Preparation method of supramolecular hydrogels and their application as biomaterials

Abstract

The present invention provides a polyrotaxane having a chemical structure in which an axial molecule represented by formula (2) passes through a cyclodextrin ring in a compound represented by formula (1). In formula (1), CD denotes a cyclodextrin ring, L2 denotes a divalent organic group, L3 denotes an ethylene group or propylene group, RA denotes a hydrogen atom or a C1-6 alkyl group, and m denotes an integer between 0 and 35. However, if L2 and N are connected by a double bond, RA is not present. In formula (2), R denotes a hydrocarbon group that is larger than the inner diameter of the cyclodextrin ring, X denotes a divalent organic group, and a denotes an integer between 100 and 1000.

Description

[Technical Field]

[0001] The present invention relates to a method for preparing supramolecular hydrogels and their application as biomaterials. [Background technology]

[0002] In the medical field, biological tissues are often injured during surgical procedures such as closing an incision, protecting a wound, and implanting or removing a medical device. Injured biological tissues begin to repair themselves in order to heal. During this repair process, other biological tissues surrounding the repaired area may become enmeshed and continuously bond with the repaired area. This condition is called adhesion. Adhesion can occur between two or more adjacent biological tissues, or between an implanted device and adjacent biological tissue. Anti-adhesion materials are used to prevent such adhesions. Furthermore, it is desirable for the material of the adhesion barrier to be biodegradable so that the placed adhesion barrier does not need to be removed.

[0003] Anti-adhesion materials in film, liquid, and spray forms have been developed (e.g., Patent Documents 1 to 4), and Seprafilm (registered trademark), Adspray (registered trademark), and other products are commercially available. Seprafilm is a bioabsorbable film containing sodium hyaluronate and carboxymethylcellulose in a weight ratio of 2:1. Adspray is a spray-type anti-adhesion material in which two types of solutions are extruded from a spray syringe and mixed uniformly by gas at the tip of the sprayer, and then sprayed in the form of a mist around the target site. The sprayed solution quickly gels, forming a physical barrier that exerts an anti-adhesion effect. [Prior art documents] [Patent documents]

[0004] [Patent Document 1] Patent No. 6403297 [Patent Document 2] Patent No. 6239771 [Patent Document 3] Patent No. 6453412 [Patent Document 4] Patent No. 6562410 [Patent Document 5] Japanese Patent Application Laid-Open No. 2016-89175 [Patent Document 6] International Publication No. 2017 / 191827 [Non-patent literature]

[0005] [Non-Patent Document 1] Maiko Ozeki et al. PLoS ONE 14 (1):e0211391. Summary of the Invention [Problem to be solved by the invention]

[0006] However, while film-type anti-adhesion materials have the advantage of being easy to use and requiring no prior preparation, they have issues with handling, such as low flexibility and stretchability, and the risk of cracking during use. Furthermore, creating the polymer that forms the film requires heating for the polymerization reaction, the use of catalysts, and acidic or alkaline conditions. These environments are significantly different from those used in living organisms, posing technical challenges for avoiding adverse effects on the living organism.

[0007] Therefore, an object of the present invention is to provide a new material that can be produced under simple conditions and can be used as an adhesion barrier. [Means for solving the problem]

[0008] The present inventors have focused on the physical properties and biodegradability of polyrotaxanes and have completed the present invention, which provides the following [1] to

[16] . [1] A polyrotaxane (polyrotaxane B) having a chemical structure in which an axis molecule represented by formula (2) passes through the cyclodextrin ring of a compound represented by formula (1). [ka] [Wherein CD represents a cyclodextrin ring, L 2 represents a divalent organic group, and L 3 represents an ethylene group or a propylene group, and R A is a hydrogen atom or C 1-6 represents an alkyl group, and m represents an integer of 0 to 35. 2 When R is connected to N by a double bond, A does not exist.] [ka] [In the formula, R represents a hydrocarbon group larger than the inner diameter of the cyclodextrin ring, X represents a divalent organic group, and a represents an integer of 100 to 1000.] [2] A polyrotaxane (polyrotaxane B) having a chemical structure in which an axis molecule represented by formula (2) penetrates the cyclodextrin ring of a compound represented by formula (1'). [ka] [Wherein CD represents a cyclodextrin ring, L 2 represents a divalent organic group, and L 3 represents an ethylene group or a propylene group, and R A is a hydrogen atom or C 1-6 represents an alkyl group, and m represents an integer of 0 to 35. [ka] [In the formula, R represents a hydrocarbon group larger than the inner diameter of the cyclodextrin ring, X represents a divalent organic group, and a represents an integer of 100 to 1000.] [3] The polyrotaxane according to [1] or [2], wherein the axial molecule further passes through the cyclodextrin ring of the compound represented by formula (3). [ka] [Wherein CD represents a cyclodextrin ring, L 3 represents an ethylene group or a propylene group, and R A is a hydrogen atom or C 1-6represents an alkyl group, and m represents an integer of 0 to 35. [4] The polyrotaxane according to any one of [1] to [3], wherein the axial molecule is a compound represented by formula (2a) or formula (2b). [ka] [In the formula, a represents an integer of 100 to 1000.] [ka] [In the formula, a represents an integer of 100 to 1000.] [5] The polyrotaxane (polyrotaxane B) according to any one of [1] to [4], wherein the compound represented by formula (1) or formula (1') is a compound represented by formula (1a-1), (1a-2) or (1a-3). [ka] [Wherein CD represents a cyclodextrin ring, L 1 represents a divalent organic group, and L 3 represents an ethylene group or a propylene group, and R A is a hydrogen atom or C 1-6 represents an alkyl group, and m represents an integer of 0 to 35. [6] The polyrotaxane according to any one of [1] to [5], which has a transmittance of 40% or more for light at a wavelength of 600 nm. [7] A medical composition comprising the polyrotaxane (polyrotaxane B) according to any one of [1] to [6]. [8] The medical composition according to [7], which is in the form of a hydrogel. [9] The medical composition according to [7] or [8], which is a cell culture material, a tissue adhesive, a tissue regeneration substrate, or an adhesion barrier.

[10] A method for producing a polyrotaxane (polyrotaxane B) having a chemical structure in which an axis molecule represented by formula (2) penetrates a cyclodextrin ring of a compound represented by formula (1), comprising the steps of: [ka] [Wherein CD represents a cyclodextrin ring, L 2 represents a divalent organic group, and L 3 represents an ethylene group or a propylene group, and R A is a hydrogen atom or C 1-6 represents an alkyl group, and m represents an integer of 0 to 35. 2 When R is connected to N by a double bond, A does not exist.] [ka] [In the formula, R represents a hydrocarbon group larger than the inner diameter of the cyclodextrin ring, X represents a divalent organic group, and a represents an integer of 100 to 1000.] A polyrotaxane (also referred to as polyrotaxane A) having a chemical structure in which an axis molecule represented by formula (2) passes through each cyclodextrin ring of one or more compounds represented by formula (3); [ka] [Wherein CD represents a cyclodextrin ring, L 3 represents an ethylene group or a propylene group, and R A is a hydrogen atom or C 1-6 represents an alkyl group, and m represents an integer of 0 to 35. [ka] [In the formula, R represents a hydrocarbon group larger than the inner diameter of the cyclodextrin ring, X represents a divalent organic group, and a represents an integer of 100 to 1000.] A step of mixing a crosslinking agent represented by formula (4) [ka] [In the formula, L 1 represents a divalent organic group, and X 1 represents a group capable of reacting with an amino group. A manufacturing method comprising:

[11] A method for producing a polyrotaxane (polyrotaxane B) having a chemical structure in which an axis molecule represented by formula (2) penetrates a cyclodextrin ring of a compound represented by formula (1'), comprising: [ka] [Wherein CD represents a cyclodextrin ring, L 2 represents a divalent organic group, and L 3 represents an ethylene group or a propylene group, and R A is a hydrogen atom or C 1-6 represents an alkyl group, and m represents an integer of 0 to 35. [ka] [In the formula, R represents a hydrocarbon group larger than the inner diameter of the cyclodextrin ring, X represents a divalent organic group, and a represents an integer of 100 to 1000.] A polyrotaxane (also referred to as polyrotaxane A) having a chemical structure in which an axis molecule represented by formula (2) passes through each cyclodextrin ring of one or more compounds represented by formula (3); [ka] [Wherein CD represents a cyclodextrin ring, L 3 represents an ethylene group or a propylene group, and R A is a hydrogen atom or C 1-6 represents an alkyl group, and m represents an integer of 0 to 35. [ka] [In the formula, R represents a hydrocarbon group larger than the inner diameter of the cyclodextrin ring, X represents a divalent organic group, and a represents an integer of 100 to 1000.] A step of mixing a crosslinking agent represented by formula (4) [ka] [In the formula, L 1 represents a divalent organic group, and X 1 represents a group capable of reacting with an amino group. A manufacturing method comprising:

[12] The method for producing a polyrotaxane (polyrotaxane B) according to

[10] or

[11] , wherein the polyrotaxane has a chemical structure in which the axis molecule represented by formula (2) passes through the cyclodextrin ring of the compound represented by formula (1) or formula (1'), and the axis molecule further passes through the cyclodextrin ring of the compound represented by formula (3). [ka] [Wherein CD represents a cyclodextrin ring, L 3 represents an ethylene group or a propylene group, and R A is a hydrogen atom or C 1-6 represents an alkyl group, and m represents an integer of 0 to 35.

[13] The method according to any one of

[10] to

[12] , wherein the step of mixing polyrotaxane A with the crosslinking agent represented by formula (4) is carried out under catalyst-free conditions.

[14] A polyrotaxane (polyrotaxane A) having a chemical structure in which an axis molecule represented by formula (2) penetrates each cyclodextrin ring of one or more compounds represented by formula (3); [ka] [Wherein CD represents a cyclodextrin ring, L 3 represents an ethylene group or a propylene group, and R A is a hydrogen atom or C 1-6 represents an alkyl group, and m represents an integer of 0 to 35. [ka] [In the formula, R represents a hydrocarbon group larger than the inner diameter of the cyclodextrin ring, X represents a divalent organic group, and a represents an integer of 100 to 1000.] A kit comprising: a crosslinking agent represented by formula (4): [ka] [In the formula, L 1 represents a divalent organic group, and X 1 represents a group capable of reacting with an amino group.

[15] The method for producing a polyrotaxane (polyrotaxane B) according to any one of

[10] to

[14] , wherein the compound represented by formula (1) or formula (1') is a compound represented by formula (1a-1), (1a-2) or (1a-3). [ka] [Wherein CD represents a cyclodextrin ring, L 1 represents a divalent organic group, and L 3 represents an ethylene group or a propylene group, and R A is a hydrogen atom or C 1-6 represents an alkyl group, and m represents an integer of 0 to 35.

[16] A polyrotaxane (polyrotaxane A) having a chemical structure in which an axis molecule represented by formula (2) passes through each cyclodextrin ring of one or more compounds represented by formula (3). [ka] [Wherein CD represents a cyclodextrin ring, L 3 represents an ethylene group or a propylene group, and R A is a hydrogen atom or C 1-6 represents an alkyl group, and m represents an integer of 0 to 35. [ka] [In the formula, R represents a hydrocarbon group larger than the inner diameter of the cyclodextrin ring, X represents a divalent organic group, and a represents an integer of 100 to 1000.] [Effects of the Invention]

[0009] The present invention provides a new material that can be produced under simpler conditions and can be used as an anti-adhesion material. Furthermore, the polyrotaxane of the present invention can be easily prepared under mild conditions (e.g., in the range of room temperature (0-30°C) to body temperature (36-37°C)), neutral conditions (e.g., pH 6-9), aqueous systems, and catalyst-free conditions), thereby avoiding acidic or alkaline conditions or the use of organic solvents, which may have adverse effects on the body. Furthermore, the anti-adhesion material containing the polyrotaxane of the present invention can be used in the form of a transparent hydrogel, which provides excellent conformability to the application site and allows the condition of the backside (the degree of adhesion) to be visually observed while the anti-adhesion material is in place. [Brief explanation of the drawings]

[0010] [Figure 1] 1 is a 1H-NMR spectrum of the polyrotaxane of Production Example 1. [Figure 2] 1 is a 1H-NMR spectrum of the polyrotaxane of Production Example 2B. [Figure 3] 1 shows photographs of the appearance of aqueous polyrotaxane solutions of Production Examples 3 and 4. [Figure 4] 1 shows total transmittance spectra in the visible light region of polyrotaxanes of Production Examples 3 and 4. [Figure 5] 1 is a graph showing the relationship between the amount of crosslinking agent used and the gelation time when the polyrotaxane of Production Example 4 was used. [Figure 6] 6(a) and 6(b) are photographs showing cell adhesion on an uncoated polystyrene dish and a dish coated with a hydrogel derived from the polyrotaxane of Preparation Example 2A, respectively. [Figure 7] 1 shows photographs of the appearance of hydrogels of Examples 1 to 5 and Comparative Example 1. [Figure 8] 1 shows photographs of the appearance of the hydrogels of Examples 7 to 10. [Figure 9] 1 shows total transmittance spectra in the visible light region of hydrogels derived from polyrotaxanes of Production Examples 3 and 4. DETAILED DESCRIPTION OF THE INVENTION

[0011] Embodiments of the present invention will be described in detail below.

[0012] First Embodiment The first embodiment of the present invention is a polyrotaxane (polyrotaxane B) that includes a chemical structure in which an axis molecule represented by formula (2) passes through the cyclodextrin ring of a compound represented by formula (1). [ka] In formula (1), CD represents a cyclodextrin ring, L 2 represents a divalent organic group, and L 3 represents an ethylene group or a propylene group, and R A is a hydrogen atom or C 1-6 represents an alkyl group, and m represents an integer of 0 to 35. 2 The terminal carbon atom of the group is connected to the adjacent nitrogen atom by a single or double bond. When the terminal carbon atom and the nitrogen atom are connected by a single bond, R A The group is a hydrogen atom or a C 1-6 It is an alkyl group. When the terminal carbon atom and the nitrogen atom are connected by a double bond, R A There is no R group. The compound represented by formula (1) has one CD ring (cyclodextrin ring) at each end. The CD rings may be the same or different from each other. A The R group is preferably a hydrogen atom. A When the group is a hydrogen atom, the crosslinking reaction proceeds more smoothly and the crosslinking density becomes higher. [ka] In formula (2), R represents a hydrocarbon group larger than the inner diameter of the cyclodextrin ring, X represents a divalent organic group, and a represents an integer of 100 to 1,000.

[0013] The following formula (1X) is a schematic chemical formula showing the overall structure of an example of polyrotaxane B. R, X, and L in formula (1X) 2 , L 3 , R A, m, and a are as defined in formula (1) or formula (2), respectively. n is the number of cross-linked CDs threaded by one axial molecule. [ka]

[0014] In particular, the following formula is a schematic chemical formula showing a cyclodextrin ring. For convenience, "OH" indicates the hydroxyl group involved in the reaction among the hydroxyl groups in the glucose that constitutes cyclodextrin. Although only one "OH" is shown in the formula, this should not be interpreted as meaning that only one hydroxyl group is present. [ka]

[0015] Polyrotaxane B contains two compounds represented by formula (2) (axle molecules) and two compounds (cyclic molecules) with modified cyclodextrins (CDs), with one axle molecule threading through each cyclodextrin ring. In formula (1X), each axle molecule represented by formula (2) threads through n CDs. The CDs are bonded to the carbonyl carbon of the urethane bond via the oxygen atoms constituting their hydroxyl groups. Both ends of the axle molecule are capped with hydrocarbon groups bulkier than the inner diameter of the CDs used to prevent the CDs from detaching.

[0016] CD may be any of αCD, βCD, γCD, and combinations thereof. A preferred CD is αCD. CD is a compound in which multiple glucose units are linked in a ring via 1,4-glycosyl bonds. αCD, βCD, and γCD are composed of 6, 7, and 8 glucose units, respectively, and their cavity diameters are 0.5 to 0.6 nm, 0.7 to 0.8 nm, and 0.9 to 1.0 nm, respectively.

[0017] Among the hydroxyl groups contained in a CD, the number of hydroxyl groups bonded to side chains (bridges) linked to other CDs at their terminals as shown in formula (1) (i.e., the number of bridges introduced into one cyclodextrin) may be 1 to 18 per cyclodextrin, preferably 1 to 10, and more preferably 1 to 6. The larger the "number of hydroxyl groups bonded to bridges," the more side chains with other CDs at their terminals extend from the CD through which one axle molecule penetrates, and the more multiple bonds each rotaxane structure (combination of an axle molecule and a CD penetrating it) forms, forming a dense network structure.

[0018] In addition, CD may have an alkyl group having 1 to 3 carbon atoms or an oxyethylene group bonded to a hydroxyl group other than the "hydroxyl group bonded to a crosslinking moiety." When these groups are bonded, the water solubility of the polyrotaxane can be further improved.

[0019] In formula (1), m is an integer of 0 to 35, and may be 0 to 33, 0 to 30, 0 to 27, 0 to 24, 0 to 20, 0 to 17, 0 to 13, 0 to 10, 1 to 35, 1 to 33, 1 to 30, 1 to 27, 1 to 24, 1 to 20, 1 to 17, 1 to 14, 1 to 10, 1 to 9, 1 to 8, 1 to 7, 1 to 6, 1 to 5, 1 to 4, 1 to 3, 1 to 2, 2 to 5, 2 to 4, 2 to 3, 3 to 5, or 3 to 4.

[0020] In formula (1), L 2 The group is a divalent organic group, and may be an aliphatic hydrocarbon group that may contain a heteroatom (e.g., oxygen atom, nitrogen atom), such as an alkylene group or an oxyalkylene group. 2 The group is preferably [ka] L 1 The L group depends on the chemical structure of the crosslinker, which will be described later. 1 The group is as defined in formula (4).

[0021] The axial molecule represented by formula (2) has a polyethylene glycol structure at the center of the molecule and an -XR group at the end. In formula (2), the R group is a hydrocarbon group larger than the inner diameter of the CD ring cavity, and the X group is a divalent organic group. The polyethylene glycol structure allows cyclodextrin to be threaded through the axial molecule.

[0022] The R group may be any hydrocarbon group having sufficient bulk to prevent the CD ring from separating from the axial molecule. For example, when the CD ring is αCD, the R group has a length of more than 0.6 nm in a direction perpendicular to the axial direction of the axial molecule. Each R group may be the same or different. The R group is preferably an adamantyl group.

[0023] The X group is a divalent organic group that connects the polyethylene glycol structure and the R group, and is not particularly limited. Examples of the X group include an alkylene group having 1 to 10 carbon atoms, an alkenylene group having 1 to 20 carbon atoms, and an alkynylene group having 1 to 20 carbon atoms. These alkylene groups, alkenylene groups, and alkynylene groups may have an oxo group, an oxy group, or an imino group at any position, or any combination thereof. For example, a carbon atom having an oxo group combined with an oxy group forms an ester bond, and a carbon atom having an oxo group combined with an imino group forms an amide bond. An alkylene group having multiple oxy groups is also called an oxyalkylene group, and includes, for example, a polyoxyethylene group having 1 to 10 carbon atoms and a polyoxypropylene group having 1 to 10 carbon atoms. Specific examples of the organic group include alkylene groups such as methylene, ethylene, propylene, butylene, pentylene, hexylene, heptylene, octylene, nonanylene, and decylene; alkenylene groups such as propynylene, butenylene, pentenylene, hexenylene, heptenylene, octenylene, nonanylene, and decenylene; and alkynylene groups such as propargyl, butynylene, pentynylene, hexynylene, heptynylene, octynylene, nonylene, and decynylene. The organic group having an oxo group, oxy group, or imino group preferably forms an ester bond, carbonate bond, or urethane bond together with an oxygen atom derived from a hydroxyl group of cyclodextrin. Examples of organic groups having an oxo group, an oxy group, or an imino group include a carbonylmethylene group (-C(=O)CH2-), a methylenecarbonyl group (-CH2C(=O)-), a carbonylmethylene group (-C(=O)CH2-), a methylenecarbonylamino group (-CH2C(=O)NH-), a carbonylaminoethylene group (-C(=O)NHCH2-), and a carbonylaminoethyleneoxyethylene group (-C(=O)NHCH2CHOCH2CH2-). The X group preferably has an amide bond, a urethane bond, or a urea bond, which is advantageous for introducing an R group into the polyethylene glycol structure and also provides superior biodegradability.Furthermore, when the X group has a disulfide bond (shown in formula (A)), an o-nitrobenzyl structure (shown in formula (B)), a hydrazone structure (shown in formula (C)), or a ketal structure (shown in formula (D)) shown below, the biodegradability is further improved. [ka]

[0024] The number of threading cyclodextrins per axle molecule can be determined independently and does not need to be uniquely determined by the molecular length of the polyethylene glycol structure. Furthermore, stoichiometrically, CDs can include two ethylene glycol units, which are the repeating units of polyethylene glycol, and therefore the number of threading cyclodextrins is limited by the molecular weight of the axle molecule used. For example, the number of threading CDs per molecule of polyethylene glycol with a number-average molecular weight of 20,000 may be 3 to 220, preferably 5 to 150, more preferably 5 to 120, and particularly preferably 5 to 100. However, the length of the cyclodextrin axle molecule in the main chain direction (when the polyrotaxane has multiple CDs, the total length of the CDs) does not exceed the molecular length of the polyethylene glycol.

[0025] The polyethylene glycol structure in formula (2) contains ethylene glycol as a monomer unit and has a molecular length sufficient to penetrate at least one cyclodextrin. The number of ethylene glycol units forming the polyethylene glycol structure may be 60 to 1000, and more preferably 65 to 950, 80 to 900, 90 to 900, 100 to 900, 110 to 900, 130 to 880, 150 to 850, 180 to 830, 200 to 800, 230 to 770, 250 to 750, 250 to 700, 270 to 680, or 350 to 650.

[0026] Specific examples of the axial molecule represented by formula (2) include compounds represented by formula (2a) or formula (2b). [ka] [In the formula, a represents an integer of 100 to 1000.] [ka] [In the formula, a represents an integer of 100 to 1000.]

[0027] In one embodiment, polyrotaxane B preferably further contains a compound represented by formula (3). In this case, the compound represented by formula (3) is contained such that the axial molecule penetrates the CD ring. The compound represented by formula (3) contained in polyrotaxane B has a side chain having a terminal amino group, and can exert the effect of further enhancing cell adhesiveness and tissue adhesiveness. When CD is αCD, the number of side chains having a terminal amino group per CD may be 1 to 17, preferably 1 to 8, and more preferably 1 to 6. [ka] In formula (3), CD represents a cyclodextrin ring, and L 3 represents an ethylene group or a propylene group, and R A is a hydrogen atom or C 1-6 It represents an alkyl group, and m represents an integer of 0 to 35.

[0028] When polyrotaxane B further contains a compound represented by formula (3), polyrotaxane B contains both a compound represented by formula (1) and a compound represented by formula (3), and is represented, for example, by formula (1Xa). [ka] R, X, and L in formula (1Xa) 2 , L 3 , R A , m and a are as defined in formula (1X). 2 The terminal carbon atom of the group is connected to the adjacent nitrogen atom by a single or double bond. When the terminal carbon atom and the nitrogen atom are connected by a single bond, R A The group is a hydrogen atom or a C 1-6 It is an alkyl group. When the terminal carbon atom and the nitrogen atom are connected by a double bond, RA The group does not exist.

[0029] The compound represented by formula (1) in polyrotaxane B is preferably a compound represented by formula (1a-1). In one embodiment, the compound represented by formula (1) further includes a compound represented by formula (1a-2) or (1a-3). In formulas (1a-1), (1a-2), and (1a-3), CD represents a cyclodextrin ring, and L represents a cyclodextrin ring. 1 represents a divalent organic group, and L 3 represents an ethylene group or a propylene group, and R A is a hydrogen atom or C 1-6 represents an alkyl group, and m represents an integer of 0 to 35. The threading number of CDs in polyrotaxane B is the total number of threading numbers of CDs contained in each compound represented by formula (1) or formula (3). [ka] In each formula, CD represents a cyclodextrin ring, and L 1 represents a divalent organic group, and L 3 represents an ethylene group or a propylene group, and R A is a hydrogen atom or C 1-6 It represents an alkyl group, and m represents an integer of 0 to 35.

[0030] In other embodiments, the compound of formula (1) can be a compound of formula (1b), (1c), (1d), (1e), or (1f), where CD represents a cyclodextrin ring and L 1 represents a divalent organic group, and L 3 represents an ethylene group or a propylene group, and R A is a hydrogen atom or C 1-6 represents an alkyl group, and m represents an integer of 0 to 35. 2 The partial structure represented by the formula (1b) corresponds to the chemical structure of the compound represented by the formula (4). A The group does not exist. [ka] In each formula, CD represents a cyclodextrin ring, and L1 represents a divalent organic group, and L 3 represents an ethylene group or a propylene group, and R A is a hydrogen atom or C 1-6 It represents an alkyl group, and m represents an integer of 0 to 35.

[0031] In polyrotaxane B, the network structure formed by the linkage of the compounds represented by formula (1) and the axle molecules represented by formula (2) becomes denser as the length of the axle molecule, the number of threading CD rings per axle molecule, and / or the number of "modified hydroxyl groups" per CD ring increase, or as the molar ratio of the compound represented by formula (1) to the axle molecule represented by formula (2) increases. More specifically, two axle molecules represented by formula (2) are linked by two CD rings at both ends of the compound represented by formula (1). Multiple compounds represented by formula (1) are threaded through one axle molecule, and at each end there is another CD ring, which is threaded by yet another axle molecule. In this way, the linkage of multiple axle molecules forms a network structure as a whole. A sufficiently dense network structure makes the hydrogel less physically brittle and easier to handle. However, if the network structure is too dense, the molecular mobility of the CD rings may decrease, resulting in a decrease in the mechanical properties or water content of the hydrogel. When the length of the axial molecule, the number of CD threads per axial molecule, and the number of "modified hydroxyl groups" per CD are within the above ranges, the density of the network structure can be suitable for use as an anti-adhesion material. Furthermore, the higher the molar ratio of the compound represented by formula (1) to the axial molecule represented by formula (2), the more crosslinked structures there are, and the denser the network structure can be.

[0032] A preferred polyrotaxane B is a polyrotaxane (polyrotaxane B) containing a chemical structure in which the axis molecule represented by formula (2) passes through the cyclodextrin ring of the compound represented by formula (1′). [ka] In formula (1'), CD represents a cyclodextrin ring, and L2 represents a divalent organic group, and L 3 represents an ethylene group or a propylene group, and R A is a hydrogen atom or C 1-6 It represents an alkyl group, and m represents an integer of 0 to 35. [ka] In formula (2), R represents a hydrocarbon group larger than the inner diameter of the cyclodextrin ring, X represents a divalent organic group, and a represents an integer of 100-1000.

[0033] Second Embodiment The second embodiment of the present invention is a medical composition containing the polyrotaxane B according to the first embodiment. The medical composition may be in any form suitable for use in the medical field, and is preferably, for example, a hydrogel, powder, sheet, or film. The medical composition may be a cell culture material, a tissue adhesive, a tissue regeneration substrate, or an adhesion barrier. Examples of the cell culture material or tissue regeneration substrate include a coating agent for polystyrene dishes for cell culture or a cell culture substrate (e.g., a coated polystyrene dish) that takes advantage of the low cytotoxicity and cell adhesiveness of the polyrotaxane. Examples of the tissue adhesive include an adhesive for bonding tissues together that takes advantage of the adhesiveness of the polyrotaxane according to the first embodiment to cells or tissues. The medical composition according to this embodiment is particularly suitable for use as an adhesion barrier.

[0034] When the medical composition according to this embodiment has excellent optical transparency, it can facilitate microscopic observation of cells cultured on a cell culture substrate. Furthermore, polyrotaxane B can be prepared in a relatively short time by mixing polyrotaxane A with a crosslinker under mild conditions (room temperature (0-30°C) to body temperature (36-37°C), neutral (e.g., pH 6-9), aqueous, no catalyst), and can be used as an injectable gel that hydrogels at the affected area. The injectable gel can also be hydrogelized by pre-encapsulating necessary drugs or active ingredients (e.g., growth factors), making it suitable for use as a cell culture substrate, tissue adhesive, regenerative medicine substrate, or adhesion barrier.

[0035] As used herein, the term "anti-adhesion effect" refers to preventing adhesion of biological tissues (for example, adhesion between organs or adhesion between an organ and the peritoneum) without substantially irritating or adversely affecting biological tissues. More specifically, this means that the material does not generate or elute substances harmful to biological tissues, and biological tissues that come into contact with the material do not recognize the material as a foreign body and do not show defensive reactions such as inflammation or blood coagulation, or the extent of such reactions is minimal.

[0036] When the X group of the axial molecule has a disulfide bond, an o-nitrobenzyl structure, a hydrazone structure, or a ketal structure, polyrotaxane B exhibits superior biodegradability, and therefore the anti-adhesion material can also exhibit biodegradability. Such an anti-adhesion material may be left in place on biological tissue while the skin is sutured. The indwelling anti-adhesion material gradually decomposes and disappears while preventing adhesion of biological tissue after surgery. Because the anti-adhesion material according to this embodiment is a translucent hydrogel, the application site can be easily visualized, and the degree of adhesion can be visually confirmed without removing the anti-adhesion material.

[0037] The adhesion barrier according to this embodiment can be used by being placed on the biological tissue exposed at the time of incision or wound creation. The site where the adhesion barrier is used is not particularly limited, and it can be used at the site of a surgical incision or wound, for example, the arm, chest, abdomen, waist, back, buttocks, thighs, legs, etc. The size of the adhesion barrier may be adjusted according to the site to which it is to be applied.

[0038] The medical composition according to this embodiment can be prepared immediately in the form of a hydrogel by mixing polyrotaxane A and a crosslinker represented by formula (4). Gelation progresses and fluidity decreases approximately 3 to 30 minutes after mixing. Therefore, polyrotaxane B can be formed by applying the two liquids to biological tissue and mixing them, making the composition suitable for use in laparoscopic surgery. Furthermore, because it takes approximately 3 to 30 minutes for the hydrogel to lose its fluidity, an anti-adhesion material that fits the shape of biological tissue can be formed. Alternatively, an anti-adhesion material prepared by gelation in advance may be placed in biological tissue. The medical composition may also be a mass, sheet, or powder obtained by gelatinizing polyrotaxane A and a crosslinker represented by formula (4) together and then freeze-drying. Medical compositions in such forms can also be used by placing or spraying them on a desired location (e.g., an affected area) and then applying physiological saline to restore the gel state. By mixing the necessary drugs or active ingredients (growth factors, etc.) into a bulk or powdered medical composition, or by wrapping the composition in a sheet, and then pouring saline over it, it can be used for a wider range of purposes.

[0039] The medical composition according to this embodiment contains the polyrotaxane B according to the first embodiment and may further contain optional components, if necessary. Examples of optional components include antioxidants, weather stabilizers, heat stabilizers, lubricants, crystal nucleating agents, UV absorbers, colorants, and antibacterial agents. The content of the optional components may be 0.5 to 10% by mass relative to the total mass of the medical composition. When the medical composition is in the form of a lump, sheet, or powder, the content of polyrotaxane B may be 30% by mass or more, and may be 50 to 99.5% by mass, 50 to 95% by mass, 55 to 90% by mass, 60 to 90% by mass, 65 to 90% by mass, 70 to 90% by mass, 75 to 90% by mass, 80 to 90% by mass, 85 to 90% by mass, 60 to 85% by mass, 60 to 80% by mass, 65 to 80% by mass, 65 to 75% by mass, or 65 to 70% by mass. When the medical composition is a hydrogel, the content of polyrotaxane B may be 0.5 to 50 mass%, 2 to 50 mass%, 2 to 45 mass%, 5 to 45 mass%, 5 to 40 mass%, 7 to 40 mass%, 7 to 35 mass%, or 10 to 35 mass% relative to the total mass of the medical composition.

[0040] The medical composition according to the present embodiment may be in the form of a hydrogel. In this specification, the term "hydrogel" refers to a swollen polymeric substance with a three-dimensional structure that is insoluble in water. The hydrogel is essentially composed of the polyrotaxane B according to the first embodiment and water. The polyrotaxane B according to the first embodiment can be produced according to the production method described below. The reaction between the polyrotaxane A and the crosslinker represented by formula (4) proceeds in a short time even under mild conditions (e.g., a temperature range from room temperature (0-30°C) to body temperature (36-37°C), a neutral pH (e.g., a pH of 6-9), an aqueous system, and no catalyst). Therefore, the polyrotaxane B according to the first embodiment can be produced in the form of a hydrogel by mixing aqueous solutions containing both raw materials. Furthermore, the higher the polyrotaxane concentration and the greater the amount of crosslinker used, the higher the storage modulus of the resulting hydrogel and the lower its swelling in water.

[0041] The medical composition according to this embodiment is preferably translucent. As used herein, "translucent" means exhibiting a transmittance of 15% or more across the entire visible light range (wavelength: 380 to 780 nm) at an optical path length of 10 mm. In other words, "translucent" can also be understood as a transmittance of 15% or more / 10 mm. The medical composition according to this embodiment preferably exhibits a transmittance of 18% or more, 20% or more, 25% or more, 30% or more, 35% or more, or 40% or more across the entire visible light range. In particular, the medical composition according to this embodiment preferably exhibits a transmittance of 40% or more, 45% or more, 50% or more, 55% or more, 60% or more, 65% or more, or 70% or more at a wavelength of 600 nm.

[0042] The light transmittance can be measured, for example, under the following conditions. For the measurement method, a general method for measuring light transmittance can be referred to, and the examples described below may also be referred to.

[0043] (Measurement conditions) Equipment: Hitachi spectrophotometer (product name: U-2910) Measurement mode: Wavelength scan Data mode: ABS mode Measurement wavelength: 210nm~830nm Scan speed: 400nm / min Cell: Polystyrene cell (Fisher Scientific) Cell length: 10mm Baseline: MilliQ water

[0044] Third Embodiment A third embodiment of the present invention is a method for producing a polyrotaxane (polyrotaxane B) having a chemical structure in which an axis molecule represented by formula (2) penetrates each cyclodextrin ring of one or more compounds represented by formula (1). [ka] [Wherein CD represents a cyclodextrin ring, L 2 represents a divalent organic group, and L 3represents an ethylene group or a propylene group, and R A is a hydrogen atom or C 1-6 represents an alkyl group, and m represents an integer of 0 to 35. 2 When N is connected with a double bond, R A does not exist.] [ka] [In the formula, R represents a hydrocarbon group larger than the inner diameter of the cyclodextrin ring, X represents a divalent organic group, and a represents an integer of 100 to 1000.]

[0045] The method for producing polyrotaxane B according to this embodiment includes the steps of: (a) producing a polyrotaxane (polyrotaxane A) having a chemical structure in which an axis molecule represented by formula (2) penetrates through each cyclodextrin ring of one or more compounds represented by formula (3); [ka] [Wherein CD represents a cyclodextrin ring, L 3 represents an ethylene group or a propylene group, and R A is a hydrogen atom or C 1-6 represents an alkyl group, and m represents an integer of 0 to 35. [ka] [In the formula, R represents a hydrocarbon group larger than the inner diameter of the cyclodextrin ring, X represents a divalent organic group, and a represents an integer of 100 to 1000.] and a crosslinking agent represented by formula (4). [ka] [In the formula, L 1 represents a divalent organic group, and X 1 represents a group capable of reacting with an amino group.

[0046] In this process, polyrotaxane A and a crosslinker represented by formula (4) are mixed together, and two molecules of polyrotaxane A react with the crosslinker represented by formula (4) to bond, producing polyrotaxane B. The reaction in this process does not require the presence of a catalyst and proceeds under mild conditions (e.g., in the range of room temperature (0-30°C) to body temperature (36-37°C), neutral (e.g., pH 6-9), aqueous, and catalyst-free conditions). In polyrotaxane A, R A When the group is a hydrogen atom, the crosslinking reaction proceeds more smoothly, resulting in polyrotaxane B with a higher crosslinking density. Because no catalyst is required for the crosslinking reaction, it can be prepared immediately and does not require purification before use. Another advantage is that even if the crosslinking reaction is carried out on the target biological tissue, the catalyst does not remain in the body.

[0047] The following is a chemical formula that schematically illustrates this process: In this process, polyrotaxane A represented by formula (3X) and crosslinker represented by formula (4) are bonded to produce polyrotaxane B. [ka] In each formula, R represents a hydrocarbon group larger than the inner diameter of the cyclodextrin ring, X represents a divalent organic group, and L 3 represents an ethylene group or a propylene group, a represents an integer of 100 to 1000, and R A is a hydrogen atom or C 1-6 represents an alkyl group, n represents an integer of 100 to 1000, m represents an integer of 0 to 35, and L 1 represents a divalent organic group, and X 1 indicates a group capable of reacting with an amino group. 2 When N is connected with a double bond, R A does not exist. 1 is defined as in equation (4). 2 teeth, [ka] Shows.

[0048] This step may be carried out in a solvent or without a solvent. It is preferable to mix polyrotaxane A and the crosslinking agent represented by formula (4) in the presence of a solvent. The solvent is not limited as long as it does not suppress or inhibit the desired reaction, and can be appropriately selected depending on the application. A preferred solvent is water. In practice, the simplest method is to prepare a solution containing polyrotaxane A and a solution containing the crosslinking agent represented by formula (4), and then mix these two solutions at the time of use. The amount of solvent is not limited as long as the reaction between polyrotaxane A and the crosslinking agent proceeds at a practical rate. The amount of solvent may be, for example, 100,000 to 10,000,000 mL per mole of polyrotaxane A used, or 500,000 to 100,000,000 mL per mole of crosslinking agent used.

[0049] In this step, the pH of the reaction system may be neutral, for example, 6 to 9, and may be preferably 6 to 8, 6.5 to 8, 6.5 to 7.5, 6.5 to 7, 6 to 7.5, or 6 to 7.

[0050] Polyrotaxane A is a polyrotaxane containing a chemical structure in which an axis molecule represented by formula (2) passes through each cyclodextrin ring of one or more compounds represented by formula (3). [ka] [Wherein CD represents a cyclodextrin ring, L 3 represents an ethylene group or a propylene group, and R A is a hydrogen atom or C 1-6 represents an alkyl group, and m represents an integer of 0 to 35. [ka] [In the formula, R represents a hydrocarbon group larger than the inner diameter of the cyclodextrin ring, X represents a divalent organic group, and a represents an integer of 100 to 1000.]

[0051] CD is defined as in formula (1) and may be any of αCD, βCD, γCD, and combinations thereof. A preferred CD is αCD. A CD is a compound in which multiple glucose molecules are linked in a ring via 1,4-glycosyl bonds. αCD, βCD, and γCD are composed of 6, 7, and 8 glucose molecules, respectively, and their cavity diameters are 0.5 to 0.6 nm, 0.7 to 0.8 nm, and 0.9 to 1.0 nm, respectively.

[0052] Of the hydroxyl groups contained in a CD, the number of hydroxyl groups having side chains linked to other CDs at their terminals as shown in formula (3) (i.e., the number of side chains introduced into one cyclodextrin) may be 1 to 18 per cyclodextrin, preferably 1 to 10, and more preferably 1 to 6. A larger "number of hydroxyl groups having side chains bound thereto" means that more side chains having other CDs at their terminals extend from a CD through which one axial molecule passes.

[0053] In CD, an alkyl group having 1 to 3 carbon atoms or an oxyethylene group may be bonded to a hydroxyl group other than the "hydroxyl group bonded to a side chain." When these groups are bonded, the water solubility of the polyrotaxane can be further improved.

[0054] In formula (3), m is an integer of 0 to 35, and may be 0 to 33, 0 to 30, 0 to 27, 0 to 24, 0 to 20, 0 to 17, 0 to 13, 0 to 10, 1 to 35, 1 to 33, 1 to 30, 1 to 27, 1 to 24, 1 to 20, 1 to 17, 1 to 14, 1 to 10, 1 to 9, 1 to 8, 1 to 7, 1 to 6, 1 to 5, 1 to 4, 1 to 3, 1 to 2, 2 to 5, 2 to 4, 2 to 3, 3 to 5, or 3 to 4.

[0055] The axial molecule represented by formula (2) has a polyethylene glycol structure at the center of the molecule and an -XR group at the end. In formula (2), the R group is a hydrocarbon group larger than the inner diameter of the CD ring cavity, and the X group is a divalent organic group. The polyethylene glycol structure allows cyclodextrin to be threaded through the axial molecule.

[0056] The R group may be any hydrocarbon group having sufficient bulk to prevent the CD ring from separating from the axial molecule. For example, when the CD ring is αCD, the R group has a length of more than 0.6 nm in a direction perpendicular to the axial direction of the axial molecule. Each R group may be the same or different. The R group is preferably an adamantyl group.

[0057] The X group is a divalent organic group that connects the polyethylene glycol structure and the R group, and is not particularly limited. Examples of the X group include an alkylene group having 1 to 10 carbon atoms, an alkenylene group having 1 to 20 carbon atoms, and an alkynylene group having 1 to 20 carbon atoms. These alkylene groups, alkenylene groups, and alkynylene groups may have an oxo group, an oxy group, or an imino group at any position, or any combination thereof. For example, a carbon atom having an oxo group combined with an oxy group forms an ester bond, and a carbon atom having an oxo group combined with an imino group forms an amide bond. An alkylene group having multiple oxy groups is also called an oxyalkylene group, and includes, for example, a polyoxyethylene group having 1 to 10 carbon atoms and a polyoxypropylene group having 1 to 10 carbon atoms. Specific examples of the organic group include alkylene groups such as methylene, ethylene, propylene, butylene, pentylene, hexylene, heptylene, octylene, nonanylene, and decylene; alkenylene groups such as propynylene, butenylene, pentenylene, hexenylene, heptenylene, octenylene, nonanylene, and decenylene; and alkynylene groups such as propargyl, butynylene, pentynylene, hexynylene, heptynylene, octynylene, nonylene, and decynylene. The organic group having an oxo group, oxy group, or imino group preferably forms an ester bond, carbonate bond, or urethane bond together with an oxygen atom derived from a hydroxyl group of cyclodextrin. Examples of organic groups having an oxo group, an oxy group, or an imino group include a carbonylmethylene group (-C(=O)CH2-), a methylenecarbonyl group (-CH2C(=O)-), a carbonylmethylene group (-C(=O)CH2-), a methylenecarbonylamino group (-CH2C(=O)NH-), a carbonylaminoethylene group (-C(=O)NHCH2-), and a carbonylaminoethyleneoxyethylene group (-C(=O)NHCH2CHOCH2CH2-). The X group preferably has an amide bond, a urethane bond, or a urea bond, which is advantageous for introducing an R group into the polyethylene glycol structure and also provides superior biodegradability.Furthermore, when the X group has a disulfide bond (shown in formula (A)), an o-nitrobenzyl structure (shown in formula (B)), a hydrazone structure (shown in formula (C)), or a ketal structure (shown in formula (D)) shown below, the biodegradability is further improved. [ka]

[0058] The number of threading cyclodextrins per axle molecule can be determined independently and does not need to be uniquely determined by the molecular length of the polyethylene glycol structure. Furthermore, stoichiometrically, CDs can include two ethylene glycol units, which are the repeating units of polyethylene glycol, and therefore the number of threading cyclodextrins is limited by the molecular weight of the axle molecule used. For example, the number of threading CDs (n) per molecule of polyethylene glycol with a number-average molecular weight of 20,000 may be 3 to 220, preferably 5 to 150, more preferably 5 to 120, and particularly preferably 5 to 100. However, the length of the cyclodextrin axle molecule in the main chain direction (when the polyrotaxane has multiple CDs, the total length of the CDs) does not exceed the molecular length of polyethylene glycol.

[0059] The polyethylene glycol structure in formula (2) contains ethylene glycol as a monomer unit and has a molecular length sufficient to penetrate at least one cyclodextrin. The number of ethylene glycol units forming the polyethylene glycol structure may be 60 to 1000, and more preferably 65 to 950, 80 to 900, 90 to 900, 100 to 900, 110 to 900, 130 to 880, 150 to 850, 180 to 830, 200 to 800, 230 to 770, 250 to 750, 250 to 700, 270 to 680, or 350 to 650.

[0060] Specific examples of the axial molecule represented by formula (2) include compounds represented by formula (2a) or formula (2b). [ka] [In the formula, a represents an integer of 100 to 1000.] [ka] [In the formula, a represents an integer of 100 to 1000.]

[0061] The crosslinking agent is a compound represented by formula (4): 1 The group represents a divalent organic group, preferably a chain or branched alkane, more preferably a C 1-10 It is an alkane. 1 The group may be an aliphatic hydrocarbon group which may contain a heteroatom (e.g., oxygen atom, nitrogen atom), and may be, for example, a carbonyl group, an alkylene group, or an oxyalkylene group. X 1 The group may be any group capable of reacting with a primary or secondary amino group, such as an oxiranyl group, a formyl group, an isocyanate group, a succinimidyloxycarbonyl group, a 3-sulfosuccinimidyloxycarbonyl group, or a carboxyimidate group. Among these, one or more groups selected from the group consisting of an oxiranyl group, a formyl group, and an isocyanate group are preferred because they do not produce by-products in the reaction of the amino group. A more preferred crosslinking agent is a compound represented by formula (4a) described below.

[0062] X 1 When the group is an oxiranyl group, the compound represented by formula (4) is represented by formula (4a) and has one oxirane structure at each end. 1 The group is L in formula (4). 1 The definition of the group is the same as above. Preferred crosslinking agents are glycidyl ethers of α,ω-alkanediols, such as ethylene glycol diglycidyl ether, propanediol diglycidyl ether, 1,4-butanediol diglycidyl ether, 1,5-pentanediol diglycidyl ether, and the like. [ka]

[0063] By reacting with a compound represented by formula (3), a compound represented by formula (1a-1), (1a-2) or (1a-3) can be obtained. 3 , m and R A corresponds to the chemical structure of the compound represented by formula (3) used, and L 1 corresponds to the chemical structure of the compound represented by formula (4a) used. [ka]

[0064] X 1 When the group is a formyl group, the compound represented by formula (4) is represented by formula (4b) and has one formyl group at each end. 1 The group is L in formula (4). 1 The definition of the group is the same as above. A preferred cross-linking agent is glutaraldehyde. [ka]

[0065] By reacting with the compound represented by formula (3), a compound represented by formula (1b) is obtained. When the compound represented by formula (4b) is used as a crosslinking agent, the compound represented by formula (3) must be a primary amine (in formula (3), R A In formula (1b), CD, L 3 , and m correspond to the chemical structure of the compound represented by formula (3) used, and L 1 corresponds to the chemical structure of the compound represented by formula (4b) used. [ka]

[0066] X 1 When the group is an isocyanate group, the compound represented by formula (4) is represented by formula (4c) and has one isocyanate group at each end. 1 The group is L in formula (4). 1The definition of the group is the same as above. A preferred crosslinking agent is 1,6-hexanediisocyanate. [ka]

[0067] By reacting with a compound represented by formula (3), a compound represented by formula (1c) is obtained. 3 , m and R A corresponds to the chemical structure of the compound represented by formula (3) used, and L 1 corresponds to the chemical structure of the compound represented by formula (4c) used. [ka]

[0068] X 1 When the group is a succinimidyloxycarbonyl group or a 3-sulfosuccinimidyloxycarbonyl group, the compound represented by formula (4) is represented by formula (4d), and has one succinimidyloxycarbonyl group or one 3-sulfosuccinimidyloxycarbonyl group at each end. 1 The group is L in formula (4). 1 The definition is the same as that of the L group. 4 The group represents a hydrogen atom or a sulfo group. Preferred crosslinking agents are disuccinimidyl suberate, bis(3-sulfosuccinimidyl)suberate, 3,3'-dithiobis(3-sulfosuccinimidyl propionate), bis(succinimidyl)penta(ethylene glycol), bis(succinimidyl)nona(polyethylene glycol), and polyethylene glycol-modified bis(sulfosuccinimidyl)suberate. [ka]

[0069] By reacting with a compound represented by formula (3), a compound represented by formula (1d) is obtained. 3 , m and R Acorresponds to the chemical structure of the compound represented by formula (3) used, and L 1 corresponds to the chemical structure of the compound represented by formula (4d) used. [ka]

[0070] X 1 When the group is a carboxyimidate group, the compound represented by formula (4) is represented by formula (4e) and has one carboxyimidate group at each end. 1 The group is L in formula (4). 1 The definition is the same as that of the group. When using the crosslinker represented by formula (4e), the reactivity of the crosslinking reaction can be higher when the pH of the reaction system is closer to 8. Preferred crosslinkers are dimethyl pimelimidate dihydrochloride or dimethyl suberimidate dihydrochloride. [ka]

[0071] By reacting with a compound represented by formula (3), a compound represented by formula (1e) is obtained. 3 , m and R A corresponds to the chemical structure of the compound represented by formula (3) used, and L 1 corresponds to the chemical structure of the compound represented by formula (4e) used. [ka]

[0072] X 1 When the group is a carbonate group, the compound represented by formula (4) is represented by formula (4f) and has one carbonate group at each end. 1 The group is L in formula (4). 1 The definition of the R group is the same as that of the R group. XThe group represents a succinimidyloxycarbonyl group or a 3-sulfosuccinimidyloxycarbonyl group. When using a crosslinker represented by formula (4f), the reactivity of the crosslinking reaction can be higher when the pH of the reaction system is closer to 8. A preferred crosslinker is disuccinimidyl carbonate. [ka]

[0073] By reacting with the compound represented by formula (3), a compound represented by formula (1f) is obtained. 3 , m and R A corresponds to the chemical structure of the compound represented by formula (3) used. [ka]

[0074] The amount of crosslinking agent used may be 0.01 to 5 mol, preferably 0.1 to 3 mol, 0.1 to 2 mol, or 0.2 to 2 mol, per mol of terminal amino groups contained in the compound represented by formula (3X), which will be described later. When the amount of crosslinking agent used per mol of terminal amino groups contained in the compound represented by formula (3X) is 0.1 to 3 mol, the number of terminal amino groups (unreacted amino groups) contained in polyrotaxane B represented by formula (1X) tends to increase, thereby further improving the cell adhesiveness or tissue adhesiveness of polyrotaxane B. When the amount of crosslinking agent used per mol of terminal amino groups contained in the compound represented by formula (3X) is 0.1 to 3 mol, a non-brittle gel with excellent handleability is obtained.

[0075] In this step, all of the terminal amino groups contained in polyrotaxane A represented by formula (3X) do not necessarily have to react with the crosslinking agent (crosslinking reaction). If the terminal amino groups do not react with the crosslinking agent, polyrotaxane B contains both the compound represented by formula (1) and the compound represented by formula (3), and has a chemical structure represented by formula (1Xa). [ka] R, X, and L in formula (1Xa) 2 , L 3 , R A , m, and a are each as defined in formula (1X), p is an arbitrary integer from 0 to n, and q is an arbitrary integer from 1 to n.

[0076] The method for producing a polyrotaxane according to this embodiment may further include a step of mixing a polyrotaxane represented by formula (5) (a polyrotaxane not modified at CD) with polyoxyethylenediamine in the presence of a carbonylating agent to obtain a compound represented by formula (3X).

[0077] In this step, polyrotaxane represented by formula (5) is reacted with diamine represented by formula (6) in the presence of a carbonylating agent to bond the oxygen atom constituting the hydroxyl group of CD with the nitrogen atom of polyoxyethylenediamine via a carbonyl group to form a urethane bond. Thereafter, optionally, the terminal primary amino group is alkylated to form R A A group may be introduced. The following formula is a chemical formula that schematically shows this step. In the formula, R, X, and L 3 , R A , m, n and a are as defined in formula (1) or formula (2). [ka]

[0078] The carbonylating agent may be any compound having two leaving groups bonded to the carbon atom of a carbonyl group, such as 1,1-carbonyldiimidazole (CDI), phosgene, or triphosgene. A preferred carbonylating agent is CDI. The amount of the carbonylating agent used may be 1 to 20 moles, preferably 2 to 15 moles, per mole of cyclodextrin contained in the polyrotaxane represented by formula (5).

[0079] The diamine represented by formula (6) is a polyoxyalkylene having amino groups at both ends, and the number m of oxyethylene units is as defined in formula (1).3 represents an ethylene group or a propylene group. Examples of the diamine represented by formula (6) include 3,3'-diaminopropyl ether and polyethylene glycol diamine. The amount of the diamine represented by formula (6) used may be 10 to 400 mol, and preferably 20 to 200 mol, per 1 mol of cyclodextrin contained in the polyrotaxane represented by formula (5).

[0080] This step is preferably carried out in an organic solvent. The organic solvent may be any solvent that does not suppress or inhibit this reaction, and examples thereof include tetrahydrofuran (THF), tert-butyl methyl ether (TBME), cyclopentyl methyl ether (CPME), and dimethyl sulfoxide (DMSO). A preferred organic solvent is DMSO. The amount of organic solvent used may be 10 to 200 mL, and preferably 20 to 100 mL, per 1 g of polyrotaxane.

[0081] This step can be carried out within a temperature range of 0 to 70° C., or may be carried out at room temperature (for example, 0 to 30° C.). The reaction solution may be heated as appropriate to promote the reaction.

[0082] The number of introduced side chains (side chains derived from the diamine represented by formula (6)) is 1 It can be calculated by H-NMR analysis.

[0083] R at the terminal amino group A The reaction for introducing the group can be carried out by N-alkylation, which is well known in the field of organic chemistry. Examples of alkylating agents include C 1-6 Alkyl halides, C 1-6 Alkanol mesylate, C 1-6 C with a leaving group such as alkanol tosylate 1-6 Alkanes include formaldehyde and C 1-5 A borohydride reagent such as sodium cyanoborohydride may be added in the presence of an alkylaldehyde to carry out a reductive N-alkylation reaction.

[0084] Fourth Embodiment A fourth embodiment of the present invention is a polyrotaxane (polyrotaxane A) comprising a chemical structure in which an axis molecule represented by formula (2) passes through each cyclodextrin ring of one or more compounds represented by formula (3). Polyrotaxane A has a chemical structure represented by formula (3X). [ka] In formula (3), CD represents a cyclodextrin ring, and L 3 represents an ethylene group or a propylene group, and R A is a hydrogen atom or C 1-6 It represents an alkyl group, and m represents an integer of 0 to 35. [ka] In formula (2), R represents a hydrocarbon group larger than the inner diameter of the cyclodextrin ring, X represents a divalent organic group, and a represents an integer of 100-1000.

[0085] The polyrotaxane A according to this embodiment has excellent water solubility and is suitable for use as a part of the kit according to the fourth embodiment.

[0086] Fifth Embodiment A fifth embodiment of the present invention is a kit comprising a polyrotaxane (polyrotaxane A) having a chemical structure in which an axis molecule represented by formula (2) passes through each cyclodextrin ring of one or more compounds represented by formula (3), and a crosslinker represented by formula (4). [ka] In formula (3), CD represents a cyclodextrin ring, and L 3 represents an ethylene group or a propylene group, and R A is a hydrogen atom or C 1-6 It represents an alkyl group, and m represents an integer of 0 to 35. [ka] In formula (2), R represents a hydrocarbon group larger than the inner diameter of the cyclodextrin ring, X represents a divalent organic group, and a represents an integer of 100-1000.

[0087] As described in the third embodiment, polyrotaxane A and the crosslinking agent represented by formula (4) are mixed to produce polyrotaxane B. For the method of using the kit according to this embodiment, see the description of the third embodiment.

[0088] The polyrotaxane A and the crosslinker represented by formula (4) contained in the kit may be in the form of a composition (e.g., a solution or dispersion) so long as they are in a separate state so that they can be mixed at the time of use. When these two components are in the form of a composition, they are easily mixed at the time of use, making the kit more practical. A preferred embodiment is a kit containing a composition containing polyrotaxane A and a composition containing the crosslinker represented by formula (4). These two compositions may be contained in separate containers or in separate compartments of a container with multiple compartments. Furthermore, when using these two compositions, they may be mixed manually or applied to the target biological tissue using a syringe equipped with a static mixer. Because the crosslinking reaction proceeds rapidly, they may be prepared just before use. Because the crosslinking reaction proceeds even at room temperature to body temperature (approximately 37°C), the composition containing polyrotaxane A and the composition containing the crosslinker represented by formula (4) may be applied to the biological tissue to be used and then mixed. [Example]

[0089] The present invention will be described in more detail below with reference to examples and comparative examples. The abbreviations used in the examples should be understood as those generally understood by those skilled in the art, and are understood, for example, as follows: αCD: α-cyclodextrin BOP: Benzotriazolyl-N-hydroxytrisdimethylaminophosphonium hexafluorophosphate CDI: 1,1-carbonyldiimidazole DMF: dimethylformamide DMSO: dimethyl sulfoxide DMT-MM: 4-(4,6-dimethoxy-1,3,5-triazin-2-yl)-4-methylmorpholinium chloride EDIPA: N,N-diisopropylethylamine EGDE: Ethylene glycol diglycidyl ether BDDE: 1,4-butanediol diglycidyl ether HOBt: 1-hydroxybenzotriazole MilliQ water: Ultrapure water purified by the Milli-Q ultrapure water system TEMPO: 2,2,6,6-tetramethylpiperidine-N-oxide THF: tetrahydrofuran

[0090] 1 H-NMR was corrected using tetramethylsilane as an internal standard and is expressed using chemical shifts (unit: ppm).

[0091] 1. Preparation of polyrotaxane Manufacturing Example 1 [ka]

[0092] [Process 1] Polyethylene glycol (20.0 g, 1.0 mmol) with a number-average molecular weight of 20,000 was dissolved in water (340 mL), and TEMPO (330 mg, 2.11 mmol), sodium bromide (330 mg, 3.21 mmol), and sodium hypochlorite (4.0%, 40 mL) were added. The reaction mixture was confirmed to be alkaline (pH = 9) and stirred at 23 °C. After 20 minutes, ethanol (40 mL) and 2 M hydrochloric acid (6 mL) were added dropwise, and the reaction mixture was confirmed to be acidic (pH = 2). The reaction mixture was extracted with methylene chloride, and the solvent in the resulting organic layer was evaporated, followed by dissolution in 50 °C ethanol (200 mL). The resulting solution was cooled to 4 °C to precipitate crystals, and the solution containing the crystals was filtered with chilled ethanol to obtain the residue. The residue was again dissolved in 50 °C ethanol, cooled to precipitate crystals, and filtered with chilled ethanol. The resulting residue was dried under reduced pressure to obtain a dicarboxylic acid (14.5 g, yield: 73%).

[0093] [Process 2] The dicarboxylic acid (10.0 g, 0.500 mmol) obtained in step 1 was dissolved in water (300 mL), and a saturated aqueous solution (45 mL) of αCD (50.0 g, 51.4 mmol) was added and stirred at 23° C. After 24 hours, the reaction solution was centrifuged to collect the precipitate, which was then freeze-dried for 3 days to obtain a crude pseudopolyrotaxane.

[0094] [Step 3] 1-Adamantanamine (13.8 g, 91.5 mmol) was dissolved in ultra-dehydrated DMF (340 ml), and the pseudopolyrotaxane (51.4 g) obtained in step 2 and DMT-MM (22.6 g, 81.5 mmol) were added to the reaction solution, followed by stirring at 23°C. After 24 hours, the crude product was washed with water and methanol, reprecipitated with aqueous DMSO, centrifuged, and freeze-dried for 4 days to obtain the polyrotaxane of Production Example 1 (32.4 g, two-step yield: 54%) as a powder. 1 The H-NMR spectrum (frequency: 500 MHz, solvent: DO+NaOD) is shown in Figure 1.

[0095] GPC analysis confirmed that the non-permeable αCD had been removed. Measurement conditions Device: Prominence-i LC-2030 Plus (Shimadzu) Detector: RID-20A refractive index detector (Shimadzu) Column: TSK gel (registered trademark) α-4000 and α-2500 (300 mm length, 7.8 mm inner diameter) (manufactured by Tosoh Corporation) Column temperature: 60℃ Flow rate: 0.35mL / min

[0096] Manufacturing example 2A~C [ka]

[0097] [Process 1] The polyrotaxane of Preparation Example 1 (8.00 g, 0.108 mmol) was dissolved in anhydrous DMSO (250 mL), CDI (7.79 g, 48.0 mmol) was added, and the reaction solution was stirred at 23°C. After one day, the reaction solution was added dropwise to 1,2-bis(2-aminoethoxyethane) (71 mL, 0.48 mol) and stirred for another day at 23°C. The reaction solution was then purified by dialysis for five days. The product was lyophilized for two days to obtain the polyrotaxane of Preparation Example 2B (7.19 g, yield: 53%) as a powder. The number of introduced side chains was 1 The molecular weight of the polyrotaxane of Production Example 2B was determined by H-NMR analysis (frequency: 500 MHz, solvent: DO+NaOD). 1 H-NMR is shown in Figure 2.

[0098] Manufacturing Example 3 [ka]

[0099] [Process 1] Polyethylene glycol (20.0 g, 1.0 mmol) with a number-average molecular weight of 20,000 was dissolved in water (340 mL), and TEMPO (330 mg, 2.11 mmol), sodium bromide (330 mg, 3.21 mmol), and sodium hypochlorite (4.0%, 40 mL) were added. The reaction mixture was confirmed to be alkaline (pH = 9) and stirred at 23 °C. After 20 minutes, ethanol (40 mL) and 2 M hydrochloric acid (6 mL) were added dropwise, and the reaction mixture was confirmed to be acidic (pH = 2). The reaction mixture was extracted with methylene chloride, and the solvent in the resulting organic layer was evaporated, followed by dissolution in 50 °C ethanol (200 mL). The resulting solution was cooled to 4 °C to precipitate crystals, and the solution containing the crystals was filtered with chilled ethanol to obtain the residue. The residue was again dissolved in 50 °C ethanol, cooled to precipitate crystals, and filtered with chilled ethanol. The resulting residue was dried under reduced pressure to obtain a dicarboxylic acid (14.5 g, yield: 73%).

[0100] [Process 2] The dicarboxylic acid (5.0 g, 0.25 mmol) obtained in step 1 was dissolved in water (20 mL), and a saturated aqueous solution (150 mL) of αCD (25.0 g, 25.7 mmol) was added and stirred at 23° C. After 24 hours, the reaction solution was centrifuged to collect the precipitate, which was then freeze-dried for 4 days to obtain a crude pseudopolyrotaxane.

[0101] [Step 3] 1-Adamantanamine (6.92 g, 45.8 mmol) was dissolved in ultra-dehydrated DMF (120 ml), and the pseudopolyrotaxane (22.0 g) obtained in step 2 and DMT-MM (11.3 g, 40.8 mmol) were added to the reaction solution, followed by stirring at 23°C. After 24 hours, the crude product was washed with water and methanol, reprecipitated with aqueous DMSO, centrifuged, and freeze-dried for 4 days to obtain polyrotaxane (15.6 g, 2-step yield: 52%) as a powder. The threading number of αCD was 1 The NMR spectrum was determined by H-NMR (frequency: 500 MHz, solvent: D2O+NaOD).

[0102] [Step 4] Polyrotaxane (4.00 g, 41.6 μmol) and CDI (5.28 g, 32.6 mmol) were dissolved in anhydrous DMSO (130 mL), and the reaction solution was stirred at 23°C. After one day, the reaction solution was added dropwise to ethylenediamine (44 mL, 0.652 mol) and stirred at 23°C for another day. The reaction solution was then purified by dialysis for five days. The product was freeze-dried for 11 days to obtain the polyrotaxane of Production Example 3 (2.78 g, yield: 51%) as a powder. The number of amino groups was 1 The NMR spectrum was determined by H-NMR (frequency: 500 MHz, solvent: D2O+NaOD).

[0103] The data for the polyrotaxanes of Production Examples 1, 2A to 2C, and 3 are shown in Table 1. The number of side chains introduced per CD is: 1 Calculated based on H-NMR analysis. [Table 1]

[0104] Production Example 4 [ka]

[0105] [Process 1] Polyethylene glycol (50.0 g, 2.5 mmol) with a number-average molecular weight of 20,000 was dissolved in THF (200 mL), CDI (1.0 g, 6.2 mmol) was added, and the reaction solution was stirred at 50°C. After 18 hours, the reaction solution was added dropwise to ethylenediamine (3.0 mL, 44 mmol) and stirred at 50°C for an additional 2 hours. Subsequently, ethanol (200 mL) was added, and the mixture was allowed to stand at -20°C for 2 hours. The mixture was washed with chilled ethanol (200 mL), filtered, and lyophilized to obtain the diamine.

[0106] [Process 2] The obtained diamine (3.0 g, 0.15 mmol) was dissolved in water (100 mL), and αCD (12.0 g, 12.3 mmol) was added and stirred at 4° C. The reaction solution was centrifuged to collect the precipitate, which was then freeze-dried to obtain a crude pseudopolyrotaxane.

[0107] [Step 3] 1-Adamantanecarboxylic acid (2.45 g, 13.6 mmol), BOP (5.25 g, 11.9 mmol), HOBt (1.75 g, 12.6 mmol), and EDIPA (2.275 mL, 17.6 mmol) were dissolved in THF (100 mL), and the pseudopolyrotaxane (14.0 g) obtained in step 2 was added. The reaction solution was stirred at 4°C. After 48 hours, the crude product was washed with DMF and methanol, reprecipitated with aqueous DMSO, centrifuged, and lyophilized to obtain the polyrotaxane as a powder. The threading number of αCD was 1 The carbon black was determined by H-NMR (frequency: 500 MHz, solvent: DMSO-d6).

[0108] [Step 4] Polyrotaxane (8.41 g, 0.108 mmol) was dissolved in anhydrous DMSO (250 mL), CDI (8.29 g, 51.1 mmol) was added, and the reaction mixture was stirred at 23°C. After one day, the reaction mixture was added dropwise to 1,2-bis(2-aminoethoxyethane) (76 mL, 0.51 mol) and stirred at 23°C for another day. The reaction mixture was then purified by dialysis for five days. The product was lyophilized to obtain the polyrotaxane of Preparation Example 4 as a powder.

[0109] The data for the polyrotaxane of Production Example 4 are shown in Table 2. The number of side chains introduced per CD is: 1 Calculated based on H-NMR analysis. [Table 2]

[0110] 2. Evaluation of water solubility of polyrotaxane 〔method〕 As shown in Table 3, a predetermined amount of polyrotaxane powder from Production Example 4 was weighed and placed in a glass bottle. MilliQ water was then added to the bottle to achieve various concentrations (80, 160, or 320 mg / mL), and the mixture was stirred at 23°C for 1 hour to prepare aqueous polyrotaxane solutions. Similarly, aqueous polyrotaxane solutions from Production Example 3 (concentration: 40 or 80 mg / mL) were prepared. The light transmittance (%T) of the polyrotaxane aqueous solutions 1 to 6 was measured using a spectrophotometer (Hitachi High-Tech Science Corporation, product name: U-2910). [Table 3]

[0111] 〔result〕 Each polyrotaxane aqueous solution was placed in an optical cell, and its appearance was photographed. The photograph is shown in Figure 3. The polyrotaxane of Production Example 1 (unmodified) was insoluble in water, so its transmittance was not measured.

[0112] The results of the light transmittance at a wavelength of 600 nm are shown in Table 4. The polyrotaxane aqueous solution of Production Example 4 was transparent even at a concentration of 320 mg / mL, demonstrating excellent water solubility. On the other hand, the polyrotaxane aqueous solution of Production Example 3 was cloudy even at a concentration of 40 mg / mL, indicating that its water solubility was not very high. The polyrotaxane aqueous solution of Production Example 3 did not become transparent even when heated to 50°C. The total transmittance spectrum in the visible light region (wavelengths of 380 to 780 nm) is shown in Figure 4. The polyrotaxane of Production Example 4 exhibited high light transmittance over the entire measured wavelength range. [Table 4]

[0113] 3. Evaluation of gelation time of polyrotaxane derivatives [Gellation] A predetermined amount of polyrotaxane powder from Production Example 4 was weighed and placed in a glass bottle. MilliQ water was added to various concentrations (80 mg / mL, 160 mg / mL, 320 mg / mL) and stirred at 23°C for 1 hour to prepare polyrotaxane aqueous solutions. The crosslinker EGDE was added to the resulting polyrotaxane aqueous solution, and the solution was stirred using a vortex mixer for 20 seconds. The solution was then stirred at 37°C using a stirrer (rotation speed: 320 rpm). 1.4 or 3.5 molar equivalents of crosslinker were added relative to the number of cyclodextrins (CD) contained in the polyrotaxane used. The time from the addition of the crosslinker until the stirring bar stopped rotating was measured and recorded as the gelation time. The gelation time was also evaluated for the polyrotaxane from Production Example 3 in the same manner.

[0114] 〔result〕 The results are shown in Table 5 and Figure 5. The gelation time decreased as the polyrotaxane concentration increased. It was found that the polyrotaxane of Production Example 4 could be gelled within 5 minutes at 37°C when the polyrotaxane concentration was 320 mg / mL. At the same concentration, when 3.5 equivalents of EGDE were added to the polyrotaxane, a slightly cloudy hydrogel was obtained. The inventors believe that this is because the EGDE used as a crosslinker is poorly soluble in water. On the other hand, when the polyrotaxane of Production Example 3 was used, the resulting hydrogel was cloudy even at a concentration of 80 mg / mL. [Table 5]

[0115] (2) Evaluation of cell adhesion of hydrogels A hydrogel was prepared by coating a polystyrene dish for cell culture with a mixed solution of the polyrotaxane aqueous solution and crosslinker prepared in Preparation Example 2A. Mouse fibroblasts were seeded on either an uncoated polystyrene dish or on the hydrogel-coated dish, and cell adhesion was evaluated by taking photographs of the cells once a day for one week.

[0116] The results are shown in Figure 6. Figure 6 is a photograph of mouse fibroblasts seeded on a polystyrene dish. On a regular polystyrene dish (uncoated, Figure 6(a)), the fibroblasts adhered and spread, and proliferation was observed after one week. On the other hand, on the coated polystyrene dish (Figure 6(b)), although the fibroblasts adhered, little spreading was observed, and even after one week, almost no spreading cells were observed, and no significant proliferation was observed. These results demonstrate that the hydrogel coating allows fibroblasts to adhere alive, while preventing their activation and proliferation.

[0117] 3. Evaluation of hydrogel tissue adhesion [Preparation of Hydrogel] A crosslinker was added to a 100 mg / mL aqueous solution of the polyrotaxane (Production Example 2A) in an amount of 3.5 molar equivalents relative to the amount of αCD contained in the polyrotaxane, and the mixture was stirred. The reaction solution was quickly poured into a mold and allowed to stand at 23°C to obtain a hydrogel. After one day, the hydrogel was removed from the mold and washed with MilliQ water.

[0118] [Tissue Adhesion Test] The tissue samples used were pig cecum, uterus, and small intestine (purchased from Tokyo Shibaura Organ Co., Ltd.). Hydrogel was attached to each tissue sample so that the hydrogel was sandwiched between the tissue samples. The adhesion and appearance of the hydrogel were observed immediately after attachment, one hour later, and one day later. For comparison, Seprafilm (registered trademark, manufactured by Sanofi K.K.) was used instead of the hydrogel.

[0119] 〔result〕 The hydrogel adhered quickly after application and remained adhered even after 1 hour and 1 day. Seprafilm adhered strongly immediately after application, but quickly changed from a sheet to a gel, and after 1 hour the material became brittle and its adhesiveness decreased.

[0120] 4. Hydrogel Transparency Evaluation 〔method〕 A predetermined amount of polyrotaxane powder from Production Example 4 was weighed and placed in a glass bottle. MilliQ water was then added to achieve various concentrations (80, 160, or 320 mg / mL). The mixture was stirred at 23°C for 1 hour to prepare aqueous polyrotaxane solutions. The aqueous polyrotaxane solution from Production Example 4 and a crosslinker (EGDE or BDDE) were mixed in a 2.0 mL tube, and 0.8 mL of the reaction solution was transferred to an optical cell and allowed to stand at 23°C for 1 day (gelation). MilliQ water was added to the reaction solution to wash the gel, and MilliQ water was added and the mixture was allowed to stand at 23°C for another 1 day.

[0121] Similarly, a predetermined amount of polyrotaxane powder from Production Example 3 was weighed and placed in a glass bottle. MilliQ water was then added to achieve various concentrations (80 mg / mL). The mixture was stirred at 23°C for 1 hour, and then immersed in a 50°C water bath for 20 minutes to prepare aqueous polyrotaxane solutions. The aqueous polyrotaxane solution from Production Example 3 and the crosslinker EGDE were mixed in a 2.0 mL tube, and 0.8 mL of the reaction solution was transferred to an optical cell and allowed to stand at 23°C for 1 day (gelation). MilliQ water was added to the reaction solution to wash the gel, and MilliQ water was added and the mixture was allowed to stand at 23°C for another 1 day.

[0122] Photographs of the appearance of each hydrogel were taken. The light transmittance (%T) of the obtained hydrogel was measured using a spectrophotometer (Hitachi High-Tech Science Corporation, product name: U-2910). The light transmittance at a wavelength of 600 nm and the total transmittance in the visible light range (380-780 nm) of each hydrogel were compared.

[0123] 〔result〕 Photographs of the appearance of each hydrogel are shown in Figures 7 and 8. When the polyrotaxane of Production Example 4 was used, a transparent hydrogel was obtained. In Examples 9 and 10, the hydrogel was highly viscous and contained air bubbles, but the hydrogel itself was transparent. On the other hand, when the polyrotaxane of Production Example 3 was used, the hydrogel was cloudy even at a low concentration.

[0124] The results of the light transmittance of each hydrogel at a wavelength of 600 nm or in the visible light region are shown in Tables 6 and 7. The total transmittance spectrum in the visible light region (wavelengths of 380 to 780 nm) is shown in Figure 9. The hydrogel derived from the polyrotaxane of Production Example 4 exhibited high transmittance even at a high concentration (320 mg / ml), but the hydrogel derived from the polyrotaxane of Production Example 3 was cloudy and had low transmittance even at 80 mg / ml. [Table 6] [Table 7]

[0125] 5. Evaluation of the adhesion prevention properties of hydrogels [Preparation of hydrogel] The crosslinker EGDE was added to the polyrotaxane aqueous solution (concentration 160 mg / mL) of Preparation Example 2B in an amount of 1.4 molar equivalents of αCD contained in the polyrotaxane, and the mixture was stirred. The reaction solution was quickly poured into a silicone rubber mold (volume: approximately 0.2 mL), filled with the mixture, covered with film to prevent bubbles, and allowed to stand at 23°C. After one day, the hydrogel was removed from the mold and washed with MilliQ water to obtain a hydrogel measuring 20 mm x 20 mm and 0.5 mm thick.

[0126] [Creating a mouse] The anti-adhesion function was evaluated using a mouse cecal abrasion adhesion model in accordance with the procedures approved by the Animal Care and Use Committee of Tokyo Medical and Dental University. Mice underwent laparotomy, and the cecum was abraded with a commercially available toothbrush. The hydrogel was then placed and the incision was sutured (n = 4). For comparison, a sham group (mice underwent laparotomy and the incision was sutured without abrasion), a control group (mice underwent laparotomy and the cecum was abraded with a toothbrush and the incision was sutured), and a Seprafilm group (mice underwent laparotomy and the cecum was abraded with a toothbrush, Seprafilm was placed, and the incision was sutured) were prepared (n = 4 per group). The anti-adhesion function was evaluated by comparing the degree of adhesion one week after suturing. Seprafilm was cut into 20 mm x 20 mm pieces, similar to the hydrogel.

[0127] 〔evaluation〕 For each treatment group, the severity score and area score of adhesions were evaluated on a five-point scale, with reference to the description in PLoS One. 2019; 14(1): e0211391. The average score was recorded. That is, the evaluation criteria are as follows:

[0128] <Criteria for assessing the degree of adhesion> 0: No adhesion 1: Mild adhesion 2: Moderate focal adhesions 3: Moderate and extensive adhesions 4: Severe adhesion that cannot be separated <Evaluation criteria for adhesion area> 0: No adhesion 1: The adhesion area is in the range of 1-24% of the abraded area 2: The adhesion area is in the range of 25-49% of the abraded area 3: The adhesion area is within 50-74% of the abraded area 4: The adhesion area is within 75-100% of the abraded area.

[0129] 〔result〕 The degree of adhesion was evaluated using a score, and the results are shown in Table 8. In the untreated group, all mice had moderate and extensive adhesions. In the Seprafilm-treated group, all mice had mild adhesions, and Seprafilm had disappeared. The hydrogel was confirmed to have an adhesion prevention function, and its function is thought to be equal to or slightly superior to Seprafilm. [Table 8]

[0130] 6. In situ hydrogelation A polyrotaxane solution and an aqueous dispersion of the crosslinker EGDE were filled into a dual syringe with a volume ratio of 1:1. Both solutions were mixed in a static mixer and applied to a glass slide to observe the fluidity of the mixed solution.

[0131] Thirty minutes after application, the mixed solution gelled and completely lost its fluidity. This result showed that mixing polyrotaxane with a crosslinker gelled in a relatively short time at room temperature in water without the need for a catalyst.

Claims

1. A polyrotaxane having a chemical structure in which an axis molecule represented by formula (2) passes through a cyclodextrin ring of a compound represented by formula (1). 【Chemistry 1】 [wherein CD represents a cyclodextrin ring, L ２ represents a divalent organic group, and L ３ represents an ethylene group or a propylene group, and R Ａ is a hydrogen atom or C １－６ represents an alkyl group, and m represents an integer of 0 to 35. ２ When R is connected to N through a double bond, Ａ does not exist.] 【Chemistry 2】 [In the formula, R represents a hydrocarbon group larger than the inner diameter of the cyclodextrin ring, X represents a divalent organic group, and a represents an integer of 100 to 1000.]

2. A polyrotaxane comprising a chemical structure in which an axis molecule represented by formula (2) passes through a cyclodextrin ring of a compound represented by formula (1'). 【Transformation 3】 [wherein CD represents a cyclodextrin ring, L ２ represents a divalent organic group, and L ３ represents an ethylene group or a propylene group, and R Ａ is a hydrogen atom or C １－６ represents an alkyl group, and m represents an integer of 0 to 35. 【Chemistry 4】 [In the formula, R represents a hydrocarbon group larger than the inner diameter of the cyclodextrin ring, X represents a divalent organic group, and a represents an integer of 100 to 1000.]

3. The polyrotaxane according to claim 1 or 2, wherein the axis molecule further passes through the cyclodextrin ring of the compound represented by formula (3). 【Transformation 5】 [wherein CD represents a cyclodextrin ring, L ３ represents an ethylene group or a propylene group, and R Ａ is a hydrogen atom or C １－６ represents an alkyl group, and m represents an integer of 0 to 35.

4. 3. The polyrotaxane according to claim 1, wherein the axis molecule is a compound represented by formula (2a) or formula (2b). 【Transformation 6】 [In the formula, a represents an integer of 100 to 1000.] 【Transformation 7】 [In the formula, a represents an integer of 100 to 1000.]

5. 3. The polyrotaxane according to claim 1, wherein the compound represented by formula (1) or formula (1′) is a compound represented by formula (1a-1), (1a-2) or (1a-3). 【Transformation 8】 [wherein CD represents a cyclodextrin ring, L １ represents a divalent organic group, and L ３ represents an ethylene group or a propylene group, and R Ａ is a hydrogen atom or C １－６ represents an alkyl group, and m represents an integer of 0 to 35.

6. 3. The polyrotaxane according to claim 1, which has a transmittance of 40% or more for light at a wavelength of 600 nm.

7. A medical composition comprising the polyrotaxane according to claim 1 or 2.

8. The medical composition according to claim 7, which is in the form of a hydrogel.

9. The medical composition according to claim 7, which is a cell culture material, a tissue adhesive, a tissue regeneration substrate, or an adhesion barrier.

10. The medical composition according to claim 8, which is a cell culture material, a tissue adhesive, a tissue regeneration substrate, or an adhesion barrier.

11. A method for producing a polyrotaxane having a chemical structure in which an axis molecule represented by formula (2) penetrates a cyclodextrin ring of a compound represented by formula (1), comprising: 【Chemistry 9】 [wherein CD represents a cyclodextrin ring, L ２ represents a divalent organic group, and L ３ represents an ethylene group or a propylene group, and R Ａ is a hydrogen atom or C １－６ represents an alkyl group, and m represents an integer of 0 to 35. ２ When R is connected to N through a double bond, Ａ does not exist.] 【Chemistry 10】 [In the formula, R represents a hydrocarbon group larger than the inner diameter of the cyclodextrin ring, X represents a divalent organic group, and a represents an integer of 100 to 1000.] a polyrotaxane (also referred to as polyrotaxane A) having a chemical structure in which an axis molecule represented by formula (2) passes through each cyclodextrin ring of one or more compounds represented by formula (3); 【Chemistry 11】 [wherein CD represents a cyclodextrin ring, L ３ represents an ethylene group or a propylene group, and R Ａ is a hydrogen atom or C １－６ represents an alkyl group, and m represents an integer of 0 to 35. 【Chemistry 12】 [In the formula, R represents a hydrocarbon group larger than the inner diameter of the cyclodextrin ring, X represents a divalent organic group, and a represents an integer of 100 to 1000.] a step of mixing a crosslinking agent represented by formula (4) 【Chemistry 13】 [In the formula, L １ represents a divalent organic group, and X １ represents a group capable of reacting with an amino group. A manufacturing method comprising:

12. A method for producing a polyrotaxane having a chemical structure in which an axis molecule represented by formula (2) penetrates a cyclodextrin ring of a compound represented by formula (1′), comprising: 【Chemistry 14】 [wherein CD represents a cyclodextrin ring, L ２ represents a divalent organic group, and L ３ represents an ethylene group or a propylene group, and R Ａ is a hydrogen atom or C １－６ represents an alkyl group, and m represents an integer of 0 to 35. 【Chemistry 15】 [In the formula, R represents a hydrocarbon group larger than the inner diameter of the cyclodextrin ring, X represents a divalent organic group, and a represents an integer of 100 to 1000.] a polyrotaxane (also referred to as polyrotaxane A) having a chemical structure in which an axis molecule represented by formula (2) passes through each cyclodextrin ring of one or more compounds represented by formula (3); 【Chemistry 16】 [wherein CD represents a cyclodextrin ring, L ３ represents an ethylene group or a propylene group, and R Ａ is a hydrogen atom or C １－６ represents an alkyl group, and m represents an integer of 0 to 35. 【Chemistry 17】 [In the formula, R represents a hydrocarbon group larger than the inner diameter of the cyclodextrin ring, X represents a divalent organic group, and a represents an integer of 100 to 1000.] a step of mixing a crosslinking agent represented by formula (4) [Chemistry 18] [In the formula, L １ represents a divalent organic group, and X １ represents a group capable of reacting with an amino group. A manufacturing method comprising:

13. 13. The method according to claim 11 or 12, wherein in a polyrotaxane having a chemical structure in which an axis molecule represented by formula (2) passes through a cyclodextrin ring of a compound represented by formula (1) or formula (1'), the axis molecule further passes through a cyclodextrin ring of a compound represented by formula (3). 【Chemistry 19】 [wherein CD represents a cyclodextrin ring, L ３ represents an ethylene group or a propylene group, and R Ａ is a hydrogen atom or C １－６ represents an alkyl group, and m represents an integer of 0 to 35.

14. The method according to claim 11 or 12, wherein the step of mixing the polyrotaxane A with the crosslinking agent represented by formula (4) is carried out under catalyst-free conditions.

15. a polyrotaxane having a chemical structure in which an axis molecule represented by formula (2) penetrates through each cyclodextrin ring of one or more compounds represented by formula (3); 【Chemistry 20】 [wherein CD represents a cyclodextrin ring, L ３ represents an ethylene group or a propylene group, and R Ａ is a hydrogen atom or C １－６ represents an alkyl group, and m represents an integer of 0 to 35. 【Chemistry 21】 [In the formula, R represents a hydrocarbon group larger than the inner diameter of the cyclodextrin ring, X represents a divalent organic group, and a represents an integer of 100 to 1000.] A kit comprising: a crosslinking agent represented by formula (4): 【Chemistry 22】 [In the formula, L １ represents a divalent organic group, and X １ represents a group capable of reacting with an amino group.

16. The method for producing a polyrotaxane according to claim 11 or 12, wherein the compound represented by formula (1) or formula (1') is a compound represented by formula (1a-1), (1a-2) or (1a-3). 【Chemistry 23】 [wherein CD represents a cyclodextrin ring, L １ represents a divalent organic group, and L ３ represents an ethylene group or a propylene group, and R Ａ is a hydrogen atom or C １－６ represents an alkyl group, and m represents an integer of 0 to 35.

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