Growth promoter for Döderlein bacilli and selective growth promoter for microorganisms

Abstract

To provide techniques for promoting the growth of Doderlein's bacillus or promoting the growth of Doderlein's bacillus without promoting the growth of Candida fungus.SOLUTION: Provided is a Doderlein's bacillus growth promoter containing glyceryl glucoside as an active ingredient. According to the present invention, the growth of beneficial microorganisms can be promoted without promoting the growth of microorganisms that are useless or harmful to the health of the delicate zone. Therefore, according to the present invention, it is possible to contribute to the acquisition or maintenance of a healthy bacterial count balance in the delicate zone.SELECTED DRAWING: Figure 2

Description

Technical Field

[0001] The present invention relates to a growth promoter for Delftia lineolata containing glyceryl glucoside as an active ingredient and a microorganism selective growth promoter, cosmetics and quasi-drugs for a delicate zone containing the same, and a method for producing cosmetics and quasi-drugs.

Background Art

[0002] The vaginas of healthy women harbor various commensal bacteria, and Lactobacillus spp. account for 75 to 90% of them. Lactobacillus spp. are Gram-positive facultative anaerobic non-spore-forming bacilli that decompose sugar to produce lactic acid. Lactobacillus spp. (Delftia lineolata) that normally inhabit the vagina maintain the vagina at an acidic pH (about 3.5 to 4.5) by producing lactic acid, preventing the invasion and abnormal growth of pathogenic bacteria. In addition, Lactobacillus spp. are suggested to play an important role in normalizing the vaginal environment due to their various abilities to suppress pathogenic bacteria, such as growth inhibition of pathogenic bacteria by bacteriocin production and adhesion inhibition of other bacteria by maintaining proper adhesion to vaginal epithelial cells. Disturbance of the vaginal flora, that is, a decrease in the amount of Delftia lineolata, is known to be a causative factor for infections with specific microorganisms such as vulvovaginal candidiasis and trichomoniasis vaginalis, as well as for bacterial vaginitis, which is a non-specific multiple bacterial infection (Non-Patent Document 1, Non-Patent Document 2).

[0003] On the other hand, Candida is a dimorphic fungus that can take either a yeast form or a filamentous form. It normally inhabits the vaginas of 15 to 20% of non-pregnant women and 20 to 40% of pregnant women. Although its pathogenicity is not high, it causes endogenous opportunistic infections (Candida vaginitis) triggered by factors such as the use of broad-spectrum antibacterial drugs or corticosteroids, diabetes, pregnancy, wearing tight and poorly ventilated underwear, etc. Candida vaginitis presents symptoms such as itching, burning sensation, irritation symptoms, dyspareunia, erythema, swelling, and epidermal exfoliation of the vagina and vulva (Non-Patent Document 3).

Prior Art Documents

Non-Patent Documents

[0004] [Non-Patent Document 1] Matsumoto, Mami et al., Special Feature: Perinatal Medicine and Microbiota - Pregnant Women: Vaginal Microbiota in Japanese Women, Perinatal Medicine, Vol. 50, No. 3, pp. 289-292, 2020. [Non-Patent Document 2] Bacterial Vaginosis, Sexually Transmitted Infections: Diagnostic and Treatment Guidelines 2008, Japanese Society for Sexually Transmitted Infections, pp. 77-80, November 30, 2008. [Non-Patent Document 3] Merck & Co., Inc., Kenilworth, NJ, USA, MSD Manual Professional Edition, 18. Gynecology and Obstetrics, Vaginitis, Cervicitis, and Pelvic Inflammatory Disease (PID), Candida Vaginitis, Last Review / Revision Date: September 2019 [Retrieved September 30, 2021], Internet <URL: https: / / www.msdmanuals.com / ja-jp / %E3%83%97%E3%83%AD%E3%83%95%E3%82%A7%E3%83%83%E3%82%B7%E3%83%A7%E3%83%8A%E3%83%AB / 18-%E5%A9%A6%E4%BA%BA%E7%A7%91%E3%81%8A%E3%82%88%E3%81%B3%E7%94%A3%E7%A7%91 / %E8%85%9F%E7%82%8E%EF%BC%8C%E5%AD%90%E5%AE%AE%E9%A0%B8%E7%AE%A1%E7%82%8E%EF%BC%8C%E3%81%8A%E3%82%88%E3%81%B3%E9%AA%A8%E7%9B%A4%E5%86%85%E7%82%8E%E7%97%87%E6%80%A7%E7%96%BE%E6%82%A3-%EF%BC%88pid%EF%BC%89 / %E3%82%AB%E3%83%B3%E3%82%B8%E3%83%80%E8%85%9F%E7%82%8E> [Overview of the Initiative] [Problems that the invention aims to solve]

[0005] From the above, it can be said that maintaining or increasing the number of Döderlein bacilli while preventing the growth of Candida is desirable for maintaining the health of the vagina, vulva, and surrounding areas (the delicate zone) and for preventing infections. The present invention has been made to solve this problem and aims to provide a technology that promotes the growth of Döderlein bacilli, or promotes the growth of Döderlein bacilli without promoting the growth of Candida. [Means for solving the problem]

[0006] As a result of diligent research, the inventors have found that glyceryl glucoside can promote the growth of Döderlein bacilli while not promoting the growth of Candida fungi. Furthermore, they have found that incorporating glyceryl glucoside into cosmetics and quasi-drugs for the delicate area can be expected to have the effect of maintaining the delicate area in a healthy state. Based on these findings, the inventors have completed the following inventions.

[0007] (1) The Döderlein bacilli growth promoter according to the present invention contains glyceryl glucoside as an active ingredient.

[0008] (2) The microbial selective growth promoter according to the present invention has the property of promoting the growth of Döderlein bacilli but not promoting the growth of Candida, and contains glyceryl glucoside as the active ingredient.

[0009] (3) The agent according to the present invention may be used as a cleansing agent or a topical agent.

[0010] (4) The cosmetic or quasi-drug for the delicate area according to the present invention contains the agent according to the present invention.

[0011] (5) A method for producing a cosmetic or quasi-drug for the delicate zone according to the present invention includes a step of blending the agent according to the present invention as a raw material. [Effects of the Invention]

[0012] According to the present invention, the growth of Döderlein bacilli can be promoted. Furthermore, according to the present invention, the growth of Döderlein bacilli can be promoted without promoting the growth of Candida fungi. In other words, according to the present invention, the growth of beneficial microorganisms can be promoted without promoting the growth of microorganisms that are useless or harmful to the health of the delicate area. Therefore, according to the present invention, it is possible to obtain or maintain a favorable balance of bacterial counts for the health of the delicate area. Furthermore, according to the present invention, it is possible to maintain a normal vaginal microbiota and contribute to the prevention or improvement of various infections.

[0013] Furthermore, glyceryl glucoside is a sugar derivative with a structure in which glucose is bonded to glycerin, and is naturally found in sake and cyanobacteria. It has been used as a cosmetic ingredient for a long time, and since no toxicity has been observed in any of the following tests, including primary skin irritation tests (rabbits), continuous skin irritation tests, skin sensitization tests (guinea pig intradermal maximization method), phototoxicity tests (guinea pigs), photosensitization tests, eye irritation tests (rabbits), mutagenicity tests (in vitro), and human patch tests, it is considered highly safe. Therefore, according to the present invention, the growth of Döderlein bacilli can be promoted without concerns about safety or side effects. [Brief explanation of the drawing]

[0014] [Figure 1] This table shows the composition of the culture medium used in the growth experiment. [Figure 2] The upper figure is a table showing the number of viable Döderlein bacilli cultured in the presence of various concentrations of the test substance (glycerin or glyceryl glucoside). The lower figure is a bar graph representing the same number of viable bacteria. [Figure 3] The upper figure is a table showing the number of viable Candida fungi cultured in the presence of various concentrations of the test substance (glycerin or glyceryl glucoside). The lower figure is a bar graph representing the same number of viable fungi. [Modes for carrying out the invention]

[0015] The present invention will be described in detail below. In this specification, Döderlein bacilli growth promoters and microbial selective growth promoters may be referred to collectively, or to either of them, as "the agent of the present invention" or "this agent."

[0016] In this invention, the term "fungus" is used as a comprehensive concept that includes organisms too small to be seen with the naked eye, such as bacteria and fungi. In other words, in this invention, "fungus" is synonymous with "microorganism."

[0017] "Promoting growth" means increasing the degree of increase in the number of microorganisms (bacteria). Similarly, "growth-promoting effect" refers to the effect of increasing the degree of increase in the number of bacteria. Furthermore, "growth promoter" refers to an agent used to promote the growth of microorganisms.

[0018] In the present invention, whether or not the agent has a growth-promoting effect can be determined by a culture test, as shown in the examples described later. In such a culture test, a sample with glycerin, which is widely used as a moisturizer and lubricant in cosmetics and pharmaceuticals, added is used as a control, and a sample with neither the agent nor glycerin added is used as a control. That is, three samples are prepared in the same type of culture medium: (a) with the agent added, (b) with glycerin added, and (c) with neither the agent nor glycerin added. The same amount of microorganisms are inoculated into these culture media and cultured for a predetermined period, after which the amount of bacteria in the culture media is measured. The amount of bacteria can be measured using known methods such as colony counting, turbidity measurement, dry cell weight method, wet weight method, and real-time PCR method, which can be appropriately selected. If the amount of bacteria in (a) is greater than in either (b) or (c), it can be determined that the growth of the microorganism has been promoted by the agent.

[0019] On the other hand, in the above culture test, if the amount of bacteria in (A) is smaller than that in either (I) or (U), it can be determined that this agent does not promote the growth of the microorganism (does not have a growth-promoting effect). That is, in the present invention, "not promoting growth" includes cases where, even if the degree of increase in the number of bacteria is large when compared only with the case where neither this agent nor glycerin is added, the degree of increase in the number of bacteria is small when compared with the case where glycerin is added.

[0020] "Microorganism-selective growth promoter" refers to an agent having a selective growth-promoting effect on microorganisms. Here, the microorganism-selective growth-promoting effect refers to an effect that promotes the growth of Delftia lineolata and does not promote the growth of Candida.

[0021] In the present invention, "delicate zone" refers to the vagina, vulva or surrounding body parts, the genital area or the groin.

[0022] In this invention, Döderlein's bacilli refer to Lactobacillus species that are naturally present in the vagina. As described above, Döderlein's bacilli are microorganisms that play a beneficial role in the health of the delicate area. Examples of Döderlein bacilli species include Lactobacillus paragasseri, Lactobacillus acidophilus, Lactobacillus casei, Lactobacillus fermenti, Lactobacillus cellobiosus, Lactobacillus crispatus, Lactobacillus iners, Lactobacillus gasseri, Lactobacillus jennsenii, Lactobacillus vaginalis, Lactobacillus reuteri, Lactobacillus rhamnosus, Lactobacillus brevis, and Lactobacillus plantarum (Non-Patent Literature 1, Non-Patent Literature 2, Tomotari Mitsuoka, Problems in the Classification of Lactobacillus, Journal of the Japan Veterinary Medical Association, Vol. 22, pp. 183-189, 1969). When verifying the effects of the present invention, strains of bacteria stored at depositary institutions such as the BioResource Center for Microbial Materials Development (JCM) (3-1-1 Takanodai, Tsukuba City, Ibaraki Prefecture) can be used (e.g., JCM5344; L. paragasseri).

[0023] As mentioned above, Candida is a microorganism that can have harmful effects on the health of the delicate area. When confirming the effects of the present invention, strains preserved at conservation institutions such as the National Institute of Technology and Evaluation (NITE) Biotechnology Center (NBRC) (2-49-10 Nishihara, Shibuya-ku, Tokyo) or the American Type Culture Collection (e.g., NBRC1594, ATCC10231) can be used.

[0024] Glyceryl glucoside (glucosylglycerol) is a sugar derivative having a structure in which one molecule of glucose and one molecule of glycerol are dehydrated and bonded (MW=254). As shown in Formula 1, there are 1-α-glyceryl glucoside, 1-β-glyceryl glucoside, 2-α-glyceryl glucoside, and 2-β-glyceryl glucoside depending on the bonding site and stereochemistry, but in the present invention, any of these may be used, or a mixture of two or more of these may be used. Glyceryl glucoside may be produced by synthesis or by microbial fermentation, or commercially available products may be used for convenience. [Formula 1] JPEG0007850889000001.jpg95165

[0025] This product can be used in the form of cosmetics, pharmaceuticals, quasi-drugs, etc. It may also be used as a raw material for cosmetics, pharmaceuticals, quasi-drugs, etc., in combination with other ingredients.

[0026] This agent exerts its effects by promoting the growth of beneficial Döderlein bacilli among the commensal bacteria of the delicate area, or by promoting the growth of Döderlein bacilli without promoting the growth of harmful Candida fungi, thereby creating a favorable bacterial flora. Therefore, this agent can be used in formulations that act on the delicate area, namely cleansers and topical preparations. Suitable examples of products containing this agent include cosmetics, pharmaceuticals, and quasi-drugs for use on the delicate area.

[0027] Examples of cosmetics, pharmaceuticals, and quasi-drugs for the delicate area include external preparations applied directly to the delicate area by application, patching, or spraying, as well as hygiene products such as cleansers, bath products, disinfectants, and antiseptics. Examples of product formulations include creams, sheets, lotions, emulsions, gels, aerosols, roll-ons, sticks, powders, liquids, foaming liquids, and tablets. Products containing this agent can be manufactured by adding glyceryl glucoside as a raw material to the raw materials commonly used in such products (for example, oils, surfactants, alcohols, preservatives, chelating agents, antioxidants, thickeners, fragrances, antiseptics, etc.) using methods known to those skilled in the art.

[0028] The glyceryl glucoside content can be set appropriately depending on the form and application of the product, but examples include 0.001% by mass or more, 0.002% by mass or more, 0.003% by mass or more, 0.004% by mass or more, 0.005% by mass or more, 0.006% by mass or more, 30% by mass or less, 29% by mass or less, 28% by mass or less, 27% by mass or less, 26% by mass or less, 25% by mass or less, 24% by mass or less, 23% by mass or less, 22% by mass or less, 21% by mass or less, and 20% by mass or less.

[0029] The present invention will be described below based on examples. However, the technical scope of the present invention is not limited to the features shown in these examples. [Examples]

[0030] <Example> Microbial growth activity of glyceryl glucoside (1) Test substance The test substances used were glycerin and glyceryl glucoside. Commercially available products shown in Table 1 were used to formulate each test substance. For glyceryl glucoside, COSARTE-2G (registered trademark) (manufactured by Toyo Sugar Refining Co., Ltd.) was used. [Table 1]

[0031] (2) Test cells and culture medium The test cells and culture media used are those shown in Table 2 below. The composition of the culture media is shown in Figure 1. [Table 2]

[0032] (3) Growth test The test bacteria were cultured on agar plates for 22 to 48 hours. Colonies that appeared were picked and suspended in sterile physiological saline to prepare a bacterial suspension. Liquid culture medium was pre-filled into a test tube, and the bacterial suspension was added to it to prepare a bacterial solution. Meanwhile, the test substance was dissolved in sterile physiological saline to prepare a series of diluted test substance solutions. The dissolved agar plates, bacterial solutions, and test substance solutions were combined and placed in a petri dish to prepare the culture medium. The final concentration of the test substance in the culture medium was set to 0.0067% by mass to 0.95% by mass. The final concentration of the test substance was calculated assuming that the concentrations of each substance in the commercially available products used in the example were 95% by mass for glycerin and 67% by mass for glyceryl glucoside. A culture medium without the test substance solution (final concentration of the test substance 0% by mass) was also prepared as a control sample. After the culture medium was solidified, it was cultured at 37°C (for Döderlein bacilli) or 30°C (for Candida) for 24 hours. Subsequently, the number of colonies was counted to calculate the concentration per culture medium (cfu / mL), which was then used as the viable cell count. The results for Döderlein bacilli are shown in Figure 2, and the results for Candida are shown in Figure 3.

[0033] As shown in Figure 2, the number of viable Döderlein bacilli remained almost constant regardless of the added concentration when using glycerin, whereas with glyceryl glucoside, the number of viable bacteria increased with increasing concentration. Furthermore, when comparing concentrations of similar magnitude (for example, 0.0095% glycerin and 0.0067% glyceryl glucoside, 0.0475% glycerin and 0.0335% glyceryl glucoside, 0.095% glycerin and 0.067% glyceryl glucoside, 0.475% glycerin and 0.335% glyceryl glucoside, and 0.95% glycerin and 0.67% glyceryl glucoside), the number of viable bacteria was higher with glyceryl glucoside even at lower concentrations than with glycerin. From these results, it is clear that glyceryl glucoside can promote the growth of Döderlein bacilli.

[0034] On the other hand, as shown in Figure 3, the number of viable Candida fungi tended to increase with increasing glycerin concentration, whereas with glyceryl glucoside, the number of viable fungi remained almost constant regardless of the concentration. Furthermore, when comparing at similar concentrations, glycerin resulted in a higher number of viable fungi than glyceryl glucoside. From these results, it became clear that glyceryl glucoside hardly promotes the growth of Candida fungi.

Claims

1. A growth promoter for Döderlein bacilli containing glyceryl glucoside as the active ingredient.

2. A microbial selective growth promoter comprising glyceryl glucoside as an active ingredient, wherein the agent promotes the growth of Döderlein bacilli without promoting the growth of Candida albicans.

3. The agent according to claim 1 or claim 2, characterized in that it is used as a cleansing agent or a topical agent.

4. A cosmetic or quasi-drug for the female intimate area, comprising the agent described in any one of claims 1 to 3.

5. A method for producing a cosmetic or quasi-drug for the female intimate area, comprising the step of incorporating the agent described in any one of claims 1 to 3 as a raw material.

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