JPH03190827A - Production of optical isomer separating agent and beta-aminoalcohol utilizing same agent - Google Patents

Abstract

(57) [Summary] This bulletin contains application data before electronic filing, so abstract data is not recorded.

Description

DETAILED DESCRIPTION OF THE INVENTION [Field of Industrial Application] The present invention relates to a novel separating agent for optical isomers and a method for separating various optically active β-amino alcohols using the separating agent.

The optical isomer separating agent of the present invention is useful for isolating various optical isomer mixtures.

[Conventional technology]

In recent years, the fields of application of optical isomers are expanding to include not only medicines and agricultural chemicals, but also food additives, ferroelectric liquid crystal materials, and even nonlinear optical materials. for example,
When pharmaceuticals contain optical isomers, each isomer may have different pharmacological activities, and as shown in the zalidomide crisis, there is a risk that the isomers may have fatal side effects. In response to this situation, the Ministry of Health and Welfare has issued a guideline stating that for pharmaceuticals containing optical isomers, it is desirable to clarify the pharmacological activity of each isomer. Up to now, a method for measuring the optical purity of optical isomers and a simple method for separating optical isomers have not been established, and the establishment of such methods is strongly desired by pharmaceutical manufacturers, particularly for the above-mentioned reasons.

Separation of optical isomers by high-performance liquid chromatography has attracted particular attention and is being actively researched because of its high efficiency and simplicity. As a result, several fillers have been applied in recent years. An example of applying peptides to the separation of optical isomers is Chroma by C. Pettersson et al.
tographia 2L 32L (1986) spear,
B.

J, Chromatograph by Rogers et al.
As stated in y 輯, 49 (1987),
This is a separation method that uses silica gel or octadecylated silica gel as the stationary phase and adds peptides to the mobile phase (3) (4) to form ion pairs or ion complexes with the compound to be separated.

Most beta blockers have a structure represented by the following formula:
3C) R3 (wherein * represents an asymmetric carbon, R3 represents a substituted or unsubstituted aliphatic hydrocarbon group or a substituted or unsubstituted aromatic hydrocarbon group, and R6 represents hydrogen or a methyl group) Catecholamines such as epinephrine and adrenaline are used as extremely effective therapeutic agents for the treatment of angina pectoris, arrhythmia, hypertension, etc., which occur via beta receptors in smooth muscles and the cardiac circulatory system. Beta-blockers have asymmetric carbon atoms, so their optical isomers may have markedly different efficacy. For example, propranolol (R5-naphthyl, R7-
It has been reported that (-) hydrogen has nearly 100 times more medicinal efficacy than (+) hydrogen.

[Problem to be solved by the invention]

However, currently commercially available columns packed with separation agents have 1) limited target compounds that can be separated, 2) low separation ability, 3) poor analytical reproducibility, and 4) limited analytical conditions. There are many problems that need to be improved, such as (for example, usable solvents), (5) low durability, and (6) high cost.

Furthermore, as an example of optical resolution using peptides, the method of adding them to the mobile phase as described above is known, and a separation agent with immobilized peptides has not been reported. Furthermore, a simple optical purity measurement method or separation method for practical use of pharmaceuticals containing asymmetric carbon, such as bake blockers, has not been established to date.

Therefore, an object of the present invention is to develop a separating agent that has good reproducibility and excellent optical separation ability for a wide variety of optical isomers.

[Means to solve the problem]

As a result of extensive research to solve the above problems, the present inventors have discovered that separation agents in which various Vepti 1- are immobilized on carriers via spacers (5) (6) - have excellent separation performance. The present invention has been completed by discovering the following.

That is, according to the present invention, there is provided an optical isomer separating agent comprising an optically active peptide having a carboxyl group at one end and a primary or secondary amino group at the other end immobilized on the surface of a carrier via a spacer. In particular, silica gel can be used as a carrier and a silane coupling agent can be used as a spacer.

According to the present invention, the formula (wherein at least one of A, , A, and A3 represents silica gel or a siloxane bonding portion with silica gel, and the remaining members each independently represent hydrogen, carbon number 1 to 2
0 alkyl group, aryl group having 6 to 20 carbon atoms, halogen, hydroxyl group, or alkoxy group having 1 to 20 carbon atoms, B represents a spacer having 1 to 30 carbon atoms, *
represents an asymmetric carbon, 0 represents an integer of 1 to 10, and R
1 represents either a substituted or unsubstituted aliphatic hydrocarbon group or a substituted or unsubstituted aromatic hydrocarbon group,
R2 represents hydrogen, a substituted or unsubstituted aliphatic hydrocarbon group, or a substituted or unsubstituted aromatic hydrocarbon group, R1 and R2 can be combined to form a cyclic structure, and R3 is a divalent The organic residue R4 represents hydrogen, a substituted or unsubstituted aliphatic hydrocarbon group, or a substituted or unsubstituted aromatic hydrocarbon group, and R3 and R4 may be combined to form a cyclic structure. The above-mentioned optical isomer separating agent is provided.

Furthermore, formula (4) (where * represents an asymmetric carbon, R9 represents a substituted or unsubstituted aliphatic hydrocarbon group or a substituted or unsubstituted aromatic hydrocarbon group, and R6 represents a hydrogen or methyl group) Provided is a method for producing an optically active β-amino alcohol, which comprises separating a mixture of optical isomers of β-aminoal (7) (representing (8)) using any of the fillers described above. be done.

The peptide forming the characteristic part of the present invention may be any optically active peptide having a carboxyl group at one end and a primary or secondary amino group at the other end, but it is represented by the following formula: It is preferable to use

(In the formula, * represents an asymmetric carbon, n represents an integer from 1 to 10, R1 represents a substituted or unsubstituted aliphatic hydrocarbon group or a substituted or unsubstituted aromatic hydrocarbon group, R2
represents hydrogen, a substituted or unsubstituted aliphatic hydrocarbon group, or a substituted or unsubstituted aromatic hydrocarbon group, R1 and R2 may be combined to form a cyclic structure, and R8 represents a divalent organic Represents a residue, R4 represents hydrogen, a substituted or unsubstituted aliphatic hydrocarbon group, or a substituted or unsubstituted aromatic hydrocarbon group, and R3 and R4 can be combined to form a cyclic structure. . ) Specifically, the substituted or unsubstituted aliphatic hydrocarbon group for R3 includes methyl, ethyl, isopropyl, isobutyl, 2-butyl, t
-butyl, neopentyl, 2-methylthiomethyl, 1-
Hydroxymethyl, ■-hydroxyethyl, 2-carbamoylethyl, 4-hydroxybenzyl, 2-indolylmethyl, 4-imidazoylmethyl, etc., and the substituted or unsubstituted aromatic hydrocarbon group of R1 is phenyl,
Examples include aromatic hydrocarbon groups such as benzyl, fluorophenyl, trifluoromethylphenyl, 4-hydroxy-2-fluorophenyl, and 4-hydroxy-3-fluorophenyl. In addition, examples of compounds in which R and R2 are combined to form a cyclic structure include pyrrolidine, piperidine,
3-Hydroxypyrrolidine, indoline, 12.3.4
Examples include -tetrahydroisoquinoline and thiazolidine. Examples of substituted or unsubstituted aliphatic hydrocarbon groups for R2 include methyl, ethyl, (9) (10) isopropyl, isobutyl, 2-butyl, t-butyl,
Examples include neopentyl, octyl, vinyl, aryl, and butenyl. The substituted or unsubstituted aromatic hydrocarbon group for R2 is phenyl, tolyl, xylyl, benzyl, phenethyl, cinnamyl, naphthyl, fluoroI
Examples include cophenyl and trifluoromethyl.

The divalent organic residue of R3 is preferably a divalent organic residue of a carboxylic acid having a primary or secondary amino group represented by the following formula.

4 H-N-R3-C-Oll 1 A specific example of the aminocarboxylic acid providing a divalent aliphatic organic residue is a 2-aminocarboxylic acid having a substituted or unsubstituted aliphatic hydrocarbon group. glycine, alanine,
2-amino-n-butyric acid, 2-aminoisoacetic acid, valine, norvaline, leucine, norleucine, isoleucine, L
-Leucine, 2-aminocaprylic acid, S-methylcysteine, serine, threonine, glutamine, 1-Lip I・
Examples of 2-aminocarboxylic acids having a substituted or unsubstituted aromatic hydrocarbon group include phenylglycine, phenylalanine, homophenylalanine, tyrosine, fluorophenylalanine, fluorotyrosine, trifluorophenylalanine, pentafluoro Examples include benylalanine and fluorotryp-1-phane, and examples of 2-aminocarboxylic acids having a cyclic structure include proline, hydroxyproline, thioproline, pipecolic acid, indoline-2-carboxylic acid, and ■.

Examples include 2.3.4-tetrahydroisoquinolinecarboxylic acid. In addition, as 3-aminocarboxylic acids having substituted or unsubstituted aliphatic hydrocarbon groups, β-alanine, 3
-amino-n-butyric acid, 3-aminoisoacetic acid, 3-amino-3-phenylpropionic acid, etc.; -n-valeric acid, 6-amino-n
-caproic acid, 7-aminohebutanoic acid, 8-amino-n
(11) (12) Caprylic acid and the like can be exemplified.

Examples of the aminocarboxylic acid that provides a divalent aromatic organic residue of R:l include aminobenzoic acid, aminotoluic acid, and aminonaphthoic acid.

The optically active peptide forming the characteristic part of the present invention can be obtained by conventionally known peptide synthesis methods. For example, Izumiya et al.: “Basics and Experiments of Peptide Synthesis” (Marukubi, 19
85) and M. Bodansky and A. Boda.
nsky:”The Practice of Pep
tide 5 synthesis” (Springer V
erlag, 1984).

As the carrier used in the present invention, any carrier may be used as long as it has excellent pressure resistance and does not swell or contract when substituted with a solvent. Specific examples include porous organic carriers made of polystyrene, polyacryloamide, polyacrylate, etc., and porous inorganic carriers made of silica, alumina, magnesia, titanium oxide, glass, silicic acid, etc.; It is preferable to use silica gel as the carrier, which has silanol groups that can be used for modification and has excellent pressure resistance. The silica gel used in the present invention is crushed or spherical, with a particle size of 0.1 to 1000 μm,
Preferably, it has a diameter of 1 to 100 μm, and is preferably porous, with an average pore diameter of 10 to 10,000, preferably 50 to 5,000.

The spacer used in the present invention refers to a group for arranging the above-mentioned peptide capable of optical resolution and the carrier at a certain distance. In other words, as a spacer, any group can be used as long as it has a functional group that can react with the carrier to generate a covalent bond at one end and a functional group that can generate a covalent bond by reacting with the amino group of the peptide at the other end. good.

When silica gel is used as the carrier of the present invention, space 4J is used to immobilize the optically active peptide on the silica gel.
As °-, a silane camping agent is used. Any known silane coupling agent can be used, and these are represented by the following formula.

A2-3i -B 3 (13) (14) (In the formula, at least one of A, , A, and A3 represents silica gel and a siloxane bonding part with silica gel, and the rest each independently represent hydrogen and the number of carbon atoms. represents an alkyl group having 1 to 20 carbon atoms, an aryl group having 6 to 20 carbon atoms, a halogen, a hydroxyl group, or an alkoxy group having 1 to 20 carbon atoms, and B is a group forming a spacer having 1 to 30 carbon atoms; It is an alkyl or aryl group having an internal halogen, amino group or oxolane group, and may contain a heteroatom. In other words, it can contain any bonding mode such as an ether bond, an amino bond, or an amide bond.) Specific details Silane coupling agents include those that have a functional group that can react with an amino group to form a covalent bond, such as chloromethyldimethylchlorosilane, bromomethyldimethylchlorosilane, chloromethyltriethoxysilane, chloromethyltrichlorosilane, 1-chlorosilane, etc. Ethylmethyldichlorosilane, 2-chloroethylmethyldichlorosilane,
Halogenated compounds such as 1-chloroethyl 1-lichlorosilane, 3-chloropropyltrimethoxysilane, 3-chloropropyldimethoxymethylsilane, 3-bromopropyldimethylchlorosilane, 3-bromopropyltrichlorosilane, p-chloromethylphenyltrimethoxysilane A silane coupling agent having an alkyl group, a silane coupling agent having a halogenated phenyl group such as p-chlorophenyl-1-rimethoxysilane, p-chlorophenyl-IJ-gishsilane, 2-(3,4-epoxy cyclohexylethyl) triethoxysilane glycidoxypropyltrimethoxysilane,
3-glycidoxypropylmethylsilane, 3-glycid F
Silane cup 'J' bonding agent with an epoxy terminal such as xyprobil dimethyl ester I xysilane, 2-
(4-chlorosulfonylphenyl)ethyltrichlorosilane, 2-(4-chlorosulfonylphenyl)ethyltrimethoxysilane, 3(+-lichlorosilyl)propylchloroformate, silane coupling agent with acid chloride at the end, 3-isocyanate Examples include silane camping agents having an isocyanate group at the end, such as propyldimethylchlorosilane, 3-isocyanatepropyl(15)(16)l-ethoxysilane, and the like.

When using the silica gel of the separation agent of the present invention as a carrier, the preparation method is a method in which silica gel treated with a silane coupling agent is reacted with an optically active peptide by a conventional method, or a method in which silica gel treated with a silane coupling agent is reacted with an optically active peptide, or a method in which silica gel treated with a silane coupling agent is reacted with an optically active peptide. Any method may be used in which the silica gel surface is prepared by reacting an optically active peptide in advance and then modifying the silica gel surface using the optically active silane coupling agent. For example, K
.

K. UnHer: Porous Silica'' (
Hlsevier, 1979) p84.

Compounds to be separated by the separation agent of the present invention include aromatic hydrocarbons, halogen compounds, alcohols, aldehydes, amines, ethers, esters, amides, amino acids, oxycarboxylic acids, and the like. In particular, it shows high selectivity in the separation of racemic mixtures of various β-amino alcohols. Specifically, it exhibits particularly excellent separation ability for Heta blockers, which are important cardiovascular treatment drugs, and their structures are shown below]7) (18) 0H CH.

H CH.

H CI(.

H CH.

(19) H CH3 (20) When performing optical resolution of a target compound using the separating agent of the present invention, the separating agent is first uniformly packed into a column by an appropriate method. There are dry and wet filling methods.
The slurry method is preferred because a column with a high number of theoretical plates can be obtained. Next, a racemic mixture of the compounds to be separated is injected into this column and developed with an appropriate eluent to perform optical resolution. There are no particular restrictions on the developing solvent used in the present invention, and a wide range of compounds can be separated by selecting commonly used normal phase or reversed phase conditions. Particularly for separating various racemic mixtures having amino groups, a developing solvent prepared by adding an appropriate amount of amine to a mixed solvent of a halogenated solvent and an alcohol is suitable. Examples of halogenated solvents include methylene chloride, chloroform, and carbon tetrachloride. Any alcohol can be used as the alcohol to be mixed, but methanol, ethanol, n-propanol, isopropanol, n-butanol, t-butanol, etc. are preferred. Further, as the amine to be added, amines which produce a chemical reaction can be used, but preferably aliphatic amines such as n-butylamine, diethylamine, diisopropylamine, and triethylamine, aliphatic cyclic amines such as pyrrolidine and piperidine, and furthermore ethanolamine,
Examples include substituted aliphatic amines such as N-methylethanolamine. By changing the concentration of the alcohol to be mixed and the concentration of the amine added, the retention time of the compound to be separated can be changed.

〔Example〕

The present invention will be explained in detail below using synthesis examples and examples.

Definitions of terms used in the examples and tables are shown below.

Retention ratio (K') - (21) (22) Separation coefficient (α) = Retention ratio (K') of shorter retained enantiomer Synthesis-Example-1 Silica gel (Develosil-3ilO manufactured by Nomura Chemical
20 g of O, 5) was dried under reduced pressure at 130°C in an oil bath for 4 hours, returned to room temperature, and then filled with nitrogen to return to normal pressure. 100ml of dry toluene and 12ml of 3-glycidoxypropyltrimethoxysilane were sequentially added, and the mixture was heated for about 20ml while removing methanol produced by the reaction from the system.
Refluxed for an hour. After returning to room temperature, glass filter =
(G-4), and dried 1-toluene at 50 mE.
After washing briefly twice and drying under reduced pressure at 80°C, 23.57 g of modified silica gel was obtained. This elemental analysis value is C
: 7.28, H: 1.47, N: <0.1,
Therefore, the 3-glycidoxyprobyl group attached to the surface was shown to be 0.87 mmo I/g.

Fruit - Toku ↓ - 512 mg of glycyl-L-proline in 15 d of methanol
An ice-cold 4N aqueous solution of thorium hydroxide was slowly added dropwise. After stirring for 2 hours, the solvent was distilled off under reduced pressure to quantitatively obtain the sodium salt of glycyl-5-proline. 3.73 g of the 3-glycidoxyprobyl group-containing silica gel synthesized in Synthesis Example 1 was dispersed in 20 ml of dry methanol, and a dry methanol solution of the thorium salt of glycyl-proline was added. The resulting reaction mixture was left at room temperature under a nitrogen atmosphere for 3 days, and then
The mixture was suction filtered using a glass filter (G4), washed twice with 20 ml of methanol, and dried under reduced pressure to obtain 3.75 g of modified silica gel.

The elemental analysis values are C: 7.38, H: 1.43, N
:0.57, and therefore, a silica gel with monoproline chemically immobilized on the surface by chemical glycyl was obtained.

river? - The obtained glycyl-L-proline-immobilized silica gel was transferred to a stainless steel column (4.6 m/m x 150 m/rr
+) by the slurry method. Optical resolution of various beta blockers was carried out using CR-64 manufactured by Kansei Co., Ltd. (23) (24) as high performance liquid chromatography and methanol-methylene diethylamine chloride as the mobile phase. The analysis results are shown in Table 1.

Synthesis Example 1 according to the method shown in Example 1 using glycyl-glycylheaproline as the optically active peptide
The surface of the silica gel prepared in was chemically modified. The elemental analysis values are C: 7.03, H: 1.37, N:
It was 0.62.

A stainless steel column (4.6 m/m
m/m) by the slurry method, and optical resolution of the beta blocker was attempted. The analysis results are shown in Table 2.

Glycyl-glycylglycyl as an optically active peptide
The surface of the silica gel prepared in Synthesis Example 1 was chemically modified using L-proline according to the method shown in Example 1. This elemental analysis value is C near, 48, H: 1.41
, N: 0.95.

The glycyl-glycylglycyl-L-proline-immobilized silica gel obtained by the above method was coated in a stainless steel column (4.6 m/mX) in the same manner as shown in Example 2.
150 m/m) by the slurry method, and optical resolution of the beta blocker was attempted. The analysis results are shown in Table 2.

Tip 1 The surface of the silica gel prepared in Synthesis Example 1 was chemically modified according to the method shown in Example 1 using glycyl-trans-4-hydroxy-■,-proline as an optically active peptide. This elemental analysis value is near C. 00, H: 1
．． 39, N: 0.57.

Symptomitis ■ The silica gel on which glycyl-trans-4-hydroxy-proline was immobilized obtained by the above method was treated in the same manner as shown in Example 2 (25) (26) and a stainless column (4.6m). /mX 150m/m) by the slurry method, and optical resolution of the beta blocker was attempted. The analysis results are shown in Table 3.

Glycyl-indoline-2 as optically active peptide
- The surface of the silica gel prepared in Synthesis Example 1 was chemically modified using carboxylic acid according to the method shown in Example 1. This elemental analysis value is C near, 14, H: 1.30,
N: 0.16.

Practical Rules The silica gel on which glycyl-I, -indoline-2-carboxylic acid obtained by the above method was fixed was coated in a stainless steel column (4.6 m/m) in the same manner as shown in Example 2.
x 150m/m) by the slurry method, and optical resolution of the beta blocker was attempted.

The analysis results are shown in Table 4.

For comparison, a separating agent was prepared in which 7-broline was chemically immobilized on the surface of silica gel having a glycidoxyprobyl group subjected to HNV in Synthesis Example 1 as shown in the following formula. The silica gel was used as shown in Example 2, and attempts were made to separate various beta blockers, but it showed no separation ability for any of the bake blockers. These results reveal that the separation agent of the present invention exhibits excellent separation ability for racemic mixtures of various β-amino alcohols, especially beta-blockers, by immobilizing optically active peptides. Ta.

(27) (28) Table 1 Optical resolution of glycyl-L β blocker o-cI (2-clI-co□-Ni+(29) Table 3 Optical resolution of glycyl-L hydroxyl by proline filler Proline series 0-C11□-C)I-C1lz-Nl+ (31) Table 2. Optical resolution of β-blocker using glycyl-■,-proline packing 11 Example 4 9.27 10.47 1.13 Mobile phase: 3% MeO1i-CHzCff 2-Et, N
(10-3M) Flow rate: 1. Omρ/min Below margin (30) Table 4. Optical resolution of β-blocker using glycyl-L-indoline-2-carboxylic acid-based packing material 11 Column K.

2 Separation factor (α) Mobile phase: 3% Merit-' C112CI! , z
-EtJ (10-3M) flow rate: 1.0 mβ/m i
[Effect of the Invention] As is clear from the above description, the separating agent of the present invention was shown to have the ability to separate a wide range of mixtures of optically active substances. In particular, it is clear that when the separating agent of the present invention is used as a packing material for chromatography, various optically active substances can be separated, and in particular, various beta blockers can be easily separated with good reproducibility.

(32)

[Brief explanation of drawings]

1 and 2 show respective liquid chromatograms when a racemic mixture of propranolol and pindolol was separated using the column prepared in Example 2.

Claims (1)

[Claims] 1. Optical isomerism characterized by immobilizing an optically active peptide having a carboxyl group at one end and a primary or secondary amino group at the other end on the surface of a carrier via a spacer. Body separation agent. 2. The optical isomer separating agent according to claim 1, wherein silica gel is used as the carrier and a silane coupling agent is used as the spacer. 3. Formulas ▲ Numerical formulas, chemical formulas, tables, etc. Alkyl group having 1 to 20 carbon atoms, aryl group having 6 to 20 carbon atoms,
represents a halogen, a hydroxyl group, or an alkoxy group having 1 to 20 carbon atoms, B represents a spacer having 1 to 30 carbon atoms, * represents an asymmetric carbon, n represents an integer of 1 to 10, and R_1 is a substituted or represents either an unsubstituted aliphatic hydrocarbon group or a substituted or unsubstituted aromatic hydrocarbon group, R_2 is hydrogen, a substituted or unsubstituted aliphatic hydrocarbon group, or a substituted or unsubstituted aromatic hydrocarbon group R_1 and R_2 can be combined to form a cyclic structure, R_3 is a divalent organic residue, R_4 is hydrogen, a substituted or unsubstituted aliphatic hydrocarbon group, or a substituted aliphatic hydrocarbon group. or an unsubstituted aromatic hydrocarbon group, and R_3 and R_4 can be combined to form a cyclic structure). agent. 4. Formulas ▲ There are mathematical formulas, chemical formulas, tables, etc. ▼ (In the formula, * represents an asymmetric carbon, and R_5 represents a substituted or unsubstituted aliphatic hydrocarbon group or a substituted or unsubstituted aromatic hydrocarbon group. , R_6 represents hydrogen or a methyl group) is separated using the separating agent according to any one of claims 1 to 3. Method for producing β-amino alcohol.