JP5831920B2 - Method for producing ferulic acid-containing fraction - Google Patents

Description

  The present invention relates to a method for producing a ferulic acid-containing fraction, comprising a step of culturing lactic acid bacteria having ferulic acid-producing ability in a plant material. The present invention also relates to lactic acid bacteria having ferulic acid producing ability, lactic acid bacteria fermented foods and drinks derived from plant materials containing ferulic acid, screening methods for lactic acid bacteria having ferulic acid producing ability, and the like.

  Various phenolic compounds are contained in the plant body. Among them, ferulic acid is widely present in the plant body and is known to be contained in a large amount in rice bran, fruits, vegetable skins and the like. It has been reported that phenolic compounds such as ferulic acid exhibit high antioxidant activity by supplementing free radicals or removing active oxygen. In addition, there have been many reports on the functionality of ferulic acid, such as antitumor (antioxidation), diabetes improvement (insulin secretion promotion, blood glucose level reduction), cardiovascular disease improvement action, etc. The effect of is attracting attention. Therefore, effective intake of ferulic acid is thought to lead to prevention and improvement of these diseases. Ferulic acid is also used as an antioxidant and discoloration inhibitor for foods due to its high antioxidant effect.

  Ferulic acid is mainly bound to dietary fiber (hemicellulose), sugar, and the like in the form of esters in food, and its absorbability in vivo is poor in that state. It is said that bound ferulic acid is decomposed by intestinal bacteria, etc., so that part of it becomes free ferulic acid and is absorbed from the intestinal tract, but the produced ferulic acid is further metabolized by intestinal bacteria. It is also known to end up. Therefore, it is important to convert ferulic acid into a free form beforehand in order to effectively ingest highly functional ferulic acid etc. from plant materials regardless of the intestinal flora of the individual. it is conceivable that.

  There are reports on methods for releasing ferulic acid from plants, such as physicochemical treatment, enzyme treatment, and microbial treatment. As a physicochemical treatment method, there is a method of producing ferulic acid by subjecting a residue obtained by extracting rice salad oil and fatty acids from rice bran to produce ferulic acid (Patent Document 1). Therefore, there is a problem that the process is complicated. A method of increasing the amount of compounds such as ferulic acid by treating lignin with a gas containing water vapor at a high temperature and pressure (Patent Document 2) is also known, but requires equipment for high temperature and pressure treatment, and is versatile. Is low.

  As a method using an enzyme, a method of releasing ferulic acid by enzymatic treatment of an ethanol extract of rice bran (Patent Document 3) is known, but there are ethanol extraction and a subsequent drying step, and the work is complicated. And energy costs are high. There is also a method for producing a fermented alcoholic beverage containing a high amount of free ferulic acid using an enzyme having ferulic acid esterase, xylanase, or cellulase activity (Patent Document 4), but considering the actual process, addition of the enzyme -The reaction process becomes complicated and there is a problem of the cost of the enzyme.

  As a method using a microorganism, a method of releasing ferulic acid by reacting a culture solution of penicillium chrysogenum with sugar beet pulp (Patent Document 5), or using a koji mold having ferulic acid esterase activity is high. Although there is a patent (Patent Document 6) relating to flavored cereal distilled liquor, these microorganisms also have strong other enzyme activities such as proteases, which may greatly change the flavor of plant materials.

  Moreover, as a method for synthesizing free ferulic acid, a method of producing ferulic acid from eugenol using Pseudomonas fluorescens (Patent Documents 7 and 8) is known. Ingestion may cause symptoms such as hematuria, convulsions, diarrhea, nausea, loss of consciousness, dizziness, palpitation, and contact with the skin may cause dermatitis due to an allergic reaction. Therefore, there is a problem in safety when left in food. A method of inducing ferulic acid by reacting a benzaldehyde derivative with malonic acid or acetic anhydride in the presence of a catalyst is also known (Patent Document 9). Therefore, versatility is low.

JP 7-78032 Patent No. 4398867 JP 2006-166834 A JP 2010-148485 JP 2006-67803 A JP2004-236634 JP-A-9-155451 JP-A-5-227980 Japanese Patent No. 3271873

  The present invention provides a method for producing a ferulic acid-containing fraction using plant materials and lactic acid bacteria, a lactic acid bacteria fermented food and drink derived from plant materials containing lactic acid bacteria and ferulic acid, and a screening method for lactic acid bacteria having ferulic acid-producing ability. And so on.

  In order to solve the above problems, the present inventors have a dietary experience, targeting lactic acid bacteria that can be eaten safely, and as a result of searching for lactic acid bacteria that release ferulic acid from plant materials, in some lactic acid bacteria, We found ferulic acid esterase activity to release ferulic acid. Surprisingly, however, many of these lactic acid bacteria had the activity of further metabolizing ferulic acid. As a result of intensive studies, the present inventors have succeeded in obtaining a lactic acid bacterium having ferulic acid esterase activity and not having ferulic acid metabolic activity or having ferulic acid metabolic activity below a certain level. By fermenting plant materials with such lactic acid bacteria, ferulic acid, which is a functional component, can be released and accumulated, and food and drink with improved absorbability and functionality of ferulic acid when ingested are provided. be able to. Furthermore, the present invention provides a screening method for lactic acid bacteria having ferulic acid-producing ability as a method for obtaining lactic acid bacteria that release and accumulate ferulic acid.

That is, the present invention provides the following (1) to (15).
(1) A method for producing a ferulic acid-containing fraction, comprising at least a step of culturing a lactic acid bacterium having ferulic acid producing ability in a plant material.
(2) The method for producing a ferulic acid-containing fraction according to (1) above, wherein the lactic acid bacterium having ferulic acid-producing ability has the following properties (A) and (B):
Property (A): a property of liberating 30% or more of the ethyl ferulate as ferulic acid when cultured for 48 hours at an optimum temperature of the lactic acid bacteria in a medium supplemented with 0.05% by weight of ethyl ferulate.
Property (B): The property that 30% or more of the ferulic acid is not liberated as dihydroferulic acid when cultured for 48 hours at the optimum temperature of the lactic acid bacteria in a medium supplemented with 0.05% by weight ferulic acid.
(3) The production method according to (1) or (2) above, wherein the lactic acid bacterium having ferulic acid producing ability is a lactic acid bacterium of the genus Lactobacillus.
(4) The method for producing a ferulic acid-containing fraction according to (3) above, wherein a lactic acid bacterium having ferulic acid producing ability is selected from the following group of lactic acid bacteria;
Lactic acid bacteria group: Lactobacillus helveticus, Lactobacillus fermentum, Lactobacillus acidophilus, Lactobacillus reuteri, Lactobacillus bucchineri, Lactobacillus gasseri, Lactobacillus kefiranofaciens, Lactobacillus gallinarum, Lactobacillus Plantarum, Lactobacillus delbruickey.
(5) The method for producing a ferulic acid-containing fraction according to any one of (1) to (4) above, further comprising a step of purifying ferulic acid from the plant material after culturing. .
(6) The above (1) to (5), wherein the plant material is derived from a material selected from the group consisting of vegetables, cereals, fruits, grasses, beans, potatoes, seeds, rice bran, bran, etc. The manufacturing method of the ferulic acid containing fraction of any one of these.
(7) The method for producing a ferulic acid-containing fraction according to any one of (1) to (6) above, wherein the plant material is enzyme-treated with a dietary fiber degrading enzyme.
(8) Lactic acid bacteria fermented food and drink derived from a plant material containing at least lactic acid bacteria having ferulic acid-producing ability and ferulic acid.
(9) Lactic acid bacteria fermented food and drink according to (8) above, wherein the lactic acid bacteria having ferulic acid producing ability have the following properties (A) and (B);
Property (A): a property of liberating 30% or more of the ethyl ferulate as ferulic acid when cultured for 48 hours at an optimum temperature of the lactic acid bacteria in a medium supplemented with 0.05% by weight of ethyl ferulate.
Property (B): The property that 30% or more of the ferulic acid is not liberated as dihydroferulic acid when cultured for 48 hours at the optimum temperature of the lactic acid bacteria in a medium supplemented with 0.05% by weight ferulic acid.
(10) The lactic acid bacteria fermented food or drink according to (8) or (9) above, wherein the lactic acid bacteria having ferulic acid producing ability are Lactobacillus lactic acid bacteria.
(11) Lactic acid bacteria fermented food and drink according to (10) above, wherein the lactic acid bacteria having ferulic acid producing ability are lactic acid bacteria selected from the following group of lactic acid bacteria;
Lactic acid bacteria group: Lactobacillus helveticus, Lactobacillus fermentum, Lactobacillus acidophilus, Lactobacillus reuteri, Lactobacillus bucchineri, Lactobacillus gasseri, Lactobacillus kefiranofaciens, Lactobacillus gallinarum, Lactobacillus Plantarum, Lactobacillus delbruickey.
(12) Any one of (8) to (11) above, wherein the plant material is a material selected from the group consisting of vegetables, cereals, fruits, grasses, beans, potatoes, seeds, rice bran, bran, etc. The lactic acid bacteria fermentation food / beverage products of description.
(13) A screening method for lactic acid bacteria having ferulic acid producing ability, comprising at least the following steps;
Step (a): culturing a specimen lactic acid bacterium in a medium containing ethyl ferulate, and quantifying the produced ferulic acid,
Step (b): culturing a specimen lactic acid bacterium in a medium containing ferulic acid, and quantifying the produced dihydroferulic acid,
Step (c): A step of evaluating the presence or absence of the following properties (C) and (D) of the sample lactic acid bacteria and determining ferulic acid production ability;
Property (C): property of producing ferulic acid at a certain ratio or more with respect to ethyl ferulate in step (a),
Property (D): Property that does not produce a certain proportion or more of dihydroferulic acid with respect to ferulic acid in step (b).
(14) Lactic acid bacteria determined to have ferulic acid producing ability by the screening method according to (13) above, and having the following properties;
Property (A): a property of liberating 30% or more of the ethyl ferulate as ferulic acid when cultured for 48 hours at an optimum temperature of the lactic acid bacteria in a medium supplemented with 0.05% by weight of ethyl ferulate.
Property (B): The property that 30% or more of the ferulic acid is not liberated as dihydroferulic acid when cultured for 48 hours at the optimum temperature of the lactic acid bacteria in a medium supplemented with 0.05% by weight ferulic acid.

  According to the production method of the present invention, the ferulic acid-containing fraction can be produced by a very simple, efficient and inexpensive method. In particular, since the production method of the present invention uses lactic acid bacteria fermentation, no special equipment is required, and the amount of ferulic acid in the vegetable material can be increased in an easily controllable temperature range of 10 ° C to 50 ° C. In addition, since a plant material is used as a raw material, the obtained ferulic acid-containing fraction has very high safety and nutritional value, is excellent in flavor due to fermentation by lactic acid bacteria, and is particularly suitable for food and drink use. The plant-derived component in the ferulic acid-containing fraction is in a form that ferulic acid and the like are easily absorbed in the living body, so it can be expected to have high functionality when ingested, as well as improved storage by free ferulic acid, Anti-discoloration effect is expected.

  The present invention further provides a screening method for lactic acid bacteria having ferulic acid producing ability. According to the screening method of the present invention, lactic acid bacteria that can be suitably used in the production method of the present invention can be efficiently screened.

The ratio of ferulic acid released to the culture supernatant after culturing the test strain for 48 hours in MRS-Broth medium supplemented with 0.05% by weight of ethyl ferulate is shown. The ratio of the amount of dihydroferulic acid released in the culture supernatant after culturing the test strain for 48 hours in MRS-Broth medium supplemented with 0.05% by weight of ethyl ferulate is shown. The ratio of ferulic acid (FA) and dihydroferulic acid (DHFA) in the culture supernatant after culturing the test strain for 48 hours in the MRS-Broth medium supplemented with 0.05% by weight of ferulic acid is shown. The amount of ferulic acid contained in the supernatant after fermenting the rice bran extract with the test strain for 48 hours is shown. The amount of ferulic acid in the unfermented rice bran extract was relatively expressed as 1. The amount of ferulic acid contained in the supernatant after fermentation of broccoli juice material treated with xylanase for 48 hours with the test strain is shown. The ferulic acid content in the unfermented broccoli juice material is relatively represented as 1. The amount of ferulic acid contained in the supernatant after fermenting the xylanase-treated rice bran extract with the test strain for 48 hours is shown. The ferulic acid content in the unfermented extract is relatively expressed as 1. The amount of ferulic acid contained in the supernatant after fermenting broccoli juice with the test strain for 48 hours is shown. The ferulic acid content in the unfermented broccoli juice material is relatively represented as 1.

(1) Production method of the ferulic acid-containing fraction of the present invention The production method of the ferulic acid-containing fraction of the present invention includes at least a step of culturing lactic acid bacteria having ferulic acid-producing ability in a plant material. It is a manufacturing method of an acid containing fraction.

  The lactic acid bacterium having ferulic acid producing ability is not particularly limited as long as it is a lactic acid bacterium capable of releasing ferulic acid when cultured under conditions suitable for production of free ferulic acid in a plant material. Specifically, it can be selected as an indicator that ferulic acid is liberated from the plant material and ferulic acid is not decomposed or the degree of decomposition is below a certain level. The lactic acid bacteria selected in this way have the activity of cleaving ester bonds of phenolic compounds contained in plants and the like, and can release ferulic acid, which is known for various functions, from plant materials.

Examples of lactic acid bacteria having ferulic acid producing ability include those having the following properties (A) and (B).
Property (A): The property that 30% or more of the ethyl ferulate is liberated as ferulic acid when cultured for 48 hours at the optimum temperature of the lactic acid bacteria in a medium supplemented with 0.05% by weight of ethyl ferulate.
Property (B): The property that 30% or more of the ferulic acid is not liberated as dihydroferulic acid when cultured for 48 hours at the optimum temperature of the lactic acid bacteria in a medium supplemented with 0.05% by weight ferulic acid.

  In the test for the presence or absence of the above properties (A) and (B), as ferulic acid ethyl and ferulic acid, known materials such as Tokyo Chemical Industry Co., Ltd. can be used. As the medium, a medium suitable for the growth of lactic acid bacteria can be used. For example, a medium most suitable for the growth of the target lactic acid bacteria is appropriately selected from, for example, MRS-Broth medium and skim milk medium. it can. By adopting culture conditions suitable for the growth of lactic acid bacteria, the number of lactic acid bacteria can be increased sufficiently in 48 hours of culture, and ferulic acid or dihydroferulic acid production ability can be appropriately evaluated. The ratio of the released ferulic acid or dihydroferulic acid can be calculated by subjecting the culture supernatant to high performance liquid chromatography and detecting the peak areas of ethyl ferulate, ferulic acid or dihydroferulic acid. More specifically, the methods described in the examples described later can be employed, such as the type of medium, the culture conditions such as the optimum temperature, the conditions of the high performance liquid chromatography method, and the like. In the property (A), “30% or more” in “the property of releasing 30% or more of the ethyl ferulate as ferulic acid” may be 30% to 100%, but 40% or more, 50% or more. The higher the ratio, the better. In the property (B), the “property that does not liberate 30% or more of the ferulic acid as dihydroferulic acid” is more preferably “property that does not liberate 20% or more of the ferulic acid as dihydroferulic acid” The property of not releasing 10% or more of ferulic acid as dihydroferulic acid is even more preferable, and it is most preferable that dihydroferulic acid is not substantially released.

Specific examples of lactic acid bacteria having ferulic acid-producing ability include lactic acid bacteria belonging to the genus Lactobacillus, Bifidobacterium, Streptococcus, Lactococcus, Leuconostoc and Oenococcus. The Lactobacillus genus includes Lactobacillus helveticus, Lactobacillus kefiranofaciens, Lactobacillus fermentum, Lactobacillus acidophilus, Lactobacillus gallinarum, Lactobacillus reuteri, Lactobacillus bukneri, Lactobacillus delbul Examples include Icky, Lactobacillus gasseri and the like. Among them, Lactobacillus helveticus, Lactobacillus fermentum, Lactobacillus acidophilus, Lactobacillus reuteri, Lactobacillus buchneri and Lactobacillus gasseri are preferred, Lactobacillus helveticus FERM-P21079 strain , FERM-BP05445 strain , FERM-P21883 strain (all strains have been deposited at the National Institute of Advanced Industrial Science and Technology under the accession numbers FERM-P21079, FERM-BP05445, and FERM-P21883. Hereinafter, FERM-P21079, FERM-BP05445, and FERM-P21883 are also registered. according to.), available as a number JCM1173 at the Lactobacillus fermentum JCM1173 strain (RIKEN microorganism strain preservation facility Noh is. Hereinafter referred to as JCM1173 both.), Lactobacillus acidophilus FERM-P14161 strain (National Institute of Advanced Industrial Science and Technology has been deposited with the National Institute of as accession number FERM-P14161. Also described as following FERM-P14161.) , Lactobacillus reuteri JCM1112 (available as the number JCM1112 at the Institute of Microbial Strains of Incorporated Administrative Agency, also referred to as JCM1112 below) , Lactobacillus bukunerii NCFB1652 ( BCCM Bacteria Collection in Belgium) It has been deposited as number LMG12000. hereinafter referred to as NCFB1652 both.) and Lactobacillus gasseri JCM5813 strain (RIKEN microbial system At presence facility it is available as a number JCM5813. Hereinafter referred to JCM5813 both.) Are more preferable, Lactobacillus helveticus FERM-P21079 strain, Lactobacillus helveticus FERM-BP05445 strain and FERM-P21883 strain being most preferred.

  The plant material is not particularly limited as long as it contains a small amount of bound or ester-type ferulic acid, but it can be used for materials such as vegetables, grains, fruits, grasses, beans, potatoes, seeds, rice bran, bran, etc. What comes from. The plant material may be the plant material or the like exemplified above, but may be obtained by appropriately processing the plant material. Examples of the processing method include pulverization with a mixer and the like, but are not particularly limited. It is preferable that there is sufficient water for lactic acid bacteria to grow, but it can also be adjusted by adding water after adjusting the plant material.

  In order to release ferulic acid from the plant material more efficiently, the plant material may be pre-enzymed with a dietary fiber degrading enzyme. Examples of the enzyme to be used include xylanase and cellulase, but are not particularly limited. For example, ferulic acid can be released more efficiently by using a plant material obtained by adding the enzyme to vegetable juice and allowing the enzyme reaction at the optimum temperature of the enzyme for several hours to 2 days.

  The method for culturing lactic acid bacteria having ferulic acid-producing ability in a plant material is not particularly limited as long as lactic acid bacteria grow in the plant material and ferulic acid can be released from the plant material. For example, it can be cultured by adding about 0.5 to 10% of a lactic acid bacteria culture to a plant material and fermenting it for 1 to 7 days in a temperature range where the lactic acid bacteria can grow, but is not limited thereto.

  The ferulic acid-containing fraction obtained by the method for producing a ferulic acid-containing fraction of the present invention may be a culture itself obtained by culturing lactic acid bacteria in a plant material, but an additive or the like is added to the culture. Alternatively, it may be obtained by removing impurities. In addition, a ferulic acid-containing fraction having a higher ferulic acid concentration can be obtained by fractionating and purifying ferulic acid from the culture.

(2) Invention lactic acid bacteria fermented food / drink The present lactic acid bacteria fermented food / drink is a plant material-derived lactic acid bacteria fermented food / drink containing at least fermentic acid-producing lactic acid bacteria and ferulic acid.

  The lactic acid bacteria having ferulic acid producing ability is not particularly limited as long as it is a lactic acid bacterium capable of releasing ferulic acid when cultured under conditions suitable for production of free ferulic acid in plant materials. (1) What was demonstrated with this ferulic acid containing fraction manufacturing method of this invention can be used.

  As the plant material, those described in (1) the method for producing a ferulic acid-containing fraction of the present invention can be used. For example, when vegetable juice, fruit juice, or the like is used, a lactic acid bacteria fermented beverage excellent in flavor and nutritional value can be provided.

  Although the ferulic acid density | concentration in this invention lactic-acid-bacteria fermented food-drinks depends on how much is contained in a plant raw material, the amount of ferulic acid in a raw material can be increased with the lactic acid bacteria of this invention.

Although the manufacturing method of this invention lactic-acid-bacteria fermented food / beverage products is not specifically limited, It can manufacture with the manufacturing method of this invention ferulic acid containing fraction.
(3) Screening method of the present invention The screening method of the present invention is a screening method for lactic acid bacteria having ferulic acid producing ability, comprising at least the following steps;
Step (a): culturing a specimen lactic acid bacterium in a medium containing ethyl ferulate, and quantifying the produced ferulic acid,
Step (b): culturing a specimen lactic acid bacterium in a medium containing ferulic acid, and quantifying the produced dihydroferulic acid,
Step (c): A step of evaluating the presence or absence of the following properties (C) and (D) of the sample lactic acid bacteria and determining ferulic acid production ability;
Property (C): property of producing ferulic acid at a certain ratio or more with respect to ethyl ferulate in step (a),
Property (D): Property that does not produce a certain proportion or more of dihydroferulic acid with respect to ferulic acid in step (b).

  According to the screening method of the present invention, lactic acid bacteria excellent in ferulic acid producing ability can be screened.

  In steps (a) to (c), the specimen lactic acid bacterium is a lactic acid bacterium to be determined for the presence or absence of ferulic acid producing ability. The culture conditions in steps (a) and (b) are not particularly limited as long as lactic acid bacteria can grow properly. For example, a suitable time for the number of lactic acid bacteria to reach a sufficient level in a medium suitable for growth of the sample lactic acid bacteria. The culture is exemplified (for example, about 12 to 48 hours). The concentrations of ethyl ferulate and ferulic acid contained in the medium are also not particularly limited, but examples include 0.01 to 1% by weight, preferably about 0.05% by weight. As a method for quantifying ferulic acid and dihydroferulic acid, for example, a high performance liquid chromatography method can be employed. For example, the method described in (1) the method for producing a ferulic acid-containing fraction of the present invention can be employed. The certain ratio in the properties (C) and (D) can be appropriately set according to the desired ferulic acid production ability. For example, 10%, 20%, 30%, 40%, 50%, 60%, 70 %, 80%, 90% or the like.

Thus, lactic acid bacteria having the desired ferulic acid producing ability can be screened. For example, lactic acid bacteria having the following properties can be obtained as lactic acid bacteria having ferulic acid producing ability.
Property (A): a property of liberating 30% or more of the ethyl ferulate as ferulic acid when cultured for 48 hours at an optimum temperature of the lactic acid bacteria in a medium supplemented with 0.05% by weight of ethyl ferulate.
Property (B): The property that 30% or more of the ferulic acid is not liberated as dihydroferulic acid when cultured for 48 hours at the optimum temperature of the lactic acid bacteria in a medium supplemented with 0.05% by weight ferulic acid.

EXAMPLES Hereinafter, although an Example demonstrates this invention further more concretely, the scope of the present invention is not limited to the following Example.

(Selection test for strains with ferulic acid releasing activity)
For the test, the strains shown in Table 1 were used and cultured in MRS-Broth medium (DIFCO) at 37 ° C. (Lactococcus lactis and Lactobacillus kefiranofaciens were 30 ° C.) for 16 hours. MRS-Broth medium supplemented with 0.05% ethyl ferulate (Tokyo Kasei Kogyo Co., Ltd.) is inoculated with 3% of the bacterial solution and incubated at 37 ° C (Lactococcus lactis and Lactobacillus kefiranofaciens are 30 ° C). For 48 hours. The culture supernatant was passed through a UF filter having a fractional molecular weight of 3 kDa, and the passing solution was analyzed by high performance liquid chromatography. Agilent technology Series 1200 HPLC system was used, and the column was Hydrosphere C18, S-5 μm, 12 nm (4.6 mm ID × 250 mm: YMC). As the solvent, A liquid: 0.05% TFA in 5% acetonitrile, B liquid: 0.05% TFA in 95% acetonitrile, the amount of B liquid is 25% from 0 to 3 minutes, 30% by 8 minutes The gradient was 100% by 9 minutes, 100% from 9 to 14 minutes, 25% by 15 minutes, and 25% from 15 to 20 minutes. The solvent flow rate was 1 ml / min, the injection amount was 10 μl, ferulic acid was detected at 320 nm, and dihydroferulic acid, which is a metabolite of ferulic acid, was detected at 210 nm. Identification was performed based on the retention time, and the concentration was calculated from the peak area value.

As shown in FIG. 1, 11 strains out of the 40 strains tested converted 30% or more of the added ethyl ferulate into ferulic acid. In particular Lactobacillus helveticus FERM-P21079 strain, FERM-BP05445 strain, FERM-P21883 strain, Lactobacillus Kefi Rano tumefaciens JCM6985 strain, 5 strains ferulic acid over 80% ethyl ferulate Lactobacillus gasseri JCM5813 strain Converted to. Otherwise, Lactobacillus fermentum JCM1173 strain , Lactobacillus reuteri JCM1112 strain , Lactobacillus acidophilus FERM-P14161 strain , Lactobacillus bukuneri NCFB1652 strain , Lactobacillus gasseri JCM1130 strain , JCM1031 strain 30% or more An acid was produced. On the other hand, the 14 strains tested metabolized ethyl ferulate but did not accumulate ferulic acid and mainly produced dihydroferulic acid in which ferulic acid was further metabolized (FIG. 2). The 14 strains tested hardly metabolized ethyl ferulate. In particular, among the 11 strains evaluated to convert 30% or more of ethyl ferulate into ferulic acid, Lactobacillus helveticus FERM-P21079 strain , FERM-BP05445 strain , FERM-P21883 strain , Lactobacillus reuteri JCM1112 strain , 6 strain of Lactobacillus acidophilus FERM-P14161 strain and Lactobacillus Bukuneri NCFB1652 strain does not substantially generate dihydro ferulic acid, Lactobacillus Farr lactofermentum JCM1173 strain and Lactobacillus gasseri JCM5813 strain small amounts of dihydro ferulic acid Only produced. Therefore, it was revealed that these strains have particularly high ferulic acid producing ability. As described above, it was found that some lactic acid bacteria have an activity to release and accumulate ferulic acid from ferulic acid esters.

(Evaluation of strains that do not have ferulic acid metabolic activity)
Among the strains evaluated in Example 1, five strains that showed ferulic acid releasing activity and five strains that produced dihydroferulic acid as a control were used in the test. The bacterial solution cultured for 16 hours at 37 ° C. in MRS-Broth medium was inoculated into 3% of MRS-Broth medium supplemented with 0.05% ferulic acid and cultured at 37 ° C. for 48 hours. The culture supernatant was treated in the same manner as in Example 1, and ferulic acid and dihydroferulic acid were quantified by HPLC.

As shown in FIG. 3, JCM1131 strain was producing only Jihirodoferura acid in Example 1, JCM1132 strain, JCM1185 strain, JCM1126 strain, the JCM2010 strain, was converted to the added ferulic acid substantially all dihydro ferulic acid. On the other hand, Lactobacillus helveticus FERM-P21079 strain , FERM-BP05445 strain , Lactobacillus acidophilus FERM-P14161 strain , Lactobacillus gasseri JCM5813 strain , Lactobacillus fermentum JCM1173 strain hardly metabolizes and decomposes ferulic acid, Dihydroferulic acid was not produced. By the above method, a strain having no ferulic acid metabolic activity or low activity could be evaluated.

(Fermentation test of plant material by selected strains)
For lactic acid bacteria having ferulic acid releasing activity selected in Examples 1 and 2 and having no ferulic acid metabolic activity, ferulic acid releasing ability when fermenting plant materials was examined. As a plant material, rice bran, which has a high ferulic acid content, was used. Fresh rice bran was suspended in 10 times the amount of water, boiled at 90 ° C. for 1 hour, and then filtered through filter paper. 55 g of MRS-Broth was dissolved in 1 L of the filtrate, and after sterilization, it was used as a rice bran extract for the fermentation test. The test strain was cultured in MRS-Broth medium at 37 ° C. for 16 hours, inoculated with 3% of rice bran extract, and fermented at 37 ° C. for 48 hours. The culture supernatant was treated in the same manner as in Example 1, and ferulic acid was quantified by HPLC.

As shown in FIG. 4, Lactobacillus helveticus FERM-P21079 strain, FERM-P21883 strain except that the amount of ferulic acid was increased 1.5-fold or more, increase 1.2 times ferulic acid Lactobacillus helveticus FERM-BP05445 strain did. On the other hand, ferulic acid decreased 0.3-fold in Lactobacillus gasseri JCM1131 strain having ferulic acid metabolic activity. From the above results, it was possible to increase the amount of free ferulic acid in plant materials by using lactic acid bacteria having ferulic acid releasing activity and no ferulic acid metabolic activity.

(Combination effect 1 with dietary fiber degrading enzyme)
In order to investigate the combined use effect with dietary fiber degrading enzymes, ferulic acid releasing ability when fermented using xylanase-treated broccoli concentrate as a raw material was examined. 100 g of commercially available broccoli turbid concentrated juice was suspended in 1 L of water and heated at 95 ° C. for 5 minutes. After cooling to 50 ° C., 5 g of xylanase (SIGMA) was added and treated at 50 ° C. for 20 hours. 55 g of MRS-Broth was added to 1 L of this enzyme-treated solution and dissolved, and then sterilized and used as a broccoli juice material in a fermentation test. The test strain was cultured in MRS-Broth medium at 37 ° C. for 16 hours, inoculated with 3% broccoli juice material, and fermented at 37 ° C. for 48 hours. The culture supernatant was treated in the same manner as in Example 1, and ferulic acid was quantified by HPLC.

As shown in FIG. 5, Lactobacillus helveticus FERM-BP05445 increased 1.5 times, Lactobacillus reuteri JCM1112 strain was 1.4 times, Lactobacillus helveticus FERM-P21883 strain was 1.3 times, Lactobacillus -Increased 1.2 times in Acidophilus FERM-P14161 strain . On the other hand, ferulic acid decreased 0.2-fold in Lactobacillus gasseri JCM1131 strain having ferulic acid metabolic activity. From the above results, it was shown that the amount of free ferulic acid can be increased even in combination with dietary fiber degrading enzyme.

(Combination effect 2 with dietary fiber degrading enzyme)
In order to investigate the combined use effect with dietary fiber degrading enzyme, ferulic acid releasing ability when fermented using xylanase-treated rice bran as a raw material was examined. 100 g of fresh rice bran was suspended in 1 L of 50 mM MES buffer and heated at 95 ° C. for 5 minutes. After cooling to 50 ° C., 5 g of xylanase (SIGMA) was added and treated at 50 ° C. for 20 hours. 55 g of MRS-Broth was dissolved in 1 L of this enzyme-treated solution, and after sterilization, it was used as an enzyme-decomposed rice bran extract in a fermentation test. The test strain was cultured in MRS-Broth medium at 37 ° C. for 16 hours, inoculated with 3% of enzyme-treated rice bran extract and fermented at 37 ° C. for 48 hours. However, Lactobacillus kefiranofaciens was cultured and fermented at 30 ° C. The culture supernatant was treated in the same manner as in Example 1, and ferulic acid was quantified by HPLC.

As shown in FIG. 6, the amount of ferulic acid was remarkably increased by about 10 times in the Lactobacillus helveticus FERM-BP05445 strain , and was also increased by 5 times or more in the Lactobacillus reuteri JCM1112 strain . Also, Lactobacillus acidophilus FERM-P14161 strain , Lactobacillus helveticus FERM-P21883 strain , FERM-P21079 strain , Lactobacillus gasseri JCM5813 strain and Lactobacillus kefiranofaciens JCM6985 strain are 4.2 to 2.4 times. Ferulic acid increased. Incidentally, ferulic acid content in the culture, Lactobacillus helveticus FERM-BP05445 strain 1.9 mg / L, Lactobacillus reuteri JCM1112 strain 1.0 mg / L, the Lactobacillus acidophilus FERM-P14161 strain 0.8 mg / L, Lactobacillus helveticus FERM-P21883 strain was 0.7 mg / L, and Lactobacillus helveticus FERM-P21079 strain was 0.5 mg / L. From the above results, it was shown that the amount of free ferulic acid can be remarkably increased by using it together with dietary fiber degrading enzyme. The fermented product thus obtained has a higher content of free ferulic acid than a normal vegetable material and can be expected to have high functionality when ingested.

(Fermentation test using vegetable material)
Next, changes in fermentability and ferulic acid content when using vegetable materials were examined. Using broccoli concentrate as a plant material, ferulic acid releasing ability when fermented with selected lactic acid bacteria was examined. 100 g of commercially available broccoli turbid concentrated juice was suspended in 1 L of water and sterilized at 95 ° C. for 30 minutes and used as a broccoli juice material in a fermentation test. The test strain was cultured in MRS-Broth medium at 37 ° C. for 16 hours, inoculated with 3% broccoli juice material, and fermented at 37 ° C. for 48 hours. However, Lactobacillus kefiranofaciens was cultured and fermented at 30 ° C. The culture supernatant was treated in the same manner as in Example 1, and ferulic acid was quantified by HPLC.

As shown in FIG. 7, in addition to the Lactobacillus gasseri JCM5813 strain having a 2-fold increase in ferulic acid, Lactobacillus fermentum JCM1173 strain was 1.6 times, Lactobacillus helveticus FERM-P21883 strain , FERM-BP05445 strain and It was increased 1.5-fold in Lactobacillus acidophilus FERM-P14161 share. Further, 1.4 times Lactobacillus reuteri JCM1112 strain, in JCM6985 strain Lactobacillus Kefi Rano tumefaciens and Lactobacillus helveticus FERM-P21079 strain 1.3 times ferulic acid was increased. From the above results, it was shown that the amount of free ferulic acid can be increased even when the broccoli juice material is fermented as a plant material.

(Prototype of fermented vegetable beverage)
Using the selected lactic acid bacteria, the effect on fermentability and flavor when actually fermenting vegetables was investigated. Commercially available carrot turbid concentrate, broccoli turbid concentrate, and kale turbid concentrate were reduced with water and sterilized by heating at 95 ° C. for 15 minutes. Lactobacillus helveticus FERM-BP05445, which had the highest ferulic acid releasing ability in Example 5 as lactic acid bacteria, was cultured in 12% reduced skim milk containing 0.5% yeast extract at 37 ° C. for 16 hours, and then vegetable juice 3% inoculated and fermented at 37 ° C. for 16 hours.

  As shown in Table 2, the selected lactic acid bacteria grew in various vegetable juices, and an appropriate increase in acidity and a decrease in pH were observed. The flavor was not so strong and very good. From the above results, it was confirmed that the lactic acid bacteria selected in this way can satisfactorily ferment vegetables.

  The present invention includes a method for producing a ferulic acid-containing fraction, a lactic acid bacterium having ferulic acid-producing ability, and a plant material containing ferulic acid, the method comprising culturing lactic acid bacteria having ferulic acid-producing ability in a plant material The present invention relates to a fermented food and drink product derived from lactic acid bacteria, a method for screening lactic acid bacteria having ferulic acid-producing ability, and the like. The present invention can be used in the food field, the pharmaceutical field, and the like.

Claims (10)

A method for producing a ferulic acid-containing fraction comprising at least a step of culturing lactic acid bacteria having ferulic acid producing ability in a plant material,
The lactic acid bacteria having ferulic acid producing ability are selected from the following lactic acid bacteria group,
Lactobacillus group: Lactobacillus helveticus, Lactobacillus acidophilus, Lactobacillus bukuneri The method for producing a ferulic acid-containing fraction, wherein the lactic acid bacterium having the ferulic acid-producing ability has the following properties (A) and (B);
Property (A): a property of liberating 30% or more of the ethyl ferulate as ferulic acid when cultured for 48 hours at an optimum temperature of the lactic acid bacteria in a medium supplemented with 0.05% by weight of ethyl ferulate.
Property (B): The property that 30% or more of the ferulic acid is not liberated as dihydroferulic acid when cultured for 48 hours at the optimum temperature of the lactic acid bacteria in a medium supplemented with 0.05% by weight ferulic acid. Lactic acid bacteria with ferulic acid producing ability is selected from the following group of lactic acid bacteria, the production method of the ferulic acid-containing fraction according to claim 1;
Lactic acid bacteria group: Lactobacillus helveticus FERM-P21079 strain, Lactobacillus helveticus FERM-BP05445 strain, Lactobacillus helveticus FERM-P21883 strain, Lactobacillus acidophilus FERM-P14161 strain, Lactobacillus bucumeri NCFB1652 strain. The manufacturing method of the ferulic acid containing fraction of Claim 1 or 2 which further includes the process of refine | purifying ferulic acid from the plant raw material after the said culture | cultivation. 4. The plant material according to any one of claims 1 to 3 , wherein the plant material is derived from a material selected from the group consisting of vegetables, cereals, fruits, grasses, beans, potatoes, seeds, rice bran, and bran. Of producing a ferulic acid-containing fraction. The method for producing a ferulic acid-containing fraction according to any one of claims 1 to 4 , wherein the plant material is enzyme-treated with a dietary fiber degrading enzyme. A lactic acid bacterium-fermented food or beverage derived from plant materials, containing at least lactic acid bacteria having ferulic acid producing ability and ferulic acid,
The lactic acid bacteria having ferulic acid producing ability are selected from the following lactic acid bacteria group,
Lactic acid bacteria group: Lactobacillus helveticus, Lactobacillus acidophilus, Lactobacillus bukuneri The lactic acid bacteria having ferulic acid-producing ability have the following properties (A) and (B):
Property (A): a property of liberating 30% or more of the ethyl ferulate as ferulic acid when cultured for 48 hours at an optimum temperature of the lactic acid bacteria in a medium supplemented with 0.05% by weight of ethyl ferulate.
Property (B): The property that 30% or more of the ferulic acid is not liberated as dihydroferulic acid when cultured for 48 hours at the optimum temperature of the lactic acid bacteria in a medium supplemented with 0.05% by weight ferulic acid. The lactic acid bacteria fermented food / beverage products of the ferulic acid containing fraction of Claim 6 from which the lactic acid bacteria which have ferulic acid production ability are selected from the following lactic acid bacteria group ;
Lactic acid bacteria group: Lactobacillus helveticus FERM-P21079 strain, Lactobacillus helveticus FERM-BP05445 strain, Lactobacillus helveticus FERM-P21883 strain, Lactobacillus acidophilus FERM-P14161 strain, Lactobacillus bucumeri NCFB1652 strain. The lactic acid bacteria fermented food and drink according to any one of claims 6 to 7 , wherein the plant material is a material selected from the group consisting of vegetables, grains, fruits, grasses, beans, potatoes, seeds, rice bran, and bran. Goods. The following steps :
Step (a): culturing a specimen lactic acid bacterium in a medium containing ethyl ferulate, and quantifying the produced ferulic acid,
Step (b): culturing a specimen lactic acid bacterium in a medium containing ferulic acid, and quantifying the produced dihydroferulic acid,
Step (c): Evaluating the presence or absence of the following properties (C) and (D) of the sample lactic acid bacteria, and determining at least ferulic acid production ability ,
A screening method for lactic acid bacteria having ferulic acid-producing ability having the following properties in the steps (a) and (b);
Property (C): property of producing ferulic acid at a certain ratio or more with respect to ethyl ferulate in step (a),
Property (D): Property that does not produce a certain proportion or more of dihydroferulic acid with respect to ferulic acid in step (b). A lactic acid bacterium determined to have ferulic acid producing ability by the screening method according to claim 9 ,
The lactic acid bacterium having ferulic acid producing ability is selected from the following lactic acid bacteria group and has the following properties:
Lactic acid bacteria group: Lactobacillus helveticus, Lactobacillus acidophilus, Lactobacillus bucumeri Properties (A): ethyl ferulate In a medium supplemented with 0.5% by weight, when cultured for 48 hours at the optimum temperature of the lactic acid bacteria, the property of releasing 30% or more of the ethyl ferulate as ferulic acid,
Property (B): The property that 30% or more of the ferulic acid is not liberated as dihydroferulic acid when cultured for 48 hours at the optimum temperature of the lactic acid bacteria in a medium supplemented with 0.05% by weight ferulic acid.