JP6890780B2 - Extraction method of fatty acid metabolism promoting component and fatty acid metabolism promoting agent - Google Patents

Description

The present invention relates to a method for extracting a fatty acid metabolism promoting component from Maitake mushroom and a fatty acid metabolism promoting agent containing the fatty acid metabolism promoting component extracted by this method as a main component.

Peroxisome proliferator-activated receptor-δ (hereinafter abbreviated as “PPARδ”), which is a type of nuclear receptor, controls gene expression using various fatty acids as ligands, and in particular, It has been shown to be involved in fat consumption in skeletal muscle. Activation of PPARδ has the effects of suppressing body weight gain due to a high-fat diet, improving insulin resistance, reducing triglyceride in blood, and suppressing fatty liver by increasing energy consumption. Therefore, PPARδ activity. The agent is considered to be useful as a preventive or ameliorating agent for obesity, insulin resistance, arteriosclerosis and fatty liver (Non-Patent Document 1).

Furthermore, since it has been reported that activation of PPARδ in skeletal muscle is involved in the transfer of skeletal muscle to slow muscle type muscle fibers, PPARδ activator is effective in increasing basal metabolism and endurance. It is believed that there is (Non-Patent Document 2). In addition, it has been reported that the muscle fiber type of skeletal muscle changes to a high-fat diet, a polysaccharide diet, and a fast muscle type with aging (Non-Patent Document 3). Therefore, it has been suggested that an increase in slow muscle type muscle fibers in skeletal muscle may be effective in preventing or ameliorating so-called locomotive syndrome caused by lifestyle and aging. From such a viewpoint, the PPARδ activator can be said to be promising as an anti-locomotive syndrome agent.

In addition, it is known that many capillaries are present in slow muscle type muscle fibers that increase due to endurance training (Non-Patent Document 4), and blood circulation is promoted by increasing the amount of capillaries due to the increase in slow muscle type muscle fibers. The effect is also expected.

Furthermore, in recent years, it has been found that the increase in slow muscle type muscle fibers increases factors related to deliciousness such as taurine and iron in meat, the amount of free amino acids in taste, and juiciness, and from the viewpoint of meat production. It is attracting attention (Non-Patent Document 5).

As mentioned above, there are many research reports on skeletal muscle of PPARδ, but since it is known that the expression level of PPARδ is also high in the brain, its function in the brain is also attracting attention. PPARδ in the brain is also expected as a new drug discovery target for neurodegenerative diseases such as Alzheimer's disease and Parkinson's disease due to its functions such as neuroprotective action and anti-inflammatory action (Non-Patent Document 6).

The search and development of PPARδ activators have been actively carried out for such a purpose, and it has been reported that some food ingredients have PPARδ activating ability.

By the way, Maitake mushroom (scientific name: Grifola frontosa), which is a kind of mushroom, is a mushroom that has been eaten for a long time and is one of the ingredients familiar to Japanese eating habits. Various studies on Maitake mushrooms, mainly polysaccharides, have been reported, such as anticancer activity, immunostimulatory activity, antimetabolix syndrome, osteoporosis prevention, and liver disease prevention (Patent Documents 1 to 5). However, the relationship between Maitake mushrooms and PPARδ activation has not been known so far.

Japanese Unexamined Patent Publication No. 5-336920 Japanese Unexamined Patent Publication No. 2007-008829 Japanese Unexamined Patent Publication No. 2011-102286 Japanese Unexamined Patent Publication No. 2001-097881 JP-A-2007-332336

Wang, Y.X., et al., "Peroxisome-proliferator-activated receptor δ activates fat metabolism to prevent obesity." Cell, 2003, 113 (2): 159-170 Wang, Y.X., et al., "Regulation of muscle fiber type and running endurance by PPARδ." PLoS Biol., 2004, 2 (10): e294 Hyatt, JPK, et al., "Muscle-Specific Myosin Heavy Chain Shifts in Response to a Long-Term High Fat / High Sugar Diet and Resveratrol Treatment in Nonhuman Primates." Front. Physiol., 2016, 7 (77): doi : 10.3389 / fphys.2016.00077 Andersen, P. & Henriksson, J., "Capillary supply of the quadriceps femoris muscle of man: adaptive response to exercise." J. Phisiol., 1977, 270 (3): 677-690.1 Kang, Y.K., et al., "Effects of myosin heavy chain isoforms on meat quality, fatty acid composition, and sensory evaluation in Berkshire pigs." Meat Sci., 2011, 89 (4): 384-389 Bordet, R. et al., "PPAR: a new pharmacological target for neuroprotection in stroke and neurodegenerative diseases." Biochem. Soc. Trans., 2006, 34 (6): 1341-1346

The problem to be solved by the present invention is to extract a fraction having a PPARδ-activating action from maitake, which is a fresh food that can stably supply raw materials throughout the year, has abundant eating experience, and ensures safety. , Thereby obtaining a fatty acid metabolism promoter.

The method for extracting a fatty acid metabolism promoting component according to the first aspect of the present application is characterized in that an organic solvent extract is obtained as a fatty acid metabolism promoting component from Maitake using an organic solvent.

The method for extracting the fatty acid metabolism-promoting component according to the second aspect of the present application is, in addition to the features of the first aspect, the organic solvent extract is an ethanol extract extracted with ethanol in water and ethyl acetate. It is characterized in that it is an ethyl acetate extract that has been partitioned and extracted as an ethyl acetate phase.

In the method for extracting the fatty acid metabolism promoting component according to the third aspect of the present application, in addition to the features of the second aspect, the organic solvent extract is prepared by using the ethyl acetate extract as a mixed solvent of hexane and ethyl acetate. Among the eluted fractions eluted by concentration gradient chromatography, the one is extracted as a fraction having a ratio of hexane: ethyl acetate of 1: 1. This concentration gradient chromatography is preferably performed using a silica gel column.

In the method for extracting the fatty acid metabolism promoting component according to the fourth aspect of the present application, in addition to the characteristics of the third aspect, the organic solvent extract has a fraction of the ratio of hexane: ethyl acetate of 1: 1. Further, among the eluted fractions eluted by the concentration gradient chromatography with a mixed solvent of chloroform and methanol, the elution fraction is characterized in that the ratio of chloroform: methanol is 200: 1. This concentration gradient chromatography is preferably performed using a silica gel column.

The fatty acid metabolism-promoting agent according to the present application contains the fatty acid metabolism-promoting component according to any one of the first to fourth aspects as a main component.

Since the fatty acid metabolism promoting component according to the present invention is an organic solvent extract extracted from Maitake mushroom, which is a food material in daily eating habits, as a fraction having a PPARδ activating action, fatty acid metabolism based on this is used. Accelerators are not likely to cause side effects. Moreover, since the fat-soluble extract is a natural substance, no labor such as synthesis is required. Furthermore, since Maitake mushrooms can be stably mass-produced throughout the year, materials can be provided in a stable manner.

The outline of the extraction process of the fatty acid metabolism promoting component from Maitake mushroom is shown. The ethyl acetate extract of Maitake mushroom has a PPARδ activating effect (A) and a PDK4 expression promoting effect (B) on the mouse skeletal muscle-derived cell line C2C12. The effect of the ethyl acetate extract of Maitake mushroom on the glucose metabolism ability is shown. It shows the effect of suppressing the weight gain of the ethyl acetate extract of Maitake mushroom. The effect of the ethyl acetate extract of Maitake mushroom on the change in tissue weight is shown. The effect of ethyl acetate extract of Maitake mushroom on liver lipids is shown. The effect of the ethyl acetate extract of Maitake mushroom on the plasma component is shown. The effect of ethyl acetate extract of Maitake mushroom on gene expression in mouse skeletal muscle is shown. Hexane from the ethyl acetate extract of Maitake mushroom: Shows the PPARδ activating action of the ethyl acetate extract fraction. Hexane: Ethyl acetate extract fraction Fr. Chloroform from 6: Shows PPARδ activating action of methanol extract fraction.

The fatty acid metabolism-promoting agent according to the present invention is composed mainly of a fatty acid metabolism-promoting component which is an organic solvent extract extracted from Maitake mushrooms with an organic solvent.

Maitake mushrooms are edible mushrooms of basidiomycetes. According to the statistics of the Ministry of Agriculture, Forestry and Fisheries in 2015, the annual production of Maitake mushrooms is about 48,800 tons, and they are cultivated throughout the year. Regarding the acquisition of Maitake mushroom, which is the material of the present invention, mycelia can be collected by tank culture or the like, but it is easier to obtain fruiting bodies for commercial use.

Further, in order to obtain the effect of the present invention, a culture of mycelium can be used, but in order to obtain the effect of the present invention sufficiently, an organic solvent extract is extracted from the fruiting body of Maitake mushroom. Is preferable.

Next, a method for extracting the organic solvent extract from Maitake mushroom will be described.

As the fruiting body or mycelium of Maitake mushroom as a material, a dry powdered one can also be used. Considering the extraction yield, it is desirable that Maitake mushrooms are dried and then pulverized. However, it is more preferable to use fresh Maitake mushrooms because the cost can be reduced by omitting the drying step and the pulverization step.

Further, for drying the fruiting bodies or mycelium of Maitake mushroom, a method such as sun drying, warm air drying, hot air drying or freeze drying can be applied.

Next, the primary extraction will be described. Fresh Maitake mushrooms or dried Maitake mushroom powder is immersed in an organic solvent, and then the organic solvent is suction-filtered to collect the extract. The organic solvent can be removed from the recovered extract under reduced pressure, and the residue can be obtained as a primary extract.

Here, as the organic solvent, a polar organic solvent or a hydrophilic organic solvent can be used. Specifically, methanol, ethanol, 1-propanol, 2-propanol, 1-butanol, acetonitrile, acetone and the like can be used alone or as a mixed solvent of two or more of them. Of these, it is more preferable to use ethanol. When these polar organic solvents or hydrophilic organic solvents are used, a mixture with water may be used as the solvent. For extraction from water-poor powder, a non-polar or low-polar organic solvent such as ethyl acetate, hexane, toluene, dimethyl ether, diethyl ether, chloroform or carbon tetrachloride can also be used.

In addition, the immersion time, the amount of the organic solvent, and the like can be appropriately changed in consideration of the extraction efficiency.

Next, the secondary extraction will be described. The primary extract itself has a PPARδ activating action, but the action can be further enhanced by further fractionation.

Distilled water and one or more organic solvents selected from low-polarity organic solvents are added to the primary extract, mixed, allowed to stand, and when the mixed solution is separated, the distilled aqueous phase is removed. Only the organic solvent phase remains. Next, the organic solvent phase is dehydrated with anhydrous sodium sulfate and then concentrated under reduced pressure to obtain a secondary extract.

Here, examples of the low-polarity organic solvent include methylene chloride, ethyl acetate, chloroform, diethyl ether, toluene, benzene, hexane and the like, preferably ethyl acetate, chloroform or hexane alone or in combination of two or more. It will be used as a mixed solvent. The most preferable of these is ethyl acetate.

Next, the tertiary extraction will be described. The secondary extract exhibits a stronger PPARδ activating effect than the primary extract, but it is desirable to further fractionate in order to further improve the purity of the active ingredient. That is, by adsorbing the secondary extract on silica gel and then eluting the silica gel adsorbate with an organic solvent, a fraction having a stronger PPARδ activating action can be obtained as a tertiary extract.

That is, the secondary extract was adsorbed on silica gel packed in an open column, and a fractionation operation was performed according to a gradient elution method using a mixed organic solvent of hexane and ethyl acetate, and hexane: ethyl acetate was 1: 1. The fraction eluted at the ratio of is obtained as a tertiary extract. The tertiary extract has a stronger PPARδ activating effect than the secondary extract.

As the silica gel filled in the open column, various silica gels for filling in normal phase column chromatography can be used, and the particle size of the silica gel is preferably 40 to 200 μm.

Here, as the mixed organic solvent for eluting the silica gel adsorbent, an organic solvent obtained by mixing two or more organic solvents selected from all of the non-polar organic solvent and the polar organic solvent is used. However, as described above, it is preferable to use an organic solvent in which hexane and ethyl acetate are mixed.

The tertiary extract may be further subjected to a quaternary extraction in order to further improve the purity of the active ingredient. That is, the tertiary extract is adsorbed on silica gel packed in an open column similar to the tertiary extraction, and a fractionation operation is performed according to a gradient elution method using a mixed organic solvent of chloroform and methanol to obtain chloroform: methanol. Fractions eluted at a ratio of 200: 1 are obtained as the quaternary extract. The quaternary extract has a stronger PPARδ activating action than the tertiary extract.

(Extraction of fatty acid metabolism promoting component)
The outline of the extraction step of the fatty acid metabolism promoting component is shown in FIG. 1, and an example of extracting the component from a fresh Maitake fruiting body is described in detail below.

41.2 kg of fresh Maitake fruiting bodies were immersed in 40 L of ethanol and left at room temperature for 1 week. Then, the ethanol solution in which the Maitake fruiting body was immersed was suction-filtered and collected to obtain an extract. Ethanol was then evaporated from the extract using an evaporator to give the residue as the primary extract.

Next, the primary extract was added to a solvent in which ethyl acetate and water were mixed at a ratio of 1: 1 and then stirred and left to stand. After a while, when the phase was separated into the ethyl acetate phase and the aqueous phase, the ethyl acetate phase was recovered. Anhydrous sodium sulfate was added to the recovered ethyl acetate phase to carry out a dehydration operation. Then, the solvent was evaporated from the ethyl acetate phase using an evaporator to obtain 8.10 g of a residue as a secondary extract (ethyl acetate extract).

(Evaluation of PPARδ activation ability of ethyl acetate extract)
The PPARδ activation ability of the ethyl acetate extract was evaluated by a luciferase assay. Human fetal kidney-derived cell line HEK293 was seeded in a 12-well plate and contained in 10% by volume of fetal bovine serum (FBS, ICN Biomedicals) and 100 units / mL penicillin and 100 μg / mL Streptomycin (Invitrogen) in Dalbeco modified Eagle's medium (DMEM, DMEM). It was cultured in SIGMA) for 24 hours. Then, a reporter gene in which the Gal4 response sequence was incorporated 5 times upstream of the luciferase gene and a fusion protein expression gene in which the ligand binding region of PPARδ was incorporated behind the Gal4 DNA binding region were introduced using the calcium phosphate method, and the luciferase assay was performed. Was performed. Six hours after gene transfer, the medium was replaced with serum-free DMEM medium containing the ethyl acetate extract (100 μg / mL), and the cells were further cultured for 18 hours.

A dual luciferase assay system (Promega) was used to measure luciferase activity. That is, the cells after culturing were lysed, a substrate solution containing luciferin was added to the lysate, and the luminescence of firefly luciferase was measured with a luminometer (MiniLumat, Berthold). In this experimental system, the value of firefly luciferase activity was divided by the value of β-galactosidase activity to obtain the value of luciferase activity. The luciferase activity was shown as a relative value with the transcriptional activity of PPARδ by dimethyl sulfoxide (DMSO), which is a solvent control, as 1. In addition, GW501516 was used as an activator of PPARδ, and GSK3787 was used as an inhibitor.

(Ethyl acetate extract enhances PPARδ target gene expression)
The effect of the ethyl acetate extract on the expression of the PPARδ target gene was analyzed. The mouse skeletal muscle-derived cell line C2C12 was seeded in a 12-well plate and cultured for 1 day in DMEM containing 10% by volume charcoal-treated FBS and 100 units / mL penicillin and 100 μg / mL streptomycin. Twenty-four hours after the start of culturing, the medium was replaced with DMEM containing 2% by volume horse serum (Equitech-Bio Inc.) to induce differentiation. DMEM containing 2% by volume horse serum was replaced with a new one every 2 days, and after induction of differentiation for 4 to 5 days, it was replaced with a medium containing the ethyl acetate extract (100 μg / mL). Total RNA was extracted from the cells 24 hours after the addition using an RNA extraction reagent (ISOGEN, Nippon Gene). A reverse transcription reaction was carried out using 125 ng of the extracted total RNA. Using the synthesized cDNA, the mRNA expression level of PDK4, which is a PPARδ target gene, was quantified by a real-time PCR method. In the real-time PCR method, after 10 minutes of initial heat denaturation at 95 ° C., 15 seconds of heat denaturation at 95 ° C. and 1 minute of annealing at 60 ° C. were set as one cycle, and this was performed for 40 cycles. As the primer of the PDK4 gene, the following sequence 1 was used as a sense primer, and the following sequence 2 was used as an antisense primer.

Sequence 1: 5'-aaaggacaggatggaaggaatca-3'
Array 2: 5'-ttttcctctgggtttgcacat-3'

As a result of the luciferase assay, as shown in FIG. 2 (A), the activity of C2C12 cells was increased by the addition of the PPARδ agonist GW501516. It was confirmed that the activation of PPARδ induced by GW501516 was decreased in a concentration-dependent manner by the PPARδ inhibitor GSK3787 (“GSK” in the figure). The numerical value in parentheses in the figure indicates the concentration (μM). As for the ethyl acetate extract (“EA” in the figure), GSK3787 showed a concentration-dependent decrease in activation, suggesting that the ethyl acetate extract has a PPARδ activating effect. Regarding the expression of PDK4, which is the target gene of PPAR, as shown in FIG. 2 (B), there is a tendency that GSK3787 reduces mRNA expression in a concentration-dependent manner, as in the graph of FIG. 2 (A). Therefore, it was inferred that the ethyl acetate extract activates PPARδ because it promotes the expression of PDK4.

(Animal experimentation)
Six-week-old male C57 / BL6J mice (Claire Japan) were used in each experiment after acclimation for one week. Mice were bred under a light-dark cycle (lights off at 21:00) every 12 hours, and feed and water (sterilized distilled water) were freely fed. As the high-fat diet used in the high-fat diet load test, a high-fat diet in which 60% of the total calories were calories by fat was used, and the diet was similarly freely fed. The test group was similarly free-fed with this high-fat diet containing 0.2% by weight or 0.4% by weight of the ethyl acetate extract. Dissection of each tissue was performed after 6 hours of fasting, and the tissue was frozen in liquid nitrogen immediately after collection and stored at -80 ° C until used in the experiment. All animal experiments were conducted based on the University of Tokyo Animal Experiment Implementation Regulations.

(Glucose tolerance test)
Six mice were divided into a test group and a control group, and the control group was fed with a high-fat diet (HF), and the test group was fed with a high-fat diet (HF + GF) containing 0.2% by weight of the ethyl acetate extract. Gave. When the amount of food consumed and the body weight were measured every 2 to 3 days after the start of feeding, no significant difference was observed between the two groups by the 12th week after the start of feeding. However, 12 weeks after the start of feeding, the glucose solution (1.5 g / kg) was orally administered to both groups of mice, and the blood glucose concentrations were measured after 0, 15, 30, 60, 90 and 120 minutes. As shown in FIG. 3, it was found that the increase in blood glucose concentration was significantly suppressed 15 to 30 minutes after the oral administration. Therefore, it was confirmed that the ethyl acetate extract of Maitake mushroom increased the glucose metabolism ability of mice.

(Obesity improving effect)
Six mice were divided into a test group and a control group, and the control group was fed with a high-fat diet (HF), and the test group was fed with a high-fat diet (HF + GF) containing 0.4% by weight of the ethyl acetate extract. It was. When the food intake and body weight were measured every 2 to 3 days for about 3 months, the total food intake during the period was not different between the two groups as shown in FIG. 4 (A), but FIG. 4 As shown in (B), significant suppression of body weight gain was observed in the test group.

(Action on lipids)
Six mice were divided into a test group and a control group, and the control group was fed with a high-fat diet (HF), and the test group was fed with a high-fat diet (GF) containing 0.4% by weight of the ethyl acetate extract. It was. Three months after the start of feeding, as shown in FIG. 5, a significant decrease in visceral fat and subcutaneous fat was observed in the test group.

In addition, the triglyceride and cholesterol levels in the liver were measured 3 months after the start of feeding. As a result, as shown in FIG. 6 (A), a significant decrease in triglyceride in the liver was observed in the test group (GF) as compared with the control group (HF). The cholesterol level in the liver also seemed to decrease in the test group as shown in FIG. 6 (B), but this was not a significant difference. Regarding glucose in plasma (FIG. 7 (A)), triglyceride (FIG. 7 (B)) and free fatty acid (FIG. 7 (D)), between the test group (HF + GF) and the control group (HF). However, there was no significant difference in cholesterol level (Fig. 7 (C)) in the test group.

(Gene expression)
Furthermore, among the skeletal muscles of mice 3 months after the start of high-fat diet containing 0.2% by weight of ethyl acetate extract, white muscle (“white”, gastrocnemius muscle) and red muscle (“red”, soleus muscle) The gene expression of PDK4 (FIG. 8 (A)), LPL (FIG. 8 (B)) and GLUT4 (FIG. 8 (C)) was examined. Specifically, the tissue obtained from mouse skeletal muscle is disrupted in an RNA extraction reagent (ISOGEN, Nippon Gene), total RNA is extracted by the same method as for C2C12 cells, and then the PDK4 gene is expressed. In the same manner as above, gene expression was observed by the real-time PCR method. As the primer of the LPL gene, the following sequence 3 was used as a sense primer, and the following sequence 4 was used as an antisense primer. As the primer of the GLUT4 gene, the following sequence 5 was used as a sense primer and the following sequence 6 was used as an antisense primer.

Sequence 3: 5'-cttcttgatttacacggaggt-3'
Sequence 4: 5'-atggcatttcacaaacactg-3'
Sequence 5: 5'-gagctgaaggatgagaaacgga-3'
Sequence 6: 5'-cattgatgcctgagagctgttg-3'

As a result, it was found that the expression of all genes in the red muscle was significantly increased in the test group (GF) as compared with the control group (HF). Here, PDK4 and LPL are PPARδ target genes, and it is presumed that lipid metabolism is activated by ingestion of ethyl acetate extract of Maitake mushrooms to cause expression fluctuations of these genes. In addition, the ethyl acetate extract of Maitake mushroom ingested from the diet can reach the skeletal muscle, which is presumed to increase the expression level of fatty acid β-oxidation-related genes (PDK4, LPL). GLUT4 is a gene related to glucose metabolism, and it is presumed that increased expression of these genes also enhances glucose metabolism.

(Summary on fat metabolism promoting action)
From the above experimental results, it was found that the ethyl acetate extract of Maitake mushroom has a PPARδ activating effect. It was also clarified that it has an effect of suppressing weight gain and an effect of reducing triglyceride in the liver when a high-fat diet is continued for a long period of time. Furthermore, it was clarified that it has an effect of improving the decrease in glucose metabolism ability due to obesity. Furthermore, it was suggested that the orally ingested Maitake component may exert an action of promoting fatty acid metabolism and glucose metabolism on skeletal muscle, either directly or by metabolites.

This indicates that the ethyl acetate extract of Maitake mushroom can function as a fatty acid metabolism promoter alone or as a food or drug containing the extract.

(Fraction of active ingredient)
For the ethyl acetate extract of Maitake mushroom having the above-mentioned action, an attempt was made to fractionate the components leading to the purification of the active ingredient.

That is, 8.10 g of ethyl acetate extract dissolved in a small amount of ethyl acetate was adsorbed on Celite and then completely dried, and then this was subjected to concentration gradient chromatography using a mixed solvent of hexane and ethyl acetate using a silica gel column. Fractionated. That is, when this Celite adsorption extract was placed on silica gel packed in an open column and elution was performed with a concentration gradient using a mixed solvent in which the concentration ratio of ethyl acetate and hexane was adjusted, Fr. 1 to Fr. Up to 10 fractions were obtained. Of these, as a fraction having a high PPARδ activating effect, Fr. With a ratio of hexane: ethyl acetate of 1: 1. 6 was selected (see FIG. 9). In the figure, "DMSO" is the DMSO as a negative control, "GW" is the GW501516 as a positive control, and "EA" is the ethyl acetate extract. This fraction Fr. The dry weight after evaporating the solvent from 6 was 80 mg. In the figure, the fraction Fr. Although the activating effect of 7 is somewhat higher, Fr. Since the activity of No. 6 was higher, the fraction Fr. 6 was selected.

This fraction Fr. It was suggested that 6 can function as a fatty acid metabolism promoter more potent than the ethyl acetate extract, alone or as a food or drug containing it.

Furthermore, this fraction Fr. 6 was dissolved in a small amount of ethyl acetate, adsorbed on Celite, and then completely dried, and then fractionated by concentration gradient chromatography using a mixed solvent of chloroform and methanol using a silica gel column. That is, when this Celite adsorption fraction was placed on silica gel packed in an open column and concentration gradient elution was performed using a mixed solvent in which the concentration ratio of chloroform and methanol was adjusted, Fr. From 51-1 to Fr. Fractions up to 51-8 were obtained. Of these, as a fraction having a high PPARδ activating effect, Fr. With a chloroform: methanol ratio of 200: 1. 51-3 was selected (see FIG. 10). In the figure, "control" is the DMSO as a negative control, "GW" is the GW501516 as a positive control, and "EA" is the ethyl acetate extract. This fraction Fr. The dry weight after evaporating the solvent from 51-3 was 30.4 mg.

From the above, the fraction Fr. It is expected that 51-3 contains a component that plays a critical role by the above-mentioned fat metabolism promoting action. Therefore, this Fr. 51-3 can be used alone or as a food or drug containing the fraction Fr. It was suggested that it could function as a more potent fatty acid metabolism promoter than 6.

The fat metabolism promoting component according to the present invention has an excellent PPARδ activating effect and is derived from food, so that it is highly safe even if it is ingested for a long period of time. In addition, since it can reduce body fat and exert endurance improving effect and anti-locomotive syndrome effect as a slow muscle increasing agent, pharmaceuticals, quasi-drugs, cosmetics and foods, and materials or compounding agents for blending them. It is useful as.

Claims (3)

An ethanol extract extracted from Maitake with ethanol is partitioned between water and ethyl acetate to obtain an organic solvent extract, which is an ethyl acetate extract extracted as an ethyl acetate phase, as a fatty acid metabolism promoting component. Extraction method of fatty acid metabolism promoting component. The organic solvent extract is a fraction in which the ratio of hexane: ethyl acetate is 1: 1 to the elution fraction obtained by eluting the ethyl acetate extract by concentration gradient chromatography with a mixed solvent of hexane and ethyl acetate. The method for extracting a fatty acid metabolism-promoting component according to claim 1, wherein the solvent is extracted as a solvent. In the organic solvent extract, the fraction of hexane: ethyl acetate of 1: 1 was further eluted by concentration gradient chromatography with a mixed solvent of chloroform and methanol, of which chloroform: methanol. The method for extracting a fatty acid metabolism promoting component according to claim 2 , wherein the solvent is eluted as a fraction of 200: 1.

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