JPS59190955A - Resolution of amino acid racemic body - 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 Biological systems have long been characterized as highly stereoselective and perhaps the most common representation of chiral environments. chirality
As a general rule, biological systems utilize only one enantiomer of these metabolites with chiral centers. Essential amino acids are an example, and only the L form is used by humans. Another example is amino at 3 a 4-dihydroxyphenylalanine, in which the L enantiomer has the only pharmacological action in the treatment of /l'-Kinson's disease.

The problem is to directly produce optically active amino acids synthetically using a chiral environment or to effectively obtain all optically active amino acids from their racemic forms. Although substantial progress has been made synthetically using chiral environments, e.g. chiral reagents, a more general method of making optically active materials from inert precursors is to combine one or both of the racemic components. A method for separating the compound from a racemic mixture containing the compound, such as a conventional optical resolution method.

One method of optical resolution is to use diastereomers with different physical properties and purify at least one diastereomer by conventional separation methods. The desired enantiomer is then obtained from this purified diastereomer by appropriate chemical conversion. Fractional crystallization is generally employed as a method for separating diastereomers. Amino acid fraction v1 by Harada and Juyo, Bull, Chem, Soc, J
apan+37.191 (1964). Harada et al. fractionated the diastereomeric hydrochloride salts of amino acid Mendelian esters by placing one of the diastereomers as a crystal seed in a supersaturated solution. Other methods include Halpern and Wθ5tley, C
An example is Hem, Comm, +421 (1965). Halper et al. found that the p-)luenesulfonate diastereomer of the amino acid menthyl ester could be easily fractionally crystallized.

Chromatographic separation of diastereomers using a chiral stationary phase is an example of "Advances in Chr.
This is a different method from separation by crystallization as described in Omatography' Vol. 16, p. 177-s, p. 3 (1978). Commercially successful chromatographic separation of diastereomers as a method for the resolution of amino acids has stringent requirements for diastereomers. One is that it must be possible to chemically and optically regenerate optically active amino acids in high yields. Another is that the process itself is relatively sensitive to changes in material supply and quality, and is easy to use with a wide range of stationary phases and different eluents so that process control is relatively undemanding. It is possible to separate diastereomers.

The basic discovery underlying the present invention is that optically active cyclohexanol and diastereomeric esters of racemic amino acids of yuzu can be easily separated under a wide range of chromatographic conditions. The above discovery is particularly successful because the optically active enantiomers of amino acids can be easily regenerated chemically and stereochemically in high yield and purity from diastereomers separated by Yuzu's method. This is the basis of the method for splitting amino acids. Furthermore, the optically active cyclohexanol used as a resolving agent can also be recovered in high yield and reused in other cycles consisting of diastereomer production, separation, and regeneration of amino acid enantiomers.

An object of the present invention is to obtain optically active amino acids from amino acid racemates. In one embodiment, the ester of an optically active amino acid racemate and cyclohexanol is chromatographically separated, the purified diastereomer is hydrolyzed, and then all of the regenerated optically active amino acids are recovered. That is what it is. In a more specific embodiment, cyclohexanol is 2-
Isopropyl-5-methylcyclohexanol.

In an even more particular embodiment, the cyclohexanol is menthol.

The invention described herein involves contacting a solution of a diastereomeric ester obtained from an amino acid racemate and an optically active cyclohexanol with a total chromatographic support, and eluting the support with a solvent under chromatographic conditions to purify the solution. The method is characterized in that at least one of the effluent fractions containing purified diastereomers is collected, the purified diastereomers are completely processed to liberate optically active amino acids, and the amino acids are recovered. This invention is made possible by the discovery that 2-isopropyl-5-methylcyclohexanol and diastereomeric esters of amino acids are readily separated on various chromatographic supports with various solvents as eluents.

Diastereomers are compounds with at least two chiral centers, at least one of which is different and at least one of which is the same. When a diastereomer contains two chiral centers, there are two sets of diastereomers, each set consisting of two enantiomers. Each set of diastereomers differs in composition in that one chiral center is different and the other chiral center is the same. For example, if A is an amino acid, M is menthol, and the symbols d and e are the rotational characteristics of total polarization, the diastereomeric silk that can be an ester of menthol and an amino acid is as follows.

(1) l-M-d-A (3)/'-M-1? −
AC2) d-M-J-A (4) d-M-d-A
Here, (1) and (2) are enantiomers like (3) and (4), but both (1) and (2) are (3), (
4) are both diastereomers. Particularly (2) and (3L) are diastereomeric esters of f-menthol and racemic amino acids.

The diastereomeric esters used in this invention can be easily separated by chromatography on a variety of chromatographic supports using a wide variety of solvents as eluents. Column selectivity is determined by the quantity . Here, t2 and 1. 11. is between the retained portions of the two diastereomers, and t is the retained portion of the unretained component. It is jI. This column selectivity α is a measure of the ease of separating components using a specific support and a specific solvent as an eluent. The larger the value of α, the easier the separation becomes. A value of α greater than about 1.3 means that separation is extremely easy. Also, the value of α is about 1.1
Is it not necessarily easy to separate the range of ~1.3?
It means possible. It has been found that the alpha value of the specific diastereomer of the amino acid racemate used in the present invention is often about 1.3 or more.

The diastereomers used in the present invention are racemates of amino acids, especially esters obtained from α-amino acids and optically active 2-isopropyl-5-methylcyclohexanol. The latter alcohol exists as four geometric isomers, each consisting of one pair of enantiomers, for a total of eight stereoisomers. The geometrical relationship between the isopropyl group and the methyl group facing the hydroxyl group is trans
, cia (menthol) * cls, tran
s (neomenthol), trans, tran
s (inmenthol), or cis, cis (neoisomenthol). Any enantiomer of the above geometric isomers can be used in the present invention. Menthol is a particularly preferred isomer because of its ready availability, and l-menthol is particularly preferred for the same reason.

Amino acids that can be split by the method of the present invention include naturally occurring amino acids, such as alanine, arginine, asparagine, aspartic acid, cysteine,
Cystine, 3. s-dizolomotyrosine, 3.5-short tyrosine, glutamic acid, glutamine, histidine,
Hydroxylysine, hydroxyproline, isoleucine, leucine, lysine, methionine, phenylalanine,
These include phlorine, serine, threonine, thyroxine, tryptophan, tyrosine and serine. Other amino acids that can be used in the present invention include 3,4-dihydroxyphenylalanine, 4-hydroxyphenylglycine, 3,5
-Dothyronine and m-tyrosine may be mentioned;
These are given for illustrative purposes only and are not limiting in any way.

A solution of this diastereomeric ester is contacted with a bachromatographic support. It has been found that a wide variety of supports can be used in practicing the entire invention. A group of preferred supports are silica, alumina and zeolites. Another group of supports that may be used in the practice of this invention consists of supports commonly used in reverse phase chromatography.

Reversed phase chromatography is a branch of chromatography in which the mobile phase is more polar than the stationary phase, ie, the support.

The support used in the reversed-phase Chromadog 7 Fee is modified silica, that is, silica modified by silanization so that the silica surface has nonpolar groups in place of the hydroxyl groups on the surface. Silicas of this type may have, for example, long chain alkyl, aryl, amino or cyano groups attached to the surface by si-containing moieties.

This polymeric support can be prepared by solvent or solvent system.
Eluted under Madograph conditions.

1 Chromatographic Conditions'' means the general principles of climatographic separation known to those skilled in the art and commonly applied to liquid-solid chromatographic operations.The general principles thus apply, e.g. The flow is uniform, preventing the formation of gas bubbles, minimizing disturbance of the solid support, etc. Solvents that can be used in the process of the invention are those typical of chromatographic methods.

Such solvents include aliphatic hydrocarbons, such as pentane, hexane, heptane, octane, nonane, decane, cyclopentane, cyclohexane, diisobutylene, pentene, hexene; aromatic hydrocarbons, such as t &1 benzene, toluene, xylene, Ethylbenzene, diethylbenzene, methylethylbenzene, etc.; halogenated hydrocarbons,
Especially chlorinated and fluorinated hydrocarbons, such as chloroform,
methylene chloride, carbon tetrachloride, chloropropane, chlorobutane, chloropentane, fluoroalkane, bromoethane, chlorobenzene, chlorotoluene and ethylene chloride: sulfides, especially carbon disulfide; ethers, such as diethyl ether, tetrahydrofuran, tetrahydrobilane and dioxane; esters, especially Acetates, such as methyl acetate, butyl acetate and about 6 carbon atoms
Esters of the following saturated carzenic acids (however, the alcohol portion is saturated and the number of carbon atoms is about 6 or less); about 8 carbon atoms
The following ketones, such as acetone, citanone, pentanone, hexanone, heptanone and octanone; Nitroalkanes, such as nitromethane, nitroethane, nitropropane and nitrobutane; Amines, especially pyridine; Nitriles, such as acetonitrile, propionitrile and butyronitrile; approximately 8 carbon atoms The following alcohols (including diols and hydriols) include acetic acid and dimethyl sulfoxide.

At least one of the purified diastereomeric effluent fractions is then collected. This effluent is often monitored for specific performance to ensure proper effluent separation. For example, specific ultraviolet or infrared absorption or refractive index may be monitored as typical properties often used to determine effluent fractions. Since the concentration of purified diastereomers can be low, differential
ntiaj) measurements are particularly useful.

After collecting the purified diastereomer effluent, the purified diastereomer is treated to liberate the optically active amino acid. The most common method of purifying diastereomers to obtain optically active 13-amino acids is hydrolysis. The most typical hydrolysis is acid- and base-catalyzed. Base-catalyzed hydrolysis is somewhat preferred because it thereby minimizes the overall optical purity of the regenerated amino acids and simultaneously minimizes racemization of the free cyclohexanol recovered.

Mineral acids are suitable examples of acid catalysts, and alkali metal hydroxides and carbonates are examples of suitable base catalysts. The optically active amino acid is then recovered by appropriate means.

The following examples are merely illustrative of the present invention and are not intended to limit the invention in any way. The chromatographic equipment used in these examples was operated at 10,000 psi.
A pump capable of obtaining a flow rate of less than 10 m13 per minute, a non-diaphragm injector with zero dead center capacity fittings, a suitable column for separation, a two-channel absorption detector for total effluent monitoring at 254 and 280 nm, and It consists of a 2-channel recorder. All pipes used are outside 1f71
It is stainless steel, 6 inches long and has an internal diameter of 0.009 inches. The column is made of stainless steel with an internal diameter of 4.6 ms and a length of 2.5 m.

All solvents used as eluents were degassed before use.
It was also filtered. The hexane was freshly distilled and the eluent of the previous phase was dried using anhydrous magnesium sulfate. As the eluent and other chemicals, commercially available products were used as they were without purification.
A round bottom flask was equipped with a reflux condenser, a magnetic stirrer and a heating mantle, and d.

, g-phenylglycine 2.509 (0,0165 mol), l-menthol 3.0011 (0,0192 mol), p-toluenesulfonic acid monohydrate 3.659 (0
,0192 mol), benzene 30 ml and toluene 13
Added m1f. The resulting slurry was gently heated under reflux for 3 days with stirring, and the produced water was removed by azeotropic distillation. After filtering the cooled slurry and treating the solids with 10% aqueous sodium carbonate, the resulting base mixture was extracted with diethyl ether. The ether phase was dried over magnesium sulfate and the solvent was removed to yield a crystalline mixture of diastereomers and unreacted menthol (4.39 g). Unreacted menthol was then removed by sublimation. In a similar manner l-
One of the diastereomers was prepared separately from menthol and l-phenylglycine.

Example 2 A diastereomeric ester was produced from menthol racemate and (-)-phenylchrysine in the same manner as in Example 1. Separation was carried out on a silica gel column using a 2-propanol-hexane solvent system as the eluent. The insensitivity of orchids to eluent compositions is shown in the table below. This table shows the α values measured for various eluents.

(Left below) α value for 2-propanol-hexane mixture on silica gel 20 1.53IQ
1.54 5 1.511
1.44 Because the pair of diastereomers used in these experiments is equivalent to the pair of diastereomers obtained from the i-potent haphenylglycine racemate and the mendol enantiomer, these experiments demonstrate that the diastereomeric esters of the present invention may be used with a change in eluent. It shows that he feels relatively little about it.

Example 3 A solution containing diastereomeric mixture 2, B9 prepared in Example 1 was eluted with silica gel (
375,9) was chromatographed. The diastereomers were circulated twice and four fractions were collected. The fractions containing relatively few 17-<n diastereomers were combined to give 0.90% of 99% pure material.

The remaining fractions were combined to yield 1.1 g of a relatively large amount of retained diastereomer with a purity of 99%.

The remainder of the substance was mostly l-menthol. Example 4 The diastereomeric ester obtained from the phenylglycine racemate and l-menthol shown in Example 1 was subjected to reverse phase chromatography using octadecyl crane-modified silica gel. Separated under graph conditions. The eluent is a solution consisting of 10% 0.1 M ammonium acetate aqueous solution and 90% acetonitrile. Under these conditions the α value was 1.08.

Example 5 Diastereomeric esters of menthol and various amino acids were prepared from 1-menthol and amino acid racemates Example 1
Manufactured by a general method. Separation of these diastereomers was carried out on a silica gel column using 20% 2-propanol-hexane as a solvent. From the table below, the diastereomeric esters of menthol and amino acids have alpha values that allow chromatographic separation, and many of these esters have alpha values that make this separation extremely easy.
It turns out that it has a value.

Phenylglycine 1.53 4-Hydroxyphenylglycine 1.39 Tyrosine 1.10 Phenylalanine 114 Tryptophan 1.30 Example 6 To l-phenylalanine-1-menthyl ester in a 100 rnl round bottom flask with a magnetic stirrer, A few equivalents of sodium hydroxide dissolved in g ml of methanol were added. After standing for 5 hours at room temperature, 15 TL of water was added to the reaction mixture.
was added and the resulting solution was extracted three times with about 20 ml of diethyl ether. The ether extract was dried over anhydrous magnesium sulfate, filtered, and concentrated to l-menthol o, 39 g.
'eT'4.

The aqueous phase was neutralized with dilute hydrochloric acid and then treated with 2-propanol, and the water and methanol were removed azeotropically to obtain a residual odor of sodium chloride and phenylalanine. The residual aroma was carefully extracted with just enough water to remove the sodium chloride, yielding phenylalanine. This phenylalanine residue was dried using azeotropic extraction with 2-propanol and further vacuum dried to obtain [α]” = −33,3±
0.36 of a white powder of 1.6° (vertical 1,23.HtO) was obtained.

Claims (1)

[Claims] 1. A solution containing a diastereomeric ester obtained from an amino acid racemate and an optically active 2-isoprogyl-5-methylcyclohexanol is brought into contact with a four-mad-grab support, and said support is Elute with a solvent under chromatographic conditions, collect at least one effluent fraction containing the purified diastereomer, process the purified diastereomer to liberate the optically active amino acid, and then process the purified diastereomer to liberate the optically active amino acid. 1. A method for producing optically active amino acids, characterized by recovering amino acids. 2. The method according to claim 1, wherein cyclohexanol is menthol. 3. The method according to claim 2, wherein the menthol is l-menthol. 4. The method according to claim 1, wherein the chromatographic support is selected from the group consisting of silica, alumina, modified silica, and zeolite. 5. The method according to claim 1, wherein the optically active amino acid is L-3,4-dihydroxyphenylalanine. 6. The method according to claim 1, wherein the purified diastereomer is hydrolyzed to liberate an optically active amino acid. 7. The method according to claim 6, wherein the hydrolysis is carried out with a base catalyst. 8. The method of claim 6, wherein said hydrolysis is carried out with an acid catalyst.