KR100551718B1 - Anti-obesity Composition for Inhibiting Activity of Fatty-Acid Synthase - Google Patents

Abstract

The present invention relates to a composition for inhibiting obesity capable of inhibiting the activity of fatty acid synthase (hereinafter referred to as "FAS"), and more specifically, to a mixture of an extract obtained from green tea and an extract from evening primrose seed. The present invention relates to a composition for inhibiting obesity, which can suppress the occurrence of obesity by inhibiting the activity of FAS involved in the synthesis of fatty acids using the composition containing as an active ingredient.

The composition of the green tea extract and evening primrose seed extract of the present invention has a very high effect in inhibiting the activity of FAS, green tea and evening primrose seed extract used in the present invention is a natural medicine harmless to the human body, without any side effects It can be usefully used for the treatment and prevention of various diseases associated with obesity.

Description

Anti-obesity Composition for Inhibiting Activity of Fatty-Acid Synthase

1 and 2 are graphs showing the effect of inhibiting FAS activity.

The present invention relates to a composition for inhibiting obesity capable of inhibiting the activity of fatty acid synthase (hereinafter referred to as "FAS"), and more specifically, to a mixture of an extract obtained from green tea and an extract from evening primrose seed. The present invention relates to a composition for inhibiting obesity, which can suppress the occurrence of obesity by inhibiting the activity of FAS involved in the synthesis of fatty acids using the composition containing as an active ingredient.

In modern society, due to economic development and the development of the food industry, the amount of exercise has been reduced, and the diet has changed from being vegetarian to eating more fat and calories than necessary. As a result, the amount of calories absorbed was greater than the amount used, the excess energy in the body was generated, and the accumulated body fat caused metabolic abnormalities, resulting in obesity.

Currently obesity is hypertension, arteriosclerosis, diabetes. Efforts to develop obesity treatments continue throughout the society, as they are directly linked to a number of serious diseases such as fatty liver, gout and cancer, and the mortality rate of obese people is reported to be 1.3 times higher than normal weight. It is becoming.

Early obesity treatments include exercise therapy to increase the use of calories, diet to reduce the absorption of calories, and behavioral therapy. Recently, drugs such as fat digestive enzyme inhibitors, appetite suppressants and lipolytic agents have been generally used. It is known to use therapy.

However, drugs such as fat digestive enzyme inhibitors, for example, Zenical, can degrade starch into disaccharides or trisaccharides by digestive enzymes during metabolism, and then inhibit the breakdown of glucose into monosaccharides in the small intestine. It lowers the absorption of sugar into the body and treats obesity. Thus, Koreans who consume a large amount of sugar, that is, starch, from calories such as sugars, lipids, and proteins have a disadvantage in that obesity is ineffective. In addition, a large amount of disaccharide suppressed during the enzyme metabolism remains in the body, causing side effects such as diarrhea and increased gas production.

In addition, as science advances, studies show that obesity is not only caused by environmental factors caused by diet, but also by genetic predisposition, such as the occurrence of genetic mutations. There is a great deal of interest in developing new obesity treatments to prevent mutations.

Among the genes in which the mutation occurs is FAS, which consists of a multifunctional polypeptide, which is a long-chain fatty acid from acetyl-CoA and malonyl-CoA. It is an important enzyme involved in energy metabolism during production, but obesity occurs as the enzyme is mutated.

Currently, a substance that inhibits the variation of the FAS is known as (+)-cerulenin of the formula (1), which includes an epoxy group therein and is limited in use as a medicine, and is a natural substance. Because of this, separation and synthesis are difficult.

[Formula 1]

SUMMARY OF THE INVENTION An object of the present invention is to provide an agent for inhibiting obesity, which effectively inhibits FAS gene mutations that cause obesity, and which can inhibit the activity of FAS, which is economical and safe in humans, without toxicity or side effects.

In order to achieve the above object, the present invention provides a green tea extract and evening primrose seed extract, which is a herbal preparation that has the effect of inhibiting the activity of FAS that is genetically modified to cause obesity, and contains a mixture of the extracts as an active ingredient. It provides a pharmaceutical composition for inhibiting obesity.

In this case, the evening primrose or the evening primrose root instead of the seed extract may be used.

In addition, for this purpose it provides a method for producing green tea extract and evening primrose seed extract.

Green tea was a common favorite food, but it contains vitamins, amino acids such as deanine, caffeine, polyphenols, and catechins of tannins, and these ingredients are anti-cancer effect, prevention of arthritis, prevention of blood pressure increase, prevention of tooth decay, anti-aging and In addition to the heavy metal removal effect, consumption is increasing rapidly according to the research results that it has a pharmacological effect of suppressing digestive enzyme action to reduce calorie absorption, and also to help burn body fat and inhibit FAS activity.

In addition, evening primrose (Oenothera odorata) is a biennial plant with a bicotyledonous cotyledon needle family originated in Chile, South America, distributed in watersides, roadsides and vacant lands all over the country. It is being used for the prescription of folk remedies.

Hereinafter, the present invention will be described in detail.

The green tea extract and the evening primrose flower extract can generally be obtained by solution extraction, that is, by heating and concentrating the green tea or evening primrose flower in water or an organic solvent. For example, the evening primrose seed extract can be obtained by adding an organic solvent of water or alcohol to the evening primrose seeds at 20 to 30 ° C. and refluxing them, followed by concentration.

At this time, the reflux time is not limited but is preferably 1 to 3 hours.

In addition, in the case of green tea can be obtained by adding an organic solvent of water or alcohol of 70 to 100 ℃, preferably 80 ℃ to the green tea leaves, and then obtained by concentrating.

At this time, the extraction time is not limited, but is preferably 5 to 20 minutes.

In the case of the composition for inhibiting obesity, which contains a mixture of green tea extract and evening primrose seed extract as an active ingredient, each extract is a natural ingredient harmless to the human body, and since the effect can be seen in a small amount, the content thereof is not particularly limited, but a generally known recommended intake amount It is possible to contain 1/3 to 3 times of the amount of evening primrose flower extract per day of human body weight intake 0.3 ~ 0.4g / kg / day, preferably 0.2 ~ 0.3g / kg / day In the case of green tea extract, the daily intake of 1kg of human body weight should be 0.09 ~ 0.13g / kg / day.

For example, when an adult with a weight of 50 to 70 kg is usually taken for 1 day to prepare a composition, the amount of evening primrose seeds extracted from the dry weight of 15 g to 18 g and the green tea leaves from 6 g to 9 g can be taken. In this case, when the experiment to ingest divided into three times to prepare a composition to include a mixed extract obtained from evening primrose flower seeds 5-6g and green tea leaves 2-3g, to obtain the best FAS inhibitory effect dried evening primrose : It is preferable to extract each raw material using the blending ratio of green tea leaf at 1.6-3: 1, and to concentrate and mix it in the same volume, and to consume it.

Experiments were provided for identifying the effects of inhibiting FAS activity using the compositions of the present invention.

First, FAS is isolated and purified from SK-Br3 breast cancer cell line, and then 0.315 mg corresponding to EC50 (Effective Concentration), which is an optimal concentration suitable for experiments on the inhibitory effect of FAS, is taken. The long chain fatty acid is synthesized by the condensation reaction of the obtained FAS with acetyl CoA and malonyl CoA to oxidize NADPH, which acts as a reducing agent. .

According to these results, it can be seen that the absorbance of NADPH is reduced at a wavelength of 340 nm, and the absorbance value was measured to determine the degree of activity inhibition of FAS by the composition of the present invention according to Equation 1 below (see FIG. 1).

[Equation 1]

Overall FAS activity (nmol NADPH / min / mg protein)

= Changed absorbance x 80.5 nmol / 20 min / mg total protein

At this time, the degree of inhibition of FAS activity was measured by the same method as described above using only green tea and evening primrose seed extract.

As a result, it can be seen that the inhibitory effect of the FAS activity of the composition of the present invention is superior to the inhibitory effect of the FAS for each of the green tea extract and the evening primrose seed extract of the comparative group. In addition, as a result of examining the sustained effect of the activity inhibitory effect of FAS after long-term storage of the composition, the effect of inhibiting the activity of FAS of the composition of the present invention than the inhibitory effect of FAS on the green tea extract and evening primrose seed extract of the comparative group It was confirmed using the following experimental example that it lasts for a long time.

At this time, when the effective amount of the extract constituting the composition is either small or increases, the FAS inhibitory effect is relatively lowered.

In addition, since the composition of the present invention uses a natural medicine that is harmless to the human body, it is a safe substance having no toxicity even when administered orally. Therefore, the extract composition may also be used in general pharmaceutical forms such as solid preparations and liquid preparations that can be administered orally.

The solid preparation for oral administration may include a powder form, and the solvent is distilled under reduced pressure to prepare an appropriate amount of lactose or dextrin in a dry residue, and liquid preparations are commonly used as emulsions and syrups. In addition to water, which is a simple diluent, various excipients may be included, for example wetting agents, sweeteners, fragrances, appetite suppressants and preservatives.

Moreover, the said composition can also be used in the form added to a drink, foodstuffs, etc.

The health food is a food which improves the function of the general food by adding the composition to the general food, and the composition can be added to the general food, or can be prepared by encapsulation, powdering and suspension.

In addition, when ingesting it brings a specific effect on health, and unlike the general medicine because the food is a raw material has the advantage that there is no side effect that can occur when taking the long term of the drug.

In addition, when the composition of the present invention is used as a food additive, the composition may be added as it is or used with other food or food ingredients, and may be appropriately used according to a conventional method. The mixed amount of the active ingredient may be appropriately determined depending on the purpose of use thereof. At this time, there is no particular limitation on the type of food.

Hereinafter, the present invention will be described in detail by examples. However, the examples are only to illustrate the invention and the present invention is not limited by the following examples.

I. Preparation of Extraction Composition for Inhibiting Obesity of the Present Invention

Example 1 Preparation of Green Tea Extract

After distilled water boiled up to 100 ° C. and cooled to 80 ° C., 100 mL of 80 ° C. distilled water was added to 2 g of green tea leaves and extracted for 10 minutes. After filtering the insoluble solid from the solution, the filtrate was distilled under reduced pressure, concentrated to 30mL of the solution, and decolorized by charcoal compounds that inhibit UV detection to prepare a green tea extract of the present invention.

Example 2. Preparation of Green Tea Extract

Tea extract of the present invention was prepared according to the method of Example 1, except that 3 g of green tea leaves were used.

Example 3. Preparation of Green Tea Extract

Tea extract of the present invention was prepared according to the method of Example 1, except that 5 g of green tea leaves were used.

Example 4 Preparation of Evening Primrose Seed Extract

5 g of evening primrose seeds were added to 100 mL of distilled water at 25 ° C and refluxed for 2 hours. After filtering the insoluble solid from the solution, the filtrate was distilled under reduced pressure, concentrated to 30mL of the solution, and decolorization treatment with compounds that inhibit UV detection to prepare Evening Primrose seed extract of the present invention.

Example 5 Preparation of Evening Primrose Seed Extract

The evening primrose seed extract of the present invention was prepared according to the method of Example 4, except that 10 g of evening primrose seed was used.

Example 6 Preparation of Evening Primrose Seed Extract

The evening primrose seed extract of the present invention was prepared according to the method of Example 4, except that 15 g of evening primrose seeds were used.

Example 7. Preparation of extract composition for inhibiting obesity of the present invention

The extracts obtained in Examples 1 and 4 were mixed to obtain a composition of the present invention (2g green tea leaf: evening primrose seed 5g = 1: 2.5).

Example 8. Preparation of Extract Composition for Obesity Inhibition of the Present Invention

The extracts obtained in Examples 1 and 5 were mixed to obtain a composition of the present invention (green tea leaves 2g: evening primrose seeds 10g = 1: 5).

Example 9 Preparation of Extract Composition for Obesity Inhibition of the Present Invention

The extracts obtained in Examples 1 and 6 were mixed to obtain a composition of the present invention (green tea leaves 2g: evening primrose seeds 15g = 1: 7.5).

Example 10 Preparation of Extract Composition for Obesity Inhibition of the Present Invention

The extracts obtained in Examples 2 and 4 were mixed to obtain a composition of the present invention (green tea leaves 3g: evening primrose seeds 5g = 1: 1.67).

Example 11. Preparation of Extract Composition for Obesity Inhibition of the Present Invention

The extracts obtained in Examples 2 and 5 were mixed to obtain a composition of the present invention (green tea leaves 3g: evening primrose seeds 10g = 1: 3.3).

Example 12 Preparation of Extract Composition for Obesity Inhibition of the Present Invention

The extracts obtained in Examples 2 and 6 were mixed to obtain a composition of the present invention (green tea leaves 3g: evening primrose seeds 15g = 1: 5).

Example 13. Preparation of the extract composition for inhibiting obesity of the present invention

The extracts obtained in Examples 3 and 4 were mixed to obtain a composition of the present invention (5g green tea leaf: evening primrose seed 5g = 1: 1).

 II. FAS inhibitory effect experiment

Experimental Example 1. Isolation of FAS

SK-BR3 cells were prepared using RPMI1640 (Inbitrogen Co., USA) medium containing 10% fetal bovine serum (FBS) and penicillin / streptomycin / fungizon. 37 ° C, supplied with 5% CO ₂ The cells were cultured in an incubator.

The cultured cells were removed from the culture vessel using a scraper, collected, washed four times with cold phosphate buffered saline (PBS) at pH 7.2, and then centrifuged (3000 rpm, 15 min). Cell precipitates were obtained. The cell precipitate was suspended in three volumes of homogenization buffer and then sonicated three times for 15 seconds to dissolve. At this time, the buffer solution was used was pH 7.0 containing 0.25M sucrose (sucrose), 10mM DTT, 1mM EDTA and 0.1mg / mL trypsin inhibitor (trypsin inhibitor).

Centrifugation (100,000 g, 4 ° C.) was performed on the cell lysate for 1 hour, and then the supernatant was separated and concentrated with a centricon 10 ultrafiltration device (Amicon) to obtain FAS. At this time, the concentrate was used for the enzyme activity measurement after protein quantification using the BCA method.

Experimental Example 2. Activity of FAS

0.315mg of FAS, 100μM NADPH and 25μM acetyl CoA isolated in Experimental Example 1 were added to Phosphate buffer at pH 7.4, and then 100μM malonyl CoA was added and the reaction was started, spectrophotometer (specrtophotometer; Beckman) After continuously measuring the absorbance at an interval of 30 seconds for 30 minutes at a wavelength of 340nm, the FAS activity obtained according to Equation 1 is shown in Table 1 below (see FIG. 1).

Experimental Example 3. FAS inhibitory effect experiment using the composition of the present invention

The composition of Example 7 (green tea leaves 2g: evening primrose seeds 5g = 1: 2.5) was further included in the reaction of Experimental Example 2, and then absorbance was measured continuously from the start of the reaction. Using the obtained absorbance, the effect of inhibiting FAS activity by the composition of the present invention was measured and shown in Table 1 below (see FIG. 1).

Experimental Example 4. FAS inhibitory effect experiment of the composition of the present invention

Instead of the composition of Example 7, the composition of Example 8 (2g green tea leaves: evening primrose seeds 10g = 1: 5) to the reaction of Experimental Example 2, and then absorbance was measured continuously from the start of the reaction . Using the obtained absorbance, the effect of inhibiting FAS activity by the composition of the present invention was measured and shown in Table 1 below (see FIG. 1).

Experimental Example 5. FAS inhibitory effect of the composition of the present invention

Instead of the composition of Example 7 in the reaction of Experimental Example 2, the composition prepared from Example 9 (2g: Evening Primrose Seed 15g = 1: 7.5) was further included, and then the absorbance was measured continuously from the start of the reaction. . Using the obtained absorbance, the effect of inhibiting FAS activity by the composition of the present invention was measured and shown in Table 1 below (see FIG. 1).

Experimental Example 6. FAS inhibitory effect of the composition of the present invention

Instead of the composition of Example 7 in the reaction of Experimental Example 2 further comprises a composition prepared from Example 10 (3g green tea leaves: evening primrose seeds 5g = 1: 1.67), and then absorbance was measured continuously from the start of the reaction . Using the obtained absorbance, the effect of inhibiting FAS activity by the composition of the present invention was measured and shown in Table 1 below (see FIG. 1).

Experimental Example 7. FAS inhibitory effect of the composition of the present invention

Instead of the composition of Example 7 in the reaction of Experimental Example 2 further comprises a composition prepared from Example 11 (3g green tea leaf: evening primrose seed 10g = 1: 3.3), then absorbance was measured continuously from the start of the reaction . Using the obtained absorbance, the effect of inhibiting FAS activity by the composition of the present invention was measured and shown in Table 1 below (see FIG. 1).

Experimental Example 8. FAS inhibitory effect experiment of the composition of the present invention

Instead of the composition prepared from Example 7 in the reaction of Experimental Example 2 further comprises a composition prepared from Example 12 (green tea leaves 3g: Evening Primrose seeds 15g = 1: 5), and then the absorbance is continuous from the start of the reaction Measured. Using the obtained absorbance, the effect of inhibiting FAS activity by the composition of the present invention was measured and shown in Table 1 below (see FIG. 1).

Experimental Example 9. FAS inhibitory effect experiment of the composition of the present invention

Instead of the composition of Example 7 in the reaction of Experimental Example 2, the composition prepared from Example 13 (5g: Evening Primrose Seed 5g = 1: 1) was further included, and then the absorbance was measured continuously from the start of the reaction. . Using the obtained absorbance, the effect of inhibiting FAS activity by the composition of the present invention was measured and shown in Table 1 below (see FIG. 1).

Comparative Example 1. FAS inhibitory effect experiment using only green tea extract

In the reaction of Experimental Example 2, instead of the composition of the present invention, only the green tea extract of Example 2 was further included, and then absorbance was measured continuously from the start of the reaction. Using the obtained absorbance, the effect of inhibiting FAS activity by the composition of the present invention was measured and shown in Table 1 below (see FIG. 1).

Comparative Example 2. FAS inhibitory effect experiment using Evening Primrose seed extract

In the reaction of Experimental Example 2 instead of the composition of the present invention instead of the evening primrose seed extract of Example 4 further absorbance was measured from the beginning of the reaction. Using the obtained absorbance, the effect of inhibiting FAS activity by the composition of the present invention was measured and shown in Table 1 below (see FIG. 1).

TABLE 1

Green tea leaf: Evening primrose combination ratio FAS activity % of control Experimental Example 2 (Control) 0.8677 100.0 Experimental Example 3 1: 2.5 0.2184 25.17 Experimental Example 4         1: 5 0.3078 35.47 Experimental Example 5         1: 7.5 0.3791 43.69 Experimental Example 6  1: 1.67 0.1511 17.40 Experimental Example 7         1: 3.3 0.2722 31.37 Experimental Example 8         1: 5 0.3319 38.25 Experimental Example 9         1: 1 0.3976 45.82 Comparative Example 1 0.5908 68.13 Comparative Example 2 0.4419 50.90

Experimental Example 10 Experiment on Persistence of FAS Activity

The FAS of Experimental Example 2 was refrigerated at −20 ° C. for 50 days, and then FAS activity was measured according to the method of Experimental Example 2 and shown in Table 2 below (see FIG. 2).

Experimental Example 11. Experiment on Persistence of FAS Inhibitory Effect of the Composition of the Present Invention

The composition of Example 10 further refrigerated at 4 ℃ for 50 days in the reaction of Experimental Example 10, and then measured the effect on the persistence of the FAS activity by the composition of the present invention according to the method of Experimental Example 10 It is shown in Table 2 below (see Figure 2).

Comparative Example 3. Experiment on Persistence of FAS Inhibitory Effect Using Only Green Tea Extract

Except for using the green tea extract of Example 2 refrigerated at 4 ℃ for 50 days instead of the composition of Example 10 in the reaction of Experimental Example 10, the persistence of the FAS activity according to the method of Experimental Example 10 The effect was measured and shown in Table 2 below (see FIG. 2).

Comparative Example 4. Experiment on Persistence of FAS Inhibitory Effect Using Evening Primrose Seed Extract

The persistence of the FAS activity according to the method of Experimental Example 10, except that the evening primrose seed extract of Example 4, which was refrigerated at 4 ° C. for 50 days instead of the composition of Example 10, was used in the reaction of Experimental Example 10. The effect on was measured and shown in Table 2 below (see FIG. 2).

TABLE 2

FAS activity % of control Experimental Example 10 (Control) 0.7283 100.0 Experimental Example 11 0.1727 23.71 Comparative Example 3 0.5622 77.19 Comparative Example 4 0.6772 92.98

As a result of measuring the inhibitory effect of FAS on the composition of the present invention as in Experiment 6 (green tea leaves: evening primrose seed = 1: 1.67), the activity of FAS on the composition of the present invention is 17.4%, the inhibitory effect of FAS Was 82.6%, whereas the FAS activity of only the green tea extract of Comparative Example 1 was 68.13% (31.87% of FAS inhibitory effect), and the FAS activity of evening primrose seed extract of Comparative Example 2 was 50.9% (41.10% of FAS inhibitory effect). It can be seen that the FAS inhibitory ability of the composition of the present invention has a synergic effect.

In addition, as a result of experiments of sustaining the inhibitory effect of FAS on the composition of the present invention, green tea extract and evening primrose seed extract refrigerated for 50 days for 50 days, the inhibitory effect of FAS of the composition of the present invention is somewhat reduced than before However, with 23.71% activity, 76.29% of FAS activity inhibitory effect, whereas the green tea extract of Comparative Example 3 FAS activity of 77.19% (22.81% FAS inhibitory effect), evening primrose seed of Comparative Example 4 The FAS activity of the extract only is 92.98% (FAS inhibitory effect 7.02%), it can be seen that the FAS inhibitory ability of the composition of the present invention has a synergistic effect.

III. Obesity Inhibition Clinical Effect Experiment

Experimental Example 12.

Body weight (kg) and waist circumference (cm) were measured in adult women having an average obesity of 18 to 19%, and then the extract composition of Example 10 (3g green tea leaves: 5g evening primrose seeds 5g = 1: 1.67) was taken daily. After taking 3 times 21 days to determine the clinical efficacy effect, it is shown in Table 3 below. At this time, even long-term administration did not have side effects.

TABLE 3

Clinical effect experiment number                                          Height (cm) Before taking After taking Weight (kg) Waist circumference (cm) Weight (kg) Waist circumference (cm) One 169 74.1 93 73 91 2 174 79.3 95 78.3 91 3 178 78.9 86 77.5 85 4 163 89.3 102 85.8 90 5 165 63.2 80 61.9 73.8 6 161 82.1 97 80.4 95 7 162 68.5 95 66.4 91 8 176 76.5 91.7 76 89 9 160 65 79 65 76 10 161 51 63 48.1 62

As a result, after 21 days of ingesting the composition of the present invention, the weight was reduced by about 1 to 3.5 kg (average 1.3 to 5.7%) and the waist circumference was about 1 to 12 cm (average 1.6 to 7.8%). Decreased. These results show that the composition of the present invention is effective in weight loss by inhibiting the activity of FAS.

 As described above, the composition consisting of a mixture of green tea extract and evening primrose seed extract of the present invention has a very high effect in inhibiting the activity of FAS, by using a natural medicine harmless to the human body of various diseases associated with obesity without side effects It can be usefully used for treatment and prevention, and can be prepared as a solid preparation or a liquid preparation product while maintaining the physiological activity of the herbal medicine to the maximum, and thus easy to take.

Claims (7)

Dried evening primrose which is the raw material for extraction: Using the blending ratio of green tea leaves in a ratio of 1.6 to 3: 1 (weight ratio), each raw material was extracted, and then the evening primrose extract and the green tea extract were mixed in the same volume. Composition. The method of claim 1, The evening primrose extract is a composition for inhibiting obesity, characterized in that the seeds, petals, roots or a mixed extract of the evening primrose. delete delete delete The method of claim 1, The composition is a composition for inhibiting obesity, characterized in that the liquid, powder or solid phase. The method of claim 6, The composition is a composition for inhibiting obesity, characterized in that the drink or health food.

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