JPH0745412B2 - Separation method - Google Patents

Description

DETAILED DESCRIPTION OF THE INVENTION [Industrial Field of the Invention] The present invention relates to the analysis of a mixture consisting of a compound having two asymmetric centers, by separating two isomers called a meso form and a racemate, Alternatively, it relates to a method for isolating and purifying a pure product from a mixture. INDUSTRIAL APPLICABILITY The separation method of the present invention can be used in the chemical industry, the pharmaceutical industry, etc. for the purity measurement or purification of products by synthesis / reaction.

[Prior Art and Problems] A meso body and a racemic body are plane or axisymmetric, and a compound having two asymmetric carbons, in which one and the other asymmetric centers have opposite absolute configurations (meso body) ) With the same absolute configuration (racemic). These are chemical,
Separation tends to be considered easy because it exhibits physically different properties.

However, in reality, the functional groups of these isomers are the same, and only the stereochemistry is different. Therefore, this separation is not easy as in the case of separating optical isomers. For example, in the case of a chromatographic method using silica gel, which is a conventional general chromatographic stationary phase, the separation of a meso isomer and a racemic mixture is far incomplete.

[Means for solving problems]

The inventors of the present invention have conducted extensive studies based on the idea that a stationary phase having a high structural regularity is necessary for distinguishing stereochemical differences, and as a result, the immobilization using a natural polysaccharide or a derivative thereof as an active ingredient. The present invention is based on the finding that a phase is effective for separating meso and racemic isomers containing a polar π bond group.

That is, in the present invention, a mixture of a meso form and a racemate of a compound having 1 to 2 polar π bond groups in the molecule and having 2 asymmetric centers is converted into each isomer by a chromatography method. In the method for separating, a polysaccharide or a derivative thereof is used as a stationary phase.

The meso form and the racemic form that are the targets of separation in the present invention are polar π
A meso form and a racemic form of a compound containing one or two bonding groups. Examples of the polar π-bonding group in the present invention include a carbonyl group (including ester and amide), a cyano group, a nitro group, a sulfinyl group, a sulfonyl group, an azido group, and other multiple bond systems containing a hetero atom.

These polar π-bonding groups have a polar interaction with the carbonyl group of the polysaccharide or its derivative used as the stationary phase, preferably the ester derivative of the polysaccharide, to induce adsorption.

Hereinafter, the racemic body and the meso body which can be separated by the stationary phase are exemplified.

One polar π bond group Two polar π bond groups In the above formula, R is an alkyl group having 6 or less carbon atoms, an aryl group or an unsaturated alkyl group; R'is H, CH ₃ or C ₂ H ₅ ; X is NO ₂ , C
H, CO ₂ CH ₃ ,, CO ₂ C ₂ H ₅ ,, CONR ' ₂ , CO ₂ C ₆ H ₅ , CONHC ₆ H ₅ , COC ₆ H
₅ , COCH ₃ , N ₃ , OCOCH ₃ , OCOC ₆ H ₅ , NHCOCH ₃ , NHCOC ₆ H ₅ ,
Or NHCOH; Y is> C = 0 or> SO ₂ .

The meso form of the above compound has plane symmetry, and the racemic form has axial symmetry.

The stationary phase used in the chromatography method of the present invention contains a polysaccharide or a derivative thereof as an active ingredient. The polysaccharide as used herein may be any one of optically active, regardless of whether it is a synthetic polysaccharide, a natural polysaccharide, or a modified natural polysaccharide, but is preferably a highly regular homoglycan and has a constant binding mode. There is something. More preferably, it is possible to easily obtain a high-purity polysaccharide, cellulose, amylose, β-1,4-chitosan, chitin, β-1,4-mannan, β-1,4-xylan, inulin, α-1, 3-glucan, β-1,3-glucan and the like, and cellulose is particularly preferable. The polysaccharide derivative is a derivative in which some or all, preferably 85% or more, of the hydrogen atoms on the hydroxyl group of the above-mentioned polysaccharide are replaced with other atomic groups.

The atomic group here is -R, R is an aliphatic group having 1 to 3 carbon atoms, 3
A cycloaliphatic group having 8 to 8 carbon atoms, an aromatic group having 4 to 20 carbon atoms or a heteroaromatic group, each of which may have a substituent. These derivatives can be easily obtained by using various known chemical reactions. These polysaccharides and their derivatives are particularly suitable for industrial chromatographic separation due to the availability of raw materials and the stability.

In the separation method of the present invention, the desired meso form and racemic form can be separated by selecting an appropriate one from these polysaccharides or derivatives thereof, and the ester derivative of the polysaccharide is particularly preferable.

Since the chromatographic separating agent is preferably granular, in order to use the polysaccharide or its derivative as a compound separating agent, it is preferable to crush the polysaccharide or its derivative or to make it into a bead form. The size of the particles varies depending on the size of the column or plate used, but is 1 μm to 10 mm, preferably 1 to 300 μm, and the particles are preferably porous.

Furthermore, in order to improve the pressure resistance of the separating agent, prevent swelling and contraction due to solvent substitution, and improve the theoretical plate number, the polysaccharide or its derivative can be held on a carrier. The size of the suitable carrier varies depending on the size of the column or plate used, but is generally 1 μm to 10 mm, preferably 1 to 300 μm.
m. The carrier is preferably porous and has an average pore diameter of 10Å to 100 µm, preferably 50Å to 10000Å. The amount of the polysaccharide or its derivative held is 1 to 100% by weight, preferably 5 to 50% by weight, based on the carrier.

The method of holding the polysaccharide or its derivative on the carrier may be a chemical method or a physical method. As a physical method, a polysaccharide or a derivative is dissolved in a soluble solvent, well mixed with a carrier, and the solvent is distilled off by an air stream under reduced pressure or heating, or a polysaccharide or a derivative thereof is dissolved in a soluble solvent. : There is also a method in which the solvent is diffused after being thoroughly mixed with a carrier and then stirred and dispersed in a liquid which is incompatible with the solvent. When crystallizing the polysaccharide or its derivative held on the carrier in this manner, a treatment such as heat treatment can be performed. In addition, it is possible to change the retention state and eventually the separability by adding a small amount of solvent to swell or dissolve the polysaccharide or its derivative and then distilling off the solvent again.

The carrier may be a porous organic carrier or a porous inorganic carrier, and is preferably a porous inorganic carrier. Suitable examples of the porous organic carrier include polymeric substances such as polystyrene, polyacrylamide and polyacrylate. Suitable porous inorganic carriers include synthetic or natural substances such as silica, alumina, magnesia, titanium oxide, glass, silicates, and kaolin, and surface treatment for improving affinity with polysaccharides or their derivatives. You may go. Examples of the surface treatment method include a silanization treatment using an organic silane compound and a surface treatment method by plasma treatment.

When a polysaccharide or a derivative thereof is used for the stationary phase, the separation characteristics may change depending on the physical state such as molecular weight, crystallinity, orientation, etc. even if the derivatives are chemically the same. After giving a shape suitable for the intended use, or in the process of giving, a heat treatment, etching or other physical or chemical treatment can be applied.

The chromatographic methods for separating the meso form and the racemic form using the above separating agent include gas chromatography, liquid chromatography, thin layer chromatography and the like.

The developing solvent for liquid chromatography or thin layer chromatography is not particularly limited, except for a liquid which dissolves or reacts with the separating agent. When the separating agent is bound to the carrier by a chemical method or is insolubilized by crosslinking, there is no limitation except the reactive liquid. Needless to say, the separation characteristics of the isomers change depending on the developing solvent, so it is desirable to study various developing solvents.

When performing thin layer chromatography, 0.1 μm
A layer having a thickness of 0.1 mm to 100 mm consisting of the separating agent of the present invention composed of particles of about 0.1 mm and a small amount of a binder, if necessary, may be formed on the support plate.

[Action]

In the present invention, the reason why the separation of the meso form and the racemic form is effectively achieved by using the polysaccharide or its derivative, particularly the ester derivative of the polysaccharide as the stationary phase is not clear, but the presumed mechanism is as follows. Is like.

FIG. 1 will be described by taking a meso form and a racemic form of 2,6-dimethylcyclohexanone as an example. This isomer is effectively separated by using cellulose tribenzoate as a stationary phase, and FIG. 1 is a schematic diagram showing a molecular structure for explaining the separation mechanism. In the figure, the solid line shows a part of the cellulose tribenzoate skeleton. Three
The benzoyl group at the position avoids the overlapping interaction with C—H at the 2-position on the glucose ring, and also prevents the adjacent 6-position of the glucoside residue.
It is presumed that it is inclined with respect to the molecular axis as shown in the figure in order to alleviate steric repulsion with the position substituent. When the carbonyl compound to be separated interacts with this benzoyl group, the arrangement as shown in the figure in which the respective dipoles are close to each other with opposite orientations is the most conceivable. At this time, racemate (R ¹ , R ⁴ = CH ₃ , R ² , R ³ = H or R ¹ , R ⁴ = H, R ² , R
³ = CH ₃ ), R ⁴ and a benzene ring of a benzoyl group,
Alternatively, steric repulsion between R ³ and the glucose ring prevents access. However, in the meso form, R ¹ = R ² = CH
_When approaching such that ₃ , R ³ = R ⁴ = H, it is considered that there is almost no steric hindrance, and thus it is more strongly retained than the racemate.

〔The invention's effect〕

According to the present invention, a meso form and a racemic form can be efficiently separated by a stationary phase using an inexpensive raw material such as a polysaccharide or a derivative thereof. Therefore, the present method can be advantageously used for the purpose of purifying both from a mixture of meso form and racemic form, which often results in a chemical synthesis reaction, or for the purpose of analysis for measuring the purity of each. Therefore, the separation method of the present invention is expected to make a great contribution to simplification of manufacturing processes, process control, research, etc. in fields such as the chemical industry and the pharmaceutical industry.

〔Example〕

The present invention will be specifically described below with reference to examples, but it goes without saying that the present invention is not limited thereto. In addition, as a column for liquid chromatography, about 20% by weight is applied to silica gel (Lichrospher Si 1000 manufactured by Merck) obtained by treating a polysaccharide ester derivative with diphenylsilane.
The packing material obtained by loading was packed in a stainless steel column having a length of 25 cm and an inner diameter of 0.46 cm. The eluting isomers were detected by using a photometer equipped with an ultraviolet detector and a flow cell.

Comparative Example 1 A chromatogram obtained by developing a mixture of 2,6-dimethylcyclohexanone meso-form and cerami-form on silica gel is shown in FIG. 2 (mobile phase: hexane-2-propanol 100: 1, 1 ml / min, 22 ℃ detection 280nm).

Example 1 A chromatogram of the same compound as in Comparative Example 1 developed using cellulose tribenzoate is shown in FIG. 3 (mobile phase: hexane-2-propanol 100: 1, 0.5 ml / min, 22
℃ detection 280nm).

The first two peaks are due to the racemate eluting with optical resolution, and the latter large peaks are the meso form. It can be seen that the racemate and the meso form are much better separated than in the comparative example.

Comparative Example 2 A chromatogram obtained by developing a mixture of 2,4-pentanediyl diacetate meso-form and racemic form on silica gel is shown in FIG. 4 (conditions are the same as in Comparative Example 1. Is 220 nm.).

It can be seen that the separation is incomplete.

Example 2 A chromatogram obtained by developing the same compound as in Comparative Example 2 using cellulose tribenzoate is shown in FIG. 5 (conditions are the same as in Example 1, but the detection wavelength is 220 nm). Also in this case, it is understood that the racemic body is eluted first with optical resolution, and the meso body is eluted as a clearly separated peak.

[Brief description of drawings]

FIG. 1 is a schematic diagram showing the molecular structure of 2,6-dimethylcyclohexanone in the separation of meso form and racemic form using cellulose tribenzoate, and FIGS. 2 and 3 are 2,6.
-In the chromatogram of dimethylcyclohexanone, the second
Figure shows silica gel, Figure 3 shows the case where cellulose tribenzoate is used as stationary phase, and Figures 4 and 5 show 2,
On the chromatogram of 4-pentanediyl diacetate,
FIG. 4 is a diagram when silica gel is used, and FIG. 5 is a diagram when cellulose tribenzoate is used as a stationary phase.

─────────────────────────────────────────────────── ─── Continuation of the front page (51) Int.Cl. ⁶ Identification code Internal reference number FI technical display area C07C 49/403 67/56 69/16

Claims (1)

[Claims] 1. Chromatography using a mixture of a meso-form and a racemate of a compound having one or two polar π-bonding groups in the molecule and having two asymmetric centers as a stationary phase of a polysaccharide or a derivative thereof. A method for separating a meso form and a racemic form, which is characterized by separation by a graphic method.