KR20170030806A - Method for producing Soybean with increased glyceollins content by infecting mycelium of Aspergillus sojae - Google Patents

Method for producing Soybean with increased glyceollins content by infecting mycelium of Aspergillus sojae Download PDF

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KR20170030806A
KR20170030806A KR1020150128174A KR20150128174A KR20170030806A KR 20170030806 A KR20170030806 A KR 20170030806A KR 1020150128174 A KR1020150128174 A KR 1020150128174A KR 20150128174 A KR20150128174 A KR 20150128174A KR 20170030806 A KR20170030806 A KR 20170030806A
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mycelium
soybean
inoculated
present
spores
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KR1020150128174A
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Korean (ko)
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KR101760184B1 (en
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김정상
김민선
유재혁
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경북대학교 산학협력단
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    • A—HUMAN NECESSITIES
    • A23—FOODS OR FOODSTUFFS; TREATMENT THEREOF, NOT COVERED BY OTHER CLASSES
    • A23L—FOODS, FOODSTUFFS, OR NON-ALCOHOLIC BEVERAGES, NOT COVERED BY SUBCLASSES A21D OR A23B-A23J; THEIR PREPARATION OR TREATMENT, e.g. COOKING, MODIFICATION OF NUTRITIVE QUALITIES, PHYSICAL TREATMENT; PRESERVATION OF FOODS OR FOODSTUFFS, IN GENERAL
    • A23L25/00—Food consisting mainly of nutmeat or seeds; Preparation or treatment thereof
    • A23L25/40—Fermented products; Products treated with microorganisms or enzymes
    • A—HUMAN NECESSITIES
    • A23—FOODS OR FOODSTUFFS; TREATMENT THEREOF, NOT COVERED BY OTHER CLASSES
    • A23L—FOODS, FOODSTUFFS, OR NON-ALCOHOLIC BEVERAGES, NOT COVERED BY SUBCLASSES A21D OR A23B-A23J; THEIR PREPARATION OR TREATMENT, e.g. COOKING, MODIFICATION OF NUTRITIVE QUALITIES, PHYSICAL TREATMENT; PRESERVATION OF FOODS OR FOODSTUFFS, IN GENERAL
    • A23L29/00—Foods or foodstuffs containing additives; Preparation or treatment thereof
    • A23L29/065—Microorganisms
    • A—HUMAN NECESSITIES
    • A23—FOODS OR FOODSTUFFS; TREATMENT THEREOF, NOT COVERED BY OTHER CLASSES
    • A23L—FOODS, FOODSTUFFS, OR NON-ALCOHOLIC BEVERAGES, NOT COVERED BY SUBCLASSES A21D OR A23B-A23J; THEIR PREPARATION OR TREATMENT, e.g. COOKING, MODIFICATION OF NUTRITIVE QUALITIES, PHYSICAL TREATMENT; PRESERVATION OF FOODS OR FOODSTUFFS, IN GENERAL
    • A23L5/00—Preparation or treatment of foods or foodstuffs, in general; Food or foodstuffs obtained thereby; Materials therefor
    • A23L5/20—Removal of unwanted matter, e.g. deodorisation or detoxification
    • A23L5/28—Removal of unwanted matter, e.g. deodorisation or detoxification using microorganisms

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  • Life Sciences & Earth Sciences (AREA)
  • Microbiology (AREA)
  • Health & Medical Sciences (AREA)
  • Nutrition Science (AREA)
  • Chemical & Material Sciences (AREA)
  • Engineering & Computer Science (AREA)
  • Food Science & Technology (AREA)
  • Polymers & Plastics (AREA)
  • Preparation Of Compounds By Using Micro-Organisms (AREA)

Abstract

The present invention relates to a soybean having increased glycerol content and a method for producing the same, and more particularly, to a method for enhancing glycerol content of a soybean inoculated with an mycelial fungal strain and a soybean produced by the method. The mycelium-inoculated soybean produced by the method according to the present invention showed an increase in glycerol content compared to soybean seeds inoculated with fungal spores, and the problem of imitation and odor derived from fungal spore inoculation was effectively improved and processed into various kinds of foods It has the advantage that it can do.

Description

FIELD OF THE INVENTION [0001] The present invention relates to a method for producing soybean which has increased glycerol content by inoculation of mold mycelium,

The present invention relates to a soybean having increased glycerol content and a method for producing the same, and more particularly, to a method for enhancing glycerol content of a soybean inoculated with an mycelial fungal strain and a soybean produced by the method.

In general, phytonalexin is a low-molecular-weight antimicrobial substance that plants produce by itself to fight fungi when various insects or fungi enter. These substances are not found in the plant before infection, but within a few hours after infection It is synthesized at a very high rate and acts on the infected area and has toxic properties against various bacteria and fungi. These phytoalexins are self-produced by protecting plants. However, recently, the functions such as antimicrobial activity of these substances have been discovered, and a lot of studies have been actively carried out to use them for stable and inexpensive medicines, dyes, flavors and directions .

There are various functional ingredients in soybeans, and many studies have been conducted on them. It has been known that when the edible fungus is treated in the germination of soybean, special functional substances which are not present in the soybean itself are produced. Among them, glyceolins, which is a kind of phytoalexin produced in soybean, are most actively studied .

If soybeans are germinated by inoculating fungus, then it is necessary to remove the fungus in order to use the soybeans in other food processing, and since it is difficult to remove completely, it may have a disadvantage .

Therefore, there is a need for a new induction method of glycerol synthase to solve the defects of soybean inoculated with mold spores.

Korean Patent No. 10-1126297 Korean Patent Publication No. 10-2010-0035843

Accordingly, an object of the present invention is to provide a soybean having increased glycerol content by using mycelia and a method for producing the same.

In order to achieve the above-mentioned object of the present invention,

The present invention relates to a method for producing soybean seeds, comprising the steps of: And culturing the soybeans inoculated at 20 to 30 DEG C for 1 to 7 days, wherein the glycerol content is increased.

In one embodiment of the present invention, the mycelium is Aspergillus scavenger (Aspergillus sojae), or separation tank crispus oligonucleotide sports Russ (rhizopus oligosporous < / RTI > strains.

In one embodiment of the present invention, the inoculation of the mycelium may be a step of inoculating the mycelium suspension onto the beans divided surface.

In one embodiment of the present invention, the mycelial suspension may be a mycelium of mycelia obtained by inoculating spores into a liquid medium in an amount of 2 × 10 8 spores / 200 ml and then culturing in a stirring incubator for 14 to 18 hours.

The present invention provides beans with increased glycerol content produced by the process of the present invention.

The mycelium-inoculated soybean produced by the method according to the present invention showed an increase in glycerol content compared to soybean seeds inoculated with fungal spores, and the problem of imitation and odor derived from fungal spore inoculation was effectively improved and processed into various kinds of foods It has the advantage that it can do.

BRIEF DESCRIPTION OF THE DRAWINGS FIG. 1 is a schematic diagram of a method of inoculating spores or mycelium for comparison of glycerol production according to the form of Aspergillus sojae of the present invention. FIG.
FIG. 2 is a photograph of a day-by-day observation of an agar bean inoculated with an Aspergillus sojae spore or mycelium of the present invention.
Fig. 3 is a micrograph of a 1.5 × 12.5 magnification of the agar bean inoculated with the spore or mycelium of Aspergillus sojae according to the present invention.
Fig. 4 is a micrograph of a 1.5 x 20 magnification of agar bean inoculated with an Aspergillus sojae spore or mycelium of the present invention.
FIG. 5 is a graph showing the distribution of the Aspergillus sojae ) spores or mycelium in the culture medium.
FIG. 6 is a graph showing the results obtained by comparing the Aspergillus sojae ) mycelium of glycerol I in HLPC standard analysis.
FIG. 7 is a photograph of Aspergillus sojae) is HLPC analysis for glycerides I uploaded content measurement of soy agar inoculated with mycelium.

Inoculating an edible fungus in the germination of soybean produces glycerol, a kind of phytoalexin in soybean. In case of germination by inoculating the fungus with soybeans, it is necessary to remove the fungus in order to utilize the soybean in other food processing, , It may affect mold odor and product color when used as other food materials. In order to utilize soybean itself containing glycerol as food material, it does not affect flavor or aroma of bean Elicitors need to be excavated.

Therefore, in the present invention, soybean having increased glycerol content was prepared. In the method for producing soybean according to the present invention, the step of inoculating mycelium with mycelium of soybeans; And culturing the soybeans inoculated at 20 to 30 DEG C for 1 to 7 days, wherein the glycerol content is increased.

In order to compare the glycerol content of soybean and fungal spores in accordance with the present invention, edible fungus spores were first prepared, and edible fungus aspergillus ( Aspergillus The cells were cultured in a complete medium (CM), 10 ml of 0.1% tween buffer per plate was collected and collected in a tube using a spreader, centrifuged and distilled water was added to the remaining pellet, (See Preparation Example 1).

According to one embodiment of the present invention, mycelia are obtained by inoculating the measured spores into a liquid medium in an amount of 2 x 10 8 spores / 200 ml, then culturing in a stirring incubator, and filtering the mycelium with a concentration of 2 × 10 5 / The results of HPLC analysis of soybean cultivated by treatment with soybeans cut in half showed that there was a higher amount of glycerolin I in germinated soybeans inoculated with mycelia than in sprouted inoculated soybeans , And more than twice as much glycerolin I was measured especially on days 3 and 5 (see Example 3, Table 2 and Figure 7).

The "glycerol" of the present invention is a kind of phytoalexin substance produced during hypersensitive reaction or fungal stress in soybean.

According to a specific embodiment of the present invention, the glycerol compound is obtained by treating the soaked soybean with an Aspergillus sojae strain and extracting the soybean extract from the soybean extract by conventional purification methods such as chromatography By separation and purification.

Glycerol I is the predominant form in the natural world and its two isomers glycerolines II and III are present in minor amounts (Clinical Cancer Research 2006; 12 (23) December 1, 2006, 7159- 7164, Antiestrogenic Glyceollins Suppress Human Breast and Ovarian Carcinoma Tumorigenesis).

The active ingredient "glycerol" in the present invention is meant to include both glycerolin I, glycerolin II, glycerolin III, or mixtures thereof.

According to a preferred embodiment of the present invention, the glycerol is glycerolin I, glycerolin II, glycerolin III, or a mixture thereof. According to a more preferred embodiment of the present invention, the glycerol is glycerolin I.

Hereinafter, the present invention will be described in more detail with reference to Examples. It will be apparent to those skilled in the art that these embodiments are merely illustrative of the present invention and that the scope of the present invention is not limited to these embodiments.

<Preparation Example 1>

Aspergillus sojae ( Aspergillus sojae ) Preparation and spore counting

Aspergillus Sojae were purchased from Gyeongsang National University and Aspergillus sojae was cultured on complete medium (CM, Complete media) for 2 days at 30 ℃. Place 10 ml of 0.1% tween buffer per plate in a 50 ml tube using a spreader, and centrifuge at 5000 RPM for 10 minutes. Discard the supernatant and add 10 ml of autoclaved distilled water to the remaining cell pellet. After vortexing, the cells are diluted 100 times (10 μl cell + 990 μl autoclave dw) and the number of spores is counted using a hemocytometer (see FIG. 1).

<Preparation Example 2>

Prepare sweet potatoes

About 2 g of Aga 3 (soybeans grown by contract in Seobu-myeon, Hongseong-gun, Chungcheongnam-do), which is high in isoflavone content, was placed in a 50 ml tube and sterilized for 1 to 2 minutes with 70% ethanol. After autoclaving, Rinse with distilled water three times. After that, the autoclave is filled with distilled water up to 45 ml and is called in the dark room at room temperature for about 5 hours. The so-called beans are peeled and then cut in half. Place on a petri dish containing autoclaved whatman 110 mm filter paper and 1 ml of autoclaved distilled water, with the cut side up.

&Lt; Example 1 >

Spore inoculation

The aspergillus scavenger ( Aspergillus) measured in Preparation Example 1 Sojae spores were treated with 2 x 10 6 spores in 10 μl of distilled water to half of the beans And cultured at 25 占 폚 for 0 days, 1 day, 3 days, 5 days, and 7 days, respectively.

&Lt; Example 2 >

Mycelium inoculation

In the preparation example 1, the spores are inoculated to 200 ml of a soybean broth + trace element (2 × 10 8 spores / 200 ml), and cultured in a 220 rpm shaking incubator at 37 ° C. for 16 hours. After culturing for 16 hours, the mycelium is filtered using funnel and miracloth, collected in a 50 ml tube, vortexed with 10 ml sterilized distilled water. The vortex mycelium was treated with 2 × 10 5 mycelia in 10 μl of distilled water and incubated at 25 ° C for 0 days, 1 day, 3 days, 5 days, 7 days.

A petri dish of soybean inoculated with the spore was inoculated with the petri dish of the soybeans inoculated with the mycelium as shown in FIG. 2, and observed with a microscope at 1.5 × 12.5, At a magnification of × 20, characteristic portions were observed closer to each other. At a magnification of 1.5 × 50, the growth of mycelium and spores was observed (see FIGS. 3 to 5).

&Lt; Example 3 >

HPLC analysis for determination of glycerol content

The aspergillus scavenger ( Aspergillus) prepared in Examples 1 and 2 sojae ) Spore inoculated beans and aspergillus ( Aspergillus Sojae Mycelia In order to measure the glycerol content of the soybeans, the soybeans contained in the Petri dishes of Examples 1 and 2 were transferred to a 50 ml tube and frozen at -70 ° C for a day. The frozen sample is lyophilized for about two days to completely dry. After lyophilization, each sample is ground into a powder with a mortar and weighed. Add 1 g / 6 ml of 80% EtOH, vortex for about 20 seconds, and extract at 50 ℃ in water bath. After extraction, cool for about 30 minutes and centrifuge at 5000 RPM for 10 minutes. After centrifugation, the extract is filtered using a 0.22 μm filter. After filtration, the solution was stored at 20 [deg.] C for subsequent HPLC analysis.

The filtered extract was diluted with 1 mM glyceollin I using a Gemini 5 μm C18 150 × 2.00 mm Phenomenex column under the following conditions (see FIG. 6). After the standard measurement, the sample was diluted and the glycerol content was measured.

HPLC analysis conditions for determination of glycerolin I content R.T. (min) 0 40 45 55 % A 90 65 90 90 % B 10 35 10 10 Mobile phase: solvent A (0.1% phosphoric acid), solvent B (acetonitrile)
Column temperature: 25 ℃
Flow rate: 0.6 ml / min
Injection volume: 10μl
Detection wavelength: 280 nm

As a result of the HPLC measurement, a larger amount of glyceolin I was detected in agar beans inoculated with mycelia than in spore-infected agar beans, and more than twice as much glycerolin I (3- glyceollin I) was measured (see Table 2 and FIG. 7).

(Unit: ug / g) control Spore Mycelia 0 day 2.1379 397.1767 9.2423 1 day 4.7205 225.9218 324.9719 3 day 4.7016 414.8554 1077.8530 5 day 18.0472 435.7371 1117.7234 7 day 5.1956 736.2531 934.0704

In general, in the present invention, Agarose No. 3, which is a highly isoflavone-rich variety, was used as an edible fungus aspergillus ( Aspergillus Sojae spores or mycelium were inoculated to induce the glycerol synthesis, and the induction of the glycerol synthesis was remarkably accomplished 1 to 5 days after the inoculation of the mycelium, and it did not affect the appearance and flavor of the soybean.

The present invention has been described with reference to the preferred embodiments. It will be understood by those skilled in the art that various changes in form and details may be made therein without departing from the spirit and scope of the invention as defined by the appended claims. Therefore, the disclosed embodiments should be considered in an illustrative rather than a restrictive sense. The scope of the present invention is defined by the appended claims rather than by the foregoing description, and all differences within the scope of equivalents thereof should be construed as being included in the present invention.

Claims (5)

Inoculating the soybean with mycelium of fungi; And
Culturing the inoculated soybeans at 20 to 30 DEG C for 1 to 7 days. The method according to claim 1,
Wherein the mycelium is a mycelial suspension using spores of Aspergillus sojae or rhizopus oligosporous strain. The method according to claim 1,
Wherein the mycelium inoculation is carried out by inoculating mycelium suspension into a soybean divided surface. 3. The method of claim 2,
Wherein the suspension of mycelium is inoculated into a liquid medium in an amount of 2 x 10 8 spores / 200 ml, and then cultured in a stirring incubator for 14 to 18 hours, and then filtered to obtain a mycelium of increased glycerol content. 6. A soybean bean having increased glycerol content produced by the method of any one of claims 1 to 4.
KR1020150128174A 2015-09-10 2015-09-10 Method for producing Soybean with increased glyceollins content by infecting mycelium of Aspergillus sojae KR101760184B1 (en)

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Cited By (1)

* Cited by examiner, † Cited by third party
Publication number Priority date Publication date Assignee Title
KR101968019B1 (en) 2017-11-29 2019-04-10 경북대학교 산학협력단 Composition comprising glyceollins with cognition-improving effects and the use thereof

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KR20230071371A (en) 2021-11-16 2023-05-23 씨제이제일제당 (주) Aspergillus oryzae capable of producing mycelium comprising all kinds of essential amino acids

Citations (2)

* Cited by examiner, † Cited by third party
Publication number Priority date Publication date Assignee Title
KR20100035843A (en) 2008-09-29 2010-04-07 (주)엔앤비 The coposition, containing glyceollins, showing eminent anti-inflammation effects.
KR101126297B1 (en) 2009-11-16 2012-03-19 한국원자력연구원 Use of Glyceollin

Patent Citations (2)

* Cited by examiner, † Cited by third party
Publication number Priority date Publication date Assignee Title
KR20100035843A (en) 2008-09-29 2010-04-07 (주)엔앤비 The coposition, containing glyceollins, showing eminent anti-inflammation effects.
KR101126297B1 (en) 2009-11-16 2012-03-19 한국원자력연구원 Use of Glyceollin

Cited By (1)

* Cited by examiner, † Cited by third party
Publication number Priority date Publication date Assignee Title
KR101968019B1 (en) 2017-11-29 2019-04-10 경북대학교 산학협력단 Composition comprising glyceollins with cognition-improving effects and the use thereof

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