JP7199063B2 - Novel lactic acid bacterium and method for producing viscous fermented product using this lactic acid bacterium - Google Patents

Description

NPMD NITE P-03243

TECHNICAL FIELD The present invention relates to a lactic acid bacterium that produces a stringy component and a method for producing a viscous fermented product using this lactic acid bacterium.

Recently, the demand for gluten-free foods and plant-based foods, which are foods made only from plant-based ingredients, has increased from the perspectives of reducing environmental impact such as carbon dioxide emissions, realizing a sustainable society, and making food barrier-free. ing. Niigata Prefecture, one of Japan's leading rice-producing areas, has promoted the development of foods such as bread, noodles, miso, and soy sauce that are made entirely from rice. However, there is a limit to the production of food materials with various textures and physical properties from rice alone, and new physical property conversion technology is required.

On the other hand, some fermenting microorganisms, such as koji mold, natto bacillus, and lactic acid bacteria, decompose plant components or produce new components, contributing to the transformation of physical properties of food ingredients such as liquefaction and viscosity. . Therefore, it is thought that a technique of converting physical properties by fermenting vegetable raw materials is useful.

Also, when producing fermented foods, attempts have been made to separate local specialty foods or lactic acid bacteria derived from the land and use them as starters to produce fermented foods. For example, a method for producing fermented foods using lactic acid bacteria isolated from Nozawana pickles in the Uonuma region of Niigata Prefecture (Patent Document 1), a method for producing fermented foods using lactic acid bacteria separated from leaf soil in Shirakami Mountains (Patent Document 2) ), and a method for producing pickles using lactic acid bacteria Lactobacillus salmon HS1 isolated from fermented kimchi (Patent Document 3).

JP 2013-236604 A JP 2007-236344 A Japanese Patent Application Laid-Open No. 2001-120173

Therefore, an object of the present invention is to provide a novel lactic acid bacterium that enables the production of food materials having various textures and physical properties even when only rice is used as a raw material.

As a result of intensive studies to achieve the above object, a novel lactic acid bacterium that produces stringy components and a method for producing a viscous fermented product using this lactic acid bacterium were discovered. Specifically, we found a new lactic acid bacterium that contributes to the transformation of physical properties by imparting viscosity and stringiness (stringiness) to food materials from ``Izekomi-na'', a preserved food that has been handed down from the Yamakoshi district of Niigata Prefecture. was identified and its properties analyzed. Furthermore, the present inventors found that by fermenting a material containing vegetable protein such as brown rice and soy milk using this strain, a food material having physical properties similar to yogurt or cheese can be obtained, leading to the present invention.

That is, the lactic acid bacterium of the present invention is Lactobacillus paraplantarum YAMAKOSHI (Accession No. NITE P-03243).

Further, the method for producing a viscous fermented product of the present invention is characterized by including a step of fermentation using the lactic acid bacteria of the present invention.

Furthermore, the viscous fermented product of the present invention is characterized by being obtained by the production method of the present invention.

INDUSTRIAL APPLICABILITY According to the present invention, it is possible to impart viscosity and stringiness to a food material containing vegetable protein such as rice, thereby producing a food material having yogurt- or cheese-like physical properties.

Molecular phylogenetic tree based on the recA base sequence. 4 is a graph showing the effect of culture temperature on growth of the lactic acid bacteria of the present invention. 1 is a graph showing the effect of medium pH on the growth of lactic acid bacteria of the present invention. It is a graph which shows the influence on the viscosity by various protein addition.

The lactic acid bacterium Lactobacillus paraplantarum YAMAKOSHI (Lactobacillus paraplantarum YAMAKOSHI) of the present invention is Lactobacillus paraplant isolated from traditional preserved food "Izekomi-na" (unsalted pickled radish) in Yamakoshi district, Nagaoka City, Niigata Prefecture. It is a new fungus belonging to Lactobacillus paraplantarum, and was transferred to the National Institute of Technology and Evaluation Patent Microorganism Depositary Center (2-5-8 Kazusa Kamatari, Kisarazu City, Chiba Prefecture) on July 7, 2020. It has been deposited under accession number NITE P-03243. Hereinafter, the lactic acid bacterium of the present invention is referred to as YAMAKOSHI strain.

The YAMAKOSHI strain of the present invention produces a viscous component from a substrate containing vegetable proteins such as brown rice and soy milk. In addition, the higher the protein concentration of the substrate, the higher the viscosity of the substrate due to the fermentation of the YAMAKOSHI strain of the present invention. Therefore, by fermenting brown rice or soy milk using the YAMAKOSHI strain of the present invention, it becomes possible to produce viscous and stringy yogurt or cheese-like foods.

In addition, lactic acid bacteria that produce highly viscous and stringy ingredients are rare, and by using the YAMAKOSHI strain of the present invention, it is possible to create gluten-free, plant-based foods with new physical properties. For example, it is possible to adjust the viscosity using only plant materials such as rice and soybeans, opening the way to the development of new processed foods.

The present invention will be described in more detail below based on examples. The present invention is not limited to the following examples, and various modifications can be made without departing from the spirit of the present invention.

Isolation and identification of the novel lactic acid bacteria of the present invention will be described.

Sterilized physiological saline was added to the traditional preserved food ``Izekomina'' (unsalted pickled radish, 25 days after pickling) of Yamakoshi district, Nagaoka City, Niigata Prefecture, and homogenized with a stomacher. A part of it was cultured on an MRS chalk agar medium for lactic acid bacteria isolation, and 40 strains of lactic acid bacteria that formed a clear zone (clear zone) by dissolving calcium carbonate added to the medium were collected.

BLAST homology analysis was performed on the partial nucleotide sequence of the 16S ribosomal DNA of the strain that showed very strong stringiness (stringiness) on the medium. As a result, it was found to be Lactobacillus plantarum, Lactobacillus pentosus, or Lactobacillus paraplantarum.

Next, in order to identify this strain, Technosuruga Labo Co., Ltd. was entrusted with recA (recombinase A protein) nucleotide sequence analysis. As a result of BLAST homology search against the international base sequence database, the recA gene partial base sequence of this strain is Lactobacillus plantarum group. It showed high homology to the recA gene nucleotide sequence of L. paraplantarum. L. As a result of molecular phylogenetic analysis performed by obtaining the recA gene nucleotide sequences (Table 1) of the bacterial group included in the L. plantarum group, this strain is L. plantarum. contained within clusters formed by L. paraplantarum (Fig. 1). Therefore, from these results, this strain is L. It was identified as L. paraplantarum and named YAMAKOSHI strain.

Next, the growth characteristics of the YAMAKOSHI strain were examined. FIG. 2 shows the effect of culture temperature on growth in MRS medium (pH 6.8). The optimum temperature for growth was around 28°C, and it was shown that although it proliferates at low temperatures, it does not grow at temperatures above 40°C. In addition, FIG. 3 shows the effects of MRS medium and medium pH at 30° C. on growth. It was shown to have proliferative properties at low pH (pH 3.5 or higher).

Next, the sugar assimilation of the YAMAKOSHI strain was examined. The YAMAKOSHI strain showed ability to assimilate L-arabinose, D-ribose, D-sorbitol, methyl-αD-glucopyranoside and D-turanose, but not escarin. The YAMAKOSHI strain was recognized as a novel strain because it had different assimilation properties from the homologous lactic acid bacterium type strain (L. paraplantarum NBRC107151) and produced stringy components.

The YAMAKOSHI strain produces a stringy component. Therefore, conditions for imparting viscosity to a sugar solution using the YAMAKOSHI strain were investigated.

Various proteins (Rice endosperm protein: Oryza Protein P-70 (manufactured by Oryza Yuka); Gluten: VITAL WHEAT GLUTEN AG-850 (manufactured by AMD); Soybean protein: Protein powder (derived from soybeans, Fujifilm) Wako Pure Chemical Industries)) is added to 1%, 3%, 5% (w / w), and 1% (v / w) of YAMAKOSHI strain suspension (OD660 nm = 1.0) is added, After fermentation at 25° C. for 24 hours, the viscosity was measured with a Brookfield viscometer (BM type, manufactured by Toki Sangyo Co., Ltd., rotor No. 3, 30 rpm). As a result, as shown in FIG. 4, the viscosities differed depending on the protein species, but in all cases the viscosities increased in a concentration-dependent manner. Among them, the addition of soy protein gave the highest viscosity and also exhibited stringiness.

Next, rice endosperm protein (manufactured by Oryza Yuka Co., Ltd.) and rice oil (manufactured by Tsuno Foods Industry Co., Ltd.) were added to the koji amazake as shown in Table 3. (v/w) and fermented at 25° C. for 24 hours, and then evaluated for viscosity with a Brookfield viscometer (rotor No. 3, 30 rpm). Also, the Zahn Cup No. The spinnability was measured by measuring the dry-out time when No. 7 was used. In order to clarify the factors affecting the viscosity of the rice sugar solution, rice endosperm protein and rice oil were added to the rice sugar solution and fermented. I didn't.

Next, 3 L of rice bran suspension (750 g of rice bran or defatted rice bran (25% (w/v)), 60 g of glucose (2% (w/v)), 30 g of citric acid (1% (w/v)) were mixed with hemi-bran. 3 g (0.1% (w/v)) of cellulase was added and allowed to react for 24 hours at 50° C. To this suspension was added 1% (v/v) of a lactic acid bacteria suspension adjusted to OD660 nm=1.0. ) and fermented for 24 hours at 25° C. Samples were taken over time to evaluate viscosity and stringiness.As shown in Table 5, defatted rice bran or rice bran liquefied with hemicellulase was As a result of fermentation with YAMAKOSHI strain as a substrate, stringiness was not exhibited in defatted rice bran, but stringiness was observed in rice bran.From this, among the components contained in rice bran, a plurality of components are stringy by lactic acid fermentation. It was presumed to be involved in imparting threadiness.

Using the YAMAKOSHI strain, a yoghurt-like food was produced using only rice as a raw material.

360 mg (0.04% (w/w)) α-amylase was added to 900 g of cooked brown or white rice, and 9 ml (1% (v/w)) of YAMAKOSHI strain suspension (OD660 = 1.0) was added. In addition, the initial lactic acid bacteria count was adjusted to 10 ⁶ cfu/g and fermented at 30° C. or 25° C. for 24 hours to obtain a fermented rice product. After straining the fermented rice through a 30-mesh (595 μm) filter, the viscosity was measured using a Brookfield viscometer, and the spinnability and sensory evaluation were performed using a Zahn cup.

The spinnability was evaluated using a Zahn cup (No. 7, manufactured by Rigosha). That is, 200 ml of a sample kept at 25° C. was placed in a 200 ml beaker, a Zahn cup set at a height of 10 cm above the liquid surface of the sample was submerged in the sample in the beaker, and the beaker was quickly pulled apart. The time from when the bottom of the cup left the liquid surface to when the flow of the sample flowing out from the orifice (hole) at the bottom of the Zahn cup was interrupted was measured, and the time during which the steady flow continued was evaluated as the spinnability.

Stringiness could not be confirmed in fermented white rice, but stringiness was observed in fermented brown rice, making it possible to produce foods with yogurt-like viscosity.

A yogurt-like food was produced using soymilk.

Put 900 ml of commercially available soy milk (solid content 10%, protein 5%, pH 6.5) in a 1 L centrifuge tube, and add 9 ml (1% (v/v)) of lactic acid bacteria suspension (OD660 = 1.0). In addition, the initial lactic acid bacteria count was adjusted to 10 ⁶ cfu/g and fermented at 25° C. for 24 hours to obtain a fermented soy milk product.

A cheese-like food was produced.

Put 900 ml of commercially available soy milk (solid content 10%, protein 5%, pH 6.5) in a 1 L centrifuge tube, and add 9 ml (1% (v/v)) of lactic acid bacteria suspension (OD660 = 1.0). In addition, the initial lactic acid bacteria count was adjusted to 10 ⁶ cfu/g and fermented at 25° C. for 24 hours to obtain a fermented soy milk product. The fermented soymilk was centrifuged (2000 rpm, 20 min) to remove the supernatant. The precipitate was transferred to a 250 ml centrifuge tube, centrifuged again (10000 rpm, 15 min), and the supernatant was removed to obtain a cheese-like solid content (cheese-like food).

The cheese-like food was subjected to sensory evaluation by a panel of 23 people, using texture (hardness, smoothness, stickiness), flavor (sourness, umami, aroma) and overall evaluation (liking) as evaluation items.

The cheese-like food using the YAMAKOSHI strain was highly evaluated for its smoothness and stickiness, had a strong sourness, and received a high overall evaluation, compared to those using other strains.

A fermented liquid bran bed was produced.

Add 2160 g of water to 750 g of rice bran (25% (w/w)), 60 g of glucose (2% (w/w)), 30 g of citric acid (1% (w/w)), mix well and autoclave ( 121° C., 15 minutes). After cooling, 3 g (0.1% (w/w)) of hemicellulase (hemicellulase "Amano" 90, manufactured by Amano Enzyme Co., Ltd.) was added and allowed to react at 50°C for 24 hours. adjusted to 5. 30 ml (1% (v/w)) of YAMAKOSHI strain suspension (OD660 = 1.0) of the present invention was added to prepare an initial lactic acid bacteria count of 10 ⁶ cfu/g, and the mixture was heated at 25°C for 24 hours. It was fermented to obtain fermented liquid rice bran.

Claims (3)

Lactobacillus paraplantarum YAMAKOSHI (Lactobacillus paraplantarum YAMAKOSHI: accession number NITE P-03243). A method for producing a viscous fermented product, comprising the step of fermenting using the lactic acid bacterium according to claim 1. A viscous fermented product obtained by the production method according to claim 2.

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