CN115845126A - Bionic collagen liquid dressing and preparation method thereof - Google Patents

Bionic collagen liquid dressing and preparation method thereof Download PDF

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CN115845126A
CN115845126A CN202211672297.7A CN202211672297A CN115845126A CN 115845126 A CN115845126 A CN 115845126A CN 202211672297 A CN202211672297 A CN 202211672297A CN 115845126 A CN115845126 A CN 115845126A
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acid
collagen
liquid dressing
skin
supernatant
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杜明春
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Shenzhen Junxing Biotechnology Co ltd
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Shenzhen Junxing Biotechnology Co ltd
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Abstract

The invention relates to the field of biological materials, in particular to a bionic collagen liquid dressing and a preparation method thereof. On the basis of ensuring that the dressing has the homogeneous flow characteristic, the invention simulates the microenvironment of skin tissues from the aspects of collagen type, component simulation, acid environment and the like, realizes the bionic effect and promotes the repair and regeneration of skin wound surfaces.

Description

Bionic collagen liquid dressing and preparation method thereof
Technical Field
The invention relates to a preparation method of a biological material, in particular to a bionic collagen liquid dressing and a preparation method thereof.
Background
The first animal experiment of doctor Winter at university of london in 1962 confirmed that wounds heal 1-fold faster in a moist environment than in a dry environment. Human studies with Hinman in 1963 confirmed the scientificity of moist healing. The concept of "wet wound healing" was gradually widely accepted in the 70's of the 20 th century. The liquid dressing is a novel medical dressing developed according to a 'moist healing theory', and compared with the traditional dressing, the liquid dressing can keep a wound surface moist, is beneficial to regeneration and repair of wound surface tissues, shortens the wound healing time, reduces the dosage of the medical dressing, and obviously shortens the nursing time.
On the other hand, collagen is the protein with the most abundant content and the most extensive distribution in vertebrates, and the collagen dressing can promote the anabolism and cell regeneration of skin tissues, improve the local temperature and humidity of the skin, accelerate the blood circulation and metabolism of local skin cells, and is beneficial to the regeneration and the replacement of the skin cells, thereby accelerating the tissue regeneration and the repair. However, the collagen dressing is often provided in the form of a solid dressing such as a film type, a sponge type, or a bonded nonwoven fabric, and the collagen component in the liquid dressing is also often provided in the form of an agglomerate or gel, in which collagen molecules do not have fluidity.
The invention provides a bionic collagen liquid dressing taking a collagen aqueous solution as a main component and a preparation method thereof from the viewpoint of solving the problem of collagen molecule mobility in the liquid dressing, and on the basis of ensuring that the dressing has a homogeneous flow characteristic, the bionic collagen liquid dressing simulates a skin tissue microenvironment from the aspects of collagen type, component simulation, acid environment and the like, realizes a bionic effect and promotes the repair and regeneration of a skin wound.
Disclosure of Invention
The invention aims to provide a bionic collagen liquid dressing and a preparation method thereof.
A preparation method of the bionic collagen liquid dressing sequentially comprises the following steps:
(1) Dissolving solid collagen material in water, fully dissolving, centrifuging, and taking supernatant; the solid collagen material is selected from one or the combination of more than two types of types I, III, V, VI, VII, VIII, XII, XIII, XIV, XVI and XXVIII; the total concentration of each type of collagen in the supernatant is 0.001wt.% or more but 5wt.% or less;
(2) Adjusting the pH value of the supernatant obtained in the step (1) by an alkaline solution or an acidic solution; the acidic solution comprises at least one of salicylic acid, lactic acid, sulfuric acid, tartaric acid, citric acid, phosphoric acid, oxalic acid, acetic acid, oxalic acid, succinic acid, hydrochloric acid, maleic acid, benzoic acid, nitric acid, malic acid, nicotinic acid, sodium dihydrogen phosphate and formic acid; the alkaline solution comprises at least one of tetramethylethylenediamine, sodium hydroxide, triethylamine, sodium carbonate, potassium hydroxide, calcium hydroxide, magnesium hydroxide, ammonia water, disodium hydrogen phosphate and sodium bicarbonate;
(3) Adding a skin-simulating component into the solution obtained in the step (2); the skin-simulating component comprises at least one of hyaluronic acid, elastin, keratin, chondroitin sulfate, mucopolysaccharide, ceramide, vitamin A, vitamin E, D-panthenol, urea, sodium lactate and lactic acid; the concentration of the skin-mimicking ingredient is 0.00001wt.% or more but 2wt.% or less;
(4) Adding a simulated body fluid component to the solution obtained in step (3), wherein the simulated body fluid component comprises Na + 、K + 、Mg 2+ 、Ca 2 + 、Cl - 、HCO 3 - 、HPO 4 2- 、SO 4 2- 、CO 3 2- 、PO 4 3- 、H 2 PO 4 - One or a combination of two or more of them;
(5) Adding a protective molecule into the solution obtained in the step (4), wherein the protective molecule comprises at least one of sodium dodecyl sulfate, sodium dodecyl benzene sulfonate, tween, triton, ethylene glycol, phenylalanine, proline, lecithin, glutamic acid, lysine, cysteine, mangiferin, glycine, aspartic acid, n-butanol, propylene glycol, sodium ethylene diamine tetracetate, stearic acid, span, glycerol and gelatin;
(6) And (4) carrying out low-temperature irradiation treatment on the solution obtained in the step (5).
The invention relates to a preparation method of a bionic collagen liquid dressing, wherein the low-temperature range of the low-temperature irradiation treatment in the step (6) is-20-5 ℃.
The invention discloses a preparation method of a bionic collagen liquid dressing, wherein the irradiation treatment comprises at least one of gamma ray irradiation, electron beam irradiation and X-ray irradiation.
The invention relates to a preparation method of a bionic collagen liquid dressing, wherein the final ion concentration range of each ion in the bionic collagen liquid dressing is 0.01-0.5M.
The invention relates to a preparation method of a bionic collagen liquid dressing, wherein the mass ratio of protective molecules to collagen is (1).
The invention relates to a preparation method of a bionic collagen liquid dressing, wherein the final pH value range of supernatant in the step (2) is more than or equal to 4.0 and less than or equal to 7.8; preferably, the final range of the pH value of the supernatant in the step (2) is more than or equal to 4.1 and less than or equal to 6.5; preferably, the final range of the pH value of the supernatant in the step (2) is greater than or equal to 4.2 and less than or equal to 5.6.
The bionic collagen liquid dressing prepared by the preparation method of any bionic collagen liquid dressing.
The bionic collagen liquid dressing can be used for the wound surface of a human body and also can be used for the wound surface of an animal body.
The invention has the beneficial effects that:
the invention provides a bionic collagen liquid dressing taking a collagen aqueous solution as a main component and a preparation method thereof from the viewpoint of solving the problem of collagen molecule mobility in the liquid dressing, wherein the pH value of the collagen aqueous solution is regulated and controlled to be weak and acidic in step (2), collagen molecules are dispersed as far as possible, and the self-assembly behavior of the collagen molecules is reduced; the skin simulation component is added in the step (3), the simulated body fluid component is added in the step (4), and the protective molecule is added in the step (5), so that the solution environment can be regulated, the self-assembly behavior of collagen molecules can be reduced in a mutual cooperation manner, the homogeneous fluidity of the dressing can be ensured, the skin tissue microenvironment can be simulated from the aspects of collagen types, component simulation, acid environment and the like on the basis of ensuring that the dressing has the homogeneous fluidity, the bionic effect can be realized, the repair and regeneration of the skin wound surface can be promoted, and the repair effect of the liquid dressing can be enhanced.
Drawings
FIG. 1 is a graph showing turbidity measurements of a collagen liquid dressing (FIG. 1A is a sample of example 1, FIG. 1B is a sample of example 2, FIG. 1C is a sample of example 3, FIG. 1D is a sample of example 4, FIG. 1E is a sample of example 5, FIG. 1F is a sample of comparative example 1, FIG. 1G is a sample of comparative example 2)
FIG. 2 is a graph showing the adhesion of a collagen liquid dressing to the skin surface (FIG. 2A is a sample of example 1, FIG. 2B is a sample of example 2, FIG. 2C is a sample of example 3, FIG. 2D is a sample of example 4, FIG. 2E is a sample of example 5, FIG. 2F is a sample of comparative example 1, FIG. 2G is a sample of comparative example 2)
FIG. 3 is a diagram showing the effect of the collagen liquid dressing on repairing the skin wound
FIG. 4 graph showing HE staining effect of wound repair in rat skin (FIG. 4A is sample of example 1, FIG. 4B is sample of comparative example 1)
Detailed Description
The invention provides a bionic collagen liquid dressing taking a collagen aqueous solution as a main component and a preparation method thereof from the viewpoint of solving the problem of collagen molecule mobility in the liquid dressing, and the specific contents are as follows:
the invention relates to a preparation method of a bionic collagen liquid dressing, which sequentially comprises the following steps:
(1) Dissolving solid collagen material in water, fully dissolving, centrifuging, and taking supernatant; the solid collagen material is selected from one or more of type I, type III, type V, type VI, type VII, type VIII, type XII, type XIII, type XIV, type XVI and type XXVIII collagen; the total concentration of each type of collagen in the supernatant is 0.001wt.% or more but 5wt.% or less;
(2) Adjusting the pH value of the supernatant obtained in the step (1) by using an alkaline solution or an acidic solution; the acidic solution comprises at least one of salicylic acid, lactic acid, sulfuric acid, tartaric acid, citric acid, phosphoric acid, oxalic acid, acetic acid, oxalic acid, succinic acid, hydrochloric acid, maleic acid, benzoic acid, nitric acid, malic acid, nicotinic acid, sodium dihydrogen phosphate and formic acid; the alkaline solution comprises at least one of tetramethylethylenediamine, sodium hydroxide, triethylamine, sodium carbonate, potassium hydroxide, calcium hydroxide, magnesium hydroxide, ammonia water, disodium hydrogen phosphate and sodium bicarbonate;
(3) Adding a skin-simulating component into the solution obtained in the step (2); the skin-simulating component comprises at least one of hyaluronic acid, elastin, keratin, chondroitin sulfate, mucopolysaccharide, ceramide, vitamin A, vitamin E, D-panthenol, urea, sodium lactate and lactic acid; the concentration of the skin-mimicking ingredient is 0.00001wt.% or more but 2wt.% or less;
(4) Adding a simulated body fluid component to the solution obtained in step (3), wherein the simulated body fluid component comprises Na + 、K + 、Mg 2+ 、Ca 2 + 、Cl - 、HCO 3 - 、HPO 4 2- 、SO 4 2- 、CO 3 2- 、PO 4 3- 、H 2 PO 4 - One or a combination of two or more of them;
(5) Adding a protective molecule into the solution obtained in the step (4), wherein the protective molecule comprises at least one of sodium dodecyl sulfate, sodium dodecyl benzene sulfonate, tween, triton, ethylene glycol, phenylalanine, proline, lecithin, glutamic acid, lysine, cysteine, mangiferin, glycine, aspartic acid, n-butyl alcohol, propylene glycol, sodium ethylene diamine tetraacetate, stearic acid, span, glycerol and gelatin;
(6) And (4) carrying out low-temperature irradiation treatment on the solution obtained in the step (5).
The invention relates to a preparation method of a bionic collagen liquid dressing, wherein the low-temperature range of the low-temperature irradiation treatment in the step (6) is-20-5 ℃.
The invention relates to a preparation method of a bionic collagen liquid dressing, wherein irradiation treatment comprises at least one of gamma ray irradiation, electron beam irradiation and X-ray irradiation.
The invention relates to a preparation method of a bionic collagen liquid dressing, wherein the final ion concentration range of each ion in the bionic collagen liquid dressing is 0.01-0.5M.
The invention relates to a preparation method of a bionic collagen liquid dressing, wherein the mass ratio of protective molecules to collagen is 1.
The invention relates to a preparation method of a bionic collagen liquid dressing, wherein the final pH value range of supernatant in the step (2) is more than or equal to 4.0 but less than or equal to 7.8; preferably, the final range of the pH value of the supernatant in the step (2) is more than or equal to 4.1 and less than or equal to 6.5; preferably, the final range of pH of the supernatant in step (2) is 4.2 or more but 5.6 or less.
The invention relates to a bionic collagen liquid dressing, which is prepared by using any one of the preparation methods of the bionic collagen liquid dressing. The bionic collagen liquid dressing can be used for the wound surface of a human body and also can be used for the wound surface of an animal body.
The preparation method of the bionic collagen liquid dressing comprises the steps of (1) selecting a collagen material in the bionic collagen liquid dressing according to the type of collagen contained in skin tissues, determining the proportion of various types of collagen in the dressing according to the specific effect of the dressing, and determining the total concentration of collagen in the dressing according to the application mode of the dressing, wherein for example, a low-concentration collagen aqueous solution is used in a spraying mode, and a high-concentration collagen aqueous solution is used in a smearing mode.
Since acidic substances such as urea, uric acid, salts, lactic acid, amino acids, and free fatty acids remain on the skin surface of a human body, the skin surface is often weakly acidic. Skin is best at absorbing nutrients only within the normal pH range, i.e., mildly acidic, where the skin's ability to resist external aggression, elasticity, luster, moisture, etc., are best. Therefore, the pH of the aqueous collagen solution in step (2) of the present invention is controlled according to the pH of the skin tissue in the application of the dressing, and the aqueous collagen solution is generally weakly acidic. Meanwhile, the acidic environment of the aqueous solution is more beneficial to fully dissolving the collagen material and keeping the bionic collagen liquid dressing homogeneous and fluid.
In order to regulate and control the pH value of the collagen water solution to be matched with the pH value of the skin tissue to be repaired, a small amount of alkaline solution or acidic solution is added in the step (2) of the method, and the concentration of the collagen in the water solution is not changed obviously; meanwhile, the alkaline solution or the acidic solution is selected as an agent with good biocompatibility as much as possible.
In the step (3), a small amount of skin simulation components are selected and added according to the components of the skin tissue, so that the components of the collagen liquid dressing are matched with the skin tissue to be repaired as far as possible, and the content of the added skin simulation components is determined according to the component content of the skin tissue and is matched with the skin tissue to be repaired as far as possible.
In order to regulate and control the ionic environment of the bionic collagen liquid dressing to be matched with the ionic environment of the skin tissue to be pre-repaired, simulated body fluid components are added in the step (4) of the preparation method of the bionic collagen liquid dressing, so that the stability of all the components in the aqueous solution is enhanced, the agglomeration phenomenon is reduced, and the biocompatibility of the bionic collagen liquid dressing is enhanced.
According to the invention, the protective molecules are added in the step (5) and are combined with each component in the aqueous solution through weak interaction force such as hydrogen bonds, so that the components in the aqueous solution are prevented from being crosslinked in the irradiation process.
In the step (6), the irradiation is carried out under low temperature conditions, so that adverse effects on components in the aqueous solution caused by overhigh temperature in the irradiation process are avoided, and cross-linking of the components in the aqueous solution in the irradiation process is avoided.
Due to the self-assembly behavior of collagen molecules, under the influence of factors such as molecular environment, pH value and temperature, the collagen molecules in the liquid dressing are usually aggregated and agglomerated and even exist in a gel form, the collagen molecules have no fluidity, and the repair effect of the collagen material is influenced even if the wound surface is kept moist. In order to enhance the repairing effect of the liquid dressing, the invention provides a bionic collagen liquid dressing taking a collagen aqueous solution as a main component and a preparation method thereof from the viewpoint of solving the problem of collagen molecule mobility in the liquid dressing, wherein the pH value of the collagen aqueous solution is regulated and controlled to be weak and acidic, the collagen molecules are dispersed as far as possible, and the self-assembly behavior of the collagen molecules is reduced; the skin simulation component is added in the step (3), the simulated body fluid component is added in the step (4), and the protective molecule is added in the step (5), so that the solution environment can be regulated, the self-assembly behavior of collagen molecules can be reduced in a mutual cooperation manner, the homogeneous fluidity of the dressing can be ensured, the microenvironment of skin tissues can be simulated from the aspects of collagen types, component simulation, acid environment and the like on the basis of ensuring that the dressing has the homogeneous flowing characteristic, the bionic effect can be realized, the repair and regeneration of skin wounds can be promoted, and the repair effect of the liquid dressing can be enhanced.
The effects of the present invention will be described below with reference to specific examples.
Example 1
A preparation method of the bionic collagen liquid dressing sequentially comprises the following steps:
(1) Placing 1g I collagen and 0.5g type III collagen material into 100g deionized water, fully dissolving, centrifuging, and taking supernatant;
(2) Adjusting the pH value of the supernatant obtained in the step (1) to 6.5 by 0.5M triethylamine or 0.1M malic acid;
(3) Adding 0.1wt.% hyaluronic acid to the solution obtained in step (2);
(4) Adding 0.02g KH into the solution obtained in the step (3) 2 PO 4 And 0.005g of Na 2 CO 3 ；
(5) Adding tween into the solution obtained in the step (4), wherein the mass ratio of tween to collagen is 10;
(6) Co treatment of the solution obtained in step (5) at 0 DEG C 60 And (5) performing irradiation treatment.
Example 2
The preparation method of the bionic collagen liquid dressing sequentially comprises the following steps:
(1) Placing 2g I collagen, 0.1g V collagen and 0.1g VII collagen material into 500g deionized water, fully dissolving, centrifuging, and collecting supernatant;
(2) Adjusting the pH value of the supernatant obtained in the step (1) to 7.8 by 0.2M sodium carbonate or 1M oxalic acid;
(3) Adding 0.05wt.% elastin and 0.00001wt.% chondroitin sulfate to the solution obtained in step (2);
(4) Adding 1g of Na into the solution obtained in the step (3) 2 HPO 4 And 0.05g Mg (HCO) 3 ) 2 ；
(5) Adding lysine and glycerol into the solution obtained in the step (4), wherein the mass ratio of the lysine to the collagen is 1;
(6) Carrying out Cs treatment on the solution obtained in the step (5) at the temperature of 5 DEG C 137 And (5) performing irradiation treatment.
Example 3
A preparation method of the bionic collagen liquid dressing sequentially comprises the following steps:
(1) Placing 0.5g I collagen, 0.01g XII collagen and 0.01g XIV collagen material into 200g deionized water, fully dissolving, centrifuging, and collecting supernatant;
(2) Adjusting the pH value of the supernatant obtained in the step (1) to 4.2 by 1M potassium hydroxide or 0.2M citric acid;
(3) Adding 0.05wt.% elastin and 0.00001wt.% chondroitin sulfate to the solution obtained in step (2);
(4) Adding 2g of sodium lactate and 1g of mucopolysaccharide into the solution obtained in the step (3);
(5) Adding gelatin and ethylene glycol into the solution obtained in the step (4), wherein the mass ratio of the gelatin to the collagen is 1:5, and the mass ratio of the ethylene glycol to the collagen is 100;
(6) And (4) carrying out electron accelerator irradiation treatment on the solution obtained in the step (5) at the temperature of-10 ℃.
Example 4
A preparation method of the bionic collagen liquid dressing sequentially comprises the following steps:
(1) Placing 0.01g I collagen material into 1000g deionized water, fully dissolving, centrifuging, and taking supernatant;
(2) Adjusting the pH value of the supernatant obtained in the step (1) to 5.6 by 0.5M triethylamine or 0.1M malic acid;
(3) Adding 2wt.% of hyaluronic acid into the solution obtained in the step (2);
(4) Adding 0.02g KH into the solution obtained in step (3) 2 PO 4 And 0.005g of Na 2 CO 3 ；
(5) Adding Tween into the solution obtained in the step (4), wherein the mass ratio of Tween to collagen is 10;
(6) Performing Co treatment on the solution obtained in the step (5) at the temperature of minus 20 DEG C 60 And (5) performing irradiation treatment.
Example 5
A preparation method of the bionic collagen liquid dressing sequentially comprises the following steps:
(1) Placing 8.5g I collagen, 0.5g XII collagen and 1g XIV collagen material into 200g deionized water, fully dissolving, centrifuging, and collecting supernatant;
(2) Adjusting the pH value of the supernatant obtained in the step (1) to 4.0 by 1M potassium hydroxide or 0.2M acetic acid;
(3) Adding 0.05wt.% elastin and 0.00001wt.% chondroitin sulfate to the solution obtained in step (2);
(4) Adding 2g of sodium lactate and 1g of mucopolysaccharide into the solution obtained in the step (3);
(5) Adding gelatin and ethylene glycol into the solution obtained in the step (4), wherein the mass ratio of the gelatin to the collagen is 1:5, and the mass ratio of the ethylene glycol to the collagen is 100;
(6) And (4) carrying out electron accelerator irradiation treatment on the solution obtained in the step (5) at the temperature of-10 ℃.
Example 6
Bionic collagen liquid dressing effect comparison test
1. Preparation of comparative collagen dressing
Comparative example 1: a preparation method of the bionic collagen liquid dressing sequentially comprises the following steps:
(1) Placing 1g I collagen and 0.5g type III collagen material into 100g deionized water, fully dissolving, centrifuging, and taking supernatant;
(2) Adjusting the pH value of the supernatant obtained in the step (1) to 6.5 by 0.5M triethylamine or 0.1M malic acid;
(3) Adding tween into the solution obtained in the step (2), wherein the mass ratio of tween to collagen is 10;
(4) Performing Co treatment on the solution obtained in the step (3) at the temperature of 0 DEG C 60 And (5) performing irradiation treatment.
Comparative example 2: a preparation method of the bionic collagen liquid dressing sequentially comprises the following steps:
(1) Placing 2g I collagen, 0.1g V collagen and 0.1g VII collagen material into 500g deionized water, fully dissolving, centrifuging, and collecting supernatant;
(2) Adjusting the pH value of the supernatant obtained in the step (1) to 7.8 by 0.2M sodium carbonate or 1M oxalic acid;
(3) Adding 0.05wt.% elastin and 0.00001wt.% chondroitin sulfate to the solution obtained in step (2);
(4) Adding 1g of Na into the solution obtained in the step (3) 2 HPO 4 And 0.05g Mg (HCO) 3 ) 2 ；
(5) Carrying out Cs on the solution obtained in the step (4) at the temperature of 5 DEG C 137 And (5) performing irradiation treatment.
2. Comparison of Effect test
(1) Collagen liquid dressing turbidity test: turbidity of the collagen liquid dressing was observed on a turbidity meter (TZD-BZ-905, suzhou DiagVita Technology Co., ltd.) at a wavelength of 625nm. The test results are shown in Table 1. The turbidity test pattern of the collagen liquid dressing is shown in fig. 1, where fig. 1A is the example 1 sample, fig. 1B is the example 2 sample, fig. 1C is the example 3 sample, fig. 1D is the example 4 sample, fig. 1E is the example 5 sample, fig. 1F is the comparative example 1 sample, and fig. 1G is the comparative example 2 sample.
TABLE 1 turbidity test of collagen liquid dressings
Figure BDA0004016349080000081
As can be seen from Table 1, since examples 1-5 can form homogeneous collagen aqueous solutions, the turbidity of the aqueous solutions is small, and the aqueous solutions of examples 1, 2, 3, 4 and 5 in FIGS. 1A, 1B, 1C, 1D and 1E are relatively transparent; while the collagen molecules in comparative examples 1-2 have aggregated to form collagen fibers, and actually form a collagen fiber suspension, the turbidity is also larger, and the aqueous solutions of comparative examples 1 and 2 in fig. 1F and 1G are more turbid.
(2) Adhesion test of collagen liquid dressing on skin surface: the test sample is evenly smeared on the surface of the skin, and whether the test sample can be evenly adhered on the surface of the skin or not and whether the aggregation and agglomeration phenomenon exists or not is observed. The test pattern is shown in fig. 2, wherein fig. 2A is the example 1 sample, fig. 2B is the example 2 sample, fig. 2C is the example 3 sample, fig. 2D is the example 4 sample, fig. 2E is the example 5 sample, fig. 2F is the comparative example 1 sample, and fig. 2G is the comparative example 2 sample.
As can be seen from fig. 2, the samples of examples 1, 2, 3, 4 and 5 are homogeneous aqueous solutions, which can be uniformly adhered to the skin surface and tightly combined with the skin, while the samples of comparative examples 1 and 2 are heterogeneous suspensions, which are formed by collagen fibers and aggregated, and thus cannot be uniformly applied to the skin surface.
(3) And (3) testing the collagen liquid dressing to repair the skin wound surface: the skin wound surface is a dermal layer bleeding wound surface, the size of the wound surface is about 1cm multiplied by 1cm, the wound is sterilized by wiping the wound surface with iodophor when the wound surface is formed, then the wound surface is smeared on the skin of the example 1, and the wound surface repairing condition is observed 2 minutes, 2 days, 6 days, 9 days and 12 days after smearing, and is photographed and recorded. The repair effect is shown in fig. 3.
As can be seen from fig. 3, the sample in example 1 can be uniformly adhered to the surface of the wound surface at the initial stage of wound surface repair, and the bionic collagen dressing can efficiently stop bleeding, isolate the wound surface from air, and prevent bacteria from contaminating the wound surface; regenerated skin tissues are formed on the wound surface after the wound surface is repaired for 2 days; the wound surface is quickly repaired in 6-12 days, and basically no scar is formed.
Similar to the sample in example 1, the samples in examples 2, 3, 4 and 5 can be quickly adhered to the wound surface at the initial stage of wound surface formation, on one hand, a physical barrier is formed to provide a moist environment for wound surface repair, on the other hand, the physical barrier can simulate the microenvironment of skin tissues to promote wound surface repair, the wound surface starts skin regeneration within 3 days, and after 10 days, the repair is basically completed without obvious scars.
(4) The rat skin wound repair effect is compared: the method comprises the steps of manufacturing a bleeding wound surface on the surface of the skin of a rat, smearing a collagen liquid dressing sample after disinfection and sterilization by iodophor, killing the experimental rat after 10 days, taking out the dressing sample, fixing by using 4wt.% paraformaldehyde fixing solution, and performing HE dyeing.
The method comprises the following specific steps:
(1) Embedding the sample in a paraffin embedding machine, then slicing by a slicing machine, putting the sliced sheet into a water bath kettle at 60 ℃, carefully inserting a glass slide into water close to the sliced sheet, transferring the floating paraffin section onto the glass slide, and if bubbles exist, picking out the paraffin section by a needle.
(2) Dehydrating xylene for 5min twice, sequentially washing with 100% alcohol, 95% alcohol, 85% alcohol, 70% alcohol, and 50% alcohol, washing with tap water, staining with hematoxylin for 5min, and washing with tap water to turn blue.
(3) Placing into 1% hydrochloric acid ethanol solution for fading for 2-10s, wherein the color turns red and lighter, and the color is restored to blue by washing with tap water.
(4) Then 50% alcohol, 70% alcohol and 80% alcohol are added for respectively 5min, and then 0.5% eosin alcohol solution is used for contrast dyeing for 1-3min.
(5) Putting the slices into 95% alcohol, washing off excessive red, then putting into 100% alcohol for 3-5min, absorbing excessive alcohol with absorbent paper, and putting the slices into dimethylbenzene I and II for 3-5min respectively.
(6) Sealing with neutral gum, and fixing.
HE staining effect of rat wound repair is shown in fig. 4, where fig. 4A is the sample of example 1 and fig. 4B is the sample of comparative example 1. As can be seen from fig. 4, the sample of example 1 is more compatible with the skin tissue of the rat and substantially free of inflammatory cells due to the closer microenvironment of the sample of example 1 with the skin tissue of the rat, whereas a large number of inflammatory cells are found in the sample of comparative example 1.
Similar to the sample of example 1, the samples of examples 2, 3, 4, 5 had better skin histocompatibility, while the sample of comparative example 2 had poor skin histocompatibility.
In conclusion, it can be known from the effect test that the sample in the embodiment can simulate the microenvironment of skin tissues from the aspects of collagen types, component simulation, acidic environment and the like on the basis of ensuring that the dressing has the homogeneous flow characteristic, so that the bionic effect is realized, and the repair and regeneration of the skin wound surface are promoted.
The above-described embodiments are merely illustrative of the preferred embodiments of the present invention, and various modifications and improvements of the technical solutions of the present invention can be made by those skilled in the art without departing from the spirit of the present invention, and the technical solutions of the present invention are within the scope of the present invention defined by the claims.

Claims (9)

1. A preparation method of the bionic collagen liquid dressing is characterized by sequentially comprising the following steps:
(1) Dissolving solid collagen material in water, fully dissolving, centrifuging, and taking supernatant; the solid collagen material is selected from one or the combination of more than two types of types I, III, V, VI, VII, VIII, XII, XIII, XIV, XVI and XXVIII; the total concentration of each type of collagen in the supernatant is 0.001wt.% or more but 5wt.% or less;
(2) Adjusting the pH value of the supernatant obtained in the step (1) by an alkaline solution or an acidic solution; the acidic solution comprises at least one of salicylic acid, lactic acid, sulfuric acid, tartaric acid, citric acid, phosphoric acid, oxalic acid, acetic acid, oxalic acid, succinic acid, hydrochloric acid, maleic acid, benzoic acid, nitric acid, malic acid, nicotinic acid, sodium dihydrogen phosphate and formic acid; the alkaline solution comprises at least one of tetramethylethylenediamine, sodium hydroxide, triethylamine, sodium carbonate, potassium hydroxide, calcium hydroxide, magnesium hydroxide, ammonia water, disodium hydrogen phosphate and sodium bicarbonate;
(3) Adding a skin-simulating component into the solution obtained in the step (2); the skin-simulating component comprises at least one of hyaluronic acid, elastin, keratin, chondroitin sulfate, mucopolysaccharide, ceramide, vitamin A, vitamin E, D-panthenol, urea, sodium lactate and lactic acid; the concentration of the skin-mimicking ingredient is 0.00001wt.% or more but 2wt.% or less;
(4) Adding a simulated body fluid component to the solution obtained in step (3), wherein the simulated body fluid component comprises Na + 、K + 、Mg 2+ 、Ca 2+ 、Cl - 、HCO 3 - 、HPO 4 2- 、SO 4 2- 、CO 3 2- 、PO 4 3- 、H 2 PO 4 - One or a combination of two or more of them;
(5) Adding a protective molecule into the solution obtained in the step (4), wherein the protective molecule comprises at least one of sodium dodecyl sulfate, sodium dodecyl benzene sulfonate, tween, triton, ethylene glycol, phenylalanine, proline, lecithin, glutamic acid, lysine, cysteine, mangiferin, glycine, aspartic acid, n-butanol, propylene glycol, sodium ethylene diamine tetracetate, stearic acid, span, glycerol and gelatin;
(6) And (5) carrying out low-temperature irradiation treatment on the solution obtained in the step (5).
2. The method for preparing a biomimetic collagen liquid dressing according to claim 1, wherein the low temperature range of the low temperature irradiation treatment in the step (6) is-20-5 ℃.
3. The method of claim 2, wherein the irradiation treatment comprises at least one of gamma irradiation, electron beam irradiation, and X-ray irradiation.
4. The method of claim 3, wherein the final ion concentration of each ion in the biomimetic collagen liquid dressing is in the range of 0.01-0.5M.
5. The method of claim 4, wherein the mass ratio of the protective molecule to the collagen is from 1.
6. The method of preparing a biomimetic collagen liquid dressing according to any one of claims 1-5, wherein the final pH of the supernatant in step (2) is in a range of 4.0 or greater and 7.8 or less.
7. The method of claim 6, wherein the final pH of the supernatant of step (2) is in the range of 4.1 or more and 6.5 or less.
8. The method of claim 7, wherein the final pH of the supernatant of step (2) is in the range of 4.2 or more and 5.6 or less.
9. A biomimetic collagen liquid dressing prepared using the method of any one of claims 1-8 for preparing a biomimetic collagen liquid dressing.
CN202211672297.7A 2022-12-26 2022-12-26 Bionic collagen liquid dressing and preparation method thereof Pending CN115845126A (en)

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