Natural silkworm cocoon medical surgical operation net and preparation method thereof
By degumming and mechanical extrusion of natural silkworm cocoons, a medical surgical network with high biocompatibility and excellent mechanical properties was prepared, which solved the shortcomings of existing surgical network materials in terms of biocompatibility and mechanical properties, and achieved safer and more effective tissue repair results.
Patent Information
- Application Number
- CN202510042916.1
- Authority / Receiving Office
- CN · China
- Patent Type
- Applications(China)
- Current Assignee / Owner
- Filing Date
- 2025-01-10
- Publication Date
- 2025-05-23
AI Technical Summary
The existing medical surgical mesh materials have shortcomings in biocompatibility, mechanical properties and long-term stability, which limits the effectiveness and scope of their clinical applications.
By degumming, cutting and mechanical extrusion of natural silkworm cocoons, a medical surgical network with excellent mechanical properties and biocompatibility was prepared. The surgical net consists of 71% silk fibroin and 29% sericin, and the mesh diameter, mass density and thickness are comparable to that of the commercial surgical net.
It achieves high biocompatibility, excellent mechanical properties and suitable porous structure, reduces postoperative infection and material toxicity problems, promotes cell adhesion and regeneration of damaged tissues, and improves the effectiveness and safety of surgical repair.
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Figure CN120022431A_ABST
Abstract
Description
Technical Field
[0001] The invention relates to the technical field of biomedical materials, and in particular to a natural silkworm cocoon medical surgical mesh and a preparation method thereof. Background Art
[0002] Medical surgical mesh is a mesh that is implanted at the wound site to reinforce weak parts, provide tension-free repair, and help fibrous collagen tissues to combine. It is widely used in the repair of surgical operations such as skull injuries, tendon repair, and eye surgery, as well as the treatment of diseases such as hernia, pelvic organ prolapse, and urinary incontinence. At present, there are many types of medical surgical mesh, and researchers have also made many studies on this, but the existing surgical mesh materials still have many deficiencies in biocompatibility, mechanical properties, and long-term stability, which limits the effect and scope of its clinical application.
[0003] At present, the mainstream surgical mesh materials mainly include two categories: metal and synthetic polymer. Due to its high density and rigidity, metal prostheses often cause discomfort to patients after surgery, and it is difficult for metal prostheses to perfectly combine with soft tissues, so there are many problems in practical application. Synthetic polymer surgical meshes, such as plastic meshes such as polypropylene and polytetrafluoroethylene, although they have the advantages of flexibility, lightness and strength, their biocompatibility issues have not yet been completely resolved, which may cause serious complications such as infection, inflammation, tissue erosion and limited movement. In order to alleviate the above problems of existing synthetic polymer surgical meshes, some researchers have developed absorbable synthetic meshes, such as materials composed of polyglycolic acid, polylactic acid, dipropyl carbonate and poly-4-hydroxybutyrate. Although these materials can be gradually absorbed by the human body after surgery, thereby reducing the risks associated with long-term implantation, their mechanical properties and clinical stability are relatively low. Especially after complete absorption, they cannot continue to provide sufficient support, which in turn causes the tissue in the repair area to lose its structural stability, which may cause secondary lesions, limiting their wide application in clinical practice.
[0004] In view of the limitations of metal and synthetic polymer meshes, in recent years, researchers have gradually paid attention to the potential application of biomaterials in surgical meshes. Silk fibroin is widely used in tissue engineering, wound care, drug delivery, surgical sutures and medical implants due to its easy processing, degradability, high tensile strength, biocompatibility and low immunogenicity. However, most of the silk fibroin surgical meshes regenerated by lithium bromide dissolution have significantly lower mechanical properties (such as strength and toughness) than natural fibers due to structural damage and loss of self-assembly ability during the regeneration process. Despite many improvements, the performance gap has not been fully bridged. In addition, the cocoon structure of common silkworms (such as silkworms, tussah silkworms and ricin silkworms) is dense and the porosity is insufficient, which also limits its direct application as surgical mesh. Therefore, in order to make medical surgical meshes widely used in clinical practice, further research on medical surgical meshes is needed. Summary of the invention
[0005] The invention provides a natural silkworm cocoon medical surgical mesh, which solves the technical problems that the existing natural silkworm cocoon medical surgical mesh has poor mechanical properties, dense cocoon structure and insufficient porosity, which limits its application as a surgical mesh.
[0006] The technical solution adopted by the present invention is as follows:
[0007] A natural silkworm cocoon medical surgical mesh is obtained by degumming, cutting and mechanically extruding natural silkworm cocoons, wherein the natural silkworm cocoons are Ginkgo biloba moth cocoons.
[0008] Preferably, the natural silkworm cocoon medical surgical mesh consists of 71% silk fibroin and 29% sericin.
[0009] Preferably, the diameter of the natural silkworm cocoon medical surgical mesh is in the range of 0.9 to 3.1 mm, and the mass density is 9.6 ± 2.6 mg / cm 2 The length of the mesh is 5.1 to 7.3 cm, and the width of the mesh is 2.5 to 3.6 cm.
[0010] The method for preparing the natural silkworm cocoon medical surgical mesh comprises the following steps:
[0011] (1) Pretreatment of silkworm cocoons: Cut the cocoons of Ginkgo moth and remove the pupa shells, and wash them with clean water until they are clear to remove surface dirt and dust;
[0012] (2) Cocoon degumming treatment: The cocoons of the Ginkgo moth obtained by pretreatment are placed in a hot alkaline solution for a one-step degumming treatment to remove part of the sericin protein, retain the natural network structure of the silk fibroin fibers and part of the sericin protein, and then rinsed with distilled water to a neutral pH;
[0013] (3) Flat extrusion: The processed cocoon pieces are cut and opened flatly with pressure and pressed flat to form a surgical mesh with a predetermined shape and thickness.
[0014] More preferably, the hot alkaline solution in step (2) is NaCO 3 One or more of solution, NaOH solution, and urea solution.
[0015] Furthermore, the alkaline substance concentration of the hot alkaline solution is 0.1-0.3 mol / L. The concentration of the alkaline solution is appropriate. If the concentration is too high, the silk fibroin in the cocoon will be damaged, while if the concentration is too low, part of the sericin cannot be effectively removed.
[0016] More preferably, the degumming treatment temperature in step (2) is 70-90° C., and the treatment time is 5-20 min. The degumming time needs to be properly controlled. Too long a time will damage the silk fibroin, while too short a time will result in the sericin not being degummed to an appropriate content, affecting the degumming effect.
[0017] More preferably, in step (3), the time for squeezing the surgical mesh is 0.5 to 1.5 hours, and the squeezing pressure is 0.1 to 1 MPa.
[0018] More preferably, in step (3), a glass slide is used to squeeze the surgical mesh.
[0019] In summary, compared with the prior art, the present invention has the following advantages and beneficial effects:
[0020] 1. The present invention provides a medical surgical mesh derived from natural silkworm cocoons. Compared with previous metal and synthetic materials, the newly invented surgical mesh has high biocompatibility, excellent mechanical properties and suitable porous morphology, and has high biosafety. It can promote cell adhesion and regeneration of damaged tissues, achieve rapid repair of traumatic sites, and is used in tissue repair and regeneration in surgical operations.
[0021] 2. The surgical mesh prepared by the preparation method provided by the present invention has a pore size, mass density and thickness comparable to those of commercial surgical mesh, and at the same time has excellent mechanical strength, high biocompatibility, outstanding mechanical properties and a suitable porous structure. The surgical mesh material provided by the present invention has high biosafety and can effectively avoid possible infection and material toxicity problems after surgical operations.
[0022] 3. The preparation process of the natural silkworm cocoon surgical mesh of the present invention is simple and efficient, and makes full use of the natural structural characteristics of the Ginkgo moth cocoon. After removing the sericin protein through degumming treatment, the high-strength silk fibroin protein is retained, and a surgical mesh with excellent mechanical properties and biocompatibility is formed through physical plasticization. This process not only effectively simplifies the production process, but also can retain the natural advantages of the silkworm cocoon to the greatest extent, thereby enhancing the clinical application potential of the product.
[0023] 4. The present invention reduces the content of sericin through degumming treatment, making the natural cocoon surgical mesh composed of 71% fibroin and 29% sericin. The surgical mesh with this component ratio can not only maintain the environmental humidity, absorb the excess leakage liquid from the wound, but also has low immunogenicity. The hydroxyl, carboxyl and amino functional groups in sericin promote cell adhesion and proliferation, thereby accelerating the tissue repair process, reducing the postoperative inflammatory reaction, and enhancing biocompatibility and safety. BRIEF DESCRIPTION OF THE DRAWINGS
[0024] Figure 1 It is the microscopic surface morphology diagram of the surgical mesh prepared in Example 13 under different magnification fields of view.
[0025] Figure 2 It is the SEM image and the fluorescence microscope image co-stained with FDA and Hoescht 33258 of the surgical mesh prepared in Example 13 after inoculating and culturing with L929 cells for 72 hours.
[0026] Figure 3 It is the cell viability diagram of L929 cells cultured on the surgical mesh prepared in Example 13 for 24h - 72h.
[0027] Figure 4 It is the stereomicroscope diagram of the surgical mesh prepared in Example 13 implanted into the peritoneal cavity of rats.
[0028] Figure 5 It is the degradation state diagram of the surgical mesh prepared in Example 13 in the body of rats. DETAILED DESCRIPTION OF THE INVENTION
[0029] The present invention will be specifically described below in combination with the specific embodiments and examples, and the advantages and various effects of the present invention will be presented more clearly therefrom. Those skilled in the art should understand that these specific embodiments and examples are used to illustrate the present invention, rather than limiting the present invention.
[0030] Throughout the specification, unless otherwise specifically stated, the terms used herein should be understood as having the meaning as commonly used in the art. Therefore, unless otherwise defined, all technical and scientific terms used herein have the same meaning as the general understanding of those skilled in the art to which the present invention belongs. In case of contradiction, this specification shall prevail.
[0031] Unless otherwise specifically stated, various raw materials, reagents, instruments and equipment used in the present invention can be obtained through market purchase or can be prepared by existing methods.
[0032] The natural cocoon medical surgical mesh of the present invention is prepared through the following steps:
[0033] (1) Pretreatment of silkworm cocoons: Cut the cocoons of Ginkgo moth and remove the pupa shells, and wash them with clean water until they are clear to remove surface dirt and dust;
[0034] (2) Cocoon degumming treatment: The cocoons of the Ginkgo moth obtained by pretreatment are placed in a hot alkaline solution for a one-step degumming treatment to remove part of the sericin protein, retain the natural network structure of the silk fibroin fibers and part of the sericin protein, and then rinsed with distilled water to a neutral pH;
[0035] (3) Flat extrusion: The processed cocoon pieces are cut and opened flatly with pressure and pressed flat to form a surgical mesh with a predetermined shape and thickness.
[0036] The present application will be described in detail below with reference to embodiments and experimental data.
[0037] Example 1
[0038] The relevant preparation parameters of the surgical mesh in this embodiment are: degumming temperature is 70°C, alkaline solution is 0.1mol / LNaOH solution, degumming time is 5min, and after degumming, it is washed with distilled water until it reaches a neutral pH value. Then, the degummed silkworm cocoon pieces are cut into about 2×5cm, opened flat between glass slides with pressure, and pressed flat for 0.5Mpa with pressure for 0.5 hours.
[0039] Example 2
[0040] Based on Example 1, the degumming temperature is limited to 80°C.
[0041] Example 3
[0042] Based on Example 1, the degumming temperature is limited to 90°C.
[0043] Example 4
[0044] Based on Example 1, the degumming temperature is limited to 100°C.
[0045] Example 5
[0046] Based on Example 2, the alkaline solution is limited to 0.2 mol / L NaOH solution.
[0047] Example 6
[0048] Based on Example 2, the alkaline solution is limited to 0.3 mol / L NaOH solution.
[0049] Example 7
[0050] Based on Example 2, the alkaline solution is limited to 0.5 mol / L NaOH solution.
[0051] Example 8
[0052] Based on Example 5, the degumming time is limited to 10 minutes.
[0053] Example 9
[0054] Based on Example 5, the degumming time is limited to 20 minutes.
[0055] Example 10
[0056] Based on Example 5, the degumming time is limited to 30 minutes.
[0057] Embodiment 11
[0058] Based on Example 8, the size of the surgical mesh is limited to 2×7 cm.
[0059] Example 12
[0060] Based on Example 8, the size of the surgical mesh is limited to 2×10 cm.
[0061] Example 13
[0062] Based on Example 8, the surgical mesh pressing time is limited to 1 hour.
[0063] Embodiment 14
[0064] Based on Example 8, the surgical mesh pressing time is limited to 1.5 hours.
[0065] In order to verify whether the present invention achieves the expected effect, the structural morphology, pore size, mass density, thickness, biological activity, degradation performance, etc. of the samples of the above embodiments are tested and analyzed, as follows:
[0066] Test Example 1 Structural morphology test of natural silkworm cocoon surgical mesh
[0067] The samples obtained in Examples 1-14 were subjected to scanning electron microscopy (SEM) tests. The results showed that the samples obtained in each example all presented a regular reticular structure with uniform pore sizes and interconnected pores. Taking the sample prepared in Example 13 as an example, the SEM test results are shown in the attached figure. Figure 1 As shown. Figure 1 It can be observed that the sample of Example 13 forms a regular network structure, in which the nano-scale silk fibroin fibers are interwoven with sericin to form microfibers, and the microfibers are further combined with sericin to form a larger fiber network.
[0068] Test Example 2: Aperture and mass density test of natural silkworm cocoon surgical mesh.
[0069] The pore size range of the samples obtained in each embodiment was tested by SEM, and the mass density of the samples in the embodiment was calculated by weighing using a high-precision balance. The pore size and mass density of each embodiment are shown in Table 1:
[0070] Table 1 Pore diameter and mass density of each sample
[0071]
[0072] Studies have shown that the pore size range of 0.9 to 3.10 mm can ensure that the surgical mesh maintains proper structure and function during the repair process while ensuring effective penetration of cells and tissues. A pore size that is too small may lead to limited cell penetration, affecting the normal growth and repair of tissues, while a pore size that is too large may make the omentum structure too loose and lose the necessary support. It is not difficult to see from the data in Table 1 that the pore size of the sample obtained in Example 4 / 7 / 10 is too large and does not meet the mechanical properties of the surgical mesh, indicating that the degumming temperature, alkali solution concentration and degumming time all affect the finished product quality of the surgical mesh, while the extrusion time has no obvious effect.
[0073] Test Example 3: The thickness and average weight per square centimeter of natural silkworm cocoon surgical mesh, commercially available polypropylene and polyethylene surgical mesh were tested.
[0074] This test example uses the sample prepared in Example 13 as a representative to compare with the commercially available surgical mesh. The thickness and average weight per square centimeter of Example 13, commercially available polypropylene and polyethylene surgical meshes are measured using a micrometer, as shown in Table 2 below:
[0075] Table 2 Thickness and average weight per square centimeter of different surgical meshes
[0076]
[0077] It is not difficult to see from Table 2 that the thickness of Example 13 is not much different from that of the commercially available polypropylene mesh, but is significantly higher than that of the commercially available polyethylene mesh; but the weight is significantly lower than that of the commercially available polypropylene surgical mesh, and is not much different from that of the commercially available polyethylene surgical mesh. It can be seen that the natural silkworm cocoon surgical mesh has achieved a good balance between thickness and weight, and can provide sufficient mechanical support while maintaining a light weight. It is particularly suitable for surgical repairs with long-term loads and high requirements for biocompatibility.
[0078] Test Example 4 In vitro biocompatibility and cytotoxicity test of natural silkworm cocoon surgical mesh.
[0079] This test example still uses the sample obtained in Example 13 as a representative for testing, and uses the MTT test to detect the in vitro biocompatibility and cytotoxicity of the surgical mesh obtained in Example 13. The specific test steps are as follows:
[0080] L929 mouse fibroblasts were cultured in a 5% CO2 The cells were cultured in a 37°C humidified incubator in Dulbecco's modified Eagle's medium (DMEM) containing 10% fetal bovine serum (FBS), penicillin (100 U / mL) and streptomycin (100 μg / mL).
[0081] For the MTT assay, L929 cells were plated at 2 × 10 4 The cells were inoculated at a density of and cultured for 24 hours. After 24 hours, the culture medium was removed from the wells, and an equal volume (100 μL) of culture medium and sterilized experimental materials were added to each well. In the control wells, only 100 μL of DMEM was added. After removing the test materials at the corresponding time points (24 hours, 48 hours, and 72 hours), MTT solution (10 μL MTT and 90 μL culture medium) was added to each well and placed in a dark environment at 37°C for 3 hours. Subsequently, the culture medium was aspirated, and 100 μL of DMSO was added to each well to dissolve the purple MTT formazan crystals and allowed to stand for 5 minutes. Afterwards, use The ELISA reader measures absorbance at 492 nm.
[0082] Scanning electron microscopy (SEM, Figure 2 A) and fluorescence microscopy ( Figure 2 B) showed that L929 mouse fibroblasts were able to attach to the surface of the sample obtained in Example 13, and this attachment did not require any chemical treatment. To evaluate the cytotoxic effect of the example, MTT analysis and real-time xCELLigence system were used for monitoring, and the results are shown in the attached Figure 3 As shown, the MTT analysis results showed that the cells incubated with the sample obtained in Example 13 for 24 hours, 48 hours and 72 hours did not show any cytotoxicity.
[0083] Test Example 5 In vivo biocompatibility test of natural silkworm cocoon surgical mesh.
[0084] This test example still uses the sample obtained in Example 13 as a representative for testing.
[0085] Experimental subjects: 14 male adult albino Wistar rats, weighing 250-270 g.
[0086] Test period: 21 days.
[0087] Experimental Method: The animals were anesthetized with ketamine (100 mg / kg) / xylazine (10 mg / kg) and a 1×1 cm sample of Example 13 was placed in the abdominal cavity of the animals. Using the same protocol, Example 13 was implanted subcutaneously bilaterally in the axilla. The animals were maintained under sterile conditions throughout the study. After implantation, the rats were placed in standard cages with free access to water and food. 21 days after implantation, the rats were anesthetized and the implanted MothMesh material was excised for macroscopic and histopathological analysis.
[0088] Test results: The stereomicroscope image of the sample obtained in Example 13 of rat intraperitoneum is shown in the attached figure. Figure 4 As can be seen from the figure, after the samples obtained in Example 13 were implanted in mice for 2 and 3 weeks, the tissue samples were subjected to histopathological examination, and no necrosis, fibrosis, foreign body reaction or chronic inflammatory cells were detected. In addition, the histopathological results showed that the mouse tissues were able to accept the samples obtained in Example 13 well and regarded them as part of their own tissues. Primary tissue healing was observed in the implanted area, which was manifested as normal reconstruction of tissue structure and functional recovery, and no obvious scar formation ( Figure 4 ). This indicates that natural silkworm cocoon surgical mesh has excellent biocompatibility and can effectively avoid foreign body reactions or inflammation caused by implants, thus providing strong support for its safety and reliability as a surgical implant material.
[0089] Test Example 6 In vivo biodegradability test of natural silkworm cocoon surgical mesh.
[0090] In vivo analysis was performed on the samples obtained in Example 13. At the end of the 6th week, the surgical mesh removed under anesthesia was dried in an oven at 37°C for 3 days and then reweighed after stripping off all tissue particles to test the degradation state of the samples obtained in Example 13.
[0091] Test results Example 13 The degradation state of the sample obtained is shown in the attached Figure 5 As can be seen from the figure, 21 days after implantation, the surgical mesh obtained in Example 13 in the rat body was observed by stereomicroscope ( Figure 5 ), its structure is intact, without deformation or obvious signs of degradation, and the mesh morphology remains regular, indicating that the surgical mesh has excellent structural stability and anti-degradation properties in the biological environment. It is further proved that the natural silkworm cocoon surgical mesh has good mechanical durability and biocompatibility in vivo, which can provide ideal conditions for long-term tissue healing (Table 3).
[0092] Table 3 Comparison of the sample obtained in Example 13 with the existing surgical mesh
[0093]
[0094] Finally, it should be noted that the terms "comprises," "comprising," or any other variations thereof are intended to cover non-exclusive inclusion, such that a process, method, article, or device comprising a series of elements includes not only those elements, but also other elements not explicitly listed, or also includes elements inherent to such process, method, article, or device. Although preferred embodiments of the present invention have been described, additional changes and modifications may be made to these embodiments by those skilled in the art once the basic inventive concepts are known. Therefore, the appended claims are intended to be interpreted as including the preferred embodiments and all changes and modifications that fall within the scope of the present invention.
[0095] The above-mentioned embodiments only express the specific implementation methods of the present application, and the descriptions thereof are relatively specific and detailed, but they cannot be understood as limiting the protection scope of the present application. It should be pointed out that, for ordinary technicians in this field, several variations and improvements can be made without departing from the technical solution concept of the present application, and these all belong to the protection scope of the present application.
Claims
1. A natural silkworm cocoon medical surgical mesh, characterized in that: The natural silkworm cocoon medical surgical mesh is obtained by degumming, cutting and mechanically extruding natural silkworm cocoons, wherein the natural silkworm cocoons are Ginkgo moth cocoons.
2. The natural silkworm cocoon medical surgical mesh according to claim 1, characterized in that: The natural silkworm cocoon medical surgical mesh consists of 71% of silk fibroin and 29% of sericin.
3. The natural silkworm cocoon medical surgical mesh according to claim 1, characterized in that: The diameter of the natural silkworm cocoon medical surgical mesh ranges from 0.9 to 3.1 mm, and the mass density is 9.6±2.6 mg / cm 2 The length of the mesh is 5.1 to 7.3 cm, and the width of the mesh is 2.5 to 3.6 cm.
4. The method for preparing the natural silkworm cocoon medical surgical mesh according to any one of claims 1 to 3, characterized in that: The steps include: (1) Pretreatment of silkworm cocoons: Cut the cocoons of Ginkgo moth and remove the pupa shells, and wash them with clean water until they are clear to remove surface dirt and dust; (2) Cocoon degumming treatment: The cocoons of the Ginkgo moth obtained by pretreatment are placed in a hot alkaline solution for a one-step degumming treatment to remove part of the sericin protein, retain the natural network structure of the silk fibroin fibers and part of the sericin protein, and then rinsed with distilled water to a neutral pH; (3) Flat extrusion: The processed cocoon pieces are cut and opened flatly with pressure and pressed flat to form a surgical mesh with a predetermined shape and thickness.
5. The method for preparing the natural silkworm cocoon medical surgical mesh according to claim 4, characterized in that: The hot alkaline solution in step (2) is one or more of NaCO3 solution, NaOH solution and urea solution.
6. The method for preparing the natural silkworm cocoon medical surgical mesh according to claim 5, characterized in that: The alkaline substance concentration of the hot alkaline solution is 0.1-0.3 mol / L.
7. The method for preparing the natural silkworm cocoon medical surgical mesh according to claim 4, characterized in that: The degumming treatment temperature in step (2) is 70 to 90° C., and the treatment time is 5 to 20 minutes.
8. The method for preparing the natural silkworm cocoon medical surgical mesh according to claim 4, characterized in that: In step (3), the time for squeezing the surgical mesh is 0.5 to 1.5 hours, and the squeezing pressure is 0.1 to 1 MPa.
9. The method for preparing the natural silkworm cocoon medical surgical mesh according to claim 4, characterized in that: Step (3) uses a glass slide to squeeze the surgical mesh.