KR20110052940A - A process for preparing an extract fraction enriched with ginsenoside rg3 from ginseng radix - Google Patents

Abstract

PURPOSE: A method for preparing ginsenoside Rg3-enhanced extract fraction is provided to ensure high yield and obtain ginseng extract containing pure Rg3. CONSTITUTION: A method for preparing ginseng extract fraction with enhanced ginsenoside Rg3 comprises: a step of reacting ginsenoside Rb1-containing spirit extract and mixture of water and acetic acid to enhance ginsenoside Rg3; a step of applying the ginseng spirit extract to an adsorbing resin; a step of removing ingredients which is not adsorbed by elution; a step of adding 30-50v/v% of spirit to the adsorbing resin to remove triol ginsenosides; and a step of adding 50-100v/v% of spirit to adsorbing resin.

Description

A process for preparing an extract fraction enriched with ginsenoside Rg3 from Ginseng Radix}

The present invention relates to a method for preparing an extract fraction in which a specific ingredient is fortified from ginseng, and more specifically, an extract enhanced with a ginsenoside Rg3 component known to have vasorelaxation and anticancer activity among saponins present in ginseng. A method for preparing a fraction from ginseng.

Ginseng (Panax ginseng CA Mayer) is botanically belonging to the genus Araliaceae ginseng, which has been used for medicinal purposes in China since BC, and has been used for trade and medicinal use in Korea since the Three Kingdoms. It is widely used as a herbal medicine or health food.

Ginseng contains about 3-6% of ginsenoside-like saponin-like substance as a species-specific component, which is a representative physiologically active substance of ginseng, depending on the structure of panaxadiol (PD). ), And panaxhatriol (PT) and oleanane (Oleanane) divided into about 33 species have been reported. The content of saponins contained in ginseng is different for each part, and in the case of specific saponins, the content of the saponins is so small that the study of individual ginsenosides is insufficient.

 PD-based saponins and PT-based saponins have different functions in the body. PD-based saponins, represented by ginsenoside Rb1, have been shown to exhibit inhibitory effects on the central nervous system, resulting in mental stability, neuroleptic relaxation, analgesia, anticonvulsions, lowering blood pressure, papaverine and the like. In contrast, PT-based saponins, represented by ginsenoside Rg1, are known to have an anti-fatigue effect by acting excitably on the central nerve. In addition, ginsenoside Rg3 is present in a very small amount in white ginseng and 50 times more than red ginseng, and ginsenoside Rg3 has a vasorelaxant effect [J. Nat. Prod. 63, 1702 (2000)], inhibiting platelet aggregation [Biol. Pharm. Bull. 21, 79 (1998), Korean J. Ginseng Sci. 21, 132 (1997)], neuroprotective neurons [J. Neuroscience Res. 53, 426 (1998), Neuro Report 9, 226 (1998), and also reported to have anticancer activity [Jpn. J. cancer Res. 87, 357 (1996), J. cancer Res. Clin. Oncol. 120, 24 (1993), Anticancer Res. 17, 1067 (1997), Cancer Letters 150, 41 (2000), Dietary Anticarcinogenesis and Antimutagenesis 274 (2000).

The general manufacturing process of red ginseng is to wash the fresh ginseng with water, put it in a regular container, and use heated steam, steam it for a certain time according to its size (蒸 蔘), and then dry it by hot air first, and then use solar heat or other methods. It consists of drying until 12.5 to 13.5% of moisture, and the removal of the malonyl group of malonyl ginsenoside, the deglycosylation of the glycosyl group and the isomerization of hydroxy group through the process Reaction occurs, which creates various saponin components specific to red ginseng. However, the manufacturing process of the red ginseng is an inefficient process that consumes a lot of time and money, and has become a factor that inhibits the popularization of red ginseng.

Therefore, if ginsenoside Rg3-enhanced extract can be obtained from ginseng, it can be developed as a medicinal or health food with enhanced pharmacological activity of Rg3, and in a more economical and convenient way without the cumbersome operation of red ginseng. Ginsenoside Rg 3 may be used and would be preferred.

Korean Patent Laid-Open No. 1999-0000458 discloses a method for preparing ginsenoside Rg3 by reacting diol-type saponin with an enzyme lactase derived from Aspergillus or Penicillium in an aqueous solvent.

In addition, Korean Patent Laid-Open Publication No. 2008-0063049 discloses ginseng or red ginseng-specific ginsenosides by inoculating ginseng or red ginseng inoculated with lactic acid bacteria and finally heat-treating the culture to obtain ginseng or red ginseng with enhanced ginsenoside Rg3. A method of converting Rg3 is disclosed.

In addition, Korean Patent Registration 0444394 discloses a method for preparing a saponin-containing extract by adsorption using an adsorptive resin, but does not disclose a method for obtaining an extract fraction enhanced with a specific saponin component.

The conventional methods for enhancing ginsenoside Rg3 may be cumbersome and costly in manufacturing process using microorganisms or enzymes derived from microorganisms. Therefore, there is a need for the development of a method for producing a ginseng extract fraction with enhanced Rg3 as well as high yield while using a simpler chemical method.

Accordingly, the present inventors have established a condition for obtaining a large amount of ginsenoside, which can be a reactant for preparing Rg3 from ginseng, and converting it into Rg3 in a high yield and developing a method capable of purely obtaining Rg3. The present invention was completed.

Accordingly, it is an object of the present invention to provide a method for producing a ginseng extract fraction with high yield as well as pure Rg3 using a simpler chemical method.

Another object of the present invention is to provide a ginseng extract fraction enhanced by Rg3 obtained from ginseng by the above method.

In order to achieve the above object,

Reacting ginsenoside Rb1-containing alcohol extract of ginseng with a mixture of water and acetic acid to enhance ginsenoside Rg3;

Adsorbing the ginsenoside Rg3-enhanced ginseng extract to an adsorption resin for adsorption;

Distilled water is passed through the adsorption resin to elute and remove the unadsorbed components;

Adding 30-50 v / v% alcohol to the adsorptive resin to remove triol-based ginsenosides other than bisaponin and ginsenoside Rg3; And

It provides a method for producing a ginsenoside Rg3 enhanced ginseng extract fraction comprising the step of obtaining an eluate by adding 50-100 v / v% alcohol to the adsorption resin.

In another aspect, the present invention provides a ginseng extract fraction of the ginsenoside Rg3 enhanced by the ginsenoside Rg3 prepared by the above method containing 200 ~ 450 mg / g.

Hereinafter, the present invention will be described in more detail.

The inventors found that diol-based ginsenosides that can be converted to ginsenoside Rg3, such as ginsenoside Rb1, can be converted to ginsenoside Rg3 in high yields by reacting the extract of ginseng with a mixture of water and acetic acid. It was. In addition, the ginseng extract converted to ginsenoside Rg3 is passed through the adsorption resin, and then eluted with 30-50 v / v% alcohol to remove non-saponin and saponin components other than ginsenoside Rg3. It was found that ginseng extract enhanced with side Rg3 could be obtained.

Accordingly, the present invention in one aspect,

Reacting ginsenoside Rb1-containing alcohol extract of ginseng with a mixture of water and acetic acid to enhance ginsenoside Rg3;

Adsorbing the ginsenoside Rg3-enhanced ginseng extract to an adsorption resin for adsorption;

Distilled water is passed through the adsorption resin to elute and remove the unadsorbed components;

Adding 30-50 v / v% alcohol to the adsorptive resin to remove triol-based ginsenosides other than bisaponin and ginsenoside Rg3; And

It provides a method for producing a ginsenoside Rg3 enhanced ginseng extract fraction comprising the step of obtaining an eluate by adding 50-100v / v% alcohol to the adsorption resin.

The ginseng extract of ginseng may be any extract containing ginsenoside Rb1 obtained by treating ginseng with alcohol, but preferably an extract having a higher content of ginsenoside Rb1 may be used. In addition to ginsenoside Rb1, it may contain any ginsenoside that can be converted to ginsenoside Rg3, and such ginsenosides may contain ginsenosides Rb2, Rc, Rd, or mixtures thereof, which are diol-type saponins. Can be. Extracts having a higher content of diol-type ginsenosides, such as Rb1, can be obtained by adding 60-80% spirits to ginseng and stirring extraction at 40-80 ° C for 24-36 hours. This extraction method has been developed by the inventors as a result of the high extraction content of diol-type ginsenosides, including ginsenoside Rb1. When the extraction temperature exceeds 80 ° C, the thermal stability of ginsenoside Rb1 is reduced and the content of ginsenoside Rb1 extracted is reduced. In addition, if less than 40 ℃ extraction efficiency is lowered. And, if it is too long out of the time range, there is a problem that the economic efficiency or stability of ginsenosides, and too short, the extraction efficiency is lowered. More preferably, a spirit extract obtained by adding 65-75 v / v% of alcohol to ginseng and stirring extraction at 65-75 ° C. for 24-36 hours may be used, and most preferably 70 v / v% of ginseng. The alcohol extract obtained by adding alcohol and stirring and extracting at 70 ° C. for 24 hours may be used. The extract is preferably used by combining the extract three or more times.

Then, the obtained ginsenoside Rb1-containing alcohol extract is reacted with a mixture of water and acetic acid to enhance the ginsenoside Rg3. Rinsing Rb2, Rc, Rd, etc., including ginsenoside Rb1 contained in the spirit extract of ginseng may be converted into ginsenoside Rg3 by reacting with a mixture of water and acetic acid, and performed at 70-90 ° C. for 24-36 hours. can do. When the temperature of the conversion reaction exceeds 90 ℃ the thermal stability of the ginsenoside Rb1 is reduced, the content of the ginsenoside Rg3 is reduced. In addition, when less than 70 ℃ conversion efficiency is lowered. And, if it is too long out of the time range, there is a problem that the economic efficiency or stability of ginsenosides, and if too short, the efficiency of the conversion reaction falls. More preferably, the reaction may be carried out at 80-90 ° C. for 24-36 hours. The mixture of water and acetic acid is not particularly limited in the ratio as long as the reactant diol-type saponin ginsenoside can be converted to ginsenoside Rg3, but the conversion reaction to ginsenoside Rg3 is performed at a higher yield. For this purpose, it is preferable to use a mixture of 30 to 70 parts by weight of water and 30 to 70 parts by weight of acetic acid with respect to 100 parts by weight of alcohol. Ginseno, which is a diol-type saponin, including ginsenoside Rb1, at a significantly higher rate when using a mixture of alcohol and water and acetic acid, a mixture of water and alcohol, or a mixture of acetic acid and alcohol. The side may be converted to ginsenoside Rg3. Most preferably, a mixed solvent having a ratio of water and acetic acid of about 3: 1 is added to a volume of about 10 times the alcoholic extract of ginsenoside Rb-containing ginseng and reacted at 90 ° C. for 24 hours to convert it into ginsenoside Rg3. You can.

Thereafter, the process of obtaining only the ginsenoside Rg3 fraction may be performed by introducing and adsorbing the alcohol extract of ginseng enhanced by the ginsenoside Rg3 into the adsorption resin. Prior to performing the adsorption process, the ginsenoside Rg3-enriched spirit extract of ginseng is concentrated to 70-90 brix, it is preferable to dilute with water, preferably distilled water and add to the adsorption resin. The degree of dilution with water is not particularly limited as long as the concentrate of the ginsenoside Rg3-enhanced alcohol extract reacts with the entire adsorbent resin to adsorb saponin, but is preferably the concentrated extract weight. Can be diluted by addition of about 4-6 times, more preferably about 5 times water. As the adsorption resin, diion HP-20 (Mitsubishi, Japan) may be used, and may be used as a column-filled form for adsorption and fractionation.

Then, water, preferably distilled water, is passed through the adsorbent resin to elute and remove the unadsorbed components. Through this process, the components not adsorbed on the adsorption resin can be eluted and removed. The amount of distilled water used to remove the non-adsorbed components may be sufficient to elute and remove the non-saponin components. For example, distilled water of about five times the capacity of the adsorption resin may be used.

Then, to remove the non-saponin component adsorbed on the adsorption resin and triol-based ginsenosides other than ginsenoside Rg3, 30 to 50 v / v% of alcohol is added to the adsorptive resin to elute. Preferably, the process of eluting with the addition of 0 to 30 v / v% of alcohol before eluting by adding 30 to 50 v / v% of the alcohol to the adsorbent resin. Through this process, most components other than ginsenoside Rg3 are removed from the adsorption resin, thereby obtaining a ginseng extract fraction which is purely enhanced with ginsenoside Rg3 in the subsequent process.

When triol ginsenosides other than the non-saponin component and ginsenoside Rg3 are removed from the adsorption resin, the desired ginsenoside Rg3-enriched ginseng extract fraction can be obtained by eluting with 50-100 v / v% of alcohol. . Ginsenoside Rg3-enhanced extract fractions obtained by the above method may contain 100-200 mg / g of ginsenoside Rg3.

The ginseng used in the preparation of the ginseng extract may be any ginseng containing diol-type ginsenosides that may be reactants of ginsenoside Rg3 including saponin Rb1, for example, white ginseng, ginseng, red ginseng , Taeguksam, black ginseng, purified ginseng (puffed ginseng), or enzyme-treated ginseng or a concentrate thereof may be used.

In another aspect, the present invention provides a ginseng extract fraction fortified with ginsenoside Rg3 containing 200-450 mg / g of ginsenoside Rg3 prepared by the method according to the present invention.

As described above, according to the method of the present invention, ginsenoside Rg3, which is contained in trace amounts of ginseng and relatively large amounts in red ginseng, has a higher yield than a conventional method using microorganisms. Not only high but purely Rg3 enhanced ginseng extract fraction can be prepared.

Hereinafter, the present invention will be described in more detail with reference to the following examples. However, these examples are only for the understanding of the present invention, and the scope of the present invention is not limited by them in any sense.

Comparative Example 1: Preparation of Red Ginseng Extract

70% of alcohol was added to 1 kg of red ginseng by 15 times the weight of the sample, and the extract of red ginseng was prepared by filtering after repeated extraction three times. High performance liquid chromatography (HPLC) was performed on the alcohol extract of red ginseng obtained by the above method. The HPLC chromatoram thus obtained is shown in FIG. 1.

Example 1: Preparation of Ginseng Extract Using 70 v / v% Alcohol

70% alcohol was added to 1 kg of dried ginseng as much as 15 times the weight of the sample, and the extraction conditions of ginseng were prepared by repeating and filtering three times while changing the extraction conditions as shown in Table 1 below. In each alcohol extract of ginseng obtained by the above method, the content of ginsenoside Rb1 was measured by high performance liquid chromatography (HPLC). The results are shown in Table 1 below.

TABLE 1

According to the experimental results, the content of ginsenoside Rb1 is significantly changed depending on the temperature conditions during extraction using a 70 v / v% alcohol, in particular, the content of Rb1 was found to be significantly high at about 70 ℃. In addition, the content of ginsenoside Rb1 was found to be significantly higher when treated for 24 hours or more in the treatment time. However, at 80 ° C, the longer the treatment time, the lower the tendency, which may be due to temperature instability.

Preparation of alcohol extract of ginseng was confirmed that the treatment for 24 hours at 70 ℃, it was shown in the HPLC results of the alcohol extract obtained at 70 ℃, 24 hours conditions.

Example 2: Ginsenoside Rg3 Enhancement Process

In Example 1, a mixed solvent (10-fold of the concentrate) under the conditions shown in Table 2 was added to the concentrated solution (75-80 Brix) of the alcohol extract of ginseng obtained at 70 ° C. and 24 hours, and heated at 90 ° C. for 24 hours. It was. The content of ginsenoside Rg3 was measured by high performance liquid chromatography (HPLC). The results are shown in Table 2 below.

In addition, using the concentrate of the same extract of ginseng and the temperature and time conditions were reacted in the same manner as in Table 3, and then the content of ginsenoside Rg3 was measured, the results are shown in Table 3 below .

TABLE 2

 [Table 3]

According to the experimental results, when adding a mixed solvent of water and acetic acid (acetic acid) to the ethanol extract of ginseng ginsenoside Rg3 in a significantly higher yield than when using ethanol, water, acetic acid alone or a combination of alcohol and acetic acid You can see that it is converted to. In addition, it can be seen that when a 3: 1 mixed solvent of water and acetic acid is added and heated at 90 ° C. for 24 hours, it is converted into ginsenoside Rg 3 in a significantly higher yield than other temperature and time conditions.

Example 3: Adsorption Resin DIION HP-20 Adsorption Process

In Experimental Example 2, a 3: 1 mixed solvent of water and acetic acid was added, and the ginsenoside Rg3 reaction product obtained by heating at 90 ° C. for 24 hours was concentrated by heating to 65 brix. Thereafter, 5 times the weight of the concentrate was added thereto to sufficiently dilute it, and then the saponin component was adsorbed by passing through the adsorption resin Dione HP-20 resin. Then, about 5 times the capacity of the resin was continuously passed through distilled water to remove unadsorbed components.

Then, to remove the non-saponin component and the triol-based saponin component, first elute through 0-30 v / v% alcohol of about 5 times the volume of resin, and then elute through 30-50 v / v% alcohol. I was.

Then, the eluate obtained by passing 50-100 v / v% alcohol was taken. High performance liquid chromatography (HPLC) was performed on the eluate obtained in each step, and the results are shown in FIGS. 3 (0-30 v / v% alcohol elution fraction) and FIG. 4 (30-50 v / v% alcohol elution). Fraction), and FIG. 5 (50-100 v / v% alcohol elution fraction).

According to Figure 3-5, 0 ~ 30 v / v% alcohol elution fraction and 30 ~ 50 v / v% alcohol dissolution fraction, Rg3 was not found, 50 ~ 100 v / v% obtained after removing this elution fraction Ginsenoside Rg3 was present as a high peak in the alcohol elution fraction.

Experimental Example 1: Determination of the content of ginsenoside Rg3

The content of ginsenoside Rg3 was determined by HPLC for the extract obtained in each of Examples 1 to 3 above. In Example 1, alcohol extracts obtained at 70 ° C. and 24 hour conditions were used. In Example 2, a product obtained by adding a 3: 1 mixed solvent of water and acetic acid to the alcohol extracts and then performing Rg 3 conversion reaction at 90 ° C. for 24 hours was used. In Example 3, the content of ginsenoside Rg3 was measured for 50 ~ 100 v / v% alcohol elution fraction.

The measurement results are shown in Table 4 below.

[Table 4]

According to the results of Table 4, ginsenoside Rg3, which was hardly detected in the ginseng extract of ginseng, was eluted with 50-100 v / v% alcohol in the adsorption resin after the conversion reaction to Rg3, and thus the content of Rg3 was greatly enhanced. It was confirmed that ginseng extract fractions could be obtained.

In addition, when compared with the alcohol extract of red ginseng in Comparative Example 1, it was shown that the ginseng extract fraction containing ginsenoside Rg3 in a higher content than the red ginseng extract by the method according to the present invention.

1 is an HPLC chromatogram of red ginseng alcohol extract obtained by adding 70% alcohol by 15 times the sample weight to 1 kg of red ginseng, and filtering after repeated extraction three times. FIG. HPLC chromatogram of alcohol extract obtained by extraction at 70 ° C. for 24 hours.

3 is added to the alcohol extract extracted at 70 ℃ / 24% alcohol for 24 hours at 70 ℃, added a 3: 1 mixed solvent of water: acetic acid and reacted at 90 ℃ for 24 hours, concentrated and diluted to die It is an HPLC chromatogram of the fraction obtained by making it adsorb | suck using an ion HP-20 adsorption resin, and eluting with 0-30 v / v% of alcohol.

Figure 4 is added to the alcohol extract extracted for 70 hours at 70 ℃ / 70% alcohol for 24 hours, and added 3: 1 mixed solvent of water: acetic acid and reacted at 90 ℃ for 24 hours, concentrated and diluted die It is an HPLC chromatogram of the fraction obtained by adsorbing with an ion HP-20 adsorption resin, eluting with 0-30 v / v% alcohol, and then eluting with 30-50 v / v% alcohol.

Figure 5 is added to the alcohol extract extracted for 70 hours at 70 ℃ for 24 hours at 70 v / v% alcohol in accordance with an embodiment of the present invention was added a 3: 1 mixed solvent of water: acetic acid and reacted at 90 ℃ for 24 hours Thereafter, the concentrated and diluted mixture was adsorbed using a dia HP-20 adsorption resin, eluted with 0-30 v / v% alcohol, eluted with 30-50 v / v% alcohol, and then 50- HPLC chromatogram of fractions obtained by eluting with 100 v / v% alcohol.

Claims (8)

Reacting ginsenoside Rb1-containing alcohol extract of ginseng with a mixture of water and acetic acid to enhance ginsenoside Rg3; Adsorbing the ginsenoside Rg3-enriched spirit extract of ginseng to an adsorption resin for adsorption; Distilled water is passed through the adsorption resin to elute and remove the unadsorbed components; Adding 30-50 v / v% alcohol to the adsorptive resin to remove triol-based ginsenosides other than bisaponin and ginsenoside Rg3; And Method of preparing a ginsenoside Rg3-enriched ginseng extract fraction comprising the step of obtaining an eluate by adding 50 ~ 100 v / v% alcohol to the adsorption resin. The method according to claim 1, wherein the ginsenoside Rb1-containing alcohol extract of ginseng is 60-80 v / v% alcohol added to ginseng and stirred extraction at 65-75 ℃ for 24-36 hours. The method of claim 1 wherein the mixture of water and acetic acid is a mixture of 60-80 parts by weight of water and 20-40 parts by weight of acetic acid. According to claim 1, wherein the step of strengthening ginsenoside Rg3 of ginseng is characterized in that the ginsenoside Rb1 containing alcohol extract of ginseng is reacted with a mixture of water and acetic acid at 80-90 ℃ for 24-36 hours Way. The method according to claim 1, wherein the ginsenoside Rg3-enriched spirit extract of ginseng is concentrated to 70-90 brix and then diluted with distilled water and added to the adsorption resin. The method of claim 1 wherein the adsorptive resin is Diion HP-20. The method of claim 1, wherein the ginseng is white ginseng, ginseng, red ginseng, taegeuk ginseng, black ginseng, purified ginseng (puffed ginseng), or enzyme-treated ginseng or a concentrate thereof. A ginseng extract fraction fortified with ginsenoside Rg3 containing 200-500 mg / g of ginsenoside Rg3 prepared by the method according to any one of claims 1 to 7.

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