KR102347233B1 - Method for producing effective microorganisms fermented solution with skin cell protecting effect and reduced odor and effective microorganisms fermentated solution produced by same method - Google Patents

Abstract

The present invention relates to a method for preparing a fermented solution of effective microorganisms (EM) having skin cell-protective activity and reduced malodor, and to a fermented solution of EM prepared thereby. The fermented solution of EM prepared by the preparation method of the present invention has excellent protective activity for damaged skin cells, and obtained good results in sensory testing to determine odor, cleansing power, skin irritation, and alleviation of scalp itching, thereby receiving high consumer preference. Therefore, the fermented solution of EM can be effectively utilized in daily life.

Description

TECHNICAL FIELD The present invention relates to a method for producing an EM fermentation broth having skin cell protective activity and reduced odor, and a method for producing an EM fermentation broth prepared by the method.

The present invention relates to a method for producing an EM fermentation broth having skin cell protective activity and reduced odor, and to an EM fermentation broth prepared by the method.

EM stands for Effective Microorganisms, which means useful microorganisms, and is mainly composed of microorganisms such as yeast, lactic acid bacteria, photosynthetic bacteria, actinomycetes, and filamentous fungi. EM is currently being used usefully in various fields. In agriculture, it is used as an organic fertilizer or organic compost to grow high-quality crops and to prevent pests and diseases by replacing pesticides. In the livestock industry, it reduces the mortality rate by improving the health of livestock by removing odors and improving the environment. In the environmental field, it is used for water purification, odor removal, oil decomposition, metal and food oxidation prevention, and food waste fermentation. In 2003, Japan's 'Ecopure' magazine reported that a total of 116 countries in Asia, Africa, America and Europe were using EM.

Recently, EM-related research has been actively conducted in agriculture, forestry, livestock, fishery, environment, and medicine in Korea, and local governments and individuals are also using EM in real life to see positive effects, but the characteristics of fermented substances Due to the strong odor of , it is difficult to use in daily life such as cosmetics and quasi-drugs.

On the other hand, Korea Patent No. 2074295 discloses 'a method for producing clear EM fermentation stock solution and clear EM fermentation stock solution prepared thereby', and Korean Patent No. 2075842 discloses "hair loss containing EM fermentation broth of herbal medicine as an active ingredient" Although the 'composition for preventing or promoting hair growth' is disclosed, there is no description of the 'method for producing EM fermentation broth having skin cell protection activity and reduced odor and EM fermentation broth prepared by the method' of the present invention.

The present invention has been derived from the above needs, and the present inventors have found that the EM fermentation broth prepared by mixing and culturing Lactobacillus plantarum and Saccharomyces cerevisiae is sodium lauryl. The present invention was completed by confirming that there is a protective activity of human fibroblasts damaged by sodium lauryl sulfate (SLS), there is no odor, there is no odor, there is excellent cleaning power, there is little skin irritation, and that scalp itching is alleviated.

In order to solve the above problems, the present invention is Lactobacillus plantarum ( Lactobacillus plantarum ) After mixing the culture medium and Saccharomyces cerevisiae ) The culture medium at a temperature of 30 ~ 40 ℃ 40 ~ 80 rpm It provides a method for producing an EM fermentation broth having a skin cell protective activity and a reduced odor, comprising the step of culturing for 40 to 56 hours at a stirring speed.

In addition, the present invention provides an EM fermentation broth having skin cell protective activity and reduced odor, prepared by the above method.

The EM fermentation broth prepared by the manufacturing method of the present invention has excellent protective activity for damaged skin cells, and excellent sensory test results for odor, cleansing power, skin irritation, and scalp itch relief. can be utilized.

In order to achieve the object of the present invention, the present invention Lactobacillus plantarum ( Lactobacillus plantarum ) After mixing the culture medium and Saccharomyces cerevisiae ) The culture medium is 40 to 80 at a temperature of 30 to 40 ℃ It provides a method for producing an EM fermentation broth having skin cell protective activity and reduced odor, comprising the step of culturing for 40 to 56 hours at a stirring speed of rpm.

The manufacturing method of the EM fermentation broth according to an embodiment of the present invention, preferably after mixing the Lactobacillus plantarum culture medium and the Saccharomyces cerevisiae culture medium in a volume ratio of 0.5 to 1.5: 0.5 to 1.5, 30 It may include the step of culturing for 40 to 56 hours at a temperature of ~40 ℃ at a stirring speed of 40 to 80 rpm, more preferably the Lactobacillus plantarum culture solution and the Saccharomyces cerevisiae culture solution 1: After mixing in a volume ratio of 1, it may include, but is not limited to, incubating for 48 hours at a temperature of 35°C and a stirring speed of 60 rpm.

In addition, the culture may be used by selecting a culture medium known in the art, preferably PB (preston broth) medium, but is not limited thereto.

In one embodiment of the present invention, the Lactobacillus plantarum may preferably have an accession number of KACC91481P, and the Saccharomyces cerevisiae may preferably have an accession number of KACC93267P. not limited

In one embodiment of the present invention, the method for producing the EM fermentation broth of the present invention is preferably a Lactobacillus plantarum (Accession No.: KACC91481P) culture solution and Saccharomyces cerevisiae (Accession No.: KACC93267P) culture solution of 0.5 After mixing in a volume ratio of ~1.5:0.5~1.5, it can be cultured in PB (preston broth) medium for 40~56 hours at a temperature of 30~40℃ at a stirring speed of 40~80rpm, but is not limited thereto. .

The present invention also provides an EM fermentation broth having skin cell protective activity and reduced odor prepared by the above production method.

The EM fermentation broth prepared according to the production method according to the present invention has protective activity against human fibroblasts damaged by sodium lauryl sulfate (SLS), does not emit odor, has excellent cleaning power, and has little skin irritation; It is characterized by relief of itchy scalp.

In one embodiment of the present invention, the EM fermentation broth is a fermentation stock solution cultured by mixing Lactobacillus plantarum and Saccharomyces cerevisiae, and may be diluted and used according to the user's preference or use.

Hereinafter, the present invention will be described in detail by way of Examples. However, the following examples are merely illustrative of the present invention, and the content of the present invention is not limited thereto.

Preparation Example 1. Preparation of EM fermentation broth

Lactobacillus plantarum ( Lactobacillus plantarum ) M10 strain (Accession No.: KACC91481P) was cultured in MRS medium at 37 ° C. for 2 days _{to prepare OD 600} = 2.0, Saccharomyces cerevisiae NRRL Y -12632 strain (Accession No.: KACC93267P) was cultured in PB (preston broth) medium at 28 ° C. for 1 day _{to prepare an OD 600} = 4.0. Thereafter, the culture medium of the strain was mixed in a volume ratio of 1:1, inoculated into a PB medium, and cultured at 35° C. at a stirring speed of 60 rpm for 48 hours to prepare an EM fermentation broth.

Example 1. Comparison of the number of viable cells in EM fermentation broth

The number of viable cells in the EM fermentation broth of the present invention and the commercially available EM fermentation broth was measured and compared. Company A's EM fermentation broth consisting of lactic acid bacteria and yeast and company B and C company EM fermentation broth consisting of lactic acid bacteria were used.

After sequentially diluting each sample, the number of viable cells of lactic acid bacteria, yeast and other bacteria was measured using 100 μl of the sample diluted ^{10 −4 .} More specifically, each sample was smeared on lactic acid bacteria selection medium (MRS plate medium) and cultured at 37°C for 3 days, spread on yeast selection medium (PDA plate medium) and cultured at 28°C for 3 days, LB plate medium After plated and cultured at 37 ° C. for 2 days, the confirmed colonies were counted to calculate the CFU (colony forming unit) value per 1 ml, respectively.

As a result, it was confirmed that other bacteria were not detected in the EM fermentation broth of the present invention, but other bacteria were detected in the commercial EM fermentation broth (Table 1).

Comparison of viable cells
division live cell count
(ratio) Lactobacillus leaven other bacteria total number of bacteria of the present invention
EM fermentation broth 3.90×10 ⁸ CFU/ml
(69.6%) 1.70×10 ⁸ CFU/ml
(30.4%) - 5.60×10 ⁸ CFU/ml
(100%) Company A
EM fermentation broth 1.30×10 ⁸ CFU/ml
(4.7%) 5.00×10 ⁷ CFU/ml
(1.8%) 2.60×10 ⁹ CFU/ml
(93.5%) 2.78×10 ⁹ CFU/ml
(100%) Company B
EM fermentation broth 8.30×10 ⁸ CFU/ml
(53.2%) - 7.30×10 ⁸ CFU/ml
(46.8%) 1.56×10 ⁹ CFU/ml
(100%) Company C
EM fermentation broth 6.50×10 ⁷ CFU/ml
(63.7%) - 3.70×10 ⁷ CFU/ml
(36.3%) 1.02×10 ⁸ CFU/ml
(100%)

Example 2. Analysis of skin cell protective activity of EM fermentation broth

The degree of inhibition of cytotoxicity by sodium lauryl sulfate (SLS), which is included in detergents, cosmetics, shampoos, etc., which is known to cause skin irritation, was analyzed using the MTT assay method.

Human fibroblasts (Hs68 cells), one of the cells constituting the dermal layer of the skin, were cultured in DMEM (Dulbecco Modified Eagle Medium) medium containing 10% (v/v) FBS (fetal bovine serum) at 37° C., 5% CO ₂ condition. incubated with The cell culture medium was replaced with a fresh medium every 48 hours, and subcultures were performed every 2-3 days. Human fibroblasts were placed in a 96-well plate at a concentration of 1×10 ⁵ cells/ml per well, EM fermentation broth diluted to 0.5% (v/v) and 0.002% (w/v) SLS were simultaneously treated and cultured for 3 days. Thereafter, 100 μg of MTT (3-(4,5-dimethylthiazol)-2,5-diphenyltetrazolium bromide) was added and incubated for 4 hours, followed by centrifugation at 1,000 rpm for 10 minutes, and the supernatant was removed. Then, 150 μl of DMSO (dimethylsulfoxide) was added to dissolve the formazan precipitate generated by the reduction of MTT, and then the absorbance was measured at 540 nm using a microplate reader. Cell viability compared to the SLS-untreated group was expressed as a percentage as the mean value ± standard deviation of three repeated experiments.

As a result, it was confirmed that the survival rate of human fibroblasts in all the EM fermentation broth-treated groups increased compared to the DMEM medium-treated group, and in particular, it was confirmed that the human fibroblast survival rate of the EM fermentation broth-treated group of the present invention was the highest (Table 2).

Based on the above results, it was found that the EM fermentation broth of the present invention was excellent in the protective activity of human fibroblasts damaged by SLS.

Human Fibroblast Viability division Survival rate (%)

SLS processing EM fermentation broth of the present invention 82.54±4.23 Company A's EM fermentation broth 62.27±5.89 Company B's EM fermentation broth 78.14±7.13 Company C EM fermentation broth 69.47±4.57 DMEM medium 32.79±2.15 SLS-free - 100

Example 3. Sensory test for odor of EM fermentation broth

After storing the EM fermentation broth of the present invention and the commercially available EM fermentation broth at room temperature for 6 months, a sensory test was performed on odors by a 5-point scoring method targeting 30 people in their 20s and 30s (1: The smell is very disgusting, 2 : Smell to an unpleasant degree, 3: Smell not to an unpleasant degree, 4: Smell slightly, 5: Smell not at all).

As a result, it was confirmed that the EM fermentation broth of the present invention exhibited a higher score in the sensory test results for odor than the commercially available EM fermentation broth (Table 3).

Sensory test results for odor division stink EM fermentation broth of the present invention 4.12 Company A's EM fermentation broth 2.88 Company B's EM fermentation broth 3.70 Company C EM fermentation broth 3.92

Example 4. Detergency and skin irritation test of EM fermentation broth

After washing the dishes with bare hands using a 1:1 mixture of the EM fermentation broth of the present invention with a commercially available dishwashing detergent, a sensory test for cleaning power and skin irritation was conducted, and 30 people in their 20s and 30s were tested. The subjects were evaluated using a 5-point rating system (1: very bad/severe skin irritation, 2: bad/severe skin irritation, 3: moderate/slight skin irritation, 4: good/no skin irritation, 5: Very good/no skin irritation). A dishwashing detergent in which nothing was mixed and a dishwashing detergent in which a commercially available EM fermented solution was mixed at a ratio of 1:1 were used as Comparative Examples.

As a result, when the EM fermentation broth of the present invention was mixed and used in dishwashing detergent, it was found that the cleaning power was similar to that of dishwashing detergent and skin irritation was weak. On the other hand, when a commercial EM fermentation broth was mixed with dishwashing liquid, skin irritation was weaker than dishwashing detergent, but it was found that the cleaning power was also weak (Table 4).

Sensory test results for cleansing power and skin irritation division detergency skin irritation Dishwashing detergent + EM fermentation broth of the present invention 4.63 4.20 Dishwashing Detergent + Company A's EM Fermented Liquid 3.30 2.82 Dishwashing Detergent + Company B's EM Fermented Liquid 3.82 3.54 Dishwashing Detergent + Company C's EM Fermented Liquid 4.26 3.98 dish detergent 4.76 2.00

Example 5. Scalp itch relief test of EM fermentation broth

After washing the hair once a day for 30 days with a 1:1 mixture of the EM fermentation broth of the present invention with a commercially available general shampoo, a sensory test for scalp itchiness was conducted, and 30 people in their 20s and 30s Subjects were subjected to a 5-point rating system (1: not improved at all, 2: not improved at all, 3: moderate, 4: slightly improved, 5: improved very much). A shampoo in which nothing was mixed and a shampoo in which a commercially available EM fermentation broth was mixed in a 1:1 ratio were used as Comparative Examples.

As a result, when the EM fermentation broth of the present invention was mixed and used in the shampoo, it was confirmed that the sensory test results for scalp itch showed a high score (Table 5).

Sensory test results for scalp itchiness division itchy scalp Shampoo + EM fermentation broth of the present invention 4.52 Shampoo + Company A's EM fermentation broth 2.37 Shampoo + B company EM fermentation broth 3.56 Shampoo + C company EM fermentation broth 2.48 shampoo 1.05

Claims (3)

Accession number KACC91481P Lactobacillus plantarum ( Lactobacillus plantarum ) and Saccharomyces cerevisiae ( Saccharomyces cerevisiae ) culture solution with accession number KACC93267P After mixing the culture solution in a volume ratio of 1:1, at a temperature of 35 ° C. A method for producing an EM fermentation broth having skin cell protective activity and reduced odor, comprising the step of culturing in PB (preston broth) medium for 48 hours at a stirring speed of 60 rpm. delete delete