KR20180057278A - Lactobacillus sakei 2-6-4 and its use - Google Patents

Abstract

According to the present invention, a Lactobacillus sakei 2-6-4 strain deposited with an accession number, KCCM11861P, can efficiently produce metabolic ginsenosides Rh2 and Rg3. Lactic acid bacteria belonging to lactobacillus are generally difficult to be solid-fermented, and there is a problem of having a non-uniform decomposition activity. However, specifically, the strain according to the present invention is lactic acid bacteria, has an Rh2 or Rg3 production activity which is very effective in solid fermentation, and effectively produces metabolic ginsenoside Rh2 or Rg3 even though only ginseng powder is used as a unique nutrient source without other nutrients. Thus, it is possible to produce a ginseng including a high concentration of Rh2 or Rg3 even without performing a conventional steaming, drying, and aging process.

Description

Lactobacillus saccharum 2-6-4 strain and its use {Lactobacillus sakei 2-6-4 and its use}

The present invention relates to a strain 2-6-4 of Lactobacillus saccharomyces. More particularly, the strain 2-6-4 of Lactobacillus saccharum is cultured in a solid medium having ginseng as a sole nutrient source to produce Rh2 or Rg3 Lt; / RTI >

Ginseng is a perennial herbaceous herbaceous plant belonging to Panax ginseng (Araliaceae) in plant taxonomy and has long been used as an important medicinal herb in oriental medicine. Ginseng is used as it is or dried, and red ginseng is also used by making ginseng by steaming. It has been reported that when red ginseng is prepared by boiling ginseng, the pharmacological action of ginseng is changed and the content of the components that existed in the ginseng is increased or the content of the components that existed in the ginseng is increased.

Recently, according to the results of the efficacy test of Korea Ginseng Products Association, Ginsenosides Rg3, which is a typical anti-cancer component and saponin component having blood pressure lowering effect, neuronal cell protection function, antithrombogenic effect, In addition, black ginseng is much more effective than red ginseng in red ginseng, which means that red ginseng rather than ginseng has a greater effect on saponin pharmacological action than red ginseng.

As described above, the ginseng is more than the red ginseng, and the ginseng is more active than the ginseng. However, the production method of black ginseng, known to date, is complicated in manufacturing process, takes a long time, and fails in the manufacturing process, so that it is industrialized at a high price and difficult to be used in everyday life.

For example, Korean Patent No. 0543862 discloses a black and ginseng concentrate with 9 active ingredients and 10 dryings, which are excellent in the content of active ingredients. Korean Patent No. 0496418 discloses a method for producing black rice flour However, there is a disadvantage that the manufacturing process is complicated and takes a long time.

In order to solve the above-mentioned problems, a method for producing black ginseng having improved ginsenoside content while remarkably reducing the number of steaming and drying times has recently been developed. However, although it has been dramatically reduced, it is still a process of aging using an aqueous solution of calcium carbonate There is still a problem in that it is necessary to fill the nitrogen gas to be diluted and dried, and to use high-pressure equipment.

In order to solve these problems, a new microorganism capable of producing Rh2 or Rg3 is cultured in a solid medium having ginseng as a sole nutrient source, and a high content of metabolic type ginsenoside is produced using an efficient and environmentally friendly fermentation process It is required to develop a method that can be used.

SUMMARY OF THE INVENTION The present invention has been made in view of the above problems, and it is an object of the present invention to provide a novel microorganism Lactobacillus saccharum 2-6-4 strain having excellent activity even in a solid medium.

Another object of the present invention is to provide a starter composition for the production of Rh2 or Rg3-fortified ginseng powder containing the strain Lactobacillus sakei 2-6-4 or a culture thereof.

Still another object of the present invention is to provide Rh 2 or Rg3-fortified ginseng powder obtained by solid-fermenting ginseng powder with the strain 2-6-4 of Lactobacillus sakei .

In order to achieve the above object, the present invention provides Lactobacillus saccharum 2-6-4 strain (Lactobacillus sakei 2-6-4) deposited with Accession No. KCCM11861P.

The strain is characterized in that it is grown in a solid medium containing ginseng.

The ginseng powder-containing medium comprises 10 to 40% by weight of ginseng powder based on the total weight of the whole solid medium.

The strain is characterized in that it grows in a solid medium containing ginseng as a sole nutritional source.

The strain is characterized by producing Rh2 or Rg3 from solid medium containing ginseng powder.

The optimum growth conditions of the strain are (a) fermentation temperature of 25-37 ° C, (b) pH 4.1-4.5, and (c) culturing time of 24-48 hours.

The present invention also provides a starter composition for the production of Rh2 or Rg3-fortified ginseng powder containing a strain Lactobacillus sakei 2-6-4 or a culture thereof.

The present invention also provides Rh2 or Rg3-fortified ginseng powder obtained by solid-fermenting ginseng powder with the strain 2-6-4 of Lactobacillus sakei .

According to the present invention, the lactobacillus saccharose 2-6-4 strain deposited with accession number KCCM11861P can efficiently produce the metabolic ginsenosides Rh2 and Rg3.

Lactobacillus belonging to the genus Lactobacillus generally has difficulty in solid fermentation and has an uneven decomposition activity. Specifically, the strain according to the present invention has activity of producing Rh2 or Rg3 which is highly effective in lactic acid bacteria and solid fermentation, Although the ginseng powder is the only nutrient source without a nutritional source, it produces the metabolic ginsenosides Rh2 or Rg3 effectively. Therefore, it is possible to produce the metabolic ginsenosides containing high concentrations of Rh2 or Rg3 without the conventional processes such as drying, Ginseng can be produced.

In addition, if it is applied to the production process of ginseng containing metabolic ginsenosides, Rh2 or Rg3 can be effectively produced from a solid medium containing ginseng powder only by a simple step of fermentation. Therefore, Addition of a device in the incubator can be omitted.

BRIEF DESCRIPTION OF THE DRAWINGS FIG. 1 is a photograph of a culture solution obtained by culturing a strain of microorganism Lactobacillus saccharum 2-6-4 according to the present invention in a solid medium containing ginseng powder at various concentrations. FIG.
FIG. 2 is a graph showing the number of viable cells of six strains according to the concentration of ginseng powder-containing solid medium measured from the procedure of Experimental Example 2. FIG.
FIG. 3 is a graph showing the results of Experimental Example 4. Specifically, FIG. 3 shows growth curves of Lactobacillus saccharum 2-6-4 strain at 2 ° Brix ginseng concentrate containing 10% MRS according to temperature.
FIG. 4 is a graph showing the results of Experimental Example 5. Specifically, FIG. 4 shows the growth curve of the strain 2-6-4 of Lactobacillus saccharum over time in a solid medium containing 20% ginseng powder.

Hereinafter, various aspects and various embodiments of the present invention will be described in more detail.

The present invention provides a strain 2-6-4 of Lactobacillus sakei deposited with Accession No. KCCM11861P.

Lactobacillus saccharides 1, 2 and 5 isolated from Kimchi and Lactobacillus saccharose 3 and 4 purely isolated from doenjang were used to isolate the metabolic ginsenosides Lactobacillus saccharide strain 2-6-4, which can produce Rh 2 or Rg 3 most efficiently, was selected.

The strain 2-6-4 of Lactobacillus saccharum was found to have a fast growth rate in the solid medium containing ginseng powder containing ginseng powder as the only nutrient source and to produce metabolic ginsenosides Rh2 or Rg3 at the highest concentration during the cultivation, Was selected.

The 16S rRNA was extracted from the selected Lactobacillus saccharum 2-6-4 strain and analyzed for its base sequence, and it was confirmed that it belonged to Lactobacillus sakei . The strain thus obtained was deposited with the Korean Center for Microbiological Conservation (KCCM) and given the deposit number KCCM 11861P.

The strain 2-6-4 of Lactobacillus sakei according to the present invention is preferably but not limited to be derived from doenjang.

The strain of the present invention is characterized in that it is grown in a solid medium containing ginseng powder. The ginseng powder-containing medium may further include a nutrient source in addition to the ginseng powder. However, the present invention is not limited to obtaining a high concentration of metabolic ginsenoside (Rh2, Rg3) It is most preferable to cultivate in a solid medium containing ginseng powder as a sole nutrient source.

That is, the strain according to the present invention is characterized in that the strain is cultured in a solid medium containing ginseng powder as a sole nutritional source, and produces Rh2 or Rg3, which is a metabolic type ginsenoside, at a higher concentration.

In general, ginseng consists of about 3 ~ 6% of saponin, about 12 ~ 15% of nitrogen compound, about 1 ~ 2% for fat - soluble component, about 50 ~ 60% for carbohydrate and about 4 ~ 6% Saponin which is an active substance. Saponins, namely ginsenosides, can be divided into three major groups depending on their molecular structure: Panaxadiol (PD), Panaxatriol (PT) and Oleanane, The paper was found.

Among them, the pyrazyldiol-based ginsenosides Rh2 and Rg3 are rarely present in common ginseng, and are newly produced from Rb1 through a steaming process or an additional hydrolysis process such as an acid and an enzyme, 1 and formula (2).

[Chemical Formula 1] Rh 2

[Formula 2] Rg3

Rh2 or Rg3 has been particularly effective for anticancer, cancer treatment, reduction of anticancer drug toxicity, immunity enhancement and prevention of thrombosis, and various techniques for strengthening these components have been developed. However, It is limited. These methods are not only costly and cumbersome in process, but also have various economical disadvantages such as difficult management.

The strain of the present invention has an advantage that Rh2 or Rg3 can be obtained from the ginseng powder-containing solid medium only by growing the strain in a solid medium containing ginseng powder as a sole nutritional source.

That is, unlike the conventional techniques, the strain according to the present invention can produce sufficient Rh2 or Rg3 by solid fermentation alone, not in liquid culture. Therefore, the process is simple and requires no separate process or apparatus, Since it uses itself as a nutrient source, no additional steps are required, such as processing, extraction or drying of the raw material, so that the cost can be significantly reduced.

The solid medium containing the ginseng powder for culturing the strain may be 10-40 wt%, preferably 15-35 wt% of ginseng powder based on the total weight of the total solid medium, , More preferably from 20 to 30 wt% of the ginseng powder, based on the total weight of the total solid medium. If the ginseng powder is contained in an amount of less than 10% by weight based on the total weight of the whole solid medium, the productivity tends to deteriorate. If it exceeds 40% by weight, the uniformity may be lowered, There is a problem.

However, in the solid medium containing the ginseng powder, it is preferred that those containing 20 to 30% by weight of ginseng powder based on the total solid weight of the solid medium have the highest productivity of Rh2 or Rg3 and the most active growth activity and uniformly Rg2 or Rg3 Is most preferable.

The optimum growth conditions of the strain are (a) a fermentation temperature of 25-37 ° C, (b) a pH of 4.1-4.5, and (c) a cultivation time of 24-48 hours. Although the fermentation temperature is in a range of room temperature And the pH is also in a weakly acidic condition, so that the cultivation conditions are not complicated, and thus the cost can be reduced because the cultivation technique is simple.

Furthermore, as has been confirmed in the experimental examples to be described later, the strain Lactobacillus saccharum 2-6-4 according to the present invention produced about 0.013 mg / g of ginsenoside Rh2 from ginseng powder within 24 to 48 hours after solid fermentation (Dry weight of fermented product) and 0.177 mg / g of Rg3 (based on dry weight of ginseng fermented product). It has an effect of producing Rg3 of 0.003 mg / g, which is known to be contained in white ginseng products, and Rg3 of about 59 times and Rh_2 of about 100 times that of Rh2 of 0 mg / g.

In other words, the strain Lactobacillus saccharum 2-6-4 can produce Rh2 or Rg3 at a high concentration through solid fermentation, which can be confirmed by the following experimental examples.

Therefore, the above strain can be applied to a solid medium containing only solid ginseng powder to produce Rh2 or Rg3 at a high concentration from the ginseng powder in spite of solid fermentation.

The solid medium does not contain a carbon source, an organic acid, an inorganic element, or the like but only a ginseng powder as a sole nutritional source. Thus, the production cost can be remarkably reduced and the production of the strain and the production of Rh2 or Rg3 can be promoted.

The strain has a high Rh2 or Rg3 production yield, and has a growth characteristic after solid fermentation, and when applied to a solid medium containing ginseng powder, excellent ginseng powder containing high concentration of ginsenoside Rh2 or Rg3 Product or powder of fermented ginseng can be obtained.

The ginseng powder is not particularly limited, but may be powder. The powder may be prepared by grinding white ginseng, fresh ginseng, direct ginseng, ginseng, bangkoku, raw ginseng, and ginseng using a pulverizer. The degree of powdering of ginseng is not particularly limited, and it is sufficient that the strain can efficiently penetrate into the tissue or fibrin of ginseng. Preferably, the degree of powdering of ginseng is about 30 to 150 mesh.

The ginseng may be one using at least one selected from roots, leaves and stem of ginseng.

Therefore, the strain 2-6-4 of Lactobacillus sakei according to the present invention is useful as a ginseng powder having enhanced effects of Rh2 or Rg3, which is a physiologically active substance of ginseng having various effects such as anti-cancer, A starter composition or a food additive composition capable of producing a starch composition. The composition may contain live bacteria, dried germs or cultures of Lactobacillus sacae according to the present invention as an active ingredient, and may further contain excipients or carriers. The culture includes a culture itself cultured in a solid medium, a processed culture obtained by filtering or drying or diluting the culture, and the like. The content of Lactobacillus saccharum 2-6-4 in the composition may vary depending on the composition use and formulation.

The compositions can be prepared and administered in a variety of formulations and methods. For example, a strain of Lactobacillus saccharae 2-6-4 or a culture thereof is mixed with a carrier and a flavoring commonly used in the pharmaceutical field to prepare tablets, troche, capsules, May be manufactured and administered in the form of elixir, syrup, powder, suspension or granule, and the like. Examples of the carrier include binders, lubricants, disintegrants, excipients, solubilizers, dispersants, stabilizers, suspending agents and the like. The administration method may be oral, parenteral or application, but is preferably orally administered. The dose may be appropriately selected depending on the degree of absorption, inactivation rate and excretion rate of the active ingredient in the body, age, sex, condition, etc. of the subject.

The strain Lactobacillus saccharum 2-6-4 according to the present invention or a culture thereof can be used as a food additive for foods such as kimchi, beverages, baby foods, and dairy products. In addition, the Lactobacillus saccharum strain 2-6-4 of the present invention can be used as a starter for the production of Rh2 or Rg3-fortified ginseng powder. The Rh2 or Rg3-fortified ginseng powder using the Lactobacillus saccharide 2-6-4 strain of the present invention can be prepared according to a conventional method known in the art, and the ginseng powder is added to the Lactobacillus sakei ) It may be a solid fermented with strain 2-6-4.

In the context of the present invention, the term "fortification" as used herein refers to a composition comprising 2 times, preferably 10 times, more preferably 20 times, more preferably 100 times Rh2 component, 1.5 times, preferably 2.0 times, Means that Rg 2 component of 2.5 times is contained.

Specifically, the strain Lactobacillus saccharum 2-6-4 or a culture thereof can be used for the production of Rh2 or Rg3-containing ginseng. The present invention relates to a method for producing fermented ginseng containing Rh2 or Rg3, which is preferably fermented by inoculating the lactobacillus saccharide strain 2-6-4 into a solid medium containing ginseng powder.

Solid fermentation in which a strain is inoculated into the solid medium does not cause serious contamination as in liquid fermentation because the growth of contaminants is limited due to low water activity. In case of performing liquid fermentation and solid fermentation, Rh2 or Rg3 productivity is high in fermentation. In addition, the metabolic type ginsenoside Rh2 or Rg3 produced by the solid fermentation method does not require a separate process, so that the stability is high. In the case of solid fermentation, the strain has a low level of metabolic inhibition and a higher amount of the metabolic ginsenoside Rh2 Or Rg3. Furthermore, since the wastewater is hardly generated during processing and the fermentation residues are low in water content, it can be used as a ginseng product fermented without additional drying and processing steps. The medium used for solid fermentation is an artificial synthetic medium used for liquid fermentation Relatively simple and relatively inexpensive. On the other hand, the production facilities required for solid fermentation are economical compared to the production facilities required for liquid fermentation, and the solid fermentation methods require relatively low energy to be used for controlling aeration and temperature as compared with liquid fermentation methods.

The solid medium to be used in the solid fermentation step is not added with carbon source, organic acid, inorganic element, etc., but only the ginseng powder is made the only nutrient source. Thus, the manufacturing cost can be remarkably reduced and the production of the strain and the production of Rh2 or Rg3 . ≪ / RTI >

The strain has a high Rh2 or Rg3 production yield, and has a growth characteristic after solid fermentation, and when applied to a solid medium containing ginseng powder, excellent ginseng powder containing high concentration of ginsenoside Rh2 or Rg3 Product or a fermented ginseng powder can be obtained.

The ginseng powder is not particularly limited, but may be powder. The powder may be prepared by grinding white ginseng, fresh ginseng, direct ginseng, ginseng, bangkoku, raw ginseng, and ginseng using a pulverizer. The degree of powdering of ginseng is not particularly limited, and it is sufficient that the strain can efficiently penetrate into the tissue or fibrin of ginseng. Preferably, the degree of powdering of ginseng can be such that the average particle size is about 1 to 500 mu m.

The ginseng may be one using at least one selected from roots, leaves and stem of ginseng.

On the other hand, the composition of the solid medium was selected on the basis of the content of ginseng powder in the solid medium, and the fermentation characteristics and Rh2 or Rg3 productivity of Lactobacillus saccharum strain 2-6-4 were evaluated in each medium. As a result, The ginseng powder contained in the culture medium may be 10-40 wt%, preferably 15-35 wt%, more preferably the most Rh2 or Rg3 productivity and has the highest growth activity The most active ginseng powder may contain 20 to 30 wt% of ginseng powder based on the total weight of the total solid medium so that Rg2 or Rg3 is uniformly produced.

For the solid medium, up to about 5 wt.% Based on the total weight of the total solid medium was found to be a universal level of addition, but in the present invention much higher levels of solid fermentation were found to be adequate levels of addition.

More specifically, the solid medium may be one prepared by suspending the ginseng powder in water.

The solid fermentation step is preferably carried out at a temperature of from 25 to 37 DEG C and the optimum fermentation temperature is found to be 30 DEG C. When the solid fermentation temperature is lower than 25 ° C or higher than 37 ° C in mass production, the growth activity of the strain may be lowered, resulting in problems such as a process delay.

The initial inoculum size of the solid fermentation step is preferably 10 ⁵ to 10 ⁸ CFU / g, and the optimal inoculum level of the strain 2-6-4 of Lactobacillus sakei is active at about 10 ⁷ CFU / g Respectively. On the other hand, in the case of Lactobacillus saccharomyces 2-6-4, the optimum fermentation time showed the highest Rh2 or Rg3 productivity in 24 to 48 hours, and the Rh2 or Rg3 content did not change in 48 hours. Over fermentation is not cost effective.

The strain is Lactobacillus saccharum 2-6-4 isolated from miso, and is harmless to human body. A strain of Lactobacillus isolated from doenjang does not produce Rh2 or Rg3 through solid fermentation unconditionally. This can be confirmed in the selection process of the experimental example described later of the present invention.

In other words, the Lactobacillus saccharide strain 2-6-4 selected in the present invention has the highest productivity of Rh2 or Rg3 as well as the growth activity in the solid fermentation of ginseng powder. Respectively.

By the above-mentioned process, the strain according to the present invention is simply inoculated into a solid medium containing ginseng powder and fermented without any cumbersome process of improving and drying specific physiologically active substances of impression like red ginseng, The side Rh2 or Rg3 can be most efficiently produced.

In other words, Rh2 or Rg3 can be produced at the same or superior concentration as white ginseng products or red ginseng products without any additional processing.

Hereinafter, the present invention will be described in more detail with reference to Examples and the like, but the scope and content of the present invention can not be construed to be limited or limited by the following Examples. It will be apparent to those skilled in the art that various modifications and variations can be made in the present invention without departing from the spirit and scope of the present invention as set forth in the following claims. It is natural that it belongs to the claims.

<Measurement method>

1) Number of living cells

The culture medium or the fermentation product to be measured for the viable cell count was diluted 10-fold, plated on an MRS plate culture medium, cultured at 30 DEG C for 24 hours, and colonies formed were counted to confirm viable cell count.

2) ginsenoside content (HPLC)

The ginsenosides were quantitatively analyzed by performing high performance liquid chromatography (HPLC) (Jasco Co., Japan) on the culture solution or the fermentation product to be assayed for ginsenoside content.

Specifically, a culture medium or a fermentation mule to be measured was obtained, lyophilized, extracted with 70% methanol, filtered again with a 0.45 μm filter, and used for analysis. The columns used were GS520 (7.5 mm x 300 mm L, Showa Denko Co., Toyko, Japan) + GS 320 (8.0 mm x 300 mm L, Showa Denko Co., Toyko, Japan) and 0.2 M NaCl The sample injection volume was 20 μl and the flow rate was 0.4 ml / min and detected with a refractive index detector (Jasco Co., Toyko, Japan).

The detected peaks were confirmed by comparison with the retention time of reference compounds.

3) polysaccharide analysis

For the molecular weight determination, 20 mg of the standard substance and 20 mg of ginseng powder extract extracted with hot water at 100 ° C for 2 hours were dissolved in 1 ml of 0.2 M NaCl, and then filtered through membrane filter paper and used for HPLC (Jasco Co., Japan) analysis.

The columns used were GS520 (7.5 mm x 300 mm L, Showa Denko Co., Toyko, Japan) + GS 320 (8.0 mm x 300 mm L, Showa Denko Co., Toyko, Japan) and 0.2 M NaCl The sample injection volume was 20 μl and the flow rate was 0.4 ml / min and detected with a refractive index detector (Jasco Co., Toyko, Japan). The molecular weight of the purified polysaccharide was measured in comparison with the standard curve obtained using the Fluka product (molecular size; 10K, 48.8K, 366K) as a standard.

Experimental Example 1: Selection of 6 strains

Traditional fermented foods such as kimchi and doenjang were suspended in sterilized fresh saline solution to obtain a supernatant, which was then plated on a BL Hanfan medium, a lactic acid bacterin cell culture medium, to select lactic acid bacteria - specific colonies. The selected strains were separated by pure separation and cultured in a sterile 5% ginseng powder-containing solid medium. The major ginsenoside metabolite changes were measured from the culture medium, and the best 5 kinds of lactobacillus such as Lactobacillus were finally selected.

For comparison, a known strain of Pediococcus pentosaceus LY-011 known in Korean Patent Registration No. 10-1240663 was also used in the experiment. Five selected strains of lactic acid bacteria and known strains were listed in Table 1 below.

The ginseng powder-containing solid medium was prepared by mixing 5 wt% of ground ginseng powder having an average particle size of 100 mu m and 95 wt% of water.

Strain
number Strain name Separation source One Lactobacillus saccharae 1 Kimchi 2 Lactobacillus casei 2 Kimchi 3 Lactobacillus casei 3 Miso 4 Lactobacillus case 4 Kimchi 5 Lactobacillus saccharum 2-6-4 Miso 6 Pediococcus pen Tossa Sius LY-011 Kimchi

Experimental Example 2: Selection of one strain (Lactobacillus saccharum 2-6-4 strain)

Among the six strains of Experimental Example 1, one strains having the highest production capacity for Rh2 or Rg3 and having excellent growth activity in a solid medium containing a high content of ginseng powder were selected.

1) Measurement of growth activity

First, the growth whether performed for a pre-MRS (lactobacilli, leuconostoc and weisella strain) before incubation (pre-culture) to the medium before inoculation of each strain-containing solid medium of 20% ginseng powder day, measure its OD ₆₀₀ value Respectively.

1% (107 CFU per gram of medium of kimchi medium) of each of the pre-cultivated strains was mixed with various concentrations of ginseng powder-containing solid medium (ginseng powder 5, 10, 15, 20, 25% by weight and the remaining weight of water), and cultured at 30 DEG C for 48 hours to obtain a ginseng fermented product of each strain from solid medium containing various concentrations of ginseng powder.

First, in order to confirm the growth activity, the viable cell count per g of the ginseng fermented product obtained through the above procedure was measured and shown in Fig.

As shown in FIG. 2, the strain 2-6-4 of Lactobacillus saccharum was found to have the best growth activity in a solid medium containing ginseng powder at a high concentration. Specifically, the Lactobacillus saccharum strain 2-6-4 according to the present invention has increased growth activity from a ginseng powder-containing medium containing 10% by weight or more of ginseng powder. In particular, the strain Lactobacillus saccharum 2-6-4 according to the present invention showed the best growth activity in the ginseng powder-containing medium containing 20% by weight or more of ginseng powder.

2) Measurement of production capacity for Rh2 or Rg3

Next, the concentrations of total ginsenosides, ginsenosides Rb1, Rh2 and Rg3 (mg / g) from the fermented ginseng obtained through the above Experimental Example 2-1) were analyzed and are summarized in the following table.

Table 2 shows the total concentrations of ginsenosides, ginsenosides Rb1, Rh2 and Rg3 (mg / g) measured in the ginseng fermentation products of each strain obtained from the solid medium containing 5% ginseng powder,

Table 3 shows the total concentrations of ginsenosides, ginsenosides Rb1, Rh2 and Rg3 (mg / g) measured in the ginseng fermentation products of each strain obtained from the solid medium containing 10% ginseng powder,

Table 4 shows the total concentrations of ginsenosides, ginsenosides Rb1, Rh2 and Rg3 (mg / g) measured in the ginseng fermentation product of each strain obtained from the solid medium containing 15% ginseng powder,

Table 5 shows the total concentrations of ginsenosides, ginsenosides Rb1, Rh2 and Rg3 (mg / g) measured in the ginseng fermentation product of each strain obtained from the solid medium containing 20% ginseng powder,

Table 6 shows the total concentrations of ginsenosides, ginsenosides Rb1, Rh2 and Rg3 (mg / g) measured in the ginseng fermentation products of each strain obtained from the solid medium containing 25% ginseng powder.

The concentrations of the total ginsenosides, ginsenosides Rb1, Rh2, and Rg3 (mg / g) were analyzed by HPLC, respectively. The concentrations were calculated by dividing the ginseng fermented product by the total ginsenosides measured for the dry solid g, (Mg) of side Rb1, Rh2, and Rg3. The above values are obtained by recording an average of the results of five repeated experiments.

In the present invention, the control group means a solid medium containing ginseng powder before fermentation.

** Total ginsenosides are represented by Rg1, Re, Rf, Rg2 + Rh1, Rb1, Rh1 (R), Rc, Rb2, Rb3, Rf1, Rd, Rf2, Rg3 CK, and Rh2.

Strain classification ** Total ginsenoside concentration (mg / g) Concentration of ginsenoside Rb1 (mg / g) Concentration of ginsenoside Rh2 (mg / g) Concentration of ginsenoside Rg3 (mg / g) Control group 1.096 0.124 0 0.083 Lactobacillus saccharae 1 1.112 0.127 0 0.089 Lactobacillus casei 2 1.080 0.12 0 0.092 Lactobacillus casei 3 1.051 0.136 0 0.079 Lactobacillus case 4 0.927 0.118 0 0.081 Lactobacillus saccharum 2-6-4 0.993 0.119 0 0.105 Pediococcus pen Tossa Sius LY-011 1.217 0.116 0 0.099

Strain classification ** Total ginsenoside concentration (mg / g) Concentration of ginsenoside Rb1 (mg / g) Concentration of ginsenoside Rh2 (mg / g) Concentration of ginsenoside Rg3 (mg / g) Control group 0.91 0.112 0 0.083 Lactobacillus saccharae 1 0.814 0.099 0 0.084 Lactobacillus casei 2 0.790 0.119 0 0.087 Lactobacillus casei 3 1.131 0.104 0 0.085 Lactobacillus case 4 0.909 0.113 0 0.075 Lactobacillus saccharum 2-6-4 0.848 0.123 0.001 0.096 Pediococcus pen Tossa Sius LY-011 1.010 0.118 0 0.101

Strain classification ** Total ginsenoside concentration (mg / g) Concentration of ginsenoside Rb1 (mg / g) Concentration of ginsenoside Rh2 (mg / g) Concentration of ginsenoside Rg3 (mg / g) Control group 0.983 0.116 0 0.084 Lactobacillus saccharae 1 0.765 0.108 0 0.078 Lactobacillus casei 2 0.743 0.115 0 0.081 Lactobacillus casei 3 1.051 0.121 0 0.085 Lactobacillus case 4 0.962 0.108 0 0.080 Lactobacillus saccharum 2-6-4 1.033 0.089 0.01 0.116 Pediococcus pen Tossa Sius LY-011 1.091 0.1 0 0.082

Strain classification ** Total ginsenoside concentration (mg / g) Concentration of ginsenoside Rb1 (mg / g) Concentration of ginsenoside Rh2 (mg / g) Concentration of ginsenoside Rg3 (mg / g) Control group 0.994 0.111 0 0.088 Lactobacillus saccharae 1 0.809 0.1 0 0.084 Lactobacillus casei 2 0.786 0.114 0 0.076 Lactobacillus casei 3 0.965 0.11 0 0.065 Lactobacillus case 4 0.982 0.109 0 0.080 Lactobacillus saccharum 2-6-4 1.171 0.033 0.013 0.177 Pediococcus pen Tossa Sius LY-011 1.103 0.105 0 0.080

Strain classification ** Total ginsenoside concentration (mg / g) Concentration of ginsenoside Rb1 (mg / g) Concentration of ginsenoside Rh2 (mg / g) Concentration of ginsenoside Rg3 (mg / g) Control group 0.908 0.113 0 0.079 Lactobacillus saccharae 1 0.842 0.101 0 0.069 Lactobacillus casei 2 0.818 0.11 0 0.073 Lactobacillus casei 3 0.824 0.109 0 0.072 Lactobacillus case 4 0.916 0.114 0 0.078 Lactobacillus saccharum 2-6-4 1.038 0.042 0.013 0.138 Pediococcus pen Tossa Sius LY-011 1.008 0.104 0 0.081

As a result of looking at Tables 2 to 6, it was confirmed that the strain Lactobacillus saccharum 2-6-4 has excellent growth activity in a solid medium containing ginseng powder at a high concentration, and also has excellent production capacity of Rh2 or Rg3.

Furthermore, the strain Lactobacillus saccharum 2-6-4 according to the present invention can confirm that the ginseng powder increases the growth activity from the solid medium containing a high content of ginseng powder. However, the productivity of Rh2 or Rg3 is the most excellent and the growth activity is most active, and Rg2 or Rg3 is uniformly produced when 20 to 30% by weight of ginseng powder is contained based on the total weight of the whole solid medium.

That is, the strain Lactobacillus saccharum 2-6-4 was finally selected in the present invention.

 The strain Lactobacillus saccharum 2-6-4 according to the present invention contains about 0.01 to 0.013 mg / g of ginsenoside Rh2 and 0.096 to 0.177 mg / g of Rg3 on the basis of the solid content of dried ginseng fermented product after solid fermentation Production. It has an effect of producing Rg3 of about 0.003 mg / g, which is known to be contained in white ginseng products, and about 59 times of Rg3 and about 100 times of Rh2, than that of Rh2 of 0.0 mg / g.

In addition, the results of Tables 2 to 6 show total ginsenosides, ginsenosides Rb1, Rh2, and Rg3 measured on the basis of the solid content of the dried ginseng fermented product. There was not much difference in case. However, from the solid medium containing 10% by weight or more of ginseng powder, it was confirmed that Rh2 and Rg3 were produced in Lactobacillus saccharum 2-6-4 strain.

Also, when a solid medium containing 20 to 25% by weight of ginseng powder was used, the Lactobacillus saccharum strain 2-4-6 according to the present invention showed a decrease in Rb1, an intermediate metabolite of metabolic ginsenosides, Of Rh2 and Rg3. In contrast, the growth activity was significantly lower in the other strains, Rh2 was not produced at all, and Rg3 was not significantly increased in the amount present in the initial ginseng powder-containing medium.

Experimental Example 3: Identification of Strain

The identification of the strains finally selected in Experimental Example 2 was carried out using 16S rDNA gene sequencing.

The identification of the strain was carried out by asking Macrogen Co., Ltd., and the nucleotide sequence of the 16S rRNA gene and the ITS region were analyzed. As a result, 1884661 nucleotide sequences of the rRNA gene of the strain were analyzed, and it was confirmed that it was 99% homologous with the complete genome sequence of Lactobacillus sakei 23K standard strain. Thus, the strain was identified as a strain of Saccharomyces cerevisiae in the genus Lactobacillus.

Therefore, the strain of the present invention was designated as Lactobacillus saccharum 2-6-4, and this strain was deposited with the Korean Culture Center of Microorganism on July 8, 2016. The accession number is KCCM 11861P.

Experimental Example 4: Optimum growth temperature of Lactobacillus saccharum 2-6-4 strain.

The final selected Lactobacillus casei strain 2-6-4 was cultured at 25, 30, 37, and 40 ℃, respectively, and the growth curve was obtained by absorbance measurement (600 nm) for 48 hours. The medium to be cultured was a 2 ° Brix ginseng concentrate containing 10% MRS.

FIG. 3 is a graph showing the results of Experimental Example 4. Specifically, FIG. 3 shows growth curves of Lactobacillus saccharum 2-6-4 strain at 2 ° Brix ginseng concentrate containing 10% MRS according to temperature.

As shown in Fig. 3, it was confirmed that the growth was remarkably lowered after 40 占 폚. Thus, it can be seen that the Lactobacillus saccharum 2-6-4 strain of the present invention is preferably fermented at a temperature of 25-37 ° C.

Experimental Example 5: Optimum Fermentation Time of Lactobacillus saccharum 2-6-4

The total ginsenosides and Rb1, Rh2, and Rg3 (Table 7) were obtained from the cultivated Lactobacillus saccharum 2-6-4 strain selected in Experimental Example 2 at various fermentation times, And the optimum growth time was confirmed. The medium to be cultured was a solid medium containing 20% ginseng powder. The ginseng powder-containing solid medium is a mixture of 20 wt% of ginseng powder and 80 wt% of water as described above.

Culture time (h) 0 24 48 72 Concentration of Rb1 (mg / g) 0.116 0.104 0.101 0.099 Concentration of Rg3 (mg / g) 0.085 0.095 0.104 0.103 Concentration of Rh2 (mg / g) 0.000 0.010 0.013 0.013 ** Ginsenoside concentration (mg / g) 1.102 0.962 1.206 1.186

(S, R), PPT (S, R), CK (R), Rc, Rb2, Rc, Rb2, Rb3, Rf1, Rd, , 16 kinds of Rh2

4 shows the results of Experimental Example 5. Specifically, FIG. 4 shows the growth curve of the strain 2-6-4 of Lactobacillus saccharum over 20% ginseng powder solid medium with time.

As shown in Fig. 4, the strain 2-6-4 of Lactobacillus saccharum was continuously increased after inoculation and remained at the highest level for 24 to 48 hours, and then rapidly decreased in viable cell count from 72 hours.

As a result, it was confirmed that the Lactobacillus saccharum strain 2-6-4 according to the present invention had a fermentation time of 24 to 72 hours, more preferably 24 to 48 hours.

However, the concentration of metabolite ginsenosides such as Rh2 or Rg3 was highest at 48 hours, and did not show any significant difference depending on the fermentation time. Therefore, considering all circumstances, fermentation for 72 hours, preferably 48 hours or more, is not cost effective.

Name of depository: Korea Microorganism Conservation Center (overseas)

Accession number: KCCM11861P

Checked on: 20160708

Claims (8)

Lactobacillus sakei 2-6-4 strain deposited with accession number KCCM11861P. The method according to claim 1,
Wherein the strain is grown on a solid medium containing ginseng powder. The method according to claim 1,
Wherein the ginseng powder-containing medium comprises 10 to 40 wt% of ginseng powder based on the total weight of the total solid medium. The method according to claim 1,
Wherein the strain is grown in a solid medium containing ginseng powder as a sole nutritional source. The method according to claim 1,
Wherein said strain produces Rh2 or Rg3 from solid medium containing ginseng powder. The method according to claim 1,
The optimum growth conditions of the strain are (a) a fermentation temperature of 25-37 ° C, (b) a pH of 4.1-4.5, and (c) a culturing time of 24-48 hours. A starter composition for the production of Rh2 or Rg3-fortified ginseng powder containing the strain Lactobacillus sakei 2-6-4 of claim 1 or a culture thereof. The ginseng powder obtained by solid-fermenting the ginseng powder with the strain 2-6-4 of Lactobacillus sakei of claim 1 as a Rh2 or Rg3-fortified ginseng powder.

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