WO2017138785A1 - Method for preparing germinated soybeans - Google Patents

Abstract

The present invention relates to a method for preparing germinated soybeans with enhanced nutrients, comprising an immersion step of immersing soybeans in an immersion liquid and a germination step of treating the soybeans that has passed through the immersion step with a germinating liquid, wherein the immersion liquid comprises a soybeans germination promoting agent. The germinated soybeans prepared according to the above preparation method have a high isoflavone content or free amino acid content.

Description

Method of manufacturing germinated beans

The present specification relates to a method for preparing a germinated bean enhanced nutrition.

Soy is an excellent natural product with many nutritional values. Previously, there were techniques for improving the quality of soybean sprouts such as the growth of soybean sprouts and the prevention of microbial contamination.

In addition, there has been a technique for increasing the growth of beans by administering a growth accelerator.

However, growth growth and nutrient change are different aspects, and when the growth of the plant is promoted, the nutrient content in the plant is exhausted, which lowers the nutritional value or functional value of the plant, that is, the beans.

Accordingly, the inventors of the present invention have studied a method for producing soybean germination, and have invented a method for producing germinated soybean having excellent nutritional or functional properties.

Prior art literature

(Patent Document 1) Japanese Laid-Open Patent No. 1994-172119

(Patent Document 2) Japanese Patent Application Laid-Open No. 1998-276578

One aspect of the present invention, a method for producing a nutrient-enhanced germinated soybean, comprising a soaking step of immersing soybeans in immersion liquid and a germination step of treating the germination liquid to the soaked soybeans, the immersion liquid To provide a method for producing a germinated bean characterized in that it comprises a soybean germination promoter.

One aspect of the present invention, a method for producing a nutrient-enhanced germinated soybean, comprising a soaking step of immersing soybeans in immersion liquid and a germination step of treating the germination liquid to the soaked soybeans, the immersion liquid The soybean germination provides a method for producing a germinated bean, characterized in that it comprises a promoter.

In one aspect of the invention, the immersion step is immersed for 5 to 24 hours, the germination step provides a method for producing germinated beans, which germinate 12 hours or more.

In one aspect of the present invention, the germination in the germination step is replaced with a new germination solution after 20 to 28 hours, provides a method for producing a germinated bean.

In one aspect of the present invention, the immersion step is immersed for 5 to 24 hours, the germination step provides a method for producing germinated beans, the germination for 36 to 60 hours.

In one aspect of the present invention, the germination step provides a method for producing germinated beans, which germinate under light or dark germination conditions.

In one aspect of the invention, the immersion liquid and the germination liquid comprises water, provides a method for producing germinated beans.

In one aspect of the present invention, the immersion liquid provides a germination bean containing 50% by volume to 100% by volume based on the total soaking promoter.

In one aspect of the present invention, the soybean germination promoter comprises a citrus water or a processed product thereof, the citrus water is a liquid obtained from the immature fruit of the plant citrus (Citrus) plants, provides a method for producing germinated beans.

In one aspect of the invention, the nutritional component provides any one or more of isoflavones, free amino acids and dipeptides, the production of germinated beans.

In one aspect of the invention, the isoflavone is dizin (Daidzin), 6 "-O-acetyldizin (6" -O-Acetyldaidzin), 6 "-O-malonyldiazine (6" -O-Malonyldaidzin) , Glycitin, 6 "-O-Acetylglycitin, 6" -O-malonylglycitin, 6 "-O-Malonylglycitin, Genistin, 6 "-O-Acetylgenistin, 6" -O-malonylgenistin, Daidzein, Glycitein, and Genistein Any one or more of

The free amino acid is Serine, Glutamine, Glycine, Glycine, Aspartic acid, Glutamic acid, Threonine, Alanine, Gammaaminobutyric acid any one or more of -aminobutyric acid, Theanine, Tyrosine, Methionine, Methionine, Valine, Leucine, Isoleucine, Phenylalanine and Trytophan Including;

The dipeptides are isoleucine-valine (Ile-Val), valine-leucine (Val-Leu), serine-tryptophan (Ser-Trp), isoleucine-leucine (Ile-Leu), isoleucine-phenylalanine (Ile-Phe ) And phenylalanine-leucine (Phe-Leu).

In one aspect of the invention, the free amino acid may include any one or more of Valine, Leucine, and Isoleucine.

Method according to one aspect of the present invention has the effect of eliciting changes in proteins, peptides, amino acids which are the core nutrients of soybean.

The method according to an aspect of the present invention has a superior effect of soybean germination under both dark and bright conditions when the germinated bean promoter is applied to soybean germination.

The method according to an aspect of the present invention can shorten the germination time compared to natural germinated beans.

The method according to one aspect of the present invention has a higher effect of the content of free amino acids of germinated beans than natural germinated beans.

Beans germinated by the method according to an aspect of the present invention is superior to natural germinated beans content of BCAA (leucine, isoleucine, valine) which is an amino acid that is beneficial to musculoskeletal health.

Soybean germinated by the method according to an aspect of the present invention has an excellent content of phenylalanine, an amino acid that is beneficial to musculoskeletal health, compared to natural germinated soybeans.

Soybean germinated by the method according to an aspect of the present invention is superior in content of isoflavones compared to natural germinated beans.

1 to 2 are photographs confirming the state of the germinated beans according to Example 1 and Comparative Examples 1 to 3 of the present invention.

The present invention is a technology for controlling the speed and direction of the nutrient conversion of the organism itself is formed based on the germination of the bean itself when the germination of beans, specifically, changes in proteins, peptides, amino acids or isoflavones that are core nutrients of soybeans It is about germination method that leads to.

Normally, germination of soybean proceeds in two stages of immersion-wetting germination and is carried out under dark conditions in which light is blocked for cultivation of bean sprouts.

The time to grow as bean sprouts usually takes 72 ~ 96 hours, including soaking time, which induces physiological changes of the beans and accelerates the change of nutrients in the beans. After about 100 hours, the nutrients accumulated for growth and the formation of the leaves after cotyledons are converted into cell walls. After overgrowth, most of the nutrients useful to the human body disappear.

However, when soybean sprouts naturally, significant nutrient change occurs about 72 hours. To do this, it grows in a single layer without stacking soybeans and requires a large area of cultivation space.

In addition, natural germination redistributes the nutrients in a favorable direction, so if the human body needs nutrient conversion that requires special purpose, it is disadvantageous in terms of yield, time and cost by additional processes such as extraction and purification.

In the present invention, a germination promoter is used as a substance for inducing change, increase, and promotion of nutrients in soybeans, and the germination promoter is added during soaking of soy to increase the content of free amino acids of soybeans in a shorter time than natural germination. It can increase BCAA, Phe., Glu., Which is helpful for musculoskeletal health, which becomes important with age. Alternatively, the content of isoflavones can be increased.

Hereinafter, the present invention will be described in detail.

One aspect of the present invention, a method for producing a nutrient-enhanced germinated soybean, comprising a soaking step of immersing soybeans in immersion liquid and a germination step of treating the germination liquid to the soaked soybeans, the immersion liquid The soybean germination provides a method for producing a germinated bean, characterized in that it comprises a promoter.

In the present specification, it will be understood by those skilled in the art that the bean used for germination may be used without limitation so long as it is capable of germination. For example, the kind of beans that can be used in the present invention may be varieties for tofu and tofu, for herbs, for rice or for soybeans. For the varieties of tofu and tofu, Daepung, Hojang, Jangwon, Rhubarb, Sodam, Songhak, Daewon, Genuine, and protein ), Soy milk, Shinpaldal, Taekwang, Great Wall, Longevity, Wuhan, Baiyun, New Egg, Golden and Long Leaf. . Varieties for herbs include mythology, wishes, Anpyeong, southwest, colorful, sorok, soho, vocation, tea garden, and fengshan Herbs, Iksan Herbs, Sobaek Herbs, Gwang'an, Single Leaf, and Galaxy. Examples of rice varieties include celadon, black blue, black sea flakes, black and white, black soybeans and one black soybeans. In addition, varieties for green soybeans include Daol, fresh green, birds, black, sorghum green beans, Hwaeum green soybeans, and large greens. In another aspect of the invention, the soybean is preferably a variety capable of germinating and resistant to pests. Such beans include, for example, myths, wishes, Anpyeong, southwest, colorful, sorok, soho, vocation, and tea gardens. ), Pungsan greens, Iksan greens, Sobaek greens, Gwangan, single leaves and galaxies. However, in the present specification, it will be apparent to those skilled in the art that the bean for germination is not limited to the type of the bean.

In the present specification, "bean germination promoter" is a substance that promotes germination of soybeans compared to natural soybean germination, and may change the nutritional content of germinated soybeans.

In one aspect of the invention, the immersion step is immersed for 5 to 24 hours, the germination step provides a method for producing germinated beans, which germinate 12 hours or more.

Specifically, the immersion step may be 5 hours or more, 8 hours or more, 10 hours or more, 13 hours or more, 14 hours or more, 15 hours or more, or 18 hours or more, and may be 24 hours or less, 20 hours or less, or 18 hours or less. have.

In addition, the germination step is 12 hours or more, 16 hours or more, 20 hours or more, 22 hours or more, 23 hours or more, 24 hours or more, 25 hours or more, 30 hours or more, 32 hours or more, 36 hours or more, 40 hours or more, At least 44 hours, at least 46 hours, at least 47 hours, at least 48 hours, at least 49 hours, at least 50 hours. In addition, the germination step may be 85 hours or less, 80 hours or less, 75 hours or less, 70 hours or less, 65 hours or less, 60 hours or less, 55 hours or less, 50 hours or less, 49 hours or less or 48 hours or less.

When the time of the immersion step and the germination step is within the above range, useful amino acids and isoflavones can be obtained through soybean germination in a short time, thereby increasing the useful product yield.

In one aspect of the present invention, the germination in the germination step is replaced with a new germination solution after 20 to 28 hours, provides a method for producing a germinated bean. Specifically, the time to replace the germination liquid may be when 22 to 26 hours, 23 to 25 hours or 24 hours. When the germination liquid replacement time is within the above range, contamination by microorganisms can be prevented, and useful amino acids and isoflavones can be obtained, thereby increasing useful material production.

In one aspect of the present invention, the immersion step is immersed for 5 to 24 hours, the germination step provides a method for producing germinated beans, the germination for 36 to 60 hours.

In one aspect of the present invention, the germination step provides a method for producing germinated beans, which germinate under light or dark germination conditions.

In one aspect of the invention, the immersion liquid and the germination liquid comprises water, provides a method for producing germinated beans.

In one aspect of the present invention, the immersion liquid provides a germination bean containing 50% by volume to 100% by volume based on the total soaking promoter. Specifically, the soybean germination promoter may be 50 vol% or more, 60 vol% or more, 70 vol% or more, 80 vol% or more or 90 vol% or more, and 100 vol% or less, 95 vol% based on the entire immersion liquid. Or less, 90% or less, 85% or less, or 80% or less. When the content of soybean germination promoter for the whole immersion liquid is within the above range, useful amino acids and isoflavones can be obtained, thereby increasing the useful material production.

In one aspect of the present invention, the soybean germination promoter comprises a citrus water or a processed product thereof, the citrus water is a liquid obtained from the immature fruit of the plant citrus (Citrus) plants, provides a method for producing germinated beans.

In the present specification, "Green Mandarin water (GMW)" is all liquid originated from the immature fruit of citrus. Specifically, this may include water obtained by distilling the juice of the citrus and the peel from the citrus and skin before or after processing, and a liquid obtained by diluting the obtained water by adding cold or ionized water. As used herein, the term “processed product” refers to all materials processed by a conventional method of a raw material, and specifically, refers to extracts, heated materials, dried products, fermented products, distillates, or dry powders of the raw materials. However, the scope of the workpiece is not limited thereto. In one aspect of the invention, the citrus plant may include any one or more of pomelo, citron, mandarin, tanza, golden persimmon, lime, grapefruit, lemon, sour orange, tangerine and sweet orange. Specifically, citrus maxima , citrus medica , mandarin ( citrus reticulate ), citrus trifoliate , citrus japonica , citrus paradise , lemon ( citrus limon ), sour orange ( citrus limon ) It may include any one or more of aurantium , citrus unshiu and sweet orange ( Citrus sinensis ). In one embodiment, the citrus water or the processed product may be 10 to 100% by weight based on the total weight of the bean germination promoter. Specifically, the citrus water or the processed product for the soybean germination promoter may be at least 10 wt%, at least 20 wt%, at least 30 wt%, at least 40 wt%, at least 50 wt% or at least 60 wt%. Also, it may be 100 wt% or less, 95 wt% or less, 90 wt% or less, 85 wt% or less, 80 wt% or less, 70 wt% or less, or 60 wt% or less. When the content of the citrus water is within the above range, the soybeans germinated using the soybean germination promoter show excellent amino acid content. In one embodiment, the citrus water or the processed product provides a soybean germination promoter in which an enzyme of immature fruit of citrus is inactivated. In one embodiment, the citrus water or the processed product is any one of linalool, terpinene-4-ol, terpineol and citronellol. It may contain the above components.

In one aspect of the invention, the nutritional component provides any one or more of isoflavones, free amino acids and dipeptides, the production of germinated beans.

As used herein, "free amino acid" is an amino acid in which an amino acid exists as a single molecule as opposed to a binding amino acid such as a protein or a peptide.

As used herein, "di-peptide" is a compound in which two molecules of free amino acids are formed by peptide bonds.

As used herein, "isoflavones" is a form of naturally occurring isoflavonoids and refers to both isoflavones as glycosides and isoflavones as nonglycosides.

In one aspect of the invention, the isoflavone is an isoflavone glycoside, diazine (Daidzin), 6 "-O-acetyldiazine (6" -O-Acetyldaidzin), 6 "-O-malonyldiazine (6"- O-Malonyldaidzin, Glycitin, 6 "-O-Acetylglycitin, 6" -O-malonylglycitin, 6 "-O-Malonylglycitin, Genistin Genistin), 6 "-O-Acetylgenistin, and 6" -O-malonylgenistin (6 "-O-Malonylgenistin), but are not limited thereto.

In one aspect of the present invention, the isoflavone may be any one or more of Dadezein, Glycitein, and Genistein as an isoflavone nonglycoside, but is not limited thereto.

In one aspect of the invention, the isoflavone is dizin (Daidzin), 6 "-O-acetyldizin (6" -O-Acetyldaidzin), 6 "-O-malonyldiazine (6" -O-Malonyldaidzin) , Glycitin, 6 "-O-Acetylglycitin, 6" -O-malonylglycitin, 6 "-O-Malonylglycitin, Genistin, 6 "-O-Acetylgenistin, 6" -O-malonylgenistin, Daidzein, Glycitein, and Genistein Any one or more of

The free amino acid is Serine, Glutamine, Glycine, Glycine, Aspartic acid, Glutamic acid, Threonine, Alanine, Gammaaminobutyric acid any one or more of -aminobutyric acid, Theanine, Tyrosine, Methionine, Methionine, Valine, Leucine, Isoleucine, Phenylalanine and Trytophan Can include,

The dipeptides are isoleucine-valine (Ile-Val), valine-leucine (Val-Leu), serine-tryptophan (Ser-Trp), isoleucine-leucine (Ile-Leu), isoleucine-phenylalanine (Ile-Phe ) And phenylalanine-leucine (Phe-Leu).

In one aspect of the invention, the free amino acid may include any one or more of Valine, Leucine, and Isoleucine.

Another aspect of the present invention provides a composition comprising a germinated bean or its processed product produced by the above production method.

In another aspect of the invention, the composition provides a food, medicament or cosmetic composition.

In another aspect of the invention, the composition provides a composition formulated in the form of powders, granules, tablets, capsules, suspensions, emulsions, syrups, aerosols, external preparations, suppositories, and sterile injectable solutions.

One embodiment of the present invention provides an extract of the germinated bean. As used herein, the term "extract" includes any material obtained by extracting a component therefrom from a natural product, regardless of the method of extraction or the kind of the component. For example, it is a broad concept that includes all the components which are dissolved in a solvent from a natural product using water or an organic solvent, and those obtained by extracting only a specific component such as an amino acid.

In one aspect of the invention, the pharmaceutical composition may further comprise, in one aspect, suitable carriers, excipients and diluents commonly used in the manufacture of pharmaceutical compositions. In one aspect the carriers, excipients and diluents that may be included in the composition include lactose, dextrose, sucrose, sorbitol, mannitol, xylitol, erythritol, maltitol, starch, acacia rubber, alginate, gelatin, calcium phosphate, calcium silicate, Cellulose, methyl cellulose, microcrystalline cellulose, polyvinyl pyrrolidone, water, methylhydroxybenzoate, propylhydroxybenzoate, talc, magnesium stearate and mineral oil.

In addition, the pharmaceutical composition may be formulated in the form of powder, granules, tablets, capsules, suspensions, emulsions, syrups, aerosols, oral formulations, external preparations, suppositories, and sterile injectable solutions according to conventional methods. .

When formulated, diluents or excipients such as fillers, extenders, binders, wetting agents, disintegrating agents, and surfactants are usually used. Solid preparations for oral administration may include tablets, pills, powders, granules, capsules and the like. Such solid preparations may include at least one excipient such as starch, calcium carbonate, sucrose or lactose, gelatin, in addition to the active ingredient. In addition to simple excipients, lubricants such as magnesium styrate talc may also be included. Oral liquid preparations include suspensions, solvents, emulsions, syrups, and the like, and various excipients such as wetting agents, sweeteners, fragrances, preservatives, etc., in addition to commonly used simple diluents such as water and liquid paraffin. Formulations for parenteral administration may include sterile aqueous solutions, non-aqueous solvents, suspensions, emulsions, lyophilized preparations, suppositories. As the non-aqueous solvent and suspending agent, propylene glycol, polyethylene glycol, vegetable oil such as olive oil, injectable ester such as ethyl oleate and the like can be used. As the base of the suppository, witepsol, macrogol, tween 61, laurin, cacao butter, glycerogelatin and the like can be used.

The dosage of the extracts disclosed herein may vary depending on the condition and weight of the patient, the extent of the disease, the form of the drug, the route of administration, and the duration and may be selected from the ranges commonly used in the art. In one aspect, the daily dose of the active ingredient may be 0.0001 to 1000 g / kg on a dry weight basis, and in one aspect, 0.001 to 1 g / kg.

In one aspect of the invention, the composition may be a cosmetic composition. Cosmetic compositions herein can be prepared in any formulation commonly prepared in the art, including, for example, solutions, suspensions, emulsions, pastes, gels, creams, lotions, powders, soaps, surfactant-containing cleansing , Oils, powder foundations, emulsion foundations, wax foundations and sprays, and the like, but are not limited thereto.

In one aspect of the present invention, the composition may be a food composition, for example, various foods, beverages, gums, teas, vitamin complexes, dietary supplements, and the like, and may be used in the form of powders, granules, tablets, capsules, or beverages. Can be. In addition to the active ingredient, the food composition of each formulation may be suitably selected by a person skilled in the art according to the formulation or purpose of use in addition to the active ingredient, and may be synergistic when applied simultaneously with other raw materials.

At this time, the amount of the active ingredient included in the food or beverage may generally include 1 to 5% by weight of the total food, if the health food composition, the health beverage composition is 0.02 to 10g based on 100ml, 0.3 in one aspect To 1 g.

Hereinafter, the present invention will be described in more detail with reference to Examples. These examples are only for illustrating the present invention, and it will be apparent to those skilled in the art that the scope of the present invention is not to be construed as being limited by these examples.

Preparation of soybean germination promoter Citrus  Produce)

Blue tangerine (unripe state of Wenzhou citrus) was purchased from Juyoung International. The purchased tangerines were washed with purified water to remove foreign substances, and stored at low temperature (5-7 ° C.) to stop the ripening of fruits. The tangerine was subjected to the juice treatment and pressure treatment using a skew type juicer. Heat treatment was performed at 70 ° C. to inactivate the enzyme of the tangerine. The obtained juice was then passed through an 18-30 mesh size net to remove residual solids. The juice obtained by removing the solids was distilled, and the generated vapor was introduced into a reduced pressure concentrator and vaporized by heat treatment at 30 mmHg and 45. The distilled solution was collected through evaporation to obtain citrus water.

Tangerine water obtained through the above process was used in the following examples and experimental examples.

Example 1 and Comparative Examples 1 to 3: germinated soybean production method using soybean germination promoter

-Pretreatment of soybeans: About 80 kg of soybeans harvested in October 2014 from a farmhouse located in Seogwipo-si, Jeju-do, were refrigerated for 3-5 ℃ to maintain germination. 50 g of the soy beans were washed with distilled water 100 ml for 2 minutes (twice).

Germinated beans were prepared by immersing the pretreated soybeans in immersion liquid and germinating using germination liquid. When the above-mentioned soybean germination promoter is not administered in both the immersion step and the germination step (Comparative Example 1), when the blue water is administered in the germination step (Comparative Example 2), the immersion step Germinated beans were prepared in the case of administration to (Example 1) and the case in which the tangerine water was administered in both the dipping step and the germination step (Comparative Example 3).

Immersion treatment: Soak sterilized and washed beans in 150ml of dipping solution (distilled water or tangerine water) and soak in 25 ° C 80% RH incubator for 15 hours.

-Germination: Discard soaking solution of soaked soybeans and wash with 100ml of distilled water (2 times). Beans are added to the balat lei of the bean cultivator (Cheongsiru, Shinchang Inc) and coated with uniform thickness. 500 ml germination (distilled water or tangerine water) is added and germinated by placing a cultivator in a 25 ° C constant temperature incubator for 24 hours. In soybean growers, the germination liquid at the bottom is automatically sprayed on the surface of soybeans every 20 minutes for 1 minute.

-Change germination during germination: Replace germination after 24 hours of germination. Remove germ contaminants from germinating soybeans and discard germination after 24 hours. Germinated beans are placed in a beaker and washed with 200ml of distilled water for 1 minute (repeat twice). Fill a new germination with an existing soybean cultivator and repeat the germination method.

-End of germination: After germination for 24 hours, take a quarter of the growing tray as a sample, and replace the germination liquid to continue germination. After 48 hours germination, one third of the grower tray (same size sampled after 24 hours germination) is taken as a sample. Check the condition of the germinated beans (photo data, germination rate, length measurement) and freeze for 2 hours at -70 ° C. The state of the germinated beans is shown in Table 1 and FIGS.

Germination rate and length comparison (excluding length of bean itself)

"Example" of Table 1 is "Example 1".

After germination, the lyophilizer is dried at 4-10 mmHg for 24 hours, and then dried to less than 10% (w / w) with LOD (loss on dry). The dried germinated soybeans are placed in a 70 ° C dryer for 1 hour to inactivate the soybean enzymes.

Experimental Example 1: Comparison of Isoflavone Contents

Analytical pretreatment: The germination is terminated and the enzyme-inactivated germinated soybeans are finely pulverized for 1 minute * 3 times using an IKA grinder (M20) to analyze the powder that has passed through a 500-micron sieve.

For the germinated beans of Example 1 and Comparative Examples 1 to 3, which were treated before the analysis, the isoflavone content was analyzed by the method of Table 2 below.

Test Methods	Corrected some conditions (mobile phase, mobile phase condition) based on Isoflavone test method
Sample Manufacturing	Ultrasonic Extraction by Adding Methanol
device	Waters alliance 2695 LC (Waters, Milford, MA, USA)
column	Mightysil C18 (250 X 4.6 mm, 5um)
Column temperature	40 ℃
Injection volume	10 μL
Flow rate	1.0 ml / min
Mobile phase	1% acetic acid aqueous solution, acetonitrile
Mobile phase condition	Concentration gradient solvent composition method

According to the isoflavone analysis, the isoflavone contents of the germinated beans of Example 1 and Comparative Examples 1 to 3 were as shown in Table 3 below.

Sample name mg / g	Daidzin	Glycitin	Genistin	Daidzein	Glycitein	Genistein	Total Isoflavones
Comparative Example 1, dark condition 1 day	0.054	0.034	0.051	0.004	0.002	0.003	0.15
Comparative Example 2, dark condition 1 day	0.104	0.059	0.107	0.003	0.001	0.003	0.28
Example 1 , Dark condition  1 day	0.097	0.054	0.097	0.005	0.002	0.003	0.26
Comparative Example 3, dark condition 1 day	0.098	0.055	0.104	0.003	0.002	0.003	0.26
Comparative Example 1, dark conditions 2 days	0.001	0.000	0.012	0.281	0.036	0.413	0.74
Comparative Example 2, dark conditions 2 days	0.000	0.000	0.014	0.291	0.000	0.436	0.74
Example 1 , Dark condition  2 days	0.000	0.000	0.027	0.350	0.000	0.492	0.87
Comparative Example 3, dark conditions 2 days	0.188	0.079	0.184	0.009	0.002	0.014	0.48

 As shown in Table 3, the isoflavone content of the embodiment of the present invention administered with the soybean germination promoter in the immersion step was excellent, in particular, it was confirmed that the isoflavone content is excellent on the second germination.

Experimental Example 2 Comparison of Free Amino Acid Content

Analytical pretreatment: The germination is terminated and the enzyme-inactivated germinated soybeans are finely pulverized for 1 minute * 3 times using an IKA grinder (M20) to analyze the powder that has passed through a 500-micron sieve.

For the germinated beans of Example 1 and Comparative Examples 1 to 3, which were treated before the analysis, the free amino acid content was analyzed by the method of Table 4 below.

Test Methods	AccQ-Tag Amino Acid Assay
Sample Manufacturing	Ultrasonic extraction with distilled water, followed by AccQ-Tag method guidelines
device	Waters AcquityTM UPLC system (Waters, Milford, MA, USA)
column	Waters AccQ-Tag Ultra column (100 x 2.1 mm, 1.7 μm)
Column temperature	55 ℃
Injection volume	1 μL
Flow rate	0.7 ml / min
Mobile phase	AccQ-Tag Ultra Eluent conc. A diluent, AccQ-Tag Ultra Elent B
Mobile phase condition	Concentration gradient solvent composition method

According to the free amino acid analysis, the free amino acid contents of the germinated beans of Example 1 and Comparative Examples 1 to 3 were as shown in Tables 5 and 6.

Free amino acid content of germination day 1

Item	Comparative Example 1 Cancer Conditions 1 Day	Comparative Example 2 Cancer Conditions 1 Day	Example 1 Dark condition 1 day	Comparative Example 4 Cancer Conditions 1 Day
Histidine	0.04	0.05	0.05	0.04
Asparagine	0.14	0.15	0.19	0.14
Serine	0.06	0.07	0.07	0.06
Glutamine	0.03	0.04	0.03	0.03
Arginine	0.18	0.21	0.17	0.17
Glycine	0.01	0.01	0.01	0.01
Aspartic acid	0.04	0.06	0.09	0.09
Glutamic acid	0.23	0.25	0.22	0.21
Throneine	0.04	0.04	0.04	0.03
Alanine	0.13	0.12	0.12	0.1
γ-aminobutyric acid	0.05	0.05	0.05	0.03
Theanine	0.01	0.01	0.01	0.01
Proline	0.03	0.03	0.03	0.03
Cystine	0	0	0	0
Lysine	0.05	0.05	0.05	0.05
Tyrosine	0.04	0.04	0.04	0.04
Methionine	0.01	0.01	0.01	0.01
Valine	0.04	0.04	0.04	0.04
Leucine	0.03	0.03	0.03	0.03
Isoleucine	0.03	0.03	0.03	0.03
Phenylalanine	0.04	0.04	0.05	0.04
Tryptophan	0.05	0.05	0.05	0.05
Total content (%)	1.3	1.4	1.4	1.3

Free amino acid content of germination day 2

Item	Comparative Example 1 Dark conditions 2 days	Comparative Example 2 Cancer Conditions 2 Days	Example 1 Dark conditions 2 days	Comparative Example 4 Cancer Conditions 2 Days
Histidine	0.16	0.14	0.2	0.11
Asparagine	0.71	0.59	0.64	0.98
Serine	0.12	0.13	0.14	0.11
Glutamine	0.13	0.13	0.16	0.05
Arginine				0.15
Glycine	0.12	0.11	0.16	0.01
Aspartic acid	0.32	0.34	0.55	0.11
Glutamic acid	0.22	0.25	0.34	0.27
Throneine	0.13	0.12	0.18	0.04
Alanine	0.25	0.21	0.32	0.11
γ-aminobutyric acid	0.4	0.42	0.53	0.07
Theanine	0.02	0.02	0.03	0.02
Proline	0.15	0.14	0.2	0.04
Cystine	0.01	0.01	0.01	0
Lysine	0.84	0.16	0.15	0.05
Tyrosine	0.14	0.13	0.16	0.03
Methionine	0.07	0.07	0.09	0.01
Valine	0.17	0.16	0.22	0.06
Leucine	0.11	0.1	0.14	0.04
Isoleucine	0.22	0.23	0.3	0.03
Phenylalanine	0.17	0.18	0.24	0.05
Tryptophan	0.09	0.09	0.12	0.07
Total content (%)	4.6	3.7	4.9	2.4

As shown in Tables 5 to 6, the free amino acid content of Example 1 of the present invention to which the soybean germination promoter was administered in the immersion step was excellent, and in particular, it was confirmed that the free amino acid content was excellent on the second germination day.

Experimental Example 3 Comparison of Di-peptide Contents

Example 2 and Comparative Example 4 were grown in Seogwipo, Jeju Island in 2015, and studied in November, and then compared to Example 1 with respect to soybean (cultivar name: mythological soybean) stored at 0-5 degrees C. It prepared in the same manner as in Example 1. After adding water to the beans of Example 2 and Comparative Example 4 and grinding to make 20% (w / w), and then added to the proteolytic enzyme solution of Bacillus marcerans 20% (v / v) at 50 degrees Hydrolysis for 16 hours, and then heat-treated at 95 degrees for 30 minutes to inactivate the enzyme. After concentration / sterilization with a 0.5 micrometer filter and spray-dried to prepare a powder.

For the germinated beans of Example 2 and the beans of Comparative Example 4, di-peptide content was analyzed by the following method.

(1) pretreatment

1) 1 ml of distilled water (10,000 ppm) per 10 mg sample was sonicated for 10 minutes in a 4 ml glass vial.

2) Centrifuge at 14000 rpm, 4 ℃ for 10 minutes

3) Analyze by taking 200 μl of supernatant

(2) UPLC analysis

The pretreated supernatant was subjected to UPLC analysis under the following conditions.

-Column: Waters Acquity UPLC BEH C ₁₈ (ID2.1X100mm, 1.7μm)

Eluent: A-0.1% formic acid water, B-0.1% formic acid in Acetonitrile

Time (min)	% A	% B
0.0	95.0	5.0
0.5	95.0	5.0
15.0	20.0	80.0
15.1	0.0	100.0
17.0	0.0	100.0
17.1	95.0	5.0
20.0	95.0	5.0

Flow rate: 0.4 mL / min

Injection volume: 1 μL

-Column Temp. : 50 ℃

(3) MS analysis

The pretreated supernatant was subjected to MS analysis under the conditions of Table 8.

Apparatus	Thermo Q-Exactive Plus System: Hybrid Quadrupole-FT / Orbitrap Mass Spectrometer
Source parameter	Ionization mode	Heated Electron Spray Ionization (HESI) mode
	Sheath gas flow rate	45
	Aux gas flow rate	10
	Spray voltage	3.50
	Capillary temperature	350 ℃
	S-Lens RF Level	70
	Aux gas heater temp	350 ℃
Scan parameter	Polarity	Positive
	Scan range	100.0 to 1500.0 m / z
	Resolution	70000
	Microscan	One
	AGC target	3e6
	Maximun inject time	100
dd-MS 2	Stepped NCE	20, 30, 40

According to the di-peptide analysis, the di-peptide contents of Example 2 and Comparative Example 4 were as shown in Tables 9 and 10 below.

No.	RT	Dipeptide	Abbrev.	[M + H] + Formula	m / z [M + H] +
One	2.37	Ile-val	IV	C11H23N2O3	231.16966
2	3.11	Val-Leu	VL	C11H23N2O3	231.16996
3	3.36	Ser-Trp	SW	C14H18N3O3	292.12872
4	3.90	Ile-Leu	IL	C12H25N3O2	245.18593
5	4.52	Ile-Phe	IF	C15H23N2O3	279.16995
6	4.82	Phe-Leu	FL	C15H23N2O3	279.16977

No.	Dipeptide	Total NL		maximum NL		Relative Quantitative Index		Increase
		Comparative Example 4	Example 2	Comparative Example 4	Example 2	Comparative Example 4	Example 2
One	Ile-val	7.8E + 08	1.5E + 09	6.6E + 07	2.8E + 08	0.085	0.191	2.25
2	Val-Leu			1.9E + 08	7.7E + 08	0.249	0.520	2.09
3	Ser-Trp			1.8E + 07	8.2E + 07	0.024	0.056	2.37
4	Ile-Leu			3.5E + 07	7.9E + 08	0.046	0.537	11.78
5	Ile-Phe			1.1E + 08	4.1E + 08	0.144	0.277	1.92
6	Phe-Leu			9.5E + 07	4.1E + 08	0.123	0.278	2.26

Relative index: maximum peak NL / total NL

** Increase Ratio: Example 2 Relative Quantitative Index / Comparative Example 4 Relative Quantitative Index

As in Table 9 and 10, it was confirmed that the di-peptide content of the embodiment of the present invention administered soybean germination promoter in the immersion step.

Claims (11)

1. As a method of producing germinated soybean with enhanced nutrition,

   And a germination step of treating the germination liquid to the soaked beans after the soaking step in the soaking solution, and the soaking solution comprises a soybean germination promoter.
2. The method of claim 1,

   The immersion step is immersed for 5 to 24 hours, the germination step is to germinate for more than 12 hours, the production method of germinated beans.
3. The method of claim 1,

   The germination solution in the germination step is replaced with a new germination solution after 20 to 28 hours, the production method of the germinated beans.
4. The method of claim 1,

   The immersion step is immersed for 5 to 24 hours, the germination step is to germinate for 36 to 60 hours, the production method of germinated beans.
5. The method of claim 1,

   The germination step is to germinate under light or dark germination conditions, the production of germinated beans.
6. The method of claim 1,

   The immersion liquid and the germination liquid containing water, the production method of the germinated beans.
7. The method of claim 1,

   The immersion liquid containing a soybean germination promoter in 50% by volume to 100% by volume based on the total immersion liquid, method for producing a germinated bean.
8. The method of claim 1,

   The soybean germination promoter includes a citrus water or a processed product thereof, wherein the citrus water is a liquid obtained from an unripe fruit of a citrus plant.
9. The method of claim 1,

   The nutritional component comprises any one or more of isoflavones, free amino acids and dipeptides, the production of germinated beans.
10. The method of claim 9,

    The isoflavones are dizin (Daidzin), 6 "-O-Acetyldaidzin, 6" -O-malonyldizin (6 "-O-Malonyldaidzin), glycidin (Glycitin) 6 "-O-Acetylglycitin, 6" -O-malonylglycitin, 6 "-O-Malonylglycitin, Genistin, 6" -O-acetylgenistine At least one of 6 "-O-Acetylgenistin, 6" -O-malonylgenistin, Daidzein, Glycitein and Genistein ,

    The free amino acid is Serine, Glutamine, Glycine, Glycine, Aspartic acid, Glutamic acid, Threonine, Alanine, Gammaaminobutyric acid any one or more of -aminobutyric acid, Theanine, Tyrosine, Methionine, Methionine, Valine, Leucine, Isoleucine, Phenylalanine and Trytophan Including;

    The dipeptides are isoleucine-valine (Ile-Val), valine-leucine (Val-Leu), serine-tryptophan (Ser-Trp), isoleucine-leucine (Ile-Leu), isoleucine-phenylalanine (Ile-Phe ) And phenylalanine-leucine (Phe-Leu), a method for producing germinated beans.
11. The method of claim 10,

    The free amino acid includes any one or more of valine, leucine, leucine and isoleucine.

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