JP6723811B2 - Method for producing sesaminol - Google Patents

Description

The present invention relates to a method for producing sesaminol. The present invention also relates to novel lactic acid bacteria having the ability to produce sesaminol from sesaminol glycosides. The present invention also relates to a fermented product of a sesaminol glycoside-containing raw material by lactic acid bacteria. The present invention also relates to novel compositions containing sesaminol.

Sesame seeds (Sesamum indicum L.) seeds (hereinafter sometimes simply referred to as "sesame seeds") contain lignans. About 80% of sesame lignans are sesamin or sesamolin, and the remaining about 20% are lignan phenols. One of the lignan phenols contained in sesame is sesaminol. Most of lignanphenols in sesame exist as glycosides, and few exist as aglycones (free forms) (Non-patent Document 1).

Unlike sesamin, sesaminol is known to have antioxidant activity by itself, and is expected to be applied in the fields of health foods, medicines and the like.

As described above, sesaminol exists as a glycoside in natural products such as sesame. The sesaminol glycoside has a structure in which a sugar chain is bonded to the phenolic hydroxyl group of sesaminol. The phenolic hydroxyl group is considered to be the active group for the antioxidative activity of sesaminol, and it is considered that no activity is exhibited in the form of sesaminol glycoside. Since sesaminol glycoside has hydrophilicity, it differs from lipophilic sesamin in that it is contained in a large amount in the pressed meal produced as a by-product when sesame oil is pressed.

The following techniques have been disclosed as techniques for producing sesaminol by releasing sesaminol from sesaminol glycoside.

Patent Document 1 discloses that an enzyme produced by a microorganism of the genus Panjebacillus has an activity of cleaving the sugar chain of sesaminol glycoside.

Patent Document 2 discloses that a crushed product of whole-grain raw sesame is decomposed with Rhizopus oligosporus crude enzyme, and lactic acid fermentation is carried out before isoleucine is produced to produce fermented sesame having an antioxidant power. .. Specific examples of lactic acid bacteria include Lactobacillus bulgaricus and Streptococcus thermophilus. Patent Document 2 describes that this fermented sesame contains sesaminol, but does not disclose at what stage and by what mechanism sesaminol is produced.

Patent Document 3 discloses a method for producing sesaminol, which comprises incubating a lignan such as sesaminol triglucoside or a substrate material containing the same in the presence of Bacillus bacteria and Enterococcus bacteria.

Patent Document 4 discloses fermenting a raw material such as sesame with an Aspergillus strain or a Bacillus strain in order to produce a sesame fermented product containing a sesame lignan glycoside and an aglycone sesame lignan.

Japanese Patent No. 4839231 Japanese Patent No. 3261075 JP, 2006-61115, A JP, 2007-319157, A

Ishiyama et al., Japan Society for Food Science and Technology, Vol. 53, No. 1, January 2006

As described above, a technique of fermenting sesame by a microorganism to produce sesaminol and enhancing antioxidant activity has been developed so far.

However, since the microorganisms or enzymes used in Patent Documents 1 to 4 are not generally edible, there is concern about the safety of the produced sesaminol.

Therefore, the present invention provides a new means for producing sesaminol from a raw material containing sesaminol glycosides using lactic acid bacteria.

The present inventors have surprisingly found that, among lactic acid bacteria, lactic acid bacteria belonging to the genus Lactobacillus have the ability to hydrolyze sesaminol glycosides to produce sesaminol. Therefore, the present inventors provide the following inventions.
(1) A sesaminol-producing step of producing a sesaminol from the sesaminol glycoside by allowing a raw material containing a sesaminol glycoside to coexist with one or more lactic acid bacteria belonging to the genus Lactobacillus. Manufacturing method.
(2) The lactic acid bacteria are DNBL1826 strain (accession number: NITE P-02225), DNBL1832 strain (accession number: NITE P-02229), DNBL1829 strain (accession number: NITE P-02226), DNBL1830 strain (accession number: NITE). P-02227), DNBL1831 strain (accession number: NITE P-02228), and Lactobacillus casei Hasegawa strain (accession number: FERM BP-10123), which is one or more and is described in (1). Method.
(3) The method according to (1), wherein the lactic acid bacterium is a combination of two or more lactic acid bacteria belonging to the genus Lactobacillus.
(4) The lactic acid bacterium has the following (A) and (B):
(A) one or more lactic acid bacteria belonging to Lactobacillus paracasei and having the ability to convert sesaminol triglucoside to sesaminol diglucoside;
(B) belongs to Lactobacillus paracasei and belongs to one or more other lactic acid bacteria, Lactobacillus pentosus, which has the ability to convert sesaminol diglucoside to sesaminol, and has the ability to convert sesaminol diglucoside to sesaminol A combination of at least one lactic acid bacterium and one or more lactic acid bacteria belonging to Lactobacillus plantarum and having the ability to convert sesaminol diglucoside to sesaminol, selected from the group consisting of at least one lactic acid bacterium (3 ).
(5) The lactic acid bacterium of (A) above is one or more selected from the DNBL1826 strain (accession number: NITE P-02225) and the DNBL1832 strain (accession number: NITE P-02229),
The lactic acid bacterium of (B) is DNBL1829 strain (accession number: NITE P-02226), DNBL1830 strain (accession number: NITE P-02227), DNBL1831 strain (accession number: NITE P-02228), and Lactobacillus casei. -The method according to (4), which is 1 or more selected from Hasegawa strain (accession number: FERM BP-10123).
(6) DNBL1826 strain (accession number: NITE P-02225).
(7) DNBL1832 strain (accession number: NITE P-02229).
(8) DNBL1829 strain (accession number: NITE P-02226).
(9) DNBL1830 strain (accession number: NITE P-02227).
(10) DNBL1831 strain (accession number: NITE P-02228).
(11) A fermented product of a raw material containing a sesaminol glycoside with one or more lactic acid bacteria belonging to the genus Lactobacillus.
(12) The lactic acid bacteria are DNBL1826 strain (accession number: NITE P-02225), DNBL1832 strain (accession number: NITE P-02229), DNBL1829 strain (accession number: NITE P-02226), DNBL1830 strain (accession number: NITE). P-02227), DNBL1831 strain (accession number: NITE P-02228), and Lactobacillus casei Hasegawa strain (accession number: FERM BP-10123), which is one or more and is described in (11). Fermented material.
(13) sesamin, and
Containing 100 mg or more of sesaminol per 100 g on a dry weight basis,
A composition containing sesaminol, wherein the weight ratio of sesaminol:sesamin is in the range of 1:0.1 to 1:20.

According to the method for producing sesaminol of the present invention, sesameminol can be produced using lactic acid bacteria of the genus Lactobacillus.
The lactic acid bacterium of the present invention can be used for producing sesaminol.
The sesaminol-containing composition of the present invention is rich in sesaminol and has a unique sesaminol:sesamin weight ratio.

FIG. 1 shows the results of confirmation by thin layer chromatography of the product in the middle of fermentation when fermented with lactic acid bacteria using a ground sesame hot water extract as a substrate in Experiment 1. FIG. 1A shows thin layer chromatography when DNBL1826 strain was fermented alone or in combination with other lactic acid bacteria. FIG. 1B shows thin layer chromatography when the DNBL1832 strain was used alone or in combination with other lactic acid bacteria for fermentation. FIG. 2 shows the results of confirmation by thin-layer chromatography of the product during fermentation when lactic acid bacteria were fermented with the sesame-pressed meal waste hot water extract as a substrate in Experiment 1. FIG. 2A shows thin layer chromatography when DNBL1826 strain was fermented alone or in combination with other lactic acid bacteria. FIG. 2B shows thin layer chromatography when DNBL1832 strain was fermented alone or in combination with other lactic acid bacteria. FIG. 3 shows the production during fermentation when the lactic acid bacterium was fermented using a hot water extract of ground sesame seeds in Experiment 1 as a substrate and a combination of the DNBL1826 strain and the A221 strain or a combination of the DNBL1832 strain and the A221 strain. The result of confirmation by thin layer chromatography of the product is shown. FIG. 4 shows the fifth day of fermentation in a test using a combination of DNBL1826 strain and A221 strain or combination of DNBL1832 strain and A221 strain in Experiment 1 and a hot water extract of ground sesame seeds as a substrate. The HPLC chart of a product is shown. FIG. 4a is a chart showing the retention time of the standard product of STG, FIG. 4b is the standard product of β1,2SDG, FIG. 4c is the standard product of β1,6SDG, and FIG. .. Fig. 4e shows an HPLC chart of the test group not treated with lactic acid bacteria, Fig. 4f shows an HPCL chart of the test group using the DNBL1826 strain and A221 strain in combination, and Fig. 4g shows a test group using the DNBL1832 strain and A221 strain in combination. An HPCL chart is shown. FIG. 5 shows the fermentation treatment on the 4th day in the test in Experiment 1 using only the DNBL1832 strain as the lactic acid bacterium, or a combination of the DNBL1832 strain and the A221 strain, and using the hot water extract of the commercially available sesame press cake as the substrate. The result of thin layer chromatography is shown. FIG. 6 shows the analysis results by HPLC of the ground sesame suspension after fermentation with lactic acid bacteria in Experiment 2. FIG. 6A shows an HPLC chart of a test plot not treated with lactic acid bacteria. FIG. 6B shows an HPLC chart of the test section using only DNBL1826 strain. FIG. 6C shows an HPLC chart of a test plot using a combination of DNBL1826 strain and DNBL1829 strain. FIG. 7 shows the analysis results by HPLC of the freeze-dried composition (the substrate is ground sesame suspension) after fermentation in Experiment 3. FIG. 7A shows an HPLC chart of a test plot not treated with lactic acid bacteria. FIG. 7B shows an HPLC chart of a test plot using a combination of DNBL1826 strain and DNBL1829 strain. FIG. 8 shows the analysis results by HPLC of the freeze-dried composition after fermentation (the substrate is a sesame-pressed meal suspension) in Experiment 4. FIG. 8A shows an HPLC chart of a test plot not treated with lactic acid bacteria. FIG. 8B shows an HPLC chart of a test plot using a combination of DNBL1826 strain and DNBL1829 strain. FIG. 9 shows the results of confirmation by thin layer chromatography of the product on the second day of fermentation when lactic acid bacteria were fermented using the ground hot sesame extract as a substrate in Experiment 5. FIG. 10 shows the confirmation results by thin layer chromatography of the product on the 7th day of fermentation when lactic acid bacteria were fermented using the ground hot sesame extract as a substrate in Experiment 5. FIG. 11 shows the results of confirmation by thin layer chromatography of the product on the second day of fermentation when lactic acid bacteria were fermented using the sesame-pressed meal waste hot water extract as a substrate in Experiment 6. FIG. 12 shows the confirmation results by thin layer chromatography of the product on the 7th day of fermentation when lactic acid bacteria were fermented using the sesame-pressed meal waste hot water extract as a substrate in Experiment 6. FIG. 13 shows the confirmation results by thin layer chromatography of the product on the 7th day of fermentation when lactic acid bacteria were fermented with a ground sesame extract as a substrate in Experiment 7. FIG. 14 shows the results of confirmation by thin layer chromatography of the product on the seventh day of fermentation when lactic acid bacteria were fermented using the sesame-pressed meal waste hot water extract as a substrate in Experiment 8.

<Sesaminol and sesaminol glycoside>
In the present invention, when simply referred to as “sesaminol”, it refers to the form of an aglycone having no sugar chain, unless otherwise specified.

Sesaminol is known to be four compounds having the same planar structure but different steric configurations, namely (+)-sesaminol, 2-epi-sesaminol, 6-epi-sesaminol, and diacesaminol. ing. In the present invention, the term "sesaminol" refers to one or more of these sesaminols, and may be a mixture of a plurality of types. Typically, sesaminol comprises at least (+)-sesaminol. The amount of sesaminol refers to the total amount of sesaminol when the compounds are included as sesaminol.

As the sesaminol glycoside, one or more species selected from the group consisting of sesaminol triglucoside, sesaminol diglucoside and sesaminol monoglucoside are known, and a mixture of multiple species may be used. Typically, the sesaminol glycoside comprises at least sesaminol triglucoside. The most abundant sesaminol glycoside in sesame is sesaminol triglucoside.

The structure of the conventionally known sesaminol triglucoside (STG) is shown in the following formula.

The sugar chain is one in which three glucoses are bound, and one glucose is bound to the central glucose via the β1,6-glycoside bond and the β1,2-linkage, and the central glucose is ( It is bonded to the phenolic hydroxyl group of +)-sesaminol by an O-glycoside bond.

<raw material>
The "raw material containing sesaminol glycoside" used in the present invention may contain sesaminol glycoside, and may consist of sesaminol glycoside alone, or sesaminol glycoside together with other components. It may be a composition containing the body. The composition containing sesaminol glycoside is more preferably a raw material derived from sesame. Examples of the sesame-derived raw material include one or more selected from sesame seeds, sesame seed crushed products, sesame seed defatted meal, and extracts from these with a solvent. As the ground product of sesame seeds, those used for food as "ground sesame" can also be used. As the defatted lees of sesame seeds, those produced as a by-product of sesame oil production can be used. An example of this by-product is sesame pressed cake. The variety of sesame is not particularly limited. Any sesame seeds that produce white sesame seeds, varieties that produce black sesame seeds, and other sesame seeds may be used. The raw material derived from sesame may be further mixed with other components. The solvent may be any solvent capable of eluting the sesaminol glycoside, and examples thereof include water and a hydrophilic solvent containing water, and water is particularly preferable.

<Method for producing sesaminol>
The method for producing sesaminol according to the present invention is a sesaminol that produces sesaminol from the sesaminol glycoside by allowing a raw material containing the sesaminol glycoside to coexist with one or more lactic acid bacteria belonging to the genus Lactobacillus. Including a production process.
In the present invention, as the lactic acid bacterium belonging to the genus Lactobacillus, a lactic acid bacterium belonging to the genus Lactobacillus having the ability to produce sesaminol from sesaminol glycoside is used.

Lactobacillus belonging to the genus Lactobacillus is a lactic acid bacterium that has been eaten, so the safety of sesaminol produced is high. Among the lactic acid bacteria belonging to the genus Lactobacillus, one or more selected from the group consisting of lactic acid bacteria belonging to Lactobacillus paracasei, lactic acid bacteria belonging to Lactobacillus pentosus, and lactic acid bacteria belonging to Lactobacillus plantarum are particularly preferable.

As a lactic acid bacterium belonging to Lactobacillus paracasei, which has the ability to produce sesaminol from sesaminol glycoside, DNBL1826 strain (accession number: NITE P-02225), DNBL1832 strain (accession number: NITE P-02229) are particularly preferable. , And one or more selected from the group consisting of Lactobacillus casei Hasegawa strain (accession number: FERM BP-10123). The Lactobacillus casei Hasegawa strain (accession number: FERM BP-10123) was initially classified as Lactobacillus casei, but is now classified as Lactobacillus paracasei, and in this specification, Lactobacillus paracasei.・Handle as lactic acid bacteria belonging to Paracasei. The Lactobacillus casei Hasegawa strain may be referred to as "A221 strain" in the present specification.

Lactobacillus belonging to Lactobacillus plantarum having the ability to produce sesaminol from sesaminol glycosides include DNBL1829 strain (accession number: NITE P-02226) and DNBL1831 strain (accession number: NITE P-02228). 1) or more selected from the group consisting of

DNBL1830 strain (accession number: NITE P-02227) is preferable as the lactic acid bacterium belonging to Lactobacillus pentosus having the ability to produce sesaminol from sesaminol glycoside.

In the present invention, the above-mentioned 6 types of specific lactic acid bacterial strains are concepts including their mutant strains. The mutant may be a mutant having the ability to produce sesaminol from sesaminol glycoside, may be a mutant produced by artificial mutation treatment, or may be a natural mutant. Good.

In the present invention, as a lactic acid bacterium belonging to the genus Lactobacillus, which has the ability to produce sesaminol from sesaminol glycoside, particularly DNBL1829 strain (accession number: NITE P-02226), DNBL1830 strain (accession number: NITE P- 02227), DNBL1831 strain (accession number: NITE P-02228) and Lactobacillus casei Hasegawa strain (accession number: FERM BP-10123), and it is preferable to use one or more thereof. Since these lactic acid bacteria have a particularly high ability to produce sesaminol from sesaminol diglucoside, they are suitable when a raw material containing sesaminol diglucoside is used as the sesaminol glycoside.

In the present invention, it is particularly preferable to use a Lactobacillus casei Hasegawa strain (accession number: FERM BP-10123) as a lactic acid bacterium belonging to the genus Lactobacillus, which has the ability to produce sesaminol from sesaminol glycoside. .. This lactic acid bacterium has a particularly high ability to produce sesaminol diglucoside from sesaminol triglucoside and an ability to produce sesaminol from sesaminol diglucoside, and can efficiently produce sesaminol.

The present inventors have surprisingly found that in the sesaminol production step, when a combination of two or more lactic acid bacteria belonging to the genus Lactobacillus is used as the lactic acid bacterium, the production of sesaminol is significantly promoted. ..

As the combination,
One or more lactic acid bacteria belonging to the genus Lactobacillus and having the ability to convert sesaminol triglucoside to sesaminol diglucoside;
A combination with one or more other lactic acid bacteria belonging to the genus Lactobacillus and having the ability to convert sesaminol diglucoside into sesaminol is preferable, and particularly, the following (1) and (2):
(1) One or more lactic acid bacteria belonging to Lactobacillus paracasei and capable of converting sesaminol triglucoside to sesaminol diglucoside,
(2) belongs to Lactobacillus paracasei and belongs to one or more other lactic acid bacteria, Lactobacillus pentosus, which has the ability to convert sesaminol diglucoside to sesaminol, and has the ability to convert sesaminol diglucoside to sesaminol A combination of one or more lactic acid bacteria and one or more lactic acid bacteria belonging to Lactobacillus plantarum and selected from the group consisting of one or more lactic acid bacteria having the ability to convert sesaminol diglucoside to sesaminol is preferred.

The mechanism by which sesamemin production is promoted by the combination of lactic acid bacteria is not always clear, but the following mechanism is presumed from the test results shown in the examples.

The reaction in which sesaminol glycoside is converted to sesaminol is the reaction in which sesaminol triglucoside is converted to sesaminol diglucoside and the reaction in which sesaminol diglucoside is converted to sesaminol via sesaminol monoglucoside. Is presumed to contain. Among the lactic acid bacteria belonging to the genus Lactobacillus, there are those with a particularly high ability to convert sesaminol triglucoside to sesaminol diglucoside, and those with a particularly high ability to convert sesaminol diglucoside to sesaminol. It is presumed that sesameminol can be efficiently produced by using two or more lactic acid bacteria belonging to the same category together. The sesaminol diglucoside produced as an intermediate here is presumed to be mainly sesaminol diglucoside (β1,6 SDG) having a sugar chain bound by β1,6 bonds, but the sugar chain bound by β1,2 bonds It may also be sesaminol diglucoside (β1,2 SDG). Therefore, "sesaminol diglucoside" in the "ability to convert sesaminol triglucoside to sesaminol diglucoside" and "ability to convert sesaminol diglucoside to sesaminol" described in the present specification is mainly β1, 6
It refers to SDG, but is not limited to this, and may be β1,2 SDG.

As the lactic acid bacterium of the above (1), one having the ability to convert sesaminol triglucoside to sesaminol diglucoside is used. As such lactic acid bacteria, it is preferable to use one or more selected from the DNBL1826 strain (accession number: NITE P-02225) and the DNBL1832 strain (accession number: NITE P-02229). As described above, the mutant strain is also included in the range of these specific strains used as the lactic acid bacterium of the above (1). The mutant strain may be an artificial mutant strain or a natural mutant strain as long as it is a mutant strain having an ability to convert sesaminol triglucoside to sesaminol diglucoside.

As the lactic acid bacterium of the above (2), one having the ability to convert sesaminol diglucoside to sesaminol is used. Examples of such lactic acid bacteria include DNBL1829 strain (accession number: NITE P-02226), DNBL1830 strain (accession number: NITE P-02227), DNBL1831 strain (accession number: NITE P-02228), and Lactobacillus casei. It is particularly preferable to use one or more selected from Hasegawa strain (accession number: FERM BP-10123). As described above, the mutant strain is also included in the range of these specific strains used as the lactic acid bacterium of the above (2). The mutant strain may be an artificial mutant strain or a natural mutant strain, as long as it is a mutant strain capable of converting sesaminol diglucoside into sesaminol.

In the following description, “lactic acid bacterium” includes both one lactic acid bacterium and a combination of two or more lactic acid bacteria unless otherwise specified.
The conditions for allowing the raw material to coexist with the lactic acid bacterium are not particularly limited as long as the sugar chain in the sesaminol glycoside in the raw material is cleaved to produce sesaminol by the action of the lactic acid bacterium.

The reaction mixture in which the raw material is allowed to coexist with the lactic acid bacterium preferably further contains a suitable liquid medium for efficiently contacting the raw material and the lactic acid bacterium. Examples of the liquid medium include water. Here, the water may be a buffer aqueous solution in which an appropriate buffer component is dissolved, or may be a liquid medium containing a nutrient component as described below.

Since the reaction that produces sesaminol is one type of fermentation by the lactic acid bacterium, it is preferable that the reaction mixture contains a nutrient component necessary for the lactic acid bacterium to grow. When a raw material containing the nutritional component in addition to the sesaminol glycoside is used as the raw material, it is not necessary to add a further component to the reaction mixture. For example, the sesame-derived raw material as described above generally contains components necessary for growth of the lactic acid bacterium in addition to sesaminol glycoside. When the raw material does not include a part or all of the components necessary for the growth of the lactic acid bacterium, the deficient components may be appropriately added to the reaction mixture.

In the step of producing sesaminol, the reaction mixture is incubated under conditions in which the lactic acid bacterium can grow and produce sesaminol from sesaminol glycoside. Incubation conditions at this time are not particularly limited, and various conditions such as temperature, time, presence or absence of shaking, and composition of atmosphere can be appropriately selected. Specifically, the temperature at the time of incubation may be 20 to 40°C. The time of incubation may be 24 to 336 hours. The reaction mixture may not be shaken, and may be continuously or intermittently shaken. Air can be exemplified as the atmosphere in which the reaction mixture comes into contact during the incubation.

The sesaminol produced in the sesaminol producing step can be isolated or purified after the sesaminol producing step and recovered, or can be obtained as a sesaminol-containing composition containing sesaminol.

The sesaminol-containing composition can be prepared from the reaction mixture containing the raw material and the lactic acid bacterium by a sesaminol-containing composition preparation step of preparing a sesaminol-containing composition after the sesaminol-producing step. The composition containing sesaminol preferably contains, in addition to sesaminol, other components derived from the raw materials. In the sesaminol-containing composition preparing step, the reaction mixture is treated by combining one or more means such as drying, concentration, extraction, separation of sesaminol-containing fraction, and sterilization to obtain a sesaminol-containing composition.

When the raw material is a sesame-derived raw material, in the sesaminol-containing composition preparation step, preferably, the sesameminol-containing composition is prepared by drying and sterilizing the reaction mixture after the sesaminol-producing step. The sesaminol-containing composition prepared in this embodiment comprises sesaminol and components derived from sesame and is suitable for ingestion. The sesaminol-containing composition prepared in this embodiment is more preferably the composition of another preferred embodiment of the invention described below.

<Lactic acid bacteria>
The present invention is also a novel lactic acid bacterium: DNBL1826 strain (accession number: NITE P-02225), DNBL1832 strain (accession number: NITE P-02229), DNBL1829 strain (accession number: NITE P-02226), DNBL1830 strain (accession number: NITE). P-02227) and DNBL1831 strain (accession number: NITE P-02228).

These specific lactic acid bacteria are a concept including their mutant strains. The mutant may be a mutant having the ability to produce sesaminol from sesaminol glycoside, may be a mutant produced by artificial mutation treatment, or may be a natural mutant. Good.

The DNBL1826 strain (accession number: NITE P-02225) and the DNBL1832 strain (accession number: NITE P-02229) are useful because they have particularly high ability to convert sesaminol triglucoside to sesaminol diglucoside. These mutants are preferably mutants having the ability to convert sesaminol triglucoside to sesaminol diglucoside.

On the other hand, DNBL1829 strain (accession number: NITE P-02226), DNBL1830 strain (accession number: NITE P-02227), and DNBL1831 strain (accession number: NITE P-02228) convert sesaminol diglucoside into sesaminol. It is useful because its ability to do is particularly high. These mutants are preferably mutants having the ability to convert sesaminol diglucoside to sesaminol.

<Fermented product>
The present invention also relates to a fermented product of a raw material containing a sesaminol glycoside by one or more lactic acid bacteria belonging to the genus Lactobacillus.
In this embodiment of the present invention, the range of the raw material containing the sesaminol glycoside is as described above, and the raw material derived from sesame is preferable.

As the one or more lactic acid bacteria belonging to the genus Lactobacillus, the same lactic acid bacteria as those used in the sesaminol production step can be used. It may be a combination of two or more lactic acid bacteria belonging to the genus Lactobacillus, and preferable combinations are as described above.

The "fermented product of a sesameminol glycoside-containing raw material with one or more lactic acid bacteria belonging to the genus Lactobacillus" refers to a product obtained by fermenting the raw material with the lactic acid bacterium, the raw material and the lactic acid bacterium Fermented by mixing with, liquid, may be in the state of the paste-like or solid fermentation reaction product as it is, one of the separated product obtained by solid-liquid separation of the fermentation reaction product (for example, a supernatant, Precipitation), a processed product thereof, and the like, and a processed product of the fermentation reaction product. The fermentation reaction product may be sterilized, and solid-liquid separation can be performed by means such as centrifugation or filtration. Examples of the processed product of the fermentation reaction product include a concentrated product, a diluted product, a dried product, a purified product (including a crudely purified product) of the fermented product or a separated product obtained by solid-liquid separation. Examples of the purified product include those obtained by purifying or roughly purifying sesaminol from the fermentation reaction product.
Suitable conditions for the fermentation treatment can be the same as those described in the sesaminol production step.

One or more lactic acid bacteria belonging to the genus Lactobacillus may be contained in the fermented product of the present invention in a living state, may be contained in a killed state after sterilization treatment, or may be contained after being removed. You don't have to.

The fermented product of the present invention preferably contains sesaminol.
The fermented product of the present invention is preferably a composition for oral ingestion. Such a fermented product can be used for health foods and nutritional supplements that have a function useful for health.

<Composition containing sesaminol>
Another preferred embodiment of the present invention comprises sesamin and 100 mg or more of sesaminol per 100 g on a dry weight basis, and the weight ratio of sesaminol:sesamin is in the range of 1:0.1 to 1:20. , A composition containing sesaminol.
The upper limit of the content of sesaminol in the composition containing sesaminol is not particularly limited, but is usually 500 mg or less, for example 250 mg or less, per 100 g on a dry weight basis.

The weight ratio of sesaminol: sesamin is more preferably 1:0.1 to 1:15, more preferably 1:0.1 to 1:10, more preferably 1:0.5 to 1:5, more preferably It is 1:0.5 to 1:2.
The obtained sesaminol-containing composition is suitable for oral ingestion as a food or medicine having an antioxidant effect.

In the following examples, the term “sesaminol” simply refers to an aglycone having no sugar chain.

<Lactic acid strain>
In Experiments 1 to 3 described later, "DNBL1826 strain", "DNBL1832 strain", "DNBL1829 strain", "DNBL1830 strain", "DNBL1831 strain", "A221 strain" were used as lactic acid bacterial strains.

The DNBL1826 strain is a lactic acid bacterium isolated by the present inventors from fermented foods and is classified as Lactobacillus paracasei based on 16s rDNA sequence analysis. The DNBL1826 strain has been deposited at the Patent Microorganisms Depositary Center (Postal Code 292-0818, Kazusa, Kazusa, Kisarazu City, Chiba Prefecture, Japan) under the Deposit Management Number: NITE.
P-02225 is given (consignment date: March 23, 2016).

The DNBL1832 strain is a lactic acid bacterium isolated by the present inventors from fermented foods and is classified as Lactobacillus paracasei based on 16s rDNA sequence analysis. The DNBL1832 strain has been deposited at the Patent Microorganism Depositary Center, National Institute of Technology for Product Evaluation, and is assigned the deposit number: NITE P-02229 (deposit date: March 23, 2016).

The DNBL1829 strain is a lactic acid bacterium isolated by the present inventors from fermented foods and is classified as Lactobacillus plantarum based on 16s rDNA sequence analysis. The DNBL1829 strain has been deposited at the Patent Microorganism Depositary Center, National Institute of Technology for Product Evaluation, and is assigned the deposit number: NITE P-02226 (deposit date: March 23, 2016).

The DNBL1830 strain is a lactic acid bacterium isolated by the present inventors from fermented foods and is classified as Lactobacillus pentosus based on 16s rDNA sequence analysis. The DNBL1830 strain has been deposited at the Patent Microorganism Depositary Center, National Institute for Product Evaluation Technology, and is assigned the deposit number: NITE P-02227 (deposit date: March 23, 2016).

The DNBL1831 strain is a lactic acid bacterium isolated by the present inventors from fermented foods and is classified as Lactobacillus plantarum based on 16s rDNA sequence analysis. The DNBL1831 strain has been deposited at the Patent Microorganism Depositary Center, National Institute for Product Evaluation Technology, and is assigned the deposit number: NITE P-02228 (deposit date: March 23, 2016).

The A221 strain refers to the Lactobacillus casei Hasegawa strain disclosed in JP-A-2005-156832 by the present applicant and belongs to the genus Lactobacillus. The A221 strain has been deposited at the Japan Patent Organism Depositary, National Institute of Advanced Industrial Science and Technology (Postal code 305-8566, 1-1, Higashi 1-1-chome, Tsukuba, Ibaraki, Japan), and has been given the deposit number: FERM BP-10123. (Trust date: August 11, 2003, trust number: FERM BP-10123).

<Preparation of lactic acid bacterium pellets and lactic acid bacterium suspension>
Pellets or suspensions of these lactic acid bacterial strains were prepared by the following procedure.

In <Experiment 1> and <Experiment 5> to <Experiment 8>, first, a streak of each strain was pre-streaked on an MRS plate to obtain one platinum loop of the colony, which was then transferred to a 15 ml centrifuge tube (IWAKI). 4 ml of the dispensed general lactic acid bacterium medium (Japanese water) was inoculated. After 3 days of stationary culture at 37° C., the medium was centrifuged at 3500 rpm, the medium was discarded, and then washed with 2 ml of sterile purified water. The cells were well suspended to give a strain suspension, 500 μl each was dispensed into a 1.5 ml microtube (manufactured by Eppendorf), and centrifuged again at 3500 rpm. In the preparation of mixed pellets of different strains, 500 μl of each strain suspension was dispensed in a form of being mixed into a 1.5 ml microtube and centrifuged. The supernatant was discarded and the resulting pellet of lactic acid bacteria strain was used for the test.

In <Experiment 2>, in the same manner as in Experiment 1, colonies and one platinum loop were picked from the plate and inoculated into 4 ml of a general lactic acid bacterium medium. Static culture was carried out at 37° C. for 3 days, and then 1 ml was transferred to a 1.5 ml microtube and centrifuged at 3500 rpm. After discarding the medium, the bacterial cell pellet was suspended in 1 ml of sterile purified water and washed. After centrifuging again at 3500 rpm and discarding the supernatant, the suspension was again suspended in 1 ml of sterilized purified water, and the suspension of lactic acid bacteria was used for the test.

In <Experiment 3> and <Experiment 4>, as in Experiments 1 and 2, colony-platinum loops were picked from the plate and inoculated into 4 ml of a general lactic acid bacterium medium. Static culture was carried out at 37° C. for 3 days, and then the whole amount was added to 45 ml of a general lactic acid bacterium medium. Subsequently, static culture was carried out at 37° C. for 3 days, and then 14 ml was transferred to a 15 ml centrifuge tube. The medium was centrifuged at 3500 rpm, the medium was discarded, and the cells were washed with 10 ml of sterile purified water. Then, it was subjected to centrifugation at 3500 rpm again to obtain a bacterial cell pellet as a precipitate. The supernatant was discarded, resuspended in 10 ml of sterilized purified water, and used for the test.

<Reagent>
In the following description of tests, sesaminol-related substances may be represented by the following abbreviations.
SMN: sesaminol SEM: sesamin STG: sesaminol triglucoside SDG: sesaminol diglucoside β1,2 SDG: SDG having a sugar chain bound by a β1,2 bond
β1,6 SDG: SDG having a sugar chain bound by β1,6 bond

In the following tests, the following commercially available reagents were used as standard products.
SMN standard product: (+) Sesaminol (Nagara Science Co., Ltd.: product number NS182102)
SEM standard product: (+)Sesamin (Nagara Science Co., Ltd.: product number NS180103)
STG standard product: Sesaminol Triglucoside (Nara Science Co., Ltd.: product number NS185102)
β1,2 SDG standard product: Sesaminol (1→2) Diglucoside (manufactured by Nagara Science Co., Ltd.: product number NS185201)
β1,6 SDG standard product: Sesaminol (1→6) Diglucoside (manufactured by Nagara Science Co., Ltd.: product number NS185301)

<Experiment 1: Sesaminol production test>
Using commercially available ground sesame (white) or commercially available sesame pressed cake as a raw material, fermentation was carried out by lactic acid bacteria using the hot water extract as a substrate to try to produce sesaminol.
The lactic acid bacterial strains used are as shown in the results column.

(Experiment 1/Procedure)
10 g of the raw material and 100 ml of purified water were placed in a 300 ml Erlenmeyer flask and autoclaved at 121° C. for 15 minutes.
The suspension after the autoclave treatment was No. 2 Filter paper (manufactured by Advantech) was filtered, and the filtrate was re-sterilized. The filtrate obtained by the re-sterilization was used as a hot water extract.

Thereafter, 1 mL of the hot water extract was added to each lactic acid bacterium pellet prepared in a 1.5 ml microtube by the method described in <Preparation of lactic acid bacterium pellet and lactic acid bacterium suspension> to suspend the lactic acid bacterium pellet.

Fermentation with lactic acid bacteria was carried out by incubating at 37°C while standing without stirring. The first day of starting the fermentation was the 0th day, and the fermentation was continued until the 7th day.
As a negative control, the hot water extract of the raw material was treated in the same procedure except that lactic acid bacteria were not added. This test section was defined as a “untreated lactic acid bacterium” test section.

On the 2nd day (in the case of the following result 1), the 5th day (in the case of the following result 2) or the 4th day (in the case of the following result 3) of the fermentation treatment, a minute amount of the lactic acid bacteria suspension is sampled from each of the microtubes. And developed by thin layer chromatography. For comparison, an aqueous solution of sesaminol standard and a sample of the test section not treated with lactic acid bacteria were similarly developed by thin layer chromatography. For the development of thin layer chromatography, a mixture of ethyl acetate:methanol:water at a ratio of 14:5:4 was used as a developing solvent. A 10% (v/v) aqueous solution of sulfuric acid was used as the color-developing solution after development, and detection was performed by spraying and heating.

On the 7th day (in the case of the following results 1 and 3) or the 5th day (in the case of the following result 2) of the fermentation treatment, the lactic acid bacteria suspension and ethanol from each of the microtubes were mixed at an ethanol concentration of 50% (v /V). Then, the mixture was centrifuged and the supernatant was passed through a 0.45 μm filter (manufactured by Advantech), and the obtained sample was analyzed by high performance liquid chromatography (HPLC). In HPLC (manufactured by Shimadzu Corporation), a reverse phase system using μ Bondasphere (ID 150×3.9 mm, C18 5 μm; manufactured by Waters) is used as a separation column, and acetonitrile and 2% acetic acid are used as mobile phases. The analysis was performed under different gradient conditions. Start with a 5% acetonitrile solution with the column oven set at 30°C, increase the concentration to 20% from 0 to 10 minutes, to 80% from 10 to 25 minutes, and then perform a washing step with 100% acetonitrile. The conditions that were followed were used. The detection was performed at 290 nm at a flow rate of 1 mL/min. The sample injection amount was 20 μl.

(Experiment 1/Result 1)
DNBL1826 strain only as lactic acid bacteria,
Combination of DNBL1826 strain and DNBL1829 strain,
Combination of DNBL1826 strain and DNBL1830 strain,
Combination of DNBL1826 strain and DNBL1831 strain,
DNBL1832 strain only,
Combination of DNBL1832 strain and DNBL1829 strain,
A combination of DNBL1832 strain and DNBL1830 strain, or a combination of DNBL1832 strain and DNBL1831 strain,
1, the results of thin-layer chromatography on the second day of the fermentation treatment in a test using a commercially available hot water extract of ground sesame or a commercially available hot water extract of sesame squeezed meal as a substrate are shown in FIGS. To
The results of HPLC on the 7th day of the fermentation treatment are shown in Tables 1 and 2 below.

As shown in FIGS. 1A and 2A, the DNBL1826 strain had a slight ability to produce sesaminol regardless of which raw material was used. When any of the DNBL1829 to DNBL1831 strains was used in combination with the DNBL1826 strain, the production of sesaminol was significantly promoted.

Similarly, as shown in FIGS. 1B and 2B, the DNBL1832 strain had a slight ability to produce sesaminol regardless of which raw material was used. When any of the DNBL1829 to DNBL1831 strains was used in combination with the DNBL1832 strain, the production of sesaminol was significantly promoted.

In Tables 1 and 2, ND means Not Detected. The same applies to the other tables.
The content of each component per 100 g of raw material powder is calculated on the assumption that the extraction efficiency is 100%. The same applies to other experiments.

The DNBL1826 strain showed a slight ability to produce sesaminol regardless of which raw material was used. When any of the DNBL1829 to DNBL1831 strains was used in combination with the DNBL1826 strain, the production of sesaminol was significantly promoted.

Similarly, the DNBL1832 strain showed a slight ability to produce sesaminol regardless of which raw material was used. When any of the DNBL1829 to DNBL1831 strains was used in combination with the DNBL1832 strain, the production of sesaminol was significantly promoted.

(Experiment 1/Result 2)
Thin layer chromatography on the fifth day of the fermentation treatment in a test using a commercially available hot water extract of ground sesame as a substrate, using a combination of the DNBL1826 strain and the A221 strain as the lactic acid bacterium or a combination of the DNBL1832 strain and the A221 strain. The results are shown in FIG. 3, the HPLC chart on the 5th day of the fermentation treatment is shown in FIGS. 4f and 4g, and the HPLC analysis results on the 5th day of the fermentation treatment are shown in Table 3 below.

In addition, FIG. 4a is a chart showing the retention time of the standard product of STG, FIG. 4b is the standard product of β1,2SDG, FIG. 4c is the standard product of β1,6SDG, and FIG. 4d is the standard product of SMN under HPLC under the same conditions. Is. Fig. 4e shows an HPLC chart of the test group not treated with lactic acid bacteria, Fig. 4f shows an HPCL chart of the test group using the DNBL1826 strain and A221 strain in combination, and Fig. 4g shows a test group using the DNBL1832 strain and A221 strain in combination. An HPCL chart is shown.

As shown in FIGS. 3 and 4f, g, both DNBL1826 strain and DNBL1832 strain produced a large amount of sesaminol when the ground sesame extract was used as a substrate when the A221 strain was used in combination. In addition, with the formation of sesaminol, sesaminol triglucoside decreased.

It is also shown by the analysis results in Table 3 that when both the DNBL1826 strain and the DNBL1832 strain were used in combination with the A221 strain, a large amount of sesaminol was produced when the ground sesame hot water extract was used as a substrate.

(Experiment 1/Result 3)
Using the DNBL1832 strain alone or a combination of the DNBL1832 strain and the A221 strain as the lactic acid bacteria, and using a hot water extract of commercially available sesame press cake as a substrate, the results of the thin layer chromatography on the fourth day of the fermentation treatment are shown. 5 shows the results of HPLC analysis on the 7th day of the fermentation treatment in Table 4 below.
As shown in FIG. 5, the DNBL1832 strain can generate a small amount of sesaminol alone, and when the A221 strain is used in combination, the sesaminol productivity is remarkably high.

The analysis results in Table 4 also show that when the DNBL1832 strain was used in combination with the A221 strain, a large amount of sesaminol was produced when the sesame-pressed meal waste hot water extract was used as the substrate.

<Experiment 2: Sesaminol-containing composition production example 1>
An attempt was made to produce a composition containing sesaminol by using commercially available ground sesame (white) as a raw material and fermenting the suspension with lactic acid bacteria as a substrate.
The lactic acid bacterial strains used are as shown in the results column.

(Experiment 2/Procedure)
For each test section described below, 1 g of the raw material and 20 mL of purified water were put into a 50 ml centrifuge tube to prepare a suspension, and autoclaved at 121° C. for 15 minutes.

To each of the containers containing the suspension after the autoclave treatment, 500 μl of the lactic acid bacterium suspension preliminarily prepared by the method described in <Preparation of lactic acid bacterium pellets and lactic acid bacterium suspension> was added and mixed well. When two kinds of lactic acid bacteria were used in combination, 500 μl of each separately prepared lactic acid bacteria suspension was added to the same 50 ml centrifuge tube and mixed well.

Fermentation with lactic acid bacteria was performed by incubating at 37°C. The first day of starting the fermentation was the 0th day, and the fermentation was continued until the 7th day. After the start of the culture, fermentation was carried out while stirring without stirring, except that the fermentation solution was mixed by shaking once every 3 days.

As a negative control, the suspension of the raw material was treated in the same procedure except that sterilized purified water was added instead of the lactic acid bacterium suspension. This test section was defined as a “untreated lactic acid bacterium” test section.

On the 7th day of the fermentation treatment, an equal amount (20 ml) of ethanol was mixed with the suspension of lactic acid bacteria in the container so that the ethanol concentration was 50% (v/v), and the mixture was centrifuged. The supernatant was passed through a 0.45 μm filter and analyzed by high performance liquid chromatography (HPLC). For the HPLC analysis, the same conditions as in <Experiment 1> were applied.

(Experiment 2/Result)
DNBL1826 strain only as lactic acid bacteria,
Combination of DNBL1826 strain and DNBL1829 strain,
Combination of DNBL1826 strain and DNBL1831 strain,
DNBL1832 strain only,
The combination of DNBL1832 strain and DNBL1829 strain or the combination of DNBL1832 strain and DNBL1831 strain was used, and the results of HPLC analysis on the 7th day of the fermentation treatment in a test using a suspension of commercially available ground sesame as a substrate are shown in the table below. 5 shows.

When both DNBL1826 strain and DNBL1832 strain were used in combination with DNBL1829 strain or DNBL1831 strain, the productivity of sesaminol was high when the ground sesame suspension was used as the substrate.
Moreover, the amount of sesamin (SEM) did not change by fermentation of lactic acid bacteria.

In the present test, the weight ratio of sesaminol:sesamin was 1:2.84 to 1:26.5 in the fermentation product when the DNBL1826 strain or the DNBL1832 strain was used alone. On the other hand, the weight ratio of sesaminol:sesamin was 1:1.45 to 1:1.67 in the fermentation product when used in combination with the DNBL1829 strain or the DNBL1831 strain.

6A shows the HPLC chart of the test group not treated with lactic acid bacteria, FIG. 6B shows the HPLC chart of the test section using only DNBL1826 strain, and FIG. 6C shows the test chart of the test group using the combination of DNBL1826 strain and DNBL1829 strain. An HPLC chart is shown. As shown in FIG. 6B, the DNBL1826 strain alone had the ability to produce sesaminol. On the other hand, as shown in FIG. 6C, when the DNBL1826 strain and the DNBL1829 strain were used in combination, the productivity of sesaminol was significantly improved. Fermentation with lactic acid bacteria had little effect on the amount of sesamin, and sesaminol and sesamin coexisted in the fermentation product.

<Experiment 3: Sesaminol-containing composition production example 2>
An attempt was made to produce a composition containing sesaminol by using commercially available ground sesame (white) as a raw material and fermenting the suspension with lactic acid bacteria as a substrate.
A combination of DNBL1826 strain and DNBL1829 strain was used as the lactic acid bacterium strain.

(Experiment 3/Procedure)
20 g of the raw material and 100 mL of purified water were placed in a 300 ml Erlenmeyer flask to prepare a suspension, which was autoclaved at 121° C. for 15 minutes.

In each of the containers containing the suspension after the autoclave treatment, a lactic acid bacterium suspension of DNBL1826 strain and a lactic acid bacterium suspension of DNBL1829 strain were prepared in advance by the method described in <Preparation of lactic acid bacterium pellets, lactic acid bacterium suspension>. 5 ml of each suspension was added.

Fermentation with lactic acid bacteria was performed by incubating at 37°C. The first day of starting the fermentation was the 0th day, and the fermentation was continued until the 7th day. After the start of the culture, fermentation was carried out while stirring without stirring, except that the fermentation solution was mixed by shaking once every 3 days.

As a negative control, the suspension of the raw material was treated in the same procedure except that sterilized purified water was added instead of the lactic acid bacterium suspension. This test section was defined as a “untreated lactic acid bacterium” test section.
On the 7th day of the fermentation treatment, the suspension of lactic acid bacteria in the container was sterilized by an autoclave treatment and freeze-dried to obtain a freeze-dried composition.

250 mg of the freeze-dried composition was weighed, extracted with 5 mL of 99.5% ethanol for 2 days, centrifuged, and the supernatant was collected. A sample obtained through a 0.45 μm filter was subjected to high performance liquid chromatography (HPLC). Was analyzed by. For the HPLC analysis, the same conditions as in <Experiment 1> were applied.

(Experiment 3/Result)
Table 6 below shows the HPLC analysis results of the freeze-dried composition obtained in the test group fermented with the DNBL1826 strain and the DNBL1829 strain and the freeze-dried composition obtained in the test group untreated with lactic acid bacteria.

FIG. 7A shows the HPLC chart of the test group not treated with lactic acid bacteria, and FIG. 7B shows the HPLC chart of the test group using the combination of DNBL1826 strain and DNBL1829 strain. When a DNBL1826 strain and a DNBL1829 strain were used together, a large amount of sesaminol was produced. Fermentation with lactic acid bacteria had little effect on the amount of sesamin, and sesaminol and sesamin coexisted in the fermentation product.
It should be noted that STG cannot be efficiently extracted with 99.5% ethanol.

<Experiment 4: Sesaminol-containing composition production example 3>
An attempt was made to produce a composition containing sesaminol by using commercially available sesame pressed cake as a raw material and fermenting the suspension with lactic acid bacteria as a substrate.
A combination of DNBL1826 strain and DNBL1829 strain was used as the lactic acid bacterium strain.

(Experiment 4/Procedure)
20 g of the raw material and 100 mL of purified water were placed in a 300 ml Erlenmeyer flask to prepare a suspension, which was autoclaved at 121° C. for 15 minutes.

In each of the containers containing the suspension after the autoclave treatment, a lactic acid bacterium suspension of DNBL1826 strain and a lactic acid bacterium suspension of DNBL1829 strain were prepared in advance by the method described in <Preparation of lactic acid bacterium pellets, lactic acid bacterium suspension> 5 ml of each suspension was added.

Fermentation with lactic acid bacteria was performed by incubating at 37°C. The first day of starting fermentation was the first day, and the fermentation was continued until the seventh day. After the start of the culture, fermentation was carried out while stirring without stirring, except that the fermentation solution was mixed by shaking once every 3 days.

As a negative control, the suspension of the raw material was treated in the same procedure except that sterilized purified water was added instead of the lactic acid bacterium suspension. This test section was defined as a “untreated lactic acid bacterium” test section.

On the 7th day of the fermentation treatment, the suspension of lactic acid bacteria in the container was sterilized by an autoclave treatment and freeze-dried to obtain a freeze-dried composition.

250 mg of the above composition was weighed, extracted with 5 mL of 99.5% ethanol for 2 days, centrifuged, and the supernatant was taken. The sample obtained through a 0.45 μm filter was analyzed by high performance liquid chromatography (HPLC). did. For the HPLC analysis, the same conditions as in <Experiment 1> were applied.

(Experiment 4/Result)
Table 7 below shows the HPLC analysis results of the freeze-dried composition obtained in the test section fermented with DNBL1826 strain and DNBL1829 strain and the freeze-dried composition obtained in the test section untreated with lactic acid bacteria.

8A shows an HPLC chart of the test group not treated with lactic acid bacteria, and FIG. 8B shows an HPLC chart of the test group using a combination of DNBL1826 strain and DNBL1829 strain. Sesaminol was produced when DNBL1826 strain and DNBL1829 strain were used in combination. Fermentation with lactic acid bacteria had little effect on the amount of sesamin, and sesaminol and sesamin coexisted in the fermentation product.
It should be noted that STG cannot be efficiently extracted with 99.5% ethanol.

<Experiment 5: Production test of sesaminol from ground hot sesame extract>
Using commercially available ground sesame (white) as a raw material, fermented with lactic acid bacteria using the hot water extract as a substrate, we tried to produce sesaminol.
The lactic acid bacterial strains used are as shown in the results column.

(Experiment 5/Procedure)
10 g of the raw material and 100 mL of purified water were placed in a 300 ml Erlenmeyer flask and autoclaved at 121° C. for 15 minutes.
The suspension after the autoclave treatment was No. 2 Filter paper (manufactured by Advantech) was filtered, and the filtrate was re-sterilized. The filtrate obtained by the re-sterilization was used as a hot water extract.

Thereafter, 1 mL of the hot water extract was added to each lactic acid bacterium pellet prepared in a 1.5 ml microtube by the method described in <Preparation of lactic acid bacterium pellet and lactic acid bacterium suspension> to suspend the lactic acid bacterium pellet.

Fermentation with lactic acid bacteria was carried out by incubating at 37°C while standing without stirring. The first day of starting the fermentation was the 0th day, and the fermentation was continued until the 7th day.
As a negative control, the hot water extract of the raw material was treated in the same procedure except that lactic acid bacteria were not added. This test section was defined as a “untreated lactic acid bacterium” test section.

On the second day and the seventh day of the fermentation treatment, a minute amount of the lactic acid bacterium suspension was sampled from each of the microtubes and developed by thin layer chromatography. For comparison, an aqueous solution of sesaminol standard and a sample of the test section not treated with lactic acid bacteria were similarly developed by thin layer chromatography. For the development of thin layer chromatography, a mixture of ethyl acetate:methanol:water at a ratio of 14:5:4 was used as a developing solvent. A 10% (v/v) aqueous solution of sulfuric acid was used as the color-developing solution after development, and detection was performed by spraying and heating.

On the 7th day of the fermentation treatment, the lactic acid bacterium suspension from each microtube and ethanol were mixed so that the ethanol concentration was 50% (v/v). After that, centrifugation was performed and the supernatant was passed through a 0.45 μm filter (manufactured by Advantech), and the obtained sample was analyzed by high performance liquid chromatography (HPLC). The analysis conditions by HPLC were the same as in Experiment 1.

(Experiment 5/Result)
DNBL1826 strain only as lactic acid bacteria,
Combination of DNBL1826 strain and DNBL1829 strain,
Combination of DNBL1826 strain and DNBL1830 strain,
Combination of DNBL1826 strain and DNBL1831 strain,
Combination of DNBL1826 strain and A221 strain,
DNBL1832 strain only,
Combination of DNBL1832 strain and DNBL1829 strain,
A combination of DNBL1832 strain and DNBL1830 strain,
Combination of DNBL1832 strain and DNBL1831 strain,
A combination of DNBL1832 strain and A221 strain,
DNBL1829 strain only,
DNBL1830 strain only,
DNBL1831 strain only, or
FIG. 9 shows the results of thin layer chromatography on the second day of the fermentation treatment in the test using only the A221 strain and a commercially available hot water extract of ground sesame as a substrate. The results are shown in FIG. The results of the HPLC analysis on the second and seventh days of the fermentation treatment are shown in Tables 8 and 9 below.

From the results shown in Tables 8 and 9, in fermentation using a ground sesame hot water extract as a substrate, DNBL1826 strain, DNBL1832 strain, DNBL1829 strain, DNBL1830 strain, and DNBL1831 strain were used alone, respectively, compared to DNBL1826 strain Alternatively, it was confirmed that when the DNBL1832 strain and one of the DNBL1829 strain, the DNBL1830 strain and the DNBL1831 strain were used in combination, the amount of sesaminol produced was significantly increased.

It was also confirmed that the A221 strain alone has the ability to produce sesaminol.
From the analysis results by TLC shown in FIGS. 9 and 10, the DNBL1829 strain, the DNBL1830 strain, and the DNBL1831 strain have a low ability to convert sesaminol triglucoside (STG) into sesaminol diglucoside (SDG), but sesaminol diglucoside (SDG). ) To sesaminol (SMN).

On the other hand, the DNBL1826 strain and the DNBL1832 strain have high ability to convert sesaminol triglucoside (STG) into sesaminol diglucoside (SDG), but have the ability to convert sesaminol diglucoside (SDG) into sesaminol (SMN). It was suggested to be low.

It was suggested that strain A221 has a high ability to convert sesaminol triglucoside (STG) into sesaminol diglucoside (SDG), and also has an ability to convert sesaminol diglucoside (SDG) into sesaminol (SMN).

<Experiment 6: Sesaminol production test from hot water extract of pressed sesame meal>
Production of sesaminol was attempted by using commercially available sesame pressed cake as a raw material and fermenting it with lactic acid bacteria using the hot water extract as a substrate.

(Experiment 6/Procedure)
In Experiment 5, lactic acid fermentation with a hot water extract of sesame pressed meal was carried out for 7 days by the same procedure as in Experiment 5 except that commercially available sesame pressed meal was used in place of commercially available ground sesame (white). It was The same lactic acid bacterium as in Experiment 5 was used.

On the second day and the seventh day of the fermentation treatment, a minute amount of the lactic acid bacterium suspension was sampled from each of the microtubes and developed by thin layer chromatography.
On the 7th day of the fermentation treatment, HPLC analysis was performed by the same procedure as in Experiment 5.

(Experiment 6/Result)
The results of thin layer chromatography on the second day of fermentation treatment are shown in FIG. 11, and the results of thin layer chromatography on the seventh day of fermentation treatment are shown in FIG. The results of the HPLC analysis on the second and seventh days of the fermentation treatment are shown in Tables 10 and 11 below.

From the results shown in Tables 10 and 11, DNBL1826 strains, DNBL1832 strains, DNBL1829 strains, DNBL1830 strains and DNBL1831 strains were used alone in fermentation using sesame-pressed meal hot water extract as a substrate, respectively. It was confirmed that when the strain or DNBL1832 strain and one of the DNBL1829 strain, the DNBL1830 strain and the DNBL1831 strain were used in combination, the production amount of sesaminol was significantly increased.
It was also confirmed that the A221 strain alone has the ability to produce sesaminol.

From the analysis results by TLC shown in FIGS. 11 and 12, DNBL1829 strain, DNBL1830 strain and DNBL1831 strain have low ability to convert sesaminol diglucoside (STG) to sesaminol diglucoside (SDG), but sesaminol diglucoside (SDG). ) To sesaminol (SMN).

On the other hand, the DNBL1826 strain and the DNBL1832 strain have high ability to convert sesaminol triglucoside (STG) into sesaminol diglucoside (SDG), but have the ability to convert sesaminol diglucoside (SDG) into sesaminol (SMN). It was suggested to be low.

It was suggested that the A221 strain has both high ability to convert sesaminol triglucoside (STG) to sesaminol diglucoside (SDG) and ability to convert sesaminol diglucoside (SDG) to sesaminol (SMN).

<Experiment 7: Production test of sesaminol from ground hot sesame extract>
Using commercially available ground sesame (white) as a raw material, fermented with lactic acid bacteria using the hot water extract as a substrate, we tried to produce sesaminol.

(Experiment 7/Procedure)
By the same procedure as in Experiment 5, lactic acid fermentation with a hot water extract of ground sesame was performed for 7 days. Each lactic acid bacterium was used alone.

On the seventh day of the fermentation treatment, a minute amount of the lactic acid bacteria suspension was sampled from each of the microtubes and developed by thin layer chromatography.
On the 7th day of the fermentation treatment, HPLC analysis was performed by the same procedure as in Experiment 5.

(Experiment 7/Result)
The result of thin layer chromatography on the 7th day of the fermentation treatment is shown in FIG. The results of the HPLC analysis on the seventh day of the fermentation treatment are shown in Table 12 below.

From the results in Table 12 and FIG. 13, the same tendency as in Experiment 5 was confirmed when each lactic acid bacterium strain was used alone.

<Experiment 8: Production test of sesaminol from hot water extract of sesame pressed cake>
Production of sesaminol was attempted by using commercially available sesame pressed cake as a raw material and fermenting it with lactic acid bacteria using the hot water extract as a substrate.

(Experiment 8/Procedure)
By the same procedure as in Experiment 6, lactic acid fermentation with a hot water extract of pressed sesame meal was carried out for 7 days. Each lactic acid bacterium was used alone.

On the seventh day of the fermentation treatment, a minute amount of the lactic acid bacteria suspension was sampled from each of the microtubes and developed by thin layer chromatography.
On the 7th day of the fermentation treatment, HPLC analysis was performed by the same procedure as in Experiment 5.

(Experiment 8/Result)
The result of thin layer chromatography on the 7th day of the fermentation treatment is shown in FIG. The results of the HPLC analysis on the seventh day of the fermentation treatment are shown in Table 13 below.

From the results of Table 13 and FIG. 14, the same tendency as in Experiment 6 was confirmed when each lactic acid bacterium strain was used alone.

According to the method for producing sesaminol of the present invention, it is possible to produce sesaminol or a sesaminol-containing composition that is useful as a component having an antioxidant effect in foods or pharmaceuticals.
The lactic acid bacterium of the present invention can be used in the method for producing sesaminol of the present invention.

Claims (6)

The feedstock containing sesaminol glycoside, by coexisting with one or more lactic acid bacteria belonging to the genus of Lactobacillus seen including sesaminol generating step of generating a sesaminol from the sesaminol glycosides,
The lactic acid bacterium has the following (1) and (2):
(1) one or more lactic acid bacteria selected from DNBL1826 strain (accession number: NITE P-02225) and DNBL1832 strain (accession number: NITE P-02229);
(2) DNBL1829 strain (contract number: NITE P-02226), DNBL1830 strain (contract number: NITE P-02227), DNBL1831 strain (contract number: NITE P-02228), and Lactobacillus casei Hasegawa strain (contract). No.: one or more lactic acid bacteria selected from FERM BP-10123)
A method for producing sesaminol , which is a combination of Lactic acid bacteria of the previous SL (2), DNBL1829 strain (accession number: NITE P-02226), DNBL1830 strain (accession number: NITE P-02227), and, DNBL1831 strain (accession number: NITE P-02228) or selected 1 or more, the method of claim 1 that. (1) One or more lactic acid bacteria selected from DNBL1826 strain (accession number: NITE P-02225) and DNBL1832 strain (accession number: NITE P-02229), and
(2) DNBL1829 strain (contract number: NITE P-02226), DNBL1830 strain (contract number: NITE P-02227), DNBL1831 strain (contract number: NITE P-02228), and Lactobacillus casei Hasegawa strain (contract). No.: one or more lactic acid bacteria selected from FERM BP-10123)
A combination of lactic acid bacteria including. The lactic acid bacterium of (2) is selected from DNBL1829 strain (accession number: NITE P-02226), DNBL1830 strain (accession number: NITE P-02227), and DNBL1831 strain (accession number: NITE P-02228) 1 The lactic acid bacterium combination according to claim 3, which is the lactic acid bacterium described above. Of the raw material containing sesaminol glycoside,
(1) One or more lactic acid bacteria selected from DNBL1826 strain (accession number: NITE P-02225) and DNBL1832 strain (accession number: NITE P-02229), and
(2) DNBL1829 strain (contract number: NITE P-02226), DNBL1830 strain (contract number: NITE P-02227), DNBL1831 strain (contract number: NITE P-02228), and Lactobacillus casei Hasegawa strain (contract). No.: one or more lactic acid bacteria selected from FERM BP-10123)
Fermented product by combination of. The lactic acid bacterium of (2) is selected from DNBL1829 strain (accession number: NITE P-02226), DNBL1830 strain (accession number: NITE P-02227), and DNBL1831 strain (accession number: NITE P-02228) 1 The fermented product according to claim 5 , which is the above lactic acid bacterium .

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