KR100529997B1 - Functional Composition Containing Extracts of Fruits of Hovenia dulcis var.koreana - Google Patents

Abstract

In the present invention, the pulp obtained by removing the seeds of the fruit of the bark tree is chopped, and 1 to 10 times water is used in comparison to the weight of the pulp to extract hot water at 1 to 2 atm and 80 to 120 ° C. for 1 to 12 hours. The extract obtained by filtration was concentrated to 65 to 75 ° Brix, and the concentrate was dried to contain a functional composition containing the powdered solid dispersion as an active ingredient.

According to the above configuration, it is possible to manufacture a functional composition effective for preventing various adult diseases. In addition, the composition containing the bark fruit extract prepared by the present invention can be formulated in various forms, such as tablets, powders, injections, etc. can enhance the added value of the farm.

Description

Functional Composition Containing Extracts of Fruits of Hovenia dulcis var.koreana

The present invention relates to a functional composition containing the bark fruit extract as an active ingredient and effective in various adult diseases, and more specifically, by cutting the flesh obtained by removing the seed of the bark fruit, 1 to 10 times the water weight of the flesh. Hot water extraction was carried out at 1 to 2 atm and 80 to 120 ° C. for 1 to 12 hours, and the extract obtained by filtration of the hot water extract was concentrated to 65 to 75 ° Brix, and the concentrate was dried and powdered solid dispersion. It relates to an anticancer functional composition of breast cancer, liver cancer and lung cancer containing a sieve.

Generally, the bark tree is a deciduous tree of the sea buckthorn family, also called the earth-bearing tree, and is distributed in Korea (mainly distributed in Gangwon and the sub-Hwanghae), Japan and China. The bark is black gray and the branches are brownish purple and have a cortex. Winter snow is covered with two snow scales and has hairs. Leaves are 8 ~ 15cm long, ovate, wide ovate or oval, 3 thick leaf veins are developed, and there are fine serrated edges. Fruits are brown, about 8 mm in diameter, shaped like chicken claws, with three seeds in each fruit.

When the fruit is ripe, the fruit tree becomes thick and rough. It has a subtle scent, sweetness, and can be eaten. According to the pharmacy, the herbaceous tree is said to have a decaying effect on liquor trees, and the juice is used to detoxify and relieve nausea.

Barn tree tastes sweet, calm and nontoxic to herbal wood, softens and lubricates the intestines, stools and urinates well, eliminates heat in the chest, quenches thirst, quenches vomiting, eliminates vomiting and detoxification It is said to heal five kinds of hemorrhoids.

So far, examples of products using the barberry fruit have been disclosed as a method for producing kimchi, but 2002-58589 has been published, but the loss of the active ingredient involved in the fermentation process by adding fermented kimchi from the concentrate of the barberry fruit. Is concerned,

Korean Patent No. 2002-22136 discloses a health food made in the form of a candy or a beverage by adding the bark and leaves of a bark tree, boiling water extract of plums and cucumbers, sugar, and herbs. As a result, there is a limit to forming a wider audience.

The present invention has been made to overcome the limitations of the conventional formulation described above, and an object of the present invention is to provide a functional composition containing an extract of the fruit of the fern as an active ingredient having effective functionality for various adult diseases.

It is another object of the present invention to provide a granule, tablet, powder or liquid formulation which is formulated using the composition as a raw material.

The present invention for achieving the above object is 1 to 2 times, 80 to 120 ℃ for 1 to 12 hours by cutting 1 to 10 times the weight of the flesh by cutting the flesh obtained by removing the seeds of the bark fruit Hot water extraction, the extract obtained by filtering the hot water extract is concentrated to 65 ~ 75 ° Brix, and the functional composition containing the dried and powdered solid dispersion as an active ingredient.

The composition may be formulated into granules, tablets, powders or injections.

Hereinafter, the configuration of the present invention will be described in more detail together with the function thereof.

The fruit of the bark which can be used in the present invention is not particularly limited, and the fruit of brown color of about 8 mm obtained from the tree native to Sanya in Gangwon-do or the National Arboretum, etc., preferably the seed is removed. good.

Although the suitable size of a pulp is not specifically limited, Preferably it is thin about 0.1-10 mm.

Extraction process using the pulp is enough to extract 1 to 10 times the water weight of the fine pulp to extract for 1 to 12 hours with hot water of 1 to 2 atm, 80 to 120 ℃.

The hot water extract obtained according to the above process is filtered using a known filtration device, and the extract obtained at this time is concentrated in vacuo to 65 to 75 ° Brix. The extract after the vacuum concentration is then spray dried under the conditions of 50 ~ 70 ℃ using a conventional spray dryer to obtain a solid dispersion.

Examples of formulated excipients when formulated into granules and other preparations using solid dispersions include microcrystalline cellulose, cellulose powder, cellulose acetate, calcium phosphate, calcium sulfate, corn starch, lactose, mannitol, polyvinylpyrrolidone And at least one component selected from the group consisting of talc, dextrin, glucose, fructose, maltose, sucrose, kaolin, magnesium carbonate, magnesium oxide, and polymethacrylate.

Examples of binders that can be blended with the solid dispersion include cellulose derivatives including methyl cellulose, ethyl cellulose, hydroxypropyl cellulose, hydroxypropyl methyl cellulose, and the like, alginic acid, sodium alginate, acacia, guar gum and polymethacrylate. And at least one or more components selected from.

The preparation of the functional composition containing the solid dispersion as an active ingredient is preferably in the range of 0.5 to 30% by weight. If it is added less than 0.5% by weight, it is difficult to fully expect the activity of the active ingredient contained in the barberry fruit extract, if it exceeds 30% by weight it is difficult to expect additional activity according to the addition, rather sensual May adversely affect.

When the functional composition according to the present invention is formulated into various formulations, conventional additives may be added. Examples of such additives may include various nutrients, vitamins, sugars, flavoring agents, lubricants (such as colloidal silicon dioxide or magnesium stearate), colorants, fillers, pH adjusting agents, stabilizers, preservatives, and the like.

Hereinafter, the content of the present invention will be described in more detail with reference to Examples. However, these examples are only presented to understand the content of the present invention, and the scope of the present invention should not be construed as being limited to these embodiments.

Example 1 Preparation of Solid Dispersion

Seedless brown fruit of about 8 mm diameter was cut into 2 mm pieces. Water of 10 times the weight of the fine flesh was inserted and hot water extraction was performed for 4 hours at 1.5 atm and hot water of 90 ° C.

The hot water extract was filtered using a known filtration apparatus, and the extract obtained at this time was concentrated in vacuo to 65 ° Brix. The extract after vacuum concentration was then spray dried under the condition of 55 ° C. using a conventional spray dryer to obtain a solid dispersion.

Example 2 Preparation of Granules

500 g of microcrystalline cellulose and 2 g of magnesium stearate were mixed with 100 g of the solid dispersion of Example 1, granules were prepared in 1000 ml of 5% PVP aqueous solution as a binder, and air-dried.

Example 3 Preparation of Tablet

500 g of microcrystalline cellulose and 2 g of magnesium stearate were mixed with 100 g of the solid dispersion of Example 1, granules were prepared in 1000 ml of 5% PVP aqueous solution as a binder, and air-dried.

Example 4 Preparation of Powder

Powder powder was prepared by mixing 50 g of lactose into 100 g of solid dispersion and filling into an airtight bag.

Example 5 Preparation of Injection

An injection was prepared by adding a residual amount of distilled water for injection and phosphate buffer as a pH adjusting agent to 100 mg of the solid dispersion.

Experimental Example 1 Anticancer and Cytotoxicity Test

The cell lines used in this experiment were Hep3B (human liver cancer cells), A549 (human lung cancer cells) and MCF7 (human breast cancer cells) as cancer cells, and WRL68, a normal human hepatocyte cell, was used as a normal cell to examine the cytotoxicity of the sample itself. . Among the cells used in the experiment, Hep3B, MCF7 and WRL68 were incubated with DMEM medium and 10% heat-inactivated FBS in RPMI 1640 medium for A549.

In Table 1 below, human breast cancer cells (% / mg / ml), Table 2 human liver cancer cells (% / mg / ml) and Table 3, respectively, anticancer activity results for human lung cancer cells (% / mg / ml) Indicated.

Table 1 Human breast cancer cells (% / mg / ml)

division MCF7 (Human Breast Cancer Cell) 0.2 0.4 0.6 0.8 1.0 90 ℃, 1 hour hot water extract 15.70 30.23 65.87 72.23 81.76 Effect criteria 50% or more Test method SRB

Table 2 Human Liver Cancer Cells (% / mg / ml)

division Hep3B (Human Cancer Cell) 0.2 0.4 0.6 0.8 1.0 90 ℃, 1 hour hot water extract 13.24 28.57 40.18 52.23 67.88 Effect criteria 50% or more Test method SRB

Table 3 Human Lung Cancer Cells (% / mg / ml)

division A549 (Human Lung Cancer Cell) 0.2 0.4 0.6 0.8 1.0 90 ℃, 1 hour hot water extract 23 38 57 68 70 Effect criteria 50% or more Test method SRB

As a result of measuring the growth inhibitory activity against the human breast cancer cells (MCF7), human liver cancer cells (Hep3B) and human lung cancer cells (A549) of the fruit tree fruit extract, it was confirmed that the growth inhibitory effect is large and has an anticancer effect.

Cytotoxicity is a method of measuring cell proliferation or toxicity by staining cellular proteins by SRB method.The cells of WRL68, Hep3B, MCF7 (10% FBS, DMEM medium) and A549 (10% FBS, RPMI 1640 medium) 100 μl of each well of a 96 well plate at a concentration of 4-5 × 10 ⁴ cells / ml was incubated for 24 hours (37 ° C., 5% CO ₂ ), and the final concentration of 0.2, 0.4, 100 μl each was added at 0.6, 0.8, 1.0 mg / ml and incubated for 48 hours. After the incubation was completed, the supernatant was removed, 100 μl of cold 10% (w / v) TCA was added, and the mixture was left at 4 ° C. for 1 hour, washed 4 to 5 times with distilled water to remove TCA and dried at room temperature. 100 μl of 0.4% (w / v) SRB solution dissolved in 1% (v / v) acetic acid was added to the wells and stained at room temperature for 30 minutes. The unbound SRB dye solution was washed 4-5 times with 1% acetic acid and dried, and then, 100 μl of 10 mM Trisbuffer was added to dissolve the dye solution. The absorbance was measured at 540 nm using a microplate reader. Table 4 shows.

TABLE 4 Normal Cytotoxicity (% / mg / mL)

division WRL68 (normal liver cells) 0.2 0.4 0.6 0.8 1.0 90 ℃, 1 hour hot water extract 2.51 3.76 6.27 10.08 12.34 Safety and Effectiveness Criteria 50% or more Test method SRB

The cytotoxicity of WRL-68, a normal human hepatocyte, was determined to have low cytotoxicity.

Experimental Example 2 Antimutagenic Experiment

Bacillus subtilis PB 1652 rec + and PB 1791 rec-, which were strains from Franca Zani, Italy by the Rec-assay, were lyophilized using a scalpel in a cleanbench using nutrient agar. Was inoculated in the solidified Petri dish and incubated for 24 to 48 hours. When the incubation was completed, platinum was added to sterile TSB liquid medium and incubated at 37 ° C. for 24 hours to use in the experiment. 10 ml of soft broth agar with 0.1 ml spore solution was added to the petri dish, solidified, and placed on a paper disk. 20 μl of hot water extract and 10 μl of MNNG, a control, were inoculated at 37 ° C. for 12 to 24 hours. Mutagenesis and antimutagenicity were measured after incubation.

Mutagenicity was added to the sterilized paper disc in the amount of 20 µg / paper disc using MNNG (2 µg / µl), which is a strong mutagenic material among various mutagenic substances, under sterile cleanbench. Was added to 4 ~ 100㎍ / paper disc and the mutagenicity was measured by the difference between the inhibition zone (cm) formed by the rec (+) and rec (-) bacteria. Antimutagenicity was determined by mutagenicity, and then mixed with MNNG (10µg / p.disc) and hot water extract (50, 100µg / p.disc) in 1: 1 (10µl: 10µl). After cultivation by means of the comparison of the inhibitory regions formed by each of the extract to determine the degree of mutagenicity of MNNG mutagenic material, the difference between the rec (+) and rec (-) bacteria (cm) and the difference rate ( sample + MNNG / MNNG). The control was treated only with MNNG. The results are shown in Table 5 below.

Table 5 Antimutagenic

division amounttest (µg / disc) Inhibition halodiameter (cm) differnce zone (cm) differnce ratio (sample / MNNG) rec ⁺ rec ^- 90 ℃, 1 hour hot water extract 50 1.2 2.2 1.0 0.58 100 1.1 1.6 0.6 0.53 MNNG 10 1.25 2.72 1.67 One Stability and Effectiveness Criteria 0.55 or less Test method Rec-Black

MNNG (20㎍ / paper disc), a control result of mutagen inhibitory effect according to the extract concentration, by mixing the fruit extract (50, 100㎍ / paper disc) and MNNG (10㎍ / paper disc) 1: 1 Compared to MNNG, MNNG and hydrothermal extracts in 1: 1 ratio showed relatively high antimutagenicity. I could figure out.

Experimental Example 3 Blood Glucose Lowering Experiment

Experiments were performed using α-glucosidase, which plays a critical role in raising blood glucose levels in vivo. First, the enzyme was prepared by dissolving the enzyme in 10 mM PIPES buffer, and then, 10 μl, 40 μl, and 10 μl of 20 mM maltose and hot water extract were mixed, and the final volume was mixed with 60 μl of hot water extract by concentration. Incubate for minutes. 1 ml DNS reagent was added to 60 µl of the reaction solution, and the reaction was stopped by boiling water treatment (10 min) at 100 ° C. water, and then absorbance was measured at 540 nm to quantify the reducing sugar produced by the enzymatic reaction. In comparison, the enzyme activity inhibition rate was calculated and the results are shown in Table 6 below.

TABLE 6 Blood sugar drop (% / mg / ml)

division Hypoglycemic effect 0.2 0.4 0.6 0.8 1.0 90 ℃, 1 hour hot water extract 24 28 35 58 73 Effect criteria 50% or more Test method α-glucosidase

As a result of the experiment, the bark extract showed high hypoglycemic ability, and thus it may be used as a functional food for controlling diabetes.

Experimental Example 4 Hypertension Inhibition

Since ACE is an enzyme that induces hypertension, the ACE reagent was used to measure the inhibitory activity of this enzyme. Before the experiment, all the reactants were maintained at 37 ° C., 1 vial of ACE reagent was dissolved in 10 ml of 37 ° C. distilled water, and 1 ml each was taken into a tube. 100 μl of extract and ACE calibrator for each concentration were added to each tube, and then reacted at 37 ° C. for 5 minutes, and the absorbance was measured at 340 nm. This was determined as the initial A value and the absorbance measured after 5 minutes was determined as final A. As a control, 100 μl of distilled water was added instead of the extract. The ACE (U / L) value of the control was measured by the change in absorbance when nothing was added. The activity calculation of the ACE was calculated according to the following formula and the results are shown in the following Table 7.

ACE (U / L) = (initial A-final A) _{experimental group} / (initial A-final A) _{control group} × active of ACE reagent (50U / L)

Table 7 Hypertension (% / mg / ml)

division Hypertension inhibitory ability 0.2 0.4 0.6 0.8 1.0 90 ℃, 1 hour hot water extract 23 30 51 62 73 Effect criteria 50% or more Test method ACE

As a result of the experiment, the larvae hot water extract showed high hypertension inhibitory ability close to about 80%.

Experimental Example 5 Immune Activity

After inoculating the experimental immune cell line Raji (RPM1-1640 medium) to reach the normal cell growth period (takes about 7 days), 96-well plate (100 μl / well of medium adjusted to 5 × 10 ⁴ Cells / ml) was 37 Incubate at 5 ° C for 24 hours with 5% CO ₂ , administer 100 μl of sample at each concentration, incubate for 48 hours at 37 ° C with 5% CO ₂ , remove supernatant and cool 10% (w / v) TCA. 100 μl was added and left at 4 ° C. for 1 hour, washed 4 to 5 times with distilled water to remove TCA, dried at room temperature and 0.4% (w / v) dissolved in 1% (v / v) acetic acid in each well. ) SRB solution was added 100 μl and stained at room temperature for 30 minutes.

The unbound SRB dye solution was washed 4-5 times with 1% acetic acid and dried, and then 100 μl of 10 mM Tris buffer was added to dissolve the dye solution, and the absorbance was measured at 540 nm. Cancer cell growth (%) = experimental group / control × 100 is to measure the extent to increase the growth of immune cells Raji suggests that the food can increase the immunity as cell growth increases. Immune function enhancement effect was verified using B cell (Raji), a human immune cell. The growth of the cells was incubated at 5% CO ₂ , 37 ℃ in RPMI 1640 medium containing 10% FBS, the immune function enhancement effect was used for SRB assay and the results are shown in Table 8.

TABLE 8 Immune Activity (% / mg / ml)

division Immune Activity 0.2 0.4 0.6 0.8 1.0 90 ℃, 1 hour hot water extract 109 112 125 138 152 Effect criteria 100% or more Test method SRB assay

As a result of the experiment, the concentration of the extract was found to increase approximately 1.8 times from 0.2 to 1.0 mg / ml.

Experimental Example 6 Antioxidant Experiment

In the search for antioxidants, 0.5 ml of the sample was added to 5 ml of 4.5 × ^10-3 linoleic acid aqueous solution, incubated at 50 ° C, and 1.1 ml was collected every 4 days in a test tube with a stopper. TCA 1 ml, TBA 2 ml, and BHT After mixing 0.1 ml and 1 ml of SDS, blowing N ₂ gas, sealing and incubating in a boiling water bath for 15 minutes, allowing it to cool, and mixing and shaking 1 ml of acetic acid and 2 ml of chloroform and centrifuging at 2,500 rpm for 10 minutes. The supernatant was measured for absorbance at 532 nm. Linoleic acid is an essential fatty acid and easily oxidized to polyunsaturated fatty acids. These foods, which exhibit antioxidant properties against linoleic acid, are effective in preventing aging and preventing adult diseases. Search for the antioxidant was used TBA method and the results are shown in Table 9 below.

TABLE 9 Antioxidant Activity (% / mg / ml)

division Antioxidative effect 0.2 0.4 0.6 0.8 1.0 90 ℃, 1 hour hot water extract 17 20 41 55 60 Effect criteria 50% or more Test method TBA

As a result of measuring the antioxidant activity of the hot water extracts of the bark tree in this experiment, it shows an antioxidant capacity of more than 60%, which is thought to be effective in preventing aging and adult diseases caused by body oxidation.

According to the present invention, a functional composition effective for preventing various adult diseases such as antioxidant activity can be prepared. In addition, the composition containing the bark fruit extract prepared according to the present invention can be formulated in various forms such as tablets, powders, injections, etc. can greatly contribute to enhance the added value of the farm.

Claims (7)

In the anticancer functional composition of breast cancer, liver cancer and lung cancer, Cut the flesh obtained by removing the seeds of the fruit of the barn, and inject water 1 to 10 times the weight of the flesh, extract the hot water at 1 to 2 atm and 80 to 120 ° C. for 1 to 12 hours, and filter the hot water extract. An anticancer functional composition of breast cancer, liver cancer and lung cancer, wherein the obtained extract is concentrated to 65-75 ° Brix, and the concentrate contains a dried and powdered solid dispersion. The anticancer functional composition according to claim 1, wherein any one or more selected from excipients and binders is blended with the solid dispersion. The excipient according to claim 2, wherein the excipients are microcrystalline cellulose, cellulose powder, cellulose acetate, calcium phosphate, calcium sulfate, corn starch, lactose, mannitol, polyvinylpyrrolidone, talc, dextrin, glucose, fructose, maltose Anti-cancer functional composition of breast cancer, liver cancer and lung cancer, characterized in that at least one component selected from the group of sucrose, kaolin, magnesium carbonate, magnesium oxide, polymethacrylate is blended. The method of claim 2, wherein the binder is selected from the group of cellulose derivatives including methyl cellulose, ethyl cellulose, hydroxypropyl cellulose, hydroxypropyl methyl cellulose, alginic acid, sodium alginate, acacia, guar gum, polymethacrylate. Anticancer functional composition of breast cancer, liver cancer and lung cancer, characterized in that at least one component. The anticancer functional composition of breast cancer, liver cancer and lung cancer according to claim 2 or 3, wherein colloidal silicon dioxide or magnesium stearate is further added as a lubricant when the raw materials are blended. The anticancer functional composition of breast cancer, liver cancer and lung cancer according to claim 1, wherein the solid dispersion is added in an amount of 0.5 to 30% by weight. The anticancer functional composition of breast cancer, liver cancer and lung cancer according to claim 1, wherein the composition is formulated into granules, tablets, powders or injections.