JPH0947296A - Optical resolution of chlorohydrin by microorganism - Google Patents

Abstract

PROBLEM TO BE SOLVED: To optically resolve the subject compound useful as a raw material for medicines and agrochemicals, physiologically active substances, liquid crystals, etc., by reacting racemic chlorohydrins with a cultured solution, microbial cell (treated material), etc., of a microorganism to decompose either one of optical isomers and leave the other isomer. SOLUTION: A racemic chlorohydrin compound [e.g. racemic 4-chloro-3- hydroxybutylonitrile] of the formula [R<1> is H or a lower alkyl; R' is a (substituted) lower alkyl (excluding hydroxymethyl group) when R<1> is H or R<2> is H when R<1> is a lower alkyl group] is reacted with a culture solution, or microbial cell of a microorganism [e.g. Pseudomonas sp. OS-K-29 (FERM BP-994), etc.] or its treated material. Thereby, either one of optical isomers is decomposed and either one of the optical isomers is left to provide the objective optically active chlorohydrin capable of providing chiral building blocks, intermediates, etc., useful in synthesis of optically active compounds such as medicines, agrochemicals, physiologically active substances and ferroelectric liquid crystals.

Description

Detailed Description of the Invention

[0001]

TECHNICAL FIELD The present invention relates to a method for optically resolving chlorohydrin by microorganisms, and more particularly, to chiral building blocks and intermediates useful in the synthesis of optically active compounds such as pharmaceuticals, agricultural chemicals, physiologically active substances and ferroelectric liquid crystals. The present invention relates to a method for producing an optically active chlorohydrin compound which can be a compound from a corresponding racemate using a microorganism by a biological optical resolution method. The present invention also provides
By the biological optical resolution of such racemic chlorohydrin, the optically active chlorohydrin compound, the optically active chlorohydrin compound, and the optically active 1,2-depending on the kind of the substituent of the chlorohydrin compound of the starting compound are used.
Also provided is a method for producing a diol compound and / or an optically active 3-hydroxy-γ-butyrolactone.

[0002]

BACKGROUND OF THE INVENTION As a method for producing an optically active compound, a chemical synthesis method for converting a corresponding optically active starting material into a target compound, and a corresponding racemic body are treated with an optical resolving agent. Is generally used to separate optically active substances, but recently, a method for separating optically active substances by a biological method utilizing asymmetric reduction or asymmetric hydrolysis reaction using microorganisms or enzymes has been reported. Has been done. For example, an asymmetric reduction method of β-ketoesters using a microbial cell or an enzyme is known, and among them, an optically active 4-chloro-3-ester by asymmetric reduction method of 4-chloroacetoacetate using baker's yeast. An asymmetric reduction method to hydroxybutyrate is known (CJ Sih et al., Ann. NY Ac
ad. Sci., Vol. 434, pp.186-193,1984; CJ Sih, BE8
98386). In addition, E. Santaniello et al., S-form ethyl 4-chloro-from ethyl 4-chloro-3-oxobutyrate by an asymmetric reduction method using baker's yeast.
Reported the manufacturing method of 3-hydroxybutyrate (J.Ch
em. Research (S), pp.132-133, 1984). Takahashi et al. Also reported a method for producing optically active ethyl 4-chloro-3-hydroxybutyrate by asymmetric reduction of ethyl 4-chloro-3-oxobutyrate using a microorganism (JP-A-61-161). 146191). In the manufacturing method using the enzyme,
Peters et al. Used an asymmetric reduction method using carbonyl reductase of Rhodococcus erythropolis to obtain S-form methyl from methyl 3-oxobutyrate or ethyl 4-chloro-3-oxobutyrate. A method for producing 3-hydroxybutyrate and R-form ethyl 4-chloro-3-hydroxybutyrate has been reported (J. Peters et al., Appl. Mi.
crobiol. Biotechnol., Vol. 38, pp.334-340, 1992;
T. Zelinski et al., J. Biotechnol., Vol. 33, pp. 28
3-292, 1994). In addition, Shimizu et al. Sporoboromyces salmonicolor AKU
R-form ethyl 4 using aldehyde reductase of 4429 strain
A production method of -chloro-3-hydroxybutyrate by an asymmetric reduction method has been reported (S. Shimizu et al., Biotech.
nol. Lett., Vol. 12., No. 8, pp.593-596, 1990 ； App
l.Environ. Microbiol., Vol. 56, No. 8, pp.2374-237
7, 1990). However, in the method for producing an optically active β-hydroxy ester form from a prochiral β-keto ester form by an asymmetric reduction method using these microorganisms or enzymes, expensive NADH (nicotinamide adenine dinucleotide) or NADPH (nicotine) is used for the reaction. Amide adenine dinucleotide phosphate) and other coenzymes are required, and further a reaction for converting the oxidant to the reductant is required, so that an enzyme such as glucose oxidase or formate dehydrogenase is separately required and the reaction step It is hard to say that it is an industrial production method because of the rate-limiting reaction.

Nakamura et al. Also used 1,3-dichloro-2-propanol or epichlorohydrin as a substrate in the presence of KCN.
ium) sp. N-1074 strain treated with dehalogenase to give R-form 4-
A method for producing chloro-3-hydroxybutyronitrile has been reported (JP-A-3-53889 and JP-A-3-53889).
53890). In addition, a method for obtaining an optically active 4-chloro-3-hydroxybutyronitrile or a corresponding ester form by allowing lipase to act in the presence of racemic 4-chloro-3-hydroxybutyronitrile and a fatty acid ester has also been reported. (JP-A-2-27995). However, these methods are not suitable for industrial production because the optical purity of the optically active substance obtained is low.

Furthermore, Lee et al. (LGLee, GM Whiteside
s) reports a method for producing R-form 1,2-propanediol and R-form 1,2-butanediol from 1-hydroxy-2-propanone and 1-hydroxy-2-butanone by reduction using glycerol dehydrogenase. (J.Org.Chem., Vol.51, pp.25-36, (198
6)). There is also known a production method in which a microbial cell is allowed to act on a 1-hydroxy-2-ketone compound to convert it into an S-form 1,2-diol (JP-A-1-320988). However, these methods are methods for producing 1,2-diol by utilizing an asymmetric reduction reaction by an enzyme or a microorganism, and only one optically active chlorohydrin is obtained from racemic chlorohydrin by using the microbial cell of the present invention. It is distinguished from the method of converting to a 1,2-diol. Moreover, in addition to the chemical synthetic method, the biological method is used in the presence of vinyl acetate in an organic solvent to produce optically active 3-hydroxy-γ-butyrolactone useful as a synthetic intermediate of an optically active compound such as a drug. At 3-hydroxy-γ
-A method of reacting butyrolactone with lipase to obtain an optically active 3-hydroxy-γ-butyrolactone and an ester compound corresponding thereto in the transesterification reaction (EP04397)
79); R-4- obtained by asymmetrically reducing ethyl 4-t-butoxy-3-oxobutyrate with baker's yeast
Method for deriving R-3-hydroxy-γ-butyrolactone from t-butoxy-3-hydroxybutyrate in the presence of fluoroacetic acid (Synthesis (1), pp.37-40, 19)
86) is known. However, in these methods, the operation is complicated, it is necessary to use an expensive reagent, or an expensive optically active substance is used as a starting material, and it is difficult to obtain it, and only one optical isomer can be produced. Etc. have problems to be solved. Therefore, it cannot be said to be a simple and inexpensive method for producing optically active 3-hydroxy-γ-butyrolactone.

The present inventors have previously proposed a method for producing an optically active form from a corresponding racemate by using a microorganism capable of decomposing and assimilating one optical isomer and using it as a single carbon source for growth and multiplication. I found it. For example, the genus Alcaligenes (Alca
ligenes) S-(+)-3-halogeno-
Production method of 1,2-propanediol (Japanese Patent Laid-Open No. 19917/1993)
95, and J. of Fermentation & Bioengineerin
g, Vol. 73, No. 6, 443-448, 1992); A method for producing R-(-)-3-halogeno-1,2-propanediol using Pseudomonas bacteria (JP-A-3-1).
91794, and Applied Microbiology and Biot
echnology, Vol. 40, 273-278, 1993); Optically active dichloropropanol using alkaline genus bacteria (R
Body) (JP-A-3-180197, and J. of
Industrial Microbiology, Vol. 10, 37-43, 1992);
Process for producing optically active dichloropropanol (S-form) using Pseudomonas bacteria (Japanese Patent Laid-Open No. 61-132196,
And Agric. Biol. Chem., Vol. 54, No. 12, 3185-31
90, 1990); and a method for producing an optically active epichlorohydrin from an optically active dichloropropanol obtained by using the above-mentioned Pseudomonas bacterium by alkali treatment (JP-A-1-55879 and Journal of Organic Synthetic Chemistry 51). Vol. 5, No. 5, 388-398, 1993) and the like.

[0006]

The present inventors have conducted various studies to find out a method for industrially producing optically active chlorohydrin from racemic chlorohydrin, and as a result, as a result, some microorganisms, particularly Pseudomonas sp., Enterobacter Certain bacteria of the genera, Chitrobacter and Bacillus degrade only one optically active form of racemic chlorohydrin, or its degradation activity results in optically active 1,2
-Converted to a diol form, and optionally further optically active 3-
It is converted into hydroxy-γ-butyrolactone, and therefore, together with the optically active substance chlorohydrin, optionally optically active 1,2-diol compound and / or optically active 3-
It has been found that hydroxy-γ-butyrolactone can be collected separately. According to the study of the present inventors, these microorganisms cannot assimilate one optically active substance of the starting compound in the reaction system, unlike the case of the above-mentioned halogenopropanediol or dichloropropanol. It is not possible to produce an optically active substance while growing a microorganism, but when the bacterial cells obtained by growing the microorganism are allowed to act on racemic chlorohydrin, only one of the optically active substances is decomposed, and the other optically active substance is decomposed. Since it remains in the system, it was learned that the desired optically active chlorohydrin can be obtained by recovering it, and depending on the type of microorganism or starting compound, only one optically active chlorohydrin was
To be able to produce an optically active 1,2-diol compound and / or 3-hydroxy-γ-butyrolactone together with the remaining optically active chlorohydrin for conversion into a 2-diol form and / or 3-hydroxy-γ-butyrolactone And came to complete the present invention.

That is, the present invention is characterized in that one optical isomer is stereoselectively decomposed from inexpensive racemic chlorohydrin by the action of a microorganism, and the other optically active isomer remaining is recovered. A method for producing an optically active chlorohydrin is provided. In the present invention, one of the optical isomers is dechlorinated from a racemic chlorohydrin by the action of a microorganism by a stereoselective decomposition reaction to obtain an optically active 1,2-diol compound, and further an optically active 3 compound.
-Hydroxy-γ-butyrolactone, and recovering the 1,2-diol compound and / or 3-hydroxy-γ-butyrolactone together with the remaining optically active chlorohydrin and the optically active chlorohydrin and 1,2- The present invention provides a method for producing a diol compound or 3-hydroxy-γ-butyrolactone or both optically active compounds.

[0008]

DETAILED DESCRIPTION OF THE INVENTION According to the present invention, the general formula

[Chemical 6] [Wherein R ¹ is a hydrogen atom or a lower alkyl group; R ² is
When R ¹ is a hydrogen atom, it is a substituted or unsubstituted lower alkyl group (excluding hydroxymethyl group (—CH ₂ OH)), or when R ¹ is a lower alkyl group, it is a hydrogen atom]. A desired optically active chlorohydrin is produced by allowing a microorganism to act on racemic chlorohydrin to selectively decompose one of the optical isomers and recover the other optical isomer remaining in the reaction system.

In the above method, depending on the type of starting material and microorganism used, one of the optical isomers has the following general formula:

[Chemical 7] [Wherein R ³ is a substituted or unsubstituted lower alkyl group] and the corresponding optically active 1,2-diol compound and the following general formula

Embedded image Is converted to an optically active 3-hydroxy-γ-butyrolactone, the optically active 1,2-diol compound and the optically active 3-hydroxy-γ-butyrolactone thus produced are produced together with the remaining optically active chlorohydrin. It

In the present specification, examples of the lower alkyl group for the R ¹ group include alkyl groups having 1 to 4 carbon atoms such as methyl, ethyl, propyl and butyl. R
Examples of the lower alkyl group in the substituted or unsubstituted lower alkyl group in the ² group and the R ³ group include methyl, ethyl,
Examples thereof include an alkyl group having 1 to 3 carbon atoms such as propyl, and examples of the substituent include a cyano group and -COOR '(R' is a direct group having 1 to 4 carbon atoms such as methyl, ethyl, propyl, isopropyl and butyl. Chain or branched alkyl group) lower alkoxycarbonyl group or -OR "
(R ″ is a linear or branched alkyl group having 1 to 4 carbon atoms such as methyl, ethyl, propyl, isopropyl, butyl, etc.) lower alkoxy group. Preferred substituted lower alkyl groups include cyanomethyl, lower alkoxycarbonylmethyl, lower alkoxymethyl.

The microorganisms used in the present invention include any microorganisms capable of selectively decomposing one optical isomer of racemic chlorohydrin represented by the formula [1], and preferably Pseudomonas. (Pseudomo
nas, Enterobacter, Citrobacter and Bacillus
s) Bacteria belonging to the genus. Of these, a particularly preferred strain is Pseudomonas sp. OS-K-29.
Strains, Pseudomonas sp. DS-K-NR818 strains, Enterobacter sp. DS-S-75 strains, Citrobacter freundii DS-S-13 strains, Citobacter freundii DS-K-40 strains, and Bacillus Examples include Bacillus sphaericus DS-ID-819 strain. Since these microorganisms do not have the ability to assimilate the chlorohydrin represented by the formula [1], they are subjected to stereoselective degradation of racemic chlorohydrin using cells obtained by growth.

Accordingly, a preferred embodiment of the present invention is a method represented by the following reaction formula-1 (provided that R ³ in compound [1a] is a lower alkoxycarbonylmethyl group, 3-hydroxy group: -Γ-butyrolactone [3a] is produced).

Embedded image [Wherein R ³ is the same as above] Among the methods represented by the above reaction schemes, a more preferred method is a method for producing a compound in which R ³ is cyanomethyl or lower alkoxycarbonylmethyl. In the above reaction formula-1, when R ³ is lower alkoxycarbonylmethyl, in addition to the optically active chlorohydrin [1a ′], in addition to the optically active 1,2-diol compound [2a], the optically active 3
-Hydroxy-γ-butyrolactone [3a] is produced, and these optically active 1,2-diol compounds [2a] are produced.
And the optically active 3-hydroxy-γ-butyrolactone [3a] are usually all R-isomers, and they are each represented by the following formula.

Embedded image [In the formula, R ³ 'is a lower alkoxycarbonylmethyl group]

Another preferred embodiment of the present invention is a method represented by the following reaction formula-2.

Embedded image [Wherein R ³ 'is the same as above] In the method of the above Reaction formula-2, another Enterobacter genus,
The bacteria exemplified above belonging to the genus Chitrobacter and the genus Bacillus are also preferably used. The optically active 3-hydroxy-γ-butyrolactone [3b] obtained by this method is usually the S-form and has a structure represented by the following formula. However, when Bacillus spericus strain DS-ID-819 is used and compound [1b] (in the formula, R ³ 'is a group other than n-butyloxycarbonylmethyl group) is used as a starting material, R The body is obtained.

[Chemical 12]

Still another preferred embodiment of the present invention is a method represented by the following reaction formula-3.

Embedded image [In the formula, R ¹ 'is a lower alkyl group]

Pseudomonas sp. OS-K-29 strain, which is a specific example of the microorganism used in the present invention, is a bacterium isolated from the soil by the present inventors, while Pseudomonas sp. DS-K-
The NR818 strain is the mutant strain. Based on the morphological and physiological properties of both strains, they were classified according to the Burjay's Manual of Systematic Bacteriology 8th Edition and 9th Edition. As a result, these were classified as Gram-negative, aerobic bacilli, Having polar flagella, oxidase positive, was identified as a strain of Pseudomonas from showing catalase positive, further these strains because there is no match with known strains, so named as above, respectively,
Deposited number FERM to Institute of Biotechnology, Institute of Biotechnology, AIST
Deposited as BP-994 and FERM BP-5491.

The mycological properties of the Pseudomonas sp. OS-K-29 strain are described in JP-A-1-55879. As a medium composition for culturing the Pseudomonas sp. OS-K-29 strain, any medium can be used as long as it is a medium in which the microorganism normally grows. For example, carbohydrates such as glucose, galactose and sucrose as carbon sources, glycerol, racemic 3-halogeno-1,2-propanediol, R or S isomer 3-halogeno-1,2-propanediol, racemic 2,3- Alcohols such as dichloro-1-propanol and R-form 2,3-dichloro-1-propanol, organic acids such as acetic acid, citric acid, malic acid, maleic acid, fumaric acid, gluconic acid and salts thereof, or a mixture thereof. Examples of the nitrogen source include inorganic nitrogen compounds such as ammonium sulfate, ammonium nitrate and ammonium phosphate, organic nitrogen compounds such as urea, peptone, casein, yeast extract, meat extract and corn steep liquor, and mixtures thereof. In addition, phosphates, magnesium salts, potassium salts, manganese salts, iron salts, zinc salts, copper salts, and the like as inorganic salts, and vitamins may be added as necessary. Further, in order to obtain bacterial cells having high enzyme activity, racemic 3-halogeno-1,2-propanediol, R or R is added to the above-mentioned medium and nutrient medium such as peptone medium and broth medium when culturing this strain. S-form 3-halogeno-1,2-propanediol, racemic 2,3-dichloro-1-propanol and R-form 2,3-dichloro-1-propanol may be added. Racemic 2,3-dichloro-1-propanol or R-isomer 2,3-
Dichloro-1-propanol, racemic 3-halogeno-
It is also effective to culture in a completely synthetic medium containing 1,2-propanediol as a single carbon source. Cultivation of the above-mentioned microorganism may be carried out by a conventional method, for example, a pH of 6 to 9, preferably 6.5.
~ 7.5, culture temperature 20 ~ 40 ℃, preferably 25 ~
It is preferable to perform aerobically at 37 ° C. for 10 to 96 hours.

The mycological properties of Pseudomonas sp. DS-K-NR818 strain are as follows. A. Morphology Cell morphology Cell size 0.4-0.6 × 1.2-1.6 μm Cell polymorphism Presence / absence of motility, presence / absence of polar flagella spores No Gram stainability-antiacidity-B ． Growth condition in each medium 1. Meat agar medium (30 ° C, 3 days culture) 1) Slow colony shape Normal 2) Colony shape Round 3) Colony surface shape Smooth 4) Colony uplift 5) Colony peripheral edge 6) Colony edge Content Homogeneous 7) Color of the colony Milky white 8) Luster of the colony Dull light 9) Transparency of the colony 10) Soluble pigment formation None 2. Broth agar slant medium (30 ° C, 3 days culture) 1) Good or bad growth 2) Growth condition Filamentous 3) Smooth colony surface 4) Flat cross-section of colony 5) Gloss of colony Dull light 6) Color of colony milky white 7) Transparency of colony Semi-transparent 3. Meat broth static culture (30 ° C, 3 days culture) 1) Growth condition Slight turbidity 2) No gas generation 3) Coloring of medium No gelatin liquefaction test No liquefaction Litmus milk No coagulation, no change C. Physiological test Lysine decarboxylation test + VP test − MR test − Nitrate reduction − Indole production − PPA reaction − Hydrogen sulfide production − Use of citric acid + Starch decomposition test − Denitrification reaction − Use of inorganic salt + Pigment Generation 1) King A medium − 2) King B medium − 3) Pseudomonas P medium − 4) Pseudomonas F medium − Catalase + oxidase + urease test + OF-test (by Hugh Leifson method. Gas generation is not observed. 1) D-glucose-2) glycerin O3) D-galactose-4) D-fructose-5) D-trehalose-PHB accumulation + utilization of carbon source 1) D-mannitol-2) D-fructose + 3) D-glucose-4) D-gluconic acid + 5) D-galactose-6) Glycerin + 7) p-Hydroxybenzoic acid + Growth optimum pH 5-9 Education optimum temperature 20~40 ℃

As the medium composition for culturing the Pseudomonas sp. DS-K-NR818 strain, any medium can be used as long as it is a medium in which this microorganism normally grows. For example, carbohydrates such as fructose as a carbon source, glycerol, alcohols such as racemic 3-halogeno-1,2-propanediol and racemic 2,3-dichloro-1-propanol, acetic acid, citric acid, malic acid, Maleic acid, fumaric acid, organic acids such as gluconic acid and its salts, or a mixture thereof, ammonium sulfate as a nitrogen source, ammonium nitrate, inorganic nitrogen compounds such as ammonium phosphate and urea, peptone, casein, yeast extract, meat extract, Mention may be made of organic nitrogen compounds such as Corn Steep Liquor and mixtures thereof. Other,
As inorganic salts, phosphates, magnesium salts, potassium salts,
If necessary, vitamins such as manganese salt, iron salt, zinc salt, and copper salt may be added. Further, in order to obtain bacterial cells having high degrading activity, the racemic 3-halogeno-1,2-propanediol, 2, 3-
Dichloro-1-propanol may be added. It is also effective to culture in a completely synthetic medium containing racemic 3-halogeno-1,2-propanediol and 2,3-dichloro-1-propanol as a single carbon source. Cultivation of the above-mentioned microorganism may be carried out by a conventional method, for example, a pH of 6 to 9, preferably 6.
5 to 7.5, culture temperature is 20 to 40 ° C., preferably 25
It is preferable to perform aerobically in the range of to 37 ° C for 10 to 96 hours.

Specific examples of other microorganisms to be used in the present invention are Enterobacter sp. DS-S-75 strain, Chitobacter freundii DS-S-13 strain, Chitrobacter freundii DS-K-40 strain, and Bacillus spericus. DS-ID
The -819 strain is also a strain isolated and collected by the present inventors from the soil, and based on the morphological and physiological properties of these strains, the Burjay's Manual of Systematic Bacteriology No. 9 As a result of classification according to the version, among the above bacterial cells, the Enterobacter sp. DS-S-75 strain has Gram-negative, facultative anaerobic bacilli, periflagellates, VP-positive, M-positive.
Since it was negative for R test and negative for DNase activity, it was identified as a strain of the genus Enterobacter. Further, as a result of identification, since this strain does not match any known strain, it has been named as described above and deposited under the deposit number FERM BP-5494 at the Institute of Biotechnology, Institute of Industrial Science and Technology. Similarly Kitrobacter Freundy DS-S-
13 strains and Chitobacter frownii DS-K-40 strain have Gram-negative, facultative anaerobic bacillus, periflagellate, negative VP reaction, and use citric acid as a single carbon source. It was identified as a strain. Further, as a result of identification, this strain was identified as Chitobacter freundii because it produces hydrogen sulfide, and the deposit number F was assigned to the Institute of Biotechnology, Institute of Biotechnology, AIST.
ERM BP-5492 and FERM BP-54
Deposited as 93. Similarly, Bacillus spericus strain DS-ID-819 was identified as a strain of Bacillus genus because it is Gram-positive, has periflagellates, forms spores in aerobic rods, and is positive for catalase. Based on various physiological properties, this strain was identified as Bacillus spericus and deposited at the Institute of Biotechnology, Institute of Industrial Science and Technology with deposit number FERM BP.
Deposited as -5495. The properties of the above strains are as shown below.

The mycological properties of Enterobacter sp. DS-S-75 strain are as shown below. A. Morphology Cell morphology Bacillus Cell size 0.6 to 0.8 x 1.5 to 2.8 μm Cell polymorphism Presence or absence of motile, Presence or absence of periflagellate spores Gram stainability-B. Growth condition in each medium 1. Meat agar medium (30 ° C, 3 days culture) 1) Colony size 3-4 mm 2) Colony shape irregularity 3) Colony shape slow and fast 4) Colony surface shape smooth and viscous 5) Colony uplifting convex Circular 6) Wavy around colony 7) Color of colony Milky white 8) Gloss of colony Dull light 9) Transparency of colony 10) Soluble pigment formation None 2. Meat broth agar slant medium (30 ° C, 3 days culture) 1) Good or bad growth 2) Growth condition Filamentous 3) Gloss of colony dull 4) Color of colony Milky white 3. Liquid static culture (30 ° C, 3 days culture) 1) Growth state Turbidity 2) Coloring of medium 3) Presence of precipitation C. Physiological test Indole production-catalase + urease + oxidase-β-galactosidase + lysine decarboxylase + production of hydrogen sulfide-utilization of citric acid + starch decomposition-liquefaction of gelatin-MR-VP + nitrate reduction + de-N reaction + DNase activity-NPTase + OF test (glucose) FD-glucose, acid + D-glucose, gas + Optimal growth temperature 30-37 ° C Optimal growth pH 5-9

[0021] Also, Kitrobacter Freundy DS-
The bacteriological properties of the S-13 strain are as shown below. A. Morphology Cell shape Bacillus Cell size 0.5-0.7 × 1.8-3.6 μm Cell polymorphism Presence or absence of motile, Presence or absence of periflagellate spores Gram stainability-B. Growth condition in each medium 1. Meat agar medium (30 ° C, 3 days culture) 1) Colony size 2-4mm 2) Colony shape circular 3) Slow and fast colony shape 4) Colony surface shape smooth 5) Colony uplifting convex circle 6 ) All around the colony 7) Color of the colony Milky white 8) Luster of the colony 9) Transparency of the colony 10) Transparency 10) No pigment formation 2. Meat broth agar slant medium (30 ° C, 3 days culture) 1) Good or bad growth 2) Growth condition 3) Glossy 4) Color tone milky white 3. Liquid static culture (30 ° C, 3 days culture) 1) Growth state Turbidity 2) Coloring of medium 3) Presence of precipitation C. Physiological test Indole production-catalase + urease + oxidase-β-galactosidase + lysine decarboxylase-Hydrogen sulfide production + Utilization of citric acid + Starch degradation-Gelatin liquefaction-MR + VP-Reduction of nitrate + DeN reaction + DNase activity-NPTase-OF test (glucose) FD-glucose, acid + D-glucose, gas + Optimal growth temperature 25-30 ° C Optimal growth pH 5-9

Also, Kitrobacter Freundy DS-
The mycological properties of the K-40 strain are as shown below. A. Morphology Cell shape Bacillus Cell size 0.5-0.7 × 1.8-3.6 μm Cell polymorphism Presence or absence of motile, Presence or absence of periflagellate spores Gram stainability-B. Growth condition in each medium 1. Meat agar medium (30 ° C, 3 days culture) 1) Colony size 2-4mm 2) Colony shape circular 3) Slow and fast colony shape 4) Colony surface shape smooth 5) Colony uplifting convex circle 6 ) All around the colony 7) Color of the colony Milky white 8) Luster of the colony 9) Transparency of the colony 10) Transparency 10) No pigment formation 2. Meat broth agar slant medium (30 ° C, 3 days culture) 1) Good or bad growth 2) Growth condition 3) Glossy 4) Color tone milky white 3. Static culture of broth (30 ° C, 3 days) 1) Growth state Turbidity 2) Coloration of medium 3) Presence of precipitation C. Yes Physiological test Indole production-catalase + urease + oxidase-β-galactosidase + lysine decarboxylase-Hydrogen sulfide production + Utilization of citric acid + Starch degradation-Gelatin liquefaction-MR + VP-Reduction of nitrate + DeN reaction + DNase activity-NPTase-OF test (glucose) FD-glucose, acid + D-glucose, gas + Optimal growth temperature 25-30 ° C Optimal growth pH 5-9

The mycological properties of the Bacillus spericus strain DS-ID-819 are as shown below. A. Morphology Cell morphology Bacillus Cell size 0.5-0.7 × 2.0-3.5 μm Cell polymorphism Presence or absence of motility, Presence or absence of periflagellate spores Gram stainability + B. Growth condition in each medium 1. Meat agar medium (30 ° C, 3 days culture) 1) Colony size 2-4 mm 2) Colony shape circular 3) Slow colony shape Normal 4) Colony surface wrinkle 5) Colony uplifting flat 6 ) All around the colony 7) Color of the colony Milky white 8) Gloss of the colony Dull light 9) Transparency of the colony 10) Dye formation None 2. Meat broth agar slant medium (30 ° C, 3 days culture) 1) Good or bad growth 2) Growth condition Spreaded 3) Gloss dull 4) Color tone Milky white 3. Liquid static culture (30 ° C., 3 days culture) 1) Growth condition Slight turbidity 2) Coloring of medium 3) Presence or absence of precipitation C. Physiological test Indole production-catalase + urease + oxidase + β-galactosidase-lysine decarboxylase + hydrogen sulfide production-utilization of citric acid + starch decomposition-gelatin liquefaction + MR-VP-nitrate reduction-de-N reaction -DNase activity + NPTase-OF test (glucose) OD-glucose, acid-D-glucose, gas- Optimal growth temperature 30-37 ° C Optimal growth pH 6-9

As the medium composition for culturing the above-mentioned microorganism, any medium can be used as long as it is a medium in which this microorganism normally grows. For example, as a carbon source, glucose, galactose, carbohydrates such as fructose, alcohols such as glycerol, acetic acid, citric acid, malic acid, maleic acid, fumaric acid, organic acids such as gluconic acid and its salts, or a mixture thereof. , Inorganic nitrogen compounds such as ammonium sulfate, ammonium nitrate, ammonium phosphate as a nitrogen source and urea, peptone, casein,
Mention may be made of organic nitrogen compounds such as yeast extract, meat extract, corn steep liquor and mixtures thereof. In addition, as inorganic salts, phosphates, magnesium salts, potassium salts, manganese salts, iron salts, zinc salts, copper salts, and the like.
Vitamins may be added if necessary. Cultivation of the above microorganisms may be carried out by a conventional method, for example, aerobically at a pH of 5 to 9, preferably 6.5 to 7.0 and a culture temperature of 20 to 40 ° C, preferably 25 to 37 ° C. It is preferably carried out for 10 to 96 hours.

By allowing a microorganism to act on racemic chlorohydrin, optically active chlorohydrin and optically active 1,2-
As the method for obtaining the diol compound and / or the optically active 3-hydroxy-γ-butyrolactone, 1) the culture solution of the microorganism obtained by the above-mentioned culture method of the microorganism of the present invention, or 2) the bacterial cells obtained by centrifugation and the cells thereof Treated cells (crushed cells or cell extract), or 3)
There is a method in which they are immobilized by a conventional method, and a substrate is added to a mixed solution of microbial cells mixed with a buffer solution.
The reaction temperature is preferably 15 to 50 ° C, and the reaction pH is 6 to 9
It is preferable to carry out. Substrate concentration in the reaction solution is 0.1-1
5% (v / v) is preferable, and the substrate may be added all at once at the initial stage, or may be added in portions. The reaction is usually carried out with stirring or shaking, and the reaction time is preferably 1 to 120 hours, although it varies depending on the substrate concentration and the amount of microbial cells. Preferably, the end point is determined by measuring the optical purity of the resulting optically active substance by analysis such as gas chromatography, and the end point is when the optical purity of the product is maximized.

The optically active chlorohydrin and the optically active 1,2-diol compound and / or the optically active 3-hydroxy-γ-butyrolactone thus remaining in the reaction solution can be recovered and purified by a general method. For example, after removing bacterial cells from the reaction solution by centrifugation, the supernatant is concentrated by an evaporator and extracted with a solvent such as ethyl acetate. After dehydrating the extract with anhydrous magnesium sulfate, the solvent is removed under reduced pressure to obtain a syrup of optically active chlorohydrin. When an optically active 1,2-diol compound is produced, it is obtained as a mixture thereof. These compounds may be further purified by distillation or column chromatography. When optically active 3-hydroxy-γ-butyrolactone is produced, the supernatant obtained by removing bacterial cells by centrifugation is extracted with ethyl acetate or the like, and chlorohydrin or 1,
After removing the 2-diol, the crude syrup of 3-hydroxy-γ-butyrolactone can be obtained by concentrating the aqueous layer fraction. Alternatively, the above aqueous layer fraction may be recovered and purified with activated carbon by a conventional method. Further, it may be purified by column chromatography using activated carbon or silica gel.

The present invention will be described in detail with reference to the following examples, but the present invention is not limited to these. In addition,% in the examples is represented by (w / v%) unless otherwise specified.

Example 1 Peptone, yeast extract and glycerol were each added to 1.0
5% containing 2.5 liters of nutrient medium (pH 7.2) containing 5%
Litter jar fermenter (incubator) at 121 ℃
It was autoclaved for 15 minutes. The Pseudomonas sp. OS-K-29 strain, which is a test bacterium, was used in a nutrient medium (pH 7.2) containing 1.0% each of peptone, yeast extract and glycerol.
Inoculum was prepared by shaking culture at 0 ° C. for 16 hours, and 2% (v / v) amount was aseptically inoculated into the above medium. And temperature 30
The culture was carried out under aeration and agitation for about 24 hours under the conditions of ℃, aeration rate of 1.0 L / min and rotation speed of 500 rpm. After the completion of the culture, the culture solution was taken out, collected by centrifugation, and washed three times with a 20 mM phosphate buffer solution (pH 7.2) containing 2 mM magnesium sulfate to prepare resting cells. The cells were suspended in a 500 ml Erlenmeyer flask with a baffle containing 100 ml of the same buffer containing 1.0% calcium carbonate, and 1.0 (v / v) of racemic 4-chloro was added to the suspension. -3-Hydroxybutyronitrile was added, and the mixture was reacted at 30 ° C for 90 hours with stirring. 4-Chloro-3-hydroxybutyronitrile remaining in the reaction solution at that time was analyzed by gas chromatography (column carrier: PEG20M, 60-80 mesh), and as a result, the residual rate was 40.1%. After completion of the reaction, the reaction solution was centrifuged to remove the cells, then concentrated to about 1 ml and extracted with ethyl acetate.
After dehydration with anhydrous magnesium sulfate, the solvent was removed under reduced pressure, and 380 mg of 4-chloro-3-hydroxybutyronitrile and 350 mg of 1,2-diol corresponding to the substrate were removed.
I got The identification and quantification of this substance was performed by the same gas chromatography and confirmed from the retention time.

4-chloro-3-hydroxybutyronitrile in this syrup was added to Astec (Advanced Separat
The optical isomers were analyzed by gas chromatography using a capillary column G-TA (0.25 mm × 30 m) manufactured by ion Technologies Inc., NJ, USA). On the other hand, the 1,2-diol compound was trifluorinated with trifluoroacetic anhydride and then analyzed for optical isomers by the above gas chromatography. As a result, it was found that the recovered 4-chloro-3-hydroxybutyronitrile was an S-isomer having an optical purity of 94.5% ee. Further, it was found that the recovered 1,2-diol was an R-isomer having an optical purity of 42.2% ee. The conditions for analyzing the optical isomer by the above-mentioned gas chromatography are as follows.
The retention times of 4-chloro-3-hydroxybutyronitrile are S-isomer, 64.1 minutes; R-isomer, 66.5 minutes.
The retention time of 1,2-diol is S body, 113.
1 minute; R form, 125.3 minutes. Analytical conditions: column temperature, 120 ° C; detector temperature, 200
° C; carrier gas, nitrogen; flow rate, 0.5 ml / mi
n; detector, FID; split ratio, 100/1.

Example 2-7 The reaction was carried out in the same manner as in Example 1 except that the substrate shown in Table 1 was changed from racemic 4-chloro-3-hydroxybutyronitrile. The various compounds obtained were identified and quantitatively analyzed by the same method as in Example 1, and the optical isomers were also analyzed by the same method. In addition, the 3-hydroxy-γ-butyrolactone produced in Examples 3 to 5 was extracted with chlorohydrin and 1,2-diol with an equal amount of ethyl acetate to extract and remove the supernatant from the cells, and the aqueous layer fraction thereof was obtained. The fraction was concentrated to obtain a crude syrup of 3-hydroxy-γ-butyrolactone. The identification and quantification of this substance was carried out by the same gas chromatography as shown in Example 1 and confirmed by the retention time. The optical purity of 3-hydroxy-γ-butyrolactone in this syrup was measured by acetylating with acetyl chloride, and then using Astec capillary column G- shown in Example 1.
It was performed by gas chromatography equipped with TA.
The retention times of 3-hydroxy-γ-butyrolactone were S-form, 11.3 minutes; R-form, 12.0 minutes. Analysis conditions: column temperature, 150 ° C; detection temperature, 200
℃; carrier gas, nitrogen; flow rate, 0.7 ml / mi
n; detector, FID; split ratio; 100/1. The physical properties of the obtained 3-hydroxy-γ-butyrolactone are the literature values (Synthetic Communications, 16 (2), pp. 183-1).
90, 1986). ¹ H-NMR (CDCl ₃ , 250 MHz), δppm: 2.54
(d, 1H, J = 18Hz); 2.79 (dd, 1H, J = 18Hz, J = 6Hz); 3.29
(s, 1H, OH); 4.34 (d, 1H, J = 10Hz); 4.46 (dd, 1H, J = 1
0Hz, J = 4.5Hz); 4.6-4.7 (m, 1H) ¹³ C-NMR (CDCl ₃ ), δppm: 177, 76.3, 67.2,
37.7 Table 1 shows the products of Examples 1 to 7 and the yield thereof.

[0031]

[Table 1]

[0032]

[Table 2]

Example 8 2.5 L of a nutrient medium (pH 7.0) consisting of peptone, yeast extract and glycerol (1% each) was placed in a 5 L jar famentor (incubator).
High-pressure sterilization was performed at ℃ for 15 minutes. Pseudomonas sp.DS-K-NR818, which is a test bacterium, was previously added to peptone, yeast extract,
Nutrient medium (pH 7.
0) at 30 ° C. for 18 hours with shaking to prepare an inoculum,
The above medium was inoculated aseptically with an amount of 2.0% (v / v).
Then, aeration and agitation culture was carried out for about 24 hours under the conditions of a temperature of 30 ° C., an aeration rate of 0.5 L / min and an agitation rotation speed of 500 rpm. After the completion of the culture, the culture solution was taken out, collected by centrifugation, and washed three times with a 20 mM phosphate buffer solution (pH 7.2) containing 2 mM magnesium sulfate to prepare resting cells. The fungus body is 20m containing 1.0% calcium carbonate
The suspension was suspended in a baffled 500 ml Erlenmeyer flask containing 100 ml of M phosphate buffer (pH 7.0). The suspension was prepared so that the concentration of magnesium sulfate was 2 mM. Racemic ethyl 4-chloro-3-hydroxybutyrate was added to the suspension at 1.0% (v / v), and the mixture was added at 30 ° C.
And reacted for 40 hours. After the reaction, ethyl 4-chloro-3-hydroxybutyrate remaining in the reaction solution was subjected to gas chromatography (column carrier: PEG20M, 60-
80 mesh). As a result, the survival rate is 41.
8%. After completion of the reaction, the reaction solution was centrifuged to remove the bacterial cells, then concentrated to about 1.0 ml and extracted with ethyl acetate. After dehydration with anhydrous magnesium sulfate, the solvent was removed under reduced pressure to obtain 406 mg of ethyl 4-chloro-3.
-Hydroxybutyrate was obtained. The identification and quantification of this substance was performed by the same gas chromatography and confirmed from the retention time. The remaining ethyl 4-chloro-3-hydroxybutyrate was analyzed for optical isomers by gas chromatography using a capillary column G-TA (0.25 mm × 30 m) manufactured by Astec. As a result, the recovered ethyl 4-chloro-3-
Hydroxybutyrate was an R form having an optical purity of 98% ee or higher. The conditions for analyzing the optical isomer by the above-mentioned gas chromatography are as follows. Ethyl 4
The retention times of -chloro-3-hydroxybutyrate were R isomer 39.5 minutes and S isomer 41.0 minutes. Analytical conditions: column temperature, 110 ° C; detector temperature, 200
℃; carrier gas, nitrogen; flow rate, 0.7 ml / mi
n; detector, FID; split ratio, 100/1. The 3-hydroxy-γ-butyrolactone produced was
Chlorohydrin was extracted and removed from the supernatant from which the bacterial cells had been removed with an equal amount of ethyl acetate, and the aqueous layer fraction was concentrated to obtain a crude syrup of 3-hydroxy-γ-butyrolactone. Identification and quantification of this substance are carried out in the same manner as in Example 3,
The optical isomers were analyzed in the same manner.

Examples 9 to 12 Example 8 except that the substrate was changed from racemic ethyl 4-chloro-3-hydroxybutyrate to the substrate shown in Table 2.
The reaction was carried out in the same manner as in. The various compounds thus obtained were identified and quantitatively analyzed in the same manner as in Example 8,
The optical isomers were analyzed in the same manner. Table 2 shows products and yields of the above Examples 8 to 12.

[0035]

[Table 3]

[0036]

[Table 4]

Example 13 (S) -3-hydroxy-γ-butyrolactone and (R)
-Methyl 4-chloro-3-hydroxybutyrate production 100 ml of a nutrient medium (pH 7.0) consisting of peptone, yeast extract and glycerol at 1% each was charged into a 500 ml Erlenmeyer flask equipped with a baffle, at 121 ° C, 15 ° C.
Autoclaved for 1 minute. The test bacteria, Enterobacter sp. DS-S-75 strain, were preliminarily shake-cultured at 30 ° C for 20 hours in a nutrient medium (pH 7.0) containing peptone, yeast extract, and glycerol at 1.0% each to give seeds. The prepared medium was aseptically inoculated into the above medium in an amount of 2.0% (v / v). Then, the culture was carried out at a temperature of 30 ° C. under stirring conditions for about 24 hours. After culturing, remove the culture solution and collect the cells by centrifugation to collect the cells for 2 m.
20 mM phosphate buffer containing M magnesium sulfate (p
H7.2) was washed 3 times to prepare resting cells. The cells were suspended in a 500 ml Erlenmeyer flask with a baffle containing 100 ml of a 20 mM phosphate buffer solution (pH 7.0) containing 1.0% calcium carbonate. Magnesium sulfate was adjusted to 2 mM in the suspension. Racemic methyl 4-chloro-3-hydroxybutyrate was added to the suspension at 1.0%, and the mixture was reacted at 30 ° C. for 24 hours. After the reaction, methyl 4-chloro-3-hydroxybutyrate remaining in the reaction solution was analyzed by gas chromatography (column carrier: PEG20M, 60-80 mesh). As a result, the residual rate was 48.0%. After completion of the reaction, the reaction solution is centrifuged to remove the bacterial cells,
The supernatant was applied to an activated carbon column and eluted with acetone. Acetone was distilled off from the eluate under reduced pressure to obtain an oily substance, and 342 mg of (R) -methyl 4-chloro-3-hydroxybutyrate (boiling point 75-80 / 1.
5 mmHg) and 214 mg of (S) -3-hydroxy-
γ-butyrolactone (boiling point 110-120 / 0.5 mm
Hg) was obtained as an oil. The obtained methyl 4-chloro-3-hydroxybutyrate was an R-form having an optical purity of 99% ee or higher. Further, the obtained 3-hydroxy-γ-butyrolactone was reacted with acetyl chloride to acetylate, and then the optical purity was measured. 3-Hydroxy-γ-butyrolactone was an S form having an optical purity of 95.9% ee.
The optical purity was analyzed by gas chromatography using a capillary column G-TA (0.25 mm × 30 m) manufactured by Astec. The analysis conditions of the optical isomers by the above-mentioned gas chromatography are as follows. Analysis conditions for methyl 4-chloro-3-hydroxybutyrate: column temperature, 110 ° C; detector temperature, 200 ° C;
Carrier gas, nitrogen; flow rate, 0.7 ml / min; detector, FID; split ratio, 100/1. Retention time is R body, 15.5 minutes; S body, 16.3 minutes. Analysis conditions for 3-hydroxy-γ-butyrolactone: column temperature, 150 ° C .; detector temperature, 200 ° C .; carrier gas, nitrogen; flow rate, 0.7 ml / min; detector, FI
D: Split ratio, 100/1. Retention time is S body, 11.3 minutes; R body, 12.0 minutes. The physical properties of the obtained 3-hydroxy-γ-butyrolactone are the literature values (Synthetic Communications, 16 (2), pp. 183-1).
90, 1986). [Α] _D ²⁰ = −76.2 ° (C = 1.95, EtOH) ¹ H-NMR (CDCl ₃ , 250 MHz), δppm: 2.54
(d, 1H, J = 18Hz); 2.79 (dd, 1H, J = 18Hz, J = 6Hz); 3.29
(s, 1H, OH); 4.34 (d, 1H, J = 10Hz); 4.46 (dd, 1H, J = 1
0Hz, J = 4.5Hz); 4.6-4.7 (m, 1H) ¹³ C-NMR (CDCl ₃ ), δppm: 177, 76.3, 67.2,
37.7

Examples 14 to 32 The substrate was changed from racemic methyl 4-chloro-3-hydroxybutyrate to the substrate shown in Table 1, and Enterobacter sp. DS-S-75 was used as a test strain for each substrate. Strains, Kitrobacter Freundy DS-S-13 Strains, Kitrobacter Freundy DS-K-40 Strains, Bacillus spericus
Reactions were carried out in the same manner as in Example 13 using the DS-ID-819 strain. The various compounds obtained were quantitatively analyzed by the same method as in Example 13, and the optical isomers were also analyzed by the same method. The yield and optical purity of the obtained compound are shown in Table 3. The physical properties of the obtained 3-hydroxy-γ-butyrolactone were the same as in Example 13 in all cases. Table 3 shows the products in Examples 13 to 32 and their yields. The abbreviations in Table 3 have the following meanings. MH: Methyl 4-chloro-3-hydroxybutyrate EH: Ethyl 4-chloro-3-hydroxybutyrate IPH: Isopropyl 4-chloro-3-hydroxybutyrate PH: Propyl 4-chloro-3-hydroxybutyrate BH: Butyl 4-chloro-3-hydroxybutyrate HL: 3-hydroxy-γ-butyrolactone

[0039]

[Table 5]

[0040]

INDUSTRIAL APPLICABILITY According to the present invention, microorganisms belonging to the genus Pseudomonas (Pseudomonas sp. OS-K-29 strain or Pseudomonas sp. DS-K-NR818 strain), and further Enterobacter genus (Enterobacter sp. DS-S) -75 strains, genus Kitrobacter (Citrobacter freundii DS-S-13
Strain, Kitrobacter frownie DS-K-40 strain), and Bacillus genus (Bacillus spericus DS-ID-819 strain)
Of the racemic chlorohydrin of the general formula [1] by decomposing only one optical isomer of the racemic chlorohydrin of the general formula [1], leaving the other optical isomer in the system, and dehalogenating the other optical isomer. To the optically active 1,2-diol or optically active 3-hydroxy-γ-butyrolactone, and the remaining other optical isomer and the optically active 1,2-diol and / or optically active 3-hydroxy- By recovering γ-butyrolactone, an optically active chlorohydrin, an optically active 1,2-diol compound and / or an optically active 3-hydroxy-γ-butyrolactone can be produced by a cheap raw material and an industrially simple method. You can The optically active chlorohydrin, the optically active 1,2-diol compound and the optically active 3-hydroxy-γ-butyrolactone produced by the method of the present invention are useful as synthetic intermediates for drugs, agricultural chemicals, physiologically active substances, ferroelectric liquid crystals and the like. Compound, especially
Optically active 4-chloro-3-hydroxybutyronitrile and optically active methyl 4 which is a carboxylic acid ester thereof
Compounds having a C4 skeleton such as -chloro-3-hydroxybutyrate and ethyl 4-chloro-3-hydroxybutyrate are optically active carnitines which are important in the pharmaceutical field, 4-
It is important as a precursor compound of chloro-3-hydroxybutyric acid, 4-hydroxy-2-pyrrolidinone, 1,2,4-butanetriol and the like. Therefore, the present invention can provide such important compounds inexpensively and conveniently,
Very useful industrially.

Continuation of front page (51) Int.Cl. ⁶ Identification number Office reference number FI Technical display location C07D 307/33 C12P 7/18 // C12P 7/18 17/04 17/04 C07D 307/32 Q (C12P 41 / 00 C12R 1:38) (C12P 41/00 C12R 1:01) (C12P 41/00 C12R 1:07) (C12P 7/18 C12R 1:38) (C12P 17/04 C12R 1:38) (C12P 17 / 04 C12R 1:01) (C12P 17/04 C12R 1:07) C07M 7:00

Claims (16)

[Claims] 1. A compound of the general formula [Wherein R ¹ is a hydrogen atom or a lower alkyl group; R ² is
A substituted or unsubstituted lower alkyl group (excluding a hydroxylmethyl group) when R ¹ is a hydrogen atom, or a hydrogen atom when R ¹ is a lower alkyl group] to a racemic chlorohydrin compound. A method for optical resolution of chlorohydrin, characterized in that one of the optical isomers is decomposed and the other optical isomer is allowed to remain by allowing the cells or the processed product thereof to act. 2. The method according to claim 1, wherein the remaining optically active chlorohydrin is recovered. 3. The method according to claim 2, wherein in the racemic chlorohydrin compound [1] as a starting compound, R ¹ is a hydrogen atom and R ² is a substituted or unsubstituted lower alkyl group. 4. The method according to claim 3, wherein R ² is an ethyl group or a methyl group substituted with a group selected from cyano, lower alkoxycarbonyl and lower alkoxy. 5. The method according to claim 3, wherein R ² is lower alkoxycarbonylmethyl. 6. The method according to claim 3, wherein R ² is methoxycarbonylmethyl, ethoxycarbonylmethyl or isopropoxycarbonylmethyl. 7. The method according to claim 2, wherein in the racemic chlorohydrin compound [1] as a starting compound, R ¹ is a lower alkyl group and R ² is a hydrogen atom. 8. The microorganism used is Pseudomonas (Pseu).
The method according to any one of claims 1 to 7, which is a bacterium belonging to the genus domonas). 9. The microorganism used is Pseudomonas sp. O.
The method according to any one of claims 3 to 6, which is SK-29 strain. 10. The method according to claim 5, wherein the microorganism used is a bacterium belonging to the genus Enterobacter, the genus Chitrobacter, or the genus Bacillus. 11. An optically active chlorohydrin compound together with the following formula: [Wherein R ³ is a substituted or unsubstituted lower alkyl]
The method according to claim 9, wherein the optically active 1,2-diol compound represented by 12. A racemic chlorohydrin compound [1] as a starting compound, wherein R ¹ is a hydrogen atom and R ² is a lower alkoxycarbonylmethyl group, and the microorganism used is Pseudomonas sp. OS-K-29 strain. Together with the optically active chlorohydrin compound, the following formula: [Wherein R ³ 'is a lower alkoxycarbonylmethyl group] and an optically active 1,2-diol represented by the following formula: The method according to claim 2, wherein the optically active 3-hydroxy-γ-butyrolactone represented by the following formula is produced. 13. The method according to claim 12, wherein R ³ 'is a methoxycarbonylmethyl, ethoxycarbonylmethyl or isopropoxycarbonylmethyl group. 14. The microorganism used is Pseudomonas sp.
The method according to claim 5, which is DS-K-NR818 strain. 15. The racemic chlorohydrin compound [1] as a starting compound, wherein R ¹ is a hydrogen atom and R ² is a lower alkoxycarbonylmethyl group, and the microorganism used is Pseudomonas sp. DS-K-NR818 strain, entero. Bactor sp.
DS-S-75 shares, Kitrobacter Frendy DS-S-13
A strain selected from the group consisting of S. strains, Kitrobacter freundii DS-K-40 strain, and Bacillus spericas DS-ID-819 strain, together with an optically active chlorohydrin compound. The method according to claim 2, wherein the optically active 3-hydroxy-γ-butyrolactone represented by the following formula is produced. 16. The method according to claim 15, wherein R ² in the starting compound [1] is a methoxycarbonylmethyl, ethoxycarbonylmethyl, propoxycarbonylmethyl, isopropoxycarbonylmethyl, or butyloxycarbonylmethyl group.