JPH02257895A - Preparation of optically active epichlorohydrin - Google Patents

Abstract

PURPOSE:To prepare an optically active epichlorohydrin by bringing a specific bacterium belonging to the genus Pseudomonas into contact with racemic 2,3- dichloro-1-propanol in a medium and further treating the product with an alkaline agent. CONSTITUTION:A bacterium belonging to the genus Pseudomonas and having an ability to utilize S-(-)-2,3-dichloro-1-propanol in a prescribed medium to produce R-(+)-2,3-dichloro-1-propanol [R-(+)-beta-DCH]. The cultured solution is filtered and the supernatant of the filtered solution is subjected to an operation such as an active carbon column treatment to fractionate the R-(+)-beta-DCH remained in the supernatant. The fractionated compound is treated with an alkaline agent, preferably a caustic alkali such as caustic soda or caustic potash to prepare optically active epichlorohydrin.

Description

Detailed Description of the Invention (Industrial Application Field) The present invention relates to a method for obtaining optically active epichlorohydrin by subjecting racemic 2,3-dichloro-1=propatol (hereinafter referred to as β-DCH) to microbial treatment. .

(Prior Art and Problems to be Solved by the Invention) Optically active epichlorohydrin is an important raw material for the synthesis of various medicines. However, a method for producing this optically active epichlorohydrin is described by Balclwin, Journal, Orr. Organic, Chemistry (J, ()rg,
chem) Volume 43, 1978.

Page 4876 or EIIIiS, Journal, Chemical Society, Chemical Communication,
, (J,CHEH,SOC,,CHEH,C0HH
UN, 1984, p. 1600, but all of them require advanced synthesis techniques, and no simple manufacturing method is known. The present inventors have already reported a method for obtaining highly pure optically active 5-(-)-β-DCH by bringing racemic β-DCH into contact with R-(10)-β-DCH-assimilating bacteria (Unexamined Patent Publication No. 1986-132196) and a method to obtain R-(-)-epichlorohydrin by reacting this product with an alkaline agent (Japanese Patent Application Laid-Open No. 62-6697), but the opposite optical isomer, namely S -(10)-A simple method for producing epichlorohydrin is not known.

(Means for Solving the Problems) The present inventors have completed the present invention by discovering that the optically active epichlorohydrin described above can be easily produced with high purity by microbial treatment.

That is, the present invention provides an R-
This is a method for producing optically active epichlorohydrin, which is characterized by reacting (10)-β-DCH with an alkaline agent to obtain S-(10)-epichlorohydrin.

The mycological properties of the microorganisms isolated and collected from soil by the present inventors and used in the present invention are shown in Table 1.

Table ■Cell pleomorphism Absent ■Mobility present, polar flagella ■Presence or absence of spores
No Durham stainability
Negative ■ Anti-acidity None, growth status in each medium ■ Broth agar plate culture! (Cultivated at 30℃ for 3 days) A) Slow colony shape Normal Approximately 3-4 am in diameter Colony shape Circular C Colony surface shape Smooth 2 Colony protrusion Convex circular E Colony periphery Toward golden edge Colony contents Homogeneous colony color tone
Milky white CH) Transparency of the colony Translucent) Gloss of the colony Dull luminosity N) Soluble e
Formation of grains None■ Broth agar slant culture (cultured at 30℃ for 3 days) A Quality of growth Good 18F, filamentous Colony shape Smooth C Colony cross section ridged lI! Squamous 2 Colony gloss Dull light E Colony surface shape Smooth Colony transparency Translucent Colony color Milky white■ Flesh liquid culture (30℃, 3 days culture) A) Growth characteristics
Membrane 0) Turbidity Slightly turbid C) Gas generation None E) Culture medium coloration None ■Meat juice gelatin puncture culture Without liquefying gelatin■Lidmus milk unchanged C1 Physiological test 1 Nitrate reduction 2 MR test 3 VP test 4 Indole production 5 Hydrogen sulfide production 6 Starch hydrolysis 7 Denitrification reaction -8 Utilization of citric acid 19 Utilization of nitrogen-free sources 110 Production of pigments Not particularly produced 11 Urease -12 Oxidase
+13 Catalase
+14 Freshman 1 (F) III
E) H5,0~9.0, temperature a20~45℃15
m degree to oxygen Aerobic 160-FT strike (by nugh tetrson method)
011 Presence or absence of acid and gas generation from sugars IW Class Acid Gas (1) D-Glucose -(2) D-
Galactose (3) Sucrose (4) Trehalose (5) Starch - (6) Glycerin 118 Arginine dihydrolase + 9 Based on the results above the accumulation of PH8, Purge Aids Manual of
Systematic Batatteriology (Bergey'
s Manual of Systematic Ba
According to the identification based on the description in Volume 1 of Cteriology, the water bottle has characteristics of the genus Pseudomonas (FERM P-10520)
゜hereinafter referred to as O8- to -38 strain).

The gist of the present invention is to optically activate the racemic β-DCH using this bacterium, and then react it with an alkaline agent by a conventional method to obtain optically active epichlorohydrin. In the present invention, the O8-38 strain may be used as it is or may be immobilized, but the above-mentioned bacterial culture method and immobilization method may be any commonly used method.

That is, the culture method involves culturing the above bacteria in a bouillon medium or a sweetened bouillon medium, etc., with a carbon source and a nitrogen source.

The cells may be cultured in a nutrient medium containing an organic nutrient source and an inorganic nutrient source, allowed to grow well, and then the resulting culture or cultured bacterial cells may be used. Carbon sources include carbohydrates such as glycerin, or organic compounds and their salts such as citric acid, maleic acid, and malic acid; nitrogen sources include inorganic nitrogen such as ammonium sulfate, ammonium chloride, ammonium nitrate, and ammonium phosphate, and peptone. casein.

Il-form nitrogen such as yeast extract and meat extract can be used. Other inorganic salts include phosphates.

is used. The culture conditions are usually between 20 and 45 degrees Fahrenheit.
C., preferably 25 to 31.degree. C., pH about 5 to 9, preferably 1) H6.0 to 1.5, and is carried out aerobically by means such as shaking or aeration stirring.

Further, the immobilization method may be, for example, a method of enclosing the shikokutai using acrylamide, carrageenan, agar, gelatin, sodium alginate, etc. After immobilization, it may be crushed into an appropriate size and shape for use. .

The reaction between this strain and racemic β-DCH is racemic β-DCH.
The above-mentioned culture or its immobilized product may be stirred and brought into contact with each other in a synthetic medium containing CH, and the contact time is usually half a day to 10 days, and the concentration of β-DCH is approximately 0.1 times the width of the medium.
It is sufficient that the amount is about 0.6% by volume.

After the reaction is completed, the reaction solution is taken out and filtered to separate the immobilized product and the supernatant, and the R-(+)-β-D remaining in the supernatant is removed.
CH is separated by operations such as activated carbon column treatment, ether extraction, and vacuum distillation. Add an alkaline agent, preferably caustic soda, to this aliquot.

It is made into epichlorohydrin by the action of a caustic alkali such as caustic potash.

Furthermore, if immobilized bacterial cells are used in the method of the present invention, operations such as centrifugation become easy, and the immobilized product can be used repeatedly.

This will be explained in detail below using Examples. In the examples, percentages are based on weighted boxes unless otherwise noted.

Example 1 Medium 20j with yeast extract 1.0%, glycerin 2.0%, polypeptone 1.0%, pH 7.0! 301! Place in a jar fermenter and heat sterilize as usual.
-38 strain was inoculated into O3- and cultured for 24 hours under the following conditions.

Temperature 30℃ pH Initial pH 7.0 Ventilation amount 20j! /min Stirring rotation speed: 30 Or, p, m.

After the culture was completed, the microbial cells and the culture filtrate were separated using a centrifuge to obtain 600 g of viable cells. Next, the viable bacterial cells are
The following (shown below) was suspended in a synthetic medium to a volume of 10.1 mm, and fixed with acrylamide in the usual manner.The immobilized material was crushed into 0.5-1 mm square pieces using a mixer and thoroughly washed with a synthetic medium. did.

Components of synthetic medium Ammonium sulfate 0.05% by weight Ammonium nitrate o, os n Potassium hydrogen phosphate
0.1 Sodium phosphate 0.2 Sodium phosphate 0.1 Magnesium sulfate 0.05 Iron sulfate, copper sulfate, manganese sulfate Trace pI
Initial Start 1) t16.8 Next, the immobilized product thus prepared was placed in a 1001-volume jar fermentor and made up to 807 ml with a synthetic medium. Furthermore, racemic β
- Add 320 d of DCH and 160 g of calcium carbonate,
Stirring was carried out under the following conditions.

Temperature 30℃ Ventilation fJ 40.1! /min rotation speed
300 r, p, m.

72 hours after the start of the reaction, the supernatant liquid and the immobilized product were separated by filtration,
The remaining β-DCH was collected from this liquid by using an activated carbon column, ether extraction, and distillation under reduced pressure, and 152 g was collected.

The substance was identified using the following method.

1) Identification column carrier PE by gas chromatography
As a result of comparison with commercially available β-DCH using G-20MP, 5%, 60-80 mesh, the retention time was exactly the same. Purity 98.2% or higher.

2) Identification by IR (infrared absorption spectrum) As shown in the chart shown in Figure 1, the absorption pattern is that of commercially available β-D.
It was exactly the same as CH.

From the above, it was clear that this substance was β-DCH. The following method was used to confirm that this substance was R-(10)-β-DCH.

1) Measurement of optical rotation The specific optical rotations of commercially available β-DCH and this substance are as follows.

Commercially available β-DCH [α] 萱 = o, o'' c = i, dichloromethane Main substance [α] 萱 = + 10.4°C = 1. Dichloromethane 2) R-(decade)-α-methoxy-α-tri Preparation of fluoromethylphenylacetate ester and analysis by high performance liquid chromatography R-(+)-α-methoxy-α-trifluoromethylphenylacetate chloride was reacted with commercially available β-DCH and this substance,
After preparing the ester derivative, the analysis results by liquid chromatography were as follows.

Analysis conditions Column carrier ZORBAXODS 4.6mmX25Cm (manufactured by DOPOnt) Eluent
Meta/-/L, : Water = 65 : 35 (V/V
) Elution ffi 11Idl/min detection method
As a result of absorbance analysis at 260 nm, commercially available β-DCH gave two peaks with the same area at retention times of 50.5 minutes and 52.0 minutes.

This substance gave a peak only at a retention time of 52.0 minutes and did not give a peak at 50.5 minutes.

3) Preparation of dichloropropyl-N-phenyl carbamate and its optical rotation Commercially available β-DCH1, 1 g of this substance, and 0.90 g of phenyl isocyanate were added to 30 d of dry acetone and 0.3 rIII of triethylamine, heated under reflux for about 3 hours, and the dichloropropyl Propyl-N-phenylcarbamate Commercially available β-DCH [α] tube = 0.0" C = 1, methanol Main substance [α] tube = + 16.4° C = 1, methanol Based on the above results, this substance has R - (+)-β-DCH
It was found that its optical purity was 99% or more.

Next, add this R-(10)-β-DCH100IJ to 1.4
The mixture was mixed with 650 mg of N caustic soda aqueous solution in a 1000 rd flask and stirred vigorously at room temperature for 80 minutes, and 20 M of ether was added and after stirring, the ether layer was separated.

Subsequently, the ether layer was dried with magnesium sulfate, the ether was distilled off, and epichlorohydrin was distilled off to give 60. I got 30. The purity of this epichlorohydrin was 99.4% or more as measured by gas chromatography. Moreover, the specific optical rotation was as follows.

[α] = +34.3° (C = 3.4, methanol), that is, the obtained product (chlorohydrin is S-(+)
-Epichlorohydrin, and its optical purity was 99% or more.

Example 2 Same as Example 1, yeast extract 1.0%, polypeptone i
．． 0%, glycerin 2.0%, pH 7.0 medium 2
g was placed in a 51-volume Jarf 1-mentor and sterilized by heat in the usual manner.The -38 strain was inoculated into OS- and cultured under the same conditions as in Example 1 for 24 hours.

This culture is then 100.1! Synthetic medium 80j shown in Example 1 for Jarf 1-Mentor! and calcium carbonate 1 60g, racemic β-DCH 32oIdl,
Put it together with polypeptone 40Q, heat sterilize it, and then inoculate it as usual, at a temperature of 30℃ and aeration volume of 40J! /min
The reaction was carried out while culturing under the conditions of rotation speed of 30 rpm.

48 hours after the start of the reaction, the reaction solution is separated into supernatant, bacterial cells, and precipitate using a centrifuge, and the remaining β is removed from the supernatant.
-DCH was fractionated in the same manner as in Example 1, and R- (10)-
148 g of β-DCH was obtained.

The specific optical rotation of the obtained R-(+)-β-DCH is [α]
萱=+10.4' (C=1.0, dichloromethane)
As a result of analysis in the same manner as in Example 1, the optical purity was 9.
It was over 9%. Further, the conversion of R-(10)-β, -DCH to S-(+)-epichlorohydrin was carried out in the same manner as in Example 1, and the specific optical rotation [α] 91 = +3
4.3° (C=3.4, methanol) Optical purity 99
% or more of S-(+)-epichlorohydrin was obtained.

(Effects of the Invention) According to the present invention, optically active R-(10)-2 can be produced more easily and with higher purity than 2,3-dichloro-1-propanol by using bacteria belonging to the genus Pseudomonas isolated from soil.
, 3-dichloro-1-propanol to optically active S
-(±)-epichlorohydrin can be obtained.

[Brief explanation of drawings]

Figure 1 shows R, the raw material of the present invention obtained in Example 1.
It is an infrared absorption spectrum of -(+)-2,3-dichloro-1-propanol and a commercially available product of the same substance. □ indicates commercially available β-DCH, and -m- indicates R-(+)-β-DCH.

Claims (2)

[Claims] (1) Bacteria belonging to the genus Pseudomonas having the ability to assimilate S-(-)-2,3-dichloro-1-propanol, or their cultured cells, are grown in a culture medium to produce racemic 2,3-dichloro-1-propanol.
R-(+)-2 obtained by reacting with 1-propanol
, 3-dichloro-1-propanol with an alkali agent to obtain S-(+)-epichlorohydrin. (2) Claim 1, which uses a bacterium belonging to the genus Pseudomonas having the ability to assimilate S-(-)-2,3-dichloro-1-propanol, or a cultured cell thereof, after being immobilized.
Manufacturing method described in section.