KR0179415B1 - Novel saponin component and purification thereof - Google Patents

Abstract

The present invention relates to novel saponins, 6-O-β - D-glucopyranosyl dammar-20 (22), 24-diene-3β, 6α, 12β-triol. This saponin was named Ginsenoside Rh ₄ . This invention also relates to a method for separating ginsenoid Rh ₄ from silica ethanol extract of Korean red ginseng by silica gel column chromatography.

Description

Novel saponin components and isolation method

FIG. 1 is a diagram showing the hydrogen nuclear magnetic resonance spectrum (400 mega hetz, deuterated pyridine) of ginsenoside Rh ₄ .

2 is a diagram showing the carbon nuclear magnetic resonance spectrum (100 mega hetz, deuterated pyridine) of ginsenoside Rh ₄ .

3 is a diagram showing the hydrogen nuclear magnetic resonance spectrum (400 mega hetz, deuterated pyridine) of the non-sugar portion of ginsenoside Rh ₄ .

FIG. 4 is a diagram showing the hydrogen nuclear magnetic resonance spectrum (100 megahealth, deuterated pyridine) of non-sugar part of ginsenoside Rh ₄ .

[Purpose of invention]

[Technical field to which the invention belongs and the prior art in that field]

The present invention is a novel saponin represented by the following structural formula (1) 6-O-β - D- glucopyranosyl dammar-20 (22), 24-diene-3β, 6α, 12β-triol (hereinafter ginsenoid Rh ₄ ).

The stated ginsenoid Rh ₄ is a novel substance that is separated from ginseng.

Korean ginseng (Panax ginseng C. A. Meyer) is a perennial herb belonging to the Araliaceae family. White ginseng is a dry processed ginseng as it is, red ginseng is a process of shame. In other words, red ginseng is steamed and dried, and the ginseng component is thought to undergo chemical change because it is heated during processing. In particular, the main purpose of this numerical process in red ginseng can be thought to be to increase the drug efficacy, to broaden the site of action of the drug and to keep the drug clean.

Studies on the structure of ginseng saponins have been conducted since the early 1960s by the Shibata and Tanaka groups in Japan. In other words, ginsenosides Ro, Ra, Rb, Rd, Re, Rf, Rg and Rh have been identified and separated in the order that the ginseng saponins were depleted by chromatographic chromatography. With the establishment of the method, the pharmacological action of individual saponins began to be examined in connection with the effects of ginseng. In particular, many pharmacological studies have been conducted on the major saponins contained in ginseng, ginseng, white ginseng, and red ginseng. Among them, saponins contained mainly in red ginseng or known only in red ginseng are the major It has been a concern. Red ginseng saponins known to date are mostly found in white ginseng, but the ginsenosides Rs ₁ , Rs ₂ and 20 (R) -ginsenosides Rh ₁ , 20 (S) -ginsenosides Rg ₃ and 20 (R) -ginsenoside Rg _{2 and the like} were also identified. In particular, the above-mentioned saponin components are considered to be prosapogenin glycosides in which the terminal 20 carbon or the terminal sugar of carbon ginseng is removed from the main saponin of ginseng by thermal decomposition during red ginseng manufacturing process. In addition, a unique saponin component, which is estimated to be a saponin compound in which a sugar of 20 carbon is desorbed and a double bond is formed due to chemical change, is released. As described above, the saponin compound in which the double bond is introduced at carbon 20 has been identified as ginsenoside Rh ₃ by separating from the leaves of ginseng, but it has not been reported that ginsenoside Rh ₃ has been separately identified from the root of ginseng.

[Technical problem to be achieved]

An object of the present invention is to provide a new ginseng saponin.

The present inventors found that there is a new saponin in which a double bond is introduced at carbon 20 in the process of tracking new active ingredients of Korean red ginseng. Compared to the compound and ginsenoside Rh ₁ binary, but the carbon nuclear magnetic resonance spectrum and the position of each peak of ginsenoside Rh ₁ is almost the same it is possible to find some difference in ambient carbon 20 times, in the case of a new saponin By showing a molecular weight of 620, one molecule of water fell on carbon number 20 of ginsenoside Rh ₁ and it was confirmed that a double bond was produced. In addition, the stereocoordination of the double bond between carbon 20 and 22 was determined by Entgen as NOE was observed between 21 methyl hydrogen and 23 methylene hydrogen from the nuclear magnetic resonance spectrum (NOESY). It was. However, the case of ginsenoside Rh ₃ isolated from the leaves of ginseng has been reported as Zusammen.

[Configuration and Function of Invention]

It consists of ginsenoside Rh4 of the above-mentioned structural formula (1) of this invention.

Ginsenoside Rh ₄ of the present invention showed a three-dimensional difference from ginsenoside Rh ₃ . Ginsenoside Rh ₄ , a novel saponin glycoside of the present invention, could be estimated to be produced by dehydration of one molecule of water from ginsenoside Rh ₁ during the preparation of red ginseng from ginseng. The physicochemical properties of the new saponin were as follows and named this material ginsenoside Rh ₄ of formula (1).

Physicochemical Properties of Ginsenoside Rh ₄

1) Colorless microcrystals (methanol-water)

2) melting point: 160-161 ℃

3) Specific light intensity: [α] _D : + 28.2 ° (c = 1.0, methanol)

4) Rf value: Chloroform-methanol-water-7: 3: 1, Rf using Kieselgel F ₆₀ thin layer chromatography; 0.64

5) Thin layer chromatography in 4) Sprayed with 20% water-soluble sulfuric acid and heated to give red color.

6) High Speed Atomic Impact Mass Spectrum: M ⁺ + H = 621

Elemental Analysis C ₃₆ H ₆₀ O ₈ Calculated: 69.64% Carbon, Hydrogen: 9.74%

Anal: 69.68% carbon, 9.80% hydrogen

8) Hydrogen Nuclear Magnetic Resonance Spectrum: same as first degree.

9) Carbon Nuclear Magnetic Resonance Spectrum: same as 2nd degree.

The present inventors then obtained Dhammara-20 (22) 24-diene-3β, 6α, 12β-triol of Structural Formula (2) by acid hydrolysis in the course of confirming the structure of the ginsenoid Rh ₄ . . As a spectroscopic result of the non-sugar part, it was confirmed that the double bond was formed by the dehydration of the hydroxyl group of carbon 20, and the stereoconfiguration of the double bond in the carbon 20 of the protopanax triol was also confirmed in the entgegen type.

Physical and chemical properties of this material are as follows.

1) White powder (methanol-water)

2) melting point: 131-132 ℃

3) Specific light intensity: [α] _D : + 40.0 ° (c = 0.2, methanol)

4) Electron impact mass spectrum: calculated value C ₃₀ H ₃₅ O ₃ : 458. 3762

Found C ₃₀ H ₃₅ O ₃ : 458. 3775

5) Hydrogen nuclear magnetic resonance spectrum: as shown in FIG.

6) Carbon Nuclear Magnetic Resonance Spectrum: same as FIG.

The present invention also relates to a method for separating ginsenoside Rh ₄ from ginseng.

For example, the separation method of ginsenoside Rh ₄ of the present invention is as follows. Ethanol extract was obtained from Korean red ginseng, and repeated silica gel column chromatography was performed to separate ginsenoside Rh ₄ into a colorless microcrystalline form. In addition, after hydrolysis of the ginsenoside Rh ₄ obtained in the microcrystalline form for structural confirmation, the non-sugar part is purified by column chromatography.

In the present invention, silica gel chromatography for separating ginsenoside Rh ₄ is preferably repeated three times. For example, a chloroform: methanol 5: 1 solution is used as an eluent and a chloroform: methanol is used twice. It is recommended to use water 20: 3: 1 solution and n-butanol: ethyl acetate 1: 3 solution three times.

Hereinafter, the present invention will be described in detail with reference to Examples.

Example 1

Isolation of Ginsenoside Rh ₄ from Red Ginseng

1Kg of Korean red ginseng powder was sonicated with 2.5 L of ethanol three times each overnight at room temperature to obtain 120 g of ethanol extract. 100 g of them are separated by silica gel column chromatography (eluent = chloroform: methanol = 5: 1) to separate the four fractions. The third fraction was silica gel column chromatography (chloroform: methanol: water 20: 3: 1), and then repeated silica gel color chromatography (n-butanol: ethyl acetate = 1: 3) to obtain ginsenoside Rh of formula (1). ₄ 94.9 mg was isolated and purified.

Example 2

20 mg of ginsenoside Rh ₄ is dissolved in 9% hydrochloric acid / methanol and refluxed for 2 hours. The reaction solution was neutralized with silver carbonate, filtered and concentrated under reduced pressure. A portion of the silica gel thin layer chromatography (chloroform-methanol-water = 6: 4: 1) was used to identify D-glucose, and the remaining portion was silica gel column chromatography (nucleic acid-ethylacetate = 2: 1). 6.3 mg of Dhammara-20 (22), 24-diene-3β, 6α, 12β-triol of formula (2) were isolated.

Claims (3)

Novel 6-O-β - D-glucopyranosyl dammar-20 (22), 24-diene-3β, 6α, 12β-triol (hereinafter referred to as ginsenoid Rh ₄ ) represented by formula (1). 6-O-β - D-glucopyranosyl Dhammara-20 (22), 24-diene-3β, 6α, 12β-triol of Structural Formula (1) as described by silica gel column chromatography from ethanol extract of Korean red ginseng (Ginsenoid Rh ₄ ). The silica gel column chromatography for separating the ginsenoid Rh ₄ of formula (1) is a primary, chloroform, methanol and water (20: 3) using chloroform and methanol (5: 1) as eluent. : A secondary silica using 1) as an eluent and tertiary silica gel column chromatography using n-butanol and ethyl acetate (1: 3) as eluents.