FORM 2THE PATENTS ACT, 1970 (39 of 1970) &The Patents Rules, 2003COMPLETE SPECIFICATION(See section 10; rule 13)1. Title of the invention. - AN IMPROVED PROCESS FOR THE PREPARATION OF L-2'-ANHYDRO-l-(B-L- ARABINO FURANOSYL)THYMINE2. Applicant(s)(a) NAME : ALEMBIC LIMITED(b) NATIONALITY: An Indian Company.(c) ADDRESS : Alembic Campus, Alembic Road, Vadodara - 390 003, Gujarat, India.3. PREAMBLE TO THE DESCRIPTIONThe following specification particularly describes the invention and the manner in which it is to be performed:

Field of the invention:

The present invention relates to an improved process for preparation of L-2,2'-anhydro-1-(P-L-arabino furanosyl)thymine represented by a structural formula (I) as given below.

The present invention also relates to a novel polymorphic form AL of L-2,2'-anhydro-1-(P-L-arabino furanosyl)thymine.
Background of the invention:
The molecular formula of L-2,2'-anhydro-1-(P-L-arabino furanosyl)thymine is C11H16N2O4 and molecular weight is 240.25. CAS registry No. of L-2,2'-anhydro-1-(P-L-arabino furanosyl)thymine is [433733-92-7].
L-2,2'-anhydro-1-(P-L-arabino furanosyl)thymine is used as an intermediate in the various API synthetic preparation such as L-nucleic acid derivatives useful as a medicine as well as to synthesize intermediates therefore. It is useful in nucleoside or nucleotide preparation.
J. Org. Chem., Vol 41, No. 10, 1976, p 1830 discloses a process for preparation. The synthetic reaction scheme for the preparation is as given below in the scheme.

In this process D-arabinose is converted to 2-amino-b-D-arabinofurano[1',2'4,5]-2-oxazoline i.e D-arabinoaminooxazoline using neat cyanamide in the presence of DMF and sodium bicarbonate. The storage of neat cyanamide is very difficult as it is prone to degrade and it requires special storage condition.

US7125983 discloses a process for preparation of L-2,2'-anhydro-1-(P-L-arabino furanosyl)thymine which is as given below in the scheme.

In the above process, ethyl-a-hydroxymethyl acrylate(ll) is reacted with thionyl chloride to give ethyl-a-chloromethyl acrylate(lll) which is further reacted with L-arabinoaminooxazoline (IV) to give L-arabinoaminooxazoline-a-chloromethyl acrylic acid ester adduct (VI). Compound (VI) is reacted with hydroquinone in the presence of water and sodium carbonate. The reaction mixture is neutralized with acetic acid to give aqueous solution of L-2,2'-anhydro-5,6-dihydrouridine-5-exomethylene(VII) which is not isolated and directly hydrogenated to give L-2,2'-anhydro-1-(|3-L-arabinofuranosyl)thymine(l). The compound (I) is isolated by evaporation of solvent after removal of hydrogenation catalyst and purification by column chromatography on silicagel. This process is commercially not feasible because it requires chromatographic purification. Sodium carbonate is neutralized with acetic acid and sodium acetate salt is formed which is carried forward in the hydrogenation step. The product(l) as well as the salt both are soluble in the water. Therefore the isolation of the product (I) is very difficult.
Hence, there is a need to develop a process which not only overcomes disadvantages of the prior art but also provide a process which is economical, operationally simple and industrially applicable.

The present inventors have directed their research work towards developing a process which overcomes the problems associated with existing art. They developed an improved process in which aqueous cyanamide is used instead of neat cyanamide in the preparation of L-arabinoaminooxazoline. This significantly reduced the problems associated with the storage of neat cyanamide. They also provided a solution to difficulty associated with the separation of the salt from the product at an industrial scale, i.e without using chromatographic purification. They modified the workup process with change of solvent which gave separation of salt from final product. The process also gives novel polymorphic form AL of L-2,2'-anhydro-1-(P-L-arabino furanosyl)thymine.
Object of the invention:
The primary object of the present invention is to provide an improved process for the preparation of L-2,2'-anhydro-1-((3-L-arabino furanosyl)thymine (I)
Another object of the present invention is to provide an improved process for the preparation of L-2,2'-anhydro-1-((3-L-arabino furanosyl)thymine having purity of at least 99%.
Another object of the present invention is to provide a process feasible at an industrial scale.
A further object of the present invention is to provide a process for recovery of the product from the mother liquor obtained after first crop isolation.
Yet another object of the present invention is to provide a novel polymorphic form AL of L-2,2'-anhydro-1 -(P-L-arabino furanosyl)thymine.
Summary of the invention:
Accordingly, the present invention provides a process for preparation of L-2,2'-anhydro-1-(P-L-arabino furanosyl)thymine (I)

HO n

(b) acidifying the filtrate obtained in step (a) with dilute acid till pH is about 6 to about 6.5 is obtained;
(c) removing solvent to its one third of the original volume and added acetone to it;
(d) removing salt from the reaction mixture obtained in step (c) by filtration ;
(e) removing solvent from the filtrate to give solid product
The present invention provides a process for preparation of L-arabinoaminooxazoline (IV)

with 50% aqueous cyanamide in the presence of potassium bicarbonate and dimethylformamide.
The present invention also provides a novel polymorphic form AL of L-2,2'-anhydro-1-(P-L-arabino furanosyl)thymine.
A process for recovery of second crop of L-2,2'-anhydro-1-((3-L-
arabinofuranosyl)thymine(l) comprising steps of
(x) concentraing the mother liquor to 2/3 of original volume obtained after first crop
isolation;
(y) adding antisolvent to the concentrated mother liquor obtained in step (x);
(z) isolating the solid as second crop.
Brief description of the drawings
FIG. 1 shows a powder X-ray diffraction (PXRD) diffractogram of AL form of L-2,2'-anhydro-1 -(P-L-arabino furanosyl)thymine.

FIG. 2 shows DSC thermogram of AL form of L-2,2'-anhydro-1-(p-L-arabino furanosyl)thymine.
Detailed description of the invention:
The present invention provides a process for preparation of L-2,2'-anhydro-1-((3-L-arabino furanosyl)thymine (I). The synthetic reaction scheme of the present invention is as shown below.

In one embodiment, the process for preparation of L-2,2'-anhydro-1-(P-L-arabino furanosyl)thymine (I) is given. Ethyl a-hydroxymethyl acrylate is reacted with thionyl chloride at about 85°C to about 95°C for 2 to 3hr. The excess thionyl chloride is distilled out from the reaction mixture and degassed the trace amount of thionyl chloride from the product. The product ethyl-a-chloromethyl acrylate (III) is used as such in the next reaction step.
L-Arabinoamino oxazoline (IV) is suspended in dimethylacetamide (DMAc). Ethyl-a-chloromethylacrylate (III) is added dropwise to the above suspension under ice-cooling. The reaction mixture is stirred at room temperature for 4hr. The progress of the reaction can be monitored on TLC. Ethylacetate is added to the reaction mixture and stirred at room temperature for 1hr. The resulting crystals are colleted by filteration, washed with ethyl acetate, and dried under vacuum to give L-arabinoaminooxazolin-a-chloro methylacrylic acidester adduct (VI).

A mixture of L-arabinoaminooxazolin-a-chloro methylacrylic acidester adduct (VI), p-methoxy phenol, DM water and base such as potassium carbonate is stirred at about 2°C till completion of the reaction. The base is organic or inorganic base. The examples of organic base includes but not limited to pyridine, triethylamine, dimethyl aniline, DBU and the like or mixture thereof. The examples of inorganic base includes but not limited to NaOH, KOH, Ca(OH)2, Ba(OH)2, Na2C03, K2C03, NaHC03, KHCO3, sodium acetate, potassium acetate, sodium methoxide, potassium tert butoxide and the like or mixture thereof. The product formed L-2,2'-anhydro-5,6-dihydrouridine-5-exomethylene (VII) in an aqueous solution is used as such as for next step. The aq. Solution is hydrogenated in the presence of hydrogenation catalyst such as 5% Pd/alumina at about 60°C to about 70°C at atmospheric pressure for 8 to 10hr. The hydrogenation is carried out in the presence of metal catalyst such as Pd, Pt, Rh, Ru or raney Nickel. The metal catalyst can be in the form of oxide, chloride or in any complex form. It can be supported on active carbon, alumina or silica. The reaction can be carried out in the presence of inert atmosphere or in hydrogen atmosphere. Inert gas such as nitrogen or argon can be used alone or in the combination with hydrogen gas. The progress of the reaction can be monitored on TLC or HPLC. The hydrogenation catalyst is removed from the reaction mixture by filtration on hyflow bed. The filtrate is acidified with dilute acid such as dilute hydrochloric acid till pH at about 6 to about 6.5 is obtained. The acid used here can be inorganic acid or organic acid. The examples of inorganic acid includes but not limited to hydrochloric acid, sulfonic acid, phosphoric acid and the like or mixture thereof. The examples of organic acid includes but not limited to formic acid, acetic acid, methane sulfonic acid, p-toluene sulfonic acid, acidic resin and the like or mixture thereof. The solvent is distilled out up to one third of the original volume under vacuum at below 60°C. Solvent such as acetone is added to it and stirred. Other solvent such as ethanol, methanol, propanol, isopropanol, ethylacetate, dichloromethane can also be used in the place of acetone. Potassium chloride salt is separated out, which is filtered off. The filtrate is distilled out completely to give solid.
In another embodiment, the present invention provides novel polymorphic of L-2,2'-anhydro-1-(b-L-arabino furanosyl)thymine (I), designated as form AL, characterized by an x-ray powder diffraction spectrum having peaks expressed as 29 at about 11.0, 12.6, 12.9, 15.8, 18.1, 19.2, 21.1, 22.1, 24.9, 26.0, 27.6 degrees. FIG. 1 shows typical form AL x-ray powder diffraction spectrum.

DSC thermogram of AL form of L-2,2'-anhydro-1-(b-L-arabino furanosyl)thymine is as shown in FIG. 2.
In accordance with the present invention, a process is provided for preparation of AL form of L-2,2'-anhydro-1-((3-L-arabino furanosyl)thymine (I), which comprises crystallizing the crude solid L-2,2'-anhydro-1-(b-L-arabino furanosyl)thymine (I) from ethanol.
In another embodiment, the present invention provides a process for recovery of the product from the mother liquor obtained after first crop isolation. The crude L-2,2'-anhydro-1-(b-L-arabino furanosyl)thymine (I) is crystallized from ethanol. The solid crystalline product is filtered to give solid as first crop. The mother liquor obtained after the filtration is concentrated and antisolvent is added to it. The suspension is stirred and filtered to give the second crop of the L-2,2'-anhydro-1-((3-L-arabino furanosyl)thymine (I). The examples of the antisolvent includes but not limited to ethylacetate, acetone, dichloromethane, diethylether, isopropyl alcohol, acetonitrile and the like or mixture thereof.
In a further embodiment, the present invention provides a process for preparation of L-arabinoaminooxazoline (IV) as shown in the scheme below.

A mixture of L-arabinose, 50% aqueous cyanamide, potassium carbonate and DMF is stirred at about 80°C to about 100°C for time sufficient for the reaction to take place. The reaction mixture is cooled to about 25°C to about 30°C. Solvent such as ethyl acetate is added over the period of time 30min. The suspension is stirred for 30min at about 25°C and then 1 to 2hr at about 0°C. The crystalline solid is filtered, washed with mixture of ethyl acetate: DMF(1:1), ethyl acetate and then suck dried. The solid is dried at 50°C to 55°C under vacuum to give L-Arabinoamino oxazoline(IV) as off-white crystalline solid.

The following examples illustrate the invention further. It should be understood, however, that the invention is not confined to the specific limitations set forth in the individual example but rather to the scope of the appended claims.
Example-1
Preparation of ethyl-a-chloromethyl acrylate (III)
Ethyl a-hydroxymethyl acrylate (60.0g) was reacted with thionyl chloride (63.62g) at
90°C for 2hr. The excess thionyl was distilled out from the reaction mixture and
degassed the trace amount of thionyl chloride from the product. The product ethyl a-
chloromethyl acrylate (III) (62.Og) is used as such in the next step.
Yield: 90%
Purity: 89.51% (by GC)
Example-2
Preparation of L-Arabinoamino oxazoline (IV)
A mixture of (200.0g) L-arabinose, 50% aqueous cyanamide(137.84g), potassium
carbonate(7.98g) and DMF(1.2L) was stirred at 90°C for 2hr. The reaction mixture was
cooled to 30°C. Ethyl acetate(800ml) was added over 30min. The suspension was
stirred for 30 min at 25°C and 1hr at 0°C. The crystalline solid was filtered .washed with
mixture of ethyl acetate:DMF(1:1) (200mlx2), ethyl acetate (300 ml) and dried at 50°C
under vacuum to give L-Arabinoamino oxazoline(IV) (122g,) as off-white crystalline solid.
Yield: 53%
Purity: 99.66% by HPLC.
1HNMR (300MHz, DMSO-d6, 6): 3.19-3.31(m,2H), 3.63(m,1H), 4.0(s,1H), 4.51-
4.53(d,1H), 4.73(br,1H), 5.43(br,1H), 5.57-5.66(d,1H), 6.33(br,1H).
Mass Spectra: 175.3(M+1)
Example-3
Preparation of L-arabinoaminooxazolin-a-chloro methylacrylic acidester adduct
(VI)
L-Arabinoamino oxazoline (IV) (150.0g) was suspended in dimethylacetamide (800ml). ethyl-a-chloromethylacrylate(152.92g) was added dropwise to the above suspension under ice-cooling. The reaction mixture was stirred at room temperature for 4hr. The progress of the reaction was monitored on TLC. Ethylacetate(1.0L) was added to the

reaction mixture and stirred at room temperature for 1hr. The resulting crystals were
colleted by filteration, washed with ethyl acetate, and dried under vacuum to give L-
arabinoaminooxazolin-a-chloro methylacrylic acidester adduct (VI) (95.0g).
Yield: 38%.
Purity: 95.98% (by HPLC)
1HNMR (300MHz, DMSO-d6, 5): 1.15(t,3H), 3.19-3.33(m,2H), 3.97(s,1H), 4.045-
4.42(m,6H), 5.12(s,1H), 5.20-5.77(d,1H), 5.77-5.79(d,1H), 5.99-6.032(m,1H), 6.21(s,1H),
9.73(br,2H).
Mass Spectra: 287.4(M+1)
Example-4
Preparation of L-2,2'-anhydro-1-(P-L-arabino furanosyl)thymine (I)
A mixture of L-arabinoaminooxazolin-a-chloro methylacrylic acidester adduct (VI) (55.0g), p-methoxy phenol(0.298g), DM water(150ml), Potassium carbonate (35.34g) was stirred at 2°C till completion of reaction checked on TLC or HPLC. The product formed L-2,2'-anhydro-5,6-dihydrouridine-5-exomethylene (VII) in an aqueous solution was used as such as for next step.
An aqueous solution of L-2,2'-anhydro-5,6-dihydrouridine-5-exomethylene (VII) was
hydrogenated in the presence of 5% pd/alumina(3.75g) in water (100ml) at 65°C at
atmospheric pressure for 8-10hr. The reaction progress was monitored on TLC or HPLC.
The hydrogenation catalyst is removed from the reaction mixture by filtration on hyflow
bed. The filtrate was acidified with dilute hydrochloric acid till pH 6 to 6.5 is obtained. The
solvent was distilled out up to its 60 % concentration of original volume under vacuum at
below 60°C. Acetone (500ml) was added to it. Potassium chloride was separated out,
which was filtered off. The filtrate was distilled out completely. The solid was crystallized
from ethanol to give final product L-2,2'-anhydro-1-((3-L-arabino furanosyl)thymine (I)
(14.0g)
Yield: 30%
Purity: 99.55% (by HPLC)
1HNMR (300MHz, DMSO-d6, 6): 1.79(s,3H), 3.12-3.29(m,2H), 4.04(m,1H), 4.37(br,1H),
4.97(t,1H), 5.18(d,1H), 5.87(d,1H),6.28(d,1H).
Mass spectra: 241.4 (M+1)
SOR: +48.7181° (1% in water)

Example-5
Preparation of AL form of L-2,2'-anhydro-1-(b-L-arabino furanosyl)thymine (I)
L-2,2'-anhydro-1-(b-L-arabino furanosyl)thymine (I) (10.0g) was dissolved in ethanol
(100.0ml) at 55°C till the solution becomes clear. If the solution is not clear, filter off the
solid and take the clear solution. The filtrate is cooled to 0°C to 5°C. The solid was
precipitated out. The solid was filtered, suck dried and then dried in oven at 45 to 55°C.
AL form of L-2,2'-anhydro-1-(b-L-arabino furanosyl)thymine (I) (4.0g) is obtained.
Purity: 99.55% (by HPLC)
1HNMR (300MHz, DMSO-d6, 5): 1.79(s,3H), 3.12-3.29(m,2H), 4.04(m,1H), 4.37(br,1H),
4.97(t,1H), 5.18(d,1H), 5.87(d,1H),6.28(d,1H).
Mass spectra: 241.4 (M+1)
SOR: +48.7181° (1% in water)
PXRD (expressed at 26) at about: 11.0, 12.6, 12.9, 15.8, 18.1, 19.2,21.1,22.1,24.9,
26.0, 27.6 degrees
Example-6
Recovery of second crop
L-2,2'-anhydro-1-(B-L-arabino furanosyl)thymine (I) (10.0g) was dissolved in ethanol (100.0ml) at 55°C till the solution becomes clear. If the solution is not clear, filter off the solid and take the clear solution. The filtrate is cooled to 0°C to 5°C. The solid was precipitated out. The solid was filtered. The mother liquor (100ml) obtained after filtration was concentrated to 66ml (2/3 of the original filtrate). Ethyl acetate (1.0L) was added to it and stirred for 30 min. The isolated product is filtered to give second crop (2.0g).

We claim:

1. A process for preparation of L-2,2'-anhydro-1 -(b-L-arabino furanosyl)thymine (I)

comprising steps of:
(i) reacting ethyl-a-hydroxymethyl acrylate(II) with thionyl chloride to give ethyl-a-chloromethyl acrylate(III);

(ii) reacting ethyl-a-chloromethyl acrylate(III) obtained in step (i) with L-arabinoaminooxazoline (IV) in the presence of dimethylacetamide to give L-arabinoaminooxazoline-a-chloromethyl acrylic acid ester adduct (VI);

(iii) reacting L-arabinoaminooxazoline-a-chloromethyl acrylic acid ester adduct (VI) with p-methoxy phenol in the presence of water and base to give aqueous solution of L-2,2'-anhydro-5,6-dihydrouridine-5-exomethylene(VII)

(iv) aqueous solution of L-2,2'-anhydro-5,6-dihydrouridine-5-exomethylene(VII) is hydrogenated in the presence of metal catalyst to give L-2,2'-anhydro-l-(p-L-arabinofuranosyl)thymine(I).

2. A process for preparation of L-2,2'-anhydro-l-(b-L-arabino furanosyl)thymine (I)

comprising steps of :
(iii) reacting L-arabinoaminooxazoline-a-chloromethyl acrylic acid ester
adduct (VI) with p-methoxy phenol in the presence of water and base to give aqueous solution of L-2,2'-anhydro-5,6-dihydrouridine-5-exomethylene(VII)

(iv) aqueous solution of L-2,2'-anhydro-5,6-dihydrouridine-5-
exomethylene(VII) is hydrogenated in the presence of metal catalyst to give L-2,2'-anhydro-1 -(b-L-arabinofuranosyl)thymine(I).
2. The process as claimed in claim 1 or 2, wherein the base is selected from the group comprising inorganic base such as NaOH, KOH, Ca(0H)2, Ba(0H)2, Na2C03, K2CO3, NaHCO3, KHCO3, sodium acetate, potassium acetate, sodium methoxide, potassium tert butoxide or mixture thereof.
3. The process as claimed in claim lor 2, wherein the base is selected from the group comprising organic base such as pyridine, triethylamine, dimethyl aniline, DBU or mixture thereof.
4. The process as claimed in claim lor 2, wherein the metal catalyst is selected from the group comprising Pd, Pt, Rh, Ru or raney Nickel.
5. The process as claimed in claim 1 or 2, wherein the metal catalyst is Pd on alumina.

6. The present invention further provides a process for isolation of L-2,2'-anhydro-
l-(b-L-arabinofuranosyl)thymine(I) from the reaction mixture obtained after
hydrogenation comprising steps of:
(a) removing of the hydrogenation catalyst from the reaction mixture by filtration;
(b) acidifying the filtrate obtained in step (a) with dilute acid till pH is about 6 to about 6.5 is obtained;
(c) removing solvent to its one third of the original volume and added second solvent to it;
(d) removing salt from the reaction mixture obtained in step (c) by filtration ;
(e) removing solvent from the filtrate to give solid product.

7. The process as claimed in claim 6, wherein the acid is selected from organic acid such as formic acid, acetic acid, methane sulfonic acid, p-toluene sulfonic acid or inorganic acid such as hydrochloric acid, sulfonic acid, phosphoric acid or acidic resin.
8. The process as claimed in claim 6, wherein the second solvent is selected from acetone, ethanol, methanol, propanol, isopropanol, ethylacetate, dichloromethane or mixture thereof.
9. A process for preparation of L-arabinoaminooxazoline (IV)

HO
comprising reacting L-arabinose(V)

with 50% aqueous cyanamide in the presence of potassium bicarbonate and dimethylformamide.

10. A crystalline L-2,2'-anhydro-l-(b-L-arabinofuranosyl)thymine(I) AL form, characterized by an x-ray powder diffraction spectrum having peaks expressed as 29 at about 11.0, 12.6, 12.9, 15.8,18.1,19.2, 21.1, 22.1,24.9, 26.0, 27.6 degrees.
11. A crystalline L-2,2'-anhydro-l-(b-L-arabinofuranosyl)thymine(I) AL form, further characterized by an x-ray powder diffraction spectrum as in FIG. 1.
12. A crystalline L-2,2'-anhydro-l-(b-L-arabinofuranosyl)thymine(I) AL form, further characterized by DSC thermogram as in FIG. 2.
13. A process for preparation of L-2,2'-anhydro-l-(b-L-arabinofuranosyl)thymine(I) AL form as defined in claim 10, which comprises a step of crystallizing crude solid L-2,2'-anhydro-l-(b-L-arabino furanosyl)thymine (I) from ethanol.
14. A process for recovery of second crop of L-2,2'-anhydro-l-(b-L-arabinofuranosyl)thymine(I) comprising steps of
(x) concentraing the mother liquor to 2/3 of original volume obtained after first crop
isolation;
(y) adding antisolvent to the concentrated mother liquor obtained in step (x);
(z) isolating the solid as second crop.

TITLE: AN IMPROVED PROCESS FOR THE PREPARATION OF L-2,2'-
ANHYDRO-1 -(B-L-ARABINO FURANOSYL)THYMINE
A B S T R A C T
The present invention relates to an improved process for preparation of L-2,2'-anhydro-1-(0-L-arabino furanosyl)thymine (I).

The present invention further provides an isolation procedure for of L-2,2'-anhydro-1-(b-L-arabino furanosyl)thymine (I) from the reaction mixture. The present invention also provides novel polymorphic form AL of L-2,2'-anhydro-1-(b-L-arabino furanosyl)thymine
(I).