KR100915288B1 - method extracting shikonin using fatty acid breakdown enzyme, an extract the same and method for using an extract the same - Google Patents

Abstract

Disclosed are a method for selectively producing sikonin from lichen root or lichen root extract using fatty acid degrading enzyme, and a method of using the same. In this method, esterase, one of fatty acid degrading enzymes, is treated with an extract of lichen root or lichen root to remove acyl groups of sikonin derivatives to selectively produce only sikonin. According to this method, by selectively producing siconin from siconin derivatives, pigments with higher polarity, brighter color and less cytotoxicity can be produced easily from the ground, and can be produced in various fields such as food, pharmaceuticals, etc. There is an effect that can be useful in the field.

Description

Method extracting shikonin using fatty acid breakdown enzyme, an extract the same and method for using an extract the same}

The present invention relates to a method for extracting sikonin using a fatty acid degrading enzyme, a method for extracting the same, and a method for using the extract. In particular, a siconin and a siconin derivative are mixed by selectively producing only siconin using a fatty acid degrading enzyme. It suggests a method that can produce pigments with higher polarity and brighter color than conventional extracts, but with less cytotoxicity.

It is a perennial herb called Chichi, Licorice, Hemostasis, and Root Root. It grows in mountains and fields all over Korea, and has white hairs on the leaves and stems. Fruit bears The roots are thick and purple, and they have been used in traditional medicine and folk medicine to promote blood circulation, prevent constipation, antibiotics, burn treatment, and antipyretic drugs. The roots of the grass contain a large amount of naphtoquionone-based compounds, such as siconin and derivatives of these substances. These components are reddish purple and most of the pharmacological effects of the grass are known to be due to these compounds. Research is underway to increase pharmacological value and yield.

These siconin and siconin derivatives are generally low-soluble fat-soluble substances and soluble in non-polar solvents, but not soluble in highly polar solvents. In particular, there are more than five kinds of siconin derivatives in the roots of the grass, but the content of highly polar siconins is usually 5% or less and the lower polarities of siconin derivatives account for 95% or more. Although it is well extracted in non-polar solvents such as acetate, it is not well extracted in solvents having high polarity such as water. Even in the production of Hongju, one of the representative foods using grass, if the alcohol content is low, siconin derivatives are not well extracted, so only products with an alcohol content of 40% or more are usually produced.

In addition, because siconin is bright red and has low cytotoxicity, while siconin derivatives are dark red and often have cytotoxicity. Although the yield is not so high, it is not commercialized.

The present invention has been made to solve the problems of the prior art as described above, the technical problem of the present invention is to search for enzymes that can convert the siconin derivatives to sikonin targeting lipase and esterase which are fatty acid degrading enzymes As a result, it was found that esterase derived from pig liver has not only the ability to decompose fatty acids but also to convert siconin derivatives to siconins. Its main purpose is to provide a method for the selective and convenient production of sigonin, a natural pigment with low color and low cytotoxicity.

In addition, the present invention has another object to provide a composition comprising a natural pigment and a natural pigment prepared according to the above method.

In order to solve the above technical problem, the present invention,

 a) adding ethanol at a concentration of 1-20% (v / v) to the herbaceous extract obtained from the herb;

b) adding 1-8 units of esterase or lipase fatty acid degrading enzyme per milligram of herbal extract to the mixture of step a);

c) reacting the mixture to which the fatty acid degrading enzyme is added at 20-30 ° C. in a sealed state; And

d) the reaction of step c) when the reaction is complete to remove ethanol to obtain the sikonin; siconin using fatty acid degrading enzyme, characterized in that extracting sikonin from fatty acid extract using fatty acid degrading enzyme Provides an extraction method.

And, a) adding ethanol with a concentration of 1-20% (v / v) to the ground powder obtained from the ground;

b) adding 1-8 units of esterase or lipase fatty acid degrading enzyme per gram of ground powder to the mixture of step a);

c) reacting the mixture to which the fatty acid degrading enzyme is added at 20-30 ° C. in a sealed state; And

d) the reaction of step c) when the reaction is complete to remove ethanol to obtain the sikonin; siconin using fatty acid degrading enzyme, characterized in that extracting sikonin from fatty acid extract using fatty acid degrading enzyme Provides an extraction method.

B) adding tris buffer of pH 6.8 to a final concentration of 10 mM to increase the activity of fatty acid degrading enzymes; It is characterized by including.

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Step d) is a step of filtering the reaction is completed, and then concentrated under reduced pressure for 10 minutes at 25 ℃-40 ℃ characterized in that the step of removing ethanol.

The d) step is characterized in that the step of removing the ethanol by filtering the reaction is completed and left for 1 hour at 50 ℃.

The step d) further comprises the step of adding and reacting the fatty acid degrading enzyme which is an esterase or lipase to the reaction is completed.

On the other hand, it provides an extract, characterized in that extracted by the above-described method.

And adding 1-10% by weight to the extract of the above-described method based on any one substance selected from liquor, beverages, pharmaceuticals, cosmetics and foods to obtain a substance containing a siconin extract. do.

In summary, by removing the acyl group (R portion) of the siconin derivatives (Formula 1 below) using esterase, only siconin, a highly polar and bright red color and low cytotoxic pigment, is selected. It provides a method for easy production, provides an extract containing such sikonin, and provides a method for using such an extract in a variety of materials.

Chemical Structure of Main Siconin Derivatives and Siconins

The esterase used in the present invention, along with lipase, is a representative fatty acid degrading enzyme, and its original action is to produce glycerol and fatty acids by hydrolyzing triglyceride, which is mainly triglyceride (Formula 2 below). It also acts on derivatives to break down acyl groups to produce siconins.

Formula 2. Actions of Fatty Acid Degrading Enzymes

First, as the extracting object used in the present invention, as shown in Figs. 1 and 2, both the organic solvent extract or the grass powder can be used. In this case, the organic solvent extract refers to an extract prepared by a general method. That is, by adding a solvent such as ethyl acetate, spirits or ethanol to extract the pigment of the ground, and then the extract is filtered and concentrated under reduced pressure to remove the solvent. To the grass extract or grass powder is added ethanol with a concentration of 5% (v / v) -20% (v / v), preferably 5% (v / v) ethanol. This ethanol is for the fatty acid degrading enzyme to react smoothly.

Subsequently, the extract is treated with 1-8 units of esterase, preferably 2 units, per 1 mg of extract, and, in the case of grass powder, 1-8 units of esterase, preferably 2 units, per 1 g of dried grass powder.

At this time, when the ethanol concentration is 30% (v / v) or more, the reaction rate is sharply reduced. In addition, in order to increase the activity of esterase, Tris buffer of pH 6.8 may be added so that the final concentration is 10 mM. As the buffer is added, the enzyme activity increases by about 10%. The reactants treated with esterase are sealed and reacted at room temperature, preferably at 25 ° C. for 2 days. In order to further increase the yield of sikonin, 2 units per mg extract of the extracts for lichen extract, and 2 units per 1 g of powder for lichen powder were further reacted for 2 days under the same conditions. The reaction product is filtered and then concentrated under reduced pressure at 35 ° C. for 10 minutes or left at 50 ° C. for 1 hour to remove ethanol, thereby obtaining an aqueous solution having a siconin content of 90% or more. In some cases, the same amount of ethyl acetate may be added to this aqueous solution to extract and concentrate the siconin to obtain a siconin extract. The composition thus prepared contains more than 90% of siconin and may be used for any harmless substance used in the human body such as food, beverage, food, pharmaceutical or cosmetics selected from beverages, alcohol, pharmaceuticals and cosmetics. .

At this time, the extract is preferably limited to 1-10% by weight based on the material. That is, 90 to 99% by weight of the substance: 1-10% by weight of the extract. This is because when the extract is more than 10% by weight may lose the properties of the material itself, when less than 1% by weight may reduce the performance of the extract.

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

Prior to this, terms or words used in the present specification and claims should not be construed as being limited to the ordinary or dictionary meanings, and the inventors should properly explain the concept of terms in order to best explain their own invention. Based on the principle that can be defined, it should be interpreted as meaning and concept corresponding to the technical idea of the present invention.

Therefore, the embodiments described in the specification and the drawings shown in the drawings are only the most preferred embodiments of the present invention, and do not represent all of the technical idea of the present invention, which can be replaced at the time of the present application It should be understood that there may be various equivalents and variations. In particular, as used herein, "unit" (unit) is a unit that defines the enzyme activity means a reaction amount of a specific substrate per unit time or the production amount of the product per unit time. One unit of an enzyme used in the present invention means an amount of enzyme (or enzymatic activity) capable of degrading (converting) 1 micromol of substrate in one minute under reference measurement conditions.

Hereinafter, exemplary embodiments of the present invention will be described in detail with reference to the accompanying drawings.

Example 1: Esterase  Used From grass extract Siconin  production

First, the siconin derivative is extracted from the ground using an organic solvent. That is, the dried liquorice root is immersed in a solvent, such as 5-15 times the weight of the lichen root, preferably 10 times or more ethyl acetate, ethanol or spirits, and left for approximately 24 hours. Subsequently, the ground roots are pulverized together with the solvent through a grinding apparatus to extract the siconin derivative, which is a red pigment contained in the ground roots. That is, by grinding the ground roots with the solvent so that the sikonin derivative is more easily extracted in the solvent. The extract thus obtained is filtered to obtain only a solvent layer containing a siconin derivative, and then concentrated under reduced pressure to remove the solvent, thereby finally obtaining a herbal extract. 5% (v / v) -20% (v / v) ethanol, preferably 5% (v / v) ethanol, was added 0.05-0.5 mL, preferably 0.1 mL per 1 mg of the extract to the extract. 1-8 units of esterase, preferably 2 units, per mg. The reason for processing 2 units of esterase per mg of extract is that this is the most economical concentration. In other words, up to 2 units of esterase per 1mg of extract, the activity is increased almost proportionally, but if it is more than that activity does not increase proportionally. For example, 1 unit of esterase per 8 mg of extract extracts 16.2% of siconin, and 20.8 units of esterase produces 30.8% of esterase. Only 49.9% is generated. At this time, when the ethanol concentration is 30% (v / v) or more, the reaction rate is drastically lowered. In some cases, in order to increase the activity of esterase, Tris buffer of pH 6.8 may be added so that the final concentration is 10 mM. . As the buffer is added, the enzyme activity of about 10% increases. The reactants treated with esterase are sealed and reacted at room temperature, preferably at 25 ° C. for 2 days. In addition, in order to increase the yield of the siconin further, the extract was treated with 2 units of esterase per 1 mg of extract, and further reacted for 2 days under the same conditions. The reaction product was filtered and concentrated under reduced pressure at 35 ° C. for 10 minutes using a vacuum concentrator or left at 50 ° C. in a shaking incubator for 1 hour to remove ethanol to obtain an aqueous solution having a siconin content of 90% or more. In some cases, the same amount of ethyl acetate may be added to this aqueous solution to extract and concentrate the siconin to obtain a siconin extract. The composition thus prepared contains more than 90% of siconin and may be used in various fields such as beverages, alcoholic beverages, pharmaceuticals, and the like.

Example 2: Esterase  Used From ground powder Siconin  extraction

First, prepare ground powder. The ground powder is powdered dried grass roots using a blender. The reason for powdering the grass is to easily elute the siconin derivative from the grass roots and to react with esterase well. To the ground powder, ethanol having a concentration of 5% (v / v) -20% (v / v), preferably 5% (v / v) ethanol, is added as in Example 1 above. At this time, the concentration of 5% (v / v) ethanol is enough to submerge the ground powder, preferably 10 mL per 1 g of the ground powder, and the esterase is treated with 1-8 units of esterase, preferably 2 units per 1 g of the ground powder. . The reactants treated with esterase are sealed and reacted at room temperature, preferably at 25 ° C. for 2 days. In order to further increase the yield of siconin, 2 units of esterase per 1 g of the ground powder may be further treated and reacted for 3 days under the same conditions. In this case, too, the ethanol concentration may not be higher than 30% (v / v) as in the case of Example 1, and the enzyme activity may be increased by adding a Tris buffer having a pH of 6.8 to a final concentration of 10 mM. The reactant is filtered to remove the powder, and then concentrated under reduced pressure at 35 ° C. for 10 minutes or left at 50 ° C. for 1 hour to remove ethanol to obtain an aqueous solution having a siconin content of 90% or more. In some cases, the same amount of ethyl acetate may be added to this aqueous solution to extract and concentrate the siconin to obtain a siconin extract. The composition thus prepared contains more than 90% of siconin and may be used in various fields such as beverages, alcoholic beverages, pharmaceuticals, and the like.

Hereinafter, the present invention will be described in more detail through the following experimental examples.

Experimental Example 1: Fatty Acid Degrading Enzyme Sikonin Generation  Search

Lipase and esterases, lipolytic enzymes, were tested for the ability to convert siconin derivatives to siconins. The fatty acid degrading enzymes used in the experiments were derived from Thermomyces lanuginosus (Lipozyme TL 100), Candida antarctica (Lipozyme CALB L), Candida rugosa- derived lipases, and pig liver from relatively low cost and low substrate specificity. A total of four fatty acid degrading enzymes, including one esterase, were selected and used. First, five kinds of siconin derivatives, such as isovaleryl siconin, acetylsiconin, dimethylacrylsiconin, isobutyl siconin, and betahydroxyisobareryl siconin, were 53.9%, 29.7%, 11.2%, and 3.2%, respectively. 10 mg per 1 mL of the reaction solvent was added to the extract of Siconin derivative containing 2.1% and reacted with four fatty acid degrading enzymes under the conditions shown in Table 1.

Table 1. Reaction Condition Using Fatty Acid Degrading Enzyme

enzyme Esterase Lipase (origin) Pig liver Candida rugosa Thermomyces lanuginosus (TL100) Candida antarctica (CALB L) Enzyme concentration (per 10 mg extract) 35unit 1000unit 1000unit 1000unit Reaction solution (1 mL) 10% (v / v) ethanol in 10mM Tris buffer (pH6.8) 10% (v / v) ethanol in 10mM Phosphate buffer (pH7.0) 10% (v / v) ethanol in 10 mM Phosphate buffer (pH 7.0) 10% (v / v) ethanol in 10mM Acetatebuffer (pH6.0) Reaction temperature 25 ℃ 40 ℃ 40 ℃ 40 ℃

Subsequently, 200 reaction products were taken every 2 hours, 4 hours, 6 hours, and 10 hours, and the mixture was added to the vial, and ethyl acetate 500 was added to extract siconin and siconin derivatives. The extracted ethyl acetate fraction was dehydrated by adding Na ₂ SO ₄ , filtered, and the solvent was removed through a rotary vacuum concentrator. Acetontrile 100 was dissolved in this sample, and the amount of siconin was confirmed through HPLC analysis (Table 2).

Table 2. HPLC Conditions for Siconin Content Analysis

ColumeMobile phaseFlow RateDetectorInjection volume   Phenomenex Luna 5u C18 Acetonitrile: 0.1% TFA = 75: 25 1.0mL / min UV 520nm 5μl

As a result, as shown in Table 3, in the case of esterase, 31.7% of siconin was produced after 10 hours even at a small amount of 2 units per mg of the siconin derivative. In comparison, other fatty acid degrading enzymes produced only 0.5 ~ 11.0% after 10 hours even when used in 1000 units, and it was confirmed that esterase had excellent siconin production ability.

Table 3. Siconin Production from Fatty Acid Degrading Enzymes

Siconin production amount (Siconin derivative amount)% Reaction time Esterase Lipase Pig liver Candida rugosa Thermomyces lanuginosus (TL100) Candida antarctica (CALB L) Before reaction 0 (100) 0 (100) 0 (100) 0 (100) 2 hours 19.7 (80.3) 0.9 (99.1) 0.2 (99.8) 0.7 (99.3) 4 hours 25.8 (74.2) 3.2 (96.8) 0.3 (99.7) 0.8 (99.2) 6 hours 27.9 (72.1) 5.0 (95.0) 0.4 (99.6) 0.9 (99.1) 10 hours 31.7 (68.3) 11.0 (89.0) 0.5 (99.5) 1.1 (98.9)

Experimental Example 2: Esterase  Find the optimal reaction condition

The optimal reaction conditions of esterases, which were found to be excellent in the ability to produce siconin from siconin derivatives, were examined.

(1) Investigation of Siconin Formation Capacity by Ethanol Concentration

First, the siconin production ability of esterases according to the ethanol concentration in the reaction solution was investigated. The reaction was carried out under the same conditions as in Experimental Example 1, that is, 10 mg of herb extract, 1 mL of 10 mM Tris buffer (pH 6.8), 20 units of esterase, and 5% alcohol (ethanol) concentration of the reaction solution. v), 10% (v / v), 20% (v / v), 30% (v / v), 40% (v / v), 50% (v / v) and then adjusted at 25 ° C The amount of siconin produced during each hour was examined.

As a result, as shown in Table 4 below, the yield of siconin was the best at 5% (v / v) ethanol concentration, and relatively good activity was also observed at 20% (v / v) ethanol concentration. It is thought that the activity of the enzyme decreases as the alcohol (ethanol) content (concentration) increases, but the activity increases as the frequency of reaction with the enzyme increases because the extraction amount of the siconin derivative increases. In addition, when the ethanol concentration is 30% (v / v) or more, it was thought that the enzyme is rapidly deactivated, reducing the yield of siconin.

table. 4. The amount of siconin produced by esterase according to ethanol concentration (%)

Reaction time Ethanol content (concentration)% (v / v) 5% 10% 20% 30% 40% 50% Before reaction 0 0 0 0 0 0 30 minutes 14.5 7.0 10.2 1.6 0.8 0.4 2 hours 31.1 19.7 24.0 4.5 1.2 0.9 4 hours 40.3 25.8 30.7 5.8 1.5 1.8 6 hours 45.1 27.9 35.4 7.8 1.9 1.8 10 hours 52.3 31.7 47.5 9.2 2.0 1.8

(2) investigation of siconin production capacity according to the amount of esterase added

In the above results, the ability to produce sikonin according to the amount of esterase added at 5% (v / v) ethanol concentration showed the best activity. The reaction was carried out under the same conditions as in the above experimental example, that is, the amount of esterase was added to 10 units and 20 units under the conditions of 10 mg of the grass extract and 1 mL of 10 mM Tris buffer (pH 6.8) containing 5% (v / v) ethanol. After adjusting to 40 units, 80 units and reacting at 25 ℃ the amount of sikonin produced by each time was investigated.

As a result, as shown in Table 5 below, the amount of esterase added up to 20 units per 10 mg of extract almost increased in proportion to activity, but above that, the activity did not increase proportionally. The addition amount was found to be 2 units per mg of extract.

 table. 5. Production amount of siconin according to the amount of esterase added (%)

Reaction time Esterase addition amount (per 10 mg of grass extract) 10unit 20unit 40unit 80unit Before reaction 0 0 0 0 30 minutes 4.3 8.5 10.6 12.7 2 hours 8.3 16.2 20.5 23.6 4 hours 12.4 23.7 30.2 34.8 6 hours 14.2 26.5 34.0 39.2 8 hours 16.2 30.8 40.1 45.9

Experimental Example 3: Sikonin  Maximize creation

Based on the results of Experimental Example 2, a method for maximizing siconin production was examined. As can be seen from the above results, the yield of siconin was best when 2 units of esterase per 1 mg of the extract were reacted in 0.1 mL of 10 mM Tris buffer (pH 6.8) containing 5% (v / v) ethanol. The hourly production of siconin increases rapidly up to the first 10 hours, but then increases slowly afterwards and hardly increases after 2 days. Therefore, 2 days after the addition of 2 units per mg extract was investigated the amount of sikonin produced. In addition, in order to commercialize the reaction product as it was, the amount of siconin produced in distilled water without Tris buffer was investigated.

As a result, as shown in Table 6, by adding 2 units of esterase per 1mg of extract after 2 days, it was confirmed that the amount of siconin produced could be increased to 90% or more. In addition, it was confirmed that the activity of the esterase was maintained in the distilled water state without the addition of Tris buffer. However, when distilled water was used, the amount of sikonin produced was reduced by 10%.

Table 6. Production of Siconin from Herb Extracts Using Esterase

In addition, the content of the siconin and the siconin derivatives of the final reactants obtained as a result of Experimental Example 3 is shown in Table 7 below.

Table 7. Contents of Siconin and Siconin Derivatives in Final Reactions of Herb Extracts by Esterase

Reaction condition   Tris buffer (pH6.8) Distilled water Total amount of esterase used (per extract mg) 2unit 4unit (2unit + 2unit) 2unit 4unit (2unit + 2unit) Sikonin 79.7 98.8 66.7 91.3 Betahydroxy isovareryl siconin 0.6 0.4 0.8 2.4 Acetyl Siconin 7.9 0.4 11.4 3.2 Isobutyl Siconin 2.4 0.1 7.4 1.0 Dimethyl Acrylic Siconin 5.8 0.3 5.6 2.1 Isovaleryl siconin 3.6 - 8.1 - Total content 100 100 100 100

Experimental Example 4: Esterase  Used From ground dry powder Sikonin  produce

In order to produce siconin more easily using esterase, a method of producing siconin using immediately lichen powder without going through solvent extraction process was examined. 10 g of dried lichen root powder was pulverized with a mixer, powdered, and then placed in an Erlenmeyer flask, and 100 mL of 10 mM Tris buffer (pH 6.8) was added. The amount of siconin produced was analyzed. At this time, adjust the amount of esterase added to 1 unit, 2 units, 4 units, 8 units per 1g of the ground powder or adjust the ethanol content (concentration) to 5% (v / v) or 20% (v / v). The reaction conditions were examined.

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As a result, as shown in Table 8 below, even when the ground powder was used, a large amount of siconin was produced by the activity of the esterase, and the addition amount of the esterase was determined to be the most economical 2 units per 1g of the ground powder. In addition, when the ethanol content (concentration) is also 5% (v / v), it was found that the best.

Table 8. Siconin Contents in Lime Powder Reactions According to Esterase and Ethanol Concentrations

Reaction time Esterase addition amount (per 1g of powder) Ethanol concentration% (v / v) 1unit 2unit 4unit 8unit 5% 20% 30 minutes 5.4 10.7 17.2 19.4 10.7 3.6 2 hours 12.4 23.0 41.6 34.7 23.0 9.4 4 hours 15.7 29.8 52.3 34.0 29.8 10.9 6 hours 17.1 33.4 48.8 43.8 33.4 11.9 10 hours 20.8 40.7 57.1 45.7 40.7 15.8

In addition, on the basis of the above results, a method for maximizing the production of siconin from the ground powder in the same manner as in Experiment 3 was examined.

Table 9. Production of Siconin from Grass Powder Using Esterase

As a result (Table 9), even when using the ground powder, it was found that the siconin production was maximized by additionally adding esterase after 2 days in the same manner as in Example 3. Through this method, about 5 mg of a composition containing 90% or more of siconin per gram of ground powder can be obtained. In addition, the content of siconin and siconin derivatives of the final reactants showed a similar pattern to the experimental results of the herbaceous extract.

Extraction method of siconin using fatty acid degrading enzyme according to the present invention, the extract and the method using the extract, as described above, by treating the esterase or one of the fatty acid degrading enzymes to the extract or grass powder only siconin The present invention relates to a method for selectively producing, according to the present invention, by removing the acyl group of the siconin derivatives by esterase, it is possible to extract (produce) sikonin in a large amount. In addition, the pigments extracted by this method can be dissolved in low alcohol, have a bright red color, and have low cytotoxicity, which will increase the applicability and application of conventionally available lipo pigments. There is an effect that can be usefully used in various fields such as food and pharmaceuticals, such as low-escape manufacturing.

As described above, although the present invention has been described by way of limited embodiments and drawings, the present invention is not limited thereto, and the present invention is provided by the person skilled in the art to which the present invention pertains. Of course, various modifications and variations are possible within the scope of equivalents of the claims to be described.

Figure 1 is a flow chart for explaining the method of producing weed sikonin according to an embodiment of the present invention.

Figure 2 is a flow chart for explaining the method of producing ground sikonin according to another preferred embodiment of the present invention.

Claims (14)

a) adding ethanol at a concentration of 1-20% (v / v) to the herbaceous extract obtained from the herb; b) adding 1-8 units of esterase or lipase fatty acid degrading enzyme per milligram of herbal extract to the mixture of step a); c) reacting the mixture to which the fatty acid degrading enzyme is added at 20-30 ° C. in a sealed state; And d) removing the ethanol when the reaction of step c) completes the reaction to obtain the sikonin; the sikonin using fatty acid degrading enzyme, characterized in that to extract the sikonin from fatty acid extract using fatty acid degrading enzyme Extraction method. a) adding 1-20% (v / v) ethanol to the ground powder obtained from the ground; b) adding 1-8 units of esterase or lipase fatty acid degrading enzyme per gram of ground powder to the mixture of step a); c) reacting the mixture to which the fatty acid degrading enzyme is added at 20-30 ° C. in a sealed state; And d) sikonin using fatty acid degrading enzyme, characterized in that to extract the sikonin from fatty acid extract using fatty acid degrading enzyme, including the step of removing the ethanol when the reaction of step c) is complete Extraction method. delete The method according to claim 1 or 2, wherein step b) comprises the steps of adding a tris buffer of pH 6.8 to a final concentration of 10mM in order to increase the activity of fatty acid degrading enzymes; Method of extracting sikonin using fatty acid degrading enzyme, characterized in that it comprises. The method according to claim 1 or 2, wherein the step d) is a step of filtering the reaction is completed, the concentration of siconin using a fatty acid degrading enzyme, characterized in that the step of concentration under reduced pressure at 25 ℃-40 ℃ 10 minutes to remove ethanol Extraction method. The method according to claim 1 or 2, wherein the step d) is a method of extracting sikonin using a fatty acid degrading enzyme, characterized in that ethanol is removed by filtering the reactant after completion of the reaction and standing at 50 ° C for 1 hour. The method according to claim 4, wherein the step d) is a step of filtering ethanol to remove the ethanol by filtering the reactant after completion of the reaction and leaving it at 50 ° C for 1 hour. The method according to claim 1 or 2, wherein the step d) further comprises the step of reacting by adding a fatty acid degrading enzyme which is an esterase or lipase to the reaction is complete reaction of the fatty acid degrading enzyme using a fatty acid degrading enzyme Extraction method. delete delete delete delete The extract extracted according to claim 1 or 2 is added to 1-10% by weight based on any one selected from alcohol, beverages, pharmaceuticals, cosmetics and foods to obtain a substance containing a siconin extract Way. The method of claim 4, wherein 1-10% by weight of the extract extracted from liquor, beverages, pharmaceuticals, cosmetics and foods is added to obtain a substance containing a siconin extract.