Recombinant collagen gel dressing and preparation method thereof

Abstract

The invention discloses a recombinant collagen gel dressing and a preparation method thereof, and relates to the technical field of biological medicines.The recombinant collagen gel dressing is prepared from 6-10 parts of modified sodium hyaluronate, 5-6 parts of recombinant human collagen, 4-5 parts of polydeoxyribonucleotide, 3-4 parts of traditional Chinese medicine extract and 1-3 parts of surface osmotic activator, the surface permeation active agent is one or a combination of more of polyoxyethylated castor oil, polysorbate and mannitol, and the polyoxyethylated castor oil, the polysorbate and the mannitol are used as the surface permeation active agent, so that skin surface solubilization can be supported, and a protein expression system can be adjusted to increase drug absorption; sodium hyaluronate and a disulfide cross-linked product are cross-linked to prepare gel, and the performance of the gel is tested, so that the wound healing speed and the tissue repair capability can be remarkably improved, and rapid vitality recovery and elasticity maintenance of skin cells and subcutaneous cells are facilitated.

Description

TECHNICAL FIELD

[0001] The present application relates to the technical field of biological medicine, and particularly relates to a recombinant collagen gel dressing and a preparation method thereof. BACKGROUND

[0002] The recombinant collagen generally refers to recombinant human collagen, which is prepared by using DNA recombination technology. The amino acid sequence can be designed and improved according to requirements. The gene sequence of the recombinant collagen is highly consistent with the human gene sequence, and the tissue compatibility is good. The recombinant collagen can be directly absorbed by the human body and participate in the construction of collagen to help tissue repair and site filling. There are various types of collagen in the human body, and the common types are type I, type II, type III, type V and type XI. In normal skin tissue, collagen mainly exists in the form of type I and type II collagen fibers. In the middle of infant skin, the content of type III collagen accounts for more than 80%, which has excellent elasticity and repair function. With the increase of age, the synthesis speed is slower than the loss speed, and the aging phenomenon such as wrinkles and relaxation occurs. The gene sequence of the recombinant human collagen is highly consistent with the human gene sequence, and the tissue compatibility is also good. The recombinant human collagen is not easy to induce the rejection reaction of the body, can smoothly enter the dermis and be absorbed and assimilated, enhance the expansion force of the damaged skin, restore the elasticity of the skin, and achieve the effects of repair, moisturizing, brightening and filling. Chinese Patent Publication No. CN115637279B discloses a recombinant human collagen for repairing skin and a preparation method thereof. The recombinant human collagen is obtained through seed culture, shake flask fermentation and centrifugal drying, and is applied to a dressing. The dressing includes a recombinant human collagen stock solution and a carrier. The recombinant human collagen stock solution and the carrier are aseptically packaged in an aluminum foil bag, and after sealing, low-temperature radiation sterilization is performed to obtain the dressing. The recombinant human collagen stock solution is composed of recombinant human collagen, a penetration enhancer, a thickening agent, lavender extract, a preservative and water. The penetration enhancer 1- (oxirane-2-ylmethyl) azepan-2-one prepared in the above-mentioned technology has a hydrocarbon chain with an epoxy group and a seven-membered ring lactam group with very strong polarity. In the mixing process with the recombinant human collagen, the epoxy group ring-opening reaction occurs with the amino group contained in the recombinant human collagen, the 1- (oxirane-2-ylmethyl) azepan-2-one is grafted to the recombinant human collagen, and a large number of hydrophilic groups are introduced. In the use process, the 1- (oxirane-2-ylmethyl) azepan-2-one can penetrate into the skin keratin layer and enter the hair follicle inner root sheath through the hair follicle pathway, react with the intercellular lipid or biological membrane lipid, increase the fluidity of the membrane ester, cause the cell structure to be loose, and promote absorption. At the same time, the presence of the hydrophilic group can increase the water content of the cell. The 1- (oxirane-2-ylmethyl) azepan-2-one can effectively fill the local collapse, improve the relaxation, tighten the skin and reduce the wrinkles. The above-mentioned technology only realizes the role of enhancing the penetration of recombinant human collagen, and has limited effect on improving the skin penetration, which is not conducive to deep and long-term skin nourishment and subcutaneous cells. Therefore, the present application provides a recombinant collagen gel dressing and a preparation method thereof to solve the above-mentioned problems. SUMMARY

[0003] The technical problem to be solved by the present application is that the acne-removing and whitening effect of the gel dressing is limited. The present application provides a recombinant collagen gel dressing and a preparation method thereof to solve the above-mentioned problems.

[0004] To achieve the above-mentioned purposes, the present application provides the following technical solutions. A recombinant collagen gel dressing and a preparation method thereof, the recombinant collagen gel dressing comprising: modified sodium hyaluronate 6-10 parts, recombinant human collagen 5-6 parts, polydeoxyribonucleotide 4-5 parts, traditional Chinese medicine extract 3-4 parts, and surface permeation active agent 1-3 parts, wherein the surface permeation active agent is one or a combination of several of polyoxyethylene castor oil, polysorbate, and mannitol. The preparation method of the recombinant collagen gel dressing comprises the following preparation steps: S1. Dissolve the modified sodium hyaluronate in an ethanol solution under light-proof conditions, and fully stir to form a homogeneous gel carrier; S2. Dissolve the recombinant collagen, polydeoxyribonucleotide, traditional Chinese medicine extract, and surface permeation active agent in purified water, and sterilize by ultraviolet rays to obtain a dressing carrier; S3. Under the condition of ultraviolet light irradiation, mix and stir the dressing carrier prepared in S2 into the gel carrier prepared in S1 to obtain a recombinant collagen gel dressing.

[0005] Further, the wavelength of the ultraviolet rays in step S3 is 150-450 nm, and the irradiation time is 5-10 min.

[0006] Further, the photoinitiator in step S3 is 1-vinyl-2-pyrrolidone and triethanolamine.

[0007] Further, the preparation method of the modified sodium hyaluronate comprises the following steps: A1. Dissolve sodium hyaluronate in water, and add a certain amount of hydrazine disulfide crosslinking agent while stirring; A2. Adjust and maintain the pH value of the mixed solution to be 4.75 by using 1.0 mo / L hydrogen chloride, and add solid carbodiimide and stir; A3. Add 1.0 mol / L sodium hydroxide until the pH value increases to 8.5, and stir until the reaction stops; A4: Continue stirring for 24 hours, adjust the pH to 3.5, and transfer the acidic solution to a dialysis tube; A5: After dialysis with dilute hydrogen chloride, centrifugation, freeze-drying, and purification, modified sodium hyaluronate is obtained.

[0008] Further, in step A1, the disulfide crosslinker is either 3,3'-dithiopropionic acid or 4,4'-dithiobutyric acid.

[0009] Furthermore, the dialysis duration in step A5 is 6-36 hours, with the water changed every 6 hours.

[0010] Furthermore, the centrifugation conditions in step A5 are: centrifugation at 8000-10000 r / min for 5-12 min.

[0011] Furthermore, the freeze-drying conditions in step A5 are: freeze-drying at -80℃ for 48-72 hours.

[0012] Compared with the prior art, the beneficial effects of the present invention are as follows: 1. The present invention provides a recombinant collagen gel dressing and its preparation method, which uses polyoxyethylene castor oil, polysorbate ester, and mannitol as surface permeability surfactants. It can not only support skin surface solubilization, but also regulate the protein expression system to increase drug absorption and improve bioavailability. 2. This invention provides a recombinant collagen gel dressing and its preparation method. Sodium hyaluronate is cross-linked with disulfide cross-linking agent to form a gel. Its performance is tested and it can significantly improve wound healing speed and tissue repair ability, which is conducive to the rapid recovery of skin cells and subcutaneous cells and maintain elasticity. 3. This invention discloses a recombinant collagen gel dressing and its preparation method, which combines recombinant human collagen, modified sodium hyaluronate, and a surfactant. The surfactant enhances the absorption capacity of the skin and subcutaneous cells of the recombinant human collagen, while the modified sodium hyaluronate enhances the self-healing ability of the skin and subcutaneous cells. The three components work synergistically to improve skin permeability, increase membrane fluidity, and loosen the cell structure. This effectively fills local skin depressions, improves sagging, tightens the skin, reduces wrinkles, and facilitates the release of medicinal extracts, deeply nourishing the skin. Detailed Implementation

[0013] The technical solutions in the embodiments of the present invention will be clearly and completely described below. Obviously, the described embodiments are only some embodiments of the present invention, and not all embodiments. Based on the embodiments of the present invention, all other embodiments obtained by those skilled in the art without creative effort are within the scope of protection of the present invention. Example

[0014] A recombinant collagen gel dressing and its preparation method are disclosed. The recombinant collagen gel dressing comprises: 10 parts modified sodium hyaluronate, 6 parts recombinant human collagen, 5 parts polydeoxyribonucleotide, 4 parts traditional Chinese medicine extract, and 3 parts surfactant. The surfactant is prepared by mixing polyoxyethylene castor oil, polysorbate, and mannitol in a 1:1:1 ratio. The preparation method of the recombinant collagen gel dressing includes the following preparation steps: S1. Dissolve the modified sodium hyaluronate in an ethanol solution under light-protected conditions, and stir thoroughly until a homogeneous gel carrier is formed. S2. Take recombinant collagen, polydeoxyribonucleic acid, traditional Chinese medicine extract and surfactant, mix and dissolve in purified water, sterilize with ultraviolet light, and use as a dressing carrier. S3. Under ultraviolet light irradiation, the dressing carrier prepared in S2 is added to the gel carrier prepared in S1 and mixed and stirred to obtain recombinant collagen gel dressing.

[0015] Furthermore, in step S3, the ultraviolet wavelength is 400nm and the irradiation time is 10min.

[0016] Furthermore, the photoinitiator in step S3 is 1-vinyl-2-pyrrolidone and triethanolamine.

[0017] Furthermore, the preparation method of the modified sodium hyaluronate includes the following steps: A1: Dissolve sodium hyaluronate in water, and add a certain amount of hydrazine disulfide crosslinking product while stirring; A2: Adjust and maintain the pH of the mixture at 4.75 with 1.0 mol / L hydrogen chloride, then add solid carbodiimide and stir. A3: Add 1.0 mol / L sodium hydroxide until the pH rises to 8.5, and stir until the reaction stops; A4: Continue stirring for 24 hours, adjust the pH to 3.5, and transfer the acidic solution to a dialysis tube; A5: After dialysis with dilute hydrogen chloride, centrifugation, freeze-drying, and purification, modified sodium hyaluronate is obtained.

[0018] Furthermore, the disulfide crosslinker in step A1 is 3,3'-dithiopropionic acid.

[0019] Furthermore, the dialysis duration in step A5 is 36 hours, with the water changed every 6 hours.

[0020] Furthermore, the centrifugation conditions in step A5 are: centrifugation at 8000 r / min for 8 min.

[0021] Furthermore, the freeze-drying conditions in step A5 are: freeze-drying at -80℃ for 60 hours. Example

[0022] Compared with Example 1, the raw material ratio is different in the preparation process of recombinant human collagen dressing in this embodiment, while the rest is the same as in Example 1.

[0023] A recombinant collagen gel dressing and its preparation method are disclosed. The recombinant collagen gel dressing comprises: 6 parts modified sodium hyaluronate, 6 parts recombinant human collagen, 5 parts polydeoxyribonucleotide, 4 parts traditional Chinese medicine extract, and 3 parts surfactant. The surfactant is prepared by mixing polyoxyethylene castor oil, polysorbate, and mannitol in a 1:1:1 ratio. The preparation method of the recombinant collagen gel dressing includes the following preparation steps: S1. Dissolve the modified sodium hyaluronate in an ethanol solution under light-protected conditions, and stir thoroughly until a homogeneous gel carrier is formed. S2. Take recombinant collagen, polydeoxyribonucleic acid, traditional Chinese medicine extract and surfactant, mix and dissolve in purified water, sterilize with ultraviolet light, and use as a dressing carrier. S3. Under ultraviolet light irradiation, the dressing carrier prepared in S2 is added to the gel carrier prepared in S1 and mixed and stirred to obtain recombinant collagen gel dressing.

[0024] Furthermore, in step S3, the ultraviolet wavelength is 400nm and the irradiation time is 10min.

[0025] Furthermore, the photoinitiator in step S3 is 1-vinyl-2-pyrrolidone and triethanolamine.

[0026] Furthermore, the preparation method of the modified sodium hyaluronate includes the following steps: A1: Dissolve sodium hyaluronate in water, and add a certain amount of hydrazine disulfide crosslinking product while stirring; A2: Adjust and maintain the pH of the mixture at 4.75 with 1.0 mol / L hydrogen chloride, then add solid carbodiimide and stir. A3: Add 1.0 mol / L sodium hydroxide until the pH rises to 8.5, and stir until the reaction stops; A4: Continue stirring for 24 hours, adjust the pH to 3.5, and transfer the acidic solution to a dialysis tube; A5: After dialysis with dilute hydrogen chloride, centrifugation, freeze-drying, and purification, modified sodium hyaluronate is obtained.

[0027] Furthermore, the disulfide crosslinker in step A1 is 3,3'-dithiopropionic acid.

[0028] Furthermore, the dialysis duration in step A5 is 36 hours, with the water changed every 6 hours.

[0029] Furthermore, the centrifugation conditions in step A5 are: centrifugation at 8000 r / min for 8 min.

[0030] Furthermore, the freeze-drying conditions in step A5 are: freeze-drying at -80℃ for 60 hours. Example

[0031] Compared with Example 1, the raw material ratio is different in the preparation process of recombinant human collagen dressing in this embodiment, while the rest is the same as in Example 1.

[0032] A recombinant collagen gel dressing and its preparation method are disclosed. The recombinant collagen gel dressing comprises: 10 parts modified sodium hyaluronate, 5 parts recombinant human collagen, 4 parts polydeoxyribonucleotides, 3 parts traditional Chinese medicine extract, and 1 part surfactant. The surfactant is prepared by mixing polyoxyethylene castor oil, polysorbate, and mannitol in a 1:1:1 ratio. The preparation method of the recombinant collagen gel dressing includes the following preparation steps: S1. Dissolve the modified sodium hyaluronate in an ethanol solution under light-protected conditions, and stir thoroughly until a homogeneous gel carrier is formed. S2. Take recombinant collagen, polydeoxyribonucleic acid, traditional Chinese medicine extract and surfactant, mix and dissolve in purified water, sterilize with ultraviolet light, and use as a dressing carrier. S3. Under ultraviolet light irradiation, the dressing carrier prepared in S2 is added to the gel carrier prepared in S1 and mixed and stirred to obtain recombinant collagen gel dressing.

[0033] Furthermore, in step S3, the ultraviolet wavelength is 400nm and the irradiation time is 10min.

[0034] Furthermore, the photoinitiator in step S3 is 1-vinyl-2-pyrrolidone and triethanolamine.

[0035] Furthermore, the preparation method of the modified sodium hyaluronate includes the following steps: A1: Dissolve sodium hyaluronate in water, and add a certain amount of hydrazine disulfide crosslinking product while stirring; A2: Adjust and maintain the pH of the mixture at 4.75 with 1.0 mol / L hydrogen chloride, then add solid carbodiimide and stir. A3: Add 1.0 mol / L sodium hydroxide until the pH rises to 8.5, and stir until the reaction stops; A4: Continue stirring for 24 hours, adjust the pH to 3.5, and transfer the acidic solution to a dialysis tube; A5: After dialysis with dilute hydrogen chloride, centrifugation, freeze-drying, and purification, modified sodium hyaluronate is obtained.

[0036] Furthermore, the disulfide crosslinker in step A1 is 3,3'-dithiopropionic acid.

[0037] Furthermore, the dialysis duration in step A5 is 36 hours, with the water changed every 6 hours.

[0038] Furthermore, the centrifugation conditions in step A5 are: centrifugation at 8000 r / min for 8 min.

[0039] Furthermore, the freeze-drying conditions in step A5 are: freeze-drying at -80℃ for 60 hours. Example

[0040] Compared with Example 1, the raw material ratio is different in the preparation process of recombinant human collagen dressing in this embodiment, while the rest is the same as in Example 1.

[0041] A recombinant collagen gel dressing and its preparation method are disclosed. The recombinant collagen gel dressing comprises: 6 parts modified sodium hyaluronate, 5 parts recombinant human collagen, 4 parts polydeoxyribonucleotides, and 3 parts traditional Chinese medicine extract. The surfactant is prepared by mixing polyoxyethylene castor oil, polysorbate, and mannitol in a ratio of 1:1:1. The preparation method of the recombinant collagen gel dressing includes the following preparation steps: S1. Dissolve the modified sodium hyaluronate in an ethanol solution under light-protected conditions, and stir thoroughly until a homogeneous gel carrier is formed. S2. Take recombinant collagen, polydeoxyribonucleic acid, traditional Chinese medicine extract and surfactant, mix and dissolve in purified water, sterilize with ultraviolet light, and use as a dressing carrier. S3. Under ultraviolet light irradiation, the dressing carrier prepared in S2 is added to the gel carrier prepared in S1 and mixed and stirred to obtain recombinant collagen gel dressing.

[0042] Furthermore, in step S3, the ultraviolet wavelength is 400nm and the irradiation time is 10min.

[0043] Furthermore, the photoinitiator in step S3 is 1-vinyl-2-pyrrolidone and triethanolamine.

[0044] Furthermore, the preparation method of the modified sodium hyaluronate includes the following steps: A1: Dissolve sodium hyaluronate in water, and add a certain amount of hydrazine disulfide crosslinking product while stirring; A2: Adjust and maintain the pH of the mixture at 4.75 with 1.0 mol / L hydrogen chloride, then add solid carbodiimide and stir. A3: Add 1.0 mol / L sodium hydroxide until the pH rises to 8.5, and stir until the reaction stops; A4: Continue stirring for 24 hours, adjust the pH to 3.5, and transfer the acidic solution to a dialysis tube; A5: After dialysis with dilute hydrogen chloride, centrifugation, freeze-drying, and purification, modified sodium hyaluronate is obtained.

[0045] Furthermore, the disulfide crosslinker in step A1 is 3,3'-dithiopropionic acid.

[0046] Furthermore, the dialysis duration in step A5 is 36 hours, with the water changed every 6 hours.

[0047] Furthermore, the centrifugation conditions in step A5 are: centrifugation at 8000 r / min for 8 min.

[0048] Furthermore, the freeze-drying conditions in step A5 are: freeze-drying at -80℃ for 60 hours.

[0049] The recombinant collagen gel dressings prepared in Examples 1 to 4 were used in acne treatment trials on four volunteers, as follows: Four volunteers, aged 20-28, were selected. All volunteers had a small amount of acne and exhibited redness and itching. Instructions for use: After cleansing the face, apply the provided dressing once daily for 90 consecutive days. The dressing was changed every Monday morning at 11:00 AM. Photos of the test area were taken under identical lighting conditions, and the results were saved for comparison. The treatment results are shown in the table below: Recovery phase Volunteer 1 Volunteer 2 Volunteer 3 Volunteer 4 Few pimples with surrounding redness 0 0 0 0 Small red pimples with redness subsiding 2 2 2 3 No redness area, partial scabbing off 3 4 4 6 Skin pimple pits lightened with post-inflammatory hyperpigmentation 6 6 7 12 Skin smooth and even with post-inflammatory hyperpigmentation / 7 9 / Skin smooth and fine without post-inflammatory hyperpigmentation 8 8 / / Experimental conclusion: The rehabilitation effect of the recombinant collagen gel dressings prepared in Examples 1 to 4 decreased progressively, proving that the content of surfactant has a significant impact on the rehabilitation effect.

[0050] Although embodiments of the invention have been shown and described, it will be understood by those skilled in the art that various changes, modifications, substitutions and alterations can be made to these embodiments without departing from the principles and spirit of the invention, the scope of which is defined by the appended claims and their equivalents.

Claims

1. A recombinant collagen gel dressing and its preparation method, characterized in that, The recombinant collagen gel dressing comprises: 6-10 parts of modified sodium hyaluronate, 5-6 parts of recombinant human collagen, 4-5 parts of polydeoxyribonucleotides, 3-4 parts of traditional Chinese medicine extract, and 1-3 parts of surfactant, wherein the surfactant is one or a combination of polyoxyethylene castor oil, polysorbate, and mannitol. The preparation method of the recombinant collagen gel dressing includes the following preparation steps: S1. Dissolve the modified sodium hyaluronate in an ethanol solution under light-protected conditions, and stir thoroughly until a homogeneous gel carrier is formed. S2. Take recombinant collagen, polydeoxyribonucleic acid, traditional Chinese medicine extract and surfactant, mix and dissolve in purified water, sterilize with ultraviolet light, and use as a dressing carrier. S3. Under ultraviolet light irradiation, the dressing carrier prepared in S2 is added to the gel carrier prepared in S1 and mixed and stirred to obtain recombinant collagen gel dressing.

2. The recombinant collagen gel dressing and its preparation method according to claim 1, characterized in that, In step S3, the ultraviolet wavelength is 150–450 nm, and the irradiation time is 5–10 min.

3. The recombinant collagen gel dressing and its preparation method according to claim 1, characterized in that, The photoinitiator in step S3 is 1-vinyl-2-pyrrolidone and triethanolamine.

4. The recombinant collagen gel dressing and its preparation method according to claim 1, characterized in that, The preparation method of the modified sodium hyaluronate includes the following steps: A1: Dissolve sodium hyaluronate in water, and add a certain amount of hydrazine disulfide crosslinking product while stirring; A2: Adjust and maintain the pH of the mixture at 4.75 with 1.0 mol / L hydrogen chloride, then add solid carbodiimide and stir. A3: Add 1.0 mol / L sodium hydroxide until the pH rises to 8.5, and stir until the reaction stops; A4: Continue stirring for 24 hours, adjust the pH to 3.5, and transfer the acidic solution to a dialysis tube; A5: After dialysis with dilute hydrogen chloride, centrifugation, freeze-drying, and purification, modified sodium hyaluronate is obtained.

5. The recombinant collagen gel dressing and its preparation method according to claim 4, characterized in that: In step A1, the disulfide crosslinker is either 3,3'-dithiopropionic acid or 4,4'-dithiobutyric acid.

6. The recombinant collagen gel dressing and its preparation method according to claim 4, characterized in that, The dialysis duration in step A5 is 6-36 hours, with the water changed every 6 hours.

7. The recombinant collagen gel dressing and its preparation method according to claim 4, characterized in that: The centrifugation conditions in step A5 are: centrifugation at 8000-10000 r / min for 5-12 min.

8. The recombinant collagen gel dressing and its preparation method according to claim 4, characterized in that: The freeze-drying conditions in step A5 are: freeze-drying at -80℃ for 48-72 hours.