KR830002765B1 - Method for preparing medicinal extract from seeds of Cardus marianus - Google Patents

Abstract

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Description

Method for preparing medicinal extract from seeds of Cardus marianus

The present invention relates to a method for preparing a medicinal extract that can be used for the treatment of liver disease from the seed of Cardus Marianus.

Cardus Marianus is a plant belonging to the Asteraceae family and is wild or cultivated in southern Europe and South America.

The seeds of Cardus marianus (hereinafter referred to as seeds) have been used for various liver diseases and biliary tract diseases as tea by themselves or like plants such as dandelion and chicory.

In the early 19th century, German physician Rademarker studied the clinical effects of these seeds physically, and as a result, the seeds were found to have an excellent effect on liver disease.

In recent years, Wagner and Hansel have succeeded in independently extracting and separating active ingredients from seeds and determining their chemical structure. In addition, the German Central Cancer Institute conducted molecular biological experiments using purely isolated materials, and found that the foreign material promotes the regeneration of destroyed hepatic cells.

This proved the plant's medicinal value.

There are many known methods for making modern pharmaceutical formulations from these seeds, and methods for quantifying the active ingredients are known and developed. As a result, methods for preparing formulations containing high concentrations of active ingredients have been developed.

The active ingredient of the seed belongs to the flavonoid material bacterium, and consists of silybin, silidianine, and silicryrin.

The conventional method is to decompress the silyboom marianum seeds in a seed press screw press as disclosed in Patent Publication No. 76-461, extract the residue with ethyl acetate, and evaporate the solvent to remove the oily dry residue. This is obtained by only distributing the countercurrent in countercurrent on the lower layer of the methanol / water / petroleum ether system, centrifuging, and concentrating the clarified liquid from which the solid component is removed to obtain sirimarin. The present invention is characterized by low extraction steps and very easy extraction, which is a method of remarkable technology and is advantageous in terms of cost.

That is, the extraction process of the present invention consists of three steps of degreasing (脫脂), dehydration (脫水), solvent extraction of the seed, which will be described in detail as follows.

First, put the seeds in the Soksteek part compression extractor (Note: means a compression extractor with Sokstete device) and squeezed 2-3 times so that the shell of the seed is destroyed, add a non-polar plaque to it, stir and compress The solvent and its dissolved solution are sent to the evaporation section of Soxteut.

The solvent is evaporated from the evaporation section of Soxtete and sent to the pressed seeds. In this way, when the amount of solvent in the pressed seeds is stirred, the seed powder is stirred with a stirrer attached to the pressed piston, and then pressed again, and the solvent and the dissolved substance are sent to the evaporator. Repeat this to complete degreasing. The solvent used at this time is a non-polar straight-chain or branched saturated hydrocarbon of 5-8 carbon atoms.

The amount of the degreasing solvent for the seed is preferably in the range of 2-7 times, and the degreasing time may be 20-180 minutes depending on the type of solvent, the amount and the amount of the seed. The advantage of using saturated hydrocarbon as a degreasing solvent is to reduce the loss of active ingredients. Although compressing the seed without the solvent can remove a considerable amount of unnecessary substances, polar substances are mixed in the compressed lipids, so that the polar active ingredients are dissolved in the lipids, resulting in loss of the active ingredients.

However, the saturated hydrocarbon solvent of the present invention selects and dissolves only lipids so that the solution itself is nonpolar, so that an active ingredient having a relatively high polarity is not eluted into the solution.

The degreasing process described above is the first element of the present invention because it has the advantage of low solvent consumption and short degreasing time.

Next, the seed powder thus degreased contains water, which may exist in a relatively free state chemically, and may have molecular bonds with polymers or polar substances.

For example, when pectin contains water, chemical molecules such as hydrogen bonds with water molecules make the molecules themselves flattened. Pectin in this state is dissolved in a significant amount in contact with alcohols or other solvents capable of hydrogen bonding. Or emulsified. In other words, when the seed is extracted with a solvent having the ability of hydrogen bonding without removing the moisture of the seed, a considerable amount of the polymer is extracted together with the active ingredient. In contrast, the solubility of dry pectin in these solvents is much lower.

Pectin-like proteins, peptides, carbohydrates, and the like belong to polymer materials having such properties.

In conclusion, in order to reduce the elution amount of the polymer material into the organic solvent, the dehydration operation of the seed part should be preceded. Dehydration operation is preferably carried out under reduced pressure at a temperature of 20-40 ℃.

Such a dehydration method can raise the concentration of the active ingredient in the primary extract by inhibiting the elution of the polymer material in the solvent extraction process, and by providing convenience to the preparation of the high concentration extract to run, this dehydration process is the second element of the present invention It's fresh.

Finally, an organic solvent was used to extract the active ingredient from the degreased and dehydrated seed meal.

Organic solvents are alcohols, kerons, or eslers.

The alcohols are R ₁ OH of Formula (I) (wherein R ₁ represents an alkyl group of C ₁ -C ₄ ), and kerons are R ₂ CO—R ₃ of Formula (II) (wherein R _{2 3} each represents an alkyl group of C ₁ -C ₃ , and the esters are R ₄ COOR of Formula (III), wherein R ₄ and R ₅ each represent an alkyl group of C ₁ -C ₃ .

A certain amount of degreased and dehydrated seed powder is extracted with a Soxter part compression extractor, which is described above with these solvents, and then the extraction solution is concentrated to a certain amount. The concentrated extract solution is left to stand at a temperature range of 5-25 ° C. and the resulting precipitate is removed. Repeat the above precipitation procedure until the minimum amount of extraction solution remains within the limit that no active ingredient is contained during the precipitation. In this way, a dry extract containing 30-45% of the active ingredient was prepared depending on the nature of the solvent.

As such, the step of preparing a high concentration of extract that can be used pharmaceutically as the monosolic treatment of the third step is to be the third element of the present invention.

In the extraction of the single solvent as described above, while increasing the extraction temperature, the extraction speed is increased, while there is a disadvantage that a lot of impurities are extracted so that the extraction stock solution is suspended.

At high temperatures, chemical changes in the active ingredient may occur. The amount of the extraction solvent is 3-10 times the amount of the used value according to the type of solvent and the extraction conditions. It is also an advantage of the present invention that no excess solvent is actually necessary because of the use of a fast steep extractor.

Various methods have been conventionally used to make extracts containing high concentrations of total polarvonoids.

That is, the chromatographic method using polyamide, Sephadex, cellulose, etc., the coprecipitation method of the active ingredient using an inorganic ion or a polymer precipitation agent, etc. are mentioned. However, conventional methods, such as chromatographic methods, are uneconomical for mass production, and use of nominal methods has the disadvantages of loss of active ingredients and fear of mixing of foreign substances.

Therefore, the present invention has been developed a method of increasing the total purity of the polar bonoids by a mixed solvent method. That is, a method of removing the high-molecular substances and the high-polar substances in the extract by mixing a low polarity solvent with a predetermined amount of a single solvent extract that already contains a considerable amount of the amount of the polar bonide.

The addition solvent used at this time is the above-mentioned ester of R ₄ COOR ₅ of general formula (III) or R ₆ -OR ₇ of general formula (IV), wherein R ₆ and R ₇ are C ₁ -C ₄ alkyl groups. Phosphorus ethers, tetrahydrofuran and dioxane are also used as addition solvents, and halogenated alkyls such as chloroform and carbon tetrachloride are also used as addition solvents.

However, it is difficult to use it as an additive solvent by causing the abrupt fall of the solvent of other halogenated alkyls. Even when a pure hydrocarbon solvent is used as an additive solvent, it is very difficult to control the optimum concentration of the active ingredient due to the drastic extreme decrease of the mixed solution. Therefore, the most suitable substance group as an additive solvent is the ester of the general formula (III) and the ethers of the general formula (IV), tetrahydrofuran, dioxane, chloroform and carbon tetrachloride.

They serve to precipitate more polar molecules, i.e., less soluble polymers, which are themselves in solution while having solubility in the active ingredient.

Thus, the operation of increasing the purity of the active ingredient by adjusting the polarity of the solvent is an important element of the present invention.

The amount of the added solvent with respect to a predetermined amount of the monosolvent extract obtained from 1 kg of seeds was shown in Table 1 below.

TABLE 1

Amount of Solvent Added to Monosol Concentrate

(Unit: ml)

When esters of general formula (III) having high solubility with respect to the active ingredient are used, it is advantageous to keep the amount of the monosolvent extract to a minimum.

For example, if the amount of extract of alcohols frozen from 1 kg of seeds is concentrated to 200-300 ml and the esters are added in a range of 1-2 times, the purity of the active ingredient can be easily increased.

When the addition solvent is added, the optimum temperature is 15-35 ℃, which is a range of room temperature, and the standing temperature after mixing is 5-25 ℃. The standing time is 3-l 2 hours depending on the type of solvent.

Thus, using the mixed solvent method can be prepared a dry extract in the range of 70-80% of the total flavonoid content, the process of the present invention will be described in more detail in the following examples.

Example 1

First process (degreasing and drying process)

1 kg of seeds are placed in a Sokster part extractor (hereinafter referred to as an extractor) and pressed 2-5 times so that the seeds are destroyed.

Next, 1.5 l of N-hexane is added to the pulverized seed, stirred well, and pressed to remove the solvent and its dissolved material.

The same operation is performed again using 1.5 L of solvent.

Run the Soxter system and continue to squeeze for 2 hours.

The seed powder from which the solvent was completely removed was put in a vacuum dryer and dried at room temperature for 12 hours to obtain 668 g of dry seed powder. (Yield 66.8%)

Second ball. Extraction process

1 kg of seeds is treated in a first step to obtain 700 g of seed powder. (Yield 77%)

This seed is placed in an extractor and press-extracted at 25 ° C. for 3 hours with 2 liters of methanol. The methanol extract solution of about 2 L obtained after the final compression was concentrated to 500 ml and left at 5 ° C. for 12 hours.

The resulting precipitate is filtered off and the filtrate is concentrated again to 300 ml and left at 5 ° C. for 12 hours.

At this time, the precipitate was removed and the filtrate was obtained 49.4g of concentrated dry extract.

3rd process (purification process)

300 ml of the methyl alcohol concentrate prepared in the second step was added to a 2 L beaker, and 500 ml of ethyl acetate was slowly added while stirring. After stirring for 10 minutes, the mixture was left for 1 hour.

The precipitate produced is filtered and the solid on the filter paper is washed with 100 ml of ethyl acetate.

The combined filtrates were evaporated to dryness to freeze dry extract 32 '.

Example 2

1 kg of seed is obtained in the same manner as in the first step of Example 1 to 600 g of seed powder. (Yield 60%)

2nd process

Seed powder was added to an extractor and extracted by compression of 2 liters of ethyl alcohol at 25 ° C. for 3 hours. About 2 L of ethyl alcohol extract was concentrated to 500 ml and left at 5 ° C for 12 hours. At this time, the resulting precipitate is removed by filtration and the filtrate is concentrated to 300ml again and left for 12 hours at 5 ℃. The resulting precipitate was filtered off and the filtrate was evaporated to dryness to obtain 47.lg of dry extract.

3rd process

300 ml of ethyl alcohol concentrate was prepared by the second step and placed in a 2 L beaker and stirred. 500 ml of ethyl acetate was added thereto, followed by stirring for 10 minutes. After standing for 3 hours, the resulting precipitate is filtered. The solid on the filter paper is washed with 100 ml of ethyl acetate. The combined filtrates were evaporated to dryness to obtain 20 g of dry extract.

Example 3

500 ml of the ethyl alkal concentrate prepared in the second step of Example 2 was added to a 2 L beaker, and 178 ml of ethyl ether was added little by little while stirring, followed by standing at 5 ° C. for 12 hours. The precipitate formed is filtered off, and 80 ml of ethyl ether is added to the filtrate, and the mixture is left at 5 ° C.

The resulting precipitate was filtered off and the filtrate was evaporated to dryness to weigh 27.4 g of the dry extract.

Example 4

500 ml of ethyl alcohol concentrate prepared in the second step of Example 2 was added to a 2 L beaker and 200 ml of ethyl ether was slowly added while stirring. After the stirring was continued for 10 minutes, the mixed solution was left at 5 ° C for 12 hours. The resulting precipitate is removed by filtration, 100 ml of ethyl ether is added to the filtrate, mixed and left to stand again at 5 ° C for 12 hours. The resulting precipitate was removed by filtration and the filtrate was evaporated to dryness to obtain a dry extract 26g.

Example 5

Using 1 kg of seeds, 692 g of seed was prepared in the first step of Example 1 (yield 69.2%).

2nd process

Seed powder is added to the extractor and extracted with 2 liters of ethyl acetate for 5 hours. The solution of about 2 L of ethyl acetate in Soxter Furask was concentrated to 500 m and then left at 5 ° C. for 12 hours. The resulting precipitate is filtered off and the filtrate is concentrated again to 180 ml and left at 5 ° C. for 12 hours. The resulting precipitate was filtered off and the filtrate was evaporated to dryness to obtain 30.3 g of a dry extract.

3rd process

500 ml of ethyl acetate concentrate obtained in the second step was added to a 2 L beaker and 210 ml of ethyl ether was slowly added while stirring. After further stirring for 10 minutes, the mixed solution is left at 5 ℃ for 12 hours. The precipitate formed is filtered off and 67 ml of ethyl ether is added to the filtrate and mixed. This mixed solution is left to stand again at 5 ° C for 12 hours. The produced precipitate was removed by filtration and the filtrate was evaporated to dryness to give a dry extract 24.7g.

Example 6

678 g of seed powder are obtained by the first step of Example 1 from 1 kg of seeds. (Yield 67.8%)

2nd process

This was put into an extractor and extracted with 2 L of acetone for 3 hours. The acetone extract was concentrated to 500 ml and left at 5 ° C. for 12 hours. The resulting precipitate is filtered off and the filtrate is concentrated again to 200 ml and then left at 5 ° C. The produced precipitate was removed by filtration and the filtrate was evaporated to dryness to obtain a dry extract 36.4.

3rd process

100 ml of ethyl ether was added to 500 ml of the acetone concentrate prepared according to the second step and mixed. At this time, the produced precipitate is removed by filtration, and the filtrate is added again to 100 ml of ethyl ether, mixed and left to stand at 5 ° C. for 12 hours. The produced precipitate is removed by filtration, and the filtrate is further added by adding 50 ml of ethyl ether, mixed and left at 5 ° C. for 12 hours. The resulting precipitate was filtered off and the filtrate was evaporated to dryness to give 26.5 g of dry extract.

The result of preparing a dry extract in the same manner as in Example 1-6 using terahydrofuran, dioxane, chloroform and carbon tetrachloride as an additive solvent is shown in Table 2 below.

TABLE 2

Comparison table of effects of using solvent

Unit: amount and concentration: ml

Drying quantity: g

Purity:%

Claims (4)

Cardus marianus seeds are degreased using a non-polar saturated hydrocarbon having 5 to 8 carbon atoms in combination with the compression method, followed by dehydration, followed by alcohols of general formula R1OH, kerons of R2-CO-R3, or esters of R4COOR5. Extraction is performed, and the extract is concentrated, prevented, and repeated filtration process to prepare a medicinal extract from the cardinalus Marianus seeds. Wherein Rl is an alkyl group of Cl-C4, and R2, R3, R4 and R5 are each alkyl groups of Cl-C3. The process according to claim 1, wherein N-nucleic acid is used as the nonpolar saturated hydrocarbon and methanol, ethanol, acetone or ethyl acetate is used as the extraction solvent. Cardus marianus seeds were combined with the compression method and degreased using non-polar saturated hydrocarbons having 5 to 8 carbon atoms, followed by dehydration and extraction with kerons with alcohols R2-CO-R3 of the general formula R1OH, or esters with R ₄ COOR ₅ . Carru which concentrates, leaves and filters the mixed solvent extract by adding esters of the general formula R ₄ COOR ₅ , the general formula R ₆ -OR ₇ ether, terahydrofuran, dioxane, chloroform, or carbon tetrachloride to the extract. A method of making medicinal extracts from Dus Marianus seeds. In the above formula, R ₄ , R ₅ are each an alkyl group of C ₁ -C ₃ , and R ₆ , R ₇ are each an alkyl group of C ₁ -C ₄ . 4. The process according to claim 3, wherein methanol, ethanol, acetone or ethyl acetate is used as the primary extraction solvent and ethyl acetate, ethyl ether, tetrahydrofuran, dioxane, chloroform or carbon tetrachloride are used as the addition solvent.