JP2009095773A - Method for producing hazardous substance removal material - Google Patents

Abstract

<P>PROBLEM TO BE SOLVED: To provide a method for manufacturing a harmful-substance removing material which contains a carrier carrying a chicken egg antibody and can be reserved for a long time. <P>SOLUTION: The method for manufacturing the harmful-substance removing material containing a carrier carrying the chicken egg antibody includes a process of filtering a solution containing the chicken egg antibody with a filter of a size of 0.5 μm or less and a process of adhering the solution after filtration to the carrier. <P>COPYRIGHT: (C)2009,JPO&INPIT

Description

  The present invention relates to a method for producing a hazardous substance removing material. More specifically, the present invention relates to a method for producing a hazardous substance removing material including a carrier carrying a chicken egg antibody that can be stably stored for a long period of time, and a harmful substance removing material obtained by the production method.

The antibody-carrying filter is useful as a material for removing viruses and bacteria in the liquid phase and gas phase. Since this filter uses an antibody that is easily denatured depending on temperature and humidity conditions, means for ensuring the long-term stability of the filter in use or the filter stored for replacement is desired. In general, for long-term storage, there are means for performing electron beam sterilization, gamma ray sterilization, gas sterilization after enclosing in a storage package. When such means are applied to an antibody-carrying filter, May cause denaturation and deterioration.
In Patent Document 1, a hazardous substance removing material carrying an antibody that removes a harmful substance in a gas phase atmosphere is stored in a sealed manner so that the storage atmosphere is 18 to 25 ° C. and humidity is 40% or less for long-term storage. There is a description of a method, coexistence of an oxygen adsorbent, and purging with an inert gas or nitrogen gas.
Patent Document 2 describes that a liquid in which a defatted egg yolk powder containing an antibody for immersing a nonwoven fabric is filtered is used for producing an air purification filter using an antibody substance. However, there is no suggestion regarding anti-corruption, and detailed conditions for filtration are not disclosed.
JP 2004-313755 A Japanese Patent Laid-Open No. 2003-210558

An object of the present invention is to provide a method for producing a hazardous substance removing material capable of long-term stable storage, which includes a carrier carrying a chicken egg antibody.

As a result of diligent research to solve the above problems, the present inventors have examined the pretreatment of the solution or carrier for supporting the egg egg antibody, and the package for storage, so that the antibody-carrying filter The inventors have found a condition for improving the storage stability, and have completed the present invention based on this finding.
That is, the present invention provides the following [1] to [5].

[1] A hazardous substance removing material comprising a carrier carrying a chicken egg antibody, comprising filtering a solution containing a chicken egg antibody with a filter having a pore size of 0.5 μm or less and attaching the filtered solution to a carrier. Production method.
[2] filtering the solution containing the egg antibody with a filter having a pore size of 0.5 μm or less,
A method for producing a harmful substance removing material comprising a carrier carrying a chicken egg antibody, the method comprising sterilizing a carrier with radiation or gas and adhering the filtered solution to the carrier after sterilization.
[3] A hazardous substance removing material obtained by the production method according to [1] or [2].
[4] The hazardous substance removing material according to [3], which is packaged in a package that does not transmit ultraviolet rays.
[5] The hazardous substance removing material according to [4], wherein the previous package is a package including an aluminum foil or an aluminum deposited film.

According to the present invention, there is provided a method for producing a harmful substance removing material including a carrier carrying a chicken egg antibody and capable of long-term stable storage.

Hereinafter, the present invention will be described in more detail.
The carrier for supporting the chicken egg antibody is not particularly limited, but a carrier composed of fibers is preferable, and the carrier can be composed of a woven fabric, a nonwoven fabric, or the like. Examples of the fibers include, but are not limited to, synthetic fibers such as polyester, cellulose ester, polyvinyl alcohol, vinylon, acrylic, polyurethane, polyamide, natural fibers such as cotton, silk, wool, and regenerated fibers such as rayon. I can do it. These fibers may be combined. For the purpose of improving the strength and dimensional stability, the carrier may be reinforced with other suitable structural materials such as metals, polymer materials, and ceramics. These reinforcing materials are desirably used for portions other than the substantially outermost surface of the side surface to which the harmful substance removing material is supplied (for example, used on the opposite surface of the side surface or a core material).

  The fiber used as a carrier in the present invention can be produced by a general production method such as melt spinning, wet spinning, dry spinning, wet drying spinning, or physical treatment (for example, strong mechanical shearing treatment using an ultra-high pressure homogenizer). In order to ensure stable quality, it is preferable to use dry spinning or wet-dry spinning. In order to produce uniform fibers with an average fiber diameter of 100 nm or less, further processing techniques, 2005, 40, No. 2, 101, and 167; Polymer International, 1995, 36, 195- 201; Polymer Preprints, 2000, 41 (2), 1193; Journal of Macromolecular Science: Physics, 1997, B36, 169, etc. It is preferable to employ the electrospinning method.

  Solvents used for spinning include chlorinated solvents such as methylene chloride, chloroform and dichloroethane, amide solvents such as dimethylformamide, dimethylacetamide and N-methylpyrrolidone, and ketones such as acetone, ethyl methyl ketone, methyl isopropyl ketone and cyclohexanone. Any solvent that dissolves the resin used for the synthetic resin fiber, such as a solvent, an ether solvent such as THF or diethyl ether, an alcohol solvent such as methanol, ethanol, or isopropanol, or water can be used. These solvents may be used alone or in combination of two or more.

  When the electrospinning method is employed, a salt such as lithium chloride, lithium bromide, potassium chloride, or sodium chloride may be further added to the resin solution.

  It is desirable that the fibers constituting the carrier have a structure in which a three-dimensional network is formed by partial adhesion. By adopting such a structure, it is possible to improve processing and practical mechanical resistance, and to improve the reliability of the hazardous substance removing material. Moreover, the retention property of an antibody can be improved. The adhesion between fibers can be observed by a method such as SEM. The density of the bonding points between the fibers is preferably 10 or more per 1 mm square with respect to the projected surface area of the harmful substance removing material, and more preferably 100 or more.

  As a method for forming an adhesion point, it may be formed by an adhesion formed by a dry spinning method or a fusion point formed by a melt spinning method, or after spinning, addition of an adhesive, a plasticizing solvent, etc. You may perform the adhesion point formation process by. From the viewpoint of production cost, it is preferable to form adhesion points by a dry spinning method using an appropriate solution formulation.

In the production method of the present invention, the carrier is preferably sterilized. The sterilization method is not particularly limited as long as the sterilization method does not deteriorate the carrier, but sterilization by radiation or sterilization by gas is preferable.
Examples of radiation sterilization methods include gamma sterilization and electron beam sterilization. Among these, the method using gamma ray sterilization is preferable.
Examples of the gas sterilization method include methods using ethylene oxide gas sterilization and chlorine dioxide gas sterilization. Among these, the method using ethylene oxide gas sterilization is preferable.

  The antibody used in the production method of the present invention is a protein that specifically reacts with a specific harmful substance (antigen) (antigen-antibody reaction), has a molecular size of 7 to 8 nm, and is a Y-shaped molecule. It has a form. Of the Y-shaped molecular structure of an antibody, a pair of branch parts is called Fab and a trunk part is called Fc, and among these, a toxic substance is captured at the Fab part.

  The type of the antibody corresponds to the type of harmful substance that can be captured. Examples of harmful substances captured by antibodies include bacteria, molds, viruses, allergens, and mycoplasmas. Specifically, examples of bacteria include gram-positive bacteria, Staphylococcus (S. aureus and Staphylococcus epidermidis), micrococcus, anthrax, cereus, Bacillus subtilis, acne, and gram-negative bacteria. And Pseudomonas aeruginosa, Serratia, Sephacia, Streptococcus pneumoniae, Legionella, and Mycobacterium tuberculosis. Examples of molds include Aspergillus, Penicillius, and Cladosporium. Examples of the virus include influenza virus, coronavirus (SARS virus), adenovirus, rhinovirus and the like. Examples of allergens include pollen, mite allergen, cat allergen and the like.

A chicken egg antibody can be obtained by purifying an egg yolk powder obtained by administering an antigen to a chicken to produce an immunized egg, and sterilizing and spray-drying the egg yolk liquid. The method of obtaining an antibody from chicken eggs can easily and in large quantities obtain an antibody, and can reduce the cost of a harmful substance removing material.

  As a method of supporting (immobilizing) the antibody on the carrier, in addition to a method of physically adsorbing the antibody on the carrier, the carrier is silanized using γ-aminopropyltriethoxysilane and the like, and then glutarylated. A method of introducing an aldehyde group onto the surface of the carrier with aldehyde or the like and covalently bonding the aldehyde group and the antibody, a method of immobilizing the antibody on the carrier by ionic bonding by immersing an untreated carrier in an aqueous solution of the antibody, a specific method A method of introducing an aldehyde group into a carrier having a functional group and covalently bonding the aldehyde group and the antibody, a method of ion-binding the antibody to a carrier having a specific functional group, and coating the carrier with a polymer having a specific functional group A method of introducing an aldehyde group later and covalently binding the aldehyde group to the antibody can be mentioned.

Here, examples of the specific functional group include an NHR group (R is any alkyl group other than H, methyl, ethyl, propyl, and butyl), NH ₂ group, C ₆ H ₅ NH ₂ group, and CHO group. , COOH group, and OH group.

  Also, there is a method in which the functional group on the surface of the carrier is converted to another functional group using BMPA (N-β-Maleimidopropionic acid) or the like, and then the functional group and the antibody are covalently bonded (SH group in BMPA). Are converted to COOH groups).

  Furthermore, there is a method in which a molecule (Fc receptor, protein A / G, etc.) that selectively binds to the Fc portion of the antibody is introduced onto the surface of the carrier and the antibody Fc is bound thereto. In this case, the Fab that captures the harmful substance faces outward with respect to the carrier, and the probability of contact of the harmful substance with the Fab increases, so that the harmful substance can be efficiently captured.

The antibody may be supported on a carrier via a linker. In this case, the degree of freedom of the antibody on the carrier is increased and access to harmful substances is facilitated, so that high removal performance can be obtained. Examples of the linker include bi- or higher-valent cross-linking reagents, specifically maleimide, NHS (N-Hydroxysuccinimidyl) ester, imide ester, EDC (1-Ethyl-3- [3-dimethylaminopropyl] carbodiimido), There is PMPI (N- [p-Maleimidophenyl] isocyanate), which is selective to target functional groups (SH group, NH ₂ group, COOH group, OH group) and non-selective. Moreover, the distance (space arm) between crosslinks also changes for every crosslink reagent, and it can select in the range of about 0.1 nm-3.5 nm according to the target antibody. From the viewpoint of efficiently capturing harmful substances, those that bind to the Fc of the antibody as a linker are preferred.

  As a method for introducing a linker, either a method in which a linker is bound to an antibody and then further bound to the antibody, or a method in which a linker is bound to a carrier and the antibody is bound to the linker on the carrier is possible. is there.

In the production method of the present invention, the antibody is supported on the carrier by attaching a solution containing the chicken egg antibody to the carrier. The method for attaching the chicken egg antibody solution to the carrier is not particularly limited, and examples thereof include a method of immersing the carrier in the chicken egg antibody solution, spray coating, ink jet coating, and curtain coating.
What is necessary is just to produce the solution containing a chicken egg antibody by melt | dissolving the chicken egg antibody obtained as mentioned above in physiological saline or phosphate buffered saline. The concentration of the chicken egg antibody in the solution may be 0.001% by mass to 10% by mass, preferably 0.005% by mass to 5% by mass, and more preferably 0.01% by mass to 1% by mass. The solution may contain additives such as stabilizers, antibacterial agents and antifungal agents.

In the production method of the present invention, a solution containing a chicken egg antibody is used after being filtered through a filter having a pore size of 0.5 μm or less. The filter for filtration is more preferably 0.3 μm or less. By filtering with a filter having a pore size of 0.5 μm or less, it is possible to suppress the growth of germs and molds in the produced harmful substance removing material.

The hazardous substance removing material obtained by the production method of the present invention can remove harmful substances in the gas phase or liquid phase. This harmful substance removing material can be used even in a dry environment where the antibody faces the gas phase, and can be used for a filter for an air cleaner, a mask, a wipe sheet, and the like. The hazardous substance removing material obtained by the production method of the present invention can be stably stored for a long period of time.
The hazardous substance removing material obtained by the production method of the present invention is preferably packaged using a package that does not transmit ultraviolet rays. By using such a package, it is possible to prevent the deterioration of the antibody due to ultraviolet rays, and further stable storage for a long period of time is possible. As a package, an aluminum foil film and an aluminum vapor deposition film are preferable, and a multilayer film containing these is also preferably used.

The present invention will be described more specifically with reference to the following examples. The following embodiments can be modified as appropriate without departing from the spirit of the present invention. Therefore, the scope of the present invention is not limited to the following specific examples.
<Preparation of antibody filter>
<Production of carrier>
(Carrier N-1)
Acetone: water (97: 3) solution (25% by mass) of cellulose acetate (manufactured by Aldrich, total substitution degree 2.4, number average molecular weight 30,000) is heated to 60 ° C., and from a nozzle having a diameter of 0.1 mm, A nonwoven fabric was formed by jetting together with air at a spinning speed of 500 m / m to obtain a nonwoven fabric carrier N-1 having a thickness of 85 μm. The spinning cylinder was heated to 100 ° C. with a heater. It was 8 micrometers when the average fiber diameter was measured by SEM.
(N-2)
The nonwoven fabric N-1 was sterilized with gamma rays to obtain a sterilized nonwoven fabric carrier N-2.

<Preparation of antibody solution>
The yolk solution of the immunized egg produced by the chicken administered with the antigen was spray-dried to obtain a dried egg yolk powder. Next, the dried egg yolk powder was degreased with ethanol to remove the degreasing component, and then dried under reduced pressure to obtain a degreased egg yolk powder as an antibody substance. When this defatted egg yolk powder was refine | purified and the purity of the influenza virus antibody (IgY antibody) was measured, it was 3 mass%. Next, the defatted egg yolk powder was dissolved in phosphate buffered saline (PBS) and adjusted to an antibody concentration of 100 ppm. Antibody solution (K-1) filtered through a filter having a pore size of 0.22 μm, antibody solution (K-2) filtered through a filter having a pore size of 0.45 μm, antibody solution filtered through a filter having a pore size of 1 μm ( K-3), an antibody solution (K-4) that was not filtered was prepared.

<Immobilization of antibody>
(Antibody filter F-1)
The carrier N-1 was immersed in the antibody solution K-1 at room temperature for 16 to 24 hours to give the antibody to the fiber surface. The obtained sample was allowed to stand for 24 hours in an environment of 25 ° C. and 20% RH, and then allowed to stand for 24 hours in an environment of 25 ° C. and 90% RH. This operation was repeated 3 times alternately for a total of 6 conditions.

(Antibody filter F-2)
Antibody filter F-2 was produced in the same manner as F-1, except that the antibody solution was changed from K-1 to K-2.
(Antibody filter F-3)
Antibody filter F-3 was produced in the same manner as F-1, except that the carrier was changed from N-1 to N-2.

(Antibody filter F-4)
Antibody filter F-4 was produced in the same manner as F-1, except that the antibody solution was changed from K-1 to K-3.
(Antibody filter F-5)
Antibody filter F-5 was produced in the same manner as F-1, except that the antibody solution was changed from K-1 to K-4.

<Forced rot test>
The antibody filters F-1 to F-5 were stored in a dark room at 25 ° C. and 80% humidity for 2 weeks, 1 month, 3 months, and 6 months, and then visually checked for appearance.

As is clear from Table 1, the antibody filter obtained by the production method of the present invention does not rot for a long time even in a high humidity environment.

<Packaging of antibody filters>
(Packaged antibody filter H-1)
The antibody filter F-1 was enclosed in a packaging bag (strong pack ALH made by Meiwa Packs) containing a moisture-proof and light-shielding aluminum foil film.
(Packaged antibody filter H-2)
The antibody filter F-2 was enclosed in a packaging bag (strong pack ALH made by Meiwa Packs) containing a moisture-proof and light-shielding aluminum foil film.
(Packaged antibody filter H-3)
The antibody filter F-4 was sealed in a packaging bag (strong pack ALH made by Meiwa Packs) containing a moisture-proof and light-shielding aluminum foil film.
(Packaged antibody filter H-4)
The antibody filter F-1 was sealed in a moisture-proof and transparent packaging bag (Strong Pack BX, manufactured by Meiwa Packs).

<Storage under UV irradiation>
The packaged antibody filters H-1 to H4 were irradiated with ultraviolet rays of 100 μW / cm ² for 2 weeks in a storage atmosphere at 25 ° C. and a humidity of 60%. The ultraviolet intensity was measured with a UV-power meter (C9536-01 / H9958) manufactured by Hamamatsu Photox.

<Evaluation of virus inactivation efficiency>
Evaluation of virus inactivation of the antibody filter after storage under ultraviolet irradiation was performed.
As the test virus solution, purified influenza virus diluted 10 times with PBS (virus concentration 200,000 plaques / mL) was used. Each of the samples was cut into 5 cm squares and attached and fixed in the center of the virus spray test apparatus. The test virus solution was put in a nebulizer installed on the upstream side, and a virus recovery device was attached on the downstream side. Compressed air was sent from an air compressor, and the test virus was sprayed from the nebulizer spray port. A gelatin filter was installed on the downstream side of the mask, and air in the test apparatus was sucked at a suction flow rate of 10 L / min for 5 minutes to collect passing virus mist.
After the test, the gelatin filter collecting the virus was collected, and the virus infectivity after passing through the sample was determined by the TCID50 method (50% cell infectious dose measurement method) using MDCK cells. From the comparison of the virus infectivity value of the gelatin filter with and without the sample, the transient removal rate of the virus of each sample was calculated.

As is apparent from Table 2, the packaged antibody filter of the present invention does not deteriorate the antibody due to ultraviolet rays, and has the ability to inactivate viruses even after long-term storage.
Furthermore, when H-1 and H-2 are compared with H-3, the transient removal rate of virus is significantly high in H-1 and H-2, and the antibody solution to be attached is filtered with a 0.5 μm filter. It can be seen that the antibody filter has an improved virus removal rate.

Claims (5)

A method for producing a harmful substance removing material comprising a carrier carrying a chicken egg antibody, comprising filtering a solution containing a chicken egg antibody with a filter having a pore size of 0.5 μm or less and adhering the filtered solution to a carrier. Filtering the solution containing the egg antibody with a filter having a pore size of 0.5 μm or less,
A method for producing a harmful substance removing material comprising a carrier carrying a chicken egg antibody, the method comprising sterilizing a carrier with radiation or gas and adhering the filtered solution to the carrier after sterilization. A hazardous substance removing material obtained by the production method according to claim 1. The hazardous substance removing material according to claim 3, which is packaged in a package that does not transmit ultraviolet rays. The hazardous substance removing material according to claim 4, wherein the first package is a package including an aluminum foil or an aluminum vapor deposition film.