JPH01217035A - Production of crosslinked polymer particle - Google Patents

Abstract

PURPOSE:To obtain the subject particles, excellent in pressure resistance and hydrophilicity and suitable as a liquid chromatographic packing, by carrying out suspension polymerization of a glycidyl monovinyl compound with a polyvinyl compound, hydrolyzing the resultant polymer and then subjecting the hydrolyzed polymer to graft polymerization treatment. CONSTITUTION:The aimed particles, obtained by dispersing (A) glycidyl monovinyl ester or ether with (B) a polyvinyl ester or ether of a polyhydric alcohol in an aqueous medium containing water mixed with an organic solvent, adding a polymerization initiator and carrying out suspension polymerization, hydrolyzing the resultant polymer in water containing, e.g., sulfuric acid, reacting the obtained particles with (C) an alkyl halide having oxirane ring, further adding and reacting (D) a compound (e.g., glycidol) having oxirane ring and >=2g/100ml solubility in water and <=1000 molecular weight with the resultant particles and having preferably 1-50mum particle diameter.

Description

[Detailed description of the invention] (Industrial application field) The present invention is a packing material for liquid chromatography.

In particular, the present invention relates to a method for producing porous crosslinked polymer particles useful as fillers for aqueous liquid chromatography or intermediates for their synthesis.

(Prior art) Conventionally, as a packing material for liquid chromatography, (1
) Copolymer gel of styrene and crosslinkable single compact [CG Moore, Journal Ope Polymer Science (J, C, Moore+ J, Polym.

Sci, ), A2, 835 (1964)], ■ Copolymer gel of polyacrylic ester of acrylic ester monomer and crosslinkable monomer polyalkylene glycol (Japanese Unexamined Patent Publication No. 55-99068), ■Gel made by crosslinking polyvinyl alcohol with epichlorohydrin (Unexamined Japanese Patent Publication No. 52-1
Publication No. 38077).

■Polyvinyl alcohol gel obtained by hydrolyzing a copolymer gel of vinyl acetate and triallyl isocyanurate (Japanese Patent Application Laid-Open No. 57-30945), ■Acrylic acid ester monomer or acrylamide and crosslinkable monomer methylenebisacrylamide copolymer gel (Suzuki Teru et al. 1 Journal of the Chemical Society of Japan, 1975゜1756), ■Gel made by crosslinking dextran with epichlorohydrin (Japanese Patent Publication No. 1983-21405), ■Chemical modification of inorganic gels such as silica gel and glass. What I did [Nis H Chan, Earl Nonil,
F.E. Regnir, Analytical Chemistry (S, a Chang * R, Noel mF'
,E,Regniers Anal,Chem,
) 48.1839 (1976)) are known.

(Problem to be Solved by the Invention) The above gel can be an excellent packing material in liquid chromatography using an organic solvent as an eluent, but it cannot be used in an aqueous system because it is not compatible with water. ■ above,
The gels (2) and (2) remain slightly hydrophobic, so if the substance to be separated is hydrophobic, hydrophobic adsorption will occur, which is inconvenient when performing single molecule sieving separation.

The gels of ■ and ■ above are not hydrophobic.

It has poor mechanical strength and is used as a column filler for high-performance liquid chromatography where high pressure is applied (it cannot be damaged.Also, the gel in (■) is not hydrophobic and has 9 mechanical strength, but is weak against alkalis and has poor durability. It had the disadvantage of being inferior to

Therefore, the present invention solves the drawbacks of the prior art and provides a material that has hydrophilicity, pressure resistance, and alkali resistance and is useful as a packing material for liquid chromatography or an intermediate for synthesizing a packing material for liquid chromatography. The object is to provide porous crosslinked polymer particles.

(Means for Solving Problem 1) The present invention provides glycidyl monovinyl ester or glycidyl monovinyl ether (CI), a polyvinyl ester of a polyhydric alcohol, or a polyvinyl ether of a polyhydric alcohol (II) in an organic solvent that is not miscible with water. Aqueous suspension polymerization in the presence of (1) and further K.

The oxirane ring of the obtained particle (A) due to component (I) is hydrolyzed, and further, the obtained particle (B) is reacted with an alkyl halide having a K oxirane ring to introduce an oxirane ring. The present invention relates to a method for producing crosslinked polymer particles, which comprises graft-polymerizing a compound having an oxirane ring onto the oxirane ring of the particle (C).

Next, each component used in the method for producing crosslinked polymer particles of the present invention will be described in detail.

Examples of the glycidyl monovinyl ester or glycidyl monovinyl ester used in the present invention include glycidyl methacrylate, glycidyl acrylate, and allyl glycidyl ether.

Considering reactivity and alkali resistance, glycidyl methacrylate is preferred.

Examples of polyvinyl esters of polyhydric alcohols and polyvinyl ethers of polyhydric alcohols used in the present invention include ethylene glycol dimethacrylate, ethylene glycol diacrylate, and propylene glycol dimethacrylate.

Alkylene glycol divinyl esters such as propylene glycol diacrylate, polyethylene glycol dimethacrylate, polypropylene glycol dimethacrylate, polyethylene glycol diacrylate,
Divinyl esters of polyalkylene glycols such as polypropylene glycol diacrylate, di- or triacrylates of glycerin, di- or trimethacrylates of glycerin, trimethylolpropane di- or trimethacrylates, trimethylolpropane di- or triacrylates, tetramethylolmethane di-, tri- or Tetramethacrylate, tetramethylolmethanedi, tri- or tetraacrylate, ethylene glycol diallyl ether,
Examples include fluoropylene gelyl diallyl ether, polyethylene glycol diallyl ether, polypropylene glycol diallyl ether, glycerin di- or triallyl ether, trimethylol pro-kundi or triallyl ether, tetramethylolmethane di, tri- or tetraallyl ether, etc. 1 reactivity and the hydrophilicity and pressure resistance of the produced porous crosslinked polymer particles, ethylene glycol dimethacrylate, tetramethylol meta/di.

Tri- or tetramethacrylates are preferred.

Green glycol monovinyl ester or green glycol monovinyl ether (IL monovalent acyal alcohol ribinyl ester or polyhydric alcohol monovinyl ether
) As the ratio of (■) increases, the hydrophilicity of the generated gel particles improves, but the pressure resistance tends to decrease.

Therefore, the ratio of (I)/(n) is 5015 in weight ratio.
It is preferably in the range of 0 to 97/3.

The organic solvent that is not miscible with water is used in the present invention.

These particles are used to make the particles porous and have a solubility of less than 15 g in 100 g of water at 25° C.1 For example, toluene, diethylbenzene, dodecane, isoamyl alcohol, chlorobenzene, ethyl acetate, propyl acetate. , butyl acetate, etc. These solvents may be used alone or in combination. The solvent is preferably used in an amount of 50 to 300% by weight based on the total amount of monomers. If there are too few organic solvents that are not compatible with water.

Too much will make it difficult to make the resulting particles porous.

The resulting particles have a large porosity and poor pressure resistance.

In the present invention, glycidyl monovinyl ester or glycidyl monovinyl ether (I),! = I-valent acyal alcohol ribinyl ester or polyvinyl ether of polyhydric alcohol (II) is suspension polymerized in an aqueous medium in the presence of an organic solvent that is not miscible with water.

Water is used as the aqueous medium, but an aqueous organic solvent (e.g. methanol,
ethanol, acetone, etc.) or inorganic salts (e.g. chloride)
Water containing dissolved IJum, sodium sulfate, etc.) may also be used. Note that the aqueous medium is the above (I), (n
) and 1 to 59 for in of organic solvents that are incompatible with water.
It is preferable to use twice the amount by weight. At this time, it is preferable to add all the organic solvents that are not compatible with water before the 9 reactions or until the polymerization rate reaches 20%. The addition method may be divided into one portion.

Suspension polymerization initiators suitable for use in the process of the invention include benzoyl peroxide, dichlorobenzoyl peroxide, dicumyl peroxide, di-tert-butyl peroxide,
．． 5-di(peroxybenzoate)hexyne-3,1
, 3-bis-(tert-butylperoxyisopropyl)benzene, lauroyl peroxide, tert-butyl peracetate, s5
-dimethylose 5-di(tert-butylperoxy)hexane-3゜2.5-dimethyl-2,5-di(tert-butylperoxy)hexane and tert-butyl perbenzoate, methyl ethyl ketone peroxide, methyl cyclohexano/beroyaside, etc. There are organic peroxides, azo compounds such as azobis-isobutyronitrile, and dimethylazodiisobutyrate, and one or more of these can be used.

The amount used is determined depending on the type of vinyl monomer and the desired molecular weight of the resulting polymer, but it is preferably used in an amount of 0.1 to 4.0% by weight based on the vinyl monomer. Ru.

In addition, in the suspension polymerization mentioned above, sparingly soluble phosphates, water-soluble polymeric protective colloids, etc. can be added to the polymerization system as dispersants.

Examples of a-soluble phosphates include tricalcium phosphate and magtosium phosphate. Polymer protective colloids include polyvinyl alcohol, water-soluble cellulose derivatives such as alkyl cellulose, hydroxyalkyl cellulose, and carboquin alkyl cellulose, and polyac II A/acid).
There are IJum etc. Slightly soluble phosphates are used within the range of 0 to 1'ffr relative to the total amount of substances present in the bipolymerization system, and water-soluble polymeric protective colloids are used within the range of 10 to 1'ffr. preferable.

When performing aqueous a-water polymerization, each component except the aqueous medium can be mixed in advance or each component can be added separately to the aqueous medium and dispersed. In this case, in order to disperse well, it is preferable to perform high-speed stirring using a homomixer or the like, and this high-speed stirring can be carried out up to the initial stage of 9-polymerization. The subsequent polymerization can be carried out under ordinary stirring using a propeller stirrer or the like.

Hydrolysis of the oxirane ring of particles (A) is carried out by dissolving the copolymer having an oxirane ring in water containing H acid or phosphoric acid from 30 to 1
Approximately 80% of the oxirane ring can be hydrolyzed by treatment at 00°C for 5 to 20 hours.

In order to introduce an oxirane ring, the alkyl halide having an oxirane ring to be reacted with the particles (B) obtained by aqueous suspension polymerization and subsequent hydrolysis is shrimp chlorohydrin/l shrimp bromohydrin/.

Examples include shrimp fluorohydrin and shrimp iodohydrin. These compounds are sodium hydroxide.

Dimethyl sulfoxide, N,N-dimethylformamide in the presence of a hydroxide such as potassium hydroxide.

Particles (
B) can be swollen and reacted.

The alkyl halide having an oxirane ring is preferably used in an amount of 10''6 mol or more per gram of well-dried porous crosslinked polymer particles.

If less than 104 moles per gram are used.

The concentration of the oxirane ring (reaction starting point for the next graft polymerization) imparted to the particles (B) is low, and the effect of improving fresh water properties tends to be reduced.

The amount of hydroxide such as IJium or potassium hydroxide may be at least equimolar to the alkyl halide having an oxirane ring, and dimethyl sulfoxide, N, N, etc.
- Polar solvents such as dimethylformamide, N,N-dimethylacetamide, etc. should be used at a concentration of 1 to 90% to stir the reaction system.
1 is sufficient, and there is no particular restriction.

The reaction is preferably carried out at a temperature in the range of 30-100°C for 1 hour or more.

Particles having oxirane rings (C
) is 0.1 to 2 me of oxirane group per 1 g of core particles.
It is preferable to adjust it so that it has q. If this value is too small, it will cause problems in the first food.

If the oxirane/ring-containing compound cannot be grafted sufficiently, and if the amount is too large, it will be difficult to introduce the oxirane ring.

In addition, particles obtained by aqueous suspension polymerization and hydrolysis (B
The point of reaction with the alkyl halide having an oxira/ring in ) is the hydroxyl group present in the particle (B).

The particles (C) into which the oxirane ring has been introduced are subjected to graft polymerization. The compound having an oxirane ring must have sufficient hydrophilicity, and it is desirable that the compound contains a large amount of hydrophilic groups, that is, hydroxyl groups and ether groups.
Mata. It is preferable to have a certain number or more of oxiras/rings so that the pore surfaces are covered with hydrophilic side chains through a copolymerization reaction.

Further, if the molecular weight of the compound having an oxirane ring is too large, the pore volume in the finally obtained porous crosslinked polymer particles is decreased, which is not preferable.

For this reason, 9 are used in the present invention. As a compound with an oxirane ring, the solubility in water is 29/100ml! water(
25°C) or more and 1,000 or less, and an epoxy group equivalent (molecular weight/number of oxirane rings) of 300 or less. For example, ethylene oxide, glycidol,
1.2.34-Jepoxybutane, diglycidyl ether, ethylene glycol (mono- or di)glycidyl ether, glycerin polyglycidyl ether, sorbitol polyglycidyl ether, diglycerol polyglycidyl ether, polyethylene glycol (7- or di)glycidyl ether ( (polymerization degree of polyethylene glycol 9 or less), polypropylene glycol (mono- or di)glycidyl ether (polymerization degree of polypropylene glycol 3 or less), 1,4-butanediol (
Mono- or di)glycidyl ether, 1.3-7'
Examples include 0-panediol (7-no or di)glycidyl ether.

The graft density of these compounds is preferably adjusted to be 0.05 to IgK relative to the particle (C)ig into which the oxirane ring is introduced. If the amount of grafting is too large, the degree of swelling of the final particles will increase, and when the particles are packed in a column for analysis, the column pressure must be increased, and the particles are likely to be destroyed. Become. If the amount of grafting is too small, the effect of imparting hydrophilicity by grafting the compound having an oxirane ring will tend to be reduced.

Sulfuric acid is used as a catalyst for graft polymerization.

Protonic acids such as hydrochloric acid, perchloric acid, and phosphoric acid; Lewis acids such as boron trifluoride, boron trifluoride etherate, and zinc boron tetrafluoride; and stannous chloride, tin chloride, titanium tetrachloride, aluminum chloride, etc. Among them, boron trifluoride etherate is particularly preferred. In addition, the amount of catalyst used is 0.01 to 3 for the compound having an oxirane ring.
Preferably it is 0% by weight.

When carrying out graft polymerization, the presence of a solvent is not essential (from the viewpoint of surface uniformity and operability).

It is usually advantageous to carry out the reaction in the presence of a solvent.

The solvent may be inert to the 9 particles (C) or the compound having an oxirane ring, and may include aliphatic or alicyclic ethers, ketones, esters, hydrocarbons, halogenated hydrocarbons, Nitriles.

Examples include sulfokides and the like. Particularly preferred solvents include ethers such as ethyl ether, diethylene glycol dimethyl ether, ethylene glycol dibutyl ether, tetrahydrofuran, 1°4-dioxane, 1.2-dichloroethane, 1.4-dibromoethane, 1.2-dichloroethane, etc. 'jc1 Hough, 1.1.
Examples include halogenated hydrocarbons such as 2-trichloroethane, sulfoxides such as dimethyl sulfoxide, and the like.

Some graft polymerizations have a fast reaction rate even at low temperatures, but in general, the reaction rate slows down at low temperatures, making them impractical; on the other hand, at high temperatures, the various optimal reaction temperatures are Although the temperature is selected depending on the type of compound, the type of solvent, the type of catalyst, etc., the temperature is usually in the range of 0 to 150°C.

The spherical particles obtained in the above manner had 9 particle sizes of 1 to 50.
Classified into μm, preferably 1 to 15 μm.

It can be used as a packing material for gel permini-7 chromatography or partition adsorption chromatography. Furthermore, by introducing ion exchange groups, glycidyl groups, long chain alkyl groups, etc., the properties as fillers can be changed.
It can also be used as a carrier for affinity chromatography and ion exchange chromatography.

Glycidyl monovinyl ester or glycidyl monovinyl ether (1) and polyvinyl ester of polyhydric alcohol or polyvinyl ether of polyhydric alcohol (II) are subjected to aqueous suspension polymerization in the presence of an organic solvent (1) that is not miscible with water, and further , (I)Ilii of the obtained particles (A)
When the oxirane ring based on the compound is hydrolyzed with water, the resulting particles (B) have both hydrophilicity and pressure resistance.

Also, by graft polymerizing a compound having an oxirane ring to particles (C) obtained by reacting 9 particles (B) with an alkyl halide.

Further, crosslinked polymer particles of the final product with increased hydrophilicity can be obtained.

(Example) Next, the present invention will be explained in detail based on examples.

The present invention is not limited to this.

Example 1 (1) Production of particles (A) and particles (B) using the same by aqueous suspension polymerization Ethylene glycol dimethacrylate) 409°Gly7
Zil methacrylate 1609. 75g n-butyl acetate,
A mixture of 125g of isoamyl alcohol and 1.0g of azobisisobutyronitrile was added to 1760ml of water! In addition to
200ml of 1% methylcellulose aqueous solution at room temperature with high speed stirring using a homomixer! was added sequentially to adjust the particle size.

The mixture was transferred to a 31-7 flask to which 220 mJ of 1-thimethylcellulose aqueous solution had been added in advance, and suspension polymerization was carried out at 85° C. for 6 hours.

The entire amount of the obtained particles (A) was reacted in 0.5N sulfuric acid at 80°C to hydrolyze the oxirane ring to obtain particles (B).

(2) Production of particles (C) by introducing ocilane rings (1)
25 g of particles (B) synthesized and classified into 8 to 15 μm,
Epichlorohydrin 17411s dimethyl sulfoxide 225 ml and 30 m sodium hydroxide aqueous solution 1
8g II! Prepared in a flask.

The reaction was carried out at 40°C for 6 hours. After the reaction, the particles were collected after 9 passes and washed successively with dimethyl sulfoxide, water and acetone to obtain particles (C) having an oxirane ring.

When the oxirane ring was quantified using the hydrochloric acid/calcium chloride method, 1. It was tmeq/g.

(3) Particles having oxirane rings synthesized by graft polymerization (2) (C) 50
g, 50 g of ethylene glycol diglycidyl ether (solubility in water Log/100 ml! water, 25°C), 5 ml of boron trifluoride ethyl ether complex and 500 ml of 1,4-dioxane! 11! The mixture was charged into a flask and reacted at 80°C for 5 hours to perform graft polymerization. After the reaction, the produced particles were collected by sieving and washed sequentially with 1,4-dioxane and acetone.

It was dried under reduced pressure at 60°C for 3 hours. The obtained crosslinked polymer particles were porous and the yield was sa, 7 g.

(4) Application The particles thus obtained were sieved to a particle size of 8 to 15 μm and packed into a stainless steel column with a diameter of 8.0 ms and a height of 300 mm. Proteins and amino acids were measured using this column. The measurement conditions were a mobile phase of 20mM phosphate buffer (pH 7) containing 0.5M sodium chloride, and a flow rate of 1.0ml! /min.

The temperature was 25°C, and the detector was a UV detector (280 nm
)It was used.

Proteins (thyroglobulin, T-globulin, bovine serum albumin, ovalbumin, α-chymotrypsinogen,
Cytochrome C, insulin.

The relationship between the molecular weight of β-alanine) and the elution volume (
The intersection curves are shown in Figure 1. Codes 1 to 8 are.

This is a plot of the substances in the order listed above.

Example 2 Particles (C) 509. having an oxirane ring synthesized in Example 1 (2). Glycidol sog, boron trifluoride ethyl ether complex 5ml! and L4-dioxa 7500 m
11 l! The mixture was charged into a flask and reacted at 80°C for 5 hours to perform graft polymerization. After reaction.

The produced particles were collected by filtration, sequentially washed with 1,4-dioxane and acetone, and dried under reduced pressure at 60° C. for 3 hours.

The resulting crosslinked polymer particles are porous.

The yield was 63.59.

The particles thus obtained were sieved to a particle size of 8-15 μm and packed into a stainless steel column with a diameter of 8.0 am+e and a height of 300#. Proteins and amino acids were measured using this column. The measurement conditions were: 20mM phosphate buffer (pH 7) containing 0.5M sodium chloride in the mobile phase;
The flow rate was 1.0 mj'/min.

The temperature was set to 25°C, and the detector was a FiUV detector (280
nm) was used.

A calibration curve showing the relationship between the molecular weight of the protein (same as in Example 1) and the elution volume is shown in FIG. Codes 9 to 16 are
Sequentially, the substances are plotted in the order listed above.

Comparative Example The particles (B) synthesized in Example 1 (1) were sieved to have a particle size of 8 to 15 μm, and the diameter & Off1b height was 3.
It was packed into a 00 mm stainless steel column. This column was used to measure white matter and amino acids.

The measurement conditions were: 20mM phosphate buffer (pH 7) containing 13 um (0.5M chloride in the mobile phase), and a flow rate of 1.0ml.
! /min, the temperature was 25℃, and the detector was equipped with a UV detector (2
80 nm> was used.

A calibration curve showing the relationship between the molecular weight of the protein (same as in Example 1) and the elution volume is shown in FIG. Reference numerals 17 to 24 are plots of substances in the order of +m.

(Effects of the Invention) According to the present invention, porous spherical particles with excellent pressure resistance and hydrophilicity can be obtained, and these porous crosslinked polymer particles can be used as a filler for liquid chromatography or as a filler for liquid chromatography. Suitable as an intermediate for filler synthesis.

[Brief explanation of the drawing]

FIG. 1 is a calibration curve measured in Example 1. FIG. 2 is a calibration curve measured in Example 2. FIG. 3 is a calibration curve measured in a comparative example. Explanation of symbols 1... Thyroglobulin 2... γ-globulin 3.
...Bovine serum albumin/4...Ovalbumin5...
・α-chymotrypsinogen 6...Cytochrome C7...Insulin 8...
β-alanine 9...Thyroglobulin 10...γ
-Globulin 11...Bovine serum albumin 12...
Ovalbumin 13...α-chymotrypsinogen 14...Cytochrome C15...Insulin 16
...β-Alanine 17... Thyroglobulin 18.
・・γ-globulin 19・・bovine serum albumin 20・
・・Ovalbumin 21・α-chymotrypsinogen 22・Cytochrome C23・Insulin 24
...β...Alanine elution volume 1 (■ Punishment 1 Figure 432 Volume Master Tsuno 2

Claims (1)

[Scope of Claims] 1. Glycidyl monovinyl ester or glycidyl monovinyl ether (I) and polyvinyl ester of polyhydric alcohol or polyvinyl ether of polyhydric alcohol (I)
I) is subjected to aqueous suspension polymerization in the presence of an organic solvent (III) that is not miscible with water, and further, (I) of the obtained particles (A) is
The oxirane ring caused by the component is hydrolyzed, and the obtained particles (B) are further reacted with an alkyl halide having an oxirane ring to introduce an oxirane ring, and the oxirane ring is added to the oxirane ring of the particle (C) having this oxirane ring. 1. A method for producing crosslinked polymer particles, the method comprising graft polymerizing a compound having the following. 2. Glycidyl monovinyl ester or glycidyl monovinyl ether (I) and polyvinyl ester of polyhydric alcohol or polyvinyl ether of polyhydric alcohol (I)
2. The method according to claim 1, wherein the weight ratio of I) is 50/50 to 97/3. 3. The manufacturing method according to claim 1 or 2, wherein component (I) is glycidyl methacrylate. 4. A compound with an oxirane ring has a solubility in water of 2.
g/100ml water (25℃) or more, molecular weight 100
0 or less, epoxy equivalent (molecular weight/number of oxirane rings)
4. The method according to claim 1, 2 or 3, wherein the compound has a compound of 300 or less.