JPH11255671A - Chromatographic separation of optical isomer - Google Patents

Abstract

PROBLEM TO BE SOLVED: To provide a chromatographic separation of optical isomers in which a polysaccharide derivative chiral stationary phase is used as a chiral stationary phase and the mobile phase that has high separation performance, particularly giving a high fractionation productivity in the fractionation process, is used as a mobile phase. SOLUTION: In this chromatographic separation of optical isomers, a polysaccharide derivative chiral stationary phase is used as a chiral stationary phase, while a mixed solvent comprising a solvent selected from the group consisting of ethers, ketones, esters, amides and halogenated solvents, and hydrocarbon solvents is used as a mobile phase solvent.

Description

DETAILED DESCRIPTION OF THE INVENTION

[0001]

The present invention relates to a chromatographic separation method having a high separation effect in the optical resolution of optical isomers, and more particularly to a chromatographic separation method capable of obtaining high preparative productivity when performing chromatographic preparative separation. Things.

[0002]

2. Description of the Related Art It is well known that polysaccharides and derivatives thereof, for example, ester derivatives such as cellulose and amylose, and carbamate derivatives exhibit high optical resolution. It is also well known that chromatographic separating agents in which these are physically adsorbed and supported on silica gel are excellent separating agents exhibiting a wide range of optical resolution, a high number of stages, and durability (Y. Okamoto). , M. Kawashima and K. Hatada, J. Am. Chem. S
oc., 106, 5357, 1984).

[0003] However, in the above-mentioned physical adsorption type separating agent, a solvent capable of dissolving or swelling the polysaccharide derivative cannot be used for a mobile phase, and there is a limitation in selection of separation conditions. Further, there is a disadvantage that the washing solvent is limited even in the washing of contaminants strongly adsorbed on the separating agent. Due to these drawbacks, the polysaccharide derivatives are directly chemically bonded to silica gel (Japanese Patent Application Laid-Open Nos. 62-270,602 and 7-138,301). After coating the polysaccharide derivatives on silica gel, the polysaccharide derivatives are crosslinked with each other ( JP-A-8-59,702 or the like, or by coating a polysaccharide derivative on silica gel and then coating the surface of the polysaccharide derivative with a synthetic polymer (Japanese Patent Application No. 9-271,064, etc.) to immobilize the polysaccharide derivative on silica gel. Also, an immobilized type separating agent having solvent resistance has been proposed.

[0004] In recent years, attention has been paid to a preparative method (production method) of optical isomers using a polysaccharide derivative-based chiral stationary phase, and various preparative methods have been proposed for specific substances to be separated. (JP-A-61-161226, JP-A-61-267,536, JP-A-2-69,456, W
O95 / 23125, etc.).

However, since the chiral stationary phases disclosed in these prior arts are physical adsorption type separating agents, the types of mobile phase solvents are limited, and most of them are hexane / alcohol solvents. In this case, there is a disadvantage that a sample having low solubility in a hexane / alcohol-based solvent (object to be separated) can obtain only low preparative productivity.

[0006] On the other hand, specific mobile phases in the case of performing chromatographic separation of optical isomers using the above-mentioned immobilized type separating agent have not been well studied, and particularly, high preparative productivity. Was not known for the mobile phase giving

[0007] Against this background, the problem to be solved by the present invention is to achieve high separation performance in a chromatographic separation method using a polysaccharide derivative-based chiral stationary phase as the chiral stationary phase, and particularly high performance in the case of fractionation. It is an object of the present invention to provide a method for chromatographic separation of optical isomers using a mobile phase solvent which gives preparative productivity.

[0008]

Means for Solving the Problems The inventors of the present invention have made intensive studies to solve the above-mentioned problems, and as a result, have reached the present invention. That is, the present invention uses a polysaccharide derivative-based chiral stationary phase as the chiral stationary phase, and as the mobile phase solvent, a solvent selected from the group consisting of ether solvents, ketone solvents, ester solvents, amide solvents, and halogen solvents. (Hereinafter referred to as solvent B) and a hydrocarbon-based solvent (hereinafter referred to as solvent A).

[0009]

Embodiments of the present invention will be described below in detail.

In the present invention, the polysaccharide derivative-based chiral stationary phase is an immobilized type separating agent. Specifically, the polysaccharide derivative is directly chemically bonded to the carrier. Or by coating a polysaccharide derivative on a carrier and then coating the surface of the polysaccharide derivative with a synthetic polymer, thereby fixing the polysaccharide derivative to the carrier and imparting solvent resistance. These polysaccharide derivative-based chiral stationary phases can be produced by a known method described in the above-mentioned literature. In addition, a polysaccharide derivative-based chiral stationary phase to which insolubility has been imparted by performing treatment such as light irradiation and radical polymerization may be used.

The polysaccharide forming the polysaccharide derivative-based chiral stationary phase of the present invention is not particularly limited to any one of synthetic polysaccharides, natural polysaccharides and modified natural polysaccharides, as long as they are optically active, but are preferably bonded. A highly regular style is desirable. For example, β-1,4-glucan (cellulose), α-1,4-glucan (amylose, amylopectin), α-1,6-glucan (dextran), β-1,6
-Glucan (Bustran), β-1,3-glucan (eg, curdlan, schizophyllan, etc.), α-1,3-glucan, β-1,2-glucan (Crown Gall polysaccharide), β-1,4
-Galactan, β-1,4-mannan, α-1,6-mannan, β-1,2-fructan (inulin), β-2,6-fractan (levan), β-1,4-xylan, β -1,3-
Xylans, β-1,4-chitosan, α-1,4-N-acetylchitosan (chitin), pullulan, agarose, alginic acid, etc., and also starches containing amylose.
Among these, cellulose, amylose, β-1,4-xylan, β-1,4-
Chitosan, chitin, β-1,4-mannan, inulin, curdlan and the like are preferred, and cellulose and amylose are particularly preferred.

The number average degree of polymerization of these polysaccharides (average number of pyranose or furanose rings contained in one molecule) is 5 or more, preferably 10 or more, and there is no particular upper limit.
It is desirable that it is 500 or less from the viewpoint of easy handling.

The polysaccharide derivatives used in the present invention include:
Compounds obtained by derivatizing a compound having a functional group capable of reacting with a hydroxyl group in a part of the hydroxyl group of the polysaccharide as described above by forming an ester bond, a urethane bond, an ether bond, or the like by a conventionally known method. . Here, as a compound having a functional group capable of reacting with a hydroxyl group,
Any isocyanic acid derivative, carboxylic acid, ester, acid halide, acid amide, halide, epoxy compound, aldehyde, alcohol or other compound having a leaving group may be used, and these may be aliphatic, alicyclic or aromatic. Group or heteroaromatic compounds can be used. Among these compounds, compounds represented by the following general formula (I) are particularly preferred.

[0014]

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(Wherein X is a halogen atom or a C 1 -C 1)
3 represents an alkyl group, and n represents a number of 1 to 3. Particularly preferred as the polysaccharide derivative used in the present invention are polysaccharide ester or carbamate derivatives having 0.1 or more ester bonds or urethane bonds per monosaccharide.

The carrier used for producing the polysaccharide derivative-based chiral stationary phase of the present invention includes a porous organic carrier or a porous inorganic carrier, and is preferably a porous inorganic carrier. Suitable as a porous organic carrier is a polymer material composed of polystyrene, polyacrylamide, polyacrylate, and the like, and suitable as a porous inorganic carrier are silica, alumina, magnesia, glass, kaolin, titanium oxide, silica, and the like. Acid salt, hydroxyapatite, etc., and a particularly preferred carrier is silica gel. The particle size of the carrier is 0.1 μm to 10 mm, preferably 1 μm to 300 μm, and the average pore size is 10 μm to 100 μm, preferably 50 μm to 50 μm.
000 Å. When silica gel is used as the carrier, it is desirable that the surface is subjected to a surface treatment in order to eliminate the influence of the residual silanol, but there is no problem even if the surface treatment is not performed at all.

In the present invention, the mobile phase solvent is a mixed solvent obtained by combining the above-mentioned solvent A and solvent B. Of the solvent B, tetrahydrofuran (T
HF) and the like, acetone and the like as ketone solvents, ethyl acetate and the like as ester solvents, N, N-dimethylformamide, N, N-dimethylacetamide and the like as amide solvents, and halogen solvents as the solvent. Chloroform, methylene chloride and the like can be mentioned. Hexane and the like are mentioned as the hydrocarbon solvent of the solvent A. The mixing ratio of the solvent A and the solvent B in the mixed solvent used in the present invention is as follows:
/ B (capacity ratio) = 95/5 to 10/90, preferably 90/10 to
20/80 is more preferred.

The mobile phase solvent in the present invention has not been widely applied to conventional physical adsorption type separating agents from the viewpoint of solubility of polysaccharide derivatives, but in the present invention, the immobilized type separating agent is not used. Applicable to use.
A small amount of diethylamine or trifluoroacetic acid can be added to the mobile phase solvent in the present invention, if necessary. The application of the mobile phase solvent in the present invention surprisingly improved the separation performance and improved the preparative productivity as compared with the conventional hexane / alcohol solvent.

The method for chromatographic separation of optical isomers of the present invention comprises:
The method can be applied to a batch process using a single column or a simulated moving bed process in which a plurality of columns are continuous.

[0020]

According to the method for chromatographic separation of optical isomers of the present invention, high separation performance, especially high preparative productivity in the case of preparative separation, can be obtained, and the optically active substance can be economically produced.

[0021]

EXAMPLES The present invention will be described in more detail with reference to the following Examples, which by no means limit the scope of the present invention. be able to. In the examples, the separation coefficient α was obtained by the following equation.

[0022]

(Equation 1)

Production Example 1 Production of a Chiral Stationary Phase in which Polysaccharide Derivatives are Coated on Silica Gel and Crosslinked by Polysaccharide Derivatives Silica gel surface inactivation treatment Porous silica gel (SP-1000, manufactured by Daiso, particle size 7 μm, The average pore diameter of 1000 °) was reacted with 3-aminopropyltriethoxysilane to give an aminopropylsilane treatment (APS treatment). 200 g of this APS-treated silica gel was mixed with 15 ml of 3,5-dimethylphenyl isocyanate in 1.0 liter of methylene chloride at room temperature.
The reaction was performed for 1.5 hours. This was collected by filtration with a glass filter, washed sequentially with methylene chloride / methanol = 2/1, and methylene chloride, ethanol, acetone, and hexane, and then dried under vacuum.

Cellulose-6-hydroxy-2,3-
Synthesis of bis (3,5-dimethylphenylcarbamate) Under a nitrogen atmosphere, 4.0 g of tritylcellulose in which 0.9 to 1.0 trityl groups were reacted with a glucose unit synthesized by a known method was dissolved in dry pyridine, and Add 10 ml of dimethylphenylisocyanate and add 25 ml at 100 ° C.
Stirred for hours. This was poured into 700 ml of methanol, and the precipitated solid was collected by filtration, washed with ethanol and hexane, dried, and then concentrated in hydrochloric acid-containing methanol (cellulose derivative).
The mixture was stirred in 0.25 ml of concentrated hydrochloric acid and 40 ml of methanol per 1.0 g to remove the trityl group. The detritylated cellulose derivative was collected by filtration, washed with ethanol and hexane, and dried to obtain 14.9 g of cellulose-6-hydroxy-2,3-bis (3,5-dimethylphenylcarbamate).

Cellulose-6-hydroxy-2,3-
Cellulose-6-hydroxy-2,3-bis (3,3) obtained by preparation of silica gel supporting bis (3,5-dimethylphenylcarbamate)
5-dimethylphenylcarbamate) 1.5 g to 8 ml of T
It was dissolved in HF and sprinkled evenly onto 5.7 g of silica gel and applied. After the solvent was distilled off, the residue was washed successively with methanol, ethanol and hexane, and dried to obtain 7.1 g of silica gel on which a cellulose derivative was supported.

Silica gel supporting cellulose derivative obtained by immobilization on silica gel by crosslinking reaction between only cellulose derivatives 6.7
35 g of dry toluene was added to the resulting mixture, and a solution of 110 mg of diphenylmethane diisocyanate and 5 ml of dry pyridine was added thereto. The mixture was heated and stirred at 110 ° C. for 5 hours. After the completion of the reaction, the resultant was collected by filtration, washed with THF, methanol, ethanol, and hexane in that order, vacuum dried, and a separating agent immobilized on silica gel by a cross-linking reaction between cellulose derivatives only.
6.8 g were obtained.

To the separating agent obtained by the modification of the unreacted hydroxyl group of the cellulose derivative immobilized on silica gel, dry toluene 25 ml, dry pyridine 25
Add 0.5 ml of 3,5-dimethylphenylisocyanate, and heat and stir at 110 ° C for 18 hours.The carbamoyl of the unreacted hydroxyl group of the cellulose derivative immobilized on the silica gel by the crosslinking reaction between the cellulose derivatives alone is added. Was performed. After completion of the reaction, the reaction solution was collected by filtration, washed sequentially with THF, methanol, ethanol, and hexane, and then dried under vacuum. As a result, the final separating agent was 6.9 g. The loading amount of the cellulose derivative on the silica gel is 20
% (Calculated as 2.7 of the three hydroxyl groups in the glucose unit in cellulose being carbamoylated).

Using a separation agent immobilized on silica gel by a cross-linking reaction between only the cellulose derivatives produced in the above, and 3.5 g of methanol as a dispersion solvent and a pressurized solvent, The slurry was packed in a stainless steel column having a length of 25 cm and an inner diameter of 0.46 cm by a slurry filling method. The pressure at this time was 250 kgf / cm ² .

Examples 1 to 3 and Comparative Example 1 Using a mixed solvent shown in Table 1 as a mobile phase solvent, a racemic laudanosine represented by the following formula (II) was optically resolved under the following conditions. Table 1 shows the results.

[0030]

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<Conditions for Optical Resolution> Chiral stationary phase: separating agent produced in Production Example 1 Mobile phase flow rate: 1.0 ml / min Temperature: 40 ° C. Detection: 254 nm Sample concentration: 2 mg / 1.0 ml Amount injected: 20 μl

[0032]

[Table 1]

Examples 4 to 6, Comparative Example 2 Using a mixed solvent shown in Table 2 as a mobile phase solvent, a pindolol racemic compound represented by the following formula (III) was optically resolved under the following conditions. Table 2 shows the results.

[0034]

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<Conditions for Optical Resolution> Chiral stationary phase: separating agent produced in Production Example 1 Mobile phase flow rate: 1.0 ml / min Temperature: 40 ° C. Detection: 254 nm Sample concentration: 2 mg / 1.0 ml Amount injected: 20 μl

[0036]

[Table 2]

Examples 7 to 9 and Comparative Example 3 Using a mixed solvent shown in Table 3 as a mobile phase solvent, a racemic propranolol represented by the following formula (IV) was optically resolved under the following conditions. Table 3 shows the results.

[0038]

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<Conditions for Optical Resolution> Chiral stationary phase: separating agent produced in Production Example 1 Mobile phase flow rate: 1.0 ml / min Temperature: 40 ° C. Detection: 254 nm Sample concentration: 2 mg / 1.0 ml Amount injected: 20 μl

[0040]

[Table 3]

Examples 10 to 12 and Comparative Example 4 Using a mixed solvent shown in Table 4 as a mobile phase solvent, a racemic warfarin represented by the following formula (V) was optically resolved under the following conditions. Table 4 shows the results.

[0042]

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<Conditions for Optical Resolution> Chiral stationary phase: separating agent produced in Production Example 1 Mobile phase flow rate: 1.0 ml / min Temperature: 40 ° C. Detection: 254 nm Sample concentration: 2 mg / 1.0 ml Amount injected: 20 μl

[0044]

[Table 4]

Example 13, Comparative Examples 5 and 6 The separating agent prepared in Production Example 1 as a chiral stationary phase (Example
13, Comparative Example 5) or a separating agent (trade name: CHIRALCEL OD, manufactured by Daicel Chemical Industries, Ltd., Comparative Example 6) in which cellulose tris (3,5-dimethylphenylcarbamate) was coated on silica gel; Using the mixed solvent shown in (1), the racemate of warfarin represented by the above formula (V) was subjected to optical resolution under the following conditions, and the preparative productivity was evaluated by the following method. Table 5 shows the results. Also, the embodiment
13 shows the chromatogram obtained in Comparative Example 5, FIG. 2 shows the chromatogram obtained in Comparative Example 5, and FIG. 3 shows the chromatogram obtained in Comparative Example 6.

<Optical Resolution Conditions> Mobile phase flow rate: 1.0 ml / min Temperature: 40 ° C. Detection: 254 nm <Preparation productivity evaluation method> The amount of the racemic substance with the maximum dissolution amount in the mobile phase was increased, and almost the baseline was obtained. Amount to be implanted when separation stops (maximum amount of racemic implant)
The maximum amount of the enantiomer per one injection (mg / time), the second from the rise of the first peak
The peak interval time until the peak returned to the baseline was defined as Δt (min / time), and the fractionation productivity (the amount of each enantiomer collected per hour; mg / hr) was determined by the following formula.

Preparative productivity (mg / hr) = maximum dose of enantiomer (mg / time) × number of shots per hour (time / hr) Here, the number of shots per hour is defined by the following formula Is done. Number of shots per hour (time / hr) = 60 (min / h)
r) / Δt (min / times)

[0048]

[Table 5]

[Brief description of the drawings]

FIG. 1 is a chromatogram obtained in Example 13.

FIG. 2 is a chromatogram obtained in Comparative Example 5.

FIG. 3 is a chromatogram obtained in Comparative Example 6.

Claims (5)

[Claims] 1. A polysaccharide derivative-based chiral stationary phase is used as a chiral stationary phase, and a solvent selected from the group consisting of ether solvents, ketone solvents, ester solvents, amide solvents and halogen solvents is used as a mobile phase solvent. Chromatographic separation of optical isomers, characterized by using a mixed solvent comprising a combination with a hydrocarbon solvent. 2. A mixed solvent comprising tetrahydrofuran, acetone, ethyl acetate, N, N-dimethylformamide,
2. The method for chromatographic separation of optical isomers according to claim 1, wherein the method comprises a combination of a solvent selected from the group consisting of N, N-dimethylacetamide, chloroform and methylene chloride and hexane. 3. The method for chromatographic separation of optical isomers according to claim 1, wherein the polysaccharide derivative-based chiral stationary phase is obtained by chemically bonding a polysaccharide derivative to a carrier. 4. The method for chromatographic separation of optical isomers according to claim 1, wherein the polysaccharide derivative-based chiral stationary phase is obtained by coating the polysaccharide derivative on a carrier and then crosslinking the polysaccharide derivatives. 5. The method for chromatographic separation of optical isomers according to claim 1, wherein the polysaccharide derivative-based chiral stationary phase is obtained by coating a polysaccharide derivative on a carrier and then coating the surface of the polysaccharide derivative with a synthetic polymer.