JPS63159756A - Adsorbent carrier for chromatography - Google Patents

Abstract

PURPOSE:To increase the hardness of a chromatography adsorbent carrier and to improve separability by bonding biotin to a porous gel-like copolymer consisting of glycidyl monovinyl ester, etc. to form the above-mentioned carrier. CONSTITUTION:The glycidyl monovinyl ester or glycidyl monovinyl ether is polymerized in such a manner that the epoxy group remains therein, then the polymer is crosslinked by alkylene glycol divinyl ester to obtain the porous gel-like copolymer. A compd. having a bond group, for example, epoxy group, amino group, hydrazino group, carboxyl group, formyl group, thiol group, etc., is bonded to the epoxy group of such gel-like copolymer. The biotin or aminoalkyl biotin is covalent-bonded to the above-mentioned gel-like copolymer via said bond group to form the adsorbent carrier. Since the adsorbent carrier for chromatography is obtd. by bonding the biotin bonding specifically to protein avidin, etc., to the gel-like copolymer, the carrier is usable for a pressure- resistant column and the shortened time for operation and the improved efficiency of refining by sepn. are attained.

Description

DETAILED DESCRIPTION OF THE INVENTION Field of Industrial Application The present invention relates to an adsorption carrier for chromatography that can be used in various chromatography including affinity chromatography.

As a type of conventional chromatography technology, affinity chromatography is a method of separating and purifying a biological material by utilizing the affinity of a pair of substances that specifically interact with each other. It is useful for identifying and purifying chemical properties, that is, a specific chemical structure on a molecule.

An adsorption carrier for affinity chromatography (Affinity Glue) is, for example, an active support obtained by bonding a binding group (spacer) to an insoluble carrier (matrix), and a bond is bonded to the spacer. An adsorption operation is performed by selecting a combination of substances that interact with this ligand.

As one of the ligands, biotin is a vitamin widely distributed in the liver, yeast, etc., and serves as a coenzyme for enzymes involved in carbonic acid fixation reactions. Soft glue specifically binds to the protein avidin in egg white, and immunology-related applications that take advantage of this characteristic are being carried out using soft glue, since the target is physiologically active substances that are unstable over time. , it is thought that there is a great advantage of speeding up the process.

For example, in separation, purification, and analysis by affinity chromatography in which biotin is used as a ligand and protein is used as the adsorption target substance, the desired properties of the active support, which is the main component of the adsorption carrier for affinity chromatography, are as follows: Low specific adsorption, high porosity, easy binding of ligand and large immobilization capacity, chemical stability and sufficient stability under a wide range of β range, salt concentration, and temperature conditions. Examples include no change in volume, sufficient mechanical strength and stability, and good flow properties, and resistance to biological contamination.

Cellulose, dextran, polyacrylamide, agarose, etc. that have been conventionally used as base materials for adsorption carriers for cyaffinity chromatography do not necessarily have these desired properties. However, since it is a so-called soft glue that lacks hardness, it has serious drawbacks such as poor flow characteristics and poor separation characteristics, and has a short shelf life.Furthermore, silica beads, which have been incorporated in recent years, have poor flow characteristics and poor separation characteristics. Although satisfactory in terms of hardness, it had the problem that it could not be used under alkaline conditions, which placed significant restrictions on the selection of separation conditions and elution/washing conditions.

Problems to be Solved by the Invention It is an object of the present invention to overcome the drawbacks of the conventional adsorption carriers for chromatography, and to provide a chromatography carrier having all the above-mentioned properties desired in an adsorption carrier for affinity chromatography. An object of the present invention is to provide an adsorption carrier for

The present invention provides an adsorption carrier for chromatography that can achieve the above object.

That is, the present invention provides a gel-like copolymer (hereinafter referred to as a gel-like copolymer) which mainly contains (A) glycidyl monovinyl ester or glycidyl monovinyl ether and (ω) alkylene glycol divinyl ester and is crosslinked with the component (A). 1. The glue-like copolymer according to the present invention is characterized in that it consists of a porous copolymer containing biotin covalently bonded to an epoxy group based on the component b) via a bonding group. This invention relates to an adsorption carrier for chromatography.

The adsorption carrier for chromatography of the present invention will be explained below.

The glue-like copolymer according to the present invention is disclosed in Japanese Patent Application Laid-Open No. 60-1042.
As described in Publication No. 56, for example, (A) glycidyl monovinyl ester or glycidyl monovinyl divinyl ester) is aqueous suspension polymerized in the presence of a water-insoluble organic diluent to form a porous material. It can be prepared by obtaining a sex guru.

Since the components (g) and 03) are sparingly soluble in water, they can be copolymerized by an aqueous suspension polymerization method, and the aqueous suspension polymerization can be carried out by a simple oil-in-water suspension method. The copolymerization is carried out in the presence of an organic diluent, and due to the presence of this organic diluent, the alkylene glycol divinyl ester component of Φ) acts as the main crosslinking component,
Most of the O epoxy groups in the glycidyl monovinyl ester or glycidyl monovinyl ether component of (A) remain in the produced copolymer as they are without ring opening, and are also useful for adjusting the pore size of the produced porous glue.

As the above component, glycidyl ester of monovinyl carboxylic acid having 3 to 12 carbon atoms or glycidyl ether of monovinyl alcohol having 3 to 12 carbon atoms is used. Among these, those having a small number of carbon atoms are particularly preferably used, and examples thereof include methacrylic acid glycidyl ester, acrylic acid f IJ cidyl ester, allyl glycidyl ether, and the like.

On the other hand, as the component (B) which acts as a crosslinking component, an alkylene glycol having 2 or 3 carbon atoms or an ester of the polyalkylene glycol with acrylic acid or methacrylic acid is preferably used. Examples include alkylene glycol divinyl esters such as ethylene glycol noacrylate, ethylene glycol dimethacrylate, zolopylene glycol diacrylate, propylene glycol dimethacrylate, and polynalkylene glycol divinyl esters such as hypopolyethylene glycol no methacrylate and polyglobylene glycol dimethacrylate. can be mentioned.

The ratio of component (A) to component (B) is 10 to 90 mol of the former to 90 to 10 mol of the latter. Preferably, the amount of component (g) is 40 to 80 mol, and the amount of component ω) is 60 to 20 mol. Up to half of component (g) can be replaced by a comonomer, ie, a lower vinyl ester such as methyl methacrylate, methyl acrylate, or vinyl acetate. Of course, as the proportion of component (a) increases, the content of epoxy groups in the copolymer increases.

The greater the proportion of component (B), the greater the degree of crosslinking of the copolymer, and therefore the denser the network structure, the lower the degree of swelling, and the harder the copolymer becomes. If the ratio of both components is outside the above range, it will not be possible to obtain a glue-like copolymer that fully satisfies the desired characteristics and properties.

The organic diluent used may be one that is inert to the polymerization reaction, insoluble or sparingly soluble in water, but capable of dissolving the raw material monomer. The amount used is at least 30 by volume based on the total amount of monomer components and organic diluent, preferably 40 to 8
0 capacity. The larger the amount used, the more suppressed is the ring opening of the epoxy group during the polymerization process. Generally speaking, about 60 to 95% of the epoxy groups contained in the amount of the raw material used can easily remain in the resulting polymer. The remainder is hydrolyzed during the polymerization process, and a portion of it is thought to be used for crosslinking.

As mentioned above, the content of epoxy groups in the produced glue can be adjusted by the amount of component (A) used, i.e. the ratio to component (B), or by partial substitution with comonomers, and also by the dose of organic diluent. Preferably, the epoxy oxygen is present in the resulting polymer in an amount of 1% by weight or more, expressed as epoxy oxygen, up to about 10% by weight.

As mentioned above, the use of organic diluents during aqueous suspension polymerization also helps control the pore size of the resulting porous glue. If an organic diluent having a low swelling property is used for the produced glue, phase separation will occur in the suspended particles during the polymerization process, and as a result, a glue with a large pore size and a so-called giant pore structure can be obtained. On the other hand, if a highly swelling diluent is used, the polymerization will proceed in a swollen state during the polymerization process.
Phase separation is difficult to occur and a polymer with relatively small pores formed by swelling is obtained. The degree of swelling can be determined based on the solubility/grammeter of the organic diluent. Examples of organic diluents that can be suitably used in the present invention include cyclohexanone, chlorobenzene, benzene, toluene, n-propyl acetate, n-butyl acetate, n-butyl acetate, and n-propyl acetate.
-Octane etc.

The aqueous suspension polymerization can be carried out in the presence of a free-radical generating catalyst according to conventional methods known per se.

The amount of the aqueous phase is approximately the same volume or more based on the size of the organic phase, and there are no particular restrictions, but an amount up to about 10 times the volume may be used.

Introduction of the bonding group into the glue-like copolymer according to the present invention can be carried out by utilizing the reactivity of the epoxy group in the binder component of the glue-like copolymer according to the present invention. In this case, it is necessary that the binding group has a functional group capable of covalently bonding to biotin or aminoalkylbiotin, but there are no particular restrictions on the structure of the binding group other than having the above-mentioned functional group. There isn't. Examples of the functional group capable of covalently bonding with the piotin or aminoalkylbiotin include an epoxy group, an amine group, a hydrazino group, a carboxyl group, a formyl group, and a thiol group. In this case, the bonding group can be easily introduced using a known method, and it is not necessarily necessary to introduce it in one reaction step;
It can also be introduced separately into more than one reaction step. A specific example of introducing a bonding group will be described below.

(i) Introduction of a bonding group having an epoxy group as a functional group, for example, a hydroxyl group produced by ring-opening modification of the epoxy group based on the carrier component of the dull copolymer according to the present invention with a hydroxyl group-containing compound such as water. A compound having a glycidyl group is bonded to the bonding group to introduce a bonding group having an epoxy group based on the glycidyl group. (Hereinafter, the fc polymer into which glycidyl groups are introduced in this way will be referred to as the modified glue-like copolymer according to the present invention.) Ring-opening modification is performed in the presence of an acid or an alkali,
For example, when carrying out in the presence of an acid, these copolymers are heated at 70°C to 10°C in water using sulfuric acid or hydrochloric acid as a catalyst.
This is done by heating to 0° C. for about 2 hours. The acid concentration is preferably 0.1N to 0.5N, and the A0 ring-opening modification is performed almost quantitatively.

Compounds having a glycidyl group that are bonded to the hyproxyl group produced by ring-opening modification include shrimp chlorohydrin, shrimp bromohydrin, shrimp halohydrins from Nato, ethylene glycol diglycidyl ether,
Diglycidyl ethers such as 1,4-butanediol sogelicidyl ether and polyethylene glycol diglycidyl ether, 1,3-butadiene diepoxide,
Examples include alkylsoene diepoxides such as 1,7-octazidine diepoxide.

To make a compound with a glycidyl group act.

For example, a ring-opening modified copolymer is reacted with shrimp chlorohydrin in an aqueous solution of an inorganic base such as sodium hydroxide or potassium carbonate, or a hydrophilic copolymer is reacted with water containing sodium borohydride. This is carried out by reacting with ethylene glycol, 1,4-butanediol sogelicidyl ether, or polyethylene glycol diglycidyl ether in an aqueous solution of an inorganic base such as sodium oxide or potassium carbonate. Repeating unit of polyethylene glycol diglycidyl ether
It is preferable to use one having a repeating number of C2H40- of 1 to 10.

(10) Introduction of a bonding group having an amino group, hydrazino group, carboxyl group, etc. as a functional group For example, as a method for introducing a bonding group having an amine group, hydrazino group, carboxyl group, etc., the r-shaped copolymer according to the present invention or an epoxy group based on the glycidyl group of the modified R-shaped copolymer according to the present invention, an epoxy group based on the component (to) of the general formula (1) (wherein R' is a hydrogen atom, an amino group, an ω- (representing an aminoalkyl group or an ω-carboxyalkyl group), or when an amine compound other than the ω-carboxyalkyl group is reacted with the amine compound, further alkylene y Cargonic acid anhydride is applied to the epoxy group or glycidyl group to form a compound of the general formula 01) NHR (wherein R is a hydrogen atom, an amino group, an ω-aminoalkyl group, ω-car is a xyalkyl group, ω-carboxy Alkanoyl group, ω-cargoxyalkanoylamino group or ω
- represents a carboxyalkanoylaminoalkyl group.

) There is a method of introducing a bonding group represented by

In this case, the reaction can be carried out according to the following reaction formula.

It represents one epoxy group or glycidyl group in the r-shaped copolymer according to the present invention or the rira copolymer according to the present invention. In the formula, m is l, 2, or n.

p=q and r are each 1 or more (preferably 1 to 1
2) is an integer.

A compound of the reaction formula (D), general formula (D') or general formula (F) is added to the epoxy group of the dull copolymer according to the present invention or the glycidyl group of the modified dull copolymer according to the present invention. The reaction can be carried out in the presence of a solvent, but if these compounds are liquid under the reaction conditions, a solvent may not be used. As a solvent, water is usually used, but other hydrocarbons such as n-hexane, benzene, and xylene nato, ethers such as tetrahydrofuran and dioxane, chloroform, chloropenzene natonochlorinated hydrocarbons, ethanol, and methyl selon rub are also used. In addition to alcohols, ketones such as acetone and methyl imbutyl ketone, dimethyl formamide and dimethyl sulfoxide are also used, and these solvents may be used alone or in a mixed solvent system.

Although no particular catalyst may be used, hydroxides or carbonates of alkali or alkaline earth metals such as sodium hydroxide and potassium carbonate may also be used.

Examples of the compound of general formula (D') include ethylenediamine,
■, 3-diaminopropane, 1,4-diaminobutane,
1,5-diaminopentane, 1,6-diamithexane, 1,7-diaminohbutane, 1,8-diaminooctane, 1,9-diaminononane, 1.10-diaminodecane, 1,12-diaminododecane, etc. diaminoalkanes.

Compounds of general formula (F) include glycine, β-alanine, 4-aminocyic acid, 6-aminocaproic acid, 8-aminocaproic acid,
-Aminocaproic acids such as amitsukazolylic acid can be exemplified.

Although there are no particular restrictions on the reaction conditions, it is generally preferable to select the following conditions to carry out the reaction.

Delicious.

The weight (,) of the dull copolymer according to the present invention or the modified dull copolymer according to the present invention and the weight (b) of the compound of formula (D), general formula (D') or general formula (F) Ratio: a:
b = 1: 0.3 to 10, preferably a: b = 1: 0.3 to 3 Reaction temperature: 0 to 150°C, more preferably room temperature to 100°C
C, reaction time = 1 to 60 hours, more preferably 1 to 30 hours, reaction pressure normal pressure to 10 atm, more preferably normal pressure.

There are no special requirements for post-treatment after the reaction, and conventional methods such as F separation and washing may be carried out as appropriate.

The chromatography carrier according to the present invention obtained above can be prepared by using a compound of formula (D) or general formula (D), and then, if necessary, as already shown in the general formula, the terminal of the active supporting group is The amino group of can be converted into an active supporting group having a carboxyl group by reacting with the acid anhydride of the general formula (E).Water is usually used as a solvent for the reaction, but other ethers such as tetrahydrofuran and dioxane can be used. carboxylic acids such as acetic acid, PIJ
Shin etc. are also used. Further, although a catalyst may not be used, the -ness of the reaction solution can be adjusted by adding an acid such as hydrochloric acid or sulfuric acid or an alkali such as sodium hydroxide or potassium carbonate.

Examples of the alkylenecarboxylic anhydride of general formula (E) or general formula (E') in the reaction formula include ni-phosphoric anhydride, geltaric anhydride, adipic anhydride, pimeline? Examples include acid anhydride, speric anhydride, azelaic anhydride, and cepatic anhydride.

The reaction conditions are not necessarily limited, but it is generally preferable to select the following conditions to carry out the reaction.

The weight (a') of the aminated product of the glue-like copolymer according to the present invention or the modified glue-like copolymer according to the present invention and the general formula (E)
Or the ratio of the weight (b') of the compound of general formula (E'): a':b'; 1:0.3-10, more preferably a':b'=1:0.3-3 Reaction temperature :0~150℃, more preferably room temperature~100℃
C, reaction time = 1 to 60 hours, preferably 1 to 30 hours, reaction pressure normal pressure to 10 atm, preferably normal pressure.

There are no special requirements for post-treatment after the reaction, and it can be carried out as appropriate by commonly used methods such as door-to-door cleaning and washing.

Another method for introducing a bonding group having a carboxyl group as a functional group is to introduce an aminocarboxylic acid having a hydroxyl group into an epoxy group based on the component (A) of the glue-like copolymer according to the present invention or a modified method according to the present invention. There is a method of binding by acting on the glycidyl groups of a glycidyl copolymer.

Examples of aminocarboxylic acids having a hydroxyl group include amino acid derivatives such as serine, homoserine, threonine, 4-hydroxyproline, 4-amino-3-hydroxy dilute acid, N-tris (hydroxymethyl), and methylglycine, and glucosaminic acid. Examples include amino sugar derivatives of.

The bonding state is such that the epoxy group of the glue-like copolymer according to the present invention or the glycidyl group of the modified glue-like copolymer according to the present invention is bonded to the amino group of the aminocarboxylic acid having a hydroxyl group. The glycidyl group opens to form a hydroxyl group and a carboxyl group having a hydroxyl group.

The action of the aminocarboxylic acid having a hydroxyl group can be carried out in the presence of a solvent.

As a solvent, water is usually used, but other hydrocarbons such as n-hexane, benzene, and xylenato, ethers such as tetrahydrofuran and dioxane, chloroform, chloropenzene, and chlorinated hydrocarbons, ethanol, and methylcerone are also used. In addition to alcohols such as , ketones such as acetone and methyl imbutyl ketone, dimethyl formamide and dimethyl sulfoxide are also used, and these solvents may be used alone or in a mixed solvent system.

Although no particular catalyst may be used, hydroxides or carbonates of alkali or alkaline earth metals such as sodium hydroxide and potassium carbonate may also be used.

There are no particular restrictions on the reaction conditions, and for example, the same conditions as in the case of reacting the compound of general formula (1) above can be used.

When a bonding group is introduced using an aminocarboxylic acid having such a hydroxyl group, nonspecific adsorption originating from hydrophobicity based on a portion other than the base material, that is, a portion of the bonding group can also be eliminated.

<r1+>Introduction of a bonding group having a formyl group as a functional group.For example, the bonding group having an epoxy group as a functional group obtained in the above (i) can be easily hydrolyzed to form 1
,2-glycol, and this 1,2-
Glycol can be converted into formyl group by the action of periodic acid.

There are no particular restrictions on the reaction conditions, and the reaction can be carried out according to a conventional method using, for example, water, ethanol, acetic acid, or a mixture thereof as a solvent at room temperature to 50° C. by reacting periodic acid or an inorganic salt thereof. .

(1■) Introduction of a bonding group having a thiol group as a functional group For example, the bonding group having an epoxy group as well as the functional group obtained in the above (1) can be prepared by applying sodium thiosulfate to the functional group, and then A thiol group can be introduced by applying hydrochloric acid or the like.

There are no particular restrictions on the reaction conditions, and the reaction can be easily carried out according to a conventional method, for example, at room temperature in an aqueous solvent.

When covalently bonding biotin or aminoalkylbiotin to a binding group, it can be carried out in an appropriate solvent using an appropriate catalyst or reaction reagent, if necessary depending on the functional group possessed by the binding group.

For example, as a catalyst, acids such as hydrochloric acid, sodium carbonate, or sodium bicarbonate, and alkalis are mainly used when the functional group is an epoxy group; When the functional group of carbodiimide is a carboxyl group, and when the condensing agent such as 1-ethyl-3-(3-dimethylaminopropyl)cardediimide has a carboxyl group, an amine group, or a hydrazino group as a functional group, For example, a reducing agent such as sodium cyanoborohydride is used when the functional group is a formyl group. Water is usually used as a solvent, but if necessary, it can also be used as a phosphoric acid or acetate buffer, or an inorganic salt such as sodium chloride can be added thereto.

The conditions for the reaction with biotin or aminoalkyl piotin are not necessarily limited, but it is generally appropriate to carry out the reaction within the following range.

Weight ratio of the R-shaped copolymer or modified R-shaped copolymer according to the present invention bound with a binding group to biotin or aminoalkyl piotin: 1:0.01 to 0.3, preferably 1:0 .05~0.
2; Reaction temperature: 0°C to room temperature, preferably 4°C to room temperature; Reaction time = 1 to 72 hours, preferably 2 to 12 hours.

There are no special requirements for post-treatment after the reaction, and it can be carried out as appropriate by commonly used methods such as door-to-door cleaning and washing.

Effects of the Invention The adsorption carrier for chromatography provided by the present invention uses a base having all of the properties desired as an adsorption carrier for 7-finity chromatography as described above, and binds it to the base through an appropriate bonding group. Since biotin is commonly bonded, the adsorption carrier of the present invention can not only be used as a packing material for affinity chromatography in the same way as adsorption carriers using conventional soft glue, but also particularly suitable for pressure-resistant columns. The biggest advantage is that it can be filled with water and used under pressure, which greatly reduces working time and therefore greatly improves the efficiency of separation and purification. Needless to say, this is essential for applications such as high performance liquid chromatography and industrial separation and purification equipment.

In particular, while the shell copolymer has many advantages as described above, the adsorption carrier to which biotin is covalently bonded via the bonding group as described above, for example, can absorb the commonly used cyanogen bromide. Compared to adsorption carriers for affinity chromatography, etc., the ligand does not fall off during use, and the length of the bonding group can be adjusted arbitrarily, so the adsorption ability of the ligand for the target substance is excellent. In addition, the adsorption carrier for chromatography provided by the present invention has great effects, including less nonspecific adsorption.

EXAMPLES Below, typical examples of the method for producing the adsorption carrier for chromatography of the present invention from known substances will be shown and explained in more detail. However, these are merely examples for explanation, and it goes without saying that the present invention is not limited thereto.

Example 1 A copolymer obtained from glycidyl methacrylate and ethylene glycol dimethacrylate was ring-opened and modified with water, and a glycidyl group was introduced with 1,4-butanediol diglycidyl ether, and then an epoxy group was modified with aqueous ammonia. To 1.09 ml (amino group: υ 0.2 mmol per 1 g of dry hydroxyl) converted into a group, 113 rnl of dimethyl sulfoxide of 243 m9 of biotin and 206 ml of dicyclocargodiimide were added, and the mixture was allowed to react at room temperature for 2 hours. By washing the colander with methanol and water, an adsorption carrier supporting 10 m9 of sybiotin corresponding to dried coll IJ was obtained from the sulfur content determined by elemental analysis.

Example 2 A copolymer obtained from glycidyl methacrylate and ethylene glycol dimethacrylate was prepared by converting the jepoxy group into an amine group using 1,3-propanthamine (amino group: dried dal 1!i). mmol) 1.0
9 and 1- in place of dicyclohexylcarbodiimide in Example 1.

Using 300ml of ethyl-3-(3-dimethylaminopropyl)carbodiimide and replacing the solvent with dimethyl sulfoxide solution, 0.1 molar sodium phosphate buffer pH 6
.

Example 3 Biotin 243m9 and N-hydroxysuccinimide 1
15 mL of dicyclohexylcarbodiimide was added in 5 ml of DMF and reacted with stirring at 10° C. for 2 hours.

After the reaction is completed, 1 g of the dal used in Example 1 is added to the reaction mixture, and the mixture is further shaken and reacted at room temperature for 2 hours. By washing the dal with methanol and water, an adsorption carrier carrying 12η of cybiotin per 1 g of dry mulle was obtained.

Example 4 Biotin 2.439 and 1,3-propanthamine 4I are reacted with dicyclohexylcarbodiimide 2.06.F in 2(1/1) dimethylformamide with stirring at 1.->10 DEG C. for 2 hours. After the reaction was completed, 2Qml of diethyl ether was added to the reaction solution to reduce the resulting dicyclohexylurea to 7ml.
After evaporation, liquid F was distilled off under reduced pressure, and the residue was recrystallized from ethanol to obtain 2.19 of the corresponding aminopropyl biotin.

Example 5 A copolymer obtained from glycidyl methacrylate and ethylene glycol dimethacrylate was open-term modified with water, and a glycidyl group was introduced with epichlorohydrin to prepare 1 g of Dal (epoxy group: 90.25 mmol per 1 g of dry Glu). Aminopropyl biotin 2 synthesized in Example 4
00 mg in 2 molar sodium carbonate solution 3d at 50°C.
Shake for 6 hours. After the reaction was completed, the dal was washed with water and methanol to obtain an adsorption carrier supporting 8 m9 of biotin per 1 g of dry dal.

Example 6 The same procedure as in Example 1 was carried out using the aminopropyl biotin prepared above, to obtain an adsorption carrier on which 10 to 100 biotin was supported per 1 g of dry glue.

Test Example 1 When the adsorbent obtained in Example 1 was packed into a 4.6φ x 75mm stainless steel column and avidin was analyzed using a high performance liquid chromatography device, a romatograph as shown in Figure 1 was obtained. It was done.

Separation conditions Eluent: 0.01M phosphate buffer (pH 7.0) (eluent ■), 0.01M phosphate buffer 0.1M Na
(:z (pH7,0) (eluent■), eluent■→■3
°min linear gradient, flow rate: Q, 5 ml/m
in, column temperature = 28°C, detector: ultraviolet spectrophotometer 2
30 m 0 Test Example 2 When egg white was analyzed in the same manner as in Test Example 1, Lomatodalum was obtained as shown in Figure 2.

[Brief explanation of the drawing]

12 and 2 are diagrams showing the separation adsorption and elution patterns of avidin or egg white using a high performance liquid chromatography column using the biotin adsorption carrier according to the present invention.

Claims (1)

[Claims] Epoxy based on component (A) of a gel-like copolymer containing (A) glycidyl monovinyl ester or glycidyl monovinyl ether and (B) alkylene glycol divinyl ester as main components, and component (A) crosslinked with component (B). 1. An adsorption carrier for chromatography comprising a porous copolymer containing biotin covalently bonded to a base via a bonding group.