KR101778853B1

KR101778853B1 - Preparation method of osmotic enzyme fermentation product using mushroom, the fermentation product prepared thereby and cosmetic, food or pharmaceutical composition comprising the same

Info

Publication number: KR101778853B1
Application number: KR1020150136956A
Authority: KR
Inventors: 옥민; 윤경훈; 박진철; 이성은; 성혜영; 이진아; 홍우종; 박성수; 강민정; 박노환; 김현성
Original assignee: (주)엠앤씨생명과학; 옥민
Priority date: 2015-09-25
Filing date: 2015-09-25
Publication date: 2017-09-14
Also published as: KR20170037397A

Abstract

The present invention relates to a method for producing a fermented osmotic enzyme using mushroom, a fermented product prepared from the fermented product, and a cosmetic, food or pharmaceutical composition containing the same.

Description

FIELD OF THE INVENTION [0001] The present invention relates to a method for producing fermented osmotic enzyme using mushroom, a fermented product prepared from the fermented product, and a cosmetic, food or pharmaceutical composition containing the same. BACKGROUND ART [0002] THE SAME}

Mushrooms are a combination of basidiomycetes and higher fungi of the carrot fungus. It grows mainly in shady land or decayed trees, and grows in horses. There are edible mushrooms and poisonous mushrooms in mushrooms, and mushrooms such as matsutake mushroom, mushroom mushroom, shiitake mushroom, mushroom mushroom, mushroom mushroom, and mushroom mushroom.

Mushrooms are abundant in dietary fiber to excrete toxins and waste products in the intestines, cleansing blood, enhancing immune function, and preventing infection and cancer.

The mushrooms having various effects can be used as foods, drinks, liquors, etc. as they are ingested or processed as they are, and in order to improve the efficacy of the mushrooms, the effective deliveries can be extracted and used. In this case, the extraction method such as hot water extraction, pressure extraction, solvent extraction, alcohol fermentation, lactic acid fermentation, and methane fermentation can be used as a method of extracting the active ingredient, and an extract containing an active ingredient of the substrate mushroom Can be prepared.

However, when hot water extraction or pressurized extraction is used, not only the active ingredient of the substrate is destroyed by high heat and pressure but also it may be difficult to exhibit the desired effect by denaturation. In case of using solvent extraction, There is a problem that it may be harmful to the human body due to a non-polar solvent, an organic solvent, etc. remaining in the extract.

Patent Publication No. 2012-0087408 discloses a mushroom fermentation extract obtained by fermentation using microorganisms and a cosmetic composition containing the same. However, when fermenting using microorganisms, it is difficult to keep conditions such as temperature, humidity, and oxygen concentration suitable for growing inoculated microorganisms constant, and desired fermentation may not occur due to contamination of other microorganisms not inoculated.

As a method for improving the shortcomings of the extraction method and the fermentation method, a direct method can be used. In the current method, a high concentration of sugar is added to elute an active ingredient together with moisture in a substrate by an osmotic pressure phenomenon. Although the active ingredient can be easily obtained while reducing the loss of the active ingredient by heat or pressure by the direct method, the immediate method does not require a long time to obtain the extract compared with the extraction method and the fermentation method, but also depends on the state and kind of the microorganism Repeatedly it may be difficult to obtain a uniform extract.

In order to solve such a problem, a method of fermenting microorganisms by inoculating the extract extracted by the conventional method is also known. However, there remains a need for a method for rapidly and uniformly obtaining an extract containing an active ingredient in a substrate.

Patent Publication No. 2012-0087408

In order to solve the above problems, the present invention provides a process for producing an osmotic enzyme fermented product capable of uniformly extracting an active ingredient of mushroom without loss by heat, a fermented product produced from the osmotic enzyme, and a cosmetic, The purpose is to provide.

(Ii) a sugar and (iii) a saccharomyces cerevisiae strain as a yeast, wherein the yeast is selected from the group consisting of: (i) at least one mushroom selected from the group consisting of a flowering mushroom, a mushroom, a mushroom, a mushroom, Preparing a first fermentation product by firstly fermenting a mixture of Saccharomyces cerevisiae and Lactobacillus fermentum at 20 to 50 ° C; (b) removing the solid content from the primary fermentation product and then aging at 0 to 10 < 0 > C, to prepare a fermented product of osmotic enzyme using mushroom.

(I) one or more mushrooms selected from the group consisting of flowering mushrooms, mushrooms, mushrooms, mushrooms, mushrooms, (ii) sugars and (iii) saccharomyces cerevisiae as yeast Preparing a first fermentation product by firstly fermenting a mixture of Saccharomyces cerevisiae and Lactobacillus fermentum at 20 to 50 ° C; (b) removing the solid content from the primary fermentation product and then performing secondary fermentation at 20 to 50 ° C to produce a secondary fermentation product; (c) removing the solid content from the secondary fermentation and then aging the fermented product at 0 to 10 < 0 > C.

(I) one or more mushrooms selected from the group consisting of flowering mushroom, mushroom mushroom, mushroom mushroom, and mushroom mushroom as a substrate, (ii) sugar, and (iii) saccharomyces as a yeast Preparing a first fermentation product by firstly fermenting a mixture of Saccharomyces cerevisiae and Lactobacillus fermentum at 20 to 50 캜; (b-1) removing the solid content from the primary fermentation product and sterilizing the product at 100 to 140 캜; (ii) Saccharomyces cerevisiae as a yeast and Lactobacillus fermentum as a lactic acid bacterium are added to the sterilized primary fermentation product (b-2) Secondary fermentation at 20 to 50 캜 to prepare a secondary fermentation product; And (c) removing the solid content from the secondary fermentation and then aging the fermented product at 0 to 10 < 0 > C.

Also, there is provided an osmotic enzyme fermented product prepared by the above-mentioned method.

Further, there is provided a cosmetic, food or pharmaceutical composition comprising the osmotic enzyme fermented product as an active ingredient.

According to the production method of the present invention, the osmotic enzyme fermented product containing the active ingredient of the mushroom uniformly can be produced by extracting the active ingredient of the mushroom without loss of the active ingredient by heat. Such an osmotic enzyme fermented product has excellent effects such as anti-inflammation, anti-allergy, skin regeneration, and antioxidation and can be used as a cosmetic, food or pharmaceutical composition.

FIG. 1 is a graph showing the inhibitory effect of NO production on osmotic enzyme fermentation according to an example of the present invention.
FIG. 2 is a graph illustrating inhibition of NO production according to the concentration of osmotic fermentation product according to an embodiment of the present invention.
FIG. 3 is a graph showing the inhibition effect of hexosaminidase secretion of the osmotic enzyme fermented according to an example of the present invention.
FIG. 4 is a graph illustrating inhibition of secretion of hexosaminidase according to the concentration of osmotic fermentation product according to an embodiment of the present invention.
FIG. 5 is a graph showing the recovery ability of the osmotic enzyme fermented product after UV irradiation according to an example of the present invention.
6 is a graph showing the antioxidant ability of the osmotic enzyme fermented product according to an example of the present invention.

BEST MODE FOR CARRYING OUT THE INVENTION Hereinafter, a method for preparing a fermented osmotic enzyme product using mushroom according to the present invention, a fermented product prepared from the fermented product, and a cosmetic, food or pharmaceutical composition containing the fermented product will be described in detail with reference to the accompanying drawings. However, these descriptions are provided only to illustrate the present invention, and the scope of the present invention is not limited by these exemplary explanations.

&Lt; Production method of osmotic enzyme fermentation product >

Osmotic enzyme fermentation water production method according to an embodiment of the present invention includes (a) (i) as a substrate mushrooms, (ii) sugar and (iii) a yeast MY process three Levy Jia a saccharide as (Saccharomyces cerevisiae) and lactic acid bacteria Lactobacillus Preparing a first fermentation product by first fermenting a mixture obtained by mixing Lactobacillus fermentum at 20 to 50 캜; (b) aging at 0 to 10 < 0 > C after removing the solid content from the primary fermentation product.

Hereinafter, the manufacturing method will be described separately for each step.

(a) a primary fermentation step

(a) is a step of firstly fermenting a mixture of a substrate (mushroom), sugar and a microorganism (yeast, lactic acid bacteria) at 20 to 50 ° C to prepare a primary fermentation product.

As a substrate, at least one mushroom selected from the group consisting of mushroom, mushroom, mushroom, mushroom, and mushroom is used, and the kind, origin and form thereof are not particularly limited.

Mushrooms can use outposts that include leaves, flowers, stems, roots, and the like. At this time, the substrate may be pulverized to a predetermined size or may be in the form of a juice using a presser to easily extract the active ingredient.

As the saccharide, there can be used any of conventional saccharides known in the art without limitation, and at least one selected from the group consisting of white sugar, sulfur sugar and raw sugar (non-saccharified sugar) can be used. As the sugar, it is preferable to use a yellow sugar or a raw sugar, and it is more preferable to use a raw sugar.

Saccharomyces cerevisiae as a yeast and Lactobacillus fermentum as a lactic acid bacterium are all used as the microorganisms to be inoculated, and yeast fermentation and lactic acid fermentation can proceed simultaneously. Saccharomyces cerevisiae MAB Y1 (KCTC 11386BP), Saccharomyces cerevisiae (KCTC 7904) can be used as the yeast, and saccharomyces cerevisiae It is preferable to use Saccharomyces cerevisiae MAB Y1 (KCTC 11386BP) . Lactobacillus fermentum Miev L1106 (KCTC 12082BP) and Lactobacillus fermentum (KCTC 3112) can be used as the lactic acid bacteria, and Lactobacillus fermentum Miev L1106 KCTC 12082BP) can be used. The microorganisms may be mixed in a ratio of yeast: lactic acid bacterium = 1: 0.5 to 2 based on the weight ratio, and the yeast and lactic acid bacteria are preferably mixed in a ratio of 1: 1.

When the substrate, the sugar and the microorganism are mixed, it is possible to extract the active ingredient of the substrate to a maximum extent by controlling the mixing ratio or the amount used. The mixing ratio of the substrate to the saccharide is 1: 0.5 to 2, preferably 1: 1 to 1: 1, based on the weight ratio. The amount of the microorganism to be used is 1 to 10% by weight, preferably 3 to 5% by weight, based on the total weight of the mixture of the substrate and the sugar.

A mixture of such a substrate, a sugar and a microorganism is prepared by primary fermentation in an incubator having appropriate temperature and aerobic conditions, and a primary fermentation product in which the active ingredient is eluted together with the moisture of the substrate.

The primary fermentation temperature is not particularly limited, but is 20 to 50 캜, preferably 25 to 45 캜.

At this time, when the mixture is fermented under anaerobic conditions in which oxygen is intercepted, alcohol is generated upon saccharification by the yeast to inhibit the enzyme activity of the microorganism, so that fermentation proceeds under aerobic conditions in which oxygen is supplied. For example, the nonwoven fabric is used to prevent the anaerobic condition by blocking the inlet of the fermentation vessel to only pass oxygen.

During the primary fermentation, the pH of the primary fermentation product is measured at regular intervals to check for contamination, and the bubbles accumulated in the upper layer of the liquid phase are removed. BCA assay, The amount of enzyme produced is estimated by protein quantification method such as Bradford assay.

The pH of the primary fermentation product is preferably 3 to 6, and if it is out of the above range, anaerobic fermentation by yeast may occur or contamination may be caused by external microorganisms other than the inoculated microorganisms. When the protein amount of the primary fermentation product is 400 to 1000 ug / ml, the enzyme is sufficiently generated, and the primary fermentation is terminated. At this time, the primary fermentation period is not particularly limited, but it may be fermented for 4 to 10 days.

(b)

Step (b) is a step of removing the solid content from the primary fermentation and aging at 0 to 10 < 0 > C.

The solid content is removed from the primary fermentation product prepared in step (a) for aging. The primary fermentation broth in which the solid content has been removed is aged. This primary fermentation broth is kept at low temperature and aged to make the components contained in the primary fermentation broth interact with each other while stopping the fermentation by the microorganism, thereby producing a final osmotic enzyme fermented product. The aging temperature is preferably 0 to 10 ° C.

Meanwhile, according to another embodiment of the present invention, after the step (a) and the step (b), the solid matter is removed from the primary fermentation product, and then the secondary fermentation product is produced by secondary fermentation at 20 to 50 ° C . &Lt; / RTI > In this case, aging proceeds after removing the solid content from the secondary fermentation product.

Meanwhile, a method for producing an osmotic enzyme fermented product according to another embodiment of the present invention comprises the steps of (a) (i) a mushroom as a substrate, (ii) sugar, and (iii) Saccharomyces cerevisiae and lactic acid bacteria Preparing a first fermentation product by first fermenting a mixture of Lactobacillus fermentum at 20 to 50 캜; (b-1) removing the solid content from the primary fermentation product and sterilizing the product at 100 to 140 캜; (ii) Saccharomyces cerevisiae as a yeast and Lactobacillus fermentum as a lactic acid bacterium are added to the sterilized primary fermentation product (b-2) Secondary fermentation at 20 to 50 캜 to prepare a secondary fermentation product; And (c) aging at 0 to 10 < 0 > C after removing the solid content from the secondary fermentation product.

Hereinafter, the manufacturing method will be described separately for each process step as follows.

(a) a primary fermentation step

(a) is a step of firstly fermenting a mixture of a substrate (mushroom), sugar and a microorganism (yeast, lactic acid bacteria) at 20 to 50 ° C to prepare a primary fermentation product. The step (a) is the same as the step (a) of the method for preparing the osmotic fermentation product according to the example of the present invention, and the first fermentation product containing the active ingredient of the substrate and the enzyme of the microorganism is prepared.

(b-1) Sterilization step

The step (b-1) is a step of removing the solid content from the primary fermentation product and sterilizing the product at 100 to 140 ° C. The primary fermentation product prepared in the step (a) is sterilized by removing the solid content using a mesh and then sterilizing the remaining primary fermentation broth. The sterilization conditions are not particularly limited, but the temperature may be 100 to 140 캜, and the time may be 5 to 30 minutes. As a result, secondary fermentation can be performed by re-inoculating microorganisms with 20 to 50% of the primary fermentation product to the substrate. At this time, the dead microorganism contained therein can be supplied with the protein source upon re-fermentation.

(b-2) Secondary fermentation product preparation step

The step (b-2) is a step of adding a saccharide and a microorganism (yeast, lactic acid bacteria) to the sterilized primary fermentation product and then performing a secondary fermentation at 20 to 50 ° C to produce a secondary fermentation product. The primary fermented product sterilized in the step (b-1) is added with sugars and microorganisms for secondary fermentation.

The sugars and microorganisms are the same as the sugars and microorganisms used in step (a) above.

When the primary fermentation product, the sugar and the microorganism are mixed, the mixing ratio or amount of the primary fermentation product, sugar, and microorganism may be adjusted to improve the reaction between the active ingredient contained in the primary fermentation product and the enzyme produced from the microorganism. The mixing ratio of the primary fermentation product to the sugar is 1: 0.5 to 2, preferably 1: 1 to 1: 1, based on the weight ratio. The amount of the microorganism to be used is 1 to 10% by weight, preferably 3 to 5% by weight, based on the total weight of the mixture of the primary fermentation product and the saccharide.

The mixture of the sterilized primary fermentation product, saccharide and microorganism is subjected to secondary fermentation under the same temperature and aerobic condition as the primary fermentation in the step (a), whereby the added microorganism is mixed with the active ingredient In addition, a dead fermented microorganism is used as a protein source to produce a secondary fermentation product by an enzymatic reaction.

(c) Aging step

(c) is a step of removing the solid content from the secondary fermentation and aging at 0 to 10 < 0 > C. The step (c) is the same as the step of aging the osmotic enzyme fermentation product according to the example of the present invention, and a final osmotic enzyme fermentation product is prepared.

As described above, the method for producing the osmotic enzyme fermented product of the present invention can rapidly and uniformly extract the active ingredient of the mushroom without loss.

<Osmotic enzyme fermented product>

The osmotic enzyme fermented product produced by the production method of the present invention contains an active ingredient of mushroom, and when used, it can exhibit the effect of mushroom.

&Lt; Cosmetic material, food, pharmaceutical composition containing osmotic enzyme fermented product >

The osmotic enzyme fermented product according to the present invention can be used as a cosmetic, food or pharmaceutical composition containing an osmotic enzyme fermented product of mushroom as an effective ingredient derived from an organism and safe for human body. At this time, the compositions may be effective for anti-inflammation, anti-allergy, skin regeneration and antioxidation by increasing immunity and regenerating function.

Specifically, the cosmetic composition containing the osmotic enzyme fermented product may be used in the form of a solution, a suspension, an emulsion, a paste, a gel, a cream, a lotion, a powder, an oil, a powder, an aerosol, Additives usually added may be added. The cosmetic composition may be used as a cosmetic composition for hair such as a shampoo, a rinse, a tonic, a hair conditioner and a hair essence or may be used as a cosmetic composition for a face or wrist, such as a body shower, a body lotion, a body oil, a body mist, a foundation, a cleanser, have.

In addition, the food composition containing the osmotic enzyme fermented product may be used in the form of various foods, beverages, gums, tea, vitamin complex, functional beverage, health supplement and the like. .

In addition, the pharmaceutical composition containing the osmotic enzyme fermented product can be used in the form of an oral preparation, a granule, a tablet, a capsule, a suspension, an emulsion, a syrup or an aerosol, an external preparation, a suppository, Additives usually added during manufacture may be added.

Hereinafter, the present invention will be described concretely with reference to Examples. However, the following Examples are intended to illustrate one embodiment of the present invention, but the scope of the present invention is not limited by the following Examples.

[Example 1]

As a substrate, the flower mushroom was washed with flowing water, and the water was completely removed and then cut into 2 cm in length, length and height. The prepared substrate and sugar were mixed in a weight ratio of 1: 1. Yeast and lactic acid bacteria were inoculated into the mixture in an amount of 5% by weight based on the total weight of the mixture of substrate and sugar. The raw materials and contents used in the following Table 1 are described.

mixture content temperament Flower mushroom 50 wt% Party Natural sugar (raw sugar) 50 wt% leaven Saccharomyces cerevisiae MABY1 (KCTC 11386BP) 5 wt% based on substrate and sugar mixture Lactobacillus Lactobacillus fermentum MieVL1106 (KCTC 12082BP) 5 wt% based on substrate and sugar mixture

The mixture was placed in a fermentation vessel, and the inlet of the fermentation vessel was blocked with a nonwoven fabric to barely pass oxygen, followed by primary fermentation in a 30 ° C incubator. The first fermentation start time was regarded as a sample on the 0th day, and the primary fermentation product was sampled at intervals of 24 hours to measure the pH, and the amount of enzyme was measured by the protein determination method (BCA assay). Bubbles formed on the surface of the primary fermentation product during the primary fermentation were removed. Approximately 7 days after confirming that the amount of enzyme in the first fermentation product was 800 ug / ml, the solid content of the first fermentation product was removed using a fine mesh (300 mesh).

The primary fermentation broth in which the solid content was removed was placed in a sterilized new container and aged at 4 캜 to prepare a fermented product of a flowering oyster mushroom osmotic enzyme.

[Examples 2 to 4]

The osmotic enzyme fermented product was prepared in the same manner as in Example 1, except that the raw mushroom, mushroom mushroom, and mushroom mushroom were used as a substrate instead of the flowering mushroom. Thus, according to Example 2, the osmotic enzyme fermented product of Pleurotus ostreatus was obtained by Example 3, the osmotic enzyme fermented product of Mushroom mushroom was produced by Example 3, and the osmotic enzyme fermented product was obtained by Example 4.

[Experimental Example 1] Anti-inflammatory effect

(1) NO production inhibitory effect

RAW 264.7 cells were cultured in 96 well microplates (1 × 10 ⁴ cells / well) for 24 hours. The culture was carried out at 37 ° C and 5% CO 2 using high glucose DMEM (Lonza, USA) containing 1% penicillin-streptomycin and 10% fetal bovine serum (FBS)

The oocyte enzyme fermented product of Example 1 in Example 1, the osmotic enzyme fermented product in Example 2, the osmotic enzyme fermented product in Example 2, the osmotic enzyme fermented product in Example 3, the osmotic enzyme of the mushroom of Example 4, Each fermented product was treated with 0.25% of each, cultured at 37 ° C in 5% CO 2 for 1 hour, treated with 1 μg / mL of LPS, and further cultured for 18 hours.

To 50 μl of the cell culture medium, 50 μl of a Griess reagent (Sigma chemical Co., USA) containing 1% sulfanilamide, 5% phosphoric acid and 0.1% naphthylethylethylene diamine was added Respectively. After incubation for 10 min, the absorbance was measured at 540 nm using an absorbance meter (TECAN / infinite M200, Switzerland). The absorbance values when treated with each sample were compared with the absorbance values obtained by treating only LPS, and the degree of NO production was expressed as a percentage, and the results are shown in Fig.

(2) Inhibitory effect of NO production on concentration

The oocyte enzyme fermented product of Example 1 in Example 1, the osmotic enzyme fermented product in Example 2, the osmotic enzyme fermented product in Example 2, the osmotic enzyme fermented product in Example 3, the osmotic enzyme of the mushroom of Example 4, The fermented broth was treated with 0.1%, 0.25%, 0.5% and 1% of each concentration. The cells were incubated at 37 ° C and 5% CO 2 for 1 hour and then treated with 1 μg / ml of LPS for 18 hours.

To 50 μl of the cell culture medium, 50 μl of a Griess reagent (Sigma chemical Co., USA) containing 1% sulfanilamide, 5% phosphoric acid and 0.1% naphthylethylethylene diamine was added Respectively. After incubation for 10 min, the absorbance was measured at 540 nm using an absorbance meter (TECAN / infinite M200, Switzerland). The absorbance value of each sample treated with the concentration was compared with the absorbance value obtained when only LPS was treated, and the degree of NO production was shown as a percentage, and the results are shown in FIG.

[Experimental Example 2] Hexosaminidase inhibitory effect

RBL-2H3 (rat basophilic leukemia cell line) was distributed from ATCC. The culture was carried out at 37 ° C and 5% CO 2 using glucose medium (Lonza, USA) containing 1% penicillin-streptomycin and 10% heat-inactivated fetal bovine serum (FBS) or gentamicin. To overcome the overcorrection caused by cell proliferation, 0.05% trypsin-EDTA solution was treated to float the cells and then passaged.

The osmotic pressure fermented product of Example 1, the osmotic pressure fermentation product of Example 2, the mushroom fermentation product of Example 3, and the osmotic fermentation product of the mushroom of Example 4 were added to the cell culture solution in an amount of 0.25 %, And Hexosaminidase secretion inhibition was measured. Specifically, the absorbance value when the osmotic fermentation product of each of the examples was treated was compared with the absorbance value when only anti-DNP IgE and DNP-BSA were treated, and the degree of hexosaminidase secretion was expressed as a percentage, Is shown in Fig. In addition, each osmotic fermentation product was treated with an amount ranging from 0.1% to 25%, and the inhibitory effect of hexosaminidase secretion was measured. The results are shown in FIG.

[Experimental Example 3] Recovery effect after UV irradiation

Human Dermal Fibroblasts, which are normal human dermal fibroblasts (NHDFs), were distributed (Lonza, USA), and then NHDF (5 × 10 ⁴ cells / well) was added to the wells of a 24-well plate Lt; / RTI > for 24 hours. The culture was performed at 37 ° C and 5% CO 2 using low glucose DMEM (Lonza, USA) containing 1% penicillin-streptomycin and 10% fetal bovine serum (FBS)

The osmotic pressure fermented product of the flowering oyster mushroom of Example 1, the osmotic pressure fermentation product of the mushroom of Example 2, the osmotic pressure fermentation product of the mushroom mushroom of Example 3, and the osmotic fermentation product of the mushroom of Example 4 were treated at a concentration of 37 , 5% CO2 for 4 hours, washed twice with PBS, and irradiated with UV-B at 100 mJ / cm2. The osmotic pressure fermented product of the flower mushroom of Example 1, the osmotic pressure fermentation product of the mushroom of Example 2, the osmotic pressure fermentation product of the mushroom mushroom of Example 3 and the osmotic fermentation product of the mushroom of Example 4 were treated at a concentration of 0.25% (MTT, Sigma) at a concentration of 1 mg / mL, and further cultured for 4 hours. 100 μl of dimethylsulfoxide (DMSO, Sigma) or 0.04 N HCl / Isopropanol was added to dissolve the formazan precipitate produced by the reduction of MTT and the absorbance was measured at 570 nm using a microplate reader. The average absorbance of each sample group was calculated and compared with the absorbance value of the control group (when UV was not irradiated), and the cell survival rate was expressed as a percentage. The results are shown in Fig.

[Experimental Example 4] Antioxidant effect

The secretion of β-hexosaminidase was measured in order to examine the inhibitory effect on degranulation, which is an index of allergic reaction. RBL-2H3 cells were suspended in DMEM containing 10% FBS and incubated in a 48-well plate (Corning, USA) at 5 × 10 ⁵ cells / ml for the measurement of β-hexosaminidase secretion in the antigen- Lt; / RTI > The cells were then sensitized with anti-DNP IgE (0.5 μg / ml) and cultured in a 37 ° C 5% CO2 incubator for 12-16 hours. Cells of each well were washed twice with Siraganian buffer (119 mM NaCl, 5 mM KCl, 0.4 mM MgCl2, 25 mM PIPES, 40 mM NaOH, pH 7.2), and then 5.6 mM glucose, 1 mM CaCl2 and 0.1% BSA Siraganian buffer was added and the fermented extracts were incubated for 1 hour at 37 ° C in a 5% CO 2 incubator. The cells were treated with DNP-BSA (20 μg / ml) for 1 hour and left in the ice bath for 10 minutes The post-reaction was terminated. 20 μl of the supernatant was transferred to a 96-well plate and 20 μl of substrate buffer (1 mM p-nitrophenyl-N-acetyl-bD-glucosaminide, 0.05 mM sodium citrate, pH 4.5) was added and incubated at 37 ° C for 30 min. The reaction was terminated by the addition of 200 μl stop solution (0.1 M Na ₂ CO ₃ / NaHCO ₃ ) per well. Absorbance was measured at 405 nm using a microplate reader (TECAN / infinite M200, Switzerland). The results are shown in Fig.

Claims

(a) (i) at least one mushroom selected from the group consisting of flowering mushroom, mushroom mushroom, mushroom mushroom, and top mushroom as a substrate,
(ii) per sugar, and
(iii) Saccharomyces cerevisiae as a yeast and Lactobacillus fermentum as a lactic acid bacterium.
Is first fermented at 25 to 45 ° C for 4 to 10 days to prepare a primary fermentation product having a protein amount of 400 to 1000 ug / ml;
(b) removing the solid content from the primary fermentation and aging at 0 to 10 < 0 > C
The present invention relates to a method for producing an osmotic enzyme fermented product using a mushroom,
In the step (a)
Wherein the mixing ratio of the substrate and the saccharide is from 1: 0.5 to 2 based on the weight ratio,
Wherein the primary fermentation comprises immediate fermentation by the sugar and microbial fermentation by lactic acid bacteria and yeast.

(a) (i) at least one mushroom selected from the group consisting of flowering mushroom, mushroom mushroom, mushroom mushroom, and top mushroom as a substrate,
(ii) per sugar, and
(iii) Saccharomyces cerevisiae as a yeast and Lactobacillus fermentum as a lactic acid bacterium.
Is first fermented at 25 to 45 ° C for 4 to 10 days to prepare a primary fermentation product having a protein amount of 400 to 1000 ug / ml;
(b) removing the solid content from the primary fermentation product and then performing secondary fermentation at 20 to 50 ° C to produce a secondary fermentation product;
(c) removing the solid matter from the secondary fermentation and aging at 0 to 10 < 0 > C
The present invention relates to a method for producing an osmotic enzyme fermented product using a mushroom,
In the step (a)
Wherein the mixing ratio of the substrate and the saccharide is from 1: 0.5 to 2 based on the weight ratio,
Wherein the primary fermentation comprises immediate fermentation by the sugar and microbial fermentation by lactic acid bacteria and yeast.

(a) (i) at least one mushroom selected from the group consisting of flowering mushroom, mushroom mushroom, mushroom mushroom, and top mushroom as a substrate,
(ii) per sugar, and
(iii) Saccharomyces cerevisiae as a yeast and Lactobacillus fermentum as a lactic acid bacterium.
Is first fermented at 25 to 45 ° C for 4 to 10 days to prepare a primary fermentation product having a protein amount of 400 to 1000 ug / ml;
(b-1) removing the solid content from the primary fermentation product and sterilizing the product at 100 to 140 캜;
(ii) Saccharomyces cerevisiae as a yeast and Lactobacillus fermentum as a lactic acid bacterium are added to the sterilized primary fermentation product (b-2) Secondary fermentation at 20 to 50 캜 to prepare a secondary fermentation product; And
(c) removing the solid matter from the secondary fermentation and aging at 0 to 10 < 0 > C
The present invention relates to a method for producing an osmotic enzyme fermented product using a mushroom,
In the step (a)
Wherein the mixing ratio of the substrate and the saccharide is from 1: 0.5 to 2 based on the weight ratio,
Wherein the primary fermentation comprises immediate fermentation by the sugar and microbial fermentation by lactic acid bacteria and yeast.

4. The method according to any one of claims 1 to 3,
Wherein the saccharide is at least one selected from the group consisting of white sugar, sulfur sugar and raw sugar.

4. The method according to any one of claims 1 to 3,
Wherein the yeast is Saccharomyces cerevisiae MAB Y1 (KCTC 11386BP) phosphorus mushroom.

4. The method according to any one of claims 1 to 3,
Wherein the lactic acid bacterium is selected from the group consisting of Lactobacillus fermentum Miev L1106 (KCTC 12082BP) and mushroom.

4. The method according to any one of claims 1 to 3,
Wherein the yeast and the lactic acid bacteria are mixed at a ratio of yeast: lactic acid bacteria = 1: 0.5 to 2 on the basis of the weight ratio.

delete

4. The method according to any one of claims 1 to 3,
Wherein the amount of yeast and lactic acid bacteria used in step (a) is 1 to 10% by weight based on the total weight of the mixture of the substrate and the sugar.

4. The method according to any one of claims 1 to 3,
Wherein the primary fermentation and the secondary fermentation are carried out under aerobic conditions.

An osmotic enzyme fermentation product using mushrooms produced by the method of any one of claims 1 to 3.

11. A cosmetic composition comprising the fermented product of claim 11 as an active ingredient.

A food composition comprising the fermented product of claim 11 as an active ingredient.

An antiallergic pharmaceutical composition comprising the fermented product of claim 11 as an active ingredient.

11. A cosmetic composition for improving inflammation comprising the fermented product of claim 11 as an active ingredient.

11. A cosmetic composition for skin regeneration comprising the fermented product of claim 11 as an active ingredient.

11. A cosmetic composition for improving allergy comprising the fermented product of claim 11 as an active ingredient.

An antioxidant cosmetic composition comprising the fermented product of claim 11 as an active ingredient.