JPH0441180B2 - - Google Patents

Description

DETAILED DESCRIPTION OF THE INVENTION [Field of Industrial Application] The present invention relates to a method for modifying the surface of polymeric materials.

[Prior art]

Currently, various polymeric materials such as polyethylene, polyethylene terephthalate, polytetrafluoroethylene, and silicone have been used as various medical materials. Among these, silicone rubber, which is chemically inert, is the most widely used as an implantation material in living organisms.The hydrophobic nature of its surface causes encapsulation, which means that the implantation material is absorbed by surrounding biological tissue. It has disadvantages such as enveloping phenomenon and poor adhesion to living tissue.

In order to improve these disadvantages, it has been proposed to make the surface of silicone rubber hydrophilic, to perform collagen treatment, etc., but a practically satisfactory method has not yet been found. For example, in a method in which a radically polymerizable monomer that provides a hydrophilic polymer is brought into contact with the surface of a material made of silicone rubber, and radiation is irradiated in this state, the polymerization of the monomer is caused by the polymerization of the material of the monomer. Since the grafting proceeds simultaneously with the grafting on the surface of the material and also along the surface of the material, the absolute amount of the polymer formed is limited and it is difficult to obtain a sufficient modifying effect. Furthermore, in the method using collagen treatment, the adhesion of collagen to the surface of the material was low and the amount of adhesion was insufficient, and as a result, the modification effect was naturally limited.

[Purpose of the invention]

The present invention has been made based on the above circumstances, and its purpose is to modify the surface of a polymer material so that the surface has extremely suitable compatibility with living organisms. We are here to provide you with a possible method.

[Structure of the invention]

The features of the present invention include the step of activating the surface of a polymeric material by electrical discharge treatment, and the step of contacting the surface of the activated polymeric material with one or more radically polymerizable monomers. The method includes a step of graft polymerization by graft polymerization, and a step of immobilizing the protein on the surface of the graft polymerized polymer material.

The present invention will be explained in detail below.

In the present invention, the surface of a polymeric material is basically modified through the following three steps to make it biocompatible.

(1) First step (discharge treatment step) In this step, discharge is performed with the polymer material whose surface is to be modified exposed in an appropriate discharge space, and the surface is exposed to the discharge. to activate the surface. Of course, the surface of the polymer material is usually subjected to a surface cleaning treatment prior to this discharge treatment step.

In the present invention, examples of the polymer material include silicone rubber, low density polyethylene, high density polyethylene, linear low density polyethylene, ethylene-vinyl acetate copolymer, or a completely or partially saponified product thereof, polypropylene, polypropylene copolymer So-called vinyl polymers such as polyvinyl chloride, polyvinylidene chloride, polystyrene, polyacrylonitrile, polytetrafluoroethylene, or polyethylene terephthalate,
Examples include so-called polycondensates such as polyethylene isophthalate, nylon 6, nylon 66, and nylon 12, polyadducts such as polyurethane, and natural polymeric substances such as cellulose and wool, which are themselves biocompatible. It is preferable to use silicone rubber, and silicone rubber is particularly preferable.

Further, the state of discharge and the mode of treatment are not particularly limited, and the polymer material may be activated by the generation of peroxide on the surface of the polymer material, and sufficient graft bonds may be formed in the next second step. All you have to do is get the status. Specifically, a discharge state such as plasma discharge, corona discharge, glow discharge, or ionizing irradiation may be formed, and the surface of the polymer material may be brought into contact with or irradiated with this state. The discharge conditions, treatment time, etc. are appropriately selected depending on the type of polymer material and other factors.

(2) Second step (graft polymerization step) In this step, 1
A species or two or more radically polymerizable monomer pairs are brought into contact and polymerized. This forms a monomeric polymer that is grafted onto the surface of the polymeric material.

For contacting the monomer, a method of applying a solution of the monomer to the surface is common, but it is not limited to this method, and there is a possibility that a gaseous monomer can be used. There is also. For polymerization, it is sufficient to create conditions that allow the monomer to polymerize; for example, heating is performed at a temperature of about 40 to 100°C.

The term "radical polymerizable monomer" as used herein refers to a compound having a carbon-carbon double bond, and is a monomer that polymerizes with radicals as propagating terminals in a chain mechanism, such as styrene, sodium p-styrene sulfonate, etc. Styrene compounds, maleic anhydride,
Maleic acid compounds such as dimethyl maleate, itaconic acid, itaconic acid compounds such as dimethyl itaconate, acrylamide, 2-acrylamide
Acrylic acid compounds such as 2-methylpropanesulfonic acid, acrylic acid, acrylic acid compounds such as methyl acrylate, methyl methacrylate, 2-methylpropanesulfonic acid, etc.
- Methacrylic acid compounds such as hydroxyethyl methacrylate, allyl compounds such as diallylamine and allyl alcohol, 2-vinylpyridine, N
Examples include vinyl compounds such as -vinyl-2-pyrrolidone and vinyl acetate. However, in the present invention, as described below, the polymer has active sites capable of immobilizing proteins, or becomes capable of having such active sites through appropriate treatment. In this respect, when the protein is collagen, acrylic acid, acrylamide, 2-
Hydroxyethyl methacrylate, N-vinylpyrrolidone and other monomers are important, with acrylic acid and acrylamide being particularly preferred. Acrylic acid has a carboxyl group that serves as an active site, and acrylamide easily becomes a carboxyl group by hydrolysis.

In addition, in this step, it is desirable that homopolymers formed simply by polymerization of monomers without being grafted onto the surface of the polymer material are removed by an appropriate cleaning treatment or the like.

In addition, when the polymeric material is silicone rubber, it is preferable that the monomer is sufficiently adhered to the surface prior to polymerization treatment, and for this purpose, it is desirable to perform a degassing treatment, for example by placing it under a vacuum. .

(3) Third step (protein immobilization step) In this step, proteins are immobilized on the surface of the polymer material having the polymer grafted in the second step. Prior to fixation of this protein,
If necessary, activation treatment of the polymer is performed.
This treatment gives the polymer necessary or desirable properties for protein immobilization, and specifically, it involves chemical treatment, heating, light irradiation, etc. in order to introduce or generate functional groups that will become active sites. Physical processing takes place.

Specific examples of proteins used for immobilization include collagen, fibrinogen, fibrin, fibronectin, and others, and among these, collagen is preferred in terms of its good biocompatibility.

Specific methods for protein fixation include:
Known methods used for enzyme immobilization can be used as they are or with some modifications; for example, a carbodiimide activation method or a promosyanide activation method in which immobilization is carried out by covalent bonding to a grafted polymer. , methods of immobilization by multiple ionic bonds, and others can be advantageously used.

〔Effect of the invention〕

The method of the present invention is as described above, and in the first step, for example, peroxide is generated and activated on the surface of the polymer material by electric discharge treatment, and in the second step, peroxide is generated and activated on the surface of the polymer material.
In the process, the radically polymerizable monomer is grafted onto the surface of the activated polymer material and polymerized, so the produced polymer is sufficiently and stably fixed to the surface of the polymer material, and each The molecules extend their chains outward from the surface. In the third step, proteins such as collagen are bonded to the polymer through covalent bonds or polyion complexes, resulting in a polymeric material that has more effective biocompatibility over a long period of time, such as high cell adhesion. can be obtained.

In the above process, in the first step, the polymeric material is activated by electric discharge treatment, so even if the polymeric material itself is chemically stable silicone rubber, the surface can be reliably activated. As a result, the subsequent second and third steps can be reliably executed without requiring special considerations.

Therefore, by using silicone rubber as a polymeric material in the present invention, in addition to the fact that the silicone rubber itself has high biocompatibility, it is possible to obtain biological materials and medical materials that are extremely useful in practice. .

〔Example〕

Examples of the present invention will be described below, but the present invention is not limited thereto.

Example 1 Using a silicone rubber sheet vulcanized with 2,4-dichloro-benzoyl peroxide as a material, this rubber sheet was placed between discharge electrodes,
A discharge treatment was performed for 2 minutes by applying a voltage of 60 Hz and 7 KV to the discharge electrode to generate corona discharge. Through this discharge treatment, peroxide groups were introduced into the surface of the sheet in an amount of 1.0×10 ⁻⁹ mol/cm ² . Next, this discharge-treated rubber sheet was vacuum degassed, and then immersed in a 10% by weight aqueous solution of acrylic acid that had been thoroughly degassed.
The mixture was heated for 1 hour for graft polymerization.

The obtained rubber sheet was thoroughly washed with water to remove the produced homopolymer, thereby obtaining an acrylic acid graft polymerized silicone rubber. When the graft amount was determined by conductivity titration, it was 27 μg/
It was warm in ^cm2 .

This rubber sheet was then coated with a solution of a water-soluble carbodiimide in physiological phosphate buffer (pH 7.4, active ingredient
1mg/ml) at 0℃ for 30 minutes,
Add fibronectin to the same physiological phosphate buffer solution (active ingredient 0.05 mg/ml) at 0°C.
for 2 hours, and unfixed fibronectin was removed by ultrasonic washing. The amount of immobilized fibronectin on the obtained modified rubber sheet was determined by the ninhydrin method and was found to be 3 μg/cm ²
It was hot.

Example 2 The same silicone rubber sheet as in Example 1 was used as the polymer material, and corona discharge treatment was performed for 2 minutes in the same manner except that the voltage was 9 KV. After the rubber sheet obtained here was vacuum degassed, it was immersed in a 10% acrylamide aqueous solution and further heated to a temperature of 50%.
Graft polymerization was carried out by heating at ℃ for 1 hour. The amount of graft obtained here was 80 μg/cm ² .
Next, this rubber sheet was immersed in an aqueous sodium hydroxide solution with a concentration of 0.5N and hydrolyzed at a temperature of 50°C for 10 minutes, converting a portion of the acrylamide into acrylic acid.

Next, in the same manner as in Example 1, the sheet was activated with carbodiimide, and then immersed in a collagen aqueous solution to fix collagen, thereby obtaining a silicone rubber sheet with a modified surface. This modified rubber sheet had 4 μg/cm ² of collagen immobilized thereon.

Further, the above-mentioned operation was repeated in exactly the same manner except that the acrylamide was not hydrolyzed or the hydrolysis time was changed to 60 minutes to obtain a modified rubber sheet. When the fixed amount of collagen was determined, it was 0 μg/cm ² for the collagen that was not hydrolyzed, and 5 μg/cm ² for the collagen that had been hydrolyzed for 60 minutes.

Application example 1 Using the modified silicone rubber sheet obtained in Example 2 with a fixed amount of collagen of 4 μg/cm ² as a substrate,
HeLa cells were cultured. The medium contains sodium bicarbonate and L-glutamine.
A mixture of Eagle'sMEM and fetal bovine serum at a concentration of 10% was used. Then, cells were suspended in this medium at a ratio of 1 x 10 ⁴ cells/ml, and this cell suspension was placed at 1 x 10 ⁴ cells in a plastic jar in which a collagen-immobilized silicone rubber sheet had been placed in advance. It was added at a rate of 2 pieces/cm ² . Then, it was incubated for 4 hours at a temperature of 37° C. in a 5% carbon dioxide atmosphere. Thereafter, the sheet was washed with physiological phosphate buffer, and the number of cells attached to the sheet was counted, and the number of attached cells was 1.5×10 ⁴ cells/cm ² .

On the other hand, when the same cell experiment as above was carried out using the silicone rubber sheet as a substrate without any modification by the method of the present invention, the number of adhered cells was less than 1×10 ² cells/cm ² .

From the above facts, it is clear that according to the method of the present invention, the surface of a polymeric material can be modified to significantly improve the adhesion of biological tissues (cells).

Example 3 Using the same silicone rubber sheet as in Example 1 as a polymeric material, this was placed between the discharge electrodes in a vacuum container, and the pressure inside the vacuum container was reduced to 0.03 to 0.05 mmHg, and argon gas was supplied at a flow rate of 20 cc/ Supply at a rate of 1 minute, in this state the frequency is 5KHz, the output
A discharge treatment was performed for 5 seconds by applying a high frequency of 0.08 W/cm ² to generate plasma discharge. Thereafter, the rubber sheet was vacuum degassed, and acrylamide was graft-polymerized in the same manner as in Example 2. After the acrylamide was hydrolyzed for 30 minutes, collagen was fixed to obtain a modified silicone rubber sheet. This modified rubber sheet had 4 μg/cm ² of collagen fixed thereon.

Claims (1)

[Claims] 1. A step of activating the surface of a polymeric material by electrical discharge treatment, and contacting the surface of the activated polymeric material with one or more radically polymerizable monomers. 1. A method for surface modification of a polymeric material, comprising a step of graft polymerization and a step of fixing a protein on the surface of the graft-polymerized polymeric material. 2. The method for surface modification of a polymeric material according to claim 1, wherein the polymeric material is made of silicone rubber and the protein is collagen.