KR101844147B1 - Method for manufacturing Flammulina velutipes fermented solution with higher GABA content and enhancing immunity activity by lactic acid fermentation - Google Patents

Abstract

The present invention relates to a method for producing a high-concentration GABA-containing immunostimulatory mushroom fermentation product by lactic acid fermentation, and more particularly, to a method for producing high-concentration GABA fermentation product by using lactobacillus Lactobacillus plantarum EJ2014 (KCCM11545P) The present invention relates to a method for producing a functional fermented mushroom lactic acid fermented product having an immunity enhancing effect. On the other hand, the present inventors have tried to develop a functional fermented product by preparing prototype such as melon or seasoning using the fermented product .

Description

FIELD OF THE INVENTION [0001] The present invention relates to a method for producing a high-concentration GABA-containing immunostimulatory mushroom fermented by lactic acid fermentation,

The present invention relates to a method for producing a high-concentration GABA-containing immunostimulatory mushroom fermentation product by lactic acid fermentation, and more particularly, to a method for producing a high-concentration GABA fermentation product by using lactobacillus Lactobacillus plantarum EJ2014 (KCCM11545P) The present invention also relates to a method for producing lactic acid fermented functional mushroom lactic acid having an immunity enhancing effect.

Edible mushrooms such as top mushroom, mushroom mushroom, and mushroom mushroom are representative typical foods, and they are a typical food material because they have rich dietary fiber, various nutrients and unique flavors. Recently, it is required to develop effective storage and processing products because domestic consumption is small compared with the size of edible mushroom produced by artificial cultivation. Raw mushrooms are usually powdered by conventional processing methods and can be stored for a long time. However, in order to utilize mushroom powder as a health food material and high value added material, it is necessary to develop a material in which a functional material is reinforced by the production of a useful substance by fermentation and utilize it as a material for food and health food.

Lactic acid bacteria are generally known as probiotics (live live bacteria that help the intestinal health) as GRAS (Generally Recognized As Safe). Lactic acid bacteria produces lactic acid from sugar in traditional fermented foods and is used as a starter for fermented foods such as kimchi and fermented milk in the east and the west. Recently, lactic acid bacteria have been reported to be beneficial to health, for example, for improving the function and immunity of lactic acid bacteria, and fermented products using lactic acid bacteria have been actively commercialized. In particular, physiologically active substances produced by lactic acid bacteria include antimicrobial substances, peptides, and GABA (gamma-amino butyric acid).

  L-glutamic acid, a neurotransmitter of the central nervous system, is known to be a substance inducing neuronal activity. Glutamate decarboxylase (GAD, EC 4.1.1.15) Aminobutyric acid (GABA). GABA is a neurotransmitter in the inhibitory system that is widely distributed in plants such as plants and plants, and increases acetylcholine and promotes brain function. GABA has been reported to have various effects such as insomnia, anti-stress, depression, blood pressure lowering, etc., mainly affecting brain function.

Korean Registered Patent No. 10-1584593 (registered on Jan. 06, 2016)

It is an object of the present invention to provide a method for producing a fermented mushroom lactic acid fermented with enhanced immunity and gamma-aminobutyric acid (GABA) using GABA-producing lactic acid bacteria.

It is also an object of the present invention to provide an immunopotentiating and GABA-enhanced topoisomeric lactic acid fermentation product prepared by the above method.

It is also an object of the present invention to provide a food composition or food additive for enhancing GABA and immunity comprising the above fermented product of mushroom lactic acid as an active ingredient.

It is also an object of the present invention to provide a method for manufacturing a fermented yeast containing the above fermented mushroom lactic acid.

It is also an object of the present invention to provide a method for preparing a seasoning containing the above fermented mushroom lactic acid.

In order to achieve the above object, the present invention provides a method for preparing a microcrystalline cellulose, comprising the steps of: (1) adding powdered mushroom powder, yeast extract, glucose and monosodium glutamate (MSG) (2) heat treating the mixture; And (3) a step of fermenting lactic acid by inoculating and incubating the heat-treated mixture with a lactobacillus starter, and then cultivating the lactic acid fermented product with gammaaminobutyric acid (GABA) to provide.

In addition, the present invention provides an immunopotentiating and GABA-enhanced topical mushroom lactic acid fermentation product prepared by the above method.

In addition, the present invention provides a food composition for promoting GABA and immunostimulating comprising the above fermented product of mushroom lactic acid as an active ingredient.

In addition, the present invention provides a food additive for enhancing GABA and immunity comprising the above fermented product of mushroom lactic acid as an active ingredient.

The present invention also relates to a method for producing a microorganism which comprises (1) 2.5 to 50 parts by weight of a powdery mushroom powder, 0.1 to 3 parts by weight of yeast extract, 0.5 to 5 parts by weight of glucose, 10 parts by weight of a polyvinyl alcohol; (2) heat treating the mixture; (3) Inoculating the heat-treated mixture with a starter of Lactobacillus plantarum EJ2014 (KCCM11545P) and culturing to ferment lactic acid to produce a fermented product of Lactobacillus acidic mushroom lactic acid; (4) mixing 5 to 50 parts by weight of the above fermented mushroom lactic acid with 100 parts by weight of whole water; And (5) mixing the mixture of step (4) with agar, sugar, salt and bean gum, heating, and then solidifying the mixture.

The present invention also relates to a method for producing a microorganism which comprises (1) 2.5 to 50 parts by weight of a powdery mushroom powder, 0.1 to 3 parts by weight of yeast extract, 0.5 to 5 parts by weight of glucose, 10 parts by weight of a polyvinyl alcohol; (2) heat treating the mixture; (3) Inoculating the heat-treated mixture with a starter of Lactobacillus plantarum EJ2014 (KCCM11545P) and culturing to ferment lactic acid to produce a fermented product of Lactobacillus acidic mushroom lactic acid; (4) mixing 1 to 50 parts by weight of roasted wheat bran with 100 parts by weight of the above fermented mushroom lactic acid; And (5) a step of hot-air drying the mixture of step (4) above, wherein the fermented mushroom lactic acid fermented product is prepared.

The present invention relates to a method for producing a high-concentration GABA-containing immunostimulatory mushroom fermentation product by lactic acid fermentation, and more particularly, to a method for producing high-concentration GABA fermentation product by using lactobacillus Lactobacillus plantarum EJ2014 (KCCM11545P) The present invention relates to a method for producing a functional fermented mushroom lactic acid fermented product having an immunity enhancing effect. On the other hand, the present inventors have tried to develop a functional fermented product by preparing prototype such as melon or seasoning using the fermented product.

FIG. 1 shows a method for optimizing lactic acid fermentation of the mushroom powder according to the addition ratio of the mushroom powder.
Fig. 2 shows the results of pH and acidity changes of lactic acid mushroom powder according to lactic acid fermentation.
Fig. 3 shows the results of the change in the viable cell count of the mushroom powder according to lactic acid fermentation.
Fig. 4 shows the results of measurement of GABA content according to lactic acid fermentation of the mushroom powder.
FIG. 5 shows a method for optimizing lactic acid fermentation of a mushroom powder according to the presence or absence of MSG.
FIG. 6 shows the results of pH and acidity changes of lactic acid mushroom powder according to fermentation with lactic acid according to presence or absence of MSG.
FIG. 7 shows the results of the change in viable counts of the mushroom powder with or without MSG according to lactic acid fermentation.
FIG. 8 shows the results of measuring the content of GABA according to lactic acid fermentation of the mushroom powder with or without MSG.
Figure 9 shows the effect of fermented mushroom lactic acid fermentation on RAW264.7 cell viability.
Fig. 10 shows the promoting effect of the fermented product of the mushroom lactic acid in the NO production.
Fig. 11 shows a process for producing melon fermented with a lactic acid fermented product of powdery mushroom powder.
Fig. 12 shows a fermented product of lactic acid fermented with powdery mushroom powder.
Fig. 13 shows the result of the storage stability evaluation of the fermented milk fermented with white mushroom powder.
Fig. 14 shows a method of producing a seasoning using fermented lactic acid of powdery mushroom powder.
Fig. 15 shows a seasoning prepared by using lactic acid fermented product of powdery mushroom powder.
16 shows the results of measurement of the GABA content of the fermented mushroom powder seasoning according to the addition of roasted wheat bran.

Accordingly, the present inventors optimized the production of functional material GABA by inoculating Lactobacillus plantarum strain EJ2014 after sterilization of the powdery mushroom powder in the range of 2.5% to 50% based on the solid content. GABA production is affected by microorganism types and culture conditions. Therefore, yeast extract, glucose, and monosodium glutamate (MSG) are added to the powdery mushroom powder at a level of 5% To prepare a fermentation medium. Lactobacillus plantarum strain EJ2014, which is excellent in GABA production ability, was inoculated at 1% level and fermented for 5 days at 30 ° C. in a stationary culture. Through this, a fermented product of lactic acid bacteria, GABA and immunity-promoted top mushroom was produced. In addition, prototype products such as fermented mushroom fermented product were developed and the present invention was completed.

The present invention relates to (1) a method for producing a microcrystalline cellulose, comprising the steps of: mixing and adding water to a mushroom powder, yeast extract, glucose and monosodium glutamate (MSG); (2) heat treating the mixture; And (3) a step of fermenting lactic acid by inoculating and incubating the heat-treated mixture with a lactobacillus starter, and then cultivating the lactic acid fermented product with gammaaminobutyric acid (GABA) to provide.

Preferably, the step (1) comprises: 2.5 to 50 parts by weight of the mushroom powder, 0.1 to 3 parts by weight of yeast extract, 0.5 to 5 parts by weight of glucose, 1 to 10 parts by weight may be added and mixed, but the present invention is not limited thereto.

Preferably, the lactic acid bacteria may be, but is not limited to, Lactobacillus plantarum EJ2014 (KCCM11545P). The Lactobacillus plantarum strain EJ2014 used in the examples of the present invention was deposited as KCCM11545P in the Korean Microorganism Conservation Center.

Preferably, the fermentation in the step (3) may be carried out at 25 to 30 DEG C for 1 to 10 days, but is not limited thereto.

In the present invention, the term "starter" refers to a culture medium of microorganisms used for producing a fermented product. Therefore, the types of starter microorganisms determine the characteristics of the product and have an important influence on the quality of the product. Bacteria, fungi, yeast, etc., which are used as starters in microorganisms, can be used alone or in combination.

In the present invention, "gama amino butyric acid (GABA)" is a 4-aminobutyric acid which is produced by a decarboxylation reaction of an L-glutamate substrate. Glutamate It has a glutamate decarboxylase (GAD) and is synthesized as a pyridoxid-5'-phosphate dependent pathway. GABA is a nonprotein amino acid that is not found in proteins. It is a neurotransmitter in the brain or spinal cord. It is known as a brain nutrient that promotes blood flow through the brain and promotes brain metabolism and brain memory by increasing oxygen supply to the brain. GABA is known to inhibit neuronal activity as opposed to glutamate activating neurons, and this function has a profound effect on the function and information processing of neurons, especially in sensory brains such as direction sensitivity, angle sensitivity It is also known to coordinate and elaborate exercise functions. GABA stimulates the cerebral blood flow and increases oxygen supply, stimulates the metabolism of brain cells, thereby improving the after effects of stroke and cerebral arterial sclerosis.

In addition, the present invention provides an immunopotentiating and GABA-enhanced topical mushroom lactic acid fermentation product prepared by the above method.

In addition, the present invention provides a food composition for promoting GABA and immunostimulating comprising the above fermented product of mushroom lactic acid as an active ingredient.

In the case of the food composition of the present invention, there is no particular limitation on the kind of the food. Examples of the food to which the fermented mushroom lactic acid fermented product can be added include dairy products including meat, sausage, bread, chocolate, candy, snack, confectionery, pizza, ramen and other noodles, gums, ice cream, Tea, a drink, an alcoholic beverage, and a vitamin complex. In the present invention, the fermented product of the above fermented mushroom lactic acid was added to the preparation of the yam.

In addition, the present invention provides a food additive for enhancing GABA and immunity comprising the above fermented product of mushroom lactic acid as an active ingredient.

The food additive of the present invention can be used as a flavoring agent such as a preservative, a bactericide, an antioxidant, a spice, a seasoning, a sweetener, a flavoring agent, a swelling agent, a strengthening agent, an improving agent, an emulsifying agent, various nutrients, a synthetic flavoring agent, A colorant, a coloring agent, a coloring agent, a thickening agent (cheese, chocolate, etc.), pectic acid and its salt, alginic acid and its salt, an organic acid, a protective colloid thickening agent, a pH adjusting agent, a stabilizer, a defoaming agent, Alcohols, and carbonating agents used in carbonated beverages. They may be added to foods by immersion, spraying or mixing. In the example of the present invention, the fermented product of the above fermented mushroom lactic acid was added to the seasoning.

The present invention also relates to a method for producing a microorganism which comprises (1) 2.5 to 50 parts by weight of a powdery mushroom powder, 0.1 to 3 parts by weight of yeast extract, 0.5 to 5 parts by weight of glucose, 10 parts by weight of a polyvinyl alcohol; (2) heat treating the mixture; (3) Inoculating the heat-treated mixture with a starter of Lactobacillus plantarum EJ2014 (KCCM11545P) and culturing to ferment lactic acid to produce a fermented product of Lactobacillus acidic mushroom lactic acid; (4) mixing 5 to 50 parts by weight of the above fermented mushroom lactic acid with 100 parts by weight of whole water; And (5) mixing the mixture of step (4) with agar, sugar, salt and bean gum, heating, and then solidifying the mixture.

The present invention also relates to a method for producing a microorganism which comprises (1) 2.5 to 50 parts by weight of a powdery mushroom powder, 0.1 to 3 parts by weight of yeast extract, 0.5 to 5 parts by weight of glucose, 10 parts by weight of a polyvinyl alcohol; (2) heat treating the mixture; (3) Inoculating the heat-treated mixture with a starter of Lactobacillus plantarum EJ2014 (KCCM11545P) and culturing to ferment lactic acid to produce a fermented product of Lactobacillus acidic mushroom lactic acid; (4) mixing 1 to 50 parts by weight of roasted wheat bran with 100 parts by weight of the above fermented mushroom lactic acid; And (5) a step of hot-air drying the mixture of step (4) above, wherein the fermented mushroom lactic acid fermented product is prepared.

Hereinafter, the present invention will be described in more detail with reference to the following examples. However, the present invention is not limited by these examples.

< Example  1> Optimization of lactic acid fermentation of top mushroom powder according to the addition ratio of top mushroom powder

1. Materials and Methods

After adding 5-20 g of powder of mushroom powder to 100 mL of distilled water, 0.5 g of yeast extract, 1 g of glucose and 5 g of MSG were added and heat-treated at 121 ° C for 15 minutes. Each sample was inoculated with 1% L. plantarum EJ2014 starter and cultured in a 30 ° C. incubator for 5 days (FIG. 1).

2. Results and discussion

(1) pH and acidity measurement

The pH of the mushroom powder before fermentation was 6.38-6.55, but it decreased to pH 4.44-4.68 on the 1st day of fermentation, and then it tended to remain similar afterwards. The acidity was 0.2-0.29%, but it was 0.86-1.1% on the 1st day of fermentation and the highest value was 1.1% in 20% of the top mushroom. From 3 days of fermentation, the acidity decreased in the 5% sample of the mushroom powder, and the 20% sample of the mushroom powder showed an increase in acidity until the 5th day of fermentation (1.6%).

(2) Measurement of viable cell count

On the first day of fermentation, viable cell counts of 10 ⁹ CFU / mL or more were observed in all the samples, but the fermentation period was maintained until the third day of fermentation. On the 5th day of fermentation, the number of viable cells decreased under all conditions, The higher the concentration of the powder, the higher the viable cell number tended to be maintained (Fig. 3).

(3) Measurement of GABA content

In order to compare the content of GABA, qualitative analysis using TLC showed that GABA was produced from the 1st day of fermentation in all samples, MSG was converted at the 3rd day of fermentation in 10% of the mushroom powder, and GABA And the change in the content was not large (Fig. 4).

< Example  2> Optimization of Lactic Acid Fermentation of Powdery Mushroom Powder with and without MSG Addition

1. Materials and Methods

After adding 10 g of powdery mushroom powder to 100 mL of distilled water, 0.5 g of yeast extract, 1 g of glucose and 0 g of MSG were added and heat-treated at 121 ° C for 15 minutes. Each sample was inoculated with 1% L. plantarum EJ2014 starter and fermented for 5 days in a 30 ° C incubator (FIG. 5).

2. Results and discussion

(1) Change in pH and acidity

The pH before fermentation was 6.70 when 0% MSG was added and 6.57 when 5% MSG was added. The pH was slightly higher when 0% MSG was added, but the fermentation with 0% MSG was abruptly dropped to about 3.9 And the fermented product added with 5% MSG decreased to 4.69 on the 1st day and gradually increased to 6.48 on the 5th day. The acidity before fermentation was about 0.3%. As the fermentation proceeded, the fermentation with 0% MSG increased to about 1%, and the fermentation with MSG 5% And decreased sharply to 0.2% on the fifth day (Fig. 6).

(2) Measurement of viable cell count

As a result of the measurement of the number of viable cells of the fermented mushroom, the number of viable cells was 2.03 × 10 ⁹ CFU / mL on the 1st day of fermentation regardless of the MSG concentration, the MSG content was 0% on the 5th day of fermentation and about 3.7 × 10 ⁸ CFU / mL, and the fermented product added with MSG 5% tended to keep the number of viable cells at 10 ⁹ (FIG. 7).

(3) Measurement of GABA content

In order to compare the content of GABA according to 0% and 5% addition of MSG, the supernatant obtained after centrifuging the fermented product was measured by TLC. As a result, there was almost no GABA content in MSG 0%, but fermentation with MSG 5% On day 5, about 2% or more GABA spots appeared in water (Fig. 8). Free amino acid analysis of the fermented mushroom fermented with 5% MSG on day 0 and day 5 showed that 3.54% of glutamic acid was converted to 2.31% of GABA (Table 1 and Table 2 ).

The 0th glutamic acid content of fermented mushroom fermented with 5% MSG **** AMINO ACID COMPOSITION TABLE **** AA Result MOL% (ug / mL) ^a (nmol / mL) ^b Cys 0.00 0.00 0.00 0.00 ASP 0.00 0.00 0.00 0.00 GLU 6010.40 100.00 35372.42 240416.12 ASN 0.00 0.00 0.00 0.00 SER 0.00 0.00 0.00 0.00 GLN 0.00 0.00 0.00 0.00 GLY 0.00 0.00 0.00 0.00 HIS 0.00 0.00 0.00 0.00 ARG 0.00 0.00 0.00 0.00 THR 0.00 0.00 0.00 0.00 ALA 0.00 0.00 0.00 0.00 GABA 0.00 0.00 0.00 0.00 PRO 0.00 0.00 0.00 0.00 TYR 0.00 0.00 0.00 0.00 VAL 0.00 0.00 0.00 0.00 MET 0.00 0.00 0.00 0.00 Cys2 0.00 0.00 0.00 0.00 ILE 0.00 0.00 0.00 0.00 LEU 0.00 0.00 0.00 0.00 PHE 0.00 0.00 0.00 0.00 TRP 0.00 0.00 0.00 0.00 LYS 0.00 0.00 0.00 0.00 TOTAL 6010.40 100.00 35372.42 240416.12

5th day GABA content of fermented mushroom fermented with 5% MSG **** AMINO ACID COMPOSITION TABLE **** AA Result MOL% (ug / mL) ^a (nmol / mL) ^b Cys 0.00 0.00 0.00 0.00 ASP 0.00 0.00 0.00 0.00 GLU 0.00 0.00 0.00 0.00 ASN 0.00 0.00 0.00 0.00 SER 0.00 0.00 0.00 0.00 GLN 0.00 0.00 0.00 0.00 GLY 0.00 0.00 0.00 0.00 HIS 0.00 0.00 0.00 0.00 ARG 0.00 0.00 0.00 0.00 THR 0.00 0.00 0.00 0.00 ALA 0.00 0.00 0.00 0.00 GABA 5609.73 100.00 23134.51 224389.00 PRO 0.00 0.00 0.00 0.00 TYR 0.00 0.00 0.00 0.00 VAL 0.00 0.00 0.00 0.00 MET 0.00 0.00 0.00 0.00 Cys2 0.00 0.00 0.00 0.00 ILE 0.00 0.00 0.00 0.00 LEU 0.00 0.00 0.00 0.00 PHE 0.00 0.00 0.00 0.00 TRP 0.00 0.00 0.00 0.00 LYS 0.00 0.00 0.00 0.00 TOTAL 5609.73 100.00 23134.51 224389.00

< Example  3> Top mushroom powder Fermented  Antioxidant activity measurement

1. Materials and Methods

After adding 10% of mushroom powder, 0.5% of yeast extract, 1% of glucose and 5% of MSG were added to distilled water and heat treated at 121 ° C for 15 minutes. L. plantarum EJ2014 starter was added to 1 % Inoculated and incubated for 5 days in a 30 ℃ incubator for 5 days. The antioxidant activity of the mushroom powder before fermentation and after fermentation were compared with each other.

(1) Total polyphenol content measurement

A mushroom powder fermentation before diluting the samples 5-fold after fermentation by taking the following 60 μL After standing 3 minutes by adding 60 μL Folin reagent (stock solution of 2-fold diluted solution) 10% Na ₂ CO ₃ Solution was added and reacted, and the absorbance of the reaction solution was measured at 700 nm. Prepare a standard curve with 0.1% (w / v) of distilled water and make final concentrations of 0, 20, 40, 60, 80 and 100 μg / mL. Respectively.

(2) Total flavonoid content measurement

Before and after fermentation, 0.1 mL of 10% aluminum nitrate and 1 M potassium acetate were added to each sample, and 4.3 mL of 80% ethanol was added. The mixture was allowed to stand at room temperature for 40 minutes and absorbance was measured at 415 nm. Prepare quercetin 0.1% (w / v) as a standard curve with distilled water and make final concentrations of 0, 50, 100, 150, and 200 μg / mL. Measure the absorbance at 415 nm Respectively.

(3) Measurement of DPPH radical scavenging activity

After fermentation and fermentation, 160 μL of sample and 40 μL of a 0.15 mM DPPH solution dissolved in ethanol were added and allowed to stand at room temperature for 30 minutes. Absorbance was measured at 517 nm. The free radical scavenging activity of each sample extract was expressed as IC ₅₀ , the concentration of the sample required to reduce the absorbance of the control without addition of the sample to 2/1. For comparison of relative activities, butylated hydroxy anisole (BHA) was used as a control.

(4) Measurement of ABTS radical scavenging activity

The final concentration of 7 mM 2,2-azino-bis (3-ethylbenzthiazoline-6-sulfonic acid) and 2.45 mM potassium persulfate were mixed and incubated at room temperature for 24 hours to form ABTS + And then diluted with phosphate buffer saline (PBS, pH 7.4) to 0.70 (± 0.02). Add 20 μL of sample to 180 μL of the diluted solution, and allow to stand for exactly 1 minute. Absorbance was measured.

2. Results and discussion

(1) Determination of polyphenol content and flavonoid content

Total polyphenol content and flavonoid content before and after fermentation of the mushroom powder were found to be 0.33 mg / mL before fermentation and 0.46 mg / mL before fermentation. Polyphenol content after fermentation was 0.25 mg / mL, and the flavonoid content was 0.35 mg / mL, indicating that both the polyphenol content and the flavonoid content showed a slightly higher antioxidant activity than the fermentation stage (Table 3).

Determination of polyphenol content and flavonoid content in fermented powder of mushroom powder Polyphenol content (mg / mL) Flavonoid content (mg / mL) Before 0.33 + 0.02 0.46 + 0.04 After 0.25 0.01 0.35 + 0.10

(2) Measurement of DPPH radical scavenging ability

The antioxidant activity of the fermented powder of mushroom powder using DPPH radical scavenging activity method is shown in Table 4. It was found that the inhibitory effect of powdery mushroom powder was 87.78%, 62.22%, and 37.78% at dilutions of 10, 50 and 100, respectively, and the IC ₅₀ value was 1.89 mg / mL. The radical scavenging activity after the fermentation of the mushroom powder was 76.67%, 56.67% and 44.44% at dilution ratios of 10, 50 and 100, respectively, and the IC ₅₀ value was 1.24 mg / mL. The DPPH radical scavenging ability of fermented fermented mushroom powder showed 50% activity at lower concentration, indicating that the fermented product was more active after fermentation.

Measurement of DPPH radical scavenging ability of fermented powder of mushroom powder Dilution factor Scavenging
activity (%) IC ₅₀ (mg / mL) Before 10 87.78 + - 11.0 1.89 ± 0.01 50 62.22 + - 3.14 100 37.78 ± 3.14 After 10 76.67 ± 1.58 1.24 0.25 50 56.67 + - 4.71 100 44.44 + - 6.29

(3) Measurement of ABTS radical scavenging ability

When ABTS potassium persulfate is incubated in a dark place, the ABTS + is cleaved by the antioxidant power of the sample to discolor the specific cyan color, which can be measured by ABTS +. The fermentation efficiency of 90%, 53.34% and 24.54% of the mushroom powder before fermentation was 10, 50 and 100, respectively, and the IC ₅₀ value was 1.89 mg / mL. The ABTS radical scavenging activity of the fermented mushroom powder was 90.27%, 51.98% and 36.17% at dilution ratios of 10, 50 and 100, respectively. The IC ₅₀ value was 1.53 mg / appear.

Comparison of ABTS radical scavenging ability of fermented mushroom powder Dilution factor Scavenging
activity (%) IC ₅₀ (mg / mL) Before 10 90.50 + 0.11 1.89 ± 0.01 50 53.34 ± 1.93 100 24.54 + - 2.47 After 10 90.27 ± 0.21 1.53 ± 0.00 50 51.98 + - 3.22 100 36.17 ± 1.72

< Example  4> RAW264 .7 cell top mushroom Fermented  Immune enhancement effect

1. Materials and Methods

(1) Culture of macrophage RAW264.7 and measurement of cell viability

The cell viability and immune activity of fermented Lactic acid fermented with mung bean mushroom were measured before and after fermentation at 20 ℃. The RAW 264.7 cell line used in this experiment was purchased from the Korean Cell Line Bank (KCLB, Seoul, Korea). RAW 264.7 cells were cultured in DMEM supplemented with 10% FBS (fetal bovine serum) and 1% penicillin, streptomycin, fungizone and amphotericin B (Gibco-BRL, Rockville, MD, USA) Lt; 0 &gt; C, 5% CO2.

(3,4-dimethyl-thiazolyl-2) -2,5-diphenyl tetrazolium bromide (MTT) assay was used to measure the toxicity of the fermented mushroom to cells. The MTT assay is based on the principle that MTT, a yellow water soluble substrate, is reduced to an insoluble purple formazan by dehydrogenase action of mitochondria. The resulting formazan absorbance reflects the concentration of living and metabolized cells. RAW264.7 cells were plated at a density of 1 × 10 ⁵ cells / mL in a 96-well plate and cultured for 24 hours at 37 ° C in a 5% CO2 incubator. After incubation, 10 μL of 5 mg / mL MTT reagent was added to each well and incubated for 4 hours in an incubator. After incubation, the supernatant was removed and 100 μL of DMSO reagent was dispensed into each well. Formazan crystals formed were dissolved and measured by absorbance at 550 nm using an ELISA reader. The cell viability was calculated based on the 100% survival rate of the untreated group in which only the cells were cultured without sample treatment.

(2) Measurement of nitric oxide (NO) production

RAW 264.7 cells were seeded in 96-well plates at a concentration of 1 × 10 ⁵ cells / mL and cultured for 24 hours at 37 ° C in a 5% CO ₂ incubator. After culturing, NO was treated with 100 ng / mL of LPS (Sigma-Aldrich Co.) for 24 hours, and 100 μL of supernatant was mixed with the same amount of Griess reagent. After 10 minutes, the absorbance at 540 nm Were measured.

2. Results and discussion

(1) cytotoxicity

To determine the optimum concentration of fermented mushroom, the viability of RAW 264.7 cells after fermentation dilution was measured. As shown in FIG. 9, the survival rate was 100% in control and 96% in LPS treatment. Survival rate of over 90% was confirmed under all conditions without any difference according to the dilution ratio of the fermented product.

(2) Measurement of NO production amount

LPS treatment is known to promote the inflammatory response and promote the production of related factors such as NO. And NO produced may lead to systemic inflammation, which may have various negative effects on the living body. However, since the production of NO is considered to be an important factor of congenital immunity, in the present invention, compared with the group treated with LPS, the ability to activate congenital immunity when the fermented mushroom is treated with normal LPS- NO production was observed using RAW 264.7 cells. As a result, compared with the control, it was confirmed that NO was expressed in the fermented condition, and it was found to be increased in a concentration dependent manner. Also, NO concentration was higher after fermentation of the fermented mushroom fermented product at the X700 concentration condition (FIG. 10).

< Example  5> Top mushroom powder lactic acid The fermented product  Used Yokan  Manufacturing 1

1. Materials and Methods

(1) Preparation of lactic acid fermented mushroom

After adding 20 g of powdery mushroom powder to 200 mL of distilled water, 1 g of yeast extract, 2 g of glucose and 4 g of MSG were added and heat-treated at 121 ° C for 15 minutes. Each sample was inoculated with 1% of L. plantarum EJ2014 starter and cultured in a 30 ° C incubator for 3 days. Then, a fermented mushroom fermented product was prepared and added to the yeast preparation (FIG. 11).

(2) Manufacture of yokes

Add 10% lactic acid fermented water and top mushroom powder to a volume of 200 mL so that the volume of the fermented product is 0, 10, 20, 30%. Add 7 g of agar, 80 g of sugar, 1 g of salt, and 200 g of bean jelly, and heat for 35 minutes while mixing.

2. Results and discussion

(1) Manufacture of yokes

The fermented product of powdery mushroom powder and lactic acid was 0, 10, 20, 30%, respectively. The paper cup was used as a frame, and the result of each was not visually apparent (Fig. 12).

(2) Stability test of fermented water yam,

Yangang of each condition was cut 1cm in length and then put in a zipper bag and observed at room temperature for one week. As a result, it was confirmed that in the case of the fermented milk added with 30% of the fermented product, mold did not grow even after storage for one week, whereas molds were produced in all other fermented milk.

(3) Measurement of physical properties

The hardness was 3247.47 dyne / cm ² and the stickiness was the highest at 1430.61% in the yin yang added with 10% fermented mushroom fermented product. The elasticity was 0.68-0.70 g and the cohesiveness was 0.39-0.44%. There was no significant difference according to the concentration of fermented product (Table 6).

Determination of the properties of Yangyang by concentration of fermented milk powder Contents of fermented mushroom (%) 0 10 20 30 Hardness
(dyne / cm ² ) 2713.00 3247.47 2479.62 2625.01 Adhesiveness
(g) -120.51 -131.12 -126.60 -138.93 Springiness
(g) 0.68 0.70 0.67 0.70 Cohesiveness
(%) 0.44 0.44 0.40 0.39 Gumminess
(%) 1170.50 1430.61 989.57 1033.11 Chewiness ¹⁾ (chewing grain growth) 797.87 1005.77 664.07 723.25

¹⁾ Chewiness = springiness × gumminess.

< Example  6> Top mushroom powder lactic acid The fermented product  Used Yokan  Manufacturing 2

1. Materials and Methods

Add 0% or 15% of 10% fermented powder of mushroom powder and add 15% of a solution of Aronia sugar to make a total volume of 100 mL. Add 3 g of agar, 20 g of sugar and 0.5 g of salt to this Add 0 g, 50 g, 100 g and 150 g of bean jam, respectively, and heat for 10 minutes while mixing. In Example 5, the heating time was reduced by using a reddish precipitate in this experiment.

2. Results and discussion

(1) pH, acidity and viable cell number change

The pH decreased to pH 4.42 on the first day of fermentation and was maintained at pH 4.43 on the third day of fermentation. The acidity increased to 1.14% on the first day of fermentation and tended to be maintained on the third day of fermentation. As a result of counting viable cell count, viable cell count was highest at 1.9 × 10 ⁹ CFU / mL on the first day of fermentation and 2.0 × 10 ⁹ CFU / mL on the third day of fermentation (Table 7).

Changes of pH, acidity and number of viable cells of lactic acid mushroom powder by lactic acid fermentation Fermentation time (days) 0 One 3 pH 6.4 4.42 4.43 Acidity (%) 0.27 1.14 1.01 Viable cell count
(CFU / mL) 1.7 × 10 ⁷ 1.9 × 10 ⁹ 2.0 × 10 ⁹

(2) Evaluation of physical properties of yam

The springiness and cohesiveness of the fermented water at 0% condition were almost the same in all conditions and the hardness of the fermented water increased with the amount of sediment. Other values did not show any particular trend (Table 8).

Measurement of Physical Properties of Yangyang by Dust Concentration without Fermentation of Powdered Mushroom Powder Sediment content (g) 0 50 100 150 Hardness
(dyne / cm ² ) 154.65 128.37 204.94 334.65 Adhesiveness
(g) -15.69 -8.49 -22.41 -14.98 Springiness
(g) 0.94 0.95 0.95 0.90 Cohesiveness
(%) 0.88 0.76 0.72 0.72 Gumminess
(%) 136.46 97.56 148.03 240.43 Chewiness ¹⁾ (chewing grain growth) 0.65 92.25 140.74 215.78

¹⁾ Chewiness = springiness × gumminess.

In case of 15% fermented water, springiness showed almost the same value under all conditions. Cohesiveness tended to decrease with increasing sediment, and hardness increased until 100 g of sediment was added. The remaining values did not show a consistent trend (Table 9).

Measurements of the physical properties of melons by the concentration of sediments added with 15% fermented powder of mushroom powder Sediment content (g) 0 50 100 150 Hardness
(dyne / cm ² ) 224.12 270.56 336.95 236.97 Adhesiveness
(g) -8.60 -24.75 -34.30 -33.6 Springiness
(g) 0.97 0.96 0.96 0.94 Cohesiveness
(%) 0.90 0.80 0.73 0.71 Gumminess
(%) 202.38 214.76 245.25 167.86 Chewiness ¹⁾ (chewing grain growth) 195.47 205.19 235.50 158.40

¹⁾ Chewiness = springiness × gumminess.

< Example  7> Top mushroom powder lactic acid The fermented product  Seasoning manufacturing

1. Materials and Methods

After adding 10 g of powder of mushroom powder to 100 mL of distilled water, 0.5 g of yeast extract, 1 g of glucose and 5 g of MSG were added and heat-treated at 121 ° C for 15 minutes. Each sample was inoculated with 1% L. plantarum EJ2014 starter and incubated in a 30 ° C incubator for 3 days.

After 5 days of fermentation, 30 g of lactic acid fermented with lactic acid was added with 0-10 g of roasted bran for powdering of seasoning during the addition of dietary fiber and hot air drying, followed by hot air drying at 50 ° C for 10 hours to prepare a seasoning (Fig. 14).

2. Results and discussion

As a result of preparing the seasoning according to the content of roasted bran, it was found that the more the roasted bran was added, the more the flavor of the roasted bran increased, but the flavor of the fermented product of the powdery mushroom powder was decreased and the fermented product alone When you make a seasoning flavor is the best, but it is difficult to make the seasoning powder. This was thought to be due to the viscosity of the mushroom powder during fermentation for 5 days. Considering the preference and flow characteristics of the fermented mushroom powder product, 1 g of roasted bran was considered to be the best condition to make the seasoning, and the completed seasoning photograph is shown in Fig.

< Example  8> Top mushroom powder lactic acid The fermented product  Sensory Evaluation of Seasoning Prepared Using

1. Materials and Methods

The sensory evaluation of the seasoning using the fermented mushroom powder was carried out after 10 food researchers were selected as the sensory test agent. The evaluation items were Color, Flavor, Umami taste, Salty taste, Flowability, Overall preference, and 5 points by using 5 point method. The preference of the characteristics was improved.

2. Results and discussion

Table 10 shows the sensory evaluation results of the seasonings using the fermented product of powdery mushroom powder. The preference for color of condiment was the highest as 3.71 ± 1.38 in Control, but Sample A was 3.86 ± 0.90 in Flavor. As the fermented mushroom powder appeared to be stronger, the distinctive fragrance of mushroom powder seemed to be stronger. Sample A and B, which had much fermented mushroom powder, showed the highest preference in umami taste.

Salty taste was also higher in Sample A (3.29 ± 0.96), suggesting that it could be used as a seasoning for low salt foods. However, the flowability was 3.00 ± 1.15, which was lower than that of Sample C, which was added with control and roasted wheat flour. The overall preference was highest in Sample A (3.71 ± 0.49), followed by Sample B> Control> Sample C. As a result, 30g of fermented mushroom powder and 1g of roasted bran were the best.

Sensory evaluation of seasoning of fermented mushroom powder sample Color
(color) Flavor
(zest) Umami
taste
(Savory taste) Salty
taste
(Salty taste) Flowability
(Flowability) Overall
preference
(All likelihood) Control ^{1 )} 3.71 ± 1.38 2.28 0.95 2.43 + - 0.53 2.86 ± 1.35 3.57 ± 1.27 2.00 0.82 A ²⁾ 2.71 ± 1.11 3.86 ± 0.90 3.00 ± 1.41 3.29 ± 0.96 3.00 ± 1.15 3.71 ± 0.49 B ³⁾ 2.86 ± 1.07 3.43 ± 1.13 3.00 ± 1.29 2.57 + - 0.98 3.14 ± 1.21 3.14 ± 1.07 C ⁴⁾ 3.57 ± 1.62 2.57 ± 0.53 1.57 + - 0.79 2.29 ± 1.25 3.57 ± 0.53 1.71 ± 1.11

1) Top mushroom powder

2) Fermented mushroom powder 30g + roasted wheat bran 1g

3) Fermented mushroom powder 30g + roasted bran 3g

4) Fermented mushroom powder 30g + roasted bran 10g

< Example  9> Top mushroom powder lactic acid The fermented product  Analysis of ingredients of seasonings prepared using

1. Materials and Methods

10% of mushroom powder, 0.5% of yeast extract, 1% of glucose and 5% of MSG were added to distilled water and heat treated at 121 ℃ for 15 minutes. Then, L. plantarum 1 g of EJ2014 starter was incubated for 5 days in a 30 ° C incubator, and then 1 g of roasted bran was added to 30 g of fermented mushroom powder fermented for 5 days. After free amino acid analysis and sodium content were measured.

2. Results and discussion

In order to compare the content of GABA according to the conditions of no added roasted bran and 1 g added to fermented mushroom powder, the fermented product was diluted 10 times, 20 times, 50 times, and centrifuged to obtain a supernatant. As a result of the measurement using TLC, it was found that the GABA content was about 18% in both of the conditions in which no roasted wheat bran was added and in both of them.

The free amino acid analysis revealed that the content of GABA was about 17%, and the sodium content of the seasoning was 2666mg / 100g. The fermented mushroom powder was prepared by adding 1 g of roasted bran.

Korea Microorganism Conservation Center (overseas) KCCM11545P 20140609

Claims (11)

delete delete delete delete delete delete delete delete delete (1) 2.5 to 50 parts by weight of powdery mushroom powder, 0.1 to 3 parts by weight of yeast extract, 0.5 to 5 parts by weight of glucose and 1 to 10 parts by weight of MSG are added to 100 parts by weight of the whole water, Mixing;
(2) heat treating the mixture;
(3) Inoculating the heat-treated mixture with Lactobacillus plantarum EJ2014 (KCCM11545P) strain starter and culturing to lactic acid fermentation to prepare immune enhancing and GABA-enhanced topoisomeric lactic acid fermented product;
(4) mixing 5 to 50 parts by weight of the above fermented mushroom lactic acid with 100 parts by weight of whole water; And
(5) A method for producing koji containing a fermented product of a lactic acid fermented with mushroom gum enhanced by immunization and GABA, comprising mixing the mixture of step (4) with agar, sugar, salt and bean gum, heating and then heating. (1) 2.5 to 50 parts by weight of powdery mushroom powder, 0.1 to 3 parts by weight of yeast extract, 0.5 to 5 parts by weight of glucose and 1 to 10 parts by weight of MSG are added to 100 parts by weight of the whole water, Mixing;
(2) heat treating the mixture;
(3) Inoculating the heat-treated mixture with Lactobacillus plantarum EJ2014 (KCCM11545P) strain starter and culturing to lactic acid fermentation to prepare immune enhancing and GABA-enhanced topoisomeric lactic acid fermented product;
(4) mixing 1 to 50 parts by weight of roasted wheat bran with 100 parts by weight of the above fermented mushroom lactic acid; And
(5) A method for preparing a seasoning containing an immunostimulatory and GABA-enhanced topical mushroom lactic acid fermented product, comprising the step of hot-air drying the mixture of step (4).

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