Title of Invention	"AN IMPROVED PROCESS FOR THE PREPARATION OF HYDROXYSTIGMASTA-22-EN/ HYDROXYSTIGMASTAN 6-ONES"
Abstract	This invention relates to an improved process for the preparation of hydroxystigmasta-22-en/hydroxystigmastan 6-ones. The process steps are: taking a solution of stigmasta -2, 22-dien-6-one in a biphase solvent system such as herein described, adding to the said solution a solution of RuCl3. 3H2O and NaIO4 in water for in situ generation of RuO4, stirring the reaction mixture at room pressure, at a temperature ranging between -5 to 30°C for at least 3 minutes, quenching the reaction mixture by adding quenching agent such as herein described, continuing the for a further period of at least 5 minutes, isolating by solvent extraction stirring and purifying the compound of formula 2 by conventional methods such as herein described .

Title of Invention

"AN IMPROVED PROCESS FOR THE PREPARATION OF HYDROXYSTIGMASTA-22-EN/ HYDROXYSTIGMASTAN 6-ONES"

Abstract

This invention relates to an improved process for the preparation of hydroxystigmasta-22-en/hydroxystigmastan 6-ones. The process steps are: taking a solution of stigmasta -2, 22-dien-6-one in a biphase solvent system such as herein described, adding to the said solution a solution of RuCl3. 3H2O and NaIO4 in water for in situ generation of RuO4, stirring the reaction mixture at room pressure, at a temperature ranging between -5 to 30°C for at least 3 minutes, quenching the reaction mixture by adding quenching agent such as herein described, continuing the for a further period of at least 5 minutes, isolating by solvent extraction stirring and purifying the compound of formula 2 by conventional methods such as herein described .

Full Text	This invention relates to an improved process for the preparation of hydroxystigmasta-22-en/ hydroxystigmastan 6-ones of formula (2). This invention particularly relates to an improved process for the preparation of (2α, 3α)-2, 3-dihydroxystigmasta-22-en-6-one of formula (2a) and (2a, 3a, 22S, 23S )-2, 3, 22, 23-tetrahydroxystigmastan-6-one of formula (2b). (Formula Removed) FORMULA-2 (Formula Removed) FORMULA-2a FORMULA-lb More particularly it relates to the cis-hydroxylation of stigmasta-2, 22-dien-6-one of formula (1) using RuO4 to give dihydroxy compound of formula (2), where R is nil and dotted line indicates the double bond at C22 - C23 and tetrahydroxy compound of formula (2), where R is OH and C22 - C23 is single bond. (Formula Removed) FORMULA-1 The compound of formula (2) serves as an important intermediate for the synthesis of (22S, 23S)-homobrassinolide (3). (Formula Removed) (22S, 23S)-Homobrassinolide of formula (3) is one of the analogues of a new class of highly potent plant growth hormone. It is active in ppm level and increases the yields of variety of agricultural crops. It may be used to increase the total food grain supply of the world. The hitherto known processes for the cis dihydroxylation of stigmasta-2, 22-dien-6-one are: (a) Oxidation of the 2, 3 double bond in ring A to get the compound (2a) using catalytic amount of OsO4 and excess N-methylmorpholine-N-oxide in aqueous acetone (Ref. : K. Mori, M. Sakakibara, Y. Ichikawa, H. Ueda, K. Okada, T. Umemura, G. Yabuta, S. Kuwahara and M. Kondo, Tetrahedron 1982,38, 2099-2109). (b) Oxidation of stigmasta-2, 22-dien-6-one using tetradecyltrimethylammonium permanganate (TDTAP) in the presence of benzyltrimethylammonium hydroxide to get compound (2a) (Ref. : B. G. Hazra, T. Pavankumar and P. L. Joshi, Liebigs Annalen 1997, 1029-1034). The method (a) uses OsO4 for hydroxylation which is very costly and highly toxic. The method (b) uses TDTAP for hydroxylation of double bond in ring A only in which lot of starting material remains unreacted. Both these methods take long time for the reaction to complete. The hitherto known processes for the tetrahydroxylation of stigmasta-2, 22-dien-6- one are: (a) Single step oxidation of stigmasta-2, 22-dien-6-one using two molar equivalents of OsO4 in benzene and catalytic amount of pyridine for 90 hr (Ref.: M. J. Thompson, W. J. Meudt, N. B. Mandava, S. R. Dutky, W. R. Lusby and D. W. Spaulding, Steroids 1982,39, 89-105). (b) Oxidation of the 2, 3 double bond in ring A using catalytic amount of OsO4 and excess N-methylmorpholine-N-oxide in aqueous acetone followed by further oxidation of the 22, 23-double bond in the side chain using equivalent amount of OsO4 in pyridine for 24 hr to get tetrol of formula (2b) (Ref. : K. Mori, M. Sakakibara, Y. Ichikawa, H. Ueda, K. Okada, T. Umemura, G. Yabuta, S. Kuwahara and M. Kondo, Tetrahedron 1982,38, 2099-2109). (c) Single step oxidation of stigmasta-2, 22-dien-6-one by lengthening the reaction time for 4 days using catalytic amount of OsO4 and excess N-methylmorpholine-N-oxide in aqueous THF (Ref. : K. Mori, M. Sakakibara, Y. Ichikawa, H. Ueda, K. Okada, T. Umemura, G. Yabuta, S. Kuwahara and M. Kondo, Tetrahedron 1982, 38, 2099- 2109). Method (a) uses two molar equivalents of OsO4 and method (b) uses one molar equivalent of OsO4 for hydroxylation of 22, 23-double bond. Method (b) involves two steps for the formation of tetraol of formula (2b). All the three methods (a), (b) and (c) use OsO4 for hydroxylation which is very costly and highly toxic. All these methods take long time (1 to 4 days) for the reaction to complete. In the process of the present invention the cis-hydroxylation of stigmasta-2, 22-dien-6-one of formula (1) using the reagent RuO4 is new and has not been reported. Stigmasta-2, 22-dien-6-one of formula (1) may be prepared from naturally occurring stigmasterol in four steps namely tosylation of stigmasterol, solvolysis of the tqsylate, oxidation of the 6ß-hydroxy obtained using Jones' reagent, acid catalysed rearrangement of the 6-keto group to afford compound (1) (Ref. : K. Mori, M. Sakakibara, Y. Ichikawa, H. Ueda, K. Okada, T. Umemura, G. Yabuta, S. Kuwahara and M. Kondo, Tetrahedron 1982, 38, 2099-2109). The main objective of the present invention therefore, is to provide an improved process for the preparation of hydroxystigmasta-22-en / hydroxystigmastan 6-ones of formula (2) in high yield, using cheap and nontoxic reagents in very short period. Accordingly, the present invention provides an improved process for the preparation of hydroxystigmasta-22-en / hydroxystigmastan 6-ones of formula (2) wherein R is nil there is double bond between C22 - C23 and when R is OH there is single bond between C22 - C23 which comprises; taking a solution of stigmasta -2, 22-dien-6-one of formula (1) in a biphase solvent system such as herein described, adding to the said solution a solution of RuCl3 3H2O and NaIO4 in water for in situ generation of RuO4, stirring the reaction mixture at room pressure, at a temperature ranging between -5 to 30°C for 3 to 30 minutes, quenching the reaction mixture by adding quenching agent such as herein described, continuing the stirring for a further period of 5 minutes, isolating by solvent extraction, stirring and purifying the compound of formula 2 by conventional methods such as herein described . In one of the embodiments of the present invention, the biphase solvent system may be such as ethyl acetate, acetone, acetonitrile, and water; ethyl acetate, carbon tetra chloride, acetonitrile and water; dichloromethane, acetonitrile and water. In another embodiment of the invention the quenching agent may be such as sodium thio sulhate, sodium bisulphite, sodium metabisulphite. In yet another embodiment the organic solvent used for extracting the product may be such as diethyl ether, ethyl acetate, chloroform, dichloromethane. In yet another embodiment of the present invention the drying agent may be such as sodium sulphate, magnesium sulphate. In a feature of the invention RuO4 may be generated in situ by adding NaIO4 to RuCl3.3H2O and the molar ratio of the compound 1: RuCl3. 3H2O : NaIO4 is 1 : 0.064 : 1.5 for compound of formula (2a) and 1: 0.128 : 3 for compound of formula (2b). The column chromatography may be carried out using silica gel, alumina column and ethyl acetate-hexane, chloroform-methanol solvent system in increasing polarity. The following examples are given to illustrate the invention and should not be considered to limit the scope of the present invention. Example 1 In a 50 ml two necked round bottomed flask, a solution of stigmasta-2, 22-dien-6-one of formula (1) (0.051 g, 0.125 mmol) in ethyl acetate : acetonitrile : acetone (3.5 ml) in the ratio 2.5 : 1 : 1 was cooled to -5° C. To this, a solution of RuCl3. 3H2O (0.002 g, 0.008 mmol) and NaIO4 (0.040 g, 0.187 mmol) in water (0.3 ml) was added at -5° C. The reaction mixture was stirred at -5° C for 5 min. To the reaction mixture saturated sodium thiosulphate solution (2 ml) was added, stirred for 5 min and it was extracted with ethyl acetate (3 x 25 ml). Ethyl acetate solution was washed with water (2 x 25 ml) and brine (2 x 25 ml). Ethyl acetate extract was dried over anhydrous sodium sulphate and the compound (2cc, 3ct)-2, 3-dihydroxystigmasta-22-en-6-one of formula (2a) was isolated by removal of ethyl acetate followed by column chromatography on silica gel using methanol : chloroform as an eluent in the ratio of 0.2 : 10. Yield 0.043 g, 78 %, m.p., 233-235° C. Example 2 In a 50 ml two necked round bottomed flask, a solution of stigmasta-2, 22-dien-6-one of formula 1 (0.103 g, 0.25 mmol) in ethyl acetate : acetonitrile : acetone (7 ml) in the ratio 2.5 : 1 : 1 was cooled to -5° C. To this, a solution of RuCl3. 3H2O (0.004 g, 0.016 mmol) and NaIO4 (0.080 g, 0.375 mmol) in water (0.6 ml) was added at-5° C. The reaction mixture was stirred at -5° C for 12 min. To the reaction mixture saturated sodium bisulphite solution (4 ml) was added, stirred for 5 min and it was extracted with chloroform (3 x 25 ml). Chloroform solution was washed with water (2 x 25 ml) and brine (2 x 25 ml). Chloroform extract was dried over anhydrous magnesium sulphate and the compound (2α, 3α)-2, 3-dihydroxystigmasta-22-en-6-one of formula (2a) was isolated by removal of solvent followed by column chromatography on silica gel using methanol : chloroform as an eluent in the ratio of 0.2 : 10. Yield 0.091 g, 84 %, m.p., 233-235° C. Example 3 In a 100 ml two necked round bottomed flask, a solution of stigmasta-2, 22-dien-6-one of formula 1 (1.03 g, 2.5 mmol) in ethyl acetate : acetonitrile : acetone (35 ml) in the ratio 2.5 : 1 : 1 was cooled to 10° C. To this, a solution of RuCl3. 3H2O (0.042 g, 0.16 mmol) and NaIO4 (0.802 g, 3.75 mmol)'in water (6 ml) was added at 10° C. The reaction mixture was stirred at 10° C for 3 min. To the reaction mixture saturated sodium metabisulphite solution (12 ml) was added, stirred for 5 min and it was extracted with dichloromethane (3 x 50 ml). Organic extract was washed with water (2 x 50 ml) and brine (2 x 50 ml) and was dried over anhydrous sodium sulphate and the compound (2α, 3α)-2, 3-dihydroxystigmasta-22-en-6-one of formula (2a) was isolated by removal of solvent followed by column chromatography on silica gel using methanol: chloroform as an eluent in the ratio of 0.2 : 10. Yield 1.02 g, 92 %, m.p., 233-235° C. Example 4 In a 50 ml two necked round bottomed flask, a solution of stigmasta-2, 22-dien-6-one of formula 1 (0.051 g, 0.125 mmol) in ethyl acetate : acetonitrile : acetone (3.5 ml) in the ratio 2.5 : 1 : 1 was cooled to -5° C. To this, a solution of RuCl3. 3H2O (0.004 g, 0.016 mmol) and NaIO4 (0.080 g, 0.37 mmol) in water (0.6 ml) was added at -5° C. The reaction mixture was stirred at -5° C for 30 min. To the reaction mixture saturated sodium bisulphite solution (2 ml) was added, stirred for 5 min and it was extracted with ethyl acetate (3 x 25 ml). Ethyl acetate solution was washed with water (2 x 25 ml) and brine (2 x 25 ml). Ethyl acetate extract was dried over anhydrous sodium sulphate and the compound (2α, 3α, 22S, 23S )-2, 3, 22, 23-tetrahydroxystigmastan-6-one of formula (2b) was isolated by removal of ethyl acetate followed by column chromatography on silica gel using methanol: chloroform as an eluent in the ratio of 0.5 : 10. Yield 0.044 g, 74 %, m.p. 205-207° C. Example 5 In a 50 ml two necked round bottomed flask, a solution of stigmasta-2, 22-dien-6-one of formula 1 (0.103 g, 0.25 mmol) in etnyl acetate : acetonitrile : carbon tetra chloride (7 ml) in the ratio 2.5 : 1 : 1 was cooled to 0° C. To this, a solution of RuCl3. 3H2O (0.008 g, 0.032 mmol) and NaIO4 (0.160 g, 0.75 mmol) in water (1.2 ml) was added at 0° C. The reaction mixture was stirred at 0° C for 15 min. To the reaction mixture saturated sodium metabisulphite solution (4 ml) was added, stirred for 5 min and it was extracted with chloroform (3 x 25 ml). Chloroform solution was washed with water (2 x 25 ml) and brine (2 x 25 ml). Chloroform extract was dried over anhydrous magnesium sulphate and the compound (2cc, 3a, 22S, 23S )-2, 3, 22, 23-tetrahydroxystigmastan-6-one of formula (2b) was isolated by removal of solvent followed by column chromatography on silica gel using methanol : chloroform as an eluent in the ratio of 0.5 : 10. Yield 0.093 g, 78 %, m.p. 205-207° C. Example 6 In a 100 ml two necked round bottomed flask, a solution of stigmasta-2, 22-dien-6-one of formula 1 (1.03 g, 2.5 mmol) in dichloromethane : acetonitrile (35 ml) in the ratio 2.5 : 1 was cooled to 20° C. To this, a solution of RuCl3. 3H2O (0.084 g, 0.32 mmol) and NaIO4 (1.6 g, 7.5 mmol) in water (6 ml) was added at 20° C. The reaction mixture was stirred at 20° C for 10 min. To the reaction mixture saturated sodium thiosulphate solution (12 ml) was added, stirred for 5 min and it was extracted with dichloromethane (3 x 50 ml). Dichloromethane solution was washed with water (2 x 50 ml) and brine (2 x 50 ml). Dichloromethane extract was dried over anhydrous sodium sulphate and the compound (2α, 3α, 22S, 23S )-2, 3, 22, 23-tetrahydroxystigmastan-6-one of formula (2b) was isolated by removal of solvent followed by column chromatography on silica gel using methanol : chloroform as an eluent in the ratio of 0.5 : 10. Yield 0.951 g, 82 %, m.p. 205-207° C. Advantages The advantages of this invention are : 1. The reactions are very fast 2. Very cheap and nontoxic reagent is used for the hydroxylation 3. The yields are very high. We Claim: 1. An improved process for the preparation of hydroxystigmasta-22-en/ hydroxystigmastan 6-ones of formula (2) wherein R is nil there is double bond between C22 - C23 and when R is OH there is single bond between C22 - C23 which comprises; taking a solution of stigmasta -2, 22-dien-6-one of formula (1) in a biphase solvent system such as herein described, adding to the said solution a solution of RuCl3 3H2O and NaIO4 in water for in situ generation of RuO4, stirring the reaction mixture at room pressure, at a temperature ranging between - 5 to 30°C for 3 to 30 minutes, quenching the reaction mixture by adding quenching agent such as herein described, continuing the stirring for a further period of 5 minutes, isolating by solvent extraction, stirring and purifying the compound of formula 2 by conventional methods such as herein described . 2. An improved process as claimed in claims 1 wherein solution of stigmasta-2, 22- dien-6-one of formula 1 is made in the biphase solvent system ethyl acetate, acetone, acetonitrile and water; ethyl acetate, carbon tetra chloride, acetonitrile and water ; dichloromethane, acetonitrile and water. 3. An improved process as claimed in claims 1 to 2 wherein quenching agent used is selected from sodium thiosulphate, sodium bisulphite, sodium metabisulphite. 4. An improved process as claimed in claims 1 to 3 wherein solvent used for extraction of product during isolation from the reaction mixture is ethyl acetate, chloroform, dichloromethane, diethyl ether. 5. An improved process for the preparation of hydroxystigmasta-22-en/ hydroxystigmastan 6-ones of formula (2) substantially as herein described with reference to the examples and drawing accompanying the specification.

Full Text

This invention relates to an improved process for the preparation of hydroxystigmasta-22-en/ hydroxystigmastan 6-ones of formula (2).
This invention particularly relates to an improved process for the preparation of (2α, 3α)-2, 3-dihydroxystigmasta-22-en-6-one of formula (2a) and (2a, 3a, 22S, 23S )-2, 3, 22, 23-tetrahydroxystigmastan-6-one of formula (2b).
(Formula Removed)

FORMULA-2
(Formula Removed)
FORMULA-2a

FORMULA-lb

More particularly it relates to the cis-hydroxylation of stigmasta-2, 22-dien-6-one of formula (1) using RuO4 to give dihydroxy compound of formula (2), where R is nil and dotted line indicates the double bond at C22 - C23 and tetrahydroxy compound of formula (2), where R is OH and C22 - C23 is single bond.
(Formula Removed)
FORMULA-1
The compound of formula (2) serves as an important intermediate for the synthesis of (22S, 23S)-homobrassinolide (3).
(Formula Removed)
(22S, 23S)-Homobrassinolide of formula (3) is one of the analogues of a new class of highly potent plant growth hormone. It is active in ppm level and increases the yields of variety of agricultural crops. It may be used to increase the total food grain supply of the world.
The hitherto known processes for the cis dihydroxylation of stigmasta-2, 22-dien-6-one are:
(a) Oxidation of the 2, 3 double bond in ring A to get the compound (2a) using catalytic amount of OsO4 and excess N-methylmorpholine-N-oxide in aqueous acetone (Ref. :
K. Mori, M. Sakakibara, Y. Ichikawa, H. Ueda, K. Okada, T. Umemura, G. Yabuta, S. Kuwahara and M. Kondo, Tetrahedron 1982,38, 2099-2109).
(b) Oxidation of stigmasta-2, 22-dien-6-one using tetradecyltrimethylammonium
permanganate (TDTAP) in the presence of benzyltrimethylammonium hydroxide to
get compound (2a) (Ref. : B. G. Hazra, T. Pavankumar and P. L. Joshi, Liebigs
Annalen 1997, 1029-1034).
The method (a) uses OsO4 for hydroxylation which is very costly and highly toxic. The
method (b) uses TDTAP for hydroxylation of double bond in ring A only in which lot of
starting material remains unreacted. Both these methods take long time for the reaction to
complete.
The hitherto known processes for the tetrahydroxylation of stigmasta-2, 22-dien-6-
one are:
(a) Single step oxidation of stigmasta-2, 22-dien-6-one using two molar equivalents of
OsO4 in benzene and catalytic amount of pyridine for 90 hr (Ref.: M. J. Thompson,
W. J. Meudt, N. B. Mandava, S. R. Dutky, W. R. Lusby and D. W. Spaulding,
Steroids 1982,39, 89-105).
(b) Oxidation of the 2, 3 double bond in ring A using catalytic amount of OsO4 and
excess N-methylmorpholine-N-oxide in aqueous acetone followed by further
oxidation of the 22, 23-double bond in the side chain using equivalent amount of
OsO4 in pyridine for 24 hr to get tetrol of formula (2b) (Ref. : K. Mori, M.
Sakakibara, Y. Ichikawa, H. Ueda, K. Okada, T. Umemura, G. Yabuta, S. Kuwahara
and M. Kondo, Tetrahedron 1982,38, 2099-2109).
(c) Single step oxidation of stigmasta-2, 22-dien-6-one by lengthening the reaction time
for 4 days using catalytic amount of OsO4 and excess N-methylmorpholine-N-oxide
in aqueous THF (Ref. : K. Mori, M. Sakakibara, Y. Ichikawa, H. Ueda, K. Okada, T.
Umemura, G. Yabuta, S. Kuwahara and M. Kondo, Tetrahedron 1982, 38, 2099-
2109).
Method (a) uses two molar equivalents of OsO4 and method (b) uses one molar equivalent of OsO4 for hydroxylation of 22, 23-double bond. Method (b) involves two steps for the formation of tetraol of formula (2b). All the three methods (a), (b) and (c) use OsO4 for hydroxylation which is very costly and highly toxic. All these methods take long time (1 to 4 days) for the reaction to complete.
In the process of the present invention the cis-hydroxylation of stigmasta-2, 22-dien-6-one of formula (1) using the reagent RuO4 is new and has not been reported. Stigmasta-2, 22-dien-6-one of formula (1) may be prepared from naturally occurring stigmasterol in four steps namely tosylation of stigmasterol, solvolysis of the tqsylate, oxidation of the 6ß-hydroxy obtained using Jones' reagent, acid catalysed rearrangement of the 6-keto group to afford compound (1) (Ref. : K. Mori, M. Sakakibara, Y. Ichikawa, H. Ueda, K. Okada, T. Umemura, G. Yabuta, S. Kuwahara and M. Kondo, Tetrahedron 1982, 38, 2099-2109).
The main objective of the present invention therefore, is to provide an improved process for the preparation of hydroxystigmasta-22-en / hydroxystigmastan 6-ones of formula (2) in high yield, using cheap and nontoxic reagents in very short period.
Accordingly, the present invention provides an improved process for the preparation of hydroxystigmasta-22-en / hydroxystigmastan 6-ones of formula (2) wherein R is nil
there is double bond between C22 - C23 and when R is OH there is single bond between C22 - C23 which comprises; taking a solution of stigmasta -2, 22-dien-6-one of formula (1) in a biphase solvent system such as herein described, adding to the said solution a solution of RuCl3 3H2O and NaIO4 in water for in situ generation of RuO4, stirring the reaction mixture at room pressure, at a temperature ranging between -5 to 30°C for 3 to 30 minutes, quenching the reaction mixture by adding quenching agent such as herein described, continuing the stirring for a further period of 5 minutes, isolating by solvent extraction, stirring and purifying the compound of formula 2 by conventional methods such as herein described .
In one of the embodiments of the present invention, the biphase solvent system may be such as ethyl acetate, acetone, acetonitrile, and water; ethyl acetate, carbon tetra chloride, acetonitrile and water; dichloromethane, acetonitrile and water.
In another embodiment of the invention the quenching agent may be such as sodium thio sulhate, sodium bisulphite, sodium metabisulphite.
In yet another embodiment the organic solvent used for extracting the product may be such as diethyl ether, ethyl acetate, chloroform, dichloromethane.
In yet another embodiment of the present invention the drying agent may be such as sodium sulphate, magnesium sulphate.
In a feature of the invention RuO4 may be generated in situ by adding NaIO4 to RuCl3.3H2O and the molar ratio of the compound 1: RuCl3. 3H2O : NaIO4 is 1 : 0.064 : 1.5 for compound of formula (2a) and 1: 0.128 : 3 for compound of formula (2b). The column chromatography may be carried out using silica gel, alumina column and ethyl acetate-hexane, chloroform-methanol solvent system in increasing polarity.
The following examples are given to illustrate the invention and should not be considered to limit the scope of the present invention.
Example 1
In a 50 ml two necked round bottomed flask, a solution of stigmasta-2, 22-dien-6-one of formula (1) (0.051 g, 0.125 mmol) in ethyl acetate : acetonitrile : acetone (3.5 ml) in the ratio 2.5 : 1 : 1 was cooled to -5° C. To this, a solution of RuCl3. 3H2O (0.002 g, 0.008 mmol) and NaIO4 (0.040 g, 0.187 mmol) in water (0.3 ml) was added at -5° C. The reaction mixture was stirred at -5° C for 5 min. To the reaction mixture saturated sodium thiosulphate solution (2 ml) was added, stirred for 5 min and it was extracted with ethyl acetate (3 x 25 ml). Ethyl acetate solution was washed with water (2 x 25 ml) and brine (2 x 25 ml). Ethyl acetate extract was dried over anhydrous sodium sulphate and the compound (2cc, 3ct)-2, 3-dihydroxystigmasta-22-en-6-one of formula (2a) was isolated by removal of ethyl acetate followed by column chromatography on silica gel using methanol : chloroform as an eluent in the ratio of 0.2 : 10. Yield 0.043 g, 78 %, m.p., 233-235° C.
Example 2
In a 50 ml two necked round bottomed flask, a solution of stigmasta-2, 22-dien-6-one of formula 1 (0.103 g, 0.25 mmol) in ethyl acetate : acetonitrile : acetone (7 ml) in the ratio 2.5 : 1 : 1 was cooled to -5° C. To this, a solution of RuCl3. 3H2O (0.004 g, 0.016 mmol) and NaIO4 (0.080 g, 0.375 mmol) in water (0.6 ml) was added at-5° C. The reaction mixture was stirred at -5° C for 12 min. To the reaction mixture saturated sodium bisulphite solution (4 ml) was added, stirred for 5 min and it was extracted with chloroform (3 x 25 ml). Chloroform solution was washed with water (2 x 25 ml) and brine (2 x 25 ml). Chloroform extract was dried over anhydrous magnesium sulphate and the compound (2α, 3α)-2, 3-dihydroxystigmasta-22-en-6-one of formula (2a) was

isolated by removal of solvent followed by column chromatography on silica gel using methanol : chloroform as an eluent in the ratio of 0.2 : 10. Yield 0.091 g, 84 %, m.p., 233-235° C.
Example 3
In a 100 ml two necked round bottomed flask, a solution of stigmasta-2, 22-dien-6-one of formula 1 (1.03 g, 2.5 mmol) in ethyl acetate : acetonitrile : acetone (35 ml) in the ratio 2.5 : 1 : 1 was cooled to 10° C. To this, a solution of RuCl3. 3H2O (0.042 g, 0.16 mmol) and NaIO4 (0.802 g, 3.75 mmol)'in water (6 ml) was added at 10° C. The reaction mixture was stirred at 10° C for 3 min. To the reaction mixture saturated sodium metabisulphite solution (12 ml) was added, stirred for 5 min and it was extracted with dichloromethane (3 x 50 ml). Organic extract was washed with water (2 x 50 ml) and brine (2 x 50 ml) and was dried over anhydrous sodium sulphate and the compound (2α, 3α)-2, 3-dihydroxystigmasta-22-en-6-one of formula (2a) was isolated by removal of solvent followed by column chromatography on silica gel using methanol: chloroform as an eluent in the ratio of 0.2 : 10. Yield 1.02 g, 92 %, m.p., 233-235° C.
Example 4
In a 50 ml two necked round bottomed flask, a solution of stigmasta-2, 22-dien-6-one of formula 1 (0.051 g, 0.125 mmol) in ethyl acetate : acetonitrile : acetone (3.5 ml) in the ratio 2.5 : 1 : 1 was cooled to -5° C. To this, a solution of RuCl3. 3H2O (0.004 g, 0.016 mmol) and NaIO4 (0.080 g, 0.37 mmol) in water (0.6 ml) was added at -5° C. The reaction mixture was stirred at -5° C for 30 min. To the reaction mixture saturated sodium bisulphite solution (2 ml) was added, stirred for 5 min and it was extracted with ethyl acetate (3 x 25 ml). Ethyl acetate solution was washed with water (2 x 25 ml) and

brine (2 x 25 ml). Ethyl acetate extract was dried over anhydrous sodium sulphate and the compound (2α, 3α, 22S, 23S )-2, 3, 22, 23-tetrahydroxystigmastan-6-one of formula (2b) was isolated by removal of ethyl acetate followed by column chromatography on silica gel using methanol: chloroform as an eluent in the ratio of 0.5 : 10. Yield 0.044 g, 74 %, m.p. 205-207° C.
Example 5
In a 50 ml two necked round bottomed flask, a solution of stigmasta-2, 22-dien-6-one of formula 1 (0.103 g, 0.25 mmol) in etnyl acetate : acetonitrile : carbon tetra chloride (7 ml) in the ratio 2.5 : 1 : 1 was cooled to 0° C. To this, a solution of RuCl3. 3H2O (0.008 g, 0.032 mmol) and NaIO4 (0.160 g, 0.75 mmol) in water (1.2 ml) was added at 0° C. The reaction mixture was stirred at 0° C for 15 min. To the reaction mixture saturated sodium metabisulphite solution (4 ml) was added, stirred for 5 min and it was extracted with chloroform (3 x 25 ml). Chloroform solution was washed with water (2 x 25 ml) and brine (2 x 25 ml). Chloroform extract was dried over anhydrous magnesium sulphate and the compound (2cc, 3a, 22S, 23S )-2, 3, 22, 23-tetrahydroxystigmastan-6-one of formula (2b) was isolated by removal of solvent followed by column chromatography on silica gel using methanol : chloroform as an eluent in the ratio of 0.5 : 10. Yield 0.093 g, 78 %, m.p. 205-207° C.
Example 6
In a 100 ml two necked round bottomed flask, a solution of stigmasta-2, 22-dien-6-one of formula 1 (1.03 g, 2.5 mmol) in dichloromethane : acetonitrile (35 ml) in the ratio 2.5 : 1 was cooled to 20° C. To this, a solution of RuCl3. 3H2O (0.084 g, 0.32 mmol) and NaIO4 (1.6 g, 7.5 mmol) in water (6 ml) was added at 20° C. The reaction mixture was

stirred at 20° C for 10 min. To the reaction mixture saturated sodium thiosulphate solution (12 ml) was added, stirred for 5 min and it was extracted with dichloromethane (3 x 50 ml). Dichloromethane solution was washed with water (2 x 50 ml) and brine (2 x 50 ml). Dichloromethane extract was dried over anhydrous sodium sulphate and the compound (2α, 3α, 22S, 23S )-2, 3, 22, 23-tetrahydroxystigmastan-6-one of formula (2b) was isolated by removal of solvent followed by column chromatography on silica gel using methanol : chloroform as an eluent in the ratio of 0.5 : 10. Yield 0.951 g, 82 %, m.p. 205-207° C.
Advantages
The advantages of this invention are :
1. The reactions are very fast
2. Very cheap and nontoxic reagent is used for the hydroxylation
3. The yields are very high.

We Claim:
1. An improved process for the preparation of hydroxystigmasta-22-en/
hydroxystigmastan 6-ones of formula (2) wherein R is nil there is double bond
between C22 - C23 and when R is OH there is single bond between C22 - C23
which comprises; taking a solution of stigmasta -2, 22-dien-6-one of formula (1)
in a biphase solvent system such as herein described, adding to the said solution a
solution of RuCl3 3H2O and NaIO4 in water for in situ generation of RuO4,
stirring the reaction mixture at room pressure, at a temperature ranging between -
5 to 30°C for 3 to 30 minutes, quenching the reaction mixture by adding
quenching agent such as herein described, continuing the stirring for a further
period of 5 minutes, isolating by solvent extraction, stirring and purifying the
compound of formula 2 by conventional methods such as herein described .
2. An improved process as claimed in claims 1 wherein solution of stigmasta-2, 22-
dien-6-one of formula 1 is made in the biphase solvent system ethyl acetate,
acetone, acetonitrile and water; ethyl acetate, carbon tetra chloride, acetonitrile
and water ; dichloromethane, acetonitrile and water.
3. An improved process as claimed in claims 1 to 2 wherein quenching agent used
is selected from sodium thiosulphate, sodium bisulphite, sodium metabisulphite.
4. An improved process as claimed in claims 1 to 3 wherein solvent used for
extraction of product during isolation from the reaction mixture is ethyl acetate,
chloroform, dichloromethane, diethyl ether.
5. An improved process for the preparation of hydroxystigmasta-22-en/
hydroxystigmastan 6-ones of formula (2) substantially as herein described with
reference to the examples and drawing accompanying the specification.

Documents:

772-del-2000-abstract.pdf

772-del-2000-claims.pdf

772-del-2000-correspondence-others.pdf

772-del-2000-correspondence-po.pdf

772-del-2000-description (complete).pdf

772-del-2000-drawings.pdf

772-del-2000-form-1.pdf

772-del-2000-form-19.pdf

772-del-2000-form-2.pdf

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Patent Number

226260

Indian Patent Application Number

772/DEL/2000

PG Journal Number

01/2009

Publication Date

02-Jan-2009

Grant Date

16-Dec-2008

Date of Filing

29-Aug-2000

Name of Patentee

COUNCIL OF SCIENTIFIC & INDUSTRIAL RESEARCH

Applicant Address

RAFI MARG, NEW DELHI-110 001, INDIA.

Inventors:

#	Inventor's Name	Inventor's Address
1	BRAJA GOPAL HAZRA	NATIONAL CHEMICAL LABORATORY, PUNE 411 008, MAHARASHTRA, INDIA.
2	ARCHNA PATIENCE MASSEY	NATIONAL CHEMICAL LABORATORY, PUNE 411 008, MAHARASHTRA, INDIA.
3	VANDANA SUDHIR PORE	NATIONAL CHEMICAL LABORATORY, PUNE 411 008, MAHARASHTRA, INDIA.

PCT International Classification Number

A61K 31/00

PCT International Application Number

N/A

PCT International Filing date

PCT Conventions:

#	PCT Application Number	Date of Convention	Priority Country
1			NA