FIELD OF THE INVENTION

The present invention relates to an improved process for the preparation of Tribenoside of formula (I). More particularly the present invention relates to the preparation of Tribenoside having purity greater than 98% by HPLC and its precursors namely compound of formula (II) and (III) in pure form.

BACKGROUND OF THE INVENTION

Tribenoside (I) is chemically known as ethyl 3,5,6-tris-0-(phenylmethyl)-D-glucofuranoside, which is a mixture of a- and B-anomers. Tribenoside (I) has been used in inflammatory and varicose disorders of the veins, including haemorrhoids. Tribenoside (I) is also used as sclerosing agent.

US 3,157,634 disjclose Tribenoside (I) and its pharmaceutical^ acceptable salts thereof. US '634 also .disclose a process for the preparation of Tribenoside (I). According to the process disclosed in US '634, Tribenoside is prepared by reacting 1,2-0- isopropylidene-a-D-glucofuranose (II) with benzyl chloride in presence of potassium hydroxide to produce
3,5,6-tri-0-benzyl-l,2-0-(l-methylethylidene-a-D-glucofuranose (Ill), which is further reacted with ethanolic hydrochloric acid. The process is as shown in Scheme-I given below: Helvetica Chimica Acta, Vol 51, Issue 6 Pages 1185-1202; 1968 reported a process for the preparation of Tribenoside (I). This process comprises reacting 1,2-0-isopropylidene-a-D-glucofuranose (II) with benzyl chloride in presence of potassium hydroxide to produce 3,5,6-tri-0-benzyl-l,2-0-(l methylethylidene-a-D-glucofuranose (III), which is further reacted with ethanol in the presence of proton donor to produce Tribenoside (I). The process is as shown in Scheme-II given below:

The reported prior art processes involve the use of impure compound of formula (II) for the preparation of compound of formula (III), which results in compound of formula (III) contaminated with undesired side products. These impurities in turn are carried forward in subsequent reaction step and results in Tribenoside (I) contaminated with the undesired impurities. Removal of these impurities in the final stage is very difficult and requires repeated crystallizations, fractional distillation which results in the low yield of Tribenoside (I) and hence is not suitable for industrial scale operations.

GB 1310382 discloses a process for the preparation of Tribenoside by reacting 1,2-cyclohexylidene-D-glucofuranose (IV) with benzyl chloride in presence of potassium hydroxide to produce 3,5,6-tri benzyl- 1,2-cyclohexylidene-D-glucofuranose (V), which is further reacted with ethanol in the presence of an acid to produce Tribenoside (I).

These processes also require repeated purification, using crystallization, fractional distillation of Tribenoside. The process is as shown in Scheme-Ill given below:

Considering the importance of Tribenoside (I) in pharmaceutical field, there is a need to have simple, easy to handle, industrially viable and cost effective process for the preparation of Tribenoside (I) with high purity i.e greater than 98.00%, preferably greater than 98.50%, more preferably greater than 99.00% by HPLC and with high yield which avoids the use of fractional distillation.

The present invention is directed towards a process for the preparation of pure Tribenoside (I), wherein 1,2-O-isopropylidene-a-D-glucofuranose (II) and 3,5,6-tri-O-benzyl-l,2-0-(l-methylethylidene-ot-D-glucofuranose (III) intermediates are purified and used in the preparation of Tribenoside. The present invention is also directed towards a process for the preparation of pure Tribenoside (I), wherein Tribenoside (I) is purified by using simple silica column chromatography.

OBJECTIVE OF INVENTION

The main objective of the present invention is to provide a simple and cost-effective process for the preparation of Tribenoside (I) with high purity on commercial scale.

Another objective of the present invention is to provide a simple and cost-effective process for purification of 1,2-O-isopropylidene-a-D-glucofuranose (II) and 3,5,6-tri-O-benzyl-l,2-0-(l-methylethylidene-a-D-glucofuranose (III).

SUMMARY OF THE INVENTION

In one aspect of the present invention provides a process for the purification of 1,2-O-isopropylidene-a-D-glucofuranose (II),
which comprises purifying the compound of formula (II) with nitrile solvent, aromatic hydrocarbon solvent or mixtures thereof.

Yet another aspect of the present invention provides a process for the preparation of Tribenoside (I) having purity greater than 98% by HPLC; which comprises the steps of; (i) optionally treating Tribenoside (I) with a polar aprotic solvent; (ii) optionally washing the step (i) mass with a hydrocarbon solvent (i); (iii) purifying Tribenoside (I) using column chromatography using an eluent; (iv) distilling the eluent followed by stripping out the residual solvent using water; and (v) isolating pure Tribenoside (I).

In another aspect of the present invention provides a process for the purification of 3,5,6-tri-O-benzyl-1,2-0-( 1 -methylethylidene)-a-D-glucofuranose (III); which comprises purifying 3,5,6-tri-0-benzyl-l,2-0-(l-methylethylidene)-a-D-glucofuranose (III) by high vacuum distillation and/or column chromatography.

DETAILED DESCRIPTION OF THE INVENTION

In an embodiment of the present invention provides a process for the purification of 1,2-O-isopropylidene-a-D-glucofuranose (II),
which comprises purifying the compound of formula (II) with nitrile solvent, aromatic hydrocarbon solvent or mixtures thereof.

In another embodiment of the present invention, the starting material 1,2-0-isopropylidene-a-D-glucofuranose (II) is prepared by conventional methods or by hydrolysis of 1,2,5,6-di-O-isopropylidene-a-D-glucofuranose (Ha) with acetic acid.

In still another embodiment of the present invention, 1,2-0-isopropylidene-a-D-glucofuranose (II) is purified by using a solvent comprises from nitrile solvent, aromatic hydrocarbon solvent or mixture thereof. The purification step employs conventional techniques such as re-crystallization, stirring the compound in the solvent, suspending the compound, washing etc, followed by optional isolation of pure 1,2-0-isopropylidene-ot-D-glucofuranose (II) by filtration.

In another embodiment of the present invention, nitrile solvent comprises from acetonitrile, propionitrile, butyronitrle, valeronitrile or mixture thereof; aromatic hydrocarbon solvent comprises from benzene, toluene, ethylbenzene, m-xylene, o-xyiene, p-xylene, chlorobenzene, flurobenzene, trifluorotoluene, or mixture thereof. The purification step is carried out at a temperature ranging between about 0°C to about 80°C for a period of about 1 to 24 hrs.

In yet another embodiment, the present invention also relates to the use of 1,2-0-isopropylidene-oc-D-glucofuranose (II), prepared by the present invention in the preparation of Tribenoside (I).

In one more embodiment of the present invention, relates to a purification of 3,5,6-tri-0-benzyl-l,2-0-(l-methylethylidene)-a-D-glucofuranose (III); which comprises purifying 3,5,6-tri-0-benzyl-l,2-0-(l-methylethylidene)-a-D glucofuranose (III) by high vacuum distillation and/or column chromatography.

Accordingly 3,5,6-tri-0-benzyl-l,2-0-(l-methylethylidene)-a-D-glucofuranose (III) is prepared by reacting 1,2-0-isopropylidene-a-D-glucofuranose (II) with benzyl halide in presence of sodium hydroxide and dimethylsulfoxide (DMSO). The prepared compound of formula (III) is optionally subjected to purification using high vacuum distillation and/or column chromatography. Though the purification of compound of formula (III) is not required, for better purity, any one of the purification or both is optionally employed.

In yet another embodiment of the present invention, 3,5,6-tri-0-benzyl-l,2-0-(l-methylethylidene)-a-D-glucofiiranose (III) is purified by high vacuum distillation to remove less boiling impurities. Said high vacuum distillation is carried out at a temperature ranging between about at 0.5-1.0 mm/Hg at 200-250°C, preferably 220°C/0.5 mm/Hg.

In still another embodiment of the present invention, 3,5,6-tri-0-benzyl-l,2-0-(l-methylethylidene)-a-D-glucofuranose (III) is purified using column chromatography such as preparative HPLC by using C|g reverse phase silica gel or column chromatography on silica gel (hereinafter may be referred to as "silica gel column") or column chromatography on vinyl alcohol copolymer and an eluent to produce pure 3,5,6-tri-0-benzyl-l,2-0-(l-methylethylidene)-a-D-glucofuranose (III). Preferably use of silica gel column makes the process commercially viable.

In still another embodiment of the present invention, eluent is selected from a solvent comprises from ethyl acetate, methyl acetate, dichloromethane, acetonitrile, tetrahydrofuran, acetone, dimethyl formamide, dimethyl sulfoxide, n-pentane, isopentane, neopentane, n-hexane, cyclohexane, isohexane, 3-methyIpentane, neohexane, n-heptane, isoheptane, neoheptane, neooctane, water or mixture thereof. Preferably the mixture of ethyl acetate and cyclohexane is used as an eluent.

In another embodiment, the present invention also relates to the use of 3,5,6-tri-O-benzyl-l,2-0-(l-methylethylidene)-a-D-glucofuranose (III), prepared by the present invention in the preparation of Tribenoside (I).

In another embodiment, the present invention provides a purification process of Tribenoside (I); which comprises; (i) optionally treating Tribenoside (I) with a solvent, (ii) optionally washing with a hydrocarbon solvent obtained in step (i), (iii) purifying Tribenoside (I) by column chromatography using an eluent; (iv) distilling the eluent followed by stripping residual solvent using water; and (v) isolating pure Tribenoside (I).

In another embodiment of the present invention, Tribenoside of formula (I) is obtained by following the process described in prior art or by reacting 3,5,6-tri-O-benzyl-l,2-0-(l-methylethylidene)-a-D-glucofuranose (III) with hydrochloric acid in presence of ethanol to produce Tribenoside (I).

In another embodiment the present invention, Tribenoside (I) is optionally dissolved with a polar aprotic solvent followed by said solution is washed with hydrocarbon solvent. This purification is optionally carried out to remove hydrocarbon soluble impurities. A polar aprotic solvent comprises from N,N-dirnethylformarnide (DMF), N,N-dimethyl acetamide, N-methyl pyrrolidine (NMP), formamide, acetamide, propnamide, hexamethyl phospharamide and hexamethyl phosphorous triamide or mixture thereof.

In another embodiment the present invention, hydrocarbon solvent used in in step (ii) comprises from n-pentane, isopentane, neopentane, n-hexane, isohexane, 3-methylpentane, neohexane, n-heptane, isoheptane, neoheptane, neooctane or mixture thereof.

In still another embodiment of the present invention, the Tribenoside (I) is purified using coloumn chromatography such as preparative HPLC by using Cis reverse phase silica gel or column chromatography on silica gel (hereinafter may be referred to as "silica gel column") or column chromatography on vinyl alcohol copolymer and an eluent to produce pure Tribenoside (I). Preferably use of silica gel column makes the process commercially viable.

In still another embodiment of the present invention, eluent is selected from a solvent comprises from ethyl acetate, methyl acetate, dichloromethane, acetonitrile, tetrahydrofuran, acetone, dimethyl formamide, dimethyl sulfoxide, n-pentane, isopentane, neopentane, n-hexane, cyclohexane, isohexane, 3-methylpentane, neohexane, n-heptane, isoheptane, neoheptane, neooctane, water or mixture thereof.

In one more embodiment of the present invention the desired fractions of eluents are collected and distilled the eleuents to get residue or oily mass. The residual solvent in residue is stripped out using water to remove the volatile impurities in the residue along with residual solvent to yield Tribenoside having HPLC chromatographic purity greater than 98.00%, preferably greater than 98.50%, more preferably greater than 99.00%. Tribenoside being a liquid presence of solvent like methanoi/propanol/i-propanol, acetone as a contamination in a eluent resulted with methyl/propyl/isopropyl Tribenoside and benzalacetone respectively. The column purification using silica gel and the process of stripping out with water helps to remove these impurities as well as to remove other volatile impurities apart from removal of residual solvents thereby able to achieve high pure Tribenoside.

The following examples illustrate the nature of the invention and are provided for illustrative purposes only and should not be construed to limit the scope of the invention.

EXAMPLES

Example-l: Process for the preparation of Tribenoside (I).

Step 1: Preparation of 3,5,6-tri-0-benzyl-l,2-0-(l-methylethylidene)-a-D- glucofuranose (III):
1,2-O-isopropylidene-a-D-glucofuranose (II; 100 g) is dissolved in dimethylsulfoxide (500 mL) in a round bottom flask under nitrogen atmosphere at 25-30°C. 1st lot of sodium hydroxide powder (48 g) was added to the reaction mixture at 25-30°C and stirred for 1 hour at 25-30°C. 1st lot of benzyl chloride (143 g) was added slowly at 25-35°C and stirred for 1 hour at 25-35°C. 2nd lot of Sodium hydroxide powder (38 g) was added at 25-35°C. The reaction mixture was cooled to 10-15°C. 2nd lot of benzyl chloride (64 g) was slowly added at 10-15°C and the reaction mass was maintained at 10-15°C for 1 hour. The reaction mass was allowed to warm to 20-25°C and maintained at 20-25°C. The reaction mass was again allowed to warm to 65-70°C and maintained for 2 hours at 65-70°C. The reaction mass was cooled to 25-30°C. Cylclohexane (500 mL) was added to the reaction mass and stirred for 10-15 min and then DM water (1000 mL) was added and stirred for 15 min. Organic layer was separated and the aqueous layer was extracted with cyclohexane (200 mL). Combined the organic layer was washed with DM water (3 X 300 mL) and evaporated completely under reduce pressure at 30-450°C to afford yellow-brownish liquid material.
Purity by HPLC: Not Less than 80% Yield: 98%
Purification of 3,5,6-tri-0-benzyI-l,2-0-(l-methyIethyIidene)-a-D-gIucofuranose (III):
S.S^-tri-O-benzyi-l^-O-O-methylethylidene^a-D-gfucofuranose crude (75 g) was subjected to column chromatography over silica gel (100-200 mesh, 413 g) using cylohexane and ethyl acetate mixture as eluent. The fractions having desired purity were combined and the solvent was concentrated under reduced pressure at temperature below 40°-45°C to produce a light yellow viscous liquid. Purity by HPLC: 99% Yield%: 75±5%

Step-2: Preparation of Ethyl 3,5,6-tris-O-benzyl-D-gIucofuranoside (Tribenoside (I)):
3,5,6-tri-0-benzyl-l,2-0-(l-methylethylidene)-a-D-glucofuranose (III) is dissolved in ethanol (4000 mL) in a round bottom flask at 25-30°C under nitrogen atmosphere. The reaction mixture is cooled to less than 20°C and hydrogen chloride solution in ethanol (212.85 g) was added to the reaction mixture and stirred at 25-30°C till completion of the reaction. The reaction mixture was quenched with 10% aqueous sodium hydroxide solution (82 g of sodium hydroxide in 750 mL of DM water). The solvent from the reaction mixture is evaporated under reduced pressure at 40°C. The organic residue was diluted with DM water (300 mL), extracted with cyclohexane (1 X 300 mL) and stirred for 15 min at 25-30°C. Layers were separated and the aqueous layer is extracted with cyclohexane (300 mL). Organic layers were combined and washed with DM-water (3 X 200 mL) and evaporated under reduced pressure at below 45°C to afford Tribenoside concentrated mass. Purity by HPLC: Not less than 98.5% Yield: 98% Purification of Tribenoside (I): Tribenoside concentrated mass (300 mL) was subjected to column chromatography over silica gel (100-200 mesh, 400 g) using cyclohexane and ethyl acetate mixture as eluent. The fractions having desired purity were combined and the solvent was concentrated under reduced pressure at temperature below 40°C. Water (50 mL) was added to the concentrated mass and distilled out under reduced pressure at 35-45°C to produce the desired compound as light yellow, viscous liquid. Purity by HPLC: Not less than 98.5% Yield: 88%

Example-2: Process for the Preparation of Tribenoside (I).
Step 1: Preparation of 1,2-O-isopropyIidene-a-D-gIucofuranose (II): 1,2,5,6-di-O-isopropylidene-a-D-glucofuranose (Ha; 150 g) and 66% aqueous acetic acid (900 mL) were charged into a round bottom flask at 25-30°C and maintained till completion of reaction. Aqueous acetic acid from the reaction mixture was evaporated under reduced pressure at 40-45°C. Organic residue was chassed with water, followed by toluene (300 mL) to remove traces of acetic acid and then water, to afford 126 g of the title compound as a solid. Purification of 1,2-O-isopropyIidene-a-D-gIucofuranose (II): 1,2-O-isopropylidene-a-D-glucofuranose crude (II) was suspended in acetonitrile (600mL) at 25-35°C. The reaction mixture was stirred for 1 hour at 25-30°C. The reaction mass was cooled to 10-15°C and maintained for 1 hr at 10-15°C. The reaction mass was filtered and washed with 100 ml acetonitrile. The obtained solid was dried at 40-45°C under reduced pressure for 6-8 hours to afford 115 g of the title compound. Purity by HPLC: 98.6% Yield: 80%

Step 2: Preparation of l,2-0-isoproylidene-3,5,6-tri-0-benzyl-a-D-glucofuranose (III):
Dimethylsulfoxide (300 mL) and 1st lot of sodium hydroxide powder (24 g) were charged into a round bottom flask under nitrogen atmosphere at 25-30°C and stirred for 15-20 minutes at 25-30°C. 1,2-O-isopropylidene-a-D-glucofuranose (II; 50 g) was added to the reaction mixture at 25-30°C and maintained for 1 hour at 25-30°C. 1st lot of benzyl chloride (71.66 g) was added slowly for about 1 hour at 25-35°C and stirred for 1 hour at 25-35°C. 2nd lot of sodium hydroxide powder (19 g) was added at 30-35°C. The reaction mixture was cooled to 10-15°C. 2nd lot of benzyl chloride (31.16 g) was slowly added for about 40-45 minutes at 10-15°C and the reaction mass was maintained at 10-15°C for 1 hour. The reaction mass was allowed to warm to 20-25°C and maintained at 20-25°C. The reaction mass was again allowed to warm to 65-70°C and maintained at 65-70°C. The reaction mass was cooled to 30-35°C. Toluene (500 mL) was added to the reaction mass and stirred for 10-15 min and then DM water (600 mL) was added and stirred for 15 min. Organic layer was separated and the aqueous layer was extracted with toluene (250 mL). Combined the organic layer and the organic layer was washed with DM water (5 X 600 mL) and brine solution (1 X 300 mL). The solvent from the organic layer was evaporated completely under reduce pressure at 45-50°C to afford pale yellowish oily mass. Purity by HPLC: 71.67%
Purification of l,2-0-isoproylidene-3,5,6-tri-0-benzyl-a-D-glucofuranose (III):
l,2-0-isopropylidene-3,5,6-tri-0-benzyl-a-D-glucofuranose crude (oily mass; 123 g) was subjected to high vacuum distillation under reduced pressure (0.5-1.0 mm Hg) at ~220°C to remove the impurities from the product. Purity by HPLC: 91.03% Yield: 90%

Step-3: Preparation of Ethyl 3,5,6-tris-0-(phenylmethyI)-D-glucofuranoside (Tribenoside) (Formula (I)): l,2-Isopropylidene-3,5,6-tri-0-benzyl-a-D-glucofuranose (90 g) and 1.0N hydrochloric acid in ethanol solution (900 mL) were charged in to round bottom flask at 25-30°C and stirred till completion of reaction. The reaction mixture was cooled to 0-10°C. The reaction mixture was quenched with 15% aq. sodium hydroxide solution (450 mL). The solvent from the reaction mixture is evaporated under reduced pressure at 45-50°C. The organic residue was diluted with DM water (1350 mL), extracted with dichloromethane (1 X 1350 mL; 1 X 450 mL). Organic layers were combined and washed with DM-water (3 X 540 mL), followed by brine solution (1 X 450 mL) and evaporated under reduced pressure at below 40°C to afford brown color oily mass. Purity by HPLC: 90.74%

First purification of Tribenoside (I):
Tribenoside crude (oily mass; 84 g) was dissolved in iV.JV-dimethylformamide (DMF) (168 mL) and washed with n-heptane (5 X 336 mL) to remove the impurities at 25-30°C. The DMF layer was diluted with demineralization water (DM water) (420 mL) and ethyl acetate (588 mL) and stirred for 5-10 minutes. Both layers were separated. The aqueous layer was extracted with ethyl acetate (252 mL). The organic layers were combined and washed with DM-water (3 X 504 mL). Activated carbon (2.52 g) was added to the organic layer and stirred for 30 min at 25-30°C. The reaction mixture was filtered through hyflow bed and washed with ethyl acetate (2 X 42 mL). Filtrates were combined and the solvent and evaporated under reduced pressure at 45-50°C to afford 71.7 g of pale yellowish oily mass. Purity by HPLC: 94.14% Second purification of Tribenoside (I): Tribenoside crude (oily mass; 38 g) was subjected to column chromatography over silica gel using ethyl acetate and hexane mixture as eluent. The fractions having desired purity were combined and the solvent was concentrated under reduced pressure at temperature below 40°C. Water (38 mL) was added to the concentrated mass and distilled out under reduced pressure at 45-50°C. Thereafter toluene (78 mL) was added and the solvents were distilled out. Further, 50 mL water was added and distilled out under reduced pressure at temperature below 50°C to produce the desired compound. Purity by HPLC: 98.38% Yield%: 29 g (76%)

Advantages: The present invention avoids the fractional distillation and yield high pure Tribenoside by employing simple silga gel column which makes the process commercially viable.

WE CLAIM:

1. A process for preparation of Tribenoside (I) having purity greater than 98.00 % by HPLC
which comprises the steps of;

(i) optionally treating Tribenoside (I) with a polar aprotic solvent;

(ii) optionally washing the step (i) mass with a hydrocarbon solvent (i);

(iii) purifying Tribenoside (I) using column chromatography using an eluent;

(iv) distilling the eluent followed by stripping out the residual solvent using water; and

(v) isolating pure Tribenoside (I).
The process as claimed in claim 1, wherein polar aprotic solvent used in step (i) is selected from N,N-dimethylformamide (DMF), N,N-dimethyl acetamide, N-methyl pyrrolidine (NMP), formamide, acetamide, propnamide, hexamethyl phospharamide and hexamethyl phosphorous triamide and hydrocarbon solvent used in step (ii) is selected from n-pentane, isopentane, neopentane, n-hexane, cyclohexane, isohexane, 3-methylpentane, neohexane, n-heptane, isoheptane, neoheptane, neooctane.

A process for the purification of 3,5,6-tri-0-benzyl-l,2-0-(l-methylethylidene)-a-D-glucofuranose (III), which comprises purifying 3,5,6-tri-0-benzyl-l,2-0-(l-methyIethyIidene)-a-D-glucofuranose (III) by high vacuum distillation or column chromatography.

4. A process for the purification of 1,2-O-isopropylidene-a-D-glucofuranose (II),
which comprises purifying the compound of formula (II) with nitrile solvent, aromatic hydrocarbon solvent or mixtures thereof.

The process as claimed in claim 1, wherein the Tribenoside (I) is obtained by converting the pure compound of formula (II) prepared according to claim 4 into 3,5,6-tri-0-benzyl-l,2-0-(l-methylethylidene)-a-D-glucofuranose (III), subjecting the compound of formula (III) purification according to claim 3 followed by treating pure compound of formula (III) into Tribenoside.

The process according to claim 4, wherein nitrile solvent comprises from acetonitrile, propionitrile, butyronitrle, valeronitrile; aromatic hydrocarbon solvent comprises from benzene, toluene, ethylbenzene, m-xylene, o-xylene, p-xylene, chlorobenzene, flurobenzene, trifluorotoluene, or mixtures thereof.

The process as claimed in claim 3 and 4, further comprising converting the compound of formula (II) or (III) into Tribenoside (I).

The process according to proceeding claims, wherein the column chromatography comprises column chromatography on silica gel, column chromatography on vinyl alcohol copolymer, or preparative HPLC with a Cjg reverse phase silica gel and an eluent.

A process according to claim 1 and 3, eluent used in the chromatography comprises a solvent selected from ethyl acetate, methyl acetate, dichloromethane, dichloroethane, acetonitrile, tetrahydrofuran, acetone, dimethyl formamide, dimethyl sulfoxide, n-pentane, isopentane, neopentane, n-hexane, cyclohexane, isohexane, 3-methylpentane, neohexane, n-heptane, isoheptane, neoheptane, neooctane, water or mixtures thereof.

A process as described herein with specification and examples.