KR20170141952A - Method for manufacturing old antler and Auricularia judae fermented solution by lactic acid fermentation and food composition for improving inflammation comprising the fermented solution - Google Patents

Abstract

The present invention relates to a method for manufacturing Cervus Nippon and Auricularia auricula-judae fermented solution through lactic acid fermentation and to a food composition for alleviating inflammation comprising the fermented solution as an active ingredient, and more specifically, to a method for manufacturing a composition for alleviating inflammation having reinforced dietary fiber, high-concentration GABA and probiotics by performing selective extraction of useful components by stepwise hydrothermal extraction of the Cervus Nippon and using lactic acid bacteria of Kimchi with a mixture of a Cervus Nippon extract and an Auricularia auricula-judae powder. In the present invention, Cervus Nippon and Auricularia auricula-judae fermented solution having reinforced GABA and dietary fiber were manufactured through the method, and a functional food material having antioxidant and anti-inflammatory effects were developed.

Description

TECHNICAL FIELD [0001] The present invention relates to a method for preparing a fermented liquid of fermented green algae and throat mushroom by lactic acid fermentation and a composition for improving inflammation comprising the fermented liquid as an active ingredient }

The present invention relates to a method for preparing a fermented liquid of Lactobacillus thuringiensis and Lactobacillus acidus through lactic acid fermentation and a food composition for improving inflammation comprising the fermentation broth as an active ingredient. More particularly, The present invention relates to a method for producing an inflammation-reducing composition having enhanced dietary fiber, high-concentration GABA and probiotics by using lactic acid bacteria of the genus Kimchi,

Edible mushrooms such as thorny mushroom are representative typical foods, and they are rich in dietary fiber, various nutritional ingredients and unique flavors, and are a typical health food material. Thirsty mushroom is very high in dietary fiber content, and is rich in vitamin D. In particular, the function of "smooth bowel activity" was recognized as a functional ingredient of health functional foods in the food industry. Recently, it is required to research and develop high value added materials through evaluation of efficacy of edible mushroom produced by artificial cultivation. In order to utilize mushroom 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 effective in treating immune diseases such as suicide, atopy and arthritis, and fermented products using lactic acid bacteria have been commercialized. In particular, physiologically active substances produced by lactic acid bacteria include antimicrobial substances, peptides and gamma-butyric acid (GABA).

  L-glutamic acid is known as a substance that induces neuronal activity and is activated by glutamate decarboxylase (GAD, EC 4.1.1.15), r-aminobutyric acid acid (GABA). GABA is an inhibitory neurotransmitter that has been known to be effective in improving cerebral blood flow, relieving stress, improving memory, lowering blood pressure, relieving depression, and obesity. The plant GABA content, which can be obtained from nature, increases the GABA content using microorganisms because it exerts a pharmacological effect and uses it in various products.

Korean Patent Publication No. 10-2015-0102327 (published on Sep. 20, 2015)

It is an object of the present invention to provide a method of producing lactic acid bacteria which comprises fermenting lactic acid using Lactobacillus plantarum EJ2014 (KCCM11545P), antioxidative activity and a functional substance such as gammaaminobutyric acid (GABA) Mushroom complex lactic acid fermentation broth.

In order to accomplish the above object, the present invention provides a method for producing a green algae extract, comprising the steps of: (1) (2) adding and mixing monosodium glutamate (MSG), glucose, and yeast extract to the above-prepared extract of green tea leaves; (3) heat treating the mixture; And (4) incubating the heat-treated mixture with a Lactobacillus starter and incubating and fermenting the lactic acid, and an antioxidant activity and gamma-aminobutyric acid (GABA) -enhanced edible oil extract, And a manufacturing method thereof.

In addition, the present invention provides a food composition for improving inflammation, comprising as an active ingredient, a green tea extract and a fermented liquid mixture of lactic acid fermented with Lactobacillus plantarum EJ2014 (KCCM11545P).

The present invention relates to a method for preparing a fermented liquid of Lactobacillus thuringiensis and Lactobacillus acidus through lactic acid fermentation and a food composition for improving inflammation comprising the fermentation broth as an active ingredient. More particularly, The present invention relates to a method for producing an inflammation-reducing composition having enhanced dietary fiber, high-concentration GABA and probiotics by using lactic acid bacteria of the genus Kimchi, In the present invention, a functional food material having antioxidative and anti-inflammatory effects was produced by producing the fermented product of GABA and dietary fiber fortified with the above-mentioned method.

Fig. 1 shows a process for extracting corn germ oil and lactic acid fermentation.
FIG. 2 shows the acidity and the pH change according to the MSG content of the extract of the green tea extract.
Fig. 3 shows changes in the number of live cells during the fermentation period of lactic acid bacteria according to MSG concentration.
Fig. 4 shows GABA production of fermented lactic acid according to the presence or absence of throat mushroom in the extract of green tea.
Figure 5 shows GABA production of lactic acid fermentations according to the MSG concentration of the mushroom and green onion extracts.
Fig. 6 shows the cell survival rate of the extract of Angelica keiskei koidz.
FIG. 7 shows the cell survival rate of the extract of the green tea extract after addition of the mushroom.
Fig. 8 shows the results of NO production of the extract from the green wing before the addition of the mushroom.
FIG. 9 shows the results of NO production of the extract of the green tea extract after addition of the mushroom.

The present inventors optimized the production of GABA, a functional material, by adding lactic acid bacteria derived from Kimchi to the extracts of green tea leaves, adding throat mushroom in the range of 2.5% to 10% based on the solid content, and sterilizing. For the production of GABA and the enhancement of functional materials, lactic acid fermentation medium was prepared by adding 5% of vitamin D and dietary fiber - rich mushroom and sodium glutamate. GABA lactic acid bacteria were inoculated at 1% level and fermented at 30 ℃ for 7 days. This resulted in fermentation of probiotics, GABA and dietary fibers, and their antioxidant and inflammation The present inventors have completed the present invention by preparing a functional fermented starch complex composition through evaluation of relaxation efficacy.

The present invention relates to (1) a method for preparing a green algae extract by hydrothermal extraction of a green algae; (2) adding and mixing monosodium glutamate (MSG), glucose, and yeast extract to the above-prepared extract of green tea leaves; (3) heat treating the mixture; And (4) incubating the heat-treated mixture with a Lactobacillus starter and incubating and fermenting the lactic acid, and an antioxidant activity and gamma-aminobutyric acid (GABA) -enhanced edible oil extract, And a manufacturing method thereof.

Preferably, the step (2) comprises, by weight, 0.1 to 10 parts by weight of mushroom, 1 to 5 parts by weight of MSG, 1 to 5 parts by weight of glucose, and yeast extract, 0.1 to 3 parts by weight may be added and mixed, but the present invention is not limited thereto.

Preferably, the fermentation in the step (4) may be performed at 25 to 30 DEG C for 1 to 10 days, but is not limited thereto.

The Lactobacillus plantarum strain EJ2014 used in the examples of the present invention was deposited with KCCM11545P at the Korean Microorganism Conservation Center.

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.

The present invention also provides an antioxidative activity, a GABA enhanced edible oil extract, and a fermented broth of Lactobacillus acidus produced by the above method.

In addition, the present invention provides a food composition for improving inflammation, comprising as an active ingredient, a green tea extract and a fermented liquid mixture of lactic acid fermented with Lactobacillus plantarum EJ2014 (KCCM11545P).

In detail, the antioxidant activity and GABA can be enhanced by the fermented liquid of the extract of the green algae and the combined fermented lycopersicum lactic acid.

In the present invention, "gamma-aminobutyric acid (GABA)" is an inhibitory neurotransmitter, which is known to be effective in improving cerebral blood flow, relieving stress, improving memory, lowering blood pressure, relieving depression, Product development is being done. The plant GABA content, which can be obtained from nature, increases the GABA content using microorganisms because it exerts a pharmacological effect and uses it in various products.

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 fermentation broth can be added include dairy products including meat, sausage, bread, chocolate, candy, snack, confectionery, pizza, ramen noodles, gums, ice cream, soups, drinks, tea, Alcoholic beverages and vitamin complexes.

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> Preparation of Extracts of Green Crab and Lactic Acid Fermentation

1. Manufacture of green algae extract

After adding 500 g of green tea to 10 L of distilled water, the solution is extracted at high temperature for 40 hours. The extracts of hot water extracts (100 mL) were mixed with 2.5 g of mushroom, 2.5 ~ 4.5% of MSG, 1.5% of glucose and 0.5% of YE and then heat treated at 121 캜 for 15 minutes. And was a Lactobacillus Planta volume (L. plantarum) EJ2014 starter (starter) on each sample inoculated with 1% to prepare a fermented product and incubated at 30 ℃ incubator 7 days (Fig. 1).

2. Results and discussion

(1) Solid content, pH, acidity and viable cell number change

The pH of the lactic acid fermented extract from the green crab tended to increase during the fermentation period with increasing concentration of MSG. In the case of MSG 2.5% addition, it decreased to pH 4.3 on the 1st day of fermentation and increased to pH 4.8 with slight increase on 3 days of fermentation and showed similar pH value during 7 days of fermentation. The acidity showed the highest 1.3% after 3 days of fermentation and then decreased rapidly on the 5th day of fermentation.

MSG concentration was increased from 3 days after fermentation to 4.5 and 6.8 and 5 days after fermentation, respectively. The acidity of the fermentation decreased rapidly with increasing fermentation period at 0.8% on the first day of fermentation, and was very low as about 0.15% on the third day of fermentation (FIG. 2).

Therefore, the pH of lactic acid extract was lowered in lactic acid fermentation process despite the acid production. This phenomenon appears to be caused by depletion of hydrogen ion during GABA production.

The viable cell counts showed the highest value at 8 × 10 ⁸ on the first day of fermentation and 1.5 × 10 ⁹ on the third day of fermentation without the addition of the mushroom. On the other hand, the number of viable cells per day of fermented mushroom was 4.5 × 10 ^9, and the highest value was 1.9 × 10 ⁹ on the third day of fermentation. Therefore, it was shown that the addition of thymus mushroom had an effect on the number of viable cells during lactic acid fermentation (Table 1).

When MSG concentration was added 2.5% ~ 4.5% at the addition of thymus mushroom, the highest viable cell count was observed on the 1st day of fermentation, and the number of viable cells tended to decrease gradually with increasing fermentation period. In particular, when the concentration of MSG was high, the number of live bacteria was highest at 7 days of fermentation (FIG. 3).

(2) GABA production (MSG 2.5-4.5%)

The results of qualitative analysis using TLC to compare changes in GABA content are shown in FIGS. 4 and 5. GABA was gradually produced from fermentation 1 to fermentation 7 days, and MSG was mostly converted at 7 days of fermentation. GABA production by lactic acid bacteria was more effective when MSG was converted on the 5th day of fermentation when thrips were present (Fig. 4).

When the MSG content was increased from 2.5% to 4.5% as a substrate to be added, most MSG was converted to GABA on the third day of fermentation of lactic acid when the thymus was added to the extract of the wax extract. Therefore, it was possible to produce high concentration GABA according to the addition of throat mushroom to the extract of the green peach (Fig. 5).

< Example  2> Antioxidant efficacy of mixed lactic acid fermented extract

1. Manufacture of fermented lactic acid complex of throat mushroom and green tea extract

5 g to 20 g of thistle mushroom powder was added to 200 mL of the green tea extract prepared in Example 1, 1 g of yeast extract, 2 g of glucose and 5 to 10 g of MSG were added and heat-treated at 121 ° C for 15 minutes. Each sample was inoculated with 1% of Lactobacillus plantarum EJ2014 starter and cultured in a 30 ° C incubator for 3 days to prepare a fermented product.

2. Total polyphenol content measurement

After the fermentation of the mushroom powder and the fermentation, dilute the sample 5 times, then add 60 μL of the Folin reagent (2 times dilution of the original solution) for 3 minutes, add 60 μL of 10% Na ₂ CO ₃ solution 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.

3. Total flavonoid content measurement

0.1 mL of 10% aluminum nitrate and 1 M potassium acetate and 4.3 mL of 80% ethanol were added to 0.5 mL of the sample before and after the fermentation of the powdery mushroom powder and allowed to stand at room temperature for 40 minutes The 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.

4. DPPH  Measurement of radical scavenging activity

Before and after the addition of thymus mushroom, 160 μL of the extract of lactic acid and 30 μL of a 0.15 mM DPPH solution dissolved in ethanol was added to the sample. The absorbance was measured at 517 nm after 30 minutes at room temperature. 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 ½. For comparison of relative activities, butylated hydroxy anisole (BHA) was used as a control.

5. ABTS  Measurement of radical scavenging activity

7 mM 2-azino-bis (3-ethylbenzthiazoline-6-sulfonic acid) and 2.45 mM potassium persulfate were mixed at a final concentration and allowed to stand in a dark room at room temperature for 24 hours to form ABTS + Was diluted with phosphate buffer saline (PBS, pH 7.4) so that the absorbance value was 0.70 (+/- 0.02). 20 μL of sample was added to 180 μL of the diluted solution, and the solution was allowed to stand for exactly one minute, and the absorbance was measured.

6. Results and discussion

(1) Determination of polyphenol content and flavonoid content

The total polyphenol contents and flavonoid contents were measured before and after fermentation. The content of polyphenol before fermentation was 22.67 mg / mL and flavonoid content was 10.90 mg / mL before fermentation. The contents of polyphenols and flavonoids were higher than those of other extracts. In addition, the content of polyphenol and flavonoid were 19.11 mg / mL and 18.25 mg / mL, respectively, before fermentation, and 21.42 mg / mL and 13.69 mg / mL, respectively, (Table 2).

(2) DPPH radical scavenging ability

The antioxidant activity of lactic acid fermented product was evaluated by DPPH radical scavenging activity according to the presence or absence of thymus mushroom in the antler extract. 50, and 41.11% at 10, 50, and 70 dilution ratios of pre - fermented green peach extract without addition of thymus mushroom, respectively. IC ₅₀ values indicating 50% antioxidant capacity were 0.17 mg / mL. The radical scavenging activity of lactic acid fermented before addition of thymus mushroom was 87.22%, 53.88%, and 50.55% at dilution ratios of 10, 50 and 70, respectively. IC ₅₀ value was 0.11 mg / mL lower than that before fermentation. The DPPH radical scavenging activity was 50% at the lower concentration of the fermented lactic acid fermented with the extract of the green algae. The radical scavenging activities were 57.86%, 58.18% and 52.22% at dilution ratios of 10, 50 and 70, respectively. The fermented lactic acid was 73.33% and 49.44% after the addition of thymus mushroom, respectively. 42.77%. IC ₅₀ values of 50% activity before and after fermentation were 0.12 mg / mL and 0.11 mg / mL, respectively. The antioxidant activity is evaluated by the DPPH radical scavenging activity while shown to the neck low IC ₅₀ values by the addition of mushroom and lactic acid fermentation to overgrown antler extract was found to have high antioxidant activity (Table 3).

(3) Measurement of ABTS radical scavenging ability

After ABTS + potassium persulfate was incubated in the dark, ABTS + was removed by the antioxidant activity of the sample. The ABTS + was discolored due to the discoloration of the specific cyan color. In the dilution ratios of 10, 50 and 100, the extracts showed 74.56%, 27.93% and 17.76%, respectively, and the IC ₅₀ value was 2.83 mg / mL. The ABTS radical scavenging activity of lactic acid fermentation extracts showed similar radical scavenging activity and IC ₅₀ value was 2.83 mg / mL.

The ABTS radical scavenging activity of the extracts of P. vannamei was 71.63%, 29.88% and 18.84% in dilution ratios of 10, 50 and 100, respectively, and the IC ₅₀ value was 2.62 mg / mL. The ABTS radical scavenging activity of lactic acid fermented with the thymus mushroom was 80.49%, 27.88%, and 15.99% at dilution ratios of 10, 50 and 100, respectively. The IC ₅₀ value was 2.71 mg / mL and showed similar antioxidative activities before and after fermentation. As shown in Table 4, ABTS radical scavenging ability was excellent regardless of fermentation depending on the addition of thistle mushroom.

< Example  3> Assessment of inflammation mitigation efficacy of fermented lactic acid mixture

1. Cell viability

The survival rate of RAW cells was not cytotoxic until 15-fold dilution, but slightly cytotoxic in 5-fold dilution, when fermented samples were not fermented. The cytotoxicity was also alleviated in the 5-fold diluted samples and the cell survival rate was increased to the control level (Fig. 6).

In the case of green peach extract added with thymus mushroom, the lactic acid fermented sample showed no cytotoxicity in the 15-fold diluted sample but cytotoxic in the 5-fold diluted sample. On the other hand, in the case of the fermented samples, the cell viability was improved by decreasing the cytotoxicity in the 5-fold diluted sample. Therefore, it was judged that the agaricus extract added with Thymus mushroom alleviated cytotoxicity of RAW cells by fermentation (FIG. 7).

2. NO generation

The production of NO produced by the induction of LPS material in RAW cells was compared. In the case of the waxy extract without addition of thymus mushroom, the pre-fermentation sample showed an inhibitory effect on NO production in the 15-fold dilution. In the case of the 5-fold dilution, the inhibition of NO production due to cytotoxicity was considered. On the other hand, in the case of fermented samples, inhibition of NO production was not observed even in a sample diluted 15 times (Fig. 8).

In the case of the green peach extract added with thymus mushroom, NO production inhibition was insignificant in samples diluted 15 times before lactic acid fermentation, and NO production was significantly inhibited in samples diluted 5 times. This was thought to be due to cytotoxicity. On the other hand, in fermented samples, NO production was inhibited in 5 times diluted samples. Considering the case where the cytotoxicity of the fermented sample was alleviated, it was judged that NO production was inhibited by the addition of thymus mushroom and lactic acid fermentation (FIG. 9).

Korea Microorganism Conservation Center (overseas) KCCM11545P 20140609

Claims (7)

(1) preparing a green leaf extract by hot water extraction of a green leaf;
(2) adding and mixing monosodium glutamate (MSG), glucose, and yeast extract to the above-prepared extract of green tea leaves;
(3) heat treating the mixture; And
(4) an antioxidative activity including lactic acid fermentation by inoculating a lactobacillus starter into the heat-treated mixture and culturing the lactic acid bacteria and gamma-aminobutyric acid (GABA) enhanced guano extract and a fermented broth Way. [3] The method according to claim 1, wherein the step (2) comprises adding 0.1 to 10 parts by weight of mushroom, 1 to 5 parts by weight of MSG, 1 to 5 parts by weight of glucose, and yeast extract extract and 0.1 to 3 parts by weight of the extract are mixed and then mixed. The method according to claim 1, wherein the Lactobacillus is Lactobacillus plantarum EJ2014 (KCCM11545P). The method of claim 1, wherein the lactic acid bacterium is Lactobacillus plantarum EJ2014 (KCCM11545P). The method according to claim 1, wherein the fermentation in step (4) is fermented at 25 to 30 ° C for 1 to 10 days, and the method for producing the fermented liquid of LABORATORY EXAMPLES. An antioxidative activity and a GABA-enhanced antioxidant extract prepared by the method of any one of claims 1 to 5 and a fermented liquid of mixed lactic acid of chestnut mushroom. A composition for improving inflammation, comprising as an active ingredient, a green tea extract fermented with a strain of Lactobacillus plantarum EJ2014 (KCCM11545P) and a lactic acid fermentation solution of a mixture of Lactobacillus plantarum and Lactobacillus japonica. [Claim 7] The composition for improving inflammation according to claim 6, wherein the antioxidative activity and the GABA are enhanced in the fermentation broth of the Lactobacillus japonica extract and the mushroom complex lactic acid.

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