Title of Invention

"PROCESS FOR PREPARING OPTICALLY PURE (S)-3, 4-DIHYDROXYBUTYRIC ACID DERIVATIVES"

Abstract The present invention provides a process to prepare optically pure (S)-3, 4-dihydroxy butyric acid derivatives from commercially available amylopectin with ease. The process also enables preparing optically pure (S)-3, 4-dihydroxy butyric acid derivatives economically in large quantities and also minimizes the formation of bi-product.
Full Text PROCESS FOR PREPARING OPTICALLY PURE
(S)-3,4-DIHYDROXYBUTYRIC ACID DERIVATIVES
BACKGROUND OF THE INVENTION Field of the Invention
The present invention relates to a process for preparing optically pure (S)-3,4-dihydroxybutyric acid derivatives expressed by the following Formula 1 and more particularly, to a process that enables preparing optically pure (S)-3,4-dihydroxybutyric acid derivatives economically in large quantities, by:
(a) Preparing α-(l,4)-linked oligosaccharide with adequate sugar
distribution by reacting amylopectin which is easily available from
natural product with enzyme under a specific condition; and
(b) Performing oxidation and esterification sequentially under a specific
condition.

(Formula Removed) Wherein, R represents linear or branched alkyl group with 1~5 carbon atoms.
Description of the Related Arts
(S)-3,4-Dihydroxy butyric acid derivatives and
(S)-3-hydroxy-y-butyrolactone are used as synthetic intermediates for preparing various chiral compounds. For example, it is well known that they act as key intermediates for preparing neuromeidator (R)-GABOB [Tetraliedron, 46, 4277(1990)], treatment for hyperlipemia (Atorvastatin; HMG-CoA reductase inhibitor) [Tetrahedron Lett., 33, 2279(1992)], (S)-oxiracetam which is improvement agent in brain metabolism [International patent publication WO93/06,826], L-carnitine which is health supplement agent [International

patent publication WO99/05,092], (S)-3-hydroxytetrahydrofuran [J. Am. Ctem. Soc., 117,1181(1995); International patent publication WO94/05,639] which is an essential intermediate of AIDS drug (Agenerase; HIV protease inhibitor), (S)-mono-betalactam [Japanese patent publication 64-13,069(1989)], ester of (S)-3-hydroxy-4-bromobutyric acid [Japanese patent publication 4-149,151(1992); Japanese patent publication 6-172,256(1994)], potentiating intermediate of satiety agent [Bull. Cliem. Soc. Jpn., 61, 2025(1988)] and neuroleptic drug [USP 4,138,484] and useful intermediates in synthetic efforts towards natural products [J. Org. Chem., 50, 1144 (1985); Can. ]. Chem., 65, 195 (1987), Tetrahedron Lett., 507 (1992)]. Optical purity is the most important factor in preparing these chiral compounds.
The conventional technologies for preparing (S)-3,4-dihydroxybutyric acid derivatives and (S)-3-hydroxy-y-butyrolactone, which are useful for preparing the said chiral compounds, are explained in detail hereunder.
Methods of preparing (S)-3-hydroxybutyric acid derivatives from the enzymatic or catalytic reduction of p-ketoester were known [J. Am. Cliem. Soc., 105, 5925-5926(1983); Teteralvdron Lett., 31, 267-270(1990); European patent publication 452,143A2]. These methods have difficulty in that the prochiral center should be reduced to one-side to generate chiral center and expensive metal catalyst should be used.
A technology preparing ester of (S)-3,4-dihydroxybutyric acid and (S)-3-hydroxy-y-butyrolactone by selective reduction of (L)-malic acid ester was known [Chem. Lett., 1389-1392(1984); USP 5,808,107]. This technology has disadvantage in that reduction should be performed selectively to only one of the two ester functional groups.
Many methods of preparing (S)-3,4-dihvdroxybutyric acid derivatives and (S)-3-hydroxy-y-buryrolactone from carbohydrate have been reported.
A technology preparing isosaccharinic acid (B) or

(S)-3,4-dihydroxybutyric acid (C) is reported [J. Qiem. Soc., 1924-1931(1960)] by
alkaline degradation of carbohydrate containing glucose substituent in the
4-position, such as 4-O-methyI-(D)-glucose, maltose, amylose and cellulose,
elimination of C-4 substituent as leaving group, forming dicarbonyl compound
(A; 4-deoxy-2,3-hexodiulose), and reacting the formed dicarbonyl compound
with base as shown in Scheme 1. However, the yield of (S)-3,4-dihydroxybutyric acid is low. Scheme 1
(Scheme Removed)
Also, it has been reported that (S)-3,4-dihydroxybutyric acid (C) and
glycolic acid (D) were obtained as major products by forming dicarbonyl
compound (A) from alkaline degradation of carbohydrate containing glucose
substituent in the 4-position, and separating the formed dicarbonyl compound
(A) and reacting it with hydrogen peroxide [J. Chem. Soc., 1932-1938(1960)].
This method has a serious problem that the product exists as small amount of
isomers due to tautomerization and a mixture of cyclic compounds and
hydrates derived from dicarbonyl compound (A). So, the dicarbonyl
compound (A) cannot be separated in good yields from the reaction mixture. Another problem is that the prepared (S)-3,4-dihydroxybutyric acid is degraded
to formic acid and glycolic acid due to the overoxidation.
A similar technology for preparing (S)-3,4-dihydroxybutyric acid from
carbohydrate either using base only or using oxygen in base was known. It
proposed that the dicarbonyl compound (A) was a synthetic intermediate for
(S)-3,4-dihydroxybutyric acid as shown in the Scheme 1. But the yield was
reported to be as low as about 30% [J. Res. Natl. Bur. Stand., 32, 45(1944); /. Am. Chem. Soc., 2245-2247(1953); /. Am. Chem. Soc., 1431-1435(1955); Carbohyd. Res., 11, 17-25(1969); /. Chromatography, 549, 113-125(1991)]. In these methods, (S)-3,4-dihydroxybutyric acid is produced with various kinds of mixtures including glycolic acid (D), isosaccharinic acid (B), formic acid, ketone, diketone and glyceric acid. Since the yield of (S)-3,4-dihydroxybutyric acid is very low, these methods are also considered as not suitable for industrial use.
A method for preparing (S)-3,4-dihydroxybutyric acid from
disaccharide (lactose) using base and oxidant has been reported [International
patent publication WO98/04543]. In this work, (S)-3,4-dihydroxybutyric
acid was cyclized to (S)-3-hydroxy-y-butyrolactone under the reaction condition and purified by protection of the two hydroxy groups to acetonide ester compound, methyl (S)-3,4-0-isopropylidene-3,4-dihydroxybutanoate, which was recyclized to (S)-3-hydroxy-y-butyrolactone under acidic media.
Preparing methods of (5)-3,4-dihydroxybutyric acid including the
process of alkaline oxidation of carbohydrate containing glucose substituent in
the 4-position have been known [USP 5,292,939, 5,319,110 & 5,374,773(1994)].
In these methods, dicarbonyl compound (A) intermediate is formed at first,
oxidized to (S)-3,4-dihydroxybutyric acid (C) and glycolyic acid (D).
However, optical purity, the most important physical property of chiral
compounds, is not mentioned at all. Also, purification of target.compound
is very difficult, considering the reaction mechanism. In the case of
disaccharides such as maltose or lactose, only one sugnr unit in the disaccharide
forms (S)-3,4-dihydroxybutyric acid and the other sugar unit functions as
leaving group, so that the target product and leaving group coexist as 1:1
mixture. Accordingly, it is very difficult to separate and purify
(S)-3,4-dihydroxybutyric acid or (S)-3-hydroxy-y-butyrolactone from the
reaction mixture. The theoretical mass conversion obtainable is 28.3 wt%.
In other words, 28.3g of (S)-3-hydroxy-y-butyrolactone can be obtained from
lOOg of disaccaride. For polysaccharides, such as maltodextrin, starch and
cellulose, mentioned in the above patents, the (1,4) and/or (1,6) glucose units
are linked complexly like nets. The problem is that the step-by-step
oxidation proceeding from the reducing end units comprising (1,4) linkage
terminates at (1,6) linkage unit. Therefore, no more target product is
formed. Also, the polysaccharides are degraded by overoxidation of
reducing end units to complex acid mixtures containing formic acid, oxalic acid, glycolic acid and erythronic acid [J. Am. Chem. Soc., 81, 3136(1959); Starch 41 Nr. 8, S. 303-309(1989); Synthesis, 597-613(1997)].
There was an attempt to improve the yield of (S)-3,4-dihydroxybutyric
acid or (S)-3-hydroxy-y-butyrolactone for polysaccharide by degradation of
higher-molecular sugars to relatively lower-molecular sugars through acid or
base hydrolysis. Though the reactivity by this method is increased to a
degree, (1,4) linkage and (1,6) linkage are not hydrolyzed selectively to afford
random distribution. Accordingly, there is a fundamental problem in
preparing (S)-3,4-dihydroxybutyric acid derivatives in high yield [Encyclopedia of Chemical Technology, 3rd ed. 492-507].
Regarding the preparation of (S)-3-hydroxy-y-butyrolactone using (1,4)
linked polysaccharide, the. step-by-step oxidation proceeds continuously from
the reducing end units to non-reducing end units to afford
(S)-3,4-dihydroxybutyric acid until the last chain unit (a leaving group)
remained. Namely, if (l,4)-linked polysaccharide is used as a source
material for preparing (S)-3-hydroxy-y-butyrolactone, the theoretical mass
conversion obtainable is 63 wt%, about two times more compared with the
method using disaccharide. In other words, 63g of
(S)-3-hydroxy-y-butyrolactone can be obtained from 100g of (l,4)-linked polysaccharide. Also, since the small amount of leaving group is produced in the reaction mixture compared with disaccharide, the target product is easily purified. Therefore, the use of (l,4)-linked polysaccharide promises the enhanced productivity.
However, regarding conventional polysaccharides, the target product and by-products (acids such as formic acid, oxalic acid, glycolic acid and erythronic acid) are formed competitively in the step-by-step oxidation due to the compact structure having random (1,4) linkage and (1,6) linkage. Thus, selective degradation technique of polysaccharide to a suitable sugar distribution range having (1,4) linkage is required.
On the other hand, there have been many reports of transforming higher-molecular sugars to lower-molecular sugars using biological enzymatic treatment process for industrial use.
The reported technologies include preparing glucose, maltose and
ethanol through enzymatic treatment of starch [USP 3,791,865(1974); USP
3,922,200(1975); USP 4,855,232(1989): Japanese patent publication
4-158,795(1992); Methods Carbohydr. Chem., 10, 231-239(1994); Methods Carbohydr.
Chem., 10, 245-248(1994)], and preparing maltodextrin with adequate dextrose
equivalent (DE) [USP 3,986,890(1976); USP 4,447,532(1984); USP 4,612,284(1986);
USP 5,506,353(1996)]. In these references, through the degradation or
transformation of high molecular polysaccarides, they are converted to adequate materials for medicines, food additives and diagnostic reagents.
But, the method for preparing a-(l,4) linked oligosaccharides with suitable sugar distribution for the production of (S)-3,4-dihydroxybutyric acid
derivatives is not known at present.
SUMMARY OF THE INVENTION
The inventors of the present invention made intensive efforts to develop
a method for preparing optically pure (S)-3,4-dihydroxy butyric acid derivatives
from commercially available amylopectin with ease. As a result, a process
which enables preparing optically pure (S)-3,4-dihydroxybutyric acid
derivatives economically in large quantities is found by preparing
oligosaccharide with structural specificity which can minimize formation of
by-products from amylopectin by enzymatic reaction. Furthermore,
oxidation reaction can be performed continuously in the same reactor without additional separation and purification of the above prepared oligosaccharide.
Accordingly, an object of this invention is to provide a method for preparing optically pure (S)-3,4-dihydroxybutyric acid derivatives in high yield without additional purification of intermediates.
Brief Description of the Drawings
Fig. la represents the optical purity analysis results by gas chromatography (GC) of racemic 3-hydroxy-y-butyrolactone.
Fig. 1b represents the optical purity analysis results by gas chromatography (GC) of 3-hydroxy-y-butyrolactone prepared from disaccharide of the conventional method.
Fig. 1c represents the optical purity analysis results by gas chromatography (GC) of 3-hydroxy-y-butyrolactone prepared from oligosaccharide of the present invention.
Detailed Description of the Invention
The present invention is characterized by comprising the following
(a) Enzymatic reaction of amylopectin to a-(l,4) linked oligosaccharide
expressed by the Formula 2; and
(b) Oxidation of the oligosaccharide with oxidant in base, and
subsequent esterification with alcohol in the presence of an acid
catalyst to afford ester of (S)-3,4-dihydroxybutyric acid derivatives
expressed by the Formula I.
(Formula Removed)
, wherein R represents linear or branched alkyl group with 1~Scarbons.
The detailed description of the present invention is given hereunder.
The fundamental inventive concept of the present invention is selective
degradation of α-(l,4) linkage and α-(l,6) linkage within amylopectin using
specific enzymes, i.e., transforming amylopectin to α-(l,4)-inked
oligosaccharide with the optimal sugar distribution for preparing the target
compound. And subsequently oxidation and esterification are performed
to prepare (S)-3,4-dihydroxybutyric acid derivatives.
Namely, focusing on the specificity of enzymes, amylopectin is degraded sequentially with specific enzymes to a-(l,4) linked oligosaccharide,
and optically pure (S)-3,4-dihydroxybutyric acid derivatives are prepared from
the transformed oligosaccharide in high yield. Optical purity of the
desired product prepared by a sequential method is over 99.9%ee.
Oligosaccharide used in the present invention is prepared with
biological enzymatic treatment of amylopectin, and amylopectin is
commercially available with ease. Especially, since amylopectin is highly
soluble in water or in buffer solution of pH 4.0~8.0, used as reaction solvent for
enzymatic reaction of the present invention, the relative reactivity to enzyme is
greatly increased compared with other polysaccharides such as starch and
cellulose. Thus, the same is very effective material for preparing of
oligosaccharide having suitable sugar distribution for the preparation of (S)-3,4-dihydroxybutyric acid derivatives.
When using pullulanase as an enzyme for selective degradation of
and reduced enzyme activity. So, rather than using pullulanase only,
a-amylase was used to improve reactivity in degrading amylopectin to a
suitable sugar distribution, and then pullulanase was used. However, in
this case, activity of the remaining a-amylase persists and thus amylopectin is degraded excessively, so that the desired oligosaccharide is not formed. Accordingly, a technology of inactivating the remaining a-amylase before the pullulanase reaction was introduced.
The detailed explanation of the preparation process of this invention is
as follow. It comprises; 1) a step preparing oligosaccharide with
characteristic a-(l,4) linkage expressed in Formula 2 by selective degradation of amylopectin using biological treatment of specific enzymes, and 2) a step preparing optically pure (S)-3,4-dihydroxy butyric acid derivatives by esterifying (S)-3,4-dihydroxybutyric acid salt formed through oxidation. Especially, the preparation process of this invention is characterized by
preparing (S)-3,4-dihydroxybutyric acid derivatives in the same reactor without additional purification of the intermediates (oligosaccharide and (S)-3,4-dihydroxybutyric acid).
The enzymatic reaction of the present invention uses a-amylase and pullulanase sequentially. α-Amylase degrades ct-(l,4) linkage and pullulanase degrades α-(l,6) linkage selectively.
The superiority of the present invention lies in that optically pure (S)-3,4-dihydroxybutyric acid derivatives are prepared in high yield under a mild reaction condition by using enzymes selectively degrading a-(l,4) linkage or a-(l,6) linkage, while the chemical hydrolysis method has no selectivity.
The enzymatic reaction of the present invention is performed in water
or buffer solution of pH 4.0-8.0 at 40-120°C. α-Amylase is used in the
range of 0.001-10 wt% of amylopectin, and enzymatic reaction of a-amylase is
performed for 30 min - 4 hr, and then remaining a-amylase is inactivated.
Inacrivation is performed under acidic (pH 2.0-4.5) and high temperature
(60-150 °C) conditions and maintained for 10 min - 4 hr. In the enzymatic
reaction of pullulanase, pullulanase is used in the range of 0.001-10 wt% of amylopectin, and most of the oligosaccharides distribute within 3-50 glucose units by 10-40 hr of the pullulanase enzymatic treatment. Reducing end units and molecular weight distribution of the prepared oligosaccharide are analyzed from reducing end units and dextrose equivalent analysis by an optical analyzer, HPLC analysis, and gel permeation chromatography (GPC) analysis.
The oligosaccharide is obtained from the selective enzymatic reaction
and has distribution mostly between 3-50 glucose units, and preferably 5-50
glucose units. Since most of the glucose units are linked with a-(l,4)
linkage, (S)-3,4-dihydroxybutyric acid derivatives can be obtained in high yield through continuous sequential reactions with minimizing the by-products (e.g.,
acid mixtures of formic acid, oxalic acid, glycolic acid and erythronic acid). Furthermore, the obtained (S)-3,4-dihydroxybutyric acid derivatives were identified to be optically very pure (>99.9%ee).
Oxidation of oligosaccharide is performed by adding base and oxidant dropwise for 6-36 hr under the condition of 30-65°C. Hydrogen peroxide, alkali metal peroxides, akaline earth metal peroxides and alkyl hydroperoxides are used for the oxiciants, and hydrogen peroxide is the most preferable. The oxidant is used in the range of 1-3 equivalents per molar glucose unit of amylopectin. The base is selected from the group consisting of alkali metal hydroxide or alkaline earth metal hydroxide, and sodium hydroxide or potassium hydroxide is preferable. The base is used in the range of 2-4 equivalents per molar glucose unit of amylopectin.
Esterification of the present invention is performed in the presence of
acid catalyst using alcohol as both a reaction solvent and reagent in the range of
30-80 °C. Inorganic acids such as hydrochloric acid, sulfuric acid,
phosphoric acid and nitric acid, or organic acids such as fluoroalkylsulfonic acid, aralkylsulfonic acid, hydrate of aralkylsulfonic acid and trifluoroacetic acid are used as acid catalyst. Linear or branched alcohol with 1-5 carbon atoms is used for the alcohol.
In order to compare the preparation yields depending on the source
material of oxidation, the prepared (S)-3,4-dihydroxybutyric acid derivatives
were cyclized (S)-S-hydroxy-y-butyrolactone as follows [Refer to Experimental
example 1.]. (S)-3,4-dihydroxybutyric acid derivatives were cyclized in the.
presence of acid catalyst by agitating the same for 2-5 hr in the range of
30-80 "C to obtain (S)-3-hydroxy-y-butyrolactone. Inorganic acids such as
hydrochloric acid, sulfuric acid, phosphoric acid and nitric acid, or organic acids such as fluoroalkylsulfonic acid, aralkylsulfonic acid, hydrate of aralkylsulfonic acid and trifluoroacetic acid are used as acid catalyst. As a
result, if maltose (disaccharide) or lactose (disaccharide) obtained from cheese by-product is used as source material, the theoretical mass conversion yield of (S)-3-hydroxy-Y-butyrolactone is not more than 28.3 wt% of the source material weight used. On the other hand, if amylose (α-(l,4) linked polysaccharide) with more than 50 glucose units is used, the theoretical mass conversion yield of (S)-3-hydrbxy-y-butyrolactone is equal to that of amylopectin. But, the double helix structure due to very strong intramolecular hydrogen bond limits the step-by-step oxidation, so the yield becomes very low. However, by using oligosaccharide of the present invention is used as source material, the yield of (S)-3-hydroxy-y-butyrolactone is very high as 57.2 wt% of the source material weight used.
As explained above, the present invention is excellent in that the low
reactivity of amylopectin to oxidation is overcome by transforming amylopectin
to oligosaccharide with the application of specific enzymes. Furthermore,
by-product formation is minimized and optically pure (S)-3,4-dihydroxy butyric acid derivatives can be prepared in high yield with very simple purification process.
The following examples are intended to be illustrative of the present invention and should not be construed as limiting the scope of this invention defined by the appended claims.
Example 1: Preparation of methyl (S)-3,4-dihydroxybutanoate
j 10 L of water and 5 kg of dried amylopectin were put into a 50 L reactor.
After heating the reactor to 55 °C, 12 g of a-amylase (BAN; EC 3.2.1.1 from
Bacillus lidteniformis, Novo Nordisk) was added. After heating this reaction
solution to 750C, the same was stirred for 2 hr at the same temperature. 5 mL of 0.1N HC1 solution was added to adjust the pH 3.0~3.5, and then the same was stirred for 1 hr at 900C to inactivate the remaining a-amylase.
After slowly cooling the reaction mixture to 300C, 3.7L of 4M acetic acid buffer solution (pH 5) and 1.3L of water were added to adjust the pH to 5. The reaction solution was heated to 60°C, and then 62.5g of pullulanase (Promozyme; EC 3.2.1.4 from Bacillus acidopullulyticus, Novo Nordisk) was added and the solution was stirred for 22 hr at the same temperature. 0.54kg of 40% NaOH solution was added to the reaction solution to neutralize acetic acid and the temperature was raised to 60°C. 40% NaOH (8.64 kg) solution and 30% H2O2 (5.25 kg) solution were added dropwise for 24 hr to the reaction solution and the same was stirred for 1 hr at the same temperature. The prepared sodium salt of (S)-3,4-dihydroxybutyric acid was identified using NMR analysis.
1H-NMR (D2O, ppm) 6 2.27 (dd, 1H), 2.39 (dd, 1H), 3.41 (dd, 1H), 3.51 (dd, 1H), 3.8-3.9 (m, 1H)
The reaction solution was concentrated/ and 10L of methanol was added. Sulf uric acid were added to adjust the pH to 4~5, and then the same was stirred for 3 hr at 50°C. Sodium carbonate was added to neutralize the solution, and the same was filtered to remove the by-product, and then methanol was concentrated to obtain methyl (S)-3,4-dihydroxybutanoate. The formation of methyl (S)-3,4-dihydroxybutanoate (conversion ratio: 92%) was identified through NMR analysis by comparison with internal standard. 1H-NMR (CDCl3, ppm) 5 2.5 (dd, 2H), 3.5 (dd, 1H), 3.6 (dd, 1H). 3.7 (s, 3H), 4.1 (m, 1H)
Example 2: Preparation of (S)-3-hydroxy-y-butyrolactone
10L of water and 5kg of dried amylopectin were put into a 50L reactor.
After heating the reactor to 55°C, 12g of a-amylase (Teramyl; EC 3.2.1.1 from
Bacillus nmyloliqitcfaciens, Novo Nordisk) was added. After heating this
reaction solution to 85°C, the same was stirred for 2 hr at the same temperature.
5mL of 0.1N HC1 solution was added to adjust the pH 3.0-3.5, and then the same was stirred for 1 hr at 90 °C to inactivate the remaining α-amylase. After slowly cooling the reaction to 300C, 3.7L of 4M acetic acid buffer solution (pH 5) and 1.3L of water were added to adjust the pH to 5. The reaction solution was heated to 600C, and then 62.5g of pullulanase (Promozyme; EC 3.2.1.4 from Bacillus acidopullulyticus, Novo Nordisk) was added and the. solution was stirred for 22 hr at the same temperature. 0.54kg of 40% NaOH solution was added to the reaction solution to neutralize acetic acid and the temperature was raised to 60 °C. 40% NaOH (8.64kg) solution and 30% H2O2 (5.25kg) solution were added dropwise for 24 hr to the reaction solution and the same was stirred for 1 hr at the same temperature. The prepared sodium salt of (S)-3,4-dihydroxybutyric acid was identified using NMR analysis.
1H-NMR (D2O, ppm) 6 2.27 (dd, 1H), 2.39 (dd, 1H), 3.41 (dd, 1H), 3.51 (dd, 1H), 3.8-3.9 (m, 1H)
The reaction solution was concentrated, and 10L of methanol was added.
In this solution, methanesulfonic acid was added to adjust the pH to 4-5, and
then the same was stirred for 3 hr at 50°C. After cooling, sodium carbonate
was added to neutralize the solution, and the same was filtered to remove the
by-product, and then methanol was concentrated to obtain methyl
(S)-3,4-dihydroxybutanoate. Formation of methyl
(S)-3,4-dihydroxybutanoate (conversion ratio: 93%) was identified using NMR analysis by comparison with the internal standard.
1H-NMR (CDCl3, ppm) 5 2.5 (dd, 2H), 3.5 (dd, 1H), 3.6 (dd, 1H), 3.7 (s, 3H), 4.1 (m, 1H)
The prepared methyl (S)-3,4-dihydroxybutanoate was cyclized at 65 °C
under reduced pressure by adding 0.5 wt% of concentrated HC1 without any
separation being performed. The resultant solution was dissolved with

ethyl acetate and the same was neutralized with sodium carbonate. After filtrating and concentrating the same, (S)-3-hydroxy-y-butyrolactone (2.86kg, 57.2 wt% of the amylopectin weight used) was obtained.
1H-NMR (CDCl3, ppm) 6 2.28 (dd, 1H), 2.74 (dd, 1H), 4.13 (dd, 1H), 4.32 (dd, 1H), 4.4-4.5 (m, 1H)
Example 3: Preparation of (S)-3-hydroxy-y-butyrolactone
As in the Example 2, however using 1 wt% of methanesulfonic acid
rather than concentrated HC1 in the cyclization of the prepared methy
(S)-3,4-dihydroxybutanoate, the same was cyclized at 65°C under reduced
pressure. The resultant solution was dissolved with ethyl acetate and the
same was neutralized with sodium carbonate. After filtering and
concentrating the same, (S)-3-hydroxy-y-butyrolactone (2.85kg, 57 wt% of the amylopectin weight used) was obtained.
iH-NMR (CDCl3, ppm) 5 2.28 (dd, 1H), 2.74 (dd, 1H), 4.13 (dd, 1H), 4.32 (dd, 1H), 4.4-4.5 (m, 1H)
Example 4: Preparation of methyl (S)-3,4-dihydroxybutanoate
As in the Example 1, however using t-butylhydroperoxide (4.16 kg) rather than H2O2 for the oxidant, methyl (S)-3,4-dihydroxybutanoate was obtained. The formation of methyl (S)-3,4-dihydroxybutanoate (conversion ratio: 91%) was identified using NMR analysis by comparison with internal standard.
1H-NMR (CDCl2, ppm) 5 2.5 (dd, 2H), 3.5 (dd, 1H), 3.6 (dd, 1H), 3.7 (s, 3H), 4.1 (m, 1H) Comparative example 1: Preparation of (S)-3-hydroxy-y-y-butyrolactbne from starch 20L of water and 5kg of dried starch were put into a 5OL reactor, and the temperature was raised to 70°C. 40% NaOH (8.64kg) solution and 30% H2O2 (5.25kg) solution were added dropwise for 48 hr to the reaction solution and the same was stirred for 1 hr at the same temperature. The same was esterified and cyclized as in Example 2 to obtain (S)-3-hydroxy-y-butyrolactone (l.lkg, 22.0 wt% of starch weight used).
Comparative example 2: Preparation of (S)-3-hydroxy^y-butyrolactone from starch
10L of 0.5N HCI solution and 5kg of dried starch were put into a 5OL
reactor, and the starch was hydrolyzed for 20 min at 100°C. After cooling
the solution to 20°C, the same was neutralized with lOOmL of 40% NaOH
solution and the temperature was raised to 70°C. 40% NaOH (8.64kg)
solution and 30% H2O2 (5.25kg) solution were added dropwise for 48 hr to the reaction solution and the same was stirred for 1 hr at the same temperature. The same was esterified and cyclized as in Example 2 to obtain (S)-3-hydroxy-y-butyrolactone (1.22kg, 24.4 wt% of starch weight used).
Comparative example 3: Preparation of (S)-3-hydroxy-y-butyroIactone from amylose
20L of water and 5kg of dried amylose were put into a 50L reactor, and
the temperature was raised to 70°C. 40% NaOH (8.64kg) solution and 30%
H2O2 (5.25kg) solution were added dropwise for 48 hr to the reaction solution
and the same was stirred for 1 hr at the same temperature. The same was
esterified and cyclized as in the Example 2 to obtain (S)-3-hydroxy-y-butyrblactone (1.35kg, 27.0 wt% of amylose weight used).
Experimental example 1: Comparison of (S)-3-hydroxy-y-butyrolactone yield
depending on the source material
For the reaction solutions containing each of the carbohydrates shown in Table 1, oxidation, esterification and cyclization were performed as in the Example 2 to obtain (S)-3,4-dihydroxybutyric acid derivatives, and subsequently cyclized to obtain (S)-3-hydroxy-y-butyrolactone. The yields of (S)-3-hydroxy-y-butyrolactone are shown in Table 1.
Table 1

(Table Removed)Table 1 shows that for disaccharide the relative mass conversion yield is
low as 23.7 wt%. On the other hand, if amylopectin is transformed to
oligosaccharide with specific enzyme treatment, the relative mass conversion yield is enhanced to 57.2 wt%, almost two times compared with disaccharide. If amylopectin is not treated with enzymes, the relative mass conversion yield is relatively low as 20.2 wt%.
Experimental example 2: Optical purity analysis of (S)-3-hydroxy-y-butyrolactone
(S)-3-Acetoxy-y-butyrolactone was synthesized by the following method
in order to analyze optical purity of (S)-3-hydroxy-y-buryrolactone prepared from the present invention and the conventional preparing method.
102mg (1mmol) of (S)-3-hydroxy-y-butyrolactone prepared from each method was dissolved in 3mL of methylene chloride, and 0.4mL (5mmol) of pyridine and 0.47mL (5mmol) of acetic anhydride were added to the same. After 3 hr, the reaction was quenched with IN HCI. (S)-3-Acetoxy-y-butyrolactone was extracted with the methylene chloride. After work up, the same was purified with silica gel column chromatography. The obtained (S)-3-acetoxy-y-butyrolactone was dissolved in methylene chloride, and 0.5µl was taken with syringe for GC analysis. The result is shown in the following Table 2 and Figs. la~lc.
Table 2

(Table Removed)
To improve the medicinal efficiency and minimize the side effect, more
than 99.5%ee of high optical purity is required for chiral compounds. Table
2 and Figs. la~lc show that the optical purity of (S)-3-hydroxy-y-butyrolactone
prepared from the present invention is very high as 99.9%ee. So, the same
is very useful for the intermediates of other chiral compounds. The results
are illustrated in Figure la, Ib, and Ic, respectively
The preparing method of the present invention gives optically pure
(S)-3,4-dihydroxybutyric acid derivatives, which is very useful for industrial
uses because the by-product formation is minimized and the purification
process is very simple. It comprises alkaline oxidation of a-(l,4)-linked
oligosaccharide from the enzymatic reaction of amylopectin in a specific condition followed by esterifying to afford the target product. The present invention has overcome the disadvantage of using expensive metal catalyst for selective asymmetric reduction, and enables easy preparation from inexpensive natural product having optically pure chiral center, thereby the industrial utility as chiral intermediates of various medicine, can be maximized. Furthermore, the relative mass conversion yield is almost double compared with disaccharides.





CLAIMS
What is claimed is:
1. A process for producing optically pure (S)-3,4-dihydroxybutyric acid derivatives expressed by the following Formula 1 from polysaccharide source, which comprises the following steps:
Enzymatic reaction of amylopectin to α-(l,4) linked oligosaccharide
expressed by the Formula 2; and
Oxidation of the oligosaccharide with oxidant in base, and subsequent
esterification with alcohol in the presence of an acid catalyst.
(Formula Removed)
, wherein R represents linear or branched alkyl group with 1-5 carbon
atoms.
2. The process according to claim 1, wherein the said oligosaccharide has
3-50 glucose units.
3. The process according to claim 1, wherein the said oxidation is
performed in the temperature range of 30-65 °C.
4. The process according to claim 1, wherein the base used in the said
oxidation is selected from alkali metal hydroxide and alkaline earth
metal hydroxide.
5. The process according to claim 4, wherein the said base is sodium
hydroxide.
6. The process according to claim 1 or claim 4, wherein the said base is
used in the range of 2~4 equivalents per molar glucose unit of
amylopectin.
7. The process according to claim 1, wherein the oxidant used in the said
oxidation is selected from hydrogen peroxide, alkali metal peroxide,
alkaline earth metal peroxide and alkyl hydroperoxide.
8. The process according to claim 7, wherein the said oxidant is hydrogen
peroxide.
9. The process according to claim 7, wherein the said oxidant is
t-butylhydroperoxide.
10. The process according to claim 1 or claim 7, wherein the said oxidant is
used in the range of 1-3 equivalents per molar glucose unit of
amylopectin.
11. The process according to claim 1, wherein the said esterification is
performed in the temperature range of 30-80 °C.
12. The process according to claim 1, wherein the acid catalyst used in the
said esterification is an inorganic acid selected from hydrochloric acid,
sulfuric acid, phosphoric acid and nitric acid.
13. The process according to claim 1, wherein the acid catalyst used in the said esterification is an organic acid selected from fluoroalkylsulfonic acid, aralkylsulfonic acid, hydrate of aralkylsulfonic acid and trifluoroacetic acid.
14. A process for producing optically pure (S)-3,4-dihydroxybutvric acid derivatives substantially as herein described with reference to the accompanying drawings.


Documents:

in-pct-2001-00053-del-abstract.pdf

in-pct-2001-00053-del-claims.pdf

in-pct-2001-00053-del-correspondence-others.pdf

in-pct-2001-00053-del-correspondence-po.pdf

in-pct-2001-00053-del-description (complete).pdf

in-pct-2001-00053-del-drawings.pdf

in-pct-2001-00053-del-form-1.pdf

in-pct-2001-00053-del-form-13.pdf

in-pct-2001-00053-del-form-19.pdf

in-pct-2001-00053-del-form-2.pdf

in-pct-2001-00053-del-form-26.pdf

in-pct-2001-00053-del-form-3.pdf

in-pct-2001-00053-del-form-5.pdf

in-pct-2001-00053-del-pct-210.pdf

in-pct-2001-00053-del-pct-301.pdf

in-pct-2001-00053-del-pct-304.pdf

in-pct-2001-00053-del-pct-308.pdf

in-pct-2001-00053-del-pct-332.pdf

in-pct-2001-00053-del-pct-409.pdf

in-pct-2001-00053-del-pct-416.pdf

in-pct-2001-00053-del-petition-137.pdf


Patent Number 219637
Indian Patent Application Number IN/PCT/2001/00053/DEL
PG Journal Number 28/2008
Publication Date 11-Jul-2008
Grant Date 12-May-2008
Date of Filing 22-Jan-2001
Name of Patentee SAMSUNG FINE CHEMICALS CO. LTD.
Applicant Address 190, YEOCHEON-DONG, NAM-KU, 680-090, ULSAN, REPUBLIC OF KOREA.
Inventors:
# Inventor's Name Inventor's Address
1 KYOUNG ROK ROH
2 YIK-HAENG CHO
3 HO SUNG YU
4 YOUNG MI PARK
5 DAE LL HWANG
6 JONG PIL CHUN
PCT International Classification Number C12P 41/00
PCT International Application Number PCT/KR99/00396
PCT International Filing date 1999-07-23
PCT Conventions:
# PCT Application Number Date of Convention Priority Country
1 98-29912 1998-07-22 Republic of Korea
2 98-29912 1998-07-24 Republic of Korea