KR102588878B1 - Composition for improving vaginitis and colpoxerosis, and Improvement agent for vaginitis and colpoxerosis containing the same - Google Patents

Abstract

The present invention relates to a composition for improving vaginitis and vaginal dryness using a complex extract of natural plants and platelet-rich plasma, and to a composition that effectively improves vaginitis while also being excellent in preventing and improving vaginal dryness, and to an agent for improving vaginitis and vaginal dryness using the same. It's an invention.

Description

Vaginitis and vaginal dryness improvement composition, and vaginitis and vaginal dryness improvement agent containing the same {Composition for improving vaginitis and colpoxerosis, and Improvement agent for vaginitis and colpoxerosis containing the same}

The present invention relates to a composition that can improve vaginal dryness while preventing and improving bacterial vaginosis while maintaining a slightly acidic environment inside the vagina that helps protect against invasion by pathogenic microorganisms, and a vaginal improvement agent using the same. will be.

The female vagina is located at the entrance to the uterus, so it is directly affected by the menstrual cycle, and is located close to the urethra and anus, so there is a high possibility of contamination by colonic microorganisms and urethral disease microorganisms.

Pathogenic microorganisms that cause infections around the female genital tract include E. coli, Tricomonas vaginalis, Candida albicans, Chlamidia, Haemophilus vaginalis, and Herpes virus. and Gardnerella sp., and vaginitis is commonly caused by single or mixed infection with these bacteria. Bacterial vaginosis (BV) generally accounts for more than 50% of infections of the female reproductive system, causing persistent chronic infection symptoms. In particular, there are cases where women cannot maintain a normal state due to the infiltration of pathogenic microorganisms during sexual activity, and not only can they suffer from various types of diseases such as local inflammation, but harmful microorganisms can also penetrate the pelvic cavity, leading to serious conditions such as infertility. I also do it.

Normal bacteria living inside a woman's vagina include Lactobacilli, Streptococcus (group β-hemolytic streptococcus), and Coliform bacteria. Among these, female hormones are used to prevent contamination by external microorganisms. Under the influence of estrogen, glycogen stored in vaginal epithelial cells is used for lactic acid fermentation. Lactic acid fermentation is achieved by lactic acid bacteria present in the vagina. The appropriate level of lactic acid bacteria maintains the pH in the vagina at 4.5 to 5.5, thereby suppressing the growth of pathogenic microorganisms invading from the outside.

Bacterial vaginitis is clinically treated by oral administration of antibiotics or administration of ointment or vaginal tablets directly into the vagina. Imidazole and nitroimidazole derivatives are often used to treat these symptoms, and other types of drugs used are There are nitrofurfuryl derivatives and various antibiotics. These active antibiotic ingredients can be formulated for oral or topical administration. Metronidazole is commonly administered by the oral route, but mixed infections are intended for topical administration, especially in pessary form containing two or more active ingredients. It is suitable for However, in the case of treatment using antibiotics, antibiotics affect the growth of not only pathogenic microorganisms that cause vaginitis, but also the normal bacterial flora, especially lactic acid bacilli, in the vagina, thereby reducing their numbers. This destroys the normal vaginal flora, resulting in chronically recurring vaginitis. can occur.

In addition, vaginal dryness is a condition in which a woman's Y zone becomes dry and causes pain. It occurs before and after menopause, after childbirth and miscarriage, long-term use of contraceptives, and repeated vaginitis, as well as vaginal dryness, itchiness, and dyspareunia. It causes symptoms such as: In the case of vaginal dryness, it can be the cause of vaginitis due to the imbalance of the vaginal environment and atrophy of the mucous membrane. If it occurs together with vaginitis, the pain becomes extreme and purulent discharge or bleeding may occur. It is effective to prevent or treat.

Accordingly, the need for efficient methods to prevent and improve vaginal dryness while also preventing, improving, and even treating vaginitis is increasing.

Republic of Korea Patent Publication No. 10-2020-0001214 (publication date 2020.01.06) Republic of Korea Patent No. 10-0862211 (announcement date 2008.10.09)

The present invention uses extracts derived from natural plants to provide a composition with an optimal composition that is not only effective in healing vaginal wounds caused by sexual intercourse, etc. without causing trouble, but can simultaneously prevent or treat vaginal dryness along with vaginitis, and We would like to provide application products using this.

The composition for improving vaginitis and vaginal dryness of the present invention for solving the above-described problems includes a complex extract of natural plants; platelet-rich plasma (PRP); hyaluronic acid; Antibacterial agents including mandelic acid; warming agent; Solubilizer; thickener; non-ionic surfactants; and water;

Additionally, the present invention is an injection-type agent for improving vaginitis and vaginal dryness, and includes the composition for improving vaginitis and vaginal dryness.

The composition for improving vaginitis and vaginal dryness of the present invention is not toxic to beneficial bacteria in the vagina, has excellent antibacterial properties against bacteria that cause vaginal inflammation, and is excellent in preventing vaginal odor and preventing or improving vaginal dryness. The composition of the present invention includes tablets, ointments, gels, creams, cleansers, water (W) type, oil (O) type, oil-in-water (O/W) type, and water-in-oil (W/O) type for intravaginal insertion. , it can be applied in various formulations such as silicone (S) type, water in silicone (W/S) type, silicone in water (S/W) type, solid form, liquid form, etc., and is preferably commercialized as an injection type.

Hereinafter, the present invention will be described in more detail.

The composition for improving vaginitis and vaginal dryness (hereinafter referred to as composition) of the present invention includes natural plant complex extract, platelet-rich plasma (PRP), hyaluronic acid, antibacterial agent, warming agent, solubilizer, thickener, and non-ionic interface. Contains activator and water.

Among the compositions of the present invention, the natural plant complex extract includes natural plant mixed extract; and a fermented product of spikenard root extract; preferably included in a weight ratio of 1:0.10 to 0.30, preferably in a weight ratio of 1:0.12 to 0.20 in terms of maintaining the vaginal environment while increasing antibacterial properties along with the antibacterial agent, and the natural Complex plant extracts are also advantageous in terms of improving Trichomonas vaginitis and/or Candida vaginitis.

The natural plant mixture extract is a mixed extract of a natural plant mixture including ratania root, elm bark, lotus root, and Virginia wildflower leaves.

Rhatany is a plant that grows in the dry desert areas of Bolivia and Peru and is known to have excellent antioxidant, antibacterial and anti-inflammatory effects. This can provide vaginal conditioning and a refreshing feeling while increasing antibacterial properties against harmful vaginal bacteria. In the present invention, the roots of Ratania are used.

Sugar elm tree ( Ulmus) 　 davidiana Planchon) is a typical continental elm type and is widely distributed on the Korean Peninsula, becoming more numerous in the central region. Among the elm trees (Ulmus spp.), it is more commonly observed on the slopes of mountain valleys in igneous or metamorphic rocks. In the present invention, the bark of the elm tree is used.

Hamamelis virginiana is a medicinal plant known as witch hazel or Western hazelnut. It is known to have wound healing properties and contains AHA ingredients, which remove dead cells from the skin surface and allow healthy cells to rise to the epidermis. It is known to help restore the skin cell regeneration cycle, and in the present invention, the leaves of Virginia wildflower are used.

In addition, the natural plant mixture may include 100 to 150 parts by weight of elm bark, 20 to 50 parts by weight of softwood, and 20 to 50 parts by weight of Virginia wildflower leaves, based on 100 parts by weight of ratania roots. Preferably, It may include 100 to 130 parts by weight of sugar elm bark, 20 to 40 parts by weight of lotus root, and 30 to 50 parts by weight of Virginia wildflower leaves, more preferably, per 100 parts by weight of ratania roots. It is good to include 100 to 120 parts by weight of elm bark, 20 to 30 parts by weight of softwood, and 35 to 45 parts by weight of Virginia wildflower leaves in terms of increasing the antibacterial effect against harmful bacteria in the vagina and preventing toxicity to beneficial bacteria.

In addition, the natural plant mixed extract is prepared by mixing 20 to 35 parts by weight of a natural plant mixture, preferably 22 to 35 parts by weight, and more preferably 25 to 35 parts by weight, with respect to 100 parts by weight of purified water; Obtaining an extract by extracting the raw material by distillation at 100 to 110°C for 4 to 6 hours; Preparing a primary filtrate obtained by primary filtering the extract; Adding and stirring 1,2-hexanediol to the primary filtrate, followed by secondary filtering to prepare a mixed extract; a product prepared by performing the following process may be used.

When preparing the mixed extract, the first filtering may be performed using a filter membrane with an average pore size of 20 μm to 30 μm, and preferably, the filtering may be performed using a filter membrane with an average pore size of 23 μm to 27 μm.

In addition, the secondary filtering may be performed using a filter membrane with an average pore size of 8㎛ to 15㎛, preferably 8㎛ to 12㎛ filter membrane.

Nardostachys chinensis , used in the production of fermented product of spikenard root extract, is a perennial herb belonging to the Matariaceae family and is native to the mountainous regions of India and Nepal. Because of its sweet taste, it is also called spikenard, spikenard, gomichi, and fragrant. It is also said that The spikenard has a cylindrical shape bent into a bow shape, the outer surface is shiny yellow-brown to brown, and the cut surface is yellowish-brown. The outer surface has 5 to 6 vertical wrinkles and irregular wrinkles between them, is hard, has a long and thick main root, has a beard root, and the quality is soft and easy to bend. The cortex is dark brown and splits into pieces, and the xylem is yellowish white. Perilla incense has been used as an ingredient in painkillers since ancient times, and it is known as nard incense, which is one of the fragrance oils used by Jews at weddings.

The fermented product of spikenard root extract provides a refreshing feeling, soothes and relieves vaginal skin, removes and prevents vaginal odor, and can play a role in alleviating cytotoxicity of mixed extracts of natural plants. The fermented product of the spikenard root extract is prepared by mixing the spikenard root powder with an aqueous ethanol solution at a concentration of 40 to 50% by volume, followed by an extraction process at 40 to 50° C. for 16 to 24 hours to produce an ethanol extract; Step 2 of mixing the ethanol extract with the microbial fermentation broth, inoculating the fermentation strain and performing a fermentation process to obtain a fermented product; The fermented product can be produced by performing a process including three steps of sequentially performing a filtration process, a sterilization process, and a purification process.

Step 2 is a process of obtaining a fermented product from the extract prepared in Step 1, and the ethanol extract is added to the microbial fermentation culture at a concentration of 20 to 40 g/L, preferably at a concentration of 20 to 30 g/L, more preferably at a concentration of 20 to 40 g/L. After mixing at ~25g/L, fermentation can be performed by inoculating the fermentation bacteria and then culturing the fermentation bacteria. At this time, if the concentration of the complex extract in the microbial fermentation culture is less than 20g/L, a sufficient amount of fermented product may not be obtained, and if it exceeds 40g/L, there may be a problem of poor fermentation because the fermenting bacteria are not cultured well. .

In addition, the microbial fermentation culture medium may use MRS medium adjusted to pH 5.5 to 7.0 with a pH adjuster such as dipocalcium phosphate.

And, the fermentation bacteria are Lactobacillus plantarum, Lactobacillus acidophilus, Lactobacillus brevis, Lactobacillus paracasei, and Leuconostoc mesenteroides. One or more species selected from (leuconostoc mesenteroides) and Enterococcus faecium can be used, preferably Lactobacillus plantarum, Leuconostoc mesenteroides and/or Enterococcus faecium, more preferably Can be used by mixing Lactobacillus plantarum and Leuconostoc mesenteroides at a ratio of 1:1.5 to 3.0 CFU, more preferably at a ratio of 1:1.8 to 2.5 CFU.

And, the inoculation amount of the fermentation strain is (1.0 ~ 8.0) × 10 ⁵ cfu/L, preferably (1.5 ~ 5.0) × 10 ⁵ cfu/L, more preferably (2.0 ~ 3.5) for the microbial fermentation culture mixed with the extract. )×10 ⁵ cfu/L is advantageous in terms of obtaining an appropriate amount of fermented product and controlling the degree of fermentation.

The fermentation process is performed by culturing the fermenting strain at pH 5.5 to 7.0 and 30 to 38°C for 36 to 52 hours, preferably at pH 5.8 to 6.5 and 32 to 38°C for 40 to 52 hours. good night. At this time, if the pH is outside the range of 5.5 to 7 and/or the temperature is 35 to 38°C, there may be a problem in which the strain is not cultured well, so it is recommended to perform the fermentation process under the above conditions.

Additionally, the fermentation process can be performed under aerobic or normal anaerobic conditions, and is preferably performed under anaerobic conditions.

Next, step 3 is a process of obtaining a purified fermented product by sequentially performing a filtration process, a sterilization process, and a purification process on the fermented product obtained in step 2.

The filtration and purification processes are processes that separate and remove foreign substances generated during the fermentation process from the fermented product through general filtration and purification methods used in the industry.

The sterilization process is a process of killing fermentation bacteria in the fermentation product, and can be performed through general sterilization methods. For example, in a preferred embodiment, the fermentation product is placed in an autoclave at a temperature of 70°C or higher and 80 to 90°C. After injection, sterilization can be performed through heat treatment for 10 to 30 minutes.

Next, among the compositions of the present invention, platelet-rich plasma (PRP) plays a role in treating vaginal wounds, etc. and preventing and improving vaginal dryness, and the PRP is pure platelet-rich plasma, white blood cell, etc. It may include deficient-platelet-rich plasma or white blood cell-rich-platelet-rich plasma, and preferably may include pure-platelet-rich plasma or white blood cell-rich-platelet-rich plasma. In addition, the amount of PRP used in the composition of the present invention can be 0.01 to 25.00 parts by weight, preferably 0.10 to 20.00 parts by weight, and more preferably 1.00 to 5.00 parts by weight, based on 100 parts by weight of the natural plant complex extract. , if the amount of PRP used is less than 0.01 parts by weight, the content in the composition is too small, so the effect of preventing and improving vaginal dryness may be minimal, and using more than 25 parts by weight is excessive use, and the increase in effect due to excessive use is no longer possible. No, it is uneconomical due to price increases.

Next, the hyaluronic acid in the composition of the present invention serves to provide a moisturizing effect and increase the feeling of use, and the amount used is 0.1 to 5.0 parts by weight, preferably 0.3 to 4.0 parts by weight, based on 100 parts by weight of the natural plant complex extract. It can be used in an amount of 0.5 to 3.0 parts by weight, more preferably 0.5 to 3.0 parts by weight. If the amount of hyaluronic acid used is less than 0.1 part by weight, the amount used is too small and the effect of improving vaginal dryness through the moisturizing effect due to its use may be minimal, and 5 parts by weight If it is used in excess of the amount, miscibility and compatibility within the composition may decrease and the viscosity of the composition may become too high, so it is better to use it within the above range.

Next, the antibacterial agent in the composition of the present invention may be one known in the art to have antibacterial properties against vaginitis-causing bacteria, and mandelic acid may be preferably used. In addition, mandelic acid is an ingredient that has antibacterial properties against vaginitis-causing bacteria, and when used with natural plant complex extracts, it can increase antibacterial properties against vaginitis-causing bacteria. In addition, specific examples of the vaginitis-causing bacteria include Gardnerella vaginalis, C. albicans, Bacillus cereus, Enterococcus faecalis, Escherichia coli, Listeria monocytogenes, Salmonella enteritidis, Salmonella gallinarum, Yersinia enterocolitica and Staphylococcus areus It may include one or more species selected from (Staphylococcus aureus).

In addition, the amount of the antibacterial agent used may be 0.5 to 5.0 parts by weight, preferably 0.5 to 3.5 parts by weight, and more preferably 0.8 to 2.5 parts by weight, based on 100 parts by weight of the natural plant complex extract. At this time, if the amount of the antibacterial agent used is less than 0.5 parts by weight, there may be a problem that the antibacterial properties against the vaginitis-causing bacteria are reduced, and if the amount of the antibacterial agent used exceeds 5.0 parts by weight, there may be a problem of the vaginal environment being broken due to the antibacterial effect on the beneficial bacteria in the vagina. It can cause itchiness, so it is better to use it within the above range.

Next, the warming agent in the composition of the present invention serves to increase the feeling of use and improve the vaginal environment by causing a warming effect after applying the composition of the present invention intravaginally. The warming agent may include one or more selected from vanillin ethyl ether, vanillin propyl ether, and vanillin butyl ether, and preferably includes vanillin butyl ether. In addition, the amount of the warming agent used is appropriately 2.0 to 8.0 parts by weight, preferably 3.0 to 6.0 parts by weight, and more preferably 3.5 to 5.5 parts by weight, based on 100 parts by weight of the natural plant complex extract. When used, the antibacterial properties against vaginitis-causing bacteria and/or the improving effect of natural plant complex extracts on Trichomonas vaginitis and/or Candida vaginitis may be reduced, and the effectiveness of preventing and improving vaginal dryness may also be reduced, so it is recommended to use within the above range. .

Next, the solubilizing agent in the composition of the present invention serves to increase the miscibility between compositions and the solubility of the composition in water, and includes C ₆ to C ₁₂ fatty acids or glycols, monoglycerides, diglycerides or triglycerides thereof. It may contain ceride ester, preferably C ₆ to C ₁₂ fatty acid monoglyceride, and more preferably C ₈ to C ₁₂ fatty acid monoglyceride. In addition, the appropriate amount of the solubilizer to be used is 0.05 to 2.00 parts by weight, preferably 0.10 to 1.50 parts by weight, and more preferably 0.20 to 1.00 parts by weight, based on 100 parts by weight of the natural plant complex extract. If the topical amount used exceeds 2.00 parts by weight, the problem of phase separation may occur when the composition for improving vaginitis and vaginal dryness is stored for a long period of time, so it is recommended to use within the above range.

Next, the thickener in the composition of the present invention plays a role in controlling the viscosity of the composition of the present invention, and any thickener known to be harmless to the human body can be used without limitation, preferably polyethylene glycol with a weight average molecular weight of 200 to 1,000. Preferably, low molecular weight polyethylene glycol with a weight average molecular weight of about 200 to 600 can be used. In addition, the amount of the thickener used can be adjusted depending on the viscosity of the composition to be prepared, and as a preferred example, based on 100 parts by weight of the natural plant complex extract, 0.10 to 10.00 parts by weight, preferably 0.50 to 8.00 parts by weight, More preferably, it is appropriate to use it within the range of about 1.50 to 6.00 parts by weight.

Next, the nonionic surfactant in the composition of the present invention serves to increase the miscibility and solubility of the composition together with the solubilizer, and includes PEG-6 (Polyethylene glycol-6) palmitostearate, ethylene glycol palmito It may contain one or more selected from stearate, sorbitan monostearate, lauroyl polyoxyl-6 and polysorbate 80, preferably sorbitan monostearate, lauroyl polyoxyl-6 and polysorbate. It may contain one or more types selected from Sorbate 80. In addition, the amount of the nonionic surfactant used is 0.10 to 3.00 parts by weight, preferably 0.20 to 2.50 parts by weight, and more preferably 0.40 to 2.00 parts by weight, based on 100 parts by weight of the natural plant complex extract. , if the amount of nonionic surfactant used is less than 0.10 parts by weight, the effect of increasing miscibility and solubility between compositions due to its use may be minimal, and if it exceeds 3.00 parts by weight, it is uneconomical due to excessive use, so it is better to use it within the above range.

Next, the water in the composition of the present invention serves as a solvent for the composition, and distilled water, ionized water, or ultrapure water can be used. Any water that has been sterilized and removed foreign substances can be used without limitation. In addition, the amount of water used can be 10 to 50 parts by weight, preferably 10 to 30 parts by weight, and more preferably 10 to 25 parts by weight, based on 100 parts by weight of the natural plant complex extract. When used in less than 10 parts by weight, Miscibility between compositions may not be achieved well, and if used in excess of 50 parts by weight, there may be a problem of excessive use and the viscosity of the composition being too low, so it is appropriate to use within the above range.

The composition of the present invention described above may further include pharmaceutically acceptable additives. Pharmaceutically acceptable additives of the present invention may include, but are not limited to, vaginal beneficial lactic acid bacteria, binders, anti-adhesion agents, stabilizers, disintegrants, excipients, conditioning agents, neutralizers, colorants, and fragrances. The content and type of the additive can be appropriately selected by a person skilled in the art within a known range as long as it does not affect the stability or efficacy of the active ingredient.

The composition of the present invention can be prepared in the form of a general emulsified formulation and solubilized formulation using commonly known production methods. For example, tablets for vaginal insertion, ointments, gels, creams, cleansers, water (W) type, oil (O) type, oil-in-water (O/W) type, water-in-oil (W/O) type, silicone. It can be manufactured in various formulations such as (S) type, water-in-silicon (W/S) type, silicone-in-water (S/W) type, solid phase, and liquid phase. Preferably, it can be produced in a liquid phase with a gel-like viscosity. It can be manufactured in a form that is injected intravaginally. Additionally, it can be applied to the vulva and used to improve vaginitis and/or vaginal dryness in the vulva.

Hereinafter, the present invention will be described in more detail through examples. However, the following examples do not limit the scope of the present invention, and should be interpreted to aid understanding of the present invention.

[Example]

Preparation Example 1: Preparation of natural plant mixed extract

Dried ratania roots, dried elm bark, dried lotus root, and Virginian wildflower leaves were prepared, and each of them was pulverized to prepare their pulverized products.

A natural plant mixture was prepared by mixing 100 parts by weight of pulverized ratania root, 115 parts by weight of pulverized linseed bark, 23 parts by weight of pulverized softwood, and 38 parts by weight of pulverized Virginia wildflower leaves on a dry mass basis.

Raw materials were prepared by mixing 32 parts by weight of the powdered natural plant mixture with 100 parts by weight of purified water.

Next, the raw material was distilled and extracted at 110°C for 4 hours to obtain an extract.

Next, the extract was filtered through a filter membrane with an average pore size of 25㎛ to obtain a primary filtrate.

Next, 2 ml of 1,2-hexanediol was added to 94.9 ml of the filtrate, and after sufficient stirring, the stirred solution was secondarily filtered using a filter membrane with an average pore size of 10 ㎛ to prepare a natural plant mixed extract. . The obtained natural plant mixed extract was liquid, and the pH was about 5.37.

Preparation Example 2: Preparation of fermented product of spikenard root extract

After washing the spikenard root, it was cut into pieces, then dried and powdered to prepare spikenard fragrance powder. Next, the spikenard root powder was mixed with an ethanol aqueous solution at a concentration of 45% by volume at a weight ratio of 1:3, and then an extraction process was performed at 42-45°C for 18 hours to prepare an ethanol extract. Next, the ethanol extract was mixed at a concentration of 22.3 g/L with the microbial fermentation broth (MRS medium), and the pH was adjusted to about 5.6-5.7 with dipocalcium phosphate.

Next, Lactobacillus plantarum and Leuconostoc mesenteroides as fermentation bacteria were inoculated into the microbial fermentation broth mixed with pH adjustment and extract at a ratio of 1:2.28 CFU. At this time, the total amount of fermented bacteria inoculated was about (3.2~3.3)×10 ⁵ cfu/L.

Next, after sealing the fermenter containing the microbial fermentation broth, the fermentation process was performed for 46 hours under conditions of pH 6.1 to 6.2 and 35 to 36°C.

Next, the microbial fermentation broth from which the fermentation process was performed was filtered to obtain a fermented product, which was then sterilized and purified in an autoclave at 80 to 82°C to prepare a fermented product of spikenard root extract.

Preparation Example 3-1: Preparation of natural plant complex extract

A natural plant complex extract was prepared by mixing the natural plant mixed extract prepared in Preparation Example 1 and the fermented spikenard root extract prepared in Preparation Example 2 at a weight ratio of 1:0.15 and stirring slowly.

Preparation Examples 3-2 to 3-3 and Comparative Manufacturing Examples 3-1 to 3-3

Natural plant composite extracts were prepared using the same natural plant mixed extract and fermented spikenard root extract as in Preparation Example 3-1, but varying the weight ratio as shown in Table 1 below.

division Weight ratio of natural plant mixed extract and fermented spikenard root extract Preparation example 3-1 1 : 0.15 Preparation example 3-2 1 : 0.20 Preparation example 3-3 1 : 0.25 Comparison preparation example 3-1 1:0 Comparison preparation example 3-2 1 : 0.05 Comparison preparation example 3-3 1:0.35

Experimental Example 1: Cytotoxicity experiment

(1) Cell culture

RAW264.7 macrophages were purchased from the Korean Collection for Type Culture (KCTC, Korea), and 10% fetal bovine serum (FBS) and 1% penicillin/streptomycin were added. It was used after subculturing 2 to 3 times in an incubator at 37°C and 5% CO ₂ using DMEM medium (Welgene, Korea).

(2) Cytotoxicity analysis

To confirm the cytotoxicity of the natural plant complex extract, MTT analysis was performed. RAW264.7 macrophages were distributed in a 96-well plate at a concentration of 1 × 10 ⁵ cells/well and cultured overnight. Then, each natural plant complex extract was treated at different concentrations (200, 500 μg/ml) and cultured for 24 hours. Afterwards, 10 μL of MTT (5 mg/mL) was added to each well and cultured for additional 4 hours at 37°C, and then 100 μL of DMSO was added to each well to lyse the cells. Afterwards, the plate was maintained at room temperature for 5 minutes, and the absorbance was measured at 550 nm using a microplate reader (multiwell spectrophotometer, Molecular Devices, Sunnyvale, CA). The results are shown in Table 2 below.

The control group was not treated with the natural plant complex extract, and the gamma PGA was treated with gamma PGA at a concentration of 500 μg/ml.

cell viability Extract treatment concentration
200 μg/ml Extract treatment concentration
500 μg/ml control group 100% 100% Gamma PGA - 99.3% Preparation example 3-1 100.2% 98.4% Preparation example 3-2 102.6% 99.3% Preparation example 3-3 103.7% 100.8% Comparison preparation example 3-1 98.2% 95.9% Comparison preparation example 3-2 99.4% 97.7% Comparison preparation example 3-3 104.0% 100.6%

Looking at the cytotoxicity measurement results in Table 2 above, in the case of Comparative Preparation Example 3-1, in which only the natural plant complex extract of Preparation Example 1 was used, cytotoxicity was shown to occur somewhat. On the other hand, Preparation Examples 3-1 to 3-3 and Comparative Preparation Examples 3-2 to 3-3, which used a mixture of fermented spikenard root extracts, showed results in which cytotoxicity was greatly alleviated or improved. However, Comparative Preparation Example 3-2, which used a small amount of fermented spikenard root extract, did not show a satisfactory cell viability, and Comparative Preparation Example 3-3 was used at 500 μg compared to Preparation Examples 3-1 and 3-3. At a concentration of /ml, the effect of increasing cell viability was minimal or showed no increase.

Experimental Example 2: Evaluation of antibacterial activity against Trichomonas vaginalis

The growth inhibitory effect on Trichomonas vaginalis of each of the natural plant complex extracts of Preparation Examples 3-1 to 3-3 and the natural plant mixed extract of Preparation Example 1 (excluding fermented spikenard root extract) is as follows. Measured together.

Trichomonas vaginalis was cultured anaerobically in ATCC medium 361 containing 10% FBS and used in the experiment. Extracts were treated at different concentrations in 96-well microplates, inoculated so that the final bacterial count was 5×10 ⁴ cells/ml, and then cultured anaerobically for 24 hours. The untreated group was used as a negative control group, and the absorbance of the cultured strain was measured at a wavelength of 600 nm to compare the relative number of bacteria compared to the negative control group to confirm the minimum inhibitory concentration. The results are shown in Table 3 below.

growth
inhibition rate 0.2
mg/ml 0.8
mg/ml 1.2
mg/ml 2.0
mg/ml 3.0
mg/ml 4.0
mg/ml Ieast
Inhibitory concentration Preparation example 3-1 80.3% 73.6% 40.3% 19.5% 13.5% 13.6% 3.0mg/ml Preparation example 3-2 77.9% 70.4% 38.5% 11.4% 11.5% 11.2% 2.0mg/ml Preparation example 3-3 75.6% 68.3% 36.7% 11.9% 12.1% 12.0% 2.0mg/ml Preparation Example 1 (Control) 90.6% 84.8% 68.4% 50.4% 39.8% 30.4% -

Looking at Table 3 above, Preparation Examples 3-1 to 3-3 using a mixture of fermented spikenard root extracts had a higher growth inhibition effect on Trichomonas vaginalis than using only the natural plant mixed extract of Preparation Example 1. It was confirmed that the natural plant complex extract had an excellent growth inhibition effect on Trichomonas vaginalis.

Experimental Example 3: Evaluation of antibacterial activity against Candida albicans

The number of Candida albicans cells was diluted with Sabouraud Dextrose medium to 2 × 10 ⁵ cells/ml, and then prepared by serial dilution in 96-wells. Example 3 The natural plant complex extract of -1 and the natural plant mixed extract of Preparation Example 1 (control group) were each treated at different concentrations. Afterwards, the culture was incubated at 28°C for 24 hours and the absorbance was measured at a wavelength of 570 nm using a microplate reader (BIO-RAD) to determine the IC ₅₀ value. The results are shown in Table 4 below.

division IC ₅₀ (mg/ml) value Preparation example 3-1 10.6 Preparation Example 1 (Control) 13.2

Through Table 3 and Table 4 above, the natural plant complex extracts of Preparation Examples 3-1 to 3-3 have excellent antibacterial activity against not only Trichomonas vaginalis but also Candita albicans, and due to the use of spikenard root fermented extract It was confirmed that there was an antibacterial effect.

Preparation Example 4-1: Preparation of antibacterial agent mixture

A mixture of 100 parts by weight of the natural plant complex extract prepared in Preparation Example 3-1 and 1.06 parts by weight of mandelic acid, an antibacterial agent, was prepared.

Preparation Example 4-2: Preparation of antibacterial agent mixture

A mixture was prepared by mixing 4.02 parts by weight of mandelic acid, an antibacterial agent, with 100 parts by weight of the natural plant complex extract prepared in Preparation Example 3-1.

Experimental Example 4: Evaluation of antibacterial activity against vaginitis-causing pathogens

The minimum inhibitory concentration of mandelic acid against vaginitis-causing pathogens was measured, and this was used as a control, and the minimum inhibitory concentration for each of the mixture prepared in Preparation Examples 4-1 and 4-2 and the natural plant complex extract of Preparation Example 3-1 was measured. Inhibitory concentration was measured using the same method.

The minimum inhibitory concentration for vaginitis-causing pathogens was measured using a microtiter plate using the NCCLS (National Committee for Clinical Laboratory Standards) method, and Lactobacillus (Lactobacillus spp.) alone or Lactobacillus and vaginitis-causing pathogens were measured at 5 After culturing at 37°C for 12 hours under anaerobic conditions in ml broth, the cells were transferred to a microcentrifuge tube and centrifuged at 15,000 rpm for 5 minutes to remove the supernatant. To remove the medium components, 1 ml of 0.85% saline was added and completely suspended, and then centrifuged for 5 minutes to remove the supernatant. At this time, the pathogens causing vaginitis are Enterococcus faecalis (E. faecalis), Y. enterocolitica, Listeria monocytogenes (L. monocytogenes), or Salmonella enteritidis (S. enteritidis), respectively. did.

Then, the precipitated cell pellet was suspended by adding 1 ml of 0.85% saline solution, and then the absorbance at 625 nm (colony forming unit) was 1 × 10 ⁷ CFU using the Macfarland method ( It was inoculated at 10% on a microtiter plate so that OD 625 nm) was 0.09. Different concentration ranges of mandelic acid or the mixture of Preparation Examples 4-1 and 4-2 (0.10 mg/ml, 0.25 mg/ml, 0.75 mg/ml, 1.00 mg/ml, 1.25 mg/ml, 1.5 mg/ml, 1.75 mg/ml, 2.0 mg/ml, 2.25 mg/ml, 2.5 mg/ml, 3.5 mg/ml, 5 mg/ml, 7.5 mg/ml, and 10 mg/ml) and treated at 37°C for 24 hours. After culturing for some time, the absorbance at 600 nm was measured using a spectrophotometer (Molecular Devices, VERSAmax microplate reader, USA). The measurement results are shown in Table 5 below.

Minimum inhibitory concentration mandelic acid Preparation example 3-1 Preparation example 4-1 Preparation example 4-2 Lactobacillus No growth reduction No growth reduction No growth reduction 7.5 mg/ml Enterococcus faecalis 1.25mg/ml 7.5mg/ml 1.0 mg/ml 0.75 mg/ml Ersinia enterocolitica 1.25mg/ml 10mg/ml 1.25mg/ml 1.0 mg/ml Listeria monocytogenes 1.25mg/ml 5mg/ml 1.0 mg/ml 1.0 mg/ml Salmonella Enteritidis 2.5mg/ml 10mg/ml 2.0mg/ml 1.75mg/ml

Looking at the experimental results in Table 5 above, in the case of the natural plant complex extract of Preparation Example 3-1, the antibacterial activity was lower compared to mandelic acid and its mixture (Preparation Example 4-1, Preparation Example 4-2). showed.

In addition, the antibacterial activity tended to increase when mixed with natural plant complex extracts rather than when mandelic acid was used alone.

In addition, mandelic acid alone, the natural plant complex extract of Preparation Example 3-1, and Preparation Example 4-1 did not inhibit the growth of Lactobacillus, which is a normal bacterium. However, in the case of Preparation Example 4-2, 7.5 mg/ml Lactobacillus Results showed that growth was inhibited.

Example 1: Preparation of an agent for improving vaginitis and vaginal dryness

Based on 100 parts by weight of the natural plant complex extract prepared in Preparation Example 3-1, 4.5 parts by weight of pure platelet-rich plasma, 2.7 parts by weight of hyaluronic acid, 1.2 parts by weight of mandelic acid (antibacterial agent), 4.0 parts by weight of vanillin butyl ether (warming agent). Parts by weight, 0.72 parts by weight of C ₁₂ fatty acid monoglyceride (solubilizer), 4.2 parts by weight of polyethylene glycol with a weight average molecular weight of 300 to 400 (thickener), 0.65 parts by weight of sorbitan monostearate (nonionic surfactant), and purified 18 parts by weight of water was added to the reactor and slowly stirred to prepare a liquid vaginitis and vaginal dryness improvement agent with a gel-like viscosity.

Experimental Example 5: Evaluation of agents for improving vaginitis and vaginal dryness

The vaginitis and vaginal dryness improvement agent prepared in Example 1 was evaluated for improvement in vaginal dryness, improvement in enlargement, improvement in itching, improvement in peculiar odor, and feeling of use in 30 adult women in their 20s to 50s (20s) with vaginitis and/or vaginal dryness. The evaluation was conducted on 5 people (19 people in their 30s, 4 people in their 40s, 2 people in their 50s), and a small amount of vaginitis and vaginal dryness improvement agent was injected into the vagina once a day, and the evaluation results were conducted after 2 weeks of use.

Previously used vaginitis treatments were set at 3 points, and their relative scores were evaluated as very good (5 points), good (4 points), average (3 points), insufficient (2 points), and ineffective (1 point). , the results showing the average values are shown in Table 6 below.

Improvement of vaginal dryness Improvement of hypertrophy Improvement of pruritus Singular odor improvement feeling of use 4.2 4.0 4.0 4.5 4.2

As a result of the post-use evaluation of the vaginitis and vaginal dryness improvement agent, it was evaluated that the improvement effect on vaginal dryness, enlargement, itching, and unusual odor was excellent, and the feeling of use was also good.

Through the above examples and experimental examples, it can be confirmed that the agent for improving vaginitis and vaginal dryness of the present invention has no special side effects, improves enlargement, itching, and specific odor caused by vaginitis, and has an excellent improvement effect on vaginal dryness. there was. Although the above example was carried out only in the form of an intravaginal injection, the vaginitis and vaginal dryness improvement agent of the present invention was processed into various dosage forms such as tablets, ointments, gels, creams, cleansers, water (W) type, oil (O) type, Available in various formulations such as oil-in-water (O/W) type, water-in-oil (W/O) type, silicone (S) type, water-in-silicon (W/S) type, silicone-in-water (S/W) type, solid, and liquid. It can also be manufactured.

Claims (4)

Natural plant complex extract; platelet-rich plasma (PRP); hyaluronic acid; antibacterial agents; warming agent; Solubilizer; thickener; nonionic surfactants; and water;
The natural plant complex extract is a natural plant mixed extract of a mixture containing ratania root, elm bark, lotus root, and Virginia wildflower leaves; and fermented spikenard root extract; at a weight ratio of 1:0.10 to 0.30,
The natural plant mixed extract contains 100 to 150 parts by weight of elm bark, 20 to 50 parts by weight of lotus root, and 20 to 50 parts by weight of Virginia wildflower leaves, based on 100 parts by weight of ratania roots,
The platelet-rich plasma includes pure-platelet-rich plasma, leukocyte-deficient-platelet-rich plasma, or leukocyte-rich-platelet-rich plasma,
The antibacterial agent includes mandelic acid,
The warming agent includes at least one selected from vanillin ethyl ether, vanillin propyl ether, and vanillin butyl ether,
The solubilizer includes C ₆ to C ₁₂ fatty acids or glycols, monoglycerides, diglycerides, or triglyceride esters thereof,
The thickener contains polyethylene glycol with a weight average molecular weight of 200 to 1,000,
The nonionic surfactant is one or more selected from PEG-6 (Polyethylene glycol-6) palmitostearate, ethylene glycol palmitostearate, sorbitan monostearate, lauroyl polyoxyl-6, and polysorbate 80. Includes,
The antibacterial agent is an ingredient that has antibacterial properties against vaginitis-causing bacteria, and the vaginitis-causing bacteria include Enterococcus faecalis, Listeria monocytogenes, Salmonella enteritidis, and Ercinia entero. A composition for improving vaginitis and vaginal dryness, comprising at least one selected from Yersinia enterocolitica.
delete delete An agent for improving vaginitis and vaginal dryness, comprising the composition for improving vaginitis and vaginal dryness of claim 1.