KR20180128781A - A method for manufacturing kolaviron - Google Patents

Abstract

The present invention relates to a method for economically and efficiently extracting high purity kolaviron from Garcinia kola in a short time without organic solvents other than water ethanol. The high purity kolaviron produced by the method can be applied as various functional health foods and pharmaceutical compositions.

Description

[0001] The present invention relates to a method for manufacturing colaviron,

The present invention relates to a process for extracting a pure columbarone ingredient from Garcinia coli fruit and a pharmaceutical effect of the collavone thus extracted.

Garcinia Kola , a plant native to Africa, is a flower tree belonging to the family Clusiaceae or Guttiferae. These trees naturally inhabit wetlands such as Cameroon, Benin, Congo, Côte d'Ivoire, Gabon, Nigeria, Liberia, Ghana, Senegal and Sierra Leone. The fruit of this plant is traditionally said to be beneficial to Africans, from folk remedies to treat cough and fever, to liver, kidney and colon. Gegenbaurs morphol. Jahrb. In Leipzig 136 (1990) p95 ~ 101, Victor B. Braide et al. Reported the effects of Garcinia cola Fruit on liver, kidney, and large intestine of rats. In Pharmacological Research Vol 42 (2000) p75 Olatunde Farombi Discloses the effect of columbarone in the genus Garcia coli as an effect of alleviating the toxicity of the plant.

Garcinia cola fruit contains about 7.8% of crude fat and about 5.9% of crude protein. The following mineral components (K, about 50g; Ca, about 10g; Mg, about 17g; N, about 125g; P , About 72 grams), sugar components (about 18.1 grams), ascorbic acid (about 13 grams), and the like. In addition, GB-1 (Garcinia biflavonoid 1, Garcinia biflavonoid 1) (II-3-4'-11-4 to -1-5-11-5-1-7-11-7-heptahydroxy- -biflavone), GB-2 (Garcinia biflavonoid 2, Gardinia biflavonoid 2) (11-3-11-3'-1-4'-11-4 to -1-5-11-5-1-7- 11-7-octahydroxy-3,8 "-biflavone), and Kolaflavone [garcinianin] (Tr-3-11-3'-I1-4" -1-5-TI-5-1- (see ORAyepola et al., Phytomedicine 21, 2014). It is known that garcinia cola fruit can be obtained from natural cola boron It corresponds to a few sources.

According to Pharmacological Research Vol 42 (2000) p75, Garcinia cola is used as a drug substance in two ways.

First, it is a method to dry Garcinia cola fruit, then grind it and consume it as it is (Victor B. Braide, Gegenbaurs morphol. Jahrb. Leipzig 136 (1990) p95-101).

Second, dry and grind Garcinia cola fruit, extract fat with Soxhlet using petroleum ether (bp 40-60 degrees) for 24 hours, dry and then extract Kolaviron with acetone. It is then concentrated, diluted with water, extracted with ethyl acetate, and then the water is removed and the solvent is blown off and vacuum dried to obtain Kolaviron (Olatunde Farombi, Pharmacological Research Vol 42 (2000) p75).

The first method is very simple, but it has a disadvantage that it consumes a very large amount of fiber and carbohydrate components remaining in the fruit and a relatively low content of cobarbone. In addition, the fact that plant species that are native to Africa, a tropical climate, detects large amounts of microbes such as bacteria and fungi in dried fruits, making this process less trustworthy.

The second method is to obtain a relatively pure columbarone, but it is disadvantageous in that the process is complicated and time-consuming and expensive. In addition, since acetone and ethyl acetate are used as the extraction solvent, it is also problematic that it can not be applied to food or the like.

In addition to the above-mentioned two methods, the present inventors have completed a process for producing clavulanic acid which can be used as a raw material for health functional food by saving more cost, completing the process quickly, and using no organic solvent other than water and ethanol, It is strongly desired to confirm the pharmacological properties of a colarbone in the field to which the present invention belongs.

SUMMARY OF THE INVENTION Accordingly, it is an object of the present invention to provide a method for producing colla- radone from Garcinia coli fruit with economical efficiency and high efficiency by minimizing the use of an organic solvent, particularly an organic solvent which is problematic because of its high toxicity.

In order to solve the above-mentioned problems, the present invention provides a method for producing ethanol, comprising the steps of: (S1) adding ethanol to a Garcinia coli fruit to obtain an ethanol extract; And (S2) performing a hydrophobic interaction chromatography on the ethanol extract. ≪ Desc / Clms Page number 5 >

In addition, the step (S2) is performed sequentially according to the following steps: (1) A method for producing cocolabrons from Garcinia coli fruit.

That is, the step (S2)

(S2-1) adding an ethanol extract to an adsorption resin column and adsorbing it;

(S2-2) washing the non-adsorbed component by passing water through the adsorption resin column; And

(S2-3) adding ethanol to the adsorption resin column to obtain an eluate.

Preferably, the present invention is a method for producing cola caniron from a garcinia cola fruit, characterized in that a reversed phase adsorbent polymerized with polystyrene and divinylbenzene is used as the adsorbent resin . More preferably, the adsorbent resin is Diaion HP-20, wherein the adsorbent resin is Diaion HP-20.

Also, the present invention provides a method for producing collarenon from Garcinia coli fruit, wherein the ethanol in step (S2) is ethanol at 80 to 98% (v / v).

Further, the present invention provides a method for producing collarenic acid from Garcinia coli fruit, wherein the ethanol in step (S1) is 80 to 95% (v / v) ethanol.

Further, the present invention provides a method for producing cola caniron from Garcinia coli fruit, characterized in that the ethanol extract of step (S2) is subjected to a step of filtration and dilution with water.

In addition, the colaboron produced by the method according to the present invention

A colaboron having a purity of 95% or more, preferably 97% or more; or

(C ₃₀ H ₂₂ O ₁₂ ), garnia biflavonoid 1 (C ₃₀ H ₂₂ O ₁₁ ), colaflavanone (C ₃₁ H ₂₄ O ₁₂ ) and binarinenin (C ₃₀ H ₂₂ O ₁₀ ) Which is characterized in that it is a clavulanate.

The present invention also relates to a process for the preparation of

A colaboron having a purity of 95% or more, preferably 97% or more; or

(C ₃₀ H ₂₂ O ₁₂ ), garnia biflavonoid 1 (C ₃₀ H ₂₂ O ₁₁ ), colaflavanone (C ₃₁ H ₂₄ O ₁₂ ) and binarinenin (C ₃₀ H ₂₂ O ₁₀ ) And a caffeine-containing food.

.

The present invention also relates to a process for producing

A colaboron having a purity of 95% or more, preferably 97% or more; or

(C ₃₀ H ₂₂ O ₁₂ ), garnia biflavonoid 1 (C ₃₀ H ₂₂ O ₁₁ ), colaflavanone (C ₃₁ H ₂₄ O ₁₂ ) and binarinenin (C ₃₀ H ₂₂ O ₁₀ ) Or a pharmaceutical composition for treating, preventing or ameliorating arthritis containing cocolabyrin as an active ingredient.

Hereinafter, the present invention will be described more specifically.

The present invention relates to (S1) a step of obtaining an ethanol extract by adding ethanol to Garcinia coli fruit; And (S2) performing a hydrophobic interaction chromatography on the ethanol extract. ≪ Desc / Clms Page number 3 >

More particularly, the present invention relates to a method for producing a garlic colostrum, comprising: (a) pulverizing garlic cola fruit; (b) drying; (c) extracting with ethanol; (d) an extractive filter stage; (e) mixing water; (f) injecting the product of (e) into an HP-20 resin column; (g) washing the HP-20 resin column with excess water; (h) extracting cola virone with ethanol to HP-20 resin column; (i) concentrating the extracted ethanol solution with a rotary concentrator; (j) vacuum drying the concentrate; And (k) pulverizing and drying the dried columbarone.

(S1)

Garcinia Kola does not limit its origin, harvest time, or breed. The most preferred parts of the Garcinia cola for use in the process according to the invention may be fruit, or some other part may be incorporated.

Garcinia cola fruit can be added to the process of pulverization and / or extraction after drying. The pulverization is carried out according to various methods known in the art, and is not limited thereto, but may be pulverized to a size of 500 mesh or more, preferably 5-100 mesh. Drying is carried out in accordance with various methods known in the art and can be carried out, for example, at a temperature of between 50 and 80 ° C, but is suitably carried out within a temperature range in which the moisture contained in the fruit is removed, And is not limited to the above temperature range. The pulverization and drying may be performed at the same time, or may be performed at this time, and the order of the two processes is not limited.

To obtain an ethanol extract of Garcinia coli fruit, ethanol is added to Garcinia coli fruit and the sufficient amount of the effective ingredient is stirred to be extracted. The ethanol may preferably be 80 to 95% (v / v) ethanol. According to a specific example, stirring can be carried out for about 1 to 2 hours, but this can be suitably adjusted according to the amount of the fruit of the garcinia cola.

(S2)

Hydrophobic interaction chromatography is performed on the ethanol extract.

The ethanol extract may be subjected to a hydrophobic interaction chromatography process after filtration and / or dilution. The filtration and dilution can be carried out according to various methods known in the art, and the diluting solvent is preferably water.

Hydrophobic interaction Chromacography can be carried out according to the following method:

- adsorbing the ethanol extract on the adsorption resin column;

Passing the water through the adsorption resin column to clean the non-adsorbed component; And

Adding ethanol to the adsorbent resin column to obtain an eluate;

The ethanol extract is applied to the adsorption resin column and brought into contact with the adsorbent resin under conditions that cobalone can bind to the adsorbent resin. Optionally, an increased concentration of a non-polar solvent is added to the adsorption resin column to reduce the water content of the colaboron adsorption resin. The column is washed using a polar solvent, preferably water (more preferably, 5 to 10 times the volume of the adsorbent resin) under the condition that the non-adsorbed component remaining in the adsorbent resin column dissociates, Remove the beard. Finally, an eluent is obtained by adding a polar solvent, preferably an alcohol, more preferably 80 to 98% (v / v) ethanol under the condition that the clavulan is dissociated from the adsorbent resin.

As the adsorbent, a reversed phase adsorbent polymerized with polystyrene and divinylbenzene may be used. For example, Diaion HP-20, Amberlite XAD-2, Bio-beads SM- 2, and so on.

The prepared cola virone

The clavaviron prepared according to the present invention is a clavulanic acid with minimal or high purity derived from or derived from garcinia cola (hereinafter referred to as impurities).

Preferably, the clavaviron produced in accordance with the present invention may be any of the following clavulanones:

A colaboron having a purity of 95% or more, preferably 97% or more; or

(C ₃₀ H ₂₂ O ₁₂ ), garnia biflavonoid 1 (C ₃₀ H ₂₂ O ₁₁ ), colaflavanone (C ₃₁ H ₂₄ O ₁₂ ) and binarinenin (C ₃₀ H ₂₂ O ₁₀ ) Lt; / RTI >

Utilization of manufactured clavabyrone

The clavulanic acid prepared according to the present invention can be utilized in various ways. For example, a health functional food, a food composition, a cosmetic composition, a pharmaceutical composition and the like.

Particularly, the clavabyrin produced according to the present invention has a high content of colavirone and a low content of other impurities, and has an advantage of being harmless to the human body since water and an organic solvent other than alcohol are not used as an extraction solvent.

As an example, a health functional food containing clavabyrone prepared according to the present invention can be provided. The health functional food is based on the pharmacological and pharmacological effects of colavirone known in the art.

As another example, a pharmaceutical composition containing the collavone prepared according to the present invention as an active ingredient can be provided. The pharmaceutical composition is based on the pharmacological effect of colavirone known in the art. Preferably, the pharmaceutical composition according to the invention can be used for the treatment, prevention or amelioration of arthritis.

The pharmaceutical composition according to the present invention may contain a pharmaceutically effective amount of collavone alone or may further comprise one or more pharmaceutically acceptable carriers, excipients or diluents.

The term " pharmaceutically acceptable " means a non-toxic composition which is physiologically acceptable and which, when administered to a human, does not inhibit the action of the active ingredient and does not normally cause an allergic reaction such as gastrointestinal disorder, dizziness, .

Examples of the carrier, excipient or diluent include lactose, dextrose, sucrose, sorbitol, mannitol, xylitol, erythritol, maltitol, starch, acacia rubber, alginate, gelatin, calcium phosphate, calcium silicate, cellulose, methylcellulose , Polyvinylpyrrolidone, water, methylhydroxybenzoate, propylhydroxybenzoate, talc, magnesium stearate, mineral oil, and the like. The pharmaceutical composition may further include a filler, an anti-coagulant, a lubricant, a wetting agent, a flavoring agent, an emulsifying agent or an antiseptic agent.

The " pharmaceutically effective amount " refers to an amount that exhibits a further reaction than the negative control, and preferably refers to an amount sufficient to exhibit the effect of improving, preventing and / or treating arthritis.

In addition, the pharmaceutical composition of the present invention may be formulated using methods known in the art so as to provide rapid, sustained or delayed release of the active ingredient after administration to the mammal. The formulations may be in the form of powders, granules, tablets, emulsions, syrups, aerosols, soft or hard gelatine capsules, sterile injectable solutions, sterile powders.

The route of administration of the pharmaceutical composition of the present invention includes, but is not limited to, oral administration or parenteral administration. Parenteral routes of administration may include, for example, various routes such as transdermal, nasal, peritoneal, muscular, subcutaneous or intravenous. When it includes oral administration, the composition may be formulated orally in the form of tablets, capsules, cachets, gel caps, liquids, suspensions, and the like. Tablets or capsules may contain a binder (for example, pregelatinized corn starch, polyvinylpyrrolidone, or hydroxypropylmethylcellulose); Fillers (e. G., Lactose, microcrystalline cellulose, or calcium hydrogen phosphate); Lubricants (e. G., Magnesium stearate, talc or silica); Disintegrants (e. G., Potato starch or sodium starch glycolate); Or a pharmaceutically acceptable excipient such as a wetting agent (e. G., Sodium lauryl sulfate). The tablets may be coated by methods well known in the art. Liquid preparations for oral administration may take the form of solutions, syrups or suspensions, but are not limited to, they may be present as anhydrous products for constitution with water or other suitable vehicle before use. Such liquid preparations may contain suspending agents (for example, sorbitol syrup, cellulose derivatives, or hydrogenated edible fats); Emulsifiers (e. G., Lecithin or acacia); Non-aqueous vehicles (e. G., Almond oil, oily esters, ethyl alcohol, or fractionated vegetable oils); And preservatives (e. G., Methyl or propyl-p-hydroxybenzoates or sorbic acid). ≪ / RTI > The formulations may also optionally contain buffer salts, flavors, colorants, and sweeteners. Formulations for oral administration may be suitably formulated for sustained release, controlled release, or sustained release of the binding protein (s) useful in the present invention to treat arthritis.

The preferred dosage of the composition of the present invention varies depending on the condition and the weight of the patient, the degree of disease, the type of drug, the route of administration and the period of time, but can be appropriately selected by those skilled in the art.

The composition of the present invention may be used alone or in combination with methods using surgery, radiation therapy, hormone therapy, chemotherapy, and biological response modifiers. For example, the pharmaceutical composition of the present invention may be administered in combination with a known compound having an effect of improving, preventing and / or treating arthritis.

The food composition according to the present invention may further comprise a pharmaceutically acceptable amount of collavone alone or in combination with one or more other pharmaceutically acceptable carriers, excipients or diluents.

The food composition of the present invention includes all natural forms of processing such as food, functional food, nutritional supplement, health feed, and food additives. Food compositions of this type can be prepared in a variety of forms according to conventional methods known in the art.

There is no particular limitation on the kind of the food. Examples of the foods to which the above substances can be added include dairy products including dairy products, meat, sausage, bread, biscuits, rice cakes, chocolate, candies, snacks, confectionery, pizza, ramen and other noodles, gums, ice cream, Beverages, alcoholic beverages and vitamin complexes, dairy products, and dairy products, all of which include health functional foods in a conventional sense.

The clavulanic acid according to the present invention can be added directly to food or used together with other food or food ingredients, and can be suitably used according to conventional methods. The amount of the active ingredient to be mixed can be suitably determined according to the intended use (for prevention or improvement). Generally, the amount of the complex in the health functional food may be 0.1 to 90 parts by weight of the total food weight. However, in the case of long-term intake intended for health and hygiene purposes or for the purpose of controlling health, the amount may be less than the above range, and since there is no problem in terms of safety, the active ingredient may be used in an amount exceeding the above range.

The food composition of the present invention is not particularly limited to the other ingredients other than the above-mentioned collavone as an essential ingredient in the indicated ratios, and may contain various flavors or natural carbohydrates as an additional ingredient such as ordinary beverages. Examples of the above-mentioned natural carbohydrates include monosaccharides such as glucose, fructose and the like; Disaccharides such as maltose, sucrose and the like; And polysaccharides, for example, conventional sugars such as dextrin, cyclodextrin and the like, and sugar alcohols such as xylitol, sorbitol and erythritol. Natural flavors (tau martin, stevia extracts (e.g., rebaudioside A, glycyrrhizin, etc.) and synthetic flavors (saccharin, aspartame, etc.) can be advantageously used as flavors other than those described above .

In addition to the above, the food composition of the present invention may contain flavoring agents such as various nutrients, vitamins, minerals (electrolytes), synthetic flavors and natural flavors, coloring agents and aging agents (cheese, chocolate, etc.), pectic acid and its salts, Salts thereof, organic acids, protective colloid thickeners, pH adjusting agents, stabilizers, preservatives, glycerin, alcohols, carbonating agents used in carbonated drinks, and the like. In addition, it may contain natural fruit juice and pulp for the production of fruit juice drinks and vegetable drinks.

INDUSTRIAL APPLICABILITY According to the present invention, it is possible to economically and efficiently extract collairone of high purity within a short period of time without using water and other organic solvents other than ethanol from Garcinia cola, and the high purity cola-veron produced by this method has various health functions Food, pharmaceutical compositions, and the like.

BRIEF DESCRIPTION OF THE DRAWINGS FIG. 1 is a schematic diagram showing one embodiment of a method for producing high purity columbarone from Garcinia cola according to the present invention. FIG.
Figure 2 shows an HPLC chart of high purity columbarone extracted from Garcinia cola according to the present invention.
FIG. 3 shows the effect of arthritis treatment of a high purity cacabyrone-containing garcinia cola extract according to the present invention. X axis: RANKL SHL (ug / ml), Y axis: TRAP activity (%)

Hereinafter, embodiments of the present invention will be described in detail to facilitate understanding of the present invention. However, the embodiments according to the present invention can be modified into various other forms, and the scope of the present invention should not be construed as being limited to the following embodiments. Embodiments of the invention are provided to more fully describe the present invention to those skilled in the art.

EXAMPLES Example 1 Extraction of Cola Viron from Garcinia Cola

Hard Garcinia Kola was pulverized to a size of 5-100 mesh, and then dried at 50-80 ° C. Approximately 95% (v / v) of ethanol was added to dried Garcinia cola ground and stirred for 1-2 hours, followed by filtration to obtain an ethanol extract. The ethanol solution was mixed with water to lower the solubility of cocolaboron, and the clavabyrone was adsorbed on the porous part of the HP-20 resin by injecting it into a column of 15 to 20 cm in diameter and 1 m in length filled with HP-20 resin Respectively. Water of about 5 to 10 times the HP-20 resin volume was repeatedly administered to remove impurities except for the desired active ingredient such as sugar starch residual oil dissolved in both ethanol and water. 95% ethanol was added to the column to elute the colaborone adsorbed on the porous portion. The eluate was concentrated to a high viscosity with a rotary evaporator to make it highly viscous. Then, the cobracon concentrate was dried in a vacuum oven at a temperature of 40 to 60 ° C under a vacuum of 1 mmHg or less for 5 to 12 hours. The completely dried columbarone was pulverized and finalized into powder form.

Example  2. Colebiron's HPLC  analysis

50 mg of the colaborone sample prepared according to the method of Example 1 and 50 mg of the standard foam (colaborone sample according to the preparation method of "Ayepola et al. BMC Complementary and Alternative Medicine 2013, 13: 363") were dissolved in ethanol, And conditions for HPLC instrument analysis were as follows:

[Instrument analysis condition]

Column: Capcellpak C18 4.6 * 150mm, 5um

Mobile phase: A-0.1% formic acid in water / B-Acetonitrile

Time A solution B solution 0 min 100 0 0 to 10 min 85 15 10 to 50 min 60 40 50 to 60 min 85 15 60 to 80 min 100 0

Flow rate: 1.0 ml / min

Injection volume: 10μl

Oven temp.

Wavelength: 295 nm

The results of LC-Mass analysis were as shown in Table 2.

Peak No. Standard product (LC-MS) Example  1 (LC-MS) Ingredients Peak 1 573.1023 572.939 Garcinia biflavonoid 2 Peak 2 557.1074 556.980 Garcinia biflavonoid 1 Peak 3 587.1178 587.124 Kolaflavanone Peak 4 557.1072 556.980 Garcinia biflavonoid 1 Peak 5 541.1123 541.079 Binaringenin Peak 6 555.0909 Not Detected Kolaflavone

Reference: Ayepola et al. BMC Complementary and Alternative Medicine 2013

The results of HPLC analysis were as shown in Table 3. The primary and secondary of the following Table 3 were obtained from G. kola obtained at the same time, respectively, according to the standard foam preparation method ("Ayepola et al. BMC Complementary and Alternative Medicine 2013, 13: 363" The results are as follows. In Table 3,% represents the area ratio of each peak when the sum of the peaks of 100% is calculated by the HPLC method described above.

Primary Secondary Peak No. Ingredients Standard form Example  One Standard form Example  One Peak 1 Garcinia biflavonoid 2 25.7% 25.6% 18.3% 19.9% Peak 2 Garcinia biflavonoid 1 55.7% 55.4% 60.4% 60.2% Peak 3 Kolaflavanone 5.8% 5.7% 8.3% 7.6% Peak 4 Garcinia biflavonoid 1 4.3%   4.2% 3.8% 3.9% Peak 5 Binaringenin 5.6% 5.5% 7.9% 6.5% Peak 6 Kolaflavone N.D. N.D. N.D. N.D. system 97.1% 96.4% 98.7% 98.1%

Reference: Ayepola et al. BMC Complementary and Alternative Medicine 2013

As can be seen from Tables 2 and 3, the comparison between the standard product and Example 1 shows that there is almost no difference between the components identified in the standard production method and the manufacturing method in Example 1, I could confirm.

Example  3. Refined Colebiron's  Arthritis treatment effect

In order to confirm the therapeutic effect of the purified collavone according to Example 1, osteoclasts were used to investigate the inhibition of TRAP (tartrate-resistant acid phosphatase) activity.

The osteoclasts have a close relationship with the cause of arthritis and have tartrate-resistant acid phosphatase (TRAP), which is an acid phosphatase that is resistant to tartrate. It is an osteoclast Is known to be used as a cytochemical labeling enzyme. Therefore, the treatment effect of arthritis was confirmed through the inhibition of TRAP activity of the osteoclast.

# Experimental animal breeding condition

5 weeks old ICR mouse (Chungju Laboratory Animal Co., Korea)

(Samyang Diet, Korea). The mice were fed with the above conditions for 7 days and adapted to the laboratory environment. The mice were fed a diet containing 20 ~ 24 ℃ of temperature, 50 ~ 55% Were used.

The tibia of the ICR mouse was harvested and both ends were cut and the bone marrow cells were collected by passing through α-MEM essential medium and cultured for 24 hours with 50 ng / mL macrophage-colony stimulation factor (M-CSF) . Untreated cells were washed with α-MEM, and then seeded in 96-wells at 5 × 10 ⁴ cells / well and cultured in α-MEM medium supplemented with 50 ng / mL M-CSF for 3 days. After that, 50 ng / mL of MCSF and 100 ng / mL of RANKL (Receptor Activator for Nuclear Factor κB Ligand) were treated together and treated for 4 days. The cells were washed with PBS, and then 100 μL of substrate solution (1.36 mg / mL 4-nitrophenyl phosphate disodium salt, 10 mM tartrate, 50 mM citrate buffer, pH 4.6) was added to the cells fixed with 10% formaldehyde. And reacted for 30 minutes. After the reaction, the enzyme reaction solution was transferred to a new plate and the reaction was stopped with 0.1N NaOH, and absorbance was measured at 405 nm. The TRAP activity is plotted as a percentage of the absorbance of the sample as a percentage of the absorbance of the control (FIG. 3).

As shown in the graph, it was confirmed that the produced colavirone inhibits osteoclast formation in a concentration-dependent manner. Specifically, TRAP activity was not observed in the control group that did not undergo RANKL treatment, whereas TRAP activity was significantly elevated in the case of differentiation through RANKL treatment. In addition, it was confirmed that TRAP activity was decreased in a concentration-dependent manner when the prepared colarbone was treated even when it was differentiated through RANKL treatment. In particular, when treated with 100 μg / ml, TRAP activity was almost similar to that of the control group . From the results, it was confirmed that the prepared cravabiron has excellent arthritis treatment, prevention or amelioration effect.

Claims (10)

(S1) adding ethanol to Garcinia coli to obtain an ethanol extract; And (S2) performing a hydrophobic interaction chromatography on the ethanol extract. The method according to claim 1, wherein the step (S2) is carried out sequentially according to the following steps:
(S2-1) adding an ethanol extract to an adsorption resin column and adsorbing it;
(S2-2) washing the non-adsorbed component by passing water through the adsorption resin column; And
(S2-3) adding ethanol to the adsorption resin column to obtain an eluate. The method according to claim 2, wherein a reversed phase adsorbent polymerized with polystyrene and divinylbenzene is used as the adsorbent resin. The method according to claim 3, wherein the adsorbent resin is Diaion HP-20. 3. The method of claim 2, wherein the ethanol is 80-98% (v / v) ethanol. The method of claim 1, wherein the ethanol in step (S1) is 80-95% (v / v) ethanol. The method of claim 1, wherein the ethanol extract of step (S2) is filtered and diluted with water. The method according to claim 1, wherein the clavulanon is any one of the following clavulanic acid:
Colavirone with a purity of greater than 95%; or
(C ₃₀ H ₂₂ O ₁₂ ), garnia biflavonoid 1 (C ₃₀ H ₂₂ O ₁₁ ), colaflavanone (C ₃₁ H ₂₄ O ₁₂ ) and binarinenin (C ₃₀ H ₂₂ O ₁₀ ) Lt; / RTI > A health functional food containing a collavone produced according to the method of any one of the following claims:
Colavirone with a purity of greater than 95%; or
(C ₃₀ H ₂₂ O ₁₂ ), garnia biflavonoid 1 (C ₃₀ H ₂₂ O ₁₁ ), colaflavanone (C ₃₁ H ₂₄ O ₁₂ ) and binarinenin (C ₃₀ H ₂₂ O ₁₀ ) Lt; / RTI > A pharmaceutical composition for the treatment, prevention or amelioration of arthritis comprising, as an active ingredient, any of the following collavones prepared according to the method of claim 1:
Colavirone with a purity of greater than 95%; or
(C ₃₀ H ₂₂ O ₁₂ ), garnia biflavonoid 1 (C ₃₀ H ₂₂ O ₁₁ ), colaflavanone (C ₃₁ H ₂₄ O ₁₂ ) and binarinenin (C ₃₀ H ₂₂ O ₁₀ ) Lt; / RTI >

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