Hyaluronic acid gel with abdominal cavity hemostasis and adhesion prevention functions as well as preparation method and application of hyaluronic acid gel

Abstract

The invention relates to hyaluronic acid gel with abdominal cavity hemostasis and adhesion prevention functions as well as a preparation method and application thereof, and relates to the technical field of biological medicines. The hyaluronic acid gel is prepared from the following raw materials: modified hyaluronic acid, metal salt and an amino high-molecular polymer, the modified hyaluronic acid comprises at least one of formylated hyaluronic acid, amidated hyaluronic acid, esterified hyaluronic acid and sulfhydrylated hyaluronic acid. Modified hyaluronic acid and an amino high-molecular compound are subjected to chemical crosslinking, metal ions in metal salt and carboxyl and hydroxyl in hyaluronic acid are coordinated to generate ionic crosslinking and chemical and physical double-channel crosslinking, and a formed three-dimensional structure is higher in crosslinking degree and better in mechanical strength and can be gradually degraded in vivo within 7-10 days.

Description

Technical Field

[0001] The present invention relates to the field of biomedical technologies, and particularly to a hyaluronic acid gel with abdominal hemostasis and anti-adhesion functions, a preparation method thereof, and an application thereof. Background Art

[0002] Postoperative adhesions generally refer to abnormal tissue hyperplasia caused by the deposition of connective tissue and fibrin around the wound after surgical operations, resulting in the adhesion of originally separated adjacent tissues or organs in a strip or sheet shape. Currently, common types of postoperative adhesions include abdominal adhesions, intrauterine adhesions, tendon adhesions, pericardial adhesions, and epidural adhesions, etc. Among them, abdominal adhesions are the most common type of adhesions. The harms of abdominal adhesions such as small intestinal obstruction, leading to symptoms such as chronic pain in the abdomen and pelvis, abnormal organ functions, etc., which will increase the economic burden on patients and the risk of future reoperations. Therefore, postoperative adhesions are a major problem that has to be faced in clinical surgical treatment. However, there are still many deficiencies in current postoperative anti-adhesion preparations, such as inconvenient use, difficult to degrade in the body, short retention time in the body, etc. In view of this, the present invention provides a hyaluronic acid gel with abdominal hemostasis and anti-adhesion functions, a preparation method thereof, and an application thereof. Summary of the Invention

[0003] The technical problem to be solved by the present invention is to provide a hyaluronic acid gel with abdominal hemostasis and anti-adhesion functions, a preparation method thereof, and an application thereof. The purpose is to provide a hyaluronic acid gel with abdominal hemostasis and anti-adhesion functions that is safe, stable, has good biocompatibility, and non-toxic and harmless degradation products.

[0004] The technical solution of the present invention to solve the above technical problems is as follows:

[0005] In the first aspect, a hyaluronic acid gel with abdominal hemostasis and anti-adhesion functions, the raw materials of the hyaluronic acid gel include: modified hyaluronic acid, metal salts, and amino high molecular polymers;

[0006] The modified hyaluronic acid includes at least one of aldehyde-modified hyaluronic acid, amide-modified hyaluronic acid, ester-modified hyaluronic acid, and mercapto-modified hyaluronic acid;

[0007] The metal salts include at least one of calcium salts, magnesium salts, zinc salts, iron salts, and barium salts.

[0008] Among them, the hyaluronic acid gel of the present invention forms a first network through chemical cross-linking of modified hyaluronic acid and amino high molecular polymers, and carboxyl groups, hydroxyl groups, etc. on the chain have coordination interactions with metal ions in the metal salts, and form a second network through ionic cross-linking. The two layers of networks interpenetrate each other to form a physical and chemical cross-linked double-network composite hyaluronic acid gel.

[0009] The beneficial effects of the present invention are:

[0010] (1) The hyaluronic acid gel of the present invention is an in-situ gel, which is easy to inject and can adaptively tamponade the wound for hemostasis after injection; the hyaluronic acid gel has a fast gelation rate and will quickly adhere to the wound surface when acting on the wound without falling off; the hyaluronic acid gel physically isolates the wound from the external environment during the wound healing process to prevent wound infection; the hyaluronic acid gel plays a lubricating role during the wound healing process, reducing friction with other organs and preventing adhesion.

[0011] (2) The present invention uses chemically modified hyaluronic acid and an amino polymer compound to undergo chemical cross-linking, and at the same time, metal ions in the metal salt coordinate with carboxyl groups, hydroxyl groups, etc. in hyaluronic acid to undergo ionic cross-linking. The double-channel cross-linking of chemistry and physics forms a three-dimensional structure with a higher cross-linking degree and better mechanical strength;

[0012] (3) The raw materials of the present invention are simple and easy to obtain, safe, and have good biocompatibility, and the hyaluronic acid gel will gradually degrade within 7-10 days in vivo.

[0013] Further, the calcium salt includes at least one of calcium lactate, barium chloride, calcium dihydrogen phosphate, calcium bicarbonate, and calcium bisulfate; the magnesium salt includes at least one of magnesium chloride, magnesium sulfate, and magnesium nitrate; the zinc salt includes at least one of zinc chloride and zinc sulfate;

[0014] The iron salt includes at least one of ferric chloride, ferric sulfate, and ferric dihydrogen phosphate; the barium salt includes at least one of barium sulfate, barium carbonate, barium chloride, barium sulfide, and barium nitrate.

[0015] The beneficial effect of adopting the above further scheme is that an appropriate amount of metal ions is helpful for obtaining a composite gel with good performance to prevent adhesion after surgery and scavenge free radicals to promote wound healing.

[0016] Further, the amino polymer includes at least one of gelatin (derived from pigskin or fish skin), chitosan, carboxymethyl chitosan, polyethyleneimine, amino-modified polyethylene glycol, and polylysine. Preferably, one or two of branched polyethyleneimine, gelatin, and carboxymethyl chitosan.

[0017] The beneficial effect of adopting the above further scheme is that the amino polymer is degradable, has mild reaction conditions, can react with modified hyaluronic acid at room temperature; has a fast cross-linking speed, good biological safety, and few usage restrictions.

[0018] Further, the mass ratio of the modified hyaluronic acid, the metal salt, and the amino polymer is (0.1-20):(0.05-5):(0.1-20);

[0019] Or / and, the solid content of the hyaluronic acid gel is 0.5-30 wt%.

[0020] The beneficial effects of adopting the above further solution are as follows: The hyaluronic acid gel within this range has good adhesiveness and can form a gel in situ rapidly.

[0021] In a second aspect, a preparation method of a hyaluronic acid gel with abdominal hemostasis and anti-adhesion functions includes the following steps: Dissolve the modified hyaluronic acid in a first solvent to obtain a first component; dissolve the amino polymer and the metal salt in a second solvent to obtain a second component; and form a gel in situ with the first component and the second component.

[0022] The beneficial effects of adopting the above solution are as follows: The preparation method of the hyaluronic acid gel with abdominal hemostasis and anti-adhesion functions of the present invention is simple and stable in operation, plays an auxiliary role in the self-repair process of the human body, and has no unknown side effects in drug use.

[0023] Furthermore, when dissolving the modified hyaluronic acid in a solvent, or when dissolving the amino polymer and the metal salt in a solvent, a stirring and dissolving method can be adopted. The stirring and dissolving method specifically includes at least one of stirring and dissolving at room temperature, stirring and dissolving at 40 - 60°C, mechanical stirring and dissolving, vortex mixing and dissolving, and ultrasonic dissolving; preferably, at least one of stirring and dissolving at room temperature, stirring and dissolving at 40 - 60°C, and vortex mixing and dissolving.

[0024] Further, when the modified hyaluronic acid is aldehyde-group modified hyaluronic acid, the preparation method of the aldehyde-group modified hyaluronic acid is as follows: Dissolve the hyaluronic acid in water to obtain a hyaluronic acid solution; dissolve sodium periodate in water to prepare a sodium periodate solution, and dropwise add the sodium periodate solution into the hyaluronic acid solution, and carry out the reaction in the dark at room temperature, then add ethylene glycol to terminate the reaction, and after dialysis and freeze-drying, obtain the aldehyde-group modified hyaluronic acid;

[0025] Among them, the dialysis is carried out using a dialysis bag with a molecular cut-off of 6000 - 8000D; the purpose of dialysis is to remove excess salt ions to obtain pure modified hyaluronic acid;

[0026] Or / and, when the modified hyaluronic acid is thiol-group modified hyaluronic acid, the preparation method of the thiol-group modified hyaluronic acid is as follows: Dissolve the hyaluronic acid in water to obtain a hyaluronic acid solution; add 1-ethyl-3-(3-dimethylaminopropyl)carbodiimide (EDC) and 1-hydroxybenzotriazole (HOBt) to the hyaluronic acid solution to activate the carboxyl group, and then add cystamine dihydrochloride to carry out the reaction at room temperature to obtain cystamine-conjugated HA (HA-cys); add dithiothreitol (DTT) to the cystamine-conjugated HA and cleave the disulfide bond at room temperature, and purify to obtain the thiol-group modified hyaluronic acid (HA-SH);

[0027] Or / and, when the modified hyaluronic acid is an esterified modified hyaluronic acid, the preparation method of the esterified modified hyaluronic acid is as follows: Dissolve hyaluronic acid in water to obtain a hyaluronic acid solution; add N-hydroxysuccinimide (NHS) and 1-ethyl-(3-dimethylaminopropyl) carbodiimide hydrochloride (EDC) to the hyaluronic acid solution for reaction, purify and dry to obtain the esterified modified hyaluronic acid;

[0028] Or / and, when the modified hyaluronic acid is an amidated hyaluronic acid, the preparation method of the amidated hyaluronic acid is as follows: React hyaluronic acid with methacrylic acid under ice bath conditions, then adjust the pH to 8-9, and precipitate in an alcoholic solution to obtain a product precipitate; Dialyze the product precipitate to obtain the amidated hyaluronic acid.

[0029] Furthermore, in the preparation of the aldehyde group-modified hyaluronic acid, the mass content of the hyaluronic acid solution is 1-5%; the mass content of the sodium periodate solution is 0.1-10%; the molar ratio of the hyaluronic acid to the sodium periodate is 1:(0.5-2); the oxidation degree of the aldehyde group-modified hyaluronic acid is 20-80%;

[0030] Or / and, in the preparation of the thiolated hyaluronic acid, the mass ratio of the hyaluronic acid, the EDC and the HOBt is 1:(0.5-10):(0.5-10); the mass ratio of the hyaluronic acid to the cystamine dihydrochloride is 1:(0.5-10):(0.5-10); the DTT accounts for 3-8 times the mass of the cystamine-conjugated HA;

[0031] Or / and, in the preparation of the esterified modified hyaluronic acid, the mass ratio of the hyaluronic acid, the NHS and the EDC is 1:(0.5-10):(0.5-10);

[0032] Or / and, in the preparation of the amidated hyaluronic acid, the mass-to-volume ratio of the hyaluronic acid to the methacrylic acid is 1 g:(2-6) mL.

[0033] Furthermore, the solvent in the first component is at least one of secondary distilled water, ultrapure water, deionized water, phosphate buffer solution, bicarbonate buffer solution, and borate buffer solution;

[0034] Or / and, the solvent in the second component is at least one of secondary distilled water, ultrapure water, deionized water, phosphate buffer solution, bicarbonate buffer solution, and borate buffer solution.

[0035] Furthermore, the mass content of the modified hyaluronic acid in the first component is 0.1-20%;

[0036] Alternatively, and / or, the mass content of the amino polymer in the second component is 0.1-20%, and the mass content of the metal salt is 0.05-10%.

[0037] Alternatively, and / or, the volume ratio of the first component to the second component is 1:(0.1-20).

[0038] The beneficial effect of adopting the above further scheme is that the modified hyaluronic acid, the amino polymer and the metal ions undergo moderate crosslinking at specific mass contents to construct a hyaluronic acid gel with both adhesiveness and continuous fluidity.

[0039] In a third aspect, the application of the hyaluronic acid gel with peritoneal hemostasis and anti-adhesion in the preparation of products for peritoneal hemostasis and anti-adhesion.

[0040] Among them, when the hyaluronic acid gel with peritoneal hemostasis and anti-adhesion is used, each component is independently packed in a double-barrel syringe and injected at the peritoneal bleeding site through a premixing head. The hyaluronic acid gel adheres to the surface of the wound, and the hemostatic effect is achieved through instant physical occlusion and stuffing. At the same time, the hyaluronic acid gel has a lubricating effect, reducing the friction between the wound surface and the outside world to achieve the anti-adhesion effect. Specifically, after the surgical operation is completed, the flowing gel is injected into the peritoneal wound surface by using a syringe and injected in a relatively low-viscosity state through in-situ injection, and rapidly forms a gel state at body temperature, which can not only stop bleeding but also adapt to wounds of different shapes and sizes; since the hyaluronic acid gel formed by the crosslinking reaction is a reaction between modified hyaluronic acid (aldehyde hyaluronic acid, amidated hyaluronic acid, esterified hyaluronic acid, mercapto hyaluronic acid) and amino groups, the formed hyaluronic acid gel slowly degrades in the peritoneal cavity, and finally the gel state gradually changes into a flowing liquid state and is naturally metabolized and excreted from the body in accordance with physiological conditions.

[0041] Furthermore, the hyaluronic acid gel with peritoneal hemostasis and anti-adhesion is used to prepare hyaluronic acid gels with peritoneal hemostasis and anti-adhesion for animals such as humans, mammals, birds, reptiles, etc. Description of the Drawings

[0042] Figure 1 This is a diagram of the hyaluronic acid gel prepared in Example 1 of the present invention. The left is the state diagram after gel formation, and the right is the state of injecting the hyaluronic acid gel into the cecal wound of a rat;

[0043] Figure 2 This is a comparison diagram of the nuclear magnetic hydrogen spectra of hyaluronic acid and modified hyaluronic acid of the present invention, where a is HA, b is OHA, c is HAMA, and d is HA-NHS;

[0044] Figure 3 This is a diagram of the in vitro anti-adhesion experiment of the hyaluronic acid gels of Examples 1, 4, 6 and Comparative Example 3 of the present invention;

[0045] Figure 4 Comparison of abdominal cavity adhesion prevention on the 7th day for Examples 1, 4, and 6 of the present invention and Comparative Example 3

[0046] Figure 5 Experimental result graph of in vitro degradation experiment for Example 1 of the present invention Detailed implementation manners

[0047] The principles and features of the present invention are described below. The examples given are only used to explain the present invention and are not intended to limit the scope of the present invention. For those without specific technical or conditions noted in the examples, the techniques or conditions described in the literature in this field or according to the product specifications are followed. For reagents or instruments without the manufacturer noted, they are all conventional products that can be purchased through regular channels.

[0048] Material description:

[0049] Hyaluronic acid (HA, molecular weight 400000 da - 3000000 da) was purchased from Macklin; sodium periodate was purchased from Macklin; N-hydroxysuccinimide (NHS) was purchased from Macklin, 1-ethyl-(3-dimethylaminopropyl)carbodiimide hydrochloride (EDC) was purchased from Macklin, gelatin (derived from pigskin or fish skin) was purchased from Sigma; chitosan was purchased from Qingdao Haidefeng; carboxymethyl chitosan was purchased from Qingdao Haidefeng; polyethyleneimine was purchased from Macklin; 1-hydroxybenzotriazole (HOBt) was from Macklin; cell culture medium was purchased from Solarbio; cells were purchased from the Chinese Cell Bank.

[0050] Example 1

[0051] This example relates to a preparation method of a hyaluronic acid gel with abdominal cavity hemostasis and adhesion prevention, including the following steps:

[0052] A 1% aqueous solution of hyaluronic acid and a 2% aqueous solution of sodium periodate, with a volume ratio of 1:0.5 between the aqueous solution of hyaluronic acid and the aqueous solution of sodium periodate, are subjected to an oxidation reaction for 16 hours under lightless conditions, and the reaction is terminated with excessive ethylene glycol to obtain a mixture; the mixture is successively dialyzed and freeze-dried, and the dialysis uses a dialysis bag with a molecular cut-off of 6000 - 8000 D to obtain aldehyde-functionalized hyaluronic acid. The aldehyde-functionalized hyaluronic acid is dissolved in water to prepare a first component with a mass content of 5%; carboxymethyl chitosan and MgCl2 are dissolved in water to prepare a second component with mass contents of 5% and 1%, and the first component and the second component with a volume ratio of 1:0.2 are mixed through a premixing head for 15 s at room temperature to obtain the hyaluronic acid gel with abdominal cavity hemostasis and adhesion prevention( Figure 1 ).

[0053] Example 2

[0054] This embodiment relates to a method for preparing a hemostatic and wound-healing promoting hyaluronic acid gel, comprising the following steps:

[0055] Dissolve hyaluronic acid in water to prepare a solution with a mass content of 1%, add NHS and EDC, and the molar ratio of the hyaluronic acid, the NHS, and the EDC is 1:1.2:1.2. Stir at room temperature for 16 h, add ethanol for precipitation, redissolve the precipitate in water, and freeze-dry to obtain esterified modified hyaluronic acid. Dissolve the esterified modified hyaluronic acid in a PBS solution to prepare a first component with a mass content of 4%; dissolve carboxymethyl chitosan and calcium lactate in water to prepare a second component with mass contents of 2% and 0.4%, and mix the first component and the second component with a volume ratio of 1:0.3 through a premixing head for 35 s at room temperature to obtain the hyaluronic acid gel with intraperitoneal hemostatic and anti-adhesion effects.

[0056] Example 3

[0057] This embodiment relates to a method for preparing a hemostatic and wound-healing promoting hyaluronic acid gel, comprising the following steps:

[0058] Dissolve hyaluronic acid in water to prepare a solution with a mass content of 1%, add EDC and 1-hydroxybenzotriazole (HOBt), and the molar ratio of the hyaluronic acid, the EDC, and the HOBt is 1:1.2:1.2. Activate the carboxyl group by stirring at room temperature for 2 h, add cystamine dihydrochloride and stir overnight at room temperature, and the mass ratio of the hyaluronic acid to the cystamine dihydrochloride is 1:2 to obtain cystamine-conjugated HA-cys; dialyze the reaction solution with deionized water to remove unreacted EDC, HOBt, and cystamine dihydrochloride. Treat HA-cys with 5-fold excess of DTT at room temperature to cleave the disulfide bond in water for 4 h. After the reaction is completed, adjust the solution to pH 3.5, add NaCl to a final mass content of 5%, and then, thiolated HA precipitates from ethanol, filter to obtain a white precipitate, redissolve the precipitate in water, filter and freeze-dry to obtain thiolated modified hyaluronic acid. Dissolve the thiolated modified hyaluronic acid in water to prepare a first component with a mass content of 4%; dissolve gelatin and calcium chloride in water to prepare a second component with mass contents of 0.5% and 0.5%, and mix the first component and the second component with a volume ratio of 1:0.2 through a premixing head for 15 s at room temperature to obtain the hyaluronic acid gel with intraperitoneal hemostatic and anti-adhesion effects.

[0059] Example 4

[0060] This embodiment relates to a method for preparing a hyaluronic acid gel with intraperitoneal hemostatic and anti-adhesion effects, comprising the following steps:

[0061] The same method as described in Example 1 was adopted to prepare aldehyde - modified hyaluronic acid. The aldehyde - modified hyaluronic acid was dissolved in water to prepare a first component with a mass content of 2%; the amino - modified four - arm polyethylene glycol derivative (4 - arm - PEG - NH2) and MgSO4 were dissolved in water to prepare a second component with mass contents of 2% and 0.5% (w / v). The first component and the second component with a volume ratio of 1:0.2 were mixed through a premixing head, and the hyaluronic acid gel with abdominal hemostasis and anti - adhesion effects could be obtained at room temperature in 35 s.

[0062] Example 5

[0063] Compared with Example 1, except that MgCl2 was replaced by ferric chloride; the rest was the same as Example 1.

[0064] Example 6

[0065] This example relates to a preparation method of a hyaluronic acid gel with abdominal hemostasis and anti - adhesion effects, including the following steps:

[0066] Hyaluronic acid with a mass content of 1% was dissolved in 100 mL of deionized water and stirred at room temperature until completely dissolved. 3 mL of methacrylic acid was slowly dropped into it in an ice - water bath, and a non - light reaction was carried out for 24 hours. After the reaction was completed, the pH was adjusted to 8 - 9 with 1M NaOH aqueous solution, and then 3 times the volume of pre - cooled absolute ethanol was added for product precipitation. After precipitation, the supernatant was removed, and the precipitate was centrifuged at 5000 rpm for 10 minutes in a high - speed centrifuge to obtain the crude product of HAMA. Finally, the crude product was dissolved in water, and after dialysis and freeze - drying, amidated hyaluronic acid was obtained. The amidated hyaluronic acid was dissolved in water to prepare a first component with a mass content of 5%; Lap and MgCl2 were dissolved in water to prepare a second component with mass contents of 0.15% and 0.5%. The first component and the second component with a volume ratio of 1:0.5 were premixed and irradiated under UV at 280 nm, and the hyaluronic acid gel with abdominal hemostasis and anti - adhesion effects was obtained at room temperature in 15 s.

[0067] Comparative Example 1:

[0068] In this comparative example, except that calcium lactate was not added, other conditions were the same as those in Example 2. The first component and the second component were mixed through a premixing head, and gelation occurred at room temperature in 1.5 min. Since the gelation speed was slow and it could not gel in situ at the wound surface, no hemostasis and anti - adhesion effects were observed.

[0069] Comparative Example 2

[0070] This example relates to a preparation method of a hyaluronic acid gel with abdominal hemostasis and anti - adhesion effects, including the following steps:

[0071] Using the same method as described in Example 1, aldehyde-modified hyaluronic acid was prepared. The aldehyde-modified hyaluronic acid was dissolved in water to prepare a first component with a mass content of 0.1%; carboxymethyl chitosan and MgCl2 were dissolved in water to prepare a second component with mass contents of 0.1% and 1%, respectively. The first component and the second component were mixed through a premixing head in the same ratio as described in Example 1. Since the solid content was low, gelation could not occur within 5 minutes at room temperature, so there was no hemostatic or anti-adhesion effect.

[0072] Comparative Example 3

[0073] In this comparative example, except that the hyaluronic acid in Component A was not modified, other conditions were the same as those in Example 1. The first component and the second component were mixed through a premixing head. Since the unmodified hyaluronic acid did not have groups for cross-linking with amino polymers, chemical cross-linking could not occur, and gelation could not occur within 5 minutes at room temperature, so there was no hemostatic or anti-adhesion effect.

[0074] Comparative Example 4

[0075] Compared with Example 1, except that MgCl2 was not added, the rest was the same as in Example 1.

[0076] Regarding Examples 1-6 and Comparative Examples 1-4, the comparison of the properties of hyaluronic acid gels under different formulations is shown in Table 1.

[0077] Table 1

[0078]

[0079]

[0080] It can be seen from the comparison between the examples and the comparative examples in Table 1 that the present invention selects modified hyaluronic acid to prepare hyaluronic acid gel and adds metal ions at a certain concentration, which can effectively improve the gelation speed of hyaluronic acid gel and its hemostatic and anti-adhesion effects.

[0081] Test Example

[0082] (1) 1H NMR spectra of the raw material hyaluronic acid (HA) and the prepared OHA.

[0083] Using the method for preparing hyaluronic acid gel, the aldehyde-modified hyaluronic acid (OHA) prepared by modifying the raw material hyaluronic acid (HA) was dissolved in deuterated water, and the 1H NMR spectrum was scanned. The results are as Figure 2 shown.

[0084] From Figure 2 it can be concluded that the HA was oxidized by sodium periodate at a certain temperature and time, Figure 2 b a new peak appeared at 4.9 - 5.3 ppm, and it can be judged that OHA was prepared, and the oxidation degree was about 36%; Figure 2A new double bond characteristic peak appears at 5.5 - 6.5 ppm for c, from which it can be judged that HAMA is prepared. Figure 2 A NHS ester characteristic peak appears at 2.5 - 3 ppm for d, from which it can be judged that HA - NHS is prepared.

[0085] (2) In vitro anti - adhesion experiment of hyaluronic acid gel.

[0086] Place the hyaluronic acid gels of Comparative Example 3 and Examples 1, 4, and 6 in a culture dish, add cell culture medium and cells for culture. Evaluate the anti - adhesion effect by observing the adhesion of cells on the surface of the hyaluronic acid gel.

[0087] From Figure 3 it can be obtained that compared with Comparative Example 4, Examples 1, 4, and 6 all have varying degrees of anti - cell adhesion effects and the effects are significant.

[0088] (3) Animal experiment on hemostasis and anti - adhesion in the abdominal cavity of SD rats.

[0089] Create a defect model in the cecum of SD rats, and apply the products of Example 1, Example 4, Example 6 and Comparative Example 3 to the modeled area respectively. Open the abdomen after 7 days to observe the anti - adhesion situation, as shown in Figure 4 . From Figure 4 it can be obtained that the unmodified hyaluronic acid in Comparative Example 3 has no anti - adhesion effect on the abdominal cavity of rats, while the hyaluronic acid gels of Example 1, Example 4, and Example 6 have significant hemostasis and anti - adhesion effects.

[0090] (4) In vitro degradation experiment.

[0091] Accurately weigh five portions of the hydrogel prepared in Example 1 with a mass of 200 mg into 50 mL centrifuge tubes, add 100 - fold volume of PBS aqueous solution with a pH value of about 7.4, and place each centrifuge tube into a constant - temperature shaking water bath at 37 °C with a shaking speed of 150 rpm / min for shaking degradation. Record the mass of the hydrogel every 24 h. Before weighing, it is necessary to absorb the moisture on the surface of the hydrogel with filter paper. The results are shown in Figure 5 . From Figure 5 it can be obtained that the hyaluronic acid gel of Example 1 is completely degraded in PBS solution in 6 days and no residue is produced.

[0092] In summary, the hyaluronic acid gel of the present invention is an in - situ gel, which is easy to inject and can adaptively tamponade the wound for hemostasis after injection; the hyaluronic acid gel has a fast gelation speed and will quickly adhere to the wound surface when acting on the wound and will not fall off; the hyaluronic acid gel physically isolates the wound from the external environment during the wound healing process to prevent wound infection; the hyaluronic acid gel plays a lubricating role during the wound healing process, reducing friction with other organs and preventing adhesion.

[0093] The present invention uses a modified hyaluronic acid complexed with an amino polymer compound. At the same time, metal ions in the metal salt coordinate with carboxyl groups, hydroxyl groups, etc. in the hyaluronic acid to undergo ionic crosslinking, achieving crosslinking through chemical and physical dual channels. The formed three-dimensional structure has a higher crosslinking degree and better mechanical strength. The raw materials of the present invention are simple and easily available, safe, have good biocompatibility, and are hyaluronic acid gels.

[0094] Although the embodiments of the present invention have been shown and described above, it can be understood that the above embodiments are exemplary and should not be construed as limiting the present invention. Those of ordinary skill in the art can make changes, modifications, substitutions, and variations to the above embodiments within the scope of the present invention.

Claims

1. A hyaluronic acid gel with abdominal hemostasis and anti-adhesion functions, characterized in that, The raw materials of the hyaluronic acid gel include: modified hyaluronic acid, metal salts, and amino polymer; The modified hyaluronic acid includes at least one of aldehyde group-modified hyaluronic acid, amidated hyaluronic acid, esterified hyaluronic acid, and mercapto-modified hyaluronic acid; The metal salts include at least one of calcium salts, magnesium salts, zinc salts, iron salts, and barium salts.

2. The hyaluronic acid gel with abdominal hemostasis and anti-adhesion according to claim 1, wherein, The calcium salts include at least one of calcium lactate, calcium chloride, calcium dihydrogen phosphate, calcium hydrogen carbonate, and calcium bisulfate; the magnesium salts include at least one of magnesium chloride, magnesium sulfate, and magnesium nitrate; the zinc salts include at least one of zinc chloride and zinc sulfate; the iron salts include at least one of ferric chloride, ferric sulfate, and ferrous dihydrogen phosphate; the barium salts include at least one of barium sulfate, barium carbonate, barium chloride, barium sulfide, and barium nitrate.

3. The hyaluronic acid gel with abdominal hemostasis and anti-adhesion according to claim 1, characterized in that The amino polymer includes at least one of gelatin, chitosan, carboxymethyl chitosan, polyethyleneimine, amino-modified polyethylene glycol, and polylysine.

4. The hyaluronic acid gel with abdominal hemostasis and anti-adhesion according to claim 1, wherein The mass ratio of the modified hyaluronic acid, the metal salt, and the amino polymer is (0.1-20):(0.05-10):(0.1-20); Or / and, the solid content of the hyaluronic acid gel is 0.5-30 wt%.

5. A preparation method of a hyaluronic acid gel with abdominal hemostasis and anti-adhesion according to any one of claims 1 to 4, characterized in that, It includes the following steps: Dissolve the modified hyaluronic acid in a first solvent to obtain a first component; dissolve the amino polymer and the metal salt in a second solvent to obtain a second component; and in-situ gel the first component and the second component.

6. The preparation method of a hyaluronic acid gel with abdominal hemostasis and anti-adhesion according to claim 5, characterized in that, When the modified hyaluronic acid is aldehyde group-modified hyaluronic acid, the preparation method of the aldehyde group-modified hyaluronic acid is as follows: dissolve hyaluronic acid in water to obtain a hyaluronic acid solution; dissolve sodium periodate in water to prepare a sodium periodate solution, and dropwise add the sodium periodate solution to the hyaluronic acid solution, react at room temperature in the dark, then add ethylene glycol to terminate the reaction, and obtain aldehyde group-modified hyaluronic acid through dialysis and freeze-drying; Or / and, when the modified hyaluronic acid is mercapto-modified hyaluronic acid, the preparation method of the mercapto-modified hyaluronic acid is as follows: dissolve hyaluronic acid in water to obtain a hyaluronic acid solution; add EDC and HOBt to the hyaluronic acid solution to activate the carboxyl group, then add cystamine dihydrochloride and react at room temperature to obtain cystamine-conjugated HA; add DTT to the cystamine-conjugated HA and cleave the disulfide bond at room temperature, and purify to obtain mercapto-modified hyaluronic acid; Or / and, when the modified hyaluronic acid is esterified modified hyaluronic acid, the preparation method of the esterified modified hyaluronic acid is as follows: dissolve hyaluronic acid in water to obtain a hyaluronic acid solution; add NHS and EDC to the hyaluronic acid solution and react at room temperature, then purify and dry to obtain esterified modified hyaluronic acid; Or / and, when the modified hyaluronic acid is amidated hyaluronic acid, the preparation method of the amidated hyaluronic acid is as follows: react hyaluronic acid with methacrylic acid under ice bath conditions, then adjust the pH to 8-9, and precipitate in an alcoholic solution to obtain a product precipitate; dialyze the product precipitate to obtain amidated hyaluronic acid.

7. The preparation method of a hyaluronic acid gel with abdominal hemostasis and anti-adhesion according to claim 6, characterized in that, In the preparation of the aldehyde-modified hyaluronic acid, the mass content of the hyaluronic acid solution is 1-5%; the mass content of the sodium periodate solution is 0.1-10%, and the molar ratio of the hyaluronic acid to the sodium periodate is 1:(0.5-2); the oxidation degree of the aldehyde-modified hyaluronic acid is 20-80%; Or / and, in the preparation of the thiol-modified hyaluronic acid, the molar ratio of the hyaluronic acid, the EDC and the HOBt is 1:(0.5-10):(0.5-10); the mass ratio of the hyaluronic acid to the cystamine dihydrochloride is 1:(0.5-10); the DTT accounts for 3-8 times the mass of the cystamine-conjugated HA; Or / and, in the preparation of the esterified modified hyaluronic acid, the mass ratio of the hyaluronic acid, the NHS and the EDC is 1:(0.5-10):(0.5-10); Or / and, in the preparation of the amidated hyaluronic acid, the mass-volume ratio of the hyaluronic acid to the methacrylic acid is 1 g:(2-6) mL.

8. The preparation method of a hyaluronic acid gel with abdominal hemostasis and anti-adhesion according to claim 5, characterized in that, The solvent in the first component is at least one of secondary distilled water, ultrapure water, deionized water, phosphate buffer solution, bicarbonate buffer solution, and borate buffer solution; Or / and, the solvent in the second component is at least one of secondary distilled water, ultrapure water, deionized water, phosphate buffer solution, bicarbonate buffer solution, and borate buffer solution.

9. The preparation method of a hyaluronic acid gel with abdominal hemostasis and anti-adhesion according to claim 5, characterized in that, The mass content of the modified hyaluronic acid in the first component is 0.1-20%; Or / and, the mass content of the amino polymer in the second component is 0.1-20%, and the mass content of the metal salt is 0.05-10%; Or / and, the volume ratio of the first component to the second component is 1:(0.1-20).

10. Use of the hyaluronic acid gel with peritoneal hemostasis and anti-adhesion according to any one of claims 1 to 4 in the preparation of a product for peritoneal hemostasis and anti-adhesion.

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