KR20200145254A - Manufacturing method for a composition rich in ginsenosides Rg3, Rg5 and Rk1, which is excellent in improving brain function and cognitive function using ginseng - Google Patents

Abstract

The present invention relates to a method for preparing a composition rich in ginsenosides Rg3, Rg5, and Rk1 having excellent brain function and cognitive function improvement effects using ginseng, wherein the method comprises the following steps: (a) pulverizing fresh ginseng to obtain a pulverized fresh ginseng; (b) putting the pulverized fresh ginseng in a microwave-only container, closing a lid of the container, and performing heat treatment using microwave; (c) inoculating Lactobacillus plantarum MH16 in a medium, followed by culturing and sterilizing the same to prepare a treatment solution for treating the fresh ginseng completed in step (b); and (d) adding fresh ginseng heat-treated through the step (b) to the treatment solution prepared in the step (c), followed by aging the same.

Description

Manufacturing method for a composition rich in ginsenosides Rg3, Rg5 and Rk1, which is excellent in improving brain by using ginseng function and cognitive function using ginseng}

The present invention is a manufacturing method for increasing the content of ginsenosides Rg3, Rg5, and Rk1, which are excellent in improving brain function and cognitive function, from ginseng, and in detail, ginsenosides having excellent effect on improving brain function and cognitive function using ginseng It relates to a method for preparing a composition rich in Rg3, Rg5, and Rk1.

Ginseng from 4 to 6 years old is harvested on the farmland and unprocessed in its original form. It is also called fresh ginseng because it is not dried. Fresh ginseng usually contains about 75% moisture, so it is difficult to store it for more than one week as it is mined. Especially, it is prone to spoilage or damage during distribution, so it is processed in a moisture deodorant type for long-term preservation of ginseng.

The component involved in various pharmacological and physiological activities of ginseng is saponin, and it is named ginsenoside as a glyco-side isolated from ginseng to distinguish it from other plant saponins. By affecting the endocrine system, immune system, and metabolic system, it exerts various effects in regulating body functions. Until now, 38 kinds of ginsenosides have been separated from Korean ginseng (fresh ginseng) by purification and separation analysis technology, and their chemical structure has been revealed.The active ingredient of ginseng does not flow out and the saponin content and Other active ingredients increase, and in this process, new ginsenosides that ginseng does not have are produced.

Fermented ginseng is obtained by fermenting ginseng with specific microorganisms to improve the absorption rate of ginsenosides in the body and to standardize efficacy. The polymeric ginsenosides are not absorbed by the human body by themselves and have a small molecular weight by the intestinal microorganisms. Since it is decomposed and absorbed by senoside metabolites, a fermentation process is used as an effective method for obtaining high molecular weight ginsenosides as low-molecular ginsenosides. However, even in the conventional fermentation method, there is a problem in that the yield of conversion to low-molecular ginsenoside is low or a specific low-molecular ginsenoside cannot be obtained.

Red ginseng and black ginseng are typically used as processed ginseng to increase ginsenoside by processing ginseng. However, red ginseng and black ginseng are steamed and dried, resulting in a rapid decrease in weight and the production of benzopyrene, a carcinogen, and complex. It has disadvantages such as an increase in manufacturing cost due to the manufacturing process, and the temperature, pressure, time, etc., are irregularly controlled during the manufacture of red ginseng and black ginseng, causing problems such as the content of ingredients, color, and flavor. Therefore, in the process of increasing ginsenoside by processing ginseng, it is harmless to the human body, and standardized processing processes such as temperature, pressure, and time are very important to prevent loss of ginsenoside.

However, until now, efficient manufacturing methods such as heat treatment methods to increase the components of ginsenosides in ginseng and processed ginseng, microorganisms for fermentation and methods of use thereof, standardization of temperature, pressure, time, etc., in particular, increase memory and cause NMDA. Since it has excellent neurotoxin protection from neurotoxins, a manufacturing method that increases the content of ginsenosides Rg3, Rg5, and Rk1, which are excellent in improving brain function, learning ability or cognitive function, from ginseng has not been reported through specific experiments or studies. Did.

1. Republic of Korea Patent Publication 10-2013-0062628 2. Korean Patent Registration 10-0665087

Under this background, the technical task to be achieved by the present invention is to enhance memory and protect nerve cells from neurotoxins caused by NMDA, without using a conventional method for producing red or black ginseng and fermenting the prepared red or black ginseng using microorganisms. It is to provide a manufacturing method that increases the content of ginsenosides Rg3, Rg5, and Rk1, which are excellent in improving brain function, learning ability or cognitive function, from ginseng.

In order to achieve the above technical problem, the present invention

(a) pulverizing fresh ginseng to obtain a pulverized fresh ginseng;

(b) placing the pulverized fresh ginseng in a microwave-only container, closing the lid of the container, and performing heat treatment using a microwave;

(c) Lactobacillus inoculated with Bacillus Francisco taryum (Lactobacillus plantarum) MH16 in the medium, and then preparing a processing liquid for processing by a sterilization treatment and incubated complete the step (b) the fresh ginseng; And

(d) ginseno having excellent brain function and cognitive function improvement effect using ginseng, comprising the step of aging the heat-treated fresh ginseng through the (b) step to the treatment solution prepared in step (c). It provides a method for preparing a composition rich in side Rg3, Rg5, and Rk1.

In the manufacturing method according to the present invention as described above, the heat treatment using a microwave in step (b) may be performed in a microwave at a frequency of 500 to 5.000 MHz for 0.5 to 5 hours.

In the production process according to the invention as described above, the Lactobacillus Francisco taryum medium of step (c) to a culture (Lactobacillus plantarum) has MH16

0.5 to 1.5 (w/v)% peptone,

0.5 to 1.5 (w/v)% beef extract,

0.5 to 1.5 (w/v)% dipotassium hydrogen phosphate,

0.5 to 1.5 (w/v)% triammonium citrate,

0.5 to 1.5 (w/v)% magnesium sulfate heptahydrate,

7.0-20.0 (w/v)% juice-free,

7.0-20.0 (w/v)% pear juice,

Balance (w/v)% of distilled water

It may be to include.

Inoculated with a manufacturing method according to the invention, the step (c) culturing the Lactobacillus Francisco taryum (Lactobacillus plantarum) 1.0 part to 10.0 parts by weight per 100 parts by weight of the medium of MH16 as described above, and then 25 to 45 It may be made for 10 to 72 hours at ℃.

In the manufacturing method according to the present invention as described above, the aging in step (d) may be performed at 4 to 30° C. for 10 to 72 hours.

The present invention in order to achieve another technical problem,

It provides a pharmaceutical composition or a health food composition excellent in improving brain function and cognitive function, characterized in that it contains a composition rich in Rg3, Rg5, and Rk1 as an active ingredient prepared by the above-described manufacturing method. .

The present invention is a manufacturing method in which ginsenosides Rg3, Rg5, and Rk1, which are excellent in improving brain function and cognitive function, are significantly increased.The manufacturing method of the present invention is a heat treatment process and microorganisms compared to the conventional red ginseng to black ginseng manufacturing process. It is not a skilled process of directly fermenting ginseng using using, but through the aging process of treating ginseng heat-treated with this culture after cultivating microorganisms that have excellent enzyme production ability and rich organic acid in a medium. Efficient and easy to manage quality,

Below. The specific details for carrying out the present invention will be described in detail.

The present invention is a manufacturing method for increasing the content of ginsenosides Rg3, Rg5, Rk1 excellent in brain function and cognitive function improvement from ginseng,

(a) pulverizing fresh ginseng to obtain a pulverized fresh ginseng;

(b) placing the pulverized fresh ginseng in a microwave-only container, closing the lid of the container, and performing heat treatment using a microwave;

(c) Lactobacillus inoculated with Bacillus Francisco taryum (Lactobacillus plantarum) MH16 in the medium, and then preparing a processing liquid for processing by a sterilization treatment and incubated complete the step (b) the fresh ginseng; And

(d) adding fresh ginseng heat-treated through step (b) to the treatment solution prepared in step (c) and aging.

In the present invention described above, the fresh ginseng means that the harvested ginseng is not dried, and when frozen for storage, it may be thawed and used, or it may be used in a frozen state. Any ginseng harvested to produce red ginseng or black ginseng can be used without special restrictions.

In the present invention, the fresh ginseng is pulverized to obtain a fresh ginseng pulverized product. In this case, the pulverization method is not particularly limited, and the size of the pulverized product is also not particularly limited, but the finer the heat treatment effect described later may be increased.

In addition, in the present invention, it is preferable to pulverize the ginseng as it is without adding additional water while pulverizing the ginseng, and the moisture generated during pulverization is used as it is in the heat treatment to be described later.

Next, the prepared pulverized fresh ginseng containing moisture is placed in a microwave-only container, covered with a lid, and then heat-treated using microwaves.

Covering the lid of the container in the present invention is to achieve the effect of steaming the fresh ginseng during a separate water replenishment business as the vapor generated in the heat treatment process does not fly away, and the water is rehydrated from the lid and flows down to the bottom of the container.

When the microwave heat treatment according to the present invention passes through, the process of extracting hot water from the ginseng by steam supplemented from the outside in the process of steaming in order to manufacture red ginseng or black ginseng in the related art is performed, so that active ingredients are lost and the drying process is performed. There is no method to use it again because it is passed through, but in the present invention, since only the moisture contained in the fresh ginseng is used without supply of additional moisture or steam, there is no loss of active ingredients. In addition, the present invention is after completing the heat treatment process in the aging process described later. Since the remaining moisture is used as it is, there is no problem of loss.

The microwave heat treatment as described above is not particularly limited as long as it is widely known in the technical field to which the present invention belongs, but may be treated for 0.5 to 5 hours in a microwave at a frequency of 500 to 5.000 MHz, and the specific selection is fresh ginseng state, treatment It can be done appropriately in consideration of the weight of fresh ginseng to be taken.

Next, when a treatment liquid for aging is prepared and the heat-treated fresh ginseng is aged using this treatment liquid, the manufacturing method according to the present invention is completed.

The treatment fluid formulation is Lactobacillus Francisco taryum made by the MH16 (Lactobacillus plantarum) process, and then cultured after sterilization inoculated to the medium, fermentation is put into the water within the heat treatment is finished ginseng and its container in the prepared treatment solution as certain It is stored under conditions, and there is no particular limitation on the container for aging, but a container for rest such as pottery is preferable.

In the production process according to the invention as described above, the Lactobacillus Francisco taryum (Lactobacillus plantarum) medium of the (c) step of culturing the MH16 is job medium commonly used for culture of the Lactobacillus bacteria, such as MRS medium can, but the present invention will be described in detail through the embodiments to be described later in the selection process and the medium composition of the gradation is straight, and the Lactobacillus Francisco taryum (Lactobacillus plantarum) MH16 strain using a medium having a composition as shown below.

The medium used in the present invention is 1.0 (w/v)% peptone, 1.0 (w/v)% beef extract, 0.4 (w/v)% yeast extract, 2.0 (w/v)% glucose, 0.5 (w/v) )% sodium acetate trihydrate, 0.1 (w/v)% polysorbate 80 (also known as Tween 80), 0.2 (w/v)% dipotassium hydrogen phosphate, 0.2 (w/v)% triammonium citrate, 0.02 (w/v) % magnesium sulfate heptahydrate, 0.005 (w/v)% manganese sulfate tetrahydrate by changing the MRS medium

0.5 to 1.5 (w/v)% peptone,

0.5 to 1.5 (w/v)% beef extract,

0.5 to 1.5 (w/v)% dipotassium hydrogen phosphate,

0.5 to 1.5 (w/v)% triammonium citrate,

0.5 to 1.5 (w/v)% magnesium sulfate heptahydrate,

7.0-20.0 (w/v)% juice-free,

7.0-20.0 (w/v)% pear juice,

Balance (w/v)% of distilled water

It is preferable to include.

The cultivation of the step (c) of the present invention is not particularly limited, but after inoculating the Lactobacillus plantarum MH16 at 1.0 to 10.0 parts by weight based on 100 parts by weight of the medium, 10 to 72 at 25 to 45°C. It can be done over time.

After such cultivation is completed, it is sterilized to be used as a treatment solution for aging. The sterilization treatment is possible without particular limitation, as long as it is known in the art to which the present invention belongs, such as heating. In order to minimize it, it is preferable to use a filtration method capable of removing germs and making them sterile, and it can be said that a conventional method is to prepare a sterilized liquid through filtration using a membrane filter.

The final step, the aging of the present invention, is to put fresh ginseng that has been heat-treated in the prepared treatment liquid together with moisture in a container and leave it to stand under certain conditions.If it is aging in the process of preparing fermentation liquid and enzyme liquid, it can be used without particular limitation. And, in the present invention, the aging of step (d) may be performed at 4 to 30° C. for 10 to 72 hours.

The present invention can be heat-treated by using a microwave without any loss of the active ingredient, the newly configured using a dedicated medium enzymes and organic acid producing ability is excellent Lactobacillus Francisco taryum (Lactobacillus plantarum) enzyme by culturing the MH16 as described above Since it has excellent activity and can prepare a culture medium rich in organic acids, it is possible to prepare a composition rich in Rg3, Rg5, and Rk1 by aging using this.

Accordingly, the present invention is a manufacturing method in which ginsenosides Rg3, Rg5, and Rk1, which are excellent in improving brain function and cognitive function, are significantly increased, and the manufacturing method of the present invention comprises a heat treatment process and a heat treatment process compared to the conventional process of manufacturing red ginseng to black ginseng. Rather than a skilled process of directly fermenting ginseng using microorganisms, it is simple and standardized through the aging process of treating ginseng heat-treated with this culture after cultivating microorganisms that have excellent enzyme-producing ability and rich organic acids in a medium. Is efficient and easy for quality control.

Hereinafter, the present invention will be described in detail through Examples and Test Examples, and the present invention is not limited by Examples to be described later.

Example 1: strain selection

1. Strain culture and storage

The lactic acid strain used in the experiment was a strain obtained from the Fermentation Industry Microbiology Promotion Agency, the Korea Microbiological Conservation Center (KCCM), and the Korea Research Institute of Bioscience and Biotechnology Biological Resource Center (KCTC) as a comparative strain with the lactic acid bacteria isolated from the traditional fermentation sample.

The obtained strains were inoculated in MRS liquid medium by 1% and cultured for 48 hours at 37°C, and after the end of use, they were stored in a deep freezer at -70°C with 60% glycerol solution.

2. Evaluation of enzyme activity

In order to evaluate the enzyme activity and select the final strain, extracellular enzyme activity and enzyme activity using API ZYM were evaluated through the following method.

First, the measurement of the extracellular enzyme activity of the primary strain selected first was investigated for protease and beta glucosidase, and a well diffusion method using 1% skim milk (Difco) and 0.5% esculin (sigma) as respective substrates. It was measured using.

In addition, 18 enzyme activities other than alkaline phosphatase were tested using API ZYM kit (bioMerieux Co., Marcy-I'Etoile, France) to investigate the enzyme activity of the primary strain. The strain was cultured in MRS solid medium to recover the cells, washed three times in sterile water, and then suspended in sterile water to prepare a sample. After dissolving 500 ul of the suspension in a 5 mL suspension medium, turbidity was adjusted with 5-6 Mcfarland (bioMerieux Co.). After inoculating the turbidity-adjusted suspension solution into each cuple of the ZYM kit and incubating for 4 hours at 37°C, drop one drop of ZYM A and B reagents to help increase expression activity and dissolution into each cuple and react for 5 minutes. The degree of enzyme activity was investigated by observing the change in color. At this time, according to the degree of color change, the values ranged from 0 to 5, where 0 is a negative response, 5 (≥40 nmoles) is the maximum intensity response, and 4-1 is the middle of 30, 20, 10, and 5 nmoles, respectively. A value of 3 or more was determined as positive.

The extracellular enzyme activity and ZYM kit results of the comparative strain and the primary strain are shown in Tables 1 and 2 below.

First, as can be seen from Table 1, it was difficult to confirm the reaction of the comparative strain KCTC 3035, and KCCM 12116 showed the activity, but it was confirmed that the activity of MH16 was the most excellent, and SRCM 102226 was confirmed that the reaction did not appear. Could

In addition, as can be seen from Table 2, it was confirmed that the enzyme activity of MH16 was remarkably higher than that of the comparative strain in the results of examining the additional enzyme activity using the ZYM kit.

Specifically, it was observed that MH16 has 11 more enzyme activities in addition to β-glucosidase through the ZYM kit.

Esterase, esterase lipase, and peptide hydrolyzing enzymes such as Leucine arylamidase, Valine arylamidase, and Cystine arylamidase, which are involved in lipolysis, showed 5~30 nmol activity. However, the activity of β-glucosidase required for ginsenoside conversion was confirmed to be 30 nmol.

Accordingly, Lactobacillus plantarum MH16, which has excellent enzyme activity, was selected as the final strain.

3. Evaluation of organic acid content

The following method was evaluated in order to select the final strain by evaluating the organic acid content of the culture solution.

The organic acid was carried out using High Perfomrnace Liquid Chromatography (HPLC), adjusted to 50 mL of distilled water in 5 g of sample, stirred at 37° C. for 3 hours, passed through a 0.45 μm membrane filter, and used as a sample.

Free acid analysis conditions were performed by setting as shown in Table 3 below.

The total organic acid content was measured for each strain, and the results are shown in Table 4 below.

As can be seen in Table 4, Lactobacillus plantarum MH16, which has a high organic acid content, was selected as the final strain among the lactic acid strains, and Lactobacillus plantarum MH16, which was selected as the final strain.

The result of identification of the strain is as follows.

Example 2. Badge evaluation

Lactobacillus Francisco taryum medium of step (c) to a culture (Lactobacillus plantarum) has MH16

0.5 to 1.5 (w/v)% peptone,

0.5 to 1.5 (w/v)% beef extract,

0.5 to 1.5 (w/v)% dipotassium hydrogen phosphate,

0.5 to 1.5 (w/v)% triammonium citrate,

0.5 to 1.5 (w/v)% magnesium sulfate heptahydrate,

7.0-20.0 (w/v)% juice-free,

7.0-20.0 (w/v)% pear juice,

Balance (w/v)% of distilled water

In order to evaluate the effect of such a medium, it was cultured in MRS medium under the same conditions, and the enzyme activity and organic acid content of the culture medium were measured to evaluate the effect of the medium according to the present invention.

The medium according to the present invention is 1.0 (w/v)% peptone, 1.0 (w/v)% beef extract, 1.0 (w/v)% dipotassium hydrogen phosphate, 1.0 (w/v)% triammonium citrate, 1.0 (w/ v)% magnesium sulfate heptahydrate, 15 (w/v)% radish juice, 15 (w/v)% pear juice, and the balance (w/v)% distilled water.

Culture was carried out at 37°C for 48 hours by inoculating 1 part by weight with respect to 100 parts by weight of the medium. After the culture was completed, filtration was performed to remove culture strains and solids using a membrane filter. The total content was measured, and the measurement method was the same in Example 1, and the results are shown in Table 5 below.

When cultured in the medium according to the present invention as shown in Table 5, it was found that the enzyme activity and the total organic acid content of the culture medium were significantly increased.

Example 3: Preparation of the composition according to the invention and Rg3 , Rg5 , Rk1 Content comparison test

1. Preparation of the composition according to the invention

1 kg of 4-year-old fresh ginseng purchased from Jinan was pulverized using a grinder, and the pulverized product was put in a microwave-only container, closed, and then cooled in a 2.000 MHz frequency microwave for 2.5 hours.

Then preparing a composition according to the present invention Lactobacillus bacteria Francisco taryum (Lactobacillus plantarum) was used for a culture medium to prepare a culture solution of the MH16 in the second embodiment, placed after the heat treatment the cooling ginseng pulverized by aging at 10 ℃ for 30 hours, I did.

2. Rg3 , Rg5 , Rk1 Content comparison test

The content of Rg3, Rg5, and Rk1 was tested against the other compositions of the present invention. The contents were compared.

The content test was performed using HPLC, the content analysis conditions were as follows, and the results are shown in Table 6.

HPLC: YL9100 HPLC system (Youngrin, Korea)

Detector: ELSD ZAM 3000 (Schambeck SFD GmbH, Germany)

Column: Luna C18 5 um, 46 x 150 mm (Phenomenex, USA)

Solvent: A: Acetonitril: Water: 5% Acetic acid in water = 15: 80: 5(v/v), B: Acetonitril :Water = 80: 20(v/v)

Flow rate: 10 mL/min

0min (min) A:B = 100:0, 6min A:B = 70:30, 18min A:B = 50:50, 30min A:B = 0:[0087] 100, 37min A: B = 0:100

As can be seen from Table 6, it was confirmed that the contents of Rg3, Rg5, and Rk1 were significantly increased in the composition according to the present invention.

Claims (7)

(a) pulverizing fresh ginseng to obtain a pulverized fresh ginseng;
(b) putting the pulverized fresh ginseng in a microwave-only container, closing the lid of the container, and performing heat treatment using a microwave;
(c) Lactobacillus inoculated with Bacillus Francisco taryum (Lactobacillus plantarum) MH16 in the medium, and then preparing a processing liquid for processing by a sterilization treatment and incubated complete the step (b) the fresh ginseng; And
(d) ginseno having excellent brain function and cognitive function improvement effect using ginseng, comprising the step of aging the heat-treated fresh ginseng through the (b) step to the treatment solution prepared in step (c). A method for preparing a composition rich in side Rg3, Rg5, and Rk1.
The method of claim 1,
The heat treatment using the microwave in step (b) is ginsenoside Rg3 having excellent brain function and cognitive function improvement effect using ginseng, characterized in that the heat treatment using a microwave at a frequency of 500 to 5.000 MHz is processed for 0.5 to 5 hours, A method for preparing a composition rich in Rg5 and Rk1.
The method according to claim 1 or 2,
The Lactobacillus Francisco taryum medium of step (c) to a culture (Lactobacillus plantarum) has MH16
0.5 to 1.5 (w/v)% peptone,
0.5 to 1.5 (w/v)% beef extract,
0.5 to 1.5 (w/v)% dipotassium hydrogen phosphate,
0.5 to 1.5 (w/v)% triammonium citrate,
0.5 to 1.5 (w/v)% magnesium sulfate heptahydrate,
7.0 to 20.0 (w/v)% juice-free,
7.0-20.0 (w/v)% pear juice,
Balance (w/v)% of distilled water
A method for preparing a composition rich in ginsenosides Rg3, Rg5, and Rk1 having excellent brain function and cognitive function improvement effects using ginseng comprising a.
The method of claim 3,
Wherein the step (c) the ginseng which comprises culturing a 1.0 part to 10.0 parts by weight per 100 parts by weight of the medium inoculated with the Lactobacillus Francisco taryum (Lactobacillus plantarum) MH16 in the next 25 to 45 ℃ for 10 to 72 hours A method for preparing a composition rich in ginsenosides Rg3, Rg5, and Rk1, which are excellent in improving brain function and cognitive function by using.
The method according to claim 1 or 2,
A composition rich in ginsenosides Rg3, Rg5, and Rk1 having excellent brain function and cognitive function improvement effects using ginseng, characterized in that the aging of step (d) is performed at 4 to 30°C for 10 to 72 hours Manufacturing method.
A pharmaceutical composition excellent in improving brain function and cognitive function, characterized in that it contains a composition rich in Rg3, Rg5, and Rk1 as an active ingredient prepared by the manufacturing method of claim 1.
A health food composition having excellent brain function and cognitive function improvement effect, characterized in that it contains a composition rich in Rg3, Rg5, and Rk1 as an active ingredient prepared by the manufacturing method of claim 1.