KR102094084B1 - Aged and complex-fermented mountain-cultivated ginseng having increased ginsenosides of human body absorption type and omega-6 fatty acid, and preparation method thereof - Google Patents

Abstract

The present invention relates to aged and complex-fermented mountain-cultivated ginseng having significantly increased contents of ginsenosides Rd2, Rg3, and F2 and compound K which are easily absorbable into a human body, omega-6 fatty acids (ω-6 fatty acid: linoleic acid and arachidonic acid), GABA, phenolics, and flavonoids, and a preparation method thereof. According to the present invention, the aged and complex-fermented mountain-cultivated ginseng has remarkably increased contents of ginsenosides Rd2, Rg3, and F2 and compound K, omega-6 fatty acids, GABA, phenolics, and flavonoid, and has excellent antioxidant effects, thereby being used as a material for functional foods and medicines. The functional foods and cosmetics according to the present invention have excellent antioxidant activity, are useful for alleviation of the reduction in the immunity due to oxidative stress by radicals or the like, enhancement of cognitive ability and fatigue recovery, and are also useful for edible cosmetics and skin beauty care.

Description

Aged and complex-fermented mountain-cultivated ginseng having increased ginsenosides of human body absorption type and omega-6 fatty acid, and preparation method thereof}

The present invention relates to a complex fermentation product of a wild-fermented ginseng-aged ginsenoside and omega-6 fatty acid, and a method of manufacturing the same, more specifically, alternately fermenting / sealing and fermenting / opening fermented goats four times Ginsenosides Rd2, Rg3, F2 and compound K, omega-6 fatty acids (ω-6 fatty acids; easy to absorb by human body) are prepared by complex fermentation with Lactobacillus Brevis BMK484 strain and Lactobacillus plantarium P1201 strain after performing Linoleic acid and arachidonic acid), gaba, phenolics, and flavonoids, and the content of the fermented wild ginseng aging complex fermentation is significantly improved.

Ginseng is divided into ginseng (人蔘), wild ginseng (山 蔘), or wild ginseng (山羊 蔘) according to the difference between cultivation environment and natural growth and morphological differences. In particular, wild ginseng belongs to the family Ogapiaceae, and refers to artificially grown wild ginseng seeds or ginseng grown wild in the wild under the mountain forest by the perennial vegetation, Panax ginseng . Also, wild ginseng grown wild in the wild has been considered the best wild ginseng and is also called camphor ginseng. In the case of ginseng, most of them have 3-7 outcrops (heads), but many of them are older and older, and the roots of ginseng are thick and short, but long and thin. It is known that goats' ginseng is more effective than cultivated ginseng in many efficacy, but there is not much scientific evidence to support this compared to ginseng.

It has been reported that Samun affects the central nervous system, endocrine system, immune system, metabolic system, etc. and has various pharmacological effects on the regulation of body functions, and skin health effects such as whitening, wrinkle improvement, moisturizing and anti-inflammatory effects are also reported. It is known that such a pharmacological effect is due to easy absorption by the human gut or skin cells, that is, human absorption type ginsenoside.

Human absorption type ginsenosides include ginsenosides Rg1, Rg3, Rd, Rf, F2, compound K, and the like. Ginsenoside Rg1 improves learning function, anti-fatigue action, ginsenoside Rf acts as a brain neuron analgesic, inhibits lipid and oxidation, ginsenoside Rg3 suppresses cancer cell metastasis, protects liver, inhibits anticancer drug resistance, ginsenoside Rd series has been shown to promote adrenal cortical hormone secretion, ginsenoside compound K has anti-cancer, liver-protecting, skin-protecting, tumor-proliferating, antioxidant, and anti-allergic effects. Ginsenoside F2 has recently been known to reduce inflammation and reduce itching of atopic skin. Ginsenoside Compound K (20-O-beta-D-glucopyranosyl-20 (S)-protopanaxadiol) is a ginsenoside metabolite, which is known to have excellent absorption rate in the body and excellent pharmacological activity. , Neuropathic pain treatment, whitening and wrinkle improvement effects have also been reported (Registration Patent 10-1539573, Patent Registration 10-1300775, Patent Publication 10-2014-0003198).

However, these human-absorbed ginsenosides are rarely contained in ginseng itself, and several techniques have been developed to convert ginsenosides contained in ginseng into human-absorbed types using enzymes to solve this. This high or human absorption type of ginsenosides has a problem of low enhancement, so there is still a need to develop a goat's ginseng processed product with improved body absorption type ginsenosides.

The fatty acid has a carboxyl group (-COOH) at one end and a methyl group (-CH ₃ ) at the other end, and depending on the number of carbons from the carbon (ω carbon) to which the methyl group is attached, omega-3 (ω- 3) or omega-6 (ω-6) fatty acids. Omega-6 fatty acids refer to unsaturated fatty acids with a double bond to the 6th carbon from ω carbon, and include linoleic acid and arachidonic acid. Since omega-6 fatty acids are not synthesized in animals, they are called essential fatty acids, and corn oil and peanut oil, vegetable oils, are known as source foods. Omega-6 fatty acids are effective in lowering blood cholesterol levels.

Gaba (Gamma-aminobutyric acid, GABA) is a non-protein amino acid composed of 4 carbons. Gaba is released by the release of carbon dioxide along with decarbonation reaction from glutamic acid (GA) by glutamate decarboxylase (GADase). Is converted. Gaba not only has a role as a neurotransmitter in the brain, but also has a wide variety of physiological functions such as promoting brain function, stabilizing action, lowering blood pressure, diuretic action, improving liver function, preventing obesity, promoting alcohol metabolism, and deodorizing action. In addition, it is registered as a medicine and is used to treat headaches, tinnitus, and decreased motivation due to stroke or sequelae of cerebral arteries. Normally, the human body produces gaba, but there is a limit to the production of gaba in the body since aging progresses due to excessive consumption of inappropriate foods or additives, or insufficient intake of vitamins or minerals. Accordingly, it is necessary to consume foods containing gaba.

In relation to the processed products of wild ginseng, the processed products of wild ginseng with significantly improved contents of human absorbed ginsenoside, omega-6 fatty acid, gaba, phenolics, and flavonoids have not been developed.

As a result, the present inventors continued research to meet the demands of the prior art, and after performing ginseng ginseng steaming and sealing ripening, steaming and open aging alternately four times, Lactobacillus brevis BMK484 strain and Lactobacillus plantarium P1201 When processed by fermentation with a strain, it was confirmed that the human absorbed ginsenosides Rd2, Rg3, F2 and compound K, omega-6 fatty acids (linoleic acid and arachidonic acid), gaba, phenolics, and flavonoids contents were significantly improved. The present invention has been completed.

Accordingly, an object of the present invention is to provide a ginsenoside Rd2, Rg3, F2 and compound K, omega-6 fatty acids, gaba, phenolics, and wild ginseng ripening complex fermentation with enhanced flavonoids content.

Another object of the present invention is to provide a method for producing a ginsenoside Rd2, Rg3, F2 and compound K, omega-6 fatty acids, gaba, phenolics, and wild ginseng ripening complex fermentation with enhanced flavonoids content.

Another object of the present invention is to provide a functional food for improving immunity, improving cognitive ability, restoring fatigue, and eating cosmetics (inner beauty) containing the wild-fermented ginseng fermentation compound of the present invention.

Another object of the present invention is to provide a functional cosmetic for improving whitening and wrinkles, including the wild-fermented ginseng fermentation compound of the present invention.

In order to achieve the above object, the present invention includes ginsenoside Rd2, Rg3, F2 and compound K, omega-6 fatty acids, gaba, phenolics, and flavonoids. Provides a method of making water:

Iv) alternately performing steaming and sealing aging of mountain goats and steaming and open aging four times; And

Ii) Fermentation with a complex strain of Lactobacillus brevis BMK484 strain and Lactobacillus plantarium P1201 strain.

The production method of the present invention may further include the step of drying and pulverizing the wild-fermented ginseng fermentation compound from step ii).

Step iv): Steaming and ripening goats

Goat ginseng is steamed and sealed and steamed and open aged alternately four times.

In the present invention, 'goat ginseng' means goat or ginseng sprouts.

In the present invention, 'goat sprout sprouts' is grown in a mountainous region of 500 m or more above sea level for more than 3 years, from May to July, the roots, leaves and stems of wild ginseng ginseng ginseng outposts or ginseng ginsengs from wild ginsengs for more than 2 years in an artificial environment for 90 days It means cultivated hydroponic or indigenous soil. On the other hand, during the cultivation of goat sprouts, it is an eco-friendly agricultural product that does not use any pesticides, and it has advantages over goats because it forms a lower price than conventionally used goats due to hydroponic or land cultivation, year-round production, and shorter cultivation period.

In the present invention, it is preferable to use goat sprouts grown in May or wild plants grown in a natural environment for 30 days in an artificial environment, and more preferably, goats sprouts cultured in an artificial environment for 30 days. Hemp is used.

Goat ginseng and goat ginseng can be used as outposts, including roots, leaves, and whole stems.

Wash 3 times in running water of goat or ginseng sprouts, remove water and steam.

Steaming can be performed using a steaming machine such as a conventional steamer, and is performed at 100 ° C for 30 to 120 minutes. If the steaming time is less than 30 minutes, sufficient steaming does not proceed, and subsequent steps such as ripening may not be smooth and contamination of germs may occur. When evaporated for more than 120 minutes, the long-term heat treatment may destroy the nutritional components of goats or ginseng sprouts.

When steaming, if necessary, water can be added to goats or goat sprouts so that the water content is 40 to 50%.

The sealed aging is carried in a sealed aging machine (container) containing steamed goat or ginseng sprouts for 2 to 3 days at 70 to 80 ℃.

Open aging is carried out for 2 to 3 days at 60 to 70 ° C after placing the boiled wild ginseng or goat sprouts in an open aging machine.

Closed aging and open aging are performed alternately (alternately), and the order of closed aging and open aging is irrelevant.

As illustrated in the following examples (manufacturing examples) as an example, wild ginseng is first steamed and first sealed matured, second steamed and second open matured, third steamed and tertiary sealed matured, fourth steamed And the fourth open aging.

The fermentation and maturation of wild ginseng or goat sprout ginseng as described above is sufficiently matured and decomposition of the produced bioactive material is minimized.

Step ii) Combined fermentation

The fermented and matured wild goat or goat sprout from step iii) is fermented with a complex seed of the Lactobacillus brevis BMK484 strain and the Lactobacillus plantarium P1201 strain.

One strain constituting the complex seed in the present invention, Lactobacillus brevis BMK484 strain, was isolated / identified from the kimchi by the present inventors and deposited on December 12, 2016 at the National Academy of Agricultural Science, Agricultural Genetic Resource Center (KACC). With the number KACC92157P), the productivity of Gaba and Ginsenoside Compound K are excellent (Patent Application 10-2017-0152033).

Another strain constituting the complex spawn in the present invention, Lactobacillus plantarium P1201 strain, was isolated / identified from the fermented foods by the present inventors on July 19, 2013 at the National Academy of Agricultural Science, Agricultural Genetic Resource Center (KACC). As a deposited strain (Accession No. KACC91848P), it has excellent probiotic performance and excellent bioactive material productivity (Patent Registration No. 10-154418).

The Lactobacillus brevis BMK484 strain and the Lactobacillus plantarium P1201 strain constituting the complex seed of the present invention exhibit synergistic effects by complementing the fermentation properties. Lactobacillus brevis BMK484 strain is a hetero-lactic acid fermentation lactic acid bacteria that does not produce sufficient lactic acid when fermented alone, and the Lactobacillus plantarum P1201 strain is a homo-lactic acid fermentation lactic acid fermentation alone The lactic acid is excessively produced. Surprisingly, the fermentation properties of both strains are complemented by the use of these two species together with wild goat or goat sprout, and human absorption type ginsenoside, omega-6 fatty acid, gaba, Phenolics and flavonoids contents were significantly enhanced (Test Examples 1-4).

In the present invention, the complex seed may be added as a separate culture medium for each of the Lactobacillus brevis BMK484 strain and the Lactobacillus plantarium P1201 strain, or may be added as a mixed culture mixture.

The mixing ratio of the two strains in the complex seed can be mixed in a ratio of 3: 1 to 1: 3 (v / v).

Fermentation may be performed by inoculating the culture medium of the complex seed with 2-10% (v / w) concentration of boiled and aged goats or goats at a concentration of 2-10% (v / w) for 2-7 days at 25-40 ° C.

When the inoculation dose is less than 2% (v / w), the fermentation rate may be delayed, and when it exceeds 10% (v / w), the cell growth rate is fast, resulting in low conversion of active ginsenosides and excessive sour taste. If the fermentation temperature is less than 25 ℃, the fermentation period becomes longer, causing contamination of germs, and when it exceeds 40 ℃, the growth of the strain may be stopped. The production of active substances and the like may be poor, and when it exceeds 7 days, the bioactive substances may be decomposed by over-fermentation.

In addition, upon fermentation, if necessary, water is adjusted and / or sterilized to 40 to 50% (v / w) by adding water to steamed and matured wild ginseng or wild goose sprouts, and then inoculated with a complex seed to ferment. You can. The fermentation efficiency is highest when the moisture content is in the above range. If the moisture content is less than 40%, the microorganism does not have sufficient moisture for survival or growth, and smooth fermentation does not proceed. In the case of more than 50%, drying time and cost may be high after the fermentation is completed. Smooth fermentation may not proceed due to contamination of various bacteria.

The fermented ginseng fermented fermented ginseng produced by processing in the above-described steps of the present invention has a ginsenoside Rd2 of 1.5 times or more, a ginsenoside Rg3 of 8.1 times or more, and a ginsenoside F2 of 11.6 times or more compared to before processing. Senoside compound K increased 12.4 times or more, and phenolic and flavonoids increased 3.6 times and 1.4 times, respectively, and omega-6 fatty acids linoleic acid and arachidonic acid were 3.1 times and 3.7 times, respectively. It was enhanced, and Gaba was more than 3.1-fold enhanced (Table 2, Figure 2, Figure 3a, Figure 3b, Table 3).

Another object of the present invention is prepared by the above production method, ginsenoside Rd2, Rg3, F2 and compound K, omega-6 fatty acids, gaba, phenolics, and flavonoids enhanced fermentation of wild ginseng aging complex fermentation to provide.

According to the present invention, the ginseng aged fermented fermented ginsenoside Rd2 is 2.75 mg / g or more, ginsenoside Rg3 is 2.90 mg / g or more, ginsenoside F2 is 3.61 mg / g or more, and ginsenoside compound K is 6.20. mg / g or more, phenolics 8.48 mg / g or more, flavonoids 2.22 mg / g or more, and omega-6 fatty acids linoleic acid and arachidonic acid each contain 282.6 mg / 100g or more and 6.6 mg / 100g or more, Gaba contains more than 100.33 mg / 100g (Table 2, Figure 2, Figure 3a, Figure 3b, Table 3).

The wild fermented ginseng fermented fermentation product according to the present invention has remarkably enhanced antioxidant activity (Test Example 4).

According to another object of the present invention, the present invention is prepared by the above preparation method, ginsenoside Rd2, Rg3, F2 and compound K, omega-6 fatty acids, gaba, phenolics, and flavonoids, goat ginseng enhanced content Provided is a functional food having antioxidant activity that contains an aged complex fermented substance as an active ingredient.

In the present invention, the functional food is dried or fermented wild-fermented ginseng fermented fermentation product of the present invention is used as it is or extracted, used as a food additive, powder or extract liquid in the form of granules, pills, drinks, jelly, etc. Can be manufactured as a product.

The food according to the present invention is useful for improving immunity, improving cognitive ability, restoring fatigue and eating cosmetics (inner beauty).

According to another object of the present invention, the present invention is prepared by the above preparation method, ginsenoside Rd2, Rg3, F2 and compound K, omega-6 fatty acids, gaba, phenolics, and flavonoids are enhanced goat fermentation It provides a cosmetic having antioxidant activity containing a complex fermented substance as an active ingredient.

The cosmetic product according to the present invention is useful for whitening and wrinkle improvement.

Goat ginseng ripening complex presentation according to the present invention is dried or powdered as it is or extracted and used as an additive in cosmetics for whitening, wrinkle improvement or moisturizing, or manufactured in the form of products such as mask packs, skins, creams, and lotions Can be.

The production method of the present invention, compared to the conventional method, produces ginsenosides Rd2, Rg3, F2 and compound K, omega-6 fatty acids, gaba, phenolics, and flavonoids, and goat's ginseng processed products with significantly improved antioxidant activity Make it.

According to the present invention, the fermented ginseng fermentation complex fermented ginsenosides Rd2, Rg3, F2 and compound K, omega-6 fatty acids, gaba, phenolics, and flavonoids are remarkably enhanced, and also have excellent antioxidant effects. , It can be used as a material for cosmetics and pharmaceuticals.

The functional food according to the present invention has excellent antioxidant activity, and is useful for improving immunity reduction due to oxidative stress, improving cognitive ability, fatigue recovery, eating cosmetics, and skin care.

1 is an example of a manufacturing process of a fermented composite fermented ginseng according to the present invention.
Figure 2 shows the ginsenoside compound HPLC chromatogram of the complex fermentation of wild ginseng according to the present invention. Figure 2a is a HPLC chromatogram of 21 kinds of ginsenoside standards, Figure 2b is a ginsenoside compound HPLC chromatogram of Comparative Example 1, Figure 2c is a ginsenoside compound HPLC chromatogram of Comparative Example 2, Figure 2d Is the ginsenoside compound HPLC chromatogram of Example 1, and 2e is the ginsenoside compound HPLC chromatogram of Example 2.
Figure 3 is a graph showing the phenolics and flavonoids content of the fermentation complex of wild ginseng according to the present invention. Figure 3a shows the total phenolics content, Figure 3b shows the total flavonoids content.
Figure 4 is a graph showing the antioxidant activity of the fermented ginseng fermentation according to the present invention. FIG. 4A shows DPPH radical scavenging activity, FIG. 4B shows ABTS radical scavenging activity, FIG. 4C shows hydroxyl radical scavenging activity, and FIG. 4D shows FRAP reducing power.

The invention is explained in more detail by the following examples. These examples are intended to illustrate the invention, and the scope of the invention should not be limited by them.

Preparation Example 1. Preparation of complex fermented fermented ginseng

After washing three times in running water of 10 kg of wild ginseng (outpost) for more than 3 years, and completely draining it, 5 kg of it was put in a steamer container and steamed at 100 ° C for 60 minutes. The primary steamed wild ginseng was placed in a sealed maturing machine for 2 kg each, and the water was evaporated, and the primary sealing maturation was performed at 75 ° C for 3 days. After adding moisture (moisture content of about 40%) to the first aged wild ginseng, put it in a steamer, and then boil it for 2 minutes at 100 ℃ for 60 minutes. After the second boiled goat was transferred to an open aging machine, the second open aging was performed at 65 ° C for 3 days. After adding water (moisture content of about 40%) to the 2nd aged wild ginseng and putting it in a steamer container, it boiled for 3rd time at 100 ℃ for 60 minutes, and put 3rd steamed wild ginseng back into a closed maturing period for 3 days at 75 ℃ Sealing aging was performed. Finally, after adding water (moisture content of about 40%) to the 3rd-aged aged ginseng, placing it in a steamer, and then maturing for 4 minutes at 100 ℃ for 60 minutes, and transferring the 4th-aged steamed goat to an open aging machine, and then 3 at 65 ℃. Fourth open aging was performed daily to prepare steamed and aged goats (FIG. 1).

500 g of cooked steamed and aged wild ginseng is placed in a fermentation container, double-washed water (1 L) is added and hydrated for 1 hour to adjust the water content to about 50%, and sterilized at 120 ° C. for 30 minutes, Lactobacillus plantarium P1201 strain and Lactobacillus brevis BMK484 strain were each cultured as strains in malt extract liquid medium and inoculated with 5% (v / w) of mixed strains in a 1: 1 ratio and fermented at 30 ° C for 7 days Was prepared to ferment the wild-fermented ginseng (Example 1).

On the other hand, fermented fermented composition was prepared by fermenting, fermenting, and fermenting wild ginseng ginseng (goat sprouts) in the same manner as above, 10 kg of wild ginseng ginseng (outpost) grown over 2 years for over 30 years. ).

For comparison, the raw material goat ginseng was prepared as comparative example 1, and as a comparative example 2, the process of drying 5 kg of wild ginseng at 100 ° C for 1 hour, putting it in a dry tray (open container), and drying it at 65 ° C for 3 days was repeated 4 times. After fermentation for 12 days, a fermented composition of wild ginseng fermented in the same manner as above was prepared.

<Physicochemical properties>

Physicochemical properties were measured for Example 1, Example 2, and Comparative Examples 1 and 2. The pH was measured using a pH meter, and the total acidity was measured by neutralization titration and expressed in terms of lactic acid, and the reducing sugar was expressed through conversion by glucose through DNS method (Mille, 1953) and shown in Table 1. .

Analysis items Comparative Example 1 Comparative Example 2 Example 1 Example 2 pH 5.05 4.21 4.31 4.26 Acidity (%, based on lactic acid) 1.81 3.02 2.91 3.12 Reducing sugar (mg / g) 5.02 6.24 5.85 5.55 * All experiments are performed with 3 repetitions and are expressed as average values.

The fermented composite fermented ginseng (Example 1 and Example 2) prepared according to the present invention has a lower pH and a higher total acidity than Comparative Example 1 (raw ginseng), and thus it is judged that the preservation and palatability are improved. There was no significant difference in reducing sugar (Table 1).

Test example  1. Ginsenoside content analysis

The fermented composition of the wild-fermented ginseng of Example 1 prepared above, the fermented wild-fermented ginseng of Example 2, the fermented wild-fermented raw material of Comparative Example 1 and the wild-fermented ginseng of Comparative Example 2 are each dried for 2 days at 50 to 55 ° C. After pulverization, it was pulverized to 100 mesh or less with a grinder and powdered.

<Analysis Sample Preparation>

The powder prepared above was extracted with trisenoside according to the health functional food analysis method. Specifically, 1 g of each dry powder was precisely taken into a 250 ml Erlenmeyer flask, 20 ml of 70% methanol was added thereto, and the mixture was allowed to stand in an 80 ° C. constant temperature bath for 1 hour and cooled. This was centrifuged to take only the supernatant and it was repeated twice. This supernatant was concentrated under reduced pressure at 60 ° C., and the residue was dissolved in 2 ml of tertiary distilled water, filtered through a 0.45 μm membrane filter, and used as an analysis sample.

<Analysis of ginsenoside compounds>

Ginsenoside compound analysis was analyzed by high-pressure liquid chromatography (HPLC) by modifying the method described in Functional Food Analysis. As an analytical column, TSKgel ODS-100Z was used to make a sample injection volume of 10 µl, a temperature of 30 ° C, a measurement wavelength of 203 nm, and a flow rate of 1.0 ml / min. As the mobile phase, A solution was used for HPLC water and B solution for acetonitrile. . As for HPLC analysis conditions, the mobile phase solution flowed to A solution 81%: B solution 19% at 0 minutes, A solution 80%: B solution 20% at 15 minutes, A solution 77%: B solution 23% at 40 minutes, A solution at 42 minutes 70%: B solution 30%, 75 minutes A solution 65%: B solution 35%, 80 minutes A solution 30%: B solution 70%, 90 minutes A solution 10%: B solution 9 0 The mobile phase was shed in%. The analyzed ginsenoside contents of Comparative Example 1, Comparative Example 2, Example 1 and Example 2 are shown in Table 2 and FIG. 2.

Content (mg / g d.w.) Comparative Example 1 Comparative Example 2 Example 1 Example 2 Ginsenoside Rd2 1.81 2.77 2.75 4.63 Ginsenoside F2 0.31 2.27 3.61 4.09 Ginsenoside Rg3 0.36 1.93 3.03 2.90 Ginsenoside C.K 0.50 4.13 6.20 6.30 * All experiments are performed with 3 repetitions and are expressed as average values.

 As shown in Table 2 and FIG. 2, in the case of the fermented composite fermented ginseng of Example 1, the ginsenoside Rd2 is about 1.5 times (2.75 mg / g) or more, compared to Comparative Example 1 (raw goats, FIG. 2b), Ginsenoside Rg3 is about 8.4 times (3.03 mg / g) or more, ginsenoside F2 is about 11.6 times (3.61 mg / g) or more, and ginsenoside compound K is about 12.4 times (6.20 mg / g) or more Enhanced (FIG. 2D).

In the case of the fermented gourd sprouts of Example 2, ginsenoside Rd2 was enhanced by about 2.6 times (4.63 mg / g) or more, and ginsenoside Rg3 was improved by about 8.1 times (2.90 mg / g) or more, particularly ginsenoside Side F2 was about 13.2 times (4.09 mg / g) or more and ginsenoside compound K was about 12.6 times (6.30 mg / g) or more (Fig. 2e).

In addition, ginsenoside Rg3, ginsenoside F2, and compound K were much more enhanced in Examples 1 and 2 than the wild-fermented ginseng or fermented ginseng sprouts fermentation composition of Comparative Example 2.

Test Example 2. Phenolic and flavonoids content analysis

The phenolics and flavonoids, the bioactive components, were analyzed.

<Analysis Sample Preparation>

10 g of 50% fermented alcohol was added to 10 g of each powder prepared as in Test Example 1, extracted at room temperature (20 ± 5 ° C.) for 24 hours, and centrifuged at 3,000 rpm to take only the supernatant to prepare an extract. Using this extract as a sample, total phenolics and total flavonoids were analyzed.

<Total phenolics content>

The total phenolics content was measured by the Folin Denis method (1912).

Specifically, 0.5 ml of each analysis sample prepared above was dispensed into a test tube, and 0.5 ml of a 25% Na ₂ CO ₃ solution was added thereto, and allowed to stand for 3 minutes. Again, 0.25 ml of 2N-Folin-Ciocalteu phenol reagent was added and mixed, followed by standing at 30 ° C for 1 hour, and absorbance was measured at 750 nm. At this time, the total phenolics content was calculated from the standard curve prepared using gallic acid and calculated as an amount equivalent to gallic acid, and the results are shown in FIG. 3A.

As shown in Figure 3a, the total phenolics content of Comparative Example 1 (goat ginseng raw material) and Comparative Example 2 (open-aged fermented wild ginseng fermentation composition), the wild-fermented ginseng fermentation compound according to the present invention (Example 1), respectively About 3.6-fold and 1.5-fold improvement, and the fermented gourd sprout fermentation compound (Example 2) was improved about 3.7-fold and 1.4-fold, respectively.

<Total flavonoids content>

Total flavonoids content was determined by Davis' method.

Specifically, 0.01 ml of 1 N NaOH was added to 0.1 ml of the sample prepared above, and 1.0 ml of diethylene glycol was added thereto, and after standing at 37 ° C. for 1 hour, absorbance at 420 nm was measured. At this time, the total flavonoids content was obtained from the standard curve prepared by preparing the final concentrations of rutin to 0, 0.25, 0.5, and 1.0 mg / ml, and the results are shown in FIG. 3B.

As shown in Figure 3b, compared to the total flavonoids content, Comparative Example 1 (goat ginseng raw material) and Comparative Example 2 (open fermented wild ginseng fermentation composition), the fermented ginseng ginseng fermentation according to the present invention (Example) respectively About 13.1 fold and 2.5 fold were improved, and the goat fermented mature fermentation compound (Example 2) was about 13.2 fold and 2.6 fold enhanced, respectively.

Test Example 3. Analysis of omega 6 fatty acids (linoleic acid and arachidonic acid) and gaba content

Omega 6 fatty acid content analysis was analyzed by gas chromatography (gas chromatograph, GC), Gaba content analysis was analyzed by an automatic amino acid analyzer.

<Omega 6 fatty acid content analysis>

It was performed in accordance with the fatty acid analysis method of the food engineering method.

Specifically, 1 g of each powder prepared as in Test Example 1 was taken, and 3 ml of methanolic 0.5 N NaOH was added and heat treated at 100 ° C. for 10 minutes. Then, 2 ml of BF ₃ was added and stirred, followed by fatty acid methylation for 30 minutes. After the reaction was completed, 1 ml of isooctane was added and mixed, and only isooctane was recovered, dehydrated with anhydrous sodium sulfite anhydrous, and filtered with a 0.45 μm membrane filter (Dismic-25CS) to be used as an analysis sample.

As the analysis, GC was used. After the SP-2560 capillary column (100 m × 0.25 mm, 0.20 μm, Sigma-Aldrich Co.) was mounted, nitrogen gas was used as the mobile phase. 20 μl of the analysis sample was injected to maintain a mobile phase rate of 1 ml / min. At this time, the temperature of the oven was raised to 200 ° C, and then maintained at a final temperature of 230 ° C for 30 minutes to detect it using a fluorescence detector and the contents thereof are shown in Table 3.

<Gaba content analysis>

After adding 4 ml of distilled water to 1 g of each ginseng powder prepared as in Test Example 1, and hydrolyzing it at 60 ° C for 1 hour, 1 ml of 10% 5-sulfosalicylic acid was added and left at 4 ° C for 2 hours. The resulting supernatant was filtered through a 0.45 μm membrane filter and concentrated under reduced pressure at 60 ° C. After concentration, 2 mL of a pH 2.2 lithium buffer was added to the dried product, dissolved, and filtered through a 0.45 μm membrane filter, analyzed by an amino acid automatic analyzer, and the contents are shown in Table 3.

Content (mg / 100 g d.w.) Comparative Example 1 Comparative Example 2 Example 1 Example 2 Linoleic acid (C18: 2c) 261.6 300.5 329.1 282.6 Arachidonic acid (C20: 4n6) 1.8 4.3 7.4 6.6 GABA 75.12 83.17 100.61 100.33 * All experiments are performed with 3 repetitions and are expressed as average values.

 As shown in Table 3, compared to Comparative Example 1 (goat ginseng raw material), the wild-fermented ginseng fermentation complex according to the present invention (Example 1) had linoleic acid and gaba each about 3.1 times (329.1 mg / 100 g: linoleic acid and 100.61). mg / 100 g: gaba), in particular, arachidonic acid was significantly enhanced to about 4.1 times (7.4 mg / 100 g), and the goat fermented mature fermentation complex (Example 2) had linoleic acid content (282.6 mg / 100 g) ) Were improved, and gaba and arachidonic acid were about 3.1 and 3.7 times higher, respectively.

In addition, compared to Comparative Example 2 (open fermented wild goose ginseng fermentation composition), arachidonic acid and gaba were much improved in the wild fermented ginseng fermented fermented products (Examples 1 and 2) according to the present invention.

Test Example 4. Antioxidant activity analysis

Antioxidant activity was analyzed by measuring DPPH radical scavenging activity, ABTS radical scavenging activity, hydroxyl radical scavenging activity and FRAP reducing power.

<Sample preparation>

10 g of 50% fermented alcohol was added to 10 g of each powder prepared as in Test Example 1, extracted at room temperature (20 ± 5 ° C.) for 24 hours, centrifuged at 3,000 rpm, and only supernatant was taken to prepare an extract. , Each extract was concentrated under reduced pressure and freeze-dried at 60 ° C, and analyzed for antioxidant activity on samples prepared at concentrations of 0.25, 0.5, and 1.0 mg / ml.

<DPPH radical scavenging activity>

DPPH radical scavenging activity was added to 0.8 ml of DPPH solution (1.5 × 10 ^-4 M) to 0.2 ml of the sample (0.25, 0.5, 1 mg / ml concentration) prepared above, and allowed to stand for 30 minutes after uniformly mixing for 525 nm. Absorbance was measured at. The negative control of DPPH radical scavenging activity was performed in the same way using distilled water instead of the sample, and the difference in absorbance was calculated as a percentage (%) by the following equation, and the results are shown in FIG. 4A.

Radical scavenging activity (%) = [1- (absorbance of negative control ÷ absorbance of experimental sphere)] × 100

<ABTS radical scavenging activity>

For ABTS radical scavenging activity, 5 ml of 7 mM ABTS reagent and 5 ml of 140 mM K ₂ S ₂ O ₈ (FW 270.3, Sigma 9392) were mixed and left in a dark place for 16 hours to generate cationic radicals, followed by mixing with methanol to 732 nm. The ABTS solution adjusted so that the absorbance value of the control was 0.7 ± 0.02 was used.

0.1 ml of the sample (0.25, 0.5, 1 mg / ml concentration) prepared above and 0.9 ml of ABTS solution were mixed and reacted for 3 minutes, and absorbance was measured at 732 nm. The ABTS radical scavenging activity was also performed in the same way using distilled water instead of the sample, and the difference in absorbance was calculated as a percentage (%) by the above formula, and the results are shown in FIG. 4B.

<Hydroxyl radical scavenging activity>

The hydroxyl (· OH) radical scavenging activity is FeSO _{4 . 2} ml of 7H ₂ 0-EDTA (10 mM) and 0.2 ml of 2-deoxyribose (10 mM), H ₂ O ₂ (10 mM) 0.2 ml, 1.4 ml of each composition of the preparation example was mixed and reacted at 37 ° C. for 4 hours to 1% thiobarbituric acid (TBA) and 2.8% trichloroaceric acid (TCA) in this mixture. ) Was added to 1 ml each, color was developed at 100 ° C for 20 minutes, cooled, and absorbance was measured at 520 nm. The hydroxyl radical scavenging activity negative control was performed in the same way using distilled water instead of the sample to calculate the difference in absorbance as a percentage (%) by the above formula, and the results are shown in FIG. 4C.

<FRAP reducing power analysis>

FRAP (Ferric reducing antioxidant power) is a method of measuring the reducing power of a compound, and is a method of measuring the power of reducing Fe ^{3 +} to Fe ^{2 +} . Specifically, when the FeⅢ-TPTZ (ferric tripyridyl triazine) is reduced to the blue FerII tripyridyl triazine (FeII-TPTZ) by the reducing power of the sample, the absorbance is measured to determine the antioxidant activity.

As a reaction solution in the FRAP reducing power analysis, 30 mM acetate buffer (pH 3.6), 10 mM 2,4,6-tripyridyl-s-triazine (TPTZ, T1253, C18H12N6, MW312.33) dissolved in 40 mM hydrochloric acid and 20 mM FeCl ₃ (F7134, MW 162.20, in DW) was prepared, and the acetate buffer, TPTZ solution and FeCl ₃ solution were mixed at 10: 1: 1 (v / v / v) to pre-react for 15 minutes at 37 ° C. Ordered.

50 µl of each assay sample (0.25, 0.5, 1 mg / ml concentration) prepared above and 950 µl of FRAP reagent were dispensed into a test tube, reacted for about 15 minutes, and 593 using a microplate reader (Biorad 3055, Sweden). Absorbance was measured at nm and the results are shown in FIG. 4D.

As shown in Figures 4a ~ 4d, compared to Comparative Example 1 (goat ginseng raw material) and Comparative Example 2 (open fermented wild ginseng fermentation composition), fermented ginseng ginseng according to the present invention (Example 1) and goat sprout The three fermented fermentation complexes (Example 2) had significantly improved DPPH radical scavenging activity, ABTS radical scavenging activity, hydroxyl radical scavenging activity, and FRAP reducing power, especially when treated with 1 mg / ml of sample 1 and 2 DPPH radical scavenging activity was 90.66% and 89.92% respectively, ABTS radical scavenging activity was 89.77% and 94.37%, respectively, hydroxyl radical scavenging activity was 64.57% and 67.72%, respectively, and FRAP reducing power was 1.752 and 1.859, respectively.

From the above results, it can be seen that the wild-fermented ginseng or wild-fermented ginseng-fermented fermentation product according to the present invention has significantly improved antioxidant activity and is useful as a material for functional food or cosmetics.

Agricultural Biotechnology Research Institute KACC92157P 20161212 Agricultural Biotechnology Research Institute KACC91848P 20130719

Claims (8)

Ginsenoside Rd2, Rg3, F2 and compound K, omega-6 fatty acids, gaba, phenolics, and flavonoids are improved fermentation of wild ginseng ripening complex fermented substances,
The above method
Iv) alternately performing 4 times of steaming and sealing aging and steaming and open aging of goats or ginseng sprouts; And
Ii) fermentation with a complex seed of the Lactobacillus Brevis BMK484 strain deposited with accession number KACC 92157P and the Lactobacillus plantarium P1201 strain deposited with accession number KACC91848P,
The steaming is performed at 100 ℃ for 30 to 120 minutes,
The sealed aging is performed for 2-3 days in a sealed aging machine at 70 to 80 ° C,
The manufacturing method of the aging is to perform aging for 2-3 days in an open aging machine at 60 to 70 ° C.
A wild-fermented fermented ginseng fermented by the method according to claim 1, wherein the ginsenosides Rd2, Rg3, F2 and compound K, omega-6 fatty acids, gaba, phenolics, and flavonoids are enhanced.
According to claim 2, wherein the ginseng aged fermentation complex is ginsenoside Rd2 2.75 mg / g or more, ginsenoside Rg3 is 2.90 mg / g or more, ginsenoside F2 is 3.61 mg / g or more, ginsenoside compound K contains 6.20 mg / g or more, phenolics 8.48 mg / g or more, flavonoids 2.22 mg / g or more, and omega-6 fatty acids linoleic acid and arachidonic acid respectively 282.6 mg / 100g or more and 6.6 mg / 100 g Ginsenosides Rd2, Rg3, F2 and Compound K, omega-6 fatty acids, gaba, phenolics, and flavonoids are enhanced. water.
[3] The ginsenoside Rd2, Rg3, F2 and compound K, omega-6 fatty acids, gaba, phenolics, and flavonoids according to claim 2, wherein the wild-fermented ginseng-fermented complex fermentation product has enhanced antioxidant activity. Enhanced fermented ginseng ginseng complex.
Food comprising the fermented ginseng fermentation according to any one of claims 2 to 4.
Cosmetic comprising the fermented ginseng fermented ginseng according to any one of claims 2 to 4.
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