KR102723348B1 - Composition having enhanced skin condition comprising extract of fermented pig placenta and hyaluronic acid - Google Patents

Abstract

The present invention relates to a composition for improving skin. The composition according to the present invention contains fermented placenta of a mammal and hyaluronic acid as active ingredients, and exhibits skin improving effects including skin moisturizing, elasticity improvement, exfoliation, and/or antioxidant effects.

Description

Composition having enhanced skin condition comprising extract of fermented pig placenta and hyaluronic acid

The present invention relates to a composition for improving skin.

The skin is an important organ that protects the human body from the external environment, acts as a barrier to prevent internal moisture and useful ingredients from leaking out, regulates body temperature, and performs various physiological functions such as excretion. However, for the following reasons, the activity of skin cells may decrease and the skin condition may worsen.

Collagen and elastin, which exist in the dermis layer of the skin, perform functions such as mechanical strength of the skin, resistance of connective tissue, maintenance of tissue cohesion, and support of cell adhesion, and thus have a great influence on the composition of the skin. This collagen is reduced by stress, aging, or photoaging caused by ultraviolet irradiation, and on the other hand, elastin is known to have its three-dimensional structure distorted by elastase, a decomposition enzyme whose activity increases after exposure to ultraviolet ray. As a result, the skin tissue becomes loose and loses elasticity.

As the cell function of the epidermis deteriorates, metabolism does not proceed smoothly, so dead skin cells do not shed well, and when exposed to ultraviolet rays while dead skin cells are excessively accumulated, elasticity decreases and wrinkles occur. Or, when a sebum film is not formed on the epidermis, moisture and sebum secretion decreases and the skin becomes dry, forming wrinkles.

When the skin is exposed to ultraviolet rays, it synthesizes melanin to protect itself. The synthesized melanin is transferred to the keratinocytes of the skin through melanosomes. These keratinocytes undergo a turnover phenomenon in a 28-day cycle. Therefore, the melanin that is created is usually lost in a 28-day cycle by the keratinocytes. However, if the skin cell cycle is not well regulated due to stress, skin aging, etc., the keratinocytes do not fall off in the proper time, resulting in pigmentation such as blemishes, freckles, and liver spots.

On the skin, sebum, sweat, and cosmetic ingredients are broken down into highly toxic substances by skin flora, which can irritate the skin and cause inflammation. In addition, irritation from ultraviolet rays can increase the production of nitric oxide (NO), an inflammatory mediator, in the skin, which can cause skin trouble. The production of nitric oxide is mostly due to iNOS, and iNOS is known to be rapidly induced by stimuli such as LPS and cytokines to produce excessive amounts of NO.

Reactive oxygen species (ROS), also called toxic oxygen species, are toxic substances that are constantly produced and destroyed in cells by physiological functions such as respiration, and exist at around 3-5% under normal conditions. These reactive oxygen species exist as free radicals (atoms or molecules that do not have pairs in their outermost electron orbitals, making them highly _unstable and reactive) such as superoxide radicals (O2-) and hydroxyl radicals (HO+), or as compounds with paired electrons such as hydrogen peroxide ( _H2O2 ) and singlet radicals. Reactive oxygen species have the advantage of having a biological defense function that sterilizes bacteria in the physiological system, but they generally have a harmful effect that causes diseases by causing oxidation in the body. It has been reported that these reactive oxygen species attack biomolecules, damage cells and tissues, and cause various diseases related to aging and various adult diseases.

Research has continued to develop substances that can suppress skin aging caused by the various factors described above and have moisturizing, elasticity-improving, exfoliating, and/or antioxidant effects.

Republic of Korea Patent No. 10-0901138

The problem to be solved by the present invention is to provide a composition use that has excellent skin improvement effects, such as skin moisturizing, elasticity improvement, exfoliation, and/or antioxidant effects, while having few side effects and being safe for the human body.

In other words, the problem that the present invention seeks to solve is to provide a composition having excellent skin improvement effects.

In order to solve the above problems, the present invention provides a composition for skin moisturizing, elasticity improvement, exfoliation and/or antioxidation containing fermented placenta of a mammal and hyaluronic acid as effective ingredients, and preferably provides a cosmetic composition, a pharmaceutical composition or a food composition.

As a result of the research efforts of the present inventors to develop a material having a skin improvement effect, they confirmed that a composition containing mammalian fermented placenta and hyaluronic acid as active ingredients has skin moisturizing, elasticity improvement, exfoliation, and/or antioxidant effects, and completed the present invention. That is, the inventors of the present invention experimentally confirmed a remarkable synergistic effect in skin moisturizing, elasticity improvement, exfoliation, and/or antioxidant effects compared to a composition containing mammalian fermented placenta or hyaluronic acid as active ingredients, respectively.

In the present invention, 'skin moisturizing' means increasing moisture in the skin and maintaining the moist state. The skin moisturizing effect can help improve wrinkles and increase elasticity of the skin.

In the present invention, ‘improving elasticity’ means alleviating the degree of sagging or stretching of the skin.

In the present invention, 'removal of dead skin cells' means the sloughing off of dead skin cells present on the skin surface. When dead skin cells are excessively accumulated and exposed to ultraviolet rays, elasticity decreases and wrinkles occur. Therefore, if the effect of removing dead skin cells is excellent, it is effective in improving wrinkles on the skin. If dead skin cells are not removed well, pigmentation such as blemishes, freckles, and liver spots may occur. Therefore, if the effect of removing dead skin cells is excellent, it also helps in whitening the skin.

In the present invention, 'antioxidant' refers to suppressing oxidative stress. In the human body, substances that cause oxidative stress can occur. When oxygen used as an energy source in the body is incompletely reduced, active oxygen is released into the cytoplasm. This active oxygen can damage surrounding cells and further promote tissue damage due to inflammation. However, although this active oxygen is generated even under normal circumstances, if there is no antioxidant defense mechanism in the body, we cannot survive. Therefore, our body has various forms of antioxidant defense systems to counter normal oxidative stress (Park Jong-yeol et al., BioWave 2004; 6 (13)). However, if oxidative stress exceeds the normal level, the body suffers serious damage, so countermeasures against oxidative stress are necessary. Antioxidants are natural or synthetic substances that function to prevent or suppress oxidative stress. In order to evaluate the antioxidant power of these substances, various methods such as oxygen radical absorbance capacity (ORAC), total polyphenol contents, total flavonoid contents, DPPH free radical scavenging activity, and ferric reducing/antioxidant power (FRAP) are used. These methods are mainly measures of the ability to remove free radicals such as active oxygen or the content of antioxidant substances. In the present invention, the antioxidant effect of the composition was confirmed through free radical scavenging activity.

In the present invention, the placenta refers to a structure that mediates the exchange of substances necessary for the survival and growth of the fetus between the fetus and the mother. In the present invention, the placenta may preferably be a mammalian placenta, and for example, the placenta of a human, pig, horse, sheep, cow, etc. may be used. However, the use of the human placenta is regulated in industries other than pharmaceuticals, and there are ethical issues, so the placenta of a pig is more preferably used.

The present inventors have confirmed that using a fermented placenta has better skin moisturizing, elasticity improvement, exfoliation, and/or antioxidant effects than using a natural placenta of a mammal, and therefore, a fermented placenta may be preferably used. The fermented placenta in the present invention refers to a placenta fermented with a fermenting microorganism, and is not limited thereto, but a commercially sold placenta may be purchased and fermented, or a commercially sold fermented placenta may be used. Any method for fermenting the placenta is not limited thereto, so long as it is a conventional method for fermenting the placenta in the art.

For example, a method for producing a fermented placenta may include (S1) a step of crushing a mammalian placenta and removing blood; and (S2) a step of adding a fermenting microorganism and fermenting, and may further include a step of adding and mixing a carbon source and/or an energy source of the fermenting microorganism added before or in step (S2), and the carbon source and/or energy source may be any one known in the art. For example, oligosaccharides, lactose, glucose, fructose, sugar (including brown sugar), galactose, maltose, starch, or a mixture of two or more thereof may be used, and preferably, a monosaccharide or a disaccharide may be used. In addition, it may be added in an arbitrary range considering the fermentation time, the degree of fermentation, etc. For example, it may be added in a range of 3 to 30 parts by weight with respect to 100 parts by weight of the crushed placenta.

In the step (S1), the size of the pulverized mammalian placenta is not particularly limited to a specific size, but is preferably a size that allows the pulverized material to be uniformly fermented by the microorganisms to be added. The size refers to the longest length when the length is measured from the particles of the pulverized material, and the size that allows the pulverized material to be uniformly fermented by the microorganisms to be added can be determined within the range of typical capabilities in the art. For example, it can be used by being pulverized to a size of 0.3 to 1 mm. Since the mammalian placenta usually has a moisture content of about 90%, there is no need to specifically supply moisture for fermentation. However, when the mammalian placenta is pulverized by a method such as freeze-drying and used as a raw material for fermentation, moisture can be supplied for fermentation.

In the above step (S2), the fermenting microorganism may be a microorganism such as lactic acid bacteria, Bacillus genus, or yeast ( Saccharomycetes class), but is not limited thereto.

The above fermentation microorganism is Aspergillus genus Any microorganism capable of fermentation, such as Penicillium sp., Rhizopus sp., Acetobacter sp., Saccharomyces sp., Torulopsis sp., and lactic acid bacteria, can be used. The amount can be added in an arbitrary range considering the fermentation time, fermentation degree, etc. For example, it can be added in a range of 3 to 30 parts by weight per 100 parts by weight of the placental pulverize.

The above lactic acid bacteria may be Lactobacillus sp., Streptococcus sp., Pediococcus sp., Leuconostoc sp., and Bifidobacterium sp. More specifically, Lactobacillus plantarum, Lactobacillus delbueckii , Lactobacillus bulgaricus , Lactobacillus casei , Lactobacillus brevis , Lactobacillus acidophilus , Pediococcus pentosaceus , Pediococcus cerevisiae cerevisiae ), Leuconostoc citreum , Leuconostoc mesenteroides , Bifidobacterium bifidum bifidum ), Bacillus coagulans , etc. can be used.

For yeast, yeasts of the genus Saccharomyces ( Saccharomyces sp.) are preferred, such as Saccharomyces lucii ( Saccharomyces rouxii ), Saccharomyces cerevisiae cereviciae ), Saccharomyces oviformis , Saccharomyces steineri , Saccharomyces ellipsoideus ellipsoideus ), Saccharomyces coreanus , etc. can be used.

For the genus Bacillus, Bacillus subtilis , Bacillus lichneformis , Bacillus megaterium , Bacillus amyloliquefaciens , Bacillus natto , Bacillus antharcis , Bacillus lentus, Bacillus pumilus, Bacillus thuringiensis , Bacillus alvei , Bacillus azotofixans , Bacillus macerans , Bacillus polymyxa , Bacillus Bacillus polilliae , Bacillus coagulans , Bacillus stearothermophilus , Bacillus pasteurii , Bacillus sphaericus , Bacillus fastidiosus , etc. can be used.

In addition, in the step (S2), it is preferable that the fermentation is carried out until the growth of the added microorganism reaches its maximum. The point in time when the growth of the added microorganism reaches its maximum can be determined according to the fermentation temperature, the amount of the fermentation raw material, the amount of the added microorganism, etc. A person skilled in the art can determine the point in time when the growth of the added microorganism reaches its maximum by considering the fermentation temperature, the amount of the fermentation raw material, the amount of the inoculated microorganism, etc. within the scope of his or her normal ability.

The fermentation of the above (S2) step can be carried out within a range of 15°C to 45°C, and preferably within a range of 20°C to 40°C. If the fermentation temperature is lower than the above range, the fermentation rate may be slowed and undesirable fermentation products may be generated, and if the fermentation temperature is higher than the above range, the fermentation rate may be slowed or undesirable fermentation products may be generated due to the death of fermentation microorganisms.

In addition, a maturation step may be additionally included after the step (S2). The maturation step is intended to stabilize the fermentation product by the microorganism even after the growth of the added microorganism has stopped, and may be performed for, but is not limited to, 10 days to 12 months.

In addition, a sterilization step may be additionally included after the step (S2). The sterilization step may be performed using a conventional method known in the art, such as heat treatment or ultraviolet irradiation.

The fermented placenta manufactured according to the above fermented placenta manufacturing method is initially obtained in a liquid form, but can be concentrated by methods such as reduced pressure concentration or freeze drying, or used in powder form. The fermented placenta can also be used by precipitating it with an appropriate precipitant such as ethanol or acetone, removing the supernatant, and then recovering the precipitate.

That is, the term "placenta" used in this specification is a term that includes all of the placenta's pulverized product, extract, extract liquid, concentrate, filtrate, filtrate, solution, etc., and preferably means an extract of the placenta.

The above fermented placenta may be included in an amount of 0.0001 to 10 wt% based on the total weight of the composition, preferably 0.001 to 5 wt%, and more preferably 0.01 to 1 wt%.

In the present invention, the hyaluronic acid is a substance having a repeating structure of a disaccharide in which N-acetyl D-glucosamine (GlcNAc) and D-glucuronic acid (GlcA) are linked via a β1-4 glycosidic bond, and the disaccharide forms hyaluronic acid via a β1-3 bond. Hyaluronic acid is present in almost all tissues of vertebrates, such as the placenta, synovial fluid, vitreous humor, and rooster comb, but is not limited thereto, and is present in large quantities in connective tissue areas, such as the skin and synovial fluid.

The hyaluronic acid may be contained in an amount of 0.0001 to 10 wt% based on the total weight of the composition, preferably 0.001 to 5 wt%, and more preferably 0.01 to 1 wt%.

The composition according to the present invention has a technical characteristic in that it has confirmed a synergistic effect in skin improvement by including both fermented placenta and hyaluronic acid of a mammal rather than including them alone, and the fermented placenta and hyaluronic acid may be included in a weight ratio (fermented placenta: hyaluronic acid) of 1:0.1 to 30, preferably 1:0.2 to 15, more preferably 1:0.5 to 8.

According to one embodiment of the present invention, the present invention provides a cosmetic composition for moisturizing the skin, improving elasticity, removing dead skin cells, and/or antioxidation, comprising fermented placenta of a mammal and hyaluronic acid as active ingredients.

Cosmetics manufactured by containing the composition as an effective ingredient as a cosmetic composition can be manufactured in the form of general emulsified formulations and solubilized formulations. For example, the formulations may be, but are not limited to, toners such as flexible toners or nourishing toners, emulsions such as facial lotions and body lotions, creams such as nourishing creams, moisture creams, and eye creams, essences, cosmetic ointments, sprays, gels, packs, sunscreens, makeup bases, foundations in liquid, solid, or spray types, makeup removers such as powders, cleansing creams, cleansing lotions, and cleansing oils, and cleansers such as cleansing foams, soaps, and body washes.

In the case of the composition being manufactured as a cosmetic, in the case of wash-off type cosmetics such as makeup removers and cleansers, in which the active ingredients remain on the skin for a short period of time, the composition of the present invention may be included in a relatively high concentration. On the other hand, in the case of leave-on type cosmetics such as toners, emulsions, creams, and essences, in which the active ingredients remain on the skin for a long period of time, the composition of the present invention may be included in a lower concentration than in wash-off type cosmetics.

In addition, the cosmetic may contain, in addition to the composition of the present invention, adjuvants commonly used in the field of cosmetology, such as fatty substances, organic solvents, solubilizers, thickening and gelling agents, emollients, antioxidants, suspending agents, stabilizers, foaming agents, fragrances, surfactants, water, ionic or nonionic emulsifiers, fillers, sequestering and chelating agents, preservatives, vitamins, blocking agents, humectants, essential oils, dyes, pigments, hydrophilic or lipophilic active agents, lipid vesicles or any other ingredients commonly used in cosmetics.

In addition, the cosmetic composition of the present invention may additionally include at least one cosmetically acceptable carrier that is incorporated into a general skin cosmetic.

The cosmetically acceptable carrier included in the cosmetic composition of the present invention varies depending on the formulation. When the formulation of the present invention is an ointment, paste, cream or gel, animal oil, vegetable oil, wax, paraffin, starch, tragacanth, cellulose derivatives, polyethylene glycol, silicone, bentonite, silica, talc, zinc oxide or mixtures thereof can be used as the carrier component.

When the formulation of the present invention is a powder or spray, lactose, talc, silica, aluminum hydroxide, calcium silicate, polyamide powder or a mixture thereof may be used as a carrier component, and particularly in the case of a spray, a propellant such as chlorofluorohydrocarbon, propane/butane or dimethyl ether may be additionally included.

When the formulation of the present invention is a solution or an emulsion, a solvent, a solubilizer, or an emulsifier is used as a carrier component, and examples thereof include water, ethanol, isopropanol, ethyl carbonate, ethyl acetate, benzyl benzoate, propylene glycol, 1,3-butyl glycol oil, and in particular, cottonseed oil, peanut oil, corn germ oil, olive oil, castor oil, and sesame oil, glycerol aliphatic esters, polyethylene glycol, or fatty acid esters of sorbitan.

When the formulation of the present invention is a suspension, liquid diluents such as water, ethanol or propylene glycol, suspending agents such as ethoxylated isostearyl alcohol, polyoxyethylene sorbitol ester and polyoxyethylene sorbitan ester, microcrystalline cellulose, aluminum metahydroxide, bentonite, agar or tragacanth, etc. can be used as carrier components.

When the formulation of the present invention is soap, alkali metal salts of fatty acids, fatty acid hemiester salts, fatty acid protein hydrolysates, isethionates, lanolin derivatives, fatty alcohols, vegetable oils, glycerol, sugars, etc. can be used as carrier components.

In addition, according to one embodiment of the present invention, the present invention provides a pharmaceutical composition for skin moisturizing, elasticity improvement, exfoliation and/or antioxidation, comprising mammalian fermented placenta and hyaluronic acid as active ingredients.

The composition may have any one dosage form selected from the group comprising a solution, a gel, an emulsion, a suspension, a microemulsion, a microcapsule, a liposome, a cream, a lotion, an ointment, an aerosol, a spray, a paste, and a patch.

The pharmaceutical composition of the present invention can be used as a single preparation, or can be manufactured and used as a combination preparation, and can be manufactured in a unit dose form by formulating it using a pharmaceutically acceptable carrier or excipient, or can be manufactured by placing it in a multi-dose container.

As used herein, the term "pharmaceutically acceptable carrier" may mean a carrier or diluent that does not stimulate a living organism and does not inhibit the biological activity and properties of the compound to be injected. The type of the carrier usable in the present invention is not particularly limited, and any carrier that is commonly used in the art and is pharmaceutically acceptable may be used. Non-limiting examples of the carrier include saline solution, sterile water, Ringer's solution, buffered saline, albumin injection solution, dextrose solution, maltodextrin solution, glycerol, ethanol, and the like. These may be used alone or in combination of two or more. The carrier may include a non-naturally occurring carrier.

In addition, if necessary, other conventional additives such as antioxidants, buffers, and/or bacteriostatic agents may be added and used, and diluents, dispersants, surfactants, binders, lubricants, etc. may be additionally added and used as injectable formulations such as aqueous solutions, suspensions, and emulsions, pills, capsules, granules, or tablets.

In addition, the pharmaceutical composition of the present invention may contain a pharmaceutically effective amount of fermented placenta of a mammal and hyaluronic acid. The term "pharmaceutically effective amount" as used herein means an amount sufficient to induce skin moisturizing, elasticity improvement, exfoliation, or antioxidant effects at a reasonable benefit/risk ratio applicable to medical treatment, and may be administered once or several times a day in an amount of generally 0.001 to 1000 mg/kg, preferably 0.05 to 200 mg/kg, more preferably 0.1 to 100 mg/kg. However, for the purpose of the present invention, a specific therapeutically effective amount for a specific patient is preferably applied differently depending on various factors including the type and degree of the response to be achieved, the specific composition including whether other agents are used depending on the case, the age, weight, general health condition, sex, and diet of the patient, the time of administration, the route of administration, and the secretion rate of the composition, the treatment period, drugs used together or simultaneously with the specific composition, and similar factors well known in the medical field.

In addition, according to one embodiment of the present invention, the present invention provides a food composition for skin moisturizing, elasticity improvement, exfoliation and/or antioxidation, comprising mammalian fermented placenta and hyaluronic acid as effective ingredients.

The term "food" used in this specification means a food prepared by adding the fermented placenta and hyaluronic acid of the mammal to food materials such as beverages, teas, spices, gums, and confectionery, or by manufacturing the same in the form of encapsulation, powder, suspension, etc., and which has a specific effect when consumed; however, unlike general drugs, it has the advantage of not having side effects that may occur with long-term use of drugs made from food. The food of the present invention obtained in this way is very useful because it can be consumed on a daily basis and can be expected to have excellent effects in moisturizing the skin, improving elasticity, removing dead skin cells, and/or antioxidation.

The food composition of the present invention may additionally contain a food-wise acceptable carrier.

There is no particular limitation on the type of food to which the composition of the present invention can be added, and examples thereof include various beverages, gum, tea, vitamin complexes, health supplements, etc. Other ingredients that do not interfere with the effects of the present invention can be added to the food composition, and the types thereof are not particularly limited. For example, as with conventional foods, various herbal extracts, food-wise acceptable food supplement additives, or natural carbohydrates can be contained as additional ingredients.

The above food additives are added in the manufacture of each type of food, and can be appropriately selected and used by a person skilled in the art. For example, various nutrients, vitamins, minerals (electrolytes), flavoring agents such as synthetic flavoring agents and natural flavoring agents, coloring agents and fillers, pectic acid and its salts, alginic acid and its salts, organic acids, protective colloid thickeners, pH regulators, stabilizers, preservatives, glycerin, alcohol, carbonating agents used in carbonated beverages, etc. are included, but the types are not limited by the above examples.

There are no special restrictions on the types of the above foods. Examples of foods to which the above substances can be added include meat, sausage, bread, chocolate, candy, snacks, confectionery, pizza, ramen, other noodles, gum, dairy products including ice cream, various soups, beverages, tea, drinks, alcoholic beverages, and vitamin complexes, and all foods in the conventional sense are included.

If the food is a beverage, it may be included in a ratio of 1 to 30 g, preferably 3 to 20 g, per 100 ml. In addition, the composition may include additional ingredients commonly used in food compositions to improve odor, taste, sight, etc. For example, it may include vitamins A, C, D, E, B1, B2, B6, B12, niacin, biotin, folate, pantothenic acid, etc. In addition, it may include minerals such as zinc (Zn), iron (Fe), calcium (Ca), chromium (Cr), magnesium (Mg), manganese (Mn), copper (Cu), etc. In addition, it may include amino acids such as lysine, tryptophan, cysteine, and valine. In addition, food additives such as preservatives (potassium sorbate, sodium benzoate, salicylic acid, sodium dehydroacetate, etc.), bactericides (bleaching powder and highly bleached powder, sodium hypochlorite, etc.), antioxidants (butylated hydroxyanisole (BHA), butylated hydroxytoluene (BHT), etc.), coloring agents (tar color, etc.), coloring agents (sodium nitrite, sodium nitrate, etc.), bleaching agents (sodium sulfite), seasonings (MSG (monosodium glutamate)), sweeteners (dulcin, cyclamate, saccharin, sodium, etc.), flavorings (vanillin, lactones, etc.), leavening agents (alum, D-potassium hydrogen tartrate, etc.), reinforcing agents, emulsifiers, thickeners (glucose), film agents, gum bases, foam suppressants, solvents, and improvers can be added. The above additives are selected depending on the type of food and used in an appropriate amount. The proportion of these additives is not particularly important, but is typically selected in the range of 0.01 to 0.1 parts by weight per 100 parts by weight of the composition of the present invention.

The food composition of the present invention can be manufactured by a method commonly used in the art, and can be manufactured by adding raw materials and ingredients commonly added in the art during the manufacturing process. In addition, unlike general medicines, it has the advantage of not having side effects that may occur when taking medicine for a long period of time by using food as a raw material, and can be highly portable.

The composition according to the present invention contains fermented mammalian placenta and hyaluronic acid as active ingredients, and exhibits skin improvement effects including skin moisturizing, elasticity improvement, exfoliation, and/or antioxidant effects.

Figure 1 is a graph showing radical scavenging ability according to examples and comparative examples of the present invention.
Figure 2 is a graph showing the skin moisturizing effect according to the intake of a composition according to the present invention.
Figure 3 is a graph showing the effect of improving skin elasticity according to intake of a composition according to the present invention.
Figure 4 is a photograph measuring the amount of skin keratin following ingestion of a composition according to the present invention.
Figure 5 is a graph showing the effect of reducing skin keratin by ingestion of a composition according to the present invention.

Hereinafter, in order to help understand the present invention, examples and the like will be described in detail. However, the examples according to the present invention may be modified in various different forms, and the scope of the present invention should not be construed as being limited to the following examples. The examples of the present invention are provided to more completely explain the present invention to a person having average knowledge in the art.

Manufacturing example 1: Fermentation Half of the money and preparation of a composition comprising hyaluronic acid ( Comparative example 1 to 4, Example 1)

Unfermented swine placenta (SPF Swine Placenta, VS-PL E type) and fermented swine placenta (Fermented Placenta Extract Powder A (K)) were purchased from HORUS Co., Ltd., Japan. To obtain fermented swine placenta extract, the enzymatically hydrolyzed swine placenta was sterilized and filtered, and Saccharomycetes and brown sugar were added, followed by fermentation at 25–30°C for 1 month. After that, it was aged for 2 months at 25°C, sterilized, and spray-dried to manufacture powder.

Hyaluronic acid was purchased from SK Bioland Co., Ltd. and used. Hyaluronic acid powder (hyaluronic acid content of 90% or more) was used by concentrating and filtering hyaluronic acid produced through Streptococcus zooepidemicus microbial fermentation, precipitating it with ethanol, and then drying it.

A mixture was prepared by adding the ingredients in the amounts shown in Table 1 below.

Comparative Example 1 Comparative Example 2 Comparative Example 3 Comparative Example 4 Example 1 Hyaluronic acid (mg/ml) 2 - - 1 1 Normal placenta (mg/ml) - 2 - 1 - Fermented pig placenta (mg/ml) - - 2 - 1 water To 100 To 100 To 100 To 100 To 100

Experimental example 1: Confirmation of antioxidant activity

To confirm the antioxidant effect of the composition, the inventors measured radical scavenging activity using DPPH (1,1-diphenyl-2-picryl-hydrazyl).

First, 4 mg of DPPH was dissolved in 50 ml of ethanol to make a DPPH solution, and hyaluronic acid, fermented pig placenta extract, and normal pig placenta extract were diluted in distilled water to make an appropriate concentration. Ascorbic acid was used as a standard, and 100 μl of the standard and each sample were dispensed into 3 wells of a 96-well plate, and 100 μl of DPPH solution was added to each well. The plate was wrapped in foil and left in a 37°C incubator for 30 minutes, and then the absorbance was measured at 516 nm. Distilled water was used as the control group, and the antioxidant power by the DPPH method was calculated using the following mathematical equation 1.

[Mathematical formula 1]

Antioxidant power (%) = 100 - (absorbance of sample/absorbance of control) × 100

The results of the antioxidant activity experiment determined through absorbance are shown in Table 2 below.

Comparative Example 1 Comparative Example 2 Comparative Example 3 Comparative Example 4 Example 1 Antioxidant power (%) 28.1 21.5 33.7 38.9 57.2

As a result, it was confirmed that Example 1 containing fermented pig placenta and hyaluronic acid had a remarkable antioxidant effect compared to Comparative Examples 1 to 3 containing each component as shown in Table 2 above and Comparative Example 4 containing general pig placenta and hyaluronic acid.

That is, the composition of the present invention contains both fermented placenta and hyaluronic acid as effective ingredients, and thus a synergistic effect in terms of antioxidant effect was confirmed.

Experimental example 2: skin cells Confirmation of proliferation effect

In order to confirm the skin cell proliferation effect of the composition shown in Table 1 above, the inventors of the present invention first cultured human dermal fibroblast (HDF) cell lines derived from human skin in a DMEM medium containing 10% fetal bovine serum (FBS) for 48 hours (37°C, 5% _CO2 incubator).

After that, the culture medium was removed, the culture components were washed with PBS solution, the cells were treated with trypsin to detach from the culture vessel, and then centrifuged. After centrifugation, the supernatant was removed, and the cells were diluted again in an appropriate amount of DMEM containing 10% FBS and seeded in a 96-well plate with a cell number of 1 × 10 ⁵ per well. After about 24 hours, when the cells were about 70% full in each well, the general medium was washed out and replaced with a serum-free medium containing each composition shown in Table 1, and then cultured for 24 hours. Then, 20 ㎕ of MTT solution (3-(4,5-dimethylthiazol-2yl)-2,5-diphenyl-2H-tetrazolium bromide) was added and cultured in a 37℃ CO ₂ incubator for 4 hours.

After that, the medium was carefully removed, 200㎕ of DMSO was added, and the plate was left on a shaker for 15 minutes. The absorbance was measured at 540 nm using an ELISA reader. The control group was set as a well that was not treated with any sample. The absorbance of the control group was set to 100, and the survival rate of each sample treatment group compared to the control group is shown in Figure 1.

As a result, as shown in Fig. 1, when the cell viability of the control group was set as 100%, the cell viability tended to increase by each sample or the combined treatment, but it significantly increased in Comparative Example 1 (115.7%, p=0.04) and Example 1 (135.7%, p=0.0046). Among them, Example 1 showed a remarkable increase, indicating that the combined treatment of hyaluronic acid and fermented placenta had a remarkable synergistic effect on skin cell proliferation.

Experimental example 3: Check skin improvement effects (skin moisturizing, elasticity improvement, exfoliation)

In order to clinically confirm the skin improvement synergy effect of fermented placenta and hyaluronic acid confirmed in Experimental Examples 1 and 2, the inventors recruited 30 clinical subjects.

A beverage with the same taste, smell, and color was manufactured, and the subjects were divided into three groups: a beverage containing hyaluronic acid, a beverage containing fermented pig placenta, a beverage containing both hyaluronic acid and fermented pig placenta, and a control group containing nothing. The subjects consumed 100 ml of the beverage per day for four weeks.

After stabilizing for 30 minutes in a constant temperature and humidity room (22–24°C, 40–50% RH) at weeks 0, 2, and 4, the keratin mass, moisture content, and elasticity were measured. The keratin mass was measured by placing a keratin tape on a designated area, pressing it with a pressure plate for 5 seconds, removing the tape, and taking a picture with a scanvisiometer. Moisture content was measured using a Corneometer (CM825), and elasticity was measured using a Cutometer (MPA580). The mixing ratio of drinks for each group is as shown in Table 3 below.

(Unit: Weight%) Control group
(n=8) Test group 1
(n=8) Test group 2
(n=6) Test group 3
(n=8) Purified water 89.8133 89.6733 89.6733 89.6733 sugar 10 10 10 10 pigment 0.0267 0.0267 0.0267 0.0267 incense 0.16 0.16 0.16 0.16 Hyaluronic acid 0 0.14 0 0.07 Fermented pork placenta 0 0 0.14 0.07 Total 100 100 100 100

1) Skin moisturizing effect

Skin hydration was measured three times using a Corneometer at a fixed location on the inside of the right arm and the average was calculated. The results are shown in Fig. 2. The units were expressed in arbitrary units (AU) assigned by the device.

It was confirmed that moisturizing tended to increase in test groups 1 and 3, and in the 4th week, test group 1 increased significantly by 113.5% (p=0.02) from an average of 29.06 to 33 compared to week 0, and test group 3 increased significantly by 121.5% (p=0.008) from an average of 30.26 to 36.76 compared to week 0. There was no significant difference between the control group and test group 2.

Therefore, it was confirmed that the skin improvement effect was improved more by combined intake of hyaluronic acid and fermented pig placenta than by intake of hyaluronic acid or fermented pig placenta alone.

2) Improves skin elasticity

Elasticity was measured using the Cutometer, which pulls the skin into the probe with negative pressure and then returns the skin to its original state when the negative pressure is removed. The results are shown in Figure 3.

The R2 (gross elasticity), which is the value obtained by measuring the same area around the right eye, was used as a variable, and the closer this value is to 1, the higher the elasticity.

As a result, test groups 1 and 3 showed a trend of increasing elasticity, and it was confirmed that elasticity significantly increased in test group 3 at the 4th week (p = 0.0442), confirming that the skin improvement effect was improved more by combined intake of hyaluronic acid and fermented pig placenta than by intake of hyaluronic acid or fermented pig placenta alone.

3) Skin exfoliation effect

The keratin mass was calculated by attaching a keratin tape to a certain area on the inside of the right arm, pressing it with a pressing plate for 5 seconds, then removing it and calculating the area of the keratin area as a % of the total area using a scanvisiometer. The area was automatically displayed by software as shown in the photograph in Fig. 4, and the results were presented as a graph in Fig. 5.

Figure 5 is a graph showing the quantitative analysis of each mass. In test group 3, a significant decrease (p=0.0458) was observed from 10.46% at week 0 to 9.06% at week 4, confirming that the skin improvement effect was improved more by combined intake of hyaluronic acid and fermented pig placenta than by intake of hyaluronic acid or fermented pig placenta alone.

Claims (11)

A food composition for moisturizing the skin, improving elasticity, removing dead skin cells or providing antioxidant effects, comprising fermented placenta of mammals and hyaluronic acid as active ingredients. In the first paragraph,
A food composition characterized in that the fermented placenta is contained in an amount of 0.0001 to 10 wt% based on the total weight of the composition. In the first paragraph,
A food composition characterized in that the hyaluronic acid is contained in an amount of 0.0001 to 10 wt% based on the total weight of the composition. In the first paragraph,
A food composition characterized in that the fermented placenta and hyaluronic acid are included in a weight ratio of 1:0.1 to 30 (fermented placenta: hyaluronic acid). In the first paragraph,
A food composition characterized in that the mammal is a pig. A cosmetic composition for moisturizing the skin, improving elasticity, removing dead skin cells or providing antioxidant effects, comprising fermented placenta of a mammal and hyaluronic acid as active ingredients. In Article 6,
A cosmetic composition characterized in that the fermented placenta of the mammal is contained in an amount of 0.0001 to 10 wt% based on the total weight of the composition. In Article 6,
A cosmetic composition characterized in that the hyaluronic acid is contained in an amount of 0.0001 to 10 wt% based on the total weight of the composition. A pharmaceutical composition for moisturizing the skin, improving elasticity, removing dead skin cells or providing antioxidant properties, comprising fermented placenta of a mammal and hyaluronic acid as active ingredients. In Article 9,
A pharmaceutical composition characterized in that the fermented placenta of the mammal is contained in an amount of 0.0001 to 10 wt% based on the total weight of the composition. In Article 9,
A pharmaceutical composition characterized in that the hyaluronic acid is contained in an amount of 0.0001 to 10 wt% based on the total weight of the composition.