JP2022139601A - Composition - Google Patents

Abstract

To provide a technique for promoting the proliferation of major species of bacteria belonging to the genus Bifidobacterium in the intestinal flora.SOLUTION: Oligosaccharides including galactooligosaccharides, lactulose, and raffinose are used as prebiotics for bacteria belonging to the genus Bifidobacterium including Bifidobacterium longum, Bifidobacterium breve, Bifidobacterium infantis, and Bifidobacterium bifidum. Provided is a composition containing the oligosaccharides and the bacteria belonging to the genus Bifidobacterium. Moreover, provided is a composition for promoting the proliferation of bacteria belonging to the genus Bifidobacterium, the composition containing the oligosaccharides.SELECTED DRAWING: None

Description

The present invention relates to compositions containing galacto-oligosaccharides, lactulose and raffinose.

It is known that the intestinal flora of infants is dominated by Bifidobacterium (hereinafter also referred to as bifidobacteria), and this is considered to be closely related to the maintenance of infant health. ing. Therefore, developing foods that promote the growth of bifidobacteria is important for maintaining the health of infants.

In recent years, research on prebiotics that proliferate bifidobacteria has been actively conducted, and sugar sources are considered to be particularly important for the proliferation of bifidobacteria, and various oligosaccharides are actively used. It is also proposed to combine various oligosaccharides and use them as prebiotics.
In Patent Document 1, when an oligosaccharide selected from lactulose, fructo-oligosaccharide and galacto-oligosaccharide and raffinose are combined and Bifidobacterium is cultured alone, Bifidobacterium longum, Bifidobacterium breve and Bifidobacterium adolescentis, respectively.
Non-Patent Document 1 describes that a combination of lactulose, raffinose and galacto-oligosaccharides significantly increased the cell numbers of Bifidobacterium breve and Bifidobacterium bacteria.

JP-A-10-175867

T. Ehara et al., British Journal of Nutrition (2016), 116, 270-278

In order to effectively utilize the ingestion of prebiotics, it is necessary that the prebiotics are assimilated by intestinal bacteria and the intestinal bacteria proliferate. However, the Bifidobacterium genus bacteria include many species, and their composition in the intestinal flora varies from individual to individual. In addition, various species of Bifidobacterium bacteria are likely to be enhanced by prebiotics. Therefore, even if prebiotics are ingested, some individuals do not have Bifidobacterium bacteria in their intestines that can assimilate the prebiotics, that is, they are "responders" of the prebiotics. (= non-responder), and the effect of prebiotics may not be obtained sufficiently.
An object of the present invention is to provide a technique for further promoting the growth of a plurality of major species of bacteria belonging to the genus Bifidobacterium in the intestinal flora.

As a result of intensive research conducted by the present inventors to solve the above problems, the combination of galacto-oligosaccharides, lactulose, and raffinose enables even growth of all four major Bifidobacterium species. and conceived that such a composition can be a prebiotic that is less likely to cause non-responders, leading to the completion of the present invention.

That is, a first aspect of the present invention is a composition containing an oligosaccharide and a bacterium of the genus Bifidobacterium, wherein the oligosaccharide contains galactooligosaccharide, lactulose, and raffinose, and the Bifidobacterium Bifidobacterium longum, Bifidobacterium breve, Bifidobacterium infantis, and Bifidobacterium bifidum.
In this embodiment, it is preferable that the mass ratio of the contents of lactulose and raffinose is 1:9 to 9:1.
In this embodiment, it is preferable that the content ratio of galacto-oligosaccharide and lactulose is 1:9 to 9:1.
In this embodiment, the content ratio of galacto-oligosaccharides to raffinose is preferably 1:9 to 9:1.
In this aspect, the total number of viable bacteria of the genus Bifidobacterium contained in the composition is preferably 1×10 ⁸ to 1×10 ¹¹ cfu per 1 g of the oligosaccharide contained in the composition.
The composition of this aspect is preferably a nutritional composition, more preferably a formula.

A second aspect of the present invention is a composition for promoting the growth of Bifidobacterium bacteria containing oligosaccharides, wherein the oligosaccharides include galacto-oligosaccharides, lactulose and raffinose, and the Bifidobacterium genus A composition wherein the bacterium comprises Bifidobacterium longum, Bifidobacterium breve, Bifidobacterium infantis, and Bifidobacterium bifidum.
In this embodiment, it is preferable that the mass ratio of the contents of lactulose and raffinose is 1:9 to 9:1.
In this embodiment, it is preferable that the content ratio of galacto-oligosaccharide and lactulose is 1:9 to 9:1.
In this embodiment, the content ratio of galacto-oligosaccharides to raffinose is preferably 1:9 to 9:1.
The composition of this aspect is preferably a nutritional composition, more preferably a formula.
The composition of this embodiment is preferably an infant formula.

A third aspect of the present invention is a composition containing an oligosaccharide and a Bifidobacterium bacterium, wherein the oligosaccharide is galacto-oligosaccharide, lactulose, raffinose, fructo-oligosaccharide, soybean oligosaccharide, or milk oligosaccharide. three or more selected from the group consisting of sugar, xylooligosaccharide, isomaltooligosaccharide, human milk oligosaccharide, coffee bean mannooligosaccharide, gluconic acid, polydextrose, and inulin, and the Bifidobacterium bacterium is Bifidobacterium A composition comprising Umm longum, Bifidobacterium breve, Bifidobacterium infantis, and Bifidobacterium bifidum.

INDUSTRIAL APPLICABILITY According to the present invention, a composition is provided that can evenly promote the growth of all four major types of bifidobacteria in the intestine. INDUSTRIAL APPLICABILITY The composition of the present invention can be an excellent prebiotic that does not easily cause "non-responders", in which the ingestion of the composition is difficult to obtain the effect of increasing Bifidobacterium bacteria.
Such a composition can be in the form of an oral composition such as food, beverages, or pharmaceuticals, or can be contained in food, beverages, or pharmaceuticals in the form of an additive or the like. By ingesting the composition of the present invention,
It is expected to have an effect of improving the intestinal flora and help maintain the health of consumers, especially infants.

The present invention will now be described in detail. However, the present invention is not limited to the following embodiments, and can be freely modified within the scope of the present invention.

The composition of the present invention contains oligosaccharides, said oligosaccharides essentially comprising galacto-oligosaccharides, lactulose and raffinose.
In the composition of the present invention, the total content of oligosaccharides is preferably 0.1 to 20% by mass, more preferably 0.5 to 10% by mass, and still more preferably It is 1 to 5% by mass. Galactooligosaccharides, lactulose, and raffinose preferably account for 20% by mass or more, more preferably 50% by mass or more, and even more preferably 80% by mass or more of the total oligosaccharides contained in the composition of the present invention.

Lactulose is a disaccharide consisting of fructose and galactose (4-O-β-D-galactopyranosyl-D-fructose, Gal β1→4 Fru). Lactulose can be produced by known methods such as those described in JP-A-3-169888 and JP-A-6-228179. Moreover, lactulose can also use a commercial item (for example, Morinaga Milk Industry Co., Ltd. etc.).
In the composition of the present invention, the content of lactulose is preferably 0.02 to 10% by mass, more preferably 0.05 to 5% by mass, still more preferably 0.05 to 5% by mass, based on the total composition (per solid content). It is 1 to 2% by mass.

Raffinose is a trisaccharide (β-D-fructofuranosyl-α-D-galactopyranosyl-(1-6)-α-D-glucopyranoside, Gal α1 →6 Glc α1→2β Fru). Raffinose can be produced by a known method, for example, the method described in "New Food Material Effective Utilization Technology Series No. 6, 'Raffinose', Page 2, Confectionery Technical Center, 1996". . A commercial product (for example, manufactured by Nippon Beet Sugar Co., Ltd., etc.) can also be used as raffinose.
In the composition of the present invention, the raffinose content is preferably 0.02 to 10% by mass, more preferably 0.1 to 5% by mass, still more preferably 0.1% to 5% by mass, relative to the entire composition (per solid content). It is 2 to 2% by mass.

Galacto-oligosaccharides (GOS) are oligosaccharides having a structure represented by Gal-(Gal)n-Glc (where n is 1 to 3, β1→4 linkage or β1→6 linkage) or mixtures thereof. Galacto-oligosaccharides are industrially produced from lactose by a transfer reaction using β-galactosidase, and the main component is 4'-galactosyl lactose (4'-GL ). Galacto-oligosaccharides can also be used as commercial products (for example, manufactured by Yakult Pharmaceutical Industry Co., Ltd.). Galacto-oligosaccharides may be of one type or a mixture of two or more types.
In the composition of the present invention, the content of galacto-oligosaccharides is preferably 0.02 to 10% by mass, more preferably 0.1 to 5% by mass, and still more preferably 0% by mass relative to the entire composition (per solid content). .2 to 2% by mass.

The composition of the present invention may contain one or more of other oligosaccharides as long as it contains galacto-oligosaccharides, lactulose and raffinose. Other oligosaccharides include fructooligosaccharides, soybean oligosaccharides, milk oligosaccharides, xylooligosaccharides, isomaltooligosaccharides, human milk oligosaccharides, coffee bean mannooligosaccharides, gluconic acid, polydextrose,
Inulin etc. are mentioned.

In the composition of the present invention, the content ratio of lactulose and raffinose is preferably 1:9 to 9:1, more preferably 2:8 to 8:2, still more preferably 3:7 to 7:3. be.
In the composition of the present invention, the content ratio of galacto-oligosaccharide and lactulose is preferably 1:9 to 9:1, more preferably 2:8 to 8:2, still more preferably 3:7 to 7:3. is.
In the composition of the present invention, the content ratio of galacto-oligosaccharides and raffinose is preferably 1:9 to 9:1, more preferably 2:8 to 8:2, still more preferably 3:7 to 7:3. is.

The composition of the present invention can promote the growth of bacteria of the genus Bifidobacterium.
Bifidobacterium bacteria whose growth is promoted by the composition of the present invention include at least Bifidobacterium longum, Bifidobacterium breve, and Bifidobacterium infantis. (Bifidobacterium infantis, reclassified as Bifidobacterium longum subspecies infantis), and Bifidobacterium bifidum. These bifidobacteria are four species (hereinafter also referred to as "infant bifidobacteria") that are particularly important for the health of the intestinal flora in infancy, and the composition of the present invention can evenly promote their growth. .
In addition, as long as the growth of the above four species is promoted, the growth of other Bifidobacterium bacteria may also be promoted. Other Bifidobacterium bacteria include Bifidobacterium adolescentis, Bifidobacterium catenulatum, Bifidobacterium pseudocatenulatum, Bifidobacterium Bifidobacterium animalis, Bifidobacterium lactis, Bifidobacterium pseudolongum and the like.

As used herein, "proliferation" of bacteria includes an increase in the absolute number of bacteria, and "promotion of growth" means that the degree of increase is greater when the composition of the present invention is applied than when it is not applied. . That is, the effect of increasing the number of bacteria of the genus Bifidobacterium when the composition of the present invention is applied in vivo or in vitro compared to when it is not applied is obtained. The degree of increase in the number of bacteria is not particularly limited, but is preferably 1.1 times or more, more preferably 1.5 times or more, relative to the number of bacteria when the composition of the present invention is not applied, More preferably, it means that the number of bacteria becomes 3 times or more.
In addition to directly measuring the number of bacteria, such an increase in the number of bacteria can be achieved, for example, by measuring the turbidity (absorbance) of the contents of the gastrointestinal tract of animals such as humans who ingested the medium in which bacteria were cultured or the composition, or short-term effects such as acetic acid. It can be confirmed by measuring the amount of chain fatty acids to see if the value increases, measuring the pH of the medium and checking if the value decreases, and the like.

In addition, "proliferation" includes an increase in the proportion of bacteria present in the intestinal flora, and "proliferation promotion" indicates a greater degree of increase when the composition of the present invention is applied than when it is not applied. Say things. That is, it includes increasing the proportion of bacteria of the genus Bifidobacterium present in the gastrointestinal tract of animals such as humans that have ingested the composition of the present invention. Here, the "presence ratio" can also be rephrased as the "occupancy rate" for the entire bacteria group detected in the intestinal flora. "Increase in abundance" means that, as long as the abundance in the intestinal flora of bacteria of the genus Bifidobacterium increases, even if the abundance in the intestinal flora of other bacteria increases or decreases at the same time. good.
Although the degree of increase in the abundance ratio is not particularly limited, it is preferably 2% or more, more preferably 5% or more, and still more preferably 20% relative to the abundance ratio of the bacteria when the composition of the present invention is not applied. % or more.

In addition, "enhancement of growth" may be an increase in the speed at which the absolute number of bacteria increases or the speed at which the proportion of the bacteria in the intestinal flora increases. The degree of increase in speed is not particularly limited, but is preferably 10% or more, more preferably 20% or more, and still more preferably 50% or more relative to the speed of the bacteria when the composition of the present invention is not applied. Say something big.
The increase in speed can be confirmed by, for example, shortening the time required to reach an arbitrary bacterial count or abundance ratio when the composition of the present invention is applied compared to when it is not applied.

The composition of the present invention may itself be in the form of foods, beverages, pharmaceuticals, etc., or may be contained in foods, beverages, pharmaceuticals, etc. as an additive.
The intake (administration) route of the composition of the present invention may be oral or parenteral, but is usually oral. In addition, parenteral intake (administration) includes rectal administration and the like.

As described above, the composition of the present invention can promote the growth of bacteria belonging to the genus Bifidobacterium, so it is preferable to include the bacterium belonging to the genus Bifidobacterium in the composition together with the oligosaccharide. That is, it is a composition according to the first aspect of the present invention, containing an oligosaccharide and a Bifidobacterium genus.
Here, Bifidobacterium bacteria contained in the composition of the first aspect of the present invention include Bifidobacterium longum, Bifidobacterium breve, Bifidobacterium infantis, and Bifidobacterium Contains Bacterium bifidum.

As Bifidobacterium longum, Bifidobacterium longum NITE
BP-02621 (alias: BB536 or Bifidobacterium longum subsp. longum ATCC BAA-999) can be used. Bifidobacterium longum BB536 has been deposited internationally under the Budapest Treaty with NPMD under the accession number NITE BP-02621 on January 26, 2018. The same bacterium Bifidobacterium longum subsp. longum ATCC BAA-999 (number: ATCC BAA-999) is A
American Type Culture Collection (ATCC: USA, 2
10801 (108) University Blvd., Manassas, Virginia 0110
01 University Boulevard, Manassas, VA 2011
0, United States of America)) as ATCC BAA-999 (see, for example, JP-A-2012-223134).
As Bifidobacterium longum, Bifidobacterium longum Subspecies longum ATCC 15707 available from ATCC can be used.

As Bifidobacterium breve, Bifidobacterium breve M-1
6V is mentioned. Bifidobacterium breve M-16V was released on January 2, 2018.
On the 6th, Budapest with the accession number NITE BP-02622 to the National Institute of Technology and Evaluation Patent Microorganism Depositary Center (NPMD) (2-5-8 122 Kazusa Kamatari, Kisarazu City, Chiba Prefecture 292-0818) An international deposit has been made under a treaty. Available as a commercial product, for example, "Bifidobacterium breve M-16" manufactured by Morinaga Milk Industry Co., Ltd.
"V" may be used.
In addition, as Bifidobacterium breve, Bifidobacterium breve
Also included is MCC1274. Bifidobacterium breve MCC1274 has 2
On August 25, 2009, the Independent Administrative Institution National Institute of Advanced Industrial Science and Technology Patent Organism Depositary Center (currently the National Institute of Technology and Evaluation Patent Organism Depositary Center (IPOD) (Kazusa Kamatari, Kisarazu City, Chiba Prefecture 292-0818) 2-5-8 Room 120), accession number FERM BP-
11175, an international deposit was made under the Budapest Treaty.

Bifidobacterium infantis also includes Bifidobacterium infantis M-63. Bifidobacterium infantis M-63 has been deposited under the Budapest Treaty under the Budapest Treaty on 26 January 2018 with the NPMD under accession number NITE BP-02623.
Also, Bifidobacterium infantis is available from ATCC, Bifidobacterium longum subspecies infantis ATCC
15697 can be used.

Bifidobacterium bifidum includes Bifidobacterium bifidum MCC1092. Bifidobacterium bifidum MCC1092 has been deposited under the Budapest Treaty on 21 February 2017 with the NPMD under the accession number NITE BP-02429.
Bifidobacterium bifidum includes Bifidobacterium bifidum MCC1319. Bifidobacterium bifidum MCC1319 has been deposited under the Budapest Treaty under the Budapest Treaty on February 21, 2017 with the NPMD under accession number NITE BP-02431.
Bifidobacterium bifidum includes Bifidobacterium bifidum MCC1868. Bifidobacterium bifidum MCC1868 has been deposited under the Budapest Treaty on February 21, 2017 with the NPMD under the accession number NITE BP-02432.
Bifidobacterium bifidum includes Bifidobacterium bifidum MCC1870. Bifidobacterium bifidum MCC1870 has been deposited under the Budapest Treaty under the Budapest Treaty on February 21, 2017 with the NPMD under accession number NITE BP-02433.

In addition, the composition of the first aspect of the present invention may also contain other Bifidobacterium bacteria as long as it contains the above four Bifidobacterium bacteria. Other Bifidobacterium bacteria include Bifidobacterium adolescentis, Bifidobacterium catenulatum, Bifidobacterium pseudocatenulatum, Bifidobacterium animalis, Bifidobacterium lactis, Bifidobacterium pseudolongum and the like.

It should be noted that the bacteria identified by the above-exemplified bacterial names are not limited to the strains themselves that have been deposited or registered with a predetermined institution under the bacterial name (hereinafter also referred to as "deposited strains" for convenience of explanation), Strains substantially equivalent thereto (also referred to as "derivative strains" or "derived strains") are also included. That is, it is not limited to the strain itself deposited with the depositary institution under the above accession number, but also includes substantially equivalent strains. For each bacterium, "a strain substantially equivalent to the above-deposited strain" means that it belongs to the same species as the above-deposited strain, has an effect of improving the intestinal flora, and has a 16S rRNA gene whose base sequence is equivalent to that of the above-deposited strain. preferably 99.86% or more, more preferably 99.93% or more, still more preferably 100% identity to the nucleotide sequence of the 16S rRNA gene of the strain, and preferably identical to the above deposited strain Strains with mycological properties. For each bacterium, a strain substantially equivalent to the deposited strain may be, for example, a derivative of the deposited strain as a parent strain. Derivative strains include strains bred from the deposited strain and strains that arise naturally from the deposited strain. Breeding methods include modification by genetic engineering techniques and modification by mutation treatment. Mutagenesis treatments include X-ray irradiation, ultraviolet irradiation, and treatment with mutating agents such as N-methyl-N'-nitro-N-nitrosoguanidine, ethyl methanesulfonate, and methyl methanesulfonate. be done. Strains naturally occurring from the deposited strain include strains naturally occurring during use of the deposited strain. Such strains include mutants naturally occurring by culturing (eg, subculturing) the deposited strain. Derivative strains may be constructed with one modification, or may be constructed with two or more modifications.

As the cells of the genus Bifidobacterium to be contained in the composition of the first aspect of the present invention, commercially available products may be used, or those obtained by appropriate production may be used.
In addition, the Bifidobacterium bacterium to be contained in the composition of this embodiment can be easily obtained by culturing the aforementioned Bifidobacterium bacterium. The culture method is not particularly limited as long as the bacteria of the genus Bifidobacterium can grow. As a culture method, for example, a method commonly used for culturing Bifidobacterium bacteria can be used as it is or after being modified as appropriate. The culture temperature may be, for example, 25-50°C, preferably 35-42°C. Culturing is preferably carried out under anaerobic conditions, for example, while passing anaerobic gas such as carbon dioxide. Cultivation can also be performed under microaerobic conditions such as liquid stationary culture. Culturing can be carried out, for example, until the Bifidobacterium spp. grows to the desired extent.

The medium used for culture is not particularly limited as long as the Bifidobacterium bacteria can grow. As the medium, for example, a medium commonly used for culturing bacteria of the genus Bifidobacterium can be used as it is or after being modified as appropriate. That is, as carbon sources, for example, sugars such as galactose, glucose, fructose, mannose, cellobiose, maltose, lactose, sucrose, trehalose, starch, starch hydrolysate, blackstrap molasses, etc. can be used depending on the assimilation. . As the nitrogen source, for example, ammonia, ammonium salts such as ammonium sulfate, ammonium chloride and ammonium nitrate, and nitrates can be used. Examples of inorganic salts that can be used include sodium chloride, potassium chloride, potassium phosphate, magnesium sulfate, calcium chloride, calcium nitrate, manganese chloride, and ferrous sulfate. Organic ingredients such as peptone, soybean flour, defatted soybean meal, meat extract and yeast extract may also be used. In addition, as the medium commonly used for culturing Bifidobacterium bacteria, specifically, reinforced Clostridial medium, MRS medium (de Man, Rogosa, and Sharpe medium), mMRS medium (modified MRS medium) ), TOSP medium (TOS propionate medium), TOSP Mup
A medium (TOS propionate mupirocin medium) can be mentioned.

As the Bifidobacterium bacterium that can be contained in the composition of this embodiment, its cells or a fraction containing it can be used without particular limitation. That is, as the Bifidobacterium bacterium, for example, the culture obtained by culturing may be used as it is, the culture may be diluted or concentrated and used, or the cells recovered from the culture may be used. good too. Further, various additional operations such as heating and freeze-drying can be performed after culturing as long as the intestinal microflora-improving effect is not impaired. It is preferable that the additional operation has a high viability of the cells. That is, specific examples of Bifidobacterium bacteria that can be contained in the composition of this embodiment include cultures of Bifidobacterium bacteria, cells collected from the cultures, and processed products thereof. Examples of the processed product include diluted products, concentrated products, dried products, and the like. In addition, it is preferable that the cells are usually used in a form containing viable cells. The cells may be, for example, viable cells, or a mixture of viable cells and dead cells.

In this aspect, the Bifidobacterium genus bacteria contained in the composition include at least viable bacteria, and the total amount in the composition is preferably 1×10 ⁴ to 1×10 ¹³ cfu/
g or 1×10 ⁴ to 1×10 ¹³ cfu/mL, more preferably 1×10 ⁵ to 1×10 ¹² cfu/g or 1×10 ⁵ to 1×10 ¹² cfu/mL, more preferably 1×10 It contains ⁶ to 1×10 ¹¹ cfu/g or 1×10 ⁶ to 1×10 ¹¹ cfu/mL of viable bacteria of the genus Bifidobacterium.
Further, in this aspect, the total number of viable bacteria of the genus Bifidobacterium contained in the composition is preferably 1×10 ⁸ to 1×10 ¹¹ cfu per gram of oligosaccharide contained in the composition, and more It is preferably 1×10 ⁸ to 1×10 ¹⁰ cfu, more preferably 1×10 ⁸ to 3×10 ⁹ cfu.
In addition, as long as live bacteria are contained, dead bacteria may also be included.
Here, cfu refers to colony forming units. In this specification, for example, the value obtained when cultured at 38° C. in a solid medium containing 10% by mass of reconstituted skim milk can be used.
These ranges may be the contents at the time of distribution as a composition or the contents at the time of oral ingestion.

As described above, the composition of the present invention can promote the growth of Bifidobacterium bacteria including infantile bifidobacteria, and thus can be preferably applied to the use of promoting the growth of Bifidobacterium bacteria. That is, it is a composition for growing bacteria of the genus Bifidobacterium containing an oligosaccharide, which is the second aspect of the present invention.

Here, the Bifidobacterium bacterium to be proliferated is not limited to the Bifidobacterium bacterium contained in the composition as in the first aspect of the present invention, and may be found in the gastrointestinal tract of animals such as humans orally ingested. Also included are Bifidobacterium that are present. The composition of the second aspect of the invention is also used to grow all four Bifidobacterium infantum.
Therefore, the composition of the second aspect of the invention can be used to improve the intestinal flora. Here, "improvement of bacterial flora" includes an increase in the number of bacteria present in the intestinal flora of Bifidobacterium, particularly infantile bifidobacteria, and the proportion of the bacteria present. In addition, as long as the abundance of Bifidobacterium spp. It may also include reducing its prevalence in the intestinal flora. Other good bacteria include lactic acid bacteria and the like. Examples of bad bacteria include Clostridium perfringens, Salmonella, Staphylococcus aureus, and pathogenic Escherichia coli.
The above-mentioned "presence ratio" can also be rephrased as "occupancy rate" for the entire bacteria group detected in the intestinal flora.

The composition of the present invention is used for subjects with diseases or pathological conditions that can be prevented or improved by improving the intestinal flora, or for subjects with diseases or pathological conditions caused by deterioration of the intestinal flora. can be useful. For example, for intestinal regulation, immunoregulation, anti-allergy, protection against bacterial and viral infections, reduction of oxidative stress, prevention and improvement of diarrhea, prevention and improvement of constipation, inflammatory bowel disease, and colorectal cancer. It can be used for preventive purposes.

A second aspect of the present invention is the use of an oligosaccharide in the production of a composition for promoting the growth of Bifidobacterium spp. Fidobacterium spp. can translate to uses including Bifidobacterium longum, Bifidobacterium breve, Bifidobacterium infantis, and Bifidobacterium bifidum.
A second aspect of the present invention is the use of oligosaccharides in promoting the growth of Bifidobacterium bacteria, said oligosaccharides comprising galactooligosaccharides, lactulose and raffinose, wherein said Bifidobacterium bacteria can also be translated to uses that include Bifidobacterium longum, Bifidobacterium breve, Bifidobacterium infantis, and Bifidobacterium bifidum.
A second aspect of the present invention is an oligosaccharide used to promote the growth of Bifidobacterium bacteria, wherein the oligosaccharide comprises galactooligosaccharide, lactulose, and raffinose, and the Bifidobacterium Bacteria of the genus include Bifidobacterium longum, Bifidobacterium breve, Bifidobacterium infantis, and Bifidobacterium bifidum.

A second aspect of the present invention is a method for promoting growth of Bifidobacterium bacteria, comprising administering an oligosaccharide to a subject, wherein the oligosaccharide comprises galacto-oligosaccharide, lactulose, and raffinose. , the Bifidobacterium genus includes Bifidobacterium longum, Bifidobacterium breve, Bifidobacterium infantis, and Bifidobacterium bifidum. .
Here, the subject is not particularly limited as long as it is an animal, but is usually a human. Note that "administering an oligosaccharide to a subject" may be synonymous with "making a subject ingest an oligosaccharide." The intake may be voluntary (free intake) or forced (forced intake). Specifically, the administration step may be, for example, a step in which the oligosaccharide is blended in a food, drink or feed and supplied to the subject, thereby allowing the subject to freely ingest the oligosaccharide.

The timing of ingestion (administration) of the composition of the present invention is not particularly limited, and can be appropriately selected according to the condition of the subject to be administered.

The intake (administration) amount of the composition of the present invention is appropriately selected depending on the age, sex, condition, other conditions, etc. of the intake (administration) target.
It should be noted that regardless of the amount and duration of intake (administration), the drug can be administered once a day or in multiple doses.

A third aspect of the present invention is a composition containing an oligosaccharide and a Bifidobacterium bacterium, wherein the oligosaccharide is galacto-oligosaccharide, lactulose, raffinose, fructo-oligosaccharide, soybean oligosaccharide, milk fruit Also provided are those containing three or more selected from the group consisting of oligosaccharides, xylooligosaccharides, isomaltooligosaccharides, human milk oligosaccharides, coffee bean mannooligosaccharides, gluconic acid, polydextrose and inulin. The three or more oligosaccharides may be in any combination, but preferably contain one or more selected from galacto-oligosaccharides, lactulose and raffinose.
Here, Bifidobacterium bacteria include Bifidobacterium longum, Bifidobacterium breve, Bifidobacterium infantis, and Bifidobacterium bifidum, and the first According to the description in the aspect of.

The composition of the present invention is preferably in the form of a food or drink.
The form and properties of the food and drink are not particularly limited as long as they do not impair the effects of the present invention, and the food and drink can be produced by ordinary methods using raw materials commonly used for food and drink.
In addition, the aspect of the additive added to foods, drinks, and pharmaceuticals is also included in the composition of the present invention. Examples of such aspects include additives added to expressed breast milk and formula milk, and it is assumed that the milk after addition is ingested by newborns and infants.
The food and drink are usually orally ingested, but are not limited to this, and may be, for example, nasally ingested, or ingested through a gastrostomy or an enterostomy. For example, it is envisioned that newborns and infants will be fed formula milk, which is the composition of the present invention described later, or mother's milk to which the composition of the present invention has been added, through a nasogastric feeding tube or the like.

Food and drink may be liquid, paste, gel-like solid, powder, etc. Examples include tablets; bread, macaroni, spaghetti, noodles, cake mixes, fried flour, bread crumbs and other flour products; instant noodles. , cup noodles, retort/prepared food, cooked canned food, microwave food, instant soup/stew, instant miso soup/soup, canned soup, freeze-dried food, other instant food; canned agricultural products, canned fruit, jam・Processed agricultural products such as marmalades, pickles, boiled beans, dried agricultural products, and cereals (processed grains); Processed marine products such as canned seafood, fish ham and sausages, fish paste products, seafood delicacies, and tsukudani; Canned livestock・Processed livestock products such as pastes, meat hams and sausages; Processed milk, milk drinks, yogurts, lactic acid bacteria drinks, cheese, ice creams, cream and other dairy products; butter, margarines , Oils and fats such as vegetable oil; Basic seasonings such as soy sauce, miso, sauces, processed tomato seasonings, mirin, and vinegar; Cooking mixes, curry ingredients, sauces, dressings, noodle soups, spices, Complex seasonings and foods such as other complex seasonings; Material Frozen foods such as frozen foods, semi-cooked frozen foods, and cooked frozen foods; caramels, candies, chewing gums, chocolates, cookies, biscuits, cakes, pies, snacks, Confectionery such as crackers, Japanese confectionery, rice confectionery, bean confectionery, dessert confectionery, jelly, and other confectionery; Soymilk, soymilk drinks, coffee drinks, tea drinks, powdered drinks, concentrated drinks, sports drinks, nutritional drinks, alcoholic drinks, other favorite drinks, baby food, furikake, ochazuke seaweed and other commercial foods, etc. nutritional compositions such as formula milk (including powdered milk, liquid milk, etc.), liquid diets, and supplements; functional foods (foods for specified health uses, foods with nutrient function claims);

Among these, nutritional compositions are preferred.
In the present invention, the "nutritional composition" is not particularly limited as one mode of food or drink, but is preferably prepared milk, liquid diet, etc., and more preferably prepared milk. Ingestion subjects may be infants, toddlers, children or adults, but preferably infants and young children.
Infant formula includes powdered infant formula and liquid infant formula.
In the Ministerial Ordinance Concerning Ingredient Standards for Milk and Dairy Products (Ministerial Ordinance for Milk, etc.), powdered milk is defined as "raw milk, cow's milk, special milk, or food products made from these raw materials that are processed or used as main raw materials, and that are necessary for infants. nutritious and powdered”.
Formulated liquid milk is defined in the above ministerial ordinance as "raw milk, cow's milk, special milk, or foods made from these raw materials, processed or made into a liquid by adding nutrients necessary for infants". .
Formulated milk is formulated with nutritional components such as various proteins, fats and oils, carbohydrates, minerals, and vitamins, and includes those processed into powder or liquid.
In addition, infant formula further includes "infant formula", "infant formula liquid formula", "pregnant and lactating women's formula" in food for special dietary use stipulated in the Health Promotion Law, infant formula, Embodiments such as nutritional powders for adults and nutritional powders for the elderly are also included.

When the composition of the present invention is in the form of a supplement, it can be formulated into solid formulations such as powders, granules, tablets, and capsules; liquid formulations such as solutions, syrups, suspensions, and emulsions; can. Such formulations can be made in accordance with the description of the components, carriers, and methods relating to the formulation of pharmaceuticals, which will be described later.

Moreover, it can also be set as feed as one aspect|mode of food-drinks. Examples of the feed include pet food, livestock feed, fish feed, and the like.
The form of the feed is not particularly limited. Bran such as defatted rice bran; Manufacturing lees such as corn gluten meal and corn jam meal; Animal feeds such as fish meal, skimmed milk powder, whey, yellow grease, and tallow; Yeasts such as torula yeast and brewer's yeast; Mineral feeds such as calcium phosphate and calcium carbonate; oils and fats; simple amino acids; sugars and the like may be contained.

When the composition of the present invention is in the form of a food or drink (including feed), it can be provided and sold as a food or drink labeled with the use of proliferating bifidobacteria in the intestine.

Such "display" acts include all acts for informing consumers of the above-mentioned use. Regardless of the object, medium, etc. to be displayed, all of them fall under the act of "display" of the present invention.
In addition, it is preferable that the "display" be performed in an expression that allows the consumer to directly recognize the use. Specifically, the act of transferring, handing over, displaying for the purpose of transfer or delivery, importing products related to food and beverages or product packaging that describes the above-mentioned use, advertisements related to products, price lists or transaction documents Examples include the act of displaying or distributing information with the above-mentioned use described, or providing information containing such information with the above-mentioned use by electromagnetic means (Internet, etc.).

On the other hand, the content of the display is preferably a display approved by the government (for example, a display that is approved based on various systems established by the government and performed in a manner based on such approval). In addition, it is preferable to attach such display contents to packaging, containers, catalogs, pamphlets, POP and other advertising materials at sales sites, other documents, and the like.

In addition, “labeling” includes health food, functional food, enteral nutrition food, food for special dietary use, food with health claims, food for specified health use, food with nutrient function claims, food with function claims, quasi-drugs, etc. Display is also included. Among these, in particular, the labeling approved by the Consumer Affairs Agency, for example, the labeling approved by the system related to food for specified health use, food with nutrient function claims, or food with function claims, or similar system. Specifically, labeling as a food for specified health use, labeling as a food for specified health use with certain conditions, labeling to the effect that it affects the structure and function of the body, labeling to reduce the risk of disease, labeling for functionality based on scientific evidence. Labeling, etc. can be mentioned, more specifically, the Cabinet Office Ordinance Concerning Permission for Special Use Labeling as stipulated in the Health Promotion Act (Cabinet Office Ordinance No. 57 of August 31, 2009) A typical example is labeling as a food for specified health use (especially labeling for health use) and similar labeling.
Such indications include, for example, "Those who want to increase bifidobacteria,""In order to increase bifidobacteria that are useful for the health of babies,""Improve intestinal flora," and "For the health of the stomach of infants." display.

When the composition of the present invention is in the form of a pharmaceutical, its administration route may be oral or parenteral, but oral is preferred. In addition, parenteral intake (administration) includes rectal administration and the like.
As for the pharmaceutical form, it can be appropriately formulated into a desired dosage form depending on the administration method. For example, in the case of oral administration, solid preparations such as powders, granules, tablets and capsules; and liquid preparations such as solutions, syrups, suspensions and emulsions can be formulated. In the case of parenteral administration, it can be formulated into suppositories, ointments, injections, and the like.
For formulation, ingredients such as excipients, pH adjusters, colorants, and corrigents that are commonly used for formulation can be used. It is also possible to use other medicinal ingredients, known or future prebiotics against bacteria of the genus Bifidobacterium, prebiotics against other bacteria, and the like.
In addition, formulation can be appropriately carried out by a known method depending on the dosage form. At the time of formulation, formulation carriers may be added as appropriate.

Excipients include, for example, sugar derivatives such as lactose, sucrose, glucose, mannitol, sorbitol; starch derivatives such as corn starch, potato starch, α-starch, dextrin, carboxymethyl starch; crystalline cellulose, hydroxypropyl cellulose, Hydroxypropylmethylcellulose, carboxymethylcellulose, cellulose derivatives such as carboxymethylcellulose calcium; gum arabic; dextran; pullulan; silicate derivatives such as light silicic anhydride, synthetic aluminum silicate, and magnesium aluminometasilicate; phosphate derivatives such as calcium phosphate; carbonate derivatives such as calcium; sulfate derivatives such as calcium sulfate;

Examples of binding agents include gelatin, polyvinylpyrrolidone, macrogol, etc., in addition to the excipients described above.

Examples of disintegrants include, in addition to the excipients described above, chemically modified starch or cellulose derivatives such as croscarmellose sodium, carboxymethyl starch sodium, and crosslinked polyvinylpyrrolidone.

Lubricants include, for example, talc; stearic acid; metal stearates such as calcium stearate and magnesium stearate; colloidal silica; waxes such as pea gum and gairou; ; carboxylic acid sodium salts such as sodium benzoate; sulfates such as sodium sulfate; leucine; lauryl sulfates such as sodium lauryl sulfate and magnesium lauryl sulfate; silicic acid anhydride and silicic acid hydrate; be done.

Examples of stabilizers include paraoxybenzoic acid esters such as methylparaben and propylparaben; alcohols such as chlorobutanol, benzyl alcohol and phenylethyl alcohol; benzalkonium chloride; acetic anhydride;

Flavoring agents include, for example, sweeteners, acidulants, flavoring agents, and the like.
In addition, solvents such as water and the like can be used as carriers for liquid formulations for oral administration.

The timing of taking the drug of the present invention is not particularly limited, such as before meals, after meals, between meals, and before bedtime.

EXAMPLES The present invention will be described in more detail below using examples, but the present invention is not limited to these examples.

<Test Example 1> Evaluation of the growth-promoting effect of infant bifidobacteria of various oligosaccharides by stool culture (1) Sample pretreatment 7 healthy infants aged 1 to 12 months (a, b, c, d, respectively) Stool samples were obtained from (e, f, g). The mice were collected immediately after defecation and immediately transferred to anaerobic conditions at 10°C or less. Within 8 hours after defecation, the collected stool was diluted with 10 volumes of physiological saline and stored at -80°C until use.

(2) Culture Test In order to simulate the environment in the large intestine, a culture test was performed according to the following procedure. Bio Jr. 8 A 100 mL×8 continuous culture device (BJR-25NA1S-8M manufactured by Viotto Co., Ltd.) was used, and the operation was performed according to the instruction manual of the device. 100 mL of YCFA medium having the composition shown in Table 1 was used as the medium. The total concentration of oligosaccharides in the medium was 1% by mass. The medium components other than the oligosaccharides were dissolved in purified water and sterilized by autoclaving, and the oligosaccharides were dissolved in purified water and then filtered and sterilized, and added to the above autoclaved solution. The culture temperature was 37° C., and filter-sterilized CO ₂ was blown into each culture vessel to maintain an anaerobic state during the culture period. After adjusting the pH to 7.0, 100 μL (10 mg as stool) of a stool sample diluted with physiological saline was added to initiate anaerobic culture. In order to mimic the pH in the colon of infants, pH was controlled by neutralization with 1 M Na ₂ CO ₃ when the pH fell below 5.5 during the culture period. After 24 hours from the initiation of culture, the medium was collected and centrifuged at 8000 rpm, 3 min, 4° C., and the supernatant and precipitate were collected.

(3) DNA extraction
Using a GENE PREP STAR PI-480 automatic extractor (Kurabo Industries Ltd Japan), DNA was extracted from the precipitate collected in (2). Using the extracted DNA as a template, Bifidobacterium bacteria and various types of Bifidobacterium (Bifidobacterium breve, Bifidobacterium Umm longum, Bifidobacterium infantis, and Bifidobacterium bifidum) were measured by quantitative PCR. Primers and PCR conditions are shown in Table 2.

(4) Results Tables 3 to 7 show the degree of proliferation of Bifidobacterium and various species of Bifidobacterium after culturing for 24 hours, as measured by quantitative PCR. The degree of growth is a logarithm of the increase rate when the number of bacteria before culture is set to 1. The number and species of Bifidobacterium bacteria carried by the test infants ranged from 1 to 4, consistent with previous reports, and the combinations varied among individuals. The growth promoting effect on Bifidobacterium genus and each Bifidobacterium species was analyzed by analysis of variance / analysis of covariance ("fit to model" menu of statistical analysis software JMP), and each group (Example / Comparative example) ) were ranked in descending order of proliferative effect, and the top groups were arranged at the top of the table. In addition, each specimen was ranked in descending order of proliferative effect, and the top specimens were arranged in the left column of the table. That is, each group and each specimen were ranked by comprehensive evaluation by statistical processing, and arranged in order from the upper left to the lower right so that the proliferation effect was weaker.

In Example 1, which includes a combination of galacto-oligosaccharides, lactulose, and raffinose, four strains of Bifidobacterium breve, Bifidobacterium longum, Bifidobacterium infantis, and Bifidobacterium bifidum Among various combinations of various oligosaccharides, the growth-promoting effect was ranked first or second for all infantile bifidobacteria. From this, it was clarified that the combination of galacto-oligosaccharide, lactulose and raffinose has the effect of evenly promoting the growth of all infantile bifidobacteria in all infants tested. That is, the combination of galacto-oligosaccharides, lactulose, and raffinose can be an excellent prebiotic that is less likely to produce non-responders.

Claims (14)

A composition containing an oligosaccharide and a Bifidobacterium bacterium,
the oligosaccharides include galacto-oligosaccharides, lactulose, and raffinose;
The composition, wherein the Bifidobacterium bacteria include Bifidobacterium longum, Bifidobacterium breve, Bifidobacterium infantis, and Bifidobacterium bifidum. The composition according to claim 1, wherein the content of lactulose and raffinose has a mass ratio of 1:9 to 9:1. The composition according to claim 1, wherein the content of galacto-oligosaccharide and lactulose has a mass ratio of 1:9 to 9:1. The composition according to claim 1, wherein the content of galacto-oligosaccharides and raffinose has a mass ratio of 1:9 to 9:1. 2. The composition according to claim 1, wherein the total number of viable bacteria of the genus Bifidobacterium contained in the composition is 1×10 ⁸ to 1×10 ¹¹ cfu per 1 g of the oligosaccharide contained in the composition. Composition. A composition according to any preceding claim, which is a nutritional composition. 7. The composition of claim 6, which is formula milk A composition for promoting the growth of Bifidobacterium bacteria containing an oligosaccharide,
the oligosaccharides include galacto-oligosaccharides, lactulose, and raffinose;
The composition, wherein the Bifidobacterium bacteria include Bifidobacterium longum, Bifidobacterium breve, Bifidobacterium infantis, and Bifidobacterium bifidum. The composition according to claim 8, wherein the content of galacto-oligosaccharide and lactulose has a mass ratio of 1:9 to 9:1. The composition according to claim 8, wherein the content of galacto-oligosaccharides and raffinose has a mass ratio of 1:9 to 9:1. The composition according to claim 8, wherein the content of lactulose and raffinose has a mass ratio of 1:9 to 9:1. A composition according to any one of claims 8 to 11, which is a nutritional composition. 13. The composition of claim 12, which is a formula. A composition containing an oligosaccharide and a Bifidobacterium bacterium,
The oligosaccharide consists of galacto-oligosaccharide, lactulose, raffinose, fructo-oligosaccharide, soybean oligosaccharide, milk oligosaccharide, xylooligosaccharide, isomalto-oligosaccharide, human milk oligosaccharide, coffee bean mannooligosaccharide, gluconic acid, polydextrose and inulin. including three or more selected from the group,
The composition, wherein the Bifidobacterium bacteria include Bifidobacterium longum, Bifidobacterium breve, Bifidobacterium infantis, and Bifidobacterium bifidum.