JP5822273B2 - Biodegradable hydrogel and method for producing the same - Google Patents

Description

  The present invention is highly hydrophilic and capable of absorbing and retaining a large amount of water, and in addition, biodegradable hydrogel exhibiting extremely excellent gel characteristics and biodegradable hydrogel and efficiently producing the hydrogel It is about the method.

  A polymer compound called a hydrogel has a large number of hydrophilic groups such as hydroxyl groups and carboxy groups in its chemical structure, and has a network-like cross-linked structure, and therefore includes a large amount of water in the network structure. In addition, some pressure does not release the water inside.

  Examples of the hydrogel include those obtained by crosslinking a hydrophilic polymer such as polyacrylic acid, polyvinyl alcohol, polyethylene oxide, and polyvinylpyrrolidone by chemical means using a cross-linking agent or the like, or physical means using γ rays. . Such a hydrogel has the property that water absorption is extremely excellent in addition to the advantage of high transparency, and its use as a wound dressing or the like is being studied. That is, since it can be said that a wound is healed more easily in a wet environment than in a dry environment, it is considered that healing can be promoted by covering the wound site with a sheet made of hydrogel. In addition, it can be used as a moisturizing agent added to cosmetics, a water retaining material used in the agricultural field, and a cell culture gel. In particular, crosslinked polyacrylates excellent in water absorption are being used as water-absorbing sheet materials such as diapers.

  However, the cross-linked product of the above-mentioned chemical conversion polymer does not exhibit biodegradability and also requires a large amount of energy for combustion treatment due to its excellent water absorption, and therefore has a large environmental load at the time of disposal after use. . Therefore, utilization of a highly hydrophilic biodegradable polymer can be considered. Such a biodegradable polymer can be decomposed by microorganisms and the like, and therefore has a small environmental load at the time of disposal. Moreover, since it is excellent in safety, it can also be used as an adhesive that can be used in vivo in, for example, a dental implant.

  However, the biodegradable polymer is inferior in transparency as compared with the chemical conversion polymer, and has a problem that the water absorption is not sufficient at most about 10 times in mass ratio. For example, there are agar and konjac as typical biodegradable hydrogels, and it can be understood by everyone that these are safe and easy to decompose, but it is difficult to absorb water once gelled. In addition, it can be sufficiently confirmed that it is difficult to use in applications that require high water absorption.

  In recent years, poly-γ-glutamic acid, which is the main component of natto yarn, has attracted attention as a biodegradable polymer. Since poly-γ-glutamic acid exhibits extremely high hydrophilicity, it can be used as a water-absorbing material by crosslinking. For example, Patent Document 1 describes a method for producing a biodegradable water-absorbent resin by irradiating a solution of poly-γ-glutamic acid with an electron beam. However, cross-linking with an electron beam loses the biodegradability that is characteristic of poly-γ-glutamic acid. On the other hand, since poly-γ-glutamic acid has a carboxy group for each monomer unit constituting the poly-γ-glutamic acid, it can be chemically crosslinked, and if it is chemically crosslinked, biodegradability can be maintained. The development of water-absorbing materials can also be expected. However, poly-γ-glutamic acid has a problem that chemical crosslinking is difficult due to its high hydrophilicity.

  That is, poly-γ-glutamic acid has high solubility in water, but hardly dissolves in organic solvents. Therefore, in order to modify the side chain carboxy group (α-carboxy group) of poly-γ-glutamic acid In the aqueous solution, 1-ethyl-3- (3-dimethylaminopropyl) carbodiimide hydrochloride (Water Soluble Carbodiimide, WSC), which is a water-soluble dehydrating condensing agent, was condensed with a crosslinking agent or a modifying agent ( Patent Documents 2 to 4). However, the dehydration reaction in an aqueous solution is naturally very inefficient. In addition, the dehydrating condensing agent is relatively expensive and may cause allergic symptoms in the living body.

  By the way, the present inventors have continued research on poly-γ-glutamic acid, and the complex of poly-γ-glutamic acid and a quaternary ammonium ion compound has a moisture retaining property against water. It has been found that it can be dissolved in an organic solvent while showing insolubility (Patent Document 5).

JP 2000-327795 A JP 2002-80593 A JP 2007-297559 A JP 2009-209362 A JP 2010-2222496 A

  As described above, a technique for crosslinking poly-γ-glutamic acid and utilizing it as a water-absorbing material or the like has been known. A method for modifying the side chain carboxy group of poly-γ-glutamic acid has also been known.

  However, the conventional method for modifying the side chain carboxy group of poly-γ-glutamic acid is very inefficient in that dehydration reaction is carried out in an aqueous solution. Moreover, the conventional poly-γ-glutamic acid side chain modified product does not exhibit sufficient water absorption. For example, the modified products described in Patent Documents 2 to 3 have only introduced a water-soluble compound into the side chain of poly-γ-glutamic acid, which is originally highly water-soluble, and do not become a hydrogel in order to exhibit water-solubility. Conceivable. The cross-linked body of Patent Document 4 is not a hydrogel but a molded body and is a hard material, and is difficult to use as a water-absorbing material. On the other hand, although the crosslinked body of patent document 1 shows very high water absorption, since it was bridge | crosslinked by irradiation of an electron beam, biodegradability has been lost.

  Under such circumstances, the present invention is based on a biodegradable hydrogel having extremely high hydrophilicity and capable of absorbing and retaining a large amount of water, and exhibiting biodegradability and extremely excellent gel characteristics, and electron beam irradiation. It aims at providing the method for manufacturing the said hydrogel efficiently.

  The inventors of the present invention have made extensive studies to solve the above problems. As a result, if a specific amine compound is allowed to act on the organic solvent solution of the poly-γ-glutamate ion complex developed by the present inventors, the amine compound and the side chain can be surprisingly used without using a dehydration condensing agent. The present invention was completed by finding that the carboxy group reacted efficiently, and that the resulting compound was a hydrogel exhibiting excellent water absorption even though it was not a crosslinked product.

  The biodegradable hydrogel according to the present invention is characterized by comprising poly-γ-glutamic acid having the following unit structure (I) in which a side chain carboxy group is modified.

[Wherein X represents a linker group, and Y represents a C _6-12 aryl group having one or more hydroxyl groups]

X represents a C _1-6 alkylene group, or an amino group (—NR— group at the Y-side end; R represents a hydrogen atom or a C _1-6 alkyl group), an ether group (—O— group) Amide group (—NHC (═O) — group or —C (═O) NH— group), carbonyl group, —O (C═O) — group, — (C═O) O— group or urea group ( C _1-6 alkylene group having —NHC (═O) NH— group), and Y can be a phenyl group having one or more hydroxyl groups.

In the present invention, the “C _6-12 aryl group” refers to a monovalent aromatic hydrocarbon group having 6 to 12 carbon atoms. Examples thereof include a phenyl group, an indenyl group, a naphthyl group, and a biphenyl group. Of these, a phenyl group or a naphthyl group is preferable, and a phenyl group is more preferable.

The “C _1-6 alkylene group” refers to a linear or branched divalent aliphatic hydrocarbon group having 1 to 6 carbon atoms. Examples thereof include a methylene group, a dimethylene group, a trimethylene group, a methyldimethylene group, a tetramethylene group, a 1-methyltrimethylene group, a 1,1-dimethyldimethylene group, a pentamethylene group, and a hexamethylene group. Of these, a C _1-4 alkylene group is preferable, and a C _1-2 alkylene group is more preferable.

The “C _1-6 alkyl group” refers to a linear or branched monovalent aliphatic hydrocarbon group having 1 to 6 carbon atoms. For example, methyl group, ethyl group, n-propyl group, isopropyl group, n-butyl group, sec-butyl group, t-butyl group, n-pentyl group, n-hexyl group and the like can be mentioned. Of these, a C _1-4 alkyl group is preferable, and a C _1-2 alkyl group is more preferable.

The “C _12-20 alkyl group” means a linear or branched monovalent aliphatic hydrocarbon having 12 to 20 carbon atoms. For example, linear C _12-20 alkyl group such as dodecyl group, tridecyl group, tetradecyl group, pentadecyl group, hexadecyl group, heptadecyl group, octadecyl group, nonadecyl group, icosyl group; isotetradecyl group, isopentadecyl group, Isohexadecyl group, isoheptadecyl group, isooctadecyl group, sec-tetradecyl group, sec-pentadecyl group, sec-hexadecyl group, sec-heptadecyl group, sec-octadecyl group, t-tetradecyl group, t-pentadecyl group, t-hexadecyl group A branched C _12-20 alkyl group such as a group, t-heptadecyl group, t-octadecyl group, neotetradecyl group, neopentadecyl group, neohexadecyl group, neoheptadecyl group, neooctadecyl group, and the like. . R ⁴ is preferably a C _15-20 alkyl group, more preferably a C _16-20 alkyl group, and most preferably a C _17-20 alkyl group. R ⁵ is preferably a C _13-20 alkyl group, more preferably a C _14-19 alkyl group, and most preferably a C _15-18 alkyl group.

  The number of hydroxyl groups of the aryl group in the definition of Y is preferably 1 or more and 5 or less. By setting the number of hydroxyl groups to 1 or more, it becomes possible to ensure water absorption and use as an in vivo adhesive. On the other hand, there is no particular upper limit on the number of hydroxyl groups, and the number of aryl groups can be substituted. However, if the number of hydroxyl groups is too large, the price of the raw material compound may increase and the production cost may increase. Is preferably 5 or less. The number of hydroxyl groups is more preferably 2 or more, more preferably 4 or less, and still more preferably 3 or less.

The method for producing a biodegradable hydrogel according to the present invention comprises:
A step of preparing an ion complex by adding a quaternary ammonium ion compound of the following formula (II) or formula (III) to a poly-γ-glutamic acid aqueous solution;

[Wherein R ^{1 to} R ³ independently represent a C _1-2 alkyl group, and R ^{4 to} R ⁵ independently represent a C _12-20 alkyl group]

  It includes a step of modifying the side chain carboxy group of poly-γ-glutamic acid by reacting the ion complex with an amine compound of the following formula (IV) in an organic solvent.

[Wherein X represents a linker group, and Y represents a C _6-12 aryl group having one or more hydroxyl groups]

  In the method of the present invention, an alcohol solvent is preferably used as the organic solvent. Alcohol solvents have high solubility in the ion complex and are safe.

  Moreover, as X and Y, the thing similar to what was mentioned above can be mentioned.

  The hydrogel according to the present invention has a structure in which an amine compound is bonded with an amide bond to a side chain carboxy group of poly-γ-glutamic acid, so that it retains biodegradability and is extremely high in safety. In addition, it does not put an impact on the environment at the time of disposal. Moreover, it has extremely high hydrophilicity and can absorb and retain a large amount of water. Furthermore, it exhibits extremely excellent gel properties. Accordingly, it can be expected to be applied to adhesives that can be used in vivo, wound dressings, cell culture gels, agricultural water retaining materials, moisturizing agents blended into cosmetics, water absorbing materials, and the like.

  In addition, the method for producing the biodegradable hydrogel according to the present invention can use an organic solvent, unlike the low-efficiency conventional method of dehydration reaction in an aqueous solution, and is expensive and harmful to living organisms. There is no need to use an agent. Therefore, the method of the present invention is very useful as a biodegradable hydrogel having the above-mentioned characteristics that can be produced at low cost and efficiently.

FIG. 1 is a photograph showing changes with time when an ion complex of poly-γ-glutamic acid is added to a saturated ethanol solution of dopamine. It can be seen that when an ion complex is added ((1)), a reaction product with dopamine is generated at the interface as the ion complex dissolves ((2) to (4)). FIG. 2 is a photograph of the biodegradable hydrogel according to the present invention, which was allowed to stand after adding 100 to 2000 mass times of water. When 2000 mass times of water is added, it becomes a viscous sol, but when 100 mass times to 1000 mass times of water is added, the gel state is maintained. FIG. 3 is a graph showing the results of subjecting the biodegradable hydrogel dilution according to the present invention to a Frequency Sweep test. From these results, it can be seen that rheologically, even a sample diluted 3000 times shows gel properties. FIG. 4 is a graph showing the results of subjecting the biodegradable hydrogel dilution according to the present invention to the strain sweep test. From these results, it can be seen that the 1000-fold dilution of the biodegradable hydrogel according to the present invention exhibits gel properties until a strain amount of 15 to 20% is added. FIG. 5 is a graph showing the results of testing the self-healing property of the biodegradable hydrogel according to the present invention, where (1) shows the change in strain applied and (2) shows the G ′ value. From these results, no decrease in G ′ value was observed even when the amount of strain was changed, and it became clear that the biodegradable hydrogel according to the present invention showed excellent self-healing properties. It was. FIG. 6 is a graph showing the results of testing hysteresis characteristics regarding water absorption between the biodegradable hydrogel according to the present invention and an agarose gel which is a general hydrogel. It was found that the agarose gel hardly absorbs water once dried, whereas the dopamil PGA gel according to the present invention does not lose the water absorption performance even after repeated drying and water absorption five times.

  Hereinafter, first, the manufacturing method of the biodegradable hydrogel which concerns on this invention is demonstrated according to the order of implementation.

(1) Ion Complex Preparation Step In this step, a quaternary ammonium ion compound of formula (II) or formula (III) is added to an aqueous solution of poly-γ-glutamic acid (hereinafter sometimes abbreviated as “PGA”). By adding, an ion complex composed of PGA and a quaternary ammonium ion compound is prepared.

  The type of PGA is not particularly limited. For example, there are those consisting only of L-glutamic acid, those consisting only of D-glutamic acid, and those containing both, and any of them can be used.

  Since it is considered that a more stable high-dimensional structure is exhibited, it is preferable that 90% or more of glutamic acid constituting PGA is L-glutamic acid or D-glutamic acid. As the said ratio, 92% or more is more preferable, 95% or more is more preferable, and 98% or more is further more preferable. Moreover, it is preferable that most of glutamic acid which comprises PGA is L-glutamic acid.

  The molecular size of the PGA used is not particularly limited, but those having an average molecular mass of 10 kDa or more are suitable. In general, the larger the molecular size, the higher the performance such as strength. On the other hand, PGA with an excessively large molecular size is expensive to manufacture and may be technically difficult to manufacture, so it is usually set to 1,000 kDa or less.

  If there exists what is marketed, PGA may use it and may manufacture it separately. However, since poly-α-glutamic acid is obtained when glutamic acid is polymerized under normal conditions, it is preferably biosynthesized using a microorganism. As a microorganism capable of producing PGA having a large molecular size, there is Natalba aegyptica which is a hyperhalophilic archaea.

  As PGA as a raw material, a salt thereof may be used. Examples of the salt include alkali metal salts such as sodium salt and potassium salt; alkaline earth metal salts such as calcium salt and magnesium salt. Further, even when a salt is used, it is not necessary that all carboxy groups are salts, and only a part thereof may be a salt. However, since polyvalent metal salts such as alkaline earth metal salts may have low solubility in water, a PGA free body or a monovalent metal salt of PGA is preferably used.

  The quaternary ammonium ion compounds (II) and (III) according to the present invention are composed of a tertiary amine and a long-chain alkyl group, and are generally used as a surfactant or a phase transfer catalyst. Is. In particular, the quaternary ammonium ion compound (III) is also used as an antibacterial agent, bactericidal agent, and antifungal agent.

  As the PGA ion complex according to the present invention, one that is sufficiently modified with a quaternary ammonium salt is suitable. More specifically, the ratio of the quaternary ammonium ion compound in the PGA ion complex is preferably 0.5 times mol or more, and 0.6 times mol or more with respect to glutamic acid constituting PGA. More preferably, it is 0.7 times mole or more.

In particular, those containing equimolar or substantially equimolar amounts of glutamic acid and quaternary ammonium ion compound constituting PGA are suitable. If the quaternary ammonium ion compound in the PGA ion complex is equimolar or substantially equimolar with the glutamic acid constituting the PGA, that is, the side chain carboxy group, the PGA can be sufficiently modified. Here, “substantially equimolar” means that the number of moles of both is substantially equal. Specifically, the quaternary ammonium ion compound with respect to glutamic acid constituting PGA is 0.8 times mole or more and 1.2 times mole. It shall be said that it is below mol, especially 0.9 times mol or more and 1.1 times mol or less.

  In addition, a quaternary ammonium ion compound may be used individually and may be used in mixture of 2 or more types. Further, a quaternary ammonium ion compound (II) and a quaternary ammonium ion compound (III) may be mixed and used. As the quaternary ammonium salt, compound (III) is more preferable.

  Quaternary ammonium ion compounds are usually present as halide salts. Therefore, in the present invention, the quaternary ammonium ion compound salt may be added directly to the reaction solution, or may be added after dissolving the salt in an aqueous solvent. The quaternary ammonium ion compound is preferably used in a sufficient amount relative to PGA in order to sufficiently modify PGA.

  The PGA ion complex of the present invention can be produced very easily only by mixing PGA and a quaternary ammonium ion compound in a solvent.

  As the solvent used here, water is suitable. This is because the raw material PGA can be dissolved well and the target compound PGA ion complex is insoluble in water, which is convenient for isolation and purification of the target product after the reaction. However, depending on the water solubility of the quaternary ammonium ion compounds, in order to increase their solubility in the reaction solution, alcohol solvents such as methanol and ethanol; ether solvents such as THF; amide solvents such as dimethylformamide and dimethylacetamide A water-soluble organic solvent such as may be added to the reaction solution. However, in consideration of recovery after completion of the reaction, it is preferable to use only water as the solvent.

  Since the PGA ion complex of the present invention is insoluble in water and can be easily separated from the aqueous solvent, the concentration of each component in the reaction solution is not particularly limited. For example, the concentration of PGA in the reaction solution is about 0.5 w / v% or more and about 10 w / v% or less, and the concentration of the quaternary ammonium ion compound is about 1.0 w / v% or more and about 10 w / v% or less. Can do.

  The reaction solution is preferably heated moderately in order to improve the dispersibility of the quaternary ammonium ion compound. The heating temperature can be, for example, about 40 ° C. or higher and 80 ° C. or lower. The reaction time may be adjusted as appropriate, but can usually be about 1 hour or more and 24 hours or less.

  Since the PGA ion complex of the present invention is insoluble in water, it can be easily separated from an aqueous solvent by filtration or centrifugation. The separated PGA ion complex can be washed with water to remove excess PGA, quaternary ammonium ion compound, and other salts. The aqueous solvent can be easily removed by washing with acetone or the like.

(2) Reaction step Next, the side chain carboxy group of poly-γ-glutamic acid is modified by reacting the PGA ion complex obtained above with the amine compound (IV) in an organic solvent. In the method of the present invention, the reaction proceeds even without using a dehydrating condensing agent such as WSC. The reason is not necessarily clear, but in the PGA ion complex solution, the ionized PGA is dissolved in an organic solvent by the action of the quaternary ammonium salt, so that the ionized PGA is more stabilized. It is considered that the reaction equilibrium is greatly inclined in the direction of reacting with an amine compound having a higher reactivity than the quaternary ammonium salt to form an amide. Moreover, since the reaction proceeds in an organic solvent, there is also a factor that the dehydration reaction easily proceeds.

  Since the amine compound (IV) has a simple chemical structure, if it is commercially available, it can be used, or can be easily synthesized from a commercially available compound by methods known to those skilled in the art.

  Conditions for reacting the PGA ion complex with the amine compound (IV) can be appropriately adjusted. For example, a solution of PGA ion complex can be used.

The solvent for dissolving the PGA ion complex is not particularly limited as long as it can sufficiently dissolve the PGA ion complex and does not inhibit the amidation reaction with the amine compound (IV). _C1-6 alcohol solvents, such as methanol, ethanol, n-propanol, isopropanol, n-butanol, can be mentioned from the outstanding solubility with respect to a complex, and safety | security. Of these, C _1-4 alcohol solvents are preferred, C _1-2 alcohol solvents are more preferred, and ethanol is even more preferred.

  The concentration of the PGA ion complex solution is not particularly limited and may be adjusted as appropriate. For example, it is preferably 0.1 w / v% or more and 20 w / v% or less. If the said density | concentration is 0.1 w / v% or more, reaction can be advanced with sufficient efficiency. On the other hand, if the concentration is too high, it may be difficult to dissolve the PGA complex. Therefore, the concentration is preferably 20 w / v% or less. The concentration is more preferably 0.5 w / v% or more, further preferably 1 w / v% or more, more preferably 15 w / v% or less, and still more preferably 10 w / v% or less.

  The addition amount of the amine compound (IV) may be appropriately adjusted depending on the use amount of the PGA ion complex, and may be 0.01 or more and 5 or less in mass ratio with respect to the PGA ion complex, for example. If the said mass ratio is 0.01 or more, reaction can be advanced with sufficient efficiency. On the other hand, if the mass ratio is too high, the reaction efficiency may possibly be lowered. Therefore, the mass ratio is preferably 5 or less. The mass ratio is more preferably 0.05 or more, further preferably 0.1 or more, particularly preferably 0.4 or more, more preferably 4 or less, still more preferably 2 or less, and particularly preferably 1 or less.

  Moreover, you may determine the addition amount of amine compound (IV) according to the desired modification rate of the side chain carboxy group in the biodegradable hydrogel (side chain carboxy group modification PGA) finally obtained. That is, for example, the number of moles of total glutamic acid constituting the PGA is calculated from the average molecular weight and amount of the PGA used, and the theoretical required amount of the amine compound (IV) capable of obtaining a desired modification rate is calculated. The amine compound (IV) may be used in an amount of 0.5 to 2 mol, more preferably 1 to 1.8 mol. In the present invention, the desired modification rate of the side chain carboxy group refers to the number of moles of the modified side chain carboxy group, the number of moles of total glutamic acid constituting the PGA used, that is, the total number of moles of the side chain carboxy group. Divide by number and say percentage multiplied by 100. Here, since PGA is a polymer, the terminal α-carboxy group is ignored. The number of moles of the modified side chain carboxy group can be determined by solid-state nuclear magnetic resonance analysis or the like.

  The amine compound (IV) may be added to the PGA ion complex solution as it is, or may be added once the solution is formed.

The solvent for dissolving the amine compound (IV) may be appropriately selected. For example, water; organic acid solvents such as formic acid and acetic acid; C _1-6 alcohols such as methanol, ethanol, n-propanol, isopropanol, and n-butanol Solvents; sulfoxide solvents such as dimethyl sulfoxide; amide solvents such as N-methylpyrrolidone, dimethylformamide and dimethylacetamide; ether solvents such as diethyl ether and tetrahydrofuran; mixed solvents of these two or more. However, depending on the type and amount of the solvent, a PGA ion complex having low solubility may be precipitated or decomposed. Therefore, the solvent may be the same as that of the PGA ion complex, or may be an amine compound (IV ) It is preferable to suppress the amount as much as possible by increasing the concentration of the solution.

  The reaction in this step proceeds very rapidly and the solubility of the resulting reaction product is low. Therefore, it is also possible to control the form of the reaction product obtained by means of adding the amine compound (IV) solution. For example, when an amine compound (IV) solution is extruded from a nozzle and added to a PGA ion complex solution, a fibrous material having a desired diameter can be obtained by adjusting the nozzle diameter.

  In contrast to the above, the PGA ion complex may be added to the amine compound (IV) solution as it is or in a solution state.

In this case, the solvent for dissolving the amine compound (IV) is preferably the same as the solvent for dissolving the PGA ion complex, such as methanol, ethanol, n-propanol, isopropanol, n-butanol, etc. It is preferable to use a C _1-6 alcohol solvent. Of these, C _1-4 alcohol solvents are preferred, C _1-2 alcohol solvents are more preferred, and ethanol is even more preferred.

  When the PGA ion complex is added as it is to the amine compound (IV) solution, the PGA ion complex dissolves in the solvent and a reaction with the amine compound (IV) occurs to precipitate a reaction product.

  What is necessary is just to adjust the reaction temperature and reaction time of this process suitably. For example, if the reaction is performed at room temperature, temperature control is not necessary, and it is suitable for industrial mass production. The reaction time can be about 10 minutes or more and about 50 hours or less after the PGA complex and the amine compound (IV) are mixed. Further, the reaction mixture may be left standing or stirred.

(3) Post-treatment step Next, the reaction product obtained in the above step, that is, the PGA in which the side chain carboxy group is amidated with the amine compound (IV) is isolated.

  PGA in which the side chain carboxy group is amidated with amine compound (IV) exhibits extremely high hydrophilicity, but has low solubility in a general solvent. Therefore, the product precipitated from the reaction mixture can be separated from the reaction solution by centrifugation or the like, or attached to an instrument such as a spatula and collected.

The isolated product is then preferably washed with an organic solvent. As the organic solvent used for washing, those showing solubility in PGA ion complex, amine compound (IV) and the like such as C _1-6 alcohol solvent are preferable.

  Furthermore, it is preferable to dry by dehydration treatment with anhydrous acetone or the like, or under reduced pressure.

  The biodegradable hydrogel produced by the method of the present invention described above is very safe because it exhibits biodegradability, and does not give an environmental load at the time of disposal. It can absorb and retain water as much as 1000 times its mass. In addition, it exhibits extremely excellent gel characteristics. Therefore, it can be expected to be applied to adhesives, wound dressings, agricultural water retention materials, water absorption materials and the like that can be used in vivo.

  The modification rate of the side chain carboxy group of the biodegradable hydrogel according to the present invention is preferably 5% or more and 90% or less. If the modification rate is 5% or more, the gel begins to take shape. Moreover, if the said modification rate is 10% or more, it will fully hydrogel. On the other hand, the upper limit of the modification rate is not particularly limited and may be 100%. However, since it may be difficult to make the modification rate 100%, it is preferably 90% or less. The modification rate is more preferably 20% or more, further preferably 30% or more, more preferably 80% or less, still more preferably 70% or less, and particularly preferably 60% or less.

  EXAMPLES Hereinafter, the present invention will be described more specifically with reference to examples. However, the present invention is not limited by the following examples, but may be appropriately modified within a range that can meet the purpose described above and below. Of course, it is possible to implement them, and they are all included in the technical scope of the present invention.

Example 1
(1) Preparation of PGA ion complex Poly-γ-glutamic acid sodium salt (10 g) derived from aegypticaca and having an average molecular weight of 1000 kD was dissolved in purified water to obtain a 2 w / v% solution. An equal amount of a 0.2M aqueous solution of hexadecylpyridinium bromide (HDPB) kept at 60 ° C. was added to the solution. After confirming that a water-insoluble material was formed immediately after HDPB addition, the mixture was further kept at 60 ° C. for 4 hours. The obtained water-insoluble material was collected by filtration and then washed 3 times with 100 mL of hot water. Furthermore, it dehydrated by washing | cleaning with acetone, Then, it vacuum-dried.

  The obtained water-insoluble material was acid-hydrolyzed and analyzed by chiral resolution HPLC according to a conventional method. As a result, 90% or more of the side chain carboxy group of the poly-γ-glutamic acid sodium salt used as a raw material was modified. I understood.

(2) Preparation of PGA ion complex solution Add the PGA ion complex (2 g) obtained in (1) above to absolute ethanol (50 mL), mix by inverting the container at room temperature until completely dissolved, and add 4 w / v % Solution.

(3) Preparation of dopamil PGA Separately, dopamine hydrochloride (0.85 g) was dissolved in absolute ethanol (100 mL) to obtain a 0.85 w / v% solution. When the PGA ion complex solution (5 mL) obtained in (2) above was filled into a syringe, poured into the dopamine solution (10 mL), and allowed to stand at room temperature, a fibrous substance was produced. The fibrous material was collected by centrifugation, washed 3 times with absolute ethanol (10 mL), and then dehydrated by immersion in acetone. The fibrous substance was separated by centrifugation and then vacuum dried to obtain a reaction product of PGA and dopamine (hereinafter referred to as “dopamil PGA”). This experiment was repeated and the standing time was changed from 30 minutes to 24 hours, but the obtained reaction product was not changed.

  The PGA ion complex obtained in the above (1) was yellowish, whereas the obtained dopamil PGA was white.

Example 2
The PGA ion complex (10 mg) obtained in Example 1 (1) was charged at room temperature into an Eppendorf tube containing a saturated ethanol solution of dopamine hydrochloride (500 μL) prepared at room temperature (FIG. 1 (1)). As a result, the PGA ion complex is dissolved in ethanol, and the modification reaction with dopamine that occurs instantaneously is accompanied by the property that the reaction product of PGA and dopamine is insoluble in alcohol. The fine fibrous material (arrow part of FIG. 1 (2)) formed by (1) was wound up with a pipette and collected (FIGS. 1 (2) to (3)). This operation was performed until the PGA ion complex disappeared (FIG. 1 (4)). The collected fine fibrous material was washed and dried in the same manner as in Example 1 (3).

Test Example 1 Gel Characteristic Test Dopamil PGA obtained in Example 2 is put in a 20 mL container with a lid, and 100 times, 250 times, 300 times, 500 times, 1000 times, or 2000 times water is added by mass ratio, respectively. The lid was shaken and allowed to stand. Fig. 2 shows a photograph of these containers turned upside down.

  As shown in FIG. 2, the dopamil PGA according to the present invention was able to hold water at least up to 1000 times in mass ratio, and did not show fluidity even when the container was inverted, and the gel state was maintained. . A general biodegradable hydrogel can only hold up to about 10 times the water inside, but the biodegradable hydrogel according to the present invention has a far greater water absorption capacity than the conventional biodegradable hydrogel. The water absorption was comparable to that of the chemical hydrogel.

  Further, when water of 2000 times by mass was added to dopamil PGA, it was transferred to a highly viscous sol having fluidity. In general, the chemical conversion polymer does not show a sol-gel transition, which is a characteristic characteristic of biodegradable biogels. If it is a sol state or a non-viscous aqueous solution state, the treatment is very easy and the load on the environment at the time of disposal is small. It has been clarified that the hydrogel according to the present invention retains such characteristics of the biodegradable hydrogel while exhibiting high water absorption as described above.

  Incidentally, the water solubility of PGA is extremely high, and it takes an aqueous solution state even at a high concentration exceeding 10 w / w%. This unusually high hydrophilicity control has been a major issue when considering the practical application of PGA to industrial materials. Therefore, the hydrogel according to the present invention is also interesting as a model for such materialization.

Test Example 2 Frequency Sweep Test Whether the diluted solution of dopamil PGA exhibited gel property or sol property was examined by the Frequency Sweep test. According to this test, the presence or absence of gel property and sol property can be clarified by measuring dynamic viscoelasticity by changing the frequency under a certain amount of strain. Specifically, the property of recovering the mechanical energy applied to the sample is called elasticity (gel property), and can be represented by storage rigidity G ′. On the other hand, when the deformation is stopped in a state where a stress proportional to the deformation rate is generated, the stress becomes 0, and the property of remaining deformed is called viscosity (sol property). In this case, all of the applied mechanical energy is dissipated as heat, and this can be measured as the loss rigidity G ″. The viscoelasticity of the sample is the loss tangent tan δ = G ″ (ω) / G ′ (ω) [formula Where ω represents an angular velocity], and in rheology, when tan δ <1, it can be said that the sample is a gel, and the tendency for the applied energy to be stored in the sample becomes strong. On the other hand, when tan δ> 1, it can be said that the sample is a sol, and the energy applied to the sample is released as heat.

  The dopamil PGA obtained in Example 2 was placed in a 20-mL container with a lid, 1000 times or 3000 times the water in mass ratio was added, and the mixture was diluted with shaking after being covered. As in Test Example 1, apparently, the gel was maintained when diluted 1000-fold, but it was sol when diluted 3000-fold. Using a stress rheometer (TA Installation, AR-G2), the storage rigidity G ′ and the loss rigidity G ″ of each sample were determined by rheological analysis. Measurement was performed on a titanium parallel plate. The thickness was 400 μm, and the temperature was maintained at 25 ° C. by Peltier control, a plate having a diameter of 4 cm was used for measurement of a 1000-fold diluted dopamil PGA sample, and a plate having a diameter of 6 cm was used for a 3000-fold diluted dopamil PGA sample. (%) Is defined as a percentage of rω / h (where r represents a radius, ω represents an angular velocity, and h represents a sample thickness), and is fixed at 1% for 1000-fold diluted dopamil PGA. On the other hand, since the 3000-fold diluted dopamil PGA sample was too soft and it was difficult to analyze the low frequency region with a distortion amount of 1%, the distortion amount was fixed at 5%. . Frequency was varied from 0.01Hz to 10 Hz. The results are shown in FIG.

  As shown in FIG. 3, the G ′ value (●) of the 1000-fold diluted dopamil PGA sample was clearly larger than the G ″ value (◯), indicating a clear gel property. In the PGA sample, the difference between the G ′ value (▲) and the G ″ value (Δ) is clearly small, but tan δ <1. Therefore, it was apparent that the 3000-fold diluted dopamil PGA sample was in the form of a sol, but in terms of rheology it was a gel.

  As described above, the dopamil PGA according to the present invention can be expected to cause a sol-gel transition depending on the water content, but it exhibits excellent gel properties even when it contains 3000 times as much water. It was proved to show gel properties.

Test Example 3 Strain Sweep Test The strain sweep test changes the amount of strain under a certain frequency, and examines how much strain the gel state is destroyed. Therefore, a strain sweep test was performed on 1000-fold diluted dopamil PGA showing clear gel properties.

  The basic experimental conditions and equipment are the same as in Test Example 2, but in this test, the frequency was fixed at 1 Hz, the strain amount was changed from 0.01% to 3000%, the storage rigidity G 'and the loss rigidity ratio G ″ was measured. The results are shown in FIG.

  As shown in FIG. 4, in the case of a 1000-fold diluted dopamil PGA sample, the G ′ value (●) began to decrease around the strain amount of 3%, and reversed to the G ″ value (◯) at the strain amount of 15-20%. It is thought that the sol state or solution state was reached.

Test Example 4 Self-healing test The self-healing test is to examine the self-healing property of the gel by changing the amount of strain greatly at constant time under a constant frequency. The self-repairing property of the 1000-fold diluted dopamil PGA according to the present invention was examined.

  Specifically, as shown in FIG. 5 (1), for the 1000-fold diluted dopamil PGA according to the present invention, the strain amount starts from 1%, the strain amount becomes 100% after 302 seconds, and the strain amount becomes 1 after 598 seconds. %, The strain amount is set to 100% after 1120 seconds, the strain amount is set to 1% after 1490 seconds, the strain amount is set to 100% after 2090 seconds, and finally the strain amount is set to 1000% after 2085 seconds, The G ′ value and the G ″ value were measured while periodically switching the strain amount between 1% and 100%. The results are shown in FIG.

  As shown in FIG. 5 (2), at the moment when the strain amount was switched from 1% to 100%, the G ″ value (◯) exceeded the G ′ value (●) and became a sol state. The G ′ value and the G ″ value were instantaneously reversed, and the gel property was recovered. Such behavior could be confirmed over and over again. Further, the deterioration of the gel is manifested by a decrease in the net G ′ value, but here it was almost 100% recovered. In general chemical gels that are widely used at present, it is said that the gel structure will never return to the gel when subjected to stress that causes the gel structure to collapse. It has been demonstrated to have a very good gel property of restorability. Finally, as a result of trying to break and break reaching the polymer chain of dopamil PGA by applying a strong strain of 1000%, the self-healing gel properties completely disappeared. It was shown that the properties were attributed to the polymer structure region.

Test Example 5 Drying / Water Absorption History Characteristic Test A property that a material released from a specific stress tries to return to an initial state is called a hysteresis characteristic. For example, in the case of a hydrogel, if water absorption near the initial state is maintained even after repeated drying, the possibility of being useful as an excellent history gel material increases. This time, hysteresis characteristics relating to drying and water absorption were compared between agarose, which is a typical biogel contained in agar powder, and dopamil PGA according to the present invention.

Specifically, 2 mL each of 1 w / v% dopamil PGA gel and 1 w / v% agarose gel was allowed to stand in a glass petri dish (φ3.5 cm). Next, after sufficiently drying for 50 hours, the water absorption after 5 minutes from the addition of 2 mL of distilled water was calculated. The water absorption rate was calculated from the ratio of the remaining water amount to the added water amount. This operation was repeated up to 5 times.
The results are shown in FIG.

  As shown in FIG. 6, the agarose gel hardly absorbs water once dried (open bar). As described above, general polysaccharide biogels such as agarose gel have a problem of large hysteresis (property that hinders history behavior). On the other hand, in the dopamil PGA gel according to the present invention, it has been found that the water absorption performance is not lost even if drying and water absorption are repeated 5 times (black bar). As described above, in the dopamil PGA according to the present invention, an excellent return behavior with respect to water absorption was recognized.

Claims (4)

A method for producing a biodegradable hydrogel comprising:
A step of preparing an ion complex by adding a quaternary ammonium ion compound of the following formula (II) or formula (III) to a poly-γ-glutamic acid aqueous solution;
[Wherein R ^{1 to} R ³ independently represent a C _1-2 alkyl group, and R ^{4 to} R ⁵ independently represent a C _12-20 alkyl group]
The manufacturing method characterized by including the process of modifying the side chain carboxy group of poly-gamma-glutamic acid by making the said ion complex and the amine compound of following formula (IV) react in an organic solvent.
[Wherein X represents a linker group, and Y represents a C _6-12 aryl group having one or more hydroxyl groups] The production method according to claim 1 , wherein an alcohol solvent is used as the organic solvent. X is a C _1-6 alkylene group, or C having an amino group, an ether group, an amide group, a carbonyl group, an —O (C═O) — group or a — (C═O) O— group at the Y-side end. The production method according to claim 1 or 2 , wherein the amine compound (IV) which is a _1-6 alkylene group is used. The process according to any one of claims 1 to 3, Y is an amine compound is a phenyl group (IV) having one or more hydroxyl groups.