KR100629314B1 - Composition comprising extract of Alnus japonica Steud. or diarylheptanoid compounds isolated from them for prevention and treatment of cardiovascular disease - Google Patents

Abstract

The present invention relates to a composition for the prophylaxis and treatment of cardiovascular disease, comprising an alder extract or a diaryl heptanoid compound isolated therefrom as an active ingredient.

Since the alder extract of the present invention or the diaryl heptanoid compound isolated therefrom is very excellent in antioxidant activity against low density lipid protein, cardiovascular diseases such as hyperlipidemia and arteriosclerosis, which are known to be caused by oxidation of low density lipid protein It can be useful for the prevention and treatment of

Description

A composition for the prevention and treatment of cardiovascular diseases, comprising an alder extract or a diaryl heptanoid compound isolated therefrom as an active ingredient. Composition comprising extract of Alnus japonica Steud. or diarylheptanoid compounds isolated from them for prevention and treatment of cardiovascular disease}

The present invention relates to a composition for the prophylaxis and treatment of cardiovascular disease, comprising an alder extract or a diaryl heptanoid compound isolated therefrom as an active ingredient.

Recently, as well as an increase in adult disease, vascular disorders such as arteriosclerosis have been greatly increased. Increased cholesterol, especially high LDL-cholesterol, is recognized as a risk factor for diseases associated with atherosclerosis. In order to prevent this disease, attempts have been made to reduce plasma LDL levels through inhibition of cholesterol absorption and inhibition of biosynthesis (Principles in Biochemistry, lipid biosynthesis, 770-817, 3rd Edition, 2000 Worth Publishers, New York; Steinberg, N. Engl. J. Med., 1989, 320, 915-924).

Recently, the production of LDL oxide in the blood as a factor of atherosclerosis has been the main concern (Circulation, 1995, 91, 2488-2496; Arterioscler. Thromb. Vasc. Biol., 1997, 17, 3338- 3346), in particular, the production of LDL peroxides as the formation of foam cells following the influx of HM-LDL (highly modified LDL), which is produced through overoxidation and structural modification, into macrophage Studies on factors and elimination are being actively conducted (Curr. Atheroscler. Res., 2000, 2, 363-372).

Plaque formation and rupture in the vessel wall are the main factors in the development of myocardial infarction, and atherosclerosis is a chronic inflammatory process for damage of the vessel wall and is suggested as a defense mechanism rather than an injury mechanism (Circ. Res. 2001, 89, 298-304).

Probucol, N, N'-diphenylenediamine (N, N'-diphenylenediamine) and phenolic synthetic antioxidants BHA (butylatedhydroxyanisol) and BHT (butylated hydroxy toluene), which are currently being used for the treatment of hyperlipidemia, It is reduced, weakens the degree of oxidation and reduces the formation of lesions, the antioxidant power is excellent, but many side effects are limited use.

Therefore, there is a growing interest in the combined administration of LDL antioxidants with lipid lowering agents in patients with hyperlipidemia or arteriosclerosis, and there is a need for development of antioxidants having fewer side effects and excellent antioxidant power.

On the other hand, Alnus japonica Steud. Is a deciduous algae tree that is attached to the birch family, and is called glass tree, or red sheep, and tea in China. Alder's bark contains 5-10% tannin, taraxolol and betulinic acid. Fruits are astringent diarrhea 약 diarrhea, used for diarrhea, gastrointestinal diseases. The skin is used as postpartum blood 약 pill, gastroenteritis, and used for eye irritation, rheumatism, throat irritation, and stroke. Alder has a taste and bitterness, and is cool in nature, reducing heat and poisoning. In particular, it is very effective in dissolving alcohol poison and has excellent therapeutic effect on various liver diseases such as hepatitis, cirrhosis and fatty liver.

Accordingly, the present inventors, while searching for a new hyperlipidemia and atherosclerosis treatment agent with fewer side effects in natural products, confirmed that the alder extract or diaryl heptanoid compound isolated therefrom has an antioxidant activity against low-density lipid protein. Completed.

The present invention is to provide a composition for the prevention and treatment of cardiac circulatory diseases comprising an alder extract or a diaryl heptanoid compound isolated therefrom.

The present invention provides a composition for the prevention and treatment of cardiovascular disease, which comprises an alder extract or a diaryl heptanoid compound isolated therefrom as an active ingredient.

Compositions of the present invention include pharmaceutical and health food compositions useful for the prevention and treatment of cardiovascular diseases.

Hereinafter, the present invention will be described in detail.

The present invention provides a composition for the prevention and treatment of cardiac circulatory diseases comprising an alder extract or a diaryl heptanoid compound represented by the following Chemical Formulas 1 and 2 as an active ingredient.

(R is methyl, beta-di-xylpyranosyl.)

The diaryl heptanoid compound of Formula 1 is hirsutanone, and the diaryl heptanoid compound of Formula 2 is hirstanonol 5-O-methane (R = methyl) and Oregonin (R = beta-di-xylopyranosyl). The diaryl heptanoid compound may be used in the form of a pharmaceutically acceptable salt, and includes all salts, hydrates, and solvates prepared by conventional methods.

The alder extract or diaryl heptanoid compound isolated therefrom used in the present invention can be obtained by all conventional methods, and commercially available reagents can be used.

Extraction, separation, and purification methods of the alder extract or diaryl heptanoid compounds separated therefrom according to the present invention are as follows.

The dried alder leaves were left in methanol for 3 weeks, filtered, and then mixed with filtrate and stirred for 12 hours at room temperature. After filtering and concentrating the solution, it is suspended in water and filtered once more. The upper layer obtained is dissolved in ethyl acetate and concentrated to give a yellow oily substance. The concentrated solution obtained above was dissolved in dichloromethane, n-hexane was added slowly to recrystallization, filtered using a filter glass, and the liquid phase was concentrated to obtain an oily substance.

The water layer obtained above is separated in the order of n-hexane, chloroform and ethyl acetate to obtain a yellow oily substance. The yellow oily substance obtained from the ethyl acetate layer shows excellent antioxidant activity against low density lipid protein.

The ethyl acetate concentrate obtained above was dissolved in ethyl acetate again, n-hexane was added slowly to recrystallize and then filtered using a filter glass, separated into powder and liquid phase.

The powder obtained above is separated by silica gel column chromatography using a mixed solvent of chloroform and methanol as a mobile phase. At this time, it is preferable to use a chloroform: methanol = 90-70: 10-30 (v / v) solvent as a mobile phase solvent.

By the above method, hirsutanone, hirstanonol 5-O-methane (R = methyl) and oregore which are pure active substances per 1.5 kg of dried alder leaves 60 mg, 90 mg and 810 mg of orein (R = beta-di-xylpyranosyl) were obtained, respectively.

The alder extract of the present invention or the diaryl heptanoid compound isolated therefrom exhibits excellent antioxidant activity against low density lipid protein.

Therefore, the composition of the present invention can be usefully used for the prevention or treatment of cardiovascular diseases such as hyperlipidemia and arteriosclerosis, which are known to be caused by oxidation of low density lipid protein.

The composition of the present invention may further contain at least one active ingredient exhibiting the same or similar function in addition to the alder extract or the diaryl heptanoid compound separated therefrom.

The composition of the present invention may be prepared by including one or more pharmaceutically acceptable carriers in addition to the above-described active ingredients for administration. Pharmaceutically acceptable carriers may be used in combination with saline, sterile water, Ringer's solution, buffered saline, dextrose solution, maltodextrin solution, glycerol, ethanol and one or more of these components, if necessary, as antioxidants, buffers And other conventional additives such as bacteriostatic agents can be added. Diluents, dispersants, surfactants, binders and lubricants may also be added in addition to formulate into injectable formulations, pills, capsules, granules or tablets such as aqueous solutions, suspensions, emulsions and the like. Furthermore, it may be preferably formulated according to each disease or component by a suitable method in the art or using a method disclosed in Remington's Pharmaceutical Science (Recent Edition), Mack Publishing Company, Easton PA.

The compositions of the present invention may be administered parenterally (eg, intravenously, subcutaneously, intraperitoneally or topically) or orally, depending on the desired method, and the dosage may be based on the weight, age, sex, health status, The range varies depending on diet, administration time, administration method, excretion rate and severity of disease. The daily dose is 10-2,000 mg / kg, and preferably 50-500 mg / kg if the alder extract. In addition, in the case of the diaryl heptanoid compound, it is about 0.1 to 100 mg / kg, preferably 0.5 to 10 mg / kg, and more preferably administered once to several times a day.

As a result of toxicity experiments by orally administering the alder extract of the present invention or the diaryl heptanoid compound isolated therefrom to the mouse, the alder extract and the diaryl heptanoid compound were 50% lethal dose by oral toxicity test (LD ₅₀ ) appears to be at least 1,000 mg / kg.

The composition of the present invention can be used alone or in combination with methods using surgery, hormonal therapy, drug therapy and biological response modifiers for the prevention and treatment of cardiovascular diseases.

The composition of the present invention can be added to health foods for the purpose of improving cardiovascular disease. When the alder extract of the present invention or the diaryl heptanoid compound separated therefrom is used as a food additive, the alder extract or the diaryl heptanoid compound separated therefrom may be added as it is or used with other food or food ingredients. Can be suitably used according to conventional methods. The mixed amount of the active ingredient may be suitably determined depending on the purpose of use (prevention, health or therapeutic treatment). In general, in the preparation of food or beverage, the alder extract of the present invention or the diaryl heptanoid compound isolated therefrom is added in an amount of 1 to 20% by weight, preferably 5 to 10% by weight based on the raw materials. However, in the case of long-term intake for health and hygiene or health control, the amount may be below the above range, and the active ingredient may be used in an amount above the above range because there is no problem in terms of safety. .

There is no particular limitation on the kind of food. Examples of the food to which the substance can be added include dairy products including meat, sausage, bread, chocolate, candy, snacks, confectionery, pizza, ramen, other noodles, gums, ice cream, various soups, drinks, tea, drinks, Alcoholic beverages and vitamin complexes, and the like and include all of the health foods in the conventional sense.

The health beverage composition of the present invention may contain various flavors or natural carbohydrates, etc. as additional components, as in the general beverage. The above-mentioned natural carbohydrates are glucose, monosaccharides such as fructose, disaccharides such as maltose and sucrose, and polysaccharides such as dextrin and cyclodextrin, sugar alcohols such as xylitol, sorbitol and erythritol. As the sweetening agent, natural sweetening agents such as tautin and stevia extract, synthetic sweetening agents such as saccharin and aspartame, and the like can be used. The ratio of the natural carbohydrate is generally about 0.01 to 0.04 g, preferably about 0.02 to 0.03 g per 100 ml of the composition of the present invention.

In addition to the above, the composition of the present invention includes various nutrients, vitamins, electrolytes, flavoring agents, coloring agents, pectic acid and salts thereof, alginic acid and salts thereof, organic acids, protective colloidal thickeners, pH regulators, stabilizers, preservatives, glycerin, alcohols, carbonic acid. Carbonating agents and the like used in beverages. In addition, the composition of the present invention may contain a flesh for preparing natural fruit juice, fruit juice beverage and vegetable beverage. These components can be used independently or in combination. The proportion of such additives is not critical but is generally selected in the range of 0.01 to 0.1 parts by weight per 100 parts by weight of the composition of the present invention.

Hereinafter, preferred examples and experimental examples are presented to help understand the present invention. However, the following Examples and Experimental Examples are provided only to more easily understand the present invention, and the contents of the present invention are not limited by the Examples.

Example  : Extraction, Separation and Purification of Diaryl Heptanoid Compounds from Alder

1. Extraction, Separation and Purification from Alder

Alder leaves 1.5kg collected from Wolaksan, Danyang, Korea were washed and dried. The dried alder leaves were placed in 4 l of 95% methanol and left to stand at room temperature for 3 weeks, filtered through a filter paper, and then mixed with filtrate and stirred at room temperature for 12 hours. The solution was filtered and concentrated under reduced pressure to give a yellow oily substance. 200 ml of water was added to this, and it suspended and filtered using filter paper.

The water layer obtained above was separated in the order of n-hexane, chloroform and ethyl acetate to obtain 35.2g, 25.3g and 41.7g of yellow oily substance, respectively. The yellow oily substance obtained from the ethyl acetate layer showed excellent antioxidant activity against low density lipid protein.

The ethyl acetate concentrate obtained above was dissolved in ethyl acetate again, and n-hexane was slowly added to recrystallize and then filtered using a filter glass, and separated into a powder and a liquid phase.

16 g of the powder obtained above was separated by silica gel column chromatography (silica gel: Merk, Art 9385, column size: φ7 × 40 cm) using a mixed solvent of chloroform and methanol as a mobile phase. At this time, the mobile phase solvent is chloroform: methanol = 7: 3 (v / v), the active substance was most effectively separated, the pure active substances hirsutanone (hirsutanone), hirstanonol 5-O-methane (hirstanonol 5- 60 mg, 90 mg and 810 mg of O-methane; R = methyl) and oregonin (R = beta-di-xylopyranosyl) were obtained, respectively.

2. Structural Analysis of Diaryl Heptanoid Compounds

The material obtained in 1 was determined by using a VG high resolution GC / MS spectrometer, Election Ionization MS, Autospec-Ultima, and the molecular weight and molecular formula were determined by using a polarizer (Jasco DIP-181 digital). polarimeter). In addition, ¹ H NMR, ¹³ C NMR, Homo-Cozy, HMQC ( ¹ H-Detected heteronuclear Multiple-Quantum Coherence) and HMBC (Heteronuclear Multiple-) through nuclear magnetic resonance (NMR) analysis (Bruker AMX 500) Bond Coherence (DEP), Distortionless Enhancement by Polarization (DEPT) spectra were obtained, and molecular structure was determined.

The measurement results are as follows. As a result of comparative analysis with the published literatures, the compound of formula 1 was selected from histoneone (Pytochemistry, 2001, 58, 1211), and the compound of formula 2 was selected from hysteronol 5-O-methane and Oregonin (Phytochemistry, 1992, 31, 967).

[Hyrstanone {1,7-di- (3 ', 4'-dihydroxyphenyl) -4-heptan-3-one}]

1) Physical property: sticky solid

2) Molecular Weight: 328

3) Molecular formula: C ₁₉ H ₂₀ O ₅

4) ¹ H-NMR (DMSO, 500 MHz) δ 2.44 (dt, J = 7.0, 7.2 Hz, 2H, H-6), 2.59 (t, J = 7.2 Hz, 2H, H-7), 2.65 (t , J = 7.4 ㎐, 2H, H-1), 2.79 (t, J = 7.4 ㎐, 2H, H-2), 3.41 (br, 1H, -OH), 6.13 (d, J = 16.0 ㎐, 1H, H-4), 6.46 (dd, J = 1.5, 9.2 ㎐, 2H, H-6 '), 6.47 (dd, J = 1.5, 9.2 ㎐, 1H, 6 "), 6.60 (d, J = 1.6 ㎐, 1H, H-2 '), 6.62 (d, J = 1.4 Hz, 1H, H-2 "), 6.64 (d, J = 8.6 Hz, 1H, H-5'), 6.66 (d, J = 8.6 Hz) , 1H, H-5 ″), 6.87 (dt, J = 16.0, 6.7 Hz, 1H, H-5).

5) ¹³ C-NMR (DMSO, 125 MHz) δ 28.9 (C-1), 33.0 (C-7), 33.8 (C-6), 41.2 (C-2), 115.4 (C-5 '), 115.5 (C-5 "), 115.67 (C-2 '), 115.7 (C-2"), 118.7 (C-6'), 118.8 (C-6 "), 130.2 (C-4), 131.7 (C- 1 '), 131.9 (C-1 "), 143.3 (C-4'), 143.4 (C-4"), 145.1 (C-3 '), 146.8 (C-3 "), 199.2 (C-3) .

6) EIMS (rel. Int.%) M / z [M] ⁺ 51 (6.5), 69 (37.7), 123 (100), 131 (10.8), 149 (10.6), 169 (5.7), 181 (13.2) ), 219 (7.3), 328 (26.8).

[Hystannonol 5-O-methane {1,7-di- (3 ', 4'-dihydroxyphenyl) -5-methoxy-heptan-3-one}]

1) Physical property: amorphous powder

2) Luminous intensity: [α] _D ²⁵ + 84 ° (c = 0.2, methanol)

3) Molecular Weight: 360

4) Molecular formula: C ₂₀ H ₂₄ O ₆

5) ¹ H-NMR (DMSO, 500 MHz) δ 1.56-1.62 (m, 2H, H-6), 2.38 (m, 2H, H-7), 2.47-2.66 (m, 6H, H-4b, H -1, H-2, H-4a), 3.22 (s, 3H, H-8), 3.40 (br, 2H, -OH), 3.61 (m, 1H, H-5), 6.39 (dd, J = 2.0, 7.8 ㎐, H-6 '), 6.40 (dd, J = 2.7, 7.8 ㎐, H-6 "), 6.53 (d, J = 2.6 ㎐, 1H, H-2'), 6.55 (d, J = 2.6 Hz, 1H, 2 "), 6.59 (dd, J = 4.0, 7.9 Hz, H-5 '), 6.61 (dd, J = 4.0, 7.9 Hz, H-5"), 8.69 (br, OH, 2H).

6) ¹³ C-NMR (DMSO, 125 MHz) δ 28.4 (C-1), 30.0 (C-7), 35.4 (C-6), 44.6 (C-2), 46.6 (C-4), 55.9 ( C-8), 75.8 (C-5), 115.4 (5 '), 115.5 (5 "), 115.6 (C-2'), 115.7 (C-2"), 118.71 (C-6 '), 118.75 ( C-6 "), 131.9 (C-1 '), 132.6 (C-1"), 143.1 (C-4'), 143.3 (C-4 "), 144.99 (C-3 '), 145.0 (C- 3 "), 208.6 (C-3).

7) EIMS (rel. Int.%) M / z [M] ⁺ 77 (17.8), 123 (100), 136 (33.1), 149 (40.2), 165 (18.4), 328 (39.9), 360 (10.6) ).

[Oregonine {1,7-di- (3 ', 4'-dihydroxyphenyl) -5- (3,4,5-trihydroxy tetrahydropyran-2-yloxy) -heptan-3- On}]

1) Physical property: Brown amorphous powder

2) Luminous intensity: [α] _D ²⁵ -5.5 ° (c = 0.3, methanol)

3) Molecular weight: FAB-MS m / z : 501 (M + Na), EI-MS m / z : 328 (M-150).

4) Molecular formula: C ₂₄ H ₃₀ O ₁₀

5) ¹ H-NMR (CDCl ₃ , 500 MHz) δ 1.67 (m, 2H, H-6), 2.40 (m, 1H, H-7a), 2.49 (m, 1H, H-7b), 2.60 (m , 3H, H-1, H-4b), 2.73 (t like, J = 7.3, 7.6 ㎐, 2H, H-2), 2.79 (dd, J = 6.2, 16.3 ㎐, 1H, H-4a), 2.95 (t like, J = 8.2㎐, 1H, H-2 "'), 3.01 (t like, J = 10.9㎐, 1H, H-5a"'), 3.12 (t like, J = 8.8㎐, 1H, H -3 "'), 3.31 (m, 1H, H-4"'), 3.74 (dd, J = 5.2, 11.2 ㎐, 1H, H-5b "'), 4.03 (m, 1H, H-5), 4.16 (d, J = 7.6 kPa, 1H, H-1 "'), 4.95 (br, 2H, -OH), 6.44 (t like, J = 9.5 kPa, 2H, H-6', 6"), 6.58 (s, 1H, H-2 '), 6.59 (s, 1H, H-2 "), 6.64 (d, J = 8.0 kPa, 2H, H-5', 5"), 8.67 (br, 2H,- OH).

6) ¹³ C-NMR (CDCl ₃ , 125 MHz) δ 28.3 (C-1), 30.2 (C-7), 37.1 (C-6), 44.8 (C-2), 47.1 (C-4), 65.8 (C-5 "'), 69.6 (C-4"'), 73.3 (C-2 "'), 74.3 (C-5), 76.7 (C-3"'), 102.6 (C-1 "') , 115.4 (C-5 ', 5 "), 115.7 (C-2', 2"), 118.7 (C-6 '), 118.8 (C-6 "), 131.9 (1'), 132.9 (C-1 "), 143.1 (C-4 '), 143.2 (C-4"), 144.95 (C-3'), 144.97 (C-3 "), 208.9 (C-3).

Experimental Example 1  Antioxidant Activity of Alder Extract or Diaryl Heptanoid Compounds Isolated from TBARS Method

In order to investigate the antioxidant activity of the alder extract of the present invention or the diaryl heptanoid compound isolated from the low density lipid protein, the following experiment was performed.

Cu ²⁺ is known to induce oxidation of low density lipid proteins (Cu ²⁺ -mediated LDL-oxidation). Therefore, in the present invention, dialdehyde (dialdehyde), which is an oxidation product of the unsaturated fatty acid produced at this time, was measured by TBA (thiobarbituric acid) method, and the antioxidant activity of the alder extract or the diaryl heptanoid compound isolated therefrom was examined ( Packer, L. Ed. (1994) Methods in Enzymology Vol. 234, Oxygen radicals in biological systems Part D. Academic press, San Diego).

300 ml of human plasma was centrifuged at 100,000 xg for 24 hours using an ultracentrifuge to remove the high-density lipoprotein (VLDL) / chylomicron layer suspended in the upper layer and the specific solution to 1.063 g / ml. After centrifugation at 100,000 xg for 24 hours, 25 ml (1.5-2.5 mg protein / ml) of low density lipid protein suspended in the upper layer was separated again.

20 μl (protein concentration, 50-100 μg / ml) of the isolated low density lipid protein was mixed with 210 μl of 10 mM phosphate-buffered saline (PBS), and the alder extract prepared in the above example or 10 µl each of the separated diaryl heptanoid compounds was added.

The alder extract or the diaryl heptanoid compound isolated therefrom was dissolved in DMSO (dimethylsulfoxide) and diluted to various concentrations before use in experiments. As the negative control, only the solvent was added, and as the positive control, the probucol was added.

10 μl of 0.25 mM CuSO ₄ was added to the solution for 4 hours at 37 ° C., and 1 ml of 20% trichloroacetic acid (TCA) solution was added to stop the reaction. 1 ml of 0.67% TBA solution dissolved in 0.05N NaOH solution was added, stirred for 10 seconds, and heated at 95 ° C. for 5 minutes to give a color reaction. The solution was cooled with ice water. The solution was centrifuged at 3000 rpm for 5 minutes to separate the supernatant, and the absorbance at 540 nm was measured by UV-vis spectrophotometry to determine the amount of malondialdehyde (MDA) produced by the color reaction. .

On the other hand, using a stock solution of tetramethoxypropane malonaldehyde bis (dimethylacetal) [tetramethoxypropane (dimethylacetal)] using a stock solution of 250 μL of PBS standard solution containing 0 to 10 mmol malondialdehyde.

The standard solution was developed in the same manner as described above, the absorbance at 540 nm was measured, and the standard curve of malondialdehyde was obtained.

The amount of malondialdehyde in this experiment using an alder extract or a diaryl heptanoid compound isolated therefrom was quantified using this standard curve.

The results are shown in Table 1.

compound IC ₅₀ (uM) Alder Extract (10µg / mL) 97% inhibition

3.3

R = methyl 1.5 R = beta-di-xylpyranosyl 2.2 Positive control group (Probucol) 1.1

As shown in Table 1, the alder extract of the present invention or the diaryl heptanoid compound isolated therefrom is excellent in antioxidant activity against low density lipoprotein.

Therefore, the alder extract according to the present invention or the diaryl heptanoid compound isolated therefrom can be usefully used for the prevention or treatment of cardiovascular diseases such as hyperlipidemia and arteriosclerosis, which are known to be caused by oxidation of low density lipid protein.

Experimental Example 2  : Acute Toxicity in Oral Administration in Mice

In order to determine the acute toxicity of the alder extract according to the present invention or the diaryl heptanoid compound isolated therefrom, the following experiment was performed.

12 females and 12 males (3 males and 3 females each) were used as specific pathogens free ICR mice at 4 weeks of age in an animal room of temperature 22 ± 3 ° C, humidity 55 ± 10%, and illumination 12L / 12D. Breeding in. Mice were allowed to acclimate for about a week before being used in the experiment. Feed for experimental animals (JeilJedang Co., Ltd., mice and rats) and negative water were supplied after sterilization and free ingestion.

The alder extract prepared in Example or the diaryl heptanoid compound isolated therefrom was prepared at a concentration of 50 mg / ml using 0.5% Tween 80 as a solvent, and then 0.04 ml (100 mg / kg) and 0.2 ml per 20 g of mouse body weight. (500 mg / kg) and 0.4 ml (1,000 mg / kg) were administered orally. Samples were administered orally once and observed for side effects or lethality for 7 days after administration. That is, on the day of administration, changes in general symptoms and the presence of dead animals were observed at least once in the morning, at least once every afternoon from 1 hour, 4 hours, 8 hours, 12 hours, and the next day after administration.

In addition, on the 7th day of administration, animals were killed and dissected, and the internal organs were visually examined. Changes in body weight were measured at daily intervals from the day of administration to observe the weight loss phenomenon of the animal by the alder extract or the diaryl heptanoid compound isolated therefrom.

As a result, all mice treated with test substance showed no clinical symptoms and no dead mice, and no toxicity change was observed in weight change, blood test, blood biochemistry test and autopsy findings.

Therefore, the alder extract according to the present invention or the diaryl heptanoid compound isolated therefrom showed no toxicity change up to 1,000 mg / kg in all mice, and the minimum lethal dose (LD ₅₀ ) was at least 1,000 mg / kg or more. It was considered safe.

Examples of preparations for the compositions of the present invention are illustrated below.

Formulation Example 1  : Preparation of Pharmaceutical Formulations

1. Preparation of powder

Diaryl heptanoid compound 2g

1g lactose

The above ingredients were mixed and filled in airtight cloth to prepare a powder.

2. Preparation of Tablets

Diaryl Heptanoid Compound 100mg

Corn Starch 100mg

Lactose 100mg

2 mg magnesium stearate

After mixing the above components, tablets were prepared by tableting according to a conventional method for producing tablets.

3. Preparation of Capsule

Diaryl Heptanoid Compound 100mg

Corn Starch 100mg

Lactose 100mg

2 mg magnesium stearate

After mixing the above components, the capsule was prepared by filling in gelatin capsules according to the conventional method for producing a capsule.

4. Preparation of Injection Solution

10 μg / ml of diaryl heptanoid compound

Dilute hydrochloric acid BP until pH 3.5

Injectable sodium chloride BP up to 1 ml

The diaryl heptanoid compound was dissolved in an appropriate volume of sodium chloride BP for injection, the pH of the resulting solution was adjusted to pH 3.5 with dilute hydrochloric acid BP, and the volume was adjusted with sodium chloride BP for injection and thoroughly mixed. . The solution was filled into a 5 ml Type I ampoule of clear glass, dissolved in glass and enclosed under an upper grid of air, sterilized by autoclaving at 120 ° C. for at least 15 minutes to prepare an injection solution.

Formulation Example 2  : Manufacture of food

Foods containing a diaryl heptanoid compound were prepared as follows.

1. Preparation of Cooking Seasonings

0.2 to 10% by weight of a diaryl heptanoid compound was prepared for cooking spices for health promotion.

2. Preparation of Tomato Ketchup and Sauce

Health-promoting tomato ketchup or sauce was prepared by adding 0.2 to 1.0 wt% of a diaryl heptanoid compound to tomato ketchup or sauce.

3. Manufacturing of Flour Foods

0.1 to 5.0% by weight of the diaryl heptanoid compound was added to the flour, and bread, cake, cookies, crackers and noodles were prepared using the mixture to prepare foods for health promotion.

4. Preparation of soups and gravy

0.1 to 1.0% by weight of a diaryl heptanoid compound was added to soups and broth to prepare meat products for health promotion, soups of noodles, and broth.

5. Preparation of Ground Beef

10% by weight of a diaryl heptanoid compound was added to the ground beef to prepare a ground beef for health promotion.

6. Manufacture of Dairy Products

0.1 to 1.0% by weight of the diaryl heptanoid compound was added to the milk, and the milk was used to prepare various dairy products such as butter and ice cream.

7. Manufacture of wire

Brown rice, barley, glutinous rice, and yulmu were alphad by a known method, and then dried and roasted to prepare a powder having a particle size of 60 mesh using a grinder.

Black beans, black sesame seeds, and perilla were also steamed and dried by a known method, and then ground to a powder having a particle size of 60 mesh.

The diaryl heptanoid compound was decompressed and concentrated in a vacuum concentrator, and the dried product obtained by drying with a spray and a hot air dryer was pulverized with a particle size of 60 mesh to obtain a dry powder.

The dry powders of the grains, seeds and diaryl heptanoid compounds prepared above were prepared in the following proportions.

Cereals (30% by weight brown rice, 15% by weight radish, 20% by weight barley),

Seeds (7% by weight perilla, 8% by weight black beans, 7% by weight black sesame),

Dry powder of a diaryl heptanoid compound (1% by weight),

Ganoderma lucidum (0.5% by weight),

Foxglove (0.5 wt%)

Formulation Example 3  : Preparation of Beverages

1. Preparation of Carbonated Drinks

5-10% of sugar, 0.05-0.3% citric acid, 0.005-0.02% caramel, 0.1-1% of vitamin C are mixed, and 79-94% purified water is mixed to make syrup, and the syrup is 85-98 After sterilizing at 20 ° C. for 20 seconds to 180 ° C., the mixture was mixed with cooling water at a ratio of 1: 4, and carbonated gas was injected at 0.5˜0.82% to prepare a carbonated beverage containing a diaryl heptanoid compound.

2. Manufacture of health drinks

Instant sterilization by homogeneously mixing minor ingredients such as liquid fructose (0.5%), oligosaccharide (2%), sugar (2%), salt (0.5%), water (75%) and diaryl heptanoid compounds This was packaged in small packaging containers such as glass bottles and plastic bottles to prepare a healthy beverage.

3. Preparation of Vegetable Juice

0.5 g of the diaryl heptanoid compound was added to 1,000 ml of tomato or carrot juice to prepare vegetable juice for health promotion.

4. Preparation of Fruit Juice

0.1 g of the diaryl heptanoid compound was added to 1,000 ml of apple or grape juice to prepare a fruit juice for health promotion.

The alder extract of the present invention or the diaryl heptanoid compound isolated therefrom is very excellent in antioxidant activity against low density lipid protein.

Therefore, the composition of the present invention can be usefully used for the prevention and treatment of cardiovascular diseases such as hyperlipidemia and arteriosclerosis, which are known to be caused by oxidation of low density lipid proteins.

Claims (6)

delete Pharmaceutical composition for the prevention and treatment of cardiovascular diseases selected from the group consisting of hyperlipidemia, coronary heart disease, atherosclerosis, and myocardial infarction, using the hysteranol 5-O-methane represented by the formula (2). <Formula 2>

(R is methyl) 3. The cardiovascular disease according to claim 2, wherein the hysteranol 5-O-methane is obtained by extracting, isolating, and purifying the alder from hyperlipidemia, coronary heart disease, atherosclerosis, and myocardial infarction. Pharmaceutical composition for the prevention and treatment of. delete delete delete