Fast-acting hemostatic medical adhesive and method of making same

Abstract

The application discloses a kind of fast hemostatic medical adhesive, by diisocyanate and guanidyl carboxymethyl chitosan under the action of hydrophilic chain extender reaction generation.It also discloses its preparation method.The fast hemostatic medical adhesive of the application has fast blood coagulation and good adhesive property, realizes the fast hemostasis after nasal cavity operation and the immediate closure of surgical incision, realizes the possibility of "day surgery";With strong bacteriostasis, compared with traditional operation, greatly reduce the bacterial infection rate in the process of postoperative hemostasis and wound recovery;And good fluidity, facilitate the use of nasal cavity postoperative hemostasis, greatly improve the operability of clinic.

Description

Technical Field

[0001] This invention belongs to the field of medical technology, and in particular relates to a rapid hemostatic medical adhesive and its preparation method. Background Technology

[0002] The nose is an important gateway and starting point of the human respiratory tract, and nasal diseases are extremely common and widespread. Many conditions often require surgical treatment because the nose is rich in blood vessels and nasal mucosa. Once surgery is performed, the need to control bleeding at the surgical incision site and seal the nasal mucosa arises. As a respiratory organ, the nasal cavity traps most external microorganisms and dust. It is estimated that there are approximately 10,000 different types of bacteria in the human nasal cavity, making it highly susceptible to infection if there is an open wound. Currently, postoperative hemostasis of the nasal cavity is generally achieved through nasal packing. This method primarily works by reducing the space in the nasal cavity and applying pressure to stop bleeding. Although it is simple and easy to operate, the bleeding is slow, and packing is generally required for 3 to 7 days. Packing often causes severe pain. After packing, the nasal cavity becomes obstructed, and patients often experience severe headaches, increased tearing, hypoxia, collapse, or even shock. It causes significant damage to the nasal mucosa, easily leading to complications such as sinusitis, nasal adhesions, and nasal septum perforation. Posterior nasal packing, in particular, causes even greater pain and more complications. Packing usually requires a second removal, which can cause further damage to the mucosa and even bleeding. Improper post-operative care can lead to bacterial infections.

[0003] Therefore, developing a rapid hemostatic adhesive with antibacterial properties is of great significance for postoperative hemostasis in the nasal cavity. Addressing the problems of poor hemostasis, unpleasant patient experience, and various complications arising from secondary removal of existing nasal packing, this invention designs a medical adhesive with rapid hemostasis and good antibacterial properties. Summary of the Invention

[0004] This invention provides a fast-acting hemostatic medical adhesive that achieves rapid hemostasis and exhibits good antibacterial properties.

[0005] The technical solution adopted in this invention is as follows:

[0006] A rapid hemostatic medical adhesive, characterized in that it is formed by polymerization of diisocyanate and guanidyl-modified carboxymethyl chitosan, wherein the molar ratio of guanidinium to isocyanate is (0.03-0.47):9.9.

[0007] Preferably, the curing time of the adhesive is 8 to 46 minutes, the viscosity is 78 mPa·s to 158 mPa·s, and the clotting time is 7 to 276 seconds.

[0008] Preferably, the adhesive has an adhesive strength of 86 to 156 kPa.

[0009] Preferably, the degree of guanidinium substitution in guanidinomethyl chitosan is 56%–91%.

[0010] Preferably, the diisocyanate refers to an isocyanate with a functional group of 2, and is one or a mixture of several of toluene diisocyanate, isophorone diisocyanate, diphenylmethane diisocyanate, dicyclohexylmethane diisocyanate, hexamethylene diisocyanate or lysine diisocyanate.

[0011] Preferably, it is formed by the polymerization reaction of diisocyanate and guanidine carboxymethyl chitosan under the action of a hydrophilic chain extender.

[0012] Preferably, the hydrophilic chain extender is one or a mixture of several of the following: lysine, serine, cystine, butanediamine, butanediol, propylene glycol, or ethylene glycol.

[0013] This invention also discloses a method for preparing guanidinomethyl chitosan, characterized by the following steps:

[0014] (1) Preparation of guanidine derivatives: Add an aqueous solution of the guanidine-modified substance to isopropanol, adjust the pH to 3.8-4.2, add a tertiary amine compound dropwise, and react in an ice bath to obtain the guanidine derivatives;

[0015] (2) Preparation of carboxymethyl chitosan derivatives: Carboxymethyl chitosan was dissolved in water, and epichlorohydrin was added dropwise to react and obtain carboxymethyl chitosan derivatives;

[0016] (3) Preparation of guanidinomethyl chitosan: Dissolve the derivative of carboxymethyl chitosan in anhydrous ethanol, and add guanidino derivative dropwise to react and obtain guanidinomethyl chitosan.

[0017] Preferably, the guanidinized substance is selected from thiourea trioxide, cyanamide, disperidin, diphenylthiourea, guanidine dodecyl acetate, or biguanide acetate.

[0018] Preferably, the tertiary amine compound is selected from N,N-dimethylaniline, N,N-diethylaniline, N,N-dimethyl-p-toluidine, N,N-dihydroxyethyl-p-toluidine, N-methyl-N-hydroxyethyl-p-toluidine, triethylamine, 1,6-hexanediamine or N,N-dimethyl-1,3-propanediamine.

[0019] Preferably, the molar ratio of the amino group of the carboxymethyl chitosan derivative to the guanidinium group of the guanidinium derivative is 1:3 to 5.

[0020] This invention also discloses a method for preparing the above-mentioned rapid hemostatic medical adhesive, characterized in that its steps include:

[0021] (1) Add diisocyanate, guanidyl carboxymethyl chitosan and reaction solvent to the reaction vessel, purge the above system with nitrogen gas, and react for a period of time with stirring.

[0022] (2) Add a hydrophilic chain extender and stir to react for a period of time;

[0023] (3) Increase the temperature and stir the reaction for a period of time;

[0024] (4) Cool down to room temperature, add neutralizing agent to carry out neutralization reaction, and stir the reaction for a period of time;

[0025] (5) Remove the solvent by rotary evaporation, add diluent, and stir until a single clear phase is obtained to obtain a fast hemostatic medical adhesive.

[0026] Preferably, the reaction solvent is one or a mixture of several of tetrahydrofuran, dichloromethane, N,N-dimethylformamide, dimethyl sulfoxide, ethyl acetate, petroleum ether, hexane, or acetone.

[0027] Preferably, the neutralizing agent is a reagent that can neutralize hydrophilic chain extenders.

[0028] Preferably, the neutralizing agent is one or a mixture of several of glacial acetic acid, phosphoric acid, hydrochloric acid, citric acid, or sulfuric acid.

[0029] Preferably, the diluent is one or a mixture of several of isosorbide dimethyl ether, soybean oil methyl ester, triacetin, or triethylene glycol dimethyl ether.

[0030] Preferably, the guanidinomethyl chitosan refers to the guanidinomethyl chitosan prepared by any one of steps 4-6.

[0031] Preferably, in step (1), the reaction temperature is 50-80℃, the reaction time is 3-8h, the stirring speed is 80-120r / min, the amount of guanidyl-modified carboxymethyl chitosan added is 2-30mg / ml, and the mass ratio of solvent to diisocyanate is 1.5:1-3.5:1.

[0032] Preferably, in step (2), the reaction temperature is 50-80℃, the reaction time is 30-45min, the stirring speed is 200-400r / min, and the amount of hydrophilic chain extender added is based on a molar ratio of isocyanate to hydroxyl group greater than 2.0:1.

[0033] Preferably, in step (3), the reaction temperature is 75-90℃, the reaction time is 2-5h, and the stirring speed is 400-500r / min.

[0034] Preferably, in step (4), the amount of neutralizing agent added is 20ppm to 80ppm, the temperature is controlled at 7 to 18°C, the stirring reaction time is 30 to 45 minutes, and the stirring speed is 500 to 600 r / min.

[0035] Preferably, in step (5), the stirring speed is 300-600 r / min, and the amount of diluent added is 30%-50% WT of the total reaction system.

[0036] The detailed preparation process of guanidinomethyl chitosan is as follows:

[0037] 1. Preparation of guanidinolated chitosan

[0038] 1.1 Preparation of guanidine derivatives

[0039] (1) Prepare a 50% aqueous solution of the guanidine compound for later use;

[0040] (2) Take 1 part of the above aqueous solution, add 10 parts of isopropanol to a three-necked flask, adjust the pH of the reaction system to 3.8-4.2, add 2 parts of tertiary amine compound at 5-8 drops / min, and react in an ice bath for 8-12 hours;

[0041] (3) Let stand for 30-45 minutes, then separate the product using a separatory funnel;

[0042] (4) Wash with isopropanol 3 to 5 times to obtain the guanidine derivative, as shown in the following reaction formula:

[0043]

[0044] 1.2 Preparation of carboxymethyl chitosan derivatives

[0045] (1) Take 1 part of carboxymethyl chitosan and add it to 30 parts of deionized water. Stir with an electric mixer until completely dissolved. At 80°C, slowly add 1 part of epichlorohydrin and react for 8-10 hours.

[0046] (2) The unreacted residue was distilled off under reduced pressure to obtain a solid product of the chitosan derivative;

[0047] (3) Wash the solid 3-5 times with anhydrous ethanol and dry under vacuum to obtain a derivative of carboxymethyl chitosan. The reaction formula is as follows:

[0048]

[0049] 1.3 Preparation of guanidinomethyl chitosan

[0050] (1) Dissolve 1 part of carboxymethyl chitosan derivative in 50 parts of anhydrous ethanol;

[0051] (2) Adjust the pH to 5-6, slowly add the guanidine derivative at room temperature, and react for 8-12 hours with electric stirring;

[0052] (3) Anhydrous ethanol was distilled off under reduced pressure;

[0053] (4) Dialyze using dialysis bags for 7-10 days, changing the deionized water every 24 hours;

[0054] (5) The dialysis product was removed, subjected to vacuum distillation, and then dried under vacuum to obtain guanidyl-modified carboxymethyl chitosan. The reaction formula is as follows:

[0055]

[0056] The guanidinolated substances mainly refer to those synthesized by reacting amines (ammonia) with guanidinolation reagents such as cyanamide, isothiourea, isourea, hydrazine, and isothiocyanate. These include, but are not limited to, thiourea trioxide, monocyanamide, dispermine, diphenylthiourea, guanidine dodecyl acetate, biguanide acetate, etc., with monocyanamide being preferred.

[0057] The tertiary amine compounds include, but are not limited to, N,N-dimethylaniline, N,N-diethylaniline, N,N-dimethyl-p-toluidine, N,N-dihydroxyethyl-p-toluidine, N-methyl-N-hydroxyethyl-p-toluidine, triethylamine, 1,6-hexanediamine, N,N-dimethyl-1,3-propanediamine and combinations thereof, preferably N,N-dimethyl-1,3-propanediamine and 1,6-hexanediamine.

[0058] The carboxymethyl chitosan has a molecular weight of less than 5000, preferably less than 2000, and a degree of deacetylation of more than 85%, preferably more than 90%, and more preferably more than 95%.

[0059] The dialysis bag can dialyze unreacted raw materials and small molecule byproducts that have not been added, and its dialysis molecular weight should be below that of chitosan.

[0060] During preparation, carboxymethyl chitosan containing anionic carboxyl groups is used. It then reacts with epichlorohydrin and subsequently with guanidine-containing substances, resulting in a chitosan guanidine derivative that is dimorphic, containing both acidic and basic functional groups. The guanidine group is a basic functional group, which would lead to extremely high viscosity and easy gelation of the adhesive. The addition of acidic carboxyl groups reduces this gelation effect during the reaction with isocyanates. One end of the carboxyl group can react with isocyanates, and the residual hydroxyl group can also react. By attaching isocyanates to the prepolymer of the polyurethane adhesive molecules, the viscosity of the adhesive is not too high.

[0061] The beneficial effects of this invention are as follows:

[0062] 1. The rapid hemostatic medical adhesive of the present invention has rapid hemostasis and good adhesion properties, realizing rapid hemostasis after nasal surgery and immediate closure of surgical incision, making "day surgery" possible.

[0063] 2. The rapid hemostatic medical adhesive of the present invention has strong antibacterial properties, which greatly reduces the bacterial infection rate during postoperative hemostasis and wound healing compared with traditional surgery.

[0064] 3. The rapid hemostatic medical adhesive of the present invention has good fluidity, which facilitates postoperative hemostasis in the nasal cavity and greatly improves clinical operability. Attached Figure Description

[0065] Figure 1 This is the result of the guanidine substitution in Example 6;

[0066] Figure 2 This is a photograph of the inhibition zone in Example 7;

[0067] Figure 3 It is the diameter of the inhibition zone in Example 7;

[0068] Figure 4 This is a photograph of cell morphology from Example 7. Detailed Implementation

[0069] To further describe the specific content of the present invention, the present invention will be described in detail below with reference to embodiments. The operating methods and reagents involved in the embodiments are well known to those skilled in the art. It should be understood that the specific embodiments described below are merely for the purpose of describing the invention in detail, but the implementation of the present invention is not limited to the following embodiments.

[0070] Example 1

[0071] 1.1 Preparation of guanidinolated chitosan

[0072] 1.1.1 Preparation of guanidine derivatives

[0073] (1) Prepare a 50% WT cyanamide aqueous solution for later use;

[0074] (2) Take 1 part of the above aqueous solution, add 10 parts of isopropanol to a three-necked flask, adjust the pH of the reaction system to 3.8-4.2, add 2 parts of N,N-dimethyl-1,3-propanediamine at a rate of 5-8 drops / min, and react in an ice bath for 8 hours;

[0075] (3) Let stand for 30 minutes, then separate the product using a separatory funnel;

[0076] (4) Wash with isopropanol three times to obtain N,N-dimethyl-N'-guanidinyl-1,3-propanediamine.

[0077] 1.1.2 Preparation of carboxymethyl chitosan derivatives

[0078] (1) Take 1 part of carboxymethyl chitosan and add it to 30 parts of deionized water. Stir with an electric mixer until completely dissolved. At 80°C, slowly add 1 part of epichlorohydrin and react for 8 hours.

[0079] (2) The unreacted residue was distilled off under reduced pressure to obtain a solid product of the chitosan derivative;

[0080] (3) The solid was washed three times with anhydrous ethanol and dried under vacuum to obtain N-(1-hydroxy-3-chloropropyl)-carboxymethyl chitosan.

[0081] 1.1.3 Preparation of guanidinomethyl chitosan

[0082] (1) Dissolve 1 part of a carboxymethyl chitosan derivative with a degree of deacetylation of 98% in 50 parts of anhydrous ethanol;

[0083] (2) Adjust the pH to 6.0, and at room temperature, slowly add N,N-dimethyl-N-guanidin-1,3-propanediamine at a ratio of 1:5 (molar ratio) of amino group of carboxymethyl chitosan derivative to guanidin group of N,N-dimethyl-N-guanidin-1,3-propanediamine, and react for 8 hours under electric stirring.

[0084] (3) Anhydrous ethanol was distilled off under reduced pressure;

[0085] (4) Dialyze using dialysis bags for 3 days, changing the deionized water every 8 hours;

[0086] (5) The dialysis product was removed, and then vacuum distilled and dried to obtain guanidinomethyl chitosan.

[0087] 1.2 Preparation of rapid hemostatic medical adhesive for nasal surgery

[0088] (1) In a four-necked flask equipped with an electric stirrer, a temperature detector and a reflux condenser, add lysine diisocyanate, guanidyl carboxymethyl chitosan and tetrahydrofuran, wherein the amount of guanidyl carboxymethyl chitosan added is 15 mg / ml.

[0089] (2) The above system was purged with nitrogen and reacted for 3 hours at a stirring speed of 300 r / min and an oil bath at 50 °C.

[0090] (3) Add lysine according to the molar ratio of isocyanate to amino group of 2.05:1, and stir at 300 r / min for 30 min.

[0091] (4) Slowly raise the temperature to 80°C and react in an oil bath for 3 hours.

[0092] (5) Cool down to 18°C, add citric acid for neutralization reaction, and stir for 30 min.

[0093] (6) The solvent was removed by rotary evaporation, and the diluent isosorbide dimethyl ether was added. The mixture was stirred at a stirring speed of 300 r / min until a single clear phase was obtained to obtain a fast hemostatic medical adhesive 1, wherein the molar ratio of guanidine to isocyanate was 0.23:9.9.

[0094] 1.3 Sterilization

[0095] The adhesive was sterilized by irradiation at a dose of 15 KGy.

[0096] Example 2

[0097] Based on Example 1, the reaction was carried out with guanidyl carboxymethyl chitosan added at a rate of 2 mg / ml to obtain rapid hemostatic medical adhesive 2, wherein the molar ratio of guanidinium to isocyanate is 0.03:9.9.

[0098] Example 3

[0099] Based on Example 1, the reaction was carried out with guanidyl carboxymethyl chitosan added at a rate of 30 mg / ml to obtain rapid hemostatic medical adhesive 3, wherein the molar ratio of guanidin to isocyanate was 0.47:9.9.

[0100] Example 4

[0101] 4.1 Preparation of guanidinolated chitosan

[0102] 4.1.1 Preparation of guanidine derivatives

[0103] (1) Prepare a 50% WT thiourea trioxide aqueous solution for later use;

[0104] (2) Take 1 part of the above aqueous solution, add 10 parts of isopropanol to a three-necked flask, adjust the pH of the reaction system to 3.8-4.2, add 2 parts of 1,6-hexanediamine at a rate of 5-8 drops / min, and react in an ice bath for 8 hours;

[0105] (3) Let stand for 30 minutes, then separate the product using a separatory funnel;

[0106] (4) Wash with isopropanol three times to obtain guanidino-1,6-hexanediamine.

[0107] 4.1.2 Preparation of carboxymethyl chitosan derivatives

[0108] (1) Take 1 part of carboxymethyl chitosan with a degree of deacetylation of 95% and add it to 30 parts of deionized water. Stir with an electric mixer until completely dissolved. At 80°C, slowly add 1 part of epichlorohydrin and react for 8 hours.

[0109] (2) The unreacted residue was distilled off under reduced pressure to obtain a solid product of the chitosan derivative;

[0110] (3) The solid was washed three times with anhydrous ethanol and dried under vacuum to obtain N-(1-hydroxy-3-chloropropyl)-carboxymethyl chitosan.

[0111] 4.1.3 Preparation of guanidinomethyl chitosan

[0112] (1) Dissolve 1 part of carboxymethyl chitosan derivative in 50 parts of anhydrous ethanol;

[0113] (2) Adjust the pH to 6.0, and at room temperature, slowly add guanidin-1,6-hexanediamine dropwise at a ratio of 1:5 (molar ratio) of amino group of carboxymethyl chitosan derivative to guanidin group of guanidin-1,6-hexanediamine, and react for 8 hours under electric stirring.

[0114] (3) Anhydrous ethanol was distilled off under reduced pressure;

[0115] (4) Dialyze using dialysis bags for 3 days, changing the deionized water every 8 hours;

[0116] (5) The dialysis product was removed, and then vacuum distilled and dried to obtain guanidinomethyl chitosan.

[0117] 4.2 Preparation of rapid hemostatic medical adhesive for nasal surgery

[0118] (1) In a four-necked flask equipped with an electric stirrer, a temperature detector and a reflux condenser, add lysine diisocyanate, guanidyl carboxymethyl chitosan and tetrahydrofuran, wherein the amount of guanidyl carboxymethyl chitosan added is 15 mg / ml.

[0119] (2) The above system was purged with nitrogen and reacted for 3 hours at a stirring speed of 300 r / min and an oil bath at 50 °C.

[0120] (3) Add lysine according to the molar ratio of isocyanate to amino group of 2.05:1, and stir at 300 r / min for 30 min.

[0121] (4) Slowly raise the temperature to 80°C and react in an oil bath for 3 hours.

[0122] (5) Cool down to 18°C, add citric acid for neutralization reaction, and stir for 30 min.

[0123] (6) Remove tetrahydrofuran by rotary evaporation, add isosorbide dimethyl ether, and stir at 300 r / min until a single clear phase is obtained to obtain rapid hemostatic medical adhesive 4, wherein the molar ratio of guanidine to isocyanate is 0.23:9.9.

[0124] 4.4 Sterilization

[0125] The adhesive was sterilized by irradiation at a dose of 15 KGy.

[0126] Example 5

[0127] 5.1 Preparation of guanidinolated chitosan

[0128] 5.1.1 Preparation of guanidine derivatives

[0129] (1) Prepare a 50% cyanamide aqueous solution for later use;

[0130] (2) Take 1 part of the above aqueous solution, add 10 parts of isopropanol to a three-necked flask, adjust the pH of the reaction system to 3.8-4.2, add 2 parts of N,N-dimethyl-1,3-propanediamine at 5-8 drops / min, and react in an ice bath for 8 hours;

[0131] (3) Let stand for 30 minutes, then separate the product using a separatory funnel;

[0132] (4) Wash with isopropanol three times to obtain N,N-dimethyl-N'-guanidinyl-1,3-propanediamine.

[0133] 1.1.2 Preparation of carboxymethyl chitosan derivatives

[0134] (1) Take 1 part of carboxymethyl chitosan and add it to 30 parts of deionized water. Stir with an electric mixer until completely dissolved. At 80°C, slowly add 1 part of epichlorohydrin and react for 8 hours.

[0135] (2) The unreacted residue was distilled off under reduced pressure to obtain a solid product of the chitosan derivative;

[0136] (3) The solid was washed three times with anhydrous ethanol and dried under vacuum to obtain N-(1-hydroxy-3-chloropropyl)-carboxymethyl chitosan.

[0137] 5.1.3 Preparation of guanidinomethyl chitosan

[0138] (1) Dissolve 1 part of a carboxymethyl chitosan derivative with a degree of deacetylation of 98% in 50 parts of anhydrous ethanol;

[0139] (2) Adjust the pH to 6.0, and at room temperature, slowly add N,N-dimethyl-N-guanidin-1,3-propanediamine at a ratio of 1:3 (molar ratio) of amino group of carboxymethyl chitosan derivative to guanidin group of N,N-dimethyl-N-guanidin-1,3-propanediamine, and react for 8 hours under electric stirring.

[0140] (3) Anhydrous ethanol was distilled off under reduced pressure;

[0141] (4) Dialyze using dialysis bags for 3 days, changing the deionized water every 8 hours;

[0142] (5) The dialysis product was removed, and then vacuum distilled and dried to obtain guanidinomethyl chitosan.

[0143] 5.2 Preparation of rapid hemostatic medical adhesive for nasal surgery

[0144] (1) In a four-necked flask equipped with an electric stirrer, a temperature detector and a reflux condenser, add isophorone diisocyanate, guanidine carboxymethyl chitosan and tetrahydrofuran, wherein the amount of guanidine carboxymethyl chitosan added is 15 mg / ml.

[0145] (2) The above system was purged with nitrogen and reacted for 3 hours at a stirring speed of 300 r / min and an oil bath at 50 °C.

[0146] (3) Add lysine according to the molar ratio of isocyanate to amino group of 2.05:1, and stir at 300r / min for 30min.

[0147] (4) Slowly raise the temperature to 80°C and react in an oil bath for 3 hours.

[0148] (5) Cool down to 18°C, add citric acid for neutralization reaction, and stir for 30 min.

[0149] (6) The solvent was removed by rotary evaporation, and isosorbide dimethyl ether was added. The mixture was stirred at 300 r / min until a single clear phase was obtained to obtain a fast hemostatic medical adhesive 5, wherein the molar ratio of guanidine to isocyanate was 0.20:9.9.

[0150] 5.3 Sterilization

[0151] The adhesive was sterilized by irradiation at a dose of 15 KGy.

[0152] Comparative Example 1

[0153] (1) In a four-necked flask equipped with an electric stirrer, a temperature detector and a reflux condenser, add lysine diisocyanate and tetrahydrofuran.

[0154] (2) The above system was purged with nitrogen and reacted for 3 hours at a stirring speed of 300 r / min and an oil bath at 50 °C.

[0155] (3) Add lysine at a molar ratio of isocyanate to amino group of 2.05:1 and stir at 300 r / min for 30 min.

[0156] (4) Slowly raise the temperature to 80°C and react in an oil bath for 3 hours.

[0157] (5) Cool down to 18°C, add citric acid for neutralization reaction, and stir for 30 min.

[0158] (6) The solvent was removed by rotary evaporation, and isosorbide dimethyl ether was added. The mixture was stirred at a stirring speed of 300 r / min until a single clear phase was obtained to obtain a fast hemostatic medical adhesive 6.

[0159] Comparative Example 2

[0160] The process was essentially the same as in Example 5, except that the molar ratio of the amino group of the carboxymethyl chitosan derivative to the guanidinium group of N,N-dimethyl-N-guanidinium-1,3-propanediamine was 1:2. N,N-dimethyl-N-guanidinium-1,3-propanediamine was slowly added dropwise and reacted under electric stirring for 8 hours. After subsequent production, a rapid hemostatic medical adhesive 7 was obtained, wherein the molar ratio of guanidinium to isocyanate was 0.14:9.9.

[0161] Example 6 Characterization of the degree of substitution of carboxymethyl chitosan derivatives

[0162] 6.1 Preparation of guanidinomethyl chitosan with different ratios

[0163] (1) Dissolve 1 part of a carboxymethyl chitosan derivative with a degree of deacetylation of 98% in 50 parts of anhydrous ethanol;

[0164] (2) Adjust the pH to 6.0 and at room temperature, slowly add N,N-dimethyl-N-guanidin-1,3-propanediamine dropwise at molar ratios of guanidinized derivative to carboxymethyl chitosan of 5:1, 4:1, 3:1, 2:1, 1:1, 1:2, and 1:3 respectively, and react for 8 hours under electric stirring.

[0165] (3) Anhydrous ethanol was distilled off under reduced pressure;

[0166] (4) Dialyze using dialysis bags for 3 days, changing the deionized water every 8 hours;

[0167] (5) The dialysis product was removed, and then vacuum distilled and dried to obtain guanidinomethyl chitosan.

[0168] 6.2 Determination of the degree of substitution of carboxymethyl chitosan derivatives by potentiometric titration

[0169] 0.1 g of purified carboxymethyl chitosan was weighed using an electronic analytical balance. The solution was dissolved in 10 mL of 0.1006 mol / L hydrochloric acid solution using magnetic stirring in a beaker. A 0.1 mol / L sodium hydroxide solution was then prepared and used for titration. Conductivity was measured using a digital conductivity meter, and the pH of the titrant was determined using a pH meter. The degree of substitution of carboxymethyl chitosan was calculated by determining the amount of sodium hydroxide used in the titration. The structures are shown in Table 1. Figure 1 As shown.

[0170] The endpoint of the excess hydrochloric acid titration is V1, and the endpoint of the carboxymethyl titration is V2. V2-V1 is the volume of sodium hydroxide solution consumed in the titration of carboxymethyl chitosan. The formula for calculating the degree of substitution DS of carboxymethyl chitosan is as follows:

[0171] Equation 1: A = (V2 - V1) c / m

[0172] Equation 2: DS = 0.161A / (1 - 0.058A)

[0173] in:

[0174] In Equations 1 and 2: c is the concentration of NaOH in the titrant; m is the net weight of the sample; A is the number of moles of carboxymethyl groups in the sample; 58 is the relative molecular weight of carboxymethyl groups; and 161 is the relative molecular weight of glucosamine residues.

[0175] Table 1. Degree of Substitution Results of Carboxymethyl Chitosan

[0176]

[0177]

[0178] Note: Experiments show that the molar ratio of guanidinium group in guanidine compounds to amino group in carboxymethyl chitosan is above 3:1, and the degree of substitution can reach above 78%, which can ensure the antibacterial and hemostatic effects of the product.

[0179] Example 7: Further Characterization of the Adhesive

[0180] 7.1 Curing time of adhesive

[0181] Take an appropriate amount of adhesive sample into a round, flat-bottomed polytetrafluoroethylene mold, and use the fluidity of the colloid itself to make it flow evenly and level. Allow it to cure naturally at room temperature, and record the time it takes for the colloid to fully cure.

[0182] 7.2 Viscosity

[0183] Take an appropriate amount of medical adhesive and test it at (25±0.1)℃ using a viscometer (60 RPM) according to the third method of viscosity determination in General Chapter 0633 of the Chinese Pharmacopoeia (2020 Edition, Part IV): rotational viscometer method.

[0184] 7.3 In vitro coagulation test

[0185] Heart blood from New Zealand white rabbits was mixed with sodium citrate (an anticoagulant) at a ratio of 9:1. 0.5 ml of medical adhesive was placed in a test tube and preheated at 37°C for 5 minutes. Then, 1 ml of anticoagulated rabbit blood was added, and the mixture was incubated at 37°C for another 3 minutes. After adding CaCl2 solution (25 mmol / L), the test tube was tilted every 10 seconds to observe blood flow until complete coagulation. The time from the addition of calcium chloride solution to complete coagulation was recorded as the whole blood clotting time.

[0186] The test results are shown in Table 2.

[0187] Table 2. Test results of curing time, viscosity, and clotting time for each adhesive.

[0188] Example 1 Adhesive 1 8min 158 mPa.s 18s Example 2 Adhesive 2 46min 101 mPa.s 276s Example 3 Adhesive 3 3min 78 mPa.s 7s Example 4 Adhesive 4 18min 256 mPa.s 185s Example 5 Adhesive 5 27min 125 mPa.s 156s Comparative Example 1 Adhesive 6 135min 1025 mPa.s -- Comparative Example 2 Adhesive 7 72min 96 mPa.s 786s

[0189] As can be seen from Table 2, the content of guanidinated chitosan introduced into the adhesive molecule directly affects the curing time, viscosity, and clotting time. The less guanidinated chitosan, the longer the curing time, the higher the viscosity, and the longer the clotting time. The more guanidinated chitosan, the shorter the curing time, the lower the viscosity, and the shorter the clotting time.

[0190] 7.4 Antibacterial test

[0191] (1) The strains were activated, bacterial suspensions were prepared and test plates were prepared in accordance with GB / T38483-2020 Determination of Antibacterial Activity of Sub-metabolites of Microbial Antibiotics by Inhibition Zone Method.

[0192] (2) Sample preparation

[0193] On a clean bench, take an appropriate amount of the adhesive from the examples and comparative examples into a round, flat-bottomed polytetrafluoroethylene mold. Utilize the fluidity of the colloid itself to make it flow evenly and level, and allow it to cure naturally. Using a sterilized punch, prepare a 1cm diameter adhesive film. At the same time, prepare a 1cm sterilized blank filter paper disc as a control.

[0194] (3) Use sterile tweezers to pick up the gel film and place it on the bacterial plate. There is one piece of each type of gel film per plate. Place a blank filter paper in the middle of the plate as a control. There are a total of 6 gel films per plate. Three sets of experiments were conducted in parallel.

[0195] (4) Place the petri dishes in a biochemical culture medium and incubate at 37℃ for 24 hours. Measure the diameter of the inhibition zone in the petri dish, record the values, and compare the antibacterial effects. The results are as follows: Figure 2 and Figure 3 As shown, the antibacterial effect becomes better with the increase of guanidinolated chitosan in the molecule.

[0196] 7.5 Bond strength

[0197] In this experiment, commercially available pigskin was used as the tensile substrate. The fat and epidermal layers of the pigskin were removed, and the pieces were cut into 2.5±0.1cm × 15±0.2cm pieces. The pigskin was then soaked in PBS buffer for 30 minutes to remove surface grease. The shear strength of the tissue adhesive was determined according to YY / T 0729.1-2009. The tensile test was conducted on an electronic tensile testing machine, with parameters adjusted before testing; the tensile speed was 10cm / min. The results are shown in Table 3. It can be seen that the adhesive strength gradually decreases with the increase of guanidinolated chitosan in the molecule.

[0198] Table 3. Bond strength test results of each adhesive.

[0199] Example 1 Medical adhesive 1 156 Example 2 Medical adhesive 2 204 Example 3 Medical adhesive 3 86 Example 4 Medical adhesive 4 124 Example 5 Medical adhesive 5 112 Comparative Example 1 Medical adhesive 6 248 Comparative Example 2 Medical adhesive 7 178

[0200] 7.6 Biological Evaluation - Cytotoxicity Assay

[0201] The extraction was performed according to GB / T16886.5 "Biological Evaluation of Medical Devices - Part 5: In Vitro Cytotoxicity Tests" at an extraction ratio of 0.2 g / ml. The results are shown in Table 4.

[0202] Table 4 Relative cell survival rate

[0203]

[0204]

[0205] Cell morphology was examined under a microscope, and the results were as follows: Figure 4 As shown in Table 4 and Figure 4 The introduction of chitosan guanidine derivatives did not produce biological toxicity, but different diisocyanates had a significant impact on cytotoxicity. Through screening of different embodiments, lysine diisocyanate had the least biological toxicity and was most suitable for use in nasal hemostasis.

[0206] The above description is merely a preferred embodiment of the present invention and is not intended to limit the present invention. Any modifications, equivalent substitutions, and improvements made within the spirit and principles of the present invention should be included within the protection scope of the present invention.

Claims

1. A hemostatic medical adhesive, characterized in that... It is produced by polymerization of diisocyanate and guanidyl-modified carboxymethyl chitosan, wherein the molar ratio of guanidinium to isocyanate is (0.03~0.47):9.9; The preparation method of guanidinomethyl chitosan is as follows: (1) Preparation of guanidine derivatives: Add the aqueous solution of the guanidine-modified substance to isopropanol, adjust the pH to 3.8-4.2, add tertiary amine compounds dropwise and react in an ice bath to obtain guanidine derivatives; (2) Preparation of carboxymethyl chitosan derivatives: Carboxymethyl chitosan was dissolved in water, and epichlorohydrin was added dropwise to react and obtain carboxymethyl chitosan derivatives; (3) Preparation of guanidine-modified carboxymethyl chitosan: Dissolve the derivative of carboxymethyl chitosan in anhydrous ethanol, and add the guanidine derivative dropwise to react and obtain guanidine-modified carboxymethyl chitosan; The molecular weight of carboxymethyl chitosan is below 5000, the degree of deacetylation is above 85%, and the degree of guanidin substitution in guanidyl-modified carboxymethyl chitosan is 56%~91%.

2. The hemostatic medical adhesive as described in claim 1, characterized in that... The adhesive has a curing time of 8 to 46 minutes, a viscosity of 78 mPa·s to 158 mPa·s, and a clotting time of 7 to 276 seconds.

3. The hemostatic medical adhesive as described in claim 2, characterized in that... The adhesive has a bonding strength of 86~156 kPa.

4. The hemostatic medical adhesive as described in claim 1, characterized in that... The diisocyanate mentioned refers to an isocyanate with a functional group of 2, and is one or a mixture of several of toluene diisocyanate, isophorone diisocyanate, diphenylmethane diisocyanate, dicyclohexylmethane diisocyanate, hexamethylene diisocyanate or lysine diisocyanate.

5. The hemostatic medical adhesive as described in claim 1, characterized in that... It is produced by the polymerization reaction of diisocyanate and guanidine carboxymethyl chitosan under the action of a hydrophilic chain extender.

6. The hemostatic medical adhesive as described in claim 5, characterized in that... The hydrophilic chain extender is one or a mixture of several of the following: lysine, serine, cystine, butanediamine, butanediol, propylene glycol, or ethylene glycol.

7. The method for preparing the hemostatic medical adhesive according to any one of claims 1-6, characterized in that... The steps include: (1) Add diisocyanate, guanidyl carboxymethyl chitosan and reaction solvent to the reaction vessel, introduce nitrogen gas into the reaction vessel, and react for 3-8 hours with stirring; (2) Add a hydrophilic chain extender and stir for 30 min to 45 min; (3) Increase the temperature and stir the reaction for 2-5 hours; (4) Cool down to room temperature, add neutralizing agent to carry out neutralization reaction, and stir the reaction for 30~45 min; (5) Remove the solvent by rotary evaporation, add diluent, and stir until a single clear phase is obtained to obtain medical adhesive.

8. The method for preparing the hemostatic medical adhesive as described in claim 7, characterized in that: The reaction solvent is one or a mixture of several of the following: tetrahydrofuran, dichloromethane, N,N-dimethylformamide, dimethyl sulfoxide, ethyl acetate, petroleum ether, hexane, or acetone.

9. The method for preparing the hemostatic medical adhesive as described in claim 7, characterized in that: The neutralizing agent is a reagent for neutralizing hydrophilic chain extenders.

10. The method for preparing the hemostatic medical adhesive as described in claim 9, characterized in that: The neutralizing agent is one or a mixture of several of the following: glacial acetic acid, phosphoric acid, hydrochloric acid, citric acid, or sulfuric acid.

11. The method for preparing the hemostatic medical adhesive as described in claim 7, characterized in that: The diluent is one or a mixture of several of isosorbide dimethyl ether, soybean oil methyl ester, triacetin, or triethylene glycol dimethyl ether.

12. The method for preparing the hemostatic medical adhesive according to any one of claims 7-11, characterized in that... In step (1), the reaction temperature is 50~80℃, the stirring speed is 80-120r / min, the amount of guanidyl carboxymethyl chitosan added is 2~30mg / ml, and the mass ratio of solvent to diisocyanate is 1.5:1~3.5:

1.

13. The method for preparing the hemostatic medical adhesive as described in claim 12, characterized in that... In step (2), the reaction temperature is 50~80℃, the stirring speed is 200~400r / min, and the amount of hydrophilic chain extender added is based on a molar ratio of isocyanate to hydroxyl group greater than 2.0:1.

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