A biological tissue sealing material and its preparation method

Abstract

The present invention provides a tissue sealing material with good biocompatibility and capable of effectively and rapidly sealing a wound site. It is characterized in that the tissue sealing material is sponge-like and is prepared by segmented freeze-drying of aldehyde-functionalized natural polysaccharide. The present invention also provides a preparation method of the tissue sealing material, including the following steps: Step S1, dissolving the aldehyde-functionalized natural polysaccharide in water to prepare an aldehyde-functionalized natural polysaccharide solution with a concentration of 0.5% to 20%, and adjusting the pH to 7 to 9; Step S2, putting the aldehyde-functionalized natural polysaccharide solution into a freeze-dryer pre-cooled to -80°C to -40°C and maintaining it for 3 to 12 hours; Step S3, setting the vacuum degree of the freeze-dryer to 0.01 to 0.5 mbar and freeze-drying for 24 to 96 hours; Step S4, maintaining the vacuum degree, heating to 0°C at a rate of 5 to 15°C per hour, and then freeze-drying for 12 to 48 hours; Step S5, restoring the vacuum degree to normal pressure, heating to 30°C to 50°C at a rate of 10 to 20°C per hour, and maintaining it for 12 to 24 hours to obtain the product.

Description

Technical Field

[0001] The present invention belongs to the field of medical materials, and particularly relates to a biological tissue sealing material and a preparation method thereof. Background Art

[0002] Medical gauze and sponges are commonly used hemostatic and filling materials. Their mechanism of action is to absorb the moisture in the blood through their own water absorption ability, enrich coagulation factors to promote blood coagulation, or produce compressive hemostasis by their own expansion and interaction with surrounding soft tissues.

[0003] However, such materials are usually applicable to situations with small amounts of bleeding such as oozing blood, and cannot effectively stop bleeding for massive bleeding. Moreover, they rely on the patient's own blood coagulation and are ineffective for patients with coagulation dysfunction. At the same time, these materials cannot effectively seal the wound site. After application, continuous bleeding and tissue fluid leakage still occur at the wound. Summary of the Invention

[0004] Based on the above problems, the present invention provides a biological tissue sealing material with good biocompatibility and capable of effectively and rapidly sealing the wound site. The characteristic is that the biological tissue sealing material is in a sponge shape and is prepared by segmented freeze-drying of aldehyde group-modified natural polysaccharides.

[0005] The biological tissue sealing material provided by the present invention may also have the following technical feature: the pores in the biological tissue sealing material are 1 - 20 μm, preferably 10 - 20 μm.

[0006] The biological tissue sealing material provided by the present invention may also have the following technical feature: the water absorption of the biological tissue sealing material is less than 5 g / g. This biological tissue sealing material is not easily swollen by water absorption. When in use, it reacts and cross-links with amino groups / sulfhydryl groups on cells or proteins in body fluids or on the tissue surface to form a physical sealing layer, thereby closing the wound site.

[0007] The present invention also provides a preparation method of the biological tissue sealing material as described in any one of the above, characterized in that it is prepared by a segmented freeze-drying method, including the following steps:

[0008] Step S1: Dissolve the aldehyde group-modified natural polysaccharide in water to prepare an aldehyde group-modified natural polysaccharide solution with a concentration of 0.5% - 20%, and adjust the pH to 7 - 9;

[0009] Step S2: Place the aldehyde group-modified natural polysaccharide solution in a freeze-dryer pre-cooled to -80°C to -40°C and hold for 3 - 12 hours;

[0010] Step S3: Set the vacuum degree of the freeze-dryer to 0.01 - 0.5 mbar and freeze-dry for 24 - 96 hours;

[0011] Step S4: Maintain the vacuum degree, heat up to 0°C at a rate of 5 - 15°C per hour, and then freeze-dry for 12 - 48 hours;

[0012] Step S5: Restore the vacuum degree to normal pressure, heat up to 30 - 50°C at a rate of 10 - 20°C per hour, and maintain for 12 - 24 hours to obtain the biological tissue sealing material.

[0013] In the preparation method of the biological tissue sealing material provided by the present invention, the pre-cooling temperature in Step S2 is preferably -55°C to -45°C.

[0014] The preparation method of the biological tissue sealing material provided by the present invention may further have the following technical feature: the aldehyde-group modified natural polysaccharide is one or a combination of more than one of aldehyde-group modified hyaluronic acid, aldehyde-group modified dextran, aldehyde-group modified pullulan, aldehyde-group modified chitosan, and aldehyde-group modified carboxymethyl cellulose.

[0015] Function and Effect of the Invention

[0016] The biological tissue sealing material provided by the present invention is prepared by freeze-drying the aldehyde-group modified polysaccharide in segments. Therefore, it has a dense structure and small pores. Such a biological tissue sealing material is not easily water-absorbed and swollen. When in use, once it contacts the body fluid or the tissue surface, it can react and crosslink with the amino groups / sulfhydryl groups on the cells or proteins in the body fluid or on the tissue surface to form a physical sealing layer, thereby effectively sealing the wound site. Description of the Drawings

[0017] Figure 1 is a physical photograph of the biological tissue sealing material of Example 1 of the present invention;

[0018] Figure 2 is an electron microscope photograph of the biological tissue sealing material of Example 1 of the present invention;

[0019] Figure 3 is a physical photograph of the biological tissue sealing material of Comparative Example 1 of the present invention;

[0020] Figure 4 is an electron microscope photograph of the biological tissue sealing material of Comparative Example 1 of the present invention;

[0021] Figure 5 is a physical photograph of the biological tissue sealing material of Example 1 of the present invention after being soaked in pure water and polylysine solution for 30 minutes;

[0022] Figure 6 is a photograph of the hemostatic performance test of the biological tissue sealing material of Example 1 of the present invention;

[0023] Figure 7 is a photograph of the hemostatic performance test of the biological tissue sealing material of Example 1 of the present invention;

[0024] Figure 8 These are the photos of the hemostatic performance test of the biological tissue sealing material in Example 1 of the present invention;

[0025] Figure 9 These are the photos of the adhesion strength test of the biological tissue sealing material in Example 1 of the present invention in different organs;

[0026] Figure 10 These are the photos of the process of the adhesion strength test of the biological tissue sealing material in Example 1 of the present invention on the muscle surface;

[0027] Figure 11 These are the photos of the test results of the adhesion strength of the biological tissue sealing material in Example 1 of the present invention on the muscle surface. Detailed implementation manners

[0028] The following examples illustrate the biological tissue sealing material of the present invention and its preparation method by way of example. In the following examples, the reagent materials without specified sources are all commercially available as usual, and the operation methods without specified specific processes refer to the conventional operation methods of the prior art.

[0029] In the following examples, aldehyde - modified natural polysaccharides are used, specifically aldehyde - modified dextran, aldehyde - modified hyaluronic acid, and aldehyde - modified pullulan; the aldehyde - modification reactions of all three use sodium periodate. In addition, the molecular weights of dextran, hyaluronic acid, and pullulan in the present invention are 10,000 - 150,000, and the molecular weights used in the examples are all about 100,000.

[0030] The specific aldehyde - modification method is as follows:

[0031] Aldehyde - modified dextran: Dissolve 5 g of dextran in 100 mL of pure water, and stir until the dextran is completely dissolved. Weigh 10 g of sodium periodate and add it to a spherical flask, and stir the reaction overnight. Subsequently, place the solution in a 3 - L beaker and dialyze it with pure water for 3 days to remove small - molecule by - products, and then lyophilize the dialyzed solution to obtain aldehyde - modified dextran.

[0032] Aldehyde - modified hyaluronic acid: Dissolve 5 g of hyaluronic acid in 100 mL of pure water, and stir until the hyaluronic acid is completely dissolved. Weigh 10 g of sodium periodate and add it to a spherical flask, and stir the reaction overnight. Subsequently, place the solution in a 3 - L beaker and dialyze it with pure water for 3 days to remove small - molecule by - products, and then lyophilize the dialyzed solution to obtain aldehyde - modified hyaluronic acid.

[0033] Aldehyde - modified pullulan: Dissolve 2 g of hyaluronic acid in 100 mL of pure water, and stir until the hyaluronic acid is completely dissolved. Weigh 5 g of sodium periodate and add it to a spherical flask, and stir the reaction overnight. Subsequently, place the solution in a 3 - L beaker and dialyze it with pure water for 3 days to remove small - molecule by - products, and then lyophilize the dialyzed solution to obtain aldehyde - modified pullulan.

[0034] <Example 1>

[0035] The preparation method of the biological tissue sealing material in this example includes the following steps:

[0036] Step S1: Redissolve the obtained aldehyde - modified dextran in water to prepare an aldehyde - modified dextran solution with a concentration of 10%, and adjust the pH to 8.5 with NaOH;

[0037] Step S2: Directly put the solution prepared in Step S1 into a freeze - dryer pre - cooled to - 40 °C and keep it for 8 hours;

[0038] Step S3: Set the vacuum degree of the freeze - dryer to 0.5 mbar and freeze - dry for 48 hours;

[0039] Step S4: Maintain a vacuum degree of 0.5 mbar, heat up to 0 °C at a rate of 10 °C / hour, and then freeze - dry for 12 hours;

[0040] Step S5: Restore the vacuum degree to normal pressure, heat up to 50 °C at a rate of 10 °C / hour, and keep it for 12 hours to obtain the biological tissue sealing material of aldehyde - modified dextran.

[0041] Figure 1 is a physical photo of the biological tissue sealing material of Example 1 of the present invention, Figure 2 is an electron microscope photo of the biological tissue sealing material of Example 1 of the present invention.

[0042] As Figure 1 shown, the biological tissue sealing material of Example 1 is sponge - like, with a dense surface. When torn, it can be seen that the structure at the cross - section is also very dense, and there are no obvious pores visible to the naked eye.

[0043] As Figure 2 shown, in the microscopic structure of the biological tissue sealing material of Example 1, most of the pore structures are closed pores, arranged closely to each other, and the pores are all less than 10 μm, indicating that the internal structure of this biological tissue sealing material is also very dense.

[0044] <Example 2>

[0045] The preparation method of the biological tissue sealing material in this example includes the following steps:

[0046] Step S1: Redissolve the obtained aldehyde - modified hyaluronic acid in water to prepare an aldehyde - modified hyaluronic acid solution with a concentration of 3%, and adjust the pH to 7 with NaOH;

[0047] Step S2: Directly put the solution prepared in Step S1 into a freeze - dryer pre - cooled to - 80 °C and keep it for 3 hours;

[0048] Step S3: Set the vacuum degree of the freeze dryer to 0.01 mbar and freeze-dry for 72 hours;

[0049] Step S4: Maintain the vacuum degree of 0.01 mbar, heat up to 0 °C at a rate of 15 °C per hour, and then freeze-dry for 12 hours;

[0050] Step S5: Restore the vacuum degree to normal pressure, heat up to 40 °C at a rate of 20 °C per hour, and hold for 24 hours to obtain the biological tissue sealing material of aldehyde-group modified hyaluronic acid.

[0051] Similar to Example 1, the biological tissue sealing material of this example is also sponge-like, with a dense surface. When torn open, the structure at the cross-section is also very dense, and there are no obvious pores visible to the naked eye.

[0052] <Example 3>

[0053] The preparation method of the biological tissue sealing material of this example includes the following steps:

[0054] Step S1: Redissolve the obtained aldehyde-group modified pullulan in water to prepare an aldehyde-group modified pullulan solution with a concentration of 0.5%, and adjust the pH to 9 with NaOH;

[0055] Step S2: Directly put the solution prepared in Step S1 into a freeze dryer pre-cooled to -50 °C and hold for 12 hours;

[0056] Step S3: Set the vacuum degree of the freeze dryer to 0.1 mbar and freeze-dry for 96 hours;

[0057] Step S4: Maintain the vacuum degree of 0.1 mbar, heat up to 0 °C at a rate of 5 °C per hour, and then freeze-dry for 24 hours;

[0058] Step S5: Restore the vacuum degree to normal pressure, heat up to 30 °C at a rate of 15 °C per hour, and hold for 18 hours to obtain the biological tissue sealing material of aldehyde-group modified pullulan.

[0059] Similar to Example 1 and Example 2, the biological tissue sealing material of this example is also sponge-like, with a dense surface. When torn open, the structure at the cross-section is also very dense, and there are no obvious pores visible to the naked eye.

[0060] <Comparative Example 1>

[0061] This comparative example provides another material prepared from aldehyde-group modified dextran, and its preparation method is as follows:

[0062] The obtained aldehyde - modified dextran was redissolved in water to prepare an aldehyde - modified dextran solution with a concentration of 10%. The prepared solution was directly placed in a freeze - dryer pre - cooled to - 40°C and maintained for 8 hours. Subsequently, the vacuum was set to 0.5 mbar and freeze - dried for 48 hours to obtain the product.

[0063] Figure 3 This is a physical photograph of the aldehyde - modified dextran material of Comparative Example 1 of the present invention. Figure 4 This is an electron microscope photograph of the aldehyde - modified dextran material of Comparative Example 1 of the present invention.

[0064] As Figure 3 shown, the obtained aldehyde - modified dextran material of Comparative Example 1 has obvious pores visible to the naked eye on the surface and obvious cracks.

[0065] As Figure 4 shown, in the aldehyde - modified dextran material of Comparative Example 1, most of the pore structures are open pores, arranged relatively loosely and irregularly, and the pores are all larger than 50 μm.

[0066] <Comparative Example 2>

[0067] This comparative example provides another material prepared from aldehyde - modified hyaluronic acid, and its preparation method is as follows:

[0068] The obtained aldehyde - modified hyaluronic acid was redissolved in water to prepare an aldehyde - modified hyaluronic acid solution with a concentration of 3%. The prepared solution was directly placed in a freeze - dryer pre - cooled to - 60°C and maintained for 3 hours. Subsequently, the vacuum was set to 0.01 mbar and freeze - dried for 72 hours to obtain the product.

[0069] The obtained aldehyde - modified hyaluronic acid material of this comparative example is similar to that of Comparative Example 1, with obvious pores visible to the naked eye on the surface and obvious cracks.

[0070] <Comparative Example 3>

[0071] This comparative example provides another material prepared from aldehyde - modified pullulan, and its preparation method is as follows:

[0072] The obtained aldehyde - modified pullulan was redissolved in water to prepare an aldehyde - modified pullulan solution with a concentration of 0.5%. The prepared solution was directly placed in a freeze - dryer pre - cooled to - 50°C and maintained for 12 hours. Subsequently, the vacuum was set to 0.1 mbar and freeze - dried for 92 hours to obtain the product.

[0073] The obtained aldehyde - modified pullulan material of this comparative example is similar to those of Comparative Example 1 and Comparative Example 2, with obvious pores visible to the naked eye on the surface and obvious cracks.

[0074] <Test Example 1>

[0075] This test example is the crosslinking test of the materials obtained in Example 1 and Comparative Example 1.

[0076] The specific test method is as follows: Take 0.05 g of the materials of Example 1 and Comparative Example 1, and immerse them in 1.5 ml of pure water and 10% polylysine solution respectively for 30 minutes.

[0077] Figure 5 It is a physical picture of the biological tissue sealing material of Example 1 of the present invention after being immersed in pure water and polylysine solution for 30 minutes. Among them, the left side is pure water and the right side is polylysine solution.

[0078] Upon observation, the biological tissue sealing material in Example 1 can completely dissolve in water within 20 minutes, but will not dissolve in 10% polylysine solution. As Figure 5 shown, after soaking for 30 minutes, the material in pure water has dissolved and is in a uniform state as a whole, while a gel-like object (within the dotted line frame in the figure) remains in the polylysine solution.

[0079] The reason why the biological tissue sealing material does not dissolve in polylysine solution is that the aldehyde groups in the material react chemically with the amino groups and / or mercapto groups on polylysine, resulting in crosslinking of the biological tissue sealing material.

[0080] In addition, upon observation, the aldehyde dextran material of Comparative Example 1 can completely dissolve in water within 5 minutes, but it will also crosslink into a gel and will not dissolve in 10% polylysine solution.

[0081] Take out the sample soaked in polylysine solution for 30 minutes for weighing test, and calculate the water absorption per g of the material by comparing the total weight difference before and after soaking. The water absorption of the material in Example 1 is 1.1 g / g, and the water absorption of the material in Comparative Example 1 is 26 g / g.

[0082] <Test Example 2>

[0083] This test example is the crosslinking water absorption test of the materials obtained in each example and comparative example.

[0084] The specific test method is as follows: Take 0.05 g of biological tissue sealing material, immerse it in an excessive amount of 10% polylysine solution, take it out and weigh it after 24 hours, and calculate the water absorption per g of the material according to the total weight difference before and after soaking. The results are as follows:

[0085] Table 1 Water absorption of biological tissue sealing materials in examples and comparative examples

[0086] Group Water absorption (g / g) Example 1 1.2 Example 2 3.1 Example 3 1.8 Comparative Example 1 54 Comparative Example 2 77 Comparative Example 3 36

[0087] It can be seen that the water absorption of the biological tissue sealing materials in Examples 1 to 3 is relatively low, with the water absorption per gram of the material being 1.2 to 3.1 g, that is, the water absorption of the biological tissue sealing materials in the examples is about 1 to 3 times their own weight.

[0088] In contrast, the water absorption of the three aldehyde - modified polysaccharide materials in Comparative Examples 1 to 3 is very high, with the water absorption per gram of the material ranging from 36 to 77 g; that is to say, the water absorption of the materials in the comparative examples reaches more than 30 times their own weight, which is also the water absorption range of common hemostatic biological materials such as gelatin sponges.

[0089] <Test Example 3>

[0090] This test example is for the hemostatic performance test of the materials obtained in Example 1 and Comparative Example 1.

[0091] The specific test method is as follows: The hemostatic performance test is carried out in a model of massive hemorrhage caused by transverse cutting of the femoral artery of a 2.5 - kg New Zealand white rabbit. After anesthetizing the rabbit, a 1 - cm incision is made at the femoral artery of the rabbit with a scalpel to cut off the femoral artery; then, 0.2 g of the prepared material is quickly pressed on the bleeding wound, kept pressed for a certain time, and then released.

[0092] Figure 6 This is a photo of the hemostatic performance test of the biological tissue sealing material of Example 1 of the present invention. Among them, the left side shows the state just after the femoral artery is cut, and the right side shows the state after the biological tissue sealing material is released after being pressed for 30 seconds.

[0093] As Figure 6 shown, after cutting the femoral artery, a large amount of blood can be seen gushing out rapidly; when the biological tissue sealing material is quickly pressed on the bleeding wound and then released, it can be seen that there is no blood flowing out at the incision of the femoral artery, and the surface of the biological tissue sealing material is not stained red by blood, proving that the sponge - like aldehyde - modified dextran biological tissue sealing material obtained in Example 1 has excellent tissue adhesion and sealing performance.

[0094] Figure 7 This is a photo of the hemostatic performance test of the aldehyde - modified dextran material of Comparative Example 1 of the present invention. Among them, the left side shows the state just after the femoral artery is cut, and the right side shows the state after being released after pressing for 3 minutes.

[0095] As Figure 7 shown, after cutting the femoral artery, a large amount of blood can be seen gushing out rapidly; when the aldehyde - modified dextran material is quickly pressed on the bleeding wound and pressed for 3 minutes, after release, it can be seen that the material is completely stained red by blood and swells due to blood absorption, and blood quickly gushes out from under the material and the surface of the material, indicating that the material obtained in Comparative Example 1 cannot block the massive hemorrhage of the main rabbit's femoral artery and cannot achieve rapid hemostasis.

[0096] <Test Example 4>

[0097] This test example is a lymphatic fistula occlusion test for the materials obtained in Example 2 and Comparative Example 2.

[0098] The specific test method is as follows: After anesthetizing the rabbit, find the lymphatic vessel of the rabbit, create a 2-mm diameter notch, and white chylous fluid flows out; press 0.2 g of the biological tissue sealing material on the notch, maintain the pressing for a certain period of time, and then release it to observe the wound condition. Then, suture the wound and place a drainage tube to observe the liquid drainage condition within 3 days.

[0099] After testing and observation, when 0.2 g of the biological tissue sealing material of Example 2 was pressed on the notch and released after maintaining the pressing for 15 minutes, and continued to observe, no liquid flowed out again within 15 minutes. After suturing and placing the drainage tube, basically no liquid drained out within 3 days, indicating that the biological tissue sealing material obtained in Example 2 has excellent tissue adhesion and sealing performance.

[0100] In addition, when 0.2 g of the aldehyde-modified hyaluronic acid material of Comparative Example 2 was pressed on the notch and released after maintaining the pressing for 15 minutes, and continued to observe, it was visible that the material significantly swelled by absorbing water and white chylous fluid slowly flowed out. After suturing and placing the drainage tube, obvious white chylous fluid drained out in the first two days, indicating that the tissue adhesion and sealing performance of the material obtained in Comparative Example 2 is limited.

[0101] <Test Example 5>

[0102] This test example is a hemostatic performance test for the material obtained in Example 1.

[0103] The specific test method is as follows: A major hemorrhage hemostatic performance test is carried out in a femoral artery injury and bleeding model of a 40-kg Bama miniature pig. After anesthetizing the pig, make a 0.5-cm long incision at the femoral artery with a scalpel; then, quickly press the prepared material with an area of 2 cm * 2 cm on the bleeding wound, maintain the pressing for a certain period of time, and then release it.

[0104] Figure 8 It is a photo of the hemostatic performance test of the biological tissue sealing material of Example 1 of the present invention. Among them, the upper side is the just-cut state, and the lower side is the state after the biological tissue sealing material is pressed and released for 30 seconds.

[0105] As Figure 8 shown, after cutting, a clear wound end face (the part pointed by the arrow in the photo) can be seen, and a large number of blood columns are ejected; when the biological tissue sealing material is quickly pressed on the wound and continuously pressed, no blood flows out at the femoral artery incision after release, and the surface of the biological tissue sealing material is not stained red by blood, proving that the sponge-like aldehyde-modified dextran biological tissue sealing material obtained in Example 1 has excellent tissue adhesion and sealing performance.

[0106] <Test Example 6>

[0107] This test example is for testing the adhesion strength of the material obtained in Example 1.

[0108] The test method is as follows: Select fresh rat organs (including heart, liver, spleen, lung, kidney, stomach, small intestine, and tail) without wiping blood or tissue fluid, immediately press a biological tissue sealing material with a size of about 1 cm × 1 cm quickly on the organ surface and continuously press for 30 s, and after releasing, clamp the biological tissue sealing material with forceps and lift it up.

[0109] Figure 9 These are the photos of the adhesion strength test of the biological tissue sealing material of Example 1 of the present invention in different organs.

[0110] As Figure 9 shown, the biological tissue sealing material of the present invention can adhere to the fresh tissue surfaces of various organs such as heart, liver, spleen, lung, kidney, stomach, small intestine, and tail, and even when the material is clamped and the whole organ is lifted, there is no phenomenon of material detachment or tearing. On the one hand, it shows that the biological tissue sealing material has good tissue adhesion performance and also has relatively high mechanical strength itself. On the other hand, it also shows that the biological tissue sealing material is applicable to the surfaces of various different organs.

[0111] <Test Example 7>

[0112] This test example is for testing the adhesion strength of the material obtained in Example 1.

[0113] The test method is as follows: After removing the skin on the back of the rat to expose the muscle surface, immediately press a biological tissue sealing material with a size of about 1.5 cm × 4 cm quickly on the muscle surface and continuously press for 30 s, and after releasing, clamp the biological tissue sealing material with forceps and lift the whole rat up. Then, use forceps to tear the sealing material forcefully and take photos to record the whole process of tearing.

[0114] Figure 10 These are the photos of lifting the rat in the adhesion strength test of the biological tissue sealing material of Example 1 of the present invention on the muscle surface, where the upper side is the state before lifting and the lower side is the state after lifting; Figure 10 These are the photos of the process of tearing the material in the adhesion strength test of the biological tissue sealing material of Example 1 of the present invention on the muscle surface, and the photos are arranged from left to right in chronological order.

[0115] As Figure 10As shown, after the biological tissue sealing material of the present invention is applied to the surface of fresh muscle tissue, it adheres to the muscle tissue. Even when lifting a rat weighing up to 486 g, there is no phenomenon of material detachment or tearing, indicating that the biological tissue sealing material has good tissue adhesion performance and high mechanical strength.

[0116] As Figure 11 shown, after the biological tissue sealing material of the present invention is applied to the surface of fresh muscle tissue, it can quickly bind to the amino groups on the tissue surface to form a strong adhesion. After forcefully tearing the sealing material with forceps, it can be seen that there is still a layer of material adhered to the tissue surface, indicating that the biological tissue sealing material has good tissue adhesion performance and high mechanical strength.

[0117] Functions and effects of the examples

[0118] As described above, in Examples 1 to 3, since the aldehyde group-containing polysaccharide is freeze-dried by a segmented method, that is, after being placed in a pre-freeze dryer, it is first kept at a low temperature for a period of time and then vacuumized for the first-stage freeze-drying treatment, then the vacuum degree is maintained and the temperature is slowly raised for the second-stage freeze-drying treatment, and finally the temperature is raised and maintained after returning to normal pressure. Therefore, the resulting biological tissue sealing material can have a dense structure and small pores; such a biological tissue sealing material is easily soluble in water but not easily swollen by water absorption. When in use, once it comes into contact with body fluid or the tissue surface, it can react and crosslink with the amino groups / sulfhydryl groups on the cells or proteins in the body fluid or on the tissue surface to form a physical sealing layer, thereby effectively closing the wound site. Moreover, the biological tissue sealing material also has good tissue adhesion performance and high mechanical strength, and can be applied to the surfaces of various different organs.

[0119] In contrast, in the aldehyde group-containing polysaccharide materials prepared by the traditional freeze-drying method in Comparative Examples 1 to 3, the structure is relatively loose and the pores are large, and the water absorption is very high, resulting in easy swelling after contact with body fluid or tissue. This swelling will cause the material to deform and the pores to further increase, making it difficult to effectively close the wound site.

Claims

1. A preparation method of a biological tissue sealing material, characterized in that, Prepared by a segmented freeze-drying method, which includes the following steps: Step S1: Dissolve the aldehyde-functionalized natural polysaccharide in water to prepare an aldehyde-functionalized natural polysaccharide solution with a concentration of 0.5% to 20%, and adjust the pH to 7 to 9; Step S2: Place the aldehyde-functionalized natural polysaccharide solution in a freeze-dryer pre-cooled to -80°C to -40°C and keep it for 3 to 12 hours; Step S3: Set the vacuum degree of the freeze-dryer to 0.01 to 0.5 mbar and freeze-dry for 24 to 96 hours; Step S4: Maintain the vacuum degree, heat up to 0°C at a rate of 5 to 15°C per hour, and then freeze-dry for 12 to 48 hours; Step S5: Restore the vacuum degree to normal pressure, heat up to 30 to 50°C at a rate of 10 to 20°C per hour, and keep it for 12 to 24 hours to obtain the biological tissue sealing material.

2. The preparation method of the biological tissue sealing material according to claim 1, wherein: Among them, The pre-cooling temperature in Step S2 is -55°C to -45°C.

3. The preparation method of the biological tissue sealing material according to claim 1, wherein: Among them, The aldehyde-functionalized natural polysaccharide is one or a combination of more than one of aldehyde-functionalized hyaluronic acid, aldehyde-functionalized dextran, aldehyde-functionalized pullulan, aldehyde-functionalized chitosan, and aldehyde-functionalized carboxymethyl cellulose.

4. The biological tissue sealing material prepared by the preparation method of the biological tissue sealing material according to any one of claims 1-3.

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