JP2604339B2 - Hydrophilic gel fine particles and method for producing the same - Google Patents

Description

Description: TECHNICAL FIELD The present invention relates to hydrophilic gel fine particles having a narrow particle size distribution and a method for producing the same.

(Prior art) Conventionally, as hydrophilic gel fine particles, natural polymers such as dextran, agarose, and gelatin are insolubilized using various cross-linking agents and formed into fine particles, or acrylamide is polymerized together with a cross-linking agent to obtain particles. And so on. Methods for producing these particles are known. For example, in the case of dextran gel, an alkali solution of dextran having an appropriate molecular weight distribution is dispersed in an appropriate organic solvent that is immiscible with water, and then a dispersion stabilizer such as nonionic soap is added. create. When epichlorohydrin is added to this system, it reacts with the dextran matrix to form a glycerin cross-linking between sugar chains to form gel particles. In the case of acrylamide gel which is a synthetic polymer, for example, it is produced by a so-called reverse phase suspension polymerization method as disclosed in Japanese Patent Publication No. 56-25921, Japanese Patent Application Laid-Open No. 57-153010, or a solution. It is produced by pulverizing and classifying a polymer mass polymerized in the above. With these methods,
Both the particle size and particle size distribution of the obtained particles, the particle shape is determined by the mechanical stirring conditions when suspending the monomer or the pulverizing conditions of the polymer mass, the particle size is usually several tens of microns or more The particle size distribution also had a certain distribution. In addition, since these gel fine particles mainly use a natural polymer as a polymer skeleton, the functional group desired by the particle user, the amount of the functional group, or
It is almost impossible to control colloidal properties such as ζ-potential.

In recent years, hydrophilic gel fine particles have been widely used with the development of the bioindustry. For example, a carrier for enzyme immobilization, a column packing material for chromatographic separation of macromolecules such as biopolymers, or a biological material such as antigen-antibody reaction. It is used as a carrier for diagnostics by the particle agglutination method utilizing a chemical reaction. However, as described above, most of the currently available hydrophilic gel fine particles have a particle size of several tens of microns or more, the total surface area of the particles is small, and the particle size distribution is wide, so that as described above. When used for various applications,
It cannot be said that it has sufficient performance in terms of reaction efficiency, reaction reproducibility, or sensitivity. Further, even when a biological substance is immobilized on hydrophilic gel particles, a desired functional group and amount of the functional group cannot be easily selected arbitrarily.

In recent years, Japanese Patent Publication No.
No. 29962 discloses that glycidyl methacrylate is
% To form particles, and then a method for producing hydrophilic particles in which an epoxy group is ring-opened with an acid or a base to convert to 2,3-dioxypropyl methacrylate has been proposed. Enzymes, antigens, antigens,
When used for the treatment of proteins such as antibodies, protein denaturation is involved. When used as a carrier for a diagnostic agent, a column packing material, a carrier for immobilizing an enzyme, etc., the part involved in the reaction is the particle surface, and it is not necessary to make the inside of the particle hydrophilic.
Although the particles obtained by this method have a particle size on the order of microns, they have a wide particle size distribution, which is not sufficient for satisfying the sensitivity of the reaction or the required basic performance.

(Problems to be Solved by the Invention) The present inventors have conducted intensive studies to develop a method for producing hydrophilic gel fine particles free from the above-mentioned disadvantages. The present inventors have found that hydrophilic gel fine particles having a particle diameter of 0.1 to 10 μm in a water-swelled state and having a sharp particle diameter distribution can be obtained, and the present invention has been completed based on this finding.

According to the present invention, the monoethylenically unsaturated amide monomer is 20 to 94.8% by weight, the crosslinkable ethylenically unsaturated monomer is 5 to 60.0% by weight, the ethylenically unsaturated Carboxylic acid 0.1-30
A crosslinked polymer particle comprising 0.1 to 50% by weight of an acrylic acid or methacrylic acid ester monomer, and 30% by weight or less of a monomer copolymerizable with the monomer, if necessary. The particle size in the water swelling state is 0.1 to 10 μm
Wherein the ratio of the weight average particle diameter to the number average particle diameter (weight average particle diameter / number average particle diameter) is 1.2 or less and the hydrophilic gel fine particles and the monoethylenically unsaturated amide monomer 20 to 94.8.
% By weight, 5-60.0% by weight of crosslinkable ethylenically unsaturated monomer, 0.1-30% by weight of ethylenically unsaturated carboxylic acid, 0.1-50% by weight of acrylic acid or methacrylic acid ester monomer 30% by weight or less of a monomer copolymerizable with the above monomer may be copolymerized using a radical polymerization initiator in a solvent in which these monomers are dissolved and the resulting polymer is not dissolved. Production of hydrophilic gel fine particles having a particle diameter in a water-swelled state of 0.1 to 10 μm and a ratio of weight average particle diameter to number average particle diameter (weight average particle diameter / number average particle diameter) of 1.2 or less. A law is provided.

The hydrophilic gel fine particles of the present invention are prepared by dispersion polymerization (Dispersion P).
olymerzation). Dispersion polymerization (see, for example, CAN. J. Chem., Vol. 63 209-216 (1985)) is a solvent for a monomer, but dissolves the monomer in an organic solvent that is a non-solvent for the polymer. Is a method of polymerizing using a radical polymerization initiator. If necessary, a dispersion stabilizer may be added to the reaction system in order to prevent aggregation of the produced polymer. Although the mechanism of the polymerization is not yet clear, when the polymerization reaction is started in the solution, the monomer is converted into a polymer, but at a certain degree of polymerization, the polymer is precipitated and becomes a particle. Thereafter, the polymerization reaction is expected to proceed in a heterogeneous system. When observing the polymerization behavior, the reaction system changes from transparent to milky white within a few minutes after the initiation of polymerization. Since the polymer is precipitated almost at the same time, particles having a uniform particle size can be obtained.

Among the monomers used in the present invention, the monoethylenically unsaturated amide monomer is a monomer that forms the skeleton of the hydrophilic gel fine particles of the present invention, and is essential for imparting hydrophilicity to the particles. Component. Examples of the monoethylenically unsaturated amide monomer include acrylamide, methacrylamide, diacetacrylamide, N-hydroxymethylacrylamide, and the like.
It can be used in combination of more than one species. The amount of the ethylenically unsaturated amide monomer used is usually 20 to
94.8% by weight. Below 20% by weight the particles obtained do not meet the purpose of the present invention. On the other hand, when the content exceeds 94.8% by weight, the obtained polymer particles are insolubilized (gelled), it becomes difficult to control the particle size distribution, and the amount of functional groups becomes insufficient. Preferably it is 20 to 88% by weight.

The crosslinkable ethylenically unsaturated monomer used in the present invention is an essential component for insolubilizing particles and making them water-swellable.
Examples of the crosslinkable ethylenically unsaturated monomer include bis (meth) acrylamide monomers such as methylenebis (meth) acrylamide, ethylene glycol di (meth) acrylate, triethylene glycol dimethacrylate, ethylene dimethacrylate, and trimethylolpropane. Examples thereof include polyfunctional (meth) acrylate monomers such as trimethacrylate. The amount used is 5 to 60% by weight of the total monomers. If it is less than 5% by weight, gel particles having good water swellability cannot be obtained, and if it exceeds 60% by weight, the monodispersity of the particle size distribution is impaired (weight average particle diameter / number average particle diameter> 1.2). Preferably it is 10 to 50% by weight.

The ethylenically unsaturated carboxylic acid used in the present invention is
It is an essential component for maintaining the monodispersity of the particle size distribution, and if polymerization is performed in a system lacking the ethylenically unsaturated carboxylic acid, monodispersed particles cannot be obtained. Examples of the ethylenically unsaturated carboxylic acid include acrylic acid, methacrylic acid, crotonic acid, cinnamic acid, itaconic acid, fumaric acid, maleic acid, butenetricarboxylic acid, 3-butenoic acid, and 4-pentenoic acid. it can. The amount of the carboxylic acid used is 0.1 to 30% by weight of the total monomers, and if it exceeds 30% by weight, the obtained particle size distribution becomes wide. Preferably, 2 to 20% by weight
It is.

The acrylic acid or methacrylic acid ester used in the present invention is an essential component for introducing a specific functional group into the obtained particles and for maintaining the monodispersibility of the particles. (Meth) acrylate monomers include ethylene oxide-containing (meth) acrylates such as ethylene glycol (meth) acrylate and triethylene glycol (meth) acrylate; hydroxymethyl (meth) acrylate, (meth) acryl Hydroxy group-containing (meth) acrylates such as hydroxypropyl acid; epoxy group-containing (meth) acrylates such as glycidyl (meth) acrylate; methylaminoethyl (meth) acrylate, t-butylaminoethyl (meth) acrylate; Amino group-containing (meth) acrylates such as dimethylaminoethyl (meth) acrylate, dimethylaminopropyl (meth) acrylate, diethylaminoethyl (meth) acrylate, and dibutylaminoethyl (meth) acrylate Examples thereof include alkyl (meth) acrylates such as methyl (meth) acrylate, ethyl (meth) acrylate, and butyl (meth) acrylate, and these monomers are used alone or in combination of two or more. be able to. The amount used is required to be adjusted depending on the purpose of use of the particles. However, in order to maintain the monodispersibility of the particles, the amount is 0.1 to 50% by weight of the total monomers, preferably 5 to 40%. % By weight.

Monoethylenically unsaturated amide monomers used in the present invention, crosslinkable ethylenically unsaturated monomers, ethylenically unsaturated carboxylic acids, in addition to ester monomers of acrylic acid or methacrylic acid, these Monomers copolymerizable with the monomers can also be used within certain limits. Examples of the type of the monomer include aromatic vinyl monomers such as styrene, α-methylstyrene, p-methylstyrene, chloromethylstyrene, and halogenated styrene, and conjugated diolefins such as butadiene and isoprene. There is no particular limitation so long as it is soluble in the solvent used for the polymerization reaction and copolymerizes with other monomer components constituting the particles. The amount used is in the range of 0 to 30% by weight of the total monomers. When the amount exceeds 30% by weight, the ratio of other components constituting the particles is reduced, and the particle size distribution of the swollen particles is widened.

The solvent used in the present invention dissolves the radical polymerization initiator and dissolves the monomer used before the start of the reaction, but is not particularly limited as long as it does not dissolve the obtained polymer. As the solvent used, lower alcohols, tetrahydrofuran, dimethylformamide, dioxane,
Methyl ethyl ketone, pyridine, dimethyl sulfoxide and the like can be exemplified. It is also possible to mix water with the above-mentioned solvent if necessary. In the present invention, the use of lower alcohols is particularly preferred. Examples of the lower alcohol include methyl alcohol, ethyl alcohol, propyl alcohols, butyl alcohols, and the like, and one or more kinds may be used.

Further, the concentration of the monomer in the solution is preferably 50% by weight or less. If the content is more than 50% by weight, it is difficult to obtain primary particles, and the particles aggregate. More preferably, it is 1 to 20% by weight.

In the present invention, conventionally known azo compounds, organic peroxides and the like are used as the radical polymerization initiator, but are not particularly limited as long as they are soluble in the above-mentioned solvents. For example, examples of the azo compound include 2,2-azobisisobutyronitrile and 2,2-azobis (2-methyl) valeronitrile, and examples of the organic peroxide include acetyl peroxide and propionyl peroxide. , Isobutyryl peroxide, benzoyl peroxide and the like, and these can be used in an appropriate combination. The amount used is also not particularly limited, and varies depending on the type of the initiator, but is usually used at a ratio of 0.005 to 5 parts by weight per 100 parts by weight of the monomer mixture. The polymerization temperature varies depending on the type of the solvent used and the type of the radical polymerization initiator, but is usually preferably in the range of 20 to 100 ° C.

In this way, the particle diameter in the water-swelled state is 0.1 to 10 μm by dispersion-polymerizing the monomer,
Hydrophilic gel fine particles having a sharp particle diameter distribution (weight average particle diameter / number average particle diameter ≦ 1.2) can be obtained.

The particles obtained after the polymerization are separated from the solution used for the polymerization reaction by filtration, decantation, or an evaporator. The obtained particles are dried and redispersed in water to form hydrophilic gel fine particles.

The polymerization may be usually carried out by dissolving all monomers and a polymerization initiator in a solvent, but when a desired particle size cannot be obtained, or when a desired amount of a functional group cannot be obtained, after polymerization is completed, It is also possible to obtain a desired particle size or functional group content by adding a polymer or monomer, a polymerization initiator or a polymerization initiator of the monomer and an organic solvent dissolving them, and further continuing the polymerization reaction. Even in that case, the concentration of the monomer in the solution is
It is preferable that the content be 50% by weight or less. If the content is 50% by weight or more, it is difficult to obtain primary particles, and the particles aggregate.

(Effects of the Invention) Thus, according to the present invention, the particle diameter and particle diameter distribution in the water-swelled state as compared with conventionally known hydrophilic gel fine particles,
It is possible to obtain a hydrophilic fine particle gel in which the functional group and the amount of the gel fine particles and the crosslinking density are controlled. Further, this hydrophilic fine particle gel can be used as a carrier for a diagnostic agent.

(Example) Hereinafter, the present invention will be described more specifically with reference to examples. The parts and percentages in the examples are on a weight basis unless otherwise specified.

Example 1 Two reactors equipped with a stirring blade, a cooling condenser, a nitrogen gas inlet tube, and a thermometer were purged with nitrogen in advance. In this reaction vessel, 750 g of ethanol, 67.1 g of acrylamide, glycidyl methacrylate (Blenmer G, manufactured by NOF Corporation) 10
g, methylene bisacrylamide 16.2 g, methacrylic acid 10
g and 0.42 g of 2,2-azobisisobutyronitrile were added, and the mixture was stirred until a homogeneous system was obtained. Then, under nitrogen bubbling
The mixture was heated to 60 ° C. to start the reaction and kept for 24 hours. Then it was cooled. Ethanol was removed from the obtained suspension using an evaporator, and the suspension was further treated with a vacuum dryer for 12 hours to completely remove ethanol. Thereafter, the obtained resin powder was added to distilled water, and 0.1N
An aqueous OH solution was added until pH 9. The particle diameter of the obtained hydrophilic gel fine particles measured with a Coulter Multisizer (manufactured by Coulter Inc.) in a state of swelling in water has a weight average of 2.
It was a spherical particle having an extremely sharp particle size distribution of 05 μm and a number average of 1.82 μm (weight average particle size / number average particle size = 1.13). Alternatively, the ζ-potential measured by a microscope-type electrophoretic velocity measuring apparatus (Mark II Electrophoretic velocity measuring apparatus manufactured by Rank Brothers) was -32.5 mV.

Comparative Example 1 A polymerization reaction was carried out in the same manner as in Example 1 except that methacrylic acid was not used. The particle diameter of the obtained hydrophilic gel fine particles in a swollen state in water (measured by Coulter Multisizer (manufactured by Coulter)) is 4.85 μm in weight average and 1.32 μm in number average, and the spherical particles have a very wide particle size distribution (Weight average particle diameter / number average particle diameter = 3.67).

Example 2 Example 1 was repeated, and after completion of the polymerization reaction, 40 g of acrylamide, 10 g of methacrylic acid, and 0.1 g of 2,2-azobisisobutyronitrile were further added, and the reaction was continued at 60 ° C. for 24 hours. The particle size of the obtained hydrophilic gel fine particles in a swelling state in water,
The spherical particles had an extremely sharp particle size distribution with a weight average of 2.54 μm and a number average of 2.30 μm (weight average particle size / number average particle size = 1.10). The ζ-potential measured with a microscope-type electrophoretic velocity measuring apparatus (Mark II Electrophoretic velocity measuring apparatus manufactured by Rank Brothers) was -52.0 mV.

Example 3 In the same manner as in Example 1, n-butanol was added to the reaction vessel.
750 g, 67.1 g of acrylamide, 10 g of glycidyl methacrylate (Blenmer G, manufactured by NOF CORPORATION), 16.2 g of methylenebisacrylamide, 10 g of methacrylic acid, and 0.42 g of 2,2-azobisisobutyronitrile were added and stirred until a homogeneous system was obtained. Thereafter, the mixture was heated to 60 ° C. under nitrogen bubbling to start the reaction and kept as it was for 24 hours. Then it was cooled.
N-Butanol was removed from the obtained suspension by an evaporator, and further treated by a vacuum dryer for 12 hours to completely remove n-butanol. Then, the obtained resin powder was added to distilled water, and a 0.1N NaOH aqueous solution was added to the suspension.
It was added until pH 9. The particle diameter of the obtained hydrophilic gel fine particles in a water-swelled state is 4.35 μm in weight average, and 4.14 in number average.
μm, and were spherical particles having a monodisperse particle size distribution (weight average particle size / number average particle size = 1.05). Alternatively, the ζ-potential measured by a microscope-type electrophoretic velocity measuring device is −3.
0.3 mV.

Example 4 In the same manner as in Example 1, ethanol 750 was placed in a reaction vessel.
g, 50.9 g of acrylamide, 16.2 g of methylenebisacrylamide (manufactured by Wako Pure Chemical Industries), polyethylene oxadiacrylate (molecular weight: 522) (KAYARAD PEG400D manufactured by Nippon Kayaku Co., Ltd.)
A) 16.2 g, methacrylic acid 20 g, and 2,2-azobisisobutyronitrile 0.42 g were added, and the mixture was stirred until a homogeneous system was obtained. Thereafter, the mixture was heated to 60 ° C. under nitrogen bubbling to start the reaction and kept as it was for 24 hours. Then it was cooled. Ethanol was removed from the obtained suspension with an evaporator, and further treated with a vacuum dryer for 12 hours to completely remove ethanol. Thereafter, the obtained resin powder was added to distilled water, and a 0.1 N NaOH aqueous solution was added to the suspension until the pH became 9. The particle diameter of the obtained hydrophilic gel fine particles in a water-swelled state was 1.64 μm in weight average and 1.61 μm in number average, and was spherical particles having a monodisperse particle diameter distribution (weight average particle diameter / number average particle). Diameter = 1.02). The ζ-potential measured by a microscope-type electrophoresis velocity measuring device was -24.7 mV.

Example 5 In the same manner as in Example 1, ethanol 750 was placed in a reaction vessel.
g, 52 g of acrylamide, 10 g of methyl methacrylate, 30.0 g of methylenebisacrylamide, 8 g of methacrylic acid, 2,2
-0.42 g of azobisisobutyronitrile was added and stirred until a homogeneous system was obtained. Thereafter, the mixture was heated to 60 ° C. under nitrogen bubbling to start the reaction and kept as it was for 24 hours. Then it was cooled. Ethanol was removed from the obtained suspension with an evaporator, and further treated with a vacuum dryer for 12 hours to completely remove ethanol. Thereafter, the obtained resin powder was added to distilled water, and a 0.1 N NaOH aqueous solution was added to the suspension until the pH became 9. The particle diameter of the obtained hydrophilic gel fine particles in a swelling state in water was 0.84 μm in weight average and 0.76 μm in number average, and was spherical particles having a monodisperse particle diameter distribution (weight average particle diameter / number average particle). Diameter = 1.11).
The ζ-potential measured by a microscope-type electrophoresis velocity measuring device was -23.3 mV.

Comparative Example 2 A polymerization reaction was carried out in the same manner as in Example 5 except that methyl methacrylate was not used, and the particle diameter of the obtained hydrophilic gel fine particles in a water-swelled state was 2.23 μm in weight average. Was a spherical particle having a number average of 0.26 μm and an extremely wide particle diameter distribution (weight average particle diameter /
Number average particle size = 8.58).

Example 6 In the same manner as in Example 1, ethanol 750 was placed in a reaction vessel.
g, acrylamide 67.1 g, dimethylaminoethyl methacrylate (Mitsubishi Rayon acrylic ester DM) 10 g,
Methylenebisacrylamide 16.2 g, methacrylic acid 10 g,
0.42 g of 2,2-azobisisobutyronitrile was added, and the mixture was stirred until a homogeneous system was obtained. Then, 60 ℃ under nitrogen bubbling
To start the reaction and kept for 24 hours. Then it was cooled. Ethanol was removed from the obtained suspension with an evaporator, and further treated with a vacuum dryer for 12 hours to completely remove ethanol. Thereafter, the obtained resin powder was added to distilled water, and a 0.1 N NaOH aqueous solution was added to the suspension until the pH became 9. The particle diameter of the obtained hydrophilic gel fine particles in the state of swelling in water was 2.45 μm in weight average and 2.33 μm in number average, and was spherical particles having a monodisperse particle diameter distribution (weight average particle diameter / number average particle). Diameter = 1.05). When the ζ-potential at each pH was measured in a saline buffer having an ionic strength of 10 ^{-3 with} a microscope-type electrophoretic velocity measuring device,
The particles were amphoteric having an isoelectric point of pH 7.2.

Example 7 In the same manner as in Example 1, ethanol 750 was placed in a reaction vessel.
g, 62.1 g of acrylamide, 10 g of glycidyl methacrylate (Blenmer G, manufactured by NOF Corporation), 16.2 g of methylenebisacrylamide, 10 g of methacrylic acid, 5 g of styrene, 2,2-
0.42 g of azobisisobutyronitrile was added, and the mixture was stirred until a homogeneous system was obtained. Then, after bubbling with nitrogen, the reactor was heated to 60 ° C. to start the reaction and kept as it was for 24 hours. Then it was cooled. Ethanol was removed from the obtained suspension with an evaporator, and the suspension was further treated with a vacuum dryer for 12 hours to completely remove ethanol. Thereafter, the obtained resin powder was added to distilled water, and a 0.1 N NaOH aqueous solution was added to the suspension until the pH became 9. The obtained hydrophilic gel fine particles were spherical particles having a monodisperse particle size distribution in a water-swelled state with a weight average of 1.66 μm and a number average of 1.45 μm (weight average particle size / number average particle size). = 1.14). The ζ-potential was −42.5 mV.

 ──────────────────────────────────────────────────の Continued on the front page (72) Inventor Masayoshi Sekiya 3-21-13 Minamimagome, Ota-ku, Tokyo

Claims (4)

(57) [Claims] (1) a monoethylenically unsaturated amide monomer (20 to 9);
4.8% by weight, crosslinkable ethylenically unsaturated monomer 5 to 60.0% by weight, ethylenically unsaturated carboxylic acid 0.1 to 30% by weight, acrylic acid or methacrylic acid ester monomer 0.1 to 50
% By weight of the crosslinked polymer particles having a particle diameter of 0.1 to 10 μm in a water-swelled state, and having a ratio of weight average particle diameter to number average particle diameter (weight average particle diameter / number average particle diameter).
Hydrophilic gel fine particles having a particle size of 1.2 or less. 2. Monoethylenically unsaturated amide monomers 20 to 9
4.8% by weight, crosslinkable ethylenically unsaturated monomer 5 to 60.0% by weight, ethylenically unsaturated carboxylic acid 0.1 to 30% by weight, acrylic acid or methacrylic acid ester monomer 0.1 to 50
% By weight, a crosslinked polymer particle comprising 30% by weight or less of a monomer copolymerizable with the above-mentioned monomer, having a particle diameter of 0.1 to 10 μm in a water-swelled state, having a weight average particle diameter and a number average particle diameter. Particle diameter ratio (weight average particle diameter / number average particle diameter) of 1.2
The following hydrophilic gel fine particles. 3. Monoethylenically unsaturated amide monomers 20 to 9
4.8% by weight, crosslinkable ethylenically unsaturated monomer 5 to 60.0% by weight, ethylenically unsaturated carboxylic acid 0.1 to 30% by weight, acrylic acid or methacrylic acid ester monomer 0.1 to 50
% By weight in a water-swelling state characterized by being copolymerized using a radical polymerization initiator in a solvent in which these monomers are dissolved and the obtained polymer is not dissolved.
A method for producing hydrophilic gel fine particles having a particle diameter of 1 to 10 μm and a ratio of weight average particle diameter to number average particle diameter (weight average particle diameter / number average particle diameter) of 1.2 or less. 4. A monoethylenically unsaturated amide monomer 20 to 9
4.8% by weight, crosslinkable ethylenically unsaturated monomer 5 to 60.0% by weight, ethylenically unsaturated carboxylic acid 0.1 to 30% by weight, acrylic acid or methacrylic acid ester monomer 0.1 to 50
% By weight, and 30% by weight or less of a monomer copolymerizable with the above monomer, in a solvent in which these monomers are dissolved and the resulting polymer is not dissolved, using a radical polymerization initiator. The particle size in the water swelling state characterized by polymerization is 0.1 to 1
0 μm, and a method for producing hydrophilic gel fine particles wherein the ratio of weight average particle diameter to number average particle diameter (weight average particle diameter / number average particle diameter) is 1.2 or less.