polymer components

Abstract

The present invention addresses the problem of providing a novel polymer composition which takes advantage of a bioaffinity attributable to the structure of 2-pyrrolidone, especially providing a polymer composition which, due to the inclusion of a novel structure including said 2-pyrrolidone structure, is water-insoluble.　The polymer composition includes a polymer comprising a given monomer unit having the structure of 2-pyrrolidone in a side chain, and is water-insoluble over a temperature region within the range of 0-100°C.

Description

[Technical Field]

[0001] The present invention relates to a polymer composition characterized by being less likely to cause foreign body reactions in a living body when brought into contact with biological-related substances such as a living body, biological tissues including blood, cells, and proteins. The present application claims priority based on Japanese Patent Application No. 2021-127929 filed with Japan on August 4, 2021, and incorporates the content thereof by reference. [Background Art]

[0002] Generally, when biological-related substances such as blood come into contact with the surface of various artificially synthesized materials, the surface of the material is recognized as a foreign substance. For example, non-specific adsorption of proteins to the material surface occurs, causing denaturation, and as a result, activation of the coagulation system, complement system, platelet system, etc. occurs. For this reason, for example, on the surface of medical devices used in contact with living bodies and biological-related substances, it is desired to impart biocompatibility to the surface of the device in order to prevent the device from being recognized as a foreign substance and causing a foreign body reaction in biological-related substances during use. Up to now, various materials exhibiting biocompatibility have been proposed and are being put into practical use in medical settings and the like.

[0003] For example, 2-methacryloyloxyethyl phosphorylcholine (MPC) polymer is a polymer obtained by polymerizing, as a monomer, a unit structure in which a structure imitating a constituent substance of a living body (phospholipid polar group) is bonded to a polymerizable group such as a vinyl group. It is known that applying a composition containing the MPC polymer to the surface of a medical device imparts biocompatibility, suppresses platelet adhesion and the like, and exhibits excellent antithrombotic properties. Furthermore, polyethylene glycol (PEG) is a polymer having -(C2H4-O)-, which is a chain ether structure, as a repeating unit. It is known that despite having a structure not similar to substances constituting a living body, it has extremely excellent biocompatibility.

[0004] Previous research is revealing the mechanism by which the aforementioned polymers and other materials exhibit biocompatibility despite being artificially synthesized. Specifically, it has been revealed that synthetic polymers and biologically derived substances that exhibit biocompatibility commonly contain water molecules in a state called "intermediate water" when they come into contact with the aqueous phase and undergo hydration. Furthermore, it has been observed that the adsorption and denaturation of proteins in the aqueous phase are prevented on the surface of materials where this "intermediate water" is present, and as a result, the activation of the coagulation system, complement system, platelet system, etc., is suppressed (see, for example, Non-Patent Document 1).

[0005] In other words, for a polymer or the like to exhibit biocompatibility, it is necessary for the hydration to occur in a form that produces water molecules in the form of intermediate water when it comes into contact with an aqueous phase. From this perspective, it is considered necessary for polymers or the like that exhibit biocompatibility to have a certain degree of hydrophilicity. Furthermore, by using the surface of a polymer or the like that contains water molecules in the form of intermediate water through hydration as a substrate for cell culture, it is possible to perform cell culture successfully (Patent Document 1).

[0006] On the other hand, polymers used for purposes such as imparting biocompatibility to the surface of medical devices are required to exhibit a certain degree of water insolubility in order to prevent the polymer from leaching out when it comes into contact with an aqueous phase such as blood. To achieve this, the polymer must possess a predetermined degree of hydrophobicity. In other words, polymers used for purposes such as imparting biocompatibility to the surface of medical devices require contradictory properties: they must be hydrophilic to contain intermediate water, while simultaneously being water-insoluble (hydrophobic) to prevent elution and dissolution.

[0007] Regarding the above-mentioned MPC polymer, since MPC homopolymer is water-soluble and difficult to use as a medical material, a technique is used to achieve both biocompatibility and water insolubility by copolymerizing it with hydrophobic constituent units (such as butyl methacrylate) (Patent Document 2). Also, since the above-mentioned PEG is water-soluble, its water resistance is improved by copolymerizing it with other constituent units (Patent Document 3) or by creating a cross-linked structure (Patent Document 4).

[0008] The above examples involve methods such as adding hydrophobic components that do not exhibit biocompatibility to biocompatible polymers in order to impart hydrophobicity to make them insoluble in water, or modifying parts of the original biocompatible polymer's structure. These methods raise concerns from a biocompatibility standpoint.

[0009] It is known that polymers having a 2-pyrrolidone structure with a five-membered ring lactam structure in their side chains exhibit good biocompatibility. Polyvinylpyrrolidone, which is commonly used as a polymer with a 2-pyrrolidone structure, exhibits a certain level of biocompatibility and, like the MPC polymer and PEG mentioned above, is water-soluble. For example, Patent Document 5 describes a technique for crosslinking polyvinylpyrrolidone by radiation in order to make it water-insoluble.

[0010] Furthermore, Patent Document 6 describes that N-2-(meth)acryloyloxyethyl-2-pyrrolidone, which is formed by providing the above-mentioned 2-pyrrolidone structure to the side chain portion of a main chain having a (meth)acrylic structure, exhibits biocompatibility and water solubility. It also states that in order to use this structure as a medical hydrogel, it is necessary to make it water-resistant by copolymerizing it with hydrophobic constituent units, but that biocompatibility decreases due to these hydrophobic constituent units.

[0011] The common characteristics exhibited by the various polymers described above suggest that the phenomenon of biocompatibility resulting from the inclusion of intermediate water through hydration of polymers is essentially similar to the phenomenon of water solubility of polymers, indicating a kind of trade-off relationship between biocompatibility and water solubility.

[0012] Furthermore, in relation to the water solubility of polymers, it is known that there are polymer compositions whose water-soluble / water-insoluble properties change depending on the temperature, as described in Patent Document 7, for example. Polymers exhibiting such properties are called temperature-responsive polymers (or temperature-sensitive polymers), and it is understood that the change in water-soluble / water-insoluble properties occurs due to a discontinuous change in the morphology of the molecular chains in the aqueous phase, caused by the temperature dependence of the interfacial energy between the polymer chains and the aqueous phase.

[0013] Regarding the temperature at which the above-mentioned water-soluble / water-insoluble transition occurs, if water solubility is observed at a temperature lower than the transition temperature, that temperature is called the Lower Critical Solution Temperature (LCST), and if water solubility is observed at a temperature higher than the transition temperature, that temperature is called the Upper Critical Solution Temperature (UCST). [Prior art documents] [Patent Documents]

[0014] [Patent Document 1] Japanese Patent Publication No. 2016-63801 [Patent Document 2] Japanese Patent Application Publication No. 3-39309 [Patent Document 3] Japanese Patent Publication No. 2005-23108 [Patent Document 4] Japanese Patent Publication No. 2005-255875 [Patent Document 5] Japanese Patent Application Publication No. 2-86838 [Patent Document 6] Japanese Patent Application Publication No. 4-28705 [Patent Document 7] WO2004 / 087228 pamphlet [Non-Patent Literature]

[0015] [Non-Patent Literature 1] Tanaka, M . et al., Journal of Biomaterials Science Polymer Edition, 2010, Vol. 21, pp. 1849-1863 [Non-Patent Literature 2] Masamichi Nakayama, Drug Delivery System, Vol. 23, No. 6, 2008, pp. 627-636 [Summary of the Invention] [Problem to be Solved by the Invention]

[0016] In medical devices and the like for which it is desired to impart biocompatibility through surface coating with various compositions or the like, it is required that not only good biocompatibility is imparted to the surface, but also a wide variety of properties suitable for the intended use are exhibited. In addition, applications other than coating the surface of medical devices and the like also demand various polymer compositions that exhibit biocompatibility and have various properties. Furthermore, in order to exhibit good biocompatibility and exert various properties suitable for various applications, it is considered necessary to appropriately optimize the degree and content of hydrophilicity of the above-mentioned polymer compositions.

[0017] An object of the present invention is to provide a novel polymer composition that utilizes biocompatibility derived particularly from the 2-pyrrolidone structure described above, and in particular, an object of the present invention is to provide a polymer composition that is water-insoluble by having a novel structure containing the 2-pyrrolidone structure. [Means for Solving the Problem]

[0018] In order to solve the above problem, the present invention provides a polymer composition comprising a polymer containing a monomer unit represented by the following structural formula (2), wherein the polymer composition has a temperature range of 0 to 100°C in which it is water-insoluble.

[0019] The present invention also provides a polymer composition comprising a polymer containing two or more structures having mutually different values of m in the following structural formula (2). The present invention also provides a polymer composition comprising a polymer containing a structure in which m in the following structural formula (2) is 5 or more.

[0020] The present invention also provides a polymer composition comprising a polymer containing a monomer unit represented by the following structural formula (2), wherein the polymer composition is water-insoluble over the entire temperature range of 0 to 100°C. The present invention also provides a polymer composition comprising a polymer containing a monomer unit represented by the following structural formula (2), wherein the polymer composition has a temperature range within 0 to 100°C where a transition between water solubility and water insolubility occurs, the polymer composition is water-insoluble at temperatures equal to or higher than the temperature range, and exhibits water solubility at temperatures equal to or lower than the temperature range. The present invention also provides a polymer composition, wherein the proportion of monomer units represented by the following structural formula (2) in the polymer is 80% or more. Effects of the Invention

[0021] According to the present invention, there is provided a polymer composition whose degree of hydrophilicity can be arbitrarily adjusted, which exhibits water insolubility and the like under predetermined conditions while maintaining biocompatibility. Brief Description of Drawings

[0022] [Figure 1] FIG. 1 is a diagram schematically showing the hydrophilic transition temperature of homopolymers obtained by changing the value of m in structural formula 2. [Figure 2] It is a photograph showing that an aqueous solution of a homopolymer with m=1, 3, 4 in structural formula 2 colloidalizes depending on temperature. [Figure 3]This graph shows the change in light transmittance when the temperature of an aqueous solution of a homopolymer with m values ​​of m=1, 3, and 4 in structural formula 2 is changed. [Figure 4] This graph shows the change in light transmittance when the temperature is changed for each aqueous solution of the homopolymer with varying solubility concentrations, where the m value in structural formula 2 is set to m=4. [Figure 5] This graph shows the change in light transmittance when an aqueous solution of a copolymer between constituent units with m values ​​of m=2 and m=6 in structural formula 2 is subjected to temperature changes. [Figure 6] This graph shows the platelet adhesion on the surface of the polymer composition, etc., according to the present invention. [Figure 7] This graph shows the hemolytic properties of the polymer composition, etc., according to the present invention in an aqueous solution. [Figure 8] This graph shows the amount of protein adsorbed on the surface of the polymer composition, etc., according to the present invention. [Modes for carrying out the invention]

[0023] As described above, conventional homopolymers containing the 2-pyrrolidone structure in their side chains are generally known to exhibit biocompatibility and water solubility, regardless of structural differences other than the 2-pyrrolidone structure. The phenomena exhibited by polymers containing the 2-pyrrolidone structure are thought to be due to the hydrophilicity of the 2-pyrrolidone structure. When the polymer absorbs water, a predetermined hydration structure is formed, containing water molecules in an intermediate water state. Furthermore, the hydrophilicity of the 2-pyrrolidone structure surpasses the hydrophobicity of other structural parts within the polymer, resulting in miscibility with the aqueous phase at the molecular level.

[0024] The inventors of the present invention have investigated various structures other than the 2-pyrrolidone structure portion of a constituent unit (monomer unit) that partially has a 2-pyrrolidone structure. As a result, they have found that a polymer containing the constituent unit shown in structural formula 2 below behaves as a temperature-responsive polymer. By utilizing this behavior, they have found a polymer composition that maintains biocompatibility derived from the 2-pyrrolidone structure while exhibiting water-insoluble properties at least within a predetermined temperature range. In structural formula 2, R1 is either H or CH3, and R2 is either O or NH. The m value indicates the number of carbon atoms in the carbon chain that connects R2 and the 2-pyrrolidone structure, and the n value indicates the number of repeats of the constituent unit.

[0025] [ka]

[0026] In other words, when a polymer containing the structural units shown in structural formula 2 above is in contact with an aqueous phase, it was found that the polymer tends to be water-insoluble and phase-separate from the aqueous phase at high temperatures, while at low temperatures it tends to be water-soluble and easily miscible with the aqueous phase. This indicates that it has a so-called lower critical solution temperature (LCST), and exhibits the behavior of a temperature-responsive polymer in which its water-soluble / water-insoluble properties reversibly change depending on the temperature.

[0027] Furthermore, by creating a polymer composition containing constituent units with various m values ​​set in structural formula 2, it is possible to set the LCST within the range of 0 to 100°C, making the polymer water-insoluble at temperatures above the LCST, and even making it water-insoluble throughout the entire range of 0 to 100°C. This makes it possible to construct a water-insoluble polymer composition that can be used in various applications where biocompatibility is required.

[0028] The mechanism by which polymers containing the structural units shown in structural formula 2 behave as temperature-responsive polymers is presumed to be as follows: In structural formula 2, the part related to the m value is the carbon chain that connects the main chain portion of the polymer to the nitrogen atom contained in 2-pyrrolidone, and this carbon chain portion is thought to be hydrophobic. On the other hand, the 2-pyrrolidone portion has strong hydrophilicity, and it is considered that the overall hydrophilicity / hydrophobicity of the polymer is determined by the balance between the two.

[0029] Furthermore, the degree of hydrophobicity of the carbon chain portion and the degree of hydrophilicity of the 2-pyrrolidone portion each have their own unique temperature dependence, and as a result, the hydrophilicity / hydrophobicity of the polymer as a whole changes with temperature, resulting in temperature responsiveness. Moreover, by changing the m value in structural formula 2, it is possible to change the degree of hydrophobicity originating from the carbon chain portion, and thus it is possible to change the transition temperature between water solubility and water insolubility of the polymer as a whole, making it possible to obtain a polymer that exhibits water insolubility in a predetermined temperature range. The carbon chain constituting the linkage portion can be saturated or unsaturated, and can be linear or branched. For example, in addition to having an alkylene group as the linkage portion, it can also be a carbon chain including branches, with a structure in which 2-pyrrolidone is bonded to at least one of the ends of the branched carbon chain.

[0030] N-2-(meth)acryloyloxyethyl-2-pyrrolidone, which is formed by providing the 2-pyrrolidone structure described in Patent Document 6 to the side chain portion of a main chain having a (meth)acrylic structure, corresponds to a constituent unit where the m value in the above structural formula 2 is m=2. While its homopolymer exhibits good biocompatibility, it is also water-soluble, making it difficult to use as a material for the surface of medical devices, etc.

[0031] On the other hand, as shown in the examples and comparative examples of the present invention below, homopolymers in which the polymer main chain of the structural unit shown in structural formula 2 has an acrylic structure and the m value is m=1, 3, and 4 exhibit the behavior of a temperature-responsive polymer with lower critical solution temperatures (LCST) of approximately 30.2°C, 65.3°C, and 29.7°C, respectively, in pure water, and it was observed that they exhibit water insolubility at temperatures higher than the LCST. Furthermore, in homopolymers with an m value of m≧5, water insolubility was observed throughout the temperature range of 0 to 100°C, and it was shown that homopolymers obtained by changing the m value in the structural unit shown in structural formula 2 exhibit a temperature range in which they are water insolubility.

[0032] Furthermore, regarding homopolymers (m=2, 5, 6, etc.) in which LCST is not observed in the temperature range of 0 to 100°C where an aqueous phase can exist, as shown in the structural formula 2 above, the present invention has found that in copolymers formed between m=2 structural units, which are water-soluble throughout the entire range, and m=6 structural units, which are water-insoluble throughout the entire range, LCST is expressed within the temperature range of 0 to 100°C (see Table 20 below, etc.).

[0033] The phenomenon in which properties change when copolymers are formed between constituent units with different m values ​​is thought to be a result of intermediate properties being exhibited regarding the properties such as the water-soluble / water-insoluble transition temperature inherent to each constituent unit, due to the coexistence of constituent units with different m values ​​within the polymer. Furthermore, the LCST observed in the above copolymer is thought to suggest that the m=2 constituent unit has a potential water-soluble / water-insoluble transition temperature above 100°C, making it water-soluble throughout the temperature range of 0 to 100°C, while the m=6 constituent unit has a potential water-soluble / water-insoluble transition temperature below 0°C, making it water-insoluble throughout the temperature range of 0 to 100°C.

[0034] In other words, the present invention is based on the discovery that a polymer containing the structural unit shown in the above structural formula 2 has a transition temperature between water-soluble and water-insoluble as a temperature-responsive polymer, and that this transition temperature can be adjusted by setting the m value. By setting various m values ​​in the form of a homopolymer or copolymer, the present invention provides a polymer that exhibits water insolubility at temperatures above 0 to 100°C by setting the transition temperature (LCST) within the range of 0 to 100°C, and further provides a polymer that exhibits water insolubility throughout the entire range of 0 to 100°C by setting the transition temperature below 0°C.

[0035] The above-mentioned transition between water-soluble and water-insoluble substances is caused by a rapid change in the solubility of polymers and other substances in the aqueous phase depending on the temperature, and this transition occurs over a predetermined temperature range. In this invention, as an index for evaluating each polymer, for example, the temperature at which significant dissolution / precipitation of the polymer is observed in an aqueous solution with a polymer concentration of 1 wt% is described as the water-soluble / water-insoluble transition temperature. In this invention, this water-soluble / water-insoluble transition temperature may also be referred to as the "hydrophilization temperature."

[0036] Furthermore, when it is stated in the present invention that a polymer is water-insoluble, it means that when the polymer is brought into equilibrium with the aqueous phase in the environment in which it is used, the phase containing the polymer can separate from the aqueous phase to form a separate phase. Typically, this means that the saturation solubility of the polymer in an aqueous solution is 0.5 wt% or less, preferably 0.1 wt% or less, which allows for the formation of a separate phase separated from the aqueous phase. Moreover, it is not limited to the case that the phase containing the polymer that exists separately from the aqueous phase is a liquid phase or a solid phase.

[0037] For example, when an aqueous solution containing the polymer composition according to the present invention is heated to a temperature above the hydrophilization temperature to make it water-insoluble, in addition to the case where the polymer composition consisting of a powdery solid phase precipitates, a form in which the hydrated polymer composition forms a fluid phase and separates from the aqueous phase and precipitates is observed. In the present invention, the formation of a separate phase by separating from the aqueous phase in such a fluid form is also included as one form of water-insolubleness. Furthermore, in this invention, the terms monomer and monomer, polymer and polymer, and high polymer are used interchangeably, and a composition containing such high polymer is referred to as a high polymer composition. In addition, the terms copolymer and copolymer are used interchangeably in reference to high polymers, and are used to distinguish them from homopolymers which are substantially formed by polymerizing a single constituent unit.

[0038] Figure 1 schematically shows the hydrophilization temperatures of homopolymers obtained by changing the m value in structural formula 2 above. As described above, for homopolymers with m values ​​of m=1, 3, and 4, a hydrophilization temperature (LCST) at which water solubility / insolubility transitions is observed within the temperature range of 0 to 100°C, where the aqueous phase can exist. On the other hand, as shown in Figure 1, the fact that the homopolymer with m=2 is water-soluble throughout the 0 to 100°C range indicates that the hydrophilization temperature at which its water solubility / insolubility changes exists above 100°C. Conversely, it is considered that homopolymers with m≧5 exhibit water insolubility because they have a potential hydrophilization temperature below 0°C.

[0039] The polymer composition according to the present invention exhibits good biocompatibility regardless of the m value in structural formula 2, and can therefore be suitably used in applications where the water-soluble / water-insoluble state is changed by temperature changes, etc., while ensuring biocompatibility, or in applications where it is used in a water-insoluble state.

[0040] In the polymer composition according to the present invention, the m value in the constituent unit shown in structural formula 2 is not particularly limited, and by including constituent units having an appropriate m value within a range that does not hinder the effects of the polymer composition according to the present invention, a homopolymer or copolymer that exhibits water insolubility in at least a predetermined temperature range can be obtained and used as a polymer composition containing such units.

[0041] The polymer composition according to the present invention can be used as a homopolymer obtained by polymerizing structural units having a predetermined m value in the structural unit shown in structural formula 2. In particular, by expanding the m value, the polymer composition according to the present invention exhibits water insolubility in the homopolymer state over the entire range of 0 to 100°C, and is therefore preferably used as a coating agent for coating the surfaces of various medical devices and the like. In the homopolymer, flexibility decreases due to polymer crystallization and the like as the m value expands, so in this embodiment, it is preferable to use structural units with an m value of m=10 or less, or m=8 or less.

[0042] On the other hand, in the polymer composition according to the present invention, by using a copolymer containing structural units shown in structural formula 2, which have particularly different m values, in any ratio, it is possible to obtain a polymer composition having intermediate properties between those exhibited by homopolymers composed of each of the structural units used. By utilizing this, it is possible to construct a polymer composition having a hydrophilization temperature at which a transition between water-soluble and water-insoluble states occurs at a desired temperature, and for example, a polymer composition that can be used as a water-insoluble film at a predetermined temperature above the hydrophilization temperature.

[0043] When synthesizing the above copolymer, for example, a component with m=2 that is water-soluble throughout the range of 0 to 100°C can be used as a component having a hydrophilic temperature of 100°C or higher, and a component with m≧5 that is water-insoluble throughout the range of 0 to 100°C can be used as a component having a hydrophilic temperature of 0°C or lower. When synthesizing the copolymer, component units with m values ​​of 10 or higher may be included, particularly for the purpose of imparting additional properties to the polymer.

[0044] The polymer composition according to the present invention only needs to have a temperature range within 0 to 100°C in which it exhibits non-water solubility in an aqueous solution used in contact with the polymer composition, and particularly includes those that are non-water solubility throughout the entire temperature range of 0 to 100°C under the usage environment. Furthermore, the polymer composition according to the present invention can include homopolymers or copolymers in which the hydrophilization temperatures at which the water-soluble / non-water-soluble state transitions are mutually different, and can be a polymer composition having multiple hydrophilization temperatures, or a polymer composition with a broad hydrophilization temperature range and exhibiting a slow water-soluble / non-water-soluble transition behavior.

[0045] The polymer composition according to the present invention is a copolymer composed solely of structural units that share a common basic structure. By adjusting the blending ratio of these structural units, it is possible to change the transition behavior between water-soluble and water-insoluble substances. This makes it less likely for microscopic phase separation to occur within a single molecule or within the composition, and enables the creation of a uniform composition.

[0046] Furthermore, in addition to polymers composed solely of the structural units shown in structural formula 2, the polymer composition according to the present invention can be imbued with various properties by adding structural units other than those shown in structural formula 2, to the extent that they do not hinder the effects of the polymer composition according to the present invention, for purposes such as adjusting the hydrophilization temperature or the degree of hydration.

[0047] Examples of structures other than the constituent units shown in structural formula 2 above include, for example, aminoalkyl acrylates such as aminomethyl acrylate, aminoethyl acrylate, and aminoisopropyl acrylate; diaminoalkyl acrylates such as diaminomethyl acrylate, diaminoethyl acrylate, and diaminobutyl acrylate; aminoalkyl methacrylates such as aminomethyl methacrylate and aminoethyl methacrylate; diaminoalkyl methacrylates such as diaminomethyl methacrylate and diaminoethyl methacrylate; methyl acrylate, ethyl acrylate, isopropyl acrylate, butyl acrylate, hexyl acrylate, 2- Examples include alkyl acrylates such as ethylhexyl acrylate, alkyl methacrylates such as methyl methacrylate, ethyl methacrylate, butyl methacrylate, and hexyl methacrylate, alkoxy(meth)acrylates such as methoxy(meth)acrylate, alkoxyalkyl(meth)acrylates such as methoxyethyl(meth)acrylate, glycidyl methacrylate, acrylamide, alkyl acrylamides such as t-butyl acrylamide, n-butyl acrylamide, i-butyl acrylamide, hexyl acrylamide, and heptyl acrylamide, N,N-dialkyl acrylamides such as N,N-dimethyl acrylamide and N,N-diethyl acrylamide, methacrylamide, N,N-dialkyl methacrylamides such as N,N-dimethyl methacrylamide and N,N-diethyl methacrylamide, and propylene.

[0048] Furthermore, it is possible to use (meth)acrylamide compounds that are generally known to exhibit LCST without containing intermediate water, N-acrylic-substituted (meth)acrylamide derivatives such as N-isopropyl(meth)acrylamide, N,N-dialkyl-substituted (meth)acrylamide derivatives such as N,N-dimethyl(meth)acrylamide, and N-heterocyclic-substituted (meth)acrylamide derivatives such as 1-(1-oxa-2-propenyl)-pyrrolidine.

[0049] In the polymer composition according to the present invention, the proportion of constituent units other than those shown in structural formula 2 can be determined according to its intended use. On the other hand, from the viewpoint of ensuring uniformity based on the fact that it is composed only of constituent units that share a common basic structure, it is preferable that the proportion of constituent units shown in structural formula 2 be, as a molar ratio between monomer units, for example, 80 mol% or more, and more preferably greater than 80 mol%, such as 85 mol% or more, 90 mol% or more, or 95 mol% or more.

[0050] Furthermore, in the polymer composition according to the present invention, the constituent units with particularly large m values ​​in structural formula 2 exhibit good biocompatibility and high water insolubility. This allows them to be used, for example, as water-insolubility agents to impart water insolubility to water-soluble polymers such as MPC polymers. Specifically, a polymer can be obtained by copolymerizing constituent units that exhibit water insolubility in a homopolymer state, such as those with an m value of m≧5, with constituent units other than those shown in structural formula 2 in an appropriate proportion.

[0051] The proportion of a homopolymer unit exhibiting water insolubility in the state of m≧5, etc., when copolymerizing it with other units is not particularly limited. For example, the molar ratio between monomer units can be 5 mol% or more. By setting this molar ratio to 10 mol% or more, 30 mol% or more, or 50 mol% or more, a polymer that better reflects the characteristics of the structural unit shown in structural formula 2 can be obtained.

[0052] Furthermore, the polymer composition according to the present invention includes not only the polymer composition in its anhydrous state, but also hydrates and the like obtained by hydrating the polymer composition. Within the scope of the objectives of the present invention, the polymer composition according to the present invention can be used by mixing polymers other than the polymer of the present invention with it. In addition, if necessary, the composition may be made by adding additives such as radical scavengers, peroxide decomposers, antioxidants, ultraviolet absorbers, heat stabilizers, plasticizers, flame retardants, and antistatic agents.

[0053] The polymer composition according to the present invention can contain intermediate water due to its hydration structure, mainly at the 2-pyrrolidone site, and exhibits good biocompatibility. Furthermore, the hydrophilization temperature at which water solubility / water insolubility transitions can be arbitrarily set. By utilizing these characteristics, it can be suitably used in various applications where the expression of biocompatibility is desired, such as those listed below.

[0054] (1) Various uses that utilize dissolution / precipitation at a predetermined temperature by having LCST The polymer composition according to the present invention can exist stably in the temperature range of 0 to 100°C in which the aqueous phase is present, and it is possible to impart an arbitrary hydrophilicity temperature (LCST). Furthermore, since it exhibits biocompatibility in both the state where it is dissolved in the aqueous phase in the water-soluble temperature range and the state where it exists separately from the aqueous phase in the water-insoluble temperature range, it is expected that when the polymer composition according to the present invention is present in an aqueous solution containing bio-related substances such as proteins, cells, and tissues, it will have little adverse effect on the bio-related substances and will produce effects such as suppression of protein denaturation.

[0055] By utilizing the above characteristics, for example, by coexisting a polymer composition according to the present invention, prepared to have a predetermined hydrophilic temperature, in a water-soluble or water-insoluble state, with a bio-related substance such as a protein in an aqueous solution, the temperature change occurring in the aqueous solution can be visually detected as the dissolution / precipitation of the polymer composition, and can be used as a means to capture temperature changes.

[0056] Furthermore, by utilizing the fact that the hydrophilic temperature (LCST) exhibited by the polymer composition according to the present invention changes depending on the acidity (pH) of the equilibrium aqueous phase and the presence of various ions, a sensor can be constructed that allows for the visual observation of changes in the solute in the aqueous phase in a state of coexistence with various bio-related substances.

[0057] (2) Use as a material for DDS The polymer composition according to the present invention can be preferably used as a material constituting a drug carrier in a so-called drug delivery system (DDS), which is used particularly for the purpose of supplying drugs to a predetermined site in a living organism. Conventionally, a technique has been developed to release drugs by using a temperature-responsive polymer such as PNIPAM (poly-(N-isopropylacrylamide)), which has an LCST near body temperature, as part of a DDS carrier, and by utilizing the fact that the polymer becomes water-insoluble near the affected area heated to a predetermined temperature (for example, Non-Patent Document 2).

[0058] In the polymer composition according to the present invention, it is possible to construct a polymer composition having a hydrophilization temperature (LCST) of an appropriate temperature near body temperature by copolymerizing, in an appropriate proportion, a structural unit in which the m value in the structural unit shown in Chemical Formula 1 above is set to m=2 or m3, and the LCST of the homopolymer is above body temperature, with a structural unit in which the m value is set to m=1 or m≧4, and the LCST of the homopolymer is above body temperature, and this can be used as a DDS carrier by a method similar to that of PNIPAM, etc.

[0059] In other words, for example, a temperature-responsive liposome can be constructed by grafting the polymer composition according to the present invention onto the liposome membrane, which mainly constitutes a drug carrier that encapsulates a drug. In a drug carrier using such a temperature-responsive liposome, heating it to a temperature above the LCST (Low Temperature Stress Point) introduces mechanical strain into the liposome structure due to morphological changes associated with the hydrophobicization of the molecules constituting the polymer composition according to the present invention, thereby destabilizing it and releasing the encapsulated drug.

[0060] Furthermore, a polymer micelle-type drug carrier can be constructed using the polymer composition according to the present invention. Specifically, by including the polymer composition according to the present invention and forming a block copolymer or graft copolymer having a heterogeneous structure such as hydrophilicity / hydrophobicity at a predetermined temperature, a polymer micelle made of the copolymer can be generated, and a polymer micelle-type drug carrier can be constructed in which a drug is physically or chemically supported inside. In this polymer micelle-type drug carrier, temperature changes across the LCST cause the polymer composition according to the present invention to become hydrophilic / hydrophobic, destabilizing the micelle structure and allowing the encapsulated drug to be released.

[0061] Furthermore, by utilizing the fact that the polymer composition according to the present invention forms a coacervate structure when precipitated from the aqueous phase, a drug carrier can be constructed in which a drug is supported on the coacervate. In a drug carrier of this structure, the polymer composition according to the present invention can dissolve in the aqueous phase by cooling to below the LCST, and the encapsulated drug can be released.

[0062] The drug carrier containing the polymer composition according to the present invention can be administered in any suitable form, for example, by oral administration, transdermal administration, intravenous or intramuscular administration, and rectal administration, depending on the dosage form such as oral preparations, patches, injections, intravenous infusions, or suppositories. In the polymer composition according to the present invention, since it is composed of copolymers between constituent units that are similar to each other except for their m values, the correlation between the polymerization composition and the hydrophilization temperature (LCST) is more linear, and it is possible to suppress the occurrence of secondary properties.

[0063] (3) Use as a protein and cell separator / concentrator and protective agent The polymer composition according to the present invention has been observed to have a higher affinity for proteins compared to conventionally known MPC polymers (see Patent Document 1, etc.) that possess biocompatibility. For example, on the surface of the polymer composition according to the present invention, which has been made water-insoluble under the usage environment, proteins in an aqueous solution are adsorbed at high density, and the proteins can be separated from the aqueous solution. Alternatively, by dissolving the polymer composition according to the present invention in an aqueous solution containing dissolved proteins, and then heating the polymer composition above its hydrophilic temperature to precipitate the water-insoluble polymer composition, the proteins can be concentrated and incorporated into the precipitated polymer composition, thereby separating the proteins from the aqueous solution.

[0064] By utilizing the above-described characteristics, the polymer composition according to the present invention can be effectively used for the separation and recovery of proteins from aqueous solutions, and at the same time, the biocompatibility exhibited by the polymer composition according to the present invention can prevent denaturation of adsorbed proteins. Similarly, by dispersing bio-related substances such as cells and tissues in an aqueous solution in which the polymer composition according to the present invention is dissolved, and then heating the solution above the hydrophilic temperature of the polymer composition to make it water-insoluble, the polymer composition according to the present invention can be precipitated on the surface of the cells, etc., and used for purposes such as maintaining cell activity. Cell culture can then be performed with the polymer composition according to the present invention attached to the surface of the cells, etc.

[0065] When the polymer composition according to the present invention is dissolved in an aqueous phase at a temperature below its hydrophilicity temperature (LCST), and then heated to a temperature above the hydrophilicity temperature to make it water-insoluble, it is observed that the hydrated polymer composition precipitates from the aqueous phase as a colloidal fluid phase. This is a liquid / liquid phase separation in which the polymer hydrate as a whole becomes hydrophobic and separates from the aqueous phase, and is considered to be the formation of a so-called coacervate.

[0066] By utilizing this characteristic, it is possible to separate / concentrate various bio-related substances and other substances to be separated, which are dissolved in the aqueous phase, into the coacervate phase based on the selectivity ratio they exhibit with respect to the aqueous phase / coacervate phase. For example, as described above, by utilizing the affinity that proteins and the like exhibit to the polymer composition according to the present invention, it is possible to incorporate them into the formed coacervate phase with a high selectivity ratio and separate and recover them from the aqueous phase.

[0067] Examples of substances to be separated include proteins, cells, nucleic acids such as RNA and DNA, hydrogen-bonding substances such as antisense nucleic acids, siRNA, miRNA, ribozymes, RNA aptamers and other nucleic acid derivatives, anticancer drugs such as paclitaxel, hydrophobic substances such as carbon nanotubes, anionic substances, cationic substances, and the like.

[0068] The separation / concentration method described above, which utilizes the coacervate formation of the polymer composition according to the present invention, can be carried out in a relatively low temperature range, for example, 5 to 36°C, and can be suitably used for the separation / concentration of bioproducts such as microorganisms and biological materials from cell cultures, as well as proteins such as enzymes, antibodies, and physiologically active substances.

[0069] (4) Use as a coating material to form a biocompatible surface When a surface formed by a polymer composition or the like, which is expected to exhibit biocompatibility, comes into contact with living organisms, biological tissues, cells, etc., it is common practice to use a temperature around 37°C, which is the temperature of living organisms. In the polymer composition according to the present invention, by setting the hydrophilization temperature to below the usage temperature, particularly below 0°C, it is possible to form a water-insoluble state by using a homopolymer obtained by setting the m value of the constituent units shown in Formula 1 above within a predetermined range, or a copolymer using constituent units having different m values ​​in a predetermined ratio, and it can be used as a coating composition that exhibits biocompatibility, in the same way as conventional biocompatible polymers.

[0070] The formation of a water-insoluble film used at the above-mentioned predetermined temperature can be achieved by using a polymer composition containing homopolymers or copolymers whose hydrophilization temperature is set below the operating temperature by setting the m value within a predetermined range, for example, by dissolving the polymer in a predetermined organic solvent to form a solution, and then applying it to the substrate surface by a coating method, spraying method, dipping method, etc., such that the film thickness of the formed film is, for example, about 0.1 μm to 1 mm, and then drying and removing the organic solvent.

[0071] Furthermore, in the polymer composition according to the present invention, when an aqueous solution in which the polymer composition according to the present invention is dissolved is heated above the hydrophilic temperature (LCST), a colloidal fluid phase (coacervate phase) composed of the hydrated polymer composition precipitates from the aqueous phase. This coacervate phase can be used as a coating agent and applied to the substrate surface, and then a film can be formed by drying and removing the hydration water. According to this method, for example, good coating can be performed on substrates with low resistance to organic solvents and aqueous phases.

[0072] Furthermore, in order to more firmly fix the polymer composition according to the present invention to the material to be coated, heat can be applied after coating to further enhance the adhesion with the polymer composition according to the present invention. Alternatively, the composition may be fixed by crosslinking the surface. As a method of crosslinking, a crosslinkable monomer may be introduced as a comonomer component. Alternatively, crosslinking may be performed by electron beam, gamma ray, or light irradiation. Examples of crosslinkable monomers include compounds having multiple vinyl or allyl groups in one molecule, such as methylenebisacrylamide, trimethylolpropane diacrylate, triallyl isocyanate, trimethylolpropane triacrylate, and tetramethylolmethane tetraacrylate, as well as polyethylene glycol diacrylate.

[0073] Among the polymer compositions according to the present invention, a polymer composition having a hydrophilization temperature of 37°C or lower, particularly 0°C or lower, and not having a temperature range in which it exhibits water solubility, exhibits excellent biocompatibility, as well as antifouling, antibacterial adhesion, and anti-inflammatory effects, when it contains a predetermined amount of water. When used as a raw material or surface coating agent in the manufacture of artificial organs, medical devices, etc., which require biocompatibility, it is possible to provide artificial organs or medical devices that have the above effects.

[0074] Examples of the above-mentioned artificial organs and medical devices include, but are not limited to, those having parts that come into contact with biological substances such as blood. Specifically, these include, but are not limited to, blood filters, artificial lungs, dialysis machines, blood storage bags, platelet storage bags, blood circuits, artificial hearts, indwelling needles, catheters, guidewires, stents, artificial blood vessels, and endoscopes.

[0075] The material and shape of the base material constituting the above-mentioned artificial organs and medical devices are not particularly limited. For example, examples of materials include natural polymers such as brocade and hemp, nylon, polyester, polyacrylonitrile, polyolefin, halogenated polyolefin, polyurethane, polyamide, polycarbonate, polysulfone, polyethersulfone, poly(meth)acrylate, ethylene-vinyl alcohol copolymer, butadiene-acrylonitrile copolymer, and mixtures thereof. Metals, ceramics, and composite materials thereof are also examples. Artificial organs and medical devices may be composed of multiple types of base materials. For example, the polymer composition according to the present invention can be applied to the surface of a substrate having a shape such as a porous body, fiber, nonwoven fabric, particle, film, sheet, tube, hollow fiber, or powder.

[0076] When imparting antibacterial properties and / or anti-inflammatory properties to artificial organs, medical devices, etc., it is preferable to apply the polymer composition according to the present invention to at least a portion, preferably substantially the entire surface, of the surface that comes into contact with in vivo tissues or blood.

[0077] The polymer composition according to the present invention can be used as a material forming the whole of artificial organs and medical devices used in contact with biological tissues and blood, or as a material forming the surface thereof. It is desirable that at least a portion, preferably almost the entire surface that comes into contact with blood, of medical devices such as implantable artificial organs and therapeutic devices, extracorporeal circulation artificial organs, surgical sutures, and catheters (cardiovascular catheters such as angiography catheters, guidewires, and PTCA catheters, digestive catheters such as gastric tubes, gastrointestinal catheters, and esophageal tubes, and urological catheters such as urethral catheters and ureteral catheters) is composed of the polymer composition according to the present invention. Furthermore, the polymer composition according to the present invention can also be used as a hemostatic agent, an adhesive for biological tissues, a repair material for tissue regeneration, a carrier for drug sustained-release systems, hybrid artificial organs such as artificial pancreases and artificial livers, artificial blood vessels, embolizing materials, and matrix materials for scaffolds in cell engineering.

[0078] These artificial organs and medical devices may be further given surface lubrication to facilitate insertion into blood vessels and tissues and to prevent tissue damage. A superior method for providing surface lubrication is to insolubilize a water-soluble polymer to form a water-absorbing gel layer on the material surface. This method provides a material surface that combines biocompatibility and surface lubrication.

[0079] Specifically, the polymer composition according to the present invention may be coated on at least a portion of the surface of the substrate constituting the blood filter. Alternatively, the polymer composition according to the present invention may be coated on at least a portion of the surface of the blood bag and the tube communicating with the blood that comes into contact with the blood. Furthermore, at least a portion of the surface of the extracorporeal circulation blood circuit that comes into contact with the blood, consisting of an instrument-side blood circuit section comprising tubes, arterial filters, centrifugal pumps, hemoconcentrators, cardioplegia, etc., and a surgical field-side blood circuit section comprising tubes, catheters, suction devices, etc., may be coated with the polymer composition according to the present invention.

[0080] When using the polymer composition according to the present invention in an indwelling needle assembly, at least a portion of the surface of the indwelling needle assembly that comes into contact with blood may be coated with the polymer composition according to the present invention. The assembly comprises an inner needle having a sharp needle tip, an inner needle hub installed on the proximal end side of the inner needle, a hollow outer needle into which the inner needle can be inserted, an outer needle hub installed on the proximal end side of the outer needle, a protector attached to the inner needle and movable in the axial direction of the inner needle, and a connecting means for connecting the outer needle hub and the protector. Furthermore, at least a portion of the surface of a catheter, which consists of a long tube and an adapter connected to its proximal end (proximal side), that comes into contact with blood may be coated with the polymer composition according to the present invention.

[0081] At least a portion of the surface of the guidewire that comes into contact with the blood may be coated with the polymer composition according to the present invention. Furthermore, at least a portion of the surface of various stents that come into contact with the blood may be coated with the polymer composition according to the present invention, such as hollow tubular bodies made of metal or polymer materials with pores on their sides, or stents formed by weaving metal wires or polymer fibers into a cylindrical shape.

[0082] When using the polymer composition according to the present invention in a cardiopulmonary bypass machine, it may be an artificial lung of the type in which a large number of porous hollow fiber membranes for gas exchange are housed in a housing, blood flows on the outer side of the hollow fiber membranes, and oxygen-containing gas flows inside the hollow fiber membranes, and the polymer composition according to the present invention is coated on the outer surface or outer layer of the hollow fiber membranes. A dialysis apparatus comprising a dialysis circuit including at least one dialysis fluid container filled with dialysis fluid and at least one drainage container for collecting dialysis fluid, and a fluid delivery means for delivering dialysis fluid starting from the dialysis fluid container or ending at the drainage container, wherein at least a portion of the surface in contact with blood may be coated with the polymer composition according to the present invention.

[0083] (5) Use as a protein adsorption membrane and cell culture substrate Conventionally, in many polymers known to exhibit biocompatibility, the mechanism by which this biocompatibility is expressed is observed to be the use of so-called antifouling properties, which prevent proteins contained in blood, etc., from adsorbing onto the polymer surface. As a result of the inability to adsorb proteins, the denaturation of proteins on the polymer surface is prevented, and various phenomena caused by denatured proteins are prevented, thus resulting in the expression of biocompatibility.

[0084] On the other hand, it has been observed that the surface of the polymer composition according to the present invention exhibits a greater adsorption amount of proteins dissolved in an aqueous solution compared to conventionally known biocompatible substances, and that the denaturation of the adsorbed proteins is suppressed. Furthermore, by changing the X value of the constituent units of the polymer composition according to the present invention, it is possible to adjust the adsorption characteristics of proteins and bio-related substances. In particular, by using constituent units with a large X value, it is possible to increase the amount of protein adsorbed, and proteins can be adsorbed efficiently.

[0085] Since the inner walls of blood vessels that come into contact with blood and the extracellular matrix that comes into contact with cells contain a large proportion of protein, it is expected that the polymer composition according to the present invention will form a surface on which proteins are adsorbed without being denatured, thereby exhibiting superior biocompatibility. Based on the characteristics described above, the polymer composition according to the present invention can be used as a filter or the like to recover and separate various proteins in an undenatured state by bringing the surface of the polymer composition into contact with an aqueous solution containing dissolved proteins such as blood or lymph.

[0086] Furthermore, by utilizing the fact that proteins present dissolved in blood, lymph, etc., are adsorbed in an undenatured state onto the surface of the polymer composition according to the present invention, it is possible to perform cell culture under conditions similar to those in vivo by using the polymer composition according to the present invention as a support during cell culture. The surface of the polymer composition according to the present invention can be widely used for culturing adherent cells that require adhesion to a substrate, regardless of the type of cell. In other words, the cell culture support using the polymer composition according to the present invention is not particularly limited as long as it contains cells that adhere to a substrate and live, including epidermal cells, gastrointestinal epithelial cells such as vascular endothelial cells, oral endothelial cells, esophageal epithelial cells, gastric epithelial cells, intestinal epithelial cells, respiratory epithelial cells such as nasal mucosal epithelial cells, tracheal epithelial cells, alveolar epithelial cells, exocrine gland cells such as sweat gland cells, sebaceous gland cells, apocrine gland cells, mammary gland cells, salivary gland epithelial cells, lacrimal gland cells, pancreatic islet cells, adrenal medullary cells, adrenal cortical cells, pineal gland cells, etc. It can be applied to cell cultures of endocrine gland cells such as pituitary cells and thyroid cells, visceral parenchymal cells such as hepatocytes, renal epithelial cells, pancreatic cells, and adrenal cells, sensory organ cells such as taste bud cells, olfactory epithelial cells, and hair cells, nerve cells, glial cells such as astrocytes and Schwann cells, muscle cells such as cardiomyocytes, skeletal muscle cells, and smooth muscle cells, mesenchymal cells such as fibroblasts, stromal cells, connective tissue cells, chondrocytes, and osteoblasts, thymic epithelial cells, uterine epithelial cells, ovarian follicular cells, fallopian tube epithelial cells, seminiferous tubule epithelial cells, and Leydig cells.

[0087] Furthermore, the cell culture support using the polymer composition according to the present invention can be used for culturing various stem cells, including pluripotent stem cells such as embryonic stem cells (ES cells), induced pluripotent stem cells (iPS cells), embryonic tumor cells (EC cells), embryonic germ stem cells (EG cells), nuclear transfer ES cells, somatic cell-derived ES cells, hematopoietic stem cells, bone marrow-derived mesenchymal stem cells, adipose tissue-derived mesenchymal stem cells, other stromal-derived stem cells, Muse cells, neural stem cells, and other tissue stem cells, as well as progenitor cells in various tissues such as liver, pancreas, adipose tissue, bone tissue, and cartilage tissue.

[0088] The stem cells described above may be subjected to treatments such as the introduction of foreign genes or gene editing on the chromosomes, or to treatments that promote proliferation under culture conditions by viral infection (such as transformation). Furthermore, in the culture of stem cells using a cell culture support containing the polymer composition according to the present invention, the degree of differentiation from stem cells to various cells changes depending on the amount of intermediate water contained in the substrate. Therefore, by selecting a substrate according to the purpose of culturing various stem cells, it is possible to promote or suppress differentiation while performing cell culture in accordance with the purpose of the culture.

[0089] Furthermore, the cell culture support using the polymer composition according to the present invention can be used to culture organ parenchymal cells that exhibit characteristic functions of organs, including but not limited to internal organs such as the liver, kidneys, pancreas, and adrenal glands, exocrine glands such as sweat glands, sebaceous glands, apocrine glands, and mammary glands, and endocrine glands such as the pineal gland, pituitary gland, and thyroid gland (metabolism, decomposition, or catabolism of waste products in the blood, alcohol, and other drugs, exocrine secretion of digestive fluids, etc., and endocrine secretion of hormones such as insulin and adrenaline into the bloodstream). The organ parenchymal cells may be somatic cells (primary cells) excised or recovered from a subject, or cultured cells (cell lines) capable of finite or unlimited proliferation under culture conditions, provided that they exhibit the characteristic functions of their respective organs. Provided that the safety of the subject is ensured, the organ parenchymal cells of the present invention may be tumor-derived cells, or they may be subjected to treatments such as the introduction of foreign genes or gene editing on chromosomes, or they may be subjected to growth promotion treatments under culture conditions by viral infection (such as transformation).

[0090] Furthermore, in cell culture using the polymer composition according to the present invention, in addition to maintaining and proliferating the cells to be cultured, it can also be used, for example, in a production bioreactor that produces useful proteins such as albumin and blood coagulation factors by culturing cells, or as part of an artificial organ that cultures and organizes predetermined cells and expresses at least one function that an organ in the human body performs, for purposes such as assisting the function of organs in patients with organ diseases. Furthermore, cells cultured using the polymer composition according to the present invention can be used to investigate the effects of various drugs on cells, or for verification of the cells themselves.

[0091] Furthermore, the polymer composition according to the present invention can be used as a substrate for immobilizing enzymes or antibodies, and can be used in the preparation of immobilized enzymes and immobilized antibodies used in qualitative or quantitative analysis of test substances (such as proteins) using immunoassays, protein purification, bioreactor construction, and the like. When immobilizing enzymes or antibodies onto the polymer composition according to the present invention, the process is carried out by coexisting the polymer composition with an aqueous phase containing the enzymes or antibodies in a temperature range in which the polymer composition exhibits water insolubility. Alternatively, an antigen that produces an antigen-antibody reaction with the target antibody can be immobilized onto the polymer composition according to the present invention, and the target antibody can then be immobilized using the antigen-antibody reaction.

[0092] When using the polymer composition according to the present invention as a coating material to form a biocompatible surface as described above, or as a substrate for protein adsorption or cell culture, setting the hydrophilization temperature (LCST) of the polymer composition according to the present invention to, for example, 0°C to 35°C, between the solidification temperature of water and the usage temperature for various purposes, makes it possible to use water as a solvent when applying the polymer composition according to the present invention to a substrate surface for coating, thereby avoiding the drawbacks that occur when using organic solvents as a solvent.

[0093] When using a polymer composition with a hydrophilization temperature (LCST) set between approximately 0°C and 35°C to coat a substrate surface with water as the solvent, an aqueous solution at a temperature below the hydrophilization temperature is prepared, applied to the substrate surface, dried, and heated as necessary to form a film with the polymer composition according to the present invention.

[0094] Furthermore, as described above, by using a cell culture support whose surface is formed using a polymer composition with a hydrophilic temperature (LCST) set between approximately 0°C and 35°C, it becomes possible to easily detach cells, cell sheets, or pseudo-tissues obtained by culture from the support, thereby reducing the burden on the cells. In other words, after performing cell culture or the like on a surface coated with the polymer composition according to the present invention at around 37°C, the polymer composition is dissolved by cooling the culture system to a temperature below the hydrophilicity temperature (LCST) of the polymer composition, making it possible to easily separate the cultured cells from the support. Utilizing the above features, a cell culture support using the polymer composition according to the present invention can be used as a scaffold for regenerative medicine using various stem cells, such as by culturing cardiomyocytes to produce a myocardial sheet.

[0095] The polymer composition according to the present invention can be produced by polymerizing the monomer using the structure shown in structural formula 3, particularly using only a single type of monomer, or by mixing multiple types of monomers with different carbon chain lengths, etc., that bond R2 and 2-pyrrolidone, using an appropriate method. R1, R2, and m in structural formula 3 are the same as R1, R2, and m in structural formula 2. Furthermore, for example, depending on the purpose of imparting various properties to the polymerized polymer composition, other (meth)acrylic acids, etc., can be mixed and used as monomers during the polymerization process, within a range that does not hinder the effects of the present invention.

[0096] [ka]

[0097] Polymerization of the above monomers can be carried out by general methods such as random polymerization, ionic polymerization, photopolymerization, and polymerization using macromers, by adding an appropriate initiator to a solution obtained by dissolving at least one monomer in a predetermined solvent. As initiators for this polymerization, for example, peroxide-based radical initiators (benzoyl peroxide, ammonium persulfate, etc.), azo-based radical initiators (2,2'-azobisisobutyronitrile (AIBN), 2,2'-azobis-dimethylvaleronitrile (ADVN), etc.), 2,2'-azobiscyanovaleric acid (ACVA), 2,2'-azobis[2-(2-imidazolin-2-yl)propane] dihydrochloride (VA-044), water-soluble or oil-soluble redox-based radical initiators (consisting of dimethylaniline and benzoyl peroxide), etc. can be used.

[0098] The amount of polymerization initiator used can be appropriately determined within the range in which the desired polymer can be obtained. For example, it is usually preferably 0.01 to 10 parts by mass, and more preferably 0.01 to 5 parts by mass, per 100 parts by mass of the above monomer. The polymerization temperature and polymerization time can be appropriately selected and determined depending on the type of polymerization initiator and the presence and type of other monomers. For example, when AIBN is used as the polymerization initiator, the polymerization temperature is 40 to 90°C, preferably 50 to 80°C, and more preferably 60 to 70°C. The polymerization time can be 1 to 48 hours, preferably 1 to 24 hours, and more preferably 2 to 24 hours.

[0099] The pressure at which the polymerization reaction is carried out is not particularly limited, but atmospheric pressure is preferred. Polymerization reactions using multiple types of monomers may be carried out by random copolymerization by mixing each monomer, or by block copolymerization by polymerizing each monomer to a certain extent and then mixing them together. The solvent used in the above polymerization reaction is not particularly limited as long as it can dissolve the monomers subjected to polymerization; general solvents can be used. For example, polar aprotic solvents such as acetone, dioxane, dimethylformamide (DMF), dimethyl sulfoxide (DMSO), and tetrahydrofuran (THF), and polar protic solvents such as propanol, ethanol, methanol, and water can be appropriately selected and used.

[0100] The molecular weight of the polymer composition that mainly constitutes the osmotic pressure regulator according to the present invention is not particularly limited and can be appropriately determined according to its application, etc. Generally, the number average molecular weight (Mn) can be about 1,000 to 100,000, and it is particularly preferable to have a molecular weight of about 10,000 to 70,000. The embodiments of the present invention described below are for illustrative purposes only and do not limit the technical scope of the invention. The technical scope of the invention is defined solely by the claims. [Examples]

[0101] (Monomer synthesis example 1) Synthesis of N-(acryloyloxy-n-methyl)-2-pyrrolidone (NAMeP) N-(acryloyloxy-n-methyl)-2-pyrrolidone (NAMeP) was synthesized as a monomer for obtaining the polymer composition according to the present invention via N-hydroxymethyl-2-pyrrolidone (NHMeP) by the following method. NAMeP corresponds to the structure in structural formula 3 where R1 is a hydrogen atom, R2 is an oxygen atom, and the m value is 1. 2-pyrrolidone (106.2 g, 1.25 mol, Kanto Chemical Co., Ltd.) was mixed with potassium hydroxide (0.3 g, 5.34 mmol, FUJIFILM Wako Co., Ltd.) and heated to 80°C. Paraformaldehyde (37.8 g, 1.25 mol (formaldehyde equivalent), FUJIFILM Wako Co., Ltd.) was added, and the mixture was stirred for 30 minutes to induce the reaction. The reaction solution was cooled to room temperature, and 200 mL of toluene was added to precipitate the target product. The obtained precipitate was collected and recrystallized three times from toluene to obtain NHMeP as a white solid (77.2 g, 53.6%). Table 1 shows the results of nuclear magnetic resonance (NMR) spectroscopy, mass spectrometry (HRMS), and elemental analysis (EA) of the obtained white solid.

[0102] [Table 1]

[0103] The NHMeP (54.0 mmol) obtained above and triethylamine (1.1 equivalents, Tokyo Chemical Industry Co., Ltd.) were dissolved in 200 mL of dichloromethane and the mixture was placed in an ice bath. Acryloyl chloride (1.0 equivalent, Tokyo Chemical Industry Co., Ltd.) dissolved in 90 mL of dichloromethane was added dropwise, and the mixture was stirred at room temperature for 12 hours after the addition was complete. The reaction mixture was washed three times with 100 mL of 1 M hydrochloric acid, three times with 100 mL of 5% potassium carbonate aqueous solution, and once with 100 mL of saturated brine. The organic layer was collected and dried over anhydrous magnesium sulfate. The solvent was removed under reduced pressure to obtain the crude product. A small amount of BMH (Seiko Chemicals, Ltd.) as a high-temperature polymerization inhibitor was added, and pure NAMeP (boiling point: 76-78°C, 0.5 mmHg, 5.64 g, 61.4%) was obtained as a colorless, transparent liquid by distillation. Table 2 shows the results of nuclear magnetic resonance (NMR) spectroscopy, mass spectrometry (HRMS), and elemental analysis (EA) for the obtained NAMEP.

[0104] [Table 2]

[0105] (Monomer synthesis example 2) Synthesis of N-(acryloyloxy-n-ethyl)-2-pyrrolidone (NAEtP) NAEtP was synthesized in the same manner as in Synthesis Example 1 above, except that commercially available NHEtP (manufactured by Tokyo Chemical Industry Co., Ltd.) was used as an intermediate in the synthesis of the monomer NAEtP (54.0 mmol). In structural formula 3, NAEtP has a structure in which R1 is a hydrogen atom, R2 is an oxygen atom, and the m value is 2. Table 3 shows the results of nuclear magnetic resonance (NMR) spectroscopy, mass spectrometry (HRMS), and elemental analysis (EA) of the obtained NAEtP (boiling point: 83-85°C, 0.5 mmHg, 6.23 g, 63.0%).

[0106] [Table 3]

[0107] (Monomer synthesis example 3) Synthesis of N-(acryloyloxy-n-propyl)-2-pyrrolidone (NAPrP) NAPrP was synthesized in the same manner as in Synthesis Example 1 above, except that a commercially available product (manufactured by Tokyo Chemical Industry Co., Ltd.) was used as an intermediate for the synthesis of the monomer NAPrP (54.0 mmol). In structural formula 3, NAPrP has a structure in which R1 is a hydrogen atom, R2 is an oxygen atom, and the X value is 3. Table 4 shows the results of nuclear magnetic resonance (NMR) spectroscopy, mass spectrometry (HRMS), and elemental analysis (EA) of the obtained NAPrP (boiling point: 94-96°C, 0.5 mmHg, 6.54 g, 61.4%).

[0108] [Table 4]

[0109] (Monomer synthesis examples 4-6) Synthesis of N-(acryloyloxy-n-butyl)-2-pyrrolidone (NABuP), N-(acryloyloxy-n-pentyl)-2-pyrrolidone (NAPnP), and N-(acryloyloxy-n-hexyl)-2-pyrrolidone (NAHxP) N-(acryloyloxy-n-alkyl)-2-pyrrolidone (NARP) (R=Bu,Pe,Hx) was synthesized as a monomer for obtaining the polymer composition according to the present invention via N-hydroxyalkyl-2-pyrrolidone (NHRP) (R=Bu,Pe,Hx) by the following method.

[0110] NABuP has a structure in structural formula 2 where R1 is a hydrogen atom, R2 is an oxygen atom, and the m value is 4. NAPeP has a structure in structural formula 2 where R1 is a hydrogen atom, R2 is an oxygen atom, and the m value is 5. NAHxP has a structure in structural formula 3 where R1 is a hydrogen atom, R2 is an oxygen atom, and the m value is 6. 0.84 mol each of 4-aminobutanol (Bu), 4-aminopentinol (Pn), and 4-aminohexanol (Hx) (manufactured by Tokyo Chemical Industry Co., Ltd.) was mixed with γ-butyrolactone (72.4 g, 0.84 mol, Tokyo Chemical Industry Co., Ltd.), stirred at room temperature for 1 hour, and then reacted in an autoclave at 250°C for 8 hours. The resulting mixture was distilled to obtain NHBuP, NHPnP, and NHHxP, respectively, as colorless, transparent liquids. Tables 5-7 show the results of nuclear magnetic resonance (NMR) spectroscopy, mass spectrometry (HRMS), and elemental analysis (EA) for each liquid obtained above.

[0111] [Table 5]

[0112] [Table 6]

[0113] [Table 7]

[0114] The monomers NABuP, NAPnP, and NAHxP were synthesized in the same manner as in Synthesis Example 1 above, except that NHBuP, NHPnP, and NHHxP (54.0 mmol each) obtained above were used. Tables 8-10 show the results of nuclear magnetic resonance (NMR) spectroscopy, mass spectrometry (HRMS), and elemental analysis (EA) for each monomer obtained above.

[0115] [Table 8]

[0116] [Table 9]

[0117] [Table 10]

[0118] (Synthesis of homopolymers) Using the NARP (R=Me,Et,Pr,Bu,Pn,Hx) synthesized above as monomers, various poly((N-acryloyloxyalkyl)-2-pyrrolidone) (PNARP) were synthesized using the following method. Each NARP (20 mmol) was dissolved in N,N-dimethylformamide (DMF) to a monomer concentration of 2 M. 2,2'-azobisisobutyronitrile (1 / 200 equivalent, Tokyo Chemical Industry Co., Ltd.) was added as a polymerization initiator, and the mixture was bubbled with argon gas for 30 minutes to remove dissolved oxygen. The mixture was then heated to 60°C and reacted for 40 minutes.

[0119] The reaction mixture was poured into a large excess of diethyl ether, and the resulting polymer was precipitated. The polymer was purified by reprecipitation using acetone as a good solvent and diethyl ether as a poor solvent, and finally the solvent was removed under reduced pressure to obtain pure polymer as a white solid (PNAMeP) or a colorless, transparent viscous substance (PNAEtP, PNAPrP, PNABuP, PNAPnP, PNAHxP). The structure of each obtained polymer was determined by NMR spectroscopy. 1 Structural analysis was performed using 1H NMR [400 MHz, CDCl3, TMS], number-average molecular weight (Mn) and molecular weight distribution (PDI, Mw / Mn) were measured by size exclusion chromatography (SEC), and the glass transition temperature (Tg) was evaluated by differential scanning calorimetry (DSC). Tables 11 and 12 show the results of NMR spectroscopy analysis for each polymer obtained above.

[0120] [Table 11]

[0121] [Table 12]

[0122] Table 13 shows the number-average molecular weight (Mn), molecular weight distribution (PDI, Mw / Mn), and glass transition temperature (Tg) of the polymers synthesized above. As shown in Table 13, various homopolymers composed of structural units with different m values ​​in structural formula 2 were synthesized with similar degrees of polymerization using the above synthesis method.

[0123] [Table 13]

[0124] (Synthesis of copolymers) Using the NARP (R=Me,Et,Pr,Bu,Pn,Hx) synthesized above as monomers, various PNARP copolymers containing NARP with different m values ​​as constituent units were synthesized using the following method. Two types of NAR1P and NAR2P used for polymerization were dissolved in DMF in a predetermined ratio (total amount; 6 mmol) to a monomer concentration of 2 M. 2,2'-azobisisobutyronitrile (1 / 200 equivalent, Tokyo Chemical Industry Co., Ltd.) was added as a polymerization initiator, and argon gas was bubbled through for 30 minutes to remove dissolved oxygen. The mixture was then heated to 60°C and held for 40 minutes to induce the polymerization reaction. The reaction solution was poured into a large excess of diethyl ether to precipitate the resulting polymer. The polymer was then purified by reprecipitation using acetone as a good solvent and diethyl ether as a poor solvent, and finally the solvent was removed under reduced pressure to obtain the polymer as a colorless, transparent, viscous substance.

[0125] The structure of each polymer obtained was determined by NMR spectroscopy. 1 Structural analysis was performed using 1H NMR [400 MHz, CDCl3, TMS], number-average molecular weight (Mn) and molecular weight distribution (PDI, Mw / Mn) were measured by size exclusion chromatography (SEC), and the glass transition temperature (Tg) was evaluated by differential scanning calorimetry (DSC). Tables 14-17 show the results of NMR spectroscopy analysis for each polymer obtained above.

[0126] [Table 14]

[0127] [Table 15]

[0128] [Table 16]

[0129] [Table 17]

[0130] Table 18 shows the composition (measured values ​​and initial composition), number-average molecular weight (Mn), molecular weight distribution (PDI, Mw / Mn), and glass transition temperature (Tg) for each copolymer. In Tables 18 and 20, each PNARP is abbreviated, for example, "PNAMeP" is written as "Me". As shown in Table 18, by mixing monomers with different m values ​​in structural formula 2 in a predetermined ratio and polymerizing them using the above synthesis method, a copolymer can be obtained that reflects the mixing ratio of the monomers and has a similar degree of polymerization.

[0131] [Table 18]

[0132] Homopolymers composed of constituent units with different m values, as listed in Table 13, were added to pure water at a concentration of 1 wt%. The temperature was then varied within the range of 0 to 100°C, and the dissolution behavior of each polymer in the aqueous phase was observed. As a result, PNAEtP, with m=2, dissolved in pure water throughout the 0-100°C range, forming a clear aqueous solution as a single phase, demonstrating its water solubility. On the other hand, PNAPnP and PNAHxP, with m=5 and 6, were observed to be water-insoluble, as the polymer phase did not mix with pure water throughout the 0-100°C range.

[0133] In contrast to the above, for PNAMeP, PNAPrP, and PNABuP, where m=1,3,4, it was observed that at low temperatures, such as around 10°C, the polymers dissolved in pure water, forming a clear aqueous solution as a single phase, thus exhibiting water solubility. On the other hand, when these aqueous solutions were heated to high temperatures, such as around 80°C, the polymers precipitated from the aqueous solution, causing it to become colloidal and cloudy. It was observed that each polymer became water-insoluble, forming a polymer phase different from the aqueous phase and separating. Furthermore, it was observed that this transition between water solubility and water insolubility was reversible.

[0134] Figure 2 compares the state of samples of the polymers related to m=1, 3, and 4, as well as PNIPAM, which is known to be a temperature-responsive polymer, when mixed with pure water to a polymer concentration of 1 wt%, after being cooled to around 10°C ((a) below LCST) and when heated to around 80°C ((b) above LCST). As shown in Figure 2, at around 10°C, all polymers, including PNIPAM, exhibit water solubility and form clear aqueous solutions, while at around 80°C, the polymers precipitated and became colloidal and cloudy.

[0135] Figure 3 shows the change in light (600 nm) transmittance when the temperature of each of the above samples is changed. In Figure 3, the change in light transmittance for each sample is shown, normalized so that the state with the highest transmittance is 100% and the state with the lowest transmittance is 0%. As shown in Figure 3, it was observed that in each sample, the change in light transmittance occurred reversibly at approximately the same temperature during both the heating and cooling processes.

[0136] Figure 4 shows the change in light transmittance of the above-mentioned m=4 polymer (PNABuP) when the temperature is changed between 20 and 50°C, using samples mixed with pure water at various polymer concentrations from 0.125 wt% to 2.0 wt%, normalized in the same way as in Figure 3. As shown in Figure 4, the light transmittance of PNABuP (m=4) changes rapidly between approximately 30 and 45°C. Below 30°C, the polymer dissolves in pure water at concentrations of 2.0 wt% or more, while above 45°C, its solubility drops to about 0.1 wt% or less, indicating that it is not water-soluble.

[0137] The evaluation results shown in Figures 2-4 above indicate that the polymers related to m=1, 3, and 4 are temperature-responsive polymers that, like PNIPAM used as a comparison, have a lower critical solution temperature (LCST), exhibiting non-water solubility at temperatures above the LCST and water solubility at lower temperatures.

[0138] Table 19 shows the LCST (Low-Critical Values) for each homopolymer shown in Table 1 in pure water and PBS(-). In Table 19, for aqueous solutions prepared by dissolving each polymer in pure water or PBS at a polymer concentration of 1 wt%, the change in light transmittance with temperature was measured, as in Figures 3 and 4, and the temperature at which the slope of the change curve was greatest was defined as the LCST. Furthermore, the maximum intermediate water content for each polymer shown in Table 19 indicates the weight percentage of water molecules contained in the form of intermediate water when each polymer is saturated with water under a pure water environment. The intermediate water content was determined by dividing the enthalpy change caused by the ordering of water molecules, particularly in the sub-zero temperature range of -60 to -20°C, during the process of changing the temperature of each sample in a DSC in the range of -100 to 30°C by the latent heat of solidification of water (see Non-Patent Document 1, etc.).

[0139] [Table 19]

[0140] As shown in Table 19, the LCST (Low Cell Saturation Temperature) values ​​of homopolymers with m values ​​of 1, 3, and 4 were observed to change in pure water and PBS, suggesting that this is influenced by ions and other factors present around the polymer. Furthermore, PNAEtP, with m=2, showed water solubility in PBS as well as in pure water across the entire temperature range of 0-100°C, and no LCST was observed. On the other hand, PNAPnP and PNAHxP, with m=5 and 6, showed water insolubility in PBS as well as in pure water across the entire temperature range of 0-100°C.

[0141] Furthermore, it was observed that each homopolymer shown in Table 19 can contain an intermediate amount of water in pure water, ranging from approximately 14.1 to 22.1 wt%. In particular, even polymers with large m values ​​that are insoluble in water (m=5,6) showed a similar amount of hydration as polymers with m=2 that are soluble in water. Therefore, it was inferred that in the constituent units shown in structural formula 2, regardless of the m value, the 2-pyrrolidone moiety at the end of the side chain undergoes hydration with a predetermined amount of water molecules.

[0142] Furthermore, when the m value is set to m=1 (methylene group), a decrease in intermediate water content is observed compared to other constituent units. This suggests that when the m value in structural formula 2 is set to m=1 and the carbon chain is shortened, the 2-pyrrolidone portion becomes closer to the main chain, reducing its degrees of freedom and resulting in a change in the hydration state. Consequently, its degree of hydrophilicity decreases, and LCST (Low-Core Stem Cellular Stem) is observed.

[0143] Homopolymers with m values ​​of 1, 3, and 4 exhibit water solubility below their respective LCSTs (Low Cell Temperatures), and when the temperature is raised above the LCST, the polymer generally precipitates in a colloidal state (Figure 2). During this precipitation, PNAMeP (m=1) produces a powdered (solid) polymer, while PNAPrP and PNABuP (m=3 and 4) precipitate through liquid / liquid phase separation of droplets composed of hydrated polymers in the aqueous phase. The change in morphology during precipitation from the aqueous solution is considered to be a result of the degree of interaction between polymer molecules and water molecules changing with the m value.

[0144] Table 20 shows the LCSTs in pure water and PBS for each copolymer shown in Table 2, determined by the method described in Table 19. It was observed that for each polymer shown in Table 20, when a homogeneous aqueous solution was prepared at a temperature below the respective LCST and then heated to precipitate the polymer, precipitation occurred in the form of coacervates for all polymers.

[0145] [Table 20]

[0146] As shown in Table 20, in copolymers between m=1, 3, and 4 constituent units (2-1 and 2-2 in Table 4) in which LCSTs are observed when homopolymerized, intermediate LCSTs were observed between those constituent units and the homopolymers composed of those units. This suggests that in copolymers containing constituent units with different m values ​​within the structure shown in structural formula 2, intermediate LCSTs are expressed in relation to the LCSTs specific to each constituent unit.

[0147] On the other hand, in the copolymer of NABuP, in which LCST is observed in the homopolymer, and NAPnP, which exhibits non-water solubility across the entire temperature range of 0 to 100°C (2-3 in Table 20), LCST was observed at lower temperatures than in the PNABuP homopolymer. This suggests that PNAPnP, which does not exhibit water solubility above 0°C, has a potential water-soluble / non-water-soluble transition temperature (hydrophilization temperature), and that the presence of this hydrophilization temperature below 0°C prevents the observation of overt LCST.

[0148] Furthermore, LCSTs were observed in copolymers of NAEtP, which is water-soluble over the entire temperature range of 0-100°C, and NAHxP, which is water-insoluble over the entire temperature range of 0-100°C (2-4 to 2-13 in Table 20), and it was observed that these LCSTs changed continuously depending on the polymerization ratio. This indicates that even water-soluble PNAetP has a potential hydrophilization temperature, and it is thought that the presence of this hydrophilization temperature above 100°C leads to water solubility over the entire temperature range of 0-100°C.

[0149] As shown in Tables 18 and 20, by using polymers (copolymers) with appropriately set m values ​​as shown in structural formula 2, or copolymers between constituent units having different m values, it is possible to obtain polymer compositions that have a hydrophilization temperature (LCST) at a predetermined temperature in the range of 0 to 100°C and exhibit water insolubility at temperatures above that hydrophilization temperature. Furthermore, by using polymers (copolymers) with a hydrophilization temperature of 0°C or lower, or copolymers between constituent units having different m values, it is possible to obtain polymer compositions that exhibit water insolubility over the entire range of 0 to 100°C.

[0150] Figure 5 shows the change in light (600 nm) transmittance of aqueous solutions prepared by dissolving each copolymer, with varying proportions of NAEtP and NAHxP from the copolymers listed in Table 18, in pure water or PBS(-) at a ratio of 1 wt%, as the temperature of each aqueous solution is changed. In Figure 5, the value of "x" indicates the molar ratio of PNAetP in the copolymer.

[0151] As shown in Figure 5, polymers in which the composition ratio of NAEtP and NAHxP is changed exhibit LCSTs corresponding to the composition. In particular, by combining NAEtP, which has a potential hydrophilization temperature of 100°C or higher in homopolymers, with PNAHxP, which has a potential hydrophilization temperature of 0°C or lower, it is possible to set a desired temperature within the entire range of 0 to 100°C in which the aqueous phase exists as the LCST.

[0152] By utilizing the characteristics described above, it is possible to obtain a polymer composition having a desired hydrophilic temperature by using copolymers of multiple structural units with mutually different m values ​​in structural formula 2 and changing their composition ratio. In particular, since the LCST can be set to any temperature around 0 to 40°C, it can be suitably used in various applications that come into contact with various bio-related substances and utilize the expression of the LCST.

[0153] (Platelet adhesion test) To evaluate the biocompatibility of the homopolymers and copolymers shown in Tables 18 and 20 above, particularly those polymers that exhibit water insolubility around 37°C, the adhesion of platelets to the surface of these polymers was assessed. It is known that platelet adhesion and activation are suppressed on surfaces exhibiting biocompatibility.

[0154] For evaluation, we used experimentally purchased human whole blood collected in the United States (within 5 days of collection). The refrigerated human whole blood was allowed to return to room temperature by leaving it at room temperature for approximately 30 minutes. After three inversions and mixing, it was centrifuged at 400 rcf for 5 minutes using a tabletop centrifuge (2420, KUBOTA). Approximately 500 μL of the supernatant (pale yellow, translucent) was collected and designated as platelet-rich plasma (PRP). Subsequently, it was centrifuged again at 2500 rcf for 10 minutes, and approximately 2 mL of the supernatant (pale yellow, transparent) was collected and designated as platelet-poor plasma (PPP). The platelet concentration in the PRP was calculated by counting platelets in the PRP diluted 800-fold with PBS(-) using a hemocytometer, and the seeding concentration was 3.0 × 10⁶. 7 cells / cm 2 Platelet suspension was prepared by diluting PRP with PPP to achieve the desired result.

[0155] Of the polymers shown in Tables 18 and 20 above, the homopolymers PNABuP, PNAPnP, and PNAHxP (1-4 to 1-6), which exhibit water insolubility at around 37°C, and the copolymer of PNAEtP and PNAHxP (Et:Hx=30:70, 2-12) were prepared. Polymer solutions were prepared by dissolving each polymer in methanol at a ratio of 0.5 wt%, spin-coating them onto a PET (polyethylene terephthalate) film surface, and then drying and removing the solvent. For comparison, samples coated with PMEA (poly(2-methoxyethyl acrylate)), known for its high biocompatibility, an uncoated PET film surface, and a sample coated with PNIPAM, known to exhibit LCST (Low-Coating Steady Saturation), were also used.

[0156] Add 450 μL (approximately 300 μL / cm³) of the platelet suspension prepared above to each sample. 2 Platelets were adhered to the substrate by incubating it at 37°C for 1 hour in the poured state. After that, the platelet suspension was removed from the sample surface, washed twice with PBS, and then the adhered platelets were immobilized on the substrate by immersion in a 1% glutaraldehyde solution (25% glutaraldehyde, polyscience, Inc. 01909 diluted to 1 / 25 with PBS(-)) and incubated at 37°C for 2 hours. After immobilization, the samples were washed by immersion once each in PBS(-) (10 minutes), PBS(-):water = 1:1 (8 minutes), and water (8 minutes, 10 minutes). After washing, the samples were air-dried for 3 hours, and then dried in a container with silica gel for more than 1 day. After drying, the number of adhered platelets on each sample surface was counted by observing the substrate surface with a scanning electron microscope (SEM, KEYENCE, 3D Real Surface View Microscope VE-9800).

[0157] Figure 6 shows the number of platelets adhering to each sample surface described above. In Figure 6, the breakdown of platelets adhering to the sample surface is shown, showing the different morphologies: Type I (normal), Type II (pseudopod formation), and Type III (extended). On PET surfaces that did not exhibit biocompatibility, a large total number of adherent platelets was observed, along with a high proportion of platelets adhering in type II and type III morphologies. On the other hand, the homopolymers according to the present invention (PNABuP, PNAPnP, PNAHxP) showed similar levels of total and breakdown of adherent platelets to PMEA, indicating high biocompatibility. Furthermore, the copolymer of PNAEtP and PNAHxP showed a tendency to exhibit higher biocompatibility compared to the homopolymers mentioned above. From the above results, it was observed that the polymer composition according to the present invention exhibits high biocompatibility on a water-insoluble surface.

[0158] (Hemolysis test) To evaluate the biocompatibility of the homopolymers and copolymers shown in Tables 18 and 20 above, particularly those that are water-soluble at around 37°C, the hemolytic properties that occur when these polymers are mixed with human whole blood were assessed. As described above, human whole blood, returned to 20°C or 37°C, was inverted and mixed three times, and then PBS(-) solutions containing each polymer dissolved in various concentrations up to 10 mg / mL were mixed in a volume ratio of 1:1 (final red blood cell concentration: approximately 1.5 × 10⁻⁶). 9 The cells (Cells / mL) were incubated at 20°C or 37°C for 2 hours, and the absorbance (540 nm) of the supernatant was measured. The hemolysis rate was then calculated from this absorbance.

[0159] For evaluation, homopolymers of PNAMeP~PNABuP (m=1~4, 1-1~1-4) and copolymers of PNAEtP and PNAHxP (Et:Hx=80:20, 70:30, 2-7, 2-8) were used, and PBS(-) solutions containing tween20 and triton X100 were used for comparison. All of the above polymers have a Lowest Temperature Set (LCST) above 20°C and dissolve uniformly in the aqueous phase at 20°C. On the other hand, at 37°C, only PNABuP shows insolubility. Therefore, in the above test, PBS in which PNABuP was dissolved at a temperature below the LCST was heated to 37°C, and the PBS in which PNABuP precipitated in a coacervate form was mixed with human whole blood for evaluation.

[0160] Figure 7 shows the hemolysis rate observed when the above polymer was mixed with human whole blood. As shown in Figure 7, at both 20°C and 37°C, in the concentration range where clear hemolysis was observed with triton X100 used as a comparison, substantially no hemolysis was observed with the above polymer. This indicates that the above polymer exhibits biocompatibility even when dissolved in the aqueous phase.

[0161] (Protein adsorption test) To evaluate the adsorption capacity of the polymers shown in Tables 18 and 20 above, which adsorb proteins in aqueous solutions, the following evaluations were performed. Of the homopolymers and copolymers shown in Tables 2 and 4 above, the homopolymer PNABuP~PNAHxP (m=4~6, 1-4~1-6) and the copolymer of PNAEtP and PNAHxP (Et:Hx=70:30~10:90, 2-8~2-14) were selected as polymers that exhibit water insolubility around 37°C. Polymer solutions (0.5 wt%) of each polymer were dissolved in methanol and spin-coated onto the surface of a PET film, after which the solvent was dried and removed to prepare samples coated with each polymer. For comparison, PET films coated with PMEA and PNIPAM, as well as uncoated PET films, were also used.

[0162] Each of the above samples was immersed in PBS(-) (300 μL) at 37°C for 1 hour to thoroughly hydrate the polymer surface. Then, 1 mL of Dulbecco's modified Eagle medium (DMEM) containing 10% fetal bovine serum (FBS) as a protein solution was added dropwise to the PBS(-), and the mixture was incubated at 37°C for 1 hour. After that, excess protein solution was removed from each polymer surface by gently washing it three times with PBS(-), and the amount of protein adsorbed on each polymer surface was calculated using the Micro BCA method.

[0163] Figure 8 shows the amount of adsorbed protein on each polymer surface calculated above. It was observed that the PNABuP~PNAHxP homopolymer according to the present invention exhibited protein adsorption equivalent to or greater than that of PMEA, and that the amount of adsorption increased as the m-value increased. Furthermore, it was shown that the amount of protein adsorption in the PNAEtP and PNAHxP copolymer changed depending on the mixing ratio, and that there was a mixing ratio in which particularly large amounts of protein adsorption were observed.

[0164] Furthermore, when comparing Figure 8 with Figure 6, for example, no significant difference in the amount of adsorbed protein is observed between PET, which exhibits no biocompatibility and causes vigorous foreign body reactions such as platelet adsorption and activation, and PMEA, which exhibits biocompatibility and suppresses platelet adsorption. On the other hand, considering that the foreign body reactions observed on the PET surface are due to the activation of the coagulation system, complement system, platelet system, etc., caused by the denaturation of proteins adsorbed on the PET surface, it can be inferred that the proteins adsorbed on the PET surface in Figure 8 are denatured, while the proteins adsorbed on the PMEA surface are not denatured.

[0165] Furthermore, while the polymer surface according to the present invention exhibits adsorption of a larger amount of protein compared to PMEA, platelet adhesion is suppressed to the same extent as with PMEA. Therefore, it is inferred that the polymer surface according to the present invention can adsorb and retain a large amount of protein in an undenatured state. [Industrial applicability]

[0166] The polymer composition according to the present invention can be made water-insoluble within a predetermined temperature range, and in particular, the LCST can be set to a predetermined temperature, and exhibits high biocompatibility, making it suitable for use as a material in contact with various bio-related substances.

Claims

1. A polymer composition characterized by comprising a polymer having two or more monomer units represented by structural formula (1), wherein m is mutually different, and having a temperature range of 0 to 100°C in which it is insoluble in water. However, in structural formula (1), R １ R is either a hydrogen atom or a methyl group. ２ represents either O or NH, m is a natural number greater than or equal to 1, and n is the number of repeating monomer units.

2. The polymer composition according to claim 1, characterized in that the polymer contains a structure in which m is 5 or more.

3. The polymer composition according to claim 1 or 2, characterized in that the polymer is water-insoluble throughout the temperature range of 0 to 100°C.

4. The polymer composition according to claim 1 or 2, characterized in that the polymer has a temperature range of 0 to 100°C in which it transitions between being water-soluble and water-insoluble, is water-insoluble above that temperature range, and is water-soluble below that temperature range.

5. The polymer composition according to any one of claims 1 to 4, characterized in that the proportion of monomer units represented by the above structural formula (1) in the polymer is 80 mol% or more.

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