JP2001019650A - Production of optically active alcohol - Google Patents

Abstract

PROBLEM TO BE SOLVED: To obtain an optically active alcohol of high optical purity that is gentle to the ecosystem and permits reduction in the reaction cost by applying ovalbumin originating from egg, particularly from egg white of chicken egg to the organic synthetic chemistry as an optical resolution catalyst. SOLUTION: Egg white and ovalbumin of egg white is spray-dried to a powder and the powder is preferably immobilized with an alginate salt extracted from seaweeds by the entrapping immobilization method. The ovalbumin powder immobilized with alginate salt is used as a catalyst to effect the enzymatic conversion of the starting material, for example, racemic alcohol or ketone molecule, preferably in an aqueous solvent. In an embodiment, an optically active alcohol, for example, 1-(4-bromophenyl)ethanol or the like or their acyl derivative is extracted with an inert organic solvent, for example, ethyl acetate or the like, preferably isolated and purified through silica gel (thin-layer) chromatography, and, when necessary, hydrolyzed to give the objective optical alcohol.

Description

DETAILED DESCRIPTION OF THE INVENTION

[0001]

The present invention relates to a method for producing an optically active alcohol from a substrate using egg white or ovalbumin isolated from egg white immobilized as a catalyst.

[0002]

2. Description of the Related Art Optically active alcohols are extremely important as raw materials or intermediate raw materials for pharmaceuticals and agricultural chemicals, and synthetic intermediates in the field of fine chemicals such as ferroelectric liquid crystals. In order to biosynthesize such optically active substances such as optically active alcohols, conventionally, 1) microbial cells, 2) enzymes derived from microbial cells, 3) enzymes derived from animal tissues, 4) plant cultured cells, etc. There is known a method of producing an optically active alcohol by reacting. As a method using the microbial cells of the above 1), there is a method of reacting the cultured cells with a substrate to obtain an optically active alcohol.
No. 578 (a method for producing optically active 1,2-diols) is known. In addition, as a method of using the microorganism-derived enzyme of 2), there is a method in which a pulverized liquid of cells cultured by introducing a gene is reacted with a substrate to obtain an optically active alcohol, for example, Japanese Patent Publication No. 10-210981. (Novel proteins that catalyze the conversion of halohydrins into optically active diols) are known.

[0003] As a method of 3) using an enzyme derived from animal tissue, there is a method of reacting a protein separated from animal tissue with a substrate to obtain an optically active alcohol. For example, Japanese Patent No. 2756790 (optically active) Production method of cyclopentenol derivative) is known. In the reaction using plant cultured cells of 4), there is a method of reacting plant cells with a substrate to obtain an optically active alcohol. For example, JP-A-4-222592 (a method for producing optically active inabenfide using plant cultured cells) is known. It is known.

[0004] 1) A method using "microbial cells" is disclosed in Japanese Patent No. 2784578 (a method for producing optically active 1,2-diols) and Japanese Patent No. 2774341 (a method for producing an optically active 2-hydroxy acid derivative). ), The microbial cells are grown in a culture solution by appropriately setting the culture conditions, and the cells are collected from the culture solution by centrifugation or filtration, and the cells are recovered at 0.1 M phosphate. Acid buffer (pH
6.5) Alternatively, the optically active alcohol is synthesized by asymmetric reduction of the ketone body of the substrate in a liquid suspended in distilled water or the like. However, due to the variety of cell-containing enzymes, since side reactions other than the substrate conversion reaction occur simultaneously, the yield of the desired optically active alcohol is low,
Further, even if the isolation and purification work is performed from a solvent containing a reaction product, the purity of the obtained optically active alcohol is low, so that it is difficult to use it as a synthetic intermediate in the field of fine chemicals.

[0005] 2) The method using "microbial enzyme" is as follows.
For example, as disclosed in JP-A-10-210981 (a novel protein that catalyzes the conversion of halohydrin to optically active diol), a large number of genes cloned by genetic recombination exist in the cells. This is a method for producing optically active epihalohydrin and optically active diol using transformed microorganisms, but this method also controls the side reactions that occur during the substrate reaction because there is no significant difference between the method using microbial cells and the reaction process itself. Therefore, it has the same problem as the method using "microbial cells". Furthermore, transgenic strains are heterogeneous varieties in nature, and have the risk of adversely affecting ecosystems, including humans.Therefore, it is necessary to bear the expense of equipment to isolate from the external environment and incineration of reaction residues. is there.

[0006] 3) "Animal tissue-derived enzymes" are known from Japanese Patent No. 2,756,790 (a method for producing an optically active cyclopentenol derivative) and the like. There is a method for producing an optically active cyclopentenol derivative used. As described in Kazuo Satake, “Organic Chemistry for Biology 3 Protein”, pp. 114-172 (published by Asakura Shoten), enzymes derived from animal tissues are used. Since is a crude enzyme, it is undeniable that a side reaction occurs to lower the yield as in the above 1) and 2).

[0007] 4) “Plant cultured cells” are disclosed in
-222592 (Method for producing optically active inabenfide using plant cultured cells) and JP-A-8-10328
No. 9 (stereoselective α-alkyl- by plant cells)
As is known in the process for producing β-hydroxycarboxylic acid esters, the yield of the obtained optically active alcohol is low due to side reactions other than the substrate conversion reaction caused by the variety of plant-containing enzymes. There's a problem. or,
The cultivation of plant cells is difficult in breeding because of the necessity of aseptic operation in the whole process, and furthermore, it requires a period of repeating subculture for 1 to 2 years. MS medium, MSK-II medium, B-5
Reaction operation is complicated, such as the necessity of preparing a medium or the like.

[0008]

SUMMARY OF THE INVENTION It is an object of the present invention to solve the above-mentioned problems by providing eggs, especially egg whites derived from inexpensive chicken eggs, which are produced and consumed in various countries around the world. By applying albumin to organic synthetic chemistry as an optical resolution catalyst, it is eco-friendly, and it is possible to greatly reduce the reaction cost. Furthermore, it is to provide a method for producing an optically active alcohol with high optical purity. .

[0009]

In the present invention, a method for producing an optically active alcohol having high optical purity by using the above-mentioned optical resolution catalyst generally includes the following method. (1) A method for producing an optically active alcohol in which one enantiomer of a racemic alcohol as a substrate is selectively oxidized to a ketone and the other enantiomer is left unreacted and separated as an optically active alcohol. (2) A method for producing an optically active alcohol by asymmetric reduction of a ketone molecule as a substrate. (3) A method for producing an optically active alcohol by asymmetric hydrolysis of an acylated racemic alcohol as a substrate. (4) An optically active alcohol which is stereoselectively acylated on one enantiomer of a racemic alcohol in an organic solvent and hydrolyzes the obtained acylated enantiomer to be separated as an optically active alcohol is produced. how to.

The inventor of the present invention has solved the above-mentioned problems and has conducted intensive studies on a method for obtaining a high-purity optically active alcohol by a safe and simple method. As a result, a protein selected from egg white and albumin separated from egg white was obtained. First powdering
A second step of immobilizing the protein, a third step of performing an enzymatic conversion reaction of a substrate using the immobilized protein as a catalyst, and the reaction substrate and the reaction converted in the third step. A fourth step of extracting the product mixture with an organic solvent and a fifth step of isolating and purifying the optically active alcohol or an acylated form of the optically active alcohol from the extract of the fourth step are combined, if necessary. By hydrolysis, high-purity R which can be sufficiently used as a raw material for synthesizing synthetic intermediates in the field of fine chemicals
The present inventors have found that optically active alcohols in the form of S or S can be obtained safely and easily, and have completed the present invention. Suitable as the optically active alcohol of the present invention is 1- (4-
(Bromophenyl) ethanol, 1- (4-chlorophenyl) ethanol, 1-phenylethanol, 1-phenylpropanol, 1-phenylbutanol, 1- (4-
Examples thereof include, but are not limited to, methylphenyl) ethanol, 1- (4-methoxyphenyl) ethanol, 1- (4-nitrophenyl) ethanol, and 1- (2-naphthyl) ethanol.

The albumen or ovalbumin which can be used in the present invention includes those derived from chicken eggs, but egg albumins of birds, amphibians and fish can also be used, and are not limited to those derived from chicken eggs. . The catalyst of the present invention may be any component containing ovalbumin, and thus egg white itself can be used. However, it is desirable to use ovalbumin separated from egg white efficiently. In the pulverization step in the first step of the present invention, ovalbumin separated from egg white by a known method such as the ammonium sulfate method or the sodium sulfate method has an ovalbumin concentration of 0.1 to 5 (volume)%. Or an aqueous solution is prepared using egg white itself such that the ovalbumin concentration is in the above range. These liquids are made of a hard alloy made of 0.3 to 4 m in diameter.
m with a pressure of 30 to 70 kg / cm ² through a hole of about 5 mm in diameter.
The powdered egg white or ovalbumin is obtained by using a spray-drying method of spraying in a state of very fine particles in hot air of 130 to 150 ° C. as droplets of 00 μm and evaporating water instantaneously to dry.

In the second step, the method of immobilizing the protein is as follows: 1) Binding of the protein to a water-insoluble carrier, for example, a polysaccharide derivative such as cellulose, dextran, agarose, or a polyacrylamide gel. 2) A cross-linking method in which the extracted protein is fixed by forming a cross-link between the proteins using a reagent having two or more functional groups, and 3) The protein is gelled, for example, alginate, starch, konjac Incorporation into a fine lattice of gels such as polyacrylamide gel and polyvinyl alcohol (lattice type) or covering with a semipermeable membrane (microcapsule type) Can be used. However, the comprehensive immobilization method using a salt of alginic acid extracted from seaweed is most preferable because it is environmentally friendly and the immobilization operation is simple.

In the third step, the enzymatic conversion method for obtaining an optically active alcohol or an acylated optically active alcohol from a substrate as a raw material comprises the steps of (1) selecting one enantiomer of a racemic alcohol as a substrate (2) a method of obtaining an optically active alcohol by asymmetric reduction of a ketone molecule as a substrate, (3) a substrate, To obtain an optically active alcohol by asymmetric hydrolysis of an acylated form of a racemic alcohol as (4) Stereoselection of one enantiomer of the racemic alcohol as a substrate with an acyl halide or vinyl acetate in an organic solvent The method includes a method of performing a selective acylation.

In the (2) asymmetric reduction reaction using a ketone molecule as a substrate, the enzymatic conversion
%, And stops halfway without reaching%, and if the reaction solution is not subjected to an extraction treatment at or before the reaction stops, the optical purity decreases with time. Therefore, it is necessary to determine an appropriate reaction stop time according to the type of egg white. Generally, when the reaction is terminated at a conversion rate of 7% or less, the configuration and optical purity of the resulting optically active alcohol are
In the case of using a racemic alcohol as a substrate in (1), the asymmetric hydrolysis reaction in the reaction using an acylated form of racemic alcohol as a substrate in (3) generally exceeds 7%. And the optical purity decreases. Therefore, it is necessary to determine an appropriate reaction stopping time depending on the type of egg white. However, although the yield is low, when the reaction is terminated at a conversion rate of 7% or less, the configuration and optical purity of the produced optically active alcohol are , (1) is the same as when the racemic alcohol is used as the substrate.

In the reaction (4) using the asymmetric acylation of a racemic alcohol in an organic solvent, the acylated product obtained by the asymmetric acylation reaction is further hydrolyzed to obtain an optically active alcohol. Is what you get. However, when the conversion rate exceeds 5%, the optical purity of the asymmetric acylation reaction deteriorates. Therefore, it is necessary to determine the reaction stop time depending on the type of egg white. Generally, the acylation reaction is terminated at a conversion rate of 5% or less. Then, the configuration and optical purity of the optically active alcohol produced by the subsequent hydrolysis are the same as those in the case of using racemic alcohol as the substrate in (1).

In the present invention, the method of (1) for obtaining an optically active alcohol by selectively oxidizing one enantiomer using a racemic alcohol as a substrate is most preferable from the viewpoint of yield and the like. The reaction temperature of the above (1) to (4) is about 25 to 4
5 ° C., preferably 30-40 ° C., is suitable,
Most preferably, it is carried out at a temperature of 0 ° C. In addition, the above (1) to
In the reaction (3), water can be used as the polar solvent, and acetone, methanol, ethanol, and the like can be used as the non-polar solvent, but water as the polar solvent is most preferable.
In the reaction (4), an organic solvent such as benzene, toluene, or isopropyl alcohol (dry) can be used, but benzene is preferably used. Further, the optically active alcohol obtained by the reaction becomes S-form or R-form under the influence of the substituent of the substrate.

In the fourth step, a non-reactive solvent such as ethyl acetate, diethyl ether or dichloromethane can be used as the extraction organic solvent. In the fifth step, the operation of performing isolation and purification is most preferably using silica gel chromatography or silica gel thin layer chromatography, but is described in Japanese Patent No. 2804247 (reaction using an immobilized biocatalyst). A part of the reaction solution rich in the product is withdrawn from the reaction tank, transferred to a crystallization tank set at the product deposition temperature to precipitate the product, separated by filtration, and then the substrate is added to the mother liquor. Then, a series of operations for returning to the reaction tank and performing the reaction are repeated, and an isolation and purification method known before the present application, such as an isolation and purification method for accumulating a suspended product in the crystallization tank, can be used.

[0018]

DESCRIPTION OF THE PREFERRED EMBODIMENTS Next, the present invention will be specifically described based on embodiments, but this is for explanation only,
Therefore, the present invention should not be limited. (1)
Method for Producing Optically Active Alcohol by Selectively Oxidating One Enantiomer of Racemic Alcohol as a Substrate of Example 1 (Ovalbumin) First, as a first step, egg white is separated from chicken eggs and subjected to ammonium sulfate method. Ovalbumin is separated in
% Ovalbumin was dissolved in water to give a concentration of ovalbumin and spray-dried to prepare ovalbumin as a powder. The aqueous sodium alginate solution was prepared by dissolving sodium alginate in the aqueous solution under autoclave conditions at a temperature of 121 ° C. for a time of 20 minutes.

Next, in the second step, 200 ml of a 10-fold equivalent of distilled water is added to 20 g of ovalbumin powder, and 250 ml of a 1.25-fold equivalent of a 5% aqueous sodium alginate solution is further added to make it uniform. To obtain a mixed solution of ovalbumin and sodium alginate. The obtained ovalbumin / sodium alginate mixed solution is
Ovalbumin-containing calcium alginate gel beads in an immobilized state were prepared by dropping into a 0.6% calcium chloride aqueous solution using a syringe or the like. Further, the beads were left standing in a 0.6% calcium chloride aqueous solution for 5 hours or more to strengthen the bead membrane.

Subsequently, in the third step, the ovalbumin-containing calcium alginate gel beads are sufficiently washed with distilled water to remove the calcium chloride aqueous solution.
Twenty-fold equivalent of distilled water (400 times the ovalbumin powder used)
ml) as a reaction solution, and the temperature of distilled water is adjusted to 35 ° C. using a thermostatic shaking incubator. Then, 1- (4-bromophenyl) ethanol,
1- (4-chlorophenyl) ethanol, 1-phenylethanol, 1-phenylpropanol, 1-phenylbutanol, 1- (4-methoxyphenyl) ethanol, 1- (4-nitrophenyl) ethanol, 1- (2
-Naphthyl) ethanol was added, and each was set to the condition of a shaking incubator at 55 rpm to convert the substrate.

After the reaction is completed, in a fourth step, the beads and the reaction solvent are roughly separated, and the beads are sufficiently washed with a solvent such as distilled water. The solvent portion was extracted with diethyl ether. Further, the ether layer was washed with a saturated saline solution, dehydrated and dried with sodium sulfate, and allowed to stand. Finally, in a fifth step, the diethyl ether layer is removed using an evaporator, and the reaction substrate and the reaction product are removed by 70%.
The desired optically active alcohol was isolated and purified using a silica gel chromatograph of ２３０230 mesh with a developing solvent of hexane to ethyl acetate 9: 1.

The isolated optically active alcohol is described in the literature as J. CHEM. SOC. PERKIN TRANS 11995 pp. 1295-1298.
And Phytochemistry, Vol. 30, No. 11, pp. 3595-3597
The configuration can be determined from a comparison between the (+ or-) value obtained with reference to the optical rotation of the obtained optically active alcohol,
Analysis conditions for high performance liquid chromatography (HPLC), Chiral Cell OB 0.46 cmφ × 25 cm (manufactured by Daicel Chemical Industries, Ltd.): 30 °, UV 254 nm, eluent: hexane: 2-propanol = 9: 1, flow rate 0.5 ml / min. Depending on the substrate (±) -1- (4-bromophenyl) ethanol, (±) -1- (4-chlorophenyl) ethanol, (±) -1-phenylethanol, (±) -1-
Phenylpropanol, (±) -1-phenylbutanol, (±) -1- (2-naphthyl) ethanol,
Chiral cell OB 0.46 cmφ × 25 cm (manufactured by Daicel Chemical Industries, Ltd.): 30 °, UV 254 nm, solvent: hexane: 2-propanol = 9: 1, flow rate 1.0 ml /
Substrate (±) -1- (4-methoxyphenyl)
The retention time of the configuration S-form and R-form of ethanol and (±) -1- (4-nitrophenyl) ethanol can be confirmed, and the difference in the integration ratio between the S-form and the R-form enantiomer appearing in HPLC can be determined optically. Purity (ee = enantiomer excess)
).

In the above instrumental analysis, (±) -1- (4-
Retention time of bromophenyl) ethanol, S
Body; 10.47, R form; 11.031, (±) -1-
Retention time of (4-chlorophenyl) ethanol, S-form; 9.936, R-form; 10.355, (±)-
1-phenylethanol retention time, S-form;
11.958, R form; 13.133, retention time of (±) -1-phenylpropanol, S form 8.19
0, R-form 8.915, (±) -1-phenylbutanol, retention time of (±) -1- (2-naphthyl) ethanol, S-form; 15.693, R-form; 17.04
9, retention time of (±) -1- (4-methoxyphenyl) ethanol, S-form; 9.165, R-form; 1
0.781, retention time of (±) -1- (4-nitrophenyl) ethanol, R form; 18.923, S
Body: 19.562 respectively. Also, 4-bromoacetophenone, 4-chloroacetophenone, acetophenone, propiophenone, butyrophenone, 4-methoxyacetophenone, and 4-bromoacetophenone formed by oxidation of each substrate
12. retention time of nitroacetophenone;
112, 10.304, 16.763, 16.845,
10.709, 17.169, and 37.208 were similarly confirmed.

A. (R) -1- (4-bromophenyl)
Synthesis of ethanol The biochemical conversion reaction of immobilized ovalbumin to (±) -1- (4-bromophenyl) ethanol (200 mg) is as follows (S) -1- (4-bromophenyl)
Via biotransformation to 4-bromoacetophenone following the stereoselective oxidation of ethanol, it takes 24 hours, yielding 54 mg and 27% yield of (R) -1- (4-bromo). Phenyl) ethanol was obtained. 86% optical purity
e. e. Met. GC conditions are HITACHI G-3
500 gas chromatograph, carrier gas, He
0.48 ml / min; spirit ratio; 1/55; oven temperature; 150 ° C., inlet temperature; 250 ° C., outlet temperature;
0 ° C., pressure. 136 kPa, flow rate value. 42 ml / min, analytical column: TC-5HT 0.25 mmI. D × 30M
The reaction tracking and the time at the end of the reaction were determined by df (GL Science Co., Ltd.).

B. (R) -1- (4-chlorophenyl)
Synthesis of Ethanol The biochemical conversion reaction of immobilized ovalbumin on the substrate (±) -1- (4-chlorophenyl) ethanol (200 mg) is as follows: (S) -1- (4-chlorophenyl) ethanol is stereoselective. 24 hours via biotransformation to 4-chloroacetophenone with strong oxidation,
In a yield of 52 mg, (R) -1- (4-
(Bromophenyl) ethanol was obtained. Optical purity is 9
6% e. e. Met.

C. Synthesis of (R) -1-phenylethanol The biochemical conversion reaction of immobilized ovalbumin on the substrate (±) -1-phenylethanol (200 mg) is as follows. Via bioconversion to acetophenone following oxidation, 48
It takes time and the yield is 82 mg and the (R) is 41%.
-1-Phenylethanol was obtained. Optical purity is 9
3.2% e. e. Met.

D. Synthesis of (R) -1-phenylpropanol The biochemical conversion reaction of immobilized ovalbumin on the substrate (±) -1-phenylpropanol (200 mg) is as follows. Via bioconversion to propiophenone following oxidation, 150 hours were required, yielding 90 mg and (R) -1-phenylpropanol in 45% yield. The optical purity is 90.2% e. e. Met.

E. Synthesis of (R) -1-phenylbutanol The biochemical conversion reaction of immobilized ovalbumin on the substrate (±) -1-phenylbutanol (200 mg) is as follows. Via biotransformation to butyrophenone following oxidation, 17
It took 8 hours to obtain (R) -1-phenylbutanol in a yield of 96 mg in a yield of 48%. The optical purity is 90.4% e. e. Met.

F. Synthesis of (R) -1- (4-methoxyphenyl) ethanol The biochemical conversion reaction of immobilized ovalbumin on the substrate (±) -1- (4-methoxyphenyl) ethanol (200 mg) is as follows (S) Via bioconversion to 4-methoxyacetophenone following the stereoselective oxidation of -1- (4-methoxyphenyl) ethanol, it takes 24 hours, yields 52 mg, 26% yield (R ) -1-
(4-Methoxyphenyl) ethanol was obtained. Optical purity is 100% e. e. Met. The HPLC condition was a flow rate of 1.0 ml / min, and the GC condition was an oven temperature of 19 ml / min.
It was set to 0 ° C.

G. (S) -1- (4-nitrophenyl)
Synthesis of Ethanol The biochemical conversion reaction of immobilized ovalbumin on the substrate (±) -1- (4-nitrophenyl) ethanol (200 ml) is as follows: (R) -1- (4-nitrophenyl) ethanol Via biotransformation to 4-nitroacetophenone with selective oxidation, it takes 3 days, yield 5
At 0 mg, (S) -1- (4-nitrophenyl) ethanol was obtained in 25% yield. Optical purity is 79%
e. e. Met. HPLC conditions were flow rate 0.5 ml /
The GC conditions were set to 190 ° C. for the oven temperature.

H. Synthesis of (R) -1- (2-naphthyl) ethanol The biochemical conversion reaction of immobilized ovalbumin with respect to the substrate 1- (2-naphthyl) ethanol (201 mg) is as follows: (S) -1- (2- Naphthyl) via bioconversion to 2-acetonaphthone with stereoselective oxidation of ethanol, requiring 24 hours, 48 mg, 24% yield of (R) -1- (2-naphthyl). 85) ethanol
% E. e. Obtained with an optical purity of.

Next, the results of the synthesis of the above optically active alcohols using immobilized ovalbumin as an optical resolution catalyst are shown in the following table.

[Table 1]

From the above, it has been found that ovalbumin is effective as an optical resolution catalyst for synthesizing a synthetic intermediate in the field of fine chemicals, and that a highly pure R-form or S-form optically active alcohol can be obtained safely and easily.

Example 2 (Efficacy of Continuous Recycling of Immobilized Ovalbumin) The first effectiveness in the second to fifth steps using the ovalbumin powder prepared in the first step as in Example 1 As for the properties, it is effective as an optical resolution catalyst that can be sufficiently used as a synthetic intermediate in the field of fine chemicals,
It has been found that high-purity R- or S-form optically active alcohols can be obtained safely and easily. Example 2 describes results on the effectiveness of continuous reuse of immobilized ovalbumin.

A. (R) -1- (4-bromophenyl)
Synthesis of Ethanol The biochemical conversion reaction of immobilized ovalbumin on the substrate (±) -1- (4-bromophenyl) ethanol (200 mg) is as follows: (S) -1- (4-bromophenyl) ethanol Via biotransformation to 4-bromoacetophenone with selective oxidation, takes 24 hours,
Yield 54 mg, yield 27%, optical purity, 86-91%
e. e. Gave (R) -1- (4-bromophenyl) ethanol. Further, the same reaction was examined again from the third step using immobilized ovalbumin used in the fourth step with the substrate (±) -1- (4-bromophenyl) ethanol. Optical purity by reaction time, 8
6-91% e. e. (R) -1- (4-bromophenyl) ethanol was biosynthesized. Furthermore, as a result of trying the same reaction for the third time, 86 to 91% in a reaction time of 20 hours
e. e. (R) -1- (4-bromophenyl) ethanol was biosynthesized. Furthermore, as a result of trying the same reaction for the fourth time, 86 to 91% e. e. (R) -1- (4-bromophenyl) ethanol was biosynthesized.

[0036]

[Table 2]

B. (R) -1- (4-chlorophenyl)
Synthesis of Ethanol The biochemical conversion reaction of immobilized ovalbumin on the substrate (±) -1- (4-chlorophenyl) ethanol (200 mg) is as follows: (S) -1- (4-chlorophenyl) ethanol is stereoselective. 24 hours via biotransformation to 4-chloroacetophenone with strong oxidation,
54 mg, 27% yield, 93-96% optical purity e.
e. Gave (R) -1- (4-chlorophenyl) ethanol. Further, the immobilized ovalbumin used in the fourth step is replaced with the substrate (±) -1- (4-chlorophenyl)
As a result of examining the same reaction again from the third step using ethanol, the optical purity was found to be 93 to 93 after a reaction time of 22 hours.
96% e. e. (R) -1- (4-chlorophenyl)
Ethanol was biosynthesized. Furthermore, as a result of trying the same reaction for the third time, the optical purity was 93 to 9 with a reaction time of 20 hours.
6% e. e. (R) -1- (4-chlorophenyl) ethanol was biosynthesized. Furthermore, as a result of trying the same reaction for the fourth time, the optical purity was 93 to 96 with a reaction time of 18 hours.
% E. e. (R) -1- (4-chlorophenyl) ethanol was biosynthesized.

[0038]

[Table 3]

C. Synthesis of (R) -1-phenylethanol The biochemical conversion reaction of immobilized ovalbumin on the substrate (±) -1-phenylethanol (200 mg) is as follows. Via bioconversion to acetophenone following oxidation, 48
It takes time, yield 82 mg, yield 41%, optical purity 93
~ 95% e. e. Thus, (R) -1-phenylethanol was obtained. Furthermore, the same reaction was examined again from the third step using immobilized ovalbumin used in the fourth step with the substrate (±) -1-phenylethanol.
Optical purity 93-95% with a reaction time of 4 hours e. e. (R) -1-phenylethanol was biosynthesized. Furthermore,
As a result of trying the same reaction for the third time, the optical purity was 93 to 95% e.g. with a reaction time of 22 hours. e. (R) -1-phenylethanol was biosynthesized. Furthermore, as a result of trying the same reaction for the fourth time, the optical purity was 93 to 9 with a reaction time of 20 hours.
5% e. e. (R) -1-phenylethanol was biosynthesized.

[0040]

[Table 4]

D. Biosynthesis of (R) -1-phenylpropanol The biochemical conversion reaction of immobilized ovalbumin on the substrate (±) -1-phenylpropanol (200 mg) is as follows. It took 24 hours via biotransformation to 4-propiophenone following the oxidation, yielding 54 mg, yield 27%, optical purity 86-91% e. e. Gave (R) -1-phenylpropanol. Further, the same reaction was repeated from the third step using immobilized ovalbumin used in the fourth step with the substrate (±) -1-phenylpropanol. 86-91
% E. e. (R) -1-phenylpropanol was biosynthesized. Further, as a result of trying the same reaction for the third time, 20
86-91% optical purity with a reaction time of e. e. (R) -1-phenylpropanol was biosynthesized. Furthermore, as a result of trying the same reaction for the fourth time, the optical purity was 86 to 91% e. e. (R) -1-phenylpropanol was biosynthesized.

[0042]

[Table 5]

E. Biosynthesis of (R) -1-phenylbutanol The biochemical conversion reaction of immobilized ovalbumin on the substrate (±) -1-phenylbutanol (200 mg) is as follows. Via the biotransformation to 4-butyrophenone following the strong oxidation,
178 hours required, yield 96 mg, yield 48%, optical purity 88-91% e. e. Thus, (R) -1-phenylbutanol was obtained. Further, the same reaction was examined again from the third step using the substrate (±) -1-phenylbutanol for the immobilized ovalbumin used in the fourth step. 88-91% e.
e. (R) -1-phenylbutanol was biosynthesized.
Furthermore, as a result of trying the same reaction for the third time, the optical purity was 88 to 91% e. e. (R) -1
-Phenylbutanol was biosynthesized. Further, as a result of trying the same reaction for the fourth time, the optical purity was 88 to 91% e. e. (R) -1-phenylbutanol was biosynthesized.

[0044]

[Table 6]

F. Biosynthesis of (R) -1- (4-methoxyphenyl) ethanol The biochemical conversion reaction of immobilized ovalbumin on the substrate (±) -1- (4-methoxyphenyl) ethanol (200 mg) is as follows (S ) Via biotransformation of 4- (4-methoxyphenyl) ethanol to 4-methoxyacetophenone following stereoselective oxidation, it takes 24 hours, yield 52 mg, yield 26%, optical purity 100%.
e. e. Gave (R) -1- (4-methoxyphenyl) ethanol. Furthermore, the same reaction was again performed from the third step using the substrate (±) -1- (4-methoxyphenyl) ethanol for the immobilized ovalbumin used in the fourth step. Optical purity 100% e. e. (R) -1- (4-methoxyphenyl) ethanol was biosynthesized. Further, as a result of trying the same reaction for the third time, an optical purity of 100 was obtained in a reaction time of 20 hours.
% E. e. (R) -1- (4-methoxyphenyl) ethanol was biosynthesized. Furthermore, as a result of trying the same reaction for the fourth time, the optical purity was 100% e.
e. (R) -1- (4-methoxyphenyl) ethanol was biosynthesized.

[0046]

[Table 7]

G. Biosynthesis of (R) -1- (2-naphthyl) ethanol The biochemical conversion reaction of immobilized ovalbumin on the substrate (±) -1- (2-naphthyl) ethanol (200 mg) is as follows: Via biotransformation of 2- (1-naphthyl) ethanol to 2-acetonaphthone following the stereoselective oxidation, it took 24 hours, yield 48 mg, yield 24%, optical purity 85% e. e. And (R) -1- (2-
Naphthyl) ethanol was obtained. Further, the immobilized ovalbumin used in the fourth step is replaced with the substrate (±) -1-
The same reaction was examined again from the third step using (2-naphthyl) ethanol, and as a result, the optical purity was 85% e.g. in a reaction time of 22 hours. e. (R) -1- (2-naphthyl) ethanol was biosynthesized. Furthermore, as a result of trying the same reaction for the third time, the optical purity was 85% with a reaction time of 20 hours.
e. e. (R) -1- (2-naphthyl) ethanol was biosynthesized. Furthermore, as a result of trying the same reaction for the fourth time,
85% optical purity with a reaction time of 18 hours e. e. (R)
-1- (2-Naphthyl) ethanol was biosynthesized.

[0048]

[Table 8]

Example 3 Synthesis of (R) -1- (4-methoxyphenyl) ethanol by Asymmetric Reduction from 4-Methoxyacetophenone Example 1 was performed using ovalbumin powder obtained in the same manner as in Example 1. Under the same reaction conditions as described above, 4-methoxyacetophenone was converted into a substrate. The reaction was performed for 8 days. Yield 5
%, Optical purity 90% e. e. Thus, (R) -1- (4-methoxyphenyl) ethanol was obtained.

Example 4 (R) -1- by asymmetric hydrolysis of an acetylated form of 1- (4-methoxyphenyl) ethanol in a benzene solvent
Example 1 Synthesis of (4-methoxyphenyl) ethanol Using ovalbumin powder obtained in the same manner as in Example 1, Example 1 was used.
Under the same reaction conditions as described above, the acetylated form of 1- (4-methoxyphenyl) ethanol was asymmetrically hydrolyzed for 4 days.
(R) -1- (4-methoxyphenyl) ethanol yield 3%, optical purity 93% e. e. Was obtained. Example 5 Acetate of 1- (4-methoxyphenyl) ethanol formed was subjected to acid treatment and hydrolyzed by asymmetric esterification of 1- (4-methoxyphenyl) ethanol and vinyl acetate in a benzene solvent. (S) -1- (4-
Synthesis of methoxyphenyl) ethanol Using ovalbumin powder obtained in the same manner as in Example 1, 1- (4-methoxyphenyl) ethanol was converted into a substrate under the same reaction conditions as in Example 1, and the resultant was dried in dry benzene. And vinyl acetate for 4 days. The obtained 1-
The hydrolysis compound of the acetate form of (4-methoxyphenyl) ethanol is (S) -1- (4-methoxyphenyl) ethanol, the yield is 3%, and the optical purity is 93.
% E. e. Met.

[0051]

Industrial Applicability According to the present invention, a highly usable raw material for synthesizing synthetic intermediates in the field of fine chemicals by performing an enzymatic conversion reaction of a substrate as a raw material using egg white or ovalbumin derived from egg white as a catalyst. An R- or S-form optically active alcohol having a high purity can be obtained safely and easily.

[Brief description of the drawings]

FIG. 1 shows the use of the immobilized ovalbumin of the present invention for the racemic substrate 1- (2-naphthyl) ethanol (S) -1.
According to the production of (R) -1- (2-naphthyl) ethanol (R-1) via bioconversion to 2-acetonaphthone accompanying stereoselective oxidation of-(2-naphthyl) ethanol (S-1). It is a graph which shows the relationship between a reaction time and a production conversion rate.

FIG. 2 shows the effectiveness when the immobilized ovalbumin of the present invention is used continuously, wherein (a), (b) and (c) show the following (a), (b) and ( It is a graph which shows the relationship between each reaction time of the 1st-3rd time when the reaction of c) was performed, and the production conversion rate. (A) via the bioconversion of the racemic substrate 1- (4-bromophenyl) ethanol to 4-bromoacetophenone following the stereoselective oxidation of (S) -1- (4-bromophenyl) ethanol -1- (4-bromophenyl)
Production reaction of ethanol [(R) -1a] (b) Substrate racemic 1- (4-chlorophenyl) ethanol to 4-chloroacetophenone accompanying stereoselective oxidation of (S) -1- (4-chlorophenyl) ethanol (R) -1- (4-chlorophenyl) via bioconversion of
Production reaction of ethanol [(R) -2a] (c) 4-methoxy accompanying stereoselective oxidation of racemic substrate 1- (4-methoxyphenyl) ethanol to (S) -1- (4-methoxyphenyl) ethanol Formation reaction of (R) -1- (4-methoxyphenyl) ethanol [(R) -3a] via bioconversion to acetophenone

Continued on the front page (51) Int.Cl. ⁷ Identification symbol FI Theme coat II (reference) C07C 43/23 C07C 43/23 C 201/16 201/16 205/19 205/19 C12N 11/04 C12N 11/04 C12P 41/00 C12P 41/00 CF // C07B 57/00 343 C07B 57/00 343 61/00 300 61/00 300 F term (reference) 4B033 NA01 NA42 NB48 NB62 NC06 ND02 ND03 4B064 AC17 AD64 CA21 CB03 CB13 CB18 CB25 CD05 CD06 CD27 DA01 DA11 DA16 4H006 AA02 AC83 AD16 BA81 FC52 FC54 FE11 GN08 GP03 4H039 CA60 CA62 CA90 CJ20

Claims (6)

[Claims] 1. a first step of powdering a protein selected from egg white and ovalbumin separated from egg white;
A second step of immobilizing the protein, a third step of performing enzymatic conversion of a substrate as a raw material using the protein as a catalyst in a non-polar solvent or a polar solvent, and a reaction mixture converted in the third step. A fourth step of extracting with an organic solvent, a fifth step of isolating and purifying the optically active alcohol or an acylated product of the optically active alcohol from the extract in the fourth step, and, if necessary, a hydrolysis step. A method for producing an optically active alcohol. 2. The method for producing an optically active alcohol according to claim 1, wherein the protein is ovalbumin. 3. The method for producing an optically active alcohol according to claim 1, wherein the substrate for the enzymatic conversion in the third step is a racemic alcohol, and one of its enantiomers is selectively oxidized. 4. The method for producing an optically active alcohol according to claim 1, wherein the substrate for the enzymatic conversion in the third step is a ketone, which is obtained by an asymmetric reduction reaction. 5. The method for producing an optically active alcohol according to claim 1, wherein the substrate for the enzymatic conversion in the third step is an acylated form of a racemic alcohol and is subjected to an asymmetric hydrolysis reaction. 6. The enzymatic conversion in the third step is an asymmetric acylation reaction of the racemic substrate alcohol with an acyl halide or vinyl acetate, and the extract in the fourth step is an acylated form of an optically active alcohol, The method for producing an optically active alcohol according to claim 1, further comprising a hydrolysis step after the fifth step.