JPH01315408A - Hydrophilic gel fine particle and production thereof - Google Patents

Abstract

PURPOSE:To obtain the title fine particles uniform in size in a water-swollen state, with narrow size distribution, useful as a carrier for diagnostic drugs by dispersing a monomer mixture of specified composition followed by polymerization. CONSTITUTION:A copolymerization is carried out, using a radical initiator, between (A) 20-94.8wt.% of a monoethylenic unsaturated amide monomer (e.g., acrylamide), (B) 5-60wt.% of a crosslinkable ethylenic unsaturated monomer (e.g., ethylenedimethacrylate), (C) 0.1-30wt.% of an ethylenic unsaturated carboxylic acid (e.g., acrylic acid), (D) 0.1-50wt.% of a (meth)acrylic ester monomer (e.g., ethylene glycol acrylate), and (E) 0-30wt.% of another copolymerizable monomer in a solvent soluble for the components A-E but insoluble for the polymer to be formed (pref. lower alcohol), thus obtaining the objective fine particles 0.1-10mum in size in a water-swollen state and <=1.2 in ratio: weight- average size/number-average size.

Description

DETAILED DESCRIPTION OF THE INVENTION (Field of Industrial Application) The present invention relates to hydrophilic gel particles with a narrow particle size distribution and a method for producing the same.

(Prior art) Hydrophilic gel microparticles have conventionally been made by insolubilizing natural polymers such as dextran, agarose, and gelatin using various crosslinking agents and turning them into microparticles, or by polymerizing acrylamide with a crosslinking agent. There are some things that have changed. These particle manufacturing methods are known. For example, in dextran gel, an alkaline solution of dextran with 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. to create a reverse phase dextran drop. When epichlorohydrin is added to this system, it reacts with the dextran matrix to form glycerin crosslinks between sugar chains, resulting in gel particles. In the case of acrylamide gel, which is a synthetic polymer, for example, Japanese Patent Publication No. 56-25921, Japanese Patent Application Laid-Open No. 57-1
It is produced by a so-called reverse-phase suspension polymerization method as disclosed in publications such as No. 53010, or by crushing and classifying a polymer mass polymerized in a solution. In all of these methods, the particle size, particle size distribution, and particle shape of the particles obtained are determined by the mechanical stirring conditions when suspending the monomer or the crushing conditions of the polymer lump, and the particle size is usually a few. The particle size was 10 microns or more, and the particle size distribution was also to some extent. In addition, since these gel particles mainly use natural polymers as the polymer skeleton, it is almost impossible for particle users to control desired functional groups, amount of functional groups, or colloidal properties such as ζ-potential. be.

In recent years, hydrophilic gel microparticles have come into widespread use with the development of the bioindustry, for example as carriers for enzyme immobilization,
It is used as a column packing material for chromatography for separating large molecules such as biopolymers, or as a carrier for diagnostic reagents using particle aggregation methods that utilize biological reactions such as antigen-antibody reactions. However, as mentioned above, most of the hydrophilic gel particles currently on the market 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 the above-mentioned problems arise. When used in such applications, it cannot be said to have sufficient performance in terms of reaction efficiency, reaction reproducibility, or sensitivity.

Further, when immobilizing biological substances on hydrophilic gel particles, there are limitations to the method because it is not possible to easily select a desired functional group or amount of functional groups.

In addition, recently, as a method to improve the above-mentioned drawbacks,
No. 29962 discloses that hydrophilic particles are prepared by forming particles using 50% or more of glycidyl methacrylate, and then ring-opening the epoxy group using an acid or base to convert it into 2,3-dioxypropyl methacrylate. Although a production method has been proposed, if the particles are not sufficiently purified and the acids or bases used in the processing are completely removed, they may cause denaturation of the proteins when used for processing proteins such as enzymes, antigens, and antibodies. It ends up. Furthermore, when used as a carrier for diagnostic reagents, a column packing material, a carrier for enzyme immobilization, etc., the part involved in the reaction is the particle surface, and there is no need to make the inside of the particle hydrophilic as well. Further, although the particles obtained by this method have a particle size on the micron order, the particle size distribution is wide, so this is not sufficient to provide sufficient reaction sensitivity or to satisfy the required basic performance.

(Problems to be Solved by the Invention) In order to develop a method for producing hydrophilic gel particles free from the above-mentioned drawbacks, the present inventors have conducted intensive research and found that by polymerizing specific monomers by dispersion polymerization, It was discovered that hydrophilic gel particles having a particle size of 0.1 to 10 μm in a water-swollen state and a sharp particle size distribution can be obtained, and based on this knowledge, the present invention was completed.

(Means for Solving the Problems) According to the present invention, 20 to 94.8% by weight of monoethylenically unsaturated amide monomer, 5 to 60% by weight of crosslinkable ethylenically unsaturated monomer, ethylene unsaturated carboxylic acid 0.
Crosslinked polymer particles consisting of 1 to 30% by weight, 0.1 to 50% by weight of an ester monomer of acrylic acid or methacrylic acid, and 0 to 30% by weight of a monomer copolymerizable with the above monomers. and the particle size in the water-swollen state is 0.1 to 10μ
hydrophilic gel fine particles and monoethylenically unsaturated amide monomer 20, which are
~94.8% by weight, crosslinkable ethylenically unsaturated monomer 5~
60.0% by weight, ethylenically unsaturated carboxylic acid 0.1~
30% by weight, 0.1 to 50% by weight of an ester monomer of acrylic acid or methacrylic acid, and 0 to 30% by weight of a monomer copolymerizable with the above monomers, by dissolving these monomers. The particle size in a water-swollen state is 0.1 to 10 μm, and the weight average particle size and number average particle size are copolymerized using a radical initiator in a solvent that does not dissolve the resulting polymer. The ratio (weight average particle diameter/number average particle diameter) is 1
．． 2 or less is provided.

The hydrophilic gel particles of the present invention are produced by dispersion polymerization (Dispe-r).
sion Polymerzation). Dispersion polymerization [e.g. CAN, J, CheI
ll,, vol 63209 216 (1985)] is a solvent for the monomer, and a method in which a monomer is polymerized using a polymerization initiator soluble in the solvent in an organic solvent that is a non-solvent for the polymer. It is.

Further, a dispersion stabilizer may be added to the reaction system if necessary to prevent agglomeration of the produced polymer.

The mechanism of polymerization is still unclear, but when the polymerization reaction starts in a solution, the monomer is converted into a polymer, but at a certain degree of polymerization, the polymer precipitates and becomes particulate. After that, it is expected that the polymerization reaction will proceed in an unrestricted manner. Observing the polymerization behavior, the reaction system changes from transparent to milky white several minutes after the start of polymerization. Since the polymers precipitate at approximately the same time, it is possible to obtain particles with uniform particle diameters.

Among the monomers used in the present invention, the monoethylenically unsaturated amide monomer is a monomer that forms the skeleton of the hydrophilic gel particles of the present invention, and is essential for imparting hydrophilicity to the particles. It is a component. Examples of monoethylenically unsaturated amide monomers include acrylamide, methacrylamide, diacetoacrylamide, N-hydroxymethylacrylamide, etc., and these monomers may be used alone or in combination of two or more. I can do it. The amount of ethylenically unsaturated amide monomer used is usually 20 to 20% of the total monomers.
It is 94.8% by weight. If it is less than 20% by weight, the particles obtained will not meet the objectives of the present invention.

If it exceeds 94.8% by weight, it becomes difficult to control the insolubilization (gelation) and particle size distribution of the resulting polymer particles, 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 the particles and making them water-swellable. Examples of crosslinked ethylenically unsaturated monomers include bis(meth)acrylamide monomers such as methylene bis(meth)acrylamide, ethylene glycol di(meth)acrylate,
Examples include polyfunctional (meth)acrylic acid ester monomers such as triethylene glycol dimethacrylate, ethylene dimethacrylate, and trimethylolpropane trimethacrylate. The amount used is 5 to 60% by weight of the total monomers. If it is less than 5% by weight, gel particles with good water swelling properties cannot be obtained, and if it exceeds 60% by weight, the monodispersity of the particle size distribution will be impaired (weight average particle size/number average particle size> 1.2). ). Preferably 10-50M
%.

The ethylenically unsaturated carboxylic acid used in the present invention is an essential component to maintain monodispersity of the particle size distribution,
If polymerization is carried out in a system lacking ethylenically unsaturated carboxylic acid, monodisperse particles cannot be obtained.

Examples of ethylenically unsaturated carboxylic acids include acrylic acid, methacrylic acid, crotonic acid, cinnamic acid, itaconic acid, fumaric acid, maleic acid, butenetricarboxylic acid, 3-butenoic acid, and 4-pentenoic acid. 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 resulting particle size distribution will become wider. Preferably it is 2 to 20% by weight.

Furthermore, the ester of acrylic acid or methacrylic acid used in the present invention is an essential component for introducing specific functional groups into the particles obtained and for maintaining the monodispersity of the particles. (Meth)acrylic acid ester monomers include ethylene oxide-containing (meth)acrylic acid esters such as ethylene glycol (meth)acrylate and triethylene glycol (meth)acrylate; hydroxymethyl (meth)acrylate, (meth)acrylate Hydroxy group-containing (meth)acrylic esters such as acid hydroxypropyl; Epoxy group-containing (meth)acrylic esters such as glycidyl (meth)acrylate; Methylaminoethyl (
meth)acrylate, L-butylaminoethyl (meth)
Acrylate, dimethylaminoethyl (meth)acrylate, dimethylaminopropyl (meth)acrylate,
Amino group-containing [meth)acrylic acid esters such as diethylaminoethyl (meth)acrylate and dibutylaminoethyl (meth)acrylate; alkyl (meth)acrylates such as methyl (meth)acrylate, ethyl (meth)acrylate, butyl (meth)acrylate, etc. Examples include acrylic esters, and these monomers can be used in combination of one or two types.
More than one species can be used in combination. The amount to be used needs to be adjusted depending on the intended use of the particles, but in order to maintain the monodispersity of the particles, the total amount of monomers should be 0.
It is 1 to 50% by weight, preferably 5 to 40% by weight.

In addition, other copolymerizable monomers used in the present invention are:
It is used as necessary to adjust the hydrophilicity or swelling degree of particles. The types of monomers are styrene, α-methylstyrene, p-methylstyrene, chloromethylstyrene,
Examples include aromatic vinyl monomers such as halogenated styrene, and diolefins such as butadiene and isoprene, but other monomers that are soluble in the solvent used in the polymerization reaction and that constitute the particles can be used. There is no particular limitation as long as it is copolymerized with body components. The amount used is in the range of 00 to 30% by weight of the total monomer, and if it exceeds 30% by weight, the ratio of other components constituting the particles decreases and the particle size distribution of the swollen particles becomes wide.

The solvent used in the present invention dissolves the polymerization initiator and the monomer used before the reaction starts, but is not particularly limited as long as it does not dissolve the obtained polymer.

Examples of the solvent to be used include lower alcohols, tetrahydrofuran, dimethylformamide, dioxane, methyl ethyl ketone, pyridine, and dimethyl sulfoxide. It is also possible to mix water with the above-mentioned solvent, if necessary. According to the invention, the use of lower alcohols is particularly preferred. Examples of lower alcohols include methyl alcohol, ethyl alcohol, propyl alcohol, butyl alcohol, etc.
It is also possible to use one species or two or more species.

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

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

By dispersing and polymerizing the above-mentioned monomers in this way, the particle diameter in the water-swollen state is 0.1 to 10 μm, and the particle diameter distribution is sharp (weight average particle diameter/number average particle diameter ≦1.2 ) hydrophilic gel particles are obtained.

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

Polymerization can usually be carried out by dissolving all monomers and a polymerization initiator in a solvent. However, if the desired particle size or the desired amount of functional groups cannot be obtained, after the completion of polymerization, The polymerization reaction is further continued by adding a monomer, a polymerization initiator, or a monomer, a polymerization initiator, and an organic solvent for dissolving them.
It is also possible to obtain a desired particle size or amount of functional groups. Even in that case, the concentration of monomer in the solution was 50
It is preferable to make it less than % by weight. At 50% by weight or more, it is difficult to obtain primary particles, and the particles tend to aggregate.

(Effects of the Invention) Thus, according to the present invention, compared to conventionally known hydrophilic gel particles, the particle size and particle size distribution in the water-swollen state, the functional groups and their amounts possessed by the gel particles, and the crosslinking density can be controlled. A hydrophilic microparticle gel can be obtained. Moreover, this hydrophilic microparticle gel can be used as a diagnostic agent.

(Example) The present invention will be described in more detail with reference to Examples below. In addition, parts and percentages in the examples are based on weight unless otherwise specified.

Example 1 Twenty-one reactors equipped with stirring blades, cooling condensers, nitrogen gas inlet tubes, and thermometers were replaced with nitrogen in advance. In this reaction vessel, 750 g of ethanol and 67.1 g of acrylamide were added.
g, glycidyl methacrylate (NOF Premer G) Log, methylene bisacrylamide 16.2 g,
methacrylic acid l.

g, 2.2-azobisisobutyronitrile 0.42 g
was added and stirred until a homogeneous system was obtained. Thereafter, the mixture was heated to 60° C. under nitrogen bubbling to initiate the reaction, and maintained as such for 24 hours. It was then cooled.

Ethanol was removed from the resulting suspension in an evaporator, and the suspension was further treated in a vacuum dryer for 12 hours to completely remove ethanol. Thereafter, the obtained resin powder was added to distilled water, and a 0.1N-Na0)I aqueous solution was added to the suspension until the pH reached 9. The particle diameter of the obtained hydrophilic gel particles in a swollen state in water was measured using a Coulter Multisizer (manufactured by Coulter Co., Ltd.), and the weight average diameter was 2.05.
μm, number average 1.82 μm, and the particles were spherical particles with an extremely sharp particle size distribution (weight average particle size/number average particle size = 1.13). Further, the ζ-potential measured with a microscope-type electrophoretic velocity measuring device (Mark 1 electrophoretic velocity measuring device 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 omitted. The particle diameter of the obtained hydrophilic gel particles in a swollen state in water (measured with a Coulter Multisizer (manufactured by Coulter)) was a weight average of 4.85μ.
m, the number average was 1.32 μm, and they were spherical particles with a very wide particle size distribution (weight average particle size / number average particle size =
3.67).

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

Example 3 In the same manner as in Example 1, n-butanol 7 was added to the reaction vessel.
50 g, acrylamide 67.1 g, glycidyl methacrylate (NOF Premer G) 10 g, methylenebisacrylamide 16.2 g.

10 g of methacrylic acid and 0.42 g of 2,2-abbisisobutyronitrile were added and stirred until a homogeneous system was obtained.

Thereafter, the temperature was heated to 60''C under nitrogen bubbling to initiate the reaction, and the reaction was maintained for 24 hours.

It was then cooled. N-butanol was removed from the resulting suspension using an evaporator, and the suspension was further dried for 12 hours using a vacuum dryer.
The n-butanol was completely removed by treatment for several hours. Thereafter, the obtained resin powder was added to distilled water, and 0.0% was added to the suspension. 1N-NaOH aqueous solution was added until the pH reached 9.

The particle diameter of the obtained hydrophilic gel microparticles in a swollen state in water was 4.35 μm in weight average and 4.14 μm in number average, and they were spherical particles with a monodisperse particle size distribution (weight average particle diameter/number Average particle size = 1.05). In addition, the ζ-potential measured with a microscope electrophoresis velocity measuring device was -30.3 mV.
Met.

Example 4 In the same manner as in Example 1, 750 ethanol was added to the reaction vessel.
g, acrylamide 50.9 g, methylene bisacrylamide (Wako Pure Chemical Industries, Ltd.) 16.2 g, polyethylene oxide diacrylate (molecular weight 522) (Nippon Kayaku Co., Ltd. KAYARAD PEG400DA) 16.2 g, methacrylic acid 20 g, 2.2-azo 0.42 g of bisisobutyronitrile was added and stirred until a homogeneous system was obtained. Thereafter, the mixture was heated to 60° C. under nitrogen bubbling to initiate the reaction, and maintained as such for 24 hours. It was then cooled. Ethanol was removed from the resulting suspension using an evaporator, and the suspension was further treated in a vacuum dryer for 12 hours to completely remove ethanol. Thereafter, the obtained resin powder was added to distilled water, and the suspension contained 0. 1N-NaOH aqueous solution was added until the pH reached 9. The weight average particle diameter of the obtained hydrophilic gel particles in a swollen state in water is 1.64 μm.
, the number average was 1.61 gm, and they were spherical particles with a monodisperse particle size distribution (weight average particle size / number average particle size =
1.02). Further, the ζ-potential measured with a microscopic electrophoresis velocity measuring device was -24.7 mV.

Example 5 In the same manner as in Example 1, 750 ethanol was added to the reaction vessel.
g, Acrylamide 52g1 Methyl methacrylate 1
0 g, methylenebisacrylamide 30, Og, methacrylic acid 8 g, 2.2-azobisisobutyronitrile 0.
42 g was added and stirred until a homogeneous system was obtained. after that,
The reaction was started by warming to 60°C under nitrogen bubbling and kept there for 24 hours. It was then cooled. Ethanol was removed from the resulting suspension using an evaporator, and the suspension was further treated in a vacuum dryer for 12 hours to completely remove ethanol. Then, the obtained resin powder was added to distilled water, and an O, IN-NaOH aqueous solution was added to the suspension.
It was added until it reached H9. The particle diameter of the obtained hydrophilic gel microparticles in a swollen state in water was a weight average of 0.84 μm and a number average of 0.76 μm, and they were spherical particles with a monodisperse particle size distribution (weight average particle diameter/number). Average particle size = 1.11
). Further, the electric potential measured with a microscopic 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 omitted. The resulting hydrophilic gel particles had a weight average particle size of 2.23 μm and a number of particles in a swollen state in water. They were spherical particles with an average particle size of 0.26 μm and an extremely wide particle size distribution (weight average particle size/number average particle size = 8.58).

Example 6 In the same manner as in Example 1, 750 ethanol was added to the reaction vessel.
g, acrylamide 67.1 g, dimethylaminoethyl methacrylate (Mitsubishi Rayon Acryester D)
M) 10g, methylenebisacrylamide 16.2g
, methacrylic acid Log, 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 initiate the reaction, and maintained as such for 24 hours. It was then cooled. Ethanol was removed from the resulting suspension using an evaporator, and the suspension was further treated in a vacuum dryer for 12 hours to completely remove ethanol. Thereafter, the obtained resin powder was added to distilled water, and the suspension contained 0. IN-Na(lf
1 aqueous solution was added until the pH reached 9. The weight average particle diameter of the obtained hydrophilic gel particles in a swollen state in water is 2.45.
μm, number average 2.33 μm, and they were spherical particles with a monodisperse particle size distribution (weight average particle size/number average particle size = 1.05 >. Microscopic electrophoresis velocity measurement device showed an ionic strength of 10 When the ζ-potential was measured at each pH in a saline buffer of -3, it was found that the particles were amphoteric particles with an isoelectric point at pH 7.2.

Example 7 In the same manner as in Example 1, 750 ethanol was added to the reaction vessel.
g, acrylamide 62.1 g, glycidyl tamacrylate (NOF Bremmer G) 10 g, methylenebisacrylamide 16.2 g, methacrylic acid 10 g, styrene 5 g, 2.2-azobisisobutyronitrile 0.
42 g was added and stirred until a homogeneous system was obtained. Thereafter, after bubbling with nitrogen, the reactor was heated to 60°C to start the reaction, and maintained as it was for 24 hours. It was then cooled. Ethanol was removed from the resulting suspension using an evaporator, and the suspension was dried for 12 hours in a vacuum dryer.
The mixture was treated for several hours to completely remove ethanol. Then, add the obtained resin powder to distilled water and add 0% to the suspension.
．． 1N-NaOH aqueous solution was added until the pH reached 9. The resulting hydrophilic gel microparticles were spherical particles with a monodisperse particle size distribution of a weight average of 1.66 μm and a number average of 1.45 μm when swollen in water (weight average particle size/
Number average particle diameter = 1.14).

The ζ-potential was -42.5 mV.

Patent applicant: Zeon Corporation

Claims (1)

[Claims] 1. Monoethylenically unsaturated amide monomer 20-94.8
Weight%, crosslinkable ethylenically unsaturated monomer 5-60.0% by weight, ethylenically unsaturated carboxylic acid 0.1-30% by weight
, ester monomer of acrylic acid or methacrylic acid 0.
1 to 50% by weight, 0 monomers copolymerizable with the above monomers
~30% by weight, the particle size in the water-swollen state is 0.1 to 10 μm, and the ratio of the weight average particle size to the number average particle size (weight average particle size/number average particle size) ) is 1.2 or less. 2. Monoethylenically unsaturated amide monomer 20-94.8
Weight%, crosslinkable ethylenically unsaturated monomer 5-60.0% by weight, ethylenically unsaturated carboxylic acid 0.1-30% by weight
, ester monomer of acrylic acid or methacrylic acid 0.
1 to 50% by weight, 0 monomers copolymerizable with the above monomers
~30% by weight is copolymerized using a radical initiator in a solvent that dissolves these monomers and does not dissolve the resulting polymer.The particle size in the water-swollen state is 0.1 A method for producing hydrophilic gel fine particles having a particle size of 10 μm and a ratio of weight average particle size to number average particle size (weight average particle size/number average particle size) of 1.2 or less.