KR20230128186A - A composition for improving, preventing and treating of metabolic disorders comprising fermented aronia and pear - Google Patents

Abstract

The present invention relates to a composition for improving, preventing or treating metabolic diseases, which contains a mixed fermented product including aronia and pear juice as an active ingredient, thereby preventing obesity, type 2 diabetes, hypertension, hypercholesterolemia, hyperlipidemia, and high triglyceride. Metabolic diseases selected from the group consisting of hyperlipidemia, fatty liver, and arteriosclerosis can be improved, prevented, or treated, and are effective in antioxidants. In addition, since it is non-toxic, it can be ingested in the form of food.

Description

A composition for improving, preventing and treating metabolic disorders comprising fermented aronia and pear}

The present invention relates to a composition capable of improving, preventing or treating metabolic diseases by containing a mixed fermented product including aronia and pear juice as an active ingredient.

Metabolic diseases include obesity, diabetes, insulin resistance, abnormal lipid metabolism, hypertriglyceridemia, increase in free fatty acids, decrease in high-density cholesterol or high blood pressure, etc. and various complications resulting therefrom, and although the cause of the disease is not well known, it is generally assumed that insulin resistance is a fundamental problem.

The insulin resistance means that the body's response to insulin, a hormone that lowers blood sugar, is reduced so that muscle and fat cells do not take in glucose well, and more insulin is secreted to overcome this, causing various problems.

The imbalance of energy metabolism in the body caused by high-calorie, high-fat, and high-sugar meals can lead to obesity, induce insulin resistance and metabolic inflammation, and cause diseases such as abnormal lipid metabolism and type 2 diabetes, so it can be said to be a lifestyle-type disease. .

Visceral fat in the abdominal cavity is metabolically very active and secretes various substances. These substances raise blood pressure and interfere with the role of insulin, a blood sugar control hormone, resulting in hyperinsulinemia, insulin resistance and blood sugar rise, thereby reducing the risk of diabetes. It is known to increase the risk of cardiovascular disease by inducing inflammation or coagulation in blood vessels, triggering arteriosclerosis, increasing the risk of cardiovascular disease, and thus induced hypertension, diabetes, and hyperinsulinemia also increase the risk of cardiovascular disease.

It is known that adipose tissue in mammals is composed of two functionally different types of fat, that is, white fat and brown fat, and thus fat cells also exist in the form of white fat cells and brown fat cells. Unlike white adipocytes that store fat as an energy source, brown adipocytes decompose stored fat and release thermal energy, which is known to be beneficial for obesity and metabolic diseases.

Recently, it has been found that when sympathetic nerves are activated by exercise, cold, or spicy food intake, uncoupling protein 1 (UCP1), which is involved in mitochondrial thermogenesis, is expressed in white adipocytes and converted into beige adipocytes. Known. Beige adipocytes have a large amount of mitochondria, many small fat globules, and generate heat by oxidizing fat. Beige adipocytes are known to be beneficial for obesity and metabolic diseases similar to brown adipocytes, and most of the beneficial brown adipocytes found in humans are known as beige adipocytes.

Therefore, there is a need for natural substances capable of improving, preventing or treating these metabolic diseases.

Republic of Korea Patent No. 2249535 Korean Registered Patent No. 2019926

An object of the present invention is to provide a food composition capable of improving or preventing metabolic diseases by containing a mixed fermented product including aronia and pear juice as an active ingredient.

In addition, another object of the present invention is to provide a pharmaceutical composition capable of preventing or treating metabolic diseases by containing a mixed fermented product including aronia and pear juice as an active ingredient.

In addition, another object of the present invention is to provide a method for producing a food composition capable of improving or preventing metabolic diseases, including a mixed fermented product containing aronia and pear juice.

The food composition capable of improving or preventing metabolic diseases of the present invention for achieving the above object may contain a mixed fermented product including aronia and pear juice as an active ingredient.

The mixed fermented product may be a mixture of aronia, pear juice and sugars fermented with lactic acid bacteria.

The mixed fermented product may be fermented by inoculating 0.05 to 0.5 parts by weight of lactic acid bacteria into a mixture of 100 parts by weight of aronia, 5 to 20 parts by weight of pear juice, and 10 to 30 parts by weight of sugars.

The lactic acid bacteria are Lactobacillus plantarum , Lactobacillus casei , It may be at least one selected from the group consisting of Lactobacillus pentosus , Lactobacillus sakei , Leuconostoc mesenteroides , and Lactobacillus buchneri .

The fermentation may be performed for 1 to 5 days at 32 to 45 °C.

The sugars include fructooligosaccharide, sucrose, maltose, glucose, allulose, xylitol, sorbitol, mannitol, lactitol, polyglycitol, erythritol, maltitol, isomalt, isomaltooligosaccharide, xylooligosaccharide, galactooligosaccharide, stevio It may be at least one selected from the group consisting of side, sucralose, luohan fruit, licorice extract, acesulfame potassium, and thomas tin.

The metabolic disease may be at least one selected from the group consisting of obesity, type 2 diabetes, hypertension, hypercholesterolemia, hyperlipidemia, hypertriglyceridemia, fatty liver and arteriosclerosis.

In addition, the pharmaceutical composition capable of preventing or treating metabolic diseases of the present invention for achieving the other object described above may contain a mixed fermented product including aronia and pear juice as an active ingredient.

In addition, the method for producing a food composition capable of improving or preventing metabolic diseases of the present invention for achieving the above other object is (A) mixing aronia, pear juice and sugars; and (B) inoculating the mixture with lactic acid bacteria and performing fermentation at 32 to 45 ° C. for 1 to 5 days to obtain a mixed fermentation product.

The composition for improving, preventing or treating metabolic diseases containing a mixed fermented product including aronia and pear juice of the present invention as an active ingredient has ACE inhibitory activity, alpha-glucosidase inhibitory activity and HMG-CoA reduction Not only is it excellent in enzyme inhibitory activity, but it also lowers the content of fat globules, triglycerides and free glycerol in cells, thereby reducing obesity, type 2 diabetes, hypertension, hypercholesterolemia, hyperlipidemia, hypertriglyceridemia, fatty liver and arteries. Metabolic diseases selected from the group consisting of sclerosis can be improved, prevented or treated.

In addition, the composition of the present invention has an antioxidant effect, is non-toxic, and can improve, prevent, or treat metabolic diseases, so that it is effective in manufacturing medicines and foods.

1 is a graph measuring DPPH radical scavenging activity when using Example 1, Comparative Examples 2 to 6 and vitamin C of the present invention.
Figure 2 is a graph measuring the ACE inhibitory activity when using Example 1 of the present invention, Comparative Examples 2 to 6 and a positive control group.
Figure 3 is a graph measuring alpha-glucosidase inhibitory activity when using Example 1, Comparative Examples 2 to 6 and a positive control of the present invention.
Figure 4 is a graph measuring HMG-CoA reductase inhibitory activity when using Example 1 of the present invention, Comparative Examples 2 to 6 and a positive control group.
Figure 5 is a graph showing the generation of adipocytes in 3T3-L1 cells when Example 1 of the present invention, Comparative Examples 2 to 6, a control group and a positive control group were used.
Figure 6 is a graph showing the content of neutral fat in 3T3-L1 cells when using Example 1, Comparative Examples 2 to 6, a control group and a positive control group of the present invention.
7 is a graph showing the content of free glycerol in 3T3-L1 cells when using Example 1, Comparative Examples 2 to 6, and a control group and a positive control group of the present invention.

The present invention relates to a composition capable of improving, preventing or treating metabolic diseases by containing a mixed fermented product including aronia and pear juice as an active ingredient.

The metabolic disease may include at least one selected from the group consisting of obesity, type 2 diabetes, hypertension, hypercholesterolemia, hyperlipidemia, hypertriglyceridemia, fatty liver and arteriosclerosis.

Hereinafter, the present invention will be described in detail.

The composition capable of improving, preventing or treating metabolic diseases of the present invention contains a mixed fermented product including aronia and pear juice as an active ingredient.

Specifically, the mixed fermented product is inoculated with lactic acid bacteria in a mixture of aronia, pear juice and sugars, and then 32 to 45 ° C., preferably 35 to 38 ° C. for 1 to 5 days, preferably 1 to 3 days. it is fermented

The aronia has excellent antioxidant activity due to its high anthocyanin content, has excellent anticancer effects, and helps prevent liver damage, relieve inflammation, and relieve eye fatigue as well as prevent stroke.

In addition, the pear is effective for bronchial diseases, so it is good for colds, relief, asthma, etc., and helps defecation and diuresis. It removes phlegm and cough, relieves symptoms when your throat is hoarse or when your stomach is cold and sore, and it not only helps treat boils, but also has a detoxifying effect, so it eliminates hangovers.

The pear juice is used in an amount of 5 to 20 parts by weight, preferably 10 to 18 parts by weight, based on 100 parts by weight of aronia. If the content of pear juice is less than the lower limit, the effect on metabolic diseases may be reduced, and if it exceeds the upper limit, the sensory properties are reduced and the desired effect on metabolic diseases cannot be obtained.

In addition, the saccharides not only help fermentation to be performed smoothly, but also can increase organic acid acidity and improve antioxidant capacity and metabolic disease effects, such as fructooligosaccharide, sucrose, maltose, glucose, allulose, xylitol, sorbitol, Mannitol, lactitol, polyglycitol, erythritol, maltitol, isomalt, isomaltooligosaccharide, xylooligosaccharide, galactooligosaccharide, stevioside, sucralose, luohan, licorice extract, acesulfame potassium and thomastine and one or more selected from the group consisting of; Preferably, fructooligosaccharides may be used for excellent effects.

The sugar is used in an amount of 10 to 30 parts by weight, preferably 15 to 25 parts by weight, based on 100 parts by weight of aronia. When the content of saccharides is out of the above range, the functionality may be remarkably reduced and desired effects on metabolic diseases may not be obtained.

Examples of the lactic acid bacteria include Lactobacillus plantarum , Lactobacillus casei , At least one selected from the group consisting of Lactobacillus pentosus , Lactobacillus sakei , Leuconostoc mesenteroides and Lactobacillus buchneri , Preferably, Lactobacillus plantarum is used in that it exhibits excellent effects on metabolic diseases. In the case of using strains other than the above strains, there may be no or low effect on metabolic diseases.

When the temperature and time during the fermentation are less than the lower limit, the content of bioactive substances in the mixture of aronia, pear juice and saccharides may be lowered, and when it exceeds the upper limit, the desired effect is exhibited due to decomposition of the physiologically active substances may not be

The lactic acid bacteria are used in an amount of 0.05 to 0.5 parts by weight, preferably 0.1 to 0.2 parts by weight, based on 100 parts by weight of aronia. If the content of lactic acid bacteria is less than the lower limit, the normal fermentation may not be performed due to a long fermentation period, and if the content exceeds the upper limit, the effect on metabolic diseases may be remarkably low.

The mixed fermented product containing aronia and pear juice of the present invention can be used as a pharmaceutical composition in addition to a food composition.

The mixed fermented product containing aronia and pear juice of the present invention is meant to include a processed product of the mixed fermented product, such as mixed fermented product powder, formulated so that the mixed fermented product can be administered to animals in a broad sense. Although the present invention proceeds with a mixed fermented product containing aronia and pear juice, it will be expected by those skilled in the art that the desired effect can be achieved even in the same form as a processed product of the mixed fermented product.

On the other hand, in the present specification, the term 'contained as an active ingredient' means that it includes an amount sufficient to achieve the efficacy or activity of a mixed fermented product containing aronia and pear juice. For example, the mixed fermentation product is used at a concentration of 10 to 1500 μg/ml, preferably 100 to 1000 μg/ml. Since mixed fermented product is a natural product and does not have side effects on the human body even when administered in excess, the upper limit of the amount of mixed fermented product included in the composition of the present invention can be selected and implemented by those skilled in the art within an appropriate range.

The pharmaceutical composition of the present invention may be prepared using pharmaceutically suitable and physiologically acceptable adjuvants in addition to the above active ingredients, and the adjuvants include excipients, disintegrants, sweeteners, binders, coating agents, expanding agents, lubricants, A lubricant or flavoring agent may be used.

The pharmaceutical composition may be preferably formulated as a pharmaceutical composition by including one or more pharmaceutically acceptable carriers in addition to the active ingredients described above for administration.

Formulations of the pharmaceutical composition may be granules, powders, tablets, coated tablets, capsules, suppositories, solutions, syrups, juices, suspensions, emulsions, drops, or injectable solutions. For example, for formulation in the form of tablets or capsules, the active ingredient may be combined with an oral, non-toxic, pharmaceutically acceptable inert carrier such as ethanol, glycerol, water, and the like. In addition, if desired or necessary, suitable binders, lubricants, disintegrants and coloring agents may also be included in the mixture. Suitable binders include, but are not limited to, starch, gelatin, natural sugars such as glucose or beta-lactose, corn sweeteners, natural and synthetic gums such as acacia, tracacanth or sodium oleate, sodium stearate, magnesium stearate, sodium benzoate, sodium acetate, sodium chloride, and the like. Disintegrants include, but are not limited to, starch, methyl cellulose, agar, bentonite, xanthan gum, and the like.

In compositions formulated as liquid solutions, acceptable pharmaceutical carriers are sterile and biocompatible, and include saline, sterile water, Ringer's solution, buffered saline, albumin injection solution, dextrose solution, maltodextrin solution, glycerol, ethanol and One or more of these components may be mixed and used, and other conventional additives such as antioxidants, buffers, and bacteriostatic agents may be added if necessary. In addition, diluents, dispersants, surfactants, binders, and lubricants may be additionally added to prepare formulations for injections such as aqueous solutions, suspensions, and emulsions, pills, capsules, granules, or tablets.

Furthermore, using a method disclosed in Remington's Pharmaceutical Science, Mack Publishing Company, Easton PA as an appropriate method in the field, it can be preferably formulated according to each disease or component.

The pharmaceutical composition of the present invention can be administered orally or parenterally, and in the case of parenteral administration, it can be administered by intravenous injection, subcutaneous injection, intramuscular injection, intraperitoneal injection, transdermal administration, etc., preferably oral administration. .

The suitable dosage of the pharmaceutical composition of the present invention varies depending on factors such as formulation method, administration method, patient's age, weight, sex, morbid condition, food, administration time, route of administration, excretion rate and reaction sensitivity, A ordinarily skilled physician can readily determine and prescribe dosages effective for the desired treatment or prophylaxis. According to a preferred embodiment of the present invention, the daily dosage of the pharmaceutical composition of the present invention is 0.001-10 g/kg.

The pharmaceutical composition of the present invention may be prepared in unit dose form by formulation using a pharmaceutically acceptable carrier and/or excipient, or prepared by being placed in a multi-dose container. In this case, the formulation may be in the form of a solution, suspension or emulsion in an oil or aqueous medium, or may be in the form of an extract, powder, granule, tablet or capsule, and may additionally contain a dispersing agent or stabilizer.

In addition, the present invention provides a food composition for improving or preventing metabolic diseases containing a mixed fermented product including aronia and pear juice as an active ingredient.

The food composition according to the present invention can be formulated in the same way as the pharmaceutical composition and used as a functional food or added to various foods. Foods to which the composition of the present invention can be added include, for example, beverages, alcoholic beverages, confectionery, diet bars, dairy products, meat, chocolate, pizza, ramen, other noodles, chewing gum, ice cream, vitamin complexes, and health supplements. etc.

The food composition of the present invention may include not only a mixed fermented product containing aronia and pear juice as active ingredients, but also ingredients commonly added during food manufacturing, such as proteins, carbohydrates, fats, nutrients, seasonings. and flavoring agents. Examples of the aforementioned carbohydrates include monosaccharides such as glucose, fructose, and the like; disaccharides such as maltose, sucrose, oligosaccharides and the like; and polysaccharides such as conventional sugars such as dextrins and cyclodextrins and sugar alcohols such as xylitol, sorbitol and erythritol. As flavoring agents, natural flavoring agents [thaumatin, stevia extract (eg, rebaudioside A, glycyrrhizin, etc.]) and synthetic flavoring agents (saccharin, aspartame, etc.) can be used. For example, when the food composition of the present invention is made into drinks and beverages, citric acid, high fructose corn syrup, sugar, glucose, acetic acid, malic acid, fruit juice, and various plant extracts may be further included in addition to the mixed fermented product of the present invention. .

The present invention provides a health functional food comprising a food composition for improving or preventing metabolic diseases containing the mixed fermented product as an active ingredient. Health functional food is a food made by adding mixed fermented material to food materials such as beverages, teas, spices, chewing gum, confectionery, etc., or made into capsules, powders, suspensions, etc. However, unlike general drugs, it has the advantage of not having side effects that may occur when taking drugs for a long time using food as a raw material. The health functional food of the present invention obtained in this way is very useful because it can be consumed on a daily basis. The addition amount of the mixed fermented product in such a health functional food cannot be uniformly defined depending on the type of target health functional food, but it can be added within a range that does not impair the original taste of the food, and is usually 0.01 for the target food. to 50% by weight, preferably 0.1 to 20% by weight. In addition, in the case of health functional foods in the form of pills, granules, tablets or capsules, it is usually added in the range of 0.1 to 100% by weight, preferably 0.5 to 80% by weight. In one embodiment, the health functional food of the present invention may be in the form of pills, tablets, capsules or beverages.

Hereinafter, preferred embodiments are presented to aid understanding of the present invention, but the following examples are merely illustrative of the present invention, and various changes and modifications are possible within the scope and spirit of the present invention. It is obvious to those skilled in the art, It goes without saying that these variations and modifications fall within the scope of the appended claims.

[Measurement by strain]

Example 1. Lactobacillus plantarum ( Lactobacillus plantarum ) use

Aronia 100 parts by weight, pear juice (sugar content: 12.9 brix) 15 parts by weight and fructooligosaccharide (sugar content: 76 brix) 20 parts by weight of a mixture was inoculated with 0.1 parts by weight of Lactobacillus plantarum and inoculated at 36 ° C. Fermented for 1 day to obtain a mixed fermentation product.

Example 2. Lactobacillus casei ( Lactobacillus casei ) use

It was carried out in the same manner as in Example 1, except that Lactobacillus casei was used instead of Lactobacillus plantarum to obtain a mixed fermented product.

Comparative Example 1. Bifidobacterium bifidum ( Bifidobacterium bifidum )

It was carried out in the same manner as in Example 1, but a mixed fermentation product was obtained using Bifidobacterium bifidum instead of Lactobacillus plantarum .

<Test Example Ⅰ>

Test Example 1. Sugar content, pH, organic acid acidity

1-1. Sugar content (brix): measured using a sugar meter (MSTER-53T, ATAGO, Japan).

1-2. pH: measured using a pH-meter (Orion 420 A+, USA).

1-3. Organic acid acidity (%): 90 ml of distilled water was added to 10 g of the sample solution to homogenize, and titrated with 0.1N-NaOH solution until the pH reached 8.3. %) to confirm the organic acid acidity.

[Equation 1]

division Fermentation period (days) 0 One 2 3 5 0 One 2 3 5 0 One 2 3 5 Sugar content pH organic acid acidity Example 1 9.0 26.2 26.5 26.5 26.4 5.72 5.80 5.83 5.71 5.63 0.656 0.405 0.319 0.373 0.33 Example 2 9.0 26.6 26.5 26.5 26.3 5.72 5.68 5.69 5.54 5.49 0.656 0.481 0.486 0.500 0.53 Comparative Example 1 9.0 26.0 26.1 26.0 25.9 5.70 5.62 5.58 5.52 5.11 0.66 0.59 0.62 0.67 0.70

As shown in Table 1 above, it was confirmed that the mixed fermented product prepared according to Examples 1 and 2 of the present invention had higher pH and lower organic acid acidity as the fermentation period elapsed compared to Comparative Example 1.

In particular, it was confirmed that the mixed fermented product of Example 1 had higher pH and lower organic acid acidity as the fermentation period elapsed compared to Example 2.

Test Example 2. Measurement of DPPH radical and polyphenol content

2-1. DPPH radical scavenging activity (%): DPPH radical scavenging activity is a method of measuring the antioxidant activity of a sample using the principle that free radicals are scavenged by donating electrons to DPPH free radicals. The test method is to add 10 μL of sample and 190 μL of 100 μM DPPH (1,1-diphenyl-2-picrylhydrazyl) solution, react at room temperature for 30 minutes, and measure the absorbance at 515 nm using a microplate reader.

[Equation 2]

2-2. Polyphenol content (mg/ml): measured by modifying the Folin-Denis method. Add 100 μL of sample and 900 μL of distilled water, mix 100 μL of Folin-Ciocalteu reagent, react at room temperature for 5 minutes, add 1 mL of 7% Na ₂ CO ₃ solution and 400 μL of distilled water, mix, and react at room temperature for 90 minutes. After that, absorbance was measured at 750 nm. The standard calibration curve calculated the total phenol content using gallic acid.

division Fermentation period (days) 0 One 2 3 5 0 One 2 3 5 DPPH radical scavenging activity polyphenol content Example 1 - 53.5 52.9 50.2 41.5 15.7 143.8 146.3 149.8 136.5 Example 2 - 53.8 50.7 47.6 38.9 15.7 140.8 141.7 141.8 124.7 Comparative Example 1 - 26.8 22.7 19.9 13.1 15.7 103.4 105.2 105.9 73.3

As shown in Table 2 above, it was confirmed that the mixed fermented product prepared according to Examples 1 and 2 of the present invention had higher DPPH radical scavenging ability and polyphenol content as the fermentation period elapsed compared to Comparative Example 1.

In particular, when compared on the same fermentation day, it was confirmed that the mixed fermented product of Example 1 had higher DPPH radical scavenging ability and polyphenol content than Example 2.

In addition, on the 5th day of fermentation, it was confirmed that DPPH radical scavenging ability and polyphenol content were low in all groups.

Accordingly, it was confirmed that the mixed fermented product of Example 1 had the best antioxidant activity compared to the other groups.

Test Example 3. Measurement of the number of lactic acid bacteria

The number of lactic acid bacteria (cfu/ml) was evaluated using 3M dry film medium for lactic acid bacteria.

division Fermentation period (days) 0 One 2 3 5 Lactobacillus count Example 1 15.7 115X10 ¹⁰ 96X10 ¹⁰ 24X10 ¹⁰ 35X10 ⁹ Example 2 15.7 3X10 ⁵ 3X10 ⁵ 14X10 ⁶ 11X10 ⁴ Comparative Example 1 15.7 16X10 ³ 7X10 ³ 58X10 ² 4x10 ²

As shown in Table 3 above, it was confirmed that the mixed fermented product prepared according to Examples 1 and 2 of the present invention had a higher number of lactic acid bacteria as the fermentation period elapsed compared to Comparative Example 1.

In particular, it was confirmed that the mixed fermented product of Example 1 had a higher number of lactic acid bacteria as the fermentation period elapsed compared to Example 2.

In addition, it was confirmed that the number of lactic acid bacteria on the 5th day of fermentation was the lowest depending on the fermentation period.

Therefore, in the following test, the test was performed using the mixed fermented product of Example 1, in particular, the mixed fermented product of Example 1 on the 1st to 3rd day of fermentation.

[Measurement by content and substance type]

Example 1.

This is the mixed fermented product of Example 1.

Comparative Example 2. Skipping fermentation

A mixture of 100 parts by weight of aronia, 15 parts by weight of pear juice (sugar content: 12.9 brix) and 20 parts by weight of fructooligosaccharide (sugar content: 76 brix) was obtained.

Comparative Example 3. Use of blueberries

In the same manner as in Example 1, but using blueberries instead of aronia and fermenting for 1 day, a mixed fermented product was obtained.

Comparative Example 4. Pear juice content

In the same manner as in Example 1, but using 30 parts by weight of pear juice and fermenting for 1 day, a mixed fermented product was obtained.

Comparative Example 5. Fructooligosaccharide content

In the same manner as in Example 1, but using 40 parts by weight of fructooligosaccharide and fermenting for 1 day, a mixed fermented product was obtained.

Comparative Example 6. Omit pear juice

It was carried out in the same manner as in Example 1, but fermentation was performed for 1 day without using pear juice to obtain a mixed fermented product.

<Test Example Ⅱ>

For statistical analysis of the experiment, a significant difference in mean values between groups was confirmed, and the other letters were significant results when they had a p value lower than 0.05 as determined by Duncan's multiple range test (Duncan test vs. Comparative Example 2). considered. In order to indicate statistical significance between each group and Comparative Example 2, a to d were indicated.

Test Example 4. Measurement of DPPH radical scavenging activity_antioxidation

DPPH radical scavenging activity was measured using the same method as in 2-1 above.

1 is a graph measuring DPPH radical scavenging activity when using Example 1, Comparative Examples 2 to 6 and vitamin C of the present invention.

Vitamin C (L-ascorbic acid, sigma) was used as a positive control.

As shown in Figure 1, it was confirmed that the fermented mixture prepared according to Example 1 of the present invention showed better antioxidant activity than the fermented product of Comparative Examples 2 to 6.

In particular, among the mixed fermented products of Example 1, it was confirmed that the 1st and 2nd fermentation days had higher antioxidant activity than vitamin C, and the 3rd fermentation day was confirmed to have lower antioxidant activity compared to the 1st and 2nd fermentation days.

Test Example 5. Measurement of antihypertensive activity

Angiotensin I converting enzyme (ACE) is an enzyme associated with an increase in blood pressure, and ACE inhibitors (angiotensin converting enzyme inhibitors) are used as a first-line treatment for hypertension.

In the present invention, the antihypertensive activity of the sample was measured by ACE inhibitory activity. The ACE inhibitory effect was measured using an ACE kit (Dojindo Molecular Technologies, Inc.). After mixing, 20 μL of each concentration sample, 20 μL of substrate, and 20 μL of enzyme working solution were mixed, reacted at 37 ° C for 1 hour, and then added to the reaction solution. After adding 200 μL of the indicator working solution and reacting at room temperature for 10 minutes, the absorbance was measured at 450 nm using a microplate reader. Antihypertensive activity was expressed as the average value after measuring the same test solution three times.

[Equation 3]

Figure 2 is a graph measuring the ACE inhibitory activity when using Example 1 of the present invention, Comparative Examples 2 to 6 and a positive control group.

CoQ Ten Pang Pang (Healing Factory) was used as a positive control group.

As shown in FIG. 2, it was confirmed that the fermented mixture prepared according to Example 1 of the present invention showed superior ACE inhibitory activity than the fermented products of Comparative Examples 2 to 6.

In addition, 1 to 3 days of fermentation of the mixed fermented product of Example 1 showed similar ACE inhibitory activity, which was confirmed to be superior to the positive control group individually recognized by the Ministry of Food and Drug Safety.

Accordingly, it was confirmed that the mixed fermented product prepared according to Example 1 of the present invention showed an excellent effect on hypertension.

Test Example 6. Measurement of antidiabetic activity

Alpha-glucosidase is an enzyme involved in the rise of diabetes, and alpha-glucosidase inhibitors reduce postprandial hyperglycemia by inhibiting glycolysis and glucose absorption in the small intestine.

The alpha-glucosidase inhibitory effect was measured using an alpha-glucosidase inhibitor screening kit (BioVision, Inc.). After adding 10 μL of each concentration sample, 3 μL of substrate, and 17 μL of assay buffer, mixing, reacting at room temperature for 1 hour, and then microplate Absorbance was measured at 410 nm using a reader. Antidiabetic activity was expressed as the average value after measuring the same test solution three times.

[Equation 4]

After measuring the same test solution three times, it was expressed as an average value.

Figure 3 is a graph measuring alpha-glucosidase inhibitory activity when using Example 1, Comparative Examples 2 to 6 and a positive control of the present invention.

Acarbose was used as a positive control.

As shown in FIG. 3, it was confirmed that the mixed fermented product prepared according to Example 1 of the present invention exhibited better alpha-glucosidase inhibitory activity than the fermented product of Comparative Examples 2 to 6.

In addition, 1 to 3 days of fermentation of the mixed fermented product of Example 1 showed similar alpha-glucosidase inhibitory activity, which was confirmed to be superior to the positive control group individually recognized by the Ministry of Food and Drug Safety.

Accordingly, it was confirmed that the mixed fermented product prepared according to Example 1 of the present invention showed an excellent effect on diabetic ability.

Test Example 7. Anticholesterol activity

Our body regulates intracellular cholesterol concentration by connecting to LDL receptors that absorb dietary cholesterol, HMC-CoA reductase, an enzyme that regulates the rate of cholesterol synthesis in hepatocytes, enzymes that regulate the rate of synthesis of bile acids, and ester linking enzymes. In this case, hypercholesterolemia appears, and if cholesterol is not controlled over a long period of time, it leads to atherosclerosis. In the mechanism of cholesterol, inhibition of HMG-CoA reductase activity has the effect of reducing cholesterol synthesis, so anticholesterol activity was evaluated by HMG-CoA reductase inhibitory activity.

After preparing each sample by dissolving it in water by concentration, 4 μL of NADPH, 12 μL of HMG-CoA, and 2 μL of HMG reductase were added to 5 μL of the sample, and the absorbance change was measured kinetically for 20 minutes. Anti-cholesterol activity was expressed as the average value after measuring the same test solution three times.

[Equation 5]

Figure 4 is a graph measuring HMG-CoA reductase inhibitory activity when using Example 1 of the present invention, Comparative Examples 2 to 6 and a positive control group.

Monacosanol was used as a positive control.

As shown in FIG. 4, it was confirmed that among the mixed fermentation products prepared according to Example 1 of the present invention, fermentation days 1 and 2 showed superior HMG-CoA reductase inhibitory activity than the fermentation products of Comparative Examples 2 to 6.

In particular, the HMG-CoA reductase inhibitory activity on days 1 and 2 of fermentation was similar to that of the positive control individually approved by the Ministry of Food and Drug Safety, and significantly superior to that of day 3 of fermentation.

Accordingly, it was confirmed that the mixed fermented product, which was fermented for only 1 or 2 days according to Example 1 of the present invention, showed an excellent effect on cholesterol.

Test Example 8. Anti-obesity

The anti-obesity function was confirmed using 3T3-L1 cells. 3T3-L1 cell is a cell line derived from mouse fibroblast, 3T3 cell, and differentiates into adipocyte when cultured under appropriate conditions. It is widely used as a model to study the metabolic process of fat cells as well as the process of fat accumulation and differentiation of fat cells because its biological properties, such as the promotion of secretion of proteins during differentiation into adipocyt, are well known. Therefore, 3T3-L1 cells differentiated by differentiation promoting factors accumulate intracellular fat and show the morphology and characteristics of fat cells, such as cholesterol and triglyceride related to fat.

3T3-L1 subadipocytes derived from mouse embryos are the most frequently used cells in the anti-obesity in vitro system and contain 4.5 g/L, 10% fetal bovine serum, penicillin/streptomycin (100 μg/mL penicillin and 100 μg/L). mL streptomycin in 0.85% saline) and 1% DMEM containing 100 μM sodium pyruvate at 37 °C in a 5% CO ₂ incubator.

8-1. Measurement of intracellular adipocyte formation

In order to confirm the production of adipocytes in 3T3-L1 cells, Oil Red O staining, which is stained by specifically reacting to fat in lipid droplets, was performed, and the fat globules stained with accumulated fat were analyzed using absorbance. . Induction of 3T3-L1 subadipocytes into adipocytes became 100% confluent in the cell culture well, and after two days, they were transformed into adipocytes with a medium containing 100 μM 3-isobutyl-1-methylxanthine, 250 μM dexamethasone, and 170 μM insulin (MDI). After 3 days of induction, the MDI medium was replaced with DMEM medium containing 10% FBS and 170 nM insulin every other day until each experimental day. On the 3rd, 6th, and 9th days of 3T3-L1 cell culture, respectively, to completely remove the components of the culture medium, wash twice with phosphate buffered saline (PBS, pH 7.0), prepare a 0.6% Oil Red O solution, and dye for about 2 hours. After washing with PBS three times, 1.5 mL of isopropanol was added and the oil red elution solution was measured for absorbance at 500 nm.

Figure 5 is a graph showing the content of fat globules in 3T3-L1 cells using Example 1, Comparative Examples 2 to 6, and a control group and a positive control group of the present invention. Untreated was used as a control group, and Cissus was used as a positive control group.

The following [Table 4] is a photograph of 3T3-L1 intracellular fat globules of Example 1 of the present invention, Comparative Example 2, control group and positive control group taken with an OLYMPUS CKK41 microscope.

As shown in FIG. 5 and Table 4, it was confirmed that the mixed fermented product prepared according to Example 1 of the present invention had a lower content of intracellular fat globules than the fermented product of the control group and Comparative Examples 2 to 6.

In addition, among the mixed fermented products of Example 1, fermentation 1 and 2 showed similar intracellular fat globules, which was confirmed to be similar to the positive control group individually recognized by the Ministry of Food and Drug Safety.

8-2. Triglyceride content measurement

The intracellular triglyceride content was measured according to the quantification method indicated in the AdipoRed ^TM assay reagent, which is known to have high specificity for triglycerides as a Nile Red staining method. Intracellular triglyceride content was measured by dispensing 1X10 ⁶ cells per well in a 6-well plate, and the cells were cultured at 100% density. After further culturing for 2 days, the differentiation induction medium was treated with the extract, and cell differentiation induction was induced for 2 days, and then the medium was replaced with 10% FBS DMEM medium every 2 days and cultured for 9 days. Samples were treated together with the culture medium to be cultured. The differentiated 3T3-L1 was removed from the medium, washed twice with PBS, treated with 1 mL of AdipoRed reagent per well, incubated at 37 ° C for 10 minutes, and fluorescence was measured at excitation 485 nm and emission 572 nm. .

Figure 6 is a graph showing the content of neutral fat in 3T3-L1 cells when using Example 1, Comparative Examples 2 to 6, a control group and a positive control group of the present invention. Untreated was used as a control group, and Cissus was used as a positive control group.

As shown in FIG. 6, it was confirmed that the fermented mixture prepared according to Example 1 of the present invention had a lower intracellular triglyceride content than the control and fermented products of Comparative Examples 2 to 6.

In addition, among the mixed fermented products of Example 1, fermentation days 1 and 2 showed similar contents of intracellular neutral fat, which was confirmed to be similar to the positive control group individually recognized by the Ministry of Food and Drug Safety.

8-3. Measurement of the degree of lipolysis

In order to determine whether the above reduction in fat accumulation was due to lipolysis by the mixed fermented product, the content of free glycerol produced by decomposition of neutral fat present in fat globules was confirmed. The effect of the samples on lipolysis was measured using glycerol reagent and glycerol standard solution.

After treating 3T3-L1 cells with differentiation induction medium and samples, 50 μL of the culture medium obtained on days 3, 6, and 9 from the point of differentiation was added to 50 μL of glycerol reagent, and reacted at 37 ° C for 10 minutes in a 96 well plate. Measured in nm absorbance.

7 is a graph showing the content of free glycerol in 3T3-L1 cells when using Example 1, Comparative Examples 2 to 6, and a control group and a positive control group of the present invention. Untreated was used as a control group, and Cissus was used as a positive control group.

As shown in FIG. 7, it was confirmed that the fermented mixture prepared according to Example 1 of the present invention had a lower content of free glycerol than the fermented product of Comparative Examples 2 to 6, the control group.

In addition, among the mixed fermented products of Example 1, fermentation days 1 and 2 showed similar contents of intracellular neutral fat, which was confirmed to be similar to the positive control group individually recognized by the Ministry of Food and Drug Safety.

Accordingly, it was confirmed that the mixed fermented product prepared according to Example 1 of the present invention showed an excellent effect on obesity.

Test Example 9. Indicator component analysis

Analysis of marker components of the mixed fermented product of Example 1 fermented under optimal fermentation conditions was performed. Standard materials were carried out with tartaric acid, malic acid, lactic acid, and citric acid, which are representative organic acids of aronia. 10 mg of each standard material was put into a 10 mL volumetric flask, filled with distilled water up to the mark, and then filtered through a 0.45 μm membrane filter.

Analysis conditions of organic acids are shown in [Table 5] below.

Item Condition Detector UV DAD detector (210 nm) Column Capcell Pak MGⅢ (4.6×250 mm, 5μm) Column temperature 25 ℃ Injection vol. 20 µL Flow rate 0.7 mL/min
A : 0.025M KH ₂ PO ₄ (pH2.5) in water
B : water Mobile phase min A (%) B (%) 0 100 5 20 100 25

division Tartaric acid (μg/mL) Lactic acid μg/mL) Example 1 1 day 834 3,141 2 days 767 5,583 Comparative Example 2 3,809 - Comparative Example 3 94 1,150

As shown in Table 6 above, peaks of tartaric acid and lactic acid were detected in the fermented mixture prepared according to Example 1 of the present invention and the fermented blueberry product of Comparative Example 3, and fermentation In Comparative Example 2, which was not performed, lactic acid was not detected.

In addition, in the case of the mixed fermented product of Example 1, it was confirmed that the content of tartaric acid decreased and the content of lactic acid increased as fermentation progressed.

Hereinafter, formulation examples of the composition containing the powder of the present invention will be described, but the present invention is not intended to limit them, but only to be specifically described.

Formulation Example 1. Preparation of powder

500 mg of mixed fermented product obtained in Example 1

Lactose 100 mg

Talc 10 mg

A powder is prepared by mixing the above ingredients and filling them in an airtight bag.

Formulation Example 2. Preparation of tablets

300 mg of mixed fermented product obtained in Example 1

Corn Starch 100 mg

Lactose 100 mg

Magnesium stearate 2 mg

After mixing the above ingredients, tablets are prepared by tableting according to a conventional tablet manufacturing method.

Formulation Example 3. Preparation of capsule formulation

200 mg of mixed fermented product obtained in Example 1

3 mg of crystalline cellulose

Lactose 14.8 mg

Magnesium stearate 0.2 mg

Capsules are prepared by mixing the above ingredients and filling them into gelatin capsules according to a conventional capsule preparation method.

Formulation Example 4. Preparation of Injections

600 mg of mixed fermented product obtained in Example 1

Mannitol 180 mg

Sterile Distilled Water for Injection 2974 mg

Na ₂ HPO _4, 12H ₂ O 26 mg

It is prepared with the above component content per 1 ampoule according to the conventional method for preparing injections.

Formulation Example 5. Preparation of liquid formulation

4 g of mixed fermented product obtained in Example 1

Isomerized sugar 10 g

5 g mannitol

Appropriate amount of purified water

According to the conventional liquid formulation manufacturing method, each component is dissolved in purified water, lemon flavor is added in an appropriate amount, the above components are mixed, and then purified water is added to adjust the total amount to 100g, and then filled into a brown bottle to be sterilized. to prepare a liquid.

Formulation Example 6. Preparation of granules

1,000 mg of mixed fermented product obtained in Example 1

Appropriate amount of vitamin mixture

Vitamin A Acetate 70 μg

Vitamin E 1.0 mg

Vitamin B1 0.13 mg

Vitamin B2 0.15 mg

Vitamin B6 0.5 mg

Vitamin B12 0.2 μg

Vitamin C 10 mg

10 μg of biotin

Nicotinamide 1.7 mg

Folic acid 50 μg

Calcium Pantothenate 0.5 mg

Appropriate amount of mineral mixture

Ferrous sulfate 1.75 mg

Zinc Oxide 0.82 mg

Magnesium Carbonate 25.3 mg

Potassium Phosphate Monobasic 15 mg

Dibasic Calcium Phosphate 55 mg

Potassium citrate 90 mg

Calcium Carbonate 100 mg

Magnesium Chloride 24.8 mg

Although the composition ratio of the above vitamin and mineral mixture was prepared by mixing ingredients suitable for granules in a preferred embodiment, the mixing ratio may be arbitrarily modified, and after mixing the above ingredients according to a conventional granule manufacturing method, It can be prepared and used for preparing a health functional food composition according to a conventional method.

Formulation Example 7. Manufacturing of functional beverages

1,000 mg of mixed fermented product obtained in Example 1

Citric Acid 1,000 mg

100 g of oligosaccharides

2 g plum concentrate

1 g of taurine

Add purified water to total 900 mL

After mixing the above ingredients according to the normal health drink manufacturing method, stirring and heating at 85 ° C. for about 1 hour, the resulting solution is filtered and collected in a sterilized 2 L container, sealed and sterilized, and then refrigerated. It is used for preparing the functional beverage composition of the present invention.

Although the composition ratio is a mixture of ingredients suitable for a relatively favorite beverage in a preferred embodiment, the mixing ratio may be arbitrarily modified according to regional and ethnic preferences such as the class of demand, the country of demand, and the purpose of use.

Claims (9)

A food composition for improving or preventing metabolic diseases containing a mixed fermented product containing aronia and pear juice as an active ingredient. The food composition for improving or preventing metabolic diseases according to claim 1, wherein the mixed fermented product is a mixture of aronia, pear juice and saccharides fermented with lactic acid bacteria. The metabolic disease according to claim 1, wherein the mixed fermented product is fermented by inoculating 0.05 to 0.5 parts by weight of lactic acid bacteria into a mixture of 100 parts by weight of aronia, 5 to 20 parts by weight of pear juice, and 10 to 30 parts by weight of sugars. Food composition for improvement or prevention of. According to claim 2, wherein the lactic acid bacteria are Lactobacillus plantarum ( Lactobacillus plantarum ), Lactobacillus casei ( Lactobacillus casei ) , Metabolism, characterized in that at least one selected from the group consisting of Lactobacillus pentosus , Lactobacillus sakei , Leuconostoc mesenteroides and Lactobacillus buchneri A food composition for improving or preventing disease. The food composition for improving or preventing metabolic diseases according to claim 2, wherein the fermentation is performed at 32 to 45 °C for 1 to 5 days. The method of claim 1, wherein the saccharide is fructooligosaccharide, sucrose, maltose, glucose, allulose, xylitol, sorbitol, mannitol, lactitol, polyglycitol, erythritol, maltitol, isomalt, isomaltooligosaccharide, xylooligosaccharide A food composition for improving or preventing metabolic diseases, characterized in that it is at least one selected from the group consisting of galacto-oligosaccharide, stevioside, sucralose, luohan, licorice extract, acesulfame potassium, and tomatine. The method of claim 1, wherein the metabolic disease is at least one selected from the group consisting of obesity, type 2 diabetes, hypertension, hypercholesterolemia, hyperlipidemia, hypertriglyceridemia, fatty liver, and arteriosclerosis. Food composition for prevention. A pharmaceutical composition for preventing or treating metabolic diseases containing a mixed fermented product containing aronia and pear juice as an active ingredient. (A) mixing aronia, pear juice and sugars; and
(B) inoculating the mixture with lactic acid bacteria and performing fermentation at 32 to 45 ° C. for 1 to 5 days to obtain a mixed fermented product; a method for producing a food composition for improving or preventing metabolic diseases, including.

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