KR101382930B1 - Method for manufacturing mushroom vinegar using liquid culture medium of sparassis crispa - Google Patents

Abstract

The present invention relates to a method for manufacturing mushroom vinegar using a liquid culture medium of Sparassis crispa, and more specifically, to a method for manufacturing mushroom vinegar comprising: a step of culturing Sparassis crispa fungus in a media which glucose is contained in water strained off coagulating bean curd, adding sucrose or potato powder in the medium, and inoculating acetobacter thereby acetic acid-fermenting. In addition, in providing Sparassis crispa vinegar manufactured by the method, the vinegar has higher antioxidant activity than conventional vinegar.

Description

Method for manufacturing mushroom vinegar using liquid culture medium of Sparassis crispa}

The present invention is a mushroom mushroom ( Sparrassis) The present invention relates to a method for preparing mushroom vinegar using a liquid culture solution of crispa ), and more particularly, to a method for producing functional mushroom vinegar, in which the mycelium of the blossom mushroom is cultured in a medium containing tofu pure water and subjected to second acetic acid fermentation.

Recently, as awareness of health has increased, research on natural products containing a large amount of physiologically active substances has been actively conducted, and interest in mushrooms containing a large amount of useful physiologically active substances among natural products is increasing. As a method for using mushrooms as a functional food material, there is a growing interest in cultivating mycelia of mass production and the extracellular functional substances secreted by mushrooms. When liquid culture is performed using the growth characteristics of mushroom mycelium, various metabolites having antimicrobial and antioxidant activities can be obtained depending on the composition and growth conditions of the medium. That is, the mushroom bacteria can bioconvert the components contained in the medium to substances having various functionalities, and secondary metabolites having special functions through mushroom culture can be produced and accumulated in the culture solution. In addition, artificial liquid culture has the advantage that it is possible to produce in a limited space without being seasonally limited in culture production. The economics of producing mushroom mycelium liquid culture solution is to secure inexpensive culture raw materials while reducing the incubation time.

Tofu was obtained as a by-product in the production of tofu as a material to replace the expensive medium raw materials used for the liquid culture of the mushroom. Tofu puree is a by-product from the compression process of protein coagulants in general tofu production, and about 50% of the total amount of water added in tofu production can be obtained as tofu puree. Tofu products from the tofu plant contain water-soluble substances in addition to the insoluble fiber of soybeans and some coagulated polymer proteins, and a small amount of coagulant used to coagulate the tofu. The solid content of tofu is 2-3%, among which nitrogen compounds such as proteins are about 16% and sugars are about 53%. Functional substances such as isoflavone, saponin and oligosaccharides It is known that it contains. The tofu puree containing such a functional substance was used for cultivating the blossom mushroom of the present invention.

The mushroom spawn used for the liquid culture of synthetic medium and tofu purify is Sparassis crispa . Blossom mushrooms are edible mushrooms distributed in Korea, Japan, China, North America, Europe, Australia, etc. Fruiting bodies are formed like cabbage by clustering on stems or stumps near the roots of living trees from summer to autumn. Flower mushrooms contain a large amount of β-glucan (43.6 g per 100 g of dried fruiting body), and research on artificial cultivation and extraction of functional substances is rapidly progressing. In addition, the mushroom mushroom is significantly higher in content of β-1,3-glucan (glucan) than other mushrooms, has been proved to have anti-cancer and anti-tumor effects, and is widely recognized as an edible as well as medicinal. In particular, it is known to produce antifungal metabolite such as sparsol, and sparsol has been studied for the treatment of human diseases and control of leaf disease.

 Vinegar is one of the longest-established fermented foods in the history of human life along with alcohol. It is a fermented food with a long history regardless of the East or West. It is used as an acidic seasoning and is closely related to our diet. Because of its strong acidity, it has the effect of inhibiting the growth of harmful microorganisms in food. In addition to acetic acid, it contains a variety of organic acids, sugars, amino acids and esters (esters) to stimulate the appetite with a unique flavor and taste, and is effective in preventing adult diseases such as arteriosclerosis and high blood pressure. In addition, cholesterol lowering effect, body fat reduction, fatigue recovery effect by lactic acid decomposition and vitamin C protection effect has been recently increased interest in vinegar has been released a variety of drinks added with drinking vinegar and vinegar. Antioxidants that inhibit oxidation by free radicals are known to be effective in preventing and alleviating diseases. Recently, researches on substances having antioxidant capacity have been actively conducted. Among them, vinegar is known to have high antioxidant activity, and 19 kinds of vinegars (honey vinegar, pomegranate vinegar, bokbunja vinegar, mulberry vinegar, red ginseng vinegar, omiza vinegar, white vinegar vinegar, blueberry vinegar, five grain black vinegar, plum fiber black vinegar) , Persimmon vinegar, lemon vinegar, pear vinegar, apple vinegar, brown rice vinegar, balsamic vinegar, white wine vinegar, red wine vinegar, edible glacial acetic acid) There is little research on mushroom vinegar using mushroom liquid culture.

On the other hand, Korean Patent Application No. 10-2009-0053175 discloses a method for producing a mushroom mushroom drink using the mushroom extract, there is no mention of a method for producing a mushroom vinegar using the mushroom mushroom liquid culture solution of the present invention.

 Therefore, the present inventors liquid cultured the mycelium of zinnia mushroom containing a large amount of β-glucan known as medicinal and edible in a medium containing tofu sprouts, and then subjected to acetic acid fermentation for a second time to evaluate the antioxidant activity of functional vinegar The present invention has been completed by developing.

An object of the present invention comprises culturing the mycelium mushroom mycelium in a medium containing glucose (glucose) in the tofu pure, adding sucrose or potato powder to the culture solution, inoculating acetic acid bacteria and inoculating acetic acid fermentation It is to provide a method for producing blossom vinegar.

Another object of the present invention to provide a mushroom mushroom vinegar produced by the above method.

In order to achieve the above object, the present invention is tofu tofu medium containing 1 to 10% by weight glucose (glucose), 1 to 15% by weight sucrose or 1 to 5% by weight potato powder relative to the total tofu pure Culturing the blossom mushroom mycelium; And inoculated with acetic acid bacteria in the mushroom mushroom mycelium culture medium, and fermentation of alcohol by adding 3 to 9% by weight to the total culture solution.

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Specifically, the acetic acid bacterium is characterized in that Gluconacetobacter hansenii ( Gluconacetobacter hansenii ), the acetic acid fermentation is characterized in that the fermentation for 3 to 7 days at 20 to 40 ℃.

In addition, the present invention provides a blossom mushroom vinegar produced by the above method.

The present invention is a mushroom vinegar manufacturing method using a liquid culture liquid of zinnia mushroom, liquid mycelium of zinnia mushroom containing a large amount of β-glucan known as medicinal and edible liquid culture in a medium containing tofu pure water, secondary acetic acid After fermentation, functional vinegar was developed by evaluating antioxidant activity.

1 is a photograph of the flower mushroom mycelium cultured in PD agar medium and the flower mushroom culture cultured in PD liquid medium.
Figure 2 is a photograph of the culture solution of the mushroom mushroom cultured in a 5L jar fermenter (jar fermenter) and 5L culture bottle.
Figure 3 Gluconacetobacter passaged in mannitol medium ( Gluconacetobacter) hansenii ) solid medium culture and liquid culture pictures.
Figure 4 shows the chemical composition of the mushroom culture medium according to the concentration of sucrose (sucrose) added to tofu pure water medium and distilled water.
Figure 5 shows the chemical components of the mushroom culture medium with the fermentation time in tofu pure water medium.
Figure 6 shows the effect of the fermentation time of the mushroom mushroom culture in the chemical component of the mixed culture.
Figure 7 shows the chemical composition of the mushroom culture medium according to the potato starch concentration added to the tofu pure medium.
Figure 8 shows the effect of the starch concentration of the flower mushroom culture in the chemical composition of the mixed culture.
Figure 9 shows the DPPH radical scavenging effect of the flower mushroom culture in the chemical component of the mixed culture. Pleurotus eryngii : Matsutake vinegar, Phellinus baumii : situation mushroom vinegar.
Figure 10 shows the ABTS radical scavenging effect of the mushroom mushroom culture in the chemical component of the mixed culture. Pleurotus eryngii : Matsutake vinegar, Phellinus baumii : situation mushroom vinegar.

Hereinafter, the present invention will be described in more detail with reference to the following examples. However, the present invention is not limited by these examples.

< Example  1> Physicochemical Properties of Cultures of Mushroom Fungus Mycelium

1. Materials

Tofu pure water was supplied from Doowon Food (Gimcheon, Korea), and KH ₂ PO ₄ , MgSO ₄ , Glucose was used by Sigma Company, Yeast Extract was manufactured by Difuco Company, Soluble Starch. (soluble starch) was used to purchase Deoksan Chemical (Ansan, Korea) products. Soy bean flour was purchased from Cheonho Food (Daegu, Korea), sealed and stored in a cow. Sucrose was purchased from CJ CheilJedang (Incheon, Korea), and white barley was purchased from Gagaho Lake (Hapcheon, Korea). Potato starch was purchased from Samjin Food (Seongju, Korea). Used. Jujube extract used for acetic acid fermentation was purchased from Eden Town F & B (Incheon, Korea). Pretanol A (prethanol A) was purchased from Ansan, Korea. SMART ™ BCA) was purchased from Intron Biotechnology INC. (Sungnam, Korea).

2. Tofu Pure  Component analysis

Moisture, crude protein, and crude fat contents of tofu pures were quantified according to AOAC method. Moisture was measured using atmospheric pressure drying method, crude protein was analyzed by nitrogen automated protein analyzer (Buchi 339, Buchi Labortechnik AG, Flawil, Switzerland), crude fat was measured by Soxhlet extraction method, and mineral analysis was performed by ICP-MS (ELAN 9000, Perkin). Elmer Inc., CA, USA).

3. Cultivation of mycelia

Sparassis from the Rural Development Administration crispa Wulf. ex Fr) was inoculated in a Potato Dextrose Agar (PDA; Difco. Co. USA) medium and cultured at a temperature of 25 ° C., pH 5.1 ± 0.2, and dark for at least 20 days (FIG. 1).

4. Mushroom Mushroom Mycelia Liquid Culture

Aseptic incisions of cultivated mushroom mushrooms in PDA agar plate medium were added to 100 mL of potato dextrose broth and homogenized with a homogenizer (10,000 rpm, 3 min). 10 mL each of the homogenized liquid was added to 100 mL of potato dextrose broth, followed by incubation for 10 days at a temperature of 25 ° C., rotational speed of 160 rpm, and used as a medium for liquid seedling (FIG. 1).

1) Culture with Restriction Medium

The liquid medium composition was sucrose 10.75 g / L, soluble starch 5.35 g / L, soybean flour 1.5 g / L, K ₂ HPO ₄ 0.5 g / L, MgSO ₄ 0.5 g / L was used in combination. As a liquid culture, a 5 L jar fermenter (jar fermenter; Fermentec Co. Ltd., Cheongwon, Korea) was used, and the culture conditions were analyzed by incubation for 10 days at a temperature of 25 ℃, air injection amount 1 vvm, rotation speed 300 rpm.

2) Cultivation using overlying barley saccharification medium

Cultivation of Cultivated Mycelium Mycelium in Liquid Culture of Cultivated Mycelium was used for culture of Cultivated Mycelium in Cactus mycelium. To prepare saccharified medium, the barley was washed with groundwater to remove debris and damaged grains, and selected using a sieve. Selected barley was soaked in water at 20 ° C. for 2 days to germinate barley, and homogenized with a homogenizer (10,000 rpm, 2 min) by adding water 1: 1 to germinated barley of 55% water content. After pressing with a cloth to separate the supernatant. Liquefaction enzyme Termamyl was added to the supernatant by 0.1% (w / w) on a solid basis and heat-treated at 85 ° C. for 1 hour to saccharify it. The saccharified liquid had a sugar content of 17.8˚Brix, diluted three times with water, autoclaved at 121 ° C. for 30 minutes, and used as a bulk liquid medium for the mushroom. As a liquid culture, a 5 L jar fermenter was used to analyze a temperature of 25 ° C., an air injection amount of 1 vvm, a rotational speed of 300 rpm, and 10 days of incubation.

3) Culture using tofu pure water

Tofu pure medium was used by adding 2% (w / v) of glucose to the weight of tofu pure. As a liquid culture, a 5 L jar fermenter was used to analyze a temperature of 25 ° C., an air injection amount of 1 vvm, a rotational speed of 300 rpm, and 10 days of incubation (FIG. 2).

5. Results

Table 1 shows the results of analyzing the physicochemical properties of the cultivated mycelium mycelium culture medium according to the composition of overlying barley saccharification medium, industrial medium, and tofu. In general, the results of analyzing the mushroom mushroom mycelium culture cultured for 10 days using the industrial medium composition used for mushroom mycelium liquid culture, pH 3.26, acidity 0.1%, the sugar content was 1.0˚Brix. The mycelium content was 1.45 g / L and the extracellular polysaccharide content was 0.23 g / L, which was very low. It was judged that the mycelium of the mushroom mushroom spawn did not grow well and the extracellular polysaccharide production rate was not suitable for cultivating the mushroom mushroom mycelium in the industrial medium. Therefore, the barley was glycosylated and used in the culture medium for the flower mushroom liquid spawn and used for the culture of the flower mushroom mycelium.

Barley generally plays a role in synthesizing various hydrolytic enzymes and secreting them into starch endosperm from the neutrophil cell wall in response to gibberellic acid (GA3) in germline cells during germination, and these enzymes are α-amylase. , β-glucanase, β-glucanase, xylanase, protease, cellulase, etc. are involved. It is preferentially degraded by the action of cell wall degrading enzymes such as beta-glucanase and xylanase, thereby exposing the starch exposed to α-amylase and β-amylase. As it decomposes, saccharification proceeds. Therefore, various enzymes are produced during barley germination, saccharified and diluted to select conditions suitable for mycelial culture, which is considered to be good for the use of mycelium mushroom mycelial growth. The barley saccharified aqueous solution was diluted 3 times with water and adjusted to 6˚Brix to be used for the liquid culture of mycelium mushroom mycelium. This test was carried out by reporting that the water-soluble sugars are good at the mycelial growth under low sugar concentrations, which not only supply energy to form fruiting bodies but also continue to use them for forming cells.

As a result of analyzing the mushroom mycelium culture medium cultured for 10 days using barley saccharification medium, pH 4.34, acidity 0.08%, and 7.5˚Brix (Table 1). The mycelium content was 22.15 g / L and the extracellular polysaccharide content was 39.39 g / L. When centrifugal separation was carried out to measure the content of the mycelium mushroom mycelium, the contents of the barley and the mycelium mushroom mycelium were precipitated together as a medium component. The polysaccharide content was also high, and the polysaccharides contained in the barley were recovered together. As a result of measuring the protein content of the cultivated mycelium mycelium culture medium using industrial medium and overlying barley saccharification medium, it was 65.27 mg% and 1.68 mg%, respectively. In addition, the beta glucan content was measured by lyophilization of the culture broth, showing 4.98 (%, w / w) and 4.34 (%, w / w), respectively. In the present invention, the cultivated crushed mushroom barley mycelium liquid was cultured, but due to the difference between the medium material and the test strain, it was determined that the content of the physicochemical analysis was low. In addition, when the germination of the barley reported that the production of low molecular sugars contained in the barley tends to increase in viscosity, it was thought that the growth of mushrooms was suppressed under the influence of the high viscosity of the medium obtained by germinating saccharin. Therefore, the mushroom mushroom mycelium was cultured using tofu pure water, which is a by-product that can replace expensive existing medium.

Tofu pure used in the culture was 1% carbohydrate, 1% protein, 0% fat, and it was judged to be a favorable medium composition for mushroom culture due to high potassium content of 107.94 mg / 100 g, especially in minerals (Table 2). As a result of analyzing tofu pure water, pH 5.7, acidity 0.12%, and sugar content were 3.28˚Brix, water content was 97.71%, and solid content was 32.83 mg / ml. In view of the results of good mycelial growth in the range of 25 ℃, pH 5.0-6.0 in culture of the mushroom mushroom liquid, the pH of the tofu was determined to be favorable for the growth of mycelial mushroom. Therefore, the results of physicochemical analysis of the cultivated mycelium mushroom mycelium culture cultured at 25 ° C. for 10 days using a liquid medium containing only 2% of glucose as a carbon source to tofu pure water are shown in Table 1. The pH of the mushroom culture was 6.91, which was close to neutral, and the acidity was 0.05% and the sugar content was 1.84˚Brix. The mycelium content of the blossom mushroom culture was 17.76 g / L, and the polysaccharide content was 0.35 g / L. The protein content of the cultivated mushroom culture was found to be 511.85 mg%. The protein content of the cultivated mycelium mushroom mycelium cultured using industrial medium and barley saccharification medium was higher than the results of 65.27 mg% and 1.68 mg%, respectively. In addition, beta glucan content was measured to be 10.64 (%, w / w), which was higher than the results of 4.98 (%, w / w) and 4.34 (%, w / w) in the culture medium using industrial medium and barley saccharification medium, respectively. High. Therefore, after fermentation of acetic acid using culture medium using tofu pure water, physicochemical analysis and anti-oxidation were evaluated, and mushroom vinegar was developed as a functional product using well-being food material.

Chemical Composition in Blossom Mushroom Culture Restricted badge Crushed saccharification badge Tofu pure water badge pH 3.26 ± 0.01 4.34 ± 0.01 6.91 ± 0.11 Acidity (%) 0.10 ± 0.01 0.08 0.03 0.05 ± 0.01 Sugar (˚Brix) 1.0 ± 0.10 7.5 ± 0.01 1.84 0.45 Mycelium Content (g / L) 1.45 ± 0.35 22.15 ± 0.21 17.76 ± 0.63 Polysaccharide Content (g / L) 0.23 + 0.11 39.39 ± 0.08 0.35 ± 0.05 Water Soluble Protein Content (mg%) 65.27 ± 14.85 1.68 ± 5.68 511.85 ± 9.54 β-glucan content (%, w / w) 4.98 ± 0.14 4.34 ± 0.72 10.64 ± 1.25

Cultures were obtained by fermentation for 10 days.

Chemical Composition of Soybean Curd Whey Tofu Pure Test Item Test result calorie 10 Kcal / 100 g carbohydrate
carbohydrate 2 g / 100 g (1%,% Nutrient Standard) sugars 1 g / 100 g protein 1 g below / 100 g (1%,% Nutrient Standard) Fat


Fat 0 g / 100 g (0%,% Nutrient Standard) Saturated fat 0 g / 100 g (0%,% Nutrient Standard) trans fat 0 g / 100 g cholesterol 0 mg / 100 g (0%,% Nutrient Standard) Dietary Fiber 0 g / 100 g (0%,% Nutrient Standard) mineral




Mg 13.646 mg / 100 g Ca 25.081 mg / 100 g K 107.942 mg / 100 g Na 50 mg / 100 g (3%) Mn 0.026 mg / 100 g P 8.294 mg / 100 g

< Example  2> Sucrose ( Sucrose ) Tofu of Blossom Mushrooms According to Concentration Pure water  Culture and Acetic Acid Fermentation

1. Acetic Acid Culture

Gluconacetobacter with Acetic Acid Bacteria hansenii ) KCCM 40230 was used by Korea Microbial Conservation Center. The medium composition used for the culture was mixed with 25 g / L of mannitol, 5 g / L of yeast extract, and 3 g / L of peptone. The acetic acid bacteria were used at a temperature of 30 ° C. and a rotation speed of 200 rpm. , 3 days or more culture was used (Fig. 3).

2. pH  And pH measurement

pH was measured by using a pH meter (Digital pH meter 420A +, Thermo Orion. Beverly, MA., USA), was taken by taking 10 mL. The titratable acidity was titrated with 0.1 N-NaOH until the pH reached 8.3 with a pH meter and the consumption was converted into tartaric acid content (%, v / v).

3. Viable cell count  Measure

Acetic acid bacteria cultured in mannitol medium sterilized at 121 ° C. for 15 minutes in 10 ⁴ , 10 ⁵ , 10 ⁶ times dilutions were diluted in 20 μL in mannitol agar medium and then incubated at 30 ° C. for at least 3 days. The number of viable cells cultured is shown as CFU / mL.

4. Determination of sugar content and solid content

The sugar content of the culture solution was measured by using an electron sugar meter (POCKET REFRACTOMETER, 0-93 ° Brix). Solid content was shown by measuring the weight of the solid content remaining after drying the moisture contained in 5 mL of the culture medium using an infrared moisture meter (KETT ELECTRIC LABORATORY, FD-720).

5. Mycelia and Polysaccharide Content Analysis

The purpose of mycelial liquid culture is to produce high yield of extracellular polysaccharides. In order to determine the content of mycelia and extracellular polysaccharides contained in the mycelium mycelium culture medium, 200 mL of the culture solution was taken and centrifuged (7,000 rpm, 20 min) using a centrifuge to separate the precipitate and the supernatant. The precipitate was washed twice with distilled water as a mycelium component and lyophilized and weighed. The separated supernatant was refrigerated by adding 200 mL of 95% ethanol to separate extracellular polysaccharides, and then stored overnight to recover the precipitated polysaccharides. After washing twice with distilled water and freeze-dried was weighed.

6. Determination of Water Soluble Protein Content

The water-soluble protein content of the mushroom mushroom culture was measured using a protein assay kit (SMARTTM BCA, Intron Biotechnology INC., Sungnam, Korea), and was calculated using bovine serum albumin (BSA) as a standard.

7. Beta Glucan  Content measurement

100 mg of the lyophilized powder of the mushroom mycelium culture was quantified and the content of beta glucan was measured using a Megazyme kit (K-BGLU, Megazyme International Ireland Ltd, wicklow, Ireland). The total glucan content was added 1.5 mL of 37% hydrochloric acid to 100 mg of the sample, followed by 45 minutes of reaction at 30 ° C. 10 mL of distilled water was added to the reaction solution and heated at 100 ° C. for 2 hours. Then, the volume was adjusted to 100 mL using 10 mL of 2 M KOH and 200 mM sodium acetate buffer (pH 5.0). After centrifugation at 1,500 × g for 10 minutes, 0.1 mL of the collected supernatant was taken, and 0.1 mL of β-glucosidase was added thereto and reacted at 40 ° C. for 60 minutes. After the reaction, 3 mL of GOPOD reagent was added, and the absorbance was measured at 510 nm after 20 minutes at 40 ° C. The alpha glucan content was added to 100 mg of the sample and 2 mL of 2 M KOH, followed by cooling for 20 minutes, followed by 8 mL of 1.2 mM sodium acetater buffer (pH 3.8) and amyloglucosidase (1,630). U / mL) 0.2 mL was added and reacted at 40 degreeC for 30 minutes. 0.1 mL of the supernatant obtained by centrifugation at 1,500 × g for 10 minutes was taken, and 3 mL of GOPOD reagent was added thereto and reacted at 40 ° C. for 20 minutes. The absorbance of the reaction solution was measured at 510 nm, and the difference between total glucan and alpha glucan content was calculated as beta glucan content.

8. Results

Sucrose was added to the tofu pure water and distilled water at 1, 5, 10 and 15% (w / v), respectively. Soybean flour 0.5%, KH ₂ PO ₄ 0.1%, MgSO ₄ 0.05% Each was added and analyzed using a 500 mL Erlenmeyer flask with a temperature of 25 ° C., a rotational speed of 160 rpm, and 30 days of incubation. A culture medium containing 15% sucrose was added to the tofu pure as an optimum condition, and the culture period was sampled at 10, 15, 20, 25, and 30 days at 5 day intervals for analysis and acetic acid fermentation. 7% (w / v) of alcohol was added to the culture solution and 10% inoculation of acetic acid bacteria was performed for the second fermentation of acetic acid. As a culture condition, a 250 mL Erlenmeyer flask was used for acetic acid fermentation for 30 days at a temperature of 30 ° C. and a rotational speed of 200 rpm for 7 days.

As shown in Figure 4, the pH of the cultivated mycelium mycelium culture medium was first cultured for 30 days in a medium to which 1, 5, 10, 15% sucrose was added to the tofu pure water and distilled water by concentration As a result, the culture solution added sucrose 1% to tofu pure water was 5.24, and the culture solution added sucrose 1% to distilled water was 5.65. There was no difference between tofu pure water and distilled water, and sucrose in tofu pure water. 5% of the culture solution was added to 5.21, sucrose (10% sucrose) was added to the culture medium was 5.22, there was no significant difference according to the sucrose (sucrose) concentration. The sugar content increased from 3.8 ˚Brix to 15.4 ˚Brix, respectively, as the concentration of sucrose contained in tofu puree increased. Distilled water was also increased to 1.1 ˚Brix, 4.9 ˚Brix, 9.5 ˚Brix, and 11.3 ˚Brix as sucrose was added by 1, 5, 10, and 15%. However, the sweetness of the tofu puree itself is about 3˚Brix, so the sugar content of the medium containing 5% sucrose in tofu puree is 8.0˚Brix and the medium added with 5% sucrose in distilled water. As the sugar content was 4.9 ˚ Brix, the medium added sucrose to the tofu was found to have higher sugar content. In addition, 1, 5, 10, and 15% sucrose was added to tofu pure water and distilled water, respectively, and the mycelia and polysaccharide contents were higher. Among them, 19.8 g / L of culture medium containing 15% sucrose in tofu pure water and 12.1 g / L of culture medium containing 15% sucrose in distilled water were found. The culture solution containing 15% sucrose in silver tofu pure was 5.87 g / L, and the culture solution containing 15% sucrose in distilled water was 2.93 g / L. The culture medium containing% was selected as the optimal condition, and the culture period was sampled at 10, 15, 20, 25, and 30 days at intervals of 5 days for analysis and acetic acid fermentation.

Chosangyun used in the acetic acid fermentation received pre-sale at the Korea Culture Center of Microorganisms gluconate acetonitrile bakteo century you (Gluconacetobacter hansenii ) KCCM 40230 strains were cultured at temperatures of 30 ° C, rotational speed of 200 rpm, for 3 days or more. As a result of physicochemical analysis, pH 5.53, acidity 0.04%, and the number of viable cells were 1.5 × 10 ¹¹ CFU / mL.

The head impurities sucrose (sucrose) at 15%, soybean meal (soybean flour) 0.5%, KH 2 PO 4 0.1%, MgSO 4 0.05% to each addition using a 500 mL flask temperature 25 ℃, rotation speed 160 rpm The cultured mushroom mushroom mycelium cultures were sampled and analyzed for 30 days at intervals of 10 days to 5 days for the culture period, as shown in FIG. 5. As the incubation period increased, the pH tended to increase to 5.34 at 10 days and 6.26 at 30 days. The acidity and sugar content did not differ according to the incubation period, and the mycelia content was not significantly different from 21.2 g / L in 10 days and 23.0 g / L in 30 days.

Secondary acetic acid fermentation was performed by adding 7% alcohol (w / v) to the culture medium sampled according to the culture period and inoculating 10% acetic acid bacteria. As a culture condition using a 250 mL flask temperature 30 ℃, rotational speed 200 rpm, for 7 days by acetic acid fermentation for 7 days sampled and analyzed as shown in FIG. As a result of pH measurement, the acetate culture fermented for 10 days was 5.31, the acetate fermentation for 5 days was 4.07, the acetate fermentation for 7 days tended to gradually decrease to 3.71. Acidity also increased to 0.30% for 3 days of acetic acid fermentation and to 1.30% for 7 days of acetic acid fermentation. However, after acetic acid fermentation for 3, 5 and 7 days depending on the culture period of the mycelium mycelium culture medium, the pH and acidity were not different according to the culture period of the mushroom mushroom culture.

< Example  3> Tofu of Blossom Mushrooms According to Potato Powder Concentration Pure water  Culture and Acetic Acid Fermentation

500 mL triangle by adding 0, 1, 3, 5% (w / v) potato powder to 0.5% soybean flour, 0.1% KH ₂ PO ₄ , 0.05% MgSO ₄ The flask was incubated for 30 days at a temperature of 25 ° C., a rotational speed of 160 rpm, and subjected to analysis and acetic acid fermentation. 7% (w / v) of alcohol, 10% (w / v) of jujube concentrate were added to the culture, and 10% inoculation of acetic acid bacteria was inoculated to perform secondary acetic acid fermentation. As a culture condition, a 250 mL Erlenmeyer flask was used for acetic acid fermentation for 30 days at a temperature of 30 ° C. and a rotational speed of 200 rpm for 7 days.

As shown in FIG. 7, as a result of pH measurement, the culture medium without the potato powder was 5.09 and the culture medium with the potato powder 5% showed 4.53, and the pH was slightly decreased as the potato powder was added for each concentration. In addition, the acidity was 0.26% in the medium without potato powder and 0.43% in the medium with the potato powder 5%. As the potato powder concentration increased, the acidity increased. The sugar content was slightly increased to 3.4 ˚Brix without potato powder and 4.9 ˚Brix with 5% potato powder. The mycelium content was 6.3 g / L for 1% potato powder and 18.7 g / L for 5% potato powder. This is thought to be very high content when the culture medium is centrifuged when measuring the content of the flower mushroom mycelium, the potato powder and the flower mushroom mycelium precipitated together as a medium component.

Fermented 7% (w / v) alcohol, 10% (w / v) jujube concentrate, and 10% inoculated acetic acid bacteria were inoculated to the cultivated mushroom mycelium culture medium according to the potato powder concentration. As a culture condition, a 250 mL flask was sampled at a temperature of 30 ° C., a rotational speed of 200 rpm, for 7 days with acetic acid fermentation for 2 days, and the results of analysis were shown in FIG. 8. As a result of pH measurement, the pH of potato powder after 3 days of fermentation was 4.88, 1% of potato powder was 4.67, 4.74 of potato powder was 4.74, and 4.59 of potato powder was 4.59. . In addition, 1% of 5% fermented potato powder was 4.68, and 1% of 7% fermented potato powder was 4.66. The results of acidity and sugar were not different according to potato powder concentration and fermentation period.

< Example  4> Glucose  And Rice bowl  Tofu with added mushrooms Pure water  Culture and Acetic Acid Fermentation

One. DPPH Radical  Measurement of scavenging activity

Scavenging activity against DPPH (1,1-diphenyl-2-picrylhydrazyl) radicals was measured according to Blois's method. 160 μL of the diluted solution by concentration and 40 μL of 0.15 mM DPPH solution dissolved in ethanol were added thereto, and the absorbance was measured at 517 nm after being allowed to stand at room temperature for 30 minutes. At this time, in order to compare the relative activity, Ryu Chung Hyun Medicinal Mushroom (Andong, Korea) was used to purchase the situation brown vinegar (100% of mushroom brown rice mycelium) and Matsutake vinegar (100% of mushroom fungus mycelium).

2. ABTS Radical  Measurement of scavenging activity

Antioxidant activity was measured by ABTS + cation decolorization assay. Absorbance value was 0.70 at 732 nm after formation of 2.45 mM potassium persulfate with 7 mM 2,2-azino-bis (3-ethylbenzthiazoline-6-sulfonic acid) (ABTS, Sigma Chemical Co., USA) Diluted with phosphate buffer saline (PBS, pH 7.4) to (± 0.02). 20 μL of the sample was added to 180 μL of the diluted solution, and the absorbance was measured after standing for exactly 1 minute. At this time, in order to compare the relative activity, Ryu Chung Hyun Medicinal Mushroom (Andong, Korea) was used to purchase the situation brown vinegar (100% of mushroom brown rice mycelium) and Matsutake vinegar (100% of mushroom fungus mycelium).

3. Results

Tofu pure 2% glucose, rice bran 5%, soluble starch 1%, soybean flour 0.5%, yeast extract 0.2%, KH ₂ PO ₄ 0.1%, MgSO ₄ Each of 0.05% was added and the temperature was 25 ° C. using a 5 L bottle, and the air was injected with 0.01 MPa using Air-pump of Gosung Valve Co., Ltd. Alcohol culture 7% (w / v), jujube concentrate 10% (w / v), citric acid (citric acid) 0.1% (w / v) was added, and 10% inoculation of acetic acid bacteria was fermented by the second acetic acid. As a culture condition, the temperature was 30 ° C., air was artificially injected to 0.01 MPa, and acetic acid fermentation was performed for 7 days, and then sampled at 2 days intervals for analysis.

The physicochemical analysis of the cultivated mycelium mycelium culture on the 10th day showed pH 5.06, acidity 0.18%, and sugar content 5.8 ˚Brix. The mycelium content was 30.8 g / L and the polysaccharide content was 3.45 g / L. The high mycelial content was thought to be very high due to the precipitation of rice bran and mycelium mycelium as a medium component when centrifugation of the culture medium when measuring the content of the mycelium mycelium.

The acidity of the mushroom mushroom mycelium cultured by adding sucrose and potato powder to the tofu pure water by concentration was about 1%. Since the acidity is very low compared to 4% or more commercially available vinegar, glucose and rice bran were added to the tofu pure in this experiment. Acid production of acetic acid bacteria was added by adding citric acid solution during fermentation of acetic acid. The jujube concentrate was added to increase the texture. The 20% solution of citric acid used was pH 1.31, acidity 20.68%.

Fermented 7% (w / v) alcohol, 10% (w / v) jujube juice, 10% (w / v) citric acid, 0.1% (w / v) citric acid, and 10% inoculated acetic acid bacteria to fermentation of acetic acid Was done. The culture conditions were 30 ℃, air was artificially injected to 0.01 MPa, and after 7 days of acetic acid fermentation was sampled every 2 days for physicochemical analysis and antioxidant evaluation. Acetic acid fermentation for 3 days resulted in pH 3.62, acidity of 2.9%, and sugar content of 16.1 ˚Brix. Acetic acid fermentation for 5 days resulted in pH 3.62, acidity 3.33%, and sugar content of 17.2 ˚Brix, and for 7 days acetic acid fermentation showed pH 3.69, acidity 3.62%, and sugar content of 19.4 ˚Brix. As the fermentation period of acetic acid continued, acidity increased.

DPPH radical scavenging activity experiment was conducted to investigate the antioxidant capacity of vinegar prepared by acetic acid fermentation of the mushroom mushroom mycelium culture cultured by adding glucose and rice bran to the tofu pure. Since DPPH (1,1-diphenyl-2-picryl-hydrazyl) receives electrons and hydrogen from an antioxidant, and forms an irreversibly stable molecule, antioxidant activity can be estimated from electron donating ability. The DPPH method is characterized by bioactive substances that exhibit antioxidant activity such as tocopherol, ascorbate, flavonoid compounds, aromatic amines, maillard browning products, and peptides. It is known that the antioxidant effect is measured by electron donating ability according to the degree to which the dark purple color is reduced by reduction, and it is known as a method generally used for the search for antioxidants. Therefore, after diluting the vinegar prepared by acetic acid fermentation of the mushroom mycelium mycelium culture medium using DPPH radical scavenging activity 5 times, the sulfated power is measured as shown in FIG. 9. At this time, it was used as a control for comparison with the purchased mushroom vinegar (100% of mushroom mushroom brown mycelium) and pine mushroom vinegar (100% mushroom mushroom mycelium) purchased. Physicochemical analysis of two commercially available vinegars showed that the pine mushroom vinegar had a pH of 2.83, an acidity of 5.6%, and a sugar content of 4.9 ˚Brix. The situation mushroom vinegar had a pH of 3.03, an acidity of 5.0%, and a sugar content of 4.9 ˚Brix. As a result of DPPH measurement, the inhibition rate was 72% in the mushroom mycelium culture medium before fermentation, 70.25% in 3 days fermentation, 69.13% in 5 days fermentation, and 77.25% in 7 days fermentation. There was no difference in inhibition rate during acetic acid fermentation. Pleurotus is also available on the market eryngii ) Vinegar is 36.22%, Phellinus baumii ) showed 39.14% of electron donating ability, and the scavenging activity of the vinegar was much lower than that of the flower mushroom vinegar. Therefore, it is determined that the anticancer mushroom vinegar has higher antioxidant activity than commercial pine mushroom vinegar and situation mushroom vinegar.

In addition, the antioxidant activity of vinegar prepared by acetic acid fermentation was investigated using ABTS + radical scavenging activity. ABTS radical scavenging ability, when ABTS + potassium persulfate is left in the cow to generate ABTS + radicals, the ABTS + radicals are scavenged by the antioxidant power of the sample to decolorize the blue-green color of the radicals. Activity can be measured. Therefore, after diluting 10-fold dilution of vinegar prepared by acetic acid fermentation of mycelium mushroom mycelium culture medium using the ABTS radical scavenging activity method, the antioxidant power was measured as shown in FIG. 10. At this time, it was used as a control for comparison with the purchased mushroom vinegar (100% of mushroom mushroom brown mycelium) and pine mushroom vinegar (100% mushroom mushroom mycelium) purchased. As a result of the ABTS measurement, the inhibition rate of 76.48% of the mushroom mycelium culture medium before the fermentation of acetic acid, 93.44% of the culture solution for 3 days of acetic acid fermentation, 92.75% for the culture solution for 5 days of acetic acid fermentation, and 91% of the culture solution for 7 days of acetic acid fermentation. As the fermentation of acetic acid progressed, the inhibition rate increased. In addition, commercially available Pleurotus eryngii vinegar is 9.98%, Phellinus baumii ) vinegar showed 22.83% of electron donating ability, and the scavenging activity was much lower than that of the flower mushroom vinegar. Therefore, the results of the DPPH and ABTS measurements showed that the matsutake vinegar produced by the anti-oxidant was much higher than the commercially available matsutake vinegar and situation mushroom vinegar.

Claims (6)

Cultivating the cauliflower mycelium on tofu pure medium containing 1 to 10 wt% glucose, 1 to 15 wt% sucrose or 1 to 5 wt% potato powder, based on the total tofu pure; And
Acetic acid bacteria inoculated in the mushroom mushroom mycelium culture, and fermented acetic acid fermentation step by adding 3 to 9% by weight based on the total culture broth mushroom mushroom vinegar manufacturing method. delete delete The method according to claim 1, wherein the acetic acid bacteria is gluconacetobacter hansenii ( Gluconacetobacter hansenii ) characterized in that the mushroom mushroom vinegar manufacturing method. The method according to claim 1, wherein the acetic acid fermentation is a mushroom mushroom vinegar production method characterized in that the fermentation for 3 to 7 days at 20 to 40 ℃. Flower mushroom vinegar prepared by the method of claim 1, 4 or 5.

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