KR100444369B1 - The memory increasing novel compound ginseno-jk from ginseng hairy roots and the manufacturing method of the ginseno-jk - Google Patents

Abstract

The present invention relates to a method for preparing a new composition ginseno-JK that enhances memory using ginseng hairy root in tissue culture and the use of the prepared ginseno-JK.

Ginseng hairy root obtained by tissue culture was extracted with 70% alcohol or ethanol, and then the extract was adsorbed on benzeneethylene resin, and eluted and removed the non-saponin component with 20% alcohol, and then the desired component was 60-80% alcohol. as the elution to prepare the composition, the components of the binary ginsenosides Rb₁, Rb₂, Rc, Rd, Re, and Rg₁, ginsenoside _{Rb 1, Rb 2, Rc,} Rd are each one-half or less of new content Rg₁ Provides the composition Ginseno-JK.

The manufactured ginseno-JK has the effect of enhancing memory and can be used as an agent for preventing and improving senile dementia symptoms, beverages, and foods.

Description

MEMORY INCREASING NOVEL COMPOUND GINSENO-JK FROM GINSENG HAIRY ROOTS AND THE MANUFACTURING METHOD OF THE GINSENO-JK}

The present invention relates to a method for preparing a novel composition ginseno-JK that enhances memory using ginseng hairy roots cultured and the use of the prepared ginseno-JK.

Ginseng has been reported to be effective in improving intellectual performance by improving learning function and memory in various memory learning experimental models using experimental animals.

The central choline nervous system plays a very important role in memory and learning donation, and senile dementia has been reported to be closely associated with the deterioration of cholinergic nervous system [Bartus RT, Dean RL et al., 1982, Science 217, 408-414; ibid; , 1985, Pharmacol. Biochem. Behav. 23; 125-135.

In vitro tests using synaptosomes isolated from cerebral cortex and brain hippocampus with high distribution of acetylcholine content, ginsenoside-Rg₁ and ginsenoside-Rb₁, the representative saponins of ginseng, It has the effect of promoting the release of acylcholine by increasing the absorption of choline, a synthetic precursor. The administration of ginsenoside-Rb₁ to rats increased choline uptake, suggesting that ginsenoside-Rb₁ could be classified as a memory-learning drug [Benishin CG, Lee R, Wang LCH and Liu HJ, 1991 , Pharmacology 42, 223-229; Bennishin CG, 1992, Neurochem INT. 21 (1): 1-5].

On the other hand, PT-based saponins and ginsenoside-Rg ₁ of Korean ginseng have been reported to have a similar effect to the brain function improving drug (Cetrophenoxine, N-methyl-D-aspartate) to increase central dopamine neuronal activity (Lee SC. You KH adn Yamamoto R, 1995, Korean J Ginseng Sci., 17 (3) 187-195).

In addition, four consecutive weeks of ginsenoside-RgR and ginsenoside-Rb₁ in weaning mice were found to be effective in promoting brain development (increase brain weight, cortical thickness, etc.) and memory learning behaviors. Zhang JT, Yang Y, and Qu Zw, Jiang XY, Liu M, (1993) Proc. 6th Int'I Ginseng Symp., 69-73].

Ginsenoside-Rg® is a Cyclic AMP phosphodiesterase in vitro studies using various brain tissues such as the cerebral cortex, striatum and hippocampus in young, and old rats. Inhibition of enzyme activity of phosphodiesterase significantly inhibits the cyclic AMP content that mediates the reaction between hormones and neurotransmitters as a second messenger, supporting the central effect of ginsenoside-RgR. Stancheva SL and Alova L (1993) Gen. Pharmacol. 24 (6), 1459-1462.

In addition, ginsenoside-Rg R has been reported to increase the plasma content of ACTH (adrenocorticotropin), a neuropeptide exhibiting anti- forgetful effect [Hiai S, Yokoyama Hand Yano S (1979), Endocr. Japon, 26, 661-665. Ginsenoside-Rg® was also found to restore the decreased corticosteroid receptor binding capacity of cerebral hippocampal cytosol in elderly rats (28 months) to the level of young rats (3 months).

As for ginsenoside-Re, an effect of reducing memory impaired by scopolamine was observed [Yoshimasa Y, Masaya H, Hideshi K, Eur. J. Pharmacol. 312 (1996) 149-151. The so-called radiation maze or Morris water maze test, which evaluates so-called spatial cognitive memory to find or recognize directions or places, is a very sensitive test method for cognitive deficits associated with cholinergic nervous system decline. (Robert K, McNamara and Ronald WS, 1992, Pharmacol. Biochem. Behav. 41, 529-538)

Meanwhile, Korean Patent Application No. 1996-020085 (Ginseng drink and its manufacturing method) of the applicant of the present invention discloses a ginseng beverage having a memory enhancing effect made by adding a vitamin, a functional material, an emulsifier, and a flavor supplement to ginseng extract. However, the components are various, the technical content is different from the present invention.

In addition, Korean Patent Application No. 1998-0016211 (Method of manufacturing saponin-containing ginseng extract) of the present inventors adsorbs the raw ginseng saponin extract to an artificial synthetic resin such as benzene ethylene, and then elutes and removes the non-saponin component with dilute alcohol. In the present invention, a method of eluting saponin high content extract with a dark spirit is shown, but since the raw material is common root, and the concentration of alcohol, which is an extraction solvent, is higher than the present invention, the purpose of the present invention is different from that of the present invention, It is different and there is no mention of pharmacological effects.

In addition, Korean patent application 1994-028987 (Method of manufacturing soft brain capsules) is added to docosa nucleated saensa (DNA) fish oil to add ginseng extract, soybean lecithin, tocopherol, soft capsule for brain activation and prevention of senile dementia. It is related to, but the technical configuration is different from the present invention.

In addition, Korean Patent Application No. 1998-0017429 (a herbal composition for preventing and treating dementia) relates to a hydrothermal or alcohol extract of a composition in which ginseng, male, cheonma, and changchangpo are mixed, but the technical configuration is different from the present invention. .

The present invention facilitates the supply of raw materials by using ginseng hairy roots cultured in large tissues, and extracts ginsenoside compositions at specific concentrations and conditions by using alcohol as an extraction solvent, which is harmless to the human body, thereby having an excellent memory enhancing effect. To provide.

1 is a method for producing ginseno-JK of the present invention

2 is a memory test results of normal rats

Figure 3 is a short-term spatial cognitive memory test results

Fig. 4 shows the daily effect of ginseno-JK by doses on short-term spatial memory improvement.

Figure 5 Effect of Day 2 by Ginseno-JK Dose on Short-term Spatial Memory Improvement

The present invention provides a ginsenoside composition excellent in memory enhancing effect from tissue cultured ginseng hairy root.

In the ginsenoside composition of the present invention, the ginseng hairy root obtained by tissue culture was extracted with 70% alcohol or ethanol, and then the extract was adsorbed onto benzeneethylene resin, and the non-saponin component was eluted and removed. A composition prepared by eluting a desired component with -80% alcohol, wherein the constituents are ginsenosides Rb ', Rb2, Rc, Rd, Re, and Rg', and the content of each ginsenoside Rb ', Rb2, Rc, and Rd is Rg'. As a novel composition of less than 1/2, we named it Ginseno-JK.

The ginsenoside composition prepared by the same method as the ginsenoside-JK preparation using ginseng root A as a raw material was named ginsenoside-JA, and its component content was analyzed and the memory enhancing effect was compared. .

The components of ginsenoside-JA are ginsenosides Rb ', Rb2, Rc, Rd, Re and Rg', which are the same as ginsenoside-JK, but the content of ginsenoside Rb 'is 1.45 times higher than that of Rg'.

The ginsenoside composition prepared by the same method as in the preparation of ginsenoside-JK, using ginseng root B as a raw material, was named ginsenoside-JB, and its component content was analyzed and a comparison experiment of memory enhancement effect was performed. It was.

The components of ginsenoside-JB were ginsenosides Rb ', Rb2, Rc, Rd, Re, and Rg', which were the same as ginsenoside-JK, but the content of ginsenoside Rb 'was 3.61 times higher than that of Rg'.

Animal experiments were conducted to compare and confirm the memory enhancing effects of the prepared ginseno-JA and ginseno-JB and ginseno-JK.

In the Morris water mage experiment using rats, the short-term spatial cognitive memory improvement test showed that the ginseno-JK group had a much higher memory effect than the ginseno-JA group or the ginseno-JB group.

The memory-improving effect of ginseno-JK in the rats was high at 4-10 mg / kg.

In addition, the ginsenoside composition ginseno-JK of this invention can also be made by extracting ginsenoside component from general ginseng root and combining them suitably.

In addition, as the adsorptive resin used for extraction, ethyl benzene ethylene, 2-methyl benzene ethylene, 2-methyl propylenic acid ester resin can also be used.

Hereinafter, the present invention will be described in detail with reference to Examples, Comparative Examples, and animal experiments, but the present invention is not limited thereto.

Example 1 Preparation of Ginsenoside Composition from Ginseng Hairy Roots

Tissue cultured ginseng hairy root was obtained and 100 g was used as a raw material.

70% alcohol was added to the ginseng hairy root 7 times the weight of the sample, and the resultant was repeatedly extracted three times at 80 ° C. for 4 hours and filtered.

The alcohol extract was concentrated under reduced pressure to remove alcohol, diluted with water, and slowly passed through benzeneethylene resin to adsorb saponin.

After washing with 10 times the weight of the resin, and then eluted with 20% alcohol of 5 times the weight of the resin to remove the non-saponin components.

Ginsenoside composition was obtained by eluting with 10% of 60 times the alcohol content of the resin weight.

The ginsenoside composition obtained was subjected to TLC qualitative analysis.

The content of each component of the obtained ginsenoside composition was analyzed by HPLC.

As a result of analysis, the components and composition ratios of the ginsenoside composition are shown in Table 1.

Ginsenoside Rb ₁ : Rg ₁ = 1: 3.12.

Table 1. Composition content of each ginsenoside composition

[Unit: dry extract (%)]

Rb ₁ Rb ₂ Rc Rd Re Rg ₁ total 0.58 0.15 0.15 0.36 2.52 1.81 5.57 Rb ₁ : Rg ₁ = 1: 3.12,

Example 2 Preparation of Ginsenoside Composition from Ginseng Hairy Roots

Tissue cultured ginseng hairy root was obtained and 100 g was used as a raw material.

It was carried out in the same manner as in Example 1 below, to obtain a ginsenoside composition with an extraction alcohol concentration of 80% to perform a TLC and HPLC analysis.

The results are shown in Table 2, and ginsenosides Rb ₁ : Rg ₁ = 1: 2.34.

Table 2. Composition content of each ginsenoside composition

[Unit: dry extract (%)]

Rb ₁ Rb ₂ Rc Rd Re Rg ₁ total 0.7 0.25 0.24 0.46 2.67 1.64 5.26 Rb ₁ : Rg ₁ = 1: 2.34

A ginsenoside composition prepared in the same manner as in Examples 1 and 2 by repeating several manufacturing experiments and animal experiments in addition to Examples 1 and 2, and the components are ginsenosides Rb ', Rb2, Rc, Rd, Re, The inventors named the present invention a new invention composition wherein Rg 'is a content of ginsenosides Rb', Rb2, Rc, and Rd equal to or less than 1/2 of Rg '.

Comparative Example 1 Preparation of Ginsenoside Composition Ginseno-JA from General Root A

A 6-year-old ginseng on the market was purchased and 100 g was used as a raw material.

Hereinafter, a ginsenoside composition was obtained in the same manner as in Example 1, named Ginseno-JA, and TLC and HPLC analysis were performed.

As a result of analysis, the composition and composition ratio of Ginseno-JA were as shown in Table 3.

Ginsenoside Rb ₁ : Rg ₁ = 1.45: 1.

Table 3. Ginsenoside Content of Ginseno-JA

[Unit: dry extract (%)]

Rb ₁ Rb ₂ Rc Rd Re Rg ₁ total 1.49 0.70 0.74 0.30 0.82 1.03 5.08 Rb ₁ : Rg ₁ = 1.45: 1

<Comparative Example 2> Preparation of ginsenoside composition ginseno-JB from common muscle B

A 6-year-old ginseng on the market was purchased, and the ginseng root body was removed, and a small root was collected and 100 g was used as a raw material.

Hereinafter, a ginsenoside composition was prepared in the same manner as in Example 1, named Ginseno-JB, and TLC and HPLC analysis were performed.

The components and composition ratios of ginseno-JB are shown in Table 4.

Ginsenoside Rb ₁ : Rg ₁ = 3.61: 1.

Table 4. Ginsenoside content of ginseno-JB

[Unit: dry extract (%)]

Rb ₁ Rb ₂ Rc Rd Re Rg ₁ total 4.76 2.63 2.72 1.13 2.71 1.32 15.27 Rb ₁ : Rg ₁ = 3.61: 1

Experimental Example

Animal experiments were performed to compare and confirm the memory enhancing effects of the composition ginseno-JK of the present invention prepared in the above example and the ginseno-JA and ginseno-JB prepared in the comparative example.

1. Experimental Materials and Animals

end. Experimental material

Physiological saline was used as the control ginseno-JK, ginseno-JA, ginseno-JB and the control prepared in Example 1, Comparative Example 1, Comparative Example 2 described above.

I. Laboratory animals

Sparague Dawley rat males (180-200 g) were assigned to each group of 10-15 animals and each experiment was performed.

All. Experiment apparatus

1) Morris water maze apparatus was used.

2. Experiment and Effect Comparison

(1) the test of memory generation after learning of rats

Each sample was tested 30 minutes after intraperitoneal administration.

The Probe test was conducted after eight sessions of training in the Morris water maze.

By removing the flat foam and performing a swimming test, the retention time of the rat in the 4/1 quadrant of the flat foam was measured to determine whether the memory for the D region was generated.

The results are shown in FIG. 2 and showed normal memory production in the control group (physiological saline), ginseno-JK (10 mg / kg), ginseno-JA (10 mg / kg), and ginseno-JB (10 mg / kg). .

(2) Short-term spatial cognitive memory test

Rats who had completed memory learning training were transferred to their original position (D) and moved to another position (B) to test their swimming time to find their new target.

As a result of the test, the improvement of memory was significant only in the group administered with ginseno-JK as shown in FIG. 3.

(3) Short-term spatial memory improvement effect test by Ginseno-JK dose

Different doses of ginseno-JK were tested to improve memory capacity.

The first day results are shown in FIG. 4, and the second day results are shown in FIG. 5.

Ginseno-JK dose-dependently improved memory effect from 4mg / kg, and the effective concentration of ginseno-JK memory effect was high at 4 ~ 10mg / kg.

The ginseng hairy root obtained by tissue culture provides a large amount of the composition ginseno-JK with an excellent memory enhancing effect.

The prepared ginseno-JK has an excellent effect of enhancing memory and can be used as an agent for preventing and improving senile dementia symptoms, beverages, and foods.

Claims (3)

A composition prepared by extracting ginseng hairy root from tissue culture, the constituents of which are ginsenosides Rb ₁ , Rb₂, Rc, Rd, Re, and Rg₁, and each of ginsenosides Rb₁, Rb₂, Rc, and Rd is 1 in Rg₁. Memory Enhancing Composition Ginseno-JK that is less than / 2. After ginseng hairy roots were cultured to obtain a large amount of cultured roots, the obtained ginseng hairy roots were extracted with 70% alcohol or ethanol, and the extract was selected from benzene ethylene resin, ethyl benzene ethylene resin, or 2-methyl benzene ethylene resin. It was adsorbed on one kind of adsorption resin and eluted and removed the non-saponin component by 20% alcohol, and then eluted by 60-80% alcohol.The components were ginsenosides Rb₁, Rb₂, Rc, Rd, Re, Rg₁. Cenoside Rb ', Rb2, Rc, Rd content of each of the memory enhancing composition ginseno-JK of less than 1/2 of Rg'. delete