KR20170137566A - THE MANUFACTURING METHOD OF KOREAN PANAX GINSENG FERMENTATION SOLUTION HAVING INCREASING OF Rg3, Rh1 AND COMPOUND K CONTENT AND ANTIOXIDANT PROPERTIES BY ISOLATED LACTIC ACID BACTERIA - Google Patents

Abstract

The present invention relates to a method of preparing a lactic acid fermentation product of wood-cultivated ginseng having antioxidant effects and enhanced ginsenosides. The method comprises a step of inoculating a sterilized wood-cultivated ginseng composition solution with Leuconostoc mesenteroides DU-B1604(KCTC 13015BP). According to the method of the present invention, a lactic acid fermentation product of wood-cultivated ginseng having an increased content of ginsenosides, Rg3, Rh1 and Compound K can be produced.

Description

BACKGROUND OF THE INVENTION 1. Field of the Invention The present invention relates to a fermented soybean ginseng fermented by fermentation of isolate lactic acid bacteria, BACTERIA}

The present invention relates to a method for preparing a fermented product of Lactobacillus acidus having antioxidant activity and a specific increase in the content of ginsenosides Rg3, Rh1 and Compound K among ginsenosides, and more specifically, ( Leuconostoc mesenteroises ) DU-B1604 (KCTC 13015BP), which had been separated into strains effective for the present technology, were inoculated and fermented in an appropriate condition. And a method for producing fermented product of lactic acid fermented soybean ginseng with specific antioxidative effect and specific ginsenoside enhancement so that the content of Rg3, Rh1 and Compound K can be utilized as functional food having enhanced contents.

In general, Korean panax ginseng is produced by natural ginseng seeds and seedlings grown by natural germination in the wild, artificially sprayed in shady and humid places such as birch trees and lacquer trees deep in the mountains as closely as possible to the growth environment of wild ginseng. After a few years or decades have passed, the exact definition of goose ginseng is unclear. In other words, goat ginseng is also known as ginseng, elder, and camphor ginseng, and its lifespan is different according to the thickness and length of roots, the presence of ginseng in the trunk, the number of ginseng according to age, and soil and climatic conditions. According to the results of analysis of ginseng and ginseng root ginseng, ginsenosides Rh1 and Rg3 were not found in ginseng but only a small amount was detected in ginseng ginseng. Compared to all ginsenosides, ginseng ginseng content was higher than ginseng There is a report. As a result, it is believed that goat ginseng has high value added in terms of efficacy and marketability compared to saturated ginseng market in market competitiveness, and it is considered to have substitute material and commerciality. When fermenting this goat ginseng, Rg1, Rb1 and Rg3 are greatly increased, Of the patients. Although the price is more than 10 times higher than that of ginseng, it is expected to be competitive in the existing ginseng market by enhancing pharmacological efficacy through fermentation process. Compared to native ginseng and natural ginseng, appearance is similar to that of natural ginseng, but prices are only 1/5 to 1/10. However, pharmacological efficacy is considered to be similar to natural ginseng, and it is known to be effective against anticancer, hypotensive, antioxidant, hepatotoxicity and lipid strengthening.

The saponin contained in ginseng is called ginsenoside. It is derived from sapona (soap) and is bubbled in aqueous solution. It is used as a cardiovascular and diuretic in the herbal medicine. It is contained in the roots, stem, leaves and bark of plants. It has anti-cancer and antioxidant effects and has a different pharmacological effect from other plants. It has a unique chemical structure and is widely used as a cosmetic and functional food have. In ginseng, ginsenoside, polyacetylene, acid polysaccharide, ginseng protein and phenolic substance were identified, and the chemical structure of ginsenoside was clearly confirmed. In addition, it has antidiabetic activity as well as anticancer activity, antioxidant activity, prevention of arteriosclerosis and hypertension, promotion of hepatic function and elimination of hangover, anti-fatigue and anti-stress action, anti-aging action, brain activity acceleration action, anti- Treatment, promotion of protein synthesis, etc. have been reported. In particular, red ginseng dried with fresh ginseng is known as a ginsenoside Rg2, Rg3, Rh1, and Rh2, which is a specific component of red ginseng produced by heat, has functions of preventing cancer, inhibiting cancer cell growth, hypotensive action, Since it has antioxidant activity, it is attracting attention as a special feature of red ginseng.

Lactic acid bacteria is one of the most beneficial microorganisms that can be used by humans. It is a non-pathogenic bacterium that can function in the intestines of humans and can function as a bacterium and bacterium. It ferments sugars to produce a large amount of lactic acid. And it requires a variety of nutrients. When vegetables are fermented by lactic acid bacteria, their flavor and taste are enhanced, and their sensory characteristics are improved. Vitamin C (vitamin C) and various physiologically active substances are well preserved by organic acids. Vitamin B12 and vitamins It is known that K is biosynthesized and inhibits the growth and proliferation of spoilage bacteria and pathogenic bacteria, and becomes a hygienic food.

Korean Patent No. 10-1133739 discloses a prior art related to the present invention. However, the present invention relates to a method for producing white ginseng extract having a high Rg ₃ content by heat hydrolysis of white ginseng extract and white ginseng extract prepared from the ginsenoside Rg ₃ And Korean Patent No. 10-1401658 discloses an antimicrobial agent comprising a ginsenoside compound K or a derivative thereof. However, there is no known method for producing the fermented product of L. acidophilus with antioxidant activity and increased content of Rg ₃ , Rh ₁ and Compound K until now.

It is an object of the present invention to provide a method for producing a fermented goat ginseng having enhanced Rg ₃ , Rh ₁ and compound K contents and a fermented product thereof.

The present invention is to provide a health functional food prepared according to the above method and containing a fermented goat ginseng having enhanced ginsenoside Rg ₃ , Rh ₁ and compound K content.

The above object of the present invention is achieved by a method for producing fermented lactic acid bacteria using a goat's ginseng extract, which comprises grinding the goat's ginseng and extracting 15 brix of the goat ginseng extracted at a predetermined condition (75 DEG C, 5 hours and 30 minutes) A first step (S1) of concentrating the water in a water bath at 50 DEG C to 20-50 bricks; A second step (S2) of adjusting the goat ginseng concentrate obtained in the step (S1) to 1 to 50 volume% of the total volume and adding the same; A third step (S3) of adding 0.1 to 30% by volume of galactooligosaccharide to the total volume obtained in the above step and adjusting the pH to 5.0 to 7.0 and then sterilizing the mixture; In the above-mentioned step, the ginseng root-treated ginseng was added to a commercially available MRS medium, which had been pre-cultured, and the separated lactic acid bacteria, Leuconostoc mesenteroides ) DU-B1604 (KCTC 13015BP) in 0.1 to 20% by volume of the composition of the goat's ginseng; The composition liquid is lactic acid obtained in Step The inoculated fermentation temperature 20 ~ 45 ℃, fermentation time of 6 to comprise a fifth step (S5) to the fermentation for 120 hours antioxidant and Rg _3, Rh ₁ and Compound K and specific jinse of ≪ / RTI >

In the present invention, the fermented goat germ obtained in the fifth step (S5) is sterilized at a sterilization temperature of 60 to 100 DEG C for 10 to 60 minutes and then filtered through a microfilter filter (S6) As shown in FIG.

The liquid goat's ginseng extract in the first step is prepared by using at least one selected from the group consisting of a goat's grind liquid, a goat ginseng extract, a goat ginseng concentrate, a dried goat ginseng product and a goat ginseng granule.

 The galactooligosaccharide in the third step is further characterized by being prepared using at least one selected from maltooligosaccharide, fructooligosaccharide and isomaltooligosaccharide.

The fermented product obtained in the fifth step or the sterilized and filtered fermented product obtained in the sixth step may be added in an amount of 0.1 to 99.9 wt% or in order to improve the taste of the poultice, apple, persimmon, blueberry, blackberry, black currant, Pomegranate, grape, pear, peach, banana, strawberry, tomato, carrot, plum, plum, cherry, A raw material fruit made of mango, lemon, pineapple, a juice or a concentrated liquid is further added in an amount of 0.1 to 99.9 parts by weight or more.

The lactic acid fermented product of the goat ginseng prepared according to the present invention has antioxidant activity and the content of ginsenosides Rg ₃ , Rh ₁ and Compound K in addition to the common ginsenosides of ginseng and red ginseng, There is an excellent effect.

FIG. 1 is a diagram showing a stepwise process for preparing a fermented product of Lactobacillus acidus according to the present invention, in which the content of Rg ₃ , Rh ₁ and Compound K, which are antioxidant and specific ginsenosides, is enhanced.
Fig. 2 is a photograph of esculin reaction in an Escherichia culture medium of primary isolated lactic acid bacteria (strain No. 25 to 29) used as a publicly-known material in the present invention.
Figure 3 is a close relational diagram of a phylogenetic tree based on the 16S rRNA gene sequence of the standard strain according to the invention and the final isolate L1 (red).
FIG. 4 is a graph showing the four-dimensional reaction surface of the pH (pH: 3.40 - 3.55 - 3.70) change of the goat ginseng fermentation product according to the present invention ginseng concentration, galactooligosaccharide concentration and time condition.
FIG. 5 is a graph showing the 4-dimensional response surface of the acidity (0.4 - 0.6 - 0.8%) change of the goat's ginseng fermentation product according to the present invention ginseng concentration, galactooligosaccharide concentration and time condition.
FIG. 6 is a graph showing the 4-dimensional response surface of viable cell content (13.0 - 13.5 - 14.0 Log CFU / mL) of the goat's germ fermented product according to the inventive ginseng concentration, galactooligosaccharide concentration and time condition .
FIG. 7 is a graph showing a 4-dimensional reaction surface for crude saponin content (crude saponin content: 0.1-0.2-0.3%) of the goat's milk fermented product according to the present invention's concentration, galactooligosaccharide concentration and time conditions.
FIG. 8 is a graph showing the 4-dimensional response surface of the total phenolics compound content (70 - 75 - 80 mg%) of the goat fermented product according to the present invention, the concentration of galactooligosaccharide and time conditions .
FIG. 9 is a graph showing the 4-dimensional response surface of the DPPH radical scavenging activity (55 - 60 - 65%) change according to the present invention ginseng concentration, galactooligosaccharide concentration and time conditions.
FIG. 10 is a graph showing the 4-dimensional response surface of the ABTS radical scavenging activity (52 - 55 - 58%) changes according to the present invention ginseng concentration, galactooligosaccharide concentration and time conditions.
FIG. 11 is a graph showing the 4-dimensional response surface of the nitrate scavenging activity (35 - 40 - 45%) of the goat's milk fermentation according to the present invention, the concentration of galactooligosaccharide and time conditions.
FIG. 12 is a graph showing the multiple overlapping of the four-dimensional reaction surface on the optimum conditions of the total phenol compound content, total saponin content and DPPH radical scavenging ability of the goat's ginseng fermented product according to the present invention ginseng concentration, galactooligosaccharide concentration and time condition .
13 is the present invention sanyangsam concentration, galactosyl sanyangsam fermentation sprocket of ginsenoside Rg ₁ content according to the oligosaccharide concentration and time conditions: showing a four-dimensional reaction surface for the _{(ginsenoside Rg 1 content 3.5 - 5.5} mg% - 4.5) Change Graph.
14 is sanyangsam ginsenoside Rb ₁ content of the fermentation sprocket according to the invention sanyangsam concentration, galacto-oligosaccharide concentration and time conditions _{(ginsenoside Rb 1 content: 16 -} 19 - 22 mg%) showing a four-dimensional response surface for a change Graph.
15 is the present invention sanyangsam concentration, galacto-oligosaccharide concentration and time conditions sanyangsam ginsenosides Rh ₁ content of the fermentation Sprocket according to: showing a four-dimensional reaction surface for the _{(ginsenoside Rh 1 content 2.7 - 3.7} mg% - 3.2) Change Graph.
16 is the present invention sanyangsam concentration, galacto-oligosaccharide concentrations, and ginsenosides Rg ₃ content of sanyangsam fermentation sprocket according to the time condition _{(ginsenoside Rg 3 content: 4.0 -} 4.5 - 5.0 mg%) showing a four-dimensional response surface for a change Graph.
17 shows the 4-dimensional response surface of the ginsenoside compound K content (ginsenoside compound K content: 3 - 5 - 7 mg%) of the goat fermented product according to the present invention ginseng concentration, galactooligosaccharide concentration and time condition Graph.
FIG. 18 is a graph showing the results of multiple overlapping of four-dimensional reaction surfaces for optimal conditions for ginsenoside Rh ₁ , ginsenoside Rg ₃ and compound K maximum content of goat juice fermentation according to the present invention ginseng concentration, galactooligosaccharide concentration, Fig.
19 is a ginsenoside chromatogram of the pre-fermentation culture according to the present invention.
20 is a ginsenoside chromatogram of a culture fermented under optimal fermentation conditions for ginsenoside content according to the present invention.

Hereinafter, preferred embodiments of the present invention will be described in detail.

The present invention relates to a method for producing a lactic acid bacteria which comprises adding a galactooligosaccharide to a liquid goat's ginseng prepared by using at least one selected from the group consisting of a goat's milk powder, a goat extract, a goat ginseng concentrate, a dried goat ginseng product, Leuconostoc mesenteroides ) DU-B1604 (KCTC 13015BP), fermented under optimal fermentation conditions to obtain antioxidant activity, and the contents of specific ginsenosides Rg ₃ , Rh ₁ and Compound K were enhanced. .

According to the present invention, the goat's ginseng extract (15 brix) extracted from a predetermined condition (70% fermented alcoholic beverage, 75 ° C, 5 hours and 30 minutes) was pulverized and the alcohol in the extract was removed at 50 ° C in a water bath using a rotary concentrator A first step (S1) of concentrating to 20-50 Bricks; A second step (S2) of adjusting the goat's ghe concentrate obtained in the above step to 1 to 50% by volume of the total volume; A third step of adding 0.1 to 30% by volume of galactooligosaccharide to the total volume obtained in the above step and adjusting the pH to 5.0 to 7.0 and then sterilizing the mixture; In the soymilk composition obtained by the sterilization treatment in the above step, the separated lactic acid bacteria pre-cultured on a commercial MRS medium, Leuconostoc mesenteroides ) DU-B1604 (KCTC 13015BP) in 0.1 to 20% by volume of the composition of the goat's ginseng; Wherein the fermentation is carried out at a fermentation temperature of 20 to 45 ° C. and a fermentation time of 6 to 120 hours. The antioxidative activity of the fermentation broth is determined by measuring the content of Rg ₃ , Rh ₁ and Compound K To provide a health functional food supplemented with a fermented product of Lactobacillus of the goat's ginseng.

The method of the present invention is further characterized by further comprising a sixth step of subjecting the fermented goat having been subjected to the fifth step to a heat sterilization treatment at a sterilization temperature of 60 to 100 DEG C for 10 to 60 minutes, and then filtering through a 1.0 microfilter filter.

The present invention is characterized in that the liquid goat's ginseng in the first step is manufactured by any one or more selected from the group consisting of a raw ginseng pulverization solution, a goat ginseng extract solution, a goat ginseng concentrate, a dried goat ginseng product and a goat ginseng granule.

According to the present invention, the galactooligosaccharide in the step 3 is prepared by using at least one selected from maltooligosaccharide, fructooligosaccharide and isomaltooligosaccharide. do.

According to the present invention, the fermented product after the fifth step or the fermented product sterilized and filtered through the sixth step may be used in an amount of 0.1 to 99.9 wt%, or in order to improve the taste of the poultry, an apple, persimmon, blueberry, blackberry, Pepper, Pomegranate, Pomegranate, Grape, Pear, Peach, Banana, Strawberry, Tomato, Carrot, Plum, Plum, Cherry, Duck, Odyssey, Mulberry, Kiwi, Melon, Tangerine, Orange, Grapefruit, Tangerine, Melon, Cherry, Melon , Black superbary, mango, lemon, pineapple, fruit juice, or concentrate is added in an amount of from 0.1 to 99.9 parts by weight or by weight.

Hereinafter, the present invention will be described in detail with reference to Examples and Experimental Examples, but the scope of the present invention is not limited thereto.

Example 1.

1) Isolation of lactic acid bacteria

The fermented food samples used for the isolation of lactic acid bacteria were collected from dozens of kimchi dipped in households and restaurants and used as sample liquids. Each sample liquid was diluted to 10-1 to 10-5 with sterilized distilled water, and 0.004% BCP 100 μl aliquots were added to the commercially available MRS agar medium and cultured for 24 hours at 37 ° C. to isolate characteristic colonies of lactic acid bacteria from isolated colonies. The purely isolated lactic acid bacteria were inoculated on MRS agar slant medium, cultured at 37 ° C for 24 hours, and stored at 4 ° C in a refrigerator.

2) Selection of β-Glucosidase active strains

For the selection of strains having beta-glucosidase activity, Escherin agar medium (esculin 0.1%, peptones 1.8%, ferric citrate 0.1%, agar 2% The strain was inoculated and the change in color in the medium was observed (Fig. 2). Esculin is separated into glucose and esculin by beta glucoside, and esculin reacts with ferric ammonium citrate to form a black complex around the colon. Therefore, strains which form black spots around colonies cultured on Escherichia agar medium were selected as strains having beta glucoside activity.

… [구조식1]

... [Structural formula 1]

3) Measurement of β-Glucosidase activity

The activity of β-glucosidase activity was measured by inoculating a commercially available MRS medium containing 1% carboxymethyl cellulose (CMC) for 12 hours before inoculation, culturing the cells at 37 ° C. for 24 hours, . The cells were removed by centrifugation for 15 minutes. 0.5 ml of the supernatant was mixed with 1 ml of 5 mM ρ-nitrophenyl-β-D-glucopynanoside (ρ-NPG) solution and reacted at 37 ° C for 30 minutes. The reaction was terminated by the addition of 1 ml of 1 M Na2CO3 solution, and the resulting p-nitrophenol was measured for absorbance at 405 nm and the concentration was calculated using a calibration curve of ρ-NP.

4) Sequence analysis and phylogeny of the final selection strains

The 16S rRNA gene of the finally selected strain was analyzed by sequencing and the similarity (%) between the isolated strain and the type strain on the database was confirmed using the NCBI database.

Example 2.

1) Fermentation conditions using goat extract

(1) Acquisition of goat ginseng extract

The goat ginseng was washed with flowing water, and the foreign material was removed, and the mixture was uniformly ground with a grinder to be used for extraction. Extracts of 15 mg of goat ginseng were obtained from the extracts of goat ginseng extracted at 75 ℃ for 5 hours and 30 minutes. This extract was concentrated to 20-50 Bricks in a constant temperature water bath at 50 ° C using a rotary condenser to obtain a concentrate. The obtained goat ginseng concentrate was used for fermentation by controlling its concentration.

(2) Effect test of oligosaccharide addition

The effects of additives were investigated for efficient lactic acid fermentation. 1% by volume of oligosaccharide was added as a supplement to 5% by volume of goat's ghrelin concentrate (25 brix), sterilized at 121 ° C for 15 minutes, sterilized for 15 minutes and then inoculated with 2% by volume of isolated lactic acid bacteria pre- The efficacy of lactic acid bacteria was investigated.

(3) Measurement of crude saponin content

The crude saponin content of the extract concentrate and the fermentation broth was determined by n-butanol extraction method. That is, 30 ml of the extract and the fermentation broth were washed three times with 30 ml of diethyl ether to remove fat-soluble components, 30 ml of water-saturated n-butanol was added to the aqueous layer three times and the n-butanol layer was concentrated to obtain crude saponin And dried in a 105 ° C drier until the weight became constant. The crude saponin content is expressed as a percentage of the amount of raw material used.

(4) Measurement of ginsenoside content

The composition and content of ginsenosides of the extract concentrate and the fermentation broth were determined by filtration through a membrane filter (0.45 μm, pore size), HPLC (Waters 2695 Separations Module, Waters Co., Milford, MA, USA) and Symmetry C18 (4.6 × 150 mm, 5 μm) column was used as a solvent and A (water) and B (Acetonitrile) were used as solvents. The flow rate of the mobile phase was 1.0 ml / min, the column temperature was 30 ° C, and the detection wavelength of the PDA detector was 203 nm.

2) Antioxidative properties and fermentation conditions of specific ginsenosides by fermentation of L. acidophilus

(1) fermentation of goat's ginseng extract by isolated lactic acid bacteria

The fermentation of goat's ginseng was carried out by firstly adding 5% by weight of concentrated soybean ginseng (25 ° brix) to distilled water, sterilized by pressurization at 121 ° C for 15 minutes, slowly cooled at room temperature, and then inoculated with five pre- After fermentation for 72 hours, fermentation characteristics were investigated and one strain with the best fermentation activity was selected.

(2) Setting the concentration of goat's ginseng, fermentation time and oligosaccharide concentration

To determine the fermentation condition of goat ginseng, the extract of goat ginseng extracted under optimal conditions was concentrated under reduced pressure and adjusted to 25 bricks and used in the experiment. The response surface methodology was used and the experimental design for the fermentation conditions was carried out using the central synthetic design method to determine the concentrations (X1), fermentation times (X2) and The test range for the concentration (X3) of supplementary oligosaccharide is set, and the encoding is performed in five steps as shown in Table 1. The fermentation experiment was carried out with 16 sections according to the central synthesis plan. In addition, the dependent variable (Yn) affected by these independent variables was measured as the titratable acidity (Y1), lactic acid bacteria number (Y2), crude saponin content (Y3), ginsenoside content (Y4) and antioxidant properties (Y5) The values were used for regression analysis. The effect of fermentation conditions on the content of ginsenoside active ingredient in fermented soybean ginseng fermented product was analyzed by 4 - dimensional reaction surface analysis using Mathematica program based on the predicted model equation.

Independent variables and code table of response surface method for determination of goat juice concentration, fermentation time and galactooligosaccharide concentration Independent variable unit Display Code value -2 -One 0 One 2 Concentration of goat's ginseng (25 Bricks) % X _One 2 4 6 8 10 Fermentation time hr X ₂ 24 36 48 60 72 Galactooligosaccharide concentration % X ₃ 0 0.25 0.5 0.75 1.0

Central Synthesis Experimental Design for Optimization Conditions of Goat Ginseng Fermentation Experiment number Fermentation condition Concentration of goat's ginseng added Fermentation time Galactooligosaccharide concentration (%) (hr) (%) One 8 60 0.75 2 8 60 0.50 3 8 36 0.75 4 8 36 0.25 5 4 60 0.75 6 4 60 0.25 7 4 36 0.25 8 4 36 0.25 9 6 48 0.50 10 6 48 0.50 11 10 48 0.50 12 2 48 0.50 13 6 72 0.50 14 6 24 0.50 15 6 48 1.00 16 6 48 0.00

(3) Measurement of sugar content, pH and titratable acidity of fermented soybean ginseng

During fermentation, the samples were taken at fixed time intervals and the sugar content of fermented goat gums was measured with a refractometer (PR-201α, ATAGO, Japan). The pH of the fermented goat's ghana fermented product was converted to a lactic acid content (%) by using a pH meter (Metter Toledo Group, Switzerland) and titrating the titrated acidity with 0.1 N NaOH solution to pH 8.3.

(4) Measurement of viable count of fermented goat ginseng

The number of viable cells of the fermented goat 's ginseng was determined by counting the number of colonies after incubation at 37 ° C. for 24 hours in a commercial MRS agar medium after the samples were aseptically taken during a certain period of time during fermentation and then diluted appropriately with physiological sterilized saline.

3) Antioxidant properties of Fermented Lactic acid bacteria

(1) Electron donating ability measurement

The electron donating ability of the goat juice fermentation sample was measured by the method using α, α'-diphenyl-β-pycrylhydrazyl (DPPH). Namely, 12 mg of DPPH reagent was dissolved in 100 ml of anhydrous ethanol, and 50% ethanol solution was added to adjust the absorbance of the DPPH solution to about 1.0 at 517 nm. Then, 0.5 ml of the fermentation broth was mixed with 5 ml of DPPH solution Respectively.

(2) Measurement of total phenolic compound content

Polyphenol content of goat juice fermented samples was determined by colorimetry by Folin-Denis method. 1 ml of phenol reagent was added to 1 ml of sample, and the mixture was allowed to stand for 3 minutes. 1 ml of 10% sodium carbonate (Na2CO3) was mixed and left at room temperature for 1 hour to measure the absorbance at 700 nm. The standard curve was prepared with a gallic acid solution and the total phenolic compound content was expressed as a percentage of the amount of raw material used.

(3) 2,2-azino-bis (3-ethylbenzthiazoline-6-sulfonic acid, ABTS)

The ABTS radical scavenging activity was determined by mixing 7.2 mM ABTS solution dissolved in distilled water and 2.6 mM potassium persulfate solution in a ratio of 1: 1, allowing to stand at room temperature for 12-16 hours and then absorbance at 734 nm with methanol Was diluted to 1.1 to 1.2. 0.1 ml of the sample prepared at a concentration of 10.0 mg / ml was mixed with 3.9 ml of the ABTS dilution solution, reacted for exactly 1 minute, and the absorbance was measured at 734 nm.

(4) Nitrite scavenging ability

1 ml of 1 mM sodium nitrite solution was added to 1 ml of fermented goat's milk. 0.1 N HCl and 0.1 M citric acid buffer solution (pH 3.0) were added to make the total volume of the reaction solution 10 ml, followed by reaction at 37 ° C for 1 hour. 1 ml of this reaction solution was taken, 2% acetic acid solution and Griess reagent were added in order, and the absorbance at 520 nm was measured to calculate the nitrite scavenging ability of the sample to which no sample was added.

Experimental Example 1

1. Selection of lactic acid bacteria

1) Isolation of lactic acid bacteria from fermented food

Of the 26 strains isolated from the fermented kimchi, 49 strains were isolated by morphologically differentiating colonies showing the characteristics of lactic acid bacteria on the MRS agar medium. The results of experiments of 49 lactic acid bacteria using the Esculin agar method are shown in Fig. Glucosidase activity was observed in 27 lactic acid bacteria around the colonies and 17 of the 27 strains reacted with Esculin were found to have black spot The size of the diameter was 5 mm or less, and five strains having black spots of about 15 mm or more were isolated through repeated experiments (Table 3).

Diameter of black spots in esculin agar medium of isolated strains Strain number 25 26 27 28 29 Diameter (mm) 16.5 15.3 16.1 15.3 15.7

2) Beta-glucosidase activity of isolated strains

Beta-glucosidase activity was measured in 5 kinds of lactic acid bacteria having a black spot diameter of 15 mm or more in Escherichia reaction, and the activity of beta glucosidease activity was measured by culturing these strains for 48 hours Are shown in Table 4. Beta - glucosidase activity was the highest at 24 hours of incubation time.

Results of measurement of beta-glucosidase activity of isolated strains Isolate strain number Beta-glucosidease (unit / ml) 0h 24h 48h 25 0.030.01 6.320.64 5.541.60 26 0.020.01 5.102.71 4.232.64 27 0.040.02 4.971.15 3.920.40 28 0.050.03 4.910.76 3.700.73 29 0.090.05 4.801.36 3.770.13

3) Selection of lactic acid strains for fermentation of goat ginseng

The goat ginseng concentrate was diluted with 10 brix, and the five lactic acid bacteria isolated were inoculated and cultured at 37 ° C for 48 hours. The results of the physicochemical test for the fermented product of the goat ginseng according to the strain are shown in Table 5. As a result of the test, the lactic acid bacteria No. 25, which showed the highest acid production power and the highest number of lactic acid bacteria in the fermented product of the goat germ, was selected as the final fermented lactic acid bacteria (FIG. 2).

Characteristics of fermented goat's milk (37 ℃, 48 hours culture) for the final selection of isolated strains Isolate strain number pH Sugar content Acidity Number of lactic acid bacteria living bacteria Brix % CFU / ml Non-vaccination 5.900.03 3.50.1 0.050.01 0 25 4.270.02 1.30.0 0.310.05 8.8010 ⁹ 26 4.270.01 1.30.1 0.230.03 6.2010 ⁸ 27 4.280.02 1.20.0 0.240.02 6.9210 ⁹ 28 4.290.03 1.20.1 0.220.04 7.7010 ⁸ 29 4.300.02 1.20.1 0.200.01 5.5010 ⁸

2. Identification of isolated lactic acid bacteria

The partial sequence of the 16S rRNA gene of the 25th strain with the highest activity among the finally selected strains was analyzed and compared with the NCBI blast DB. As a result, it showed 99% similarity with the strain of Leuconostoc mesenteroides such as Leuconostoc mesenteroides strain ATCC 8293 as a comparative strain of the database. The relationship of the phylogenetic tree based on the 16S rRNA gene sequence of the database-related strain and the isolate is shown in FIG. Based on these results, the final isolate was identified as Leuconostoc mesenteroides and finally named as Leuconostoc mesenteroides DU-B1604. The strain was deposited with the BRC and deposited with KCTC 13015BP on July 25, 2016 .

16 rDNA partial nucleotide sequence analysis of the final isolate was compared with the standard strain Major standard strains Difference / Comparison Similarity Leuconostoc mesenteroides strain ATCC 8293 3/1404 99% Leuconostoc mesenteroides subsp. dextranicum strain NBRC 100495 3/1404 99% Leuconostoc mesenteroides strain JCM 6124 3/1404 99% Leuconostoc mesenteroides subsp. suionicum strain LMG 8159 3/1404 99% Leuconostoc mesenteroides subsp. cremoris strain NCFB 543 3/1404 99%

Experimental Example 2. Optimization of Fermentation Conditions of Lactic Acid Bacteria Using Concentrated Goat Yeast Extract

1) Effect of supplementary oligosaccharide addition

To investigate the effect of the addition of oligosaccharide as a raw material on the fermentation of lactic acid bacteria in the goat ginseng, 1 wt% of various oligosaccharides were added to 5 wt% of a concentrated goat gum (25 BRICKS), and then inoculated with the isolated lactic acid bacteria Leuconostoc mesenteroides DU-B1604 And the fermentation was carried out at 37 DEG C for 48 hours, and the viable cell count, pH and acidity were examined. After fermentation, galactooligosaccharide was lowest at 3.66 and isomaltooligosaccharide was the highest at pH 4.70. The highest acidity was obtained with 0.50% of galacto - oligosaccharides and the lowest with 0.34 of isomaltooligosaccharides. The number of lactic acid bacteria was increased to 1010 after fermentation at the initial dose of 107, and the addition of oligosaccharide to the growth of lactic acid bacteria was effective. Especially, the number of galacto - oligosaccharides was 7.90 × 1010, Therefore, galactooligosaccharide was most effective as a supplement to lactic acid bacteria fermentation of goat ginseng.

Effect of oligosaccharide addition on the fermentation of goat ginseng oligosaccharide pH Acidity Number of lactic acid bacteria living bacteria unit % CFU / ml Galactooligosaccharide Before fermentation 5.200.03 0.0630.05 2.62 x 10 ⁷ After fermentation 3.660.04 0.500.04 7.90 x 10 ¹⁰ Isomaltooligosaccharide Before fermentation 5.180.02 0.0660.03 2.45 x 10 ⁷ After fermentation 4.700.01 0.340.05 2.88 x 10 ¹⁰ Maltooligosaccharide Before fermentation 5.190.04 0.0620.04 2.47 x 10 ⁷ After fermentation 4.280.02 0.380.05 2.90 x 10 ¹⁰ Fructooligosaccharide Before fermentation 5.180.03 0.0640.02 2.41 x 10 ⁷ After fermentation 3.710.04 0.410.06 3.12 x 10 ¹⁰

2) Monitoring fermentation characteristics of goat juice extract according to fermentation condition

In order to establish the optimal fermentation conditions for the extract of L. acidophilus L. lactobacillus Leuconostoc mesenteroides DU-B1604, the fermentation conditions of sixteen sows, designed by the central synthesis program, were used as independent variables for the concentration of goat juice added, fermentation time and sub- The results of pH, acidity and number of lactic acid bacteria are shown in Table 8. The pH and total acidity of the fermented goat ginseng were 3.43 ~ 3.73, 0.37 ~ 0.80%, respectively, and the log CFU / ml and crude saponin content were 0.06 ~ 0.34%, respectively. .

Changes of Physicochemical Composition according to Fermentation Conditions of Goat Ginseng by Central Synthetic Planning Experiment number Experimental data pH Acidity Viable cell count Crude saponin content %  Log CFU / ml % One 3.48 0.79 14.09 0.22 2 3.72 0.47 13.86 0.20 3 3.53 0.64 13.27 0.34 4 3.73 0.41 13.50 0.16 5 3.41 0.74 13.68 0.15 6 3.66 0.41 14.00 0.08 7 3.48 0.60 13.16 0.06 8 3.66 0.37 12.71 0.09 9 3.54 0.55 13.58 0.21 10 3.56 0.55 13.50 0.23 11 3.47 0.62 13.37 0.24 12 3.60 0.53 12.89 0.06 13 3.50 0.61 14.18 0.14 14 3.65 0.48 12.29 0.11 15 3.43 0.80 13.68 0.20 16 3.69 0.41 12.91 0.15

The reaction surface regression analysis using each result was performed and regression polynomials for each dependent variable, namely, pH, acidity, number of lactic acid bacteria and crude saponin content, were obtained and shown in Table 9. The R2 of the fermentation characteristics according to the fermentation condition of the goat ginseng showed a correlation of pH 0.8221, total acidity 0.9682, lactic acid bacteria number 0.8608 and crude saponin content 0.8595. On the other hand, in case of pH, polynomials were recognized to have significance within 10%, polynomials of lactic acid bacteria and crude saponin contents were found to be within 5%, and polynomials of total acidity were considered to be significant except 1% Respectively.

Second order polynomial of goatsam according to fermentation condition Response variable Second order polynomial R ² Significance level
pH Y = 3.718125 + 0.035625X ₁ 0.002396X ₂ 0.098750X ₃ 0.003750X ₁ ² + 0.000069444X ₁ X ₂ + 0.000019290X ₂ ² 0.002500X ₁ X ₃ 0.001528X ₂ X ₃ + 0.010000X ₃ ² 0.8221 0.0921 Acidity Y = 0.455000 0.048125X ₁ 0.001007X ₂ 0.003750X ₃ + 0.006250X ₁ ² + 0.000208X ₁ X ₂ 0.000003858X ₂ ² 0.002500X ₁ X ₃ + 0.002639X ₂ X ₃ + 0.055000X ₃ ² 0.9682 0.0008 Number of lactic acid bacteria Y = 6.443125 + 1.426875X ₁ + 0.075104X ₂ + 1.093750X ₃ 0.102500X ₁ ² 0.004375X ₁ X ₂ 0.000235X ₂ ² 0.032500 X ₁ X ₃ 0.004306X ₂ X ₃ 0.245000X ₃ ² 0.8608 0.0494 Crude saponin content Y = -0.771250 + 0.100625X ₁ + 0.022396X ₂ + 0.145000X ₃ 0.004375X ₁ ² 0.000833X ₁ X ₂ 0.000165X ₂ ² + 0.040000X ₁ X ₃ 0.002500X ₂ X ₃ 0.180000X ₃ ² 0.8595 0.05603

Based on the polynomial equation for the fermentation characteristics according to the fermentation conditions of goat ginseng, a four-dimensional reaction surface graph as shown in FIGS. 4 to 7 can be confirmed. The pH and total acidity of the fermentation broth increased as the concentration of the submaterial increased, and as the fermentation time increased, the pH value tended to be lower and the total acidity was higher. The fermentation time of lactic acid bacteria was in the range of 4.5 ~ 5.5%.

Table 10 shows the effect of the response variables on the lactic acid fermentation of goat ginseng. The pH value was found to be within 5% of the sub - ingredient concentration but was not affected by the concentration of goat 's ginseng and the fermentation time. The total acidity was found to be most influenced by the sub ingredient concentration. The significance within 5% was recognized. The number of lactic acid bacteria was within 5% of the fermentation time, but it was not affected by the other two conditions. The crude saponin content was found to be the most influenced by the concentration of goat juice and was not affected by the other two conditions.

Regression analysis of cyanobacterial fermented product according to fermentation condition Response variable F-Ratio Goat ginseng concentration  Fermentation time Galactooligosaccharide concentration %  hr % pH 0.01 0.70 6.26 ** Acidity 1.59 5.30 ** 39.43 *** Viable cell count 1.31 7.85 ** 0.65 Crude saponin content 7.60 ** 1.58 1.58

The maximum conditions and maximum values of the fermentation characteristics as shown in Table 11 were predicted by ridge analysis of the results of the fermentation of goat ginseng. The pH was in the form of ginseng under the condition of the ginseng concentration 5.04%, the fermentation time 69.09 hours and the galactooligosaccharide concentration 1.90%. The total acidity of the ginseng was 5.26%, the fermentation time 68.44 and the galactooligosaccharide concentration 1.90% The maximum number of lactic acid bacteria was 4.9% in the ginseng concentration, 89.84 hr in the fermentation time and 1.07% in the galactooligosaccharide concentration. The crude saponin content was the highest at 9.05% of the ginseng concentration, 42.00 hr of fermentation time and 0.79% of the additive concentration. Also, the maximum values were estimated to be 3.37, 0.87%, 14.20 Log CFU / ml and 0.32%, respectively.

Optimal fermentation conditions for the physicochemical properties predicted by ridge analysis Response variable Predictive response shape Reaction value Goat ginseng concentration  Fermentation time Galactooligosaccharide concentration Result value %  hr % pH at least 5.04 69.09 1.90 3.37 Benefits maximum 5.03 54.33 0.00 3.74 Acidity at least 4.46 57.61 0.04 0.35 Benefits maximum 5.26 68.44 1.90 0.87 Viable cell count at least 3.98 24.47 0.73 12.27 Maximum value maximum 4.90 89.84 1.07 14.20 Crude saponin content at least 2.62 35.13 0.51 0.004 Maximum value maximum 9.05 42.00 0.79 0.32

3) Monitoring of antioxidant properties of goatsam according to fermentation conditions

The antioxidative properties of the fermented goat yams were investigated by using 16 isolates of Leuconostoc mesenteroides DU-B1604. The total phenolic compounds content of the fermented Lactobacillus acidic L. acidus was 41.53 ~ 69.97 mg%, DPPH radical scavenging activity was 52.26 ~ 70.14%, ABTS radical scavenging activity was 26.99 ~ 50.44 and nitrite scavenging activity was 16.81 ~ 32.57% And showed the difference according to the concentration.

Total Phenolic Compound Contents, DPPH and ABTS Radical Scavenging Ability and Nitrite Scavenging Activity by Goat Fermentation Conditions Based on the Central Synthetic Plans Experiment number Experimental value  Total phenolic compound content DPPH radical scavenging ability ABTS radical scavenging ability Nitrite scavenging performance (mg%) (%) (%) (%) One 62.00 64.65 41.99 27.59 2 59.62 62.59 39.23 24.69 3 60.96 64.00 42.77 25.10 4 58.17 62.01 40.66 24.55 5 48.00 56.64 31.76 20.89 6 46.43 54.46 29.99 19.43 7 49.19 57.93 34.71 18.05 8 46.30 56.52 33.53 17.22 9 55.12 59.58 37.17 25.52 10 54.12 60.22 36.50 25.80 11 69.97 70.14 50.44 32.57 12 41.53 52.26 26.99 16.81 13 51.53 59.42 40.71 22.82 14 48.11 56.37 37.91 20.82 15 62.13 65.56 43.46 27.87 16 49.95 58.12 38.35 23.86

Table 13 shows the polynomials for each antioxidant property of fermented soybean milk fermentation broth according to fermentation conditions. The R2 of the polynomials showed a relatively high correlation with total phenolic compound content of 0.9929, DPPH radical scavenging activity of 0.9063, ABTS radical scavenging activity of 0.8541 and nitrite scavenging activity of 0.9853. The contents of total phenolic compounds, DPPH radical scavenging activity and nitrite scavenging activity were within 1%, and crude saponin content and ABTS radical scavenging activity were within 5%.

Antioxidant properties of fermented products by fermentation conditions of goat ginseng Response variable Second order polynomial R ² Significance level  Total phenolic compound content Y = -8.791875 + 15.582500X ₁ + 0.347604X ₂ + 0.130000X ₃ 0.536563X ₁ ² 0.053854X ₁ X ₂ 0.001589X ₂ ² 0.750000X ₁ X ₃ + 0.034167X ₂ X ₃ + 6.740000X ₃ ² 0.9929 <.0001 DPPH radical scavenging ability Y = -28.460625 + 12.242187X ₁ + 1.252656X ₂ + 1.364500X ₃ 0.495313X ₁ ² 0.040156X ₁ X ₂ 0.010460X ₂ ² 0.677500X ₁ X ₃ 0.145417X ₂ X ₃ + 18.660000X ₃ ² 0.9063 0.0171 ABTS radical scavenging ability Y = 14.695625 + 8.106875X ₁ 0.039688X ₂ 8.220000X ₃ 0.264688X ₁ ² 0.027292X ₁ X ₂ + 0.001892X ₂ ² + 0.895000X ₁ X ₃ 0.091667X ₂ X ₃ + 13.960000X ₃ ² 0.8541 0.0558 Nitrite scavenging performance Y = 25.720625 + 2.513125X ₁ 0.468854X ₂ 10.220000X ₃ 0.095937X ₁ ² + 0.052604X ₁ X ₂ + 0.003290X ₂ ² + 1.190000X ₁ X ₃ 0.205000X ₂ X ₃ + 13.20000X ₃ ² 0.9853 <.0001

Based on the polynomial equation for the antioxidant properties of Table 13 according to fermentation conditions of lactic acid bacteria of goat ginseng, a four-dimensional reaction surface graph as shown in Figs. 8 to 11 was obtained. The antioxidative properties of goat ginseng showed a tendency to be affected by the concentration of goat ginseng and increased with increasing concentration of goat ginseng.

Table 14 shows the effect of condition variables on the fermentation of goat ginseng. The DPPH radical scavenging ability, ABTS radical scavenging ability and nitrite scavenging ability were within 1 ~ 5% in the concentration of goat juice. But not in the range of the other two conditions.

Regression analysis of antioxidant properties of goat juice according to fermentation conditions Response variable F-Ratio Goat ginseng concentration Fermentation time Galactooligosaccharide concentration  % hr %  Total phenolic compound content 203.51 *** 3.42 * 1.62 DPPH radical scavenging ability 12.04 *** 0.60 1.14 ABTS radical scavenging ability 8.03 ** 0.12 0.44 Nitrite scavenging performance 97.62 *** 2.07 1.26

The optimum conditions and the maximum value of the fermentation as shown in Table 15 were predicted through ridge analysis of the fermentation result of goat ginseng. As a result, the DPPH radical scavenging activity was found to be 8.08%, the fermentation time was 43.41 hr, the galactooligosaccharide concentration was 0.91%, the total phenolic compound content was 9.52%, the fermentation time was 36.75 hr and the galactooligosaccharide concentration was 0.52% The nitrite scavenging activity of ABTS was 9.75%, the fermentation time was 57.20 hr and the galactooligosaccharide concentration was 0.55%. The nitrite scavenging activity was the highest at 81.57 mg %, 64.81%, 63.44% and 52.36%, respectively.

Optimal fermentation conditions for the antioxidant properties predicted by ridge analysis Response variable Predictive response shape Reaction value Goat ginseng concentration Fermentation time Galactooligosaccharide concentration Result value  % hr %  Total phenolic compound content at least 2.01 46.55 0.46 29.69 Benefits maximum 9.52 36.75 0.52 81.57 DPPH radical scavenging ability at least 2.06 45.15 0.42 26.83 Benefits maximum 8.08 43.41 0.91 64.81 ABTS radical scavenging ability at least 2.00 47.06 0.47 27.70 Benefits maximum 9.00 40.47 0.79 63.44 Nitrite scavenging performance at least 2.05 51.46 0.54 14.58 Benefits maximum 9.66 57.20 0.55 52.36

As a result of superimposing each reaction surface as shown in FIG. 12 in order to set optimum conditions for crude saponin, total phenolic compound content and DPPH radical scavenging ability by optimum lactic acid fermentation of goat ginseng, It was predicted that the concentration of goat's ghrelin concentrate (25 brix) was 8 to 9 wt%, the fermentation time was 35 to 45 hours, and the concentration of galactooligosaccharide was 0.6 to 0.9%. Table 16 shows the antioxidant properties before and after fermentation. The antioxidant properties were increased by 60.67 ~ 165.20% after fermentation, indicating that the antioxidant properties were enhanced by fermentation.

Prediction of optimal fermentation conditions for the antioxidant properties of multi-lapped reaction surfaces Antioxidant properties Before fermentation (A) After fermentation (B) Growth Rate (B / A) x100  Total phenolic compound content (mg%) 49.11 81.13 165.20 DPPH radical scavenging ability 39.57 65.23 60.67 ABTS radical scavenging ability 54.93 62.89 87.34 Nitrite scavenging performance 42.60 52.36 81.35

4) Monitoring the change of ginsenoside content of goatsam according to fermentation condition

Sixteen strains were isolated by centrifugation using the extract of goat extract Leuconostoc mesenteroides The content of ginsenoside compounds in the fermented samples after fermentation with DU-B1604 was measured, and the results are shown in Table 17. The ginsenoside Rg ₁ content of the goat's fermentation broth was 1.757.21 mg%, the ginsenoside Rb ₁ content was 6.5322.81 mg%, the ginsenoside Rh ₁ content was 1.273.76 mg%, the ginsenoside Rg ₃ content was 2.915 . The content of ginsenoside compound K was 3.568.30 mg%, indicating the difference according to the concentration of goat juice, fermentation time and additive concentration.

Experimental value of ginsenoside content by the fermentation conditions of goat ginseng based on the central synthetic design method Experiment number Experimental value (mg%) Rg ₁ Rb ₁ Rh ₁ Rg ₃ Compound K One 6.17 21.34 3.76 5.33 6.66 2 5.73 20.91 3.59 4.10 5.74 3 6.28 18.75 3.54 5.06 6.10 4 4.75 17.84 3.34 4.41 5.74 5 5.17 17.71 3.20 4.76 6.25 6 4.43 16.20 3.13 4.02 4.68 7 4.95 16.23 2.90 4.20 5.58 8 3.82 15.01 2.56 3.95 4.04 9 5.21 17.96 3.36 4.78 7.96 10 5.17 17.61 3.29 4.65 8.30 11 7.21 22.81 3.75 5.17 6.56 12 1.75 6.53 1.59 2.91 3.56 13 5.08 17.85 3.38 4.99 6.32 14 4.32 15.23 1.27 3.50 3.61 15 5.21 18.60 3.51 4.27 6.24 16 3.85 17.76 3.23 3.32 5.24 Control 4.50 17.19 0.73 0.38 0.068

* Control: ginsenoside content before fermentation of 5% sample of goat's ginseng

Table 18 shows the polynomials for the change in ginsenoside content according to the fermentation conditions of goat ginseng. R ² of the polynomial equation is 0.8483 for ginsenoside Rg ₁ , 0.8487 for ginsenoside Rb ₁ , 0.8455 for ginsenoside Rh ₁ , 0.8415 for ginsenoside Rg _{3 and} 0.9225 for ginsenoside compound K Respectively. On the other hand, the content of ginsenoside compound K is lower than 1% at the level of ginsenoside Rg ₁ , ginsenoside Rb _{1 ,} ginsenoside Rh ₁ and ginsenoside Rg ₃ The contents were within the range of 10%.

Changes of ginsenoside content of fermented products by the fermentation conditions of goat ginseng Response variable Second order polynomial R ² Significance level Rg ₁ Y = -4.681250 + 0.993750X ₁ + 0.128021X ₂ + 7.090000X ₃ 0.044375X ₁ ² + 0.000208X ₁ X ₂ 0.000851X ₂ ² + 0.025000 X ₁ X ₃ 0.061667X ₂ X ₃ 2.640000X ₃ ² 0.8483 0.0616 Rb ₁ Y = -2.736875 + 3.207813X ₁ + 0.188698X ₂ + 2.322500X ₃ 0.194688X ₁ ² + 0.015573X ₁ X ₂ 0.002161X ₂ ² 0.347500X ₁ X ₃ 0.007917X ₂ X ₃ + 1.580000X ₃ ² 0.8487 0.0612 Rh ₁ Y = -5.693750 + 0.807500X ₁ + 0.214375X ₂ + 0.815000X ₃ 0.040937X ₁ ² 0.002083X ₁ X ₂ 0.001736X ₂ ² 0.010000X ₁ X ₃ 0.012500X ₂ X ₃ + 0.180000X ₃ ² 0.8455 0.0646 Rg ₃ Y = -1.468750 + 0.769062X ₁ + 0.096406X ₂ + 1.417500X ₃ 0.042500X ₁ ² 0.003490X ₁ X ₂ 0.000825X ₂ ² + 0.222500X ₁ X ₃ + 0.044583X ₂ X ₃ 3.700000 ₃ ² 0.8415 0.0689 Compound K Y = -20.360625 + 3.021563X ₁ + 0.576615X ₂ + 12.722500X ₃ 0.191875X ₁ ² 0.003906X ₁ X ₂ 0.005495X ₂ ² 0.457500X ₁ X ₃ + 0.024583X ₂ X ₃ 9.560000X ₃ ² 0.9225 0.0101

Based on the polynomial equation for the ginsenoside content in Table 18 according to the fermentation conditions of the goat ginseng, a four-dimensional reaction surface graph as shown in Figure 1317 was obtained. The content of ginsenoside in the fermentation broth was affected by the concentration of goat ginseng and the fermentation time and showed a tendency to increase with increasing concentration of goat 's ginseng and longer fermentation time.

Table 19 shows the effect of condition variables on the fermentation of goat ginseng. Ginsenosides Rg ₁ and Rb ₁ were found to have a large effect on the concentration of goat ginseng and were not affected by the other two conditions. Ginsenoside Rh ₁ was affected by the concentration of goat ginseng and fermentation time And there was little effect on the concentration of galactooligosaccharide within the set range. Also, ginsenoside Rg ₃ The contents were affected by the order of goat juice concentration and galactooligosaccharide concentration, and the effect of fermentation time was not affected within the set range. The content of Compound K was affected by all three conditions.

Regression analysis of ginsenoside content of goat ginseng according to om fermentation condition Response variable F-Ratio Goat ginseng concentration Fermentation time Galactooligosaccharide concentration  % hr % Rg ₁ 6.76 ** 0.51 1.48 Rb ₁ 7.45 ** 0.73 0.13 Rh ₁ 4.17 * 3.74 * 0.16 Rg ₃ 4.26 * 1.60 3.30 * Compound K 11.10 *** 9.46 *** 6.13 **

* Significant at 10% level; ** significant at 5% level; *** significant at 1% level.

The optimum ratios and maximum values of fermentation as shown in Table 20 were predicted through ridge analysis of the fermentation results of goat ginseng. It lost the ginsenoside Rg ₁ content sanyangsam concentration of 9.65%, the fermentation time 49.78 hr, galacto-oligosaccharide concentration 0.69% condition, ginsenoside Rb ₁ content sanyangsam concentration of 9.15%, the fermentation time 62.66 hr, galacto-oligosaccharide concentration in the 0.54% Conditions , ginsenoside Rh ₁ content sanyangsam concentration of 8.02%, the fermentation time 53.01 hr, in galacto-oligosaccharide concentration 0.91% condition, ginsenoside Rg ₃ content sanyangsam concentration of 8.44%, the fermentation time 60.66 hr, galacto-oligosaccharide concentration 0.79% in the condition, ginsenoside compound K content sanyangsam concentration 6.39%, the fermentation time 58.89 hr, the galacto-oligosaccharide concentrations, respectively of 0.94% at the maximum value condition Rg ₁ 6.83 mg%, Rb ₁ 22.31 mg% L, Rh ₁ 3.96 mg%, Rg ₃ 5.45 mg% and compound K 6.64 mg%. The predicted values of the ginsenosides of the samples before fermentation in Table 17 are shown in Table 21. Rg ₁ increased by 1.52 times and Rb ₁ increased by 1.30 times. However, Rh ₁ increased 5.42 times, Rg ₃ increased 14.34 times, and compound K increased 94.86 times. When fermenting under optimal fermentation conditions, Rh _{1 and} Rg ₃ and the compound K are increased.

In order to set optimal conditions for lactic acid bacteria fermentation of goat's ginseng, each reaction surface was superimposed on each reaction surface as shown in FIG. 18 in order to set the optimum conditions for maximizing the contents of ginsenosides Rh ₁ , Rg ₃ and compound K, As shown in Table 22, the fermentation conditions were predicted to be 68% of goat juice concentration, 5565 hours of fermentation time and 0.50.8% of additive concentration. FIG. 19 shows the chromatogram obtained before the fermentation (FIG. 18) and after the fermentation of the ginsenoside in these predicted optimal fermentation conditions.

Optimal fermentation conditions for ginsenoside contents estimated by ridge analysis Response variable Predictive response shape Reaction value Goat ginseng concentration Fermentation time Galactooligosaccharide concentration Result value  % hr % Rg ₁ at least 2.53 41.65 0.28 2.53 Maximum point maximum 9.65 49.78 0.69 6.83 Rb ₁ at least 2.02 47.44 0.44 9.41 Benefits maximum 9.15 62.66 0.54 22.31 Rh ₁ at least 3.65 28.69 0.45 1.44 Benefits maximum 8.02 53.01 0.91 3.96 Rg ₃ at least 2.53 36.15 0.53 3.15 Maximum point maximum 8.44 60.66 0.79 5.45 Compound K at least 2.92 35.45 0.31 2.88 Maximum point maximum 6.39 58.89 0.94 6.64

Comparison of estimated ginsenoside content and ginsenoside content of pre-fermented goat ginseng under optimal fermentation conditions Response variable Estimated value (mg%) (A) Before fermentation (mg%) (B) Increase ratio (A / B) Rg ₁ 6.83 4.50 1.52 Rb ₁ 22.31 17.19 1.30 Rh ₁ 3.96 0.73 5.42 Rg ₃ 5.45 0.38 14.34 Compound K 6.64 0.07 94.86

Prediction of Optimum Fermentation Conditions of Maximum Content of Ginsenosides by Multiple Lapping Reaction Surface Fermentation condition unit Predicted condition Goat ginseng concentration % 68 Fermentation time hour 5565 Galactooligosaccharide content % 0.50.8

The present invention is an extremely useful invention in the health functional food industry because it provides an antioxidant effect and a specific ginsenoside Rg ₃ , Rh ₁ and Compound K content enhanced lean crude ginseng fermented product using Leucomostic genus DUB 1604.

Korea Biotechnology Research Institute KCTC13015BP 20160425

Claims (7)

A first step (S1) of concentrating the goat's ginseng extract; A second step (S2) of adjusting the goat's ghe concentrate obtained in the above step to 1 to 50% by volume of the total volume; A third step (S3) of adding 0.1 to 30% by volume of galactooligosaccharide to the total volume of the adjustment solution obtained in the above step, adjusting the pH to 5.0 to 7.0, and sterilizing the solution; The leuconostoc mesenteroides DU-B1604 (KCTC 13015BP), which was pre-cultured in a commercially available MRS medium, was inoculated into the soymilk composition obtained by sterilizing in the above step, Step S4; The fifth step (S5) of fermenting the composition liquid obtained by inoculating the lactic acid bacterium obtained in the above step at a fermentation temperature of 20 to 45 ° C and a fermentation time of 6 to 120 hours and having an antioxidative activity and containing ginsenosides Rg ₃ , Rh ₁ and compound K Wherein the content of the lactic acid bacteria is increased.
[6] The method of claim 1, further comprising a sixth step of sterilizing the fermented goat gum obtained in the fifth step at a sterilization temperature of 60 to 100 DEG C for 10 to 60 minutes, filtering the microorganism through a microfilter filter Wherein the fermented product is a fermented product.
[Claim 2] The method according to claim 1, wherein the liquid goat ginseng obtained in the first step is selected from at least one selected from the group consisting of a ginseng root liquor, a goat ginseng extract, a goat ginseng concentrate, a dried goat ginseng product, and a goat ginseng granule.
The method according to claim 1, wherein the galactooligosaccharide in step (3) is prepared by selecting at least one selected from maltooligosaccharide, fructooligosaccharide, and isomaltooligosaccharide Wherein the fermented product is a fermented product.
The method of claim 2, wherein the fermentation product obtained in the fifth step or the sterilization filtration fermentation product obtained in the sixth step is 0.1 ~ 99.9 wt% or an apple, persimmon, blueberry, blackberry, black currant, Pomegranate, grape, pear, peach, banana, strawberry, tomato, carrot, plum, plum, cherry, A raw material fruit made of mango, lemon, pineapple or a juice or a concentrate thereof is added in an amount of 0.1 to 99.9 parts by weight or as part of a skin.
A fermented product of lactic acid bacteria of the goat ginseng having antioxidant activity produced by the method of any one of claims 1 to 5, wherein the content of ginsenosides Rg ₃ , Rh ₁ and compound K is enhanced.
A health-functional food composition comprising the fermented product of Lactobacillusthus sinensis having the antioxidant activity of claim 6 and having enhanced content of ginsenosides Rg ₃ , Rh _1, and Compound K as an active ingredient.

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