KR20230026758A - Bacillus siamensis HDB8077 strain isolated from Dendrobium candidum flower and haircare cosmetic composition contained culture of Dendrobium candidum flower using thereof - Google Patents

Abstract

The present invention relates to a Bacillus siamensis HDB8077 strain (accession number: KCCM12989P) isolated from Dendrobium candidum Wall. ex Lindley flowers and a hair care cosmetic composition containing Dendrobium candidum Wall. ex Lindley flower culture. The culture is cultured by adding Dendrobium candidum Wall. ex Lindley flower powder to the culture medium of Bacillus siamensis HDB8077 strain, and has hair root strengthening effect, hair growth effect, androgenic alopecia alleviation effect, and enhanced expression of ceramide biosynthesis enzyme. Accordingly, the culture can be easily used as a multi-functional hair composition.

Description

Bacillus siamensis HDB8077 strain isolated from Dendrobium candidum flower and haircare cosmetic composition contained culture of Dendrobium candidum flower using its}

The present invention relates to a hair care cosmetic composition containing a strain of Bacillus siamensis HDB8077 (Accession No. KCCM12989P) isolated from a flower of sagebrush and a culture of sageflower of the strain.

Human hair is very important because it not only plays a primary role in protecting the skin and scalp, but also plays a unique role in social and sexual communication.

The hair cycle consists of an anagen phase, a catagen phase, and a telogen phase, and hair is maintained by repeating hair growth and hair loss while repeating the cycle. The growth period of hair is 3-5 years for men and 4-6 years for women, the degeneration period is 30-45 days, and the resting period is about 3-4 months. At the final stage of the telogen phase, the hair growth period in which new hair is naturally generated begins and the hair is maintained. Since hair loss is a natural phenomenon, alopecia, which is recognized as a hair loss disease, refers to a condition in which hair appears to be small due to relatively large amounts of hair in a dormant state.

The cause of hair loss is not yet known exactly, but it is a major factor affecting hair growth, suppression of proliferation or functional decline of dermal papilla related to hair cycle control, abnormality of hair cycle by male hormone action, scalp Abnormal changes in the hair cycle due to a decrease in blood flow to the hair follicle, anti-hypertensive drugs, mental stress, physical stimulation, and environmental pollution have been discussed.

In the past, hair loss was common among middle-aged and elderly people, but recently, hair and scalp damage caused by mousse, perm, dyeing, dry heat, increase in social and environmental stress, and changes in food due to westernization of diet have been the causes. Hair loss is notably occurring in young people in their 30s, and female pattern hair loss as well as male pattern hair loss is increasing.

For this reason, many treatments for preventing hair loss are on the market, but 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 application 'minoxidil' or the oral drug 'Propecia' (finasteroid ingredient) in the early stage of hair loss, but for already advanced hair loss, it is known to be restored to normal. There are difficulties, and side effects such as hair growth in unwanted areas, erectile dysfunction, decreased sexual function, dizziness, headache, swelling, skin rash, etc. may occur. There are problems that cannot be.

Among them, 'Minoxidil' is a drug approved by the US FDA. Although the exact treatment mechanism is not yet clear, dermal papilla cells (DPCs) are reported to affect the hair cycle through the secretion of proteins such as various growth factors. ) It is known to have the effect of proliferating the growth of dermal papilla cells (DPCs) through the 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, to prevent androgenetic hair loss from progressing. known to be known, measuring the effect of inhibiting the activity of 5-alpha reductase or the growth and proliferation effect of dermal papilla cells (DPCs) on a substance is how effectively the substance prevents hair loss and contributes to hair growth. It can be used as an indicative indicator.

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

Recently, various research institutes are actively conducting research on many regulators involved in hair growth and hair loss mechanisms. are reported continuously. 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 act decisively on the proliferation or differentiation of hair follicle stem cells in the bulge region.

On the other hand, the number of hair loss continues to increase due to environmental stress in society along with side effects caused by environmental pollution and abuse of drugs, etc., and hair loss is becoming more frequent even in the younger generation, and the change in social trend that places importance on appearance Due to this, the development of new agents for preventing hair loss is on the rise.

In addition, among currently developed anti-hair loss preparations, especially in the case of synthetic compound preparations, there is a problem of causing various side effects on the skin, and recently, attempts have been made to obtain a preparation having an anti-hair loss effect from nature.

Accordingly, the present inventors found that the Bacillus siamensis HDB8077 strain isolated from the flowers of spp. spp. and the culture of its spp. The present invention was completed by confirming that it is available as.

Republic of Korea Patent Registration No. 10-2268243 (Name of invention: Composition showing hair loss prevention and hair growth stimulating effect, Applicant: JS Trading Co., Ltd., registration date: June 17, 2021) Republic of Korea Patent Registration No. 10-1880057 (Title of Invention: Hair Care Cosmetic Composition for Preventing Hair Loss Containing Chojeong Carbonated Water, Applicant: Fanipin Korea Co., Ltd., Registration Date: July 13, 2018) Republic of Korea Patent Publication No. 10-2019-0026175 (Title of Invention: Cosmetic Composition Containing Chrysanthemum Flower Extract, Applicant: LG Household & Health Care Co., Ltd., Date of Publication: March 13, 2019) Republic of Korea Patent Publication No. 10-2019-0094642 (Title of Invention: Cosmetic Composition Containing Fermented Iron Piseoggok and Manufacturing Method Thereof, Applicant: Rabio Co., Ltd., Publication Date: August 14, 2019) Republic of Korea Patent Publication No. 10-2006-0008662 (Title of Invention: Hair Growth Promoter Composition, Applicant: LG Household & Health Care, Publication Date: January 27, 2006) Republic of Korea Patent Publication No. 10-2021-0002005 (Title of invention: Composition for strengthening hair roots, promoting hair growth or preventing hair loss containing buckwheat sprout extract, Applicant: Hyundai Bioland Co., Ltd., publication date: January 06, 2021) Republic of Korea Patent Registration No. 10-1017709 (Title of Invention: A cosmetic composition containing extracts of complex herbal medicines such as ginseng to prevent hair loss, inhibit dandruff and promote hair growth, Applicant: Hyundai Bioland Co., Ltd., registration date: Feb. 2011 18th of the month) Republic of Korea Patent Registration No. 10-1626319 (Title of Invention: Composition for promoting hair growth or preventing hair loss containing fermented camellia oil or its extract and manufacturing method thereof, Applicant: Hyundai Bioland Co., Ltd., Registration date: May 26, 2016 Day)

An object of the present invention is to provide a hair care cosmetic composition containing a Bacillus siamensis HDB8077 strain (Accession No. KCCM12989P) isolated from a stylized spp.

The present invention relates to a strain of Bacillus siamensis HDB8077 (Accession No. KCCM12989P) isolated from the flowers of Stylophyte.

The strain is characterized in that it enhances the hair care function of stylized rhizomatous flowers. More preferably, the strain is characterized in that it enhances the hair root strengthening, hair growth promotion or hair loss prevention efficacy of stylized spp.

In addition, the present invention relates to a hair care cosmetic composition containing a flower culture of the above strain.

The culture is incubated for 20 to 48 hours at 30 to 40 ° C. after adding stylized calico flower powder to the culture medium of the Bacillus siamensis HDB8077 strain.

The hair care cosmetic composition is characterized in that it has an effect of strengthening hair roots, promoting hair growth or preventing hair loss.

The stylized staphylococcus flower culture may be filtered through a filter having pores of 0.45 μm or less. Preferably, the filter may be a filter having pores of 0.20 to 0.45 μm.

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

Water can be removed by hot-air drying, reduced pressure drying or freeze-drying of the flower culture of the Bacillus siamensis HDB8077 strain. In addition, the fermented sage flower culture can be dissolved in various solvents such as water, 1,3-butylene glycol, or a mixture thereof to prepare various types of formulations.

Prior to culturing stylized rhizomatous flowers, the culture conditions of the Bacillus siamensis HDB8077 strain of the present invention for preparing a pre-culture solution are preferably 60 to 72 hours under conditions of 100 to 200 rpm and 30 to 40 ° C. Bacillus siamensis HDB8077 strain can be used by liquid culture in tryptic soy broth (TSB) after solid culture in tryptic soy agar (TSA).

The stylized spp. flower culture may include all of the components generally used in hair or scalp products. For example, general auxiliary ingredients such as emulsifiers, thickeners, emulsions, surfactants, lubricants, alcohols, water-soluble polymers, gelling agents, stabilizers, vitamins, inorganic salts, emulsifiers, and fragrances may be included.

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 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 propellants 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, such as water, ethanol, isopropanol, ethyl carbonate, ethyl acetate, benzyl alcohol, benzyl benzoate, propylene fatty acid esters of glycol, 1,3-butylglycol oil, glycerol aliphatic esters, polyethylene glycol or sorbitan. When the formulation of the cosmetic composition of the present invention is a suspension, a liquid diluent such as water, ethanol or propylene glycol, an ethoxylated isostearyl alcohol, a suspending agent such as polyoxyethylene sorbitol ester and polyoxyethylene sorbitan ester, Microcrystalline cellulose, aluminum metahydroxide, bentonite, agar or tracanth and the like may be used. When the formulation of the cosmetic composition of the present invention is surfactant-containing cleansing, as carrier components, aliphatic alcohol sulfate, aliphatic alcohol ether sulfate, sulfosuccinic acid monoester, acethionate, imidazolinium derivative, methyl taurate, and sarcosinate , fatty acid amide ether sulfates, alkylamidobetaines, fatty alcohols, fatty acid glycerides, fatty acid diethanolamides, vegetable oils, linolin derivatives, or ethoxylated glycerol fatty acid esters. The cosmetic composition of the present invention may further contain excipients including fluorescent substances, fungicides, hydrotropes inducing substances, moisturizers, fragrances, fragrance carriers, proteins, solubilizers, sugar derivatives, sunscreens, vitamins, plant extracts, and the like. . The amount of the components added according to the formulation or purpose of use may be selected within a range that does not impair the inherent 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 preventing hair loss comprising a culture of stellar pimento flower. The pharmaceutical composition may be formulated and used as an external agent. Carriers, excipients, and diluents that may be included in the pharmaceutical composition include lactose, dextrose, sucrose, sorbitol, mannitol, xylitol, erythritol, maltitol, starch, gum acacia, alginate, gelatin, calcium phosphate, calcium silicate, and cellulose. , methyl cellulose, microcrystalline cellulose, polyvinyl pyrrolidone, water, methylhydroxybenzoate, propylhydroxybenzoate, talc, magnesium stearate and mineral oil. When formulated, 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. Determination of dosage based on these factors is within the level of those skilled in the art, and generally dosages 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 dosage is not intended to limit the scope of the present invention in any way.

The pharmaceutical composition of the present invention can be administered to mammals such as rats, livestock, and humans through various routes. All modes of administration can be envisaged, but application through skin application is most preferred in the present invention.

The present invention relates to a hair care cosmetic composition containing a strain of Bacillus siamensis HDB8077 (Accession No. KCCM12989P) isolated from a flower of sagebrush and a culture of sageflower of the strain. The culture is cultured by adding stylized calico flower powder to Bacillus siamensis HDB8077 strain culture medium, and has hair root strengthening effect, hair growth effect, androgenetic hair loss mitigation effect, ceramide biosynthetic enzyme expression enhancement effect, etc., so as a multifunctional hair composition readily available

Figure 1 shows the 16s rRNA base sequence (SEQ ID NO: 1) of the Bacillus siamensis HDB8077 strain of the present invention.
Figure 2 shows the activity of increasing the expression of VEGF mRNA for the flower cultures in the general hair growth model.
Figure 3 shows the activity of increasing the mRNA expression of Wnt10b (Fig. 3a) and KGF (Fig. 3b) for the flower cultures of stylized spp. in the general hair growth model.
Fig. 4 shows the activity of suppressing the expression of AR (Fig. 4a) and DKK-1 (Fig. 4b) in the male pattern baldness model against stylized rhizome flower cultures.
Figure 5 shows the activity of increasing the expression of SPT, a cell barrier protective factor in scalp cells of stylized septum flower culture.
Figure 6 shows the effect of removing active oxygen in the scalp cells of the philiphistridium staphylococcus flower culture.

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 to sufficiently convey the spirit of the invention to those skilled in the art, so that the disclosure herein will be thorough and complete.

<Preparation Example 1: Isolation and identification of strain>

After putting 1 g of the flowers of sagebrush in 10 mL of sterilized saline solution, it was suspended, serially diluted, and smeared on a BCP (bremocresol purple) solid medium. After culturing at 37 ° C for 24 hours, colonies were selected according to their shape and continuously passaged on BCP solid medium to obtain pure isolation.

The isolated strain was dissolved in water to form 1.5% peptone (w/v), 0.5% soybean peptone (w/v), 0.5% sodium chloride (w/v), and 1.5% agar (w/v) per liter. It was cultured using a sterilized one at 121 ~ 123 ℃ for 60 minutes.

In order to confirm the morphological characteristics of the strain, when cultured for 1 day at 37 ° C. in LB agar plate medium, the cell form was bacillus (serial), and it was confirmed that there was no motility. The sporulation ability of the strain is not confirmed, and it is a Gram-positive strain. As for the shape of the bacterial strain, it is confirmed that it builds a semi-transparent grayish circular smooth colony. When confirming the physiological characteristics of the strain, it can be cultured at 25 ~ 50 ℃, and the optimum growth temperature is 37 ℃. It was found that the pH available for growth was 6.0 to 7.5, and the optimum pH for growth was 6.0 to 6.5. The optimal growth time is 18 hours, and it can be cultured for a minimum of 16 hours and a maximum of 2 days. As a facultative aerobic strain, it was confirmed that no gas was produced.

To identify the isolated strain HDB8077, phylogenetic analysis was performed through 16S rRNA gene sequence comparison. As a result of analyzing the 16S rRNA (SEQ ID NO: 1, FIG. 1) of the HDB8077 strain using the Blast similarity search program (National Institute of Biotechnology Information), the sequence of the Bacillus siamensis strain had the highest rate of 99%. showed homology.

In addition, after multiple sequence alignment was performed with the 16S rRNA gene sequences of the same series of strains registered in GenBank, the location in the phylogenetic tree was determined, and the 16S rRNA gene on the phylogenetic tree was Bacillus sciatica. Placed closest to Mensis. As described above, using the result of molecular biological identification of the strain of the present invention, as Bacillus siamensis, HDB8077 strain having 16s rRNA of SEQ ID NO: 1 ( Bacillus siamensis HDB8077 strain, accession number KCCM12989P) was obtained from the Korean Microorganism Conservation Center (Korean Culture Center of Microorganisms, KCCM, http://www.kccm.or.kr/).

<Example 1. Preparation of a culture solution for staphylococcus aureus flowers>

A single colony of Bacillus siamensis HDB8077 strain was inoculated into sterilized TSB (Tryptic Soy Broth, BD Difco) and then incubated at 37° C. 170 rpm for 5 hours. Thereafter, 5 g of the autoclaved sterilized pyrophyllum flower powder was added to 100 g of the liquid culture medium and cultured at 37 ° C., 170 rpm, for 24 hours to prepare a culture medium of pyrophyllus trifolium to be used as a sample in the present invention. The culture solution of the flower culture of Stylus spp. was filtered through a 0.3 μm filter to remove the cells, and then completely concentrated and powdered.

<Comparative Example 1. Preparation of reactant for dead fungal solution of stylized staphylococcus flowers>

A single colony of Bacillus siamensis HDB8077 strain was inoculated into sterilized TSB Broth (Difco) and then incubated at 37°C and 170 rpm for 5 hours. Thereafter, the culture medium itself was sterilized for 15 minutes at 121 ° C using an autoclave, and 5 g of the sterilized iron pimento flower powder was added to 100 g of the sterilized culture medium and reacted at 37 ° C, 170 rpm, for 24 hours in the present invention. A reactant of dead bacteria in the flower of Stylus staphylococcus to be used as a sample was prepared. The reactant of the dead bacteria in the flower of Stylus strifolium was filtered through a 0.3 μm filter to remove the cells, and then completely concentrated and powdered.

<Comparative Example 2. Manufacture of philodendron flower extract>

After extracting 5 g of philipsodium flower powder compared to 100 g of purified water at 37 ° C., 170 rpm, for 24 hours, filtering with a 0.3 μm filter and completely concentrating to powder, to prepare an extract of pyrophyllum elegans flower to be used as a sample in the present invention.

<Experimental Example 1. Cell culture>

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

<Experimental Example 2. Confirmation of VEGF Expression for Confirmation of Hair Root Strengthening Efficacy>

To evaluate the hair root strengthening effect, VEGF (Vascullar Endotherial Growth Factor) gene expression experiments were performed through qRT-PCR. DMEM medium containing 10% (v/v) bovine serum and dermal papilla cells (DPCs, cell bio) were inoculated in 2 ml each at 2.5×10 ⁵ cells/well in a 6 well multi plate (corning) and cultured for 24 hours, then each The wells were replaced with serum free medium. Samples were treated in serum free medium and cultured for 24 hours. At this time, each sample was treated to be 25 to 100 μg/ml, and Minoxidil used for efficacy evaluation was treated to a total concentration of 1 μg/ml in the medium.

Next, after washing with PBS, the cultured cells were treated with QIAzol ^® (QIAGENL ^® , USA), and RNA was isolated using the protocol provided by the manufacturer (QIAGENL ^® ) after proceeding with cell lysis. The isolated RNA was quantified using a Qubit ^® fluoremeter with RNA BR Assay kit, and 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 was performed by amplifying the gene using a Real-time PCR Kit (2x qPCRBIO SyGreen Blue mix Lo-ROX, PCRBIOSYSTEMS, London, UK) and quantitatively analyzing the amplified product.

VEGF and β-actin primers used for PCR were synthesized and used by Cosmogenetech (Korea), and the primer sequences are shown in Table 1. The results for this are shown in Figure 2.

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 AT C CAC ACG GAG TAC TTG-3'

The reaction conditions for cDNA synthesis were 95°C for 5 seconds and 60°C for 25 seconds, 40 cycles for both VEGF and β-actin. Control was not subjected to sample treatment, and Minoxidil, used as a positive control, is known to induce hair root strengthening and hair growth.

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

As a result of the analysis in FIG. 2 , it can be seen that the culture of the flower of St. pisces of Example 1 of the present invention strengthens the hair root by supplying nutrients to the hair root through improvement of blood circulation.

<Experimental Example 3. Confirmation of gene expression of hair growth factors Wnt10b and KGF>

Regarding Wnt10b and KGF, factors related to hair growth qRT-PCR was performed as in Experimental Example 2 using HHGMC (Human Hair Germinal Matrix Cell), but the primer sequences were performed with reference to the sequences in Table 2 below. The results for this are shown in Figure 3.

Primer Sense Antisense Wnt10b 5'-CAT CCA GGC ACG AAT GCG A-3' 5'-CGG TTG TGG GTA TCA ATG AAG A-3' KGF 5'-AAG GGA CCC AGG AGA TGA AGA-3' 5'-TGC CAC AAT TCC AAC TGC C-3' β-actin 5'-GGC ACC CAG CAC AAT GAA G-3' 5'-CCG ATC CAC ACG GAG TAC TTG-3'

As a result of confirming FIGS. 3a and 3b, it can be seen that the hair growth efficacy of the culture of the stylized spp.

<Experimental Example 4. Confirmation of androgenetic hair loss factor AR and DKK-1 gene expression>

For AR and DKK-1, which are factors related to male pattern baldness, experiments were performed using HDP (Human Dermal papilla Cell) located in the center of the hair follicle. When DHT (Dihydrotestosteron) is delivered to hair follicles by AR (Androgen receptor), hair loss occurs, and thus, inhibiting AR expression inhibits DHT from being delivered to hair follicles, thereby alleviating early hair loss. In addition, as the expression of DKK-1 (dickkopf-1) increases during hair loss, which induces the death of hair follicles, it was confirmed whether the culture of stylized spp. flowers inhibits these factors.

For DKK-1 and AR, qRT-PCR was performed as in Experimental Example 2, but the primer sequences were performed with reference to the sequences in Table 3 below. The results for this are shown in Figure 4.

Primer Sense Antisense AR 5'-CCT GGC TTC CGC AAC TTA CAC-3' 5'-GGA CTT GTG CAT GCG GTA CTC A-3' DKK-1 5′-ATT CCA ACG CTA TCA AGA ACC-3′ 5′-CCA AGG TGC TAT GAT CAT TAC C-3′ β-actin 5'-GGC ACC CAG CAC AAT GAA G-3' 5'-CCG ATC CAC ACG GAG TAC TTG-3'

4a and 4b, the flower culture of Example 1 inhibited AR (Androgen Receptor) expression and showed the ability to inhibit the expression of DKK-1 (dickkopf-1), which induces hair follicle death, and showed male pattern hair loss alleviation efficacy can be seen to indicate

<Experimental Example 5. Verification of gene expression of scalp cell barrier protective factor SPT>

In order to confirm the efficacy of strengthening the scalp barrier, an experiment was conducted on Serine palmitoyl transferase (SPT), a factor inducing the expression of enzymes involved in ceramide biosynthesis in human normal keratinocytes, and the results are shown in FIG. 5 .

Primer Sense Antisense SPT 5'-TGG TCA TTT GGC CCA GGT C-3' 5'-TCC CAA CCA TTG GCT TCA CAT C-3' β-actin 5'-GGC ACC CAG CAC AAT GAA G-3' 5'-CCG ATC CAC ACG GAG TAC TTG-3'

Referring to FIG. 5, it can be seen that the culture of the stylized spp. flower of Example 1 increases the expression of SPT, a ceramide biosynthetic enzyme, which is a moisturizing factor for scalp cells, thereby improving the scalp cell barrier.

<Experimental Example 6. Confirmation of scalp antioxidant efficacy through inhibition of ROS activity>

In order to evaluate the antioxidant efficacy, active oxygen inhibition efficacy was evaluated using H ₂ O ₂ in the HaCaT cell line, which is a human-derived keratinocyte. 100 μl of HaCaT cell line and DMEM medium containing 10% bovine serum were inoculated into a 96-well multi-plate (Thermo) at a concentration of 3X10 ⁴ cells/96 well, cultured for 24 hours, and then each well was replaced with serum-free medium, and samples were collected. It was treated by serial dilution method. After culturing for 24 hours, wash using HBSS (Hanks' Balanced Salt Solution, Handok Biotech). After treating the cultured cells with 20 μM H ₂ DCFDA (2′,7′-Dichlorodihydrofluorescein diacetate, Sigma) for 30 minutes, wash the excess stained fluorescence using HBSS (H ₂ O ₂ treated group and non-treated plate) After incubating the group for 10 minutes, wash with HBSS and measure the fluorescence value at 492 nm using a microplate reader (spark 10M Elisa reader) After removing the buffer, add 50 μl of 0.5% Triton X-100 (Duksan) each, and BCA Add 50 μl of solution (Sigma) and measure the level of protein expression using the BCA protein quantification method ROS activity is calculated using the previously measured fluorescence value and protein amount, and a standard curve is prepared using BSA. Experimental results are shown in Figure 6.

Referring to FIG. 6 , it can be seen that the culture of the fermented spp. flower of Example 1 suppresses radical production in keratinocytes to exhibit excellent antioxidant efficacy.

<Formulation Example 1. Preparation of hair lotion>

A hair lotion was prepared with the composition shown in Table 5 below, using as a raw material the dissolution of the culture of the flower of the present invention with purified water and 1,3-Butylene Glycol to 2.0% (w/w).

ingredient weight(%) Cetostearyl Alcohol 2.0 Stearyltriethylammonium chloride 0.5 Hydroxyethylcellulose 0.1 Phytoxtoneolamine 0.1 Example 1 2% (w/w) aqueous solution of complex extract 2.0 Spices 0.1 pigment 0.1 Purified water balance Total 100

< formulation example 2. hair tonic manufacturing>

A hair tonic was prepared with the composition shown in Table 6 below, using as raw materials the dissolution of the culture of the flower of the present invention with purified water and 1,3-Butylene Glycol to a concentration of 2.0% (w/w).

ingredient weight(%) resorcinol 0.1 menthol 0.05 panthenol 0.2 salicylic acid 0.1 tocopheryl acetate 0.1 Pyridoxine hydrochloride 0.1 castor oil 5.0 2% (w/w) aqueous solution of stridium strifolium flower culture 2.0 pigment 0.1 Spices 0.1 ethanol 0.3 Purified water balance Total 100

< formulation example 3. hair Manufacture of Conditioner>

A hair conditioner was prepared with the composition shown in Table 7 below, using as a raw material the dissolution of the culture of the flower of the present invention with purified water and 1,3-Butylene Glycol to 2.0% (w / w).

ingredient weight(%) cetanol 3.5 Self-emulsifying glycerin monostearate 1.5 propylene glycol 2.5 Stearylmethylbenzyl Ammonium Chloride (25%) 7.0 Methyl paraoxybenzoate 0.3 2% (w/w) aqueous solution of stridium strifolium flower culture 5.0 pigment 0.1 Spices 0.1 Purified water balance Total 100

[Entrusted institution]

Name of Depository Organization: Korea Microbial Conservation Center

Accession number: KCCM12989P

Entrusted date: 20210520

<110> HYUNDAI BIOLAND CO., LTD. <120> Bacillus siamensis HDB8077 strain isolated from Dendrobium candidum flower and haircare cosmetic composition contained culture of Dendrobium candidum flower using its <160> 1 <170> KoPatentIn 3.0 <210> 1 <211> 1490 <212> DNA <213> unknown <220> <223> Bacillus siamensis <400> 1 gtcaggacga acgctggcgg cgtgcctaat acatgcaagt cgagcggaca gatggggagct 60 tgctccctga tgttagcggc ggacgggtga gtaacacgtg ggtaacctgc ctgtaagact 120 gggataactc cgggaaaccg gggctaatac cggatggttg tctgaaccgc atggttcaga 180 cataaaaggt ggcttcggct accacttaca gatggacccg cggcgcatta gctagttggt 240 gaggtaacgg ctcaccaagg cgacgatgcg tagccgacct gagagggtga tcggccacac 300 tgggactgag acacggccca gactcctacg ggaggcagca gtagggaatc ttccgcaatg 360 gacgaaagtc tgacggagca acgccgcgtg agtgatgaag gttttcggat cgtaaagctc 420 tgttgttagg gaagaacaag tgccgttcaa atagggcggc accttgacgg tacctaacca 480 gaaagccacg gctaactacg tgccagcagc cgcggtaata cgtaggtggc aagcgttgtc 540 cggaattatt gggcgtaaag ggctcgcagg cggtttctta agtctgatgt gaaagccccc 600 ggctcaaccg gggagggtca ttggaaactg gggaacttga gtgcagaaga ggagagtgga 660 attccacgtg tagcggtgaa atgcgtagag atgtggagga acaccagtgg cgaaggcgac 720 tctctggtct gtaactgacg ctgaggagcg aaagcgtggg gagcgaacag gattagatac 780 cctggtagtc cacgccgtaa acgatgagtg ctaagtgtta gggggtttcc gccccttagt 840 gctgcagcta acgcattaag cactccgcct ggggaggtacg gtcgcaagac tgaaactcaa 900 aggaattgac gggggcccgc acaagcggtg gagcatgtgg tttaattcga agcaacgcga 960 agaaccttac caggtcttga catcctctga caatcctaga gataggacgt ccccttcggg 1020 ggcagagtga caggtggtgc atggttgtcg tcagctcgtg tcgtgagatg ttgggttaag 1080 tcccgcaacg agcgcaaccc ttgatcttag ttgccagcat tcagttgggc actctaaggt 1140 gactgccggt gacaaaccgg aggaaggtgg ggatgacgtc aaatcatcat gccccttatg 1200 acctgggcta cacacgtgct acaatggaca gaacaaaggg cagcgaaacc gcgaggttaa 1260 gccaatccca caaatctgtt ctcagttcgg atcgcagtct gcaactcgac tgcgtgaagc 1320 tggaatcgct agtaatcgcg gatcagcatg ccgcggtgaa tacgttcccg ggccttgtac 1380 acaccgcccg tcacaccacg agagtttgta acacccgaag tcggtgaggt aacctttatg 1440 gagccagccg ccgaaggtgg gacagatgat tggggtgaag tctaaaaggg 1490

Claims (7)

Bacillus siamensis HDB8077 strain (Accession No. KCCM12989P). According to claim 1,
The strain is a strain of Bacillus siamensis HDB8077, characterized in that isolated from the flower of Stylus staphylococcus. According to claim 1,
The strain is Bacillus siamensis HDB8077 strain, characterized in that it enhances the hair care function of stylized staphylococcus flowers. A hair care cosmetic composition characterized in that it contains a culture of the strain of claim 1. According to claim 4,
The composition is a hair care cosmetic composition, characterized in that there is an effect of strengthening the hair root, promoting hair growth or preventing hair loss. According to claim 4,
The culture is a hair care cosmetic composition, characterized in that incubated for 20 to 48 hours at 30 ~ 40 ℃ after adding stylized calico flower powder to the culture solution of the strain of claim 1. A pharmaceutical composition for preventing hair loss, characterized in that it contains a culture of the strain of claim 1.

Images