KR20170138262A - Soybean seed extract, method for producing the same and uses thereof - Google Patents

Abstract

The present invention relates to a soybean seed extract, a method for producing the same and uses of the soybean seed extract to promote wound healing, promote proliferation of nerve cells and/or treat brain diseases and/or neurodegenerative diseases, treat breast cancer, reduce side effects of drugs interfering DNA and/or RNA replication and/or reinforce medicinal effects of drugs interfering DNA and/or RNA replication.

Description

TECHNICAL FIELD The present invention relates to a soybean seed extract, a soybean seed extract,

The present invention relates to a soybean seed extract, a method for producing the same, a method for the treatment of wound healing, promotion of neuronal cell proliferation and / or treatment of brain diseases and / or neurodegenerative diseases, treatment of breast cancer, deterioration of side effects of drugs interfering with DNA and / / RTI > and / or the use of soybean seed extract in enhancing the drug effect of drugs which interfere with DNA and / or RNA replication.

Soybeans containing soybeans and black beans (Glycine max (L.) Merrill) are one of the most important sources of oils and proteins worldwide. For example, soybeans can be processed to obtain edible oils used as salad oil, or to make margarines and shortenings. Soybean oil can also be used to make paints, linoleum, spreading, printing inks, soaps, pesticides and disinfectants. Lecithin phospholipids obtained from the by-products of the oil industry can also be used as wetting and stabilizing agents in foods, cosmetics, pharmaceuticals, leather, paints, plastics, soaps and detergents. Soy meal is a very rich feed product for livestock. In addition, the soy protein may be used in the manufacture of synthetic fibers, adhesives, fabric sizing, waterproofing, fire-fighting foam, and the like.

For medical purposes, soy has been reported to affect a number of diseases.

Soybean can be used as a nutritional supplement to control bowel, heart, kidney, liver and stomach functions. Soybean oil contains a large amount of unsaturated fatty acids and can be used to treat hypercholesterolemia. Medical lecithin derived from soybean acts as an anti-fat lipidic agent. In addition, thiagramester, also known as the antistiffness factor, and sitosterol, which is used as an alternative to diosgenin in some hypertensive treatments, are also prepared from soy. It has been suggested that isoflavones and phyto-estrogens found in soybeans have a preventive effect on various cancers including breast, prostate, and colorectal cancer (Adlercreutz, H., Phyto-oestrogens and cancer. The Lancet Oncology, 2002, Vol.3, pp. 364-373). In another document, it has been stated that in order to achieve the preventive effect of isoflavone on the development of breast cancer, 100 mg or more is to be continuously consumed in a daily dose of 1 month, and isoflavones are administered only in a high- (LLJ, Anderson KE, Grady JJ, Nagamani M; Effects of soya consumption for one month on steroid hormones in premenopausal women: implications for breast cancer risk reduction. Cancer Epidemiol Biomarkers Prev. 1996 Jan; 5 (1): 6370). Phytosterol-supplemented margarine intake is also found to lower plasma total cholesterol and LDL-cholesterol levels in middle-aged hypercholesterolemic subjects (Matvienko, OA, Lewis, DS, Swanson, M., Aendt, B., Am J Clin Nutr., 2002, 76, p. 57 64). In the present study, the serum levels of total cholesterol and LDL cholesterol in young, mildly hypercholesterolemic men, .

Some extracts from soybeans have also been reported to have pharmacological effects. The radical scavenging activity of 1,1-diphenyl-2-picrylhydrazide (DPPH) in an aqueous solution of 70% acetone to the seed coat of Akita-Zairai, a brown soybean variety, Takahata, Y., O.-Kameyama, M., Furuta, S., Takahashi, M., and Suda, I .; Highly polymerized procyanidins in brown soybean seed coat with a high radical-scavenging activity. Agric. Food Chem., 2001, 49, p. 5843 5847). Soybean, rice bran, tear grass, sesame, wheat, citron, etc., which are slowly roasted at a temperature lower than 60 ° C and fermented by adding aspergillus oryzae for 3 days to transform each component into a low molecular weight substance , Green tea, green tea leaf extract and rice malted rice extracts were found to have antioxidant effects (Minamiyama, Y., Yoshikawa, T., Tanigawa, T., Takahashi, S., Naito, Y., Ichikawa, H., and Kondo, M .; Antioxidative effects of a processed grain food, J. Nutr. Sci. Vitaminol., 1994, 40, p. The water extract of black soybeans was also tested for activity against acetaminophen-induced liver damage by measuring serum glutamate-oxalate-transaminase (sGOT) and serum glutamate-pyruvate-transaminase (sGPT) (Wu, S.-J., Wang, J.-S. and Chang, C.-H.; Evaluation of hepatoprotective activity of legumes. Phytomedicine, 2001, Vol.8 (3), p.213 219).

Some specific extracts derived from soy are known to be used as cosmetics or medicines in the treatment of some skin disorders. US Patent Application Publication No. US2002 / 0009509 A1 discloses the use of soya extract containing sphingomyelin and phospholipids in specified proportions in cosmetics for treating dry skin. This extract is prepared by treating ripe whole soya bean or oil-free soybean flour with aliphatic alcohol alone or as a mixture with water, treating with aliphatic hydrocarbons and treating with aliphatic ketones . That is, the extract is fat-soluble.

Whey and Phellodendeon amurense Ruprecht extract and selected from the group consisting of Scutellaria baicalensis Georgi, Symphytum officinale Linne and Glycine max (L.) Merrill. An acne remedy, a cosmetic production inhibitor or a cosmetic composition containing an additional one or more extracts which have been found to be effective in preventing or treating skin diseases such as rough inflammatory skin caused by acne or acne has been found 2001097842).

In addition, the fermented soybean products by microorganisms can also be used as compositions, preparations, foods, cosmetics and medicines for inhibiting the action of active oxygen (for example, Japanese Patent No. 4139132).

Although a number of uses for soy have been reported, various uses of extracts are still under development.

The present invention relates to a soybean seed extract, a method for producing the same, a method for the treatment of wound healing, promotion of neuronal cell proliferation and / or treatment of brain diseases and / or neurodegenerative diseases, treatment of breast cancer, deterioration of side effects of drugs interfering with DNA and / / RTI > and / or < RTI ID = 0.0 > DNA < / RTI > and / or RNA replication

The present invention provides an extract composition comprising a soybean seed extract prepared by a process comprising the steps of:

(a) providing a soybean seed and an extracting solution, wherein the extracting solution is water or an alcoholic solution comprising alcohol at a concentration of less than about 90 wt%;

(b) extracting the soybean seeds using an extraction solution at a pressure of less than about 1 atm and a temperature of less than about 60 캜 to obtain a crude extract; And

(c) removing the solid from the crude extract to obtain a liquid fraction.

The present invention also provides a process for preparing an extract composition as described above comprising a process for preparing a soybean seed extract comprising the steps of:

(a) providing a soybean seed and an extracting solution, wherein the extracting solution is water or an alcoholic solution comprising alcohol at a concentration of less than about 90 wt%;

(b) extracting the soybean seeds using an extraction solution at a pressure of less than about 1 atm and a temperature of less than about 60 캜 to obtain a crude extract; And

(c) removing the solid from the crude extract to obtain a liquid fraction.

The present invention also provides the use of the above-described extraction composition in the manufacture of a medicament for promoting wound healing.

The present invention also provides the use of the above-described extraction composition in the manufacture of a medicament for the promotion of neuronal cell proliferation and / or for the treatment of brain diseases and / or neurodegenerative diseases.

The present invention also provides the use of the above-described extraction composition in the manufacture of a medicament for the treatment of breast cancer.

The present invention also relates to the use of the above-mentioned extraction composition in the manufacture of a medicament for lowering the adverse effects of a drug interfering with DNA and / or RNA replication and / or enhancing the pharmaceutical effect of a drug interfering with DNA and / or RNA replication Lt; / RTI >

The invention is described in more detail in the following paragraphs. Other features, objects and advantages of the invention can be found in the detailed description and claims.

Figs. 1 to 3 show ion chromatographic spectrograms of the soybean seed extract (GMA1) according to the present invention.
4-6 illustrate ion chromatographic spectrograms of the vapor fraction of soybean seeds (GMC1) according to the present invention.
Figure 7 shows a high performance liquid chromatograph (HPLC) spectrogram of isoflavone standards.
Figure 8 shows an HPLC spectrogram of the soybean seed vapor fraction (GMC1).
Figure 9 shows an HPLC spectrogram of an extract composition comprising 0.3 part by weight of soybean seed extract (GMA1) and 1 part by weight of soybean seed vapor fraction (GMC1).
Figure 10 shows the effect of GMA1 and GMC1 on diabetic injury healing. To investigate whether GMC1 and GMA1 are effective in improving wound closure, an STZ-induced diabetic rat model was used. For comparison, two groups treated with cream base and CGS-21680 were also performed. As a result, both 0.3% GMA1 and GMC1 were effective for wound closure compared to cream base alone, and 0.3% GMA1 treated group showed better performance in wound closure than GMC1.
Figure 11 shows the effect of various doses of GMA1 in combination with GMC1 in diabetic injury healing. Experiments were conducted to investigate the best combination of GMA1 and GMC1 for the treatment of diabetic injury. As shown in Fig. 11, GMA1 was combined with GMC1 at various concentrations, 0.009%, 0.045% and 0.09%, respectively. For comparison, in all of the experimental studies, two groups treated with cream base and CGS-21680 were also performed. As a result, the combined effect of GMA1 and GMC1 at 0.009%, 0.045% and 0.09% capacity levels in wound healing was superior to GMC1 alone. The most effective combination treatment appears at the highest GMA1 dose (0.09%).
Figure 12 shows the effect of combinations of soybean seed extract and soybean seed vapor fraction on HaCaT cell migration.
Figure 13 shows cell viability of IMR-32 cells treated with GMAl. Data are expressed as mean ± SEM of 7 groups. Viability of IMR-32 cells and the positive control group treated with 25 mg / ml GMA1 was very significant, and increased significantly (p = 0.0008 GMA1, positive control, p = 0.0246). As a result, it was confirmed that GMA1 promotes cell proliferation (*** p < 0.001).
14 shows the cell survival rate of IMR-32 cells treated with GMA1. Data are expressed as mean ± SEM of 7 groups. Survival rates of IMR-32 cells treated with 25 mg / ml GMC1 and positive control were significantly increased (GMC1 p = 0.0255, positive control p = 0.0246). As a result, it was confirmed that GMC1 promotes cell proliferation (* p < 0.05).
15 shows the cell survival rate of IMR-32 cells treated with GMA1 and GMC1 (GM). Data are expressed as mean ± SEM of 7 groups. The survival rate of IMR-32 cells treated with 25 mg / ml GM and positive control was significantly increased (GMC1 p = 0.0059, positive control p = 0.0246). As a result, it was confirmed that GM promotes cell proliferation (** p < 0.01).
Figure 16 shows a diagram of a radial arm maze.
17 shows the tracing path of the demented rat in the radial maze. According to the rat's track path on day 4, the rats in the AlCl ₃ + N2 group could reach the bait position in memory after training, suggesting that the rats in the AlCl ₃ group and AlCl ₃ + cream base group were not.
18 shows a total number of error (TME) diagram of a demented rat in a radioactive maze test. AlCl ₃ group and AlCl ₃ + cream TME value of the base group has no significant difference (p> 0.05), soybean seed extract and TME value of soybean seeds day 4 not treated with the extraction composition containing vapor fraction is AlCl ₃ Group (AlCl ₃ + N 2 p = 0.0128). Extract compositions comprising soybean seed extract and soybean seed vapor fraction are found to be effective in treating dementia in rats. Data are expressed as mean ± SEM. Student's t-test and repeated ANOVA are used for statistical analysis. When comparing the AlCl ₃ group with the AlCl ₃ + cream base group, p <0.05 is denoted by * and p <0.01 is denoted by **, which means that there is a statistically significant difference.
Figure 19 shows a reference (long-term) memory error (RME) diagram of a demented rat in a radioactive maze test. AlCl ₃ group and AlCl ₃ + cream base group of RME values are not significantly different (p> 0.05), extraction compositions RME value of not the fourth day handle comprising a soybean seed extract and soybean seeds vapor fraction is AlCl ₃ Group ( p = 0.0046). The extract composition containing the soybean seed extract and the soybean seed steam fraction has an effect of restoring long-term memory of the rat. Data are expressed as mean ± SEM. Student's t-test and repeated ANOVA are used for statistical analysis. p <0.05 is denoted by *, and p <0.01 is denoted by **, which means that there is a statistically significant difference.
20 shows a total number of error (TME) diagram of vascular demented rats treated with cream base (M), GMC1 (M1) and GMC1 + 0.5% GMA1 (M3) in a radioactive maze test. The TME of the bilateral carotid artery ligation (2VO) group was significantly higher than that of the simulated group (8 days, p = 0.0013 ), suggesting that vascular dementia was successfully induced in the rat by bilateral carotid artery ligation. As a result, it was confirmed that after the treatment with M1 and M3, the TME value was remarkably improved as compared with the 2VO group (8 days, M1 p = 0.0019; M3 p = 0.0355). This suggests that M1 and M3 have efficacy in treating vascular dementia in rats. Data are expressed as mean ± SEM. Student's t-test and repeated ANOVA are used for statistical analysis. p <0.05 is denoted by *, and p <0.01 is denoted by **, which means that there is a statistically significant difference.
Figure 21 shows a reference memory (long term memory) error (RME) diagram of vascular demented rats treated with cream base (M), GMC1 (M1) and GMC1 + 0.5% GMA1 (M3) in a radioactive maze test. RME of the bilateral carotid artery ligation (2VO) group was significantly higher than that of the simulated group (8 days, p = 0.0016 ), suggesting that bilateral carotid artery ligation led to successful vascular dementia in the rat. As a result, it was confirmed that after the treatment with M1 and M3, the RME value was remarkably improved as compared with the 2VO group (8 days, M1 p = 0.0029; M3 p = 0.0171). This suggests that M1 and M3 have efficacy in treating RME of vascular dementia in rats. Data are expressed as mean ± SEM. Student's t-test and repeated ANOVA are used for statistical analysis. p <0.05 is denoted by *, and p <0.01 is denoted by **, which means that there is a statistically significant difference.
Figure 22 shows a run memory (short term memory) error (WME) diagram of vascular demented rats treated with cream base (M), GMC1 (M1) and GMC1 + 0.5% GMA1 (M3) in a radioactive maze test. The WME of the bilateral carotid artery ligation (2VO) group was significantly higher than that of the simulated group (8 days, p = 0.0111 ), indicating that bilateral carotid artery ligation resulted in successful vascular dementia in the rat. As a result, it was confirmed that, after the treatment with M1 and M3, the WME value was remarkably improved as compared with the 2VO group (8 days, M1 p = 0.0111; M3 p = 0.0139). This suggests that M1 and M3 have the efficacy of treating WME of vascular dementia in rats. Data are expressed as mean ± SEM. Student's t-test and repeated ANOVA are used for statistical analysis. p <0.05 is denoted by *, and p <0.01 is denoted by **, which means that there is a statistically significant difference.
23 shows the tracing path of vascular dementia (VD) rats in the radioactive maze. On the 8th day, the tracking pathway in the VD rats suggests that the GMO1 (M1) and GMC1 + 0.5% GMA1 (M3) groups can reach the bait position after training, depending on memory, but the 2VO and M groups are not .
24 shows a statistical diagram of brain damage rating of vascular dementia rats. Classify the degree of brain damage according to the CT image of the rat brain as 0-4 grade. As a result, it was confirmed that severe brain damage in the 2VO ( p = 0.0000 ) group was remarkably alleviated by GMC1 (M1) and GMC1 + 0.5% GMA1 (M3) treatment (M1 p = 0.0030; M3 p = 0.0238 ). Data are expressed as mean ± SEM. Student's t-test and repeated ANOVA are used for statistical analysis. p <0.05 is denoted by *, and p <0.01 is denoted by **, which means that there is a statistically significant difference.
CT grade of brain damage:
0: Obvious error in CT scan
1: There are some abnormal intensity areas in brain tissue.
2: Abnormal intensity region of more than 25% of unilateral brain tissue, or small area of abnormal intensity region in both brain tissues
3: Abnormal intensity range of more than 50% of unilateral brain tissue, midline shift, or abnormal intensity area of more than 25% of bilateral brain tissue
4: Abnormal intensity range of more than 50% of bilateral brain tissue, or presence of obvious midline shift / distortion.
Figure 25 shows a diagram of the tumor growth rate of nude mice. Tumors were induced in nude mice, grouped in mice, and then the extract composition was administered at a dose of 0.1 g / day to the tumor area and the entire skin once the tumor reached the specified volume. As a result, it is confirmed that the extract composition can lower the tumor growth rate as compared with the tumor control.
Figure 26 shows a diagram of the tumor growth rate of nude mice. Tumors were induced in nude mice, grouped in mice, and then the extract composition was administered at a dose of 0.1 g / day to the tumor area and the entire skin once the tumor reached the specified volume. As a result, it is confirmed that the extract composition can lower the tumor growth rate as compared with the tumor control.
Figure 27 shows a diagram of the tumor growth rate of nude mice. After induction of tumors in nude mice and grouping of mice, tumors were injected with CTX or GMC1 and GMA1 at a dose of 0.1 g / day to the tumor area and the entire skin when the tumor reached the specified volume. As a result, it is confirmed that the extract composition can lower the tumor growth rate as compared with the CTX control.

The present invention provides an extract composition comprising a soybean seed extract prepared by a process comprising the steps of:

(a) providing a soybean seed and an extracting solution, wherein the extracting solution is water or an alcoholic solution comprising alcohol at a concentration of less than about 90 wt%;

(b) extracting the soybean seeds using an extraction solution at a pressure of less than about 1 atm and a temperature of less than about 60 캜 to obtain a crude extract; And

(c) removing the solid from the crude extract to obtain a liquid fraction.

The invention may be more readily understood by reference to the following detailed description, examples and chemical drawings and tables of various embodiments of the invention, with reference to the accompanying description. It will be apparent to those skilled in the art that, unless explicitly stated to the contrary, the invention is not limited to specific methods of manufacture, carriers or formulations, or to the extract of the invention as a topical, oral or parenteral But are not to be construed as limited to the particular manner of formulation into the intended product or composition for administration. It is also to be understood that the terminology used herein is for the purpose of describing particular embodiments only and is not intended to be limiting.

As used in accordance with the present invention, the following terms will be understood to have the following meanings, unless otherwise stated.

Usually, ranges are expressed herein from " about "one particular value and / or" about "to another particular value. When expressed in this scope, an embodiment may encompass a range from one specific value and / or to another specific value. Likewise, when values are expressed as approximate values using the term "about ", it will be understood that the particular value constitutes another embodiment. It will be further understood that both ends of the range are meaningful to each other, and independently of the other end, as well.

"Optional" or "optionally" means that the subsequently listed phenomenon or circumstance may or may not occur and that the description includes instances where such phenomenon or circumstance occurs and instances where it does not. For example, "optionally including material" means that the material may or may not be present.

It should be noted that, as used in the specification and the appended claims, the singular forms "a," "an," and "the" include plural referents unless the context clearly dictates otherwise. Accordingly, unless the context requires otherwise, the singular terms encompass a plurality, and the plural terms encompass singular.

As used herein, the term "individual" means all animals, preferably mammals, more preferably humans. Examples of individuals include humans, primates except humans, rodents, guinea pigs, rabbits, sheep, pigs, goats, cows, horses, dogs and cats.

The term "effective amount" of the active ingredient provided herein means the amount of the ingredient sufficient to provide the desired modulation of the desired function. As will be described later, the actual required amount will vary depending on the disease state, the physical condition of the individual, the age, sex, race and body weight, the specific composition of the composition, and the formulation. Dosage regimens may be adjusted to induce optimal therapeutic response. For example, it may be administered several times in divided doses, or the dose may be proportionally reduced as prescribed for the treatment situation. That is, it is not possible to specify the actual "effective amount ". However, the appropriate effective amount can be determined by those skilled in the art through simple experimentation.

As used herein, the term " treating "or" treatment "refers to reversing, alleviating, inhibiting progression of, or reversing, alleviating, inhibiting the progression of one or more symptoms of such disorder, it means.

The term "carrier" or "excipient" as used herein is not a therapeutic agent per se, but may be used to deliver a therapeutic agent to an individual or to improve handling or storage characteristics, Refers to any substance used as a carrier and / or diluent and / or as a reinforcing agent or vehicle, which is added to a formulation to enable it to be made into a separate article such as tablets or to be easily manufactured. Suitable carriers or excipients are well known to those skilled in the art of pharmaceutical formulation or manufacture of food products. The carrier or excipient can be, for example and without limitation, a buffer, diluent, disintegrant, binder, adhesive, wetting agent, polymer, lubricant, fluidizing agent, substance added to conceal or counteract an unpleasant taste or aroma, Perfumes, and materials added to improve the appearance of the composition. Acceptable carriers or excipients include but are not limited to cytotoxic buffers, phosphate buffers, acetate buffers, bicarbonate buffers, stearic acid, magnesium stearate, magnesium oxide, sodium and calcium salts of phosphoric and sulfuric acid, magnesium carbonate, talc, gelatin, (Such as cellulose esters and cellulose alkyl esters of alkanoic acids), low melting waxes, cocoa butter, amino acids, urea, alcohols, ascorbic acid, Such as, for example, acids, phospholipids, proteins (e.g., serum albumin), ethylenediaminetetraacetic acid (EDTA), dimethylsulfoxide (DMSO), sodium chloride or other salts, liposomes, mannitol, sorbitol, glycerol or powder, Vinyl-pyrrolidone, polyvinyl alcohol, and polyethylene glycol), And other pharmaceutically acceptable materials. The carrier should not destroy the pharmacological activity of the therapeutic agent and should be non-toxic when administered at a dosage sufficient to deliver the therapeutic amount of the agent.

The extract composition according to the present invention comprises a soybean seed extract. According to the present invention, depending on the color of the seed coat of a seed, soybean may be referred to as yellow bean, vegetable soybean, white bean, peel bean, green bean, black bean; Preferably yellow soybeans or black beans. According to the present invention, soybean and belonging to the genus Soybean, preferably soybean (Glycine max (L.) Merrill), glycine formosana Hosokawa or glycine soyaoct. non Sieb. & Zucc ( Glycine soja auct. Non Sieb. & Zucc).

The seed according to the present invention preferably means a seed obtained by removing a shell from a bean pod. Generally, soybeans are hairy pods, and the shell of the pods covers the seeds. The shell of the pod is very hard and waterproof to protect the inner seeds. Methods for obtaining soybean seeds from soybeans, methods for removing peels from pods are known to those skilled in the art. Preferably, the seed according to the present invention comprises a seed coat, cotyledon and hypocotyl.

The soybean seed extract according to the present invention is prepared by a method comprising the steps of:

(a) providing a soybean seed and an extracting solution, wherein the extracting solution is water or an alcoholic solution comprising alcohol at a concentration of less than about 90 wt%;

(b) extracting the soybean seeds using an extraction solution at a pressure of less than about 1 atm and a temperature of less than about 60 캜 to obtain a crude extract; And

(c) removing the solid from the crude extract to obtain a liquid fraction.

The extraction solution for extracting soybean seeds according to the present invention is water or an alcohol solution containing alcohol at a concentration of less than about 90 wt%. Preferably, the alcohol is a C1 to C7 alcohol. The term "C1 to C7 alcohol ", as used herein, is a linear or branched, substituted or unsubstituted, mono- or polyunsaturated alcohol; Preferably, it is an unsubstituted monovalent saturated alcohol. In a preferred embodiment of the present invention, the C1 to C7 alcohols are selected from the group consisting of methanol, ethanol, n-propanol, isopropanol, n-butanol, iso-butanol, sec-butanol, tert- Butanol, 3-methyl-1-butanol, 2,2-dimethyl-1-propanol, 1-hexa 2,4-hexadien-1-ol, 2-methyl-cyclopentanol, cyclohexanol, 1-heptanol, 2-heptanol or cycloheptyl alcohol. More preferably, the C1 - C7 alcohol is methanol or ethanol; Most preferably, the C1 - C7 alcohol is ethanol. The C1 - C7 alcohol can be used alone or in combination.

The alcohol is herein preferably less than about 90% (v / v); Preferably about 5% (v / v) to about 90% (v / v); More preferably about 30% (v / v) to about 85% (v / v); , More preferably from about 50% (v / v) to about 75% (v / v).

The method according to the present invention comprises the step of (b) extracting the soybean seed using an extracting solution at a pressure of less than about 1 atm and a temperature of less than about 60 캜 to obtain a crude extract. The manner in which a portion of the seed is extracted with a solvent is well known to those skilled in the art. For example, the crude extract can be obtained by cutting the soybean seed into small pieces by any conventional means such as grinding, stirring, disturbing, cutting or finishing, and immersing the pieces in an extraction solution for extraction. The seed-cutting method in the technical field can be applied to the present invention. In a preferred embodiment of the present invention, the soybean seeds are ground into a powder. In a preferred embodiment of the present invention, the soybean seed is immersed in an extractive solution for extraction, more preferably the soybean seed is immersed in an extractive solution and subjected to ultrasonic vibration extraction.

In the method of the present invention, the soybean seed is preferably dried prior to step (b).

In the present invention, the ratio (w / v) of the seed seed to the extraction solution is not particularly limited. In one preferred embodiment of the present invention, the ratio (w / v) of the soybean seed to the extraction solution is about 1: 1 to about 1:30; More preferably from about 1: 5 to about 1: 20; Most preferably about 1:10.

According to the present invention, the extraction temperature in step (b) is less than about 60 DEG C, preferably from about 25 DEG C to about 55 DEG C; More preferably from about 30 캜 to about 50 캜; More preferably about 45 &lt; 0 &gt; C.

In one preferred embodiment of the invention, the extraction step (b) can be repeated and the extracts are collected by merging.

The process according to the invention comprises (c) removing the solids from the crude extract to obtain a liquid fraction. Methods of removing solids to obtain liquid fractions are well known to those skilled in the art and include, but are not limited to, filtration, centrifugation or precipitation.

Preferably, the process according to the invention further comprises (d) concentrating the liquid fraction obtained in step (c) to obtain a concentrated solid portion. The concentration method is well known to those skilled in the art, such as by means of a vacuum condenser.

Preferably, the process according to the invention further comprises (e) drying the concentrated solids obtained in step (d). Drying methods are well known to those skilled in the art, such as by air-drying or freeze-drying.

In one preferred embodiment of the invention, the soybean seed extract is subjected to an ion chromatographic analysis using a CarboPac PA1 Analytical (4 x 250 mm) column. The mobile phase is 87% of water and 13% of 500 mM NaOH, and the internal standard substance is maltose monohydrate. The isocratic elution is carried out at a low flow rate of 1.0 ml / min and a 0.5 second cycle. In each cycle, the analysis was performed for 0.00 seconds to 0.2 seconds, reference potential 0.1 V; 0.2 sec - 0.1 V to 0.4 sec; 0.41 seconds to -2.0 V in 0.42 seconds; 0.43 seconds, to 0.6 V; 0.44 sec - 0.5 s at -0.1 V, and the total analysis time is 55 min.

The obtained spectrogram is shown in Figs. 1-3. Peak time is shown in Table 1.

Peak time (minutes) Peak 1 Peak 2 Peak 3 Peak 4 Peak 5 Internal reference material 1 5.913 6.660 10.120 18.010 20.510 28.390 2 5.660 6.420 10.244 18.600 20.784 28.597 3 5.857 6.590 10.100 18.167 20.847 28.304

Preferably, the extract composition according to the present invention further comprises a soybean seed vapor fraction prepared by a process comprising the steps of:

(i) providing the soybean seed to a second extraction solution which is water or an alcohol solution comprising alcohol at a concentration of less than about 15 wt%; And

(ii) extracting the soybean seeds with a second extraction solution at an atmospheric pressure of less than about 1 atm and a temperature of less than about 110 &lt; 0 &gt; C, and collecting the vapor fraction.

The second extraction solution for producing the soybean seed vapor fraction according to the present invention is water or an alcohol solution containing less than about 15 wt% alcohol, preferably water. The alcohol species may be the same as the extract solution for the preparation of the soybean seed extract and is not repeated here.

The alcohol as the second extraction solution is an alcohol solution having a concentration of less than about 15% (v / v), preferably less than about 10% (v / v), more preferably less than about 5% (v / v).

The process for producing a soybean seed vapor fraction according to the present invention comprises the step of (ii) extracting the soybean seed with a second extraction solution at a pressure of less than about 1 atm and a temperature of less than about 110 캜, and collecting the vapor fraction . The extraction method may be the same as the method of producing the soybean seed extract as long as the soybean seed steam fraction is evaporated at a pressure of less than about 1 atm and a temperature of less than about 110 캜. The vapor fraction can be collected in liquid form by cooling the vapor material.

In a preferred embodiment of the present invention, the process of evaporating the soybean seed at a predetermined pressure and temperature and cooling the steam to collect the steam fraction is performed by evaporating the steam into a condensing tube to which cold water is supplied, In a rotary evaporator, which collects the vapor fraction in liquid form by passing and cooling it. This operation is simple and low cost.

According to the present invention, the ratio (w / v) of the soybean seed to the second extraction solution is not particularly limited. In a preferred embodiment of the present invention, the ratio (w / v) of the soybean seed to the second extraction solution is about 1: 1 to about 1:30; More preferably from about 1: 5 to about 1: 20; Most preferably about 1:10.

The extraction temperature in step (ii) according to the invention is less than about 110 캜; Preferably about 60 &lt; 0 &gt; C to about 110 &lt; 0 &gt; C.

In a preferred embodiment of the invention, the extraction step (ii) can be repeated and the soybean seed vapor fraction is collected by merging.

In one preferred embodiment of the invention, an ion chromatographic analysis is performed on a CarboPac PA1 Analytical (4 x 250 mm) column for the soybean seed vapor fraction. The mobile phase is 87% of water and 13% of 500 mM NaOH, and the internal standard substance is maltose monohydrate. The isocratic elution is carried out at a low flow rate of 1.0 ml / min and a 0.5 second cycle. At each cycle, the analysis was performed at a reference potential of 0.1 V at 0.00 second to 0.2 second; 0.2 sec - 0.1 V to 0.4 sec; 0.41 seconds to -2.0 V in 0.42 seconds; 0.6 V at 0.43 second; 0.44 sec - 0.5 s at -0.1 V, and the total analysis time is 55 min.

The obtained spectrogram is shown in Figs. 4-6. The peak times are shown in Table 2.

Peak time 4 2.690 5 2.587 6 2.664

In one embodiment of the invention, the content of the soybean seed extract for the extract composition is about 0.001% wt to about 10% wt; Preferably from about 0.01% wt to about 5% wt; More preferably from about 0.001% wt to about 1.5% wt. In another aspect, the content of the soybean seed vapor fraction for the extract composition is from about 0.04% wt to about 99.999% wt; Preferably from about 10% wt to about 99.9% wt; More preferably from about 30% wt to about 99% wt.

The extract composition according to the present invention is preferably a pharmaceutical composition, a food composition or a cosmetic composition.

The pharmaceutical compositions according to the invention are preferably administered locally or systemically by any method known in the art and include, but are not limited to, intramuscular, intradermal, intravenous, subcutaneous, intraperitoneal, intranasal, Oral, mucosal, or external routes. Appropriate routes, formulations, and administration schedules may be determined by those skilled in the art. In the present invention, the pharmaceutical composition may be administered orally or parenterally in the form of a liquid, a suspension, an emulsion, a syrup, a tablet, a pill, a capsule, a sustained-release formulation, a powder, a granule, an ampule, Lotions, liniments, creams, or combinations thereof. &Lt; RTI ID = 0.0 &gt; If desired, they may be sterilized or mixed with the addition of any pharmaceutically acceptable carrier or excipient, many of which are well known to those skilled in the art.

External routes of administration are also referred to herein as topical administration and include, by way of non-limiting example, administration by insufflation and inhalation. Examples of various types of formulations for topical administration include ointments, lotions, creams, gels, formulations, delivery formulations by transdermal patches, powders, sprays, aerogels, capsules or cartridges for use in inhalers or insufflations , Or suspending agents (e. G., Eye drops or nasal drops), sprayable solutions / suspensions, suppositories, pessaries, retention enema and chew or swallow tablets or pellets or liposomes or microencapsulation preparations .

Ointments, creams and gels may be formulated with an aqueous or oily base, for example, by the addition of suitable thickening, gelling and / or solvents. These bases may thus comprise, for example, oils such as water and / or oils such as liquid paraffin or arachis oil or vegetable oils such as castor oil, or solvents such as polyethylene glycol. The thickener and gelling agent that can be used according to the characteristics of the base include soft paraffin, aluminum stearate, cetostearyl alcohol, polyethylene glycol, woolfat, beeswax, carboxypolymethylene derivative, cellulose derivative, and / or glyceryl monostearate Rate and / or non-ionic emulsifiers.

Lotions may be formulated with an aqueous or oily base and will in general also include one or more emulsifying, stabilizing, dispersing, suspending or thickening agents.

External powders may be prepared using any suitable powder base, for example, talc, lactose or starch as a supplement. The drip agent may be formulated using an aqueous or non-aqueous base also comprising one or more dispersing agents, solubilizers, suspending agents or preservatives.

The spray composition may be formulated as an aqueous solution or suspension or aerosol formulation delivered using a suitable liquefied propellant from a pressurized pack, such as a metered dose inhaler. Aerosol compositions suitable for inhalation may be suspensions or solutions. The aerosol composition may optionally comprise additional surfactants, for example, oleic acid or lecithin and additional formulating agents well known in the art, such as co-solvents, for example, ethanol.

The topical preparation may be administered by application to the site of onset at least once a day; It may be beneficial to use a closed dressing over the skin area. Continuous or continuous delivery can be achieved through an adhesive reservoir system.

The cosmetic composition according to the present invention may be an aqueous formulation which is essentially composed of water; Also suitable are mixtures of water and water-miscible solvents (which may be miscible with water at 50% by weight or more at 25 DEG C), for example lower alcohols containing 1-5 carbon atoms such as ethanol or isopropanol, , Glycols containing 2-8 carbon atoms such as ethylene glycol, 1,3-butylene glycol or dipropylene glycol, water mixtures of C3-C4 ketone with C2-C4 aldehyde, and glycerin. Such an aqueous formulation is preferably in the form of an aqueous gel or a hydrogel formulation. Hydrogel formulations include thickening agents to increase the viscosity of the liquid solution. Examples of thickeners include, but are not limited to, carbomers, cellulosic materials, gums, algin, agar, pectin, carrageenan, gelatin, minerals or modified mineral thickeners, polyethylene glycols and polyalcohols, polyacrylamides and other polymeric thickeners . Thickening agents are preferably used which impart stability to the composition and provide optimal flow characteristics.

The cosmetic composition according to the present invention may be in the form of an emulsion or cream formulation. The composition may comprise an emulsifying surfactant. Such surfactants can be selected from anionic surfactants and non-ionic surfactants. The content and function (emulsification) of surfactants are described in Encyclopedia of Chemical Technology, Kirk-Othmer, volume 22, pp. 333-432, 3rd edition, 1979, Wiley, especially pages 347-377, for anionic surfactants and non-ionic surfactants.

Surfactants preferably used in the cosmetic composition according to the present invention include nonionic surfactants; Fatty acids, fatty acid alcohols, polyethoxylated or polyglycerolated fatty alcohols, such as polyethoxylated stearyl or cetylstearyl alcohol, fatty acid esters of sucrose, alkylglucose esters, especially poly Oxyethylenated fatty acid esters, and mixtures thereof; Anionic surfactants; Amines, C16-C30 fatty acids neutralized with aqueous ammonia or alkali salts, and mixtures thereof. Surfactants which enable the production of oil-in-water or water-in-oil wax emulsions are preferably used.

The cosmetic composition according to the present invention may further comprise an effective amount of a physiologically acceptable antioxidant selected from the group consisting of butylated p-cresol, butylated hydroquinone monomethyl ether and tocopherol.

The cosmetic composition according to the present invention may further comprise natural or modified amino acids, natural or modified sterol compounds, natural or modified collagen, silk protein or soy protein.

The cosmetic compositions according to the present invention are preferably formulated for topical application to keratinous materials such as skin, hair, eyelashes or nails. This may be achieved in all presentation forms commonly used in this application type, especially in the form of aqueous or oily solutions, oil or water emulsions, silicone emulsions, microemulsions or nanoemulsions, aqueous or oily gels or liquids, pastes or solid anhydrous products Lt; / RTI &gt;

The cosmetic compositions according to the present invention may have high or low fluidity and may have the appearance of white or colored creams, ointments, milks, lotions, serums, pastes, mousses or gels. The composition may optionally be topically applied to the skin in the form of an aerosol, patch or powder. It may also be in solid form, for example in the form of a stick. It can be used as a skin care product and / or as a makeup product. As another example, it can be made with a shampoo or conditioner.

The cosmetic composition according to the present invention comprises, in a known manner, conventional additives and adjuvants in cosmetics, for example, hydrophilic or lipophilic gelling agents, preservatives, antioxidants, solvents, flavoring agents, fillers, pigments, .

The extract composition can be added to a conventional food composition (i.e., edible food or beverage or its precursor) in the process of manufacturing the food composition. The extract composition of the present invention can be added to almost all food compositions. Food compositions to which the extract compositions of the present invention may be supplemented include but are not limited to candies, baked good, ice cream, dairy products, sweet and savory snacks, snack bars, meal substitutes, fast food, , Cotton, canned foods, frozen foods, dried foods, refrigerated foods, oils and fats, infant foods, or soft foods applied to breads.

The present invention also provides the use of the above-described extraction composition in the manufacture of wound healing accelerators.

The invention also provides a method of promoting wound healing in an individual comprising administering an effective amount of the above-described extraction composition, optionally, to a subject in need of accelerated wound healing with a pharmaceutically acceptable carrier or excipient do.

Preferably, the extract composition according to the present invention promotes wound healing by promoting skin cell migration. In one preferred embodiment of the invention, the extract composition can promote migration of keratinocytes, wherein the skin cells are preferably keratinocytes. In another aspect, in an animal model according to the present invention, the extract composition can promote wound healing in a diabetic patient, wherein the wound is preferably a diabetic injury.

In one embodiment of the invention, the composition comprising the soybean seed extract and the soybean seed vapor fraction has a significant effect on wound closure (p < 0.5) compared to the cream base, and the soybean seed extract has a It exhibits slightly better performance in wound closure.

In another embodiment of the present invention, the soybean seed extract is used at various dose levels in combination with the soybean seed vapor fraction to investigate its effect on diabetic wound healing. As a result, it was confirmed that the combination effect of the soybean seed extract and the soybean seed steam fraction was superior to the soybean seed steam fraction alone in wound healing. The most effective combination treatments are found in the case of using the soybean seed extract at the highest dose.

In one embodiment of the invention, the soybean seed extract and the soybean seed vapor fraction are used alone or in combination to analyze the effect of promoting migration of dermal keratinocytes. As a result, the combined effect of the soybean seed extract and the soybean seed steam fraction was found to be superior to the soybean seed extract or the soybean seed steam fraction alone in promoting migration of keratinocytes.

The present invention also provides the use of the above-described extraction composition in the manufacture of a medicament for the promotion of neuronal cell proliferation and / or for the treatment of brain diseases and / or neurodegenerative diseases.

The present invention also relates to a method of promoting the proliferation of neurons and / or preventing brain diseases and / or neuropathies in a necessary individual, comprising administering to said individual an effective amount of the aforementioned extract composition and optionally a pharmaceutically acceptable carrier or excipient A method for treating a degenerative disease.

Preferably, the nerve cell is a central nervous system cell; More preferably, it is a brain nerve cell; Even more preferably, it is a neuroblastoma cell.

The brain diseases and / or neurodegenerative diseases according to the present invention are preferably selected from the group consisting of vascular dementia, Alzheimer's disease, Parkinson's disease, cerebrovascular disease, inflammatory brain injury, inflammatory reaction around brain injury, brain lesion, swelling ventricle, and ventricular dilatation.

In one embodiment of the invention, human neuroblastoma cells are treated using an extract composition comprising a soybean seed extract, a soybean seed vapor fraction, and a soybean seed extract and a soybean seed vapor fraction. As a result, after treating the extract composition containing the soybean seed extract, the soybean seed steam fraction, and the soybean seed extract and the soybean seed vapor fraction, the cell viability is remarkably increased. Therefore, the extract composition containing the soybean seed extract, the soybean seed steam fraction and the soybean seed extract and the soybean seed steam fraction has an effect of promoting neuronal cell proliferation.

In another embodiment of the present invention, in a radioactive labyrinth test of a demented rat model, after treating an extract composition comprising a soybean seed extract and a soybean seed vapor fraction, total memory errors and reference memory errors were significantly Respectively. This shows that the extract composition containing the soybean seed extract and the soybean seed vapor fraction has the effect of treating demented rats.

In another embodiment of the present invention, when the extract composition containing the soybean seed extract and the soybean seed vapor fraction was treated in the radioactive maze test of the demented rat animal model, total memory errors and reference memory errors were significantly Low. This shows that an extract composition containing a soybean seed extract and a soybean seed vapor fraction is effective for treating demented rats.

In the vascular dementia model, the results showed that memory impairment due to vascular dementia improved significantly after treatment of the extract composition according to the present invention. Both the soybean seed steam fraction and the extract composition including the soybean seed extract and the soybean seed vapor fraction improve the performance memory error, the reference memory error, and the total memory error. Extraction compositions comprising a soybean seed extract and a soybean seed vapor fraction have a better effect in treating memory disorders due to vascular dementia compared to soybean seed vapor fraction alone.

The present invention also provides the use of the above-described extraction composition in the manufacture of a medicament for the treatment of breast cancer.

The present invention also provides a method of treating breast cancer in a subject in need thereof, comprising administering to the subject an effective amount of the aforementioned extract composition and optionally a pharmaceutically acceptable carrier or excipient.

Preferably, the breast cancer is a breast cancer of the p53 mutation type.

In one preferred embodiment of the invention, an animal model is established for simulating the situation at a primary site for observing tumor growth in mice, using a human breast cancer cell line of p53 mutation type. When simulating administration in humans, the extract composition according to the present invention is administered at various concentrations. The extract composition according to the present invention is confirmed to have an effect of suppressing tumor growth rate in a tumor-bearing mouse.

The present invention also relates to the use of the above-mentioned extraction composition in the manufacture of a medicament for lowering the adverse effects of a drug interfering with DNA and / or RNA replication and / or enhancing the pharmaceutical effect of a drug interfering with DNA and / or RNA replication Lt; / RTI &gt;

The present invention also relates to a method for the prevention and / or amelioration of side effects of drugs interfering with DNA and / or RNA replication, and / or the administration of a medicament, In a subject in need of enhancing the effect of a drug interfering with the DNA and / or RNA replication of the drug, and / or enhancing the medicinal effect of interfering with DNA and / or RNA replication of the drug, and / . &Lt; / RTI &gt;

The term "drug interfering with DNA and / or RNA replication" refers herein to a drug that kills tumor cells by interfering with DNA and / or RNA replication processes that are essential in mitosis. Preferably, drugs that interfere with DNA and / or RNA replication include, but are not limited to, cyclophosphamide, temozolomide, hexamethylmelamine, platinum complex, bleomycin, vinblastine, vincristine, ), Paclitaxel, docetaxel, polynic acid antagonists, purine antagonists or pyrimidine antagonists.

In one preferred embodiment of the invention, human breast cancer cell lines are used to establish animal models for simulating conditions at primary sites. Compared with tumor-bearing animals that received cyclophosphamide, the extract compositions according to the present invention enhance the effect of inhibiting tumor growth of chemotherapeutic agents in tumor-bearing mice. The extract composition according to the present invention inhibits tumor growth, enhances the tumor size reduction effect of the chemotherapeutic agent, and alleviates the pain by relieving tumor pressure, so that the patient can have a better quality of life.

The following examples are provided to aid those skilled in the art in practicing the present invention.

Example

Soybean seed extract (GMA1)

Seeds of soybean (Glycine max (L.) Merr.) Were pulverized into powders and 70% by weight of ethanol or distilled water was used as the extraction solution; The ratio (w / v) of the soybean seed to the extract solution was about 1:10. The soybean seeds were extracted at about 1 atm atmospheric pressure and about 45 캜 to give crude extracts. The solids were removed from the crude extract to give a liquid fraction. The liquid portion was further concentrated using a pressure-reducing condenser to obtain a concentrated solid. The concentrated solids were further dried at 70 &lt; 0 &gt; C.

The soybean seed steam fraction (GMC1)

Seeds of soybean (Glycine max (L.) Merr.) Were ground into powder and 2% by weight of ethanol or distilled water was used as the second extraction solution; The ratio (w / v) of the soybean seed to the second extraction solution was about 1:10. The soybean seeds were evaporated using a rotary evaporator (EYELA N-1000S, 1000S-W) at a pressure of less than about 1 atm and a temperature of 90 캜 and passed through a condensing tube to which cold water was supplied to obtain a vapor fraction.

Analysis of soybean seed extract (GMA1) and soybean seed steam fraction (GMC1)

The resulting soy seed extract (GMA1) and soybean seed vapor fraction (GMC1) were subjected to ion chromatography analysis on a CarboPac PA1 Analytical (4 x 250 mm) column. The mobile phase was 87% of water and 13% of 500 mM NaOH; The internal standard was maltose monohydrate. The isocratic elution was carried out at a low speed of 1.0 ml / min with a 0.5 second cycle. At each cycle, the analysis was performed at a reference potential of 0.1 V at 0.00 second to 0.2 second; 0.2 sec - 0.1 V to 0.4 sec; 0.41 seconds to -2.0 V in 0.42 seconds; 0.6 V at 0.43 second; 0.44 seconds - 0.5 seconds at -0.1 V, and the total analysis time was 55 minutes.

The spectrogram of the soybean seed extract (GMA1) is shown in Figs. 1-3. Peak time is shown in Table 1.

The spectrogram of the soybean seed steam fraction (GMC1) is shown in Figures 4-6. The peak times are shown in Table 2.

Soybean seed extract (GMA1) contains very small amounts of isoflavones, and soybean seed steam fraction (GMC1) does not contain isoflavones.

High performance liquid chromatography (HPLC) was used to analyze whether soybean seed extract (GMA1) and soybean seed steam fraction (GMC1) contained isoflavone.

The HPLC conditions are listed below:

Apparatus: Hitachi HPLC CM5000 Series; Pump: CM5110; Detector: CM5430 (DAD); Automatic supply system: CM5210; Column oven: CM5310; Software: OpenLab.

Column: RP C ₁₈ , 4.6 x 250 mm 5 탆; Detection wavelength: UV 254 nm; Flow rate: 0.8 min / ml; Column oven temperature: 30 占 폚; Concentration Gradient: Shown in Table 3.

Time (minutes) Solvent (%) Dissolution 1 Dissolution 2 0 95 5 10 85 15 25 77 23 35 72 28 40 20 80 45 20 80 46 95 5 55 95 5

Elution 1: 1% formic acid + 0.01% trifluoroacetic acid (TFA) / water

Elution 2: 1% formic acid + 0.01% trifluoroacetic acid / acetonitrile

The HPLC spectrogram of the isoflavone standard material is shown in FIG. 7, and peaks were observed at the retention times of 18.853 minutes and 24.693 minutes.

The HPLC of the soybean seed steam fraction (GMC1) is shown in Fig. 8, and there was no peak.

The HPLC spectrogram of the ointment agent containing 0.3 part by weight of the soybean seed extract (GMA1) and 1 part by weight of the soybean seed steam fraction (GMC1) is shown in FIG. 9, and the retention time was 17.307 minutes, 19.313 minutes, 20.267 minutes, 20.853 minutes, And a peak at 25.247 min.

Comparing FIGS. 7 to 9, in FIG. 8 there is no peak in the HPLC spectrogram of the soybean seed vapor fraction (GMC1), the soybean seed vapor fraction (GMC1) is free of isoflavone; In Fig. 9, the peak corresponding to the isoflavone fingerprint absorption peak appears to be minimal in the HPLC spectrogram of the extract composition containing 0.3 part by weight of the soybean seed extract (GMA1) and 1 part by weight of the soybean seed vapor fraction (GMC1). After the conversion, the amount of daidzin was 4.8 μg / ml and the amount of genistin was 8.23 μg / ml, which was far from the pharmacologically effective amount of isoflurane. Therefore, only soybean seed extract (GMA1) contains very small amounts of isoflavone, and its pharmaceutical effect is not expressed by a very small amount of isoflavone.

Extraction composition promoting wound healing

Materials and Methods:

SD male rats weighing 250 g - 300 g, weighing about 8 weeks, were obtained from the Taiwan National Laboratory Animal Center. After 7 days of isolation, they were moved when their body weight was over 350 g. The rat was identified as the ear notch of the animal. Each cage tee shows the cage number, experiment number, gender, and group name. Rats were housed in polycarbonate cages at two AAALAC accredited animal facilities, two per cage.

The environmental conditions are as follows: temperature 21 ± 2 ° C; Humidity: 50 ± 20%; Light cycle: 12 hours, 12 hour dark conditions. During the entire study period, experimental rodent feed 5001 (PMI ^R Nutrition International, Inc., MO, USA) was fed ad libitum. The tap water was fed freely through a bottle attached to the cage.

Experimental Method:

Samples (test articles) were prepared according to Table 4.

Experiment 1 group Animal / Male / Sex Test article and dose One Diabetic rat / 5 / male Cream base 2 Diabetic rat / 5 / male CGS-21680 10 μg / wound 3 Diabetic rat / 5 / male GMC1 4 Diabetic rat / 5 / male 0.3% GMA1 Experiment 2 group Animal / Male / Sex Test article and dose One Diabetic rat / 5 / male Cream base 2 Diabetic rat / 5 / male CGS-21680 6.7 / / wound 3 Diabetic rat / 5 / male GMC1 4 Diabetic rat / 5 / male GMC1 + 0.009% GMA1 5 Diabetic rat / 5 / male GMC1 + 0.045% GMA1 6 Diabetic rat / 5 / male GMC1 + 0.09% GMA1

CGS-21680: A specific adenosine A _2A subtype receptor agonist, prepared by adding 1.67 mg of CGS-21680 in 248.4 g of a cream base.

Diabetic rats: Streptozotocin (STZ, 65 mg / kg) was administered in a single dose via intravenous injection into rats weighing 300 g or more. STZ-induced rats exhibiting high blood glucose (> 300 mg / dL) for 2 months were selected for wound closure testing.

Trauma Surgery for Diabetic Rats Diabetic rats weighing less than 300 g were excluded and the rats were randomly extracted into six groups. Thereafter, pentobarbitalolate was anesthetized and the site to be operated (epidermis) was removed. Three rats (4, 6, and 8 cm apart from the two scapulae) in the dorsal central area of each rat were incised (full thickness) using a round cutting blade.

Medication and wound suture measurements: Animals were weighed and wound area measured at 2-day intervals to assess wound healing. Twice a day, the drug was applied to each wound site and the wound was covered with cheesecloth. The hood was put on the neck of the rat to prevent the animal from scratching the wound. Photographs of wound sites were taken at 4, 6, 8, 10, 12 and 14 days for wound closure measurements. At the time of shooting, a standard ruler was placed next to the wound. Before analyzing the wounds with Image pro (Media cybernetics), standard lengths were standardized in the photographs to avoid errors due to differences in photographic distance.

Data analysis and statistics: Three scars located on the back of the rat were analyzed with Image pro. The original wound area was the area of day 0. The wound area of each time zone was limited in the original wound area, and then divided into the original wound area, and the wound suture ratio was obtained. The wound sutures of each rat are shown by the average suture rate of the three wounds of each rat. Data were expressed as mean ± standard error (SEM). The p-values for the test results were calculated by performing a t-test with statistical software (SYSTAT, Systat software Inc). P <0.05 means that there is a significant difference, which is marked with *. P <0.01 means that there is a significant difference, indicated as **. P <0.001 means that there is a very significant significant difference, denoted by ***.

result:

After the drug treatment, the wound area was measured at various time intervals, and the wound closure rate was analyzed by statistical software and shown in Table 5 and FIG. On day 4, the wound closure rates of each group, cream base, CGS-21680, GMC1 and 0.3% GMA1 were -83.33 ± 6.60 (%), 21.87 ± 5.61 (%), -11.44 ± 4.34 3.65 (%). There was a significant difference between the experimental group and the cream base group during the experimental period. From 8th to 10th day, wound closure rates were significantly decreased from -4.10 ± 3.04 (%) to 40.15 ± 6.47 (%) in GMC1 group and 3% GMA1 group and from 62.19 ± 3.85% ). At 14 days, the wound closure rates of the GMC1 and 0.3% GMA1 groups were significantly increased to 86.20 ± 1.88 (%) and 91.21 ± 2.23 (%), respectively, compared to 58.92 ± 13.14% of the cream base.

Table 5: Comparison of wound closure rates of diabetic rats treated with GMC1 and 0.3% GMA1

Day Cream base CGS-21680 GMC1 0.3% GMA1 Mean ± SEM Mean ± SEM Mean ± SEM Mean ± SEM 4 -83.33 + - 6.60 21.87 ± 5.61 ^*** -11.44 + 4.34 ^*** -7.09 ± 3.65 ^*** 6 -65.84 ± 11.19 25.45 + - 4.46 ^*** 6.74 ± 4.86 ^*** 4.15 ± 3.19 ^*** 8 -63.32 + - 23.22 45.88 ± 4.28 ^*** 4.10 ± 3.04 ^* 21.21 + - 5.52 ^** 10 -10.08 + - 16.87 93.80 ± 2.79 ^*** 40.15 + - 6.47 ^* 62.19 + - 3.85 ^** 12 30.85 ± 20.08 91.15 ± 2.72 ^** 74.36 ± 2.04 ^* 82.71 + - 2.82 ^* 14 58.92 + - 13.14 98.74 ± 0.60 ^** 86.20 ± 1.88 ^* 91.21 + - 2.23 ^*

Note: Each figure is expressed as mean ± SEM. The comparison between the two groups was significant at ** p <0.01 and *** p <0.001 by Student's t test.

These results show that both 0.3% GMA1 and GMC1 are effective in wound closure compared to cream base alone and 0.3% GMA1 treatment group exhibits slightly better wound closure performance than GMC1.

The best combination of GMA1 and GMC1 was investigated in promoting diabetic wound closure. GMA1 was combined with GMC1 at different dosage levels, i.e. 0.009%, 0.045% and 0.09%. For comparison, Cream Base and CGS-21680 groups were also involved.

After the drug treatment, the wound area was measured at various time points, and the wound closure rate was analyzed by statistical software and shown in Table 6 and FIG. As shown in Table 6 and FIG. 11, a statistically significant increase was observed with increasing dose levels of GMA1, i.e. 0.009%, 0.045% and 0.09%, in the combination drug group compared to the cream base group . This result shows that the combination effect of GMC1 with 0.009%, 0.045% and 0.09% of various dose levels of GMA1 in wound healing is more effective than GMC1 alone. The most effective combination treatment is identified at the highest dose (0.09%) of GMA1.

Table 6: Comparisons of wound closure rates in diabetic rats treated with 0.009% to 0.09% GMA1 and GMC1

Note: Each figure is expressed as mean ± SEM. The comparison between the two groups showed a significant difference at ** p <0.01 and *** p <0.001 by Student's t test.

An extract composition for promoting migration of skin cells

HaCaT cells (human dermal keratinocytes) were used to analyze the effect of the extract composition on promoting skin cell migration.

Samples for cell migration analysis were as follows:

A1-0.3: GMA1 containing 0.3 占 퐂 / mL

A1-0.3-CSTC-2500X: containing 2500-fold diluted GMC1 and GMA1 at 0.3 占 퐂 / mL

Cell line: HaCaT

Medium: DMEM supplemented with 10% fetal bovine serum

Positive control: CGS-21680

Cell culture: HaCaT cells were cultured in DMEM supplemented with 10% fetal bovine serum at 37 ° C and 5% CO ₂ , and sub-cultured twice a week.

Wound healing analysis: Oris cell migration assay kit was used. The cells were inoculated into 96-well plates ( ⁴ x 10 ⁴ cells / well) equipped with a stopper, and cultured in a carbon dioxide incubator. After overnight incubation, the stopper was removed to create a wound, and a fresh medium, test article, or medium containing CGS-21680 (positive control) was added. After treatment for 0, 16, and 24 hours, the cells were observed under a light microscope at 100x magnification and photographed for wound healing monitoring.

Analysis: We used image j software to quantify wound healing area. The area of the 0 time zone was set as the wound original size. After a certain period of time, the wound area gradually decreased, and the degree of wound healing was determined by calculating the wound area (recovery ratio = area difference between T0 and T16 or T24 time / T0 area) relative to the original area at time 0, The effect of the drug on the activity was evaluated.

Statistics: Each group was repeated more than 4 times. Data are expressed as mean ± standard deviation (SEM). The significance of test results was calculated by t-test.

result:

HaCaT cell migration assay was used to compare the effects of GMC1 and GMA1 in promoting wound healing. The combination effect of 2500-fold diluted GMC1 and 0.3 / / mL of GMA1 (A1-0.3-CSTC-2500X) was superior to A1-0.3 alone in promoting HaCaT cell migration (Fig. 12).

Extract composition for promoting proliferation of nerve cells

Materials and Methods:

Cell line: human neuroblastoma cell IMR-32 (purchased from Taiwan Industry Research and Development Institute)

Reagents: HyClone ™ DMEM / High Glucose medium (Thermo Scientific), fetal bovine serum (FBS, Gibco®), HyClone® phosphate buffered saline (PBS, Thermo Scientific), antibiotic antifungal solution (Pen / Strep / Fungizone 100x; (Thermo Scientific), trippine blue, 3- (4,5-dimethylthiazol-2-yl) -2,5-diphenyltetrazolium bromide (MTT), and dimethylsulfoxide (DMSO, Sigma).

Equipment: Laminar Flow (VERTICAL HF-4BH).

Cell culture: IMR-32 cells were cultured in DMEM / high glucose medium (HyClone ™) supplemented with 10% fetal bovine serum at 37 ° C, humidified air and 5% CO ₂ .

IMR-32 cells were inoculated into 96-well plates at 5 x 10 ⁴ density and cultured in 100 [mu] l of culture medium supplemented with 10% FBS for 24 hours. After dosing, the culture medium was replaced with 100 μl of medium supplemented with 5% FBS, and then 25 mg / ml GMC1, 25 mg / ml GMA1, GMC1 + 0.3% GMA1 (GM) Control) and 10% FBS (positive control). Each group was repeated seven or more times in 96 well plates and incubated for 72 hours for MTT analysis.

MTT analysis: The culture dish was taken out from the incubator and moved to the laminar flow hood. In each well, the medium was replaced with 100 μl of serum-free medium supplemented with 0.5 mg / ml MTT reagent, and cultured for 30-60 minutes. The same volume of DMSO was added to each well and shaken for 5 minutes. Absorbance was measured at 570 nm in an ELISA reader.

Data Analysis and Statistics: The results were analyzed with GraphPad Prism 6 and statistically analyzed by T-test. Data are expressed as the cell survival rate ± SEM.

result:

Cell viability is shown in Tables 7-9 and 13-15. Cell viability was measured after treatment with 25 mg / mL GMA1 (Table 7 and Figure 13), 25 mg / mL GMC1 (Table 8 and Figure 14) or GMC1 + 0.3% GMA1 (GM, Table 9 and Figure 15) In comparison, it increased remarkably.

GMA1 Group Control group 25 mg / mL Positive control group Mean ± SEM Mean ± SEM Mean ± SEM One 99.2 ± 17.2 106.7 8.3 137.9 ± 9.8 2 110.1 ± 6.9 119.7 ± 6.7 128.7 13.8 3 86.3 ± 5.6 123.2 ± 6.3 ** 94.5 ± 2.8 4 98.3 ± 1.8 130.2 ± 7.8 ** 119.0 + - 7.7 * 5 88.6 ± 2.6 114.2 ± 5.8 ** 90.7 ± 1.6 6 94.8 ± 1.9 111.4 ± 6.1 * 105.7 ± 3.1 * 7 92.1 ± 5.7 105.7 ± 10.2 128.2 ± 2.9 *** Average 95.6 ± 3.0 115.9 ± 3.4 *** 115.0 + - 6.9 *

GMC1 Group Control group 25 mg / mL Positive control group Mean ± SEM Mean ± SEM Mean ± SEM One 99.2 ± 17.2 94.1 ± 3.3 137.9 ± 9.8 2 110.1 ± 6.9 144.1 ± 8.2 * 128.7 13.8 3 86.3 ± 5.6 120.0 + - 11.0 * 94.5 ± 2.8 4 98.3 ± 1.8 149.2 ± 6.4 *** 119.0 + - 7.7 * 5 88.6 ± 2.6 106.4 ± 7.2 90.7 ± 1.6 6 94.8 ± 1.9 101.5 ± 3.1 105.7 ± 3.1 * 7 92.1 ± 5.7 107.8 ± 6.3 128.2 ± 2.9 *** Average 95.6 ± 3.0 117.6 ± 8.1 * 115.0 + - 6.9 *

GM Group Control group 25 mg / mL Positive control group Mean ± SEM Mean ± SEM Mean ± SEM One 99.2 ± 17.2 117.2 ± 11.6 137.9 ± 9.8 2 110.1 ± 6.9 115.1 ± 12.5 128.7 13.8 3 86.3 ± 5.6 103.0 ± 6.0 94.5 ± 2.8 4 98.3 ± 1.8 118.2 ± 10.1 119.0 + - 7.7 * 5 88.6 ± 2.6 106.1 ± 5.5 * 90.7 ± 1.6 6 94.8 ± 1.9 99.9 ± 2.0 105.7 ± 3.1 * 7 92.1 ± 5.7 105.7 ± 1.6 * 128.2 ± 2.9 *** Average 95.6 ± 3.0 109.3 ± 2.8 ** 115.0 + - 6.9 *

Comments in Table 7 - 9: Each figure is expressed as mean ± SEM. Comparisons between different doses of the two groups were statistically analyzed using a T-test. p <0.05 means that there is a significant difference and is marked with *. p <0.01 means that there is a significant difference, indicated by **. p <0.001 means that there is a very significant difference, indicated by ***.

Extract compositions for treating brain and / or neurodegenerative diseases

(I) Induced dementia:

Materials and Methods:

Male rats weighing 250 g - 300 g at about 6 weeks of age were obtained from BioLASCO, Taiwan. After 7 days of isolation, they were moved when their body weight was over 300 g. Rats were identified as tail tattoos. Each cage tee shows the cage number, experiment number, gender, and group name. Rats were placed in polycarbonate cages in animal facilities with two cages per cage.

The environmental conditions are as follows: temperature 25 ± 1 ° C; Humidity: 60 ± 5%; Light cycle: 12 hours, 12 hour dark conditions. Altromin 1324 FORTI (Germany) was fed ad libitum throughout the experiment. The tap water was fed freely through a bottle attached to the cage.

The groups are shown in Table 10.

group Dementia induction Test article normal Do not none AlCl ₃ AlCl ₃ AlCl ₃ control AlCl ₃ + cream base AlCl ₃ Cream base AlCl ₃ + N 2 AlCl ₃ 0.5% GMA1 + GMC1

Dementia induction:

8-week-old rats were orally administered with 15 mg / mL AlCl ₃ solution daily at a dose of 100 mg / kg for 11 weeks to induce dementia mimicry symptoms. The rats were divided into the groups shown in Table 10 and topically applied topically to the nasal mucosa for 5 to 11 weeks by massaging the test article in the head and neck area for 30 seconds. Training was started at 11 weeks, and the memory was evaluated with a radioactive maze at 11 weeks. AlCl ₃ was continuously supplied to the rats during the treatment period.

Radioactive Labyrinth: A radioactive labyrinth is one of the most common methods of measuring spatial learning and memory in the rat. The radioactive labyrinth was designed by Olton in the seventies. The hungry rats are trained to remember the feed position in the maze for a period of time, based on the driving force to check the feed at the end of each passage. This can simultaneously measure the run memory (referred to as short-term memory) and the reference memory (referred to as long-term memory). The radioactive maze is shown in Fig. Size: (A) 122 cm; (B) 47 cm; (C) 30 cm; (D) 10 cm; (E) 20 cm.

The rats were adapted to the environment for one week. The weight of each rat was measured and fasted for 24 hours. Prior to the experiment, the rats were kept at 80-85% of their original weight; 60% of normal diets were given daily (twice a day) after training. On days 1 and 2, bait was sprinkled on each aisle arm and central platform. Four rats were placed on the central platform at the same time, and the bait was searched for 10 minutes. On days 3 and 4, bait was placed at the end of each passage arm. Each rat was separated from the central platform and allowed to navigate the maze until the feed was found. If the rat could not find the feed within 10 minutes, the training was stopped. On days 5-14, baits were placed at the end points of four fixed arms. Each rat was placed on a central platform and the maze was searched until all the baits in the four arms were found. When the rats could not find the feed within 10 minutes, the training was stopped. Paths recorded in the arms of the labyrinth were recorded and analyzed automatically. Each rat was trained twice a day, and training was conducted at 1 hour intervals.

The following three indicators were analyzed:

a. Execution memory (short-term memory) error (WME): Number of re-entry into the bait arm.

b. Reference memory (long-term memory) error (RME): Number of re-entry into the baitless arm.

c. Total Memory Error (TME): WME + RME

These three indicators represent the learning and memory capacities of the rat, and will increase markedly with brain damage levels.

Data analysis and statistics: Student t-test and repeated ANOVA were used for statistical analysis. Data are presented as mean ± SEM for each group. The data of AlCl ₃ group were compared with the normal group and the statistical significance was calculated. In addition, the data in the group compared to the AlCl ₃ AlCl ₃ + N2 groups, statistical significance was calculated. If p < 0.05, it is indicated by *. If p < 0.01, ** is indicated.

result:

It has been reported that heavy metal-induced dementia is similar to Alzheimer's disease. Both of these induce the formation of amyloid precursor proteins, which produce the nervous and nerve fiber knots. Therefore, in order to understand the effect of an extract composition containing a soybean seed extract and a soybean seed vapor fraction on Alzheimer-like disease, heavy metal-induced dementia was adopted as an animal model. Results are shown in Figures 17-19. In FIG. 17, according to the tracking path on the 4th day of rats, the rats of the AlCl ₃ + N 2 group could reach the bait position as memory after training and not the rats of the AlCl ₃ group and the AlCl ₃ + cream base group. According to FIG. 18, the TME values of the AlCl ₃ group and the AlCl ₃ + cream base group were not significantly different (p> 0.05). After treating the extract composition containing the soybean seed extract and the soybean seed vapor fraction, Value was significantly lower than that of the AlCl ₃ group (AlCl ₃ + N 2 p = 0.0128). Extracted compositions comprising a soybean seed extract and a soybean seed vapor fraction appear to be effective in treating demented rats. According to FIG. 19, there was no significant difference (p> 0.05) between the RME values of the AlCl ₃ group and the AlCl ₃ + cream base group, and the extract composition containing the soybean seed extract and the soybean seed vapor fraction was treated. Value was significantly lower than the AlCl ₃ group (p = 0.0046). This means that the extract composition containing the soybean seed extract and the soybean seed steam fraction is effective for the long-term memory recovery of rats. According to the results, errors in the AlCl ₃ group in the radio-maze test were significantly higher than in the control, indicating that AlCl ₃ -induced dementia was successfully induced in the rat. Memory impairment associated with reference memory has also been significantly improved following treatment of the extract composition comprising the soybean seed extract and the soybean seed vapor fraction. This means that the extract composition comprising the soybean seed extract and the soybean seed vapor fraction is effective for treating dementia.

(I) Vascular dementia

Materials and Methods:

The groups are shown in Table 11.

group Dementia induction Test article imitation Do not imitation 2VO Ligation of both carotid arteries Both carotid artery ligation controls M Ligation of both carotid arteries Cream base M1 Ligation of both carotid arteries GMC1 M3 Ligation of both carotid arteries GMC1 + 0.5% GMA1

Rat and animal facilities are identical to the dementia induction test of (I).

The rats were anesthetized using a 1: 1.5 ketamine-lrompun mixture (0.1 mL / 100 g, i.p.) and fixed on the operating table. Both sides of the carotid artery were exposed by midline incision in the dorsal neck area and double sutured with a silk suture. The wound was sutured and the rats were placed under a warm light until they were restored.

The cream base (M), GMC1 (M1) and GMC1 + 0.5% GMA1 (M3) were massaged for 30 seconds while topically applied to the nose area and the nasal mucosa (2g / rat). 4 weeks of treatment was continued and the memory of the rat was measured in the last two weeks of the radioactive maze. The radioactive maze test was performed in the same manner as described in (I) induction of dementia.

Data analysis and statistics: Unpaired Student t-test and 2-way ANOVA were used for statistical analysis. Data are the mean ± SEM of each group. The data of the simulation group were compared with the 2VO group and statistical significance was calculated. The drug treatment groups were compared statistically with 2VO groups. p <0.05 means that there is a significant difference and is marked with *. p <0.01 means that there is a very significant difference, indicated by **.

result:

The number of errors in the 2VO group was significantly higher than in the simulated group (Figures 20-23), indicating that vascular dementia was successfully induced in the rat by both carotid artery ligation, After weekly treatment, it was remarkably improved. Computed tomography (CT) images of the VD rats showed that bilateral occlusion of both carotid arteries (2VO) resulted in swelling, softening, tissue degradation, or even clotting of the brain tissue, leading to dementia. Brain damage is markedly reduced after M1 and M3 treatment.

Extract compositions that treat breast cancer and lower the side effects of drugs that interfere with DNA and / or RNA replication, and / or enhance the drug efficacy of drugs that interfere with DNA and / or RNA replication

material:

Cell lines and samples: MDA-MB-231 (purchased from Food Industry Research and Development Institute, Taiwan); Penicillin-streptomycin-neomycin mixture (100x), fetal bovine serum (FBS) and modified Dulbecco's medium (DMEM) of Dulbecco were purchased from Gibco®; Cyclophosphamide (cyclophosphamide) (CTX, Endoxan®).

Animal facility: BALB / cAnN.Cg-Foxn1 ^nu / CrlNarl female mice approximately 6-8 weeks old were obtained from Taiwan National Laboratory Animal Center. The environmental conditions are as follows: temperature 25 ± 1 ° C; Humidity: 60 ± 5%; Independent air conditioning; Adjusted photocycle. During the whole experiment, water and feed were fed to be free. Mice were raised according to the standard procedure of the Laboratory Animals Committee.

Tumor-bearing mice: MDA-MB-231 cells (human breast cancer cells) were cultured to 5 x 10 ⁶ required for injection into female nude mice, treated with trypsin, . Nude mice were injected subcutaneously with 100 μl of the cell solution to induce tumors and sent back to the cage to provide a normal diet. When the tumors reached about 300 mm ³ , the extract composition was topically administered at various doses and grouped into CTX and non-CTX groups. To see the efficacy of the extract composition, body weight, appetite, blood sugar and tumor size growth were observed weekly to analyze the normal physiological state expression of nude mice.

Tumor-bearing mice and treatment Grouping: When the tumors were about 300 mm ³ , mice were grouped as shown in Table 12. A chemotherapeutic agent (one injection per week) was administered in addition to the extract composition. The treatment group was topically applied to the tumor site and surrounding skin, and to the dorsal skin at a dose of 0.1 g / day. After 5 weeks of treatment, body weight, appetite, blood glucose and tumor size growth of nude mice were observed.

group Tumor induction Chemotherapeutic agent Test article normal Do not No treatment No treatment tumor MDA-MB-231 induction No treatment No treatment J1 MDA-MB-231 induction No treatment GMC1 J2 MDA-MB-231 induction No treatment 0.3% GMA1 NO CTX + L5 MDA-MB-231 induction No treatment GMC1 + 0.5% GMA1 CTX MDA-MB-231 induction CTX / 4 injections No treatment CTX + L1 MDA-MB-231 induction CTX / 4 injections Cream base CTX + L5 MDA-MB-231 induction CTX / 4 injections GMC1 + 0.5% GMA1

Clinical evaluation of tumor-bearing mice: Physiological conditions such as body weight, appetite, blood sugar and tumor size of tumor-bearing mice and tumor size were evaluated. The tumor size was calculated as tumor volume ^{(mm 3) = ab 2/} 2, was evaluated for tumor jeungsikyul.

result:

In the tumor-bearing nude mouse model induced by human breast cancer cell MDA-MB-231, after treatment, extract compositions J1 and J2 showed the effect of inhibiting tumor growth in nude mice (Table 13 and Figure 25). In evaluating physiological conditions, the extract composition group showed better appetite and blood glucose levels.

Tumor size (mm ³ ) 0 weeks 1 week 2 weeks 3 weeks J1 0.0 63.40 126.23 369.51 J2 0.0 -2.02 26.83 182.19 tumor 0.00 72.69 380.71 696.90

In the GMC1 + 0.5% GMA1 (NO CTX + L5) group, the extract composition inhibited tumor growth in nude mice (Figure 26).

The physiological status and tumor proliferation of tumor + chemotherapeutic group (CTX group), extract composition + chemotherapeutic group (CTX + L1 group and CTX + L5 group) were evaluated compared with normal group. Referring to Table 14 and Figures 26 and 27, the results of the CTX + L1 and CTX + L5 groups enhance the efficacy of chemotherapeutic agents that shrink tumors in tumor-bearing mice receiving chemotherapeutic agents, Average feed intake rate and average blood sugar were shown to be insignificant (Table 15 - 17)

Table 14: Tumor size reduction (mm ³ )

4/28 5/2 5/5 5/9 5/12 5/16 5/19 5/23 5/26 5/30 normal 0.00 0.00 0.00 0.00 0.00 0.00 0.00 0.00 0.00 0.00 tumor 0.00 108.05 364.31 428.36 627.45 874.62 1295.63 1961.51 2727.56 2938.30 CTX 0.00 99.61 78.95 -19.52 -45.11 -157.57 -176.41 -264.85 -293.68 -391.42 CTX + L1 0.00 89.22 113.36 21.27 -46.75 -138.97 -179.55 -224.38 -294.03 -384.43 CTX + L5 0.00 149.85 173.28 48.54 -56.91 -170.01 -220.34 -274.15 -338.79 -510.04

Table 15: Average weight (g)

4/28 5/2 5/5 5/9 5/12 5/16 5/19 5/23 5/26 5/30 normal 20.61 21.34 20.91 21.16 20.31 21.13 20.41 21.90 21.63 21.96 tumor 19.64 19.11 20.03 18.75 18.57 20.22 20.48 21.36 21.74 22.67 CTX 19.64 20.04 18.46 19.00 19.51 20.99 20.42 21.16 20.21 22.18 CTX + L1 20.05 20.59 18.72 19.34 18.79 19.23 19.98 20.08 19.94 21.31 CTX + L5 19.12 19.51 18.45 19.09 19.23 19.61 19.93 19.69 19.03 20.52

Table 16: Average feed intake (g)

5/5 ~ 5/9 5/9 ~ 5/12 5 / 12-5 / 16 5/16 - 5/19 5/19 ~ 5/23 5 / 23-5 / 26 5/26 ~ 5/30 normal 4.95 4.75 5.18 4.69 4.52 4.37 4.50 tumor 4.94 4.48 4.56 4.75 4.25 4.65 4.20 CTX 4.89 5.61 5.38 5.21 4.70 4.76 5.22 CTX + L1 4.45 4.58 4.88 4.64 4.08 4.01 4.46 CTX + L5 4.60 5.12 4.47 5.09 4.36 4.21 4.62

Table 17: Mean blood glucose (dL)

5/2 5/9 5/16 5/23 5/30 normal 110 84 99 97 98 tumor 60 66 82 89 74 CTX 113 93 109 122 108 CTX + L1 110 93 92 99 101 CTX + L5 110 109 104 106 102

Clinically, one of the main reasons for abandoning cancer treatment is cancer pain caused by cancer. Cancer pain is known to be caused in part by tumors, mainly due to inflammation, and nerve compression caused by tumors invading the internal organs, peripheral nerves and bones. The extract composition can effectively inhibit the proliferation of the tumor, inhibit cancer pain due to some cancers, and can be helpful for treatment.

While this invention has been described in connection with what is presently considered to be the most practical and preferred embodiment, it is evident that many alternatives, modifications and variations will be apparent to those skilled in the art. All such alternatives, modifications and variations are considered to be within the scope of the present invention.

Claims (20)

An extraction composition comprising a soybean seed extract prepared by a process comprising the steps of:
(a) providing a soybean seed and an extracting solution, wherein the extracting solution is water or an alcoholic solution comprising alcohol at a concentration of less than about 90 wt%;
(b) extracting the soybean seed using the extraction solution at a pressure of less than about 1 atm and a temperature of less than about 60 캜 to obtain a crude extract; And
(c) removing the solid from the crude extract to obtain a liquid fraction. The extraction composition according to claim 1, wherein the method for producing the soybean seed extract further comprises the step (d) of concentrating the liquid fraction obtained in step (c) to obtain a concentrated solid fraction. 3. The extraction composition according to claim 2, wherein the process for producing the soybean seed extract further comprises a step (e) of drying the concentrated solid obtained in step (d). The extraction composition of claim 1, further comprising a soybean seed vapor fraction produced by a process comprising the steps of:
(i) providing the soybean seed to a second extraction solution which is water or an alcohol solution comprising alcohol at a concentration of less than about 15 wt%; And
(ii) extracting said soybean seeds with said second extraction solution at an atmospheric pressure of less than about 1 atm and a temperature of less than about 110 &lt; 0 &gt; C, and collecting the vapor fraction. The extraction composition of claim 4, wherein the alcohol solution in step (i) comprises less than about 5 wt% alcohol. 5. The composition of claim 4, wherein the content of the soybean seed extract is about 0.001 wt% to about 5 wt%; Wherein the content of the soybean seed vapor fraction is from about 95 wt% to about 99.999 wt%. Process for the preparation of an extract composition according to any one of claims 1 to 6, comprising a process for producing a soybean seed extract comprising the steps of:
(a) providing a soybean seed and an extracting solution, wherein the extracting solution is water or an alcoholic solution comprising alcohol at a concentration of less than about 90 wt%;
(b) extracting the soybean seed using the extraction solution at a pressure of less than about 1 atm and a temperature of less than about 60 캜 to obtain a crude extract; And
(c) removing the solid from the crude extract to obtain a liquid fraction. 8. A process according to claim 7, wherein the process for producing the soybean seed extract further comprises the step (d) of concentrating the liquid fraction obtained in step (c) to obtain a concentrated solid fraction. 8. The process according to claim 7, wherein the process for producing the soybean seed extract further comprises a step (e) of drying the concentrated solids obtained in step (d). Use of an extract composition according to any one of claims 1 to 6 in the manufacture of a medicament for promoting wound healing. 11. Use according to claim 10 for the preparation of a medicament for promoting skin cell migration. 12. The use according to claim 11, wherein the skin cells are keratinocytes. 11. The use according to claim 10, wherein the wound is a diabetic injury. Use of an extract composition according to any one of claims 1 to 6 in the manufacture of a medicament for promoting proliferation of neuronal cells and / or for treating brain and / or neurodegenerative diseases. 15. The use according to claim 14, wherein said nerve cell is a neuroblastoma cell. 15. The method of claim 14, wherein said brain and / or neurodegenerative disease is selected from the group consisting of vascular dementia, Alzheimer's disease, Parkinson's disease, cerebrovascular disease, inflammatory brain injury, inflammatory reaction around brain injury, ventricle, and ventricular dilatation. Use of an extract composition according to any one of claims 1 to 6 in the manufacture of a medicament for the treatment of breast cancer. 18. Use according to claim 17, wherein the breast cancer is breast cancer of the P53 mutation type. The use of any one of claims 1 to 6 in the manufacture of a medicament for lowering the side effects of drugs interfering with DNA and / or RNA replication and / or enhancing the medicinal effects of drugs interfering with DNA and / or RNA replication Use of the extract composition according to one aspect. 20. The method of claim 19, wherein the drug interfering with DNA and / or RNA replication is selected from the group consisting of cyclophosphamide, temozolomide, hexamethylmelamine, platinum complex, bleomycin, vinblastine, vincristine, paclitaxel, docetaxel, polynic acid antagonist, purine antagonist or pyrimidine antagonist.

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