KR102154148B1 - A composition for immune regulation or enhancement comprising Dendropanax morbifera Lev. extract or an active ingredient isolated therefrom - Google Patents

Abstract

The present invention relates to a composition for immunomodulation or immunity enhancement comprising an extract of Hwangchil tree leaf ( Dendropanax morbifera Lev. leaf), a fraction thereof, or an immune active component isolated therefrom. The extracts, fractions, and immune-active components isolated therefrom of the present invention increase the proliferative ability of splenocytes and the production of IL-2 and IFN-r cytokines. In particular, the ethyl acetate fraction has the effect of proliferating CD4 ⁺ T cells and EL-4 T cells. In addition, DMW-2 single component isolated from ethyl acetate fraction significantly increases IL-2 cytokine production, and transcription factors IL-2, IL-4, NF-AT, AP-1, and NF are involved in T cell proliferation. -kB activation effect is very good.

Description

A composition for immune regulation or enhancement comprising Dendropanax morbifera Lev. Hwangchil tree extract or an active ingredient isolated therefrom. extract or an active ingredient isolated therefrom}

The present invention relates to a composition for immunomodulation or immunity enhancement comprising a hwangchil tree extract. In addition, the present invention relates to a composition for immunomodulation or immunity enhancement, comprising as an active ingredient DMW-2 or salts thereof, an active ingredient isolated from Hwangchil tree leaf extract.

In general, the human body protects the body and maintains homeostasis through immunity to identify and eliminate the invasion of an external pathogen. Immune response is a series of reactions caused by activated immune cells to antigens. The immune system recognizes infection during infection and plays a role in removing pathogens through humoral and cellular responses. When the immune system fails to function properly, the immune system is infectious. It induces various diseases as well as diseases.

Immunity refers to self-defense by defending the host against the antigen and removing the antigen, and is largely divided into innate immunity and acquired immunity. Innate immunity is a non-specific reaction to an antigen and causes an immune response by involving macrophages, natural killer cells, and complement. Acquired immunity remembers the antigen that first invaded, and when the antigen invades again, it triggers a specific immune response to remove the antigen. Acquired immunity is divided into humoral immunity and cellular immunity. In humoral immunity, B lymphocytes are involved in recognizing antigens and then secreting antibodies to trigger an immune response. In cellular immunity, T lymphocytes derived from the thymus gland recognize antigens and secrete cytokines or activate macrophages to induce an immune response.

In particular, in the immune response by T cells, an initial signal is transmitted into the cell from another signal and a TCR complex of T cells that recognize the antigen. This signal transduction process appears by the action of the PKC activator and ionophore, increasing Ca ⁺⁺ ions in the cytoplasm of T cells. In addition, the signal is eventually transmitted to the nucleus through the process of phosphorylation of various proteins. This initial signal transmitted from the cell membrane to the nucleus appears in the nucleus by the expression of various genes required for proliferation and differentiation of T cells. This process is called transcriptional activation, and transcription of NF-AT, NF-kB and AP-1, which plays a major role in T cell proliferation, occurs. In addition, transcription of the IL-2 cytokine and the IL-2 receptor gene occurs. Through this, new cytokines are secreted, and T cells proliferate and differentiate. T cell activity is linked to the synthesis of IL-2, and secreted IL-2 induces the proliferation of T cells. IL-2 is a growth factor of T cells, which activates immune cells such as T cells, B cells, and natural killer cells to exhibit an immune response. IL-2 induces various intracellular responses by binding to IL-2R present on the surface of T cells. When IL-2 binds to IL-2R of T cells, differentiation of T cells occurs, and cytokines such as IFN-r (interferon-gamma) and TNF (tumor necrosis factor) are released, thereby affecting the immune response. It also increases the secretion of IL-1, IL-6, and TNF from monocytes.

However, when this immune function is deteriorated, an impairment in immune regulation occurs due to an imbalance of cytokines in the body and an imbalance of proliferation of T/B cells. As a result, various immune diseases such as asthma, rhinitis, allergic rhinitis, atopic dermatitis, irritable dermatitis, colds, and chronic fatigue can occur. Therefore, the immune response of the body must be balanced and regulated normally to maintain health, so immune regulation is more important than treatment. However, the currently marketed therapeutic agent is an immunity enhancer with the potential for side effects to be taken for prevention, and thus a substance capable of preventing and treating immune dysfunction is required.

To this end, research is being actively conducted to find a component capable of regulating immune activity from natural products with few side effects, and to develop a formulation capable of regulating immune activity by using the same. Republic of Korea Patent Publication No. 10-2015-0051174 discloses a composition for enhancing immunity comprising an alcohol extract of elm bark as an active ingredient, and Korean Patent No. 10-1536652 discloses a composition for enhancing immunity comprising turmeric extract Is disclosed.

From ancient times, the Hwangchil tree ( Dendropanax morbifera Lev. ) is a genus of the Arlyaceae ( Ogalpiaceae ) Hwangchil tree. As a species of Korean endemic, it is an evergreen large creeping tree that grows in the beaches of Jeollanam-do and the beaches and islands of Jeju-do and in the shady and humid broad-leaved forests or shrubs on the slopes of the mountain. The leaves have no hairs on the surface, are smooth, alternate, oval or oval, and the body of the leaves does not split at all, or there are some that are deeply split like fingers in 3-5 branches. Looking at the distribution map of the Hwangchil tree in Korea, it grows a lot in the coastal areas such as Jeju Island, Jeonnam Wando, Daeheuksan Island, and Geomun Island. The generic name of Hwangchil, Dendropanax , is a compound word of the Greek words dendro (tree) and panax (all-powered medicine). The scientific name of the hwangchil tree, dendropanax mobifer, means a tree of panacea, and is also called tree ginseng. In addition, the name Hwangchil is named because a yellow liquid comes out like lacquer lacquer when a wound is made to the bark of Hwangchil. Hwangchil is used to process or color the surface in the process of making crafts, and the emperor's clothes, Gonryongpo and Najeon It has been used in various ways such as lacquerware, Goryeo Buddha, and various furniture and metals. According to the Korean herbal book, the roots and stems of Hwangchil trees are sweet in taste and warm in nature, and are effective in geopung habits and hwanghyeolmaek.

In addition, according to recent studies, a single ingredient in Hwangchil and Hwangchil has been used as a material with more interest as pharmacological effects such as antibacterial, anticancer, hyperlipidemia, antithrombotic anti-inflammatory effect and antioxidant effect have been revealed. Regarding the immunity enhancing effect of Hwangchil, Korean Patent Application Laid-Open No. 10-2015-0144061 discloses a composition for immunomodulation or immunity enhancement containing Hwangchil tree stem extract as an active ingredient. However, the immunomodulatory effect of single components in Hwangchil tree has not been accurately reported.

Research results showing that the fraction of Hwangchil tree and a single component of Hwangchil tree genus help proliferation of T cells related to acquired immunity and increase immune activity are insignificant. Accordingly, the inventors of the present invention attempted to confirm whether Hwangchil wood is helpful in enhancing immunity by activating the immune function through the proliferation of T cells.

The present inventors have made intensive research efforts to develop a plant-derived immune activating composition for immune regulation or immunity enhancement. As a result, the present inventors have developed a composition for immunomodulation or immunity enhancement comprising DMW-2, an active substance isolated from Hwangchil tree leaf extract or Hwangchil tree leaf extract.

The present inventors found that the ethyl acetate fraction isolated from the Hwangchil tree leaf extract and extract activates the proliferation of T cells, increases the transcription factors involved in T cell proliferation, NF-AT, AP-1, NF-kB, and increases IL-2 cytokine production. As an effect, it was found that it is very effective in immunomodulation or immunity enhancement.

On the other hand, the compound isolated from the ethyl acetate fraction, DMW-2 ((10E)-(-)-10,17-octadecadiene-12,14-diin-1,9,16-triol) is a T cell It does not activate proliferation, but significantly increases IL-2 cytokine production. In addition, the effect of activating the transcription factors IL-2, IL-4, NF-AT, AP-1, and NF-kB involved in T cell proliferation was very excellent.

Accordingly, an object of the present invention is to provide a composition for immunomodulation or immunity enhancement.

Another object of the present invention is to provide a composition for enhancing antitumor immunity or an anticancer composition.

Another object of the present invention is to provide a method for separating (10E)-(-)-10,17-octadecadiene-12,14-diine-1,9,16-triol compounds.

One aspect of the present invention relates to a composition for immunomodulation or immunity enhancement comprising an extract of Hwangchil tree leaf ( Dendropanax morbifera Lev. leaf).

The present inventors have made intensive research efforts to develop a plant-derived immune activating composition for immune regulation or immunity enhancement. As a result, the present inventors have developed a composition for immunomodulation or immunity enhancement comprising DMW-2, an active substance isolated from Hwangchil tree leaf extract or Hwangchil tree leaf extract.

The present inventors believe that Hwangchil tree leaf extract and ethyl acetate fraction isolated from this extract activate T cell proliferation, increase transcription factors NF-AT, AP-1, NF-kB, and IL-2 cytokine involved in T cell proliferation. As an increasing effect, it was found that it is very effective in immunomodulation or immunity enhancement.

On the other hand, the immune enhancing active substance isolated from the ethyl acetate fraction is as follows.

DMW-2: (10E)-(-)-10,17-Octadecadiene-12,14-diyne-1,9,16-triol; (10E)-(-)-10,17-octadecadiene-12,14-diine-1,9,16-triol.

DMW-2 does not activate T cell proliferation, but significantly increases IL-2 cytokine production. In addition, the effect of activating the transcription factors IL-2, IL-4, NF-AT, AP-1, and NF-kB involved in T cell proliferation was excellent.

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

Hwangchil tree leaf extract of the present invention, after drying and slicing hwangchil tree leaf ( Dendropanax morbifera Lev.leaf ), and adding a polar solvent (eg, distilled water, methanol, ethanol, propanol, butanol, etc.) as an extraction solvent, 80 It can be extracted by heating to ℃-120 ℃. According to one embodiment of the present invention, it may be extracted by heating at 80°C-120°C for 2-8 hours.

The polar solvent used in the extraction method of the present invention is (i) water, (ii) alcohol (preferably, methanol, ethanol, propanol, butanol, normal-propanol, iso-propanol, normal-butanol), (iii) Acetic acid, (iv) dimethyl-formamide (DMFO), and (v) dimethyl sulfoxide (DMSO). According to one embodiment of the present invention, the polar solvent is one or more solvents selected from the group consisting of water and lower alcohols having 1 to 3 carbon atoms. According to another embodiment of the present invention, the polar solvent is water, methanol or ethanol.

According to one embodiment of the present invention, it can be extracted for 3-5 hours at 90° C.-110° C. by adding a polar solvent to hwangchil tree leaves.

According to a specific embodiment of the present invention, it may be extracted for 3-5 hours at 95°C-105°C by adding a polar solvent to hwangchil tree leaves.

According to a specific embodiment of the present invention, it can be extracted for 4 hours at 95°C-105°C by adding a polar solvent to hwangchil tree leaves.

In the present invention, when extracting hwangchil tree leaves, hot water extraction, reflux extraction, room temperature extraction, or ultrasonic extraction may be used. According to one embodiment of the present invention, a hot water extraction method or a reflux extraction method may be used.

According to a specific embodiment of the present invention, hwangchil tree leaves were extracted with distilled water, filtered, concentrated under reduced pressure, and dried.

Meanwhile, the composition of the present invention may include an ethyl acetate extract fraction obtained by purifying only an extract soluble in an ethyl acetate solvent from a water extract or an alcohol extract from Hwangchil tree leaf.

For example, after adding water of about 0.001 to 100 times, preferably 1 to 50 times volume (v/w%) of the weight of Hwangchil tree leaf water extract or alcohol extract, hexane, chloroform, or ethyl acetate is used. By performing a fractionation process, a non-polar solvent-soluble extract fraction soluble in a non-polar solvent can be obtained.

Then, a polar solvent-soluble extract fraction soluble in a polar solvent such as butanol and water can be obtained.

The present inventors confirmed that the water extract of Hwangchil tree leaf and five fractions (hexane, chloroform, ethyl acetate, butanol and water fraction) activated T cell proliferation through an EL-4 T cell proliferation reactivity measurement experiment. Among them, it was confirmed that the ethyl acetate fraction was very excellent in proliferation of CD4 ⁺ T cells and EL-4 T cells.

The composition of the present invention may be implemented as a pharmaceutical composition or a food composition comprising a hwangchil tree leaf extract or a fraction thereof as an active ingredient.

Another aspect of the present invention relates to a composition for immunomodulatory or immunity enhancement comprising at least one compound selected from the group consisting of a compound represented by the following Formula 1 or salts thereof:

[Formula 1]

.

.

The compound of Formula 1 isolated from the ethyl acetate fraction of the above-described Hwangchil tree leaf ( Dendropanax morbifera Lev. leaf) extract did not activate T cell proliferation, but significantly increased IL-2 cytokine production. In addition, the effect of activating the transcription factors IL-2, IL-4, NF-AT, AP-1, and NF-kB involved in T cell proliferation was excellent. Based on this, the composition of the present invention can be used as an immunomodulatory or immunity enhancing composition comprising the compound of Formula 1.

The compound names of Formula 1 are as follows:

DMW-2: (10E)-(-)-10,17-Octadecadiene-12,14-diyne-1,9,16-triol; (10E)-(-)-10,17-octadecadiene-12,14-diine-1,9,16-triol.

The upper limit of the quantity of the compound or salts thereof included in the composition of the present invention can be selected and carried out by a person skilled in the art within an appropriate range.

The composition of the present invention is a drug containing (10E)-(-)-10,17-octadecadiene-12,14-diine-1,9,16-triol, or salts thereof as an active ingredient It can be implemented as a pharmaceutical composition or a food composition.

In the present specification, the term "comprising as an active ingredient" means including an amount sufficient to achieve the efficacy or activity of Hwangchil tree leaf extract, a fraction thereof, a compound of Formula 1 isolated from the fraction, or salts thereof do. Hwangchil tree leaf extract contained in the composition of the present invention, its fraction or the upper limit of the quantity of the compound can be carried out by selecting within an appropriate range by a person skilled in the art.

The compound used as an active ingredient in the present invention may be used by itself or in the form of salts, preferably in the form of a pharmaceutically acceptable salt. As the salt, an acid addition salt formed by free acid is preferable. Organic acids and inorganic acids may be used as the free acid. The organic acids include citric acid, acetic acid, lactic acid, tartaric acid, maleic acid, fumaric acid, formic acid, propionic acid, oxalic acid, trifluoroacetic acid, benzoic acid, gluconic acid, metasulfonic acid, glycolic acid, succinic acid, 4-toluenesulfonic acid, glutanoic acid and aspartic acid, etc. Includes, but is not limited to. In addition, the inorganic acids include, but are not limited to, hydrochloric acid, bromic acid, sulfuric acid, and phosphoric acid.

(10E)-(-)-10,17-octadecadiene-12,14-diin-1,9,16-triol contained in the composition of the present invention is a natural plant material, Hwangchil leaves ( Dendropanax morbifera Lev. leaf). Hwangchil tree leaves have long been used as edible and folk medicine Since it came, the compound of the present invention isolated therefrom can also be expected to have no problems such as toxicity and side effects. Thus, (10E)-(-)-10,17-octadecadiene-12,14-diin-1,9,16-triol, or salts thereof, can be used as a pharmaceutical or food composition.

According to one embodiment of the present invention, the composition of the present invention may be a pharmaceutical composition.

The pharmaceutical composition of the present invention may contain a pharmaceutically acceptable carrier. The pharmaceutically acceptable carrier is commonly used in the formulation, lactose, dextrose, sucrose, sorbitol, mannitol, starch, gum acacia, calcium phosphate, alginate, gelatin, calcium silicate, microcrystalline cellulose, Polyvinylpyrrolidone, cellulose, water, syrup, methyl cellulose, methylhydroxybenzoate, propylhydroxybenzoate, talc, magnesium stearate, mineral oil, and the like, but are not limited thereto. The pharmaceutical composition of the present invention may further include a lubricant, a wetting agent, a sweetening agent, a flavoring agent, an emulsifying agent, a suspending agent, a preservative, and the like in addition to the above components. Suitable pharmaceutically acceptable carriers and formulations are described in detail in Remington's Pharmaceutical Sciences (19th ed., 1995).

The pharmaceutical composition of the present invention may be administered orally or parenterally, and in the case of parenteral administration, it may be administered by intravenous injection, subcutaneous injection, intramuscular injection, intraperitoneal injection, transdermal administration, or the like.

A suitable dosage of the pharmaceutical composition of the present invention varies depending on factors such as formulation method, mode of administration, age, weight, sex, pathological condition, food, administration time, route of administration, excretion rate and response sensitivity of the patient, Usually, the skilled practitioner can readily determine and prescribe the dosage effective for the desired treatment or prophylaxis. According to a preferred embodiment of the present invention, the daily dosage of the pharmaceutical composition of the present invention is 0.001-10000 mg/kg.

The pharmaceutical composition of the present invention is prepared in unit dosage form by formulating using a pharmaceutically acceptable carrier and/or excipient according to a method that can be easily carried out by a person having ordinary knowledge in the art. Or it can be made by incorporation into a multi-dose container. At this time, the formulation may be in the form of a solution, suspension, or emulsion in an oil or aqueous medium, or may be in the form of an extract, powder, granule, tablet or capsule, and may additionally include a dispersant or a stabilizer.

According to another embodiment of the present invention, the composition of the present invention may be a food composition.

When the composition of the present invention is a food composition, it may be prepared in the form of powder, granules, tablets, capsules or beverages. For example, there are various foods such as candy, beverages, gum, tea, vitamin complexes, or health supplement foods.

The food composition of the present invention is Hwangchil tree leaf ( Dendropanax morbifera Lev.leaf ) extract, ethyl acetate fraction, (10E)-(-)-10,17-octadecadiene-12,14-diin-1, 9,16-triol, or its salts, as well as ingredients that are commonly added in food preparation, include, for example, proteins, carbohydrates, fats, nutrients, flavoring agents and flavoring agents. do. Examples of the aforementioned carbohydrates include monosaccharides such as glucose, fructose, and the like; Disaccharides such as maltose, sucrose, oligosaccharides, and the like; And polysaccharides, for example, common sugars such as dextrin and cyclodextrin, and sugar alcohols such as xylitol, sorbitol, and erythritol. As flavoring agents, natural flavoring agents [taumatin, stevia extract (eg, rebaudioside A, glycyrrhizin, etc.]) and synthetic flavoring agents (saccharin, aspartame, etc.) can be used. For example, when the food composition of the present invention is prepared as a drink, in addition to the extract or compound of the present invention, citric acid, liquid fructose, sugar, glucose, acetic acid, malic acid, fruit juice, headworm extract, jujube extract, licorice extract, etc. I can.

Another aspect of the present invention relates to an immunomodulatory or immunity enhancing method comprising administering to a subject an immunomodulatory or immunity enhancing composition comprising an extract of Hwangchil tree leaf ( Dendropanax morbifera Lev. leaf). .

Another aspect of the present invention is in the group consisting of (10E)-(-)-10,17-octadecadiene-12,14-diine-1,9,16-triol, and salts thereof. It relates to a method for immunomodulation or enhancement of immunity, comprising administering to a subject a composition comprising at least one compound of choice.

As used herein, the term "administering" means providing a given substance to a subject in any suitable way. The route of administration of the composition for treating, inhibiting or improving skin pigmentation of the present invention may be administered orally or parenterally through any general route as long as it can reach a target tissue. In addition, the composition of the present invention may be administered using any device capable of delivering an active ingredient to target cells.

In the present specification, the term "subject" is not particularly limited, but for example, human, monkey, cow, horse, sheep, pig, chicken, turkey, quail, cat, dog, mouse, mouse, rabbit, or It includes guinea pigs, preferably a mammal, more preferably a human.

Another aspect of the present invention relates to a composition for enhancing antitumor immunity comprising an extract of Hwangchil tree leaves ( Dendropanax morbifera Lev. leaf).

Another aspect of the present invention is (10E)-(-)-10,17-octadecadiene-12,14-diine-1,9,16-triol ((10E)-(-)-10, It relates to a composition for enhancing antitumor immunity, including 17-Octadecadiene-12,14-diyne-1,9,16-triol) or salts thereof.

The extracts, fractions, and immune-active components isolated therefrom of the present invention increase the proliferative capacity of splenocytes and the production of IL-2 and IFN-γ cytokines. In particular, the ethyl acetate fraction has the effect of proliferating CD4 ⁺ T cells and EL-4 T cells. In addition, DMW-2 single component isolated from ethyl acetate fraction significantly increases IL-2 cytokine production, and transcription factors IL-2, IL-4, NF-AT, AP-1, and NF are involved in T cell proliferation. The -kB activation effect was very good.

IL-2 is a T-cell growth factor that activates immune cells such as T cells, B cells, NK cells, or lymphokine-activated killer cells to enhance immunity and have anti-cancer effects. Represents.

Another aspect of the present invention relates to an anticancer composition comprising an extract of Hwangchil tree leaf ( Dendropanax morbifera Lev. leaf).

Another aspect of the present invention is (10E)-(-)-10,17-octadecadiene-12,14-diine-1,9,16-triol ((10E)-(-)-10, It relates to an anti-cancer composition comprising 17-Octadecadiene-12,14-diyne-1,9,16-triol) or salts thereof.

The cancer is stomach cancer, lung cancer, breast cancer, ovarian cancer, liver cancer, bronchial cancer, nasopharyngeal cancer, laryngeal cancer, pancreatic cancer, bladder cancer, colon cancer, colon cancer, cervical cancer, brain cancer, prostate cancer, bone cancer, head and neck cancer, skin cancer, kidney cancer, drainage It includes polyploid carcinoma, thyroid cancer, parathyroid cancer and ureter cancer.

The antitumor immunity enhancement or anticancer composition of the present invention can be administered orally or parenterally, and in the case of parenteral administration, intravenous injection, subcutaneous injection, intramuscular injection, intraperitoneal injection, transdermal administration, intratumorally administration, etc. Can be administered.

Anti-tumor immunity enhancement or anti-cancer composition of the present invention is the above-described present invention Hwangchil tree leaf ( Dendropanax morbifera Lev. leaf) extract or (10E)-(-)-10,17-octadecadiene-12,14-di Phosphorus-1,9,16-triol ((10E)-(-)-10,17-Octadecadiene-12,14-diyne-1,9,16-triol) is used, and the common content between the two is In order to avoid excessive complexity of the present specification, the description thereof is omitted.

Another aspect of the present invention relates to a method for separating (10E)-(-)-10,17-octadecadiene-12,14-diine-1,9,16-triol compound comprising the following steps: will be:

(a) extracting Hwangchil tree leaves ( Dendropanax morbifera Lev. leaf) with a polar solvent;

(b) purifying only the extract soluble in ethyl acetate solvent from the Hwangchil tree leaf extract of step (a) to obtain an ethyl acetate extract fraction; And

(c) Purifying the ethyl acetate extract fraction to obtain (10E)-(-)-10,17-octadecadiene-12,14-diine-1,9,16-triol.

Hereinafter, a method for separating the (10E)-(-)-10,17-octadecadiene-12,14-diine-1,9,16-triol compound will be described in detail.

Step (a): Hwangchil tree leaves ( Dendropanax morbifera Lev. leaf) extract preparation

First, it is extracted by adding a polar solvent to Hwangchil tree leaves ( Dendropanax morbifera Lev. leaf).

The method of obtaining the hwangchil tree leaf extract is as described above.

According to one embodiment of the present invention, the polar solvent is water or a low-hydric alcohol having 1 to 3 carbon atoms.

According to a specific embodiment of the present invention, water or methanol is added to Hwangchil tree leaves and heated to 80°C-120°C to obtain an extract.

Step (b): Preparation of ethyl acetate extract fraction

Subsequently, by purifying only the extract soluble in ethyl acetate solvent from the Hwangchil tree leaf extract of step (a), an ethyl acetate extract fraction was obtained.

According to an embodiment of the present invention, after adding water of about 0.001 to 100 times, preferably 1 to 70 times, more preferably 50 times volume (v/w%) of the weight of Hwangchil tree leaf extract, hexane, chloroform , A non-polar solvent-soluble extract fraction soluble in a non-polar solvent can be obtained by performing a conventional fractionation process using a non-polar solvent such as ethyl acetate.

The non-polar solvent used in the separation method of the present invention is hexane, chloroform, ethyl acetate, acetone, acetonitrile, methyl acetate, fluoroalkane, pentane, 2,2,4-trimethylpentane, decane, cyclohexane, cyclopentane. , Diisobutylene, 1-pentene, 1-chlorobutane, 1-chloropentane, o-xylene, diisopropyl ether, 2-chloropropane, toluene, 1-chloropropane, chlorobenzene, benzene, diethyl ether , Diethyl sulfide, dichloromethane, 1,2-dichloroethane, aniline, diethylamine, ether, carbon tetrachloride, and THF, but are not limited thereto. According to an embodiment of the present invention, the non-polar solvent is hexane, chloroform, ethyl acetate, or dichloromethane. According to a specific embodiment of the present invention, in step (b), (i) hexane, chloroform, and ethyl acetate are sequentially used as a non-polar organic solvent to obtain a partitioned fraction; Or (ii) dichloromethane and ethyl acetate are sequentially used to obtain a partitioned fraction.

On the other hand, from the Hwangchil tree leaf extract remaining after obtaining a fraction using a non-polar solvent, a polar solvent-soluble extract fraction soluble in a polar solvent (eg, butanol, water, etc.) can be obtained.

The polar solvent used in the separation method of the present invention is (i) water, (ii) alcohol (preferably, methanol, ethanol, propanol, butanol, normal-propanol, iso-propanol, normal-butanol, 1-pentanol. , 2-butoxyethanol or ethylene glycol), (iii) acetic acid, (iv) DMFO (dimethyl-formamide) and (v) DMSO (dimethyl sulfoxide), but are not limited thereto. According to one embodiment of the present invention, the polar solvent is water or alcohol. According to a specific embodiment of the present invention, fractions of a butanol layer and an aqueous layer were obtained after adding butanol to the Hwangchil tree leaf extract.

According to an embodiment of the present invention, step (b) may be carried out as follows:

(b-1) separating and removing the extract fraction containing only the extract soluble in at least one non-polar solvent selected from the group consisting of dichloromethane, hexane, and chloroform from the hwangchil tree leaf extract of step (a); And

(b-2) Separating the extracted fraction of step (b-1) and from the remaining hwangchil tree leaf extract

Obtaining an ethyl acetate extraction fraction containing only the extract soluble in ethyl acetate solvent.

Step (c): Purification of (10E)-(-)-10,17-octadecadiene-12,14-diine-1,9,16-triol

Then, the ethyl acetate extracted fraction obtained in step (b) is purified to obtain (10E)-(-)-10,17-octadecadiene-12,14-diine-1,9,16-triol.

According to an embodiment of the present invention, in the purification step (c), various known purification methods may be used, for example, column chromatography and thin layer chromatography may be used.

According to an embodiment of the present invention, after loading the ethyl acetate extracted fraction of step (b) onto a column, column chromatography purification may be performed using a solvent gradient of a non-polar solvent-polar solvent.

According to an embodiment of the present invention, a purified ethyl acetate fraction may be obtained using a solvent gradient of chloroform-methanol.

The purified ethyl acetate fraction can be separated from compound DMW-2 by repeating column chromatography using a non-polar solvent as a mobile phase. According to an embodiment of the present invention, the purified ethyl acetate fraction was repeatedly subjected to silica gel and YMC RP-18 column chromatography to separate compound DMW-2.

According to one embodiment of the present invention, hexane-acetone may be used as a non-polar organic solvent.

Meanwhile, the purified ethyl acetate fraction was prepared by repeating column chromatography using a polar solvent as a mobile phase to obtain compound DMW-1 (methyl(10E)-9,16-dihydroxyoctadeca-10,17-dien-12,14-diynoate). Can be separated. According to an embodiment of the present invention, the purified ethyl acetate fraction was repeatedly subjected to YMC RP-18 column chromatography to isolate compound DMW-1.

According to an embodiment of the present invention, methanol-water may be used as a polar organic solvent.

The polar solvent or non-polar solvent usable in step (c) is as described above.

The features and advantages of the present invention are summarized as follows:

The present invention relates to a composition for immunomodulation or immunity enhancement comprising an extract of Hwangchil tree leaf ( Dendropanax morbifera Lev. leaf), a fraction thereof, or an immune active component isolated therefrom.

The extracts, fractions, and immune-active components isolated therefrom of the present invention increase the proliferative capacity of splenocytes and the production of IL-2 and IFN-γ cytokines.

In particular, the ethyl acetate fraction has the effect of proliferating CD4 ⁺ T cells and EL-4 T cells. In addition, DMW-2 single component isolated from ethyl acetate fraction significantly increases IL-2 cytokine production, and transcription factors IL-2, IL-4, NF-AT, AP-1, and NF are involved in T cell proliferation. -kB activation effect is very good.

Figure 1 shows the effect on the proliferation ability of T cells in the mouse spleen after administration of Hwangchil tree leaf extract for 3 weeks and the degree of IL-2 and IFN-γ cytokine production.
Figure 2a shows the effect of Hwangchil tree leaf extract and fractions on EL-4 T cell proliferation ability.
Figure 2b-2c shows the effect of the hwangchil tree leaf extract and fractions on the proliferation ability of splenocytes or mouse CD4 ⁺ T cells, respectively.
W-Ex: Hwangchil tree leaf extract
W-Hex: n-hexane fraction obtained from Hwangchil tree leaf extract
W-CH: Chloroform fraction obtained from Hwangchil tree leaf extract
W-EA: Ethyl acetate fraction obtained from Hwangchil tree leaf extract
W-Bu: n-butanol fraction obtained from Hwangchil tree leaf extract
WW: Water fraction except W-Hex, W-CH, W-EA, and W-Bu
Figure 3a shows the structure of DMW-2, a single component in Hwangchil tree leaf extract (W-Ex) and ethyl acetate fraction (W-EA).
Figure 3b-3c shows the LC/MS/MS analysis results of DMW-2, a single component in Hwangchil tree leaf extract W-Ex) and ethyl acetate fraction (W-EA).
Figure 4a shows the effect of Hwangchil tree leaf extract (W-Ex), ethyl acetate fraction (W-EA), and DMW-2, a single component in the hwangchil tree leaf extract, on T cell proliferation ability.
Figure 4b-4c is Hwangchil tree leaf extract (W-Ex), ethyl acetate fraction (W-EA), and DMW-2, a single component in the hwangchil tree leaf extract, produces IL-2 and IFN-γ cytokines in T cells Indicate the impact on the level.
Figures 5a-5e show the effect of a single component in Hwangchil tree leaf extract on activation of transcription factors NF-AT, AP-1, NF-kB and IL-2, and IL-4 involved in T cells.

Hereinafter, the present invention will be described in more detail by the following examples. However, these examples are for illustrative purposes only, and the scope of the present invention is not limited by these examples.

Throughout the present specification, "%" used to indicate the concentration of a specific substance is (weight/weight)% for solids/solids, (weight/volume)% for solids/liquids, and Liquid/liquid is (vol/vol) %.

Example 1: Preparation of Hwangchil tree leaf extract

Hwangchil tree leaves were purchased and used from the origin of Jangheung, Jeollanam-do, Korea. 10 L of distilled water was added to 1 kg of dried Hwangchil tree leaves, heated at 100° C., extracted once, filtered through a filter paper, and concentrated under reduced pressure (~ 40° C., 670 mmHg). Hwangchil tree leaf extract concentrated under reduced pressure was freeze-dried and stored at -20°C (hereinafter referred to as “W-Ex”).

Example 2: Preparation of polar solvent and non-polar solvent-soluble fractions from water extract of Hwangchil tree leaf

2-1. Preparation of n-hexane fraction from water extract of Hwangchil tree leaf

20 g of Hwangchil tree leaf water extract (W-Ex) obtained in Example 1 was suspended in 1 L of distilled water. Thereafter, the suspension was put into a fractionation funnel, 1 L of n-hexane was added, shaken well, and fractional extraction was performed. After about 12 hours, the n-hexane fraction extracted supernatant was taken to obtain an n-hexane fraction (hereinafter referred to as "W-Hex"). The fractionation process was repeated twice, after which all the supernatant was concentrated under reduced pressure and lyophilized to obtain 533 mg.

2-2. Preparation of Chloroform Fraction from Water Extract of Hwangchil Tree Leaf

In Example 2-1, the n-hexane layer was separated, and 1 L of chloroform was added to the remaining water extract suspension of Hwangchil tree leaves, and then the fractions were extracted by shaking well. After about 12 hours, a chloroform fraction extract was obtained (hereinafter referred to as "W-CH"). The fractionation process was repeated 3 times, after which all fractions were concentrated under reduced pressure and freeze-dried to obtain 62.2 mg.

2-3. Preparation of Ethyl Acetate Fraction from Water Extract of Hwangchil Tree Leaf

In Example 2-2, the chloroform layer was separated, and 1 L of ethyl acetate was added to the remaining water extract suspension of Hwangchil tree leaves, and then fractionated by shaking well. After about 12 hours, an ethyl acetate fraction extract was obtained (hereinafter referred to as "W-EA"). The fractionation process was repeated three times, and then all fractions were concentrated under reduced pressure and freeze-dried to obtain 350 mg.

2-4. Preparation of n-butanol fraction from water extract of Hwangchil tree leaf

In Example 2-3, the ethyl acetate layer was separated, and 1 L of n-butanol was added to the remaining water extract suspension of Hwangchil tree leaves, and then fractionated by shaking well. After about 12 hours, the n-butanol fraction extract was obtained (hereinafter referred to as "W-Bu"). The fractionation process was repeated twice, and afterwards, both the fraction and the remaining aqueous layer were concentrated under reduced pressure, and freeze-dried to obtain 2.08 g of an n-butanol layer and 13 g of an aqueous layer.

Example 3: Isolation of compounds

3.0 kg of dried and shredded hwangchil leaf ( Dendropanax Morbifera Leveille leaf ) was extracted under reflux for 3 hours with 10 L of 100% methanol. Then, the extract obtained by filtering the extract was concentrated under reduced pressure to obtain 470 g of methanol X.

2 L of distilled water and 2 L of dichloromethane were added to 470 g of the obtained methanol extract, and the mixture was shaken to divide into a dichloromethane layer (A) and an aqueous layer (B). Thereafter, the dichloromethane layer was concentrated under reduced pressure to obtain 159 g of a dichloromethane fraction.

2 L of ethyl acetate was added to 2 L of the fractionated aqueous layer (B), and then divided into an ethyl acetate layer and an aqueous layer (C) by shaking, and then the ethyl acetate layer was concentrated under reduced pressure to obtain 21 g of an ethyl acetate fraction.

2 L of butanol was added to 2 L of the fractionated aqueous layer (B), and then divided into a butanol layer and an aqueous layer by shaking, and then the butanol layer and the aqueous layer were respectively concentrated under reduced pressure to obtain 56 g of a butanol fraction and 145 g of a water-soluble fraction.

After loading the obtained ethyl acetate fraction on a silica gel column, a solvent gradient (chloroform-methanol; 1:0, 60:1, 30:1, 15:1, 10:1, 7:) using chloroform and methanol as mobile phases. 1, 5:1, 3:1, 1:1, 0:1) to obtain 10 ethyl acetate fractions (EA1 ~ EA10).

After loading 900 mg of the obtained EA3 fraction into a silica gel column, 8 fractions (EA3A ~ EA3H) were obtained by gradient elution with a solvent gradient (hexane-ethylacetate; 3:1 ~ 1:6) using hexane and ethyl acetate as mobile phases. Got it.

70 mg of the obtained EA3C fraction was repeatedly subjected to YMC RP-18 column chromatography with MeOH-water (3:1) to obtain 5.3 mg of compound DMW-1.

238 mg of the obtained EA3D fraction was subjected to silica gel and YMC RP-18 column chromatography repeatedly with n-hexane-acetone (1:1) to obtain 6.2 mg of compound DMW-2.

Example 4: Structure analysis of compounds DMW-1 and DMW-2

By measuring the HR-MS spectrum (Agilent 6530 accurate mass QTOF LC/MS) of the compound DMW-1 obtained in Example 3 ([M ⁺ NH ₄ ] ⁺ m/z 336.2167, calcd for 336.2175), C ₁₉ H The molecular formula of ₂₆ O ₄ was estimated. ^{Through 1} H-NMR, ¹³ C-NMR, HMBC, HSQC, and COSY spectra, the compound DMW-1 was identified as a novel substance isolated for the first time in nature. DMW-1 was identified as a new polyacetylene compound, methyl (10E)-9,16-dihydroxyoctadeca-10,17-diene-12,14-diinoate.

By measuring the HR-MS spectrum of compound ^{DMW-2 ([M + HCOO} ] - m / z 335.1872, calcd for 335.1858), estimated a molecular formula of C ₁₈ H ₂₆ O _3. (10E)-(-)-10,17-octadecadiene-12,14-diine-1,9,16-triol, via ¹ H-NMR, ¹³ C-NMR, HMBC, HSQC and COSY spectra, That is, it was found to be fruticotriol.

Table 1 shows the physical properties of the compounds DMW-1 and DMW-2.

- - Compound 1 - Compound 2 location δ _C δ _H ( J in Hz) δ _C δ _H ( J in Hz) One 176.0 - 63.0 3.56, t (6.7) 2 34.8 2.34, t (7.5) 33.7 1.59*, m 3 26.0 1.62, m 26.9 1.37*, m 4 30.4 1.35*, m 30.7 1.37*, m 5 30.3 1.35*, m 30.6 1.37*, m 6 30.1 1.35*, m 30.5 1.37*, m 7 26.3 1.43*, m 26.4 1.54*, m 8 37.8 1.51, dt (11.7, 3.6) 37.8 1.53*, m 9 72.4 4.13, dtd (7.2, 5.7, 1.6) 72.5 4.14, m 10 151.9 6.34, dd (15.9, 5.7) 151.9 6.34, dd (15.9, 5.6) 11 108.4 5.79, ddd (15.9, 1.6, 0.9) 108.4 5.79, ddd (15.9, 1.6, 0.8) 12 78.0 - 78.0 - 13 74.0 - 74.0 - 14 70.6 - 70.6 - 15 82.2 - 82.2 - 16 64.0 4.96, dd (5.4, 1.1) 64.0 4.93, dd (5.5, 1.1) 17 138.1 5.94, ddd (17.1, 10.2, 5.4) 138.1 5.95, ddd (17.0, 10.2, 5.5) 18 116.6 5.42, d (17.1) 5.22, d (10.2) 116.6 5.43, d (17.1)
5.22, d (10.2) OCH ₃ 52.0 3.67, s - -

* Signals overlap.

Example 5: Experimental animals and breeding conditions

In the present invention, 5-week-old and 7-week-old magnetic BALB/c mice were purchased from Damul Science, acclimated for 1 day, and then used for experiments. The environment of the laboratory animal room is standard breeding conditions, the temperature is 23±3℃, the humidity is 40-60%, and the light/dark cycle is maintained for 12 hours, and feed and drinking water for experimental animals are supplied without restrictions. All animal experiments of the present invention were performed under the approval of Chonnam National University Animal Experimental Ethics Committee.

Example 6: Effect of Hwangchil tree water extract on cellular immune response

After administration of Hwangchil tree water extract for 3 weeks, the effect on the cellular immune response by T cells was investigated in order to investigate whether it exhibits an immune enhancing effect. In this example, 10 5-week-old magnetic BALB/c mice were purchased from Damul Science, acclimated for 1 day, and then used for experiments. Experimental animals and breeding conditions are the same as in Example 3 mentioned above.

In the present invention, in order to investigate whether the water extract of Hwangchil tree leaves exhibited an immune enhancing effect when periodically administered, the cellular immune response of T cells in the mouse spleen was investigated. This experiment consisted of 2 groups, a normal control group and a 200 mg/kg administration group of Hwangchil tree leaf extract, and 5 animals per group were tested. All diets and negatives were supplied freely. The experiment was conducted for a total of 3 weeks, and sacrifices were made after 3 weeks. Hwangchil tree leaf extract was injected intraperitoneally every day for 3 weeks.

At 3 weeks, the mouse spleen was removed, the spleen tissue was transferred to a sterilized Petri dish, and the spleen was ground using a 50 μm cell strainer to separate cells from the tissue. All contents were transferred to a 15 mL tube, filled with RPMI medium, and centrifuged at 1200 RPM for 5 minutes. Thereafter, 3 mL of red blood cell lysis buffer was added to the pellet from which the supernatant was removed, mixed well to hemolyze red blood cells, and then centrifuged for 15 minutes at 1500 RPM. The pellet from which the supernatant was removed was washed twice with PBS, and then suspended in RPMI medium to separate splenocytes.

The isolated splenocytes were dispensed into a 24-well plate at 2 X 10 ⁶ cells/mL and 0.5 X 10 ⁵ cells/mL in a 96 well plate, and treated with ConA (5 μg/mL) for 48 hours. Thereafter, in the case of a 96-well plate, MTT reagent was treated to confirm the cell proliferation ability, and a supernatant was taken in a 24-well plate, and the supernatant was measured using an R&D system ELISA kit (Minneapolis, MN, USA).

As a result, as shown in Figure 1a, it was confirmed that the proliferation capacity of cells was significantly increased in the group administered with 200 mg/kg of hwangchil tree leaf extract compared to the normal control group. In addition, as shown in FIGS. 1b and 1c, IL-2 and IFN-r were significantly increased in the group treated with 200 mg/kg of hwangchil tree leaf extract compared to the control group.

Example 7: Splenocytes and CD4 of experimental animals ⁺ Isolation and proliferation of T cells

7-1. Isolation of splenocytes from mouse spleen

In order to confirm the ability of splenocyte culture and T cell proliferation in animals, spleens isolated from animals were washed with PBS, and then a single cell suspension was prepared using a 50 μm cell strainer (BD Falcon, San Diego, CA, USA). Single cell suspension was added with 10% FBS (Gibco BRL, Grand Island, NY, USA), 0.1% penicillin/streptomycin (Gibco BRL, Grand Island, NY, USA) with RPMI-1640 (Welgene, Daegu, Korea). After washing, red blood cells were hemolyzed with an erythrocyte lysis buffer to make a splenocyte suspension, and 150 μL of 1 X 10 ⁶ cells/mL per well was dispensed into a 96-well plate.

7-2. Isolation of CD4 ⁺ T cells from mouse spleen and lymph nodes

In order to confirm CD4 ⁺ T cell culture and T cell proliferation in animals, spleen and lymph nodes isolated from animals were separated, washed with PBS, and then used with a 50 μm cell strainer (BD Falcon, San Diego, CA, USA). A single cell suspension was made. Single cell suspension was added with 10% FBX (Gibco BRL, Grand Island, NY, USA), 0.1% penicillin/streptomycin (Gibco BRL, Grand Island, NY, USA) with RPMI-1640 (Welgene, Daegu, Korea). After washing, the red blood cells were hemolyzed with the red blood cell lysis buffer to make a suspension. CD4 ⁺ T cells were isolated from the suspension using CD4 ⁺ beads.

As for the isolated T cells, 150 μL of 1×10 ⁷ cells/mL per well were dispensed into a 96-well plate coated with CD3 ⁺ antibody and CD28 ⁺ antibody in advance, and cultured at 37° C. for 48 hours. After the cultivation was completed, 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide (MTT, Sigma-Aldrich Co., St. Louis, MO, USA) was treated and incubated for 4 hours. The absorbance was then measured at 570 nm.

7-3. Confirmation of proliferation ability of splenocytes and CD4 ⁺ T cells

To measure the proliferative ability of T cells, splenocytes were treated with 1 μg/mL of concanavalin A (concanavalin A, ConA, Sigma-Aldrich Co., St. Louis, MO, USA), and then at 37°C for 48 hours. Cultured. In addition, cells were cultured by dividing the CD3 ⁺ and CD28 ⁺ cells in the CD4 ⁺ T-coated 96-well plate for 48 hours. After the cultivation was completed, MTT was treated and incubated for 4 hours, and the absorbance was measured at 570 nm.

As a result, it was found that the fraction of the ethyl acetate layer of Hwangchil tree leaves did not show a proliferation effect in other cells, but had very excellent proliferation effect specifically for T cells (FIGS. 2b and 2c). In addition, it was confirmed that the single component, DMW-2, had no significant effect on the proliferation of T cells (FIG. 4A).

Example 8: LC/MS/MS analysis of DMW-2, a single component, in water extract of Hwangchil tree leaf

LC/MS/MS analysis was performed to confirm the content of DMW-2 in the water extract of Hwangchil tree leaf and the ethyl acetate fraction. LC/MS/MS analysis was performed under the following conditions, and the results are shown in FIG. 3.

1. MS conditions: Turbo Ion Spray MRM scan type DMW-2 m/z 273.213/55.1 Spray voltage 5500V, temperature 500℃, positive mode, CG 20, GS1 60, GS2 60.

2. LC conditions:

1) Column: Gemini C18 5 μm, 50 mm Х 2.0 mm Gemini C18 (4.0 mm Х 2.0 mm) guard cartridge, 2) Column oven: 40°C, 3) Autosampler: 15°C, 4) Mobile phase-A: water, 0.1 % Formic acid B: acetonitrile, 0.1% formic acid, 5) Gradient: 1-4 minutes 15-60% B, 5 minutes 60% B, 5.1 minutes 15% B, 6) Flow rate: 0.3 mL/min, 7) Injection volume: 10 μL

Experimental Example 1: IL-2, IFN-γ cytokine concentration measurement using ELISA

IL-2 and IFN-γ cytokines related to T cell growth were measured using an R&D system ELISA kit (Minneapolis, MN, USA). The splenocyte suspension isolated from the mouse was dispensed into a 48-well plate with 500 μL of 5 X 10 ⁶ cells/mL per well. Thereafter, Hwangchil tree water extract (W-Ex: 100 μg/mL), ethyl acetate fraction (W-EA: 10 μg/mL), and DMW-2 (1 μM or 3 μM) were each treated for 72 hours. Cultured. Thereafter, the supernatant (culture solution) was centrifuged at 1200 RPM for 15 minutes, and then only the supernatant was taken again to conduct the experiment.

Treatment of the Hwangchil tree water extract and the ethyl acetate fraction resulted in an excellent effect of increasing IL-2 and IFN-γ cytokine production. In addition, DMW-2, a single component, had a very good effect of increasing IL-2 cytokine production, and also increased the production of IFN-γ at low concentrations (FIGS. 4B and 4C ).

Experimental Example 2: Confirmation of NF-AT, NF-kB, and AP-1 factors and IL-2, IL-4 transcription factors involved in T cells using reporter assay

After dispensing the EL-4 cell line, which is a mouse (Mus musculus) lymphoma cell, into a 6-well plate at 5 X 10 ⁵ cells/well, 4 μg of each reporter was added to Lipofectamine 3000 (Invitrogen, Carlsbad, CA, USA). ) Was used for transfection and cultured in a 37°C, CO ₂ incubator for 48 hours.

Then, Hwangchil tree water extract (W-Ex: 100 μg/mL), ethyl acetate fraction (W-EA: 10 μg/mL), and DMW-2 (1 μM or 3 μM) were treated, respectively. After 4 hours, PMA/Io was treated and incubated in a 37° C., CO ₂ incubator for 24 hours. After 24 hours, the plate was centrifuged at 1200 RPM for 15 minutes and dissolved, and measured using a luminometer using a luciferase reporter system (Promega).

As a result, in the Hwangchil tree water extract and ethyl acetate fraction treatment group, the effect of increasing the expression of NF-AT and AP-1 transcription factors was observed, and no significant effect was observed on NF-kB (FIGS. 5b-5d). In addition, the IL-2 transcription level also showed an increase effect (Fig. 5a).

Meanwhile, DMW-2, a single component, significantly increased the expression of transcription factors of NF-AT, NF-kB, and AP-1 (FIGS. 5b-5d). In addition, treatment with a low concentration of DMW-2 (1 μM) significantly increased IL-2 and IL-4 transcription levels (FIGS. 5A and 5E ).

Claims (12)

Hwangchil tree leaf ( Dendropanax morbifera Lev.leaf ) extract,
The extract is (10E)-(-)-10,17-octadecadiene-12,14-diine-1,9,16-triol ((10E)-(-)-10,17-Octadecadiene-12 ,14-diyne-1,9,16-triol) or salts thereof,
Pharmaceutical composition for immunomodulation or immunity enhancement. According to claim 1, wherein the hwangchil tree leaf extract is one or more solvents selected from the group consisting of water and a lower alcohol having 1 to 3 carbon atoms as an extraction solvent, immunomodulatory or immunity enhancing pharmaceutical composition. The pharmaceutical composition for immunomodulation or immunity according to claim 2, wherein the composition comprises an ethyl acetate extract fraction obtained by purifying only an extract soluble in an ethyl acetate solvent from a water extract or alcohol extract of Hwangchil tree leaf. . Hwangchil tree leaf ( Dendropanax morbifera Lev.leaf ) extract,
The extract is (10E)-(-)-10,17-octadecadiene-12,14-diine-1,9,16-triol ((10E)-(-)-10,17-Octadecadiene-12 ,14-diyne-1,9,16-triol) or salts thereof,
Food composition for immune regulation or immunity enhancement. The method of claim 4, wherein the hwangchil tree leaf extract is one or more solvents selected from the group consisting of water and a lower alcohol having 1 to 3 carbon atoms as an extraction solvent, immune regulation or immunity enhancing food composition. The food composition of claim 5, wherein the composition comprises an ethyl acetate extract fraction obtained by purifying only an extract soluble in an ethyl acetate solvent from a water extract or an alcohol extract of Hwangchil tree leaf. Hwangchil tree leaf ( Dendropanax morbifera Lev.leaf ) extract,
The extract is (10E)-(-)-10,17-octadecadiene-12,14-diine-1,9,16-triol ((10E)-(-)-10,17-Octadecadiene-12 ,14-diyne-1,9,16-triol) or salts thereof,
Pharmaceutical composition for enhancing antitumor immunity. Hwangchil tree leaf ( Dendropanax morbifera Lev.leaf ) extract,
The extract is (10E)-(-)-10,17-octadecadiene-12,14-diine-1,9,16-triol ((10E)-(-)-10,17-Octadecadiene-12 ,14-diyne-1,9,16-triol) or salts thereof,
Food composition for improving antitumor immunity. Hwangchil tree leaf ( Dendropanax morbifera Lev.leaf ) extract,
The extract is (10E)-(-)-10,17-octadecadiene-12,14-diine-1,9,16-triol ((10E)-(-)-10,17-Octadecadiene-12 ,14-diyne-1,9,16-triol) or salts thereof,
Anticancer pharmaceutical composition. The method of claim 9, wherein the cancer is gastric cancer, lung cancer, breast cancer, ovarian cancer, liver cancer, bronchial cancer, nasopharyngeal cancer, laryngeal cancer, pancreatic cancer, bladder cancer, colon cancer, colon cancer, cervical cancer, brain cancer, prostate cancer, bone cancer, head and neck cancer, Skin cancer, kidney cancer, ploidy cancer (polyploid carcinoma), thyroid cancer, parathyroid cancer or ureteral cancer. Hwangchil tree leaf ( Dendropanax morbifera Lev.leaf ) extract,
The extract is (10E)-(-)-10,17-octadecadiene-12,14-diine-1,9,16-triol ((10E)-(-)-10,17-Octadecadiene-12 ,14-diyne-1,9,16-triol) or salts thereof,
Anticancer food composition. The method of claim 11, wherein the cancer is stomach cancer, lung cancer, breast cancer, ovarian cancer, liver cancer, bronchial cancer, nasopharyngeal cancer, laryngeal cancer, pancreatic cancer, bladder cancer, colon cancer, colon cancer, cervical cancer, brain cancer, prostate cancer, bone cancer, head and neck cancer, Skin cancer, kidney cancer, ploidy cancer (polyploid carcinoma), thyroid cancer, parathyroid cancer or ureteral cancer.

Images