JP2875644B2 - Separation method using optical splitting membrane - Google Patents

Description

DETAILED DESCRIPTION OF THE INVENTION

[0001]

BACKGROUND OF THE INVENTION 1. Field of the Invention The present invention relates to a method for separating an optically active substance using an optical resolution membrane, and more particularly, to a method for separating an optically active substance using an optical resolution membrane comprising a polymer containing a polyamino acid having an α-helical structure as a racemate. To a novel separation method which enables efficient separation when separating into D, L-optically active substances.

[0002]

2. Description of the Related Art Many organic compounds have asymmetric centers, and optical isomers derived therefrom exist. The optical isomers hardly differ in physical properties such as boiling point and solubility. But,
Many differences in bioactivity are often found. Therefore, obtaining one of the optical isomers (D-form or L-form) is very useful in the fields of pharmaceuticals, agricultural chemicals, foods and the like. For example, in the case of glutamic acid, the L-form (S) has an umami but the D-form (R) has no umami, and in the case of aspartame sweetener, the S-form shows a sweet taste but R
The body is said to have a bitter taste. Even in the case of pharmaceuticals, the D-form and L-form may show significant differences in their efficacy, toxicity, etc., and the Ministry of Health and Welfare states in the 1985 edition of the Pharmaceutical Manufacturing Guidelines that “if the drug is a racemic form, It is desirable to study absorption, distribution, metabolism, and excretion kinetics of "."

Various means for obtaining an optically active substance from a racemic body have been devised based on such social demands. As a method for obtaining an optically active substance from a racemate, there are presently a preferential crystallization method, a diastereomer method, an enzymatic method, a chromatography method and the like. The preferential crystallization method is a method in which a desired crystal is inoculated in a racemic supersaturated solution, and only the crystal is grown and precipitated. Despite the excellent method, the track record is low for the following reasons. That is, in order to resolve a racemate by the preferential precipitation method, first measure the solubility of the racemate and both actives, racemate> active, melting point is higher in the active than the racemic It is necessary to confirm that the active substance is not soluble in a racemic saturated solution, and that the infrared absorption spectra of the racemic form and the active form match (Hiroshi Yamanaka, Yasuhisa Tashiro, Quarterly Chemistry Review) ,
No. 6, 1989, pp. 4-5). In addition, the priority crystallization method can be applied only to a special crystal because the timing of the solid-liquid separation and the reduction of the filtration time are technically important issues.

The diastereomer method is a method in which an optically active acid or base is allowed to act on a racemate, and the resulting diastereomer salt is separated by fractional crystallization according to the difference in solubility. The difficulty in selecting a resolving agent is associated with the fact that the resolving agent must readily form a salt or derivative with the racemate. In addition, there are limitations that it is considerably difficult to obtain a high-purity optically active substance and that an equal amount of a resolving agent as that of a racemic form is required.

The optical resolution method using an enzyme utilizes the stereospecificity of the enzyme for a substrate. This method is suitable as a method for obtaining a large amount of an optically active substance, and for example, an industrial scale production technique of D-amino acid by an enzymatic method combining a hydantoinase reaction and a chemical decarbamylation reaction has been established [S. TAKAHASHI, “Biotechnologyof A
minoacid Production ”, H. YAMADA et al (eds), Koda
nsha Ltd, (1986), p269]. Or U.S. Pat.No. 4,800,1
No. 62 describes a method for obtaining an optically active substance by immobilizing an enzyme on a capillary type membrane. However, in the case of the enzymatic method, it is extremely difficult to find an enzyme that is compatible with the target racemate to be resolved, and thus has the disadvantage that it can be applied only to a very limited racemate.

The chromatographic method using a chiral compound as a stationary phase is a method of optical resolution by the interaction between a D-form and an L-form and a filler. Advances in HPLC (High Performance Liquid Chromatography) and the development of packing materials with great optical recognizability have expanded the range of target compounds and improved processing capacity, but are still economical on an industrial scale. Has not reached the limit.

[0007] The separation method using a membrane is a novel technique that overcomes the inherent disadvantages of each of the above-mentioned preferential crystallization method, diastereomer method, enzymatic method, chromatographic method, etc. It can be said to be a revolutionary technology that economically obtains the body. The present inventor has previously described Japanese Patent Application No.
An object of the present invention is to provide a novel method for dramatically improving the separation performance of a membrane in a racemic separation method using a membrane.

[0008]

Means for Solving the Problems The present inventor has disclosed in Japanese Patent Application No. Hei.
When a racemate is separated into D, L-optically active substances using a membrane comprising a polymer having a polyamino acid having an α-helix structure as disclosed in the specification of JP-A-334352, the racemate located on the surface side of the membrane is separated. It has been found that the addition of an inorganic salt to the permeated water side located on the body side and / or the opposite side thereof dramatically improves the separation performance of the membrane, and has completed the present invention. That is, the present invention provides a method for separating a racemate into D, L-optically active substances using an optical resolution membrane composed of a polymer containing a polyamino acid having an α-helical structure as a component, and / or It is an object of the present invention to provide a method for separating an optically active substance using an optical splitting membrane, characterized by adding an inorganic salt to a racemic body.

In the present invention, the inorganic salt may be added to both the permeated water side and the racemic side, but the same effect can be obtained by simply adding it to the racemic side or the permeated water side. As the inorganic salt used in the present invention, a monovalent or divalent metal salt is preferable. Specifically, as the cation,
Na ⁺ , K ⁺ , Li ⁺ , Ca ⁺⁺ , Mg ⁺⁺ , Ba ⁺⁺ , Sr ⁺⁺ are valid,
These counter ions _{^{Cl -, Br -, NO 3}} -, HPO 3 -, H
PO ₄ ^- and SO ₄ ^- are used. The addition amount of the inorganic salt is 1 to 100 per liter of permeated water or a racemic solution.
A range of mmol is preferred.

The optical resolution film used in the present invention comprises a polymer containing a polyamino acid having an α-helical structure as disclosed in Japanese Patent Application No. 2-334352, preferably α-helix. It consists of a polymer obtained by reacting a polyamino acid having a helical structure with a compound having an amphiphilic group. Examples of the polyamino acid having an α-helical structure used in the present invention include poly (γ-methyl-L-glutamate), polylysine, polyalanine, polyphenylalanine, polyleucine and the like.
The degree of polymerization of the polyamino acid is not less than the lowest degree of polymerization having an α-helical structure, preferably not less than 15. Suitable compounds having an amphiphilic group used in the present invention are polyalkylene oxide compounds having an aromatic derivative at the terminal, and include, for example, compounds represented by the following formula (I).

[0011]

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(Wherein n = 1 to 50, R represents an alkyl group having up to 20 carbon atoms.) The reaction between a polyamino acid having an α-helical structure and a compound having an amphipathic group is carried out by using paratoluene sulfone. A normal transesterification reaction may be performed using a catalyst such as an acid. The method for producing a film comprising a polymer containing a polyamino acid having an α-helix structure as described above is not particularly limited, and the polymer may be dissolved in a solvent and formed into a film by an ordinary film forming method. it can.

[0013]

EXAMPLES Hereinafter, the present invention will be described in more detail with reference to Examples, but the present invention is not limited to these Examples. Example 1 According to the method described in Example 1 of Japanese Patent Application No. 2-334352, a polymer represented by the following structural formula (II) (hereinafter referred to as NON6-PL
G).

[0014]

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That is, first, the average molecular weight as a polyamino acid
Using 100,000 poly (γ-methyl-L-glutamate) (manufactured by Ajinomoto Co., Inc.) (PMLG), a 15% by weight solution of PMLG in 1,2-dichloroethane (DCE) was azeotroped with purified DCE and dehydrated. Was. The water content after dehydration is preferably 0.1% by weight or less. DCE was prepared by treating a commercially available reagent of special grade with an alkali to remove acid components, followed by washing with water, drying and distillation. As a poly (oxyethylene) nonylphenol ether having an amphiphilic group to be reacted with PMLG, Nonipol 60 having an average number of oxyethylene groups (nominal value) of 6 (Sanyo Chemical Industry Co., Ltd.)
Was used. Nonipol 60 was also dehydrated to a water content of 0.1% or less by azeotropic distillation with purified DCE.

The polymer was synthesized according to the following procedure.
A three-necked flask was charged with a 5% by weight solution of PMLG in DCE, and a commercially available reagent, primary grade p-toluenesulfonic acid (PTS) was used as a catalyst.
Was added at 1% by weight of PMLG. While continuing stirring with a three-one motor, a DCE solution of Nonipol 60 was added little by little to proceed the reaction. The amount of Nonipol 60 added was stoichiometrically 1.2 times that of PMLG. During the reaction, the solution temperature
Maintained at 65 ° C. Methanol generated as the reaction proceeded was removed from the system together with DCE by reducing the pressure in the flask. After this operation was continued for 4 to 5 hours every day for 18 days, a DCE solution of Nonipol 60 was added so that the molar ratio between the PMLG repeating unit and Nonipol 60 was 1:10. After removing DCE remaining in the flask as much as possible by operating the inside of the flask under reduced pressure, methanol of the first grade reagent was added, and the mixture was vigorously stirred to precipitate the polymer. The polymer was further washed with methanol, then dissolved in DCE solution, and reprecipitated in primary reagent n-hexane. The reprecipitate was dried under reduced pressure to obtain NON6-PLG.

This polymer was dissolved in a dichloroethane solution purified by a conventional method to prepare a dope of 0.0025% by weight. The film was formed by forming a circular hollow 2 having a diameter of 32 mm on an 8 mm thick sheet 1 made of silicon rubber as shown in FIG. 1. A polyimide ultrafiltration membrane (Nitto Denko Corporation)
(Fraction molecular weight: 20,000) was sandwiched in such a manner that the surface was on the top, and was used as a base material. That is, 0.5 cc of the prepared dope was dropped on the base material and dried at room temperature under a nitrogen atmosphere for 20 to 30 minutes. Next, a further 0.5 cc of dope was again dropped on this film and dried overnight at room temperature under a nitrogen atmosphere. Subsequently, the film was dried at room temperature for 24 hours under a reduced pressure of 1 to 5 Torr, and then immersed and washed in purified water for 1 week to obtain a film. The thickness of the film was calculated to be 25 nm.

The optical resolution experiment of the racemate was performed using the apparatus shown in FIG. The inner volumes of the undiluted liquid side chamber 3 and the permeated water side chamber 7 are 10
At 0 cm ³ and 4 cm ³ , the effective area of the membrane 9 was 8.04 cm ² .
Both chambers were equipped with a water jacket 6, which was maintained at 34 ° C. by circulating constant temperature water. Stock solution inlet 4
, A racemic tryptophan (Trp) with a concentration of 5 × 10 ⁻³ mol / dm ³ was added to the stock solution side, and purified water was added to the permeate side.
Permeation was performed by a concentration gradient. The undiluted solution side was stirred by a magnetic stirrer 5 and the permeated water side was circulated using the same method or by using an external pump 8 so that the concentration in each chamber was always uniform.

The permeate-side solution was withdrawn at regular intervals and the mass was measured. The concentrations of D-tryptophan and L-tryptophan were calculated by determining the peak area ratio between an aqueous solution of tryptophan whose concentration was known in advance and the sampled solution by liquid chromatography.
From the previously measured mass and the concentrations of the D-form and the L-form, the respective transmission amounts were determined. The result is shown in FIG. The permeated water side after sampling was refilled with purified water each time, and the experiment was continued. As shown in FIG. 3, neither the D-form nor the L-form permeated to the permeated water side until 560 hours after the start of the permeation experiment.
After 525 hours, 50 mmol / liter NaCl was added to the undiluted solution side and the permeated water side. As a result, D
-Only tryptophan began to permeate. After the elapse of 755 hours, the membrane was once removed from the apparatus, washed sufficiently with water, and then reset. Thereafter, the permeation experiment was carried out without adding NaCl to both the undiluted solution side and the permeated water side chamber.
Almost no permeation was observed with tryptophan. Thus, it is clear that the addition of NaCl enables efficient separation of the optically active substance.

Example 2 A NON6-PLG polymer prepared according to the formulation shown in Example 1 was dissolved in a dichloroethane solution purified by a conventional method to prepare a 1% by weight dope. A4 as base material
A dope is dropped onto a large polyimide ultrafiltration membrane (manufactured by Nitto Denko Corporation) (fraction molecular weight: 20,000), and the dope is leveled with a thin rod wrapped with a stainless steel wire to coat the dope to a thickness of about 50 μm. To form a film. The obtained film was dried overnight at room temperature under a nitrogen atmosphere, and then dried at room temperature under a reduced pressure of 1 to 5 Torr for 24 hours. Then put 1 in purified water for washing.
Soaked for a week.

This membrane was set in the filtration experiment apparatus shown in FIG. 2, and purified water was placed in both the undiluted solution side and the filtered water side at 65 ° C.
For 50 hours. After the completion of annealing, natural cooling (gradual cooling) was performed for 2 to 3 hours to lower the temperature to 34 ° C. A racemic tryptophan at a concentration of 5 mmol / liter was placed in the undiluted solution side chamber, and a permeation experiment was performed. In other words, D and L-tryptophan respectively permeated up to 500 hours, but after 500 hours, 25 mmol /
When 1 liter of Na ₂ HPO ₄ was added, the permeation of L-tryptophan stopped and only D-tryptophan began to permeate. After 1500 hours, the membrane was removed from the experimental apparatus, washed thoroughly with water, reset, and the experiment was continued in a state where both chambers did not contain Na ₂ HPO ₄ .L-tryptophan did not permeate, only D-tryptophan. Transmitted.

From the above results, it was found that the addition of Na ₂ HPO ₄ on the racemic side and the permeated water side resulted in a difference in the permeation speed of the D and L forms and good separation performance.

Example 3 A dope prepared in the same manner as in Example 2 was leveled with a rod of stainless steel wire wound so as to have a thickness of 20 μm to form a film. The thickness of the film was calculated to be 200 nm.

This membrane was set in the experimental apparatus shown in FIG.
Put 5 mmol / L of racemic tryptophan in the undiluted solution chamber and put purified water on the permeate side, and adjust the temperature of both chambers.
The film was kept at 65 ° C. for 50 hours to perform annealing of the film. The mixture was allowed to stand at room temperature for natural cooling (gradual cooling), and the liquid temperature was lowered to 34 ° C. In this state, a transmission experiment was started, and the results shown in FIG. 5 were obtained.

That is, until 550 hours have passed, D, L
-Tryptophan permeated at approximately the same rate, at which time NaCl was added to both chambers to a concentration of 50 mmol / l, and selective permeation of D-tryptophan was observed. After the elapse of 850 hours, the membrane was removed from the experimental apparatus, thoroughly washed and reset, and 5 mmol of racemic tryptophan containing no NaCl was placed on the stock solution side of the experimental apparatus.
/ L concentration solution and Na ₂ HPO to 25mmol / L concentration
₄ was added. Add purified water to the permeate side and add Na
₂ HPO ₄ was added to adjust the concentration to 25 mmol / l and the permeation experiment was continued. As a result, almost no permeation of L-tryptophan was performed, and the permeation speed of D-tryptophan was significantly increased.

[Brief description of the drawings]

FIG. 1 is a perspective view of a silicon rubber sheet used for forming a film in Example 1.

FIG. 2 is a cross-sectional view of an apparatus used in an optical splitting experiment of an example.

FIG. 3 is a graph showing the results of an optical splitting experiment of Example 1.

FIG. 4 is a graph showing the results of an optical splitting experiment of Example 2.

FIG. 5 is a graph showing the results of an optical splitting experiment of Example 3.

[Explanation of symbols]

 Reference Signs List 1 Silicon rubber sheet 2 Circular hollow 3 Stock solution side chamber 4 Stock solution inlet 5 Magnetic stirrer 6 Water jacket 7 Permeated water side chamber 8 Pump 9 Membrane

Continuation of the front page (51) Int.Cl. ⁶ identification code FI C07D 209/20 C07D 209/20 // C07M 7:00 (58) Field surveyed (Int.Cl. ⁶ , DB name) C07B 57/00 C07B 63/00 B01D 61/00 B01D 71/56 B01D 71/74 C07D 209/20

Claims (3)

(57) [Claims] 1. When separating a racemate into D, L-optically active substances using an optical resolution membrane comprising a polymer comprising a polyamino acid having an α-helix structure as a constituent, the permeate side and / or the racemate A method for separating an optically active substance using an optical splitting membrane, characterized by adding an inorganic salt to a body side. 2. The method according to claim 1, wherein the inorganic salt is a salt of a monovalent or divalent metal. 3. The method according to claim 1, wherein the amount of the inorganic salt added is 1 to 100 mmol per liter of permeated water or a racemic solution.