TWI680772B - A use of lactobacillus plantarum gmnl-6 composition for caring skin - Google Patents

Abstract

The present invention provides the use of a probiotic for preparing a skin health care composition, wherein the probiotic contains Lactobacillus plantarum GMNL-6, and its deposit number is BCRC 910777 or CCTCC M 2017765. The composition can be an external rub, an external medicine, an external care product, or a cosmetic, and can improve the skin condition or aesthetic appearance by promoting collagen secretion or expression of neuroamide synthase.

Description

Use of Lactobacillus plantarum GMNL-6 composition for skin health care

The invention relates to a lactobacillus isolate used in the technical field of skin care.

At present, the market emphasizes that probiotic products for skin use are quite limited. In the retrospective literature, there are two ways for probiotics to affect skin characterization. One is to directly apply probiotics to the affected part, which can affect the local immune response or directly inhibit the growth of skin bad bacteria. ; Second, after oral administration of probiotics to the intestine, it affects the intestinal bacterial phase and immune response, which in turn affects the changes of the remote skin ( Jeong JH et al , 2016 ) , for example, oral probiotics can improve or prevent ectopic Dermatitis (Atopic Dermatitis AD) has been widely used. A retrospective study (Baquerizo Nole KL et al , 2014, Fuchs-Tarlovsky V et al , 2016, Jeong JH et al , 2016) also found that certain lactic acid bacteria use the acetic acid, lactic acid, butanedione, and sphingomyelin produced by Phospholipase, hyaluronic acid or its surface substances... etc., to achieve the effect of promoting skin health. Overall, probiotics for skin mainly have several effects: for example: improving acne (Kim J, Ko Y et a l, 2010), the ability to resist ultraviolet UV light damage (Ra J et al, 2014), promoting wounds Healing effect (Peral MC et al , 2009), anti-melanin formation effect (Chan CF et al , 2014).

In the past, there have been few clinical human trials on the use of probiotics to improve skin health, and most of them are mainly oral. The research on the use of probiotics in external skin dosage forms is quite small. One of them is prepared by the extract of Phellinus rhizogenes. The resulting emulsion has a significant effect on moisturizing sensitive skin after 29 days of wiping (N=66 persons, female) (Gueniche A et al, 2010), and clinical reports on the application of Lactobacillus are still lacking.

The main function of collagen in skin is to provide complete moisture replenishment of skin, increase skin elasticity and surface gloss. Neuroamide is a lipid that naturally exists in the intercellular spaces of the stratum corneum of the body, and is the most important and highest proportion of the stratum corneum lipid. The main function is to maintain the lipid integrity of the stratum corneum in close succession, prevent the evaporation of water, and resist foreign germs and injuries. The invention mainly uses an in vitro model to screen probiotics that stimulate human skin fibroblasts to secrete and produce collagen and synthesize neuroamides, and further uses skin detection instruments to confirm the health promotion effect of probiotic emulsions on the skin.

Because the research on probiotics in the skin care field is not extensive, it is urgent to find probiotics that can be effectively improved. Since collagen and neuroamide have been widely used to discuss the skin condition and its mechanism of action, this experiment used collagen and neuroamide to explore the effects of probiotics on skin health improvement, and plant milk that may have improved skin condition has been identified Lactobacillus plantarum GMNL-6); further test the skin parameters of the subject with VISIA high-order digital skin quality tester.

The purpose of the present invention is to provide a use of a probiotic composition for skin health care, wherein the probiotic comprises Lactobacillus plantarum GMNL-6, and its deposit number is BCRC 910777 or CCTCC M2017765.

To achieve the aforementioned object of the invention, the Lactobacillus plantarum GMNL-6 is a dead cell.

In order to achieve the aforementioned object of the invention, the dead cells are obtained by treating the bacteria-containing liquid under high temperature and high pressure.

To achieve the aforementioned object of the invention, the composition is an external rub, an external pharmaceutical, an external care product, or a cosmetic.

To achieve the aforementioned object of the invention, the composition may further comprise a pharmaceutically or cosmetically acceptable carrier.

In order to achieve the aforementioned object of the invention, the external care product is a lotion, lotion, skin cream, moisturizing liquid, lip balm, cosmetic liquid mask or facial cleanser.

To achieve the aforementioned object of the invention, the skin care refers to improving the skin condition or aesthetic appearance.

To achieve the aforementioned object of the invention, the improvement of the skin condition is through the promotion of collagen secretion or the expression of neuroamide synthase.

In order to achieve the aforementioned object of the invention, the skin health care system is to reduce the number of skin spots, pores, wrinkles, ultraviolet spots, or viologen.

To achieve the aforementioned object of the invention, wherein the improvement of human skin condition or aesthetic appearance includes: (1) treatment, reduction, and/or prevention of fine lines or wrinkles; (2) reduction of skin pore size; (3) improvement of skin thickness and fullness Degree, and/or firmness; (4) improve skin softness and/or softness; (5) improve skin tone, gloss and/or clarity; (6) increase procollagen and/or collagen production; (7) Improve elastin maintenance and remodeling; (8) Improve skin texture and/or promote remodeling; (9) Improve skin barrier repair and/or function; (10) Improve skin contour appearance; (11) Restore skin Luster and/or brightness; (12) Supplement essential nutrients and/or components in the skin; (13) Improve skin appearance that deteriorates due to menopause; (14) Improve skin moisture retention; (15) Increase skin elasticity and /Or elasticity; (16) treatment, reduction, And/or prevent sagging skin; (17) reduce pigment spots; (18) treat, reduce, and/or prevent acne or acne; or any combination thereof.

To achieve the aforementioned object of the invention, each gram of the composition contains 1×10 ⁷ to 1×10 ¹¹ GMNL-6 dead cells.

To achieve the aforementioned object of the invention, the preferred number of dead cells per gram in the composition is 1×10 ⁸ to 1×10 ¹⁰ .

Figure 1A Collagen protein expression

Figure 1B Gene expression of neuroamide synthase

Figure 2 Analysis of the number of spots on the subject by VISIA high-order digital skin quality detector

Figure 3 Analysis of the subject's pore volume with VISIA high-order digital skin quality detector

Figure 4 Analysis of the number of wrinkles on the subject with VISIA high-level digital skin quality detector

Figure 5 Analysis of the number of UV spots on the subject by VISIA high-level digital skin quality detector

Figure 6 Analysis of the amount of purple pigment in the subjects with VISIA high-level digital skin texture detector

All technical and scientific terms described in this specification, unless otherwise defined, are in the field and have the meaning commonly understood by those skilled in the art.

The purpose of the present invention is to provide a use of a probiotic composition for skin care, wherein the probiotic comprises Lactobacillus plantarum GMNL-6, and its deposit number is BCRC 910777 or CCTCC M2017765. In addition, the present invention also relates to a novel use of the composition or the Lactobacillus for skin care. Its mechanism of action is to promote collagen secretion and neuroamide expression to achieve the effect of skin care.

The Lactobacillus isolates also include the progeny of their subculture, or mutant strains, but they still have the same characteristics, genomics, or uses (for the suppression of enteroviruses) as described in the present invention.

The compositions described herein may include, but are not limited to: foods, beverages, health foods, animal water additives, animal feed additives, animal and human medical compositions, food additives, beverage additives, etc. Application form.

The term "pharmaceutically or cosmetically acceptable carrier" means that the substance or composition must be compatible with the other ingredients of the formulation and not harmful to the patient.

The term "Cell Lysate" may be a single extract obtained from a specific extraction step or series of extraction steps, or the extract may also be a combination of several extracts obtained from several independent extraction steps. These combined extracts are therefore also covered by the term "extract".

The pharmaceutically or cosmetically acceptable carrier may include one or more agents selected from the group consisting of solvents, emulsifiers, suspending agents, decomposers, and binders ( binding agent, excipient, stabilizing agent, chelating agent, diluent, gelling agent, preservative, lubricant , Surfactant (surfactant), and other similar or applicable carriers of the present invention.

One or more kinds of dissolution aids, buffers, preservatives, colorants, flavors, flavors, etc. commonly used in the field of formulations can also be appropriately added to the above composition as needed.

In addition, the novel strains discovered in the present invention can also be included in a composition together with other conventional strains.

The composition provided by the present invention and its use for skin health care, its dosage form can be adjusted according to needs, and is not particularly limited, and the external skin dosage form is preferred.

The present invention is exemplified and illustrated by the following examples, but the present invention is not limited by the following examples. The drugs and biological materials used in the present invention are all commercially available, and the following are only examples of available channels.

The present invention is exemplified and illustrated by the following examples, but the present invention is not limited by the following examples.

Select the Lactobacillus plantarum with skin health care effect from the bacterial species library. The deposit number, deposit date and strain name of the lactobacillus are shown in Table 1.

Example 1 GMNL-6 can promote collagen secretion and the expression of neuroamide synthase

1-1 Dead bacteria preparation steps:

Lactobacillus plantarum (GMNL-6) 1 ml MRS broth was inoculated from the freezing tube and incubated at 37°C for 20 hours under aerobic conditions. The next day, take 15μl of overnight culture solution to 1.5ml MRS broth (1% secondary activation), incubate at 37°C for 20 hours under aerobic culture, wash once with PBS, estimate the number of bacteria with OD600nm, and adjust the number of bacteria It is 1×10 ¹⁰ cells/ml, and is autoclaved at 121°C, and is ready for use after 15 minutes.

1-2 Human skin fibroblast (Hs68) cell model:

Steps to analyze collagen in cell culture fluid: Seed Hs68 cells to 6well plate (2×10 ⁵ cells/well), wash twice with PBS overnight, and change to serum free medium (medium), and add 1×10 ⁹ cells/ml GMNL-6 dead bacteria solution, after 24 hours of treatment, collect the supernatant.

After diluting the supernatant 10-fold, analysis was performed using Procollagen type I C-peptide EIA kit (TaKaRa, #MK101).

Steps for analyzing intracellular neuroamide synthase gene (Serine palmitoyltransferase small subunit A, SPTSSA):

Hs68 cells are seeded to 6well plate (2×10 ⁵ /well), washed twice with PBS overnight, and replaced with serum free medium (medium), and added 1×10 ⁹ cells/ml of dead bacteria liquid for treatment After 24 hours, the cell extracts were collected, RNA was extracted and converted into cDNA, and quantitative PCR analysis of the neuroamide synthase gene (SPTSSA) and housekeeping gene (β-actin) was performed.

Experimental results:

Previously with human skin fibroblasts in vitro screening platform model, from dozens of Lactobacillus strains were screened Lactobacillus (L. plantarum) having a skin care effect of GMNL-6, its capacity to promote secretion of collagen. Further in the same mode, the experimental results (Figure 1) found that GMNL-6 has both the ability to promote collagen secretion and the ability to stimulate the production of intracellular neurosynthetase (SPTSSA), which is a nerve in the body. The necessary molecule in the synthesis process of amides, its rise also represents an increase in the production of neuroamides in the body. Neuroamides in skin beauty have the functions of strengthening skin moisturizing, increasing skin defense, and inhibiting the production of tyrosinase.

Example 2: Lactobacillus plantarum GMNL-6 probiotic emulsion has the function of improving the skin quality of the entire face

2-1 Experimental steps:

1. Number of participants: five women (30-40 years old).

2. Evaluation location: Functional Cosmetics Development and Evaluation Research Center of Jianan Pharmaceutical University.

3. Before the test, first clean the face with facial cleanser, perform a full-face skin diagnosis using VISIA high-level digital skin detector, and record the value (day0)

4. Apply test samples after washing your face in the morning and evening every day during the test (except for lotion, other maintenance conditions of the left and right faces are the same):

-Use on the left half of the face: control lotion (matrix)

-Use on the right half of the face: probiotic emulsion (matrix + GMNL-6 dead bacteria)

5. After four weeks of use, perform a full-face skin diagnosis (day 28), first cleanse the face with facial cleanser, then perform a full-face skin diagnosis, and record the value.

2-2 Emulsion ingredients and probiotic concentration:

1. Full composition of emulsion (matrix): Water, Polyglyceryl-3/Methyl glucose/Distearate, GlycerylTrioctanoin, Macadamia Ternifolia Seed Oil, Propanediol, PEG/PPG/Polybutylene Glycol-8/5/3 Glycerin, Cetyl Alcohol, MyristylMyristate, Glycerin, GlycosylTrehalose/Hydrogenated Starch Hydrolysate, Rice Ferment Filtrate(Sake), Saccharomyces/Daylily Flower Ferment Filtrate, Bis-PEG/PPG-20/15 PEG/PPG-20/5Dimethicone/Methoxy PEG/PPG-25/4 DimethiconeCaprylic/Capric Triglyceride, Sodium Polyacrylate, Phenoxyethanol, o-Cymen-5-ol, Xanthan Gum, Potassium Hydroxide.

2. GMNL-6 concentration in emulsion: 1×10 ⁹ cells/g emulsion

2-3 Evaluation method:

1. Use VISIA high-level digital skin quality detector for skin quality detection, take a high-resolution photo of the face with a 12-megapixel camera and three different light sources (general light, UV light, polarized light), and then use the software to analyze the face fixation The range of skin quality can accurately obtain skin parameters such as spots, pores, wrinkles, texture, purple spots and purple matter to evaluate the underlying condition of the skin basal layer. In order to understand the skin problems, we can evaluate the data changes before and after using the lotion. The test results are compared with a qualified normal range data based on gender and age, which can be used to evaluate the effectiveness of the product.

2. Evaluate each person's own left face (control lotion) and right face (probiotics) through skin quality parameters (including: spots, pores, wrinkles, purple spots, and purple texture) measured by the full-face skin quality diagnostic instrument Emulsion) The difference in scores before use (day0) and after use (day 28). The calculation formula of the score change (%) is as follows:

Experimental results:

Five subjects (NO.1~5) used the control lotion (left face) and probiotics respectively Emulsion (right face), 28 days later, the skin parameters of five subjects were analyzed by VISIA high-level digital skin tester, including: spots (Figure 2), pores (Figure 3), wrinkles (Figure 4), purple spots ( Figure 5) and Violet (Figure 6). After comparing each person's left face with each other, the use of probiotic lotion group can be evaluated to obtain better skin quality improvement data. At least three or more people improved in each evaluation item, especially spots and Pores, up to 80~100%, have been significantly improved. Therefore, in general, probiotics have the effect of improving skin texture, especially in lightening spots and reducing pores.

Collagen and neuroamide in the skin will gradually lose with the increase of age or the influence of unhealthy living habits. Therefore, supplement and restore the content of collagen and neuroamide in the skin, which is good for skin health, anti-wrinkle and anti-UV Damage and anti-aging are very important. The evaluation of this patent found through in vitro experiments that Lactobacillus plantarum GMNL-6 can simultaneously promote collagen secretion and the expression of neuroamide synthase, which represents that GMNL-6 has a special skin effect. Observed by human clinical experiments through VISIA high-level digital skin quality tester, Lactobacillus plantarum GMNL-6 probiotic emulsion has the function of improving the skin quality of the entire face, including: spots, pores, wrinkles, purple spots and purple texture, at least three people in each item The above is improved, especially to lighten the spots and reduce the pores, up to 80-100% of people are effective; in addition, the reduction of porphyrin indicates that the growth of acne bacilli is inhibited, which should help improve acne and acne. The clinical application of probiotics in human skin is mostly oral, and clinical trials of directly adding dead bacteria of Lactobacillus to emulsion are few, and the evaluation method of comparing their left and right faces at the same time is used to reduce interference with other skin care products and other individual differences. Cause of change. And five people who used GMNL-6 probiotic emulsion did not show any symptoms of allergy and discomfort after using it for four weeks. Therefore, the use of Lactobacillus plantarum (GMNL-6) for skin care products is quite safe and has no side effects. It should be a better choice for promoting skin health and preventing skin aging in the future.

The above-mentioned multiple functions are in full compliance with the statutory invention patent requirements for novelty and progress. You must file an application in accordance with the law and urge your office to approve this application for an invention patent to encourage invention.

【Biological Material Storage】

Domestic storage information [Please note in order of storage institution, date, number]

Biological Resources Preservation and Research Center of Hsinchu Food Industry Development Institute, the deposit date is April 14, 2017, and the deposit number is BCRC 910777.

Overseas hosting information [Please note in order of hosting country, institution, date, number]

China, deposit date is November 3, 2017, deposit number is CCTCC M 2017765

Claims (8)

An external composition with functions of lightening skin spots and reducing pores, wherein the external composition comprises Lactobacillus plantarum GMNL-6, and its registration number is BCRC 910777 or CCTCC M2017765; wherein the Lactobacillus plantarum GMNL- 6 is dead cells obtained by high temperature and high pressure treatment. A use of the composition for external use as described in item 1 of the patent application scope for skin health care, wherein the Lactobacillus plantarum GMNL-6 is a dead cell obtained by high temperature and high pressure treatment; wherein the skin health care is reduced The number of skin spots, pores, ultraviolet spots, or purple matter. The composition for external use as described in item 1 of the scope of the patent application, wherein the composition for external use is an external rub, a pharmaceutical for external use, a cosmetic for external use, or a cosmetic. The composition for external use as described in item 1 of the scope of the patent application, wherein the composition for external use may further comprise a pharmaceutically or cosmetically acceptable carrier. The external use composition as described in item 3 of the patent application range, wherein the external care product is a lotion, lotion, skin cream, moisturizing liquid, lip balm, cosmetic liquid mask or facial cleanser. The use as described in item 2 of the patent application scope, wherein the skin health care refers to the promotion of skin collagen secretion or the expression of neuroamide synthase. The use as described in item 2 of the patent application scope, wherein the external composition contains 1×10 ⁷ to 1×10 ¹¹ GMNL-6 dead cells per gram. The use as described in item 2 of the patent application scope, wherein the number of dead cells per gram in the external composition is 1×10 ⁸ to 1×10 ¹⁰ .

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