FORM2
THE PATENTS ACT 1970
(39 of 1970)
AND
The Patents Rules, 2003
COMPLETE SPECIFICATION
(See section 10 and rulel3)
1. TITLE OF THE INVENTION:
"PROCESS FOR THE PREPARATION OF 6-[(l-METHYL-4-NITRO-lH-IMIDAZOL-5-YL)THIO]-lH-PURINE"
2. APPLICANT(S):
(a) NAME: NEON LABORATORIES LIMITED
(b) NATIONALITY: Indian Company incorporated under the Companies
Act, 1956
(c) ADDRESS: Damji Shamji Industrial Complex, Mahakafi Caves Road,
Andheri (East), Mumbai - 400093, Maharashtra, India
3.PREAMBLE TO THE DESCRIPTION:
The following specification particularly describes the invention and the manner in which It is to be performed

Technical Field:
The present invention reiates to a novel, one-pot process for preparing highly pure 6-[(l-methyl-4-nitro-IH-imidazol-5-yl)thio]-lH-purine by reacting 6-mercaptopurine with 5-halo-l-methyl-4-nitro-l H-imidazole in presence of a base in aqueous medium.
Background and Prior art:
6-[(l-Methyl-4-nitro-lH-imidazol-5-yl)thio]-lH-purine commonly known as azathioprine, is an immunosuppressive and antineoplastic drug. It is used to prevent organ rejection following organ transplantation and to treat a vast array of autoimmune diseases, including rheumatoid arthritis, pemphigus, inflammatory bowel disease (such as Crohn's disease and ulcerative colitis), multiple sclerosis, autoimmune hepatitis, atopic dermatitis, myasthenia gravis, neuromyelitis optica, restrictive lung disease and others.
Azathioprine can be used as a chemotherapeutic agent to inhibit lymphocyte purine metabolism. In the body, azathioprine is converted to mercaptopurine and thus has the same effects as that.
Azathioprine was first described in U.S. patent number 3056785. The patent discloses the preparation of azathioprine by reacting 6-mercaptopurine with 5-chIoro-l-methyI-4-nitro-I H-imidazole in presence of anhydrous sodium acetate in dry dimethyl sulfoxide for 7 hours at 100°C; maintaining the reaction mixture overnight at room temperature; followed by recrystallization from hot acetone. Hence time consuming procedure and need of purification are the main drawbacks of this process. The patent does not disclose yield and purity of azathioprine.
A polish patent PLI54611 discloses preparation of azathioprine by condensation of 6-mercaptopurine with 5-chforo-l-methy]-4-nitro-1 H-imidazole in presence of sodium carbonate and 1-methylimidazole or tert. amine or benzyitrimethylammonium hydroxide. The crude product was dissolved in aq. ammonia, contacted for 15 minutes with activated carbon and hot-filtered. The filtrate was cooled to 20°C, treated with water/acetic acid (1:1) and dried at 105-1lOo C to give azathioprine. Thus the process requires additional reagents and purification.

Polish Journal of Pharmacology and Pharmacy (J 978), 30(4), 593-9 discloses preparation of azathioprine by condensation of 6-mercaptopurine hydrate with 5-chloro-l-methyl-4-nitro-lH-imidazole in presence of dimethylformamide, tetrahydrofuran and ammonium hydroxide; followed by purification to provide pure azathioprine, however in 80% yield. In the manufacture of drugs for human therapy high standards of purity has to be met and the process to meet the required specification is often costly. Thus it is important in any of the steps of such process to keep costs as low as possible without compromising on yield, purity of product, and ease on manipulation.
Like any synthetic compound azathioprine can contain extraneous compounds or impurities that can come from any source. The impurities can be in the form of unreacted starting materials, by-products of the reaction, products of side reactions, or degradation products. Impurities in azathioprine are undesired and might even be harmful to a patient being treated with a dosage form containing the same.
The reported methods for preparation of azathioprine are time consuming and require unavoidable multi-step purification procedure.
Needless to say it is advantageous to develop a process for preparation of highly pure 3-azathioprine which eliminates the necessity of additional purification step.
Object of the invention:
An object of the invention is to overcome or ameliorate atleast one disadvantage of the prior art or to provide a useful alternative.
Another object of the invention is to provide a novel, concise, high yielding, commercially viable and industrially applicable process which eliminates the need of additional step of purification and provides highly pure azathioprine having desirable pharmacological activity, broad safety margins, without toxicity or unfavourable side effects.

Summary of the invention:
In accordance with the above objectives, the present invention provides a process for preparation of azathioprine comprising reacting 6-mercaptopurine and 5-chloro-l-methyl-4-nitro-lH-imidazoIe in presence of atleast one base and water.
Detailed description of the invention:
Unless specified otherwise, all technical and scientific terms used herein have the same meaning as commonly understood by one of ordinary skill in the art, to which this invention belongs. Although any methods and material or equivalent to those described herein can be used in the practice or testing of the present invention, the preferred methods and materials are described. To describe the invention, certain terms are defined herein specified as follows:
Unless stated to the contrary, any of the words 'having', 'including', 'includes', 'comprising' and 'comprises' mean 'including without limitations' and shall not be construed to limit any general statement that it follows to the specific or similar items or matters immediately following it. Embodiments of the invention are not mutually exclusive, but may be implemented in various combinations. The described embodiments of the invention are given for the purpose illustration rather than limitation of the invention as set forth the appended claims.
Accordingly, the present invention provides a novel process for preparing highly pure 6-[(1-methyl-4-nitro-lH-imida2ol-5-yl)thio]-lH-purine of formula (I) commonly known as azathioprine, by reacting 6-mercaptopurine of formula (II) and 5-halo-l-methyI-4-nit.ro-lH-imidazole of formula (III)


wherein X is halogen, in presence of atleast one base in aqueous medium. Thus, the process of the present invention is short, simple and highly economical, The process of the present invention is carried out in presence of sodium hydroxide, potassium hydroxide, triethylamine or mixture thereof.
The process of the present invention is advantageously carried out by reacting 6-mercaptopurine and 5-chloro-l-methyl-4-nitro-lH-imidazole in presence of sodium hydroxide.
In another process variant, the present invention is advantageously carried out by reacting 6-mercaptopurine and 5-chloro-l-methyl-4-nitro-lH-imidazole in presence of a mixture of an inorganic and organic base. The mixture of an inorganic and organic base may be selected from combination of sodium hydroxide with triethylamine or combination of potassium hydroxide with triethylamine.
The base is conveniently used in amount, relative to 6-mercaptopurine of formula (It), preferably in a range between 1 to 5 equivalents. 7'he most preferred quantity of the base is 1 to 2 equivalents.
The process of the present invention may be carried out at suitable temperature. To minimize the decomposition of products and impurity formation the reaction is carried out at 15°C to 80°C. The most preferred reaction temperature is 25°C to 70°C.
The process of the present invention may be carried out for 0.25 to 4 hours, preferably for 0.5 to 2 hours.
The process further comprises washing the reaction mixture with water immiscible solvent followed by acidification.
The solvent used for washing the reaction mixture is dichloromethane or toluene. Washing eliminates 5-chloro-l-methyl-4-nitro-lH-imidazole (can be recovered later) and

other organic impurities leaving behind azathioprine which can be isolated by
acidification.
Azathioprine with greater than 99.90% HPLC purity is obtained using the process of
invention and hence does not require further purification (which was essential in the prior
art processes).
As azathioprine with greater than 99.90% HPLC purity is obtained in one-pot procedure using water which is the cheapest solvent, the process of the invention is highly economical.
The pure azathioprine obtained by the process of the invention may be formulated into a dosage form by combining with one or more pharmaceutically acceptable excipients using known techniques. Further the dosage form may be immediate release or extended release.
Further details of the process of the present invention will be apparent from the examples presented below. Examples presented are purely illustrative and are not limited to the particular embodiments illustrated herein but include the permutations, which are obvious as set forth in the description.
Examples:
Example: 1
Preparation of 6-[(1-methyl-4-nitro-lH-imidazol-5-yl)thio]-lH-piirine
A 2000 ml four neck round bottom flask was charged with sodium hydroxide (26 gm),
water (500 ml), 6-mercaptopurine (50 gm) and 5-chloro-1-rnethyI-4-nitro-IH-imidazole
(55 gm). The reaction mixture was stirred for 1 hour at 25 to 30°C. The reaction mixture
was washed with dichloromethane, followed by charcoal treatment and acidification to
get a solid. The solid was filtered, washed with water and dried to get 6-[(l-methyl-4-
nitro-lH-imidazol-5-yl)thio]-lH-purine.
Yield : 79 gm (86%)
HPLC Purity : 99.98%

Example: 2
Preparation of 6-[(l -methyI-4-nitro] H-imidazol-5-yl)thio]-l H-purine
A 100 ml four neck round bottom flask was charged with potassium hydroxide (0.92 gm),
triethyl amine (0.25 ml), water (30 ml) and 6-mercaptopurine (2.5 gm). The reaction
mixture was heated to 70°C for 0.5 hour. The flask was charged with 5-chloro-l-methyl-
4-nitro-lH-imidazole (2.66 gm) and the reaction mixture was maintained for 2 hours at
70°C. The reaction mixture was cooled to 10 to 15°C, washed with dichloromethane,
followed by charcoal treatment and acidification to get a solid. The solid was filtered,
washed with water and dried to get 6-[(l-methyl-4-nitro-lH-imidazol-5-yl)thio]-IH-
purine.
Yield : 4 gm (87.9%)
HPLC Purity : 99.10%

We claim,
1. A process for preparing 6-[(l-methyl-4-nitro-JH-imidazol-5-yl)thio]-lH-purine of formula (I) comprising, reacting 6-mercaptopurine of formula (II) with 5-halo-l-methyl-4-nitro-lH-imidazole of formula (III)

wherein X is halogen, in presence of at least one base in aqueous medium.
2. The process as claimed in claim }, wherein X is CI.
3. The process as claimed in claims 1, wherein the base is sodium hydroxide, potassium hydroxide, triethyl amine or mixture thereof.
4. The process as claimed in claim 3, wherein the base is used in amount, relative to 6-mercaptopurine of formula (II), in a range between 1 to 5 equivalents.
5. The process as claimed in ciaim 4, wherein the base is used in J to 2 equivalents with reference to 6-mercaptopurine of formula (II).
6. The process as claimed in claim 1 further comprising a step of washing the reaction mixture with water immiscible solvent followed by acidification.
7. The process as claimed in the claim 1, wherein the process is carried out at 10°C to 80°C.
8. The process as claimed in the claim 7, wherein the process is carried out at 25°C to 70°C.
9. The process as claimed in the claim 1, wherein the process is carried out for 0.25 to 4 hours.
10. The process as claimed in the claim 6, wherein the process is carried out for 0.5 to 2
hours.