JP2002253294A - Method for producing optically active aliphatic amino acid amide - Google Patents

Abstract

PROBLEM TO BE SOLVED: To provide an industrial method for producing an (S)-α-amino acid amide. SOLUTION: A live microbial cell or a treated substance of the live microbial cell of a microorganism having an activity for stereoselectively hydrolyzing an (R)-amino acid amide is made to act on a racemic mixture of α-amino acid amides to produce an (R)-α-amino acid. The (S)-α-amino acid amide is left unreacted and the (R)-α-amino acid is then derivatized to a racemic mixture of the α-amino acid amides and reused for stereoselective hydrolysis.

Description

DETAILED DESCRIPTION OF THE INVENTION

[0001]

The present invention relates to a method for producing an optically active aliphatic α-amino acid amide represented by the formula (1). Specifically, the asymmetric hydrolysis of a racemic mixture of α-amino acid amides is biochemically asymmetrically hydrolyzed to produce the corresponding (R) -α-amino acid, leaving the (S) -α-amino acid amide unreacted. Characteristic optically active aliphatic (S) -α
-It relates to a process for producing amino acid amides. More specifically, α
Asymmetric hydrolysis of a racemic mixture of amino acid amides to produce the corresponding (R) -α-amino acid,
A method for producing an optically active aliphatic (S) -α-amino acid amide, which comprises leaving (S) -α-amino acid amide unreacted, wherein the (R) -α-amino acid produced is α-amino acid.
The present invention relates to a method for producing an optically active aliphatic (S) -α-amino acid amide which is derived into a racemic mixture of amino acid amides and is used again as a raw material. Optically active aliphatic (S) -α-amino acid amide is an important substance as an intermediate for producing various industrial chemicals, agricultural chemicals, and pharmaceuticals.

[0002]

2. Description of the Related Art As a method for producing an optically active aliphatic (S) -α-amino acid amide, an (S) -α-amino acid is used as a starting material, esterified by a known method, and further produced by amidation ( J. Med. Chem., 14, 484.
(1971)) is known. However, in this method, even after the esterification and amidation reaction, unreacted amino acids or amino acids generated by hydrolysis of the ester formed once remain in a low yield, and under the esterification reaction conditions and the amidation reaction, Since the ester and the amide are partially racemized under the reaction conditions, the obtained optically active aliphatic (S) -α-amino acid amide has a low optical purity and cannot be said to be a method which is necessarily satisfactory in terms of yield and quality. . In addition, asymmetric hydrolysis is carried out biochemically from a racemic mixture of α-aminonitrile to obtain the corresponding optically active α-aminonitrile.
There is a method for producing an amino acid and / or an α-aminoamide (JP-A-2-31694). However, since this method is strongly affected by substrate inhibition of a nitrile compound, a reaction substrate for performing a biochemical asymmetric hydrolysis reaction is used. It is difficult to increase the concentration of the racemic mixture of α-aminonitrile
It is not an industrially advantageous method.

[0003]

SUMMARY OF THE INVENTION An object of the present invention is to solve the above-mentioned problems in the prior art and to provide an optically active aliphatic (S)-which is important as an intermediate for the production of various industrial chemicals, agricultural chemicals and pharmaceuticals. An object of the present invention is to provide a production method for producing α-amino acid amide with high quality and at low cost.

[0004]

Means for Solving the Problems The present inventors have conducted intensive studies on a production method for producing an optically active aliphatic (S) -α-amino acid amide at a high quality and at a low cost.
A racemic mixture of α-amino acid amides is biochemically asymmetrically hydrolyzed to produce (R) -α-amino acids, and (S)
By leaving the -α-amino acid amide unreacted and deriving the resulting (R) -α-amino acid into a racemic mixture of α-amino acid amides and using it again as a raw material,
The present inventors have found that an optically active aliphatic (S) -α-amino acid amide can be produced with high quality and at low cost, and have reached the present invention. The starting material used in the present invention does not need to be an expensive optically active substance, but is an inexpensive racemic mixture of α-amino acid amide, and (R) -α formed by asymmetric hydrolysis.
-Deriving the amino acid into a racemic mixture of α-amino acid amide and using it again as a raw material allows the entire amount of the racemic mixture of α-amino acid amide to be converted to the optically active aliphatic (S)
According to the present invention, it can be converted into an optically active aliphatic (S) -α-amino acid at very low cost because it can be converted to an α-amino acid amide.
It is possible to produce amino acid amides.

[0005] The reaction for obtaining (S) -α-amino acid amide from a racemic mixture of α-amino acid amide is only one step of a biochemical asymmetric hydrolysis reaction. (S) obtained by setting the conversion of amino acid amide to (R) -α-amino acid to 100%.
The optical purity of -α-amino acid amide is 100%, and it is possible to produce very high quality optically active aliphatic (S) -α-amino acid amide in terms of quality.

That is, the present invention relates to a method for producing a racemic mixture of an α-amino acid amide represented by the formula (1):
Acting a viable cell of a microorganism having an activity of stereoselectively hydrolyzing an amino acid amide or a processed product of the viable cell,
(R) -α-amino acid amide represented by the chemical formula (2), and (S) -α-amino acid amide is left unreacted. This is a method for producing an optically active aliphatic (S) -α-amino acid amide, in which the (R) -α-amino acid to be produced is derived into a racemic mixture of α-amino acid amides and used again as a raw material. .

[0007]

BEST MODE FOR CARRYING OUT THE INVENTION In the general formula (1) representing α-amino acid amide as a raw material of the present invention, R is a lower alkyl group,
A substituted lower alkyl group, a cyclohexyl group, and a substituted cyclohexyl group. The lower alkyl group is not particularly limited, but is preferably, for example, a linear or branched lower alkyl group having 1 to 4 carbon atoms such as methyl, ethyl, propyl, isopropyl, butyl, isobutyl, secbutyl and tbutyl. is there. The substituent contained in each of the substituted lower alkyl group and the substituted cyclohexyl group is, for example, hydroxy, methoxy, mercapto, methylmercapto, amino, carboxy, halogen and the like.

The racemic mixture of the α-amino acid amide represented by the chemical formula (1) used in the present invention is not particularly limited in its production method, quality, etc., for example, when α-aminonitrile easily obtained by the Strecker method is used. It can be obtained by a method of hydrolysis or the like. As a method for partially hydrolyzing α-aminonitrile, a method for partially hydrolyzing α-aminonitrile in an aqueous medium in the presence of a basic substance and a ketone, for example, JP-B-58-1774
The method described in JP-A No. 1 is preferred. This method, for example,
Using a basic substance such as an inorganic base such as sodium hydroxide and potassium hydroxide, and an organic base such as an organic quaternary ammonium compound such as tetramethylammonium hydroxide, the pH of the reaction solution should be higher than 14. Then, ketones such as acetone, methyl ethyl ketone, diethyl ketone, methyl isopropyl ketone and cyclohexanone are added to the reaction system, and the reaction temperature is maintained at 40 ° C. or lower, and the pH of the reaction solution is maintained at a pH exceeding 14. While performing the hydrolysis reaction. The crude α-aminonitrile hydrolysis reaction solution is concentrated to remove ketones, and the crude α-amino acid amide-containing solution obtained is purified as it is or by recrystallization or the like as necessary. , Can be used as a raw material. In addition, inorganic acids such as hydrochloric acid and sulfuric acid,
Even a salt with an organic acid such as acetic acid and oxalic acid can be used.

The microorganism used for the biochemical asymmetric hydrolysis of the α-amino acid amide of the present invention comprises a stereoselectively hydrolyzing (R) -α-amino acid amide corresponding to (R) -α-amino acid. Any microorganism may be used as long as it has the activity of performing the activity described above. Examples of such microorganisms include microorganisms belonging to the genera Rhodococcus, Pseudomonas, Ochrobactrum, and Serratia, and specifically, Rhodococcus erythropolis (Rhodococcus erythropolis).
s) NR23 (FERM P-8937), Rhodococcus erythropolis (Rhodococcus ery)
Tropolis) NR28 (FERM P893)
8), Rhodococcus erythropolis (Rhodoco
ccus erythropolis) JCM320
1. Pseudomonas florescens (Pseudo
monas fluorescens) IFO1205
5. Ochrobactrum anthropi
trumanthropi ATCC49237, Serratia marce sense
scens) IAM12143, but is not limited thereto. Further, any strain such as a mutant strain derived from these microorganisms by an artificial mutation means or a recombinant strain derived by a genetic technique such as cell fusion or gene recombination has the above ability. Anything can be used in the present invention.

[0010] Culture of these microorganisms is usually carried out using a medium containing assimilable carbon sources and nitrogen sources, inorganic salts essential for each microorganism, nutrients, and the like. It is also effective to add α-amino acid amide to the medium in advance. The α-amino acid amide to be added at this time is preferably an intended α-amino acid amide, but may be a general α-amino acid amide such as alanine amide or valinamide, and is not particularly limited.
The pH during the culture is in the range of 4 to 10, and the temperature is 20 to
50 ° C. Culture is performed aerobically for about one day to one week. The microorganisms cultured in this manner are viable cells or processed viable cells, such as a culture solution, isolated cells, crushed cells,
Further, it is used in the reaction as a purified enzyme. In addition, cells or enzymes can be immobilized and used according to a conventional method.

The conditions for the biochemical hydrolysis of α-amino acid amide are as follows: α-amino acid amide concentration: 1 to 40% by weight;
The amount of the microorganism used relative to the α-amino acid amide is 0.0001-3 by weight as a dry cell, and the reaction temperature is 20-70.
C, pH 5-13.

(S)-which has not reacted with (R) -α-amino acid formed by the biochemical hydrolysis of α-amino acid amide.
The α-amino acid amide is obtained by removing the microbial cells from the reaction-completed solution by a conventional solid-liquid separation means such as centrifugation or a filtration membrane, concentrating the mixture under reduced pressure, and adding an organic solvent to obtain (R)-.
An α-amino acid is precipitated, and the precipitated (R) -α-amino acid is collected by filtration, or after removing the microbial cells, water is removed under reduced pressure, and an organic solvent is added to the residual solid to remove the microorganism. The (S) -α-amino acid amide in the reaction can be dissolved and the insoluble (R) -α-amino acid can be easily separated by filtration. At this time, to precipitate the (R) -α-amino acid or unreacted (S) -α
The organic solvent added to dissolve the amino acid amide may be any solvent having low solubility of (R) -α-amino acid and high solubility of unreacted (S) -α-amino acid amide, and is not particularly limited. However, alcohols such as ethanol, 2-propanol, 2-methyl-1-propanol, 1-butanol and 2-butanol are preferably used. Further, unreacted (S) -α-amino acid amide can be separated from the reaction-terminated liquid from which the microbial cells have been separated by a method such as solvent extraction. At this time, as a solvent for extracting (S) -α-amino acid amide, for example, hexane,
Non-polar solvents such as benzene, toluene, and xylene. In addition, a method in which only amino acids to be removed are dialyzed and separated using ion exchange electrodialysis, or a separation method utilizing adsorption and desorption by an ion exchange column is also effective.

The isolated (S) -α-amino acid amide is
It can also be isolated as a salt with an acid. The kind of the acid at this time may be an inorganic acid such as hydrochloric acid or sulfuric acid, or an organic acid such as acetic acid or oxalic acid. There is no particular limitation, but hydrochloric acid is preferably used. α-amino acid amide is itself basic and hygroscopic, has low stability and is susceptible to decomposition during storage, but by being isolated as a salt with an acid,
Stability can be greatly improved.

The separated (R) -α-amino acid is esterified, aminated, and further racemized to obtain a racemic mixture of α-amino acid amides, which is used again as a raw material. The esterification can be carried out by a method known per se, for example, by heating with an alcohol under acidic conditions. The type of acid used for esterification is hydrochloric acid,
An inorganic acid such as sulfuric acid or an organic acid such as benzenesulfonic acid may be used. There is no particular limitation, but sulfuric acid is particularly preferably used. The alcohol used for the esterification is not particularly limited, but an aliphatic primary alcohol having 1 to 4 carbon atoms is preferably used, and methanol is particularly preferable. Esterification conditions can be performed within a range usually performed, and there is no particular limitation.
It can be carried out favorably by heating at 50 to 120 ° C. in a molar amount of alcohol in the presence of 2 to 5 equivalents of an acid based on (R) -α-amino acid. In addition, a method of removing water generated in the esterification reaction to the outside of the reaction system to further improve the yield is suitably performed. The amidation of the ester can also be carried out in a manner known per se, for example by reaction with ammonia. The ammonia used may be liquid ammonia or aqueous ammonia, but usually 10 to 30% aqueous ammonia is preferably used. The amidation reaction can be carried out satisfactorily by, for example, reacting at 20 to 50 ° C. in 25% aqueous ammonia containing 2 to 20 moles of ammonia with respect to the ester. The (R) -α-amino acid amide thus obtained can be easily racemized by heating in the presence of a strongly basic substance to obtain a racemic mixture of α-amino acid amide.

The strongly basic substance used in the racemization reaction is
Any organic or inorganic strong basic substance may be used. Representative examples include organic quaternary ammonium compounds such as tetramethylammonium hydroxide and tetraethylammonium hydroxide, and sodium hydroxide, potassium hydroxide, sodium methylate and sodium ethylate. And alkali metal compounds such as sodium amide and sodium hydride, and alkaline earth metal compounds such as barium hydroxide. In the reaction system, it is also possible to add a substance which can be converted into the above-mentioned strong basic substance, for example, an alkali metal simple substance such as sodium and potassium, and an alkaline earth metal simple substance such as barium. The amount of these strong basic substances used is
0.001 to 1 mol of (R) -α-amino acid amide
To 0.5 mol, preferably 0.01 to 0.1 mol.
It is a mole ratio. The racemization reaction can be carried out without using a solvent.However, when a solvent is used, the reaction temperature can be lowered, so that the risk of by-products being formed can be reduced, which is more preferable. It is. The solvent used at this time may be any solvent as long as it is inert to each of (R) -α-amino acid amide and a strongly basic substance. For example, hexane, heptane, cyclohexane,
Hydrocarbons such as benzene, toluene and xylene, 2
-Propanol, 2-methyl-1-propanol, 2-
Alcohols such as butanol, 1-butanol and 1-pentanol, and isobutyronitrile. The amount of the solvent used is not particularly limited, but practically, (R)
It is not necessary to increase the weight of the α-amino acid amide by more than 100 times, and preferably about 1 to 20 times. It is preferable that the amount of water in the racemization reaction solution is as small as possible. However, there is almost no problem if the water content is about 1 wt% or less, and there is substantially no problem if the water content is 0.1 wt% or less. The temperature of the racemization reaction is 20 to 2
00 ° C, preferably 50 to 150 ° C. The racemization reaction is usually performed under normal pressure, but does not prevent it from being performed under reduced pressure or under increased pressure.

Thus, the (R) -α-amino acid is converted to α
The whole amount of the racemic mixture of α-amino acid amides can be converted to optically active (S) -α-amino acid amides by forming a racemic mixture of amino acid amides and circulating the mixture in a biochemical asymmetric hydrolysis reaction system.

According to the method of the present invention, specifically, for example, (S) -alaninamide, (S) -valinamide,
Optically active aliphatic (S) -α-amino acid amides such as (S) -leucinamide, (S) -isoleucinamide and (S) -2-aminobutyric acid amide can be produced.

[0018]

EXAMPLES The present invention will be described more specifically with reference to examples, but the present invention is not limited to these examples. Example 1 A medium having the following composition was prepared, and 200 ml of this medium was placed in a 1-L Erlenmeyer flask, sterilized, and then subjected to Rhodococcus erythropolis (Rhodococcus erythropolis).
(Poris) NR28 (FERM P-8938), and cultured with shaking at 30 ° C. for 48 hours. Medium composition (pH 7.0) Glucose 10g polypeptone 5g yeast extract _{5g KH 2 PO 4 1g MgSO 4} · 7H 2 O 0.4g FeSO 4 · 7H 2 O 0.01g MnCl 2 · 4H 2 O 0.01g (R) - Valinamide 5 g Water 1 L Next, viable cells were obtained from the culture solution by centrifugation. The viable cells were suspended in 100 ml of water, placed in a 1 L Erlenmeyer flask, and 5.00 g (4%) of a racemic mixture of valinamide was added.
3.1 mmol) and shaken at 40 ° C. for 5 hours.
After the reaction, the reaction solution was centrifuged to obtain a supernatant. After water was distilled off from the supernatant under reduced pressure using an evaporator, 20 ml of 2-methyl-1-propanol was added thereto, and unreacted (S) was added.
-Valinamide was dissolved and the insoluble (R) -valine was filtered off as a white solid. To the filtrate after filtering off valine, 2.25 g (21.6 mmol) of concentrated hydrochloric acid was added, and 2.82 g (1) of a white solid (V) valinamide hydrochloride precipitated.
8.5 mmol) were collected by filtration. The filtrate was concentrated to obtain a second crystal (0.33 g, 2.2 mmol) for a total of 3.15 g (2 mmol).
0.7 mmol) of (S) -valinamide hydrochloride was obtained. The isolation yield from (S) valinamide in the racemic mixture charged to the reaction was 96 mol%, and the isolation yield from valinamide racemic mixture was 48 mol%. The solid was analyzed by liquid chromatography using a chiral column for optical resolution, and as a result, the optical purity was 99%.
e. e. That was all.

2.50 g of (R) -valine formed in the reaction
(21.4 mmol) in 10.0 g of methanol (310
mmol) and concentrated sulfuric acid (3.00 g, 30.0 mm
ol), and the mixture was heated under reflux for 23 hours to carry out an esterification reaction. The reaction solution was cooled with ice, 15.0 g (220 mmol) of 25% aqueous ammonia was added thereto with stirring, and the mixture was added at room temperature for 2 hours.
The mixture was stirred for 3 hours to perform an amidation reaction. After the reaction, 2.40 g (60.0 mmol) of sodium hydroxide was added to dissolve and excess ammonia, methanol and water were distilled off under reduced pressure. Then, 20 ml of 2-methyl-1-propanol was added, and the insoluble solid was filtered off. did. To the filtrate was added 0.08 g (2.00 mmol) of sodium hydroxide, and the mixture was heated at 110 ° C. for 3 hours.
Stirred for 0 minutes and racemized. After completion of the reaction, the reaction solution was cooled, and ammonium chloride was added in an amount of 0.11 g (2.00 mmol).
l) was added to neutralize the mixture, 2methyl 1-propanol was distilled off under reduced pressure, and water was added to obtain an aqueous solution containing 2.10 g (18.1 mmol) of a racemic mixture of partially purified valinamide. The yield from (R) -valine was 85 mol%.

An aqueous solution containing 2.10 g (18.1 mmol) of the thus-recovered racemic mixture of partially purified valinamide, and a new racemic mixture of valinamide 2.1.
Of Rhodococcus erythropolis (Rhodococcus erythropolis).
opoli) NR28 (FERM P8938) was reacted again with live cells obtained from a culture solution at 40 ° C. for 5 hours, treated in the same manner, and 2.28 g of (S) -valinamide hydrochloride as a white solid ( 15.0 mmol) were collected by filtration. The filtrate was concentrated to give a second crystal (0.25 g, 1.6 mm).
ol), a total of 2.53 g (16.6 mmol) of (S)
-Valinamide hydrochloride was obtained. The isolation yield from (S) -valinamide in the racemic mixture charged to the reaction was 92 mol%, and the isolation yield from the racemic mixture of newly charged valinamide was 92 mol%. The solid was analyzed by liquid chromatography using a chiral column for optical resolution, and as a result, the optical purity was 99% e.g. e. That was all.

Example 2 A medium having the following composition was prepared and sterilized, and then Ochrobactrum anthropi (Ochrobactrum a) was prepared.
nthropi) inoculate ATCC49237, 30 ° C
For 18 hours. Medium composition (pH 7.0) Glucose 1 g Tryptone 5 g Yeast extract 5 g K ₂ HPO ₄ 1 g Water 1 L After culture, the culture was centrifuged to obtain viable cells, which were suspended in phosphate buffer (pH 7.0). Was subjected to ultrasonic treatment to disrupt the cells, and the disrupted cells were removed by centrifugation to obtain a cell-free extract. This cell-free extract was subjected to ammonium sulfate fractionation (30 to 60).
%), Anion exchange column chromatography (DEA
E Toyopearl, 0 → 1M NaCl gradient and hydrophobic column chromatography (butyl Toyopearl,
(30 → 0% ammonium sulfate gradient) to obtain an enzyme solution.

Racemic mixture of 2-aminobutyric acid amides5.
Dissolve 00 g (49.0 mmol) in 100 ml of water,
The above purified enzyme solution was added thereto, and the mixture was kept at 30 ° C. for 5 hours. After the reaction, water was distilled off under reduced pressure with an evaporator, and then 20 ml of 2-methyl-1-propanol was added to dissolve unreacted (S) -2-aminobutyric acid amide and to remove insoluble (R) -2- Aminobutyric acid was filtered off as a white solid. 2
-2.56 g of concentrated hydrochloric acid was added to the filtrate after filtering out aminobutyric acid.
(24.5 mmol) was added to precipitate (S) -2-
2.85 g of aminobutyric acid amide hydrochloride white solid (20.
6 mmol) were collected by filtration. The filtrate was concentrated to obtain a second crystal (0.28 g, 2.0 mmol), for a total of 3.13 g (2 mmol).
(2.6 mmol) of (S) -2-aminobutyric acid amide hydrochloride was obtained. (S) -2 in the racemic mixture charged to the reaction
The isolation yield from -aminobutyric amide was 92 mol%, and the isolation yield from a racemic mixture of valinamide was 46 mol%. The solid was analyzed by liquid chromatography using a chiral column for optical resolution, and as a result, the optical purity was 99% e.g. e. That was all.

(R) -2-aminobutyric acid produced by the reaction
40 g (23.3 mmol) of methanol 10 g (31
0 mmol), 3 g (30 mmol) of concentrated sulfuric acid was added, and the mixture was heated under reflux for 23 hours to carry out an esterification reaction. The reaction solution was cooled, 15 g (220 mmol) of 25% aqueous ammonia was added thereto with stirring, and the mixture was stirred at room temperature for 23 hours to perform an amidation reaction. After the reaction, 2.4 g (60 mmol) of sodium hydroxide was added and dissolved, and excess ammonia, methanol and water were distilled off under reduced pressure.
-Methyl-1-propanol (20 ml) was added, and the insoluble solid was filtered off. 0.08 g of sodium hydroxide in the filtrate
(2 mmol) was added and the mixture was stirred at 110 ° C. for 30 minutes to be racemized. After completion of the reaction, the reaction solution was cooled and neutralized by adding 0.11 g (2 mmol) of ammonium chloride. After distilling off 2-methyl-1-propanol under reduced pressure, water was added to the residue to give crude purified 2-amino acid. Racemic mixture of butyric amides
An aqueous solution containing 97 g (19.3 mmol) was obtained.
The yield from (R) -2-aminobutyric acid was 83 mol%.

1.97 g (19.3 mmol) of the racemic mixture of the crude 2-aminobutyric acid amide thus recovered
l) and 1.97 g (19.3 mmol) of a new racemic mixture of 2-aminobutyric acid amide containing
After reacting again at 30 ° C. for 5 hours using the above purified enzyme solution, the same treatment was carried out, and 2.27 g (16.4 mmol) of a white solid of (S) -2-aminobutyric acid amide hydrochloride was collected by filtration. did. The filtrate was concentrated to obtain a second crystal (0.20 g, 1.4 g).
mmol), giving a total of 2.47 g (17.8 mmol) of (S) -2-aminobutyric acid amide hydrochloride. The isolation yield from (S) -2-aminobutyric acid amide in the racemic mixture charged in the reaction was 92 mol%, and the isolation yield from the racemic mixture of freshly charged valinamide was 92 mol%. The solid was analyzed by liquid chromatography using a chiral column for optical resolution, and as a result, the optical purity was 99% e.g. e. That was all.

Example 3 Various microorganisms were cultured in the same manner as in Example 1 to obtain viable cells. 5.00 g of a racemic mixture of leucinamide (38.
The same operation as in Example 1 was performed using 5 mmol) as a substrate and live cells of various microorganisms. Table 1 shows the results. In addition, the optical purity of (S) -leucinamide hydrochloride obtained after the enzymatic reaction was analyzed by liquid chromatography using a chiral column for optical resolution.
9% e. e. That was all.

Table 1 Bacterial name Yield 1 Yield 2 Yield 3 Yield 4 Rhodococcus erythropolis NR-28 (FERM P-8938) 48% 95% 92% 92% Pseudomonas florescens (IFO 12055) 44% 88% 87% 87% Serratia marcescens (IAM 12143) 41% 82% 80% 80% Yield 1: Isolation yield of (S) -leucinamide hydrochloride relative to racemic mixture of leucinamide charged in the first enzymatic reaction ( Mol%). Yield 2: Isolation yield (mol%) of (S) -leucinamide hydrochloride relative to (S) -leucine amide in the racemic mixture charged for the first enzyme reaction. Yield 3: Isolation yield (mol%) of (S) -leucinamide hydrochloride relative to (S) -leucine amide in the racemic mixture charged for the second enzyme reaction. Yield 4: Isolation yield (mol%) of (S) -leucinamide hydrochloride with respect to the racemic mixture of leucinamide newly charged in the second enzyme reaction.

[0027]

According to the method of the present invention, various industrial chemicals,
An optically active aliphatic (S) -α-amino acid amide, which is important as an intermediate for producing agricultural chemicals and pharmaceuticals, can be produced at high quality and at low cost.

──────────────────────────────────────────────────続 き Continued on the front page (51) Int.Cl. ⁷ Identification symbol FI Theme coat ゛ (Reference) (C12P 41/00 (C12P 41/00 A C12R 1: 425) C12R 1: 425) (72) Inventor Kayama Consideration 182 Niigata, Niigata-shi, Niigata-shi Niigata Research Laboratory Mitsubishi Gas Chemical Co., Ltd. (72) Inventor Atsushi Inoue 182 Niigata-shi, Niigata-shi Niigata City Niigata Research Institute 4B064 AE03 CA02 CB05 CD27 DA01

Claims (5)

[Claims] 1. A viable cell of a microorganism having an activity of stereoselectively hydrolyzing (R) -α-amino acid amide in a racemic mixture of α-amino acid amide represented by the general formula (1), or the viable cell The treated substance of the body is allowed to act to produce (R) -α-amino acid represented by the general formula (2), and (S) -α
-Leaving the amino acid amide unreacted, the (R) -α-
A method for producing an optically active aliphatic (S) -α-amino acid amide, comprising deriving an amino acid into a racemic mixture of α-amino acid amide and subjecting it to stereoselective hydrolysis again. Embedded image (R is a lower alkyl group, a substituted lower alkyl group, a cyclohexyl group, or a substituted cyclohexyl group.) (R is a lower alkyl group, a substituted lower alkyl group, a cyclohexyl group, a substituted cyclohexyl group) 2. Derivation of (R) -α-amino acid into a racemic mixture of α-amino acid amide,
The method according to claim 1, wherein the α-amino acid is converted into (R) -α-amino acid ester, and then the (R) -α-amino acid ester is converted into (R) -α-amino acid amide, followed by racemization. 3. The process according to claim 1, wherein the optically active aliphatic (S) -α-amino acid amide is isolated as a salt with an acid. Method 4. The microorganism according to claim 1, wherein the microorganism having the activity of stereoselectively hydrolyzing the (R) -α-amino acid amide is a microorganism belonging to the genus Rhodococcus, Pseudomonas, Ochrobactrum or Serratia. The production method according to any one of the above. 5. The method according to claim 1, wherein R in the general formulas (1) and (2) is an ethyl group.