Water-soluble saccharide for carrier and use thereof

Abstract

To provide a water-soluble saccharide that is capable of supporting various proteins such as enzymes and also capable of forming a carrier with high shape flexibility.SOLUTION: A water-soluble saccharide for a carrier according to the present disclosure is constituted by modifying at least some of hydroxyl groups possessed by the saccharide with (meth)acryloyl groups, the water-soluble saccharide serving for forming a carrier supporting a protein.SELECTED DRAWING: None

Description

[Technical Field]

[0001] The present invention relates to a water-soluble saccharide for use as a carrier and its use. [Background technology]

[0002] Research and development has been conducted on the use of carriers containing water-soluble sugars to support or immobilize target substances. For example, Non-Patent Document 1 discloses the production of vinegar using immobilized yeast and immobilized acetic acid bacteria on calcium alginate gel as a carrier. [Prior art documents] [Non-patent literature]

[0003] [Non-Patent Document 1] Nippon Shokuhin Kogyo Gakkaishi Vol.37, No.9, 727-725 (1990) Summary of the Invention [Problem to be solved by the invention]

[0004] On the other hand, the carrier containing water-soluble saccharides disclosed in Non-Patent Document 1 is a gel, and therefore has limitations on the support of various proteins such as enzymes. For example, since it is difficult to process the surface of the carrier on which the enzyme is supported, the degree of freedom in the shape of the carrier is low, and the range of applications is narrow.

[0005] An object of one aspect of the present invention is to provide a water-soluble saccharide that can support various proteins such as enzymes and can be formed into a carrier with a high degree of freedom in shape. [Means for solving the problem]

[0006] As a result of extensive research into solving the above problems, the present inventors discovered that the above problems could be solved by using water-soluble saccharides having specific functional groups, and thus completed the present invention.

[0007] A water-soluble saccharide for a carrier according to one embodiment of the present invention is a water-soluble saccharide for a carrier in which at least a portion of the hydroxyl groups of the saccharide have been modified to (meth)acryloyl groups, and which is used to form a carrier for carrying a protein. [Effects of the Invention]

[0008] According to one aspect of the present invention, it is possible to provide a water-soluble saccharide that can be formed into a carrier that can support various proteins such as enzymes and has a high degree of freedom in shape. [Brief explanation of the drawings]

[0009] [Figure 1] FIG. 1 shows the results of HPLC analysis in Evaluation Example 2. [Figure 2] FIG. 1 shows the results of HPLC analysis of TOM-D in Evaluation Example 3. [Figure 3] FIG. 1 shows the results of HPLC analysis of TOM-F in Evaluation Example 3. [Figure 4] FIG. 10 is a schematic diagram of an exposure mask used in Evaluation Example 4. [Figure 5] The image shows the observation results of a silicon wafer (dot pattern with 100 μm spacing) after patterning. [Figure 6] The image shows the observation results of a silicon wafer (dots spaced 50 μm apart) after patterning. [Figure 7] The image shows the observation results of a silicon wafer (dots spaced 25 μm apart) after patterning. [Figure 8] The image shows the observation results of a silicon wafer (dots spaced 10 μm apart) after patterning. [Figure 9] FIG. 1 shows the results of HPLC analysis in Evaluation Example 4 (enzyme reaction at 20° C.). [Figure 10] FIG. 1 shows the results of HPLC analysis in Evaluation Example 4 (enzyme reaction at 40° C.). [Figure 11] FIG. 1 shows the results of HPLC analysis in Evaluation Example 4 (enzyme reaction at 60° C.). [Figure 12] FIG. 1 shows the results of HPLC analysis in Evaluation Example 4 (enzyme reaction at 80° C.). [Figure 13]FIG. 1 shows the results of HPLC analysis in Evaluation Example 4 (enzyme reaction at 100° C.). [Figure 14] FIG. 1 shows the results of SDS-PAGE in Evaluation Example 5. [Figure 15] FIG. 10 is a diagram showing the calculation results of amino acid concentrations in Evaluation Example 5. DETAILED DESCRIPTION OF THE INVENTION

[0010] <Terminology> As used herein, a numerical range indicated using "to" indicates a range that includes the numerical values ​​before and after "to" as the minimum and maximum values, respectively.

[0011] As used herein, unless otherwise specified, the term "saccharides" refers to "saccharides" that are raw materials for the "water-soluble saccharides" according to one embodiment of the present invention. The term "saccharides" encompasses carbohydrates in the field of food science.

[0012] Furthermore, as used herein, the term "water-soluble saccharide" refers to a saccharide in which at least a portion of the hydroxyl groups of the saccharide have been modified with a polymerizable group, and which is water-soluble. That is, as used herein, the term "water-soluble saccharide" is written as an abbreviation for "a water-soluble saccharide in which at least a portion of the hydroxyl groups have been modified with a polymerizable group." Furthermore, unless otherwise specified, the term "saccharide" modified with "water-soluble" refers to a specific example of a water-soluble saccharide according to one embodiment of the present invention.

[0013] Additionally, as used herein, the term "(meth)acrylic" refers to either or both of acrylic and methacrylic. Specifically, the term "(meth)acrylic acid" refers to either or both of acrylic acid and methacrylic acid, and the term "(meth)acrylate" refers to either or both of "acrylate" and "methacrylate." Additionally, as used herein, the term "(meth)acryloyl group" refers to either or both of "acryloyl group" and "methacryloyl group." Additionally, as used herein, the term "(meth)acrylamide" refers to either or both of "acrylamide" and "methacrylamide."

[0014] <Water-soluble sugars for carriers> The water-soluble saccharide for carrier use according to one embodiment of the present invention (hereinafter sometimes abbreviated as "water-soluble saccharide") is a saccharide for carrier use in which at least a portion of the hydroxyl groups of the saccharide have been modified to (meth)acryloyl groups, and is used to form a carrier for supporting a protein. In other words, the water-soluble saccharide has a sugar chain derived from the raw material sugar and a (meth)acryloyl group bound to the sugar chain.

[0015] The water-soluble saccharide-containing molded article according to one embodiment of the present invention can support various proteins, such as enzymes, while maintaining the activity of the proteins. Furthermore, the molded article has a high degree of freedom in shape and can be used for various purposes. The molded article will be described later.

[0016] [Sugars] In one embodiment of the present invention, the water-soluble saccharide raw material is a saccharide that forms the backbone of the water-soluble saccharide. The saccharide that forms the backbone of the water-soluble saccharide is typically one or more selected from the group consisting of disaccharides, trisaccharides, tetrasaccharides, oligosaccharides, polysaccharides, carbohydrates, and dietary fiber. The saccharide raw material for the water-soluble saccharide may be a single type or a combination of two or more types. The saccharide raw material may be an artificially synthesized saccharide or a natural saccharide. Furthermore, when the saccharide raw material is poorly water-soluble, a saccharide obtained by hydrolyzing the saccharide may be used as the raw material. Furthermore, when the saccharide raw material is poorly water-soluble, a saccharide whose water solubility has been increased by introducing a hydrophilic group such as a hydroxyalkylene group into the saccharide may be used as the raw material. The saccharide raw material is not limited, but may be a saccharide having a solubility in water at 20°C of 5% by weight or more.

[0017] Examples of disaccharides include sucrose, lactose, maltose, trehalose, turanose, and cellobiose. Examples of trisaccharides include raffinose, melezitose, and maltotriose. Examples of tetrasaccharides include acarbose and stachyose. Examples of oligosaccharides include fructooligosaccharides, galactooligosaccharides, mannanoligosaccharides, and lactofructose oligosaccharides. Examples of polysaccharides include glycogen, starch, pullulan, dextrin, cyclodextrin, dextrose, cellulose, glucan, fructan, and chitin; and polysaccharides in which some of the hydroxyl groups of these polysaccharides have been modified with hydrophilic groups such as hydroxyalkylene groups.

[0018] According to one embodiment of the present invention, the water-soluble saccharide raw material may have a crosslinked structure in the sugar chain derived from a polyhydric alcohol or a polycarboxylic acid. Examples of polyhydric alcohols include sugar alcohols such as sorbitol, mannitol, xylitol, maltitol, and erythritol; glycerin; and the like. Examples of polycarboxylic acids include citric acid.

[0019] The saccharides that are the raw material for the water-soluble saccharide according to one aspect of the present invention are preferably pullulan, dextrin, or cyclodextrin.

[0020] The sugars used as the raw material for the water-soluble sugars according to one embodiment of the present invention are preferably water-soluble dietary fibers. Water-soluble dietary fibers have multiple hydroxyl groups in their molecules, which not only provide high water solubility, but also allow the hydroxyl groups to be modified into (meth)acryloyl groups. Furthermore, water-soluble dietary fibers have many branched structures in their sugar chains, making them suitable as raw materials for water-soluble sugars with high polymerizability. From the viewpoint of easily preparing a low-viscosity aqueous composition, the sugars used as the raw material for the water-soluble sugars are preferably water-soluble dietary fibers that form low-viscosity aqueous solutions, among water-soluble dietary fibers. Examples of water-soluble dietary fibers that form low-viscosity aqueous solutions include indigestible dextrin, isomaltodextrin, dextrose, and polydextrose. The sugars used as the raw material for the water-soluble sugars are preferably indigestible dextrin or dextrose.

[0021] [Examples of water-soluble sugars] For example, the water-soluble saccharide according to one embodiment of the present invention may have, at least in part, a structure represented by the following formula (A).

[0022] [ka]

[0023] In formula (A), C * is a carbon atom contained in the sugar chain, and is selected from the carbon atoms at positions 1, 2, 3, 4, 5, and 6 of the sugar chain, as well as carbon atoms at other positions. p is a (meth)acryloyl group. In other words, the water-soluble saccharide is a saccharide in which at least a part of the hydroxyl groups -OH of the saccharide is replaced with -OR p The water-soluble sugars are substituted with multiple (meth)acryloyl groups R p Each atom is connected to a carbon atom C at a specific position via an oxygen atom. * may be bonded to carbon atoms C at two or more different positions.* For example, multiple (meth)acryloyl groups R p All carbon atoms are C6 * and a plurality of (meth)acryloyl groups R p are independently selected from the 2-, 4- and 5-position carbon atoms C * It may be bonded to either of the following:

[0024] [Characteristics of water-soluble sugars] In order to more easily maintain the activity of the protein supported on the water-soluble saccharide, the weight-average molecular weight (Mw) of the water-soluble saccharide for carrier is preferably 500 or more, more preferably 1000 or more, and even more preferably 1300 or more. In order to improve the coatability of a composition containing the water-soluble saccharide for carrier, the weight-average molecular weight (Mw) of the water-soluble saccharide for carrier is preferably 500,000 or less, more preferably 20,000 or less, and even more preferably 15,000 or less.

[0025] As used herein, weight average molecular weight (Mw) refers to a value measured by gel permeation chromatography (GPC) in terms of standard polystyrene.

[0026] In this specification, the total number of hydroxyl groups in the raw material saccharide is defined as 100 mol%, and the proportion of the hydroxyl groups modified to (meth)acryloyl groups is referred to as the modification rate. In the water-soluble saccharide according to one embodiment of the present invention, the modification rate is preferably 5 mol% or more, more preferably 10 mol% or more. This allows suitable polymerization of water-soluble saccharide molecules via the (meth)acryloyl groups, facilitating the reaction between the protein to be supported on the water-soluble saccharide and the substrate. Furthermore, in the water-soluble saccharide according to one embodiment of the present invention, the modification rate is preferably 60 mol% or less, more preferably 50 mol% or less, and even more preferably 40 mol% or less. This allows a suitable proportion of unmodified hydroxyl groups to be present in the water-soluble saccharide, thereby ensuring good water solubility, and therefore allowing the content of water-soluble saccharide in the carrier composition to be suitably adjusted. Furthermore, by having the denaturation rate be equal to or less than the above-mentioned preferred upper limit, the time required for the process of denaturing the raw material saccharides, which is included in the method for producing water-soluble saccharides, is shortened, and therefore the carrier composition containing the water-soluble saccharides can be produced in a shorter time.

[0027] The modification rate is 13 It can be determined by C-NMR (nuclear magnetic resonance spectroscopy).

[0028] The solubility of the water-soluble saccharide according to one embodiment of the present invention in water at 20° C. may be 10% by weight or more, preferably 20% by weight or more, and more preferably 30% by weight or more. In particular, a water-soluble saccharide having a solubility in water at 20° C. of 10% by weight or more can be suitably used to prepare a carrier composition containing the water-soluble saccharide as an aqueous composition.

[0029] In this specification, the solubility in water at 20°C of a water-soluble saccharide according to one embodiment of the present invention refers to the solid content of a gel of water-soluble saccharide obtained by concentrating the water-soluble saccharide containing excess water until it becomes saturated and precipitates, and then heating the gel at 120°C for 1 hour.

[0030] In this specification, saccharides in which the hydroxyl groups of the saccharides have been modified to (meth)acryloyl groups and which have a solubility in water at 20°C of less than 5% by weight are "poorly water-soluble saccharides" and are distinguished from the water-soluble saccharides of one embodiment of the present invention.

[0031] [Method for producing water-soluble saccharides] The water-soluble saccharide according to one embodiment of the present invention can be produced, for example, by reacting the saccharide with a modifying agent, which is a compound having a (meth)acryloyl group, in the presence of an organic solvent, thereby modifying at least a portion of the hydroxyl groups of the saccharide to (meth)acryloyl groups.

[0032] (denaturant) The electrophilic group of the modifying agent can be an electrophilic group commonly used in the reaction mechanism in which the modification reaction proceeds. Examples of the electrophilic group of the modifying agent include halogen groups such as chlorine, bromine, and iodine; isocyanate groups; acid anhydride groups; and the like. Among these, the electrophilic group of the modifying agent is preferably a halogen group.

[0033] More specifically, examples of the modifying agent include (meth)acrylic acid chloride, (meth)acrylic acid anhydride, 2-(2-(meth)acryloyloxyethyloxy)ethyl isocyanate, and 2-isocyanatoethyl (meth)acrylate. Among these, the modifying agent is preferably (meth)acrylic acid chloride or (meth)acrylic acid anhydride. This allows the modification reaction to proceed more efficiently.

[0034] The raw material saccharides may be saccharides that have been previously modified by enzymatic hydrolysis, acid treatment, etc. Alternatively, the raw material saccharides may be saccharides that have been previously purified and fractionated by reprecipitation, etc.

[0035] The organic solvent used in the method for producing water-soluble saccharides can be appropriately selected from those that can dissolve the saccharides and the denaturant but do not react with the saccharides and the denaturant. The organic solvent is preferably an aprotic polar solvent. Examples of aprotic polar solvents include N-methylpyrrolidone (NMP); N,N-dimethylformamide (DMF); dimethyl sulfoxide (DMSO); dimethylacetamide (DMAc); ketones such as acetone and methyl ethyl ketone; and ethers such as dioxane and tetrahydrofuran.

[0036] When the electrophilic group of the modifying agent is a halogen group, the organic solvent used as the reaction solvent is preferably NMP. This allows hydrochloric acid, which is generated as a by-product by the reaction between the halogen group of the modifying agent and the hydroxyl group of the sugar, to have affinity with the lone electron pair of the nitrogen atom of NMP, thereby reducing the effect of the hydrochloric acid on the sugar. Therefore, using NMP as the reaction solvent allows the modifying agent to modify the sugar more efficiently.

[0037] (Reaction conditions with denaturant) In the reaction with the modifying agent, the temperature in the system is preferably -10°C or higher, more preferably 0°C or higher. This allows for easy temperature control in the reaction with the modifying agent. In addition, in the reaction with the modifying agent, the temperature in the system is preferably 100°C or lower, more preferably 60°C or lower. This reduces the polymerization reaction of the (meth)acryloyl groups in the modifying agent and the water-soluble saccharides produced by modifying the saccharides in the reaction with the modifying agent.

[0038] The reaction with the modifier may be carried out in air or in an inert gas atmosphere. Although not limited thereto, the reaction with the modifier is preferably carried out in an inert gas atmosphere. Examples of the inert gas include nitrogen gas and argon gas.

[0039] In the reaction with the modifying agent, the mixing weight ratio of the saccharide to the modifying agent in the system may be appropriately selected depending on the amount of hydroxyl groups possessed by the saccharide and the desired modification rate.

[0040] In the reaction with the modifying agent, after the reaction is completed, an amine may be added to the system to neutralize the acid generated as a by-product of the modification reaction. Examples of the amine include tertiary amines such as triethylamine.

[0041] <Carrier composition> A carrier composition according to one embodiment of the present invention includes the water-soluble saccharide according to one embodiment of the present invention and a polymerization initiator.

[0042] Examples and preferred embodiments of the water-soluble saccharide in the carrier composition according to one embodiment of the present invention have already been explained, and will not be repeated here.

[0043] [Polymerization initiator] The polymerization initiator contained in the carrier composition according to one embodiment of the present invention may be a radical polymerization initiator, and the polymerization initiator may be one type or a combination of two or more types.

[0044] In terms of ease of microfabrication of a molded article containing the carrier composition according to one embodiment of the present invention, the polymerization initiator is preferably a photosensitizer.

[0045] Examples of the photosensitizer include a photoradical polymerization initiator.

[0046] Examples of photoradical polymerization initiators include α-hydroxyketone-based photopolymerization initiators, α-aminoketone-based photopolymerization initiators, benzyl ketal-based photopolymerization initiators, oxime ester-based photopolymerization initiators, acridine-based photopolymerization initiators, benzophenone-based photopolymerization initiators, acetophenone-based photopolymerization initiators, aromatic ketoester-based photopolymerization initiators, benzoic acid ester-based photopolymerization initiators, acylphosphine oxide-based photopolymerization initiators, and titanocene-based photopolymerization initiators. The photoradical polymerization initiator may be emulsified or solubilized in an aqueous system using, for example, a surfactant and / or a polymerizable monomer. An example of a commercially available photoradical polymerization initiator is FAI-101L (manufactured by Fujifilm Corporation).

[0047] Examples of α-hydroxyketone photopolymerization initiators include 1-(4-isopropylphenyl)-2-hydroxy-2-methylpropan-1-one, 2-hydroxy-2-methyl-1-phenylpropan-1-one, 1-hydroxycyclohexylphenyl ketone, 1-[4-(2-hydroxyethoxy)-phenyl]-2-hydroxy-2-methylpropan-1-one, and 2-hydroxy-1-[4-[4-(2-hydroxy-2-methylpropionyl)benzyl]phenyl]-2-methylpropan-1-one. Examples of commercially available α-hydroxyketone photopolymerization initiators include Omnirad 2959 (manufactured by IGM Resins BV).

[0048] Examples of the photoradical polymerization initiator include organic peroxides such as methyl ethyl ketone peroxide and cyclohexanone peroxide; and azo compounds such as 2,2'-azobis(isobutylamidine) dihydrochloride (AIBA).

[0049] In order to more easily maintain the activity of the protein supported by the water-soluble saccharides, the content of the water-soluble saccharides contained in the carrier composition according to one embodiment of the present invention is preferably 5% by weight or more, more preferably 20% by weight or more, and even more preferably 30% by weight or more, based on the total weight of the water-soluble saccharides. Furthermore, in order to reduce the viscosity of the carrier composition according to one embodiment of the present invention and improve its coatability, the content of the water-soluble saccharides is preferably 50% by weight or less, more preferably 40% by weight or less.

[0050] In order to more easily maintain the activity of the protein supported on the water-soluble saccharide, the content of the polymerization initiator contained in the carrier composition according to one embodiment of the present invention is preferably 0.01 parts by weight or more, and more preferably 0.1 parts by weight or more, per 100 parts by weight of the water-soluble saccharide. Furthermore, in order to reduce the production cost of the carrier composition according to one embodiment of the present invention, the content of the polymerization initiator contained in the carrier composition according to one embodiment of the present invention is preferably 5 parts by weight or less, and more preferably 3 parts by weight or less, per 100 parts by weight of the water-soluble saccharide.

[0051] 〔protein〕 The carrier composition according to one aspect of the present invention may contain a protein. By containing a protein, a substrate that binds to the protein can be easily recovered.

[0052] Examples of the protein include enzymes, antibodies, cell adhesion proteins, microorganisms such as cells, etc. Examples of the enzyme include hydrolases such as proteases, amylases, and esterases, oxidoreductases such as glucose oxidase, transferases such as glucosyltransferase, isomerases such as glucose isomerase, and synthetic enzymes such as polyhydroxyalkanoate synthase.

[0053] Examples of the substrate include a substrate that reacts with an enzyme, an antigen, a cell, and the like.

[0054] [Other ingredients] The carrier composition according to one embodiment of the present invention may contain, as other components, a diluting solvent, a crosslinking agent, a surfactant, a reaction terminator, and other additives such as an adhesion aid, a filler, and a water-soluble resin. The other components may be one type or a combination of two or more types.

[0055] (Dilution solvent) The dilution solvent is not limited to, but may be, water, an amphipathic solvent, or a combination thereof, preferably water. Examples of amphipathic solvents include ethanol, methanol, acetonitrile, and N-methyl-2-pyrrolidone. The water content in the carrier composition can be appropriately selected depending on the desired viscosity of the carrier composition and the film thickness of the cured product obtained by polymerizing the carrier composition.

[0056] (Crosslinking agent) By including a crosslinking agent in the carrier composition, polymerizable groups of the water-soluble saccharides contained in the carrier composition can be crosslinked with each other via the crosslinking agent. The crosslinking agent preferably has two or more polymerizable groups.

[0057] Examples of crosslinking agents include radically polymerizable crosslinking agents. Examples of radically polymerizable crosslinking agents include crosslinking agents having a vinyl group, such as 1,3-bis(vinylsulfonyl)-2-propanol; and crosslinking agents having a (meth)acryloyl group. Examples of crosslinking agents having a (meth)acryloyl group include (meth)acrylic acid esters having a polyoxyalkylene group and (meth)acrylic acid esters formed from polyhydric alcohols.

[0058] In the carrier composition according to one embodiment of the present invention, the content of the crosslinking agent is preferably 0.01 parts by weight or more, more preferably 0.10 parts by weight or more, based on 100 parts by weight of the water-soluble saccharides. Also, in the carrier composition according to one embodiment of the present invention, the content of the crosslinking agent is preferably 5.00 parts by weight or less, more preferably 3.00 parts by weight or less, based on 100 parts by weight of the water-soluble saccharides.

[0059] (surfactant) By including an appropriate amount of surfactant in the carrier composition, the surface tension of the carrier composition can be adjusted as desired, improving leveling during coating and improving the thickness uniformity of the coating film.

[0060] Examples of surfactants contained in the carrier composition include fluororesin-based surfactants, silicone-based surfactants, polyoxyalkylene ether-based surfactants, and acrylic resin-based surfactants.

[0061] In the carrier composition according to one embodiment of the present invention, the surfactant content is preferably 0.01 parts by weight or more, more preferably 0.30 parts by weight or more, based on 100 parts by weight of the water-soluble saccharide content. In addition, in the carrier composition according to one embodiment of the present invention, the surfactant content is preferably 10.00 parts by weight or less, more preferably 5.00 parts by weight or less, based on 100 parts by weight of the water-soluble saccharide content.

[0062] (reaction stopper) By including a reaction terminator in the carrier composition, overpolymerization can be suppressed when the carrier composition is photopolymerized.

[0063] In the carrier composition according to one embodiment of the present invention, the content of the reaction terminator is preferably 0.01 parts by weight or more, more preferably 0.10 parts by weight or more, based on 100 parts by weight of the water-soluble saccharides. Also, in the carrier composition according to one embodiment of the present invention, the content of the reaction terminator is preferably 5.00 parts by weight or less, more preferably 1.00 parts by weight or less, based on 100 parts by weight of the water-soluble saccharides.

[0064] [Uses of the carrier composition] The carrier composition according to one embodiment of the present invention can be used in applications such as cell culture and biosensors.

[0065] <Manufacturing kit> A production kit according to one embodiment of the present invention is a production kit for producing a carrier for supporting a protein, comprising a water-soluble saccharide for a carrier according to one embodiment of the present invention and a protein. Examples and preferred embodiments of the water-soluble saccharide for a carrier and the protein have already been described, and therefore will not be repeated here.

[0066] As used herein, the term "kit" refers to a package containing containers (e.g., bottles, tubes, vials, etc.) containing specific materials. A production kit according to one embodiment of the present invention may be in a form in which each material contained therein exists independently, or in a form in which multiple materials are mixed (e.g., in the form of a composition). A production kit according to one embodiment of the present invention preferably includes instructions for using each material.

[0067] In addition to the water-soluble sugars and proteins for carriers, the production kit according to one embodiment of the present invention may further include materials for preparing the carrier composition (e.g., solvents), substrates, base plates, etc.

[0068] <Molded body> A molded article according to one aspect of the present invention comprises the carrier composition according to one aspect of the present invention and a protein.

[0069] The molded article may be molded on a substrate. Examples of the substrate include a silicon wafer substrate, a glass substrate, a plastic substrate, etc. The substrate surface may be subjected to a surface treatment such as an etching gas treatment to improve the wettability of the substrate surface.

[0070] For example, a mixture of a carrier composition and a protein is supplied onto a substrate, and the mixture on the substrate is exposed to light, thereby forming a molded article on the substrate. Exposure will be described later.

[0071] <Substrate treatment method> A substrate processing method according to one embodiment of the present invention includes the steps of: supplying a mixture of a carrier composition according to one embodiment of the present invention and an enzyme onto a substrate, and forming a molded body made of the mixture on the substrate (hereinafter sometimes referred to as a "molded body forming step"); and supplying a substrate to the molded body (hereinafter sometimes referred to as a "substrate supplying step"). Examples and preferred embodiments of the carrier composition, protein, and substrate have already been described, and will not be repeated here.

[0072] [Molded object forming process] In the compact formation step, a mixture of the carrier composition according to one embodiment of the present invention and the enzyme is supplied onto a substrate by a conventionally known method, for example, spin coating using a spinner.

[0073] After supplying the mixture of the carrier composition according to one embodiment of the present invention and the enzyme onto a substrate, the mixture may be baked (heat-dried) to remove water from the mixture. The baking temperature is preferably 50°C or higher and 300°C or lower, more preferably 60°C or higher and 100°C or lower. If a volatile organic solvent is used during baking, the water in the mixture can be removed at low temperatures.

[0074] In the compact formation step, a mixture of the carrier composition according to one embodiment of the present invention and an enzyme supplied onto a substrate may be exposed to light to form a compact made of the mixture on the substrate. This has the advantage that the substrate can be treated with the enzyme carried on the compact while the substrate and the enzyme-carrying compact can be removed together.

[0075] Examples of actinic radiation used to expose the mixture include ultraviolet light, visible light, and electron beams. Since ultraviolet light curing is superior in productivity and requires less equipment cost than electron beam curing, ultraviolet light is preferred as the actinic radiation. The exposure dose can be appropriately selected depending on the content of the polymerization initiator contained in the carrier composition, etc. The exposure dose is, for example, 50 to 200 mJ / cm. 2The exposure time can be, for example, about 10 to 30 minutes, but is not limited to these. The wavelength of the ultraviolet light to be irradiated may be selected depending on the type of polymerization initiator contained in the carrier composition.

[0076] The exposure may be performed on the entire surface of the mixture of the carrier composition and the enzyme supplied onto the substrate. The molded product of the mixture of the carrier composition and the enzyme may be used for treating the substrate while it is formed on the substrate, or the cured product of the mixture formed on the substrate may be pulverized and used for treating the substrate as a powder molded product.

[0077] When exposing the mixture, an exposure mask may be used to micro-process the molded body, and a micro-processed molded body can be obtained by development after exposure. Examples of micro-processing include dot-shaped, slit-shaped, and honeycomb-shaped shapes. The micro-processed molded body can have an increased specific surface area on the substrate, thereby increasing the amount of substrate processed, and after processing, the molded body carrying the enzyme can be removed together with the substrate.

[0078] The developer used for development may be an aqueous solvent such as water, etc. After development, baking (heat drying) may be carried out to remove the developer.

[0079] [Substrate supply process] In the substrate supplying step, a substrate is supplied to the molded body obtained in the molded body forming step. For example, the substrate can be supplied to the molded body by contacting the molded body with a solution of the substrate. In the substrate supplying step, the enzyme supported on the molded body reacts with the substrate, causing the substrate to bind to the enzyme, and the product produced from the substrate can be easily recovered.

[0080] The temperature and pH in the substrate supplying step can be selected depending on the type of enzyme supported on the molded article. In order to further promote the enzymatic reaction, the temperature in the substrate supplying step is preferably 20°C or higher and 100°C or lower. In order to further promote the enzymatic reaction, the pH in the substrate supplying step is preferably 5 or higher and 7 or lower.

[0081] <Summary> The water-soluble saccharide for a carrier according to aspect 1 of the present invention is a water-soluble saccharide for a carrier in which at least a portion of the hydroxyl groups of the saccharide have been modified to (meth)acryloyl groups, and is used to form a carrier for supporting a protein.

[0082] The water-soluble saccharide for a carrier according to aspect 2 of the present invention may be the same as that of aspect 1, wherein the weight-average molecular weight of the water-soluble saccharide is 500 or more and 500,000 or less.

[0083] The water-soluble saccharide for carriers according to aspect 3 of the present invention may be a water-soluble saccharide in aspect 1 or 2, in which 5 mol % or more and 60 mol % or less of the hydroxyl groups contained in the saccharide are modified to (meth)acryloyl groups, assuming that the total number of hydroxyl groups contained in the saccharide is 100 mol %.

[0084] The water-soluble saccharide for a carrier according to Aspect 4 of the present invention may be in any one of Aspects 1 to 3, and may have a solubility in water at 20° C. of 10% by weight or more.

[0085] A carrier composition according to a fifth aspect of the present invention comprises the water-soluble saccharide for a carrier according to any one of the first to fourth aspects and a polymerization initiator.

[0086] A carrier composition according to a sixth aspect of the present invention may be the same as that of the fifth aspect, in which the polymerization initiator is a photosensitizer.

[0087] The carrier composition according to aspect 7 of the present invention, in accordance with aspect 5 or 6, may further contain a protein.

[0088] A carrier composition according to an eighth aspect of the present invention is the composition according to the seventh aspect, wherein the protein is an enzyme.

[0089] A production kit according to a ninth aspect of the present invention is a production kit for producing a carrier for supporting a protein, which contains the water-soluble saccharide for a carrier according to any one of the first to fourth aspects and a protein.

[0090] A molded article according to a tenth aspect of the present invention is a molded article comprising the carrier composition according to any one of the fifth to eighth aspects and a protein.

[0091] The molded article according to an eleventh aspect of the present invention may be the molded article of the tenth aspect, which is molded on a substrate.

[0092] A substrate processing method according to aspect 12 of the present invention includes the steps of supplying a mixture of a carrier composition according to any one of aspects 5 to 8 and an enzyme onto a substrate, forming a molded body made of the mixture on the substrate, and supplying a substrate to the molded body.

[0093] A thirteenth aspect of the present invention relates to the substrate processing method of the eleventh aspect, and may be characterized in that the mixture supplied onto the substrate is exposed to light to form the molded body.

[0094] <Additional Notes> The present invention is not limited to the above-described embodiments, and various modifications are possible within the scope of the claims. Embodiments obtained by appropriately combining the technical means disclosed in different embodiments are also included in the technical scope of the present invention. [Example]

[0095] In the examples, room temperature refers to 20° C. to 30° C. Unless otherwise specified, % refers to % by weight.

[0096] In the graph showing the results of high performance liquid chromatography (HPLC) analysis, the horizontal axis indicates retention time (seconds) and the vertical axis indicates detection intensity (mV).

[0097] Example 1: Preparation of acrylic acid-modified pullulan (hereinafter referred to as TOM-P) The raw materials used in the preparation of Example 1 are listed below. Pullulan (Hayashibara) Acrylic acid chloride (Tokyo Chemical Industry Co., Ltd.) N-methyl-2-pyrrolidone (NMP; Mitsubishi Chemical)

[0098] 50.0 g of pullulan and 450.0 g of N-methylpyrrolidone were added to a 1 L reaction vessel equipped with a thermometer, a stirrer, and a condenser, and the mixture was heated to 80°C and dissolved in NMP. The temperature in the system was then raised to 40°C under a vacuum of 0 mmHg, and any remaining water in the system was removed. The system was then cooled to 15°C, and 25.1 g of acrylic acid chloride was added dropwise over 2 hours while maintaining the system temperature at 15°C. After the dropwise addition, the reaction was continued for 2 hours at a system temperature of 15°C.

[0099] Next, 28.0 g of triethylamine was added to neutralize the mixture, and the neutralized salt and N-methylpyrrolidone were purified and removed. Next, ion-exchanged water was added so that the content of the acrylic acid-modified product in the system became 10% by mass, and TOM-P with a modification rate of 30% (theoretical value) to acrylic groups was obtained.

[0100] <Evaluation Example 1> Evaluation of enzyme immobilization feasibility An evaluation was conducted to determine whether the resin prepared in Example 1 can immobilize an enzyme. The resin and enzyme used in Evaluation Example 1 are shown below.

[0101] 〔resin〕 Resin (TOM-P) prepared in Example 1 PHS / PEES (copolymer of parahydroxystyrene and paraethoxyethoxystyrene, organic solvent-based positive resin, hereinafter sometimes referred to as "positive resin") A mixture of 4,4',4''-(ethane-1,1,1-triyl)triphenol / formaldehyde and parahydroxystyrene / paraethoxystyrene (organic solvent-based negative resin, hereinafter sometimes referred to as "negative resin")

[0102] 〔enzyme〕 LpHera (Novozymes, α-amylase that breaks down starch and dextrin) Proteax (Amano Enzyme, a protease that breaks down proteins)

[0103] For Sample No. 1, 10 g of the resin (TOM-P) prepared in Example 1 was placed in a light-shielding bottle, and 0.06 g (3% based on the solid content) of a photosensitizer was added and dissolved. Omnirad 2959 (manufactured by IGM Resins BV) was used as the photosensitizer. Next, 1 mL of LpHera or 1.0 g of Proteax was added and dissolved in a light-shielded container. The resins evaluated and the evaluation results are listed in Table 1.

[0104] For sample No. 2, 10 g of a positive resin (PHS / PEES) was placed in a light-shielding bottle, and 0.06 g of a photosensitizer (3% of the solid content) was added and dissolved. The photosensitizer used was diphenyl 2,4,6-trimethylphenylsulfonium-p-toluenesulfonate. Next, 1 mL of LpHera or 1.0 g of Proteax was added and dissolved in a light-shielded container. The resins evaluated and the evaluation results are listed in Table 1.

[0105] For Sample No. 4, 10 g of negative resin (4,4',4''-(ethane-1,1,1-triyl)triphenol / formaldehyde and parahydroxystyrene / paraethoxystyrene) was placed in a light-shielding bottle, and 0.06 g of photosensitizer (3% of the solid content) was added and dissolved. Diphenyl 2,4,6-trimethylphenylsulfonium-p-toluenesulfonate was used as the photosensitizer. Next, 1 mL of LpHera or 1.0 g of Proteax was added and dissolved in a light-shielded container. The resins evaluated and the evaluation results are listed in Table 1.

[0106] [Table 1]

[0107] In Table 1, sample No. 3, "positive resin (10% solids, NV preparation)," is a sample obtained by diluting sample No. 2 with a soluble solvent to a solids content of 10%. Sample No. 5, "negative resin (10% solids, NV preparation)," is a sample obtained by diluting sample No. 4 with a soluble solvent to a solids content of 10%. "Good (dissolved)" indicates that the enzyme was uniformly dissolved in the resin and that the resin was able to immobilize the enzyme. "Poor (separated)" indicates that the enzyme was not uniformly dissolved in the resin and that the resin was unable to immobilize the enzyme.

[0108] As shown in Table 1, it was found that the TOM-P prepared in Example 1 was capable of immobilizing LpHera and Proteases.

[0109] Example 2: Preparation of TOM-P gel with LpHera A TOM-P gel supporting LpHera was prepared according to the following steps 1 to 8.

[0110] (Step 1) 10 g of TOM-P was placed in a light-shielding bottle, and a photosensitizer (3% of the solid content) was added and dissolved. Omnirad 2959 was used as the photosensitizer. (Procedure 2) LpHera was added in an amount that was 10% of the volume of the solution in (Procedure 1) and dissolved. (Step 3) TOM-P in which LpHera had been dissolved was applied to a silicon wafer by spin coating. (Step 4) The silicon wafer coated with TOM-P was baked (80°C, 1 minute). (Step 5) Exposure was carried out for 30 minutes. No exposure mask was used. (Step 6) Developed in pure water for 1 minute. (Step 7) The developed silicon wafer was baked (80°C, 1 minute). (Step 8) The gelled substance formed on the wafer surface was collected.

[0111] Comparative Example 1: Preparation of calcium alginate gel with LpHera A calcium alginate gel supporting LpHera was prepared according to the following steps 1 to 5.

[0112] (Procedure 1) 10.0 g of a 3% sodium alginate solution and 100.0 g of a 5% calcium chloride solution were prepared. Sodium alginate (Fujifilm Wako Pure Chemical Industries, Ltd.) was used to prepare the sodium alginate solution, and calcium chloride dihydrate (Fujifilm Wako Pure Chemical Industries, Ltd.) was used to prepare the calcium chloride solution. (Procedure 2) LpHera (1 mL) was added to a 3% aqueous solution of sodium alginate and dissolved. (Procedure 3) The liquid obtained in (Procedure 2) was added dropwise to a 5% aqueous solution of sodium alginate to precipitate a gel. (Step 4) The gel was collected by filtration. (Step 5) The gel was washed with pure water.

[0113] Evaluation Example 2: Enzyme reaction of gelled material A 5% aqueous starch solution was prepared using dissolved starch (Nacalai Tesque). 50 mL of the prepared 5% aqueous starch solution and 1.0 g of the gelled product obtained in Example 2 or Comparative Example 1 were placed in a 50 mL tube and subjected to an enzymatic reaction at 80°C. Samples for HPLC analysis were taken after 3 hours, 5 hours, 7 hours, and overnight.

[0114] The sample for HPLC analysis was filtered through a 0.45 μm filter and placed in a 1.5 mL vial, and the sugar composition of the sample was analyzed under the following HPLC analysis conditions.

[0115] (HPLC analysis conditions) Equipment: Alliance (manufactured by Waters) Column: Ultron-80N (Shimadzu GLC) Eluent: Ultrapure water Column temperature: 60℃ Flow rate: 0.6mL / min

[0116] The results of the HPLC analysis are shown in Figure 1. As shown in Figure 1, it was found that the gelled product obtained in Example 2 and the gelled product obtained in Comparative Example 1 exhibited the same enzymatic reactivity.

[0117] Example 3: Preparation of acrylic acid-modified dextrin (hereinafter referred to as TOM-D) TOM-D was used in the same manner as in Example 1, except that NSD500 (manufactured by Sanei Saccharification, dextrin) was used instead of pullulan as the sugar. However, foaming was observed during vacuum concentration, so Ryoto TM Ester CA-H1 (Mitsubishi Chemical Corporation, Food & Healthcare Division) was used as an antifoaming agent. The final solids content of TOM-D was adjusted to 35%.

[0118] Example 4: Preparation of acrylic acid-modified indigestible dextrin (hereinafter referred to as TOM-F) TOM-F was used in the same manner as in Example 1, except that Fibersol 2 (manufactured by Matsutani Chemical Industry, indigestible dextrin) was used instead of pullulan as the sugar. The final solid content of TOM-F was adjusted to 35%.

[0119] Example 5: Preparation of a wafer for enzyme reaction on which a TOM-D molded body carrying LpHera is formed Following the steps 1 to 7 below, TOM-D in which LpHera had been dissolved was dropped onto a silicon wafer and exposed to light.

[0120] (Step 1) 10 g of TOM-D was placed in a light-shielding bottle, and 0.105 g of a photosensitizer (3% of the solid content) was added and dissolved. Omnirad 2959 was used as the photosensitizer. (Step 2) 1 mL of LpHera was added in the dark and dissolved. (Step 3) Approximately 1.0 g of TOM-D in which LpHera had been dissolved was dropped onto a silicon wafer, and the silicon wafer was coated with the solution by spin coating. (Step 4) The silicon wafer coated with TOM-D was baked (80°C, 1 minute). (Step 5) Exposure was carried out for 30 minutes. No exposure mask was used. (Step 6) Developed in pure water for 1 minute. (Step 7) The developed silicon wafer was baked (80°C, 1 minute).

[0121] Example 6: Preparation of a wafer for enzyme reaction on which a TOM-F molded body carrying LpHera is formed The same procedure as in Example 5 was followed, except that TOM-D was changed to TOM-F, and TOM-F in which LpHera had been dissolved was dropped onto a silicon wafer and exposed to light.

[0122] <Evaluation Example 3> Enzyme reaction using the enzyme reaction wafers of Examples 5 and 6 A 5% aqueous starch solution was prepared using dissolved starch (Nacalai Tesque). The prepared 5% aqueous starch solution and the enzyme reaction wafer from Example 5 or 6 were placed in a 1 L beaker and subjected to an enzyme reaction at 20°C. Samples for HPLC analysis were taken after 3 hours, 5 hours, 7 hours, and overnight.

[0123] The sample for HPLC analysis was filtered through a 0.45 μm filter and placed in a 1.5 mL vial. The sugar composition of the sample was then analyzed under the same HPLC analysis conditions as in Evaluation Example 2. The analytical results of Example 5 are shown in FIG. 2, and the analytical results of Example 6 are shown in FIG. 3.

[0124] As shown in Figures 2 and 3, the peak area on the low molecular weight side increased as the enzyme reaction time increased. These results demonstrate that TOM-D and TOM-F can support the enzyme while maintaining its enzymatic activity.

[0125] Example 7: Preparation of a wafer for enzyme reaction on which a TOM-P molded body carrying LpHera is formed The gel obtained in Comparative Example 1 is a carrier containing water-soluble saccharides, as disclosed in Non-Patent Document 1. Because this carrier is a gel, it is difficult to perform surface processing when an enzyme is loaded onto it, and the degree of freedom in shape is low. In this example, we investigated whether the TOM-P molded body loaded with an enzyme has degree of freedom in shape while maintaining enzymatic activity.

[0126] TOM-P was applied to a silicon wafer for patterning, and a wafer for enzyme reaction was prepared, on which a TOM-P molded body carrying LpHera was formed. A dotted exposure mask was used. Figure 4 shows a schematic diagram of the exposure mask. The patterning process was carried out according to steps 1 to 5 below.

[0127] (Step 1) Approximately 1.0 g of TOM-P in which LpHera had been dissolved was dropped onto a silicon wafer and spin-coated. Omnirad 2959 was used as the photosensitizer. (Step 2) The silicon wafer coated with TOM-P was baked (80°C, 1 minute). (Step 3) Use an exposure mask and expose to 50 mJ / cm 2 The exposure machine used was LTCET-500 (manufactured by Litho Tech Japan). (Step 4) Developed in pure water for 1 minute. (Step 5) The developed silicon wafer was baked (80°C, 1 minute).

[0128] The silicon wafers after patterning were observed with a laser microscope to confirm and photograph the patterned shapes. The observation results are shown in Figures 5 to 8. As shown in Figures 5 to 8, good patterning was confirmed on all silicon wafers. These results demonstrate that enzyme-loaded TOM-P can be surface-processed and has a high degree of freedom in shape.

[0129] The measurement results of the surface area of ​​the silicon wafers in Figures 5 to 8 are shown in Table 2. The control in Table 2 shows the measurement results of a silicon wafer in which a TOM-P molded body with LpHera dissolved therein was formed on the entire surface without dot patterning.

[0130] [Table 2]

[0131] As shown in Table 2, it was found that the surface area increases as the spacing between dots decreases and the number of dots on the silicon wafer increases.

[0132] <Evaluation Example 4> Enzyme reaction using the enzyme reaction wafer of Example 7 The enzyme reaction was evaluated using the enzyme reaction wafer prepared in Example 7. Using a diamond cutter, the enzyme reaction wafer prepared in Example 7 was cut to have the same area for each of the "dots spaced 100 μm apart," "dots spaced 50 μm apart," "dots spaced 25 μm apart," and "dots spaced 10 μm apart" shown in Figure 4.

[0133] A 5% aqueous starch solution was prepared using dissolved starch (Nacalai Tesque). The prepared 5% aqueous starch solution and the cut silicon wafer were placed in a plastic petri dish and subjected to an enzymatic reaction at 20°C, 40°C, 60°C, 80°C, and 100°C. Samples for HPLC analysis were taken at 3 hours, 5 hours, 7 hours, and 4 days.

[0134] The sample for HPLC analysis was filtered through a 0.45 μm filter and placed in a 1.5 mL vial. The sugar composition of the sample was then analyzed under the same HPLC analysis conditions as in Evaluation Example 2.

[0135] The results of the HPLC analysis are shown in Figures 9 to 13. In Figures 9 to 13, "Control" indicates the measurement results of a sample recovered from a silicon wafer on which a TOM-P molded body with LpHera dissolved over the entire surface was formed without dot patterning. 10 μm, 25 μm, 50 μm, and 100 μm indicate the measurement results of samples recovered from silicon wafers patterned with "dots spaced 10 μm apart," "dots spaced 25 μm apart," "dots spaced 50 μm apart," and "dots spaced 100 μm apart" as shown in Figure 4, respectively.

[0136] The results of the HPLC analysis are summarized in Tables 3 to 5. The values ​​in Tables 3 to 5 indicate the area ratio (%) when the total area of ​​the peaks detected in the HPLC analysis is taken as 100%. G1 indicates an oligosaccharide composed of four or more glucose units, such as glucose, G2 indicates maltose, G3 indicates maltotriose, and G4 indicates maltotetraose.

[0137] In Tables 3 to 5, "temperature" refers to the enzyme reaction temperature. "Control" indicates the measurement results of a sample recovered from a silicon wafer on which a TOM-P molded body with LpHera dissolved over the entire surface was formed without dot-patterning. 10 μm, 25 μm, 50 μm, and 100 μm indicate the measurement results of samples recovered from silicon wafers patterned with "dots spaced 10 μm apart," "dots spaced 25 μm apart," "dots spaced 50 μm apart," and "dots spaced 100 μm apart," respectively, as shown in Figure 4.

[0138] [Table 3]

[0139] [Table 4]

[0140] [Table 5]

[0141] 9 to 13 and Tables 3 to 5, it was found that the peak area on the low molecular weight side increased as the enzyme reaction time increased. This result demonstrated that TOM-P can support enzymes while maintaining their enzymatic activity.

[0142] Furthermore, it was found that the peak area for low molecular weight molecules increased as the spacing between the dots decreased. From these results, it was found that the surface area increased as the number of dots on the wafer increased, and the enzyme reaction was accelerated.

[0143] Example 8: Preparation of a wafer for enzyme reaction on which a TOM-P molded body carrying Proteax is formed Next, we investigated whether enzymes other than LpHera could be supported on TOM-P while maintaining their activity. Following the steps 1 to 5 below, we prepared a wafer for enzyme reactions, which had a TOM-P molded body supporting ProteaX.

[0144] (Step 1) Approximately 1.0 g of TOM-P in which Proteax was dissolved was dropped onto a silicon wafer and applied to the silicon wafer by spin coating. (Step 2) The silicon wafer onto which TOM-P had been dropped was baked (50°C, 1 minute). (Step 3) UV exposure was carried out for 30 minutes. (Step 4) Developed in pure water for 1 minute. (Step 5) The developed silicon wafer was baked (50°C, 1 minute).

[0145] <Evaluation Example 5> Enzyme reaction using the enzyme reaction wafer of Example 2 A 1% albumin aqueous solution was prepared using albumin (derived from eggs, manufactured by Fujifilm Wako Pure Chemical Industries, Ltd.). The Proteax-immobilized TOM-P and the prepared 1% albumin aqueous solution were placed in a 1 L beaker and subjected to an enzymatic reaction at 60°C. The reaction solution was sampled after 1 hour, 5 hours, and overnight, and the sampled solution was boiled for several minutes. The molecular weight of the protein in the sampled solution was measured by polyacrylamide gel electrophoresis (SDS-PAGE), and the concentration of amino acids produced by the enzymatic reaction was measured and calculated using the formol method.

[0146] [SDS-PAGE] The sampling solution (5 μL each) and EzApply (5 μL, manufactured by ATTO) were mixed and boiled for 5 minutes. After boiling, the solution was confirmed to have cooled to room temperature (20-30°C), and then added to the wells of an SDS-PAGE gel (PAGRL HR) for electrophoresis. The electrophoresed gel was collected, immersed in EzStrain Aqua, and shaken overnight at room temperature. The shaken gel was collected and washed several times with pure water, after which the gel bands were confirmed.

[0147] [Formol method] 10 mL of the reaction mixture and 20 mL of pure water were added to a 50 mL beaker and stirred using a magnetic stirrer. After stirring, the mixture was adjusted to pH 8.3 with 0.1 N sodium hydroxide solution. Next, 10 mL of neutral formalin (pH 8.3) was added, and titration was initiated with 0.1 N sodium hydroxide solution, with the endpoint being pH 8.3. After titration, the amino acid concentration was calculated using the following formula:

[0148]

number

[0149] [Molecular weight of protein in collected liquid] The results of SDS-PAGE are shown in Figure 14. The left side of Figure 14 shows the results for the sampling solution that was subjected to an enzymatic reaction with albumin, and the right side shows the results for the sampling solution that was not subjected to an enzymatic reaction with albumin. In Figure 14, the band indicated by the arrow represents albumin.

[0150] As shown in Figure 14, the albumin band disappeared in the sample solution that had been subjected to an enzymatic reaction with albumin, confirming that an enzymatic reaction had occurred between albumin and the proteases supported on TOM-P.

[0151] [Concentration of amino acids produced by enzymatic reaction] The calculated results of the amino acid concentrations are shown in Figure 15. As shown in Figure 15, it was found that the amino acid concentrations of the sample solution that had been subjected to an enzyme reaction with albumin increased as the reaction time increased.

[0152] The results of Example 8 demonstrate that TOM-P can support proteases while maintaining enzymatic activity.

[0153] <Evaluation Example 6> Characteristic evaluation of TOM-P, TOM-D, and TOM-F The weight average molecular weight (Mw), solubility (%) in water at 20°C, and acryloyl group modification rate of TOM-P, TOM-D, and TOM-F were measured.

[0154] [Measurement of weight average molecular weight (Mw)] The GPC measurement device used was manufactured by Tosoh Corporation under the trade name "HLC-8120GPC" ("HLC" is a registered trademark of the company). The column used was a column consisting of three connected columns (manufactured by Tosoh Corporation under the trade names "TSKgel G3000HHR," "TSKgel G2000HHR," and "TSKgel" are registered trademarks of the company).

[0155] [Measurement of solubility (%) in water at 20°C] A gel-like water-soluble saccharide was obtained by concentrating a water-soluble saccharide containing excess water until it precipitated, and the weight change (%) of the water-soluble saccharide was calculated by heating it at 120°C for 1 hour, and this weight change was taken as the solubility (%) in water at 20°C. The weight change was determined according to the following steps 1 to 6. (Step 1) Water was removed from the water-soluble saccharides under vacuum to concentrate them until a gel of water-soluble saccharides was precipitated. With the gel of water-soluble saccharides precipitated, the liquid temperature was adjusted to 20°C, and then the precipitate was filtered to obtain a gel of water-soluble saccharides. (Step 2) The weight of the aluminum dish (empty) was measured. (Step 3) A gel-like water-soluble saccharide was added to an aluminum dish, and the amount added was measured. (Step 4) The aluminum dish containing the water-soluble sugars was heated at 120°C for 1 hour. (Step 5) The weight of the aluminum dish after heating was measured. (Step 6) The weight change rate was calculated using the following formula (B). {(Weight of aluminum dish after heating) - (Weight of aluminum dish (empty))} / Amount of water-soluble sugar added × 100 Formula (B)

[0156] [Acryloyl group modification rate] 13 The acryloyl group modification rate was measured by C-NMR. 13 The C-NMR conditions were as follows: NMR measurement device: JEOL Ltd., product name "ECZ-500R / S1" Solvent: Deuterium oxide (DO)

[0157] The measurement results for each property are shown in Table 6.

[0158] [Table 6] [Industrial Applicability]

[0159] The water-soluble saccharide for carriers according to one embodiment of the present invention can support various proteins such as enzymes and can be formed into carriers with a high degree of shape freedom. Therefore, the water-soluble saccharide for carriers can be used in applications such as cell culture and biosensors.

Claims

1. At least a part of the hydroxyl groups of the saccharide has been modified with a (meth)acryloyl group, Water-soluble sugars for use as carriers, for forming carriers that support proteins.

2. 2. The water-soluble saccharide for use as a carrier according to claim 1, wherein the weight-average molecular weight of the water-soluble saccharide is 500 or more and 500,000 or less.

3. The water-soluble saccharide for carrier use according to claim 1, wherein 5 mol% or more and 60 mol% or less of the hydroxyl groups of the water-soluble saccharide are modified to (meth)acryloyl groups, with the total number of hydroxyl groups of the water-soluble saccharide being 100 mol%.

4. 2. The water-soluble saccharide for use as a carrier according to claim 1, which has a solubility in water at 20°C of 10% by weight or more.

5. A carrier composition comprising the water-soluble saccharide for a carrier according to any one of claims 1 to 4 and a polymerization initiator.

6. The carrier composition according to claim 5 , wherein the polymerization initiator is a photosensitizer.

7. The carrier composition of claim 5 , further comprising a protein.

8. The carrier composition of claim 7, wherein the protein is an enzyme.

9. A production kit for producing a protein-carrying carrier, comprising the water-soluble saccharide for a carrier according to any one of claims 1 to 4 and a protein.

10. A molded article comprising the carrier composition according to claim 5 and a protein.

11. The molded article according to claim 10 , which is molded on a substrate.

12. A step of supplying a mixture of the carrier composition according to claim 5 and an enzyme onto a substrate, and forming a molded body made of the mixture on the substrate; and providing a substrate to the molded body.

13. The method for treating a substrate according to claim 12 , wherein the mixture supplied onto the substrate is exposed to light to form the molded body.

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