Novel lactiplantibacillus plantarum and use thereof

Abstract

To provide novel Lactiplantibacillus plantarum which allows improvement of flavor and / or taste in a cereal saccharified solution.SOLUTION: The present invention was accomplished by discovering Lactiplantibacillus plantarum (Lactiplantibacillus plantarum subsp. plantarum) strain KB-11 (Accession No.: NITE P-04017), which is utilizable for improving flavor and / or taste in a cereal saccharified solution.SELECTED DRAWING: None

Description

[Technical Field]

[0001] The present invention relates to a novel lactiprancid Bacillus plantarum and uses thereof. [Background technology]

[0002] Generally, saccharified grain solutions are known which are obtained by converting grains containing high-molecular-weight carbohydrates such as starch and cellulose into low-molecular-weight sugars by the action of enzymes, etc. Saccharified grain solutions are widely used as sweeteners for foods, beverage compositions, etc.

[0003] Grain saccharified liquids such as those disclosed in Patent Document 1 have been known for some time. Patent Document 1 discloses a beverage composition made from brown rice, which is obtained by adding protease, cellulase, pectinase, and amylase to brown rice for hydrolysis. More specifically, milled brown rice is first mixed with protease, cellulase, and pectinase, and then subjected to enzymatic hydrolysis, followed by heating to obtain a gelatinized expanded rice flour liquid. Next, this rice flour liquid is mixed with α-amylase, β-amylase, and glucoamylase, respectively, for enzymatic hydrolysis, yielding a water-soluble brown rice composition. [Prior art documents] [Patent documents]

[0004] [Patent Document 1] Japanese Patent Application Publication No. 5-137545 Summary of the Invention [Problem to be solved by the invention]

[0005] The present invention is based on the discovery of a novel Lactiprancibacillus plantarum that can be used to improve, for example, the flavor and / or aroma of a cereal saccharified liquid, more specifically, to improve the unique cereal odor of a cereal saccharified liquid, the unique fermented odor and acidic odor that a fermented cereal saccharified liquid has, and the imbalance in aroma.An object of the present invention is to provide a novel Lactiprancibacillus plantarum and uses thereof. [Means for solving the problem]

[0006] As a result of research aimed at solving the above problems, the present inventors have discovered a novel Lactiplantibacillus plantarum that can be used to improve the flavor and / or aroma of cereal saccharified liquid. Various aspects for solving the above problems will be described.

[0007] The novel Lactiplantibacillus plantarum of embodiment 1 is Lactiplantibacillus plantarum subsp. plantarum KB-11 strain (accession number: NITE P-04017).

[0008] In a second aspect, the novel lactiprancid Bacillus plantarum according to the first aspect is used to improve the taste and / or flavor of a grain saccharified liquid by fermentation. A composition of embodiment 3 is a composition comprising the lactipranci Bacillus plantarum of embodiment 1.

[0009] The fermented composition of Aspect 4 is a fermented composition obtained using the lactiprancid Bacillus plantarum of Aspect 1. Aspect 5 is the fermentation composition according to Aspect 4, which comprises a fermentation product obtained by using Lactiplantibacillus plantarum a grain saccharified liquid obtained by saccharifying grain.

[0010] A sixth aspect is the fermented composition according to the fifth aspect, wherein the grain is at least one selected from oats, rice, and millet. Aspect 7 is the fermented composition according to aspect 6, wherein the grain is oats.

[0011] Aspect 8 is the fermented composition according to aspect 6, wherein the grain is rice or millet. A method for improving the flavor and / or aroma of a grain saccharified liquid according to aspect 9 is a method for improving the flavor and / or aroma of a grain saccharified liquid, which comprises fermenting the grain saccharified liquid using Lactiprunch Bacillus plantarum according to aspect 1. [Effects of the Invention]

[0012] The novel Lactipranchi Bacillus plantarum of the present invention can improve the taste and / or flavor of grain saccharified liquid. [Brief explanation of the drawings]

[0013] [Figure 1] Figure 1 shows a simplified molecular phylogenetic tree based on the partial 16S rDNA sequence of Lactiplantibacillus plantarum subsp. plantarum strain KB-11. [Figure 2] Figure 2 shows a micrograph of Lactiplantibacillus plantarum subsp. plantarum KB-11 strain after Gram staining. DETAILED DESCRIPTION OF THE INVENTION

[0014] First Embodiment A first embodiment of the novel lactic acid bacterium of the present invention will be described below. (Novel lactic acid bacteria) The novel lactic acid bacterium is Lactiplantibacillus plantarum subsp. plantarum KB-11 strain (accession number: NITE P-04017) (hereinafter referred to as "lactic acid bacterium KB-11 strain"). This lactic acid bacterium is a novel strain of Lactiplantibacillus plantarum isolated from bee bread collected from a honeycomb. This lactic acid bacterium was deposited with the Patent Microorganisms Depositary Center of the National Institute of Technology and Evaluation (2-5-8 Kazusa Kamatari, Kisarazu City, Chiba Prefecture) under the above accession number on November 22, 2023.

[0015] The new lactic acid bacteria strain was identified as Lactiplantibacillus plantarum by BLAST homology search against DB-BA and international base sequence databases (DDBJ / ENA (EMBL) / GenBank) using the microbial identification system "ENKI" (Techno Suruga Lab Co., Ltd.) and evaluation of morphological and physiological properties (sugar fermentation ability, etc.).

[0016] (How to obtain lactic acid bacteria) The lactic acid bacteria strain KB-11 can be obtained from the Patent Microorganisms Depositary using the above accession number, or can be obtained by using beekeeping products such as bee bread as a material for isolating the bacteria. For example, the novel lactic acid bacteria can be obtained by using beekeeping products such as bee bread as a material for isolating the bacteria using the isolation method for the novel lactic acid bacteria described in the section "Test Example 1: Isolation and Identification of Novel Lactic Acid Bacteria" below.

[0017] (others) The lactic acid bacteria KB-11 strain of this embodiment may be formulated as a composition containing the lactic acid bacteria KB-11 strain prepared as a solid together with an excipient, or may be formulated as a composition containing the lactic acid bacteria KB-11 strain mixed with a raw material for fermentation.

[0018] (Effects of this embodiment) The effects of the lactic acid bacteria strain KB-11 of the first embodiment will be described. (1-1) The lactic acid bacterium strain KB-11 of the first embodiment is a novel lactic acid bacterium isolated from bee bread, a beekeeping product. The lactic acid bacterium strain KB-11 has a novel, never-reported property that differs from the conventionally known Lactiplantibacillus plantarum subsp. plantarum in that it does not ferment L-arabinose or D-turanose. For example, the lactic acid bacterium strain KB-11 differs from the lactic acid bacterium strain NITE P-03374 and the lactic acid bacterium strain KCTC11403BP in that it ferments α-methyl-D-mannoside. The lactic acid bacterium strain KB-11 differs from the lactic acid bacterium strain KCTC11403BP in that it ferments arbutin and gluconate.

[0019] Second Embodiment A second embodiment of the fermentation composition of the present invention will now be described. The fermented composition of this embodiment is a fermented composition obtained using the lactic acid bacteria strain KB-11 of the first embodiment. Because the lactic acid bacteria strain KB-11 is Lactiplantibacillus plantarum, it can be used to produce a variety of known lactic acid bacteria-fermented foods and beverages that use lactic acid bacteria. Examples of lactic acid bacteria-fermented foods and beverages include fermented foods such as processed grain products, dairy products, seafood, processed animal products, processed plant products, bee products, and fungal materials. The lactic acid bacteria strain KB-11 is particularly preferably used in the fermentation of a grain saccharified liquid obtained by saccharifying grains. By applying the lactic acid bacteria strain KB-11 to the fermentation of a grain saccharified liquid, the flavor and / or aroma of the resulting fermented composition of the grain saccharified liquid can be improved.

[0020] (grain saccharified liquid) Grain saccharified liquid is a liquid produced by saccharifying and liquefying grains as a raw material, and is used, for example, as a seasoning for foods and beverages, sweeteners, and the like.

[0021] The raw materials for the grain saccharified solution are grains and water. The grains are not particularly limited as long as they contain polysaccharides such as starch and dietary fiber and can be saccharified and liquefied to produce low-molecular-weight sugars. Specific examples of grains include rice, wheat, corn, sorghum, barnyard millet, foxtail millet, millet, barley, oats, rye, and millet.

[0022] Among these, saccharified grain liquid obtained from oats, rice, or millet is improved in flavor and / or aroma by fermentation with lactic acid bacteria KB-11, particularly by reducing the distinctive grain odor, fermented odor, and sour odor of the saccharified grain liquid, improving the balance of aromas, reducing the grainy feel and sourness, and improving the palatability. Furthermore, saccharified grain liquid obtained from oats is improved in flavor and / or aroma by fermentation with lactic acid bacteria KB-11, particularly by reducing the grain odor, fermented odor, and sour odor, reducing the grainy feel and sourness. Furthermore, saccharified grain liquid obtained from rice or millet is improved in flavor and / or aroma by fermentation with lactic acid bacteria KB-11, particularly by improving the balance of aromas, reducing the sourness, and improving the palatability.

[0023] The grain saccharified liquid may be a commercially available product, or may be produced using the above-mentioned grains as a raw material by a known method for producing a grain saccharified liquid. The method for producing the grain saccharified liquid is not particularly limited, and can be carried out, for example, by the following method.

[0024] First, a predetermined volume of water is added to the grain raw material, which has been pulverized if necessary, and high-temperature treatment is performed to swell the grain and gelatinize the starch. Next, polysaccharide-degrading enzymes such as amylase, cellulase, and pectinase, and protease if necessary, are added, and saccharification and liquefaction are carried out by enzymatic treatment at a predetermined temperature. In this process, saccharification and liquefaction can be promoted by combining micronization or pulverization treatment using a homogenizer or the like.

[0025] (Method for fermenting grain saccharified liquid) The fermentation method for the grain saccharified liquid can be carried out by inoculating a predetermined amount of the lactic acid bacteria KB-11 strain into the grain saccharified liquid obtained as described above and culturing it at a predetermined temperature. The culture temperature is set appropriately, but is preferably 15°C to 35°C, more preferably 25°C to 30°C. The culture time is set appropriately depending on the culture temperature, but is preferably 5 hours to 40 hours, more preferably 15 hours to 30 hours. The pH of the grain saccharified liquid is preferably near neutral, specifically, preferably 5 to 8, and more preferably 6 to 7.5.

[0026] (Effects of this embodiment) The effects of the fermentation composition of the second embodiment will be described. (2-1) The fermented composition of this embodiment is a fermented composition obtained using the lactic acid bacteria strain KB-11 of Embodiment 1. The lactic acid bacteria strain KB-11 is Lactiprunculus plantarum, and therefore can be used to produce a variety of known lactic acid bacteria-fermented foods and beverages that use lactic acid bacteria.

[0027] (2-2) When the fermentation composition of this embodiment is a fermentation composition in which a grain saccharified liquid obtained by saccharifying grains is fermented with the lactic acid bacteria KB-11 strain, the taste and / or flavor of the grain saccharified liquid can be improved.

[0028] (Example of change) The above embodiment may be modified as follows: The above embodiment and the following modifications may be combined with each other within the scope of technical compatibility.

[0029] In the above embodiment, the raw material grain saccharified liquid or the fermented composition obtained by fermentation may be appropriately blended with acidulants such as citric acid, sugars such as sucrose, sweeteners, vitamins, minerals, flavorings, stabilizers, thickeners, pH adjusters, etc., within the scope that does not impair the effects of the present invention.

[0030] The form of the fermented composition of the above embodiment is not particularly limited, and examples thereof include liquids, dried powders, flakes, granules, and pastes. Furthermore, the form of foods and beverages using the fermented composition of the above embodiment is not particularly limited, and examples thereof include hard capsules, soft capsules, supplements, chewable tablets, beverages, powdered beverages, granules, and films. When used as foods and beverages, the fermented composition can be in the form of beverages such as tea drinks, sports drinks, beauty drinks, fruit juice drinks, carbonated drinks, alcoholic drinks, soft drinks, jelly drinks, concentrated beverages to be diluted with water, hot water, carbonated water, etc.; dry solids such as powders, granules, and tablets to be dissolved or suspended in water, hot water, etc.; tablet confectioneries, jellies, snacks, baked goods, fried goods, cakes, chocolates, gums, candies, gummy candies, soups, noodles, rice dishes, cereals, and other foods. [Example]

[0031] The present invention will be described in more detail below with reference to examples. Note that 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.

[0032] (Test Example 1: Isolation and identification of novel lactic acid bacteria) The isolation and identification of the novel lactic acid bacteria of the present invention will now be described. (1) Isolation of new lactic acid bacteria After crushing the bee bread, a predetermined amount was suspended in water, and the supernatant was serially diluted. The mixture was then spread onto YP chalk agar medium (containing 10 ppm each of sodium azide and cycloheximide) containing various sugars (mannose, mannitol, maltose, and fructose) as carbon sources. A large number of colonies that formed clear zones after dissolving calcium carbonate were isolated. If necessary, enrichment cultures were performed in YP liquid medium of the same composition, followed by the above isolation procedure.

[0033] Among the obtained microbial strains, one was selected that was a non-motile, Gram-positive, anaerobic bacillus, did not form spores, fermented glucose, showed negative catalase and oxidase reactions, could grow on MRS agar medium, and showed high bacterial cell growth.The obtained strain (lactic acid bacteria strain KB-11) was identified by analysis of the 16S rDNA gene and various biochemical properties.

[0034] (2) Genetic characteristics The culture conditions and reagents used are as shown in Tables 1 and 2. Analysis of the 16S rDNA gene revealed that the partial 16S rDNA sequence (1491 bases in total) of the lactic acid bacteria strain KB-11 showed 99.9% identity with Lactiplantibacillus plantarum (L. plantarum subsp. plantarum) NBRC15891 (LC064896). The partial 16S rDNA gene sequence of the lactic acid bacteria strain KB-11 obtained is shown in SEQ ID NO: 1. SEQ ID NO: 1 represents the region from bases 21 to 1511 (1491 bases in total) of the 16S rDNA gene of the lactic acid bacteria strain KB-11.

[0035] Figure 1 shows a simplified molecular phylogenetic tree analyzed based on the partial 16S rDNA sequence of the lactic acid bacterium strain KB-11 obtained by a homology search against DB-BA. The line in the upper left corner of Figure 1 is the scale bar, the numbers at the phylogenetic branches are bootstrap values, and the T at the end of the strain name indicates the type strain for that species.

[0036] (3) Biochemical property analysis The results of morphological observation and physiological properties of the lactic acid bacteria strain KB-11 are shown in Table 3. Photographs of morphological observation using an optical microscope after Gram staining are shown in Figure 2. The results of the sugar fermentation test using API 50 CHL (manufactured by BioMérieux) are shown in Table 4. These properties of the KB-11 strain were almost identical to those of Lactiplantibacillus plantarum subsp. plantarum.

[0037] [Table 1]

[0038] [Table 2]

[0039] [Table 3]

[0040] [Table 4] (4) Specificity of Lactic Acid Bacteria Strain KB-11 The DNA encoding the 16S rDNA of the lactic acid bacterium strain KB-11 was confirmed to differ from the type strain Lactiprantibacillus plantarum (L. plantarum subsp. plantarum) NBRC15891 (LC064896) in the following two respects.

[0041] (i) The T (thymine) at position 214 of the reference strain has been replaced with a C (cytosine). (ii) The 229th C (cytosine) of the reference strain is substituted with Y (C, T: cytosine or thymine).

[0042] Table 4 shows the results of sugar fermentation tests for Lactobacillus KB-11, Lactibacillus plantarum NBRC 15891, Lactibacillus plantarum KCTC 11403BP, and Lactibacillus plantarum NITE P-03374. Lactibacterium KB-11 differs from previously known Lactibacillus plantarum (L. plantarum subsp. plantarum) in that it does not ferment L-arabinose or D-turanose. It was therefore inferred to be a novel Lactibacillus plantarum with unprecedented novel properties. For example, Lactibacterium KB-11 differs from Lactibacterium NITE P-03374 and Lactibacterium KCTC 11403BP in that it ferments α-methyl-D-mannoside. The lactic acid bacteria strain KB-11 differs from the lactic acid bacteria strain KCTC11403BP in that it ferments arbutin and gluconate.

[0043] (Test Example 2: Test to improve the flavor and aroma of cereal saccharified liquid) Lactic acid bacteria strain KB-11 was used to ferment grain saccharified liquid, and its effect on improving the flavor and aroma of the grain saccharified liquid was evaluated.

[0044] The grain saccharification liquids used were oat saccharification liquid (OAT saccharification liquid: manufactured by API Co., Ltd.), rice saccharification liquid (RICE saccharification liquid: manufactured by API Co., Ltd.), and miscellaneous grain saccharification liquid (miscellaneous grain saccharification liquid: manufactured by API Co., Ltd.), all adjusted to a pH of around 7.

[0045] As the lactic acid bacteria, the Lactobacillus KB-11 strain was used as an example, and the Lactiprantibacillus plantarum NBRC15891 strain was used as a comparative example. Each strain was inoculated into MRS medium and subjected to static culture at 30°C in the atmosphere for 24 hours. A 1 / 100 volume of the pre-culture solution was added to the raw material grain saccharified liquid, and static culture was performed at 30°C in the atmosphere for 24 hours. The fermented grain saccharified liquid obtained was evaluated for its effect of improving flavor and aroma based on the following criteria. To evaluate the improvement in flavor and aroma more specifically, the aroma was evaluated based on the strength of the grain odor, the strength of the fermented odor or sour odor, and the balance of the aroma. The taste was evaluated based on the strength of the grain flavor, sweetness, sourness, astringency or bitterness, and deliciousness.

[0046] Seven panelists rated the flavor and aroma of the fermented liquid on a scale of -5 (very weak) to +5 (very strong), with the non-fermented control being given a rating of 0. The average value was then calculated, excluding the maximum and minimum values ​​from the seven panelists. The results are shown in Tables 5 to 7. The difference indicates the difference between the value for the Example and the value for the Comparative Example.

[0047] [Table 5]

[0048] [Table 6]

[0049] [Table 7] As shown in Tables 5 to 7, the fermented products of all grain saccharification liquids showed a tendency to improve in the effects of reducing fermentation odor and acidic odor, improving the balance of aroma, reducing grain flavor and acidity, and improving palatability.

[0050] When the oat saccharified solution in Table 5 was fermented, it was confirmed that the Examples were more effective in reducing grain odor, fermented odor, and acidic odor than the Comparative Examples, and that the Examples were more effective in reducing grain flavor and acidity than the Comparative Examples.

[0051] When the rice saccharified solution in Table 6 was fermented, it was confirmed that the Examples had a particularly improved effect on the aroma balance compared to the Comparative Examples, and that the Examples also had a particularly improved effect on reducing sourness and improving palatability compared to the Comparative Examples.

[0052] When the saccharified cereal liquids in Table 7 were fermented, it was confirmed that the Examples had a particularly improved effect on improving the balance of aroma compared to the Comparative Examples. It was also confirmed that the Examples had a particularly improved effect on reducing sourness and improving palatability compared to the Comparative Examples.

Claims

1. A novel Lactiplantibacillus plantarum, which is Lactiplantibacillus plantarum subsp. plantarum strain KB-11 (accession number: NITE P-04017).

2. The novel Lactipruncibacillus plantarum according to claim 1, which is used to improve the taste and / or flavor of a grain saccharified liquid by fermentation.

3. A composition comprising the Lactiprunci Bacillus plantarum of claim 1.

4. A fermented composition obtained using the Lactiprunci Bacillus plantarum according to claim 1.

5. The fermentation composition according to claim 4, comprising a fermentation product obtained by using Lactiplantibacillus plantarum to ferment a grain saccharified liquid obtained by saccharifying grain.

6. The fermented composition according to claim 5, wherein the grain is at least one selected from oats, rice, and millet.

7. The fermented composition according to claim 6, wherein the grain is oats.

8. The fermented composition according to claim 6 , wherein the grain is rice or millet.

9. A method for improving the flavor and / or aroma of a cereal saccharified liquid, which comprises fermenting the cereal saccharified liquid using the Lactiprunci Bacillus plantarum according to claim 1.

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