JPH06102151B2 - Ligand exchange type silica gel filler - Google Patents

Description

TECHNICAL FIELD The present invention relates to a ligand exchange type silica gel packing material for liquid chromatography having excellent durability. Particularly, the filler of the present invention is useful as a separating agent for optical resolution.

(Prior art and problems) In general, silica gel packing has excellent performance as a packing material for liquid chromatography and is widely used today, but its use in the basic region is remarkable because of its acidic property. Be restricted. Further, the use of a protic solvent such as water or alcohol also causes deterioration due to dissolution of the silica gel. Therefore, in order to prevent deterioration to some extent, a method of using a silane coupling agent to remove acidic silanol has been used. However, even with these methods, it is difficult to completely eliminate silanol, and deterioration over a long period of time cannot be avoided because of the attack of the protic solvent on the silane treating agent. In particular, in the case of a packing material for reversed phase liquid chromatography and a packing material for ligand exchange type liquid chromatography which use a polar solvent, a problem occurs because the retention capacity changes with time.

(Means for Solving the Problems) The present inventors substantially inactivate the surface silanol by reacting with the silanol on the surface of the silica gel or forming a thin film on the surface of the silica gel, and then silane coupling. As a result of diligent search for compounds capable of avoiding long-term deterioration due to attack of a protic solvent on a silane treating agent by reacting with an agent, an element compound of Group IVb of the periodic table excluding silicon was found. The present invention has been accomplished by obtaining a ligand-exchange type silica gel filler having excellent durability by treatment.

That is, the present invention provides a layer of an element or an elemental compound of Group IVb of the periodic table excluding silicon on the surface of silica gel, and maintains a compound capable of forming a complex with a transition metal ion thereon. The present invention relates to a characteristic ligand-exchange type silica gel filler.

The ligand exchange type liquid chromatography packing material in the present invention is for carrying out mutual separation or optical resolution of water-soluble compounds such as amino acids and hydroxy acids, and the separation target is a base material which is a stationary phase. Separation is carried out by interacting with the ligand supported on (i.e., the residue of a compound having a group involved in complex formation with a transition metal ion) via the transition metal ion.

The deterioration of the silica gel filler is caused by the silica gel itself being dissolved in the solvent, or the bound sila coupling agent being desorbed. Therefore, it is necessary to eliminate the exposure of silanol in order to avoid dissolution of silica gel, and to strengthen the bond in order to prevent desorption of the silane coupling agent. In order to satisfy both of them at the same time, as a result of diligent study, Periodic Table IVb excluding silicon
It has been found that a layer of a group element or elemental compound is provided on the surface of silica gel. Preferable examples of the elements of Group IVb of the periodic table excluding silicon are titanium and zirconium. Examples of the compound include titanium chloride, titanium-n-propoxide, titanium-iso-isopropoxide, titanium-n-butoxide, zirconium chloride, zirconium oxychloride, zirconium-n.
-Propoxide, zirconium-iso-propoxide,
Zirconium-n-butoxide and the like can be mentioned. These compounds may be used alone or as a mixture, and if necessary during the reaction with the silane coupling agent, the reaction can be facilitated by treating under a water-containing condition.

In the present invention, as a method of providing a layer of an element or an element compound of Group IVb of the periodic table excluding silicon on the surface of silica gel, there is a method of reacting with silanol on the surface of silica gel. In this case, any element or elemental compound of Group IVb of the periodic table excluding silicon may be any as long as it reacts with the silanol.

As another method, there is a method of physically adsorbing a thin film of an element or an elemental compound of Group IVb of the periodic table excluding silicon by sputtering or the like. The silica gel as the base material has a particle size of 1 to 100 μm and a pore size distribution peak of 10 to 2
000Å is preferable, and the shape may be spherical or crushed.

In the present invention, the specific compound capable of forming a complex with a transition metal ion includes a residue which, after being supported by reacting with the above-mentioned surface-treated silica gel, acts as a ligand. Examples thereof include optically active or optically inactive amino acids or transition metal salts thereof, or optically active or inactive amino alcohols, α-hydroxycarboxylic acids or transition metal salts thereof, diamines, dicarboxylic acids or transition metal salts thereof, and the like. To be

In the present invention, as a method for supporting this specific compound on silica gel provided with a layer of an element or an element compound of Group IVb of the Periodic Table excluding silicon, it is preferable to react via a silane coupling agent.

As the silane coupling agent, 3-glycidoxypropyltrimethoxysilane, 3-glycidoxypropyldiethoxymethylsilane, 3-glycidoxydimethylethoxysilane, 2- (3,4-epoxycyclohexylethyl) trimethoxy Examples thereof include silane, 3-chloropropyltrimethoxysilane, 3-chloropropyltrichlorosilane, and silane compounds having a linear alkyl group having 6 to 24 carbon atoms.

These silane coupling agents may be used alone, or may be used in combination with these or other silane coupling agents. Further, after reacting these silane coupling agents, it is possible to eliminate the remaining hydroxyl groups by reacting them with the residual hydroxyl groups on the surface of the substrate using a silane coupling agent having higher reactivity.

In the case of actually synthesizing a separation filler, after reacting a silane coupling agent, a specific compound is further reacted with a reactive residue of the silane coupling agent, or a specific compound is previously added to the silane coupling agent. It may be reacted with a ring agent and then reacted with silica gel provided with a layer of an element or an element compound of Group IVb of the periodic table excluding silicon. When the compound containing a residue acting as a ligand has a hydrophobic group such as a long-chain alkyl group, it can be supported via a silane coupling agent by a hydrophobic bond that does not involve a reaction.

The reaction of the element compound of Group IVb of the periodic table excluding silicon with silica gel can be carried out by dissolving the above compound in a solvent capable of dissolving it and contacting it with silica gel, but if necessary, heating Can accelerate the reaction. Further, it is preferable to add a dehydrochlorinating agent if necessary. On the other hand, when these compounds are physically adsorbed on the silica gel surface or a thin film is formed, the solvent is distilled off after contacting these compounds with the silica gel surface, and then heating is performed. It can be more or less accompanied by polymerization of the above compounds.

Silica gel reacted with an element compound of Group IVb of the periodic table excluding silicon can have a hydroxyl group on the surface by contact with water, and can be easily reacted with a silane coupling agent. The reaction with the silane coupling agent may be carried out by an ordinary method, for example, by heating under reflux in benzene or toluene, or by reacting in a polar solvent such as water, methanol, acetonitrile, etc., depending on the silane coupling agent. The method of adding the hydrochloric acid agent may be appropriately selected.

However, in the case of using a ligand exchange type filler, after reacting a silane coupling agent, a compound having a residue acting as a ligand is further reacted with a reactive residue of the silane coupling agent, or A compound having a residue acting as a ligand is previously reacted with the silane coupling agent, and this is reacted with an element or an element compound of Group IVb of the periodic table excluding silicon by the above method or a thin film thereof is physically formed. It can be appropriately selected whether to react with silica gel that has been adsorbed.

In addition, as a metal ion that is the center of ligand exchange in actual chromatography, divalent copper, nickel,
Examples of zinc, trivalent cobalt, iron, etc., these metal ions may be complexed to the stationary phase in advance,
Further, it may be added to the solvent at the time of analysis.

(Effects of the Invention) A ligand exchange type liquid chromatography packing material is used for mutual separation or optical resolution of water-soluble compounds such as amino acids, hydroxy acids, amino alcohols, etc. The packing material obtained by reacting the agent has a problem that the hydrogen ion concentration of the eluent can be used only in a narrow range, and thus the separation ability cannot be fully exhibited. However, since the highly durable filler obtained by the present invention can use an eluent having a wider range of hydrogen ion concentration, it is expected to be applicable to the separation of a wider range of compounds.

(Examples) Hereinafter, the present invention will be described in detail with reference to Examples, but it goes without saying that the present invention is not limited thereto.

Synthesis Example 1 3.22 g of zirconium oxychloride octahydrate was added to 100 parts of methanol.
Dissolve in silica gel and add silica gel (E.MERCK Lichrosor
b SI100,10μm) 2.1g triethylamine after adding 10g
Is added and the mixture is heated under reflux for 8 hours. After cooling this, it is filtered with a glass filter, and the surface is hydroxylated by washing with water, and then dried. The presence of Zr element was confirmed by fluorescent X-ray analysis in the silica gel thus obtained. The silica gel was further silanized by heating to reflux with 5 ml of 3-glycidoxypropyltrimethoxysilane in dry benzene for 5 hours.

Synthesis Example 2 3.40 g of tetra-n-butyl titanate was dissolved in 50 ml of ethyl acetate, and this was dissolved in silica gel (Lichrosorb S manufactured by E.MERCK).
10 g of I100, 10 μm) was suspended in 50 ml of ethyl acetate, and the mixture was heated under reflux for 8 hours. After cooling this, it is filtered with a glass filter, and the surface is hydroxylated by washing with water, and then dried. In the silica gel thus obtained, the presence of Ti element was confirmed by fluorescent X-ray analysis.
The silica gel was further silanized by heating to reflux with 5 ml of 3-glycidoxypropyltrimethoxysilane in dry benzene for 5 hours.

Synthesis Example 3 3.22 g of zirconium oxychloride octahydrate was added to methanol 100
Dissolve in silica gel and add silica gel (E.MERCK Lichrosor
b SI100,10μm) 2.1g triethylamine after adding 10g
Is added and the mixture is heated under reflux for 8 hours. After cooling this, it is filtered with a glass filter, and the surface is hydroxylated by washing with water, and then dried. The presence of Zr element was confirmed by fluorescent X-ray analysis in the silica gel thus obtained. 13.7 g of L-proline Na salt was dissolved in 150 ml of methanol to give 3-glycidoxypropyltrimethoxysilane 11.8 g.
react with g. After the methanol was distilled off, 150 ml of acetonitrile was added and dissolved, to which silica gel treated with zirconium oxychloride was added and reacted at room temperature for 2 days to silanize and carry the ligand.

Synthesis Example 4 3.40 g of tetra-n-butyl titanate was dissolved in 50 ml of ethyl acetate, and this was dissolved in silica gel (Lichrosorb S manufactured by E.MERCK).
10 g of I100, 10 μm) was suspended in 50 ml of ethyl acetate, and the mixture was heated under reflux for 8 hours. After cooling this, it is filtered with a glass filter, and the surface is hydroxylated by washing with water, and then dried. In the silica gel thus obtained, the presence of Ti element was confirmed by fluorescent X-ray analysis.
13.7 g of L-proline Na salt was dissolved in 150 ml of methanol,
React with 11.8 g of 3-glycidoxypropyltrimethoxysilane. After distilling off methanol, acetonitrile
150 ml was added and dissolved, to which silica gel treated with the above tetra-n-butyl titanate was added and reacted at room temperature for 2 days to silanize and carry a ligand.

Example 1 L-proline was added to the silanized silica gel obtained in Synthesis Example 1.
L-proline Na salt is bound to silica gel by reacting Na salt in methanol at room temperature, and further converted into a copper salt by transferring into sodium sulfate aqueous solution. The silica gel packing thus obtained was filled in a stainless steel column having a length of 25 cm and an inner diameter of 0.46 cm, and 1 ml / min of a 0.25 mM copper sulfate aqueous solution was passed as an eluent, and most of the racemic amino acid was optically resolved. The holding time did not change over 300 hours.

Example 2 L-proline was added to the silanized silica gel obtained in Synthesis Example 2.
L-proline Na salt is bound to silica gel by reacting Na salt in methanol at room temperature, and further converted into a copper salt by transferring into sodium sulfate aqueous solution. The silica gel packing thus obtained was filled in a stainless steel column having a length of 25 cm and an inner diameter of 0.46 cm, and 1 ml / min of a 0.25 mM copper sulfate aqueous solution was passed as an eluent, and most of the racemic amino acid was optically resolved. The holding time did not change over 300 hours.

Example 3 Using a packed column obtained in the same manner as in Example 1, 0.25
When Tris buffer was used as the eluent instead of the mM copper sulfate aqueous solution, most of the racemic amino acid was optically resolved as in Example 1, and the retention time did not change for 300 hours or more.

Example 4 Using a packed column obtained in the same manner as in Example 2, 0.25
When Tris buffer was used as the eluent instead of the mM copper sulfate aqueous solution, most of the racemic amino acid was optically resolved as in Example 1, and the retention time did not change for 300 hours or more.

Example 5 The Na salt filler obtained in Synthesis Example 3 is converted into a copper salt by further transferring it into an aqueous solution of copper sulfate. The packing material thus obtained was packed in a stainless steel column having a length of 25 cm and an inner diameter of 0.46 cm, and 0.25 mM copper sulfate aqueous solution was used as an eluent for 1 ml / min.
When shed, many of the racemic amino acids were optically resolved,
Its holding time did not change over 300 hours.

Example 6 The Na salt filler obtained in Synthesis Example 4 is converted to a copper salt by further transferring it into an aqueous solution of copper sulfate. The packing material thus obtained was packed in a stainless steel column having a length of 25 cm and an inner diameter of 0.46 cm, and 0.25 mM copper sulfate aqueous solution was used as an eluent for 1 ml / min.
When shed, many of the racemic amino acids were optically resolved,
Its holding time did not change over 300 hours.

Comparative Example 1 Silica gel (Lichrosorb SI100, 10 μm manufactured by E.MERCK) 1
Silanization was carried out by heating 0 g of the mixture with 5 ml of 3-glycidoxypropyltrimethoxysilane in dry benzene under heating for 5 hours, and L-proline Na salt was reacted with this at room temperature in methanol to give L-proline on silica gel.
A silica gel filler obtained by binding Na salt and converting it into a copper sulfate solution by transferring it into an aqueous solution of copper sulfate was prepared to have a length of 25c.
0.25mM, packed in a stainless steel column with an inner diameter of 0.46cm
When an aqueous solution of copper sulphate was run as an eluent, the retention time decreased with time, it decreased to about 1/2 at 50 hours, then continued to decrease gradually, and after 200 hours, the optical resolution ability was considerably lost. It was Moreover, when Tris buffer was used as an eluent, the filler was dissolved.

Synthetic Example 5 Tetra n-butyl titanate (3.40 g) was dissolved in ethyl acetate (50 ml) and this was dissolved in silica gel (Develosil 100 manufactured by Nomura Chemical Co., Ltd.).
-5,) 10 g was suspended in 50 ml of ethyl acetate, and the mixture was heated under reflux for 8 hours. After cooling this, it is filtered with a glass filter, and the surface is hydroxylated by washing with water, and then dried. In the silica gel thus obtained, the presence of Ti element was confirmed by fluorescent X-ray analysis. 13.7 g of L-proline Na salt is dissolved in 150 ml of methanol and reacted with 11.8 g of 3-glycidoxypropyltrimethoxysilane. After distilling off methanol, acetonitrile 150 ml
Was added and dissolved, and silica gel treated with the above tetra-n-butyl titanate was added thereto, and the mixture was silanized by carrying out a reaction at room temperature for 2 days to carry a ligand, and then the Na salt was transferred to an aqueous solution of copper sulfate to form copper. I changed to salt. The filler was further dried in vacuum and then endcapped with N, O-bistrimethylsilylacetamide in dry toluene.

Example 7 The filler obtained in Synthetic Example 5 was used in a length of 25 cm and an inner diameter of 0.46.
It was packed in a cm stainless steel column, and 1 ml / min was run with 0.25 mM copper sulfate aqueous solution or phosphate buffer as an eluent. In both cases, most of the racemic amino acid was optically resolved, and the peak was extremely sharp, and the theory was high. A column with a plate number was obtained. The retention time of each sample did not change for more than 400 hours.

Synthesis Example 6 Zirconium oxychloride octahydrate 3.22 g was added to methanol 100
Dissolve it in ml and add silica gel (Develosil manufactured by Nomura Chemical
100-5,) 10 g was added, then triethylamine 2.1 g was added, and the mixture was heated under reflux for 8 hours. After cooling this, it is filtered with a glass filter, and the surface is hydroxylated by washing with water, and then dried. The presence of Zr element was confirmed by fluorescent X-ray analysis in the silica gel thus obtained. L
-Dissolve 13.7 g of proline Na salt in 150 ml of methanol,
Reacting with 11.8 g of glycidoxypropyltrimethoxysilane. After distilling off methanol, acetonitrile 15
Add 0 ml to dissolve, add silica gel treated with zirconium oxychloride to this and react at room temperature for 2 days to silanize and carry ligand, and then transfer Na salt to copper sulfate aqueous solution to form copper salt. Changed to. The filler was further dried in vacuum and then endcapped with N, O-bistrimethylsilylacetamide in dry toluene.

Example 8 The filler obtained in Synthetic Example 6 was used in a length of 25 cm and an inner diameter of 0.46.
It was packed in a cm stainless steel column, and 1 ml / min was run with 0.25 mM copper sulfate aqueous solution or phosphate buffer as an eluent. In both cases, most of the racemic amino acid was optically resolved, and the peak was extremely sharp, and the theory was high. A column with a plate number was obtained. In addition, the retention time of each sample did not change for more than 500 hours.

Claims (1)

[Claims] 1. A layer of an element or an element compound of Group IVb of the periodic table excluding silicon is provided on the surface of silica gel, and a compound capable of forming a complex with a transition metal ion is maintained thereon. Ligand exchange type silica gel filler.