JPH03236796A - Optical resolution method by enzyme - Google Patents

Abstract

PURPOSE:To efficiently obtain useful agricultural chemical in high yield by reacting a specific compound with a hydrolytic enzyme or enzyme mixture containing one or more kind of hydrolytic enzymes. CONSTITUTION:A microorganism of the genus Candida such as Candida cylindracea strain is cultured to afford an asymmetric hydrolytic enzyme (A) such as lipase OF. Then methyl 2-substituted-2-benzyloxy-2-methyl-carboxylate (C) expressed by formula I (R is 2-12C saturated or unsaturated alkyl) and the above-mentioned enzyme A are added to a phosphoric acid buffer of pH5-8 to give the reaction mixture liquid (D). Then the ingredient D is subjected to reciprocating shaking reaction at about 30 deg.C for 2-3 day to provide a reaction product (E). Then the ingredient E is controlled to about pH9 and extracted and purified with diethyl ether, etc., to provide an optically active ester expressed by formula II (asteric C is optically active carbon) and/or optically active carboxylic acid expressed by formula III in an optical resolution method.

Description

DETAILED DESCRIPTION OF THE INVENTION (Field of Industrial Application) The present invention relates to a method for enzymatic optical resolution of ester derivatives. More specifically, the present invention is useful as an intermediate for the production of physiologically active substances that can be used as agricultural chemicals, etc. In the formula c, R is a saturated or unsaturated alkyl group having 2 to 12 carbon atoms, and R' is hydrogen or a methyl group.

Furthermore, a method for optically resolving and isolating a compound represented by the following formula (carbon atoms marked with the title are optically active carbon atoms) by asymmetric hydrolysis by the action of an enzyme on an ester derivative represented by the formula It is related to.

(Prior Art) Optically active forms of compounds having the general formula (wherein R is a saturated or unsaturated alkyl group) are important intermediates that can be derived, for example, into frontalin or other physiologically active substances. Such tertiary alcohol derivatives cannot be synthesized by reduction reaction. Furthermore, when considering the subsequent conversion reaction, it is difficult to protect the hydroxyl group with a benzyl group or the like that is resistant to acids and bases. For this reason, if an optically active substance of the type (2) is to be obtained by a biochemical method, a method using hydrolysis of a carboxylic acid ester may be considered.

However, there are some successful examples of derivatives in which the 0-position is 2-substituted [Tetrahedron Letters].
n Letters), Vol. 28, p. 1303, (1
987)], no effective method for optical resolution by asymmetric hydrolysis of sterically crowded 3M substituted carboxylic acid derivatives is known.

(1211 to be solved by the invention) An object of the present invention is to obtain optically active 0-position compounds represented by the general formula [1] that can be useful starting materials for the production of optically active substances such as various physiologically active substances and agricultural chemicals. A method for producing a carboxylic acid derivative having two carbon functional groups in addition to the hydroxyl group protected by The object of the present invention is to provide a method for obtaining a compound efficiently and in high yield.

(Means for Solving the Problems) The present inventor conducted studies to find a method for easily and efficiently producing an optically active compound represented by the general formula [I], which is an intermediate for producing various physiologically active substances. Result, formula [I]
The present inventors have discovered that the optically active form [I[] can be easily separated by asymmetrically hydrolyzing the ester derivative represented by the following with a certain enzyme, and have completed the present invention.

That is, methyl 2-benzyloxy-2-methylcarboxylate represented by the formula of the present invention is asymmetrically hydrolyzed by the action of a hydrolase or an enzyme mixture containing them, and R in the formula c has 2 to 2 carbon atoms. 12 saturated and unsaturated alkyl groups, and each carbon atom with a prefix is an optically active carbon atom) and/or an optically active carboxylic acid represented by the formula The present invention relates to a method for collecting optically active esters [11] represented by saturated and unsaturated alkyl groups, and carbon atoms with a prefix are optically active carbon atoms.

An example of the enzyme used in the method of the present invention is an enzyme produced by Candida eylindracea, a bacterium belonging to the genus Candida, which can asymmetrically hydrolyze the ester derivative represented by the formula [11]. is most preferred.

Enzymatic hydrolysis is desirably carried out at around pH 5 to 8, particularly preferably around pH 7. For this purpose, it is preferable to use a suitable buffer such as a phosphate buffer prepared from an aqueous solution of disodium phosphate or monosodium phosphate. Add the substrate ester represented by formula [I] and lipase to such a buffer solution, and add 3
Culture with reciprocal shaking is performed at 0°C for 2 to 3 days. After the reaction, add saturated aqueous sodium bicarbonate solution to adjust the pH to 9, extract with a suitable organic solvent such as diethyl ether, treat and purify by conventional methods to obtain optically active ester [111].

After separating the ester [11] from the carbon M[I] having the opposite configuration using the above method, the reaction mixture is made acidic;
It can be isolated by extraction with a suitable organic solvent, such as ethyl acetate, and treatment and purification according to conventional methods.

The optically active compound of formula [I] separated and purified by the method of the present invention can be reacted according to the method listed in the reference example to lead to frontalin, which is a type of insect pheromone that can be effectively used as a pesticide. I can do it.

(Examples) The present invention will be explained in more detail with reference to the following examples, but the present invention is not limited thereto.

Example 1 Compound [I
] (R=-CHCH=CH2, 10g) and lipase O
F (10 g) was added and the mixture was heated at 30° C. for 96 hours. After the reaction, a saturated aqueous sodium bicarbonate solution was added to adjust the pH to 9, and sodium chloride was added for salting out. This was extracted with diethyl ether and dried over anhydrous sodium sulfate. The solvent was distilled off under reduced pressure, and the residue was distilled under reduced pressure to obtain the compound (+)-[J”1 (R=-CH,, CH=CH2
) #4J (yield 40%). After the extraction, concentrated hydrochloric acid was added to the aqueous layer to adjust the pH to 2, and the aqueous layer was extracted twice with ethyl acetate. The organic layer was dried over anhydrous sodium sulfate. By distilling off the solvent under reduced pressure, the compound [old (R=-CH,C
4.7 g (yield 52%) of H=CH,) was obtained.

Compound (+)-[I”] (R=-CH2CH=CH2)
Physical property values and spectral data: b, p, 160-170°C 10.9 pus.

[a]. +3.80° (c=1.63.chloroform).

v,,,erll-' 3000.2950.174
0.1450.1215.1155.1110°102
0,920.735.700゜'HNMRδ (400MHz, weight o roform) 1.4
9(3[(.

s), 2.60 (2H, d, J=7.0Hzl,
3.75 (3H, s), 4.49 (2H, s), 50
1 to 5.21 (2H, m), 5.63 to 6.10 (I
H, m), 7.27-743 (5EI, m).

Elemental analysis calculated value: C,4H1,03: C,71,77; H
,7,69% Actual value: C,71,88,H,7,50
% This is an optical activity shift reagent, Tris[3-
Trifluoromethylhydroxymethylene-(-)-campholatcoeuropium (IN) [Eu(tfc)]
It was confirmed to be optically pure within the measurement error of the instrument by measuring a 400 MHz NMRX vector in the presence of a 400 MHz NMRX vector.

The IR spectrum of compound [II] (R=-CH2CH=CH2) supported the structure.

v,,,am-” 3100.3050.3000.
2950.1710.1B40.1500°1455,
1390.1280.1160.1120.1030.
920.745.7000 The compound (-)-[I
"1 (R=-CH2CH=CH2) and measured the physical property values and spectral data: b, p, 160 ~
170°C 10.9M0 [a]. -2,65° (c
=0.98. chloroform).

Its IR and NMR spectra show that the remaining unhydrolyzed (+)-[I”1(R=-CH2CH=C
It was consistent with those of H2).

Elemental analysis calculated value: C, 4H, 03: C, 71, 77; H
,7,69% Actual value: C,72,10,H,7,41
%This one is 400MHz in the presence of Eu(tfe)3
Optical purity 82 by measuring NMRX vector
It was confirmed that %.

Examples 2-5 In the same manner as in Example 1, the results shown in Table 1 below were obtained.

Table 1 ■ Example R Compound [I"1 Compound [nl 2H5 C0H7 CH 6 2 2 1 0 8 1 5 〉99 4 8 5 Reference Example Compound (-)-[I"] (obtained according to Example 1) R=
-CH2CH=CH2) was further subjected to asymmetric hydrolysis using lipase OF to obtain >99% e, e, [II]
(R=-CH2CH=CH2) was obtained.

Using this product as a starting material, frontalin, a type of insect pheromone that can be effectively used as an agricultural chemical, was synthesized with a total yield of 382%.

Pan/acetone (93,9% L! dlp-toluenesulfonyl chloride 1:/pyridine (93,8%), (e1
Lithium bromide, carbonated water: Natrisom/acetone (94
, 2% L (fl magnesium/tetrahydrofugin;
Acetaldehyde: Rinnn/-acid pyrinnium/chicken (65,5%), (g) 2N tr
Aa/') Echi/L, x -te/L, (822
%).

Note that in the above, Lipase OF is the product name of Meito Sangyo (Mori).

(-)　-Fronta　l　i　n

Claims (3)

[Claims] (1) Formula ▲ Numerical formulas, chemical formulas, tables, etc. ▼ 2-benzyloxy-2-methylcarboxylic acid represented by [I] (in the formula, R is a saturated or unsaturated alkyl group having 2 to 12 carbon atoms) To optically resolve methyl, a hydrolase or an enzyme mixture containing at least one kind of hydrolase is applied to the above compound, and the formula ▲ includes mathematical formulas, chemical formulas, tables, etc. ▼ [ I ^ *] (in the formula, R is carbon 2 to 12 saturated and unsaturated alkyl groups, and the carbon atoms marked with * are optically active carbon atoms) and/or optically active esters represented by the formula ▲
There are mathematical formulas, chemical formulas, tables, etc. ▼ [II] (In the formula, R is a saturated or unsaturated alkyl group with 2 to 12 carbon atoms, and the carbon atoms marked with * are optically active carbon atoms.) An optical resolution method characterized by collecting optically active carboxylic acids. (2) The formula in which the enzyme is produced by a microorganism belonging to the genus Candida [
The optical resolution method according to claim 1, wherein the compound represented by [I] is an asymmetric hydrolyzing enzyme. (3) The optical resolution method according to claim 1 or 2, wherein the enzyme is lipase OF.