KR102282725B1 - Weissella cibaria strain having rice cake anti-retrogradation activity and use thereof - Google Patents

Abstract

The present invention relates to a Weissella ciberia strain having an activity to inhibit aging or spoilage of food using rice, and the Weissella cibaria JNU 29 [Accession No.: KCTC14070BP] strain has excellent extracellular polysaccharide production. In addition, it is possible to secure the maximum storage properties by simultaneously suppressing the aging and spoilage of the rice cake. In addition, the Weissella cibaria JNU 29 strain is a lactic acid bacterium derived from kimchi. Since it is non-toxic in the body, it can be used as food and is very stable in gastric juice or bile, so the consumption of processed rice cake products using it. Various functionalities can also be provided.

Description

Weissella ciberia strain having anti-aging and decay inhibitory activity and use thereof {Weissella cibaria strain having rice cake anti-retrogradation activity and use thereof}

The present invention relates to a Weissella siberia strain having an activity of inhibiting aging or spoilage of food using rice, a composition for inhibiting aging and spoilage using the same, and processed rice food with improved storage properties.

Tteok is a unique Korean food that has been developed over a long period of time along with the development of agriculture of our people. There are hundreds of types. A general method of making rice cakes is prepared by immersing grains in water, then kneading the grains after removing water and kneading them and increasing them by extrusion or punching.

Although the location of rice cakes has withered in the past due to the westernization of eating habits, interest in rice cakes is gradually increasing as the nutritional superiority and stability of rice cakes and the convenience that they can be used as a meal substitute have been re-examined recently. However, in the case of rice cakes, there is a big problem in storage and distribution. Specifically, due to the aging phenomenon of rice cakes, the statutory shelf life is limited to one day, and even if they are not aged, they are easily spoiled by mold and bacteria, so it is difficult to store them for a long period of time.

In order to solve the above problems, a method for producing rice cakes with oriental medicinal extract added (Patent Document 1) has been known so as to maintain flexibility and preservation for a long period of time, but this has a strong bitter taste and flavor due to oriental medicinal extracts, and has a unique taste of rice cakes. There is a problem that you cannot feel the .

On the other hand, in the method for producing rice cake dough (Patent Document 2) for prolonging the shortening of the shelf life by suppressing deterioration and aging of starch in the state that no additives are used, the aging of the rice cake manufactured through the above invention may be delayed. However, there is a problem in that it cannot be applied to mass production because the conditions of the manufacturing process are sensitive, so a lot of initial equipment cost must be invested, and it can be applied only to a small capacity. In addition, disadvantages such as a decrease in the texture of the rice cake itself are known.

Therefore, the present inventors have made an earnest effort to develop a method to improve the above-described problems, and as a result, the present inventors have found a new strain capable of imparting various functions while suppressing aging and spoilage of rice cake among numerous lactic acid bacteria. By selection and identification, the present invention was completed.

Patent Literature 1. Korean Patent Registration No. 10-1063973 Patent Document 2. Republic of Korea Patent Registration No. 10-1332580

The present invention has been devised in view of the above problems, and an object of the present invention is to provide a Weissella cibaria JNU 29 [Accession No.: KCTC14070BP] strain having anti-aging and spoilage-inhibiting activity of rice cakes.

Another object of the present invention is to provide an extracellular polysaccharide (exopolysaccharide) produced from Weissella cibaria JNU 29 [Accession No.: KCTC14070BP] strain.

In addition, another object of the present invention is the Weissella cibaria JNU 29 [Accession No.: KCTC14070BP] strain-derived extracellular polysaccharide (exopolysaccharide) containing as an active ingredient a composition for inhibiting aging of rice cakes and a composition prepared through the to provide bread.

The present invention provides a strain of Weissella cibaria JNU 29 [Accession No.: KCTC14070BP] having anti-aging and decay-inhibiting activity of rice cakes in order to achieve the above object.

The strain may have a 16S rRNA represented by SEQ ID NO: 1.

The strain may be cultured under conditions containing sucrose as a carbon source.

In order to achieve the above other object of the present invention, it provides an extracellular polysaccharide (exopolysaccharide) produced from Weissella cibaria JNU 29 [Accession No.: KCTC14070BP] strain.

The extracellular polysaccharide may be prepared by culturing Weissella cibaria JNU 29 [Accession No.: KCTC14070BP] strain in a medium containing sucrose as a carbon source.

In addition, the present invention provides a composition for inhibiting aging and spoilage of processed rice containing a Weissella cibaria JNU 29 [Accession No.: KCTC14070BP] strain or a culture solution thereof as an active ingredient in order to achieve the above other object do.

In addition, the present invention provides a rice processed food with improved storage properties, including the Weissella siberia JNU 29 strain in order to achieve the above other object.

The rice processed food may further include sucrose.

According to the present invention, the Weissella cibaria JNU 29 [Accession No.: KCTC14070BP] strain not only has excellent extracellular polysaccharide production, but also can maximize storage by inhibiting the aging and decay of rice cakes at the same time. . In addition, the Weissella cibaria JNU 29 strain is a lactic acid bacterium derived from kimchi. Since it is non-toxic in the body, it can be used as food and is very stable in gastric juice or bile, so the consumption of processed rice cake products using it. Various functionalities can also be provided.

Figure 1a is a process flow chart schematically showing the manufacturing process of the rice cake using the microorganism Weissella cibaria JNU 29 according to the present invention.
Figure 1b is a photograph showing each manufacturing process of the rice cake using Weissella cibaria JNU 29.
2 is a result of primary selection of colonies forming mucilage from among 520 isolated strains.
3 is a representative phenotype of strains having the ability to produce extracellular polysaccharides selected for the first time.
4A and 4B show the results of TLC analysis of a culture medium in which 40 strains of the second selected from Experimental Example 2 were cultured in an MRS medium, and a culture medium in which sucrose was added to the MRS medium. 'Std 1' means Standards 1, which has glucose (glucose, G) and maltose (M) spots. 'Std 2' means Standards 2 and has fructose (F) and sucrose (S) spots. 'Strain number' is the culture medium cultured in MRS medium, and 'strain number + s' is the culture medium cultured in MRS medium to which sucrose is added. 'M' is MRS liquid medium, and 'M-S' is MRS liquid medium to which sucrose is added.
5 shows the results of TLC analysis of the culture medium in which six strains (JNU9, JNU29, JNU224, JNU265, JNU465, JNU506) were cultured in sucrose-added MRS broth. 'IMO' stands for Isomalto-oligosaccharide, 'MO' stands for Malto-oligosaccharide, 'M' stands for maltose (M), 'S' stands for sucrose (S), and 'G' stands for glucose ( glucose, G) and 'F' are fructose (F) spots. Strain number' is a culture medium cultured in MRS liquid medium to which sucrose is added.
6 is after culturing 6 strains (JNU9, JNU29, JNU224, JNU265, JNU465, JNU506) in MRS medium to which sucrose is added, the phenotype of the strain and the viscosity of the culture medium were visually confirmed.
Figure 7 is after culturing each of the four strains (JNU29, JNU224, JNU465, JNU506) in the normal MRS medium and sucrose-added MRS medium, the phenotype of the strain and the viscosity of the culture medium were visually confirmed.
8 is a graph showing the number of viable cells counted when five strains (JNU9, JNU29, JNU224, JNU465, JNU506) were added to artificial gastric juice, 10% sucrose-added artificial gastric juice, and controls 1 and 2 and cultured.
9 is a graph showing the number of viable cells counted when cultured by adding 5 strains (JNU9, JNU29, JNU224, JNU465, JNU506) to artificial bile, 10% sucrose-added artificial bile, and controls 1 and 2. FIG.
10 is a result of measuring the viscosity of the culture medium of five strains (JNU9, JNU29, JNU224, JNU465, JNU506).
11 is a TLC analysis result of extracellular polysaccharides generated from the three strains finally selected (JNU 9, JNU 29, JNU 506).
12 is a TLC analysis result for extracellular polysaccharides generated from the three strains finally selected (JNU 9, JNU 29, JNU 506).
13 to 16 are HPLC analysis results for extracellular polysaccharides generated from the three finally selected strains (JNU 9, JNU 29, JNU 506).
17 is a graph showing the time-dependent hardness change of seolgitteok prepared in Example 2 and Comparative Examples 1 and 2 was measured.
18 is a photograph showing the degree of decay according to time (1 day, 7 days) when the seolgitteok prepared in Example 2 and Comparative Examples 1 and 2 was stored at room temperature.

Hereinafter, various aspects and various embodiments of the present invention will be described in more detail.

The present invention provides a strain of Weissella cibaria JNU 29 [Accession No.: KCTC14070BP] having anti-aging and decay inhibitory activity of rice cake.

After collecting traditional fermented foods from each household in Jeollanam-do, 520 species of bacteria were isolated from them. As a result of confirming the viscous material-forming ability of the 520 types of bacteria, a total of 262 strains were first selected. As a result of selecting strains exhibiting extracellular polysaccharide production ability among them, 5 strains ( W. cibaria JNU 9, W. cibaria JNU 29, B. amyloliquefaciens JNU 224, B. stratosphericus JNU 465, B. subtilis JNU 506) were selected. was selected. Since the strain has various physiological functions by extracellular polysaccharide, it is preferable to reach the intestine. Therefore, as a result of confirming the resistance to gastric juice and bile, it was confirmed that the W. cibaria JNU 29 strain had high resistance to both gastric juice and bile. In addition, as a result of analyzing the viscosity and extracellular polysaccharide production ability of the culture medium, when sucrose was added as a carbon source, the same genus had significantly superior extracellular polysaccharide production ability compared to W. cibaria JNU 9, and heat resistance when added to rice cakes In addition, it can significantly inhibit the aging and spoilage of rice cakes, so Weissella siberia The JNU 29 strain was finally selected. In general, the Bacillus subtilus strain produces a lot of mucilage and was known to be very useful in vivo, but in the present invention, Weissella siberia rather than Bacillus subtilus JNU 506 strain. It was confirmed that the JNU 29 strain not only produced more extracellular polysaccharides, but also showed a lower rate of increase in the hardness of rice cakes over time even when making rice cakes. there is.

The Weissella siberia As a result of extracting 16S rRNA from JNU 29 strain and analyzing the nucleotide sequence, it was confirmed that it belongs to Weissella cibaria. The strain thus obtained was deposited at the Korean Collection for Type Cultures and was given an accession number of KCTC14070BP. The strain is characterized in that it has 16S rRNA represented by SEQ ID NO: 1.

Weissella siberia according to the present invention The JNU 29 strain is preferably derived from kimchi, but is not limited thereto.

Weissella siberia of the present invention JNU 29 strain provides exopolysaccharide. The extracellular polysaccharide produces a high concentration of the extracellular polysaccharide of homopolysaccharide composed of glucose, which can be clearly confirmed through TLC and HPLC.

As used herein, the term "extracellular polysaccharide (EPS)" refers to a substance that is part of the cell wall of microorganisms such as fungi, and the polysaccharide is secreted out of the cell to form a capsule around it, or as a viscous substance, which is secreted around the cell or into the medium. . The extracellular polysaccharide is known to be secreted by microorganisms to protect themselves from external environments such as antibodies, toxic substances, protozoa and bacteriophages.

The Weissella siberia The JNU 29 strain is preferably cultured under conditions including sucrose as a carbon source, because extracellular polysaccharides are produced only when sucrose as a carbon source is included. Preferably, the sucrose is contained in a concentration of 2 to 20 g/l, more preferably 5 to 15 g/l, and even more preferably 9 to 11 g/l.

The Weissella siberia JNU 29 strain, the suspension of the strain, and the culture solution of the strain can inhibit or delay aging and spoilage of the product when added to processed rice products such as rice cakes. In addition, the Weissella siberia Since the JNU 29 strain has heat resistance and does not die during the manufacturing process of rice cakes, it can not only maintain storage stability in the short term, but also deliver the functional effect of extracellular polysaccharides to the intestines. It has the great advantage of being able to

That is, the Weissella siberia according to the present invention JNU 29 strain adds sucrose (sugar) to rice or a pulverized product thereof, and Weissella siberia By inoculating the JNU 29 strain, or a culture solution of the strain, it is possible to produce food having an effect of simultaneously inhibiting aging and spoilage of food using rice.

In terms of functionality and nutrition, various foods such as rice cakes have been developed, but the shelf life is very short (1 day), which causes great problems in storage and distribution, limiting market expansion. In other words, long-term storage of rice cakes is difficult, and it is susceptible to mold and bacteria, which leads to rapid decay, and even if spoilage does not progress, aging progresses in which the hardness of rice cakes, which greatly affects taste and texture, increases over time. It is difficult to store for a long time. To solve this, various technologies have been discussed, and most of these relate to new methods for packaging rice cakes, or to processing rice cakes to enable long-term storage by sterilization. The above-described methods have disadvantages in various processes, such as cumbersome and costly manufacturing, and difficult to manage.

Most rice processed foods use fermentation technology only to improve functionality or to give various flavors and textures. Since it is focused on solving a single problem, it does not deal with improving the functionality of various foods. Therefore, there are some difficulties in practical application.

Accordingly, the present inventors have discovered a new strain having an activity that can suppress not only decay but also aging so that it can be stored for a long time by extending the shelf life of rice processed foods such as rice cake, and through this, long-term storage is significantly increased, and various physiological As it was confirmed that it is practically possible to manufacture rice-processed foods such as rice cakes and the like having a function, the present invention has been completed.

That is, the present invention prevents/prevents/delays aging and spoilage of processed rice foods through a simple process of adding a strain or a culture solution of a strain to rice, which is a raw material of rice cake, unlike various conventional techniques, and has heat resistance. It has a great advantage in that it survives in the manufacturing process and continues to show activity, and it has excellent effects on gastric juice and bile, so that it can even secure the functionality of the strain when ingesting food.

Furthermore, the Weissella siberia JNU 29 strain of the present invention, as well as the strain itself, suppresses excellent aging and spoilage of the strain suspension or the culture solution of the strain, and can impart various functionalities to food, so the range of application is wide. In addition, when sucrose is provided as a carbon source, a high concentration of extracellular polysaccharide is secreted. Therefore, a separate fermentation process is not required when making rice cakes, so it is possible to easily prepare rice cakes with improved storability and storability. In the rice or its pulverized product, 1 to 5 parts by weight of the strain is added based on 100 parts by weight of the rice or its pulverized product to have the best antibacterial activity.

The optimal fermentation condition of the strain is 0.1 to 5 days at 30 to 45 ° C., but is not particularly limited thereto, but the culture technique is simple because the fermentation temperature is not high and the growth conditions are not difficult in that the pH is also a neutral condition. cost can be reduced.

In addition, as confirmed in the experimental example to be described later, the Weissella cyberia JNU 29 strain according to the present invention can be added to rice or a pulverized product of which sucrose is added as a carbon source to directly prepare rice cakes, and the conventional JUN 9 of the same cognate It was confirmed that it was possible to significantly prevent aging and decay than when the strain was used. Specifically, the rice cakes prepared with the Weissella cyberia JNU 29 strain did not rot almost even on the 9th to 12th days, which increased 3 to 4 times more than the rice cakes prepared with the JNU 9 and JNU 506 strains, and the hardness, taste, and quality were also 7 It was confirmed that it was maintained until the day. On the other hand, it was confirmed that the rice cake prepared with JNU 9 and JNU 506 strains deteriorated rapidly even after 1 to 2 days, so that the hardness was high, and the taste and quality could not be maintained, and the preference was significantly lowered.

Therefore, the Weissella cibaria JNU 29 of the present invention is a use for enhancing the storability of food using rice, that is, a starter composition that can produce extracellular polysaccharides having various effects, such as storage and convenience, or It can also be used as a composition for food additives or a composition for inhibiting aging and spoilage of rice cakes for inhibiting aging and spoilage. The composition may include live cells, dried cells, and cultures of Weissella cyberia JNU 29 of the present invention as an active ingredient, and may further include an excipient or carrier. The culture includes the culture itself cultured in rice or a pulverized product thereof, and a processed culture obtained by filtering, drying, or diluting the culture. The content of the Weissella siberia JNU 29 strain in the composition may vary depending on the use and formulation of the composition, preferably 0.0001 to 50% by weight, more preferably 0.1 to 20% by weight relative to the total weight of the composition. , the content is not limited thereto, and the content may be appropriately adjusted according to the intended use.

The composition may be prepared and administered in various formulations and methods. For example, Weissella siberia JNU 29 strain or a culture thereof may be mixed with a carrier and flavoring commonly used in the food field.

The Weissella siberia JNU 29 strain or a culture thereof according to the present invention may be used as a food additive for various foods using rice (eg, rice cakes, rice crackers, porridge, etc.). In addition, the Weissella cyberia JNU 29 strain of the present invention can be used as a starter for the production of extracellular polysaccharide.

The processed rice food may be rice cakes, porridge, seonsik, noodles or bread, but is not limited thereto.

In addition, another aspect of the present invention relates to a rice processed food with improved storability comprising the Weissella siberia JNU 29 strain.

The rice processed food may be one containing the cells of Weissella cyberia JNU 29 strain. Containing the cells of the Weissella siberia JNU 29 strain may be the culture medium of the Weissella siberia JNU 29 strain.

In the rice processed food of the present invention, when sucrose is added together with rice or a pulverized product of rice, and the Weissella cyberia JNU 29 strain is inoculated thereto, the strain grows to produce extracellular polysaccharides, and the strain is heat resistant It can suppress aging and spoilage of processed rice foods for a long time by producing extracellular polysaccharides while continuously remaining in rice processed foods.

In addition, when Weissella cyberia JNU 29 strain or its culture solution is added than the effect that can be obtained by adding simple oligosaccharides or sugars, in addition to the remarkable effect of 1.5 to 2 times or more in the effect of inhibiting aging and decay, simply adding sugar Unlike the case, it is possible to obtain a sustained aging and decay effect, and a significantly increased effect can be obtained in that a useful function in the intestine can be obtained when food is ingested.

Since the Weissella cyberia JNU 29 strain produces extracellular polysaccharides of homopolysaccharide containing glucose as a constituent sugar, it was confirmed that it was effective in enhancing the storage properties of processed rice foods such as rice cakes and food compositions prepared using the Weissella cyberia JNU 29 strain. Therefore, the present invention can be applied as a processed rice food with improved storage properties including the Weissella siberia JNU 29 strain or culture medium as an active ingredient.

In the present invention, the processed rice food or food composition with improved storability means that the period of use as a food is maintained for a long time by delaying/suppressing/preventing aging and spoilage of the processed rice food when stored under the same conditions. The Weissella cyberia JNU 29 strain produces an extracellular polysaccharide containing homopolysaccharide containing glucose as a constituent sugar, thereby inhibiting the increase in hardness of processed rice food and inhibiting and delaying food spoilage, Since it is recognized as harmless, it satisfies all the conditions for functioning as an adjuvant for improving the storability of the food composition.

The content of the Weissella cyberia JNU 29 strain contained in the food composition may be appropriately selected in consideration of the purpose and method of use, and preferably 0.01 to 99 parts by weight, more preferably based on the entire composition. It may be 0.1 to 90 parts by weight.

In the present invention, food means a processed product that can be manufactured using rice, and preferably means a state that can be eaten directly through a certain processing process, and examples include chips, noodles, dairy products, and fermented foods. , grain food, microbial fermented food, confectionery, and the like, but is not limited thereto.

The food of the present invention is characterized in that it further contains sucrose as a carbon source of the strain in addition to the rice or pulverized product thereof as an essential ingredient in the indicated ratio, and there is no particular limitation on other ingredients other than that, and various flavors like ordinary food. It may contain an agent or carbohydrate as an additional component, but is not limited thereto. The sucrose may be added to raw materials in the production of foods, beverages, health drinks, gum, tea, vitamin complexes, health functional foods, or added by a method of appropriately mixing with cooked foods, and is a food-logically acceptable food supplement additive. can be added together with In this case, the amount of sucrose in the food or beverage may be added in an amount of 0.01 to 90% by weight of the total food weight, and is preferably included in a concentration of 2 to 20 g/l, more preferably 5 to 15 g/l, more More preferably, it may be 9 to 11 g/l.

Another aspect of the present invention relates to a method for producing processed rice food with improved storage properties by inoculating the Weissella cyberia JNU 29 strain. The manufacturing method may be described in more detail with reference to FIGS. 1A and 1B .

First, the rice is thoroughly washed, and then the pulverized product is obtained by primary grinding.

The rice is washed cleanly to remove foreign substances such as pests, dust, or stones. After immersing and soaking the washed rice in water (water immersion), remove the water and set aside. Thereafter, as necessary, it is used as it is or a pulverized product obtained by pulverizing is used. The drying is not particularly limited as long as it is a drying method of rice commonly used. The pulverized pulverized product can be obtained by pulverizing 80 to 100 mesh and passing it through a sieve having a mesh to pulverize the powder of a desired size.

If the size of the pulverized product of the rice is less than 80 mesh, it is better, but since it is difficult to pulverize it to less than 80 mesh with a general grinder, it takes a lot of cost to pulverize it to less than 80 mesh, and thus it is not economical. In addition, when it exceeds 100 mesh, the effect of pulverizing the rice cannot be obtained because the dietary fiber tissue of the rice is not destroyed. Here, as the grinding equipment, a grinding equipment for fine grinding, such as a ball mill, a colloid mill, and a micro air classified mill, may be used, but the present invention is not limited thereto.

The Weissella cyberia JNU 29 strain is inoculated into the prepared pulverized material. It is preferable to inoculate 1 to 5 parts by weight of the Weissella cyberia JNU 29 strain based on 100 parts by weight of the pulverized product, but when inoculated with less than 1 part by weight, a problem that the extracellular polysaccharide cannot be sufficiently produced may occur, 5 In case of inoculation in excess of parts by weight, chewy texture may decrease during the production of foods such as rice cakes later.

Weissella siberia in the pulverized product Before inoculating the JNU 29 strain, it is preferable to further add sucrose as a carbon source for effective production and growth of the extracellular polysaccharide. The sucrose is preferably contained in a concentration of 2 to 20 g/l, more preferably 5 to 15 g/l, and even more preferably 9 to 11 g/l.

In addition, in addition to the sucrose, a lactobacillus nutrient source such as a protein source, a carbohydrate source, a vitamin or a mineral for promoting its growth may be further added. As the lactic acid bacteria nutrient source, commercially available media may be used or only necessary nutrient sources may be individually added.

In addition, the pulverized product may further include functional additives such as yeast powder, malt powder, purified enzyme, and white rice flour, and specifically, 0.5 to 5 parts by weight of yeast powder, malt powder based on 100 parts by weight of the rice or its pulverized product. It is preferable to mix 0.1 to 1 parts by weight, 0.01 to 0.1 parts by weight of purified enzyme, and 10 to 30 parts by weight of white rice.

In addition, the pulverized product, or a mixture of the pulverized product and the lactic acid bacteria nutrient source is the Weissella siberia It can be heat-sterilized prior to inoculation with JNU 29 strain.

The strain is a Weissella cyberia JNU 29 strain isolated from kimchi, and is harmless to the human body. Even a strain of the Weissella genus isolated from kimchi cannot unconditionally produce extracellular polysaccharides with anti-aging and decay-inhibiting activity in rice or its pulverized product. This can be confirmed in the selection process of the experimental examples to be described later of the present invention.

That is, when the Weissella cyberia JNU 29 strain selected in the present invention is inoculated into rice or a pulverized product thereof, a high concentration of extracellular polysaccharide is produced. It was confirmed that the activity was easily lost at high temperature, there was no resistance to gastric juice and bile, or the storage period was not properly improved when the actual product was applied.

Since the Weissella cyberia JNU 29 strain has heat resistance, it is possible to produce rice cakes without going through a separate fermentation step. In particular, for the distribution of rice cakes, it is most preferable to use the culture medium of the Weissella cyberia JNU 29 strain in which extracellular polysaccharides are sufficiently produced.

The Weissella cyberia JNU 29 strain may be a culture medium, and the culture medium preferably contains cells. When the cells of the Weissella cyberia JNU 29 strain are removed, it is not preferable because extracellular polysaccharide production does not occur.

Thereafter, the pulverized rice is secondarily pulverized. At this time, pulverization may be a pulverizing method commonly used in the rice cake manufacturing process, and a desired pulverized product can be obtained by adjusting the size of the roller and the number of pulverization of the pulverizer according to the type of rice cake to be manufactured. Finally, after kneading the pulverized rice, it is steamed in steam at 85~100℃. At this time, the steaming time may vary depending on the type of rice cake to be prepared, but generally may be 1 to 1 hour. Before steaming with the steam, the rice dough is left at room temperature for 5 to 1 hour, Weissella siberia The extracellular polysaccharide from the JNU 29 strain can be sufficiently produced.

Weissella siberia according to the present invention In the case of manufacturing rice-processed food using JNU 29 strain, that is, rice cake, it is possible to manufacture rice cake with improved storability because aging or spoilage is suppressed, and mass production and long-term distribution are possible.

Hereinafter, the present invention will be described in more detail by way of Examples and the like, but the scope and content of the present invention may not be construed as being reduced or limited by the Examples below. In addition, based on the disclosure of the present invention including the following examples, it is clear that a person skilled in the art can easily practice the present invention for which no specific experimental results are presented, and such modifications and modifications are included in the attached patent. It goes without saying that they fall within the scope of the claims.

Example 1: Weissella cyberia JNU 29 strain culture

Weissella siberia JNU 29 was added to MRS (Difico Co., Detroit MI, USA) liquid medium with 10% (w/v) sucrose, inoculated with 3% of each liquid seed, and cultured for 24 hours at 37 ° C. (IL-21, Jeio tech., Daejeon, Korea) to culture the strain, and then the culture medium was used for the production of rice cake.

Example 2: Preparation of seolgitteok of Bacillus subtilus JNU515 strain.

First, the rice is washed to remove foreign substances such as pests, dust, and stones. The washed rice is soaked in water, and the soaking time can be adjusted appropriately considering the stickiness of the rice cake to be prepared. In this example, it was soaked for 2 hours. Drain the water from the soaked rice and dry it sufficiently (dehydration). 10 kg of dried rice was mixed with 120 g of salt and first milled to prepare rice flour. 1,800 g of the Weissella siberia JNU 29 strain culture solution prepared in Example 1 was mixed with 10 kg of the first pulverized rice flour, followed by second milling. Put 100 g of sugar in the pulverized mixed rice flour, sieve it through a 100 mesh sieve (sieving), and put the rice flour that passed through the sieve into a 260 mm×260 mm×20 mm mold, then cut into 9 equal parts of a certain size (molding) ). After leaving it at room temperature for about 1 hour, it was steamed (steamed) at about 100 ° C. for 30 minutes, cooled (cooled), cut along the knife edge, put in a mold, and packaged to prepare seolgitteok (packaging). The overall process thereof is shown in FIGS. 1A and 1B .

Comparative Example 1: Preparation of fermented rice flour of Weissella siberia JNU 9 strain.

Fermented rice flour was prepared in the same manner as in Example 1 except that the culture solution of the JNU 9 strain was used instead of the culture solution of the Bacillus subtilus JNU 29 strain.

Comparative Example 2: Preparation of seolgitteok of Bacillus subtilus JNU 506 strain.

Seolgitteok was prepared in the same manner as in Example 2 except that the culture solution of the JNU 506 strain was used instead of the culture solution of the Bacillus subtilus JNU 29 strain.

Experimental Example 1: Isolation and culture of strains from fermented products

Traditional fermented foods (raw materials) such as kimchi and soybean paste were collected from each household in Jeollanam-do, and strains were isolated from them. Specifically, the food (about 10 g) was diluted in sterile physiological saline, spread on the medium, and cultured at 37 ° C. for 1-2 days to isolate the strain (in the case of fermented broth or soy sauce, it was used as a sample). In this case, LB (BD, USA) for general bacteria, YM (MBcell, korea) for yeast, and MRS (MBcell, korea) for lactic acid bacteria were used. A total of 520 strains were purely isolated from a total of 69 raw materials into 222 common bacteria through LB medium, 191 yeast through YM medium, and 107 lactic acid bacteria through MRS medium.

Experimental Example 2: Selection of strains having viscous material-forming ability

1) 1st selection

In Experimental Example 1, strains producing extracellular polysaccharides (Exopolysaccharides, EPS) were selected from 520 strains isolated from pure water. Prepare LB, YM, MRS solid medium to which 10% (w/v) sucrose was added as a carbon source, and 20 μl of the isolated strain was added dropwise to each of the above solid medium and cultured at 37° C. for 2 days to colonize colonies. As shown in Table 1 below, 262 strains of slime bacteria showing viscosity were primarily selected.

division number of isolated bacteria Number of bacteria that were first selected with EPS-producing ability Fermented broth 24 6 (25%) Kimchi 235 132 (56%) Soybean paste 143 65 (45%) Red pepper paste 69 43 (62%) Soy sauce 43 12 (28%) Vinegar 6 4 (66%) Total 520 262

2 is a result of primary selection of colonies forming mucilage from among 520 isolated strains. 3 is a representative phenotype of strains having the ability to produce extracellular polysaccharides selected for the first time.

As shown in Table 1 and FIG. 2, when cultured in a solid medium using sucrose as a carbon source, it was confirmed that 262 strains formed a viscous material, and the strains were first selected. Specifically, the strains were cultured in a solid medium using 10% sucrose as a carbon source, and strains having a phenotype as shown in FIG.

2) 2nd selection

The 262 strains that were first selected were again cultured in a liquid medium pumped with 10% (w/v) sucrose, and 40 strains secreting viscous substances with the naked eye were secondarily selected, which is shown in Table 2. In Table 2 below, No. Of isolates, JNU was omitted and only the number of the strain was written.

No. of isolates isolated source MRS 10% sucrose MRS agar 10% sucrose MRS broth 9 Fermented broth - ++++ ++++ 29 Kimchi - ++++ ++++ 59 Kimchi - ++++ +++ 67 Kimchi - +++ +++ 68 Kimchi - ++++ +++ 69 Kimchi - ++++ +++ 81 Kimchi - +++ +++ 88 Kimchi - ++++ +++ 100 Soybean paste - ++++ +++ 108 Soybean paste - +++ ++ 136 Soybean paste - + +++ 147 Soybean paste - + +++ 171 Soybean paste - ++++ +++ 206 Red pepper paste - ++++ + 224 Soy sauce - +++ ++++ 249 Vinegar + +++ +++ 257 Soybean paste - +++ +++ 265 Kimchi +++ +++ - 270 Kimchi - +++++ +++ 276 Kimchi - +++++ +++ 283 Kimchi - +++++ +++ 285 Kimchi - + + 302 Kimchi - ++++ +++ 327 Kimchi - + +++ 335 Red pepper paste - + + 338 Red pepper paste - ++++ +++ 349 Red pepper paste +++ +++ + 356 Fermented broth + ++ +++ 368 Kimchi - +++ + 386 Kimchi + ++++ + 409 Kimchi +++ + ++ 465 Red pepper paste + ++++ +++ 493 Kimchi - +++++ +++ 496 Kimchi - +++ +++ 497 Kimchi - +++ +++ 500 Kimchi + +++ +++ 503 Kimchi + +++ +++ 506 Red pepper paste - +++ +++ 511 Soybean paste + ++++ +++ 517 Red pepper paste + +++ +++ `+~+++++ : showed mucoid, - : not detected.

Experimental Example 3: Third selection and identification of strains having extracellular polysaccharide production ability

1) TLC (Thin layer chromatography) analysis

When the 40 strains secondarily selected from Experimental Example 2 were cultured, a strain having excellent sucrose degradability and high polysaccharide and oligosaccharide contents was selected. The 40 strains were each instilled in a liquid medium to which 10% (w/v) sucrose was added or a general MRS liquid medium, and then cultured at 37 ° C. for 2 days, and 1 μl of the recovered culture medium was loaded onto a TLC plate. Next, it was developed three times with a developing solvent (a mixture of acetonitrile (ACN) and water in a weight ratio of 85:15). The color was developed with a color-developing solvent (0.5% naphtal, 5% H ₂ SO ₄ in ethanol) and baked in an oven at a temperature of 105° C. to confirm a spot ( FIG. 4 ).

4A and 4B show the results of TLC analysis of a culture medium in which 40 strains of the second selected from Experimental Example 2 were cultured in an MRS medium, and a culture medium in which sucrose was added to the MRS medium. 'Std 1' means Standards 1, which has glucose (glucose, G) and maltose (M) spots. 'Std 2' means Standards 2 and has fructose (F) and sucrose (S) spots. 'Strain number' is the culture medium cultured in MRS medium, and 'strain number + s' is the culture medium cultured in MRS medium to which sucrose is added. 'M' is MRS liquid medium, and 'M-S' is MRS liquid medium to which sucrose is added.

4, including three strains of fast decomposition of glucose and fructose (JNU9, JNU29, JNU224), two strains of high oligosaccharide content (JNU265, JNU506), and a strain with fast decomposition of sucrose (JNU465) A total of 6 strains were selected.

2) Comparison of extracellular polysaccharide forming ability for 6 strains

After each dropwise addition of 6 strains having extracellular polysaccharide-producing ability to a liquid medium supplemented with 10% (w/v) sucrose, incubated at 37° C. for 2 days, and 1 μl of the recovered culture was loaded onto a TLC plate. Then, it was developed three times with a developing solvent (a mixture of acetonitrile (ACN) and water in a weight ratio of 85:15). The color was developed with a color-developing solvent (0.5% naphtal, 5% H ₂ SO ₄ in ethanol) and baked in an oven at a temperature of 105° C. to confirm a spot ( FIGS. 5 and 6 ).

5 shows the results of TLC analysis of the culture medium in which six strains (JNU9, JNU29, JNU224, JNU265, JNU465, JNU506) were cultured in sucrose-added MRS broth. 'IMO' stands for Isomalto-oligosaccharide, 'MO' stands for Malto-oligosaccharide, 'M' stands for maltose (M), 'S' stands for sucrose (S), and 'G' stands for glucose ( glucose, G) and 'F' are fructose (F) spots. Strain number' is the culture medium cultured in MRS liquid medium to which sucrose is added.

6 is after culturing 6 strains (JNU9, JNU29, JNU224, JNU265, JNU465, JNU506) in MRS medium to which sucrose is added, the phenotype of the strain and the viscosity of the culture medium were visually confirmed.

5, it can be seen that all of the strains JNU9, JNU29, JNU224, JNU265, JNU465, and JNU506 of the present invention show fast decomposition of sugar.

6, it was confirmed that the culture medium of the strains JNU29 and JNU506 secreted a large amount of mucilage, and the viscosity of the culture medium was greatly increased to the extent that it could be confirmed with the naked eye.

3) sympathy

To identify the six strains, the genomic DNA of each strain was extracted (Thermo fisher). The extracted genomic DNA was identified by amplifying the 16S rRNA base sequence. Identification of the strain was commissioned by Macrogen Co., Ltd., and the base sequence of the 16S rRNA gene was compared and analyzed using the BLAST program. As a result, JNU9 and JNU29 showed 99% homology with Weissella cibaria, and JNU265 and JNU506 were confirmed to have 99% homology with Bacillus subtilis. In addition, JNU224 showed 99% homology to Bacillus amyloliquefaciens , and JNU465 was confirmed to show 99% homology to Bacillus stratosphericus . Identification results for 6 strains are shown in [Table 3].

division Origins Description Identities (%) JNU9 Fermented broth Weissellacibaria 99 JNU29 Kimchi Weissellacibaria 99 JNU224 Soy sauce Bacillus amyloliquefaciens 99 JNU265 Kimchi Bacillus subtilis 99 JNU465 Red pepper paste Bacillus stratosphericus 99 JNU506 Red pepper paste Bacillus subtilis 99

As shown in Table 3, the strain identification results were named W. cibaria JNU 9, W. cibaria JNU 29, B. amyloliquefaciens JNU 224, B. subtilis JNU 265, B. stratosphericus JNU 465, B. subtilis JNU 506, and these were named. Finally, useful strains with high extracellular polysaccharide production ability were selected. Among them, B. subtilis JNU 265 was excluded as it was found to be the same strain as the existing strain.

Figure 7 is after culturing each of the four strains (JNU29, JNU224, JNU465, JNU506) in the normal MRS medium and the sucrose-added MRS medium, the phenotype of the strain and the viscosity of the culture medium were visually confirmed.

As shown in FIG. 7 , the four strains showed viscosity even in a solid medium, and it was confirmed that they exhibit viscosity in a liquid medium as well. In particular, strains JNU29 and JNU506 secreted a large amount of viscous material in the liquid medium and were confirmed to have a viscosity comparable to starch syrup.

Experimental Example 4: Gastric juice and bile resistance analysis

It was confirmed that the strains in the present invention are lactic acid bacteria and produce high concentrations of extracellular polysaccharides unlike other lactic acid bacteria. Extracellular polysaccharides are known to have various physiologically active functions. Therefore, when the strain of the present invention is added to food, it is possible not only to ensure long-term storage of food, but also to reach the intestine and perform useful functions.

1) gastric juice resistance

First, in order for the strains of the present invention to reach the intestine in a live state, they must pass through the stomach in which hydrochloric acid and various enzymes exist. Specifically, the pH of the stomach is very large, ranging from 2 to 8, depending on whether or not food is ingested.

Herein, with respect to the five strains selected tertiarily through Experimental Example 3, it was attempted to determine whether they were viable in vivo, and for this purpose, a resistance test to artificial gastric juice was performed. First, an MRS liquid medium prepared to have a pH of 3 by adding HCl (or MRS liquid medium containing 10% sucrose) was prepared, filtered through a 0.22 um membrane (Millipore), and 1,000 unit/ml pepsin was added to ' Artificial gastric juice (or artificial gastric juice containing 10% sucrose) was prepared. After culturing each of the five strains in MRS broth medium at 37° C. for 1 day, centrifugation was performed to remove the supernatant, and the artificial gastric juice was added as much as the removed supernatant. Incubated at 37° C. for 2 hours, and the number of viable cells was counted. In this case, the control 1 (MRS medium) is a non-artificial gastric juice, but a general MRS liquid medium that is not pH adjusted, and the control 2 (MRS + sucrose medium) is an MRS liquid containing 10% sucrose that is not pH adjusted, not an artificial gastric juice. medium was used.

8 is a graph showing the number of viable cells counted when five strains (JNU9, JNU29, JNU224, JNU465, JNU506) were added to artificial gastric juice, 10% sucrose-added artificial gastric juice, and controls 1 and 2 and cultured.

As shown in FIG. 8 , in Controls 1 and 2, there was almost no difference in the number of viable cells in all of the 5 strains. In addition, B. amyloliquefaciens JNU 224 and B. stratosphericus JNU 465 strains did not show a difference in the number of viable cells in both artificial gastric juice and artificial gastric juice supplemented with 10% sucrose. It can be seen that B. amyloliquefaciens JNU 224 and B. stratosphericus JNU 465 strains have resistance to artificial gastric juice. In particular, it was confirmed that the B. stratosphericus JNU 465 strain showed a tendency to slightly decrease the number of viable cells when sucrose was treated.

In W. cibaria JNU 29 and B. subtilis JNU 506 strains, the number of viable cells was hardly measured in artificial gastric juice, but the number of viable cells was more than four times higher in artificial gastric juice containing 10% sucrose.

In the W. cibaria JNU 9 strain, it was confirmed that the number of viable cells was decreased in Control 2 treated with sucrose rather than Control 1, and the number of viable cells was slightly increased in the artificial gastric juice treated with sucrose under the condition of artificial gastric juice.

2) bile resistance

After the strains of the present invention pass through the stomach, in order to perform a normal function in the intestine, it must be resistant to growth in a medium containing much more oxgall than the concentration of intestinal bile (0.6 g/L). For the artificial bile resistance experiment, first, 0.3% yellow sand bile (oxgall) filtered through a 0.45 um membrane (Millipore) was added to MRS liquid medium (or MRS liquid medium to which 10% sucrose was added) to 'artificial bile (oxgall) ( or 10% sucrose-added artificial bile)'.

After culturing each of the five strains in MRS broth medium at 37° C. for 1 day, centrifugation was performed to remove the supernatant, and the artificial gastric juice was added as much as the removed supernatant. Incubated at 37 °C for 2 hours, the culture medium was centrifuged again to remove the supernatant, treated as much as the supernatant from which artificial bile was removed, and then cultured at 37 °C for 24 hours to count the number of viable cells. In this case, the control 1 (MRS medium) is a non-artificial gastric juice, but a general MRS liquid medium that is not pH adjusted, and the control 2 (MRS + sucrose medium) is an MRS liquid containing 10% sucrose that is not pH adjusted, not an artificial gastric juice. medium was used.

9 is a graph showing the number of viable cells counted when cultured by adding 5 strains (JNU9, JNU29, JNU224, JNU465, JNU506) to artificial bile, 10% sucrose-added artificial bile, and controls 1 and 2. FIG.

As shown in FIG. 9 , as a result of examining the resistance to bile acids, in both artificial bile and artificial bile containing 10% sucrose, five strains (JNU9, JNU29, JNU224, JNU465, JNU506) all had high viable cell counts. was confirmed to be maintained. That is, it can be seen that the strain of the present invention is excellent in bile fluid resistance.

As such, the high resistance in both gastric juice and bile fluid is when 10% sucrose is added, and when 10% sucrose is added, extracellular polysaccharide (EPS) is produced from the strain and acts as a protective film around the cell wall. It is considered

Through the above results, the five strains finally isolated through the selection process of the present invention not only produce extracellular polysaccharides at a high concentration, but also have excellent resistance to gastric juice and bile, so that they can survive to the intestinal environment, It can survive in the intestine, so it can be seen that it is a very useful strain as a probiotic material.

Experimental Example 5: Analysis of culture medium viscosity and extracellular polysaccharide production capacity

1) Viscosity

To analyze the characteristics of extracellular polysaccharides generated from five strains (JNU9, JNU29, JNU224, JNU465, JNU506) in more detail. To this end, the above five strains were cultured in MRS broth medium and MRS broth supplemented with 10% sucrose at 37° C. for 2 days, respectively, and then the viscosity of the separated culture was measured. Viscosity of the culture solution of each strain was measured at 25° C. at 20 rpm using a spindle RV (3) of a viscometer (Brookfield viscometer, DV2T).

10 is a result of measuring the viscosity of the culture medium of five strains (JNU9, JNU29, JNU224, JNU465, JNU506). As shown in FIG. 10, the culture medium obtained by culturing in the MRS medium did not differ significantly by strain. However, when culturing in MRS broth containing 10% sucrose, it was confirmed that the cultures of W. cibaria JNU 9 and W. cibaria JNU 29 strains had the highest viscosity. In particular, it was confirmed that the W. cibaria JNU 29 strain had twice the viscosity than the W. cibaria JNU 9 strain.

2) Analysis of extracellular polysaccharide content

The content of extracellular polysaccharide (EPS) finally produced from five strains (JNU9, JNU29, JNU224, JNU465, JNU506) was measured. Five strains (JNU9, JNU29, JNU224, JNU465, JNU506) were inoculated into MRS liquid medium supplemented with 10% sucrose, respectively, and cultured at 37 ° C. for 2 days. After centrifugation of the culture medium, the supernatant was recovered. . A cooled 95% ethanol aqueous solution was added twice by weight with respect to the total weight of the supernatant of the recovered culture solution. At this time, a cooled 95% ethanol aqueous solution was slowly added, and the resultant was precipitated at 4° C. for 16 hours, and the precipitate was recovered by centrifugation. The recovered precipitate was dried to remove ethanol, and then lyophilized to obtain extracellular polysaccharides produced from each strain.

The obtained extracellular polysaccharide was added with 4% trichloroacetic acid (sigma), reacted at 4° C. for 2 hours, and then centrifuged to remove the precipitated protein. The recovered supernatant was filtered with a 0.22um membrane (Millipore) to remove the remaining protein. To the total weight of the supernatant, two times the weight of a cooled 95% aqueous ethanol solution was added, and the mixture was precipitated at 4°C for 16 hours. This was centrifuged to remove the supernatant, and only the precipitate was recovered, then the remaining ethanol was removed through the drying process, dissolved in sterile water, put in a dialysis sack (MW cut off 10,000, Thermo), and dialyzed at 4 °C for 48 hours. and then freeze-dried. After culturing 5 strains for 2 days, and separating the extracellular polysaccharide from the culture medium, the content of the finally produced extracellular polysaccharide was expressed as g/l, and the results are summarized in Table 4.

strains Final Produced EPS Content (g/L) Weissell acibaria JNU 9 11.58 ± 0.4 Weissell acibaria JNU 29 16.72 ± 0.26 Bacillus amyloliquefaciens JNU 224 8.57 ± 0.8 Bacillus stratosphericus JNU 465 15.38 ± 045 Bacillus subtilis JNU 506 8.55 ± 0.07

As shown in Table 4, as a result of measuring the content of the extracellular polysaccharide final produced from each strain, it was confirmed that the W. cibaria JNU 29 strain produced the most extracellular polysaccharide at 16.72 g/l. B. subtilis JNU 506 strain produced the least extracellular polysaccharide at 8.55 g/l.

Interestingly, the W. cibaria JNU 9 strain identified as the same species as the W. cibaria JNU 29 strain showed a difference not only in the viscosity of the culture medium, but also in the production content of extracellular polysaccharides.

B. stratosphericus JNU 465 strain also had a very high content of extracellular polysaccharide (15.38 g/l), but it was excluded from subsequent experiments because it was a strain that could not be used as food. In addition, the B. amyloliquefaciens JNU 224 strain was not only low in production of extracellular polysaccharides but also low in resistance to artificial gastric juice and artificial bile, so the B. subtilis JNU 506 strain was finally selected.

Therefore, W. cibaria JNU 29 strain, W. acibaria JNU 9 strain, and B. subtilis JNU 506 strain having high extracellular polysaccharide content were finally selected.

Experimental Example 6: Analysis of extracellular polysaccharide constituents

Extracellular polysaccharides are largely divided into homopolysaccharides and heteropolysaccharides according to strains and types of sugars constituting them. Homopolysaccharides are mainly composed of only one type of sugar, such as fructose and glucose. Heteropolysaccharide refers to a substance composed of two or more monosaccharides, such as glucose, galactose, fructose, and rhamnose, in different ratios, and in some cases, non-sugar substances such as N-acetyl-amino sugar, phosphate, acetyl, and glycerol. may exist In general, heteropolysaccharides show lower productivity than homopolysaccharides, and viscous production shows temporary characteristics.

TLC and HPLC analysis were performed to confirm the constituent sugars of the extracellular polysaccharide produced by the three strains selected from the present invention. The constituent sugars of the extracellular polysaccharide were hydrolyzed with 2N sulfuric acid at 100 °C for 3-6 hours. The hydrolyzate was neutralized with 1N NaOH, filtered using a 0.45 um filter, and used as a TLC and HPLC sample.

1) TLC analysis-1

1 μl of the hydrolyzate obtained through the above procedure was dripped onto the TLC plate. It was developed 4 times with a developing solvent (acetonitrile: distilled water = 85:15 (v/v)). The color was developed with a color-developing solvent (0.5% naphtal, 5% H ₂ SO ₄ in ethanol) and baked in an oven at 105° C. to confirm the constituent sugar spots.

11 is a TLC analysis result for the extracellular polysaccharide produced from the three strains finally selected (JNU 9, JNU 29, JNU 506), according to the results of the three strains finally selected (JNU 9, JNU 29, JNU). 506) existed in the form of extracellular polysaccharide before hydrolysis, but only one spot was present upon hydrolysis. It can be seen that this produces an extracellular polysaccharide in the form of a homopolysaccharide. In this case, ‘IMO’ means isomalto-oligosaccharide, and ‘MO’ means malto-oligosaccharide.

2) TLC analysis-2

1 μl of the hydrolyzate obtained through the above procedure was dripped onto the TLC plate. It was developed twice with a developing solvent (nitromethane: 1-propanol: distilled water = 2:5:1.5 (v/v/v)). The color was developed with a color-developing solvent (0.5% naphtal, 5% H ₂ SO ₄ in ethanol) and baked in an oven at 105° C. to confirm the constituent sugar spots.

12 is a TLC analysis result for the extracellular polysaccharide produced from the three strains finally selected (JNU 9, JNU 29, JNU 506), according to the results of the three strains finally selected (JNU 9, JNU 29, JNU). 506) existed in the form of extracellular polysaccharide before hydrolysis, but only one spot was present upon hydrolysis. It can be seen that this produces an extracellular polysaccharide in the form of a homopolysaccharide. Both TLC results were measured equally, and in the case of homopolysaccharide, it is commercially very useful, and the field of application is also very wide. In this case, ‘IMO’ means isomalto-oligosaccharide, and ‘MO’ means malto-oligosaccharide.

3) HPLC analysis

In order to accurately analyze the sugar constituents of the extracellular polysaccharide produced from each strain, the hydrolyzate of each of the three strains obtained through the above procedure was analyzed by HPLC using a carbohydrate analysis column. .

13 to 16 are HPLC analysis results for extracellular polysaccharides generated from the three finally selected strains (JNU 9, JNU 29, JNU 506). As a result of hydrolysis of the polysaccharide, only a single peak was identified for glucose. That is, it can be seen that the W. cibaria JNU 9, W. cibaria JNU 29, and B. subtilis JNU 506 strains finally selected in the present invention produce an extracellular polysaccharide of homopolysaccharide composed of glucose.

Experimental Example 7: Aging and decay inhibitory activity analysis in seolgitteok

The W. cibaria JNU 29 (Weissella ciberia JNU 29) strain isolated in the present invention has an activity of producing a homopolysaccharide extracellular polysaccharide composed of glucose through sucrose metabolism, which is a raw material of many foods. When ingested, it can pass through the stomach and reach the intestine safely, and it can maintain high production of extracellular polysaccharides due to excellent survival in the intestinal environment. Next, the purpose of this study was to analyze whether roaring and spoilage are suppressed during the production of foods such as rice cakes.

1) Total number of lactic acid bacteria

Seolgitteok prepared in Example 2 and seolgitteok prepared in Comparative Examples 1 and 2 were prepared, and the total number of bacteria was measured on day 0 and day 2. The results are shown in Table 5 below. To measure the total number of lactic acid bacteria, each seolgitteok was homogenized with saline, diluted and spread on MRS solid medium, incubated at 37 ° C. for 24 hours, and then the number of colonies was counted. In this case, as a control (con), a general seolgitteok prepared without any culture medium was used.

storage date Total number of lactic acid bacteria (CFU/g) con JNU9 JNU29 JNU506 0 day 0 2 × 10 ² 2 × 10 ² 3 × 10 ² 2 days 0 3.8 × 10 ⁷ 4.6 × 10 ⁷ 1.8 × 10 ⁶

As shown in Table 5, as a result of measuring the surviving lactic acid strains in seolgitteok, it was confirmed that the strains did not die and survived during the rice cake manufacturing process. That is, it can be seen that all three strains finally selected in the present invention have heat resistance. In addition, after 2 days of manufacture, as a result of comparing the total number of lactic acid bacteria, it was confirmed that seolgitteok (JNU 29) prepared in Example 2 contained abundant lactic acid bacteria.

2) Aging analysis of rice cake

After cutting the seolgitteok prepared in Example 2 and Comparative Examples 1 and 2 to have a width, length, and height of 8 cm × 7 cm × 3 cm, hardness was measured using a texture analyzer.

17 is a graph showing the time-dependent hardness change of seolgitteok prepared from Example 2, Comparative Examples 1 and 2, and according to this, seolgitteok prepared from Example 2 ( W. cibaria JNU 29) is a control and Comparative Example 1 , it can be seen that, unlike seolgitteok (W. cibaria JNU 9 and B. subtilis JNU 506) prepared from 2, the increase in hardness is significantly lower. In FIG. 17, a, b, and c show statistical differences between treatment groups (Duncan P≤0.05).

It was confirmed that the seolgitteok of Comparative Example 1 prepared with a strain of the same genus as W. cibaria JNU 29 had a higher hardness than the control. The seolgitteok prepared in Comparative Example 2 was prepared by adding JNU 506, a Bacillus subtilis strain widely known as a lactic acid strain, but it was also confirmed that the hardness increased more than the control. Therefore, when W. cibaria JNU 9 and B. subtilis JNU 506 strains are added instead of W. cibaria JNU 29 strain, a problem may occur that the rice cake progresses faster than normal seolgitteok.

3) Corruption analysis of rice cake

The seolgitteok prepared in Example 2, Comparative Example 1 and Comparative Example 2 were stored at room temperature to determine the degree of decay.

18 is a photograph of the degree of decay according to time (1 day, 7 days) when the seolgitteok prepared in Example 2 and Comparative Examples 1 and 2 was stored at room temperature. It can be confirmed that seolgitteok affects not only aging but also decay caused by mold. Specifically, it was confirmed that the general seolgitteok (control group) began to deteriorate from the 3rd day, but the seolgitteok prepared in Example 2 did not significantly deteriorate even on the 7th day.

On the other hand, it can be seen that the seolgitteok prepared in Comparative Examples 1 and 2 has a higher degree of decay than the seolgitteok prepared in Example 2, although the degree of decay progresses less than that of the control group.

In conclusion, the W. cibaria JNU 29 strain finally isolated through the present invention produces a high concentration of extracellular polysaccharides composed of homopolysaccharides, and can significantly delay the aging rate of rice cakes during rice cake production, and at the same time can significantly inhibit spoilage. This enables long-term storage and long-term distribution. In addition, it does not die in the high-temperature steaming process and remains in the rice cake, so it has a great advantage that it can add physiological functions to the rice cake by the W. cibaria JNU 29 strain. In addition, the W. cibaria JNU 29 strain has resistance to gastric juice and bile, so it can safely reach the intestine when ingested, and can survive in the intestine, so it is a useful prebiotic by the W. cibaria JNU 29 strain. Even activity can be achieved.

Korea Institute of Bioscience and Biotechnology KCTC14070BP 20191212

<110> INDUSTRY FOUNDATION OF CHONNAM NATIONAL UNIVERSITY <120> Weissella cibaria strain having rice cake anti-retrogradation activity and use thereof <130> HPC9088 <160> 1 <170> KoPatentIn 3.0 <210> 1 <211> 1513 <212> DNA <213> Unknown <220> <223> 16S rRNA of Weissella cibaria JNU29 <400> 1 gctcaggatg aacgctggcg gcgtgcctaa tacatgcaag tcgaacgctt tgtggttcaa 60 ctgatttgaa gagcttgctc agatatgacg atggacattg caaagagtgg cgaacgggtg 120 agtaacacgt gggaaaccta cctcttagca ggggataaca tttggaaaca gatgctaata 180 ccgtataaca atagcaaccg catggttgct acttaaaaga tggttctgct atcactaaga 240 gatggtcccg cggtgcatta gttagttggt gaggtaatgg ctcaccaaga cgatgatgca 300 tagccgagtt gagagactga tcggccacaa tgggactgag acacggccca tactcctacg 360 ggaggcagca gtagggaatc ttccacaatg ggcgaaagcc tgatggagca acgccgcgtg 420 tgtgatgaag ggtttcggct cgtaaaacac tgttgtaaga gaagaatgac attgagagta 480 actgttcaat gtgtgacggt atcttaccag aaaggaacgg ctaaatacgt gccagcagcc 540 gcggtaatac gtatgttcca agcgttatcc ggatttattg ggcgtaaagc gagcgcagac 600 ggttatttaa gtctgaagtg aaagccctca gctcaactga ggaattgctt tggaaactgg 660 atgacttgag tgcagtagag gaaagtggaa ctccatgtgt agcggtgaaa tgcgtagata 720 tatggaagaa caccagtggc gaaggcggct ttctggactg taactgacgt tgaggctcga 780 aagtgtgggt agcaaacagg attagatacc ctggtagtcc acaccgtaaa cgatgagtgc 840 taggtgtttg agggtttccg cccttaagtg ccgcagctaa cgcattaagc actccgcctg 900 gggagtacga ccgcaaggtt gaaactcaaa ggaattgacg gggacccgca caagcggtgg 960 agcatgtggt ttaattcgaa gcaacgcgaa gaaccttacc aggtcttgac atcccttgac 1020 aactccagag atggagcgtt cccttcgggg acaaggtgac aggtggtgca tggttgtcgt 1080 cagctcgtgt cgtgagatgt tgggttaagt cccgcaacga gcgcaaccct tattactagt 1140 tgccagcatt tagttgggca ctctagtgag actgccggtg acaaaccgga ggaaggtggg 1200 gatgacgtca aatcatcatg ccccttatga cctgggctac acacgtgcta caatggcgta 1260 tacaacgagt tgccaacccg cgagggtgag ctaatctctt aaagtacgtc tcagttcgga 1320 ttgtaggctg caactcgcct acatgaagtc ggaatcgcta gtaatcgcgg atcagcacgc 1380 cgcggtgaat acgttcccgg gtcttgtaca caccgcccgt cacaccatga gagtttgtaa 1440 cacccaaagc cggtggggta accttcggga gccagccgtc taaggtggga cagatgatta 1500 gggtgaagtc gta 1513

Claims (8)

Having the 16S rRNA base sequence represented by SEQ ID NO: 1, and having acid-resistance, bile-resistant and heat-resistant Weissella cibaria JNU 29 [Accession No.: KCTC14070BP] strain having anti-aging and decay-inhibiting activity of rice cakes. delete According to claim 1,
The strain is a strain characterized in that it is cultured under conditions containing sucrose as a carbon source. Has the 16S rRNA base sequence represented by SEQ ID NO: 1, and has acid-resistance, bile-resistant and heat-resistant Weissella cibaria JNU 29 [Accession No.: KCTC14070BP] strain having anti-aging and decay inhibitory activity of rice cakes Extracellular polysaccharide (exopolysaccharide). 5. The method of claim 4,
The extracellular polysaccharide is an extracellular polysaccharide, characterized in that it is prepared by culturing Weissella cibaria JNU 29 [Accession No.: KCTC14070BP] strain in a medium containing sucrose as a carbon source. Has the 16S rRNA nucleotide sequence represented by SEQ ID NO: 1, and has acid-resistance, bile-resistant and heat-resistant Weissella cibaria JNU 29 [Accession No.: KCTC14070BP] strain or a culture solution thereof having anti-aging and decay-inhibiting activity of rice cake A composition for inhibiting aging and spoilage of rice processed food containing as an active ingredient. Having the 16S rRNA nucleotide sequence represented by SEQ ID NO: 1, and having acid-resistance, bile-resistant and heat-resistant Weissella cibaria JNU 29 [Accession No.: KCTC14070BP] strain having anti-aging and decay-inhibiting activity of rice cakes Rice processed food with improved storage. 8. The method of claim 7,
The processed rice food, rice processed food, characterized in that it further comprises sucrose.

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