Polymer Material Kit for Hemostasis

Abstract

The present invention aims to provide a safe and efficient method for hemostasis, vascular occlusion, tissue coating, or body fluid coagulation using a polymeric material that is a synthetic compound of non-biological origin. A solution (pre-gel solution) containing a hydrophilic polymer that can form a hydrogel through intermolecular cross-linking under specific concentration conditions and having specific pH conditions and ionic strength is prepared, and is caused to gel in situ in an environment where blood exists, such as at the bleeding site and within blood vessels.

Description

Technical Field

[0001] The present invention relates to a polymer material kit for hemostasis, vascular occlusion, tissue covering, or body fluid coagulation, and also relates to a hemostasis method, vascular occlusion method, tissue covering method, or body fluid coagulation method using the polymer material kit. Background Art

[0002] As conventional blood coagulation means, mainly a method of accelerating the hemostasis reaction by utilizing the pharmacological action of blood coagulation factors of biological origin typified by fibrin glue (for example, Patent Document 1), or a method of physically blocking blood flow and causing blood to coagulate by the blood coagulation ability inherent in the blood itself is adopted.

[0003] However, in the former method using biological source materials, it is difficult to control the infection of animals or providers as raw materials, so there is an infection risk. In fact, problems such as drug-induced AIDS incidents and iatrogenic Creutzfeldt-Jakob disease have occurred. On the other hand, in the latter method of physically blocking, due to the characteristics of the hemostasis principle, a sheet-like material needs to be used, and it is difficult to apply to affected parts with complex structures. In addition, as a common problem of both, the following points can also be cited: Since the final hemostasis state depends on the blood coagulation ability of the patient, it is difficult to obtain a blood coagulation effect for patients who have been given drugs such as anticoagulants.

[0004] Prior Art Documents

[0005] Patent Documents

[0006] Patent Document 1: Japanese Patent Laid-Open No. 2017-66150 Summary of the Invention

[0007] Technical Problem to be Solved by the Invention

[0008] Therefore, the present invention aims to provide a safe and efficient hemostasis method, vascular occlusion method, tissue covering method, or body fluid coagulation method using a polymer material as a non-biological source synthetic substance.

[0009] Technical Solution for Solving the Technical Problem

[0010] The inventors of the present invention conducted intensive studies to solve the above problems and found that by preparing a solution (pre-gel solution) containing a hydrophilic polymer that can form a hydrogel through intermolecular cross-linking at a specific concentration condition and having a specific pH condition and ionic strength, and causing it to gel in situ in an environment where blood exists such as at the bleeding site and in blood vessels, a good blood coagulation effect and hemostasis effect can be obtained, thus completing the present invention.

[0011] That is, in one aspect, the present invention relates to a kit for forming a hydrogel in an environment where blood is present, and specifically provides the following inventions.

[0012] <1> A polymer material kit formed from a polymer solution A containing a first polymer and a second polymer solution B containing a second polymer, wherein the first polymer and the second polymer are a combination of hydrophilic polymers having a polyalkylene glycol backbone or a polyethylene backbone that can form a hydrogel by crosslinking with each other, and the first polymer and the second polymer have a weight average molecular weight (Mw) in the range of 1×10 3 ~1×10 5 . The concentrations of the first polymer and the second polymer in the polymer solutions A and B are in the range of 10 to 300 g / L. The pH of the mixture obtained by mixing the polymer A and B is 3 or more and less than 7, and the ionic strength is in the range of 10 to 100 mM.

[0013] <2> The polymer material kit according to <1> above, wherein the first polymer and the second polymer are di-branched, tri-branched or tetra-branched polyethylene glycols.

[0014] <3> The polymer material kit according to <1> or <2> above, wherein the first polymer has one or more nucleophilic functional groups in the side chain or at the end, and the second polymer has one or more electron-withdrawing functional groups in the side chain or at the end.

[0015] <4> The polymer material kit according to any one of <1> to <3> above, wherein the nucleophilic functional group is selected from a mercapto group and an amino group, and the electron-withdrawing functional group is selected from a maleimido group, N-hydroxysuccinimide (NHS), sulfosuccinimide, phthalimido group, imidazolyl group, acryloyl group, nitrophenyl group, and -CO 2 PhNO 2 .

[0016] <5> The polymer material kit according to any one of <1> to <4> above, wherein the pH of both the polymer solutions A and B is in the range of 3 or more and less than 7.

[0017] <6> The polymer material kit according to any one of <1> to <5> above, wherein in an environment where a liquid having a pH of 6.5 to 8.0 is present, the polymer solution A and the polymer solution B form a mixed state, and a hydrogel obtained by crosslinking the first polymer and the second polymer with each other is formed.

[0018] <7> The polymer material kit according to <6> above, wherein the gelation time of the hydrogel formed by the first polymer and the second polymer is in the range of 1 to 30 seconds.

[0019] <8> The polymer material kit according to <6> or <7> above, wherein the hydrogel formed by the first polymer and the second polymer has an equilibrium swelling degree in the range of 0.9 to 3.5.

[0020] <9> The polymer material kit according to any one of <6> to <8> above, wherein the hydrogel formed by the first polymer and the second polymer has a Young's modulus in the range of 0.1×10 4 ~4×10 4 Pa.

[0021] <10> The polymer material kit according to any one of <1> to <9> above, wherein the kit is used for hemostasis, vascular occlusion, tissue coating, or body fluid coagulation.

[0022] <11> A hemostatic agent, which is formed from the polymer material kit according to any one of <1> to <10> above.

[0023] <12> A vascular occluding agent, which is formed from the polymer material kit according to any one of <1> to <10> above.

[0024] <13> A tissue coating agent, which is formed from the polymer material kit according to any one of <1> to <10> above.

[0025] <14> A body fluid coagulant, which is formed from the polymer material kit according to any one of <1> to <10> above.

[0026] In another aspect, the present invention relates to a method for manufacturing a hemostatic agent or the like containing a hydrogel using the above kit, and specifically provides the following inventions.

[0027] <15> A method for manufacturing a hemostatic agent, a vascular occluding agent, a tissue coating agent, or a body fluid coagulant containing a hydrogel, which includes a step of applying a mixture of a polymer solution A containing a first polymer and a second polymer solution B containing a second polymer to an environment where there is a liquid with a pH of 6.5 to 8.0. The first polymer and the second polymer are a combination of hydrophilic polymers having a polyalkylene glycol skeleton or a polyethylene skeleton that can form a hydrogel by crosslinking with each other, and the first polymer and the second polymer have 1×10 3 ~1×10 5The weight-average molecular weight (Mw) is within the range of, the concentrations of the first polymer and the second polymer in the polymer solutions A and B are within the range of 10 to 300 g / L, the pH of the mixed solution obtained by mixing the polymer A and B is 3 or more and less than 7, and the ionic strength is within the range of 10 to 100 mM.

[0028] <16>According to the method described in the above <15>, wherein, it includes a step of mixing the polymer solution A and the polymer solution B in an environment where there is a liquid with a pH of 6.5 to 8.0.

[0029] <17>According to the method described in the above <15>, wherein, it includes a step of, after dropping the polymer solution A and the polymer solution B onto a carrier, bringing the carrier into contact with an environment where there is a liquid with a pH of 6.5 to 8.0.

[0030] <18>According to the method described in any one of <15> to <17>, wherein, the pH of both the polymer solutions A and B is within the range of 3 or more and less than 7.

[0031] In still another aspect, the present invention relates to a hemostasis method using the above kit, etc., and specifically provides the following inventions.

[0032] <19>A hemostasis method, wherein, the polymer material kit described in any one of the above <1> to <10> is used.

[0033] <20>A method for occluding blood vessels, wherein, the polymer material kit described in any one of the above <1> to <10> is used.

[0034] <21>A method for covering tissues, wherein, the polymer material kit described in any one of the above <1> to <10> is used.

[0035] <22>A method for coagulating body fluids, wherein, the polymer material kit described in any one of the above <1> to <10> is used.

[0036] <23>A medical device, which is formed from the polymer material kit described in any one of the above <1> to <10>.

[0037] <24>According to the medical device described in the above <23>, wherein, at least one of the polymer solution A and the polymer solution B is stored in a nebulizer.

[0038] Effects of the invention

[0039] If the polymer material kit of the present invention is used, by applying two polymer solutions to an environment where blood exists, such as a bleeding site and inside blood vessels, the pH of the polymer solutions changes and an in-situ gelation reaction occurs, and a gel-blood complex incorporating blood can be formed in a short time. Thus, while having a good blood coagulation effect by incorporating blood into the gel, it can be provided in a case where it also has a hemostatic effect in a physical manner of covering a bleeding site or the like with the gel.

[0040] The polymer material used in the present invention is not a material derived from animals as in the prior art, so the risk of infection and the like can be avoided. In addition, in the present invention, after the two polymer solutions are mixed, they can maintain a liquid state for a certain period of time, so it has the advantage of being applicable to tissues with complex structures. In addition, different from the conventional methods, it can provide good blood coagulation ability without relying on the coagulation ability of blood itself, so it also has the advantage of being applicable to patients given anticoagulants and the like.

[0041] Regarding its application target, the in-situ gel formation of the present invention can also be applied to blood vessels such as veins and arteries for vascular occlusion. In addition, since it is a gel formation mechanism that utilizes the pH change generated by applying to a living body environment, it can exert a coagulation effect not only on blood but also on body fluids having a pH near neutrality.

[0042] Brief Description of the Drawings

[0043] Figure 1 It is a graph showing the influence of concentration and pH on the gelation time.

[0044] Figure 2 It is a graph showing the change in gelation time based on the ionic strength.

[0045] Figure 3 It is a graph showing the change in pH based on the ionic strength.

[0046] Figure 4 It is a graph showing the relationship between the gelation time and pH.

[0047] Figure 5 It is a graph showing the change in Young's modulus based on the mixing with milk.

[0048] Figure 6 It is a graph showing the change over time in the degree of swelling in milk.

[0049] Figure 7 It is a graph showing the equilibrium swelling degree at each prepolymer concentration in water.

[0050] Figure 8 It is an image of applying the gel to the venous blood vessel of a rat's thigh.

[0051] Figure 9 It is an image of applying a gel to the arterial blood vessels of a rat's abdomen. Detailed implementation mode

[0052] Hereinafter, embodiments of the present invention will be described. The scope of the present invention is not limited to these descriptions, and for those other than the following examples, it can also be implemented with appropriate modifications within the scope not hindering the gist of the present invention.

[0053] 1. Polymer material kit of the present invention

[0054] The polymer material kit of the present invention is characterized in that it is formed by a polymer solution A containing a first polymer and a second polymer solution B containing a second polymer. The polymer solution contains a hydrophilic polymer that can form a hydrogel through intermolecular crosslinking under specific concentration conditions, and has specific pH conditions and ionic strength.

[0055] By adopting the above solution conditions, simply mixing these polymer solutions A and B directly cannot cause the gelation reaction to proceed in a short time. However, by mixing the two solutions in a solution with a pH near neutral such as blood, the gelation is promoted, and in-situ gelation occurs in a shorter time. At the same time, a hydrogel that incorporates the blood into the gel interior can be formed (also referred to as a "gel-blood complex" or a "gel-body fluid complex"). Through the formation of the in-situ gel-blood complex, it is possible to provide both a coagulation effect based on incorporating the blood itself into the gel and a hemostatic effect in a physical manner such as coating the bleeding site with the gel. In this regard, it can be said to be an unprecedented new method.

[0056] Hereinafter, the polymer materials and solution conditions used in the polymer material kit of the present invention will be described in detail.

[0057] (1-1) Polymer materials

[0058] Both the first polymer and the second polymer used in the polymer solutions A and B of the present invention are hydrophilic polymers having a polyalkylene glycol backbone or a polyethylene backbone that can form a hydrogel through crosslinking with each other. For this hydrophilic polymer, as long as it can form a hydrogel through a gelation reaction (such as a crosslinking reaction) in an aqueous solution, polymers known in the art can be used. More specifically, it is preferably a polymer that can form a mesh structure, especially a three-dimensional mesh structure, through crosslinking of the polymers in the final gel.

[0059] As a polymer having a polyethylene glycol backbone, a polymer having a plurality of polyethylene glycol backbones, particularly preferably a di-branched, tri-branched or tetra-branched polyethylene glycol, can be exemplified. In particular, a gel formed of a tetra-branched polyethylene glycol backbone is generally known as a tetra-branched PEG (Tetra-PEG) gel, and a mesh structure network is constructed by an AB-type cross-end-coupling reaction between two tetra-branched polymers having an electron-withdrawing functional group such as an active ester structure and a nucleophilic functional group such as an amino group at the ends, respectively (Matsunaga et al., Macromolecules, Vol. 42, No. 4, pp. 1344-1351, 2009). In addition, the tetra-branched PEG gel can be prepared on the spot by simply mixing two polymer solutions, and the gelation time can also be controlled by adjusting the pH and ionic strength during gel preparation. And since the gel is mainly composed of PEG, its biocompatibility is also good.

[0060] In addition, as hydrophilic polymers having a polyethylene backbone, polyalkyl methacrylates such as polymethyl methacrylate, polyacrylates, polyvinyl alcohol, poly-N-alkylacrylamide, polyacrylamide, etc. can be exemplified.

[0061] The first polymer and the second polymer have a weight average molecular weight (Mw) in the range of 1×10 3 ~1×10 5 , preferably in the range of 0.5×10 4 ~5×10 4 , more preferably in the range of 1×10 4 ~2×10 4 .

[0062] Preferably, the first polymer and the second polymer are a combination of a polymer having one or more nucleophilic functional groups in the side chain or at the end and a polymer having one or more electron-withdrawing functional groups in the side chain or at the end. For example, preferably, the first polymer has one or more nucleophilic functional groups in the side chain or at the end, and the second polymer has one or more electron-withdrawing functional groups in the side chain or at the end. A gel is formed by crosslinking the nucleophilic functional group and the electron-withdrawing functional group. Here, the total of the nucleophilic functional group and the electron-withdrawing functional group is preferably 5 or more. These functional groups are more preferably present at the ends.

[0063] As the nucleophilic functional groups present in the first and second polymers, mercapto group (-SH), amino group, etc. can be exemplified, and as long as those skilled in the art, known nucleophilic functional groups can be appropriately used. Preferably, the nucleophilic functional group is -SH group. The nucleophilic functional groups can be the same or different, preferably the same. By having the same functional groups, the reactivity with the electron-withdrawing functional groups forming the crosslinking bond is uniform, and it is easy to obtain a gel having a uniform three-dimensional structure.

[0064] As the electron-withdrawing functional group present in the first and second polymers, an active ester group can be used. As such an active ester group, examples include maleimido, N-hydroxysuccinimido (NHS), sulfosuccinimido, phthalimido, imidazolyl, acryloyl, nitrophenyl, -CO 2 PhNO 2 (wherein Ph represents ortho, meta or para phenylene), etc. As long as those skilled in the art can appropriately use other known active ester groups. Preferably, the electron-withdrawing functional group is maleimido. The electron-withdrawing functional groups can be the same or different, preferably the same. By having the same functional groups, the reactivity with the nucleophilic functional groups forming crosslinked bonds is uniform, and it is easy to obtain a gel having a uniform three-dimensional structure.

[0065] Preferred non-limiting specific examples of the polymer having a nucleophilic functional group at the terminal include, for example, a compound represented by the following formula (I) having 4 branches of polyethylene glycol skeletons and a mercapto group at the terminal.

[0066] [Chemical formula 1]

[0067]

[0068] In formula (I), R 11 ~R 14 are the same or different and represent C 1 -C 7 alkylene, C 2 -C 7 alkenylene, -NH-R 15 -, -CO-R 15 -, -R 16 -O-R 17 -, -R 16 -NH-R 17 -, -R 16 -CO 2 -R 17 -, -R 16 -CO 2 -NH-R 17 -, -R 16 -CO-R 17 -, or -R 16 -CO-NH-R 17 (wherein R 15 represents C 1 -C 7 alkylene, R 16 represents C 1 -C 3 alkylene, R 17 represents C 1 -C 5Alkylene group.

[0069] n 11 ~n 14 May be the same or different respectively. n 11 ~n 14 The closer the values of n~n are, the more uniform the three-dimensional structure can be formed and the higher the strength. Therefore, in order to obtain a gel with high strength, it is preferably the same. If the value of n 11 ~n 14 is too high, the strength of the gel becomes weak. If the value of n 11 ~n 14 is too low, it is difficult to form a gel due to the steric hindrance of the compound. Therefore, n 11 ~n 14 Can be exemplified by integer values from 5 to 600, preferably 25 to 250, more preferably 50 to 120, and even more preferably 110 to 120.

[0070] In the above formula (I), R 11 ~R 14 Is the connecting group part connecting the functional group and the core part. R 11 ~R 14 May be the same or different respectively. In order to manufacture a high-strength gel with a uniform three-dimensional structure, it is preferably the same. R 11 ~R 14 Represents C 1 -C 7 Alkylene group, C 2 -C 7 Alkenylene group, -NH-R 15 -, -CO-R 15 -, -R 16 -O-R 17 -, -R 16 -NH-R 17 -, -R 16 -CO 2 -R 17 -, -R 16 -CO 2 -NH-R 17 -, -R 16 -CO-R 17 -, or -R 16 -CO-NH-R 17 -. Here, R 15 Represents C 1 -C 7 Alkylene group. R 16 Represents C 1 -C 3 Alkylene group. R 17 Represents C 1 -C 5 Alkylene group.

[0071] Herein, "C 1 -C 7 alkylene" means an alkylene having 1 or more and 7 or less carbon atoms which may have branches, i.e., a straight-chain C 1 -C 7 alkylene or a C 2 -C 7 alkylene having 1 or 2 or more branches (including those having 2 or more and 7 or less carbon atoms in the branch). C 1 -C 7 Examples of alkylene are methylene, ethylene, propylene, and butylene. C 1 -C 7 Examples of alkylene may include -CH 2 -, -(CH 2 ) 2 -, -(CH 2 ) 3 -, -CH(CH 3 )-, -(CH 2 ) 3 -, -(CH(CH 3 )) 2 -, -(CH 2 ) 2 -CH(CH 3 )-, -(CH 2 ) 3 -CH(CH 3 )-, -(CH 2 ) 2 -CH(C 2 H 5 )-, -(CH 2 ) 6 -, -(CH 2 ) 2 -C(C 2 H 5 ) 2 -, and -(CH 2 ) 3 C(CH 3 ) 2 CH 2 - etc.

[0072] "C 2 -C 7 alkenylene" means an alkenylene having 1 or 2 or more double bonds in the chain or a branched alkenylene having 2 to 7 carbon atoms, and examples thereof may include a divalent group having a double bond formed by removing 2 to 5 hydrogen atoms from adjacent carbon atoms of the alkylene.

[0073] On the other hand, non-limiting specific examples of preferred polymers having an electron-withdrawing functional group at the terminal include, for example, a compound represented by the following formula (II) having 4 branches of polyethylene glycol skeletons and an N-hydroxysuccinimide (NHS) at the terminal.

[0074] [Chemical formula 2]

[0075]

[0076] In the above formula (II), n 21 ~n 24 may be the same or different, respectively. The closer the values of n 21 ~n 24 are, the more uniform three-dimensional structure the gel can form and the higher the strength. Therefore, it is preferred that they are the same. If the values of n 21 ~n 24 are too high, the strength of the gel becomes weak. If the values of n 21 ~n 24 are too low, it is difficult to form a gel due to the steric hindrance of the compound. Therefore, n 21 ~n 24 can be exemplified by integer values of 5 to 600, preferably 25 to 250, more preferably 50 to 120, and even more preferably 110 to 120.

[0077] In the above formula (II), R 21 ~R 24 is the connecting group part connecting the functional group and the core part. R 21 ~R 24 may be the same or different, respectively. In order to produce a high-strength gel with a uniform three-dimensional structure, it is preferably the same. In formula (II), R 21 ~R 24 represents C 1 -C 7 alkylene, C 2 -C 7 alkenylene, -NH-R 25 -, -CO-R 25 -, -R 26 -O-R 27 -, -R 26 -NH-R 27 -, -R 26 -CO 2 -R 27 -, -R 26 -CO 2 -NH-R 27 -, -R 26 -CO-R 27 -, or -R 26 -CO-NH-R 27-, where R 25 represents C 1 -C 7 alkylene. R 26 represents C 1 -C 3 alkylene. R 27 represents C 1 -C 5 alkylene.

[0078] In the present specification, the alkylene and alkenylene may have one or more arbitrary substituents. Examples of such substituents include, but are not limited to, alkoxy groups, halogen atoms (any one of fluorine atom, chlorine atom, bromine atom, or iodine atom), amino groups, mono- or disubstituted amino groups, substituted silyl groups, acyl groups, or aryl groups. When the alkyl group has two or more substituents, these substituents may be the same or different. The same applies to the alkyl moiety of other substituents containing an alkyl moiety (such as alkoxy groups or aralkyl groups).

[0079] In addition, in the present specification, when a certain functional group is defined as "may have a substituent", the type, substitution position, and number of substituents are not particularly limited. When there are two or more substituents, these substituents may be the same or different. Examples of substituents include, but are not limited to, alkyl groups, alkoxy groups, hydroxyl groups, carboxyl groups, halogen atoms, sulfo groups, amino groups, alkoxycarbonyl groups, oxo groups, etc. Substituents may further be present in these substituents.

[0080] As another form, one of the first polymer or the second polymer may be replaced with a low-molecular compound. In this case, the low-molecular compound has one or more nucleophilic functional groups or electron-withdrawing functional groups in the molecule. Thus, for example, by using a low-molecular compound having a nucleophilic functional group in the molecule instead of the first polymer and reacting it with the second polymer having one or more electron-withdrawing functional groups in the side chain or at the end, the second polymer can be gelled. Examples of the "low-molecular compound having a nucleophilic functional group in the molecule" include compounds having a mercapto group in the molecule, and dithiothreitol can be used, for example.

[0081] (1-2) Conditions of the polymer solution

[0082] The polymer solution A and the polymer solution B constituting the polymer material kit of the present invention satisfy the following conditions such as polymer concentration, pH, ionic strength, etc.

[0083] The concentrations of the first polymer and the second polymer in polymer solutions A and B are respectively in the range of 10 to 300 g / L, preferably in the range of 30 to 200 g / L, and more preferably in the range of 50 to 150 g / L. By adjusting the polymer concentrations as described above, the gelation time can be within the desired range. Among them, as long as the above ranges are satisfied, the concentrations of the first and second polymers can be the same or different respectively, and preferably they are the same concentration.

[0084] Polymer solutions A and B are adjusted according to the conditions of the mixed solution obtained by mixing these solutions in the acidic region (pH lower than 7), preferably adjusted to a range where the pH is 3 or higher and lower than 7, and more preferably in the range of 3.2 to 5.0. In addition, preferably, the pH of either polymer solution A or B is in the range where the pH is 3 or higher and lower than 7, and preferably the pH is in the range of 3.2 to 5.0. However, as long as the mixed solution of polymer solutions A and B satisfies the above acidic pH range, the pH of the other solution can also exceed 8. In a typical form, preferably, both polymer solutions A and B are within their pH ranges. By using the pH on the acidic side as described above, simply mixing polymer solutions A and B directly will not cause a gelation reaction in a short time (preferably, a gelation reaction cannot occur), but when the two solutions are mixed in an environment where there is blood or the like and the pH is near neutral (including the case of using the mixed solution in an environment with a pH near neutral), the pH of the polymer solution rises, and a gelation reaction can occur. Thus, in an environment where there is blood or the like and the pH is near neutral, a gel can be formed in situ in a relatively short time. As long as the above ranges are satisfied, the pH values of polymer solutions A and B can be the same or different respectively, and preferably they are the same pH.

[0085] The pH of polymer solutions A and B can use pH buffers well-known in the art. For example, by using a citric acid - phosphate buffer (CPB) and changing the mixing ratio of citric acid and disodium hydrogen phosphate, the pH can be adjusted to the above range.

[0086] In addition, for polymer solutions A and B, the ionic strength of the mixed solution obtained by mixing these solutions is in the range of 10 to 100 mM, preferably adjusted to the range of 10 to 40 mM. By adjusting the ionic strength as described above, the gelation time can be within the desired range. As long as the above ranges are satisfied, the ionic strengths of the respective polymer solutions A and B can be the same or different respectively, and preferably they are the same ionic strength.

[0087] By making the conditions of the polymer concentration, pH, and ionic strength of polymer solutions A and B within the above ranges, and mixing these solutions in an environment where there is a liquid with a pH of 6.5 to 8.0 equivalent to body fluids such as blood, a hydrogel in which the first polymer and the second polymer are crosslinked with each other can be formed in situ.

[0088] The gelation time at this time is preferably in the range of 1 to 30 seconds, more preferably in the range of 1 to 10 seconds. Similarly to the above, the gelation time can be mainly adjusted by appropriately setting the polymer concentration, pH, and ionic strength of the polymer solution. Here, the "gelation time" refers to the time required until the storage elastic modulus G' and the loss elastic modulus G" reach G' = G".

[0089] The solvents in polymer solutions A and B are water, and depending on circumstances, a mixed solvent containing alcohols such as ethanol or other organic solvents can also be used. Preferably, polymer solutions A and B are aqueous solutions with water as the sole solvent.

[0090] The volumes of polymer solutions A and B in the polymer material kit of the present invention are appropriate according to conditions such as the area and structural complexity of the bleeding site and blood vessels to which they are applied, etc., but typically are in the range of 0.1 to 20 ml respectively, more preferably 1 to 10 ml.

[0091] (1 - 3) Hydrogel

[0092] As described above, a hydrogel can be formed by crosslinking the first polymer and the second polymer. In this specification, a "gel" generally refers to a dispersion system of a polymer that has lost fluidity and is in a state where the storage elastic modulus G' and the loss elastic modulus G" have a correlation of G’≥G”. In addition, a "hydrogel" is a gel containing water.

[0093] The hydrogel formed by the first polymer and the second polymer preferably has a swelling equilibrium degree in the range of 0.9 to 3.5, more preferably in the range of 0.9 to 2.5. Thus, after forming a hydrogel at the bleeding site and blood vessels, etc., the gel will not swell excessively, and the undesirable effects when staying at the affected part can be inhibited within a certain period. Here, the "swelling equilibrium degree" refers to the value of the swelling degree when the change in the swelling degree with the passage of time reaches an equilibrium state after the gel is formed. The swelling degree can be measured by methods commonly used in this technical field. The swelling degree can use the value measured at 25°C.

[0094] In addition, the hydrogel formed by the first polymer and the second polymer preferably has a Young's modulus in the range of 0.1×10 4 ~4×10 4 Pa, more preferably in the range of 0.5×10 4 ~2×10 4 Pa. Thus, the hydrogel formed at the bleeding site and blood vessels, etc., can have a strength suitable for staying at the affected part within a certain period.

[0095] 2. Hemostatic agents, etc. of the present invention, and hemostasis methods, etc.

[0096] From another aspect, the present invention also relates to a hemostatic agent, a vascular occluding agent, a tissue coating agent, or a body fluid coagulant formed from a polymer material kit.

[0097] As described above, by using the polymer material kit of the present invention, the polymer solutions A and B are applied to an environment where blood exists, such as a bleeding site and within a blood vessel, and a gel is formed in situ, thereby forming a gel-blood complex incorporating blood. Thus, it can be provided while having a good blood coagulation effect produced by incorporating blood into the gel and also having a hemostatic effect in a physical manner of coating the bleeding site etc. with the gel. In addition, by applying the in situ gel formation to blood vessels such as veins and arteries, it can be used for the purpose of vascular occlusion, and is not limited to blood, but can also be used for the purpose of coagulating a body fluid having a pH near neutrality.

[0098] Here, the "tissue" which is the object of the present invention can broadly include living tissues and living organs in which a liquid having a pH near neutrality exists, and examples thereof can include organs, nerves, muscles, and parts thereof. However, even a site where the general surface is originally considered to be acidic can be included in the tissue which is the object of the present invention when it is temporarily or permanently modified to a pH near neutrality by prior treatment. Examples thereof include the gastric mucosa etc., but are not limited thereto.

[0099] In addition, from another aspect, the present invention also relates to a method for manufacturing the above-mentioned hemostatic agent, vascular occluding agent, tissue coating agent, or body fluid coagulant. The manufacturing method is characterized by including a step of applying a polymer solution A containing a first polymer and a second polymer solution B containing a second polymer to an environment where a liquid having a pH near neutrality, that is, a pH of 6.5 to 8.0 exists. In addition to this, the types of the first and second polymers and the conditions of the polymer solutions A and B are as described above.

[0100] Here, the "environment where a liquid having a pH of 6.5 to 8.0 exists" is preferably a place where blood and body fluids exist, and can be, for example, blood vessels such as arteries and veins, or tissues where blood or body fluids exist. The pH range is preferably 6.5 to 7.5.

[0101] The step of "applying" the mixture of the polymer solution A and the second polymer solution B to an environment where a liquid with a pH of 6.5 to 8.0 exists can be typically exemplified by mixing the polymer solution A and the second polymer solution B in such a pH environment. For example, it includes directly dropping or spraying the polymer solution A and B in sequence at the affected part where blood exists. Depending on the situation, the polymer solution A and B can be dropped or sprayed simultaneously. In addition, as described above, the polymer solution A and B are set to solution conditions where gelation reaction does not occur within a short time (preferably, gelation reaction cannot occur) only by direct mixing. Therefore, the polymer solution A and B can also be pre-mixed into one solution and then applied to an environment where a liquid with a pH of 6.5 to 8.0 exists.

[0102] As another form of the "applying" step, an example can be the step of dropping the polymer solution A and the polymer solution B onto a carrier and then bringing the carrier into contact with an environment where a liquid with a pH of 6.5 to 8.0 exists. In this case, it includes the step of temporarily holding each polymer solution in the carrier and then covering or protecting the affected part where blood exists (such as the affected part after a suture operation) with the carrier. As such a carrier, as long as it is a carrier made of a material that can hold the polymer solution, there is no particular limitation. Examples can include cloth-like members such as gauze and absorbent members such as sponges. In addition, as a further modified example, it is also possible to keep either the polymer solution A or B in the state of the carrier, bring the carrier into contact with the affected part, etc., and then drop or spray the remaining polymer solution onto the carrier.

[0103] As a means for mixing the polymer solution A and B by dropping, for example, a two-liquid mixing syringe disclosed in International Publication WO2007 / 083522 can be used. The temperature of the two liquids during mixing is not particularly limited, as long as the precursor units are dissolved respectively and the temperature is such that each solution has fluidity. For example, the temperatures of the two liquids can be different, and when the temperatures are the same, the two liquids are more easily mixed, so this is preferred.

[0104] As a means for spraying the polymer solution A and / or B, a sprayer containing the solution can be used. The sprayer can be appropriately a sprayer well-known in the technical field, preferably a medical sprayer. Therefore, as the container in the kit of the present invention, the above-mentioned sprayer can be used. In this case, as one form of the present invention, it can be a medical device containing the polymer solution A and B, preferably a sprayer.

[0105] Implementing the manufacturing method of the present invention can also be considered as a hemostasis method, blood vessel occlusion method, tissue covering method, and body fluid coagulation method using the above-mentioned polymer material kit. Among them, the tissues and blood vessels, etc. that can be used as objects are not limited to those in vivo, and the method also includes applications to tissues, etc. taken out from the body through surgery, etc.

[0106] Example

[0107] Hereinafter, the present invention will be described in more detail by way of examples, but the present invention is not limited by these examples.

[0108] 1. Preparation of Polymer Solution

[0109] As the starting polymers, four-arm PEG-SH (tetrathiol-polyethylene glycol) having a -SH group at the end and four-arm PEG-MA (tetramaleimide-polyethylene glycol) having a maleimide group at the end were used. These starting polymers were respectively polymers commercially available from NOF Corporation. The weight-average molecular weight (Mw) was 20,000 for both.

[0110] As a buffer for the polymer solution, citric acid-phosphate buffer (CPB) was used. The pH was adjusted by changing the mixing ratio of citric acid and disodium hydrogen phosphate. In addition, the buffering capacity was adjusted by changing the molar concentrations of citric acid and disodium hydrogen phosphate (referred to as ionic strength).

[0111] When preparing CPB with a pH of 3.8 and an ionic strength of 200 mM, each polymer solution was prepared according to the following steps.

[0112] 1. Prepare 200 mM aqueous solutions of citric acid and disodium hydrogen phosphate.

[0113] 2. Mix them in a ratio of citric acid aqueous solution:disodium hydrogen phosphate aqueous solution = 32.3:35.4.

[0114] At pH 5.8, mix them in a ratio of 19.7:60.6; at pH 3.0, mix them in a ratio of 39.8:20.4.

[0115] When finely adjusting the pH, it was carried out by mixing these CPBs in a certain ratio. In addition, the pH of each measurement sample was measured with a pH meter (HORIBA, Ltd.).

[0116] 2. Gelation Experiment

[0117] Using milk (pH = 7.4) having a pH equivalent to that of blood, the gelation behavior of the above four-arm PEG polymer was evaluated.

[0118] 1 mL of each of the two prepolymer solutions was prepared, and the time from when they were mixed until gelation was measured to examine the gelation time. Here, simply the time until the solution did not drop even when the container filled with the solution was inverted was defined as the gelation time. The concentration of the polymer, the pH of the buffer, and the ionic strength were changed.

[0119] The results of the polymer and pH dependence in the gelation time are shown in Figure 1 , and the results of the ionic strength dependence in the gelation time are shown in Figure 2 . It can be seen that the gelation time depends on the concentration of the prepolymer and the pH of the buffer. In addition, even when the ionic strength of the buffer (an index of the ability to maintain pH) is changed, although the gelation time changes, this is only because as the ionic strength decreases, the pH can no longer be maintained. In fact, after plotting the relationship between pH and gelation time obtained from Figure 2 , 3 in a graph of the relationship between pH and gelation time at the same concentration, the same relationship can be obtained ( Figure 4 ).

[0120] Next, a gelation test of milk was carried out. First, a prepolymer solution for this milk gelation test was prepared (prepolymer concentration: 150 g / L, solvent: pH 3.4, 200 mM). This prepolymer solution was added to milk (milk 10: prepolymer solution 1, volume ratio), and the fluidity of the milk was macroscopically evaluated. The gelation time at this time was 3 minutes. In order to adjust the gelation time, another prepolymer solution was prepared (prepolymer concentration: 200 g / L, solvent: pH 3.4, 40 mM). After carrying out the above test using this prepolymer solution, the gelation time was shortened to 10 seconds.

[0121] A gelation test of rat whole blood was carried out. First, a prepolymer solution for this gelation test was prepared (prepolymer concentration: 200 g / L, solvent: pH 3.4, 40 mM). Heparin treatment was pre-implemented on the whole blood collected from rats. The heparin-treated whole blood was divided into two parts, and the previously prepared prepolymer solution was added to one part (whole blood 10: prepolymer solution 1, volume ratio). The prepolymer solution was not added to the other part. At this time, gelation was confirmed only in the group to which the prepolymer solution was added.

[0122] A gelation test was carried out by replacing the four-branched PEG-SH with dithiothreitol having two SH groups (molecular weight 154.253 g / mol). First, a prepolymer solution of four-branched PEG-MA was prepared (prepolymer concentration: 200 g / L, solvent: pH 3.4, 40 mM). A new dithiothreitol solution was prepared under the condition that the terminal MA groups of the prepolymer and the SH groups of dithiothreitol reached the same molar concentration. These two solutions were mixed in equal volume, and as a result, fluidity was lost and gelation was confirmed.

[0123] A gelation test was carried out by replacing the four-arm PEG-SH with a linear PEG-SH having two branches and an SH group at the end. First, a prepolymer solution of four-arm PEG-MA was prepared (prepolymer concentration: 200 g / L, solvent: pH 3.4, 40 mM). A new linear PEG-SH solution was prepared under the condition that the molar concentrations of the terminal MA groups of the prepolymer and the SH groups of the linear PEG-SH were the same. These two solutions were mixed in equal volumes, and as a result, the fluidity was lost, and gelation was confirmed.

[0124] A gelation test was carried out by replacing the four-arm PEG-MA with a linear PEG-MA having two branches and an MA group at the end. First, a prepolymer solution of four-arm PEG-SH was prepared (prepolymer concentration: 200 g / L, solvent: pH 3.4, 40 mM). A new linear PEG-MA solution was prepared under the condition that the molar concentrations of the terminal SH groups of the prepolymer and the MA groups of the linear PEG-MA were the same. These two solutions were mixed in equal volumes, and as a result, the fluidity was lost, and gelation was confirmed.

[0125] 3. Measurement of Young's Modulus

[0126] After the two-component mixed prepolymer solution was injected into the body, it was mixed with blood and gelated. The change in hardness (Young's modulus) based on the content of blood at this time was investigated. According to the above steps, a prepolymer solution was prepared (prepolymer concentration: 50 g / L, solvent: pH 3.8, 200 mM). After mixing the two prepolymers, milk was further mixed in different proportions. After gelation and the end of the reaction, its Young's modulus was obtained through a compression test ( Figure 5 ). For example, when an equal volume of milk was added to the prepolymer solution, it was expressed as 50%. The more milk was added, the lower the final Young's modulus. This is because the prepolymer solution was diluted by adding milk, and the crosslinking point density of the gel decreased.

[0127] 4. Measurement of Equilibrium Swelling Degree

[0128] Experimental procedure

[0129] Solvent: CPB with pH 3.0, 20 mM

[0130] Prepolymer concentration: 60, 120 g / L (6, 12 wt%)

[0131] The steps for preparing the gel were the same as above. After mixing the two components and gelation and the end of the reaction, it was added to milk, and the swelling degree was investigated. All gels reached the equilibrium swelling state in about 4 hours ( Figure 6 ). The equilibrium swelling degree was 1.7 at 60 g / L and 2.3 at 120 g / L, and this value was the same as the case of swelling in water (Figure 7 ), suggesting that the results from the swelling experiment in water can be used.

[0132] 5. Application to Rat Blood Vessels

[0133] Experimental procedure

[0134] Solvent: CPB with pH 4.6 and 20 mM (mixed with pH 3.8, 20 mM CPB:pH 5.8, 20 mM CP = 1:2)

[0135] Prepolymer concentration: 50 g / L (5 wt%)

[0136] The steps for preparing the gel are the same as above. After mixing the two solutions, gelation occurs in about 3 minutes when standing still (this gelation time can be changed according to pH). Therefore, about 100 μL is injected into the femoral vein of the rat thigh before that. At this time, the blood flow is stopped by compressing the upstream part of the blood vessel to prevent the gel from flowing immediately after injection. In addition, by reducing the ionic strength of the buffer solution to 20 mM, after injection, the solution mixes with the blood, so the pH rises easily and gelation occurs immediately. After maintaining the compressed state for about 30 seconds, release the hand and confirm that the gel has solidified and does not flow. In addition, after a certain period of time, open the affected area again and confirm that the gel does not flow. The image at this time is shown in Figure 8 . Confirm that the effect of gel-based vascular occlusion lasts for at least 2 - 3 weeks

[0137] When using the conditions of solvent: CPB with pH 3.0 and 20 mM, prepolymer concentration: 100 g / L (10 wt%), after mixing the two solutions, gelation takes more than 10 minutes when standing still, but due to the low ionic strength, after mixing with the rat's blood, the pH rises and gelation occurs immediately.

[0138] 6. Experimental procedure for application to the abdominal aorta of rats

[0139] Solvent: CPB with pH 3.4 and 40 mM

[0140] Prepolymer concentration: 200 g / L (20 wt%)

[0141] Puncture the abdominal aorta of the rat with an injection needle with an outer diameter of 0.2 mm to make it bleed. For the bleeding point, add the prepolymer solution after mixing the two solutions ( Figure 9 ). Compression hemostasis was performed for 1 minute in this state, and as a result, hemostasis was confirmed. At this time, as a comparison group, compression hemostasis was performed for 1 minute without adding the prepolymer solution, and as a result, no hemostasis was confirmed.

Claims

1. A medical device for body fluid coagulation, the medical device comprising a polymer material kit, the kit being formed from a polymer solution A containing a first polymer and a second polymer solution B containing a second polymer, wherein, the first polymer and the second polymer are hydrophilic polymers having a polyalkylene glycol backbone, and when the first polymer and the second polymer crosslink with each other, the combination of the first polymer and the second polymer can form a hydrogel, wherein the first polymer has one or more nucleophilic functional groups in the side chain or at the end, and the second polymer has one or more electron-withdrawing functional groups in the side chain or at the end, Wherein, the one or more nucleophilic functional groups are selected from a mercapto group and an amino group, and the one or more electron-withdrawing functional groups are selected from a maleimido group, an N-hydroxysuccinimide (NHS), a sulfo-succinimide, a phthalimido group, an imidazolyl group, an acryloyl group, a nitrophenyl group, and -CO 2 PhNO 2 , The first polymer and the second polymer have a weight-average molecular weight (Mw) in the range of 1×10 3 to 1×10 5 . the concentrations of the first polymer and the second polymer in the polymer solution A and the polymer solution B are in the range of 10 - 300 g / L, the pH of the mixture obtained by mixing the polymer solution A and the polymer solution B is in the range of 3.2 to 5.0, and the ionic strength is in the range of 10 - 100 mM, wherein when the polymer solution A and the second polymer solution B are mixed in an environment where there is a liquid with a pH of 6.5 to 8.0, a hydrogel in which the first polymer and the second polymer crosslink with each other is formed, the gelation time of the formed hydrogel is in the range of 1 - 30 seconds, and the hydrogel has an equilibrium swelling degree in the range of 0.9 - 3.5; wherein, under the condition of a pH value of 4.6, the gelation time of the hydrogel formed by the first polymer and the second polymer is about 3 minutes.

2. The medical device according to claim 1, wherein, the first polymer and the second polymer are dibranched, tribranched or tetrabranched polyethylene glycols.

3. The medical device according to claim 1 or 2, wherein, the pH values of the polymer solution A and the polymer solution B are both above 3 and below 7.

4. The medical device according to claim 1, wherein, In an environment where there is a liquid with a pH of 6.5 to 8.0, the gelation time of the hydrogel formed by the first polymer and the second polymer is in the range of 1 to 30 seconds, and the hydrogel has a Young's modulus in the range of 0.1×10 4 ~4×10 4 Pa.

5. A method for manufacturing a hemostatic agent, vascular occluding agent, tissue coating agent, or body fluid coagulant containing a hydrogel, wherein, it includes the steps of applying a polymer solution A containing a first polymer and a second polymer solution B containing a second polymer to an environment where there is a liquid with a pH of 6.5 - 8.0, and forming a hydrogel when the first polymer and the second polymer crosslink with each other, the first polymer and the second polymer are hydrophilic polymers having a polyalkylene glycol backbone, wherein the first polymer has one or more nucleophilic functional groups in the side chain or at the end, and the second polymer has one or more electron-withdrawing functional groups in the side chain or at the end, Wherein, the one or more nucleophilic functional groups are selected from a mercapto group and an amino group, and the one or more electron-withdrawing functional groups are selected from a maleimido group, an N-hydroxysuccinimide (NHS), a sulfosuccinimide, a phthalimido group, an imidazolyl group, an acryloyl group, a nitrophenyl group, and -CO 2 PhNO 2 , The first polymer and the second polymer have a weight-average molecular weight (Mw) in the range of 1×10 3 to 1×10 5 . the concentrations of the first polymer and the second polymer in the polymer solution A and the polymer solution B are in the range of 10 - 300 g / L, the pH of the mixture obtained by mixing the polymer solution A and the polymer solution B is 3.2 to 5.0, and the ionic strength is in the range of 10 - 100 mM, Among them, when the polymer solution A and the second polymer solution B are mixed in an environment where there is a liquid with a pH of 6.5 to 8.0, a hydrogel in which the first polymer and the second polymer are cross-linked with each other is formed. The gelation time of the formed hydrogel is in the range of 1 to 30 seconds, and the hydrogel has an equilibrium swelling degree in the range of 0.9 to 3.5; Among them, under the condition of a pH value of 4.6, the gelation time of the hydrogel formed by the first polymer and the second polymer is about 3 minutes.

6. The method according to claim 5, wherein, comprises mixing the polymer solution A and the polymer solution B in an environment where there is a liquid with a pH of 6.5 to 8.

0.

7. The method according to claim 5, wherein, comprises, after dropping the polymer solution A and the polymer solution B onto a carrier, bringing the carrier into contact with an environment where there is a liquid with a pH of 6.5 to 8.

0.

8. The method according to any one of claims 5 to 7, wherein, the pH values of both the polymer solution A and the polymer solution B are in the range above 3 and below 7.

9. The medical device according to claim 1, wherein, at least one of the polymer solution A and the polymer solution B is stored in a nebulizer.

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