JPS6254479B2 - - Google Patents

Description

[Detailed description of the invention]

The present invention relates to a biochemical method for producing (S)-(-)-α-cyano-3-phenoxybenzyl alcohol. More specifically, it belongs to the genus Chromobacterium and is an organic carboxylic acid (saturated or unsaturated organic carboxylic acid having 1 to 18 carbon atoms) ester of (R S)-α-cyano-3-phenoxybenzyl alcohol. On the other hand, an esterase produced by a microorganism with asymmetric hydrolysis ability is allowed to act on the ester at a pH of 7 or less to preferentially asymmetrically hydrolyze the (S)-(-) alcohol ester, resulting in high optical purity. (S)-(-)-α-cyano- by obtaining the ester of (S)-(-)-α-cyano-3-phenoxybenzyl alcohol and its enantiomer.
The present invention relates to a biochemical production method of 3-phenoxybenzyl alcohol. α-Cyano-3-phenoxybenzyl alcohol is known as a novel alcohol component of a group of ester compounds called synthetic pyrethroids, which have excellent insecticidal activity. Synthetic pyrethroids, such as fuenvalerate, cypermethrin, and deltamethrin, which contain this alcohol as an alcohol component, have dramatically increased efficacy and are resistant to sunlight, so they are useful not only for household use and epidemic prevention, but also for agricultural chemicals. It is also attracting attention as an insecticide, and therefore the importance of α-cyano-3-phenoxybenzyl alcohol is increasing. Since the α-cyano-3-phenoxybenzyl alcohol has an asymmetric carbon at the α-position, two types of optical isomers exist, and the insecticidal efficacy as an ester is higher than that of the S-form alcohol. is known to be far superior to the corresponding R-isomer (Kosuke Yoshioka, Organic Synthetic Chemistry, Vol. 38, No. 12, 1152-
1162 pages (1980)). Therefore, (R.S)-α-cyano-3 obtained by ordinary synthetic chemical production methods
It has been desired to develop a technique for optically resolving -phenoxybenzyl alcohol using an industrially advantageous method to obtain the S-form. By the way, the α-cyano-3-phenoxybenzyl alcohol is an unstable compound, and it is difficult to employ the conventional optical resolution method of alcohol via ester or half-ester with a normal optically active organic acid. Until now, (S)-(-)-α-cyano-
As a method for obtaining 3-phenoxybenzyl alcohol, (i) (R S)-α-cyano-3-phenoxybenzyl alcohol is cis-
2,2 dimethyl-3S- (dihydroxymethyl)
reacting with cyclopropane-1R-carboxylic acid lactone to give an ether compound and separating the resulting two isomer mixtures by physical means;
A method of hydrolyzing an ether compound containing a (S)-(-) alcohol component in an acidic medium to obtain the desired (S)-(-)-α-cyano-3-phenoxybenzyl alcohol (Unexamined Japanese Patent Publication No. Showa 54-109944
No.) and (ii) (S) of chiral and cyclopropane carboxylic acids
-(-)-α-Cyano-3-phenoxybenzyl alcohol ester is reacted with boron halide and then water is reacted with (S)-(-)-α-
Method for obtaining cyano-3-phenoxybenzyl alcohol (JP-A-56-12355). is only known. However, these methods also require steps, expensive optically active reagents, and low overall yields. Furthermore, in method (ii), optically active α-cyano-3- This method is not necessarily sufficient because it requires an ester of phenoxybenzyl alcohol. The present inventors have conducted repeated research to overcome these problems and find an industrially advantageous method for producing (S)-(-)-α-cyano-3-phenoxybenzyl alcohol. , racemate i.e. (R・S)
-By using ester of α-cyano-3-phenoxybenzyl alcohol as a raw material and biochemically asymmetrically hydrolyzing it, (S)-(-)-α-cyano-3-phenoxybenzyl It was discovered that alcohol can be efficiently separated into the corresponding R-alcohol ester, and after further various studies, the present invention was completed. That is, the present invention belongs to the genus Chromobacterium, and (R.S)-α-
(S)-
Esterases that have the ability to preferentially asymmetrically hydrolyze esters of (−)-isomer alcohols (Esterases in the present invention refer to esterases in a broad sense, including lipases that are active against water-insoluble ester substrates. ) to asymmetrically hydrolyze the ester of the racemic alcohol at a pH of 7 or less to produce (S)-(-)-α-cyano-3-phenoxybenzyl alcohol and its enantiomer ester. A method for producing (S)-(-)-α-cyano-3-phenoxybenzyl alcohol by resolution is provided. Used as a raw material in the present invention (R・
Examples of the organic carboxylic acid component of the ester of S)-α-cyano-3-phenoxybenzyl alcohol include formic acid, acetic acid, propionic acid, butyric acid, citric acid, isochenric acid, caproic acid, caprylic acid, caproic acid, Examples include lauric acid, myristic acid, palmitic acid, stearic acid, oleic acid, linoleic acid, etc., and esters of organic carboxylic acids having 2 to 18 carbon atoms are preferred from the viewpoint of ease of handling, optical purity of the reaction product, etc. Furthermore, ease of handling
Ester of acetic acid is more preferable from an economic point of view. These esters can be easily produced by a conventional method for producing esters, for example, by reacting (R.S)-α-cyano-3-phenoxybenzyl alcohol with an anhydride of an organic carboxylic acid or an organic carboxylic acid halide. can do. Also,
It can also be produced by reacting 3-phenoxybenzaldehyde with sodium cyanide and a halide of the organic carboxylic acid instead of (R.S)-α-cyano-3-phenoxybenzyl alcohol. Esterases that can be used in the present invention are esterases in a broad sense, including lipases produced by microorganisms belonging to the genus Chromobacterium. Specific examples of this microorganism include the following strains. Chromobacterium viscosum ATCC6918 Chromobacterium viscosum This strain is American Type Culture
Collection (ATCC) and can be obtained from this institution. The above-mentioned microorganisms can be cultured in a liquid culture according to a conventional method, for example, by inoculating the microorganisms into a sterilized bouillon liquid medium and culturing with reciprocating shaking at 20 to 40°C for 2 to 3 days, or if necessary. Solid state culture may also be used. Furthermore, some of these microbial-derived esterases are commercially available. This commercially available enzyme can also be used for purposes of the present invention. Commercially available enzymes that can be used in the present invention include those shown below.

[Table] When carrying out the method of the present invention, (R・S)-α-
The asymmetric hydrolysis of the organic carboxylic acid ester of cyano-3-phenoxybenzyl alcohol is carried out using an esterase-containing solution such as a culture solution, a culture filtrate, an esterase extract or a concentrate obtained by culturing the microorganism,
Alternatively, it may be carried out by mixing an aqueous solution containing crude esterase or purified esterase with the organic carboxylic acid ester of the (R·S)-alcohol, and stirring or shaking the mixture. If necessary, a non-ester surfactant may be added. It is also possible to immobilize the enzyme and use it. Further, the reaction temperature at this time is suitably 10 to 65°C, and preferably 20 to 50°C since the alcohol produced is relatively unstable to heat and the reaction rate is slow at too low a temperature. The reaction time is usually 3 to 48 hours, but the reaction time can be shortened by increasing the reaction temperature or using a highly active enzyme. Furthermore, since α-cyano-3-phenoxybenzyl alcohol is particularly easily decomposed by bases, it is important to control the pH of the aqueous solution during the enzymatic reaction. That is, even if asymmetric hydrolysis by esterase proceeds in a basic medium, the resulting alcohol will be denatured, so it is important to carry out the enzymatic reaction at a pH of 7 or lower. Furthermore, since the enzyme is inactivated if the pH is too low, it is preferable to carry out the reaction within the range of PH4.5 to PH6.5. Furthermore, in order to prevent the pH from decreasing due to organic acids such as acetic acid produced by hydrolysis, it is desirable to control the pH to a constant level by using a buffer solution or the like. As the buffer solution, either an inorganic acid salt buffer solution or an organic acid salt buffer solution can be used. The concentration of the ester substrate used is in the range of 1 to 80 w/w%, preferably in the range of 5 to 35%, based on the reaction solution, and the reaction can be carried out even at a high substrate concentration. Next, after performing the asymmetric hydrolysis reaction in this way, the liberated (S)-(-)-α-cyano-3-
Phenoxybenzyl alcohol and unreacted enantiomer ester are separated and recovered. For this separation and recovery, operations such as solvent extraction, fractional distillation, column chromatography, etc. can be appropriately employed. For example, the reaction solution is extracted with an organic solvent such as ether, benzene, or toluene, and this extract is fractionally distilled under reduced pressure to produce (S)-(-)-α-cyano-3-phenoxybenzyl alcohol and its opposite compound. Free (S)-(-)-α-cyano-3 can be obtained by separating the ester from the body or by separating the extract by column chromatography using silica gel or the like.
- Phenoxybenzyl alcohol can be obtained. The unreacted ester thus separated and removed can be epitomized by contacting with a basic compound such as ammonia, pyridine, triethylamine, etc., and can be used again as a raw material in the method of the present invention. Next, the present invention will be explained in more detail with reference to Examples, but the present invention is not limited thereto. Example 1 8.0 g of acetate ester of (R・S)-α-cyano-3-phenoxybenzyl alcohol and 20 mg of Chromobacterium esterase (Lipase Toyo, manufactured by Toyo Jozo Co., Ltd.) were mixed with acetate at a concentration of 0.2M. The mixture was added to 2 ml of buffer solution (PH4.5) and reacted at 30°C with stirring. In addition, during the reaction, a concentration of 1M was added as the reaction progressed.
The NaOH aqueous solution was made into minute water droplets using a drop controller, and the PH was controlled at a constant level using a PH controller while carefully adding the solution. After reacting for 24 hours, the reaction product was extracted with toluene. The extract was subjected to high performance liquid chromatography (HPLC) [Lichrosorb RP-
18, methanol-water (6:4), 254 nm, UV detection), acetic acid ester of α-cyano-3-phenoxybenzyl alcohol and α-cyano-3
- The hydrolysis rate was calculated from the peak area ratio with phenoxybenzyl alcohol. The extract was concentrated and subjected to silica gel column chromatography, eluted with a cyclohexane-ethyl ether (94:6) solution to decompose and remove unreacted acetic acid ester of α-cyano-3-phenoxybenzyl alcohol. Elute with methanol containing a trace amount (10 ^-5 %) of para-toluenesulfonic acid.
Free α-cyano-3-phenoxybenzyl alcohol was obtained. After distilling off the elution solvent, a portion of the alcohol was taken and its specific rotation was measured in chloroform. In addition, 10 mg of the obtained free α-cyano-3-phenoxybenzyl alcohol was added to 1 portion of toluene.
ml, add equimolar amounts of (S)-(+)-2-(4-chlorophenyl)-isovaleric acid chloride and pyridine, and react with α-cyano-3-phenoxybenzyl alcohol ( S)-(+)-2-(4
-chlorophenyl)-isovaleric acid diastereomer and gas chromatography (10% DOQF-
Optical isomer analysis was performed at 1, 2.5 m and 250°C).
The results are shown in Table 1.

[Table] Example 2 Put 100 ml of sweetened bouillon liquid medium (dissolve 10.0 g of glucose, 5.0 g of peptone, 5.0 g of meat extract, and 3.0 g of NaCl in 1 part of water to adjust the pH to 7.2) in a 500 ml flask with a shoulder. After sterilization, two platinum loops of Chromobacterium violaceum (IFO-12614) were inoculated from the slant culture, and cultured with reciprocal shaking at 30°C for 72 hours. Next, the pH of the culture solution was adjusted to PH5.0 using a 2M HCl aqueous solution, and (R・S)-α-cyano-
2.0 g of acetic ester of 3-phenoxybenzyl alcohol was added, and the mixture was reacted for 30 hours with stirring at 30°C. During the reaction, the pH was controlled to be constant in the same manner as in Example 1. Thereafter, the reaction products were separated in the same manner as in Example 1, and the optical isomer ratio and hydrolysis rate of the obtained α-cyano-3-phenoxybenzyl alcohol were measured. The results are shown in Table 2.

【table】

Claims (1)

[Claims] 1 Chromobacterium
belongs to the genus, and has asymmetric hydrolysis ability for organic carboxylic acid (saturated or unsaturated carboxylic acid having 1 to 18 carbon atoms) ester of (R・S)-α-cyano-3-phenoxybenzyl alcohol. Esterase produced by microorganisms is allowed to act on the ester at a pH of 7 or lower, and the ester is asymmetrically hydrolyzed to produce (S)-
(S)-(-)-α-cyano-3-phenoxybenzyl alcohol characterized by its separation into (-)-α-cyano-3-phenoxybenzyl alcohol and its enantiomer ester. Biochemical manufacturing method.