DESCRIPTION

HAZARDOUS SUBSTANCE-REMOVING MATERIAL AND METHOD OF REMOVING HAZARDOUS SUBSTANCES

Technical Field

The present invention relates to a hazardous substance-removing material and to a method of removing hazardous substances. More particularly, the present invention relates to a highly stable hazardous substance-removing material which comprises a support having antibodies supported thereon, and to a method of removing hazardous substances employing the same.

Background Art

Antibody-supporting filters are desirable materials for removing viruses and various bacteria from liquid and gas phases. In antibody-supporting filters, denaturation of the antibodies compromises filter performance. Therefore, employing such filters to remove the bacteria and the like in a gas phase in particular requires a highly stable antibody-supporting filter in which denaturation of the antibodies is prevented.

Japanese Patent No. 3,642,340 relates to a hazardous substance-removing material which comprises a support having antibodies supported thereon, and describes storage under prescribed conditions of temperature and humidity to maintain antibody activity, and storage by immersion in water containing an activity-stabilizing agent such as glycerol. Both are attempts to maintain activity by maintaining humidity, and thus do not prevent denaturation of the antibodies.

Japanese Unexamined Patent Publication (KOKAI) No. 2003-302404 discloses a method of stably immobilizing antibodies in the presence of other proteins, but this presents the risk of competitive adsorption on the protein support and physical adsorption inhibition.

Disclosure of the Invention

The present invention has for its object to provide a hazardous substance-removing material which comprises a support having antibodies supported thereon, in which the antibodies are highly stable.

The present inventors conducted extensive research into solving the above-stated problems, resulting in the discovery of antibody stabilizing agents that solved the above-stated problems. The present invention was devised on that basis.

That is, the present invention provides [1] to [16] below: [I] A hazardous substance-removing material which comprises a support having antibodies supported thereon, in which at least one member selected from the group consisting of sugars, amino acids, SH group-protecting agents, and surfactants is incorporated as an antibody-stabilizing agent.

[2] The hazardous substance-removing material according to [1], wherein said antibody-stabilizing agent is one or more sugars selected from the group consisting of glucose, galactose, fructose, saccharose, lactose, maltose, trehalose, maltotriose, raffinose, mannitol, and sorbitol.

[3] The hazardous substance-removing material according to [1], wherein said antibody-stabilizing agent is one or more amino acids selected from the group consisting of glycine, alanine, valine, leucine, serine, proline, lysine, and glutamic acid. [4] The hazardous substance-removing material according to [1], wherein said antibody-stabilizing agent is an SH group-protecting agent selected from the group consisting of 2-mercaptoethanol and dithiothreitol.

[5] The hazardous substance-removing material according to [1], wherein said antibody-stabilizing agent is 3-[(3-cholamidopropyl)dimethylammonio]-l-propane sulfonate (CHAPS).

[6] The hazardous substance-removing material according to any one of [1] to [5], wherein the antibody-stabilizing agent content is 1 to 100 weight percent of the antibody content.

[7] The hazardous substance-removing material according to [1] or [2], wherein said antibody-stabilizing agent is a disaccharide or trisaccharide and the antibody-stabilizing agent content is 1 to 1, 000-fold by weight the antibody content.

[8] The hazardous substance-removing material according to any one of [1] to [7], wherein said support is at least one polymer containing a carbonyl group and/or an ether group.

[9] The hazardous substance-removing material according to any one of [1] to [8], wherein said support comprises at least one vinylon-containing fiber.

[10] The hazardous substance-removing material according to any one of [1] to [9] wherein said support is a fiber with an average fiber diameter of 100 nm or less.

[11] The hazardous substance-removing material according to any one of [1] to [10] wherein said antibodies are bird egg-derived antibodies.

[12] The hazardous substance-removing material according to any one of [l] to [ll], wherein said antibodies are chicken egg antibodies.

[13] The hazardous substance-removing material according to any one of [1] to [11], wherein said antibodies are ostrich egg antibodies.

[14] The hazardous substance-removing material according to any one of [1] to [13], manufactured by immersing said support in a solution obtained by dissolving or dispersing said antibody and said antibody-stabilizing agent.

[15] The hazardous substance-removing material according to any one of [1] to [14], wherein said antibodies are in contact with a gas phase.

[16] A method of removing a hazardous substance by employing the hazardous substance-removing material according to any one of [1] to [15] to remove a hazardous substance from a gas phase.

Brief Description of the Drawings

Fig. 1 : A schematic of the electrostatic fiber-forming device employed in the examples of the present invention and comparative examples.

Explanation Numerals in the Drawings

11 Power source device

12 Syringe

13 Needle

14 Collector

15 Polymer solution

16 Nanofiber

Best Mode of Carrying Out the Invention

The present invention is described in greater detail below.

The hazardous substance-removing material of the present invention comprises a support having antibodies supported thereon, in which at least one member selected from the group consisting of sugars, amino acids, SH group-protecting agents, and surfactants is incorporated as an antibody-stabilizing agent. The incorporation of such an antibody-stabilizing agent prevents denaturation of the antibodies supported on the support and maintains the activity of the antibodies for extended periods without the antibodies being affected by the use environment.

The sugars that can be employed are not specifically limited, and examples include monosaccharides such as glucose, galactose, and fructose; disaccharides such as saccharose, maltose, trehalose, cellobiose, and lactose; oligosaccharides such as fructooligosaccharides, isomaltooligosaccharides, galactooligosaccharides, xylooligosaccharides, soy oligosaccharides, lactosucrose oligosaccharides, maltotriose, and raffinose; sugar alcohols such as erythritol, xylitol, mannitol, sorbitol, and maltitol; and polysaccharides such as starch, dextrin, and glycogen. Of these, glucose, galactose, fructose, saccharose, lactose, maltose, trehalose, maltotriose, raffinose, mannitol, and sorbitol are preferable. These sugars may be employed singly or in combinations of two or more. They may also be employed in combination with other antibody-stabilizing agents.

The sugars may be employed irrespective of starting material and manufacturing method. Sugars obtained by enzymatic degradation of starches (glucose, maltose, dextran, and the like), sugars obtained by starch reduction (sorbitol, maltitol), sugars obtained by enzymatic transfer of starches (trehalose, coupling sugar, and the like), palatinose obtained by enzymatic transfer of sucrose, palatinit obtained by reduction of sucrose, lactinol obtained by reduction of lactose, xylose obtained by enzymatic degradation of xylan, xylitol obtained by reduction of xylan, and the like can be employed without specific limitation.

The quantity of sugar that is preferably added in the present invention varies with the type of antibody, type of sugar, objective, and application. However, a quantity of 1 to 200 weight percent, preferably 2 to 100 weight percent, and more preferably 5 to 50 weight percent of the quantity (by weight) of antibody is generally added. When adding a disaccharide or trisaccharide, the quantity is 1 to 1, 000-fold, preferably 10 to 500-fold, and more preferably, 20 to 200-fold the quantity of antibody by weight. When the amount added falls below these quantities, the stabilizing effect is minor, and when the amount exceeds these quantities, there are problems in that antibody activity diminishes, or the viscosity increases and content control and handling become difficult.

The amino acids that can be employed are not specifically limited. Any natural amino acid or modified amino acid can be employed. Glycine, alanine, valine, leucine, serine, proline, lysine, and glutamic acid are preferably employed. These amino acids may be employed singly or in combinations of two or more. They may also be employed in combination with other antibody-stabilizing agents.

The quantity of amino acid that is preferably added in the present invention varies with the type of antibody, type of amino acid, objective, and application. However, a quantity of 1 to 50 weight percent, preferably 2 to 20 weight percent, and more preferably 5 to 10 weight percent of the quantity (by weight) of antibody is generally added. When the amount added falls below these quantities, the stabilizing effect is minor, and when the amount exceeds these quantities, there are cases where antibody activity diminishes and desired performance cannot be achieved.

The SH group-protecting agents that can be employed are not specifically limited. Examples include 2-mercaptoethanol, dithiothreitol, glutathione, and ethylenediamine tetraacetate. Of these, 2-mercaptoethanol and dithiothreitol are preferable. One or more of these SH group-protecting agents can be employed, and they can be employed in combination with other antibody-stabilizing agents. An SH group-stabilizing agent can be employed alone as the antibody-stabilizing agent. However, since there are many cases in which deactivation of the antibodies is compounded by causes other than the oxidation of SH groups, use in combination with another type of antibody-stabilizing agent such as a sugar, amino acid, or surfactant is preferable.

The quantity of SH group-protecting agent that is preferably added in the present invention varies with the type of antibody, type of SH group-protecting agent, objective, and application. However, a quantity of 1 to 20 weight percent, preferably 2 to 15 weight percent, and more preferably 5 to 10 weight percent relative to the quantity (by weight) of antibody will generally suffice. When the amount added falls below these quantities, the stabilizing effect is minor, and when the amount exceeds these quantities, there are problems in that antibody activity sometimes diminishes and an odor is sometimes generated.

Cationic, anionic, betaine, and nonionic surfactants can all be employed as surfactants. Examples of cationic surfactants include dodecyl trimethyl ammonium chloride and hexadecyl trimethyl ammonium chloride. Examples of anionic surfactants include sodium dodecyl sulfate, sodium laureth sulfate, and sodium cholate. Examples of betaine surfactants include palmitoyl lysolecithin and 3-[(3-cholamidopropyl)di- methylammonio]-l -propane sulfonate. Examples of nonionic surfactants include polyoxyethylene (7) decyl ether and polyoxyethylene (10) isooctyl phenyl ether. Betaine surfactants or nonionic surfactants are preferably employed as surfactants; the use of 3-[(3-cholamidopropyl)dimethylammonio]-l -propane sulfonate (CHAPS) is preferred. One or more surfactants may be employed, and they may be employed in combination with other antibody-stabilizing agents.

Since surfactants sometimes interfere in the antibody support step (inhibiting antibody adsorption and the like), during use (inhibiting the adsorption of hazardous substances), and in the course of evaluation (affecting the elution and cell toxicity of antibodies and captured hazardous substances, and affecting chromatography), greater care must be exercised than for other stabilizing agents in the selection of the type and quantity of surfactant employed.

The quantity of surfactant that is preferably added in the present invention varies with the type of antibody, type of surfactant, objective, and application. However, a quantity of 5 to 20 weight percent, preferably 8 to 15 weight percent, and preferably, more preferably 10 to 12 weight percent of the quantity (by weight) of antibody may be added. When the amount added falls below these quantities, the stabilizing effect is minor, and when the amount exceeds these quantities, there are problems in that antibody activity sometimes diminishes and the reaction between antigen and antibody is sometimes inhibited.

The support is not particularly limited, and fiber is preferable. Preferable examples of the fiber include a fiber comprising, as a main component, at least one selected from the group consisting of cellulose ester, vinylon, acrylic, and polyurethane. In addition, a fiber comprising, as a main component, polyamide is also preferable. According to the present invention, the term "main component" means a component that accounts for 25% or more in terms of mass fraction with respect to the total mass of fibers.

The term "cellulose ester" refers to a cellulose derivative obtained by esterifying a hydroxyl group of cellulose with an organic acid. Examples of an organic acid used for esterification include fatty carboxylic acids such as acetic acid, propionic acid, and butyric acid and aromatic carboxylic acids such as benzoic acid and salicylic acid. They may be used alone or in combination. The rate of substitution of a hydroxyl group of cellulose with an ester group is not particularly limited; however, it is preferably 60% or more.

According to the present invention, a cellulose acylate fiber is preferable among the group of main materials which form a support. The term "cellulose acylate" used herein refers to cellulose ester in which some or all of hydrogen atoms of a hydroxyl group of cellulose are substituted with an acyl group. Examples of an acyl group include an acetyl group, a propionyl group, and a butylyl group. In terms of structure, a single group among the above examples may be substituted, or two or more acyl groups may be subjected to mixed substitution. The total sum of degrees of acyl group substitution is preferably 2.0 to 3.0, more preferably 2.1 to 2.8, and particularly preferably 2.2 to 2.7. Among them, cellulose acetate, cellulose acetate propionate, or cellulose acetate butylate capable of achieving such degree of substitution is preferable, and cellulose acetate is most preferable. In general, it has been known that a solvent for cellulose acylate varies depending on the degree of esterification. It is also possible to produce a support with cellulose acylate having a high esterification rate in advance and then subject the support to alkali hydrolysis treatment or the like for hydrophilicization of the surface thereof.

It is possible to form a sufficiently practical hazardous substance removing material consisting of a cellulose acylate fiber. However, in order to further improve strength, dimensional stability, and the like, a support may be formed with a mixed fiber (e.g., polyester-based fiber / polyolefin-based fiber / polyamide-based fiber / acrylic-based fiber). When a mixed fiber is used, the mass fraction of a cellulose acylate fiber is preferably 50% or more and more preferably 70% or more.

According to the present invention, a polyamid fiber is preferable among the group of main materials which constitutes a support.

According to the present invention, the term "polyamide" refers to a fiber comprising a linear polymer having a chemical structure unit comprising an amide bond.

Among polyamides, a linear aliphatic polyamide, which is a combination of an aliphatic diamine such as ethylenediamine, 1-methylethylenediamine, 1,3-propylenediamine, or hexamethylenediamine and an aliphatic dicarboxylic acid such

as malonic acid, succinic acid, or adipic acid, is preferable. Nylon 66 is particularly preferable.

In addition to the above diamine and dicarboxylic acid, aliphatic polyamide comprising a single component or copolymer components selected from among the following examples can be used: lactams such as ε-caprolactam and laurolactam; aminocarboxylic acids such as aminocaproic acid and aminoundecanoicacid; and para-aminomethyl benzoic acid. Nylon 6 produced using ε-caprolactam alone is particularly preferable.

In addition to the above, the following may be used: an aliphatic polyamide in which cycloaliphatic diamine such as cyclohexanediamine, l,3-bis(aminomethyl)cyclohexane, or l,4-bis(aminomethyl)cyclohexane is partially or entirely used as a material aliphatic diamine; and/or an aliphatic polyamide in which cycloaliphatic dicarboxylic acid such as 1,4-cyclohexane dicarboxylic acid, hexahydroterephthalic acid, or hexahydroisophthalic acid is partially or entirely used as dicarboxylic acid.

Further, examples of the above polyamide further include a polyamide with decreased water absorbability and an improved elastic modulus in which aromatic diamine such as aliphatic paraxylylene diamine (PXDA) or metaxylylene diamine (MXDA) and aromatic dicarboxylic acid such as terephthalic acid are partially used as starting materials. Moreover, a polymer having a side chain comprising an amide bond such as polyacrylic acid amide, poly(N-methylacrylic acid amide), or poly(N,N-dimethylacrylic acid amide) may be used.

Among polyamides, nylon 66 or nylon 6 is most preferable. This is because the following properties of such polyamide are preferable to be used as the support of the present invention: appropriate hygroscopic properties derived from amide bonds; ease of inducing fiber axis orientation of a molecular chain comprising a long-chain fatty acid having an appropriate length that results in relatively high extensibility; a dynamic and kinetic tendency not to be melted due to high melting temperature and thermal capacity (resistance to melting); flexibility of a molecular chain comprising a long-chain fatty acid;

and a tendency to not cause fibrillation or kink band formation (such tendency being imparted as a result of formation of a hydrogen bond between amide bonds), that is to say, repetitive bending and stretching properties.

Likewise, in order to improve strength or dimensional stability, the support can be reinforced with another suitable structural material such as a metal, polymer, or ceramic. The reinforcement material is desirably employed on a portion other than the portion that is essentially the outermost surface of the side on which the hazardous substance-removing material is applied (for example, the reinforcement material is preferably employed on the opposite side, or used as a core material or the like).

According to the present invention, the term "vinylon" refers to a fiber comprising a linear polymer containing vinyl alcohol units (65% by mass or more) and having a moisture regain of less than 7% obtained at least 1 week after placement of such fiber in an environment at a temperature of 20°C and at a humidity of 65%. Such fiber may be obtained by formalizing a hydroxyl group of vinyl alcohol. Also, it may be a polymer obtained by subjecting a hydroxyl group to boric acid crosslinking or a non-formalized fiber subjected to a waterproof treatment by a known method such as an alkaline spinning method or a cooled gel spinning method. The above fiber may contain, as non- vinyl-alcohol-unit component, an ethylene chain or a vinyl acetate chain. However, it is preferably a fiber formed with a vinyl alcohol support. Further, it is most preferably a non-formalized fiber obtained by cooled gel spinning. This is because a non-formalized fiber has uniform properties and high degree of orientation/crystallization and thus excellent mechanical properties and reliability can be obtained.

In general, vinylon is superior to other fibers in terms of high strength, high elastic modulus, appropriate hydrophilicity, weather resistance, chemical resistance, adhesiveness, and the like. Thus, the preferable properties thereof can be used for the support of the present invention support.

According to the present invention, the term "acrylic" refers to a fiber comprising recurring units of an acrylonitrile group (mass percentage: 40% or more). Examples thereof include a homopolymer of acrylonitrile; a copolymer of acrylnitrile and

a nonionic monomer such as acrylic ester, methacrylic ester, or vinyl acetate; a copolymer of acrylonitrile and an anionic monomer such as vinylbenzenesulfonate or allylsulfonate; and a copolymer of acrylonitrile and a cationic monomer such as vinylpyridine or methylvinylpyridine.

In general, an acrylic fiber is produced by an organic solvent wet spinning method. In this method, when a spinning stock solution is formed into a coagulated thread in a coagulating bath, water as a coagulant is mixed with the spinning stock solution that is spinning-twisted from a nozzle and a spinning solvent is externally diffused from the spinning-twisted stock solution. At such time, water and an organic solvent (e.g., DMF or DMAc) are mutually diffused such that a polymer deposits, resulting in the formation of a line of coagulated thread having a structure in which many cavities are connected to each other in a net form. In addition, such thread is characterized by deformation of a fiber section caused by volume contraction as a result of diffusion of a solvent into a coagulating bath during coagulation and by formation of concave-convex portions as a result of macrofibril structure formation on the surface thereof. Such fine structure is preferable as a structure of a support used in the present invention in terms of an increase in specific surface area or the ease of antibody loading.

An acrylic fiber used in the present invention varies depending on the composition of a starting material polymer, a spinning method, post-treatment conditions during production, and the like. However, in general, a bulky fiber having appropriate hydrophilicity and high weather resistance can be obtained, which is advantageous.

The term "polyurethane" used in the present invention refers to a fiber comprising a linear synthetic polymer in which bonds between monomers or basic substrate polymer units are mainly urethane bonds. Preferably such fiber contains a polyurethane segment at a mass percentage of 85% or more. Preferably, such polyurethane is a block copolymer of segmented polyurethane comprising a soft segment that is soft and have a molecular weight of several thousands and a low melting point and a hard segment that is rigid and have high cohesion and a high melting point. For a soft segment, polyether such as polypropylene glycol or polytetramethylene glycol can be

used. For a hard segment, a urethane group formed with 4,4'-diphenylmethane diisocyanate, m-xylene diisocyanate, or the like can be used. Polyurethane is generally characterized by a high elasticity. Also, it is further characterized by good extensibility, high restoring force upon expansion and contraction, antidegradation properties better than those of rubber materials, formation into thin fibers, and the like, although the characteristics thereof vary depending upon differences in terms of a primary structure of a high-molecular chain such as the distribution and chemical structure of each segment and upon differences in terms of a secondary structure derived from different spinning conditions. Thus, when polyurethane is used as a support of the present invention, such characteristics can be utilized.

Further, the volume swelling degree in pure water at 20 ⁰ C of a fiber constituting the support used in the present invention is not less than 1.1% to less than 10%, preferably not less than 1.1% to less than 8%, particularly preferably not less than 1.1% to less than 6%, and most preferably not less than 1.1% to less than 5%. In addition, the volume swelling degree in pure water at 2O ⁰ C of a fiber according to the present invention refers to the volume swelling degree which is obtained by measuring the fiber density of each sample in a dried state before immersion of the sample in pure water at 2O ⁰ C for 1 hour and that after such immersion. Such values are obtained by the density gradient tube method (JIS-K7112).

Regarding mechanical and physical properties and dimensional stability of a fiber constituting a support, the tensile elastic modulus in a dried state is preferably 25% or more. The term "tensile elastic modulus in a dried state" used herein refers to the degree of elongation at break of a fiber in a tensile test at 20 ⁰ C ₅ provided that such fiber has been dried for a sufficiently long period of time. In general, a fiber having a tensile elastic modulus in a dried state of 10% or more is preferable for processing such as fabric formation. In order to prevent breakage upon filter processing or practical use (such breakage leading to reduction in filtration efficiency), the tensile elastic modulus is preferably 25% or more, more preferably 30% or more, and most preferably 35% or more.

The official moisture regain of the fiber constituting the support is preferably not less than 1.0% to less than 7%, more preferably not less than 3.0% to less than 6.5%, most preferably not less than 5.0% to less than 6.5%. When a disaccharide or trisaccharide is added, the official moisture regain is preferably not less than 1.0% to less than 7%, more preferably not less than 1.5% to less than 6.5%, most preferably not less than 2.0% to less than 6.5%. Within the above range of official moisture regain, the expression of the activity of a supported antibody and the mechanical strength, rigidity, dimensional change stability in a use environment (particularly humidity) of a support can be achieved. Further, a filter obtained therewith can exhibit high performance and reliability.

In addition, the term "moisture regain" used herein refers to an official moisture regain. The term "official moisture regain" refers to a moisture regain of a fiber that has been left in an environment at 20°C and at a relative humidity of 65% for long period of time. Moreover, when a fiber is a mixed fiber further containing a different fiber, the term refers to the official moisture regain of the total mixed fibers.

Preferably, the surface of a fiber constituting a support has fine concave-convex portions several tens nanometers to several micrometers in size. The shape of a concave-convex portion may be a three-dimensionally shaped groove or ridge which is formed in the direction parallel to the fiber direction or in the direction vertical to the same, that is to say, in a concentric direction with respect to the fiber axis. Such three-dimensionally shaped groove or ridge may exist at an arbitrary proportion or density, provided that an arbitrary angle is formed in the direction between the parallel direction and the vertical direction to the fiber direction. A sample obtained by a known method for cellulose acetate fiber spinning is known to have a fiber section having a variable chrysanthemum-like shape as a result of skin layer formation on the surface thereof and depression of a skin layer due to solvent drying. In a preferred embodiment, such concave-convex portions are used for the present invention.

The above fine concave-convex portions several tens nanometers to several micrometers in size may have holes and/or projections. Preferably, such holes or

projections have an average diameter of 50 nm to 1 μm. For example, such holes and projections can be formed by cavitation of a solution, or they can be formed in a spinning step of a method using a solution in which a fine dispersoid is dispersed (e.g., a mixture containing a slurry in which barium sulfate particles are dispersed), or in a subsequent step by a method involving hydrolysis of an acyl group, surface oxidation treatment, or the like (e.g., the exposure of a cellulose portion on the fiber surface with the use of an alkaline water solution followed by generation of microcraters by an enzyme treatment).

The average fiber diameter of a fiber used for the hazardous substance removing material of the present invention is preferably 50 μm or less, more preferably 10 μm or less, particularly preferably 1 μm or less, and most preferably 100 nm or less. When vinylon is used as the support, the average fiber diameter is preferably 50 μm or less, more preferably 45 μm or less, and most preferably 35 μm or less. The average fiber diameter of the present invention is obtained by measuring the diameters of fibers in arbitrarily selected 300 sites on a scanning electron microscope (SEM) image for observation and averaging the results by calculation.

As to the method for producing the fiber used in the present invention, there are typical production methods such as melting spinning, wet spinning, dry spinning, and dry-wet spinning, and methods in which the fiber is made fine by a physical process (such as strong mechanical shearing using an ultrahigh pressure homogenizer), although dry spinning or dry-wet spinning is preferably employed for obtaining a stable quarity of product. For producing an uniform fiber having an average fiber diameter of 100 nm or less, the electrospinning method disclosed in "Kakou Gijyutsu (Processing Technology)", 2005, Vol. 40, No.2, p.lOl and p.167; "Polymer International", 1995, Vol. 36, pp.195-201; "Polymer Preprints", 2000, Vol. 41(2), p.1193; "Journal of Macromolecular Science: Physics", 1997, B36, p.169; and the like is preferably used.

Regarding the solvent used for the spinning, any solvent may be used as long as it dissolves the resin used for synthetic resin fibers. Examples thereof include: chloride-based solvents such as methylene chloride, chloroform, and dichloroethane; amide-based solvents such as dimethylformamide, dimethylacetamide, and

N-methylpyrrolidone; ketone-based solvents such as acetone, ethyl methyl ketone, methyl isopropyl ketone, and cyclohexanone; ether-based solvents such as THF and diethyl ether; alcohol-based solvents such as methanol, ethanol, and isopropanol; and water. These solvents may be used either singularly, or in mixtures of a plurality of types thereof.

The resin solution used for the electrospinning method may be added with a salt such as lithium chloride, lithium bromide, potassium chloride, and sodium chloride.

Preferably, fibers constituting a support of the hazardous substance removing material of the present invention partially adhere to each other such that a structure forming a three-dimensional network is obtained. The use of such structure results in the improvement of mechanical tolerance upon processing or practical use, leading to the improvement of reliability of the hazardous substance removing material. Further, antibody-supporting properties can be improved. Adhesion between fibers can be observed by a method involving SEM or the like. The density of fiber adhesion points is preferably 10 adhesion points or more in a 1-mm square on the projected surface area of the hazardous substance removing material and preferably 100 adhesion points or more in the same.

Regarding a method for forming adhesion points, adhesion points may be formed by a dry spinning method or by a melt spinning method. After spinning, adhesion point formation treatment may be carried out by heating or adding an adhesive/plasticizing solvent or the like. In view of production cost, it is preferable to form adhesion points by a dry spinning method with the use of an appropriate solution formulation.

The fiber employed as support in the present invention may form a nonwoven cloth. The method of manufacturing the nonwoven cloth is not specifically limited. Depending on the objective and application, a web obtained by a dry method, wet screening method, melt-blown method, span bond method, flash fiber-forming method, or air-laid method can be used to manufacture nonwoven cloth by suitably combining methods of achieving strength by physical methods such as water stream entangling

methods, needle punch methods, or stitch bond methods; adhesion methods based on heat, such as thermal bonding methods; and bonding methods based on bonding agents, such as resin bonding.

The antibody used for the hazardous substance removing material of the present invention is a protein which is reactive (antigen-antibody reaction) specifically to a specific hazardous substance (antigen), has a molecule size of 7 to 8 nm, and is in a Y-shaped molecular form. A pair of branch portions of the antibody in the Y-shaped molecular form is called Fab, and a stem portion thereof is called Fc, among which the Fab portions capture the hazardous substance.

The type of the antibody corresponds to the type of the hazardous substance to be captured. Examples of the hazardous substance to be captured by the antibody include bacteria, fungi, viruses, allergens, and mycoplasmas. Specifically, the bacteria include, for example: the genus Staphylococcus (such as Staphylococcus aureus and Staphylococcus epidermidis), Micrococcus, Bacillus anthracis, Bacillus cereus, Bacillus subtilis, and Propionibacteήum acnes, as gram-positive bacteria; and Pseudomonas aeruginosa, Serratia marcescens, Burkholderia cepacia, Streptococcus pneumoniae, Legionella pneumophilia, and Mycobacterium tuberculosis, as gram-negative bacteria. The fungi include, for example, Aspergillus, Penicillins, and Cladosporium. The viruses include influenza viruses, coronavirus (SARS virus), adenovirus, and rhinovirus. The allergens include pollens, mite allergens, and cat allergens.

Examples of methods for producing the antibody include: a method in which an antigen is administered to an animal such as a goat, a horse, a sheep, and a rabbit, and a polyclonal antibody is refined from the blood thereof; a method in which spleen cells of an animal to which an antigen is administered and cultured cancer cells are subjected to cell fusion and a monoclonal antibody is refined from a culture medium thereof or from a humor (such as ascites) of an animal in which the fussed cells are implanted; a method in which an antibody is refined from a culture medium of genetically modified bacteria, plant cells, or animal cells to which antibody producing gene is introduced; a method in which a bird, particularly a chicken or ostrich, to which an antigen is administered is

allowed to lay an immune egg and a chicken antibody or ostrich antibody is refined from yolk powder obtained by sterilizing and spray-drying the yolk of the immune egg; and a method in which a liquid containing a chicken antibody or ostrich antibody is refined by purifying the yolk with defatting, ammonium sulfate fractionation, or dialysis. Of all the above methods, the method for obtaining the antibody from a chicken egg enables easy mass production of the antibody, reducing the cost of the hazardous substance removing material.

The antibody used for the hazardous substance removing material of the present invention is preferably an antibody derived from a bird egg, more preferably a chicken egg antibody or ostrich egg antibody.

Regarding methods for immobilizing (fixing) the antibody to the support, there are: other than a method in which the antibody is physically adsorbed to the support, a method in which, after a support is subjected to silane treatment using γ-aminopropyl-triethoxysilane or the like, an aldehyde group is introduced on the surface of the support by glutaraldehyde or the like, to effect a covalent bond between the aldehyde group and an antibody; a method in which an untreated support is immersed into an aqueous solution of an antibody to cause ion boding, thereby immobilizing the antibody to the support; a method in which an aldehyde group is introduced to a support having a specific functional group to effect a covalent bond between the aldehyde group and an antibody; a method in which a support having a specific functional group is ion-bonded to an antibody; and a method in which a support is coated with a polymer having a specific functional group, followed by an introduction of an aldehyde group to effect a covalent bond between the aldehyde group and an antibody.

Here, the specific functional group includes an NHR group (R represents an alkyl group of any of methyl, ethyl, propyl, and butyl except H), an NH ₂ group, a C ₆ HsNH ₂ group, a CHO group, a COOH group, and an OH group.

Moreover, there is also a method in which a functional group on the surface of the support is replaced with another functional group using BMPA (N-β-Meleimidopropionic acid) or the like, to effect a covalent bond between the

resultant functional group and an antibody (an SH group is replaced with a COOH group by BMPA).

Further, there is another method in which a molecule (such as an Fc receptor and a protein A/G) which is selectively bindable to the Fc portion of the antibody is introduced on the surface of a support, to cause it to be bonded to the Fc of the antibody. In this case, the Fab for capturing a hazardous substance are arranged outwards from the support to cause increase in contact possibility of the hazardous substance to the Fab, resulting in efficient capturing of the hazardous substance.

The antibody may be supported on the support through a linker. In this case, the degree of freedom of the antibody on the support increases, so that the antibody can readily reach the hazardous substance, attaining a high removal performance. Divalent or multivalent crosslinking reagent may be used as the linker. Specifically, there are listed maleimide, NHS (N-Hydroxysuccinimidyl) ester, imide ester, EDC (l-Ethyl-3-[3-dimetylaminopropyl] carbodiimido), and PMPI

(N-[p-Maleimidophenyl]isocyanete), which are selectively or non-selectively bonded to a target functional group (an SH group, an NH ₂ group, a COOH group, and an OH group). Moreover, crosslinking agents have different crosslinking distances (spacer arm), and therefore, the distance can be selected within the range between about 0.1 nm to about 3.5 nm according to the target antibody. In view of efficient capturing of the hazardous substance, the linker is preferably bindable to the Fc of the antibody.

Regarding the method for introducing the linker, either one of the following methods may be possible: a method in which a linker is bonded to an antibody, and then the resultant linker is further bonded to the antibody; and a method in which a linker is bonded to a support, and then an antibody is bonded to the linker on the support.

The antibody-stabilizing agent can be incorporated into the hazardous substance-removing material of the present invention by methods such as causing the antibody-stabilizing agent to physically adsorb onto or impregnate the support, or by methods of supporting the antibody-stabilizing agent on the above-described support. Examples of methods of supporting the antibody-stabilizing agent on the above-described

support include the method of causing a specific functional group in the support to form a covalent bond with the above material as set forth above, and the method of immobilizing the above material on the support by means of an ionic bond.

The method of manufacturing the hazardous substance-removing material of the present invention is not specifically limited. One example of the manufacturing method is to immerse the support in a solution containing the antibodies (support solution) to cause the antibodies to bond to the support. The antibody- stabilizing agent can be dissolved or dispersed in the support solution, or a solution in which the antibody-stabilizing agent has been dissolved or dispersed can be separately prepared and the support immersed in this solution before attaching the antibodies. Of these, it is preferable to employ a support solution containing both the antibodies and the antibody-stabilizing agent to make it possible to simultaneously attach the antibodies and antibody-stabilizing agent to the support.

The solution may contain antibacterial agents, antifungal agents, and the like.

The ratio of the contents of the antibody and antibody-stabilizing agent in the support solution can be generally thought of as reflecting the ratio of the contents of the antibodies and antibody-stabilizing agent supported on the support in the hazardous substance-removing material of the present invention.

The hazardous substance-removing material of the present invention can remove hazardous substances from gas and liquid phases. The hazardous substance-removing material of the present invention can be employed in air purifier filters, masks, wipes, and the like.

In the hazardous substance-removing material of the present invention, both antibody activity and antibody stability are high even in dry environments where the antibodies come into contact with a gas phase.

Examples

The present invention is described in greater detail below through embodiments. The embodiments described below can be suitably modified without departing from the

scope or spirit of the present invention. Accordingly, the scope of the present invention is not limited to the specific examples given below. [Example 1]

An acetone:water (97:3) solution (10 weight percent) of cellulose acetate (total degree of substitution 2.4, number average molecular weight 30,000, made by Aldrich) was employed to conduct electrostatic fiber formation with a syringe feed rate of 0.05 mm/minute at an applied voltage of 15 kV. The fiber was then dried for 8 hours at 8O ⁰ C under a vacuum to prepare fine nonwoven cloth with a film thickness of 50 micrometers. SEM measurement of the nonwoven cloth revealed an average fiber diameter of 80 nm. Influenza antibody (IgY antibody) prepared by purifying immunized eggs laid by a chicken to which antigen had been administered was dissolved in phosphate-buffered saline (PBS) and the antibody concentration of the solution was adjusted to 100 ppm. Saccharose was added and stirred to a concentration of 10 ppm and the mixture was passed through a 0.45 micrometer filter to prepare a support solution. The above nonwoven cloth was immersed for 16 to 24 hours in the support solution to attach the antibody to the fiber surface, yielding a sample filter (N-I). [Example 2]

Sample filters (N-2 to 31) were prepared by the same method as in Example 1 with the exception that antibody-stabilizing agents given in Table 1 in the quantities recorded in Table 1 were added instead of the saccharose in Example 1. [Comparative Example 1]

A sample filter (H-I) was prepared by the same method as in Example 1 with the exception that none of the saccharose employed in Example 1 was added. [Comparative Example 2]

A sample filter (H-2) was prepared by the same method as in Example 1 with the exception that an equivalent quantity of purified CEA antibody manufactured by Dako Corp. was added instead of the saccharose in Example 1. [Evaluation of virus deactivating effect]

Purified influenza virus diluted 10-fold with PBS (virus concentration of 200,000 plaques/mL) was employed as the test virus solution. The various above samples were cut into 5 cm squares, and securely mounted in the center of a virus spraying test device. The test virus solution was loaded into a nebulizer positioned upstream, and a virus recovery device was mounted downstream. Compressed air was supplied by an air compressor, and sample virus was sprayed from the nozzle of the nebulizer. A gelatin filter was positioned downstream from the sample, aspirating air within the test device for 5 minutes at an aspiration flow rate of 10 L/minute to trap passing virus mist. Following the test, the gelatin filter that had trapped the virus was recovered and the virus infection value following passage of the sample was calculated by the TCID50 method (method measuring the quantity required to infect 50 percent of the cells) using MDCK cells. The single-pass virus removal rate of the various samples was calculated by comparing the virus infection value of the gelatin filter when the sample was present and when it was absent. [Test of high-temperature storage properties]

The above virus spray test was conducted before and after placing N-I to N-31 and H- land H-2 for a week in an environment of 5O ⁰ C and 90 percent relative humidity. The results are given in Table 1.

From the results in Table 1, it will be understood that employing a support supporting a sugar, amino acid, SH group-protecting agent, or surfactant in addition to antibodies maintained the virus removal rate by stabilizing the antibodies without a reduction in initial capacity such as that observed in Comparative Example 2. It will be further understood from the results of N- 1 , N-2, N-9 to N- 11 , and N- 13 to N-31 that the effect of maintaining a high virus removal rate was achieved even at an added quantity weight ratio of 1 to 1000-fold sugar to antibody for disaccharides such as saccharose and trehalose and trisaccharides such as raffmose.

[Table 1]

[Example 3]

Employing vinylon fiber as the principal fiber, a dry nonwoven cloth with a basis weight of 100 g/m ² containing polyester fiber was prepared by resin bonding with an acrylic adhesive. Measurement of the nonwoven cloth by SEM revealed an average fiber diameter of 25 micrometers. An influenza antibody (IgY antibody) prepared by purifying immunized eggs laid by an ostrich to which antigen had been administered was diluted with PBS to prepare a solution with an antibody concentration of 100 ppm. Saccharose was added and stirred to 10 ppm and the mixture was passed through a 0.45 micrometer filter to prepare a support solution. The above nonwoven cloth was immersed for 5 minutes at room temperature in the support solution to attach the antibody to the fiber surface, yielding a sample filter (B-I). [Example 4]

Sample filters (B-2 to B-30) were prepared by the same method as in Example 3 with the exception that the stabilizing agents given in Table 2 were added in the quantities recorded in Table 2 instead of the saccharose in Example 3. [Comparative Example 3]

A sample filter (H-3) was prepared by the same method as in Example 3 with the exception that none of the saccharose employed in Example 3 was added. [Comparative Example 4]

A sample filter (H-4) was prepared by the same method as in Example 3 with the exception that an equivalent quantity of purified CEA antibody manufactured by Dako Corp. was added instead of the saccharose in Example 3. [Evaluation of virus deactivating effect]

Purified influenza virus diluted 10-fold with PBS (virus concentration of 200,000 plaques/mL) was employed as the test virus solution. The various above samples were cut into 5 cm squares, and securely mounted in the center of a virus spraying test device. The test virus solution was loaded into a nebulizer positioned upstream, and a virus recovery device was mounted downstream. Compressed air was supplied by an air compressor, and sample virus was sprayed from the nozzle of the nebulizer. A gelatin filter was positioned downstream from the sample, aspirating air within the test device for 5 minutes at an aspiration flow rate of 10 L/minute to trap passing virus mist. Following the test, the gelatin filter that had trapped the virus was recovered and the virus infection value following passage of the sample was calculated by the TCID50 method (method measuring the quantity required to infect 50 percent of the cells) using MDCK cells. The single-pass virus removal rate of the various samples was calculated by comparing the virus infection value of the gelatin filter when the sample was present and when it was absent.

[Test of high-temperature storage properties] The above virus spray test was conducted before and after placing B-I to B-30 and H-3 and H-4 for a week in an environment of 5O ⁰ C and 90 percent relative humidity. The results are given in Table 2.

From the results in Table 2, it will be understood that employing a support supporting a sugar in addition to antibodies maintained the virus removal rate by stabilizing the antibodies without a reduction in initial capacity such as that observed in Comparative Example 4. It will be further understood that the effect of maintaining a high virus removal rate was achieved even at an added quantity weight ratio of 1 to 1, 000-fold sugar to antibody for disaccharides such as saccharose and trehalose and trisaccharides such as raffmose, and that the stabilizing effect was greater than for monosaccharides such as glucose.

[Table 2]

Industrial Applicability

The present invention provides a hazardous substance-removing material which comprises a support having highly stable antibodies supported thereon.