KR102350526B1 - Method of preparing wound dressing composition for treating skin wounds having improved aggregation - Google Patents

Abstract

The present invention relates to a method for preparing a wound dressing composition and a wound dressing prepared according to the method. More specifically, the present invention relates to a method for producing a wound dressing composition with improved cohesiveness by controlling the content ratio of hyaluronic acid and collagen. The wound dressing composition prepared by the method according to the present invention not only does not cause aggregation of hyaluronic acid and collagen, but also has an excellent wound site relief effect, so it can be used as a pharmaceutical composition for wound treatment or a quasi-drug composition. .

Description

Method of manufacturing a wound dressing composition with improved cohesiveness {METHOD OF PREPARING WOUND DRESSING COMPOSITION FOR TREATING SKIN WOUNDS HAVING IMPROVED AGGREGATION}

The present invention relates to a method for preparing a wound dressing composition for wound treatment and a wound dressing prepared according to the method. More specifically, the present invention relates to a method for producing a wound dressing composition for wound treatment with improved cohesiveness by controlling the content ratio of hyaluronic acid and collagen. The wound dressing composition prepared by the method according to the present invention not only does not cause aggregation of hyaluronic acid and collagen, but also has an excellent wound healing promoting effect, so it can be used as a pharmaceutical composition for wound treatment or a quasi-drug composition. .

A wound refers to a state in which the continuity of the tissue is destroyed by external pressure. The wound healing process is generally divided into three stages: an inflammatory stage, a proliferation stage, and a maturation stage. For wound healing, it is an ideal covering material (dressing) to prevent infection by blocking exposure to the external environment, and the inflammatory reaction should be suppressed. As a biological dressing material for wound treatment, many dressing materials using materials such as collagen and elastin in the dermis, which are components of human skin, are being developed. In the case of collagen, it plays an important role as a major protein constituting tissues such as bones and skin of the human body, and 70% of the skin is composed of collagen, which plays an important physiological role in tissue remodeling and wound healing. . In addition, several studies have shown that collagen is effective in wound healing. As an example of such a biological dressing material, Korean Patent Application Laid-Open No. 10-2010-0009305 discloses a chitosan sponge with increased adhesion of cell adhesion proteins by binding a hydrophobic fatty acid group to chitosan and a wound coating material using the same.

In addition, hyaluronic acid (HA) is a biopolymer material composed of N-acetyl-D-glucosamine and D-glucuronic acid and the repeating units are linearly connected. And it is present in a lot of chicken peat and the like. Because of its excellent biocompatibility and viscoelasticity, hyaluronic acid is being developed for various uses such as post-operative adhesion prevention agent, wrinkle improvement agent, cosmetic aid, joint function improvement agent, drug delivery system, and cell culture scaffold (F. Manna, M. Dentini, P. Desider, O. De Pita, E. Mortilla, B. Maras, Journal of European Academy of Dermatology and Venereology, 13(1999) 183-192).

On the other hand, with respect to wound dressings using collagen and hyaluronic acid, Korean Patent Laid-Open Patent No. 2013-0009651 discloses a chondrocyte therapeutic agent comprising collagen, a hyaluronic acid derivative, and mammalian umbilical cord-derived stem cells. There is a problem in that it is not possible to formulate to achieve the purpose without using a chemical crosslinking material in a mixed manner. In addition, the used formulation has a disadvantage that it can only be used as a dry formulation, so there is a limitation in that it cannot provide various methods in the field of use and the method of use.

As a result, the present inventors have made diligent efforts to develop products that can significantly improve the quality and reliability of wound dressings, and as a result, not only have an optimal wound site relief effect by adjusting the content ratio of hyaluronic acid and collagen, but also hyaluronic acid And it has been found that the aggregation phenomenon of collagen does not occur, so that a good image can be planted to consumers, and thus the present invention has been completed.

An object of the present invention is to provide a method for preparing a wound dressing composition and a wound dressing prepared by the method.

As used herein, the term “hyaluronic acid” is used to include both hyaluronic acid itself and hyaluronic acid salts. Therefore, the term "aqueous solution of hyaluronic acid" used hereinafter is a concept including both an aqueous solution of hyaluronic acid, an aqueous solution of a hyaluronic acid salt, and a mixed aqueous solution of hyaluronic acid and a hyaluronic acid salt. The hyaluronic acid salt includes inorganic salts such as sodium hyaluronate, potassium hyaluronate, calcium hyaluronate, magnesium hyaluronate, zinc hyaluronate, cobalt hyaluronate, and organic salts such as tetrabutylammonium hyaluronate. In some cases, two or more thereof may be used in combination.

the present invention

(a) Disclosed is a method for preparing a wound dressing composition comprising mixing an aqueous solution of hyaluronic acid or a salt thereof at a concentration of 5 mg/ml to 10 mg/ml and an aqueous solution of collagen at a concentration of 1 mg/ml to 60 mg/ml do.

In the method for preparing the wound dressing composition according to the present invention, the hyaluronic acid or its salt is 5 mg/ml to 10 mg/ml, preferably 5 mg/ml to 8 mg/ml, more preferably 5.85 to 7.15 It is characterized in that it is mixed at a concentration of mg/ml, most preferably 6.5 mg/ml.

In the method for preparing the wound dressing composition according to the present invention, the collagen is 1 mg/ml to 60 mg/ml, preferably 5 mg/ml to 15 mg/ml, more preferably 8.1 to 9.9 mg/ml, Most preferably, it is characterized in that it is mixed with hyaluronic acid or a salt thereof at a concentration of 9 mg/ml.

In the method for preparing the wound dressing composition according to the present invention, the molecular weight of the hyaluronic acid is 100,000 to 5,000,000 Da.

In the method for manufacturing the wound dressing composition according to the present invention, the aqueous solution is deionized water (DW), water for injection, or phosphate buffered solution (PBS).

In the method for preparing the wound dressing composition according to the present invention, the mixing is characterized in that it is performed at a stirring speed of 80 to 300 rpm under vacuum for 24 to 120 hours.

In the method for preparing the wound dressing composition according to the present invention, the preparation method is an aqueous solution of hyaluronic acid or a salt thereof in an amount of 5 mg/ml to 15 mg/ml, preferably 10 mg/ml before step (a). Stirring at a speed of 300 to 350 rpm for 6 hours to 24 hours after addition to; autoclaving the stirred aqueous solution of hyaluronic acid or a salt thereof at 110° C. to 130° C. for 10 to 20 minutes; And after cooling the stirred aqueous solution of hyaluronic acid or its salt at room temperature, it characterized in that it further comprises the step of storing at 3 ℃ to 10 ℃ for 3 to 24 hours.

In the method for manufacturing the wound dressing composition according to the present invention, the manufacturing method is characterized in that it further comprises the step of adjusting the pH of the mixed solution to 5 to 9 after step (a).

In the method for manufacturing a wound dressing composition according to the present invention, the wound dressing composition is a pharmaceutical composition, a quasi-drug composition or a cosmetic composition.

When the wound dressing composition according to the present invention is a pharmaceutical composition, it can be applied by parenteral administration or topical application by application, and a suitable dosage of the pharmaceutical composition of the present invention is determined by the patient's condition and weight, disease It may vary depending on the degree, drug form, route and duration of administration, and may be appropriately selected by those skilled in the art.

Also, the present invention

Disclosed is a wound dressing in which an aqueous solution of hyaluronic acid or a salt thereof having a concentration of 5 mg/ml to 10 mg/ml and an aqueous solution of collagen having a concentration of 1 mg/ml to 60 mg/ml are mixed.

In the wound dressing according to the present invention, the sodium hyaluronate content of the wound dressing is 5.85 to 7.15 mg/ml and the collagen content is 8.1 to 9.9 mg/ml.

In the wound dressing according to the present invention, the pH of the wound dressing is 5 to 9, preferably 7, characterized in that.

In the wound dressing according to the present invention, the viscosity of the wound dressing is characterized in that 1000 to 10000 mPa·s.

In the wound dressing according to the present invention, the injection solution for the syringe of the wound dressing is 0.5 cc to 5 cc.

In the wound dressing according to the present invention, the wound dressing is characterized in that it is translucent or opaque.

1 shows a photograph of a wound dressing comprising HA and Collagen prepared according to Example 1 of the present invention.
Figure 2 is a graph showing the wound shrinkage rate of the wound dressing material containing HA (6.5 mg / ml) and Collagen (9 mg / ml) prepared according to Example 1 of the present invention.
3 is a graph showing the results of histopathological analysis of the wound dressing containing HA (6.5 mg/ml) and Collagen (9 mg/ml) prepared according to Example 1 of the present invention.
Figure 4 shows the tissue adhesion evaluation results of the wound dressing containing HA (6.5 mg / ml) and Collagen (9 mg / ml) prepared according to Example 1 of the present invention.
5 shows the results of visual evaluation and histopathological observation of a wound dressing containing HA (6.5 mg/ml) and Collagen (9 mg/ml) prepared according to Example 1 of the present invention.

Hereinafter, various examples are presented to help the understanding of the invention. The following examples are provided for easier understanding of the invention, and the protection scope of the invention is not limited to the following examples.

<Example>

Example 1. Preparation of wound dressings containing 6.5 mg/ml of HA and 9 mg/ml of Collagen

First, sodium hyaluronate was added to water for injection at a concentration of 10 mg/ml, and stirred at a speed of 300 to 350 rpm for 6 to 24 hours. The stirring solution was autoclaved at 121° C. for 15 minutes, cooled at room temperature, and stored at 4° C. for 3 to 24 hours. Sodium hyaluronate and porcine type I collagen were mixed with the stirring solution and 3% porcine type I collagen, respectively (A) HA (9 mg/ml), Collagen (3 mg/ml), (B) HA (8 mg/ml) ), Collagen (6 mg/ml), and (C) HA (6.5 mg/ml), Collagen (9 mg/ml) at the level of the present invention by stirring at 80-100 rpm for 24 to 120 hours under vacuum. The wound dressing was completed according to the The wound dressing was filled in a syringe with a volume of 1 cc to 3 cc.

Example 2. Selection of the optimal mixing ratio of HA and Collagen

In order to examine whether aggregation occurred according to the content ratio of HA and collagen in the wound dressing prepared in Example 1, photographs were taken and the results are shown in FIG. 1 .

As a result, (A) a wound dressing containing HA (9 mg/ml) and Collagen (3 mg/ml) and (B) a wound dressing containing HA (8 mg/ml) and Collagen (6 mg/ml) In the case of aggregation, (C) it was confirmed that aggregation did not occur when HA and collagen were mixed in amounts of 6.5 mg/ml and 9 mg/ml, respectively.

Example 3. Confirmation of the performance of the wound dressing prepared with the optimal mixing ratio of HA and Collagen

According to Example 2, the pH of the wound dressing prepared in an amount of 6.5 mg/ml and 9 mg/ml of HA and Collagen, respectively, was confirmed to be 7.0±1, and the result of viscosity measurement using a rotational viscometer (Brookfield, DW2T) , was confirmed to be 5261±121.

<Experimental example>

Experimental Example 1. Confirmation of the wound healing effect of the wound dressing according to the present invention

1-1. experimental method

The wound dressing material containing HA (6.5 mg/ml) and Collagen (9 mg/ml) prepared in Example 1 was applied to the wound site a total of 10 times to the Sprague-Dawley rat used as a wound animal model, and the wound was wound for 14 days. Healing was observed while checking for wound healing. At this time, as the placebo group, only commercially available transparent sterile film dressings were used, and commercially available gel-type wound healing agents and transparent sterile film dressings were used as other companies' products.

substance to be administered gender number of animals route of administration number of applications Control M 8 - - placebo M 8 - - HA (6.5 mg/ml)-
Collagen (9 mg/ml) M 8 apply 10 third-party products M 8 apply 10

1-2. result

(1) Confirmation of wound contraction rate through image analysis

As a result of observing the change in wound size due to application of the wound dressing according to the present invention, the wound dressing according to the present invention was compared with the placebo group for 2 days (p <0.01) 4 days (p <0.05) and 9 days (p < 0.05) showed a healing effect with statistical significance, respectively. In addition, in the case of the wound dressing material according to the present invention, it was confirmed that the same or better wound area shrinkage than other products was observed (FIG. 2).

(2) histopathological analysis

As a result of measuring the thickness of the entire skin layer, when the wound is completely recovered, the entire skin layer is restored. In the case of the placebo group, compared to the control group, only about 67% of the thickness of the entire skin is recovered, whereas in the case of the wound dressing according to the present invention, 81% level It was confirmed that full-thickness recovery was observed that is equivalent to or superior to that of other products (FIG. 3).

Experimental Example 2. Confirmation of the anti-adhesion effect of the wound dressing according to the present invention

1-1. experimental method

The anti-adhesion efficacy of the wound dressing containing HA (6.5 mg/ml) and Collagen (9 mg/ml) prepared in Example 1 was evaluated using SD rats. Adhesion was induced between the epithelial tissue of the rat abdominal wall and the cecum tissue, and the test substance was applied to the site of adhesion induction. Control, commercially available competitor products 1 and 2, and wound dressings according to the present invention were applied to 15 experimental animals each. No weight loss or specific symptoms were observed in all experimental animals for 2 weeks of application. After the application period, all animals used in the experiment were euthanized and intraperitoneal adhesions were evaluated according to the Lauder Score shown in Table 2 below.

Adhesion scoring scheme by Lauder et al. (2011) Score Description 0 No adhesion One thin filmy adhesion 2 More than one thin adhesion 3 Thick adhesion with local point 4 Thick adhesion with planar attachment 5 Very thick adhesions or more than one planar

1-2. result

(1) Tissue adhesion evaluation

In the case of the wound treatment agent according to Example 1 of the present invention, the adhesion rate was 14%, and it was confirmed that the anti-adhesion effect was significantly improved compared to 93% of the negative control group and 60% and 47% of the commercial products, respectively (FIG. 4) .

[Table 3]

(2) Visual evaluation and histopathological observation results

In the case of the negative control group, most of the adhesions occurred between the peritoneum and the cecum due to the formation of granulation tissue. 5).

As described above in detail a specific part of the present invention, for those of ordinary skill in the art, this specific description is only a preferred embodiment, and it is clear that the scope of the present invention is not limited thereto. Accordingly, the substantial scope of the present invention will be defined by the appended claims and their equivalents.

Claims (9)

(a) mixing an aqueous solution of hyaluronic acid or a salt thereof at a concentration of 5.85 to 7.15 mg/ml and an aqueous solution of collagen at a concentration of 8.1 to 9.9 mg/ml,
The mixing is characterized in that it is carried out at a stirring speed of 80 to 300 rpm under vacuum for 24 to 120 hours,
Method for producing a wound dressing composition.
According to claim 1,
The molecular weight of the hyaluronic acid is characterized in that 100,000 to 5,000,000 Da, the method for producing a wound dressing composition.
According to claim 1,
The aqueous solution is deionized water (deionized water: DW), water for injection or phosphate buffer solution (PBS), characterized in that the, wound dressing composition manufacturing method.
delete According to claim 1,
The method for preparing a wound dressing composition, characterized in that it further comprises the step of adjusting the pH of the mixed solution to 5 to 9 after step (a).
According to claim 1,
The wound dressing composition is a pharmaceutical composition, a quasi-drug composition or a cosmetic composition.
As a wound dressing prepared by the method for preparing a wound dressing composition according to any one of claims 1 to 3, and any one of claims 5 to 6,
The sodium hyaluronate content of the wound dressing is 5.85 to 7.15 mg/ml and the collagen content is 8.1 to 9.9 mg/ml,
The pH of the wound dressing is 5 to 9,
The viscosity of the wound dressing is 1000 to 10000 mPa · s, characterized in that the wound dressing.
8. The method of claim 7,
The injection solution for the syringe of the wound dressing is characterized in that 0.5 cc to 5 cc, the wound dressing.
8. The method of claim 7,
The wound dressing is, characterized in that the translucent or opaque, wound dressing.

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