JPH0421637A - High polymer fine grain whose surface is modified with chitin derivative or chitosan and its production - Google Patents

Abstract

PURPOSE:To obtain high polymer fine grains having a hydrophilic and positively ionized surface and improved in affinity to a biological high polymer by modifying the surface of high polymer fine grains with a chitin derivative or chitosan. CONSTITUTION:High polymer fine grains having an active groups reactive with a carbodiimide compound, preferably a carboxyl latex having 0.2-10mum grain size is made to react with a compound expressed by the formula RN=C= NR' (R is ethyl and R' is dimethylaminopropyl or R and R' are cyclohexyl together) at ordinary temperature and successively subjected to a reaction with a chitin derivative (preferably diethylaminoethylchitin) or chitosan added at ordinary temperature. The resultant modified high polymer fine grains are used as a latex for immobilization of bioaffinitive ingredient, physiologically active substance and medicine, etc., latex for immunodiagnostic agent and endotoxin removing agent.

Description

[Detailed Description of the Invention] [Industrial Application Field] The present invention is applicable to chromatography adsorbents used for the separation and purification of biological components such as proteins and nucleic acids, carriers for drug delivery systems, and immunodiagnosis. The present invention relates to polymer particles whose surfaces are modified with chitin or chitosan, which are useful for latex, etc., and a method for producing the same.

[Conventional technology]

Conventionally, polystyrene latex with a hydrophobic surface has been used for diagnostic reagents for immunological clinical tests that utilize antigen-antibody reactions (JColloid LnterfaceSc
i, 71.350 (1979).

However, since polystyrene latex is hydrophobic,
Because of the lack of sensitization stability and storage stability, research and development are also being conducted on particulate polymers made by copolymerizing styrene with methacrylic acid, etc., in order to modify the hydrophobicity.

[Problem to be solved by the invention]

The above-mentioned styrene latex lacks sensitization stability and storage stability, and even styrene latex copolymerized with methacrylic acid etc. not only has insufficient sensitization stability and storage stability; It has a non-specific agglutination reaction and is not satisfactory as a latex for highly sensitive and quantitative analysis of immunological serum, plasma, urine, etc. on which antigens or antibodies are immobilized.

The present invention improves latex to be hydrophilic, and more preferably converts it to positively charged property to improve its affinity for biopolymers such as animal cells and immune proteins, and to specifically adsorb latex to biocomponents such as proteins and nucleic acids. The object of the present invention is to provide polymeric microparticles that have properties and are useful as carriers for drug delivery systems, latex for immunodiagnosis, etc., and a method for producing the same.

[Means to solve the problem]

The present invention etc. are bioaffinity components, physiologically active substances,
Chitin, which has selective adsorption and affinity for biopolymers such as proteins and enzymes, is used for the purpose of developing polymeric microparticles used as immobilization of drugs, diagnostic agents for immunological clinical tests, and endotoxin removal agents. Chitin derivatives and chitosan, which have glucosamine as a constituent unit and preferably also have a positive charge, can be modified on the surface of polymeric particles and used as carriers for drug delivery systems, latex for immunodiagnostic agents, etc. The present invention was completed by discovering that chitin derivatives and chitosan can be easily provided in the form of fine polymer particles, and a method for modifying chitin derivatives and chitosan into polymers.

The present invention is a polymer fine particle whose surface is modified with a chitin derivative or chitosan. Then, the carboxyl latex fine particles are reacted with a carbodiimide compound represented by the formula RN=C=NR' (wherein R is an ethyl group, R' is a dimethylaminopropyl group, or both R and R' are a cyclohexyl group), and then chitin This is a method for producing polymer fine particles whose surface is modified with a chitin derivative or chitosan, which is characterized by reacting the derivative or chitosan.

The polymer fine particles used in the present invention are polymer compounds having an active group that reacts with a carbodiimide compound, and carboxyl latex is preferable. The particle size is selected appropriately depending on the application of the product, but it is approximately 0.2 to 10 μm.
is commonly used.

The chitin derivatives modified into these polymer particles include:
Water-soluble chitin derivatives such as diethylaminoethyl chitin, carboxymethyl chitin, hydroxypropyl chitin, hydroxyethyl chitin, sulfate ester chitin, phosphate ester chitin, etc. are preferably used, and the diethylaminoethyl chitin previously announced by the present inventors Chitin [Pol
ymer Bulletin, 21.13-17 (1
989) E is the most suitable. Chitosan is also a substance obtained by deacetylating chitin, contains glucosamine, which is a constituent unit of chitin, and is a positively charged substance, and can be suitably applied to the present invention.

As described above, the polymer particles of the present invention have surfaces modified with chitin derivatives or chitosan, so the surfaces are modified to have positively charged and hydrophilic properties, and because they are fine particles, the surface area is small. In addition, the surface charge density is high, and by utilizing the surface activity, it can be usefully used as a diagnostic agent by adsorbing and desorbing biopolymers such as various proteins and enzymes, and by immobilizing antigens or antibodies. It is also expected to be used as a selective adsorbent and remover for endotoxin, a pyrogenic substance that is one of the causes of side effects of pharmaceuticals. Moreover, it is extremely useful because it can be used as a cosmetic component material such as a hair styling agent due to its water-retaining properties.

In order to produce the above-mentioned polymer fine particles having a chitin derivative or chitosan, polymer fine particles having an active group such as a carboxyl group, preferably carboxyl latex, are used, and this material is made by polymerizing, for example, a styrene monomer alone and forming a core particle. Thereafter, acrylamide is added and the styrene-acrylamide copolymer latex purified by a two-stage polymerization method is hydrolyzed.

The water-soluble general formula RN
=C=NR' (wherein R, R' have the same meanings as above), a carbodiimide compound, 1-ethyl-5-(3-dimethylaminopropyl)carbodiimide, 1,3-dicyclo React xylcarbodiimide at room temperature, then add chicken derivative or chitosan for about 12 hours.
When the reaction is carried out at room temperature for about a period of time, polymer fine particles whose surfaces are modified with the chitin derivative or chitosan of the present invention are obtained.

The appropriate amount of the chitin derivative and chitosan used in this method is 1 to 8 times the molar amount of the amino group of the chitin derivative or chitosan relative to the carboxyl group of the polymer, and 4 to 8 times the molar amount of the amino group of the chitin derivative or chitosan.
An 8-fold molar amount is preferred. Further, the amount of water-soluble carbodiimide compound to be used is suitably 0.5 to 1.5 times the molar amount, preferably 0.8 to 1.2 times the molar amount of the amino group of the chitin derivative or chitosan. .

Next, the method for producing the fine particles of the present invention and the properties of the obtained materials will be described.

〔Example〕

Example 1 Surface charge: 89.5μ equivalent/g latex, particle size: 0.
Carboxyl latex dispersion with a specific surface area of 10 rri/g/latex (concentration 0.0157 g/mf)
f) Add 0.0108 g of 1-ethyl-3-(3-dimethylaminopropyl)carbodiimide hydrochloride to 400 d and stir at room temperature for 2 hours. Next, 63 g of a 1% diethylaminoethyl chitin aqueous solution (twice the molar amount as the amine group of diethylaminoethyl chitin relative to the carboxyl group in the latex) was added, and the mixture was reacted for 12 hours.
Add M acetate buffer (PH=4), stir for 2 hours, and separate the resulting product. Washed with condensed water and dried under reduced pressure to obtain diethylaminoethyl chitin modified latex 0.68
I got g. The surface charge amount of this material is 24.6μ equivalent/
g. It was latex. Further, the contact angle (θ) of the obtained latex was 26.4. Note that the surface charge amount was measured by colloid titration, and the contact angle was measured using an automatic PHW type contact angle meter.

Examples 2-3 Same as Example 1 except that 1-ethyl-5-(3-dimethylaminopropyl)carbodiimide hydrochloride (WSC) and 1% diethylaminoethyl chitin aqueous solution (DEAE-chitin aqueous solution) were added in the amounts shown in Table 1. Perform the same operation and DEA
E-chitin modified latex was obtained. The yield, surface charge amount, and contact angle of this example were as shown in Table 1. In addition, 1%
The amount of DEAE-chitin is 4 to 8 times the molar amount of the amine group of DEAE-chitin relative to the carboxyl group in the latex.

Table Examples 4 to 6 The same operation as in Example 1 was performed except that the 1% DEAE-chitin aqueous solution was replaced with a 1% chitosan aqueous solution and the amounts of WSC and 1% chitosan aqueous solution shown in Table 2 were added. Obtained modified latex. The yield, surface charge amount, and contact angle of this example were as shown in Table 2. The amount of the 1% chitosan aqueous solution is 2 to 8 times the molar amount of the amino groups of chitosan relative to the carboxyl groups in the latex.

Below is an example of the table in the margin 7 Surface charge amount 38.5μ equivalent/g latex, concentration 0.
Using carboxyl latex aqueous dispersion 40-0175 g/d, WSC000052 g, 1% DEAE-
Chitin aqueous solution 3. The same operation as in Example 1 was carried out except that Og (twice the molar amount as the amino group of DEAE-chitin relative to the carboxyl group in the latex) was added.
0.81 g of AE-chitin modified latex was obtained. The surface charge amount of this product was 21.6 μequivalent/g·latex.

Example 8 1% DEAE-chitin aqueous solution and 1% chitosan aqueous solution 1.
Example 7 except that 1 g (twice the molar amount of the amino group of chitosan relative to the carboxyl group in the latex)
Perform the same operation as above to obtain chitosan-modified latex 0.97
I got g. The surface charge amount of this material is 4.3μ equivalent/g・
It was latex.

[Results of invention]

The present invention is a microparticle modified with a substance having a hydrophilic, preferably positively charged constituent element of chitin on the microparticle, which has a specific affinity for biopolymers and is therefore a bioaffinity component and a physiologically active substance. This is a useful invention that provides a substance that has an extremely wide range of uses, such as fixation of drugs, latex for immunodiagnostics, and endotoxin removal agent.

Claims (1)

[Scope of Claims] 1. Polymer fine particles whose surface is modified with a chitin derivative or chitosan. 2. The polymer fine particles according to claim 1, wherein the chitin derivative is diethylaminoethyl chitin. 3. The polymeric fine particles according to claim 1, wherein the polymeric fine particles are carboxyl latex. 4. Carboxyl latex fine particles have the formula RN=C=NR
' (in the formula, R is an ethyl group, R' is a dimethylaminopropyl group, or both R and R' are a cyclohexyl group), and then a chitin derivative or chitosan is reacted. A method for producing polymer fine particles whose surfaces are modified with a derivative or chitosan. 5. The method for producing fine polymer particles according to claim 4, wherein the chitin derivative is diethylaminoethyl chitin.