KR20150064521A - Fermented composition comprising bamboo salt and rhynchosia nulubilis - Google Patents

Abstract

The present invention relates to a fermented composition comprising bamboo salt and rhynchosia nulubilis and a manufacturing method thereof and usage of the same for a fatigue recovery and a hangover relief and more specifically relates to a fermented composition comprising bamboo salt and rhynchosia nulubilis manufactured by mixing bamboo salt, salicis radicis cortex, ground garlic and a sulfur fed duck with first fermented rhynchosia nulubilis and fermenting the mixture, which yields the fermented composition with an excellent sensory preference and nutrition and good effects on anti oxidation, DNA damage protection, fatigue recovery and hang over relief, so that the fermented composition can be used for fatigue recovery and hang over relief drug and health foods.

Description

FIELD OF THE INVENTION [0001] The present invention relates to a fermentation composition containing bamboo salt and bean curd,

The present invention relates to a fermentation composition prepared by fermenting bamboo salt and rice husk, a method for producing the fermented composition, and a use of the fermented composition as a medicine or a health food.

'Bamboo salt' is a process in which Korean sunbeams are poured into bamboo trees of Jiri Mountain and burned with pine wood, and the ninth is the material melted with high heat of 1,000 ℃ or higher by the rosin.

Bamboo salt is made by bamboo shoots that have been cut into ash pellets, crushed into bamboo shoots, and then crushed into bamboo shoots. And then heated to a temperature of 1000 ° C or higher while air is blown through the liquefied rosin as fuel to melt the salt and flow down into the liquid, And removing the silicate.

Bamboo salt can be divided into bamboo, bamboo, bamboo, bamboo, bamboo, and bamboo, depending on the number of times of grilling and grinding and the baking temperature. Several kinds of bamboo salt are commercially available in the market, and studies on the functional characteristics such as purity, mineral composition, structure analysis and pharmacological effect of bamboo salt have been gradually announced.

Recent trends in health-oriented eating habits have been dominated by lower salt intake and processed foods with bamboo salt, which is a functional material that can compensate for the drawbacks of salt, are emerging in the market.

The bean is called Seomoktae, and its scientific name is Rhynchosia It is Nulubilis . It is a perennial herbaceous plant of beans and its stems and leaves are brown. A yellow flower blooms in July, and a black and round fruit about 5 ~ 7 ㎜ in diameter is harvested in an elliptical pod. According to the 'primordial gangmok', it is better to use a small bean curd (豆 豆) as a medicine while the light is black. Sulfur spraying and growing are known to be more beneficial. In particular, the nature is warm, the taste is sweet, and there is no poison.

A typical reason for the superiority of rice husk beans is that they are found in black pigment in the shell. This drug is particularly excellent for antioxidant, vitreous and collagen functions, which inhibit the active oxygen toxicity caused by anthocyanin. It also promotes blood circulation, helps diuretic action, lowers edema, and has a strong detoxifying action. However, in spite of the excellent merits of these beans, it is very difficult to digest and difficult to eat because the cooked bean is cooked just like the New Year 's meal.

The present inventors have made intensive efforts to develop products using bamboo salt and bamboo salt that are safe as well as palatability and functional effects in order to promote the excellence of bamboo salt and to promote local development. As a result, the fermented composition prepared by fermenting bamboo salt and rice bran And has antioxidant activity, DNA damage-protecting activity, fatigue recovery effect and hangover resolution effect, and thus, the present invention has been completed.

The object of the present invention is to provide a method for the production of Rhynchosia Nulubilis ), bamboo salt, yugunipi , field garlic, and sulfur duck.

It is still another object of the present invention to provide a method for producing the above fermented composition.

It is another object of the present invention to provide a pharmaceutical or health food using the above fermented composition.

In order to achieve the above object, the present invention jwinunyi bean (Rhynchosia Nulubilis ), bamboo salt, rhizome , field garlic and sulfur duck.

In addition,

1) washing and growing the bean sole and cooling;

2) Inoculating the yeast into the legs of the step 1), then fermenting the yeast, and then drying it;

3) mixing the bamboo shoots, yucci, field garlic, and sulfur ducks with dried legs of step 2);

4) first aging the mixture of step 3), filtering and then heating;

5) secondary aging the fermented product of step 4)

The present invention also provides a method for producing a fermentation composition comprising a bean curd, a bamboo salt, a rhizome, a field garlic, and a sulfur duck.

In addition, the present invention provides a pharmaceutical composition containing a fermented composition containing the above fermented composition.

In addition, the present invention provides a pharmaceutical composition for hangover breakage containing the above fermented composition.

In addition, the present invention provides a health food containing a fermented composition containing the fermented composition.

In addition, the present invention provides a health food for hangover marrow containing the above fermented composition.

Fermented compositions produced by fermenting the bamboo salt and jwinunyi beans of the present invention was confirmed to have an in vitro (In vitro) test has a total phenolic content excellent in, and have excellent antioxidant activity, DNA damage protection activity, in vivo (In vivo experiment showed that the fatigue model animal induced by exercise and the hangover model animal induced by alcohol administration had antioxidative activity and DNA damage protective activity, respectively. Thus, the fermentation composition can be used as a medicine or health food for relieving hangover and restoring fatigue , And particularly useful for the development of functional fermented beverages.

FIG. 1 is a diagram showing a process for producing a fermented composition produced by bamboo salt and bean fermentation. FIG.
Fig. 2 is a diagram showing an ingredient analysis chart of the snow bean bean.
Fig. 3 is a diagram showing an ingredient analysis table of fermented dried beans. Fig.
FIG. 4 is a view showing an ingredient analysis table of the fermentation composition. FIG.
Fig. 5 is a diagram showing an ingredient analysis table of the fermentation composition (aging). Fig.
Figure 6 is a graph showing the total phenolic content of the fermentation composition produced by bamboo salt and bean fermentation:
All figures are mean ± standard deviation;
ac, significant difference at P <0.05 was indicated by ANOVA;
(A) fermented whole bean beans;
(B) Fermented chicken legs + sun salt;
(C) Fermented chicken legs + bamboo salt; And
(D) Fermented chicken legs + bamboo salt + yuu pee + field garlic + sulfur duck.
Figure 7 is a graph showing the peroxy radical scavenging activity of the fermentation composition produced by bamboo salt and bean fermentation:
Values are mean ± standard deviation;
The significant difference at ad, P < 0.05, is indicated by Duncan's multiple range test;
(A) fermented whole bean beans;
(B) Fermented chicken legs + sun salt;
(C) Fermented chicken legs + bamboo salt; And
(D) Fermented chicken legs + bamboo salt + yuu pee + field garlic + sulfur duck.
8 is a graph showing the cellular antioxidant capacity of the fermentation composition produced by bamboo salt and soybean fermentation against oxidative stress induced by AAPH in the HepG2 cell model:
Control group: HBSS-treated negative control;
AAPH: positive control treated with 80 [mu] M AAPH;
Values are mean ± standard deviation;
The significant difference at ad, P < 0.05, is indicated by Duncan's multiple range test;
(A) fermented whole bean beans;
(B) Fermented chicken legs + sun salt;
(C) Fermented chicken legs + bamboo salt; And
(D) Fermented chicken legs + bamboo salt + yuu pee + field garlic + sulfur duck.
9 is a graph showing the effect of treatment of the fermentation composition produced by bamboo salt and soybean fermentation on DNA damage induced in 200 μM H ₂ O ₂ in human leukocytes:
Values are mean ± standard deviation;
Bars with different subscripts are represented by Duncan's multiple range test;
NC: positive control treated with 1% DMSO (no oxidative stimulation);
(A) fermented whole bean beans;
(B) Fermented chicken legs + sun salt;
(C) Fermented chicken legs + bamboo salt; And
(D) Fermented chicken legs + bamboo salt + yuu pee + field garlic + sulfur duck.

Hereinafter, terms used in the present invention are defined.

That the term "jwinunyi beans" are seomoktae (鼠目太) used in the present invention, and the scientific name is Rhynchosia It is Nulubilis . It is a perennial herbaceous plant of beans and its stems and leaves are brown. A yellow flower blooms in July, and a black round fruit about 5-7 mm in diameter is harvested in an oval-shaped pod. According to the 'primordial gangmok', it is better to use a small bean curd (豆 豆) as a medicine while the light is black. Sulfur spraying and growing are known to be more beneficial. The nature is warm, the taste is sweet, and there is no poison.

The term "bamboo salt" used in the present invention refers to a substance which is made by bamboo bark of Jiri Mountain and burned with pine wood in 8 times, and 9th is melted with high heat of around 1,400 ℃ by rosin.

The term &quot; sulfur duck &quot; used in the present invention means sulfur fed to ducks, sulfur ducks as ducks, and sulfur contained in ducks is toxic and accumulated strong toxicity.

The term "field garlic" used in the present invention is a garlic raised in a field, which is excellent in sterilization, insecticidal action, gigantic birth, and hard-boiled ingredients of garlic juice make the bones hard. On the other hand, rice paddy can not be used as medicine because of accumulated pesticide poison.

As used herein, the term &quot; yugunipi &quot; refers to a bark or root bark of elm and elm, which improves wound scarring by virtue of its efficacy and is very good for nasal diseases, and in particular is useful for the treatment of gastric ulcer, duodenal ulcer, small intestine, Esophageal ulcers, and other ulcers.

The term &quot; hangover &quot; used in the present invention means that unusual discomfort, headache, or decline in working ability of the body and mind, etc., lasts for 1 to 2 days after being awakened from the surface after drunkenly drinking.

The term &quot; fatigue &quot; as used in the present invention means a depressed state of mental and physical functions caused by continuous and repeated mental or physical work. It is a state in which fatigue manifests itself as mental fatigue and physical fatigue, central fatigue and peripheral fatigue, Fatigue, and chronic fatigue. In addition, there is an orgasm with severe deterioration in function, and it is divided into muscle, nerve fatigue and heart fatigue.

Hereinafter, the present invention will be described in detail.

The present invention relates to the use of Rhynchosia Nulubilis ), bamboo salt, rhizome , field garlic and sulfur duck.

In the fermentation composition, it is preferable that the ratio is 1: 1: 1: 1 to 1: 1: 0.5 to 1: 0.5 to 1: 0.5 to 1, and the ratio of beans: bamboo salt: More preferably in a weight ratio of 0.7: 0.8.

Preferably, the above-mentioned chicken legs are fermented fermented soybeans, and 1) washing and boiling the soybeans with the chicken legs; 2) cooling; 3) inoculating the yeast; 4) fermenting step; And 5) drying the fermented soybeans.

It is preferable that the bamboo salt is a bamboo salt of bamboo which is used for 9 times, but not limited thereto, all bamboo salts can be used.

The bamboo salt is formed by 1) placing the sun salt in a bamboo container and blocking the inlet with yellow soil; 2) Bamboo bamboo in bamboo shoots and heating (800 ℃) with pine wood; 3) a step of cooling and bifurcating the bamboo salt; 4) pulverizing the baked salt and repeating steps 1) to 3) 8 times; 5) It is more preferable to use a product made by a manufacturing method comprising heating the salt baked 9 times (1400 ° C) so that the salt is melted and flowed down into the liquid to remove the ash and the silicate.

In addition,

1) washing and growing the bean sole and cooling;

2) Inoculating the yeast into the legs of the step 1), then fermenting the yeast, and then drying it;

3) mixing the bamboo shoots, yucci, field garlic, and sulfur ducks with dried legs of step 2);

4) first aging the mixture of step 3), filtering and then heating;

5) secondary aging the fermented product of step 4)

The present invention also provides a method for producing a fermentation composition comprising a bean curd, a bamboo salt, a rhizome, a field garlic, and a sulfur duck.

In the above method, in step 1), the soybean is preferably dipped for 10 hours to 15 hours at room temperature, and then heated for 3 hours in a high temperature and for 3 hours in a low temperature.

In the above production method, the fermentation in step 2) is preferably fermented at 32 to 40 DEG C for 15 to 20 hours, more preferably at 36 to 38 DEG C for 18 hours.

The drying in step 2) may be carried out using a natural drying method or a drying machine. The drying conditions are preferably 30 to 40 ° C for 10 to 20 hours, and 35 ° C for 12 to 15 hours Is more preferable.

In the above production method, it is preferable to pre-treat the roots, field garlic and sulfur ducks of the step 3) before mixing, and it is preferable that the pretreatment rods are subjected to hot water extraction after being subjected to purified water for 24 hours each.

In the above production method, the mixing of step 3) is preferably carried out at a weight ratio of 1: 1: 0.5 to 1: 0.5 to 1: 0.5 to 1 in the ratio of soybean: bamboo salt: yuukee: field garlic: More preferably in a weight ratio of 1: 1: 1: 0.7: 0.8.

In the above production method, the heating in step 4) is preferably a heat treatment step for preventing contamination of the aged fermented composition with microorganisms and the like, preferably at 80 to 100 캜 for 30 minutes to 1 hour, It is more preferable to heat for 30 minutes to 40 minutes.

The present invention also provides a pharmaceutical composition containing a fermented composition according to the present invention.

The present invention also provides a pharmaceutical composition for hangover use containing the fermentation composition according to the present invention.

The pharmaceutical composition of the present invention may contain, in addition to the fermentation composition according to the present invention, one or more active ingredients exhibiting the same or similar functions.

The pharmaceutical composition of the present invention may further comprise a pharmaceutically acceptable additive, wherein the pharmaceutically acceptable additives include starch, gelatinized starch, microcrystalline cellulose, lactose, povidone, colloidal silicon dioxide, calcium hydrogen phosphate Starch glycolate, starch glycolate, starch glycolate, carnauba wax, synthetic aluminum silicate, stearic acid, magnesium stearate, aluminum stearate, sodium carboxymethylcellulose, sodium carboxymethylcellulose, sodium carboxymethylcellulose, Calcium stearate, white sugar, dextrose, sorbitol and talc may be used. The pharmaceutically acceptable additives according to the present invention are preferably included in the composition in an amount of 0.1 to 90 parts by weight, but are not limited thereto.

The pharmaceutical composition of the present invention may be administered orally or parenterally in various formulations at the time of actual clinical administration. In the case of formulation, a diluent such as a filler, an extender, a binder, a wetting agent, a disintegrant, . &Lt; / RTI &gt; Solid formulations for oral administration include tablets, pills, powders, granules, capsules and the like, which may contain at least one excipient such as starch, calcium carbonate, sucrose, lactose (Lactose), gelatin, and the like. In addition to simple excipients, lubricants such as magnesium stearate talc may also be used. Examples of the liquid preparation for oral use include suspensions, solutions, emulsions and syrups, and various excipients such as wetting agents, sweetening agents, fragrances, preservatives and the like may be included in addition to water and liquid paraffin, which are simple diluents commonly used . Formulations for parenteral administration may include sterile aqueous solutions, non-aqueous solutions, suspensions, emulsions, freeze-dried preparations, and suppositories. Propylene glycol, polyethylene glycol, vegetable oil such as olive oil, injectable ester such as ethyl oleate, and the like can be used as the non-aqueous solvent and suspension agent. Examples of the suppository base include witepsol, macrogol, tween 61, cacao butter, laurin, glycerogelatin and the like.

The pharmaceutical composition of the present invention may be administered orally or parenterally in accordance with the desired method, and may be administered orally, parenterally or intraperitoneally, rectally, subcutaneously, intravenously, intramuscularly, It is preferable to select the injection method. The dosage varies depending on the patient's body weight, age, sex, health condition, diet, administration time, administration method, excretion rate, and disease severity.

The dosage of the pharmaceutical composition of the present invention varies depending on the patient's body weight, age, sex, health condition, diet, administration time, administration method, excretion rate and severity of the disease, 0.001 to 1000 mg / kg, preferably 0.01 to 100 mg / kg, and can be administered 1 to 6 times a day.

The pharmaceutical composition of the present invention can be used alone for improvement and treatment of fatigue or hangover, or in combination with methods using hormone therapy, chemotherapy and biological response modifiers.

In addition, the present invention provides a health food containing the fermented composition according to the present invention.

In addition, the present invention provides a health food for hangover poultry containing the fermented composition according to the present invention.

The health food of the present invention can be used as it is or in combination with other food or food ingredients according to the present invention, and can be suitably used according to conventional methods.

The health food of the present invention includes both health functional food and health supplement food, and there is no particular limitation on the kind of these health food. Examples of the foods to which the composition can be added include dairy products including beverages, tea, drinks, alcoholic beverages, vitamin complexes and various kinds of spices, meat, sausages, bread, chocolates, candies, snacks, confectioneries, noodles, gums, Products, etc., and includes all the health foods in the ordinary sense.

Among the health foods of the present invention, the health beverage composition may contain various flavors or natural carbohydrates as additional components such as ordinary beverages. Such natural carbohydrates are monosaccharides such as glucose and fructose, disaccharides such as maltose and sucrose, and polysaccharides such as dextrin and cyclodextrin, and sugar alcohols such as xylitol, sorbitol and erythritol. Examples of sweeteners include natural sweeteners such as tau martin and stevia extract, synthetic sweeteners such as saccharin and aspartame, and the like. The ratio of the natural carbohydrate is generally about 0.01 to 0.10 g per 100 ml of the composition of the present invention.

In addition to the above, the health food of the present invention may contain various nutrients, vitamins, electrolytes, flavors, colorants, pectic acids and salts thereof, alginic acid and its salts, organic acids, protective colloid thickeners, pH adjusters, stabilizers, preservatives, glycerin, , A carbonating agent used in carbonated drinks, and the like. It may also contain flesh for the production of natural fruit juices, fruit juice drinks and vegetable drinks. These components may be used independently or in combination. Although the ratio of such additives is not critical, it is generally selected in the range of 0.01 to 0.1 parts by weight per 100 parts by weight of the fermentation composition according to the present invention.

Hereinafter, the present invention will be described in more detail with reference to Examples and Preparation Examples.

However, the following examples and preparative examples are provided only for the understanding of the present invention, and the scope of the present invention is not limited to the following examples and preparative examples.

< Example  1> Bamboo salt A little bean  Preparation of fermentation composition produced by fermentation

<1-1> Screening and storage of raw materials

Selection criteria for each raw material required for manufacturing are set as follows.

Bamboo salt is a 9-fold bamboo salt (Phosphate Bamboo Salt Village, Korea). It has high transparency, relatively uniform size, hard stone, strong salty taste and sweet taste.

The husked beans were only used domestically, and the beans were rounded without any cracks, 3 to 5 mm in size, black, moist and pest-free and clean.

Ukeunpi was only used domestic, well dried and clean to avoid mold. The distinctive scent of yugun pei was bloomed deeply and used as a lot of progress in water.

The field garlic uses only domestically produced whole red light, one small one 4 cm, the big one 6 cm thick, the skin thin and well dried, and the six garlic.

Sulfur ducks were fed with natural sulfur for more than 2 years. There was no smell of peculiar birds. Only ducks were used.

The guideline for raising the sulfur duck is as follows.

Breeding guide of sulfur duck Food selection criteria Rice bran Rice bran made by crushing non-chemical rice bran finely Medicinal residue High-grade medicinal residues used in phosphoric acid clinics brimstone Natural sulfur imported from the Indonesian volcanic area,
Sulfur with fine powder particles with a pale yellow color Food mix ratio The ratio of rice bran, medicinal material, and sulfur is 1: 1: 0.04 Feeding time Twice a day (8:00 am / 18:00 pm) Food intake Feed per duck: Average 500 to 700 g / day,
Sulfur Intake: Average 10 g / day Breeding period More than 2 years Duck weight Weight 2.5kg or more Slaughter method With built-in hair,
Clean and clean

<1-2> Twigs of beans  Fermentation process

First, Korean domestic bean was selected according to the selection criteria. The selected bean curd was immersed at room temperature for 10 ~ 15 hours for 3 hours in high temperature and 3 hours in low temperature. After the increase, the puppy was inoculated with yeast. The optimum temperature of the soybean at the time of inoculation with koji was 35 ~ 38 ℃, so the soybean was boiled at 38 ℃ ± 2 and the koji was inoculated. The husks inoculated with the yeast were fermented at 32 ~ 38 ℃ for 18 hours. Optimum Fermented Rats The characteristics of the beans are that the sticky threads are stretched between the beans, the fermented beans smell, the smell disappears, and the white mold appears on the surface. Fermented dry bean was dried using a drier. At this time, the temperature was adjusted to 35 캜 and dried at a constant temperature for 12 to 15 hours. The well-dried crust is so dry that the beans are so rusty that the beans are almost dry.

<1-3> Pretreatment of raw materials

The pretreatment process of the raw materials was carried out as follows.

Sulfur duck was added to purified water for 24 hours to prepare hot water extract. The extracts of hot water extracts were prepared by suspending the yugi cakes in purified water for 24 hours. The field garlic was boiled in purified water for 19 hours to produce hot water extract. Bamboo salt was used in the form of fine powder and used directly. After the yeast inoculation, the soybeans were dried by a fermentation process.

<1-4> Setting the mixing ratio of raw materials

The pretreated raw materials were mixed at the following mixing ratios.

Setting the mixing ratio of raw materials Condition 1 Condition 2 Condition 3 Sulfur duck 1,000 g 1,000 g 1,000 g Yugunpi 1,000 g 1,000 g 1,000 g Field garlic 500 g 1,000 g 1,000 g Bamboo bamboo 700 g 700 g 1,000 g A little bean 800 g 800 g 800 g

Sensory evaluation of raw material mixing ratio was carried out by 5 point scale sensory evaluation for employees of phosphoric acid bamboo salt village co.

Sensory evaluation of raw material mixing ratio Extremely
I hate it. I hate it. is average Likes Extremely
Likes. Condition 1 One 4 3 2 Condition 2 2 5 3 Condition 3 2 6 2

As a result, the condition 1 was less than that of the other mixture, and the bamboo salt was too salty. Condition 2 was less salty than the condition 1, but the five kinds of raw materials were well mixed and the complex taste was felt. Condition 3 contained bamboo salt more than other mixtures and the salty taste was very strong. Therefore, the final selection of the raw material mixing ratio was determined as condition 2.

<1-4> Heating process of raw materials

A heat treatment process was performed to prevent contamination of the aged fermentation composition with microorganisms and the like. Care was taken for volatilization of the odor component due to decomposition of the fat component and loss of the active ingredient, followed by heating at 80 to 98 ° C for about 30 minutes.

&Lt; 1-5 > Preparation of fermentation composition

The fermentation composition produced by the fermentation of bamboo salt and the bean curd of soybean was firstly fermented with the yeast and the step of making the soybean fermented soybeans, and the second step was the fermentation of the soybean fermented soybean with bamboo salt, sulfur duck, Followed by aging to prepare a final fermentation composition.

Specifically, the beans were firstly washed and then boiled, cooled at a constant temperature of 40 ° C, and then fermented to obtain fermented soybeans by first inoculation with yeast, followed by drying. Then, fermented dried soybeans, which were prepared in the second step, were mixed with bamboo salt, sulfur duck, yukeeji, field garlic, and water and then aged. After filtration, the fermented liquid was recovered by heating and then second- (Fig. 1).

< Example  2> Bamboo salt A little bean  General composition analysis of fermented composition produced by fermentation

The general component test method is a test method for ingredients generally contained in food. It is a test method for food standard, purity and nutritional value. It is used for testing of moisture, ash, nitrogen compounds, carbohydrates and lipids, . As a general test in ordinary foods, it is necessary to test for appearance, taste, moisture, ash, crude protein, crude fat and crude fiber, and in special cases, specific gravity, amino acid content, various saccharides and lipids. In general, carbohydrate is expressed as the amount obtained by subtracting the amount of moisture, crude protein, crude fat, crude fiber and ash from 100 g of the sample, and the test result of the common ingredient in food and drink is usually expressed as a percentage. The following experiment was carried out using the general component analysis according to the Food Code of the Food and Drug Administration.

<2-1> Sugar measurement

Test solutions were prepared as follows. Specifically, about 5 g of the sample was precisely weighed and placed in a 50 ml volumetric flask. Then, 25 ml of water was added to dissolve, and then 50 ml of acetonitrile was added. The test solution was then filtered through a 0.45 um membrane filter.

The standard solution was prepared as follows. Specifically, 100 ml of each standard of fructose, glucose, sucrose, and maltose was precisely weighed in a measuring flask and dissolved in 50 ml of water, and then filled up to 100 ml with acetonitrile.

LC (Agilent 1200) was used as the measuring device, and Shodex Asahipak NH2P 50-4E was used as the column.

The test result (g / 100g) was substituted into the following equation.

Sugar (g / 100g) = C x V / S x D

C: detection concentration (mg / L);

V: final volume of test solution (mL);

S: Amount of sample (g); And

D: Dilution of test solution.]

<2-2> Crude protein  Measure

The degradation of the specimen was performed as follows. Specifically, about 1 g of the sample was precisely taken and 2 pieces of the decomposition promoter were added (the ratio of H ₂ SO ₄ and K ₂ SO ₄ was 1.4 to 2.0: 1 for the decomposition promoter). Then add 12 mL of concentrated sulfuric acid to the digestion tube and add 15 mL of concentrated sulfuric acid if the fat content of the sample is 10% or more. Then, it was decomposed in a decomposition apparatus at 420 ° C for 45 to 60 minutes. When the color of the decomposition liquid became transparent pale blue or transparent yellow, it was cooled to room temperature.

Distillation and titration were carried out as follows. Specifically, a Kjeldahl flask containing an experimental solution was mounted on an automatic nitrogen analyzer, and then an automatic distillation and titration apparatus was used.

At this time, SOX 416 Macro was used as a proteolysis apparatus and Gerhardt Vapodest 45s was used as a distillation and titration apparatus.

The crude protein content was calculated by substituting the titration value into the following equation.

(%) = 1.4007 x (a-b) x (100 / sample weight (mg)) x Nitrogen Coefficient

a: the number of mL of 0.1 N sulfuric acid required for neutralization in this test;

b: the number of mL of 0.1 N sulfuric acid consumed for neutralization in the blank test; And

1 mL of 0.1 N sulfuric acid = 1.4007 mgN.

<2-3> Crude fat  Measure

1 ~ 10 g of specimen was precisely weighed and placed in a fat extraction tube. Water was added to make about 11 mL, heated to 40 ~ 50 ℃ and completely mixed by shaking. Then, 1.5 mL of ammonia water (2.0 mL when the liquid was acidic) was added thereto, followed by mixing, followed by mixing with 10 mL of alcohol. Then, add 25 mL of ether, mix gently, open the stopper, blow off the ether, close the stopper again, and shake hard for about 1 minute. Then, it was centrifuged or left at 600 rpm, and when the supernatant liquid became completely transparent, the supernatant was transferred to an Erlenmeyer flask. Then, 15 mL of the ether was added to the remaining solution in the tube, and the above operation was carried out. Then, it was extracted three times with 15 mL of ether again. Then, the outlet of the tube and the funnel were cleaned with ether, and the filtrate and the washed solution were combined with Erlenmeyer flask and the solvent was blown off in a water bath. Then, the Erlenmeyer flask was placed in a drier at 100 ± 2 ° C, dried until the weight became constant, and the content of crude fat was substituted into the following equation.

Crude fat (%) = (W1-W2) / S * 100

W1: weight of container (g);

W2: weight of container after drying (g); And

S: Weight of sample (g).

<2-4> Trans fat measurement

Test solutions were prepared as follows.

Specifically, the sample was weighed in a predetermined amount, placed in a cylindrical filter paper, dried at 105 ° C, extracted for 8 hours with a syringe extractor, and evaporated in a water bath. Then, a constant amount was added at 100 캜 and weighed to determine the crude fat content. Then, the lipid was called 0.025 g, and then 1.5 mL of 0.5 N methanolic sodium hydroxide solution was added, followed by 1 mL of IS. Then, nitrogen was blown and immediately the lid was closed. After mixing, the mixture was heated at 100 ° C in a heating brock for 5 minutes. After cooling, 2 mL of a 14% trifluoro-borane methanol solution was added. Then, nitrogen was blown and immediately the lid was closed and mixed. The mixture was heated at 100 ° C for 30 minutes in a heating brock, cooled to 30 to 40 ° C, and 1 ml of isooctane was added. Then, nitrogen was blown in and immediately the lid was closed. After shaking vigorously at this temperature, 5 mL of a saturated sodium chloride solution was added, and the mixture was blown with nitrogen, covered with a lid and shaken. Then, after cooling to room temperature, the iso-octane layer was taken in a new glass tube, dehydrated with anhydrous sodium sulfate, and then nitrogen was blown into it and used as a test solution.

The standard solution used was a SIPELCO 37 Component FAME Mix (Part NO, 47885-U) from SUPELCO, and 40 mg of undecanoic acid was used as an internal standard solution.

 GC (Perkin Elmer GC-Clarus 600) was used as the measuring apparatus, and SP-2560 was used as the column.

The test result (g / 100g) was substituted into the following equation.

Saturated fat (g / 100g food) = crude fat content (g / 100g food) x saturated fatty acid content (g / 100g fatty acid) / 100; And

Trans fat (g / 100g food) = crude fat content (g / 100g food) x trans fatty acid content (g / 100g fatty acid) / 100.

<2-5> Cholesterol measurement

Test solutions were prepared as follows.

Specifically, about 30 mL of water and 1 mL of 5 g cholestane were added to 1 to 10 g of a sample, and homogenized (a cholesterol content of 20 to 1000 mg / 100 g was taken as a sample amount). Then, this solution was transferred to a separatory funnel, and 200 mL of chloroform: methanol (2: 1) was mixed and extracted with shaking for about 5 minutes, and then allowed to stand. After the chloroform layer was clinched, the solution was repeatedly extracted twice. The combined chloroform layers were combined and washed with 100 mL of 0.5% sodium hydroxide solution. The mixture was dehydrated with anhydrous sodium sulfate, filtered, concentrated under reduced pressure at 40 ° C or lower, and 20 mL of 2N potassium hydroxide ethanol solution was added to the residue. After refluxing, the mixture was saponified at 85 ° C for 1 hour, cooled, transferred to a separatory funnel along with about 20 mL of water, and extracted with ethyl acetate four times with about 20 mL each. The ether extracts were combined and washed with about 20 mL of water, washed with phenolphthalein indicator until no red color appeared, and the ether layer was dehydrated with anhydrous sodium sulfate, filtered and concentrated under reduced pressure. The residue was dissolved in 2 mL of hexane Dissolved and used as a test solution.

Cholesterol standard solution was prepared by dissolving 25 mg of cholesterol in hexane to 50 mL, and using 25 mL of 5a-cholestane as an internal standard solution in 100 mL of hexane.

 GC (Perkin Elmer GC-Clarus 600) was used as the measuring apparatus and HP-5 was used as the column.

The test result (mg / 100g) was substituted into the following equation.

Cholesterol (mg / 100 g) = C 占 V / S 占 D 占 1/10

C: detection concentration (mg / L);

V: final volume of test solution (mL);

S: Amount of sample (g); And

D: Dilution of test solution.

<2-6> Measurement of sodium

The sample pretreatment (dry method) was carried out as follows.

Specifically, a specimen (an amount corresponding to 5 to 20 g as a dried product) was taken in a crucible, dried and carbonized, and then calcined at 450 ° C for 20 hours. The ash was wetted with water, 3 mL of hydrochloric acid was added, and the mixture was dried in a water bath. Then, 2 mL of hydrochloric acid was added and dissolved by heating. The mixture was filtered through a filter paper and 50 mL of water was used as a test solution.

The standard solution was prepared as follows.

Specifically, sodium was used as a standard solution for STD1, STD2, STD3 and STD4 by diluting 1,000 ug / mL of sodium standard solution to 0.0 ppm, 1.0 ppm, 5.0 ppm, and 10.0 ppm with tertiary distilled water. All the piglets used for the analysis of heavy metals were immersed in 5% HNO3 solution for 24 hours to remove pollutants, and then lapped with third distilled water several times to dry.

 The measurement instrument was an inductively coupled plasma emission spectrometer (ICP-OES, Perkin-Elmer OPTIMA 7000 DV).

The test result (mg / 100g) was substituted into the following equation.

Na concentration (mg / kg) = instrumental concentration (mg / L) × final volume (mL) / sample volume (g) × dilution factor

<2-7> Moisture measurement

Moisture content was measured by atmospheric pressure heating drying method.

Specifically, weigh accurately 3 to 5 g of specimen in a weighing dish which has been preliminarily heated to a constant weight (but if it is difficult to dry, 20 g of refined margarine 20 g and a glass rod are put into a constant volume, After mixing, the glass rods were left in place. The lid was slightly opened and placed in a drier of the prescribed temperature according to the application range of the test method application range and dried for 3 to 5 hours. After drying, it was cooled in a desiccator for about 30 minutes and weighed. The weighing dish was again dried for 1 to 2 hours and the same operation was repeated until the weight became constant.

The test result (%) was substituted into the following equation.

Water (%) = (b-c) / (b-a) 100

a: weight of the weighing dish (g);

b: Weight of weighing dish and sample (g); And

c: Weight after drying (g).

<2-8> Measuring pots

The weight of the crucible was measured as follows.

Specifically, a clean crucible was heated in a painting furnace at a temperature of 600 ° C or higher for several hours, transferred to a desiccator, cooled to room temperature, weighed, heated again for 2 hours and then weighed dry and the operation was repeated until the weight became constant.

The painting was carried out as follows.

Specifically, after the pretreatment, the container was transferred directly to a painting furnace, and the mixture was continuously heated at 550 to 600 ° C. until a white to off-white ash was obtained, and after the painting was completed, the heating was stopped and the mixture was cooled to about 200 ° C. Transferred to a desiccator, allowed to cool, and weighed.

The test result (%) was substituted into the following equation.

Ash (%) = (W1-W2) / S x 100

W1: weight of crucible (g);

W2: Crucible weight after painting (g); And

S: Amount of specimen (g).

The results of the general compositional analysis of the fermented composition produced by the fermentation of bamboo salt and the bean curd soybean were as follows.

General composition of fermented composition by interval Test Items A little bean Fermented dried beans Fermentation composition Fermentation composition
(Aged 6 months) Calories (kcal) 355.8 366.1 32.1 34.0 Carbohydrate (g / 100 g) 40.3 34.5 6.8 6.4 Sugar (g / 100g) 2.2 0.3 0.3 1.4 Protein (g / 100 g) 36.3 38.7 0.9 1.9 Fat (g / 100g) 5.5 8.2 0.2 0.1 Saturated fat (g / 100g) 0.5 0.7 0.0 0.0 Trans fat (g / 100g) 0.0 0.0 0.0 0.0 Cholesterol (mg / 100g) 0.0 0.0 0.0 0.0 Sodium (mg / 100g) 8.8 14.2 6483.5 8648.8 Water (g / 100g) 13.0 14.6 72.5 72.1 Ash (g / 100g) 4.9 4.1 19.7 19.5

< Example  3> Bamboo salt A little bean  Physicochemical experiment of fermented composition produced by fermentation

<3-1> Bamboo salt pH  And ORP  Measure

10 g of bamboo salt was added to 100 ml of distilled water, and the mixture was mixed well with a stirrer for 10 minutes or more in order to reduce the error range. The ORP and the pH value were recorded when the ORP value was the most stable.

As a result, the pH of bamboo salt was 10.5 and the ORP was -400 mV.

&Lt; 3-2 > pH  And ORP  Measure

A sample of the fermented composition produced by fermentation of bamboo salt and bean curd beans, a 6-month aged sample, and a 3-year aged sample were prepared, and ORP and pH values were recorded when the ORP value was the most stable.

As a result, the pH of the immersed sample of the fermented composition was 6.67 and the ORP was 12.3 mV. The pH of the aged sample for 6 months was 5.74 and the ORP was 65.6 mV. The pH of the fermented composition aged 3 years was 5.25 ORP was 93.8 mV.

<3-3> Salinity Measurement of Fermentation Composition

Salinity was measured using a salinity meter, taking into consideration the temperature - dependent errors of the fermented composition sample, six - month aged sample, and three - year aged sample produced by the fermentation of bamboo salt and rice husk.

As a result, the salinity of the sample immersed in the fermented composition was 22.6%, the salinity of the fermented composition aged 6 months was 35%, and the salinity of the fermented composition aged 3 years was 40%.

< Example  4> Bamboo salt A little bean  Of the fermentation composition produced by fermentation In vitro In vitro ) In the model  Functional Analysis

<4-1> Preparation of sample

Each sample was classified as follows, and all of these samples were supplied through a phosphate buffered saline solution.

A: Extracted by adding 50.5 mL of water to 18.52 g of fermented twigs;

B: fermented composition prepared by using 20 kg of sun dried salt and 54 L of water in 20 kg of fermented twigs;

C: fermented composition prepared by using 20 kg of phosphoric acid bamboo salt and 54 L of water in 20 kg of fermented twigs;

D: A fermented composition prepared by hot water extraction of 20 kg of fermented dried bean curds, 20 kg of phosphoric acid bamboo salt, 1.05 kg of yuukeeji, 1.4 kg of field garlic and 2.6 kg of sulfur duck, in 60 L of water.

<4-2> Analysis of Total Phenol Content

The total phenol content was determined by taking 1 mL of the sample, adding 1 mL of 2% (w / v) Na ₂ CO ₃ solution and allowing to stand for 3 minutes, and then adding 0.2 mL of 50% Folin-Ciocalteu reagent And allowed to stand at room temperature for several minutes. The mixture was centrifuged at 13,400 × g for 10 minutes, and 1 mL of the supernatant was taken and absorbance was measured at 750 nm using a spectrophotometer (Shimadzu UV-1601, Kyoto, Japan). The total phenolic content was expressed in mg / g GAE (gallic acid equivalents) as a standard curve using gallic acid.

As a result, Composition B prepared by adding sun salt to chicken legs showed the highest content, followed by Composition C prepared by adding bamboo salt, followed by rice bran self fermentation A, bamboo salt and field garlic, sulfur duck, Showed the same level of D (FIG. 6).

<4-3> DPPH Radical  Analysis of scavenging activity

The DPPH radical scavenging activity was measured by adding 0.8 mL of 0.04 mM DPPH solution to 0.2 mL of the sample, reacting at room temperature for 30 minutes, and measuring the absorbance at 517 nm. The DPPH radical scavenging activity of each sample was calculated by the following formula and expressed as a percentage.

DPPH radical scavenging activity = (1-B / A) x 100

A: Absorbance of untreated water; And

B: Absorbance of sample.

As a result, the activity of each composition was increased in a concentration-dependent manner. The IC ₅₀ value indicating the concentration of the sample inhibiting 50% of the radical was calculated. As a result, in the case of the composition B, the DPPH radical scavenging ability was 7.5 mg D was 11.2 mg, C was 11.8 mg, and A was 15.7 mg in that order.

DPPH radical scavenging activity and IC ₅₀ μg / mL A ¹⁾ B C D 0.1 19.2 ± 0.7 ^{2) a3 )} 18.3 ± 1.6 ^a 23.5 ± 1.2 ^a 15.5 ± 0.5 ^a 0.5 22.0 ± 0.3 ^b 18.1 ± 1.9 ^a 21.1 ± 0.6 ^a 16.9 ± 0.8 ^a One 23.7 ± 0.5 ^c 22.5 ± 1.3 ^b 34.3 ± 0.3 ^b 31.1 ± 2.3 ^b 10 39.7 ± 0.6 ^d 61.3 ± 1.0 ^c 81.0 ± 1.5 ^c 57.7 ± 0.9 ^c 20 58.0 ± 0.4 ^e 65.6 ± 1.0 ^d 86.5 ± 3.1 ^d 72.1 ± 0.9 ^d IC ₅₀ ^{4 )} (mg / mL) 15.7 + - 0.2 ^{D3 )} 7.5 ± 0.3 ^A 11.8 ± 0.0 ^C 11.2 ± 0.3 ^B

¹⁾ A: Fermented chicken legs; B: Fermented chicken legs + sun salt; C: Fermented chicken legs + phosphoric acid bamboo salt; And D: fermented wheat bean + phosphoric acid bamboo salt + yuga pea + field garlic + sulfur duck.

²⁾ Data are mean ± standard deviation (n = 3);

³⁾ Significant differences at P <0.05 are indicated by different subscripts (ad, AD) in the column; And

4) IC ₅₀ (mg / mL) is the concentration of mg / mL required for 50% removal of the radical.

<4-4> ORAC  Analysis of activity

The antioxidative activity of the sample against the change of the fluorescence reduction rate due to the formation and disappearance of the peroxy radical was measured by the peroxyl radical scavenging capacity (ORACROO) assay. For the generation of peroxidation radicals, 5 mM 2, 2'-azobis (2-methylpropionamidine) dihydrochloride (AAPH) was used and 40 nM fluorescent standard solution was prepared and applied to GENios fluorescence plate reader (Tecan, Salzburg, Austria) An excitation wavelength of 485 nm, and an emission wavelength of 535 nm. The results were compared with the curve area protected by 1 μM Trolox (6-hydroxy-2,5,7,8-tetramethylchroman-2-carbonyl acid) g).

As a result, the concentration-dependent increase in activity tended to be observed in all four samples. At the concentrations of 1, 5, and 10 μg / mL, the four samples exhibited similar activity, and the highest activity was observed at the highest concentration of 50 μg / mL (13.3 in the C sample) 6.9 and 6.6, and B to 5.9 TE (FIG. 7).

<4-5> CAC ( Cellular antioxidant capacity ) Analysis of activity

The CAC assay for measuring the antioxidative activity of samples in living cells using HepG2 cells was determined by modifying the DCFH-DA assay. In a 96-well plate, 5 × 10 4 cells / mL of HepG2 was incubated at 37 ° C in a 5% CO ₂ incubator with 10% fetal bovine serum (FBS), 1% penicillin / streptomycin penicillin / streptomycin) in Dulbecco's modified Eagle's medium (DMEM). After 24 hours, the medium was removed and 200 μL of fluorescence-stable Hank's balanced salt solution (HBSS) was dispensed into each well. Each sample was treated with a final concentration of 100 μg / mL and incubated for 30 minutes. After incubation, the cells were washed with HBSS, and 2 μL of 80 mM AAPH was added to 200 μL of HBSS for peroxyl radical generation, followed by incubation for 30 minutes. After incubation for 30 min with excitation of light, 40 μM DCFH-DA was added to the fluorescent probe. The excitation wavelength was adjusted to 485 nm using a GENios fluorescence plate reader (Tecan Trading AG, Salzburg, Austria) Emission wavelength The degree of fluorescence was measured at 535 nm. AAPH and DCFH-DA were treated for positive control and 100% for control, respectively, and the relative fluorescence values of AAPH and AAPH were compared.

As a result, when the activity of the negative control without AAPH was 100%, the positive control treated with AAPH alone was not treated with 150% of the sample, High CAC activity. There was no significant difference between the concentrations of each sample and all concentrations were at the same level as NC (FIG. 8).

<4-6> DNA  damaged Inhibitory  analysis

5 mL of fresh whole blood was separated from leukocytes using histopaque 1077 and used in this experiment. The prepared leukocyte cells were treated with a sample and reacted at 37 ° C for 30 minutes. Then, the leukocytes were washed with PBS and 200 μM H2O2 was reacted at 4 ° C for 5 minutes to induce oxidative stress artificially. After the reaction, the cells were washed with PBS. Positive control was treated with 1% DMSO instead of sample and 200 μM H2O2. For the Comet assay, the reaction was completed by mixing the reacted leukocytes with 75 μL of 0.7% LMA. The suspension was then spread over a precoated slide with 1.0% NMA and covered with a cover glass And stored in a refrigerator at 4 ° C. When the gel was hardened, the cover glass was peeled off and a layer of 75 μL of a 0.7% LMA solution was further coated thereon. To the cold alkali lysis buffer (2.5 M NaCl, 100 mM Na 2 EDTA, 10 mM tris) After mixing X-100, the slide was immersed in a dark room at room temperature for 1 hour to loosen the double strand of DNA. After lysis, the slides were placed in an electrophoresis bath and filled with cold electrophoresis buffer (300 mM NaOH, 10 mM Na 2 EDTA, pH> 13) at 4 ° C and unwound for 20 minutes to reveal alkaline labile sites of DNA Then, a voltage of 25 V / 300 3 mA was applied and electrophoresis was performed for 20 minutes. In order to prevent DNA from being damaged by light, the above procedure was carried out in dark dark room conditions. After the electrophoresis was completed, washing was repeated 3 times in 0.4 M Tris buffer (pH 7.4) And the nuclei were stained with ethidium bromide at a concentration of 20 μL / mL, covered with a cover slip, and observed under a fluorescence microscope (Leica, Wetzlar, Germany). Each nucleus image sent through a CCD camera (Nikon, Tokyo, Japan) was analyzed using a computer equipped with a Komet 5.0 comet image analyzing system (Kinetic Imaging, Liverpool, UK). DNA damage caused by hydrogen peroxide in the leukocyte and inhibition of damage by the thyroid extract was calculated as 100% by measuring the DNA content in the tail portion which migrated from the nucleus and fell off to the tail portion. Two slides were made in each treatment, and DNA damage of each of 100 cells was measured. Each treatment was repeated twice.

As a result, in the case of soybean fermented, the inhibition effect of DNA damage was not detected at 95.5% at 1 μg / mL concentration but 68.6, 66.1 and 44.4% at the other concentrations, respectively. , 76.4, 52.4, and 48.5%, respectively, for C with Bamboo salt, and 71.4, 62.5, 53.0, and 40.0% with B, respectively. In the case of Sample D, which was made by adding bamboo salt, , 72.7, 55.8, and 53.2%, respectively, indicating that they inhibit DNA damage at all concentrations (FIG. 9).

< Example  5> Bamboo salt A little bean  Verification of fatigue recovery effect of fermentation composition produced by fermentation

<5-1> Preparation and breeding of experimental animals

Six - week old SD rats were used for one - week adaptive training. To adapt to water, swimming adaptation exercise was performed at 30-33 ℃ for 10 minutes. In this experiment, five times a week (20 minutes / once) was carried out and the animals were bred for 2 weeks. The temperature of water was maintained at 30 ℃ and the exercise was carried out for a certain period of time in order to make the experimental conditions of all groups the same.

The experimental diets were divided into two groups according to the control group, the negative control group (exercise group + physiological saline), the fermentation composition administration group (exercise + low concentration fermentation composition, exercise + high concentration fermentation composition) Animals were sacrificed and blood and liver were collected and used for analysis.

The composition of the experimental diet NC ^{1 )} PC 0.5% 5% Casein 20 20 20 20 Corn starch 53 53 53 53 Sugar 10 10 10 10 Corn oil 7 7 7 7 cellulose 5 5 5 5 Vitamin mixture ²⁾ One One One One Mineral mixture ³⁾ 3.5 3.5 3.5 3.5 Choline bitartarate 0.3 0.3 0.3 0.3 L-cystein 0.3 0.3 0.3 0.3 t-butylhydroquinone (t-butylhydroquinone) 0.014 0.014 0.014 0.014 BHT 0.001 0.001 0.001 0.001 Sample 0 0 0.5 5

¹⁾ NC: negative control group, PC: exercise induced fatigue, 0.5%: exercise induced fatigue + 0.5%

Fermented composition (sample) of fermented peanut, bamboo salt, rhizome, field garlic and sulfur duck, 5%: exercise induced fatigue + 5% sample.

²⁾ AIN 93 vitamin mixtures include (in g / kg of mixture): thiamine HCl 0.6; Riboflavin 0.6; Pyridoxine HCl 0.7; Niacin 3) d-calcium pantothenate 1.6; Folic acid 0.2; 0.0 &gt; d-biotin &lt; / RTI &gt;0.02; Cyanocobalamin (vitamin B ₁₂ ) 2.5; Dry vitamin A palmitate (500,000 U / d) 0.8; Dry vitamin E acetate (500 U / d) 15; Vitamin D ₃ Baesan _{(vitamin D 3 trituration) (400,000} U / g) 0.25; Vitamin K1 0.075 Menadione sodium bisulfite complex 0.15; Sucrose finely powdered 981.08.

³⁾ AIN 93 The mineral mixture includes the following (in g / kg of mixture): calcium phosphate, dibasic 500; Sodium chloride 74; Potassium citrate, monohydrate 220; Potassium sulfate 52; Magnesium oxide 24; Manganous carbonate (43-48% Mn) 3.5; Ferric citrate (16-17% Fe) 6; Zinc carbonate (70% ZnO) 1.6; Cupric carbonate (53-55% Cu) 0.3; Potassium iodate 0.01; Sodium selenite 0.01; Chromium potassium sulfate 0.55; Sucrose (finely powdered) 118.03.

The animals were fasted for 12 hours before sacrifice, and then anesthetized with CO ₂ from dry ice. Blood was collected from the abdominal aorta and coagulated with heparin. At the time of sacrifice, the remaining blood was analyzed by centrifugation at 3000 rpm for 30 minutes. Plasma and erythrocytes were separated from each other and stored frozen at -70 ° C until analysis. The organs of the experimental animals were extracted and washed with physiological saline, the water was removed from the absorbent paper and the weight was measured. The liver was rapidly frozen in liquid nitrogen and stored at -70 ° C until analysis.

<5-2> Weight gain, Dietary intake  And checking diet efficiency

Weight gain and dietary intake during the experimental period. There was no significant difference between the groups (Table 7).

Exercise-Induced Fatigue Model Effect of Sample Supply on Weight Gain, Dietary Intake, and FER in SD Rats Weight (g) Dietary intake (g) FER ^{2 )} (%) NC ^{1 )} 104.4 ± 2.6 ^{3) ns 4 )} 17.4 ± 0.3 ^ns 0.17 ± 0.00 ^{a5 )} PC 98.0 ± 3.6 17.3 ± 0.2 0.18 ± 0.01 ^ab 0.5% 93.7 ± 2.9 17.3 ± 0.4 0.19 ± 0.00 ^b 5% 98.2 ± 5.5 17.1 ± 0.5 0.17 ± 0.01 ^ab

¹⁾ NC: negative control group, PC: exercise induced fatigue, 0.5%: exercise induced fatigue + 0.5%

Fermented composition (sample) of fermented peanut, bamboo salt, rhizome, field garlic and sulfur duck, 5%: exercise induced fatigue + 5% sample.

²⁾ FER: Food efficiency rate.

³⁾ Mean ± SE (Mean ± SE).

⁴⁾ ns: Not significant.

⁵⁾ Numbers with various subscripts in the column are significant at p <0.05.

<5-3> Identification of organ weights of experimental animals

During the experimental period, weights of liver, kidney, and heart were not different between groups, but PC showed a significant decrease in spleen weight compared to NC. However, there was no significant difference between PC and low-concentration and high-concentration sari-intake groups (Table 8).

Exercise-induced fatigue model Effect of sample supply on liver, heart, spleen and kidney in SD rats liver kidney Heart spleen NC ^{1 )} 2.87 ± 0.07 ^{2) ns 3 )} 0.71 ± 0.03 ^ns 0.39 + 0.01 ^ns 0.25 + 0.01 ^b PC 2.87 ± 0.04 0.71 + 0.02 0.43 + 0.02 0.22 + 0.01 ^a 0.5% 2.86 ± 0.03 0.74 ± 0.01 0.43 + - 0.01 0.23 ± 0.01 ^ab 5% 2.72 + 0.07 0.70 + 0.02 0.44 + 0.02 0.23 ± 0.01 ^ab

¹⁾ NC: negative control group, PC: exercise induced fatigue, 0.5%: exercise induced fatigue + 0.5%

Fermented composition (sample) of fermented peanut, bamboo salt, rhizome, field garlic and sulfur duck, 5%: exercise induced fatigue + 5% sample.

²⁾ Mean ± SE (Mean ± SE).

³⁾ ns: Not significant.

⁴⁾ Numbers with various subscripts in the column are significant at p <0.05.

<5-4> Antioxidant activity and lipid peroxidation in blood of experimental animals

The total antioxidant potential (TRAP) was measured as follows. Specifically, peryl myoglobin produced by activating ABTS (2,2-azinobis (3-ethylbenzothiazo-line 6-sulfonate), 150 mM) and metmyoglobin (25 mM) with H ₂ O ₂ Is a method for measuring the degree of inhibition of absorption of an ABTS radical cation formed by interaction with a radical species (ferryl myoglobin radical species). This is proportional to the antioxidant capacity contained in the sample (0.84% plasma). The samples were incubated for 8 min at 30 ° C. and then absorbance was measured at a wavelength of 740 nm using a UV / VIS spectrophotometer. Plasma TRAP concentrations were calculated using the calibration curve of trolox and expressed as TEAC (Trolox equivalent antioxidant capacity, mM).

The plasma lipid peroxide concentration was measured as follows. Specifically, plasma was treated with 0.064 M trisodium citrate and heparin, centrifuged, and LDL-cholesterol was isolated. To extract lipids from LDL-cholesterol, chloroform-methanol (chloroform-methanol) was treated with cyclohexane and the absorbance was measured at 234 nm.

As a result, TRAP showed a significantly lower activity in PC than NC, and it was found to be improved to the level of NC by ingesting the low concentration fermentation composition. CD showed higher activity in PC than in NC, which was decreased by ingesting high-concentration fermentation composition (Table 9).

Exercise-induced fatigue model Effect of sample supply on TRAP and CD in SD rats TRAP (mM) CD (μM) NC ^{1 )} 1.18 ± 0.02 ^{2) b3 )} 5.2 ± 0.3 ^{ns4 )} PC 1.07 0.02 ^a 5.9 ± 0.3 0.5% 1.18 ± 0.02 ^b 5.9 ± 0.2 5% 1.05 + 0.01 ^a 5.7 ± 0.2

¹⁾ NC: negative control group, PC: exercise induced fatigue, 0.5%: exercise induced fatigue + 0.5%

Fermented composition (sample) of fermented peanut, bamboo salt, rhizome, field garlic and sulfur duck, 5%: exercise induced fatigue + 5% sample.

²⁾ Mean ± SE (Mean ± SE).

³⁾ Numbers with various subscripts in the column are significant at p <0.05.

⁴⁾ ns: Not significant.

<5-5> Analysis of antioxidant enzyme in blood of experimental animals

The principle of catalase enzyme activity is catalyzed by the decomposition of hydrogen peroxide into water and oxygen to measure the amount of hydrogen peroxide reduction. The reaction rate is proportional to the concentration of hydrogen peroxide. If the concentration of hydrogen peroxide is more than 0.1 M, the reaction by catalase is inhibited. Therefore, a small concentration of about 0.01 M should be used. For the reaction to proceed well, 50 mM phosphate buffer (pH 7) and hydrogen peroxide were added to the red blood cells at the latest 30 seconds after the start of the reaction, and the amount of hydrogen peroxide reduction was measured at 240 nm for 30 seconds.

As a result, catalase activity was significantly decreased in PC compared with NC, and increased in low concentration group (Table 10).

Exercise-induced fatigue model Effect of sample supply on catalase in SD rats Catalase (K / g Hb) NC ^{1 )} 1169.2 ± 60.2 ^{2) bc3 )} PC 829.9 + - 40.0 ^a 0.5% 1006.5 ± 24.6 ^bp 5% 1305.2 + - 120.6 ^c

¹⁾ NC: negative control group, PC: exercise induced fatigue, 0.5%: exercise induced fatigue + 0.5%

Fermented composition (sample) of fermented peanut, bamboo salt, rhizome, field garlic and sulfur duck, 5%: exercise induced fatigue + 5% sample.

²⁾ Mean ± SE (Mean ± SE).

³⁾ Numbers with various subscripts in the column are significant at p <0.05.

⁴⁾ ns: Not significant.

&Lt; 5-6 > DNA  Check for damage

20 μL of whole blood was collected and mixed with 75 μL of 0.7% 0.5% low melting agarose gel (LMA), and then 0.5% normal melting agarose (NMA) was precoated A fully frosted slide over the whole blood and LMA suspension was spread over the cover glass and then stored in a refrigerator at 4 ° C. When the gel was hardened, the cover gum was peeled off and another layer was covered with 75 μL of a 0.7% LMA solution. Add 1% Triton X-100 immediately before use to cold lysis buffer (2.5M NaCl, 100 mM EDTA, 10 mM tris) prepared beforehand for cell lysis. For 1 hour to release the double strand of DNA. The lysed slides were placed in an electrophoresis tank and filled with cold buffer (300 mM NaOH, 10 mM Na2EDTA) at 4 ° C. The blood was unwound for 20 minutes and then applied with a voltage of 25 V / 300 ± 3 mA And electrophoresis was carried out for 20 minutes. After electrophoresis, the cells were washed thoroughly with 0.4 M Tris buffer (pH 7.4), stained with ethidium bromide at a concentration of 20 μL / mL, observed with a fluorescence microscope, Was analyzed with a computer equipped with a comet image analyzing system. The effect of leukocyte and H ₂ O ₂ -induced leukocyte DNA damage on continuous exercise and ingestion of the fermentation composition was examined.

As a result, tail DNA, tail torque, and tail length indicating the degree of DNA damage of leukocytes were partially increased in PC compared to NC, and a tendency to decrease in low and high concentration groups was shown (Table 11).

Exercise-induced fatigue model The effect of sample supply on leukocyte DNA damage in SD rats NC1) PC 0.5% 5% Untreated DNA damage Tail DNA (%) 7.5 ± 0.3 ^{2) ns 3 )} 7.4 ± 0.2 8.4 ± 0.6 8.5 ± 0.7 Tail moment 1.6 ± 0.1 ^ns 1.7 ± 0.1 2.1 ± 0.3 2.1 ± 0.3 Tail length (μm) 12.6 ± 0.4 ^ns 12.6 ± 0.4 14.4 ± 1.4 13.7 ± 1.0 DNA damage induced by 200 μM H ₂ O ₂ Tail DNA (%) 16.3 ± 1.1 ^ns 16.8 ± 1.2 16.0 ± 1.3 15.7 ± 0.8 Tail moment 9.0 ± 1.1 ^ns 10.3 ± 1.4 9.2 ± 1.4 8.2 ± 0.6 Tail length (μm) 33.6 ± 2.6 ^ns 35.1 ± 2.9 33.1 ± 3.7 31.2 ± 1.5

¹⁾ NC: negative control group, PC: exercise induced fatigue, 0.5%: exercise induced fatigue + 0.5%

Fermented composition (sample) of fermented peanut, bamboo salt, rhizome, field garlic and sulfur duck, 5%: exercise induced fatigue + 5% sample.

²⁾ Mean ± SE (Mean ± SE).

³⁾ ns: Not significant.

< Example  6> Bamboo salt and A little bean  Verification of hangover elimination effect of fermented composition produced by fermentation

<6-1> Preparation and breeding of experimental animals

Six weeks old male Sprague-Dawley (SD) was adapted for 2 weeks. The experimental animals were divided into 4 groups (normal control, alcohol consumption group, alcohol + low concentration fermentation composition intake group, alcohol + high concentration fermentation composition intake group) The experimental diet was given for 5 weeks. The experimental diet consumed an amount of calories not contained in ethanol and 25% of the total calories was fed into ethanol. The ethanol supply gradually increased to 5, 10, 15, 20, and 25%, so that sufficient adaptation was made. Five weeks after the empirical formula was given, animals were sacrificed and blood and liver were collected and used for analysis

The composition of the experimental diet NC ^{1 )} PC 0.5% 5% Casein 18 18 20 20 Corn starch 50.5 15.5 53 53 Sugar 10 10 10 10 Soybean oil 10 10 7 7 cellulose 6.5 6.5 5 5 Vitamin mixture ²⁾ One One One One Mineral mixture ³⁾ 3.5 3.5 3.5 3.5 Choline bitartarate 0.2 0.2 0.3 0.3 L-cystein 0.3 0.3 0.3 0.3 BHT 0.001 0.001 0.014 0.014 Alcohol 0 25 25 25 Sample 0 0 0.5 5

PC: 25% alcohol supply, 0.5%: 25% alcohol supply + 0.5% sample, 5%: 25% alcohol supply + 5% sample ^, NC: negative control (without alcohol)

²⁾ AIN 93 vitamin mixtures include (in g / kg of mixture): thiamine HCl 0.6; Riboflavin 0.6; Pyridoxine HCl 0.7; Niacin 3) d-calcium pantothenate 1.6; Folic acid 0.2; 0.0 &gt; d-biotin &lt; / RTI &gt;0.02; Cyanocobalamin (vitamin B ₁₂ ) 2.5; Dry vitamin A palmitate (500,000 U / d) 0.8; Dry vitamin E acetate (500 U / d) 15; Vitamin D ₃ Baesan _{(vitamin D 3 trituration) (400,000} U / g) 0.25; Vitamin K1 0.075 Menadione sodium bisulfite complex 0.15; Sucrose finely powdered 981.08.

³⁾ AIN 93 The mineral mixture includes the following (in g / kg of mixture): calcium phosphate, dibasic 500; Sodium chloride 74; Potassium citrate, monohydrate 220; Potassium sulfate 52; Magnesium oxide 24; Manganous carbonate (43-48% Mn) 3.5; Ferric citrate (16-17% Fe) 6; Zinc carbonate (70% ZnO) 1.6; Cupric carbonate (53-55% Cu) 0.3; Potassium iodate 0.01; Sodium selenite 0.01; Chromium potassium sulfate 0.55; Sucrose (finely powdered) 118.03.

The animals were fasted for 12 hours before sacrifice and blood was drawn from the abdominal aorta by anesthesia using CO ₂ generated from dry ice to prevent clotting with heparin. After the Comet assay, the blood was centrifuged at 3000 rpm for 30 minutes, and plasma and erythrocytes were separated and stored at -70 ° C until analysis. After removing the water from the absorbent paper and measuring the weight, 0.5 g of the liver was extracted and immediately used by immersing it in HBSS buffer for analysis of comet. The rest of the animals were immediately frozen with liquid nitrogen And stored frozen at -70 ° C until analysis.

<6-2> Weight gain, Dietary intake  And checking diet efficiency

Weight gain and dietary intake during the experimental period. There was no significant difference between the groups (Table 13).

Effect of Sample Supply on Weight Gain, Dietary Intake, and FER in 25% Alcohol-Fed Rats Weight (g) Dietary intake (g) FER ^{2 )} (%) NC ^{1 )} 101.1 ± 2.8 ^{3) ab4 )} 14.5 ± 0.1 ^a 0.14 ± 0.00 ^ab PC 106.0 ± 7.1 ^ab 16.5 ± 0.2 ^b 0.16 ± 0.01 ^ab 0.5% 127.0 ± 7.9 ^b 16.5 ± 0.2 ^b 0.13 + 0.01 ^a 5% 84.7 ± 12.8 ^a 16.4 ± 0.7 ^b 0.23 ± 0.05 ^b

PC: 25% alcohol supply, 0.5%: 25% alcohol supply + 0.5% sample, 5%: 25% alcohol supply + 5% sample ^, NC: negative control (without alcohol)

²⁾ FER: Food efficiency rate.

³⁾ Mean ± SE (Mean ± SE).

⁴⁾ Numbers with various subscripts in the column are significant at p <0.05.

<6-3> Identification of organ weights of experimental animals

Long-term weight measurements during the experiment did not affect liver, kidney, heart, and spleen weights (Table 14).

Effect of Sample Supply on Liver, Heart, Spleen and Kidney in 25% Alcohol Fed Rats liver kidney Heart spleen NC ^{1 )} 2.70 ± 0.05 ^{2) NS3 )} 0.69 ± 0.02 ^ns 0.38 ± 0.01 ^ns 0.21 ± 0.01 ^ns PC 2.78 ± 0.05 0.77 + 0.03 0.38 ± 0.01 0.22 ± 0.01 0.5% 2.70 ± 0.03 0.71 + - 0.03 0.37 ± 0.01 0.22 ± 0.01 5% 2.82 + 0.15 0.79 + 0.04 0.37 ± 0.01 0.21 ± 0.01

PC: 25% alcohol supply, 0.5%: 25% alcohol supply + 0.5% sample, 5%: 25% alcohol supply + 5% sample ^, NC: negative control (without alcohol)

²⁾ Mean ± SE (Mean ± SE).

³⁾ ns: Not significant.

<6-4> Identification of Liver Damage Index in Laboratory Animals

Serum aspartate aminotransferase (AST) and alanine aminotransferase (ALT) were measured using a kit (Asan Pharmaceutical, Korea) which was prepared by colorimetric assay using substrate and enzyme reaction using kit (Asan Pharmaceutical, Korea).

As a result, both AST and ALT enzymes were significantly higher in PC group than NC group, and it was confirmed that alcohol consumption for 5 weeks affected liver damage. In the low concentration group, the activity of the enzyme was decreased compared to PC (Table 15).

Effect of Sample Supply on AST and ALT in 25% Alcohol-Fed Rats AST (U / L) ALT (U / L) NC ^{1 )} 50.2 ± 1.6 ^a 22.5 ± 1.2 ^a PC 67.0 ± 3.2 ^b 39.7 ± 2.7 ^c 0.5% 55.3 ± 2.2 ^a 31.3 ± 2.2 ^b 5% 69.2 ± 3.6 ^b 42.8 ± 4.6 ^c

PC: 25% alcohol supply, 0.5%: 25% alcohol supply + 0.5% sample, 5%: 25% alcohol supply + 5% sample ^, NC: negative control (without alcohol)

²⁾ Mean ± SE (Mean ± SE).

³⁾ Numbers with various subscripts in the column are significant at p <0.05.

⁴⁾ ns: Not significant.

<6-5> Identification of erythrocyte antioxidant enzyme activity in experimental animals

Glutathione peroxidase (GSH-Px) catalyzes the oxidation of glutathione by t-butylhydroperoxide. At this time, the oxidized glutathione is reduced again to glutathione in the presence of glutathione reductase and NADPH, and NADPH is oxidized to NADP. Glutathione, glutathione reductase and NADPH were added to hemolyzed red blood cells, and the mixture was allowed to stand at 37 ° C for 10 minutes and then reacted with t-butylhydroperoxide. The reduced NADPH concentration was 340 nm for 90 seconds to measure the degree of antioxidation of GSH-Px (U / g HB).

As a result, GSH-Px activity was significantly lowered in PC than in NC, and activity of both low and high concentration groups was increased (Table 16).

Effect of Sample Supply on GSH-Px in 25% Alcohol-Fed Rats GSH-Px (U / g Hb) NC ^{1 )} 38.1 ± 3.7 ^b PC 18.5 ± 1.2 ^a 0.5% 32.3 ± 2.4 ^b 5% 42.6 + 4.8 ^b

PC: 25% alcohol supply, 0.5%: 25% alcohol supply + 0.5% sample, 5%: 25% alcohol supply + 5% sample ^, NC: negative control (without alcohol)

²⁾ Mean ± SE (Mean ± SE).

³⁾ Numbers with various subscripts in the column are significant at p <0.05.

< Manufacturing example  1> Preparation of pharmaceutical preparations

<1-1> Sanje  Produce

2 g of the composition of the present invention

Lactose 1 g

The above components were mixed and packed in airtight bags to prepare powders.

<1-2> Preparation of tablets

100 mg of the composition of the present invention

Corn starch 100 mg

100 mg of milk

Magnesium stearate 2 mg

After mixing the above components, tablets were prepared by tableting according to a conventional method for producing tablets.

&Lt; 1-3 > Preparation of capsules

100 mg of the composition of the present invention

Corn starch 100 mg

100 mg of milk

Magnesium stearate 2 mg

After mixing the above components, the capsules were filled in gelatin capsules according to the conventional preparation method of capsules.

&Lt; 1-4 >

1 g of the composition of the present invention

Lactose 1.5 g

Glycerin 1 g

0.5 g of xylitol

After mixing the above components, they were prepared so as to be 4 g per one ring according to a conventional method.

<1-5> Preparation of granules

150 mg of the composition of the present invention

Soybean extract 50 mg

Glucose 200 mg

Starch 600 mg

After mixing the above components, 100 mg of 30% ethanol was added and the mixture was dried at 60 ° C to form granules, which were filled in a capsule.

< Manufacturing example  2> Manufacture of health food

<2-1> Production of flour food

0.5-5.0 parts by weight of the composition of the present invention was added to wheat flour, and the mixture was used to prepare bread, cake, cookies, crackers and noodles.

<2-2> soup  And juicy ( gravies )

0.1 to 5.0 parts by weight of the composition of the present invention was added to soups and gravies to prepare meat products for health promotion, soup of noodles and juices.

<2-3> Ground Beef  Produce

10 parts by weight of the composition of the present invention was added to ground beef to prepare ground beef for health promotion.

<2-4> Dairy products ( dairy products )

5-10 parts by weight of the composition of the present invention was added to milk, and various dairy products such as butter and ice cream were prepared using the milk.

<2-5> Solar  Produce

Brown rice, barley, glutinous rice, and yulmu were dried by a known method and dried, and the mixture was granulated to a powder having a particle size of 60 mesh.

Black soybeans, black sesame seeds, and perilla seeds were steamed and dried by a conventional method, and then they were prepared into powder having a particle size of 60 mesh by a pulverizer.

The composition of the present invention was concentrated under reduced pressure in a vacuum concentrator, dried by spraying, and dried with a hot-air drier, and the resulting dried product was pulverized to a particle size of 60 mesh with a pulverizer to obtain a dried powder.

The grains, seeds, and the composition of the present invention prepared above were blended in the following proportions.

(30 parts by weight of brown rice, 15 parts by weight of yulmu, 20 parts by weight of barley)

Seeds (7 parts by weight of perilla, 8 parts by weight of black beans, 7 parts by weight of black sesame seeds)

The composition of the present invention (3 parts by weight)

(0.5 part by weight),

Rhubarb (0.5 parts by weight).

< Manufacturing example  3> Health drink  Produce

<3-1> Production of health drinks

5 g of the composition of the present invention was homogeneously mixed with a sub ingredient such as liquid fructose (0.5%), oligosaccharide (2%), sugar (2%), salt (0.5%) and water (75% It was packaged in small containers such as glass bottles and plastic bottles.

<3-2> Preparation of vegetable juice

5 g of the composition of the present invention was added to 1,000 tomato or carrot juices to prepare vegetable juice.

<3-3> Production of fruit juice

1 g of the composition of the present invention was added to 1,000 apples or grape juices to prepare fruit juice.

Claims (14)

Rye beans ( Rhynchosia Nulubilis ), bamboo salt, rhizome , field garlic and sulfur duck.
The fermentation composition according to any one of claims 1 to 3, wherein the dry husk comprises soybean: bamboo salt: yuukee: field garlic: sulfur duck in a weight ratio of 1: 1: 0.5 to 1: 0.5 to 1: 0.5 to 1.
2. The fermentation composition according to claim 1, wherein the soybean bean is a fermented soybean fermented first.
The fermentation composition according to claim 1, wherein the bamboo salt is 9-fold bamboo salt.
1) washing and growing the bean sole and cooling;
2) Inoculating the yeast into the legs of the step 1), then fermenting the yeast, and then drying it;
3) mixing the bamboo shoots, yucci, field garlic, and sulfur ducks with dried legs of step 2);
4) first aging the mixture of step 3), filtering and then heating;
5) secondary aging the fermented product of step 4)
A method for producing a fermented composition comprising a legged bean, a bamboo salt, a rhizome, a field garlic, and a sulfur duck.
The fermented composition according to claim 5, wherein the fermentation of step 2) is performed at 32 to 38 ° C for 15 to 20 hours.
6. The method according to claim 5, wherein the drying in step 2) is performed at 30 to 40 DEG C for 12 to 15 hours.
[Claim 6] The method according to claim 5, wherein the yugi, field, and duck ducks of step 3) are extracted with hot water and then extracted with purified water.
6. The fermented composition according to claim 5, wherein the mixing of step 3) is carried out in a weight ratio of 1: 1: 1: 0.7: 0.8 in the ratio of bean, bamboo salt, .
The fermented composition according to claim 5, wherein the heating in step 4) is performed at 80 to 100 ° C for 30 minutes to 1 hour.
6. A pharmaceutical composition for chewing a fatigue containing the fermented composition according to any one of claims 1 to 4.
A pharmaceutical composition for hangover use containing the fermentation composition according to any one of claims 1 to 4.
A health food for eating a fatigue containing the fermented composition of any one of claims 1 to 4.
A health food for hangover habit containing the fermented composition according to any one of claims 1 to 4.

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