JPH0475209B2 - - Google Patents

Description

DETAILED DESCRIPTION OF THE INVENTION The present invention provides a separating agent, particularly a packing material for liquid chromatography that is capable of optically resolving racemic compounds. Compounds that can be optically resolved using the separating agent of the present invention include, for example, amino acids, but amino acids usually require only one enantiomer, and such optical resolving fillers are optically active. It is in high demand as an important analytical tool for determining the purity of amino acids in the production of amino acids. (Prior art) Until now, fillers obtained by supporting amino acids or metal salts of amino acids on polymers have been
VADavankov et al. (Dokl.Akad.Nauk
SSSR, 986 (1971)) with cross-linked chloromethylated polystyrene and L-proline or L-
- A method that performs optical resolution using an eluent containing divalent copper or nickel ions in the polymer obtained by reacting methyl ester of hydroxyproline and then hydrolyzing the ester, or C.Quivoron.
L-proline was bonded to polyacrylamide via formaldehyde by J. et al.
Chromatogr., 204 , 185 (1981)). On the other hand, K. Sugden et al. (Reference: J.
Chromatogr, 192 , 228 (1980)), G. Gu¨bitz et al. (Reference: J. High. Resolut, Chromatogr. &
Chromatogr.Comm., 2 , 145 (1979)), VA
Davankov et al. (Reference: Angew.Chem.Int.Ed.Engl,
21, 930 (1982)). In addition, VADavankov et al. (Reference: Chromatographia 13 , 677 (1980)) and JP-A-Sho have used reverse-phase silica gel columns to retain amino acid derivatives.
57-176936, JP-A-58-96062, etc. Among these, when an amino acid such as L-proline or a derivative thereof is supported on chloromethylated polystyrene and polyacrylamide gel, a long reaction time is required to obtain a sufficient amount of support due to steric hindrance and poor reactivity. Further, in the method of supporting the silane compound on silica gel via a silane compound, there is a problem that the silane compound is detached by hydrolysis or the like. Furthermore, stability is a problem in the method of retaining in a reverse phase column. (Objective of the invention) The present invention is directed to the addition of metal salts of various amino acids to the glycidyl groups of a polymer obtained by copolymerizing glycidyl methacrylate, which has extremely reactive glycidyl groups, with other polymerizable monomers. The object of the present invention is to make it possible to obtain a filler having an excellent optical resolution ability and carrying an optically active group. (Structure of the Invention) The polymer used in the present invention is a copolymer of glycidyl methacrylate and other polymerizable monomers. Other polymerizable monomers include styrenic vinyl monomers such as styrene and para-methylstyrene; acrylic acid ester vinyl monomers such as methyl methacrylate, ethyl methacrylate, n-butyl methacrylate, tertiary-butyl methacrylate, hydroxyethyl methacrylate, 2-ethylhexyl methacrylate, Other vinyl monomers such as stearyl methacrylate, acrylonitrile, 4-
Examples include vinylpyridine, 2-vinylpyridine, acrylamide, and methacrylamide. These vinyl monomers can be used alone or in combination of two or more. The copolymerization composition of glycidyl methacrylate and these polymerizable monomers has a molar ratio of 0.001 to 1000, preferably
0.01 to 100, more preferably 0.1 to 10
It is. Examples of polymerization methods for these copolymers include radical polymerization using peroxides, azobisisobutyronitrile, and the like as an initiator, and ionic polymerization using Grignard reagents and alkyl metals. On the other hand, although the amino acid to be reacted with this needs to be optically active, the optical purity does not need to be strictly pure. Generally, it is preferable to use one of the naturally occurring optical isomers because of the ease of obtaining raw materials, but the invention is not limited to this. The polymer having an optically active group, which is a characteristic part of this filler, can be synthesized by adding a metal salt of an amino acid to a copolymer containing glycidyl methacrylate in a suitable solvent such as methanol, methylene chloride, etc.
Although it can be obtained by reacting alone or in a mixed solvent such as dimethylformamide, the method is not limited to this method, and a polymer may also be obtained by reacting glycidyl methacrylate with an amino acid metal salt and then polymerizing it. The obtained polymer has a partial structure represented by the following general formula (1) or (2). In the above formula, R ¹ , R ² , and R ³ are groups defined by the amino acid used, and in the case of proline or hydroxyproline, they form a ring structure represented by formula (2), and R ³ is -CH ₂ −CH ₂ −CH ₂ , −CH ₂ −
CH(OH) _CH2- . In the case of other amino acids, it is expressed by formula (1),
R ¹ and R ² are as shown in Table 1 below. [Table] In the above general formula, * represents an asymmetric carbon. Also, M represents a metal, and in the reaction, an alkali metal such as Na, K, Li or Ca,
Examples include alkaline earth metals such as Mg, but when used as a filler for optical resolution,
It is necessary to substitute transition metals such as Cu, Ni, and Co. The obtained polymer may be pulverized as it is and packed into a column, but it is usually supported on a carrier by physical adsorption or chemical bonding. It is possible to apply or chemically bond to an inorganic carrier such as silica gel or glass beads or an organic carrier such as polystyrene gel or polyacrylamide. In this case, it is preferable to apply or chemically bond the polymer containing glycidyl methacrylate and then react with the amino acid metal salt, but the invention is not limited thereto. When the polymer is used as it is, it is preferably crosslinked. (Effects of the Invention) The separating agent provided by the present invention can be used when optically resolving amino acids by liquid chromatography. In particular, when used in high performance liquid chromatography, a packing material with excellent pressure resistance can be obtained. The composition of the glycidyl methacrylate copolymer, which is another feature of the present invention, is closely related to the optical resolution ability, and by appropriately adjusting the composition, it is possible to significantly improve the optical resolution ability. Ta. The present invention will be described in detail below with reference to Examples, but the present invention is not limited thereto. Synthesis example 1 10.4g of glycidyl methacrylate and styrene
1.0 g of polymer obtained by copolymerizing 18.2 g (GMA/St molar ratio = 1/2.4) using azobisisobutyronitrile as an initiator was mixed with 5 ml of methylene chloride.
Dissolve in a mixed solvent of 10 ml of DMF, add 1.0 g of sodium salt of L-proline dissolved in 5 ml of methanol, and react for 2 days at room temperature. After distilling off the solvent, 7 ml of methanol was added again to dissolve it, and this was treated with diphenyldimethoxysilane to form silica gel 5 with an average particle size of 10 μm and a pore size of 1000 Å.
After adding g, methanol is distilled off and coated. After washing the obtained solid with water, it is converted into a copper salt by adding 2 g of copper sulfate to an aqueous solution of 50 ml of water to obtain a blue filler. In the following examples, the separation coefficient is given by the following equation. Separation coefficient = Volume ratio of more strongly adsorbed enantiomer / Volume ratio of more weakly adsorbed enantiomer Example 1 The packing material obtained in Synthesis Example 1 was packed with an inner diameter of 0.46 cm and a length of 0.46 cm.
When the racemic forms of various amino acids were optically resolved by filling a 25 cm stainless steel cylindrical column, the following results were obtained. Using 0.25 mmol copper sulfate as eluent at 50 °C.
The separation coefficients at a flow rate of 1.0 ml/min were as shown in Table 2. [Table] Synthesis Example 2 10.4 g of glycidyl methacrylate and 18.7 g of methyl methacrylate (GMA/MMA molar ratio = 1/
1.0g of polymer obtained by copolymerizing 2.6) using azobisisobutyronitrile as an initiator, into 15ml
To this, 1.0 g of the sodium salt of L-proline dissolved in 10 ml of methanol was added, and the mixture was allowed to react at room temperature for 2 days. After the solvent was distilled off, 7 ml of methanol was added again to dissolve it, and the solution was treated with diphenyldimethoxysilane to obtain the average particle size.
After adding 5 g of silica gel with a diameter of 10 μm and a pore size of 1000 Å, methanol is distilled off to form a coating. After washing the obtained solid with water, it is converted into a copper salt by adding 2 g of copper sulfate to an aqueous solution of 50 ml of water to obtain a blue filler. Example 2 The filler obtained in Synthesis Example 2 was packed with an inner diameter of 0.46 cm and a length of
When the racemic forms of various amino acids were optically resolved by filling a 25 cm stainless steel cylindrical column, the following results were obtained. Using 0.25 mmol copper sulfate as eluent at 50 °C.
The separation coefficients at a flow rate of 1.0 ml/min were as shown in Table 3. [Table] Synthesis Example 3 7.1 g of glycidyl methacrylate and 11.8 g of p-methylstyrene (GMA/PMS molar ratio = 1/2.0)
1.0 g of a polymer obtained by copolymerizing using azobisisobutyronitrile as an initiator was dissolved in 15 ml of methylene chloride, and 1.0 g of sodium salt of L-proline dissolved in 10 ml of methanol was added thereto.
Allow to react for 2 days at room temperature. After distilling off the solvent, 7 ml of methanol was added again to dissolve it, and this was treated with diphenyldimethoxysilane, with an average particle size of 10 μm.
After adding 5 g of silica gel with a pore size of 1000 Å, methanol is distilled off to form a coating. After washing the obtained solid with water, it is converted into a copper salt by adding 2 g of copper sulfate to an aqueous solution of 50 ml of water to obtain a blue filler. Example 3 The filler obtained in Synthesis Example 3 was packed with an inner diameter of 0.46 cm and a length of
When the racemic forms of various amino acids were optically resolved by filling a 25 cm stainless steel cylindrical column, the following results were obtained. Using 0.25 mmol copper sulfate as eluent at 50 °C.
The separation coefficients at a flow rate of 1.0 ml/min were as shown in Table 4. [Table] Synthesis Example 4 15 ml of chlorinated 1.0 g of polymer obtained by copolymerizing 14.2 g of glycidyl methacrylate and 5.3 g of acrylonitrile (GMA/AN molar ratio = 1/1) using azobisisobutyronitrile as an initiator. Dissolve in methylene, add 1.0 g of sodium salt of L-proline dissolved in 10 ml of methanol, and allow to react at room temperature for 2 days. After the solvent was distilled off, 7 ml of methanol was added again to dissolve it, and to this was added 5 g of silica gel with an average particle size of 10 μm and a pore size of 1000 Å treated with diphenyldimethoxysilane, and the methanol was distilled off to coat it. After washing the obtained solid with water, it is converted into a copper salt by adding 2 g of copper sulfate to an aqueous solution of 50 ml of water to obtain a blue filler. Example 4 The filler obtained in Synthesis Example 4 was prepared with an inner diameter of 0.46 cm and a length of
When the racemic forms of various amino acids were optically resolved by filling a 25 cm stainless steel cylindrical column, the following results were obtained. Using 0.25 mmol copper sulfate as eluent at 50 °C.
The separation coefficients at a flow rate of 1.0 ml/min were as shown in Table 5. [Table] Synthesis Example 5 10.0 g of glycidyl methacrylate and 10.0 g of butyl methacrylate (GMA/BuMA molar ratio = 1/
1.0 g of the polymer obtained by copolymerizing 1) using azobisisobutyronitrile as an initiator was dissolved in 15 ml of methylene chloride, and 1.0 g of the sodium salt of L-proline dissolved in 10 ml of methanol was added thereto at room temperature. Let it react for 2 days. After the solvent was distilled off, 7 ml of methanol was added again to dissolve it, and the solution was treated with diphenyldimethoxysilane to obtain the average particle size.
After adding 5 g of silica gel with a pore size of 10 μm and a pore size of 1000 Å, methanol is distilled off to form a coating. After washing the obtained solid with water, it is converted into a copper salt by adding 2 g of copper sulfate to an aqueous solution of 50 ml of water to obtain a blue filler. Example 5 The filler obtained in Synthesis Example 5 was packed with an inner diameter of 0.46 cm and a length of
The racemic forms of various amino acids were optically resolved by filling a 25 cm stainless steel cylindrical column, and the following results were obtained. Using 0.25 mmol copper sulfate as eluent at 50 °C.
The separation coefficients at a flow rate of 1.0 ml/min were as shown in Table 6. [Table] [Table] Synthesis example 6 14.2g of glycidyl methacrylate and styrene
1.0 g of a polymer obtained by copolymerizing 10.4 g (GMA/St molar ratio = 1/1) using azobisisobutyronitrile as an initiator was dissolved in 15 ml of methylene chloride, and sodium salt of L-proline was added to the solution. 1.0
Dissolve g in 10 ml of methanol, add and react at room temperature for 2 days. After the solvent was distilled off, 7 ml of methanol was added again to dissolve it, and this was treated with diphenyldimethoxysilane, with an average particle size of 10 μm and a pore size of 1000.
After adding 5 g of silica gel, methanol is distilled off to coat the mixture. After washing the obtained solid with water, it is converted into a copper salt by adding 2 g of copper sulfate to an aqueous solution of 50 ml of water to obtain a blue filler. Example 6 The filler obtained in Synthesis Example 6 was packed with an inner diameter of 0.46 cm and a length of
When the racemic forms of various amino acids were optically resolved by filling a 25 cm stainless steel cylindrical column, the following results were obtained. Using 0.25 mmol copper sulfate as eluent at 50 °C.
The separation coefficients at a flow rate of 1.0 ml/min were as shown in Table 7. [Table] Synthesis Example 7 4.7 g of glycidyl methacrylate and 11.8 g of paramethylstyrene (GMA/PMS molar ratio = 1/3.0)
1.25 g of a polymer obtained by copolymerizing with azobisisobutyronitrile as an initiator was dissolved in 15 ml of methylene chloride, and 0.8 g of the sodium salt of L-proline dissolved in 10 ml of methanol was added thereto at room temperature. Allow to react for several days. After the solvent was distilled off, 7 ml of methanol was added again to dissolve, and this was treated with diphenyldimethoxysilane, with an average particle size of 10 μm.
After adding 5 g of silica gel with a pore size of 1000 Å, methanol is distilled off to form a coating. After washing the obtained solid with water, it is converted into a copper salt by adding 2 g of copper sulfate to an aqueous solution of 50 ml of water to obtain a blue filler. Example 7 The filler obtained in Synthesis Example 7 was packed with an inner diameter of 0.46 cm and a length of
When the racemic forms of various amino acids were optically resolved by filling a 25 cm stainless steel cylindrical column, the following results were obtained. Using 0.25 mmol copper sulfate as eluent at 50 °C.
The separation coefficients at a flow rate of 1.0 ml/min were as shown in Table 8. 【table】

Claims (1)

[Claims] 1. A separating agent obtained by adding an optically active metal salt of an amino acid to the glycidyl group of a copolymer containing glycidyl methacrylate.