KR20030004040A - Extract of herb for promoting release of growth hormone - Google Patents

Abstract

PURPOSE: A medicinal plant extract containing as an effective ingredient a Liriopis Tuber extract which is extracted in alcohol or a mixed solvent of alcohol and water and promotes the release of growth hormone is provided. Therefore, the Liriopis Tuber extract can be effectively used in prophylaxis and treatment of various diseases related with growth hormone because it exhibits high growth hormone releasing activity and is harmless to a human body. CONSTITUTION: Liriopis Tuber is added with alcohol or a mixed solvent of alcohol and water selected from the group consisting of 10 to 100% ethyl alcohol, 10 to 100% methyl alcohol and spirit, hydrolyzed with a 0.1 to 2N acid or base solution, neutralized and then extracted with alcohol or a mixed solvent of alcohol and water. For an example, 1g of Liriopis Tuber extract is mixed with 0.6g of NaCl, 0.1g of ascorbic acid, and an appropriate amount of distilled water to produce an injectable solution.

Description

Extract of herb for promoting release of growth hormone}

The present invention relates to a method for producing Liriopis Tuber extract, to the extract obtained by the method, and to pharmaceutical compositions and health foods comprising the same, more specifically, to add the alcohol or a mixed solvent of alcohol and water Method for preparing ganmundong extract extracted by hydrolyzing and neutralizing with an acid or base solution and adding alcohol or a mixed solvent of alcohol and water, ganmundong extract obtained by the above method and a pharmaceutical composition and health food comprising the same as an active ingredient It is about.

Growth hormone (GH) is a protein secreted by the pituitary gland of animals and is a peptide hormone consisting of 191 amino acids. These growth hormones have been isolated from animals for a long time and have been used mainly for the treatment of dwarfism patients. They have various effects because they inhibit protein loss and delay physiological aging in older adults or the elderly. It can be used to treat diseases (Horber, FF and Haymond, MW, J. Clin. Invest ., 1990, 86, 265-272).

Continuous administration of growth hormone increases body weight, muscle, and fat weight to prevent aging-related diseases. Even when administered temporarily, plasma insulin and IGF-1 (Insulin-like growth factor-1) It is known to increase the concentration and decrease the protein concentration in urine (Kohrt, WM et al., Med. Sci. Sports Exerc ., 1992, 24, 832-837; Zachweija, JJ et al., Am. J. Physiol , 1994, 266 (Endocrinol. Metab. 29), E840-844), as well as increasing the weight and size of the liver and improving the synthesis of proteins in terminal tissues such as muscle, bone and connective tissue (Kaiser, FE). et al., J. Am. Geriatr. Soc ., 1991, 39, 235-240; Marcus, R. et al., J. Clin. Endocrino. Metab ., 1990, 70, 519-527).

Until now, the effects of simultaneous growth hormone releasing hormone (GHRH) and arginine on children and adults with congenital hypopituitarism have been shown to be effective. Growth hormone response was higher in children than in adults, and growth hormone deficiency was higher in patients with deficiency of many types of pituitary hormone than in patients with pituitary gland. Obtained higher results. Responsiveness of growth hormone to GHRH and arginine tends to decrease with age ( J. Clin. Endocrinol. Metab ., 2001, 86 (4), 1574-79), and endogenous growth hormone secretagogues. Ghrelin, a ligand, was effective when administered to growth hormone deficient rats ( Diabetes , 2001, 50 (2), 227-32). The administration of alrondron (alendronate) increases bone mineral density in postmenopausal women with postmenopausal osteoporosis because it maintains a relatively higher rate of formation than when arendronate is administered alone. The effect of MK-677 and arendronate on IGF-1 levels was increased in both MK-677 alone or in combination with arendronate and increased IGF-1 levels and alleviated decreased bone formation. The bone mineral density of the neck increased while the bone mineral density of the lumbar spine and hip did not increase. The metabolic effect of growth hormone through MK-677 reduced the direct inhibitory effect of arendronate on bone formation ( J. Clin. Endocrinol. Metab ., 2001, 86 (3), 1116-25).

In the past, growth hormone was mainly extracted from the cadaveric pituitary gland, but in the process it was found to cause the disease Creutzfeldt-Jakob disease in patients receiving growth hormone. As such, there is a risk that other diseases may be introduced into the patient during the separation process, and growth hormone isolated from the carcass has been limited in its use. With the development of genetic engineering, recombinant growth hormone produced using microorganisms can be produced and supplied in large quantities, and the risk of infection of other diseases is low, so the protein preparation including recombinant growth hormone occupies most of the market for dwarfism. The method of administering the recombinant recombinant growth hormone into the body is made by intramuscular injection using a syringe because the growth hormone itself is a huge protein and cannot be orally administered. As such, although recombinant growth hormone can be mass produced, attempts to develop oral preparations have been actively conducted in recent years due to the high cost required for a certain treatment period and the inability to use injection because it is impossible to orally administer. It is becoming.

The basic requirement for growth hormone preparations for oral administration is to have high absorption rate in the body and to have biological activity that can selectively stimulate growth hormone secretion. In order to have a high absorption rate in the body by oral administration, the smaller the molecular weight is advantageous, and to perform the physiological activity in the body smoothly, it must have a special physical and chemical properties.

Representatives that have been studied as substances with these basic requirements and have been developed up to the clinical trial stage include organically synthesized L-692, 429 ( Horm. Res. , 1993, 40, 109-115) and L-163, 191 ( PNAS). , 1995, 92, 7001-7005), and the peptide compounds GHRP-2 ( Endocrinology , 1994, 140, R9-R13), GHRP-1 ( J. Neuroendocrinology , 1994, 6, 185) and hexarelin (Hexarelin: His -D-2-methyl-Trp-Ala-Trp-D-Phe-Lys-NH2). The materials are newly researched and developed using the basic structure of GHRP-6 (His-D-Trp-Ala-Trp-D-Phe-Lys-NH2), a peptide compound commonly known in the early 1980s.

Recently, the approach of developing a new material has been in the spotlight by using a chemical library to search for a large amount of candidates, and in addition to the peptide library composed of short peptides and the natural world, Natural libraries consisting of many existing materials are also used to develop these new materials. Developing new drugs using such a wide variety of libraries is considerably more efficient than traditional approaches in terms of the time and cost of development, so developed pharmaceutical companies have long been active in developing new drugs using their libraries. come. Specifically, L-163, 191, and hexarelin may be referred to as substances developed by this approach.

Originally it varies from animal to animal, but in humans there is 44 amino acids of GHRH. The regulation of growth hormone secretion in the human body is a hormone called GHRH which promotes the secretion of growth hormone and somatostatin which inhibits the growth hormone secretion. It has been known to act to secrete

GHRP-6 is a useful synthetic peptide that is less effective than GHRH in biological activity but is highly compressed in terms of molecular weight size ( Journal of Endocrinology , 1995, 10 (1), 1-9). Since the development of GHRP-6, many researchers have reported that GHRP-6 does not act on the same receptors as GHRH, and the possibility of the presence of new receptors via these synthetic growth hormone secretagogues. Has been raised ( Endocrinology , 1989, 124, 2791). This possibility is demonstrated by the discovery of the cDNA sequence of growth hormone secretagogue receptors present in the pituitary archromediate and hypothalamus of pigs and humans by Merck researchers. ( Science , 1996, 273, 974-977). This fact proved the synergism between the previously found growth hormone secretagogue and the synthetic compound growth hormone secretagogue, and is more interested in the identification of substances that exist in the body and are expected to play the same role as GHRH. This situation is increasing.

L-692, 429 ( Science , 1993, 260, 1640-1643), developed by Merck, USA, has been shown to selectively secrete growth hormone, and has been shown to have an effective body absorption rate even in animal testing. However, the results of human experiments, unlike that of animals had a problem that the absorption is not very high when administered orally. Therefore, L-163, 191, which improved these problems, was developed, which is a compound that increased the oral absorption rate, which is the biggest problem of the growth hormone secretagogues that have been developed previously, and has been reported to have an absorption rate of more than 60% when orally administered to dogs. Clinical trials are underway ( PNAS , 1995, 92, 7001-7005; Endocrinology , 1996, 137, 5284-5289). The results of these studies suggest that substances that stimulate the pituitary gland to secrete growth hormone may be useful clinically as growth hormone replacements, while developing new growth hormone secretagogues is not easy. Shows.

As part of such research, researches on finding substances that promote growth hormone secretion using natural extracts that are harmless to the human body are being conducted. Examples include alcoholic extracts of Astragalus root and 7-hydroxy-4'-methoxy-isoflavone, formononetin and stigmas-4-en-6β-ol-3-one (stigmast-4-en-6β-ol-3-one) and the like have been confirmed by the present inventors to stimulate growth hormone secretion (Korean Patent Nos. 257840 and 251519; Kim, JS and Kim, CS, Kor. J. Pharmacogn ., 1997, 28 (2), 75-79; Kim, JS et al., Kor. J. Pharmacogn. , 1996, 27 (4), 336-341), Regarding the growth hormone secretagogue composition containing one or more extracts selected from the group consisting of extracts from the ground, extracts from the dead, extracts from lilies, and gilyeong extracts (Korean Patent Application No. 1998-0053921), growth hormone secretion stimulating factor from natural herbal Method and extract of water-soluble fast extract (Korean Patent Application No. 1998-014589) and glycoalkaloids It promotes secretion of growth hormone by the spur saponin H1 (H1 asperosaponin) factors have been reported studies on (Republic of Korea Patent Application No. 1998-014590 No.).

Liriope Tuber is the bulge of the root of Liriope platyphylla Wanget Tang or Ophiopogon japonicus Ker-Gawler, a so-called perennial herbaceous genus belonging to the Malmung Swallow family. . The herbal medicine is called Liriope Tuber using the bulge in the root.

The broadleaf vegetation is widely distributed in Korea, Jeju, Jeonnam, Jeonbuk, Gyeongan, Gyeongbuk, and Gangwon. The swelling part of the root of the plant is medicated as Liriope Radix (Liriope Tuber). The bulging part of the bulging root reaches 2.5-4 cm in length and has a width of about 1 cm. The leaf lobe Macmun-dong grows in the forests of mountains and is distributed in Jeju, Gyeongnam, Gyeongbuk, and Gyeonggi.

In Chinese Pharmacopoeia, it is called pulsation and is called Maidong and belongs to the leaf lobe gram-mundong family. In Japan, the bulge portion of the root of Ophiopogon japonicus is medicated as pulmonary sinus. However, the root bulge of lobular vein sinus is about 1 ㎝ long and 0.5-0.8 ㎝ in width, so it is not only lobe pulmonary sinus. In Korea, both broadleaf vegetation and leaflet vegetation are medicinal, and there is a big difference in the length, width, and size of root bulge of these two plants. do.

The chemical composition of Macmundong is Opiopogon amide VI, 5,7-dihydroxy-6-formyl-8-methyl-3- (3,4-methylenedioxybenzyl) chroman-4-one. (5,7-Dihydroxy-6-formyl-8-methyl-3- (3,4-methylen dioxybenzyl) chroman-4-one), Azetidine-2-carboxylic acid, bise Vicenin 2, Tulipinin, 6-Aldehydo-isoophiopogonone A (6-Aldehydo-isoophiopogonone A), 6-Aldehydo-isoopiopogonone B (6-Aldehydo-isoophiopogone B 6-Aldehydo-ophiopogonone A, Opiopogon, Homoisoflavonoid I, Opiopogon homoisoflavonoid II, Opiopogon homoisoflavonoid II, Opiopogon homoisoflavonoid II Opiopogon homoisoflavonoid III, Opiopogon homoisoflavonoid IV, Opiopogon homoisoflavonoid V homoisoflavonoid Ⅴ), Opiopogonanone A, Methylophiopogonanone A, Methyl ophiopogonanone B, Dacosterol, 7-O-β- 7-O-β-D-glucopyranosyl-borneol, 7-O-α-L-arabinofumanosyl (1-6) -β-D-glucopyranosyl boron Neool (7-O-α-L-Aravinofuranosyl (1-6) -β-D-glucopyranosylborneol), Opiopogonin A, Opiopogonin B, Opiopogonin D , Opiopogonin B ', Opiopogonin C, Opiopogonin C', Opiopogonin D ', 3-O-α-L Ramponopyranosyl (1-2) -β-D-glucopyranosyl opiogenin (3-O-α-L-Rhamnopyranosyl (1-2) -β-D-glucopyranosylophiogenin) and the like.

Macmundong's medicinal effect is viscous anti-inflammatory, whooping cough, diuresis, Jinhae, mind and body anxiety, expectoration, nourishing tonic is famous. Representative oriental medicine prescription is Mcmundongtang, so dry cough, when the neck is restless, sputum is very mucus, it is used for skin drying.

According to the recent scientific results, the effect of goji berry contains lectin, which acts as a mitogen for lymphocytes and has antitumor effects (Yakhak Hoeji, 1996, 40 (4), 387-393). It has been reported that herbal compositions administered in combination with chemotherapy in patients with osteogenic sarcoma after surgery have the effect of reducing toxicity and side effects (Zhongguo Zhhong Xi Yi Jie Za Zhi, 1993, 13 (3). ), 150-2).

This is known to have a variety of efficacy of munmundong until now has been prepared in the form of a mixture with other herbal medicines. Sports scientific ginseng products and its manufacturing method (registration No. 211718), the composition for enhancing oral hygiene using herbal extracts (1998, 6. 5), pharmaceutical composition for circulatory system and its preparation method (Registration No. 249934) There is a method for producing diabetic drugs and herbal medicine-derived heterogeneous fiber (Registration No. 053436), and foreign patents include components of whey blue cheongsimhwan (US 5190757, US 5164184, US 5133964), and a combination agent (JP 104886A2) used for atherosclerosis, Anti-aging is also being made with a combination.

Thus, the present inventors have completed the present invention by studying the physiological activity of the extracts of the mammundong, revealing that wormwood extract acts as a growth hormone secretagogue to increase the production of growth hormone.

SUMMARY OF THE INVENTION An object of the present invention is to provide a method for preparing extracts of Macmundong having growth hormone secretagogue activity, extracts obtained by the method, and pharmaceutical compositions and health foods containing the same as an active ingredient.

1 is a graph showing the growth hormone was measured at regular time intervals from the tail vein after administering the pulmonary sinus extract of the present invention to the jugular vein of the rat.

In order to achieve the above object, the present invention provides a method for producing the extract.

In addition, the present invention provides a wormwood extract having growth hormone secretagogue activity prepared by the above method.

The present invention also provides a pharmaceutical composition for promoting growth hormone secretion containing the extract as an active ingredient.

In another aspect, the present invention provides a growth hormone secretion promoting health food containing the extract as the active ingredient.

Hereinafter, the present invention will be described in detail.

The present invention provides a method for producing the extract.

Specifically, the present invention provides a method for preparing the pulmonary tract extract by adding ganmundong to alcohol or a mixed solvent of alcohol and water and immersing at 5 to 40 ° C. The aqueous alcohol solution may be selected from the group consisting of 10 to 100% ethyl alcohol (ethyl alcohol), 10 to 100% methyl alcohol (methyl alcohol) and spirits.

In another aspect, the present invention provides a method for preparing the extract of McMoon Dong by hydrolyzing and neutralizing the pulse moon dong with an acid or a base, and then adding an alcohol or a mixed solvent of alcohol and water. At this time, the acid or base is preferably in a concentration of 0.1 to 2 N, the acid may be selected from the group consisting of hydrochloric acid, sulfuric acid, nitric acid and phosphoric acid and the base may be selected from the group consisting of sodium hydroxide and potassium hydroxide. have. According to this method, the bound compound of the plant, which has been bonded to various substituents such as sugar or other alkyl groups, is hydrolyzed, and the extract of Macmundong is included as a compound of the free form of the plant. The aqueous alcohol solution may be selected from the group consisting of 10 to 100% ethyl alcohol (ethyl alcohol), 10 to 100% methyl alcohol (methyl alcohol) and spirits.

In addition, the present invention provides a wormwood extract having growth hormone secretagogue activity prepared by the above method.

The present inventors confirmed whether the extract of the present invention munmundong has growth hormone secretagogue activity. First, pituitary cells were removed from Sprague-Dawleyrat rats at 4-5 weeks of age under aseptic manipulation, then gently washed and crushed to separate pituitary cells. The pulmonary tract extract of the present invention was added to the pituitary cells isolated from the above, and after the rat growth hormone secretion factor (rat GRF, Bachem Co., Torrance, CA, USA), which is a reference substance for growth hormone secretion, was added, 37 Incubated at ℃. After incubation, the supernatant was separated by centrifugation, and the amount of growth hormone was measured using a rat growth hormone RIA kit.

As a result, the mammundong extract of the present invention acted on the pituitary cells to promote the secretion of growth hormone, and the growth hormone secretion was much superior to the rat growth hormone secretion factor used as a comparison group was confirmed that the effect of promoting the secretion was excellent (see Table 1 ). .

In addition, growth hormone secretion experiments using an animal model, the extract was found to be a strong increase in growth hormone in vitro ( in vitro ) as well as in vivo (see Table 2 ).

The present invention also provides a pharmaceutical composition for promoting growth hormone secretion containing Macmundong extract as an active ingredient.

The pharmaceutical composition for promoting growth hormone secretion of the present invention contains the extract of Macmundong as an active ingredient. The Macmundong extract can be administered orally or parenterally during clinical administration and can be used in the form of a general pharmaceutical formulation. That is, the extract of the present invention may be administered in various oral and parenteral dosage forms in actual clinical administration, and when formulated, diluents such as fillers, extenders, binders, humectants, disintegrating agents and surfactants that are commonly used, or Formulated using excipients. Solid preparations for oral administration include tablets, pills, powders, granules, and capsules, and the like, which may contain at least one excipient such as starch, calcium carbonate, sucrose, lactose and gelatin, etc. Mix is prepared. In addition to simple excipients, lubricants such as magnesium styrate talc are also used. Liquid preparations for oral administration include suspensions, solutions, emulsions and syrups, and may include various excipients such as wetting agents, sweeteners, fragrances and preservatives, in addition to commonly used simple diluents such as water and liquid paraffin. have. Formulations for parenteral administration include sterile aqueous solutions, non-aqueous solvents, suspensions, emulsions, lyophilized preparations and suppositories. As the non-aqueous solvent and the suspension solvent, propylene glycol, polyethylene glycol, vegetable oil such as olive oil, injectable ester such as ethyl oleate, and the like can be used. As the base of the suppository, witepsol, macrogol, tween 61, cacao butter, laurin butter, glycerol and gelatin may be used.

Dosage units may contain, for example, one, two, three or four times the individual dosage, or they may contain 1/2, 1/3 or 1/4 times. The individual dosage preferably contains an amount in which the effective drug is administered at one time, which usually corresponds to all, 1/2, 1/3 or 1/4 times the daily dose.

The effective dose of the pulsed extract is 0.1 to 40 mg / kg, preferably 0.4 to 4 mg / kg, and may be administered 1-6 times a day.

As a result of conducting oral administration, intraperitoneal administration, and subcutaneous injection of the pulmonary sinus extract of the present invention to mice, 50% lethal dose (LD ₅₀ ) by intraperitoneal administration and subcutaneous injection toxicity test was at least a Liliope. The pulmonary vegetation extract of 134.34 ± 12.50 g / kg and the leaflet vegetative extract of Ophiopogon were found to be safe substances of 20.61 ± 7.08 g / kg.

In addition, the present invention provides a growth hormone secretion promoting health food containing the extract as an active ingredient.

In the present specification, the health food is a food that improves the function of the general food by adding the medicinal extract to the general food, the medicinal extract can be added to the general food, or can be prepared by encapsulation, powdered, suspension and the like. When ingested, it brings a specific effect on health, and unlike the general medicine because the food is a raw material has the advantage that there is no side effect that can occur during long-term use of the drug.

When using the medicinal extract of the present invention as a food additive, the medicinal extract may be added as it is or used with other food or food ingredients, and may be appropriately used according to a conventional method. The mixed amount of the active ingredient can be suitably determined according to the purpose of use (prevention, health or therapeutic treatment). In general, the extract may be added in an amount of 0.0001 to 10% by weight, preferably 0.1 to 5% by weight based on the raw materials in the manufacture of food or beverage. However, in the case of prolonged ingestion for health and hygiene purposes or health control purposes, the amount may be below the above range. In addition, the health food of the present invention contains the wormwood extract within the toxic range measured when used in the pharmaceutical composition.

There is no particular limitation on the kind of food. Examples of foods to which the extract may be added include meat, sausage, bread, chocolate, candy, snacks, confectionery, pizza, ramen, other noodles, gums, dairy products including ice cream, various soups, beverages, teas, drinks Alcoholic beverages and vitamin complexes. Specifically, the health foods containing the extract of Megmun-dong include health foods and favorite products such as juice, tea, jelly, juice, etc., which are made mainly of Megmun-dong, and are used for the purpose of viscous anti-inflammatory, Jinhae, sputum, and jigal. Therapeutic agents etc. are mentioned.

Hereinafter, the present invention will be described in detail by way of examples.

However, the following examples are merely to illustrate the invention, but the content of the present invention is not limited to the following examples.

Example 1 Preparation of Macmundong Extract 1

50 g of dried powder powder was added to 500 ml of 70% aqueous methanol solution and stirred at room temperature for 2 days. The solution obtained through the stirring process was filtered through a filter paper, concentrated under reduced pressure to remove methanol, and then lyophilized to remove moisture to prepare the extract of the present invention.

Example 2 Preparation of Macmundong Extract 2

0.1 g of dried powder powder was added to 1 ml of 1 N HCl aqueous solution and reacted at 100 ° C. for 2 hours. After hydrolysis for 2 hours and neutralized, the reaction solution was lyophilized to remove moisture, and thus, the extract of Macmundong was prepared in the same manner as in Example 1.

Preparation Example 1 Preparation of Injection Solution

Injection solution containing 10 mg of the active ingredient was prepared by the following method.

100 g of 1 g of Macmundong extract, 0.6 g of sodium chloride and 0.1 g of ascorbic acid were dissolved in distilled water. The solution was bottled and sterilized by heating at 20 ° C. for 30 minutes.

The components of the injection solution are as follows.

Megmundong extract 1 g

0.6 g sodium chloride

0.1 g of ascorbic acid

Distilled Water Determination

Preparation Example 2 Preparation of Syrup

Syrup containing 2% (weight / volume) of the active ingredient of McMangdong extract of the present invention is prepared by the following method.

Macmundong extract, saccharin, and sugar were dissolved in 80 g of warm water. After the solution was cooled, a solution consisting of glycerin, spices, ethanol, sorbic acid and distilled water was prepared and mixed therein. Water was added to this mixture to 100 ml.

The components of the syrup are as follows.

Megmundong Extract 2 g

Saccharin 0.8 g

25.4 g per

Glycerin 8.0 g

0.04 g of spices

Ethanol 4.0 g

0.4 g of sorbic acid

Distilled Water Determination

Preparation Example 3 Manufacturing Method

A tablet containing 15 mg of active ingredient is prepared by the following method.

It was mixed with 250 g of Macmundong extract, 175.9 g of lactose, 180 g of potato starch, and 32 g of colloidal silicic acid. 10% gelatin solution was added to the mixture, which was then ground and passed through a 14 mesh sieve. It was dried and the mixture obtained by adding 160 g of potato starch, 50 g of talc and 5 g of magnesium stearate was made into a tablet.

The components of the tablet are as follows.

250 g of Macmundong extract

Lactose 175.9 g

180 g potato starch

32 g of colloidal silicic acid

10% gelatin solution

Potato Starch 160 g

50 g of talc

5 g of magnesium stearate

Preparation Example 4 Manufacturing Method of Food and Beverage

The present inventors prepared the food or beverage composition containing the extract as the active ingredient as follows.

<4-1> Preparation of Biscuits

Force Class 1 88 kg

Gravity First Class 76.4 ㎏

16.5 kg per white

2.5 kg of salt

2.7 kg of glucose

Palm shortening 40.5 kg

5.3 kg of ammo

Medium kg 0.6 kg

0.55 kg sodium bisulfite

Rice flour 5.0 kg

Vitamin B1 0.003 kg

0.003 kg of vitamin B2

Milk Flavor 0.16 ㎏

71.1 kg of water

Whole milk powder 4 kg

Substitute powder 1 kg

0.1 kg of calcium phosphate

Spray salt 1 kg

25 kg of spray oil

Megmundong extract 0.1-0.5 kg

Biscuits were prepared using conventional methods with the above composition and content.

<4-2> Preparation of Beverage

522 mg of honey

Chioctosanamide 5 mg

Nicotinamide 10 mg

Riboflavin Sodium Hydrochloride 3 mg

Pyridoxine hydrochloride 2 mg

Inositol 30 mg

Orthoic acid 50 mg

Megmundong Extract 0.48-1.28 mg

200 ml of water

With the above composition and content, drinks were prepared using conventional methods.

Experimental Example 1 Growth Hormone Secretion Activity Test Using Macmundong Extract

<1-1> Pituitary Cell Culture

The pituitary gland was removed from Sprague-Dawley rats at 4-5 weeks of age under aseptic manipulation, followed by collagenase (Gibco co., Grand Island, NY) and hyaluronidase (Sigma co). , St. Louis, Mo.) rat rat pituitary cells were isolated. The isolated pituitary cells were suspended in Dulbecco's Modified Eagle's medium containing 10% horse serum, 2.5% fetal bovine serum (FBS), and antibiotics. Incubated at 37 ° C., 5% CO ₂ ).

<1-2> Growth hormone secretion promoting activity experiment

The pituitary cells isolated in Experimental Example <1-1> were cultured and then suspended in a test solution. Medicinal herbs were added to pituitary cells at a concentration of 1 mg / ml, and rat growth hormone secretion factors (rat GRF, Bachem Co., Torrance, CA, USA), which are reference substances for growth hormone secretion, were 300 nM and 500 nM. , 1,000 nM was added and the test solution was added to a total volume of 1 ml and incubated at 37 ° C. for 15 minutes. After incubation, the supernatant was separated by centrifugation, and stored at -70 ° C to measure growth hormone.

Growth hormone measurement was performed using a rat growth hormone RIA kit (rat GH RIA kit, Amersham, Buckinghamshire, England). The growth hormone contained in a constant amount of ¹²⁵ I-labeled growth hormone and the test substance stored in the above was tested by different reactions of the growth hormone-antibody complex (GH) by a competitive response to an anti-GH antibody. -Ab complex) was formed and precipitated by addition of a second antibody, and the radioactivity of ¹²⁵ I was measured from the precipitate. The results are shown in Table 1 below.

Added concentration Growth hormone concentration (ng / ml) Control 0 23.7 Comparative group ^* 300 nM 38.2 500 nM 57.7 1,000 nM 146.6 Macmundong 1mg medicine / ml 364.6

The rat growth hormone secretion factor was used as a comparison group.

As a result, as can be seen in Table 1 , the extract of the present invention acts on the pituitary cells to promote the secretion of growth hormone, and the growth hormone secretion effect is superior to the rat growth hormone secretion factor used in the comparison group. It was confirmed.

Experimental Example 2 Growth Hormone-induced Experiment of Rat 1

Six-week-old Sprague-Dawley rats were administered intraperitoneally with Macmundong extract to determine the concentration of induced growth hormone. Several rats in each experimental group were dissolved in an injection solution of Macmundong extract in an intraperitoneal injection at a dose of 4mg / kg / ml, followed by blood collection from the tail vein at intervals of 10 minutes for several hours after anesthesia. In the control group, blood was injected for the same time after only the injection of the corresponding amount. Measurement of the concentration of growth hormone in the blood in the same manner as in Experimental Example 1 Rat growth hormone RIA kit (rat GH RIA kit, Amersham, Buckinghamshire, England) was performed. A certain amount¹²⁵Growth hormones contained in the I-labeled growth hormone and the test substance stored above are different amounts of growth hormone-antibody complex (GH-Ab complex) by a competitive reaction to the anti-GH antibody. And precipitated by adding a second antibody to the precipitate¹²⁵The radioactivity of I was measured and the corresponding concentration was calculated using a standard curve.

As a result, it was confirmed that the group administered with the extract of Magmundong increased the growth hormone concentration about 10-15 times compared to the control within 1 hour after injection. This proved that Macmundong extract strongly increased growth hormone compared to the control in vitro and in vivo .

Experimental Example 3 Induction of Growth Hormone in Rats 2

Eight weeks-old Sprague-Dawley rats were anesthetized at a concentration of 50 mg / kg of pentobarbital, and the extracts of McMoon-dong were dissolved in physiological saline at a concentration of 1 mg / kg of dry medicine and administered to the jugular vein of rats. Physiological saline was administered. After the blood was collected from the tail jugular vein at a predetermined time interval, growth hormone secreted using the RIA kit was measured in the same manner as in Experimental Example 2.

Minutes Control group (ng / ml) Macmundong (ng / ml) 0 10.524 13.455 10 6.064 3.960 20 4.513 3.084 30 3.861 2.723 45 3.306 4.586 60 3.255 16.178 90 2.987 4.643

As shown in Table 2 above, the medicinal extract of the present invention was administered to the jugular vein of the rat at a concentration of 1 mg / kg of dry medicinal herbs after the blood was collected from the tail vein at regular intervals to measure the growth hormone of the separated plasma. As a result, it was increased about 5 times after 60 minutes of drug administration compared with the control group, and the growth hormone secretion ability of the wormwood extract was measured within 90 minutes ( FIG. 1 ).

Experimental Example 4 Acute Toxicity in Rats

Acute toxicity test was performed using 6-week-old specific pathogen-free (SPF) SD rats. Two animals per group were suspended in 0.5% methylcellulose solution, respectively, in the munmundong extract of the present invention and administered once orally at a dose of 1 g / kg / ml. After administration of the test substance, mortality, clinical symptoms, and changes in body weight were observed. Hematological and hematological examinations were performed. Necropsy was performed to observe abdominal and thoracic organ abnormalities.

As a result, no significant clinical symptoms or dead animals were noted in all animals treated with the test substance, and no toxic changes were observed in weight changes, blood tests, blood biochemical tests, and autopsy findings. As a result, all of the extracts of the present invention did not show a toxic change up to 30 g extract / kg in mice, and the minimum lethal dose (LD ₅₀ ) of the oral administration was determined to be a safe substance having an extract of 134.34 ± 12.50 g / kg or more.

As described above, the mammundong extract of the present invention exhibits a very high growth hormone secretagogue activity, and the raw material of this extract is harmless to humans and has good absorption rate for a long time. And can be usefully used for prevention.

Claims (6)

A method for producing ganmun dong extract, which is extracted by adding alcohol or a mixed solvent of alcohol and water to Liriopis Tuber, or hydrolyzing and neutralizing with an acid or base solution, and then adding a mixed solvent of alcohol or alcohol and water. The alcohol according to claim 1, wherein the alcohol or the mixed solvent of alcohol and water is selected from the group consisting of 10 to 100% ethyl alcohol, 10 to 100% methyl alcohol and spirits. Method of Preparation of Extracts. The method of claim 1, wherein the acid or base solution is characterized in that the concentration of 0.1 to 2 N method for producing a munmundong extract. Macmundong extract prepared by the method of any one of claims 1 to 3. A growth hormone secretagogue pharmaceutical composition comprising the extract of claim 4 as an active ingredient. Growth hormone secretion health food containing the extract of claim 4 as an active ingredient.