KR102295754B1 - Composition comprising extract of soybean embryo for strengthening hair root, activating hair growth or preventing alopecia - Google Patents

Abstract

The present invention relates to a composition for strengthening hair roots, hair growth or preventing stress-related hair loss, comprising a soybean embryo extract as an active ingredient. The soybean embryo extract of the present invention increases the expression of a VEGF gene, known as a factor related to hair root strengthening, and an IFG-1 gene, which is known to be a hair growth-related factor, and has a function of alleviating the stress-related hair loss.

Description

Composition comprising extract of soybean embryo for strengthening hair root, activating hair growth or preventing alopecia }

The present invention relates to a composition for strengthening hair roots, promoting hair growth or preventing hair loss, containing soybean germ extract.

Human hair is very important because it plays an intrinsic role in social and sexual communication, as well as its primary role in protecting the skin and scalp.

The hair cycle consists of an anagen phase, a catagen phase, and a telogen phase, and the hair is maintained by repeating hair growth and hair loss while repeating the cycle. Hair growth period is 3 to 5 years for men, 4 to 6 years for women, 30 to 45 days for hair regression, and 3 to 4 months for resting period. When the final stage of the telogen phase is reached, the generator naturally begins to generate new hair and the hair is maintained. Since hair loss is a natural phenomenon, alopecia, which is recognized as a hair loss disease, refers to a state in which there are relatively many hairs in a telogen state and there are few hairs.

The cause of hair loss is not yet known precisely, but it is a major factor that affects hair growth. It inhibits or decreases the function of dermal papilla, which is related to the regulation of the hair cycle, abnormal hair cycle due to male hormone action, and scalp. Abnormal changes in the hair cycle due to a decrease in blood flow to the hair follicles, antihypertensive drugs, mental stress, physical stimulation, and environmental pollution are being discussed.

In the past, hair loss mainly occurred in middle-aged and elderly people, but recently, hair and scalp damage due to mousse, perm, dyeing, and dry heat, increase in social and environmental stress, and changes in food due to westernization of diet, etc. Hair loss is remarkably occurring in young people in their 30s, and not only male pattern hair loss but also female pattern hair loss is on the rise.

For this reason, although many therapeutic agents for preventing hair loss are on the market worldwide, there are still few drugs or technologies that can satisfy the therapeutic effect. Currently, it is known that the progress of hair loss can be slowed down to some extent by using the topical 'minoxidil' or the oral drug 'Propecia (finasteroid)' in the early stages of hair loss. There are difficulties, and body hair grows in unwanted areas, sexual dysfunction such as erectile dysfunction and loss of libido, and side effects such as dizziness, headache, edema and skin rash may occur. There is a problem that cannot be.

Among them, 'Minoxidil' is a drug approved by the US FDA, and the exact treatment mechanism is not yet clear, but dermal papilla cells (DPCs) that are reported to affect the hair cycle through the secretion of proteins such as various growth factors. ) is known to have the effect of proliferating the growth of dermal papilla cells (DPCs) through reduction of apoptosis. In addition, drugs such as 'Propecia', a 5-alpha reductase inhibitor, inhibit the production of dihydrotestosterone (DHT), a male hormone that causes hair loss, and prevent the progression of male pattern hair loss. It is known that measuring the activity inhibitory effect of 5-alphareductase or the growth and proliferation effect of dermal papilla cells (DPCs) for a certain substance shows how effectively the ingredient prevents hair loss and contributes to hair growth. It can be used as an indicator to know.

Hair is made up of keratin protein and grows from hair follicles in the dermis. Scalp hair follicles are composed of dermal papilla cells, keratinocytes, inner and outer hair root sheath cells, and melanocytes. In addition, hair follicle dermal papilla cells are known to be closely related to hair growth.

Recently, studies on many regulatory factors involved in hair growth and hair loss mechanisms are being actively conducted at various research institutes. continuously reported. Growth factors that affect hair growth include EGF and EGFF, VEGF and VEGFR, HGF/SF and c-MET, FGF family and FGFR, IGF and IGF-IR, TGF-β and TGF-βR, etc. It is known to affect the hair cycle by regulating the activity of In addition, it is known that the Wnt pathway and Bmp signaling play a decisive role in the proliferation or differentiation of hair follicle stem cells in the bulge region.

On the other hand, recently, the number of hair loss population continues to increase due to environmental stress, etc. along with side effects due to environmental pollution and misuse of drugs. The development of new agents for preventing hair loss is emerging.

In addition, among the currently developed anti-hair loss agents, especially synthetic compound formulations, there is a problem in causing various side effects on the skin.

Looking at these prior arts, Republic of Korea Patent No. 10-0800359 (composition showing hair loss prevention and hair growth promoting effect) discloses the use of natural plants such as silk grass, ginseng, jujube, goji berry and cornus oil, and a Korean registered patent No. 10-0773457 (composition having a hair growth promoting effect and a method of using the same) discloses a composition effective for preventing hair loss and promoting hair growth using ginseng, white sorghum, angelica, astragalus, safflower jasmine, hyssop, and headworm as main raw materials.

As such, there are many developments as a mixed extract using various kinds of herbs to promote hair growth, but research on the discovery of substances having effects of hair growth, hair root strengthening, and hair loss prevention from soybean germ components is still insufficient. the current situation. As soybean germ is known to give a bitter taste in soymilk, it is a part that is removed during the selection and processing process of soybean.

Therefore, while the present inventors were studying various physiological activities of the soybean germ extract, the buckwheat sprout extract has the effect of strengthening hair roots, promoting hair growth or preventing hair loss, so a pharmaceutical composition, cosmetic composition or health for hair growth or hair loss prevention The present invention was completed by confirming that it can be used as a functional food.

Republic of Korea Patent Registration No. 10-0800359 (composition showing hair loss prevention and hair growth promoting effect) Republic of Korea Patent Registration No. 10-0773457 (composition having the effect of promoting hair growth and method of using the same) Republic of Korea Patent No. 10-1908679 (Fermented composition for preventing hair loss and promoting hair growth containing natural extract) Korean Patent Registration No. 10-1907902 (Liposome composition containing peptide)

It is an object of the present invention to provide a composition for strengthening hair roots, promoting hair growth, or preventing stress-related hair loss containing soybean germ extract. The soybean embryo extract is characterized in that it promotes the expression of growth factors in hair root-related cells, and the growth factors include insulin-like growth factor (IGF) or vascular endothelial growth factor; VEGF) and the like.

The present invention relates to a cosmetic composition for strengthening hair roots, promoting hair growth, or preventing hair loss containing soybean germ extract.

The soybean germ extract may be a soybean germ extracted with water, C2-C4 alcohol or a mixed solvent thereof. The solvent may be 20-40% (w/w) ethanol aqueous solution.

The soybean germ extract can be used by dissolving it in 40-60% (w/w) 1,3-butylene glycol aqueous solution.

That is, the soybean germ extract may be obtained by dissolving or diluting the soybean germ in an aqueous solution of 1,3-butylene glycol after extracting and concentrating the soybean embryo with an aqueous solution of 20 to 40% by weight of ethanol.

The soybean germ extract dissolved in 1,3-butylene glycol aqueous solution may be filtered through a filter having pores of 0.45㎛ or less. Preferably, the filter may be a filter having pores of 0.20 to 0.45 μm.

In addition, the soybean germ extract dissolved in 1,3-butylene glycol aqueous solution contains 900 to 1200 ppm of daizgene compound, and the weight ratio of daidgene to genistein is 100:0.3 to 100:0.9, characterized in that do. In addition, it is characterized in that the total concentration of daidzein, glycitin, genistine, daidzein, glycitein, and genistein is about 2000-2600ppm.

The soybean embryo extract can be prepared in the form of liposomes, and the liposomes are polysorbate 80 (Polysorbate 80), hydrogenated lecithin, lysolecithin (Lysolecithin), MCT (Medium Chain Triglyceride) oil soybean germ As prepared by mixing with the extract, polysorbate 80 0.5-2 wt%, hydrogenated lecithin 4-6 wt%, risolecithin 0.3-0.7 wt%, MCT oil 3-5 wt% soybean germ extract (1 ,3-Butylene glycol (dissolved in aqueous solution) is mixed with 1 to 5% by weight, and may contain an additional 85 to 90% by weight of water. When the liposome raw material such as polysorbate 80 is out of the above range, the liposome formation is not well or even if the liposome is formed, the phase separation occurs over time, which is not suitable for use as a cosmetic.

The soybean embryo can be used as long as it is removed from soybean types, preferably used as a raw material for soymilk, and more preferably soybean or seoritae embryo can be used.

The liposome may be formulated into various cosmetic compositions for hair growth, hair loss improvement, scalp improvement, and the like. Preferably, it can be prepared into hair soap, hair shampoo, hair conditioner, hair treatment, hair lotion, hair mousse, hair spray, hair gel, hair pack, hair ampoule, hair oil, and the like.

MCT as used herein means medium chain (6-12 carbons) aliphatic fatty acid esters of glycerol. In at least one embodiment the MCT oil mainly comprises C8-C10 triglycerides, such as capric and caprylic triglycerides. Thus, as an example, MCT oil may be derived from capric acid and caprylic acid. More preferably, the MCT oil may contain 50 to 80% caprylic acid and 20 to 50% capric acid. More preferably the MCT may comprise about 56% caprylic acid and about 44% capric acid, or about 50-60% caprylic acid and about 40-50% capric acid. In the present invention, MCT is a liquid at room temperature.

In another aspect, the present invention relates to a pharmaceutical composition for strengthening hair roots, promoting hair growth or preventing hair loss.

The manufacturing temperature of the soybean germ extract may be 20 to 100 ℃, but is not limited thereto. The extraction time is not particularly limited, but extraction is preferably performed within 10 minutes to 2 days, and as an extraction device, a conventional extraction device, an ultrasonic crushing extractor or a fractionator may be used. Soybean germ extract prepared in this way can be removed by hot air drying, drying under reduced pressure or freeze-drying. In addition, the soybean germ extract can be prepared in various forms by dissolving it in various solvents such as 1,3-butylene glycol.

The soybean germ extract can be formulated as the extract itself or liposomes, and may include all of the ingredients generally used for hair or scalp products. For example, it may include general auxiliary ingredients such as emulsifiers, thickeners, emulsifiers, surfactants, lubricants, alcohols, water-soluble polymers, gelling agents, stabilizers, vitamins, inorganic salts, emulsifiers, and fragrances.

More specifically, when the formulation of the cosmetic composition of the present invention is a paste, cream or gel, animal fiber, vegetable fiber, wax, paraffin, starch, tragacanth, cellulose derivative, polyethylene glycol, silicone, bentonite, silica as a carrier component , talc or zinc oxide may be used. When the formulation of the cosmetic composition of the present invention is a powder or a spray, lactose, talc, silica, aluminum hydroxide, calcium silicate or polyamide powder may be used as a carrier component, and in particular, in the case of a spray, additional chlorofluorohydro It may contain a propellant such as carbon, propane-butane or dimethyl ether. When the formulation of the cosmetic composition of the present invention is a solution or emulsion, a solvent, solvating agent or emulsifying agent is used as a carrier component, for example, water, ethanol, isopropanol, ethyl carbonate, ethyl acetate, benzyl alcohol, benzyl benzoate, propylene fatty acid esters of glycol, 1,3-butylglycol oil, glycerol fatty esters, polyethylene glycol or sorbitan. When the formulation of the cosmetic composition of the present invention is a suspension, as a carrier component, a liquid diluent such as water, ethanol or propylene glycol, a suspending agent such as ethoxylated isostearyl alcohol, polyoxyethylene sorbitol ester and polyoxyethylene sorbitan ester; Microcrystalline cellulose, aluminum metahydroxide, bentonite, agar, or tracanth may be used. When the formulation of the cosmetic composition of the present invention is surfactant-containing cleansing, aliphatic alcohol sulfate, aliphatic alcohol ether sulfate, sulfosuccinic acid monoester, acetionate, imidazolinium derivative, methyl taurate, sarcosinate as carrier components. , fatty acid amide ether sulfate, alkylamidobetaine, fatty alcohol, fatty acid glyceride, fatty acid diethanolamide, vegetable oil, linoline derivative or ethoxylated glycerol fatty acid ester, etc. may be used. The cosmetic composition of the present invention may further contain excipients including fluorescent substances, fungicides, hydrophobicity-inducing substances, moisturizers, fragrances, fragrance carriers, proteins, solubilizers, sugar derivatives, sunscreens, vitamins, plant extracts, and the like. . According to the formulation or purpose of use, the amount of the ingredients may be selected within a range that does not impair the intrinsic effect of the cosmetic composition. The amount of the components added may be, for example, 0.1 to 10% by weight, preferably 0.1 to 6% by weight, based on the total weight of the composition, but is not limited thereto.

In addition, the present invention provides a pharmaceutical composition for strengthening hair roots, promoting hair growth or preventing hair loss, comprising a soybean germ extract or a liposome containing the same.

The pharmaceutical composition may be formulated and used as an external preparation. Carriers, excipients and diluents that may be included in the pharmaceutical composition include lactose, dextrose, sucrose, sorbitol, mannitol, xylitol, erythritol, maltitol, starch, acacia gum, alginate, gelatin, calcium phosphate, calcium silicate, cellulose , methyl cellulose, microcrystalline cellulose, polyvinyl pyrrolidone, water, methylhydroxybenzoate, propylhydroxybenzoate, talc, magnesium stearate and mineral oil. In the case of formulation, it is prepared using diluents or excipients, such as commonly used fillers, extenders, binders, wetting agents, disintegrants, and surfactants.

The dosage of the pharmaceutical composition of the present invention will vary depending on the age, sex, and weight of the subject to be treated, the specific disease or pathological condition to be treated, the severity of the disease or pathological condition, the route of administration, and the judgment of the prescriber. Dosage determination based on these factors is within the level of the skilled artisan, and dosages generally range from 0.01 mg/kg/day to approximately 2000 mg/kg/day. A more preferred dosage is 1 mg/kg/day to 500 mg/kg/day. Administration may be administered once a day, or may be administered in several divided doses. The above dosage does not limit the scope of the present invention in any way.

The pharmaceutical composition of the present invention may be administered to mammals such as mice, livestock, and humans by various routes. Any mode of administration can be anticipated, and in the present invention, it is most preferred to apply through skin application.

The present invention relates to a composition for strengthening hair roots, preventing hair growth or stress-related hair loss, comprising bean germ extract as an active ingredient. The soybean embryo extract of the present invention increases the expression of the VEGF gene, known as a factor related to hair root strengthening, and the IFG-1 gene, which is known as a factor related to hair growth, and functions to alleviate stress-related hair loss.

1 shows the activity of increasing VEGF mRNA expression of soybean embryo extract in a general hair growth model.
Figure 2 shows the activity of increasing the expression of IGF-1 mRNA of the soybean embryo extract in a general hair growth model.
3 shows the expression inhibitory activity of TGF-β2 mRNA of soybean embryo extract in a model of stress hair loss.
Figure 4 shows the IL-6 mRNA expression increasing activity of the soybean embryo extract in the stress hair loss model.
Figure 5 shows a projection transfer microscope (TEM) image of the nanoliposome containing the soybean embryo extract.

Hereinafter, preferred embodiments of the present invention will be described in detail. However, the present invention is not limited to the embodiments described herein and may be embodied in other forms. Rather, it is provided so that this disclosure will be thorough and complete, and will fully convey the spirit of the invention to those skilled in the art.

<Preparation Examples 1 to 7. Preparation of soybean germ extract>

In order to remove the astringent taste during the production of soymilk from Jeongfood Co., Ltd., embryos separated from soybeans were purchased. Purchased soybean embryos were divided into 7 groups: water, ethanol aqueous solution for each concentration, and ethanol solvent alone (purity 99.9%; hereinafter, abbreviated as 100%) by separating the extraction solvent, and as shown in Table 1, It was prepared as an extract.

To this end, 10 kg of each solvent was added to 1 kg of soybean germ, extracted by heating at 60° C. for 8 hours, cooled to room temperature (25° C.), and filtered through 400 mesh filter cloth to obtain only the liquid phase. After that, the liquid was filtered again with a 0.45 μm filter, and the extract obtained was concentrated in a vacuum and dried, and finally, the dried extract for each experiment was added to a final concentration of 50% (w/w) 1,3-butylene glycol aqueous solution. was dissolved to 2% by weight.

On the other hand, the isoflavone non-glycol material, which is not readily soluble in 1,3-butylene glycol, is agglomerated, precipitated, etc. after the addition of the 50% (w/w) 1,3-butylene glycol aqueous solution and filtered. Most of them are removed in the process. Aggregation, precipitation, and filtration remove substances appear from when the alcohol concentration of the extraction solvent is 70% (w/w) or more in the initial extraction process.

experimental group Preparation Example 1 Soybean germ water extract Preparation 2 30% (w/w) ethanol aqueous solution extract of soybean germ Preparation 3 50% (w/w) ethanol aqueous solution extract of soybean germ Preparation 4 70% (w/w) ethanol aqueous solution extract of soybean germ Preparation 5 100% ethanol extract of soybean germ Preparation 6 20% (w/w) ethanol aqueous solution extract of soybean germ Preparation 7 40% (w/w) ethanol aqueous solution extract of soybean germ

For each extract prepared in Table 1, the active ingredients were confirmed as shown in Table 3 by the following quantitative method.

quantitative method

Weigh about 500mg of each sample precisely, put it in a 20mL volumetric flask, add 15mL of 100% MeOH to dissolve it, adjust it to the mark, and use it as the sample solution. Separately, purchase a standard from Sigma-Aldrich, weigh about 5 mg precisely, put it in a 200 mL volumetric flask, add 150 mL of 100% MeOH to dissolve it, adjust to the mark, and use it as the standard solution. 10 μl of each of the test solution and the standard solution was tested according to the liquid chromatography method under the following operating conditions to confirm the amount of each.

operating conditions

Detector: UV absorbance spectrometer (measurement wavelength: 260 nm, soysaponin)

Column: Capcellpak C ₁₈ 4.6 × 250mm

Column temperature: 35 ℃

Injection volume: 10 μL

Flow rate: 1.0 mL/min

Time: 60 minutes

This statue: described in Table 2 below

Time(min) A Pump
(0.05% Formic acid in Water) B Pump
(100% Acetonitrile) 0.01 90% 10% 10 90% 10% 35 60% 40% 45 60% 40% 50 90% 10% 60 90% 10%

extract Isoflavone concentration (ppm) in each extract dissolved in 50% (w/w) 1,3-butylene glycol aqueous solution Daidzin Glycitin Genistin Daidzein Glycitein Genistein soysaponin total Preparation Example 1 552 444 264 46 34 0 0 1340 Preparation 2 1043 839 419 53 37 7 0 2398 Preparation 3 1324 923 408 63 48 14 215 2995 Preparation 4 327 221 133 168 52 21 830 1752 Preparation 5 40 33 49 252 85 26 1200 1685 Preparation 6 987 819 387 55 36 4 0 2288 Preparation 7 1063 823 438 51 41 9 0 2425

Looking at Table 3, the water extract does not contain genistein and soisaponin among isoflavone compounds. In addition, it can be confirmed that the higher the alcohol concentration in the solvent, the higher the content of soisaponin.

Characteristically, when the alcohol content in the solvent is 20-40% (w/w) (extracts of Preparation Example 2, Preparation Example 6 and Preparation Example 7) daidjin, glycitin, genistine, daidzein, glycitein , it can be seen that the total concentration of genistein is about 2300-2450 ppm, and soy saponin is not included.

In addition, as a result of confirming the results of Table 2 again several times, the extracts prepared under the conditions of Preparation Examples 2, 6 and 7 had a total concentration of isoflavone compounds from daidzine to genistein of about 2000-2600ppm (μg). /ml) can be confirmed, and soy saponin was not repeatedly confirmed.

Next, the weight distribution ratio of each compound shown in Table 3 is shown in Table 4 below when daidzine is 100.

extract Dissolved in 50% (w/w) 1,3-butylene glycol aqueous solution
Weight distribution ratio of isoflavone compounds in each extract (Standard: Daidzin 100) Daidzin Glycitin Genistin Daidzein Glycitein Genistein soysaponin Preparation Example 1 100 80.43 47.83 8.33 6.16 0.00 0.00 Preparation 2 100 80.44 40.17 5.08 3.55 0.67 0.00 Preparation 3 100 69.71 30.82 4.76 3.63 1.06 16.24 Preparation 4 100 67.58 40.67 51.38 15.90 6.42 253.82 Preparation 5 100 82.50 122.50 630.00 212.50 65.00 3000.00 Preparation 6 100 82.98 39.21 5.57 3.65 0.41 0.00 Preparation 7 100 77.42 41.20 4.80 3.86 0.85 0.00

Through the results of Table 4 and repeated experiments, in the extracts (Extracts of Preparation Example 2, Preparation Example 6 and Preparation Example 7) prepared to have an alcohol content of 20-40% (w/w) in the solvent, Daidzin: The weight distribution of Glycitin is [100:70 to 100:85], Daidzin:Genistin is [100:35 to 100:45], Daidzin:Daidzein is [100:3 to 100:10], Daidzin:Glycitein is [100: 2 to 100:10], and it can be confirmed that Daidzin:Genistein is [100:0.3 to 100:0.9].

[Experimental Example 1] Cell culture

DPCs (Human Dermal Papilla cells) and HHGMC (Human Hair Germinal Matrix Cells: hair cells) cells were used, and the DPCs cell line was 10% (v/v) Fetal Bovine Serum (FBS), 1% (v/v) DMEM (Dulbecco Modified Eagle Medium) medium containing anti-biotics was used, and the HHGMC cell line was 5% (v/v) Fetal Bovine Serum (FBS), 1 MSCM with % (v/v) penicillin/streptomycin solution (S/P), 1% (v/v) mesenchymal stem cell growth supplement (MSCGS) ( Mesenchymal Stem Cell Medium) was cultured at 37° C., 5% CO _{2 conditions.}

[Experimental Example 2] Investigation of VEGF expression level of hair root strengthening gene

In order to evaluate the effect of strengthening the hair root, a VEGF (Vascullar Endotherial Growth Factor) gene expression experiment was performed. Inoculate 2 ml of DMEM medium containing 10% (v/v) bovine serum and dermal papilla cells (DPCs, cell bio) at 2.5 × 10 ⁵ cells/well in a 6 well multi plate (corning) and incubate for 24 hours, each The wells were replaced with serum-free medium. Samples were treated in serum-free medium and then cultured for 24 hours. At this time, each sample was treated so that the liquid sample dissolved in 1,3-butylene glycol solution became 3% (w/w) in the medium, and the total concentration of Minoxidil used to evaluate the efficacy was 1 μg/ml in the medium. treated to become

Next, after washing with PBS, the cultured cells were treated with QIAzol ^® (QIAGENL ^® , USA), and after cell lysis, RNA was isolated using the protocol provided by the ^{manufacturer (QIAGENL ® ).} After the isolated RNA was ^{quantified using the Qubit ®} fluoremeter with RNA BR Assay kit, cDNA was synthesized and real-time PCR was performed. For cDNA synthesis, a cDNA synthesis kit (qPCRBIO cDNA Synthesis Kit, PCRBIOSYSTEMS, London, UK) was used, and the experiment was performed according to the kit's method. Real-time PCR amplified the gene using Real-time PCR Kit (2x qPCRBIO SyGreen Blue mix Lo-ROX, PCRBIOSYSTEMS, London, UK) and then quantitatively analyzed the amplified product.

VEGF and β-actin primers used for PCR were synthesized and used by Cosmogenetec (Korea), and the primer sequences are summarized in <Table 5>.

Primer Sense Antisense VEGF 5'-TGC AGA TTA TGC GGA TCA AAC C-3' 5'-TGC ATT CAC ATT TGT TGT GCT GTA G-3' β-actin 5'-GGC ACC CAG CAC AAT GAA G-3' 5'-CCG ATC CAC ACG GAG TAC TTG-3'

The cDNA synthesis reaction conditions were 5 seconds at 95°C, 25 seconds at 60°C, and 40 cycles for both VEGF and β-actin. Control was not treated with samples, and Minoxidil used as a positive control is known to be a substance that strengthens hair roots and induces hair growth.

The results of gene expression levels are shown in <Table 6> and <Figure 1>.

test group VEGF mRNA expression rate (%) Stdev. Non-irradiated group (Control) 100.00 0.00 Minoxidil (1㎍) 134.85 5.28 Preparation Example 1 (3%) 118.59 2.39 Preparation Example 2 (3%) 139.69 8.72 Preparation Example 3 (3%) 120.58 8.65 Preparation Example 4 (3%) 123.96 7.76 Preparation Example 5 (3%) 113.15 3.27 Preparation 6 (3%) 142.5 2.34 Preparation 7 (3%) 139.2 1.04

As a result, the VEGF gene expression was the best in Preparation Example 2, Preparation Example 6 and Preparation Example 7 (20-40% (w/w) ethanol aqueous solution extract treatment group of soybean embryos) compared to the control group.

VEGF induces the formation of blood vessels in the hair follicle, supplies nutrients to the hair growth phase through the formation of blood vessels around the hair follicle, and induces cell proliferation and migration.

[Experimental Example 3] IGF-1 expression level investigation of hair growth gene

To evaluate the hair growth efficacy, IGF-1 (Insulin-like growth factor 1) gene expression experiment was performed. The experimental method was the same as in Experimental Example 2, and the IGF-1 and β-actin primers used for PCR were synthesized and used by Cosmogenetec (Korea), and the primer sequence is shown in <Table 7>. Minoxidil, used as a positive control, is known to be a substance that strengthens hair roots and induces hair growth. The results of gene expression levels are shown in <Table 8> and <Figure 2>.

Primer Sense Antisense IGF-1 5'-AGC CTG TCC ACC CTT GAG AA-3' 5'-CCC TGG AGC CAC AGA GCA T-3' β-actin 5'-GGC ACC CAG CAC AAT GAA G-3' 5'-CCG ATC CAC ACG GAG TAC TTG-3'

test group IGF-1 mRNA expression rate (%) Stdev. Non-irradiated group (Control) 100.00 0.00 Minoxidil (1 μg/ml) 320.95 6.61 Preparation Example 1 (3%) 154.50 7.63 Preparation Example 2 (3%) 238.69 5.97 Preparation Example 3 (3%) 140.24 8.58 Preparation Example 4 (3%) 153.64 7.02 Preparation Example 5 (3%) 140.86 5.56 Preparation 6 (3%) 225.23 2.31 Preparation 7 (3%) 249.24 3.82

As a result of the analysis, the expression of IGF-1, a hair growth factor, was increased in each sample compared to Control (comparative group), but Preparation Example 2, Preparation Example 6, and Preparation Example 7 samples (20-40% (w/w) of soybean embryos) It can be seen that the ethanol aqueous solution extract) exhibits the best efficacy.

[Experimental Example 4] Investigation of the expression levels of TGF-β2 and IL-6 inducing genes for stress-related hair loss

In order to evaluate the efficacy of stress-related hair loss, MSCM medium containing 5% (v/v) bovine serum and hair cells (Human Hair Germinal Matrix Cell, HHGMC, ScienCell) ^{were inoculated at 2.5×10 5} cells/well by 2 ml each. After incubation for 24 hours, each well is replaced with a serum-free medium. Substance P acetate salt hydrate (Sigma aldrich, S6883), which is secreted when stress occurs after changing the medium, was treated with the sample and incubated for 24 hours. After that, the experiment was carried out in the same manner as in Experimental Example 2 from RNA extraction, and TGF-β2, IL-6, and β-actin primers used in PCR were synthesized and used by Cosmogenetec (Korea), and the primer sequence is shown in Table 9>.

Primer Sense Antisense TGF-β2 5'-CAC CAT AAA GAC AGG AAC CTG-3' 5'-GGA GGT GCC ATC AAT ACC TGC-3' IL-6 5'-CCA GGA GCC CAG CTA TGA AC-3' 5'-CCC AGG GAG AAG GCA ACT G-3' β-actin 5'-GGC ACC CAG CAC AAT GAA G-3' 5'-CCG ATC CAC ACG GAG TAC TTG-3'

The cDNA synthesis reaction conditions were 5 seconds at 95°C, 25 seconds at 60°C, and 40 cycles of TGF-β2, IL-6, and β-actin. Control was treated with purified water used as a dissolution solvent, and Minoxidil used as a positive control is known to be a substance that strengthens hair roots and induces hair growth and inhibits hair loss. The results of expression levels are shown in <Table 10>, <Table 11>, and <Fig. 3> and <Fig. 4>.

test group TGF-β2 mRNA expression rate (%) Stdev. Non-irradiated group (Control) 64.66 5.16 Substance P (0.1 nM) 100.00 0.00 Substance P (0.1 nM) + Minoxidil (1 μg/ml) 54.55 2.97 Substance P (0.1nM) + Preparation Example 1 (3%) 74.74 3.81 Substance P (0.1nM) + Preparation Example 2 (3%) 64.93 7.12 Substance P (0.1nM) + Preparation Example 3 (3%) 82.18 1.17 Substance P (0.1nM) + Preparation Example 4 (3%) 78.21 1.70 Substance P (0.1nM) + Preparation Example 5 (3%) 74.10 3.44 Substance P (0.1nM) + Preparation Example 6 (3%) 59.23 2.42 Substance P (0.1nM) + Preparation Example 7 (3%) 62.31 3.21

test group IL-6 mRNA expression rate (%) Stdev. Non-irradiated group (Control) 46.41 2.31 Substance P (0.1 nM) 100.00 0.00 Substance P (0.1 nM) + Minoxidil (1 μg/ml) 44.52 2.38 Substance P (0.1nM) + Preparation Example 1 (3%) 43.24 3.78 Substance P (0.1nM) + Preparation Example 2 (3%) 18.84 1.88 Substance P (0.1nM) + Preparation Example 3 (3%) 51.23 7.77 Substance P (0.1nM) + Preparation Example 4 (3%) 45.90 2.20 Substance P (0.1nM) + Preparation Example 5 (3%) 56.35 2.13 Substance P (0.1nM) + Preparation Example 6 (3%) 15.23 1.41 Substance P (0.1nM) + Preparation Example 7 (3%) 20.23 0.98

As a result, expression of cytokine IL-6 inhibiting the expression of TGF-β2 involved in hair degeneration and inhibiting hair extension in 20-40% (w/w) ethanol aqueous solution extract of soybean embryos even under stress hair loss conditions It can be confirmed that this is the greatest decrease and the extract has the effect of alleviating hair loss caused by stress.

<Experimental Example 5. Experiment on the effect of promoting percutaneous absorption of soybean germ extract>

This time, liposomes, which are lipid vesicles, were prepared using hydrogenated lecithin, polysorbate 80, lyzolecithin, and the like, and compared to the conventional emulsion, the efficacy of the liposome composition, which is stable and can increase skin penetration, was compared.

To this end, liposomes and emulsions containing Daidzin were prepared in comparison with the composition shown in <Table 12> below. As MCT oil, Waglinol MCT was used.

To prepare liposomes, each component is heated to 80° C., completely dissolved, and mixed at 1,500 to 2,000 rpm for 5 minutes using a Homo-Mixer (PRIMIX Mark-II, Japan), followed by a pressure of 1200 bar and a flow rate of 500 m/s. It was passed three times through a high pressure emulsifier (Microfluidizer M210EH, Microfluidies, USA) under a high pressure to obtain liposomes.

To prepare an emulsion, each component was heated to 80° C., completely dissolved, and mixed at 1,500 to 2,000 rpm for 5 minutes using a Homo-Mixer (PRIMIX Mark-II, Japan), followed by cooling to room temperature to obtain an emulsion. On the other hand, in the emulsion, liposomes were not formed even though a high-pressure emulsifier was used as a liposome preparation method by mixing each raw material component under the same conditions.

Liposome (g) emulsion (g) Polysorbate 80 1.0 0.5 Glyceryl Monostearate - 1.0 PEG-100 Stearate & Glyceryl Stearate - 1.5 Hydrogenated Lecithin 3.0 - lyzolecithin 0.5 - MCT Oil 5.0 5.0 Soybean germ extract of Preparation Example 2 2.0 2.0 Purified water To 100 To 100

In order to compare the skin absorption effects of the prepared liposome and lotion, artificial skin Neoderm (Tego Science) was installed in a Franz-type diffusion cell (Lab fine instruments, Korea) and tested.

To this end, 50 mM phosphate buffer (pH 7.4, 0.1 M NaCl) was put into the receptor container (5 ㎖) of the Franz-type diffusion cell, and then the diffusion cell was mixed and dispersed at 32 ° C., 600 rpm. 50 μl of each of the liposomes and the emulsion were put into the donor container. Absorption and diffusion were given according to a predetermined time, and the skin where absorption and diffusion occurred was 0.64 cm ² . After absorption and diffusion of the active ingredient, Daidzin, is finished, wash the emulsion remaining on the skin without being absorbed with 10 ml of ethanol. Daidzin was extracted using 4 ml of dichloromethane. Then, the dichloromethane extract was filtered through a 0.45 μm nylon membrane filtration membrane, and the content of Daidzin as an indicator material was measured.

Transdermal absorption (㎍) Percutaneous absorption increase rate (%) Soybean germ extract liposome 0.4187 52.31 Soybean Germ Extract Emulsion 0.1426 -

As a result, as can be seen in <Table 13>, it can be confirmed that the active ingredient can reach the skin more efficiently when the soybean germ extract is prepared as a liposome.

On the other hand, various liposomes were prepared under conditions in which the content of MCT oil included in the liposome and the emulsion was changed to 1, 3, 5, 7, and 9 g each, and the MCT oil was used to form liposomes between 3 g and 5 g, Other conditions were confirmed that the liposome formation was not well.

Claims (8)

Soybean germ extract, polysorbate 80 (Polysorbate 80), hydrogenated lecithin (Hydrogenated lecithin), lysolecithin (Lysolecithin) and MCT (Medium Chain Triglyceride) oil, characterized in that prepared in the form of liposomes, anti-hair loss cosmetic composition. According to claim 1,
The soybean germ extract is a cosmetic composition for preventing hair loss, characterized in that the soybean germ is extracted with water, C2-C4 alcohol or a mixed solvent thereof. 3. The method of claim 2,
The soybean germ extract is a cosmetic composition for preventing hair loss, characterized in that it is dissolved in a 1,3-butylene glycol aqueous solution after extraction with the solvent. 4. The method of claim 3,
A cosmetic composition for preventing hair loss, characterized in that the soybean germ extract dissolved in 1,3-butylene glycol aqueous solution is filtered through a filter having pores of 0.45 μm or less. 5. The method of claim 4,
The soybean germ extract contains 900 to 1200 ppm of the daizgene compound, and the weight ratio of daidgene: genistein is 100:0.3 to 100:0.9. A cosmetic composition for preventing hair loss. For hair root strengthening, characterized in that it is prepared in the form of a liposome, including soybean germ extract, polysorbate 80, hydrogenated lecithin, lysolecithin and MCT (Medium Chain Triglyceride) oil cosmetic composition. Soybean germ extract, polysorbate 80 (Polysorbate 80), hydrogenated lecithin (Hydrogenated lecithin), lysolecithin (Lysolecithin) and MCT (Medium Chain Triglyceride) oil for preventing hair loss, characterized in that it is prepared in the form of liposomes A pharmaceutical composition for external use. Promotes hair growth, characterized in that it is prepared in liposome form, including soybean germ extract, polysorbate 80, hydrogenated lecithin, lysolecithin and MCT (Medium Chain Triglyceride) oil cosmetic composition for

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