Gel dressing containing recombinant collagen as well as preparation method and application of gel dressing

Abstract

The invention belongs to the technical field of dressings, and particularly relates to a gel dressing containing recombinant collagen as well as a preparation method and application of the gel dressing. The gel dressing containing the recombinant collagen is prepared from the following raw materials in percentage by weight: 0.1 to 1 percent of the recombinant collagen, 0.5 to 1 percent of sodium hyaluronate, 0.08 to 0.15 percent of radices stellariae dichotomae cyclopeptide, 0.5 to 1 percent of carbomer, 0.03 to 0.06 percent of cryptomeria fortunei bud extract, 15 to 20 percent of poloxamer, 0.1 to 0.5 percent of xanthan gum, 3 to 8 percent of glycerol, 0.04 to 0.08 percent of edetate disodium, 0.1 to 0.3 percent of triethanolamine, 0.08 to 0.12 percent of preservative and the balance of deionized water. The gel dressing has excellent biocompatibility, good hemostasis, cell growth promotion and tissue repair performance, and can be used for repairing vaginal mucous membrane chap and hemostasis of diffuse superficial bleeding, and improving the symptoms of elasticity reduction and the like caused by vaginal atrophy.

Description

A gel dressing containing recombinant collagen, its preparation method and application Technical Field

[0001] This invention belongs to the field of dressing technology, and particularly relates to a gel dressing containing recombinant collagen, its preparation method, and its application. Background Technology

[0002] As the body's first line of defense, the skin is frequently damaged by mechanical, physical, chemical, or biological factors. Once damage occurs, timely dressing is crucial for healing. Damage to the vaginal mucosa and its accompanying atrophy, dryness, and decreased elasticity are widespread in various physiological and pathological states. If these problems are not properly addressed, they may disrupt the local microecological balance, weaken the mucosa's physical barrier and immune defense functions, increase the risk of infection, and impose long-term physiological and psychological burdens on patients.

[0003] Ideal dressings should not only cover the wound but also regulate the healing environment and promote tissue regeneration. Traditional dressings such as gauze and cotton pads, while absorbing exudate, often cause secondary trauma during dressing changes due to adhesion to new tissue. Furthermore, their poor moisture retention makes it difficult to maintain wound moisture, failing to meet the high clinical standards of modern wound repair, especially for chronic, refractory wounds. Collagen, as the most abundant structural protein in the human body, is crucial for maintaining the mechanical properties and functional integrity of tissues such as skin. It not only provides support for cells but also guides cell migration, proliferation, and differentiation, thereby actively promoting healing. Therefore, it is considered an ideal wound repair material and has become a research frontier in the field of biomedical materials.

[0004] However, the function of collagen alone remains limited when faced with the complex repair of vaginal mucosal injuries. An ideal repair dressing needs to address multiple objectives, including structural support, maintaining a moist environment, promoting hemostasis, and accelerating cell proliferation, in order to comprehensively shorten the healing process. Therefore, scientifically combining recombinant collagen with other functional ingredients with specific biological activities to achieve synergistic effects has become a key aspect of improving the overall performance of dressings. Summary of the Invention

[0005] To overcome the shortcomings of existing technologies, the primary objective of this invention is to provide a gel dressing containing recombinant collagen. This gel dressing is a vaginal mucosal repair dressing that combines high safety, excellent biocompatibility, and repair functions. Using recombinant collagen as a safe and reliable basic framework and integrating other natural active ingredients, it exhibits significant advantages in hemostasis, cell growth promotion, and ultimately tissue repair. It can be used for the repair of vaginal mucosal fissures and hemostasis of diffuse superficial bleeding, and to improve discomfort symptoms such as decreased elasticity caused by vaginal atrophy.

[0006] The second objective of this invention is to provide a method for preparing a gel dressing containing recombinant collagen.

[0007] A third objective of this invention is to provide an application of a gel dressing containing recombinant collagen in the preparation of vaginal gels.

[0008] The objective of this invention is achieved through the following technical solution: a gel dressing containing recombinant collagen, comprising, by weight percentage, the following raw materials: 0.1-1% recombinant collagen, 0.5-1% sodium hyaluronate, 0.08-0.15% stellaria dichotoma cyclic peptide, 0.5-1% carbomer, 0.03-0.06% cedar bud extract, 15-20% poloxamer, 0.1-0.5% xanthan gum, 3-8% glycerin, 0.04-0.08% disodium edetate, 0.1-0.3% triethanolamine, 0.08-0.12% preservative, with the balance being deionized water.

[0009] Further, the preparation method of the cedar bud extract is as follows: take cedar buds, wash and dry them, and pulverize them to 20-40 mesh to obtain cedar bud powder; add 10-15 times the mass of distilled water to the cedar bud powder, heat and extract, filter and collect the filtrate and filter residue; add 10-15 times the mass of distilled water to the filter residue, continue to heat and extract, and filter; then combine the filtrates, concentrate them, and freeze-dry to obtain the extract.

[0010] Furthermore, the heating and extraction temperature is 90-100℃, and the time is 1-2 hours.

[0011] Furthermore, the recombinant collagen is recombinant type III humanized collagen with a molecular weight of 5 × 10⁻⁶. 4 -8×10 4 Da.

[0012] Furthermore, the carbomer is carbomer 980, and the poloxamer is poloxamer 407.

[0013] Furthermore, the molecular weight of the sodium hyaluronate is 1×10⁻⁶. 6 -2×10 6 Da.

[0014] Furthermore, the preservative is methylparaben or phenoxyethanol.

[0015] A method for preparing a gel dressing containing recombinant collagen includes the following steps: (1) Take half of the total amount of deionized water, add glycerin, then add carbomer, poloxamer and xanthan gum, stir and mix evenly to obtain a gel matrix; (2) Add disodium edetate to the gel matrix of step (1) and stir and mix evenly at 70-80℃ for 10-20 min to obtain mixture A, and cool to room temperature for later use; (3) Take the remaining amount of deionized water, add recombinant collagen, silver stellaria cyclic peptide, sodium hyaluronate, cedar bud extract and preservative, stir and mix evenly to obtain mixture B, and set aside for later use; (4) Add mixture B of step (3) to mixture A of step (2), stir and mix evenly, add triethanolamine, fill and sterilize to obtain the final product.

[0016] Application of a gel dressing containing recombinant collagen in the preparation of vaginal gel.

[0017] Compared with the prior art, this invention has the following advantages: 1. This invention discloses a gel dressing containing recombinant collagen, using a combination of recombinant collagen, *Stellaria media* cyclic peptide, and *Cryptomeria japonica* bud extract. The recombinant collagen has good biocompatibility, while the *Stellaria media* cyclic peptide and *Cryptomeria japonica* bud extract have good hemostatic, healing-promoting, and repairing effects. Sodium hyaluronate and glycerin have good moisturizing effects. The synergistic effect of these components can accelerate the repair of vaginal mucosal damage, improve vaginal elasticity, and alleviate discomfort caused by vaginal atrophy. This gel dressing is non-cytotoxic and safe to use.

[0018] 2. This invention discloses a method for preparing a gel dressing containing recombinant collagen, which is easy to operate and suitable for industrial-scale production.

[0019] 3. This invention discloses the application of a gel dressing containing recombinant collagen in the preparation of vaginal gel, which can be used for the repair of vaginal mucosal fissures and hemostasis of diffuse superficial bleeding, and to improve discomfort symptoms such as decreased elasticity caused by vaginal atrophy. Attached Figure Description

[0020] Figure 1 shows the in vitro cytotoxicity results of the gel dressing obtained in Example 1 of the present invention. Detailed Implementation

[0021] The present invention will be further described in detail below with reference to embodiments, but the implementation of the present invention is not limited thereto. Unless otherwise specified, the reagents, methods, and equipment used in the present invention are conventional reagents, methods, and equipment in this technical field. Test methods in the following embodiments that do not specify specific experimental conditions are generally performed according to conventional experimental conditions or experimental conditions recommended by the manufacturer. Unless otherwise specified, the reagents and raw materials used in the present invention are commercially available.

[0022] Example 1: A gel dressing containing recombinant collagen, comprising the following raw materials by weight percentage: recombinant collagen (recombinant type III humanized collagen, molecular weight 5-8 × 10⁻⁶). 4 Da) 0.5%, sodium hyaluronate (1-2×10 6 The ingredients are: Da 0.8%, Stellaria dichotoma cyclic peptide 0.12% (CAS 137476-73-4), Carbomer 980 0.8%, Cryptomeria japonica bud extract 0.05%, Poloxamer (Poloxamer 407) 18%, Xanthan gum 0.3%, Glycerin 5%, Disodium edetate 0.06%, Triethanolamine 0.2%, Preservative (Methylparaben) 0.1%, and the balance being deionized water.

[0023] The preparation method of the cedar bud extract is as follows: take cedar buds, wash and dry them, then pulverize them to 30 mesh to obtain cedar bud powder. Add 12 times the mass of the cedar bud powder with distilled water, and then extract at 95℃ for 1.5h. Filter and collect the filtrate and filter residue. Add 12 times the mass of the filter residue with distilled water to the filter residue, and continue to extract at 95℃ for 1.5h. After filtration, combine the two filtrates, concentrate, and freeze-dry to obtain the extract.

[0024] A method for preparing a gel dressing containing recombinant collagen includes the following steps: (1) Take half of the total amount of deionized water, add glycerin, then add carbomer, poloxamer and xanthan gum, stir and mix evenly to obtain a gel matrix; (2) Add disodium edetate to the gel matrix of step (1) and stir and mix evenly at 75°C for 15 min to obtain mixture A, and cool to room temperature for later use; (3) Take the remaining amount of deionized water, add recombinant collagen, silver stellaria cyclic peptide, sodium hyaluronate, cedar bud extract and preservative, stir and mix evenly to obtain mixture B, and set aside for later use; (4) Add mixture B of step (3) to mixture A of step (2), stir and mix evenly, add triethanolamine, fill and sterilize to obtain the final product.

[0025] Example 2: A gel dressing containing recombinant collagen, comprising the following raw materials by weight percentage: recombinant collagen (recombinant type III humanized collagen, molecular weight 5-8 × 10⁻⁶). 4 Da) 0.1%, sodium hyaluronate (1-2×10 6 The ingredients are: Da 0.5%, Stellaria dichotoma cyclic peptide 0.08%, Carbomer 980 0.5%, Cryptomeria japonica bud extract 0.03%, Poloxamer (Poloxamer 407) 15%, Xanthan gum 0.1%, Glycerin 3%, Disodium edetate 0.04%, Triethanolamine 0.1%, Preservative (Phenoxyethanol) 0.08%, and the balance is deionized water.

[0026] The preparation method of the cedar bud extract is as follows: take cedar buds, wash and dry them, then pulverize them to 20 mesh to obtain cedar bud powder, add 10 times the mass of distilled water to the cedar bud powder, and then extract at 90℃ for 2 hours. Filter and collect the filtrate and filter residue; add 10 times the mass of distilled water to the filter residue, continue to extract at 90℃ for 2 hours, filter and combine the two filtrates, concentrate, and freeze dry to obtain the extract.

[0027] A method for preparing a gel dressing containing recombinant collagen includes the following steps: (1) Take half of the total amount of deionized water, add glycerin, then add carbomer, poloxamer and xanthan gum, stir and mix evenly to obtain a gel matrix; (2) Add disodium edetate to the gel matrix of step (1) and stir and mix evenly at 70°C for 20 min to obtain mixture A, and cool to room temperature for later use; (3) Take the remaining amount of deionized water, add recombinant collagen, silver stellaria cyclic peptide, sodium hyaluronate, cedar bud extract and preservative, stir and mix evenly to obtain mixture B, and set aside for later use; (4) Add mixture B of step (3) to mixture A of step (2), stir and mix evenly, add triethanolamine, fill and sterilize to obtain the final product.

[0028] Example 3: A gel dressing containing recombinant collagen, comprising the following raw materials by weight percentage: recombinant collagen (recombinant type III humanized collagen, molecular weight 5-8 × 10⁻⁶). 4 Da) 1%, Sodium hyaluronate (1-2×10) 6 The ingredients are: Da 1%, Silver Buckwheat Cyclopeptide 0.15%, Carbomer 980 1%, Chinese Fir Bud Extract 0.06%, Poloxamer (Poloxamer 407) 20%, Xanthan Gum 0.5%, Glycerin 8%, Disodium edetate 0.08%, Triethanolamine 0.3%, Preservative (Phenoxyethanol) 0.12%, and the balance is deionized water.

[0029] The preparation method of the cedar bud extract is as follows: take cedar buds, wash and dry them, then pulverize them to 40 mesh to obtain cedar bud powder, add 15 times the mass of the cedar bud powder with distilled water, and then extract at 100℃ for 1 hour. Filter and collect the filtrate and filter residue; add 15 times the mass of the filter residue with distilled water to the filter residue, continue to extract at 100℃ for 1 hour, filter and combine the two filtrates, concentrate, and freeze dry to obtain the extract.

[0030] A method for preparing a gel dressing containing recombinant collagen includes the following steps: (1) Take half of the total amount of deionized water, add glycerin, then add carbomer, poloxamer and xanthan gum, stir and mix evenly to obtain a gel matrix; (2) Add disodium edetate to the gel matrix of step (1) and stir and mix evenly at 80°C for 10 min to obtain mixture A, and cool to room temperature for later use; (3) Take the remaining amount of deionized water, add recombinant collagen, silver stellaria cyclic peptide, sodium hyaluronate, cedar bud extract and preservative, stir and mix evenly to obtain mixture B, and set aside for later use; (4) Add mixture B of step (3) to mixture A of step (2), stir and mix evenly, add triethanolamine, fill and sterilize to obtain the final product.

[0031] The difference between Comparative Example 1 and Example 1 is that no silver-scented bupleurum cyclic peptide was added; otherwise, they are the same as Example 1.

[0032] The difference between Comparative Example 2 and Example 1 is that no Japanese cedar bud extract was added; otherwise, they are the same as Example 1.

[0033] The difference between Comparative Example 3 and Example 1 is that: no *Cephalotaxus fortunei* bud extract was added, and the amount of *Stellaria media* cyclic peptide was increased; otherwise, it remained the same as Example 1.

[0034] Experimental Example 11, Cytotoxicity Experiment: 1.1 Preparation of Experimental Samples The gel dressing obtained in Example 1 was immersed in serum-containing MEM medium at a ratio of 0.1 g / mL and extracted by shaking at 37°C for 24 h to prepare the corresponding extract for Example 1 group; the serum-containing MEM medium was used as a blank control solution and the MEM medium containing serum and 5% phenol was used as a positive control solution.

[0035] 1.2 Preparation of cell suspension: L929 cells were digested with trypsin and then prepared into 1×10⁻⁶ cells using fresh culture medium. 5 A single-cell suspension of cells per mL.

[0036] 1.3 Cell Culture: 100 μL of cell suspension (1×10⁻⁶ cells / mL) was cultured. 4 Cells (cells / mL) were seeded in 96-well plates, with three groups: Example 1 group, blank control group, and positive control group, each with 5 replicates. Cells were incubated at 37°C and 5% CO2 for 24 h to allow adhesion. The original culture medium was discarded, and the experimental groups were treated with the extract obtained in Example 1, the blank control group with blank control solution, and the positive control group with positive control solution. Cells were then cultured for another 24 h. Cell morphology was observed under a microscope. The results are shown in Figure 1.

[0037] As can be seen from Figure 1, the gel dressing obtained by this invention has good cell compatibility and can be used for subsequent experimental research.

[0038] Experiment 2: In vitro coagulation time determination: Accurately weigh 10 mg of each of the gel dressings prepared in Examples 1-3 and Comparative Examples 1-3. Place each sample in a clean test tube, add 1 mL of fresh blood, immediately and gently shake well, and start timing. Observe and record the time required for complete blood coagulation. The experimental results are summarized in Table 1.

[0039] Table 1 As shown in Table 1, the gel dressings obtained in Examples 1-3 exhibited better hemostatic effects compared to Comparative Examples 1-3. Comparative Example 1 did not contain *Stellaria media* cyclic peptide; Comparative Example 2 did not contain *Cryptomeria japonica* bud extract; and Comparative Example 3 did not contain *Cryptomeria japonica* bud extract but increased the amount of *Stellaria media* cyclic peptide. The clotting time of all three examples increased to varying degrees compared to Example 1, indicating that both *Stellaria media* cyclic peptide and *Cryptomeria japonica* bud extract play important roles in promoting hemostasis.

[0040] Experimental Example 3: Cell proliferation capacity assay: Vaginal epithelial cells were cultured at a density of 5 × 10⁶ cells per well. 3 Cells were seeded at a density of 1000 cells / well in a 96-well plate, and then serum-free culture medium containing the gel dressings obtained in Examples 1-3 and Comparative Examples 1-3 was added. The serum-free culture medium was set as a blank control. After culturing for 72 hours, the cell proliferation of each group was detected using a CCK-8 kit. The experimental results are shown in Table 2.

[0041] Table 2 As shown in Table 2, the control group did not contain any active ingredients, and the cells grew in the basal environment, with the proliferation rate serving as a baseline reference. After 72 hours of treatment with the gel dressings obtained in Examples 1-3, the cell survival rate of vaginal epithelial cells was significantly higher than that of Comparative Examples 1-3. This is because the recombinant collagen, *Stellaria dichotoma* cyclic peptide, and *Cephalotaxus fortunei* bud extract worked synergistically to promote cell proliferation. Comparative Example 1 did not contain *Stellaria dichotoma* cyclic peptide; Comparative Example 2 did not contain *Cephalotaxus fortunei* bud extract; and Comparative Example 3 did not contain *Cephalotaxus fortunei* bud extract, but the amount of *Stellaria dichotoma* cyclic peptide was increased. The cell-promoting effect of the three was weakened, indicating that *Stellaria dichotoma* cyclic peptide and *Cephalotaxus fortunei* bud extract can work synergistically in this formulation to promote the proliferation of vaginal epithelial cells.

[0042] The above results indicate that the gel dressing of the present invention is non-cytotoxic and helps repair vaginal mucosal damage.

[0043] Example 4: Evaluation of Skin Wound Healing: Healthy SD rats were used to create full-thickness skin defects on both sides of the spine on their backs, with a wound size of approximately 1 cm × 1 cm. The rats were randomly divided into 6 groups of 5 rats each, and treated with the gel dressings obtained in Examples 1-3 and Comparative Examples 1-3, respectively. Each group received the dressing once daily for 12 hours. Wound healing was observed on days 7 and 12 after application, and the wound healing rate was calculated using the following formula: Wound healing rate (%) = (Initial wound area − Measured wound area) / Initial wound area × 100%. The experimental results are summarized in Table 3.

[0044] Table 3 As shown in Table 3, Examples 1-3 demonstrated the best performance in promoting wound healing: the healing rate of Examples 1-3 was significantly higher than that of Comparative Examples 1-3 at both 7 and 12 days, and near-complete healing was achieved at 12 days. This further confirms the excellent healing-promoting effect of the synergistic formulation of recombinant collagen, *Stellaria media* cyclic peptide, and *Cryptomeria japonica* bud extract.

[0045] The wound healing effects of Comparative Examples 1-3 were all weaker than those of Comparative Examples 1-3. This indicates that the absence of either Stellaria dichotoma cyclic peptide or Cryptomeria japonica bud extract would severely weaken the effect. Even increasing the dosage of Stellaria dichotoma cyclic peptide could not compensate for the loss caused by the lack of Cryptomeria japonica bud extract, further emphasizing the importance of the synergistic effect of the two.

[0046] The above embodiments are merely preferred embodiments of the present invention and should not be construed as limiting the scope of protection of the present invention. Any non-substantial changes and substitutions made by those skilled in the art based on the present invention shall fall within the scope of protection claimed by the present invention.

Claims

1. A gel dressing containing recombinant collagen, characterized in that, By weight percentage, it includes the following ingredients: 0.1-1% recombinant collagen, 0.5-1% sodium hyaluronate, 0.08-0.15% stellaria dichotoma cyclic peptide, 0.5-1% carbomer, 0.03-0.06% cedar bud extract, 15-20% poloxamer, 0.1-0.5% xanthan gum, 3-8% glycerin, 0.04-0.08% disodium edetate, 0.1-0.3% triethanolamine, 0.08-0.12% preservatives, with the balance being deionized water.

2. The gel dressing containing recombinant collagen according to claim 1, characterized in that, The preparation method of the cedar bud extract is as follows: take cedar buds, wash and dry them, and pulverize them to 20-40 mesh to obtain cedar bud powder; add 10-15 times the mass of distilled water to the cedar bud powder, heat and extract, filter and collect the filtrate and filter residue; add 10-15 times the mass of distilled water to the filter residue, continue to heat and extract, and filter; then combine the filtrates, concentrate them, and freeze dry to obtain the extract.

3. The gel dressing containing recombinant collagen according to claim 2, characterized in that, The heating and extraction temperature is 90-100℃, and the time is 1-2 hours.

4. The gel dressing containing recombinant collagen according to claim 1, characterized in that, The recombinant collagen is recombinant type III humanized collagen with a molecular weight of 5 × 10⁻⁶. 4 -8×10 4 Da.

5. The gel dressing containing recombinant collagen according to claim 1, characterized in that, The carbomer is carbomer 980, and the poloxamer is poloxamer 407.

6. The gel dressing containing recombinant collagen according to claim 1, characterized in that, The molecular weight of the sodium hyaluronate is 1×10⁻⁶. 6 -2×10 6 Da.

7. The gel dressing containing recombinant collagen according to claim 1, characterized in that, The preservative is methylparaben or phenoxyethanol.

8. A method for preparing a gel dressing containing recombinant collagen according to any one of claims 1-7, characterized in that, The process includes the following steps: (1) Take half of the total amount of deionized water, add glycerin, then add carbomer, poloxamer, and xanthan gum, and stir to mix evenly to obtain a gel matrix; (2) Add disodium edetate to the gel matrix of step (1) and stir to mix evenly at 70-80℃ for 10-20 minutes to obtain mixture A, and cool to room temperature for later use; (3) Take the remaining amount of deionized water, add recombinant collagen, silver stellaria cyclic peptide, sodium hyaluronate, cedar bud extract, and preservative, and stir to mix evenly to obtain mixture B, and set aside for later use; (4) Add mixture B from step (3) to mixture A from step (2), stir to mix evenly, add triethanolamine, fill and sterilize to obtain the final product.

9. The use of a gel dressing containing recombinant collagen according to any one of claims 1-7 in the preparation of a vaginal gel dressing.

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