KR100711954B1 - Extract of tissue cultured mountain ginseng(panax schinseng ness) having high content saponin - Google Patents

Abstract

The present invention relates to a ginseng cultured root extract containing a large amount of saponin, and more particularly, inoculated in wild ginseng roots grown by tissue culture in a medium and cultured in a bioreactor, but depleted of nitrogen in the medium around 10 days after harvesting. Next, the ginseng culture root powder obtained by culturing is extracted by a warm extraction method or a reflux extraction method and relates to a ginseng culture root extract having high saponin content.

The ginseng cultured root extract according to the present invention is a functional food material containing a large amount of saponin, which is a major pharmacologically active ingredient, and not only various health benefits derived from wild ginseng saponin, but also an effect that can effectively remove active oxygen species that occur a lot in modern people Was evaluated.

Wild ginseng, culture root, extract, saponin

Description

Extract of Tissue Cultured Mountain Ginseng (Panax schinseng NESS) Having High Content Saponin}

1 is a chromatogram showing the irradiated ponson characteristics of wild ginseng cultured root extract of the present invention.

Figure 2 is a chromatogram of saponin content control of wild ginseng culture root by adjusting the culture process of wild ginseng culture root extract of the present invention.

Figure 3 is a graph showing the DPPH scavenging activity of wild ginseng cultured root extract of the present invention.

The present invention relates to a ginseng cultured root extract containing a large amount of saponin, and more particularly, inoculated in wild ginseng roots grown by tissue culture in a medium and cultured in a bioreactor, but depleted of nitrogen in the medium around 10 days after harvesting. Next, the ginseng culture root powder obtained by culturing is extracted by a warm extraction method or a reflux extraction method and relates to a ginseng culture root extract having a high saponin content.
Panax ginseng is a perennial herb belonging to the Araliaceae family, and its root is called Ginseng Radix in oriental medicine. Tonic, Gangjeong, Hematopoietic, Warmth, It is known to have the effects of healthy stomach, fatigue, mental stability, sedation (Korea Ginseng Efficacy Summary, Korea Ginseng Research Institute, Daesung Printing Co., p143, 1985). The main active ingredients of ginseng include saponins, sapongenins, polyacetylenes, pyrazine derivatives, maltol, and the like, and ginseng saponins are based on the chemical structure of protopanaxadiol and protopanaxatriol. It is divided into system and oleanonane-based saponins. To date, 19, 10 and 1 compounds have been separated and purified, and diol and triol are known to have different pharmacological effects in the body. Ginseng saponins, unlike other plant saponins, are not toxic and do not show hemolytic activity below 0.001% (Taiki, K. et al., P43, 1981). al., J. Pharmacol. 33: 245, 1972), protein synthesis (Yokozawa, T. and Oura, H., J. Nat. Prod. 53: 1514, 1990), corticosteroid secretion (Pearec) , PT et al., Endocrinol. 29: 567, 1982) and immunopotentiation (Kim, YS et al., Korean J. Ginseng Sci. 15: 13, 1991).
On the other hand, wild ginseng ( Panax schinseng NESS) is a ginseng naturally occurring in the mountains, indications and benefits are similar to ginseng, but its medicinal effect is excellent, and the representative effect is the homeostasis maintenance function of the body control function. Anti-fatigue and antistress action, blood pressure control action, anticancer action, arteriosclerosis and hypertension prevention, gastrointestinal function enhancement, immune function enhancement, antiviral action and the like have been reported based on this action.
Ginseng or wild ginseng having various pharmacological effects as described above is usefully used in medicines, foods, and cosmetics, but natural medicinal ginseng or wild ginseng is not economical because it is impossible to mass produce. Therefore, studies have been conducted to mass-reduce non-expensive medicinal ginseng and wild ginseng using plant tissue culture technology, and Korean Patent Application No. 2001-0003285 by the present inventors discloses mass growth of ginseng, camphor ginseng, and wild ginseng root muscle by tissue culture. The method is disclosed. In addition, the inventors of the present invention, in order to improve the problem that the mass of the negative root obtained through tissue culture is possible to mass production, but the saponin content is lower than that of natural ginseng, and the ratio of diol and triol is significantly different than that of natural ginseng, tissue culture The method for improving the saponin content of ginseng, camphor ginseng, and wild ginseng roots by the Korean Patent Application No. 2001-0003284 has been disclosed.

The present invention has been made in accordance with the requirements as described above, an object of the present invention is to provide a ginseng culture root extract with high saponin content improved functionality according to the saponin properties as a health functional food material.
Another object of the present invention is to provide a ginseng cultured root extract with high saponin content having excellent health functional food characteristics as a health functional food material.

The present invention for achieving the above object relates to a wild ginseng culture root extract containing a large amount of saponin, more specifically, inoculated into the medium and cultured in a bioreactor inoculated wild ginseng root roots grown by tissue culture, harvesting 10 days The ginseng culture root powder obtained by depleting nitrogen in the foreground medium and then cultivating is extracted by a warm extraction method or a reflux extraction method, and relates to a ginseng culture root extract having a high saponin content.
The wild ginseng cultured root powder of the present invention is yellow to yellowish brown powder, the average content of irradiated saponin is 6.0 ~ 8.0% by weight, the total saponin content is 2-3% by weight, and the dry content is 70-80% by weight. It is done.
In addition, the wild ginseng cultured powder is characterized in that the average acidity (pH) is 4.0 ~ 6.0.
In addition, the heavy metal standard of the wild ginseng cultured powder is characterized in that less than 3 ppm.
In addition, the evaporation residue of the wild ginseng cultured root powder is characterized in that 4 to 6% by weight.
Hereinafter, the configuration of the present invention will be described in more detail with reference to preferred embodiments, but these embodiments are only for illustrating the present invention specifically, the scope of the present invention is not limited by these examples, The scope shall be defined only by what is stated in the claims.
Example 1 Preparation of Wild Ginseng Cultured Root Extract
The 100-year-old wild ginseng harvested from Odaesan, Gangwon-do on October 25, 2002 was separated into brains, roots, and roots, sterilized and sterilized, and then slices were made into 2 ~ 3 mm2, followed by 2,4-dichlorophenoxy acetic acid. (2,4-dichlorophenoxyacetic acid), pichloram, and NAA (naphthalemeacetic acid) were added in amounts of 1-10 mg / l, and then inoculated in MS medium to induce callus. Induced callus was grown in MS medium to which 1.0 mg / L of 2,4-dichlorophenoxy acetic acid was added as a growth regulator, and then subcultured every two weeks to MS medium to which IBA was added in an amount of 5.0 mg / L. Transferred to form a root muscle. The formed root muscle is cut into 0.5 ~ 1 cm size and inoculated into MS medium with mineral concentration 1/2, pH 5.7, and sugar concentration 3% using a 10 ~ 20 L bioreactor. Proliferation was then used as seed for large-volume bioreactor culture. The seed was cut to a size of 1 to 2 cm and inoculated in a large-capacity air flotation balloon bioreactor of 200, 500, 1,000, 10,000 L scale. At this time, the medium composition was a liquid MS medium containing 5% sugar (Duchefa, Netherlands), and as a growth regulator was added 1.0-5.0 mg / l 3-indolbutyl acid (IBA). The initial pH of the medium was maintained at 5.8-6.0, the culture temperature was 22 ± 2 ° C, and the air injection amount was 0.1-0.3 vvm (volume / volume / min).
After 2 weeks of culture, the new root muscle began to form in the root muscle section, and the maximum growth occurred at 35-40 days of culture. After this period, nitrogen was depleted in medium to increase the total saponin content, and then cultured for 10 more days. Cultured roots derived from the brain head, main muscle, and root muscle were harvested and named as CBN-1, CBN-2, and CBN-3, respectively. The harvested sample was washed two to three times with tap water, dried at 50 ° C. for 24 hours, and then crushed with a grinder to obtain a powder raw material.
Example 2 Health Functional Food Characteristics of Tissue Cultured Wild Ginseng Powder
Tissue cultured wild ginseng powder was recognized for its quality as a health functional food. Since there was no reference to the health functional food process and process of cultured wild ginseng domestically and internationally, first of all, cosmetics were manufactured from cultured ginseng as raw materials. The permit was assessed for the health functional food characteristics by setting the test items, such as properties, identification, total saponin content, acidity, purity, evaporation residue, and drying loss, which were obtained from the KFDA.
The culture root material obtained in Example 1 was extracted using a warm extraction method. That is, 10 kg of wild ginseng root cultured from tissue culture was extracted at a temperature of 80-100 ° C for 6-8 ° C. At this time, the ratio of raw material and hot water was 1:20. Ethanol was removed with Hg and filtered to obtain about 10 kg of wild ginseng root extract.
The test items were examined as follows for the wild ginseng cultured root powder and extract.
O The appearance was observed by naked eye and judged as smell.
⑵ For the saponin test, 2 ml of acetic anhydride was added to 0.2 g of raw material, and heated for 2 minutes in a bath. Then, 0.5 ml of sulfuric acid was added to 1 ml of the filtrate. To confirm the amino acid, 1 ml of ninhydrin solution was added to 2 ml of the raw material, and the mixture was shaken and left to stand in boiling boiling water for 5 to 10 minutes. To confirm triterpenoids, 10 ml of the above extract was concentrated under reduced pressure, 10 ml of chloroform was added to the residue, followed by shaking. The mixture was filtered, and 0.5 ml of anhydrous acetic acid was added to the residue. Add to the surface.
확인 To confirm the saccharide, 2 ~ 3 drops of 5% α-naphthol ethanol TS was added to 0.5 ml, mixed, and added to 2 ml of sulfuric acid.
⑷ The total saponin content was analyzed according to the standard method of food industry.
⑸ Acidity (pH) was determined by measuring 5 g of the raw material and adding 50 ml of freshly distilled water after cooling to room temperature.
⑹ Purity test was carried out in accordance with the second method by taking 1 g of the raw material to check the heavy metal content. 2.0 ml of lead standard solution was added to the comparative solution (20 ppm or less) .In order to detect arsenic, 1 g of raw material was taken to make a sample solution according to the third method, and the test was carried out using the apparatus A. ).
⑺ evaporation residue test was to take 10 ㎖ of the raw material and evaporated at 105 ℃ in water bath for 6 hours and confirmed to be 4 ~ 6% by weight.
⑻ Loss on drying was maintained at 80% when 5g of raw material was heated at 105 ℃ for 4 hours to reach 70 ℃.
The characteristics of the tissue culture wild ginseng powder as a health functional food confirmed using the test method described above are shown in Table 1.
TABLE 1
Test Items Test Methods medium Constellation 4. 1) Yellowish brown powdery powder with an unusual smell Test ① 4. 2) Item ① Maroon confirmed ② 4. 2) Clause ② Purple checked ③ 4. 2) Item ③ Maroon confirmed ④ 4. 1) Clause ④ Fuchsia confirmed Total Saponin Content (%) 4. 3) 7.0 pH 4. 4) 5.3 Purity test Pb 4.5) Item ① 1.0 ppm As 4. 5) Clause ② 2 ppm or less Evaporation residue 4. 6) 5.1 wt% Dry content 4. 7) 73.2% by weight
Appearance is yellow to yellow brown powder with peculiar smell. As a result of confirmation test The interface is reddish brown (saponins); I. Solution is purple (amino acid); All. The interface is reddish brown (triterpenoids); la. The interface is reddish violet (saccharide). The irradiated phosphonine content was set to 6.0 to 8.0 wt% of the average irradiated phosphonine in the raw material, and the pH (pH) was set to 4.0 to 6.0 as the average pH of the sample. Purity test showed that the minimum value of the test value was 0.6 ppm, the maximum value was 1.6 ppm for the heavy metals, and the reference level of the heavy metals of this raw material was less than 3 ppm, and arsenic was set to 2 ppm or less for this raw material. Evaporation residue is a tissue culture production three powder extracted raw material with hot water of 80 ~ 100 ℃, the average of the evaporation residue of the sample was set to 4 to 6% by weight as the average of 5.1% by weight. The drying loss of the sample of tissue culture ginseng powder was set as the average of 73.2% by weight, 70 to 80% by weight.
Example 3 Ginsenoside Analysis of Wild Ginseng Cultured Root Extract
The characteristics of ginsenosides in wild ginseng cultured roots were compared with those of cultivated ginseng and red ginseng according to the standard analysis method of Food Code and functional food standard. Briefly, 2 g of dry cultured wild ginseng powder was extracted twice with 40 ml of 80% alcohol for 1 hour at 60 ° C., filtered and distilled under reduced pressure, and then dissolved in 50 ml of distilled water for HPLC. HPLC (Waters 2690 separation module; The analysis was performed using a Waters 996 photodiode array detector (Waters millenium 2010 chromatography manager). At this time, by adjusting the ratio of water and acetonitrile (acetonitrile) was initially analyzed at 86:14, after increasing the content of acetonitrile.
Looking at the ginsenoside characteristics of the ginseng culture root extract in Figures 1 and 2, the ginseng and red ginseng irradiated with 4 ~ 5% by weight, the wild ginseng cultured 6 ~ 8% by weight, especially the content of Rd The content was found to be high (FIG. 1). In total saponin analysis using wild ginseng root HPLC, wild ginseng root contains 2-3% by weight, more than 1-2% by weight of ginsenosides of cultivated ginseng and red ginseng. 2). In addition, the wild ginseng culture root can increase the content of physiologically active substances by physicochemical stress during the culture process, and has the advantage of controlling the content ratio of saponin diol and triol.
Example 4 Antioxidant Effect of Wild Ginseng Cultured Root Extract
The culture root material, cultivated ginseng and red ginseng obtained in Example 1 were extracted by reflux cooling. That is, 40 g of cultured wild ginseng root ginseng was placed in a round flask with a reflux condenser tube, 280 ml of 70% fermented alcohol was added thereto, and reflux extraction was performed at 70 ° C. for 2 hours. After extracting twice more with fermentation alcohol of 30%, 10% by the same method as above, and filtered with filter paper (Advantitic, Tokyo, Japan) and concentrated under reduced pressure to obtain a wild ginseng root extract. The extract was treated with concentrations of 0.05, 0.1, 0.25, 0.5, 1.0, 2.0 to determine the scavenging ability of reactive oxygen species, ie, electron donating ability (EDA%) for DPPH (MS Blois, Nature). 26: 1199, 1958).
2, the sulfated effect of DPPH scavenging activity is significantly higher in wild ginseng cultured root extracts CBN-2 and CBN-3, and the wild ginseng cultured root effectively removes free radicals that occur in modern people due to environmental pollution or stress. It was evaluated that there was an effect that could be eliminated.

As described above, the wild ginseng cultured root extract according to the present invention is a functional food material containing a large amount of saponin, which is a major pharmacologically active ingredient, and not only various health benefits derived from wild ginseng saponin, but also active oxygen species, which are particularly produced in modern people. It was evaluated to have an effect that can be effectively removed.

Claims (4)

The wild ginseng root roots grown by tissue culture were inoculated into the culture medium and cultured in a bioreactor, and the wild ginseng cultured root powder obtained by culturing the nitrogen in the medium around 10 days before harvesting was extracted by a warm extraction method or a reflux extraction method. In this high ginseng cultured root extract, The wild ginseng cultured powder is yellow to yellowish brown powder, the average content of the irradiated saponin 6.0 ~ 8.0 wt%, the total saponin content is 2-3 wt%, characterized in that the dry content is 70 ~ 80 wt% Ginseng culture root extract with high content of ginsenoside Rd and high saponin content. The wild ginseng cultured root extract of claim 1, wherein the content of ginsenoside Rd is high and saponin content is characterized in that the average acidity (pH) of the wild ginseng cultured root powder is 4.0 to 6.0. The ginseng cultured root extract according to claim 1 or 2, wherein the content of ginsenoside Rd is high and saponin content is characterized in that the heavy metal standard of the wild ginseng cultured root powder is less than 3 ppm. According to claim 3, The evaporation residue of the wild ginseng cultured root powder is characterized in that the content of ginsenoside Rd is high and saponin content of high ginseng culture, characterized in that 4 ~ 6% by weight.

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