JPH1146793A - Production of optically active glycidol - Google Patents

Abstract

PROBLEM TO BE SOLVED: To provide a method for efficiently producing optically active glycidol important for synthesizing physiologically active compounds. SOLUTION: This method for producing optically active glycidol comprises a process for allowing an enzyme capable of optically selectively hydrolyzing a (R,S)-glycidol ester of the formula (R is a 1-20C linear, branched or cyclic saturated or unsaturated hydrocarbon group substituted or not substituted by a halogen atom, hydroxyl group or alkoxy group) to act on the (R,S)-glycidol ester in a reaction solution containing an organic solvent-water mixture solvent, a process for separating the unreacted optically active glycidol ester from the reaction solution, and a process for subjecting the separated optically active glycidol ester to a transesterification to produce the optically active glycidol.

Description

DETAILED DESCRIPTION OF THE INVENTION

[0001]

The present invention relates to a method for producing optically active glycidol which is useful as an intermediate for synthesizing optically active physiologically active compounds used for medicines and agricultural chemicals and liquid crystal materials. More specifically, the present invention relates to a method for obtaining an optically active glycidol by hydrolyzing (R, S) -glycidol ester optically selectively by an enzymatic method.

[0002]

2. Description of the Related Art Hitherto, many physiologically active compounds have been used as a mixture of their optical isomers. However, the desired activity often exists in only one optical isomer. Further, it is known that the other optical isomer may have toxicity to a living body without having a desired activity. Therefore, in order to provide an effective and safe drug or bioactive compound, it is strongly desired to develop a method for producing a compound having high optical purity.

[0003] Optically active glycidol is an important compound as a key material in the synthesis of various useful physiologically active substances, pesticides, liquid crystal materials and the like. As a method for synthesizing optically active glycidol, a synthesis method from D-mannitol by Fischer et al. Has been known for a long time. However, this method required a long process for preparation and was not suitable for industrial production. Recently, Sharpless et al.'S method for asymmetric epoxidation of allyl alcohol (Asymmetric Synthesi
s, Vol. 5, Academic Press, Inc., London, 1985, p. 24
7) has been reported. This is a method that can be prepared in a short stage, but has the disadvantage that the optical purity of the product is low during mass production.

On the other hand, there has been reported a method for producing optically active glycidol using microorganisms or enzymes utilizing the optically selective catalytic properties of living organisms. For example, asymmetric oxidation by an alcohol dehydrogenase such as Acetobacter or Gluconobacter has been reported (EP 0 464 905 A1). Although these methods can provide glycidol having relatively high optical purity as compared with the synthetic method, they have problems that are not suitable for industrial production, such as expensive enzymes and difficulty in mass production. Was.

Also, a method for optically selective hydrolysis of racemic glycidyl butyrate with porcine pancreatic lipase (J. Am. C.
hem. Soc., 106,7252 (1984), US Patent No. 4728553,
JP-A-1-285199), a method for optically selective hydrolysis of racemic glycidol ester by various microbial ester-degrading enzymes such as Rhodotorula, Pseudomonas, Rhizopus, Klebsiella, Mucoa, Achromobacter, and Alcaligenes. 7-99993 and JP-A-8-84597), or an optically selective esterification of racemic glycidol with porcine pancreatic lipase in an organic solvent (Biotechnology and Bioengineering, 42,
465-468 (1993), J. Am. Chem. Soc., 110, 7200-7205 (198
8)) has been reported. However, none of these methods can produce glycidol having sufficient optical purity.

[0006] From such a viewpoint, there is a strong demand for a method for efficiently producing optically active glycidol having high optical purity.

[0007]

DISCLOSURE OF THE INVENTION The present invention has been made to solve the above-mentioned problems, and an object of the present invention is to provide a method for optically selecting glycidol ester by an enzymatic method using a reaction solvent for improving the catalytic properties of the enzyme. An object of the present invention is to provide a method for industrially efficiently producing an optically active glycidol having high optical purity by hydrolysis.

[0008]

The present invention is a process for producing optically active glycidol, which comprises the following general formula:

[0009]

Embedded image

(Wherein R is a linear, branched or cyclic saturated or unsaturated hydrocarbon group substituted or unsubstituted by a halogen atom having 1 to 20 carbon atoms, a hydroxyl group or an alkoxyl group) (R, S) -glycidol ester represented by the above formula (R, S)-
Reacting an enzyme that optically selectively hydrolyzes glycidol ester with a reaction solution containing an organic solvent / water mixed solvent; and separating the produced optically active glycidol from the reaction solution. About.

The present invention is a process for producing optically active glycidol, which comprises the following general formula:

[0012]

Embedded image

(Wherein R is a linear, branched or cyclic saturated or unsaturated hydrocarbon group substituted or unsubstituted by a halogen atom having 1 to 20 carbon atoms, a hydroxyl group or an alkoxyl group) (R, S) -glycidol ester represented by the above formula (R, S)-
Reacting an enzyme that optically selectively hydrolyzes glycidol ester with a reaction solution containing a mixed solvent of an organic solvent and water; separating unreacted optically active glycidol ester from the reaction solution; A method comprising transesterifying the optically active glycidol ester to produce optically active glycidol.

In a preferred embodiment, the enzyme selectively degrades the (S) -glycidol ester of the (R, S) -glycidol ester, and the resulting optically active glycidol is (R) -glycidol. .

In a preferred embodiment, the unreacted optically active glycidol ester is (R) -glycidol ester, and the optically active glycidol formed is (S) -glycidol.

In a preferred embodiment, (R,
(S) -glycidol ester includes (R, S) -glycidyl acetate, (R, S) -glycidyl propionate,
It is selected from the group consisting of (R, S) -glycidyl butyrate, (R, S) -glycidyl pentylate, (R, S) -glycidyl hexalate.

In a preferred embodiment, the enzyme is an esterase derived from porcine pancreas.

In a preferred embodiment, the enzyme is immobilized on a carrier for immobilization.

In a preferred embodiment, the organic solvent / water mixed solvent is methyl acetate / acetone / tetrahydrofuran / water mixed solvent, methyl acetate / acetone / water mixed solvent, methyl acetate / tetrahydrofuran / water mixed solvent,
Acetone / tetrahydrofuran / water mixed solvent, tetrahydrofuran / water mixed solvent, ethanol / water mixed solvent, n
It is selected from the group consisting of -propanol / water mixed solvent, n-butanol / water mixed solvent, acetonitrile / water mixed solvent, and 1,4-dioxane / water mixed solvent.

[0020]

DESCRIPTION OF THE PREFERRED EMBODIMENTS The present invention will be described below in detail.

The method of the present invention comprises reacting a racemic mixture with an enzyme having optically selective hydrolysis activity in a reaction solution containing a mixed organic solvent of a hydrophilic organic solvent and water,
Based on obtaining optically active compounds.

More specifically, in the method of the present invention, (R, S) -glycidol is used in a reaction solution containing a mixed organic solvent of water and various hydrophilic organic solvents for improving the reaction specificity of the enzyme. An enzyme having the activity of enantioselective hydrolysis of the ester is allowed to act on the ester to hydrolyze either the (R) or (S) glycidol ester. Thereby, the (R, S) -glycidol ester is converted to a mixture of optically active glycidol and optically active glycidol ester. Optically active glycidol or unreacted optically active glycidol ester is fractionated by utilizing the difference in their chemical or physical properties. The fractionated optically active glycidol ester can be easily converted to optically active glycidol by transesterification.

As used herein, “(R, S) -glycidol ester” refers to the following general formula:

[0024]

Embedded image

(Where R is a straight-chain, branched or cyclic saturated or unsaturated hydrocarbon group having 1 to 20 carbon atoms which is substituted or unsubstituted by a halogen atom, a hydroxyl group or an alkoxyl group) Is a racemic mixture of (R) -glycidol ester and (S) -glycidol ester. The racemic mixture typically contains the (R) form and the (S) form at a ratio of 1: 1, but the mixing ratio of the (R) form and the (S) form is necessarily 1: 1.
It is not limited to. The number of carbon atoms of R in the general formula is generally 1 to
20, preferably 1 to 10, more preferably 1 to
5

(R, S) to be a substrate for the enzyme reaction of the present invention
-Glycidol ester is used for the substrate specificity of the enzyme used,
It is appropriately selected from the solubility in the enzyme reaction solvent and the ease of separation after the enzyme reaction. For example, glycidyl acetate, glycidyl propionate, glycidyl butyrate, glycidyl pentylate, and glycidyl hexalate can be suitably used. These compounds can be prepared by known esterification methods (eg, CM Lok et al, Chem. Phys. Li
pids., 16, 115 (1978)). For example, it can be obtained by reacting racemic glycidol with butyric anhydride or butyric halide in the presence of pyridine or an amine such as triethylamine, tributylamine or dimethylaminopyridine.

The enzymes used in the method of the present invention include:
Any enzyme capable of photoselectively hydrolyzing the (R, S) -glycidol ester or of photoselectively esterifying glycidol can be used. An example of such an enzyme includes an esterase derived from pig pancreas. In addition, any form of preparation that contains the enzymatic activity described above can be used in the methods of the present invention. For example, a crude enzyme and an enzyme preparation can be used. Examples of the crude enzyme include microbial cells, cell cultures, and processed cells that produce the enzyme. Examples of the enzyme preparation include lipase Type II (porcine pancreas, Sigma) and pancreatin F (porcine pancreas, Amano Pharmaceutical). These enzymes have the catalytic activity of selectively hydrolyzing the (S) -glycidol ester of (R, S) -glycidol ester to produce (R) -glycidol, but select the (R) -glycidol ester. An enzyme having a catalytic activity of hydrolyzing to form (S) -glycidol can also be used.

Further, an immobilized enzyme in which the above enzyme is adsorbed on a carrier for immobilization may be used. As a method of immobilization, a carrier binding method (for example, a covalent binding method, an ion binding method,
Alternatively, a method known to those skilled in the art such as a physical adsorption method, a crosslinking method, or an entrapment method (lattice type or microcapsule type) can be used. The carrier for immobilization can be easily selected by those skilled in the art in view of the ability to immobilize the enzyme used in the present invention, reusability, ease of handling, and the like. For example, Chitopearl (manufactured by Fuji Spinning Co., Ltd.) or the like can be used.

The reaction solution used in the optically selective glycidol hydrolysis reaction of the present invention contains a mixed solvent of an organic solvent and water. The organic solvent / water mixed solvent can remarkably improve the substrate selectivity of the enzyme reaction when compared with a commonly used inorganic solvent such as water or a phosphate buffer. Organic solvents are selected based on physical parameters such as dielectric constant, dipole moment, and hydrophobicity. Generally, a solvent having a large dielectric constant and a large dipole moment and a solvent having a low hydrophobicity are suitably used. In general, solvents having a dielectric constant of 20 or more tend to increase the substrate selectivity of the enzymatic reaction. Solvents that exhibit more than one dipole moment also tend to increase the substrate selectivity of the enzymatic reaction. Further, if logP (a parameter relating to the distribution of n-octanol between the organic solvent and water), which is a parameter indicating the hydrophobicity of the organic solvent, is lower than 0,
It tends to increase the substrate selectivity of the enzymatic reaction.

Examples of such an organic solvent include methyl acetate, acetone, THF, ethanol, n-propanol, n-butanol, acetonitrile, 1,4-dioxane and the like.

The above organic solvent is used by mixing with water. Preferably, the water and organic solvent are between 20:80 and 50: 5.
It is mixed at a volume ratio of 0. When the amount of water is lower than 20% by volume, the reaction rate becomes slow. When the amount of water exceeds 50% by volume, the substrate selectivity of the enzymatic reaction decreases.

The organic solvent / water mixed solvent of the present invention can be used by mixing two or more kinds of organic solvents and mixing with water. Examples of such a mixed solvent include, for example, methyl acetate, acetone / THF / water (2: 1: 1: 1), methyl acetate / THF / water (1: 3: 2), methyl acetate / acetone / water ( 3: 1: 1), THF / water (2: 1), ethanol / water (2: 1), n-propanol / water (2:
1), n-butanol / water (2: 1), acetonitrile / water (2: 1), 1,4-dioxane / water (2: 1), and the like, but are not limited thereto. The numbers in parentheses indicate the volume ratio of each solvent (the same applies hereinafter in this specification).

[0033] The reaction solution of the present invention may further contain a cofactor promoting the enzymatic reaction. Such cofactors include, for example, surfactants.

The optically selective hydrolysis of the (R, S) -glycidol ester of the present invention is carried out by adding the above esterase to a reaction solution containing the (R, S) -glycidol ester and stirring the mixture. .

The reaction temperature is from -20 ° C to 50 ° C, preferably from 0 ° C to 20 ° C. Although the amount of the enzyme to be used is not particularly limited, the preferable range is usually about 0.1 to 10 g / mol of the substrate with respect to the glycidol ester of the reaction substrate. The pH of the reaction solution is 4-9, preferably 5-8. Reaction time depends on the amount of enzyme used, reaction temperature, reaction pH
Etc., but usually completes in about 1 to 24 hours.

After completion of the reaction, the produced glycidol and unreacted glycidol ester are separated by a commonly used separation operation such as a solvent extraction method, a crystal precipitation method, and a column chromatography method. In the solvent extraction method, when n-hexane, heptane, toluene, cyclohexane, isooctane, dichloromethane, isopropyl ether, and t-butyl methyl ether are used, only the unreacted optically active glycidol ester is recovered at a rate of 90% or more from the reaction solution. It is possible to collect at.

Next, the fractionated optically active glycidol ester is converted into optically active glycidol. Ordinary chemical hydrolysis using an acid, a base, or the like is not suitable for the method of the present invention because side reactions such as ring opening of an epoxy ring and racemization easily occur. In the present invention, optically active glycidol is produced by transesterification of the fractionated optically active glycidol ester. For example, by performing a transesterification reaction in excess methanol, optically active glycidol can be easily obtained.

As is known to those skilled in the art, the optically selective hydrolysis reaction of a racemic ester compound using an enzyme such as lipase is a method for quantitatively processing biochemical data.
The optical selectivity can be numerically indicated by the E value. The E value is given by the following equation.

E = ln {(1-c) (1 + ee (P))} / ln {(1-c) (1-ee (P))} where E: enantiomeric ratio c: enzymatic reaction Conversion rate (%) ee (P): Indicates the enantiomeric excess (ee) (optical purity) of the product.

In the present invention, it is preferable that the optically selective hydrolysis is achieved at an E value of 10 or more. More preferably, optically selective hydrolysis is achieved at an E value of 20 or more, and most preferably, optically selective hydrolysis is achieved at an E value of 30 or more.

Hereinafter, the present invention will be described more specifically with reference to examples, but the present invention is not limited to these examples. In the examples, quantification of glycidol and glycidol ester in the reaction solution and measurement of optical purity were performed by gas chromatography (SHIMADZU GC-14A) under the following analysis conditions.

(Gas Chromatographic Analysis Conditions) Column: Chiraldex G-TA, 30 m (manufactured by Astec) Temperature: initial column temperature: 44 ° C. (30 min), heating rate: 10 ° C./min, final temperature: 80 ° C. Injector temperature; 250 ° C, detector temperature; 250 ° C Carrier gas: helium, 1 ml / min Detector: FID

[0043]

【Example】

Example 1 Enantioselectivity and physicochemical properties of a solvent (dielectric constant, dipole moment, and l) in an optically selective hydrolysis reaction of (R, S) -glycidyl butyrate
ogP) was analyzed. Where logP
Is a parameter for the partitioning of n-octanol between the organic solvent and water. As the value of logP is larger,
It indicates that the organic solvent has high hydrophobicity.

THF / water (2: 1), acetonitrile /
Water (2: 1), ethanol / water (2: 1), acetone /
Water (2: 1), 1,4-dioxane / water (2: 1), n-
200 ml of a mixed solvent of butanol / water (5: 1), n-propanol / water (2: 1), methyl acetate / water (10: 1), or isopropyl ether / water (200: 1)
Lipase Type II (porcine pancreas, Si
gma) 2 g and glycidyl butyrate 2.5 mL (1
(7.5 mmol) and stirred at 10 ° C. for 4 hours. After completion of the reaction, the reaction solution was analyzed by gas chromatography. The results are shown in FIGS. The horizontal axes in FIGS. 1 to 3 are the dielectric constant, dipole moment, and l of each organic solvent, respectively.
ogP and the ordinates respectively show the E values of the above-mentioned optically selective hydrolysis reactions.

From the results of FIGS. 1-3, (i) the higher the dielectric constant of the reaction solvent, the higher the optical selectivity (FIG. 1); (ii)
The higher the dipole moment, the higher the optical selectivity (FIG. 2); and (iii) the lower the logP, the higher the optical selectivity, ie, the more hydrophilic the organic solvent, the higher the optical selectivity. (FIG. 3) The tendency was shown.

Example 2 Based on the results obtained in Example 1, screening of various organic solvents / water mixed solvents was performed.

(R, S) -glycidyl butyrate was prepared in a reaction solution containing various mixed solvents shown in Table 1 by lipase Type II (derived from porcine pancreas, manufactured by Sigma) in the same manner as in Example 1. The reaction is performed and the optical activity (S)
-Glycidol was obtained. As the reaction time, the reaction time shown in Table 1 was used. The results are summarized in Table 1. As shown in the table, methyl acetate, acetone, THF,
When a mixed solvent of water (2: 1: 1: 2) was used, (R) -glycidyl butyrate having an optical purity of 99% ee or more was obtained in a yield of 39%. To this reaction solution, add n-heptane 1
After adding 00 mL, (R) -glycidyl butyrate was extracted. This extract is fed to a rotary evaporator for 50 m.
L, and then concentrated to 150 mL of methanol and 1.5 g.
(S) -glycidol having an optical purity of 99% ee or more was obtained in a yield of 36%. The E value of this optically selective hydrolysis reaction was 32. This is about three times that in the case where water is used as the reaction solvent (E value = 11), indicating that the organic solvent / water mixed solvent of the present invention can significantly improve the optical selectivity of the enzyme reaction. .

[0048]

[Table 1]

Example 3 Methyl acetate / acetone / THF / water (2:
Various enzymes were screened using a mixed solvent of 1: 1: 2). Various enzymes 2 shown in Table 2 were added to a reaction solution of 200 ml of a mixed solvent of methyl acetate / acetone / THF / water (2: 1: 1: 2) or a reaction solution of 200 ml of water.
g and glycidyl butyrate 2.5 mL (17.5 mmol
l) was added, and an enzyme reaction was carried out in the same manner as in Example 1. Table 2 shows the results. As shown in the table, Lipase Type II
It was shown that the best results were obtained when using (porcine pancreas, Sigma).

[0050]

[Table 2]

(Example 4) 25 mg / ml lipase Ty was added to 100 g of the immobilization carrier Chitopearl (manufactured by Fuji Boseki Co., Ltd.).
100 ml of peII enzyme solution and 300 ml of water were added, and 20
After shaking at 時間 30 ° C. for 2 hours, suction filtration was performed. After the carrier for immobilization was sufficiently washed with water, the enzyme activity of the filtrate was measured, and the immobilized lipase Type was detected at an adsorption rate of 60%.
II was obtained.

(Example 5) Methyl acetate / acetone / TH
The immobilized lipase Ty obtained in Example 3 was added to a reaction solution obtained by adding 2.5 mL (17.5 mmol) of glycidyl butyrate to 200 mL of a mixed solvent of F and water (2: 1: 1: 2).
20 g of peII was added, and the enzymatic reaction and product extraction were performed in the same manner as in Example 1. As a result, (S) -glycidol having an optical purity of 99% ee was obtained in the same yield as in Example 1.

[0053]

According to the present invention, there is provided a method for efficiently producing optically active glycidol, which is a synthetic intermediate for pharmaceuticals, agricultural chemicals or liquid crystal materials, with high optical purity.

[Brief description of the drawings]

FIG. 1 is a graph showing a correlation between a dielectric constant of an organic solvent / water mixed solvent and an E value in an optically selective hydrolysis reaction of (R, S) -glycidyl butyrate.

FIG. 2 is a graph showing a correlation between an E value and a dipole moment of an organic solvent / water mixed solvent in an optically selective hydrolysis reaction of (R, S) -glycidyl butyrate.

FIG. 3 shows log P of an organic solvent / water mixed solvent in an optical selective hydrolysis reaction of (R, S) -glycidyl butyrate.
6 is a graph showing a correlation between the E value.

Continuation of the front page (72) Inventor Kotaro Otsuka 2-2-2 Muroya, Nishi-ku, Kobe-shi, Hyogo Nagase Sangyo R & D Center

Claims (8)

[Claims] 1. A method for producing optically active glycidol, which comprises the following general formula: (Where R is a linear, branched, or cyclic saturated or unsaturated hydrocarbon group substituted or unsubstituted by a halogen atom having 1 to 20 carbon atoms, a hydroxyl group, or an alkoxyl group) An (R, S) -glycidol ester represented by the formula (I) and an enzyme that hydrolyzes the (R, S) -glycidol ester optically selectively are combined with an organic solvent
Acting in a reaction solution containing a water mixed solvent; and separating the produced optically active glycidol from the reaction solution. 2. A method for producing optically active glycidol, which comprises the following general formula: (Where R is a linear, branched, or cyclic saturated or unsaturated hydrocarbon group substituted or unsubstituted by a halogen atom having 1 to 20 carbon atoms, a hydroxyl group, or an alkoxyl group) An (R, S) -glycidol ester represented by the formula (I) and an enzyme that hydrolyzes the (R, S) -glycidol ester optically selectively are combined with an organic solvent
Acting in a reaction solution containing a water mixed solvent; separating unreacted optically active glycidol ester from the reaction solution; and transesterifying the separated optically active glycidol ester to produce optically active glycidol. A method comprising the steps of: 3. The method according to claim 1, wherein the enzyme selectively decomposes the (S) -glycidol ester of the (R, S) -glycidol ester, and the resulting optically active glycidol is (R) -glycidol. The method described in. 4. The method according to claim 2, wherein the unreacted optically active glycidol ester is (R) -glycidol ester, and the optically active glycidol formed is (S) -glycidol. 5. The (R, S) -glycidol ester is (R, S) -glycidyl acetate, (R, S) -glycidyl propionate, (R, S) -glycidyl butyrate, (R, S) ) -Glycidyl pentylate, (R,
The method according to any one of claims 1 to 4, wherein the method is selected from the group consisting of S) -glycidyl hexalate. 6. The method according to claim 1, wherein the enzyme is an esterase derived from porcine pancreas. 7. The method according to claim 1, wherein the enzyme is immobilized on a carrier for immobilization. 8. The mixed solvent of an organic solvent and water is a mixed solvent of methyl acetate / acetone / tetrahydrofuran / water, a mixed solvent of methyl acetate / acetone / water, a mixed solvent of methyl acetate / tetrahydrofuran / water, and a mixed solvent of acetone / tetrahydrofuran / water Selected from the group consisting of tetrahydrofuran / water mixed solvent, ethanol / water mixed solvent, n-propanol / water mixed solvent, n-butanol / water mixed solvent, acetonitrile / water mixed solvent, and 1,4-dioxane / water mixed solvent The method according to claim 1, wherein the method is performed.