Wound covering material composition containing fucosan and olive leaf extract

Abstract

The present invention relates to a wound covering material composition capable of promoting the healing effect of a skin wound by means of the growth of stem cells on the epidermal basal layer of the skin. The invention provides a wound covering material composition, which comprises the following components in percentage by weight: 60 to 70 percent of glycerol; 28 to 39 weight percent of honey; 0.2 wt% to 3.0 wt% of fucosan; 0.2 to 3.0 wt% of an olive leaf extract; and 0.1 to 0.3 wt% of xanthan gum.

Description

Technical Field

[0001] The present invention relates to a wound covering material composition which can achieve skin wound healing effect by promoting the growth of stem cells in the epidermal basal layer of the skin. Background Art

[0002] Korean authorized patent 10-2088767, as an invention related to a "composition for increasing the biological activity of stem cells using mixture 4F", provides a composition for inhibiting the aging of stem cells, promoting proliferation or inducing differentiation, which includes fucoidan, tauroursodeoxycholic acid, oleuropein and vascular endothelial growth factor as active ingredients.

[0003] The "stem cells treated with the mixture 4F" proposed in the above-mentioned authorized patent not only acquire undifferentiated characteristics (stemness), but also have the effect of improving cell proliferation and mobility. Therefore, after the stem cells are transplanted into the body, cell survival and implantation survival rates can be improved, and the ability of blood vessels and tissue regeneration can be further improved, thereby being widely used in the field of stem cell differentiation, prevention or treatment of ischemic diseases.

[0004] On the other hand, human skin tissue is composed of the epidermis, dermis, and subcutaneous tissue. The epidermis can be divided into the stratum corneum (stratum corneum) composed of dead cells and the living epidermis. The living epidermis is further divided into four layers (stratum lucidum), stratum granulosum, stratum spinosum, and stratum basale) (stratum basale).

[0005] If a skin wound occurs, stem cells from the basal layer are needed to heal the wound through stem cell division and growth. Therefore, applying patent 10-2088767 and related research findings, we have developed a wound dressing material composition that can promote the growth of basal layer stem cells to achieve skin wound healing. Summary of the Invention

[0006] Technical issues

[0007] The present invention aims to provide a wound dressing material composition that can utilize fucoidan and olive leaf extract to promote the growth of stem cells in the epidermal basal layer of the skin to achieve a skin wound healing effect.

[0008] Technical Solution

[0009] The present invention provides a wound dressing material composition, comprising: 60-70 wt% of glycerol; 28-39 wt% of honey; 0.2-3.0 wt% of fucoidan; 0.2-3.0 wt% of olive leaf extract; and 0.1-0.3 wt% of xanthan gum.

[0010] The fucoidan can be obtained by mixing dried spore leaf powder or extract with pure water, centrifuging, filtering, and recovering. The olive leaf extract can be obtained by sterilizing by heating a liquid obtained by filtering the powder of crushed olive leaves or a liquid extracted from olive leaves at 120-125° C. for 15-18 minutes.

[0011] The viscosity of the wound dressing composition can be adjusted within the range of 500 to 10,000 mPa·s according to the content of the xanthan gum.

[0012] Technical Effects

[0013] The present invention achieves the following effects by optimizing the content ratio of the components of the wound dressing material composition:

[0014] 1) The effect of exudate discharge through the osmotic pressure of glycerol; 2) The moisturizing effect of honey and the antibacterial and antimicrobial effects through the action of preservative enzymes; 3) The basal layer stem cell growth promotion effect of fucoidan; and 4) The organic effects of the antioxidant, anti-inflammatory, antimicrobial and inflammation-relieving effects of olive leaf extract can be used to achieve a skin wound healing effect; and xanthan gum can be used to achieve the appropriate viscosity of the film at the wound site. BRIEF DESCRIPTION OF THE DRAWINGS

[0015] Figure 1 This is a graph showing the results of measuring the cytotoxicity of fucoidan and olive leaf extract (oleuropin: oleuropein) at various concentrations on human dermal fibroblasts and epidermal keratinocytes.

[0016] Figure 2 The present invention shows scanning electron microscope (SEM) photographs and graphs showing the results of measuring the wound recovery effects of fucoidan and olive leaf extract (oleuropin) at various concentrations on human dermal fibroblasts.

[0017] Figure 3 These are SEM photographs and graphs showing the results of measuring the wound recovery effects of fucoidan and olive leaf extract (oleuropin) at various concentrations on human epidermal keratinocytes.

[0018] Figure 4 This is a graph showing the wound recovery rates of the control group and the test group by date.

[0019] Figure 5 It is a graph showing the recovery rate of the experimental group compared with the control group by day.

[0020] Figure 6 The photos are taken of the wound recovery status of the control group and the experimental group according to the date.

[0021] Best Practice

[0022] A wound dressing material composition, characterized by comprising: 60-70 wt% glycerol; 28-39 wt% honey; 0.2-3.0 wt% fucoidan; 0.2-3.0 wt% olive leaf extract; and 0.1-0.3 wt% xanthan gum.

[0023] The fucoidan is obtained by mixing dry spore leaf powder or extract of Undaria pinnatifida with purified water, centrifuging, filtering and recovering the mixture.

[0024] The olive leaf extract is obtained by sterilizing a liquid obtained by filtering a powder of pulverized olive leaves or a liquid extracted from olive leaves by heating the liquid at 120 to 125° C. for 15 to 18 minutes. DETAILED DESCRIPTION

[0025] The present invention relates to a wound dressing material composition that utilizes a substance known to contribute to skin regeneration when the skin barrier is damaged due to trauma, burns, ulcers, contusions, post-surgical wounds, childbirth, chronic wounds, dermatitis, etc., and utilizes a relatively easily available, safety-assured substance to prevent wound infection and exhibit an antioxidant effect.

[0026] Stem cells are cells that have the ability to self-replicate and differentiate into two or more cell types. The proliferative potential of basal stem cells maintains the homeostasis of stratified epithelial tissue. Therefore, if a skin wound occurs, stem cells from the basal layer at the epidermal-dermal junction are needed to heal the wound through stem cell division and growth.

[0027] The present invention applies fucoidan and olive leaf extract, which are culture medium compositions of stem cell therapeutic agents studied in Patent No. 10-2088767, to wound dressing compositions.

[0028] However, when a wound occurs on the skin, the supply and proliferation of stem cells from the basal layer may be limited by exudates, microorganisms, and other contaminants at the wound site. Therefore, it is particularly important to use wound dressing materials to clean the wound site, relieve the burning sensation or pain caused by the wound, and ensure a predetermined viscosity of the wound dressing material so that it is not easily lost when applied to the wound site.

[0029] The present invention, which is made in consideration of the above matters, provides a wound dressing material composition comprising: 60-70 wt% of glycerin; 28-39 wt% of honey; 0.2-3.0 wt% of fucoidan; 0.2-3.0 wt% of olive leaf extract; and 0.1-0.3 wt% of xanthan gum.

[0030] The wound dressing material composition provided by the present invention uses glycerin and honey as base materials, contains fucoidan and olive leaf extract as active ingredients for medicinal effects, and contains xanthan gum as a thickener.

[0031] Glycerin (C3H8O3) is a non-irritating substance that is widely used as a main component of enemas, lubricants, creams, eye drops, injections, etc., and is also widely used as a component of wound dressing materials.

[0032] Glycerol, due to its high osmotic pressure, causes cells in the body to contract, allowing exudate to be expelled from the wound. This process also separates and expels microorganisms and other contaminants from the wound. Furthermore, glycerol prevents exudate, microorganisms, and other contaminants from reentering the body, thereby cleansing the wound and promoting wound healing.

[0033] Furthermore, glycerin has a strong binding force, so when applied to a wound, it does not penetrate into the body tissues but remains outside the wound. This glycerin is mixed with xanthan gum (described later) and forms a thin film on the wound in a state of increased viscosity.

[0034] The wound dressing material composition of the present invention contains 60 to 70 wt% of the glycerin.

[0035] Honey is the liquid expelled by bees after they feed on sucrose secreted by the nectar glands of flowers. It is a viscous liquid broken down by bee enzymes into fructose and glucose. Honey resists spoilage due to the osmotic effect of its high sugar content and the presence of several antiseptic enzymes in the nectar. Therefore, when honey is applied to skin wounds, the antiseptic enzymes and osmotic effect cause moisture from bacteria to migrate to the honey, thereby performing antibacterial and antimicrobial functions, killing bacteria and pathogens that have been deprived of moisture.

[0036] In addition, honey can form a skin protective film to perform the following functions: skin soothing function to relieve skin burning or pain caused by wound irritation, and moisturizing function to relieve skin discomfort such as burning, itching, dryness, etc. by retaining skin moisture and making the skin tender.

[0037] The wound dressing material composition of the present invention contains 28 to 39 wt% of the honey.

[0038] Fucoidan is a substance that gives brown algae their flexibility and protects them from violent currents. Fucoidan, extracted from brown algae, is a polysaccharide composed of a monosaccharide called fucose linked to sulfate groups. It is known to have anticoagulant, antitumor, gastric ulcer treatment-promoting, antibacterial, anti-inflammatory, blood pressure suppression, induction of hepatocyte growth factor (HGF) production, blood sugar suppression, immune cell regulation, anti-allergic, and antiviral effects.

[0039] Fucoidan, due to its high hydrophilic sulfate groups, enhances skin regeneration and can assist and activate the proliferative potential of basal layer stem cells, thereby contributing to maintaining homeostasis in stratified epithelial tissues. Specifically, it promotes wound healing by aiding the division and growth of basal layer stem cells at the epidermal-dermal junction.

[0040] The fucoidan may have the following structure [Chemical Structure 1].

[0041]

Chemical Structure 1

[0042]

[0043] The present invention utilizes fucoidan, which is recovered by mixing dried Undaria pinnatifida spore leaf powder or extract with purified water, centrifuging the mixture three times, and then filtering and recovering it. The fucoidan has a molecular weight ranging from 1,000 to 2,000,000 Da. The wound dressing composition of the present invention contains 0.2 to 3.0 wt% of the fucoidan.

[0044] Olive leaf extract, a component of olive oil called oleuropein, is extracted from olive leaves. It has excellent antioxidant, anti-inflammatory, and antibacterial properties, and is also effective in relieving inflammation. Therefore, it is widely used as a skin protectant and natural antibiotic. The polyphenols found in olive fruits and leaves are generally referred to as oleuropein, and the content of oleuropein in olive leaf extract is higher than that in olive oil. The oleuropein in olive leaf extract can be used with the following structure [Chemical Structure 2].

[0045]

Chemical Structure 2

[0046]

[0047] In the present invention, the wound healing effect is maximized by sterilizing the liquid obtained by filtering the powdered olive leaf or the liquid extracted from the olive leaf by heating at 120 to 125° C. for 15 to 18 minutes.

[0048] The wound dressing material composition of the present invention contains 0.2 to 3.0 wt% of the olive leaf extract.

[0049] Xanthan gum is made by pure-culture fermentation of carbohydrates using Xanthomonas campestris, and the resulting high-molecular-weight polysaccharide gum is refined, dried, and crushed in isopropyl alcohol. It is a mixture of glucose, mannose, and sodium, potassium, and calcium salts of glucuronic acid. As an exopolysaccharide (EPS), its main chain is β-1,4-glucose (cellulose), and every two glucose residues are bound to a trisaccharide (mannose-glucuronic acid-mannose) as a side chain. The mannose bound to glucose is acetylated. These xanthan gums are used as food additives in foods as stabilizers, thickeners, adhesives, emulsifiers, coagulants, and foaming agents.

[0050] Although the viscosity of the wound dressing material composition of the present invention varies depending on the concentration and content of glycerol and the content of honey, the viscosity of the wound dressing material composition can be adjusted within the range of 500 to 10,000 mPa·s by adjusting the addition amount of the xanthan gum within the range of 0.1 to 0.3 wt%.

[0051] The wound dressing material composition is stirred at 80-300 rpm for 24-120 hours to achieve homogenization, allowing it to be used not only as a pharmaceutical agent but also as a quasi-drug or cosmetic composition. Depending on the intended use, the pH of the wound dressing material composition can be adjusted within the range of 5-9 and can be prepared to be translucent or opaque.

[0052] The wound dressing material composition can be commercialized by injecting 0.1 to 500 cc of the wound dressing material composition into a container, and can be sterilized by irradiating the wound dressing material composition with the composition injected into the container with gamma rays.

[0053] The following describes the results of experiments using human dermal fibroblasts and keratinocytes.

[0054] 1. Measurement of Cytotoxicity in Human Dermal Fibroblasts and Epidermal Keratinocytes

[0055] In order to confirm the effect of the mixture of fucoidan and olive leaf extract (oleuropin) on the proliferation of human dermal fibroblasts (adult HDFa cells) and human epidermal keratinocytes (HaCaT cells), MTT assay was performed. Human dermal fibroblasts and epidermal keratinocytes were cultured at 1×10 3 Cells / well~3×10 3 After cells were plated per well in 96-well plates and cultured for 24 hours, a 1000-fold concentrated stock solution of a 1:1 weight ratio of fucoidan and olive leaf extract (oleuropein) was diluted individually with sterile distilled water. 50-fold, 25-fold, 10-fold, 1-fold, and 0.1-fold concentrations of the mixture (working solution) were then added to the wells for treatment and cultured for 24 hours. Following incubation, MTT solution was added for reaction, and absorbance was measured at 540 nm.

[0056] As a result, for human dermal fibroblasts, when the survival rate (proliferation rate) in the negative control group to which the mixed substance was not added was taken as 100%, when the mixed substance was treated at each concentration, the 24-hour survival rate (proliferation rate) was 98.38% at a 0.1-fold concentration, 98.10% at a 1-fold concentration, 88.40% at a 10-fold concentration, and 84.17% at a 25-fold concentration, confirming that there was no significant difference in cytotoxicity. For human epidermal keratinocytes, the survival rate at a 0.1-fold concentration of the mixed substance was 98.96%, the survival rate at a 1-fold concentration was 97.65%, the survival rate at a 10-fold concentration was 99.42%, the survival rate at a 25-fold concentration was 93.79%, and the survival rate at a 50-fold concentration was 86.36%, confirming that there was no significant difference in cytotoxicity (refer to Figure 1 ).

[0057] 2. Measurement of Wound Recovery Efficacy in Human Dermal Fibroblasts and Epidermal Keratinocytes

[0058] In order to confirm the effect of a mixture of fucoidan and olive leaf extract (oleuropin) on wound damage (Wound) of human dermal fibroblasts and epidermal keratinocytes, a wound healing assay was performed. Fucoidan and olive leaf extract were each added at 0.5×10 5 ~3×10 5 Cells were dispensed into 12-well plates at 1:1 ratio (weight) and cultured for 24 hours. A horizontal scratch was made using a yellow-tip pipette and the cells were washed twice with culture medium. A 1000-fold concentrated stock solution of a 1:1 weight ratio of fucoidan and olive leaf extract (oleuropein) was diluted individually with sterile distilled water. 50-, 25-, 10-, 1-, and 0.1-fold concentrations of the mixture (working solution) were then added to the wells and cultured for 24 hours. The surface area of the horizontal scratch was photographed microscopically at 0 and 24 hours after culture and measured using Image J.

[0059] The results showed that when the mixed substance of fucoidan and olive leaf extract (Oleuropin) was used to treat the scratched area of human dermal fibroblasts, the recovery rate was 76.6% (damage recovery rate 23.4%) in the negative control group, and it was confirmed that after 24 hours, it was 18.8% (damage recovery rate 81.2%) at 0.1 times the concentration, 25.0% (damage recovery rate 75%) at 1 times the concentration, and 34.3% (damage recovery rate 65.7%) at 10 times the concentration. This shows that the wound recovery effect is maximized when the mixed substance is at a low concentration (refer to Figure 2 ).

[0060] The scratch area of human epidermal keratinocytes showed the following results: compared with 39.8% in the negative control group (damage recovery rate of 60.2%), after 24 hours, it showed 5.6% at 0.1 times the concentration (damage recovery rate of 94.4%), 8.6% at 1 times the concentration (damage recovery rate of 91.4%), and 11.7% at 10 times the concentration (damage recovery rate of 88.3%). In this case, it was also shown that the wound recovery effect was greater when the mixed substance was at a low concentration (reference Figure 3 ).

[0061] Therefore, in the present invention, a small amount of 0.2 to 3.0 wt% of fucoidan and 0.2 to 3.0 wt% of olive leaf extract are added to the entire wound dressing material.

[0062] The following is a toxicity test conducted in a recognized testing institution for an arbitrarily selected example composition (the midpoint of each component, glycerin 65wt%, honey 33wt%, fucoidan 0.9wt%, olive leaf extract 0.9wt% and xanthan gum 0.2wt%) that satisfies the following conditions: glycerin 60-70wt%; honey 28-39wt%; fucoidan 0.2-3.0wt%; olive leaf extract 0.2-3.0wt%; and xanthan gum 0.1-0.3wt%; as shown in Table 1.

[0063]

Table 1

[0064]

[0065]

[0066] The contents of each test and the composition of the present invention showed no harmfulness. The results of each test are as follows.

[0067] 1. Test 1: Skin sensitization test using guinea pigs

[0068] This test is intended to evaluate whether a test substance (composition of examples of the present invention) exhibits skin sensitization after administration to test animals (guinea pigs, female).

[0069] A negative control substance and a test substance were used in a group of 5 or 10 test animals. One intradermal induction, two topical inductions, and three inductions were performed. During the test period, mortality, symptoms, and body weight were observed and measured. Furthermore, skin reactions at all application sites were evaluated 24 ± 2 hours (h) and 48 ± 2 hours after the third patch removal.

[0070] The test results showed that none of the animals administered with the negative control solution or the test solution died, nor were there any symptoms, abnormal behaviors, or weight changes attributable to the administered substances observed. No erythema or swelling was observed at the induced site in any of the animals, and the Magnusson-Kligman Scale grade was zero. The test substance was therefore assessed as not causing skin sensitization.

[0071]

Table 2

[0072]

[0073]

Table 3

[0074]

[0075]

Table 4

[0076]

[0077]

[0078] 2. Experiment 2: Single-dose acute systemic toxicity test in mice

[0079] The test substance dissolution solution and negative control substance in this test were the same as those in the above-mentioned "Test 1".

[0080] This study evaluated the potential for toxic reactions following a single intraperitoneal administration of a test substance to male mice (specific pathogen-free (SPF) mice). Five mice per test group were administered a single dose of a negative control substance and a solution of the test substance. During the study, mortality, symptoms, and body weight were observed and measured, and a gross autopsy was performed on the necropsy day.

[0081] The test results showed that none of the test animals administered with the negative control substance or the test substance died during the experimental period. No abnormal symptoms, abnormal behaviors, or weight changes attributable to the administered substance were observed in any group. No abnormal findings attributable to the administered substance were observed during the macroscopic autopsy.

[0082] 3. Experiment 3: Cytotoxicity assay using L929 cells

[0083] This test is designed to confirm whether the test substance has cytotoxicity using L929 cells.

[0084] L929 cells were seeded into 6-well plates and cultured in a culture medium at 37±1°C and 5±1% CO₂ for 24±2 hours. Agar-containing culture medium (prepared by adding approximately 50 ml of fetal bovine serum (FBS) and 10 ml of penicillin-streptomycin to 440 ml of Minimum Essential Medium (MEM)) was dispensed and allowed to solidify before staining the cells with neutral red.

[0085] The RM-C high-density polyethylene film selected according to ISO 10993 was cut into 1.0 cm 2 , and sterilized at 121±2°C for 20 minutes, thereby preparing three negative control materials, and cutting RM-B (0.25% zinc dibutyldithiocarbamate polyurethane film (ZDBC polyurethane film)) selected according to ISO 10993 into 1.0 cm 2 , and sterilized at 121±2°C for 20 minutes, thereby preparing three positive control substances. The composition of the embodiment of the present invention was fully soaked in a 1.0 cm 2 Three test substances were prepared using a 0.45 μm filter.

[0086] The negative control substance, positive control substance and test substance prepared as above are applied to the L929 cells of dyeing respectively and are observed. As a result, in the negative control group, no cell morphology changes occur at the bottom or periphery of sample, and no dissolving is confirmed. In addition, owing to not finding the position of decolorization, it is zero grade that the cytotoxic reaction grade is presented. In the positive control group, the morphology of the cell is rounded, and the dissolving and decolorization position of the cell are expanded to 0.4cm to the periphery of sample, so it is three grades that the cytotoxic reaction grade is presented. In the test group, because the abnormality of the cell and the decolorization position are limited to the immediately adjacent below of sample, it is two grades that the cytotoxic reaction grade is presented.

[0087] When the results of the negative control group and the positive control group are confirmed, the test process is reasonable, and the comprehensive test results show that the cytotoxic reaction level of the test substance is level 2, so it is judged that there is no cytotoxicity.

[0088]

Table 5

[0089]

[0090] 4. Test 4: Intradermal reaction test using rabbits (direct contact)

[0091] The test substance dissolution solution and negative control substance in this test were the same as those in the above-mentioned "Test 1".

[0092] This study was designed to evaluate the potential for irritation induced by the test substance when a single dose of the test substance solution was administered intradermally (intradermally) to rabbits (species: New Zealand White (NZW), SPF, female).

[0093] Three rabbits were administered solutions of the test substance dissolved in physiological saline (SC) and cottonseed oil (CSO). 0.2 ml of the test substance solution was intradermally administered at five sites (10 total) on the left side of the rabbit's back, centered on the dorsal spine, using each solvent (SC, CSO). 0.2 ml of the negative control substance was administered at five sites (10 total) on the right side of the rabbit's back, using each solvent (SC, CSO).

[0094] During the experiment, the mortality, symptoms and body weight of the experimental animals were observed and measured.

[0095] The results of the trial showed that no subjects died during the trial period, and none of the subjects developed specific symptoms that could be attributed to the administered substance. No weight changes were observed that could be attributed to the administered substance.

[0096] Intradermal reactions at the administration site of the test substance dissolution solution and the negative control substance were observed, and the intradermal reaction score at each administration site was calculated based on the observation criteria 24 ± 2 hours, 48 ± 2 hours, and 72 ± 2 hours after administration. The difference in the average score at the administration site for each dissolution solution was 1.0 or less, and there was no difference between the test substance and the negative control substance when the dissolution solution was the same.

[0097] Since no specific irritation was observed at the test substance administration site, it was considered that the test substance did not cause a local reaction in the rabbit skin.

[0098]

Table 6

[0099]

[0100]

Table 7

[0101]

[0102] 5. Experiment 5: Fever test using rabbits

[0103] This study aimed to verify the biological safety of the test substance by evaluating the potential for febrile reactions following a single administration of a test substance solution into the ear vein of rabbits (breed: NZW, SPF, female). SC was used as the solvent for the solution.

[0104] Control body temperature was measured before administration of the test substance solution. A single dose of 10 ml / kg of the test substance solution was administered into the ear vein of three rabbits. The experimental groups were assigned so that the control body temperature of the individual rabbits differed by less than 1.0°C. Rabbits with control body temperatures exceeding 39.8°C were excluded from the study.

[0105] One hour after the administration of the test substance, the body temperature was measured five times at 30-minute intervals, and the difference between the body temperature and the control was measured, and the mortality rate and symptoms during the test were observed.

[0106] The test results showed that none of the test animals administered the test substance solution showed a temperature increase of more than 0.5°C compared to the control body temperature, thus confirming that the test substance was non-pyrogenic (two of the three rabbits showed no temperature increase at all, while one rabbit had a maximum temperature increase of 0.2°C). Furthermore, no animals died during the test period, and no animals were observed to exhibit abnormal symptoms.

[0107] 6. Test 6: Skin irritation test using rabbits (direct contact)

[0108] This test is designed to evaluate the potential of test substances to induce irritation when in direct contact with the skin of rabbits (breed: NZW, SPF, female).

[0109] The test substance and negative control substance were applied to the backs of hairless rabbits. For the test substance, according to ISO 10993-23, a 5.0 cm x 5.0 cm sterile gauze was divided into four equal parts and made into four 2.5 cm x 2.5 cm sterile gauze pads. These were then folded into twelve layers, and 0.5 ml of the test substance was added dropwise using a syringe and applied directly to the skin. For the negative control substance, also according to ISO 10993-23, a 5.0 cm x 5.0 cm sterile gauze was divided into four equal parts and made into four 2.5 cm x 2.5 cm sterile gauze pads. These were then folded into twelve layers, and 0.5 ml of sterile saline (SC) was added dropwise using a syringe and applied.

[0110] Wrap the application site with appropriate pressure that does not cause irritation to the abdomen and secure with a bandage to prevent loss of the applied test substance and negative control substance. Remove the test substance and negative control substance from the application site 4 to 24 hours after application.

[0111] During the test, mortality, symptoms, and body weight were observed and measured. Skin reactions at the application site were observed 1 ± 0.1 h, 24 ± 2 h, 48 ± 2 h, and 72 ± 2 h after removal of the applied substance, and the skin irritation score was calculated.

[0112] The test results showed that no individuals died during the test period, and all individuals did not show any abnormal symptoms or weight changes that could be attributed to the administered substances.

[0113] The Primary Irritation Index (PII) at the application site of the test substance and the negative control substance was 0.0 (negligible). Ultimately, it was determined that the test substance did not cause skin irritation to the rabbit skin.

[0114]

Table 8

[0115]

[0116] 7. Test 7: Endotoxin test using lysate (colorimetric method)

[0117] This test is designed to quantitatively determine the endotoxin content of the test substance using the Limulus Amebocyte Lysate (LAS).

[0118] Endotoxin-free reagent water (LAL reagent water (LRW)) was selected as the elution solvent for the test substance according to USP 40 and USP 43, and elution was performed under the conditions shown in Table 9 below according to ISO 10993-12.

[0119]

Table 9

[0120]

[0121] The test substance dissolution solution was applied to a portable test system (PTS) cartridge and measured using a PTS reader. The channel information of the PTS cartridge is as follows.

[0122]

Table 10

[0123]

[0124] The endotoxin limit of the eluate (Endotoxin Limit) was calculated as follows using the endotoxin limit of the test substance (20 EU / device).

[0125] ERL (Endotoxin Release Limit)

[0126] =K×N / V=20EU×1 / 40ml=0.5EU / ml

[0127] K: Endotoxin limit per device (20EU)

[0128] N: the number of devices tested

[0129] V: the total volume of water used to extract the endotoxin from the devices tested

[0130] Using the endotoxin limit of the dissolution solution calculated above, the maximum effective dilution factor was calculated as follows.

[0131] Maximum effective dilution factor = ERL × concentration of test substance / λ (maximum sensitivity of PTS kit) = 0.5 EU / ml × 1 / 0.005 EU / ml = 100 times

[0132] The most sensitive sensitivity of PTS cartridge: 0.005EU / ml

[0133] Concentration of test substance: 1 (100% eluate)

[0134] The experimental groups are as follows.

[0135]

Table 11

[0136]

[0137] Group A was applied to channels 1 and 3 of the PTS cartridge, and Group B was applied to channels 2 and 4 of the PTS cartridge. Endotoxin recovery was measured to confirm the presence of interference factors. Groups C and D validated the information contained in the PTS cartridge by verifying the reliability of the calibration curve and LRW.

[0138] The test substance dissolution solution was prepared as a sample solution. The pH of the sample solution was then measured using pH indicator paper (MERCK), indicating a pH of 6.0 to 7.0. 25 μl of the sample solution was applied to each of the four channels of the PTS cartridge. The absorbance was measured at 395 nm using a PTS reader at 37 ± 1°C.

[0139] The test results showed that the coefficient of variation (Rxn time CV) for the sample and spike reactions was less than 25%, confirming the validity of the test. Furthermore, the endotoxin recovery rate was within the range of 50% to 200%, indicating the absence of interfering factors in the sample solution. Furthermore, the endotoxin concentration in the sample solution was less than 0.5 EU / ml. Therefore, under these experimental conditions, the endotoxin concentration in the test substance solution used in the test was determined to be less than 0.5 EU / ml.

[0140]

Table 12

[0141]

[0142]

[0143] In addition, the viscosity of the wound dressing material composition should be 500 to 10,000 mPa·s (Ministry of Food and Drug Safety Notice No. 2023-2, Korean Pharmacopoeia General Test Methods, 57. Viscosity Measurement Method)

[0144] The viscosity of the test example of the composition of the present invention was 5,420 mPa·s, which was measured to be within the standard range.

[0145]

Table 13

[0146]

[0147] The following are the results of a wound induction test on experimental animals. This test was intended to relatively evaluate the recovery ability of the wound dressing material composition of the present invention.

[0148] Test animals (specific pathogen-free (SPF) mice) were anesthetized (isoflurane inhalation anesthesia) and their back skin disinfected. After inducing wounds with a 5mm biopsy punch (diameter 5mm), the skin tissue was exfoliated. A control group, in which the natural healing process was observed, and a test group, in which the wound-covering material composition (test substance) provided by the present invention was applied and the healing process observed were compared. The test substance was soaked in sterile gauze (3cm x 5cm) (0.2g per application) and evenly applied to the wound lesion and the skin surrounding the lesion. An adhesive band (3cm x 10cm) was cut to fit around the abdomen and back of the test animal, and the bandage was secured to the abdomen and back of the test animal, with the gauze partially contacting the wound.

[0149] From the first to the tenth day of the experiment, the wound area was measured once a day, and based on the measured values, the wound recovery rate and the recovery ratio of the experimental group relative to the control group were derived by date.

[0150] To measure the wound area, use sterile surgical scissors to cut and remove the gauze and bandage. To measure the correct size of the wound area (lesion site), use sterile tweezers to remove the scab (scab), and use sterile gauze to stop bleeding for one minute when bleeding occurs. The wound area is calculated by measuring the long axis length and short axis length of the wound lesion using a vernier caliper.

[0151] The composition of the wound dressing material composition used in the test group is shown in Table 14 below, and was applied once a day to each wound lesion and the skin surrounding the lesion in the above manner.

[0152]

Table 14

[0153] Classification Content of each component (wt%) glycerin 65 Honey 33 Fucoidan 0.5 Olive Leaf Extract 1.3 Xanthan gum 0.2

[0154] Figure 4 This is a graph showing the wound recovery rates of the control group and the experimental group by date. Figure 5 This is a graph showing the recovery rate of the experimental group compared to the control group by date. Figure 6 The photos are taken of the wound recovery status of the control group and the experimental group according to the date.

[0155] exist Figure 4 In the study, compared with the control group, the experimental group showed a faster recovery rate until the fifth day of recovery. After the fifth day, the recovery rates of the experimental group and the control group were almost similar. Figure 5 The results showed that the experimental group showed a rapid increase in the recovery rate compared to the control group until the fourth day of recovery, and the difference in recovery rate gradually decreased after the fourth day. Therefore, it can be seen that the experimental group contributed to the rapid recovery of trauma.

[0156] While the present invention has been described above with reference to experimental examples, various modifications and variations are possible without departing from the spirit of the present invention, and the present invention can be applied to a variety of fields. Therefore, the scope of the claims of the present invention encompasses modifications and variations that fall within the true scope of the invention.

[0157] Industrial applicability

[0158] The invention can be used as a wound covering material for healing wounds and burns of human skin or animal skin and improving scars.

Claims

1. A wound dressing material composition, characterized in that: include: Glycerol, 60-70 wt%; Honey, 28-39 wt%; Fucoidan, 0.2-3.0 wt%; Olive leaf extract, 0.2-3.0 wt %; and Xanthan gum, 0.1-0.3wt%.

2. The wound dressing material composition according to claim 1, characterized in that The fucoidan is obtained by mixing dry spore leaf powder or extract of Undaria pinnatifida with purified water, performing centrifugal separation, and then filtering and recovering the mixture.

3. The wound dressing material composition according to claim 1, characterized in that The olive leaf extract is obtained by sterilizing a liquid obtained by filtering a powder of pulverized olive leaves or a liquid extracted from olive leaves by heating the liquid at 120 to 125° C. for 15 to 18 minutes.

4. The wound dressing material composition according to any one of claims 1 to 3, characterized in that The viscosity is adjusted within the range of 500 to 10,000 mPa·s according to the content of the xanthan gum.

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