KR102054122B1 - Fermented Gastrodia eleata that improved taste and manufacturing method thereof - Google Patents

Abstract

The present invention relates to a fermented scoop with improved flavor and a method for manufacturing the same, the fermented scoop with improved flavor according to the present invention and its manufacturing method are conventional scoops that have limitations for use in foods due to discomfort due to a unique bitter taste. In addition to improving the degree of preference, and without adversely affecting the survival of the cell, it shows an inhibitory effect of inflammation-related factors can increase the utilization of cheonma.

Description

Fermented sorghum and its manufacturing method with improved flavor {Fermented Gastrodia eleata that improved taste and manufacturing method

The present invention relates to a fermented poncho with improved flavor and a method for producing the same.

Gastrodia elata Blume) do not have chlorophyll a herbaceous perennial plant belonging to that indigenous Orchidaceae (Orcbidaceae in Korea, Japan, China), so the carbon assimilation is impossible basidiomycete giant amphipods mulberry mushroom (Armillaria grow together with mellea ). The clinical efficacy of Chunma has been widely recorded in various materials, including herbal tree, synonym and sympathy, and is known to be effective for adult diseases such as hypertension, headache, numbness, neurological diseases, diabetes, and symptoms such as stress and fatigue. Even in the early years, cheon horses have been used for the treatment of headaches, dizziness, hand and hand paralysis, strokes, and seizures.

Most active ingredients of cheonma are phenolic compounds, which include gastrodin, vanillin, vanillyl alcohol, p-hydroxybenzyl alcohol (Gastrodigenin) and parahydroxybenzyl aldehyde. (p-hydroxybenz aldehyde), and the pharmacological action of each component is being actively studied, especially gastrodine, vanillin, vanillyl alcohol and parahydroxybenzyl. The main focus is research on alcohols (p-Hydroxybenzyl alcohol).

Gastrodine is an active ingredient of the cheonma horse is known to be effective in improving cardiovascular disease by increasing memory and cognition, inhibit neuropathy such as depression, convulsions, lower blood pressure and suppress ischemic diseases, and also activate liver cells. It is also known to have glycemic control, anti-inflammatory and anticoagulant effects. Benillin and its precursor, berylyl alcohol, are known to have antioxidant, antiepileptic, antiviral, anticancer and antimicrobial effects. Parahydroxybenzyl alcohol is a metabolite of gastrodine, which has antioxidant and anti-inflammatory effects, and has been reported to have a skin whitening effect along with the effects of inhibiting neurological and ischemic diseases.

Due to these functionalities, Chunma has been registered as a food raw material since 2000, and has been developed as a processed food, but its use is limited due to the bitterness, fishy taste, and unpleasant taste peculiar to Chunma. In particular, the bitter and fishy taste of cheonma can be lost through drying, but the discomfort does not disappear even after drying. Thus, in addition to the fermentation research to increase the functionality of the cheonma recently research to reduce the unpleasant taste and smell, and the Republic of Korea Patent No. 10-0877482 No. 10 in the stevia and ocher immersed in cheonma and unpleasant flavor unique to And a method for improving taste.

Republic of Korea Patent No. 10-0877482

An object of the present invention is to provide a fermented cheonma horse with improved flavor and a method for producing the same.

In order to achieve the above object, the present invention provides a method for producing a fermented cheonma comprising the following steps:

1) immersing cheonma in vinegar; And

2) inoculating lactic acid bacteria into the vinegar dipping fermentation cheonma of step 1), and fermenting.

In addition, the present invention comprises the steps of 1) immersing cheonma in vinegar; And 2) inoculating the lactic acid bacteria into the vinegar-immersed fermentation cheonma of step 1) and providing a fermented cheonma, which is prepared by the method comprising fermentation.

In another aspect, the present invention provides a pharmaceutical composition for anti-inflammatory comprising a fermentation cheonma provided by the method as an active ingredient.

The method for producing a fermented poncho with improved flavor according to the present invention can remove the bitterness, fishy taste, unpleasant smell, etc. peculiar to cheonma, and increase the utilization of cheonma.

1 is a result showing the change of amino acids related to the taste of cheonma according to the immersion treatment method.
Figure 2 is a result showing the change of amino acids related to the taste of cheonma by vinegar dipping and lactic acid bacteria fermentation.
Figure 3 is a result of comparing the free sugar content of the cheonma extract according to the vinegar immersion, fermentation period and extraction solvent.
4 to 5 is a result of comparing the active ingredient content of the cheonma extract according to the vinegar immersion, fermentation period and extraction solvent.
Figure 4: Total Active Ingredient Content
Figure 5: A: Gastrodin, B: p-hydroxybenzyl alcohol (Gastrodigenin), C: Vanylyl alcohol, D: parahydroxybenzaldehyde (p-hydroxybenzaldehyde) , E: vanillin
6 is a result showing the inhibitory effect of NO and PGE2 in the treatment of the vinegar immersion, fermentation period and extract solvent in RAW264.7 cells induced inflammatory response (A: NO, B: iNOS)
7 is a result showing the inhibitory effect of NF-kB upon treatment of fermentation cheonma extract according to vinegar immersion, fermentation period and extraction solvent in RAW264.7 cells induced inflammatory reaction. (A: 80% ethanol extract, B: water extract)

Hereinafter, the present invention will be described in detail.

The present invention provides a method for producing a fermented cheonma comprising the following steps.

1) immersing cheonma in vinegar; And

2) inoculating lactic acid bacteria into the vinegar dipping fermentation cheonma of step 1), and fermenting.

Chunma of step 1) may be dried, but is not limited thereto.

The horse of step 1) may be ground, but is not limited thereto.

Immersion of step 1) may be to be immersed for 1 to 9 hours in vinegar of 1 to 20% concentration, preferably may be to be immersed for 3 to 5 hours in 5 to 15% concentration of vinegar. If the concentration of vinegar is lower than 1%, the effect of changing the components related to the flavor and the active ingredient may be reduced, and if higher than 20% may damage the flavor of the cheonma by the taste of the vinegar itself. In addition, if the immersion time is less than 1 hour, the effect of changing the components related to the flavor and the active ingredient may be reduced, and if it is higher than 9 hours may be damaged due to excessive reaction with vinegar.

Step 1) may further include a step of hot water for 1 to 3 hours after immersion.

The lactic acid bacteria of step 2) may be Lactobacillus brevis E3-8 strain deposited with accession number KACC 92213P, but is not limited thereto.

The fermentation process of step 2) may be a fermentation of 0.5 to 5 days at 35 to 40 ℃, preferably one to three days at 36 to 38 ℃. If the fermentation temperature is lower than 35 ℃, or higher than 40 ℃ may inhibit the activity of lactic acid bacteria. In addition, if the fermentation period is shorter than 0.5 days, the effect of changing the flavor-related components and useful components may be reduced, and if it is longer than 5 days, the reduction of useful components may occur in the metabolic process, which may lower economic efficiency.

The fermentation process of step 2) may further include the step of extracting using a solvent after fermentation.

In a specific embodiment of the present invention, cheonma immersed in vinegar has been shown to improve the flavor by reducing the amino acids associated with the pungent and bitter and components associated with an unpleasant aroma (unpleasant) (Fig. 1 and Table 1). In addition, it was confirmed that the lactic acid bacteria fermentation of cheonma lowers the pH (Table 2), causing an increase in useful components (Table 3), especially gastrodine, a major component of cheonma when fermented with Lactobacillus brevis E3-8 strain. Increasing (Table 4) and decreasing tendency of amino acids related to bitter taste (Table 5) were shown. As a result of immersion with vinegar and fermentation of L. brevis E3-8, amino acids related to bitterness (Fig. 2 and Table 8) among the quality characteristics of fermented cheonma were reduced as the fermentation progressed. In addition, the phenolic components that cause skin and respiratory irritation among the fragrance components in cheonju treated with vinegar and fermentation showed a tendency to decrease (Table 10), and flavonoids known as p-hydroxybenzyl alcohol and antioxidants as useful components. It was confirmed that the ingredients increased (Tables 9 and 11). In addition, even after extracting the vinegar immersion fermentation of the cheonma, it is confirmed whether the fermentation through the decrease in pH (Table 12) by extracting the solvent with ethanol and water free sugar (Figs. 14 and 15) it was confirmed that the content of ethanol extract is high. The fermented cheonma extract was shown to inhibit the production of NO and PGE2, which are major inflammatory factors (Table 6 and Table 17) without cytotoxicity (Table 16), and also appeared to inhibit the inflammatory cytokine NF-κB. (FIG. 7), the fermented cheonma prepared by the method of the present invention is reduced bitterness and unpleasant to reduce the degree of preference, improved Since it has an anti-inflammatory effect, it can increase the utilization of the horse.

In addition, the present invention comprises the steps of 1) immersing cheonma in vinegar; And 2) inoculating the lactic acid bacteria into the vinegar-immersed fermentation cheonma in step 1), and providing a fermentation cheonma having improved anti-inflammatory effect.

In addition, the present invention provides an anti-inflammatory pharmaceutical composition comprising the fermented cheonma as an active ingredient.

The composition may be to inhibit the production of NO, Prostaglandin E2 (PGE2) and NF-kB.

The anti-inflammatory pharmaceutical composition comprising the fermentation cheonma of the present invention as an active ingredient contains 0.1 to 99.9% by weight of the fermentation cheonma extract of the present invention as an active ingredient based on the total weight of the composition for administration, Carrier, excipient or diluent. Pharmaceutically acceptable carriers may be used in combination with saline, sterile water, Ringer's solution, buffered saline, dextrose solution, maltodextrin solution, glycerol, ethanol and one or more of these components, as necessary, including antioxidants, buffers, Other conventional additives such as bacteriostatic agents can be added. In addition, diluents, dispersants, surfactants, binders and lubricants may be additionally added to formulate main formulations, powders, tablets, capsules, pills, granules or injections, such as aqueous solutions, suspensions, emulsions and the like. Furthermore, it may be preferably formulated according to each disease or component by a suitable method in the art or using a method disclosed in Remington's Pharmaceutical Science (Mack Publishing Company, Easton PA, 19th, 1990).

The anti-inflammatory pharmaceutical composition comprising the fermentation cheonma of the present invention as an active ingredient may be formulated into tablets, capsules, powders, granules, suspensions, emulsions, syrups, and other liquids by conventional methods.

Specifically, the composition of the present invention may be administered orally or parenterally, and the formulation for oral administration may be a tablet, a troche, a lozenge, a water-soluble or a dominant suspension, a powder or granules, an emulsion, a hard or It may be formulated as a soft capsule, syrup or elixir. Lactose, saccharose, sorbitol, mannitol, erythritol, starch, amylopectin, binders such as cellulose or gelatin, excipients such as dicalcium phosphate, disintegrants such as corn starch or sweet potato starch, stearic acid for formulation into tablets and capsules Lubricants such as magnesium, calcium stearate, sodium stearyl fumarate or polyethylene glycol wax may be contained. The capsule formulation may contain a liquid carrier such as fatty oil in addition to the above-mentioned substances. In addition to the formulation of acacia rubber, calcium phosphate, alginate, gelatin, calcium silicate, microcrystalline cellulose, polyvinylpyrrolidone, cellulose, syrup, methyl cellulose, methylhydroxybenzoate, propylhydroxybenzoate, Talc, magnesium stearate and mineral oil may be added, but are not limited thereto.

In addition, the anti-inflammatory pharmaceutical composition comprising the fermented cheonma of the present invention as an active ingredient can be administered orally or parenterally, and when parenteral administration is selected subcutaneous injection, intravenous injection, intramuscular injection or intraperitoneal injection injection method It is preferable. To formulate into a parenteral formulation, the mixture of the present invention is mixed with water to prepare a suspension, which is formulated in unit dosage form of ampoules or vials.

The anti-inflammatory pharmaceutical composition comprising the fermented cheonma of the invention as an active ingredient is administered in a pharmaceutically effective amount. In the present invention, “pharmaceutically effective amount” means an amount sufficient to treat a disease at a reasonable benefit / risk ratio applicable to medical treatment, and an effective dose level means the type of disease, the severity, the activity of the drug, The sensitivity to the drug, time of administration, route of administration and rate of release, duration of treatment, factors including concurrent use of the drug, and other factors well known in the medical art can be determined. The compositions of the present invention may be administered as individual therapeutic agents or in combination with other therapeutic agents, may be administered sequentially or simultaneously with conventional therapeutic agents, and may be single or multiple doses. Taking all of the above factors into consideration, it is important to administer an amount that can obtain the maximum effect in a minimum amount without side effects, which can be easily determined by those skilled in the art.

The dosage of the active ingredient according to the present invention will consider safety and efficiency when used in the human body, it is also possible to estimate the amount used in humans from the effective amount determined through animal experiments. Such considerations when determining the effective amount include, for example, Hardman and Limbird, eds., Goodman and Gilman's The Pharmacological Basis of Therapeutics, 10th ed. (2001), Pergamon Press; And E.W. Martin ed., Remington's Pharmaceutical Sciences, 18th ed. (1990), Mack Publishing Co. For oral administration, generally 0.0001 to 500 mg of the composition of the present invention per kg of body weight per day The amount may be administered once to several times, preferably in an amount of 0.001 to 100 mg, more preferably 30 to 500 mg / kg. However, the dosage may be increased or decreased depending on the route of administration, the severity of arthritis diseases, sex, weight, age, etc., and the above dosage does not limit the scope of the present invention by any method.

The composition of the present invention may be used alone or in combination with methods using surgery, radiation therapy, hormone therapy, chemotherapy and biological response modifiers.

Compositions of the present invention may also include carriers, diluents, excipients, or combinations of two or more commonly used in biological agents. Pharmaceutically acceptable carriers are not particularly limited so long as they are suitable for in vivo delivery of the composition, for example, Merck Index, 13th ed., Merck & Co. Inc., saline solution, sterile water, Ringer's solution, buffered saline solution, dextrose solution, maltodextrin solution, glycerol, ethanol and one or more of these components can be mixed and used. And other conventional additives may be added. In addition, diluents, dispersants, surfactants, binders and lubricants may be additionally added to formulate into main dosage forms, pills, capsules, granules or tablets such as aqueous solutions, suspensions, emulsions and the like. Furthermore, it may be preferably formulated according to each disease or component by a suitable method in the art or using a method disclosed in Remington's Pharmaceutical Science (Mack Publishing Company, Easton PA, 18th, 1990).

The composition of the present invention may further contain one or more active ingredients exhibiting the same or similar functions. The composition of the present invention may comprise 0.0001 to 10% by weight of the compound, preferably 0.001 to 1% by weight, based on the total weight of the composition.

In a specific embodiment of the present invention, fermented cheonma extract immersed in vinegar has been shown to inhibit the production of NO and PGE2, which are major inflammatory factors without cytotoxicity (Table 16) (Fig. 6 and Table 17), inflammatory cytokines. It appears that also inhibits the production of the Cain NF-κB (Fig. 7), the fermented cheonma after vinegar dipping of the present invention is confirmed to have an anti-inflammatory effect can be usefully used as an anti-inflammatory pharmaceutical composition .

Hereinafter, the present invention will be described in detail by way of examples.

However, the following Examples and Experimental Examples are only for illustrating the present invention, the present invention is not limited by the following Examples and Experimental Examples.

< Example  1> celestial Immersion  process

We tried to compare the changes in the components related to taste and aroma after immersion treatment. Chunma was grown in Jeonbuk Muju-gun, washed, dried and pulverized. The pulverized dried cheonma were immersed in 10% solution of makgeolli, honey, vinegar and ginger extract, respectively, for 4 hours, and then dried in a freeze dryer (Bondiro, Ilshin) for 2 hours.

< Experimental Example  1> Immersion  Flavor Comparison of Treated Horse Horse

Experimental Example  1-1. Immersion  Comparison of Aroma Components in Treated Cheonma

The aroma component of the freeze-dried cheonma obtained in Example 1 was analyzed using a GC-MS analyzer (Clarus 600 series + TurboMatrix HSS Trap, PerkinElmer).

The mass range was m / z 1 to 1,200, the ionization method was Electron Ionization (EI), and the mobile phase gas was helium. The GC column was ELITE-FFAP (0.25 μm ID × 0.32 μm OD × 30 m length). The inlet temperature was maintained at 280 ° C., and the initial oven temperature was set at 50 ° C. and then maintained for 2 minutes. After that, the temperature was increased by 10 ° C. per minute and then maintained at 300 ° C. for 2 minutes. The injected sample amount was 1.0 μl, the split ratio was 5: 1, and the flow rate of the mobile phase gas was 1.0 mL / min.

compound RT Mass relative area (x10 ⁸ ) No treatment Makgeolli honey vinegar ginger Hydrazinecarboxamide 1:23 44 4.434 4.536 5.961 3.906 4.164 1,2-Propanediol 1:31 31 4.146 23.740 2.174 1.265 3.679 Acetone 1:35 42 0.592 1.593 0.638 0.206 0.523 Acetic acid 2:21 60 9.819 8.136 18.955 70.461 11.608 Propanoic acid, 2-oxo-, methyl ester 2:55 31 0.625 1.029 0.030 - 0.008 p-Cresol 9:17 79 0.039 0.036 0.049 0.107 0.044 Phenol, 3-methyl- (m-Cresol) 9:21 108 14.388 15.745 17.287 14.952 15.078 Phenylethyl alcohol 9:57 63 0.008 0.473 0.037 0.010
0.006 Phenol, 4- (ethoxymethyl)- 13:27 124 0.273 0.477 1.643 0.502 0.486 Tridecane, 3-methyl- 13:38 122 - - 1.035 0.446 0.326 Cyclohexene, 3- (1,5-dimethyl-4-hexenyl) -6-methylene-, [S- (R *, S *)]- 15:46 69 - - 0.717 1.147 1.605 Ar-tumerone 17:24 83 20.416 16.032 8.407 16.024 19.125 Tumerone 17:28 105 6.548 5.131 3.421 4.812 6.210

As a result, most of the immersion treatments did not show a significant reduction in the amount of 3-methyl-phenol and p-Cresol, which gave the unique phenolic aroma. Was reduced by immersion treatment, especially in honey and vinegar treatment groups (Table 1).

Experimental Example  1-2. Immersion  Comparison of Taste-related Amino Acids in Chunma

Free amino acid analysis was performed by modifying the method using trichloroacetic acid (TCA) (Aristoy and Toldra, 1991). One gram of freeze-dried cheonma sample was ground finely using a mortar and dissolved in 20 mL of 3% TCA. After mixing for 2 hours in a shaking incubator at 28 ℃, and centrifuged for 15 minutes at 4 ℃, 13,000 rpm to obtain a supernatant. After the supernatant of the centrifuged samples was filtered with a 0.2 μm filter, a total of 0.4 mL filtered samples were transferred to a new tube, and 1.6 mL of pre-filtered 3% TCA was added (2 mL total). Free amino acids were analyzed by L-8900 Amino acid analyzer (Hitachi). Reagents of analytical levels were used at Sigma-Aldrich as a standard sample of each component.

As a result, the content of free amino acids was lower in the immersion-treated cheonma than in the untreated cheonma as shown in Figure 1, in particular, the processing of the spicy amino acid Cysteine (Cystein) was reduced, of which 78 % Decreased to the lowest. In addition, valine related to bitterness was reduced by 11% in the vinegar treatment group.

< Example  2> Fermentation of Chunma Using Microorganisms

To investigate the fermentation effect of L. brevis by various strains of Lactobacillus brevis , Leuconostoc mesenteroides and Lactobacillus plantarum E3-8, L. mesenteroides N12-4, L. plantarum N56-12, L. mesenteroides N58-5, L. brevis N70-9, L. plantarum Six species of N76-10 were inoculated in MRS liquid medium which is commonly used for culture, and incubated at 37 ° C for 2 days. The dried cheonma was pulverized, and mixed with distilled water (w / v) corresponding to 10 times of the pulverized cheonma, and sterilized for 15 minutes at 1.5 atm and 121 ° C. in a high-temperature high-pressure sterilizer (Autoclave 80L, Hanyang Science). The sterilized cheonma mixture was cooled to room temperature, and then inoculated with the lactic acid bacteria (1%, w / v), and fermented at 37 ° C. and 150 rpm in a shaker incubator for 3 days, and samples were obtained at 1, 2, and 3 days. Lyophilized.

< Experimental Example  2> Comparison of Fermentation Chunma Using Fermented Microorganisms

Experimental Example  2-1. Fermentation Microorganism Fermentation  pH vs. acidity

The pH and acidity of the fermentation cheonma according to various fermentation strains described in Example 2 were investigated.

The pH was measured by using a pH meter (A211, Thermo Orion) was put 9mL distilled water in 1g fermented cheonma sample. Acidity was measured by dropping 2-3 drops of 1% (v / v) phenolphthalein indicator into 10 mL of fermented cheonmae sample, and then using a burette with 0.1 N (w / v) sodium hydroxide (NaOH) solution. Titration was made until it became reddish. After measuring the appropriate consumption (mL), the total acid in the sample was converted into lactic acid concentration, which is a general standard for measuring acidity, by the following formula.

[Calculation 1]

Acidity (% Lactic Acid) = ((0.009 × NaOH Consumption (mL) × NaOH Titer) / Volume of Sample) × 100

Strain Cheonma (1:10) pH Acidity Day 1 Day 2 Day 3 Day 1 2 days Day 3 CONT 4.75 0.22 LB E3-8 4.29 4.21 4.22 0.28 0.20 0.14 LM N12-4 4.75 4.34 4.18 0.20 0.14 0.16 LP N56-12 4.75 4.68 4.19 0.09 0.13 0.20 LM N58-5 4.76 4.75 4.16 0.12 0.15 0.17 LB N70-9 4.09 3.89 3.87 0.18 0.18 0.27 LP N76-10 4.75 4.75 4.2 0.12 0.12 0.17

(CONT: Day 0 of fermentation)

As a result, pH was lowered as the fermentation proceeded in all strains (Table 2).

Experimental Example  2-2. Fermentation Microorganism Fermentation  Component Change Comparison

In order to compare the change of parahydroxybenzyl alcohol (p-hydroxybenzyl alcohol) in the active ingredient of cheonma was analyzed using HPLC (High Performance Liquid Chromatography, Waters TM 2695).

100 g of 80% ethanol to 5 g of the lyophilized cheonma powder prepared in Example 2 was sonicated twice with an ultrasonic grinder (ultrasonicator, Power Sonic 420, 50/60 HZ, 700 W, Hwashin Co.) twice for 1 hour. (sonication). The ultrasonic treatment product Whatman No. 200 ml of the filtrate obtained by filtration with 2 (Whatman) filter paper was concentrated under reduced pressure with a rotary vacuum evaporator (EYELA), and then dried with a lyophilizer to prepare an extract sample. The detection was performed at 270 nm using a UV detector, the column was YMC-pack pro C18 (5 μm, 4.6 × 250 mm) and the mobile phase composition was water (solvent A) and acetonitrile (acetonitrile (solvent B)). The concentration gradient method over time was used and eluted at a rate of 1 mL per minute. The concentration gradient was formed by varying the volume ratio of solvents A and B over time.

Strain Cheonma (1:10), (mg / 100 g, building) Day 1 Day 2 Day 3 CONT 463.06 LB E3-8 543.34 596.54 480.66 LM N12-4 410.20 555.79 477.15 LP N56-12  509.95 503.98 514.92 LM N58-5 500.78 525.21 561.35 LB N70-9 547.06 540.96 583.91 LP N76-10 590.46 523.65 494.16

As a result, the parahydroxybenzyl alcohol content was significantly increased overall in the fermentation cheonma on the second day of fermentation, especially L. brevis The highest content was shown in fermentation Day 2 cheonma using E3-8 (LB E3-8) strain (Table 3).

The strain named ' Lactobacillus brevis E3-8' was deposited on 06 December 2017 at the Korean Agricultural Culture Center (KACC) in the National Academy of Agricultural Science (Accession Number: KACC 92213P).

Experimental Example  2-3. L. brevis  By E3-8 strain Fermentation  Ingredient and free amino acid changes

Changes in gastrodin, vanilyl alcohol and free amino acids in cheonma fermented by LB E3-8 strain with the highest parahydroxybenzyl alcohol content were performed in the same manner as in Experiment 2-2. It was investigated using.

ingredient
relative ion Non-fermentation Day 1 of Fermentation Day 2 of Fermentation Day 3 of Fermentation Gastrodin 15921.01 17489.03 25501.52 18998.39 Benilyl alcohol 1405.33 1249.98 1562.60 1243.88

As a result, as shown in Table 4, the gastrodine and the benyryl alcohol showed a higher content than the non-fermented group on the second day of fermentation.

In addition, the free amino acids related to taste were compared by the method disclosed in Experimental Examples 1-2.

flavor amino acid relative ion Non-fermentation Day 1 of Fermentation Day 2 of Fermentation Day 3 of Fermentation bitterness His 1325.01 1443.11 1475.70 1795.01 Arg 3545.23 1003.07 1149.95 1049.33 Tyr 7836.53 7622.11 8522.43 7709.30 Lys 1443.86 1386.14 1484.05 1375.01 Val 3150.30 3032.39 3619.70 3096.82 Ile 10131.98 8466.48 10566.00 8126.06 Phe 23171.09 20283.00 26599.94 20850.44 sweetness Ser 1011.38 1527.24 1744.83 1602.73 Pro 1783.68 2205.89 2387.85 2215.75 Umami Glu 6986.71 6608.38 6834.31 6991.51 Amino acid whole 60385.78 53577.81 64384.76 54811.96

As a result, arginine (Arg), tyrosine (Tyr), lysine (Lys), valine (Val), isoleucine (Ile) and phenylalanine (Phe) among the amino acids related to bitter taste as shown in Table 5 were reduced on the first day of fermentation, Some showed an increase in the second day of fermentation and then decreased again on the third day of fermentation. On the third day of fermentation, arginine, isoleucine and phenylalanine were significantly decreased compared to the non fermented group. In addition, serine (Pro) and proline (Pro) associated with the sweet taste was significantly increased compared to the non-fermented group regardless of the fermentation period.

< Example  3> Preparation of Fermented Chunma

Experimental Example 1 and Experimental Example 2 and L. brevis immersed in vinegar to enhance the flavor of cheonma It was found that the method of fermentation using E3-8 (LB E3-8) is useful, and after crushing the pulverized cheonma in vinegar as described in Example 1, as described in Example 2 L. brevis E3-8 It was fermented for 3 days after inoculation (LB E3-8).

< Experimental Example  3> Immersion  Analysis of Treated Fermented Chunma

Experimental Example  3-1. Immersion  Comparison of Quality Characteristics of Chunma with Different Treatments and Fermentations

Quality characteristics were investigated to compare the changes of the fermented cheonma with immersion treatment.

Each cheonma powder was divided into experimental and control groups and dissolved in sterile water. The experimental group was used fermented cheonma after vinegar soaked in the method of Example 3, the control group was L. brevis disclosed in Example 2 Chunma was inoculated with E3-8 (LB E3-8) and fermented for 3 days. Both the experimental and control groups were sampled before fermentation, day 1, day 2 and day 3 of fermentation, respectively.

The change of pH, total acidity, reducing sugar, free sugar, color and free amino acid according to immersion treatment and fermentation period of fermented cheonma was compared. Specifically, pH and total acidity were measured by the method described in Experimental Example 2-1, and the reducing sugar content was measured by adding 1 mL of DNS solution to 1 mL of the sample by DNS (dinitrosalicylic acid) method using a UV-visible spectrophotometer at a wavelength of 546 nm. Absorbance was measured. At this time, the quantification of sugar was measured by converting from the standard curve using glucose (glucose) as a standard material.

[Calculation 2]

Reducing sugar (%) = Amount of reducing sugar (mg) X dilution factor X 1 / sample (g)

Free sugar was analyzed by HPLC as described in Experimental Example 2-2, and chromaticity was measured by using a colorimeter (CR-400, Konica-Minolta).

Effective date pH Total acidity
(%) Sugar content
(Brix%) Reducing sugar
(%) GR 0 5.02 ± 0.05 0.24 ± 0.003 10.8 0.30 ± 0.000 One 4.61 ± 0.10 0.23 ± 0.003 12.8 0.23 ± 0.000 2 4.49 ± 0.16 0.28 ± 0.006 12.7 0.24 ± 0.000 3 4.47 ± 0.25 0.20 ± 0.006 12.5 0.23 ± 0.001 PGR 0 4.60 ± 0.03 0.23 ± 0.006 12.3 0.32 ± 0.000 One 4.41 ± 0.12 0.31 ± 0.006 11.5 0.29 ± 0.000 2 4.24 ± 0.13 0.32 ± 0.003 12.7 0.21 ± 0.000 3 4.22 ± 0.14 0.31 ± 0.009 12.8 0.18 ± 0.001

(GR: Fermented Chunma, PGR: Vinegar Soaked Fermented Chunma)

As a result, both pH and sugar content were lowered as the fermentation proceeded, regardless of immersion (Table 6).

Effective date Free sugar (g / 100g) Chromaticity Rhamnose fruit sugar glucose saccharose L a b GR 0 0.56 2.93 3.60 6.22 67.70 ± 1.01 5.32 ± 0.59 14.03 ± 1.48 One 0.99 3.21 4.47 8.72 66.46 ± 0.77 5.43 ± 0.27 15.20 ± 1.17 2 0.86 2.88 3.23 8.19 66.86 ± 1.10 5.31 ± 0.25 14.63 ± 0.77 3 0.81 2.67 3.09 7.30 71.57 ± 1.31 4.93 ± 0.04 14.96 ± 0.61 PGR 0 0.76 4.35 4.49 7.10 69.32 ± 0.43 5.30 ± 0.12 14.78 ± 0.26 One 0.83 3.36 3.60 6.74 69.81 ± 0.97 5.15 ± 0.12 14.83 ± 0.14 2 1.19 2.04 3.53 7.23 68.33 ± 0.12 5.27 ± 0.04 14.51 ± 0.20 3 1.20 1.76 2.68 7.61 66.97 ± 0.52 5.20 ± 0.04 13.76 ± 0.04

In the case of free sugar content, rhamnose tended to increase as the fermentation progressed in the experimental group, and fructose (fructose) showed the highest content in the non-fermented state in the experimental group. (sucrose) was found to have the highest content on day 1 of fermentation in the control (Table 7).

Free amino acids Fermentation period (days), (mg / 100g) GR-0 GR-1 GR-2 GR-3 PGR-0 PGR-1 PGR-2 PGR-3 Asp 19.45 21.53 23.36 44.43 35.31 35.73 36.6 19.64 Thr 14.14 6.19 5.14 13.2 17.79 14.08 11.55 5.38 Ser 44.19 26.82 23.59 44.83 45.03 39.81 37.58 23.55 Glu 122.1 136.47 143.5 163.35 131.53 138.26 138.64 122.71 Gly 25.36 30.33 32.7 61.7 46.8 51.38 51.58 28.57 Ala 61.98 62.09 64.81 79.82 75.27 75.92 66.91 58.05 Cit ND 5.57 5.72 1.99 ND ND ND 2.27 a-ABA 0.29 0.18 0.19 0.39 0.33 0.18 0.3 0.16 Val 12.33 12.66 12.67 14.82 11.9 13.34 12.25 11.44 Met 0.33 ND ND ND 0.59 ND ND ND Ile 6.66 6.94 6.91 7.05 5.7 6.43 5.68 6.26 Leu 5.77 5.63 5.64 7.5 5.76 6.73 5.77 5.11 Tyr 8.35 9.12 9.4 11.2 12.07 10.97 9.78 8.24 Phe 13 14.88 14.52 15.36 12.12 14.04 13.09 12.92 g-ABA 5.24 6.73 7.52 5.81 4.21 4.27 4.42 6.89 NH3 6.57 9.66 9.95 16.2 9.36 14.48 13.49 8.81 Orn 0.28 5.07 5.86 11.6 0.24 8.25 9.99 5.04 Lys 4.34 4.2 4.37 3.83 5.28 4.14 3.42 3.89 His 2.48 ND ND ND 3.74 ND ND ND Arg 10.25 0.62 0.22 0.5 18.36 0.19 0.39 0.25 Amino acid total amount 397.06 402.22 414.82 600.16 516.96 516.54 502.02 361.97 Total amount of essential amino acids 56.57 50.51 49.26 61.74 59.13 58.75 51.76 44.99

In the case of free amino acid content in the experimental group of vinegar-immersed fermented cheonma (PGR), the bitter taste was found to decrease significantly with the progress of the fermentation (Fig. 2 and Table 8).

Experimental Example  3-2. Immersion  Comparison of Active Ingredients and Flavor Compounds of Chunma during Treatment and Fermentation

In order to compare the change in the active ingredient and the flavor component according to the immersion treatment and fermentation period of the fermented cheonma was analyzed using HPLC as in Experimental Example 2-2.

Effective date p-hydroxybenzyl alcohol Gastrodin GR 0 592.67 333.58 One 657.63 327.30 2 602.28 307.88 3 562.05 292.77 PGR 0 550.53 209.13 One 554.11 203.77 2 537.84 194.20 3 609.15 215.33

In the experimental group, vinegar-immersed fermentation Chunma (PGR) showed a high content of parahydroxybenzyl alcohol on the third day of fermentation, and gastrodine was maintained at a constant level (Table 9).

RT Mass Classification Area (10 ⁶ ) GR-0 GR-1 GR-2 GR-3 PGR-0 PGR-1 PGR-2 PGR-3 Furan, 2-pentyl- 10: 16.6 81 furan 11 11 10 8 13 7 4 4 Diethylene glycol 11: 41.8 45 glycol 20 4 ND ND 23 12 ND 8 2-Propanone, 1-hydroxy- 12: 04.3 43 ketone 83 9 ND ND 67 19 5 ND Acetic acid 16: 02.6 60 acid 142 2953 3131 2429 3788 4941 6224 3619 Furfural 16: 23.3 95 furfural 745 65 24 13 737 74 36 16 Ethanone, 1- (2-furanyl)- 17: 24.4 95 furan ketone 23 13 ND ND 26 26 15 11 2,3-Butanediol 19: 18.6 45 glycol 159 75 47 29 219 192 150 60 Hexanoic acid 25: 20.4 60 acid 43 45 44 36 38 36 37 33 Creosol 27: 13.3 123 alcohol 18 43 72 86 12 29 50 50 Phenol 28: 30.4 94 phenol 26 21 20 15 20 17 15 16 2-Methoxy-4-vinylphenol 32: 04.7 150 phenol 18 15 13 9 20 12 13 8 Ar-tumerone 33: 09.2 83 5 ND ND ND 25 17 22 20 Hexadecanoic acid, ethyl ester 33: 10.4 88 ester 32 14 21 25 42 6 30 84 4H-Pyran-4-one 2,3-dihydro-3,5-dihydroxy-6-methyl- 33: 26.1 144 pyranketone 15 17 13 11 12 13 8 4 5-Hydroxymethylfurfural 37: 19.0 97 furfural 33 ND ND ND 63 23 6 3 Phenol, 4- (methoxymethyl)- 37: 23.6 107 phenol 20 22 17 14 13 16 13 9 Vanillin 38: 17.8 151 22 25 16 10 22 27 23 15

In addition, as a result of analysis of various fragrance components of fermented Chunma, phenolic components that cause discomfort and irritate skin and respiratory tract decreased as fermentation progressed, and ester-based components showed a sharp decrease on day 1 of fermentation. 10).

Experimental Example  3-3. Immersion  Comparison of Antioxidant Substances in Chunma by Different Treatments and Fermentations

The difference between the active ingredients of polyphenols and flavonoids, which are antioxidant substances, and the antioxidant activity were compared in the difference of the active ingredients of fermented cheonma according to the immersion treatment.

Total polyphenol content was measured by applying the Folin-Denis method. 1 ml of Folin reagent was added to 100 µl of 80% ethanol extract of fermented Chunma, and then allowed to stand at room temperature for 3 minutes, and 1 ml of 10% Na ₂ CO ₃ solution was added and allowed to stand at room temperature for 1 hour, followed by a microplate reader. Absorbance at 765 nm was measured. The absorbance was measured using gallic acid to prepare a standard curve, and the total phenol content was calculated. Total flavonoid content was determined using the Moreno method. To 0.5 ml of 80% ethanol extract of fermented Chunma, 0.1 ml of 10% aluminum nitrate, 0.1 ml of 1 M potassium acetate, and 4.3 ml of ethanol were mixed and left to stand at room temperature for 30 minutes, followed by a microplate reader. Absorbance was measured at 415 nm using. The absorbance was measured using a quercetin (Quercetin) to prepare a standard curve, the total flavonoid content was calculated. Antioxidant activity was observed through DPPH scavenging ability. After adding 180 mM 0.2 mM DPPH (1,1-diphenyl-2-picrylhydrazyl, Sigma-Aldrich Co.) solution to 20 µl of fermented Chunma extract, it was allowed to stand at room temperature for 10 minutes, and then absorbed at 517 nm using a microplate reader. Was measured. In the control group, 180 μl of DPPH solution was added to 20 μl of 80% ethanol, and the absorbance was measured at 517 nm after standing at room temperature for 10 minutes.

Effective date DPPH (%) Total Polyphenols (mg / g) Flavonoids (mg / g) GR 0 43.34 ± 0.42 15.39 ± 0.08 0.96 ± 0.019 One 46.80 ± 0.54 17.48 ± 0.02 0.71 ± 0.040 2 39.98 ± 0.74 18.03 ± 0.07 0.88 ± 0.113 3 44.53 ± 3.30 15.67 ± 0.05 0.99 ± 0.062 PGR 0 39.47 ± 0.31 15.39 ± 0.25 0.37 ± 0.041 One 43.02 ± 0.72 16.55 ± 0.20 0.47 ± 0.043 2 35.00 ± 0.03 13.81 ± 0.13 0.84 ± 0.086 3 40.52 ± 0.91 16.56 ± 0.23 0.80 ± 0.091

As a result, the total polyphenols of both the control fermentation cheonma (GR) and the experimental group vinegar immersion fermentation cheonma (PGR) almost increased compared to before the fermentation (Table 11).

< Example  4> Preparation of Fermented Chunma Extract

After extracting the fermented Chunma powder as in Example 3 using 80% ethanol and water for 1 hour, the supernatant was collected and Whatman No. 2 (Whatman) filter paper, and the filtrate was concentrated under reduced pressure using a rotary vacuum evaporator (YEYELA), and dried using a lyophilizer.

<Experiment 4> Comparison of the dipping treatment and Gastrodia elata extract the fermentation process

Experimental Example  4-1. Immersion  Comparison of Quality Characteristics of Chunma Extracts with Different Treatments and Fermentations

The pulverized cheonma was immersed in vinegar as described in Example 1, and then inoculated with L. brevis E3-8 by the method described in Example 2, and fermented for 5 days to be an experimental group (PGR), which was not immersed in vinegar. L. brevis E3-8 was inoculated on the sperm and fermented for 5 days to serve as a control (GR). Each cheonma powder is divided into experimental group (PGR) and control group (GR), and all samples are collected before fermentation, 1st fermentation, 2nd fermentation, 3rd fermentation, 4th fermentation and 5th fermentation, and dissolved in sterile water. did. Changes of pH, total acidity, reducing sugar, and free sugar according to the immersion treatment and fermentation period of the fermented cheonma sample were compared.

pH and total acidity were measured by the method disclosed in Experimental Example 2-1, and reducing sugars were measured by the method disclosed in Experimental Example 3-1.

Effective date pH Total acidity (%) Reducing Sugar (%) GR 0 5.27 ± 0.01 0.09 ± 0.006 0.14 ± 0.001 One 5.08 ± 0.02 0.13 ± 0.028 0.14 ± 0.001 2 4.81 ± 0.02 0.09 ± 0.022 0.14 ± 0.001 3 4.81 ± 0.02 0.12 ± 0.018 0.14 ± 0.001 4 4.78 ± 0.01 0.12 ± 0.010 0.15 ± 0.001 5 4.74 ± 0.01 0.12 ± 0.006 0.14 ± 0.001 PGR 0 4.80 ± 0.01 0.12 ± 0.003 0.26 ± 0.002 One 4.78 ± 0.02 0.20 ± 0.015 0.34 ± 0.004 2 4.66 ± 0.02 0.16 ± 0.009 0.27 ± 0.004 3 4.59 ± 0.03 0.22 ± 0.006 0.32 ± 0.003 4 4.55 ± 0.01 0.19 ± 0.016 0.39 ± 0.031 5 4.37 ± 0.01 0.24 ± 0.027 0.17 ± 0.011

As a result, both the fermented cheonma (GR) and vinegar soaked fermented cheonma (PGR) reduced the pH as the fermentation by lactic acid bacteria inoculated, which is consistent with the known fact that lactic acid and various organic acids are produced during fermentation by lactic acid bacteria. As a result, it was confirmed that fermentation proceeded normally (Table 12).

In addition, the ethanol extract and the water extract were prepared by the method disclosed in Example 4, and the free sugar content of the extract of Chunma (GR) and vinegar immersion Chunma (PGR) was measured by HPLC as described in Experimental Example 2-2, and fructose ( fructose), glucose (glucose) and sucrose (sucrose) content was compared.

Effective date Free sugar (g / 100g raw material) 80% EtOH Extract Water extract fructose glucose sucrose fructose glucose sucrose GR 0 0.14 0.21 0.30 0.15 0.25 0.13 One 0.27 0.38 0.57 0.08 0.11 0.16 2 0.24 0.39 0.56 0.12 0.18 0.26 3 0.13 0.22 0.36 0.08 0.11 0.16 4 0.18 0.30 0.50 0.13 0.21 0.34 5 0.15 0.26 0.43 0.08 0.12 0.21 PGR 0 0.32 0.45 0.18 0.06 0.10 0.18 One 0.33 0.49 0.19 0.15 0.20 0.08 2 0.46 0.67 0.26 0.21 0.30 0.13 3 0.32 0.50 0.22 0.15 0.21 0.10 4 0.32 0.49 0.21 0.11 0.16 0.07 5 0.32 0.53 0.28 0.15 0.23 0.11

As a result, the free sugar content of fermented cheonma (GR) and vinegar soaked fermented cheonma (PGR) was higher than that of water extract in 80% ethanol extract. It was found to be the highest (Figure 3 and Table 13).

Experimental Example  4-2. Immersion Of Active Compounds of Chunma Extracts According to Differentiation, Fermentation and Extraction Solvent

Changes in the amount of active ingredient according to the fermentation period of cheonma and vinegar dipping cheonma extract. Gastrodin, P-hydroxybenzyl alcohol (Gastrodigenin), Benylyl alcohol, Parahydroxy among the active ingredients contained in the cheonma by HPLC as described in Experimental Example 2-2 Benzyl aldehyde (p-hydroxybenzaldehyde) and benilin (Vanilin) were analyzed.

Effective date
80% EtOH Extract (g / 100g Original) Gastrodin Gastrodigenin Vanillyl alcohol P-hydroxybenzaldehyde vanillin Total GR 0 16.02 35.00 2.82 0.58 0.03 54.45 One 30.77 69.24 2.72 1.02 0.04 103.79 2 32.59 77.96 3.39 1.18 0.04 115.16 3 19.26 40.98 1.91 0.65 0.02 62.82 4 25.13 52.78 2.66 0.82 0.02 81.41 5 22.56 47.28 2.26 0.69 0.02 72.81 PGR 0 34.19 39.76 1.92 1.09 0.06 77.02 One 33.47 40.61 1.99 1.23 0.06 77.36 2 56.71 63.65 3.39 1.33 0.06 125.14 3 40.02 43.70 2.60 0.90 0.03 87.25 4 40.01 47.13 2.67 1.12 0.04 90.97 5 65.12 80.62 4.31 1.41 0.04 151.50

As a result, the 80% ethanol extract showed a tendency to increase after the fermentation cheonma (GR) up to the second day of fermentation, and the greatest increase on the fifth day of fermentation of vinegar-immersed fermentation cheonma (PGR) (FIG. 4, FIG. 5 and Table 14).

Effective date
Water Extract (g / 100g Raw Material) Gastrodin Gastrodigenin Vanillyl alcohol P-hydroxybenzaldehyde vanillin Total GR 0 19.31 14.79 1.63 0.19 0.01 35.93 One 24.71 19.46 1.92 0.28 0.02 46.39 2 35.16 30.56 2.96 0.47 0.03 69.18 3 24.15 18.68 1.92 0.23 0.02 45.00 4 42.93 35.07 3.87 0.52 0.03 82.42 5 34.89 24.22 2.69 0.19 0.03 62.02 PGR 0 36.42 15.83 1.91 0.34 0.02 54.52 One 41.61 18.64 2.19 0.44 0.03 62.91 2 52.27 23.97 3.06 0.50 0.03 79.83 3 48.95 22.08 2.77 0.43 0.03 74.26 4 36.37 16.37 2.03 0.33 0.02 55.12 5 46.84 21.41 2.81 0.34 0.02 71.42

As a result, in the case of the water extract, the fermentation cheonma (GR) was found to have the highest increase of the components on the 4th day of fermentation and the vinegar immersion fermentation cheonma (PGR) on the second day of fermentation (Fig. 4, Fig. 5 and Table 15).

<Experiment 5> Anti-inflammatory Effect of Gastrodia elata extract according to the immersion, fermentation and extraction solvent

Experimental Example  5-1. Immersion Cytotoxicity of Chunma Extracts According to Different Fermentations, Fermentations, and Solvents

Cell viability was measured by MTT assay in order to compare the cytotoxicity of the fermented cheonma extract according to immersion treatment, fermentation period and extractant. The MTT assay measures the extent to which MTT tetrazolium is reduced to MTT formazan [3- (4,5-dimethylthiazol-2-yl) -2,5-diphenyl-tetrazoliumbromide] by dehydrogenase activity in mitochondria of living cells. In this experimental example, human umbilical vein endothelial cells (HUVECs) were used.

Specifically, 100 extracts of Example 4 in which the density of cells was immersed in vascular endothelial cells (HUVEC) of 80% or more (GR / PGR) and the fermentation period (0/1/2/3/4/5 days) were different. Treatment was performed by μg / mL. 10% FCS (cal serum; GE Healthcare Bio-Sciences Co.) diluted with 2 mg / ml (in PBS) of Sigma-Aldrich Co. (MTT) to 200 μg / ml and penicillin at 100 unit / ml ) Was replaced with DMEM medium containing streptomycin and incubated for 1 hour in a 37 ° C. incubator. After the incubation was completed, the medium was completely removed, and purple formazan was dissolved by adding 300 µl of DMSO. The dissolved formazan was placed in a 96-well plate in 100 μl and the absorbance was measured at 595 nm using a microplate reader. Using the absorbance of untreated vascular endothelial cells as a control, the cell viability was calculated (% of Control) and the cytotoxicity of cheonma extract was evaluated.

Effective date
Cell viability (% of control) 80% EtOH Extract
(100 µg / ml) Water extract
(100 µg / ml) Control 99.85 ± 6.66 GR 0 98.22 ± 0.59 106.07 ± 2.81 One 103.99 ± 3.11 102.37 ± 1.18 2 106.95 ± 0.15 99.11 ± 2.37 3 95.27 ± 2.07 110.65 ± 0.89 4 93.20 ± 1.48 107.84 ± 0.15 5 79.88 ± 0.00 95.27 ± 2.07 PGR 0 97.04 ± 0.89 109.02 ± 1.04 One 96.60 ± 3.70 105.62 ± 0.89 2 108.28 ± 3.70 106.21 ± 1.78 3 100.59 ± 2.66 106.51 ± 2.66 4 97.49 ± 2.51 110.21 ± 1.04 5 91.57 ± 1.63 100.89 ± 1.18

As a result, as shown in Table 16, the cytotoxicity of the fermentation cheonma (GR) and vinegar-immersed fermentation cheonma (PGR) extracts with different fermentation periods and extraction solvents was ascertained. It was confirmed that there was no cytotoxicity by more than%.

Experimental Example  5-2. Immersion Comparison of NO and PGE2 Inhibitory Effects of Cheonma Extracts with Different Fermentations, Fermentations and Solvents

In order to compare the anti-inflammatory effects according to the immersion treatment, fermentation period and extraction solvent of cheonma extract, NO (Nitric Oxide) production and PGE2 (Prostaglandin E2) production were measured.

Specifically, 100 μg / mL of the extract of Example 4 having different immersion (GR / PGR) and fermentation period (0/1/2/3/4/5 days) was used to prepare RAW264.7 cells having a cell density of 80% or more. Pretreated for 4 hours. After pretreatment RAW264.7 cells were stimulated with LPS (1 μg / mL) for 20 hours and the medium was obtained by centrifugation (2,000 g, 4 ° C., 10 minutes). The treatment group was compared with the control group without any treatment and the inflammation induction group induced inflammation with LPS without pretreatment with cheonma extract. The concentration of NO was reacted with the same ratio of the medium (100 µl) and Griess reagent (0.1% naphthylethylenediamine dihydrochloride + 1% sulfanilamide + 5% phosphoric acid). Absorbance was measured at a wavelength of 540 nm with a microplate reader, and PGE2 was measured using PGE2 enzyme immunoassay (EIA) kit (Cayman Chemical Company) based on the concentration of the standard solution.

Effective date NO production (% of control) PGE2 production (% of control) 80% EtOH Extract
(100 µg / ml) Water extract
(100 µg / ml) 80% EtOH Extract
(100 µg / ml) Water extract
(100 µg / ml) Control 100.1 ± 1.07 99.85 ± 6.66 LPS 149.2 ± 1.07 123.53 ± 2.59 GR 0 129.0 ± 0.00 127.9 ± 1.07 98.22 ± 0.59 106.07 ± 2.81 One 122.6 ± 0.00 117.2 ± 1.07 103.99 ± 3.11 102.37 ± 1.18 2 135.4 ± 0.00 135.4 ± 4.72 106.95 ± 0.15 99.11 ± 2.37 3 130.0 ± 1.07 132.2 ± 1.07 95.27 ± 2.07 110.65 ± 0.89 4 122.6 ± 0.00 123.6 ± 1.07 93.20 ± 1.48 107.84 ± 0.15 5 131.1 ± 2.14 124.7 ± 0.00 79.88 ± 0.00 95.27 ± 2.07 PGR 0 129.0 ± 2.14 122.6 ± 2.14 97.04 ± 0.89 109.02 ± 1.04 One 115.1 ± 1.07 116.2 ± 0.00 96.60 ± 3.70 105.62 ± 0.89 2 127.9 ± 1.07 129.0 ± 0.00 108.28 ± 3.70 106.21 ± 1.78 3 123.6 ± 1.07 120.4 ± 0.00 100.59 ± 2.66 106.51 ± 2.66 4 118.3 ± 0.00 115.1 ± 1.07 97.49 ± 2.51 110.21 ± 1.04 5 126.8 ± 0.00 124.7 ± 0.00 91.57 ± 1.63 100.89 ± 1.18

As a result. Both NO and PGE2 induced by LPS showed an inhibitory effect by fermentation cheonma extract. Especially, after the fermentation of normal cheonma, ethanol extract showed a significant inhibitory effect of PGE2 after the second day of fermentation, and water extract was relatively No change in PGE2 inhibitory effect was apparent. Ethanol extracts and water extracts from vinegar-immersed fermented cheonma showed strong NO reduction effect on the 1st and 4th day of fermentation, and the inhibitory effect was repeated. In the case of PGE2, the inhibitory effect of the ethanol extract tended to be stronger after the second day of fermentation of the general fermentation Chunma, and the water extract showed a relatively small increase in the inhibitory effect according to the fermentation period. Through this, NO and PGE inhibitory effect was judged that the ethanol extract is larger than the water extract (Fig. 6 and Table 17).

Experimental Example 5-3. Comparison of NF-κB Inhibitory Effects of Chunma Extracts with Dipping, Fermentation and Solvents

The inhibitory effects of NF-κB on immersion treatment, fermentation period and extraction solvent of cheonma extract were compared.

Specifically, the extract of Example 4, which differs in immersion (GR / PGR) and fermentation period (0/1/2/3/4/5 days), is fermented cheonma extract in raw-blue cells having a cell density of 80% or more. Final concentrations of (FCL) were treated differently at 10, 50, 100 μg / mL, and the induction of inflammatory responses with LPS was followed by production of NF-κB compared to the control. The amount of NF-κB produced was measured three times at 620 nm after 20 μl of RAW BLUE cells and 200 μl of QUANTI-Blue (InvivoGen) were reacted in the dark at 37 ° C. for 15 minutes.

As a result, the ethanol extract and the ethanol extract of vinegar immersion fermentation of the general cheonma at 100 ㎍ / mL appeared to increase the inhibitory effect of NF-κB as the fermentation period has elapsed (Fig. 7).

Claims (11)

1) after washing the horse, drying and grinding;
2) step 1) immersing the ground cheonma in 1-20% vinegar solution for 1-9 hours;
3) after immersion in step 2), water bath for 1 to 3 hours, and dried to prepare vinegar dipping cheonma powder; And
4) Inoculated with lactic acid bacteria of Lactobacillus brevis E3-8 strain of accession number KACC 92213P deposited in Korean Agricultural Culture Collection (KACC) in National Agricultural Research and Development Institute Inoculation to ferment for 2 days at 35 to 40 ℃;
Component containing an unpleasant, spicy and bitter taste is reduced, the sweetness component is increased, the content of the pharmacologically active ingredient is increased, the method of producing fermented cheonma showing an antioxidant effect and an anti-inflammatory effect,
The discomfort component is Ar-tumerone or tumerone,
The spicy ingredient is Cystein,
The bitter taste component is at least one amino acid selected from the group consisting of arginine (Arg), tyrosine (Tyr), lysine (Lys), valine (Val), isoleucine (Ile) and phenylalanine (Phe),
The sweet component is an amino acid of Serine (Ser) or Proline (Pro),
The pharmaceutical active ingredient is gastrodin, gasodine, p-hydroxybenzyl alcohol (Gastrodigenin), benzyl alcohol, vanylyl alcohol, parahydroxybenzyl aldehyde (p-hydroxybenzaldehyde), and vanillin (Vanilin). At least one selected from the group consisting of, characterized in that the content of the benzyl alcohol is increased, the method for producing a fermented cheonma.
The method of claim 1,
The fermentation process of step 4) further comprises the step of extracting using a solvent after fermentation, fermentation cheonma. delete delete delete delete delete delete delete delete delete

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