is represented by formula I,

There are number of literatures available which describe the process for preparation of Ursodeoxycholic acid. Ursodeoxycholic acid is first disclosed in Journal of Biochemistry (1927), 7, 505-517. According to this article, Ursodeoxycholic acid is prepared by reacting Chenodeoxycholic acid with barium salt. Chenodeoxycholic acid on reaction with barium salt produces Chenodeoxycholic acid barium salt, which on hydrolysis to produce Chenodeoxycholic acid hemihydrate, which is further reacted with acid to produce crude

Ursodeoxycholic acid. Crystallization of crude Ursodeoxycholic acid from dilute alcohol or acetone yields Ursodeoxycholic acid.
US patent 4834919 discloses a process for the preparation of Ursodeoxycholic acid by treating Potassium salt of 7-ketolithocholic acid with potassium tertiary butoxide in isopropanol, further the above reaction mass was hydrogenated using Raney nickel under hydrogen pressure to produce crude Ursodeoxycholic acid. Further, the obtained crude Ursodeoxycholic acid is purified using acetone and . triethylamine to produce pure Ursodeoxycholic acid.
The main drawback of above process is the use of Raney nickel (100% w/w) this leads to the more impurity formation i.e, iso Ursocholic acid is 7-10 % in crude Ursodeoxycholic acid. The elimination of this impurity requires multiple purification steps. Hence, this process is not a suitable chemical process.
US patent 4282161 discloses a process for the purification of Ursodeoxycholic acid by treating crude Ursodeoxycholic acid with methanol to produce Ursodeoxycholic acid methyl ester, which is further crystallized by ethyl acetate, followed by saponification using alkali base to produce Ursodeoxycholic acid alkali metal salt. Further, the obtained Ursodeoxycholic acid alkali metal salt is treated with an acid in the presence of chloroform to produce crystalline Ursodeoxycholic acid.
The main drawback of above purification process is the more number of steps such as esterification, saponification and hydrolysis steps. As the number of steps is more in a chemical process there is apparent lowering of the overall yield and also required time cycle for production is more. Thus, this process is not a suitable chemical process. Further, this process involves the usage of huge volume of chloroform, which is not suitable on commercial scale.
Lipids (1981), 16(11), 863-865 reports the purification of crude Ursodeoxycholic acid by crystallizing from ethyl acetate to produce pure Ursodeoxycholic acid. The disadvantage of the above purification process that, it does not eliminate Chenodeoxycholic acid impurity, which is formed during reduction of 3a-hydroxy-7-oxo cholanic acid.

JP patent 58146597 discloses a process for the purification of Ursodeoxycholic acid, by treating crude Ursodeoxycholic acid with N-alkylmorpholines to produce corresponding N-alkylmorpholine salt, which is further hydrolyzed to produce Ursodeoxycholic acid.
JP patent 3810610 discloses the purification of Ursodeoxycholic acid by treating crude Ursodeoxycholic acid with triethylamine or 4-dimethylaminopyridine to produce corresponding triethylamine and 4-dimethylaminopyridine salts of Ursodeoxycholic acid which are further hydrolyzed to produce Ursodeoxycholic acid. The major disadvantage of the above processes is that achieving pharmaceutically acceptable limits of triethylamine and DMAP are difficult.
There are several known processes for preparing ursodeoxycholic acid. All the known processes have the disadvantage of leading to the production of a mixture of ursodeoxycholic, chenodeoxycholic, lithocholic, cholic and isourso in variable amounts from process to process.
Hence, there is a need to develop a process for preparing high purity ursodeoxycholic acid, which provides isolation of pure Ursodeoxycholic acid, which reduces the unwanted impurities with less no of steps as well as with higher yield and the cost of which are as contained as possible.
Thus it is highly desirable to develop a process which overcomes the drawbacks of the prior art. The present inventors have developed a very cost effective process for the preparation of Ursodeoxycholic acid of formula I, which gives better purity, better yield and avoids the formation of unwanted impurities.
Summary of the invention
According to the principal aspect, the present invention provides a novel process for the preparation of Ursodeoxycholic acid of formula I which comprises:
a) reducing 7-ketolithocholic acid of formula II using a reducing agent and an alkoxide in presence of a solvent to obtain crude Ursodeoxycholic acid of formula I; and

b) purifying crude Ursodeoxycholic acid of formula I using a base and in presence of solvent to obtain pure crude Ursodeoxycholic acid of formula I;
The present invention can be illustrated by the below reaction scheme 1:
Reaction Scheme 1


Detail Description of the Invention
Accordingly in an embodiment of the invention, 7-ketolithocholic acid of formula II in step (a) is reduced using a reducing agent selected from a group consisting of palladium on carbon, sodium borohydride, sodium cyanoborohydride and Raney nickel, preferably raney nickel and an alkoxide preferably alkali metal alkoxide, more preferably potassium tertiary butoxide in presence of a solvent selected from the group comprising of methanol, ethanol, propanol, isopropanol, butanol and the mixture thereof, most preferably isopropanol to obtain crude Ursodeoxycholic acid of formula I. The reduction is preferably carried out using 5-100% of Raney nickel, preferably 20% of raney nickel, 0.5 to 2.5 equivalent of potassium tertiary butoxide, preferably 1.2 equivalent potassium tertiary butoxide, at about 1-15 kg /cm2 of hydrogen pressure, preferably at 4-5 kg /cm2 of hydrogen pressure and at a temperature in the range of about 25-85°C preferably at 45-50°C for about 8 to 12 hours preferably for about 10 hours, to form crude Ursodeoxycholic acid of formula I.
In another embodiment of the invention, the unreacted 7-Ketolithocholic acid of formula II (<1.0%) in step a) is further reduced after hydrogenation to form Chenodeoxycholic acid using sodium borohydride (1% w/w). Conversely, if the percentage of 7-Ketolithocholic acid is greater than 0.5%, then it is very difficult to remove by further purification.
In yet another embodiment of the invention, the crude Ursodeoxycholic acid of formula I obtained in step (a) is >96% pure containing Chenodeoxycholic acid preferably in the range of 3.0 - 3.5% and Iso ursodeoxycholic acid preferably in the range of 0.02 -0.06% as impurities.
In another embodiment of the invention, crude Ursodeoxycholic acid of formula I obtained in step (a) is purified as in step (b) using a base selected from a group comprising triethylamine, trimethylamine, diisopropylethyl amine, pyridine etc., preferably triethylamine in presence of solvent preferably ketonic solvent, selected from a group comprising acetone, acetophenone, ethyl isopropyl ketone, methyl ethyl ketone etc., preferably acetone, preferably at reflux temperature for about 2 hours to obtain

Ursodeoxycholic acid triethylamine salt. Finally, the hydrolysis of obtained Ursodeoxycholic acid triethylamine salt is carried out at the pH range of 2-5 preferably at pH 4 at a temperature in the range of about 25-60°C preferably at 40 to 50°C to form crude Ursodeoxycholic acid of formula I.
In other embodiment of the invention, wet crude Ursodeoxycholic acid of formula I obtained in step (b) is purified using a ketonic solvent selected from a group comprising acetone, methyl isobutyl ketone, ethyl isopropyl ketone, methyl ethyl ketone etc., preferably methyl isobutyl ketone, by re-crystallization using 1 to 25 volume of methyl isobutyl ketone, preferably using 6-8 volume and at a temperature in the range of 50-118°C, preferably at 80-95°C, cooling to a temperature in the range of 0 to 50°C, preferably to 10-20°C and filtering to get pure Ursodeoxycholic acid of formula I.
In further embodiment of the invention, the crude Ursodeoxycholic acid of formula I obtained in step (a) is further purified as a process described in step (b) to obtain pure Ursodeoxycholic acid of formula I with a purity of >99% containing Chenodeoxycholic acid preferably in the range of 0.2 - 0.5% and Iso ursodeoxycholic acid preferably in the range of 0.02 - 0.06% as impurities.
In still further embodiment, the process of present invention has following advantages:
1. The process of prior art uses potassium salt of 7-Ketolithocholic acid, which takes 20 hours for the completion of keto reduction, whereas the process of present invention involves 7-ketolithocholic acid as a starting material, completing the reaction within 10 hours. Hence reaction time is reduced.
2. In the process of the present invention, the percentage of iso ursodeoxycholic acid in the crude Ursodeoxycholic acid is less than 0.2%, which can be removed easily during the further purification step. However, in the prior art processes the percentage of iso ursodeoxycholic acid in the crude Ursodeoxycholic acid is about 7-10%, which is very difficult to remove.

3. In the process of the present invention the use of Raney nickel is in the range of 5 - 30%, resulting iso ursodeoxycholic acid in the crude Ursodeoxycholic acid is less than 0.2%, which otherwise approximately 7-10% in prior art processes. Further, in the present invention iso ursodeoxycholic acid can be removed easily during further purification step.
4. The process of the present invention involves the use of sodium borohydride (1% w/w) to reduce unreacted 7-Ketolithocholic acid (<1.0%) to Chenodeoxycholic acid. If the percentage of 7-Ketolithocholic acid is greater than 0.5%, then it is very difficult to remove by further purification.
5. The process of the present invention involves the pH adjustment in the final purification step is carried out at 40-50°C to obtain free solid of pure Ursodeoxycholic acid which otherwise forms fine solid leading to slow filtration.
The present invention is illustrated by the following examples, which are not to limit the scope of the invention.
Examples:
a) Preparation of crude Ursodeoxycholic acid:
7-Ketolithocholic acid (40g, 0.1024 mole) was slurried using isopropanol (600 mL) and potassium tertiary butoxide (13.8g, 0.12 mole) for 30 minutes. Raney nickel (8g, 20% w/w) was added and the reaction mass is maintained at 45-55° C under 4-5kg/cm2 of hydrogen pressure in an autoclave for 10 hours. After completion of reaction, the reaction mass was cooled to 25-30°C, the catalyst was filtered and isopropanol was distilled. The residue was dissolved in water, sodium borohydride (1% w/w) was added, stirred and the pH was adjusted to 2-3 at 40-50° C. The 39.2g of crude Ursodeoxycholic acid was isolated by filtration.
b) Preparation of pure Ursodeoxycholic acid:
The 39.2g of crude Ursodeoxycholic acid was taken in 315 mL of acetone, heated to 35°Triethyalmine (13.8 mL 1.0 eq)was added in drops, reaction mass was further heated

to reflux for 2 hours, cooled to 25-35°C and TEA salt of Ursodeoxycholic acid was filtered. The TEA salt is dissolved in water, the pH was adjusted to 2-4 at 40-50°C and ursodeoxycholic acid was isolated by filtration. The wet cake is dissolved in 6 volume of Methyl isobutyl ketone, heated to 80-95°C, cooled to 10-20°C. The 32g of Ursodeoxycholic acid was isolated by filtration.

We claim:
1. A process for the preparation of Ursodeoxycholic acid formula I, which comprises:

a) reducing 7-Ketolithocholic acid of formula II using a reducing agent and an alkoxide in presence of a solvent to obtain crude Ursodeoxycholic acid of formula I; and

b) purifying crude Ursodeoxycholic acid of formula I using a base and in presence of solvent followed by recrystallization to obtain pure crude Ursodeoxycholic acid of formula I;
2. A process according to claim 1, wherein reducing agent in step (a) is selected from the group comprising of palladium on carbon, sodium borohydride, sodium cyanoborohydride and Raney nickel, preferably Raney nickel.
3. A process according to claim 1, wherein an alkoxide in step (a) is alkali metal alkoxide, preferably potassium tertiary butoxide.

4. A process according to claim 1, wherein solvent in step (a) is selected from the group comprising methanol, ethanol, propanol, isopropanol, butanol and the mixture thereof, preferably in isopropanol.
5. The quantity of Raney nickel of which with respect to 7-Ketolithocholic acid is in the range of 5-30% preferably 20%.
6. A process according to claim 1, wherein the reduction in step (a) is carried out at temperature in the range of 25-85°C preferably 45-55°C.
7. A process according to claim 1, wherein the base in step (b) is selected from triethylamine, trimethylamine, diisopropylethyl amine, pyridine etc., preferably triethylamine.
8. A process according to claim 1, wherein solvent in step (b) is a ketonic solvent selected from acetone, methyl isobutyl ketone, ethyl isopropyl ketone, methyl ethyl ketone etc., preferably acetone.
9. A process according to claim 1, wherein solvent for recrystallization in step (b) is a ketonic solvent selected from acetone, methyl isobutyl ketone, ethyl isopropyl ketone, methyl ethyl ketone etc., preferably methyl isobutyl ketone.
10. The pure Ursodeoxycholic acid of formula I obtained by a process according to claim 1, having purity > 99.5% containing Chenodeoxycholic acid of formula III preferably in the range of 0.2 - 0.5%) and Iso ursodeoxycholic acid of formula IV preferably in the range of 0.02 - 0.06% and other specified and unspecified impurities <0.05%.