JP7368484B2 - Composition, food and beverage compositions containing the composition, and formula milk - Google Patents

Description

ATCC 29521 NPMD NITE BP-02429 NPMD NITE BP-01252 NPMD NITE BP-02431 NPMD NITE BP-02648 NPMD NITE BP-02432 NPMD NITE BP-02433 ATCC 15700 NPMD NITE BP-02460 IPOD FERM BP-11175 NPMD NITE BP-02622 ATCC 15697 NPMD NITE BP-02623

The present technology relates to a composition capable of promoting the growth of probiotics that have a low ability to assimilate κ-casein glycomacropeptide (hereinafter also referred to as "GMP").

Bifidobacterium is a resident bacteria in the intestinal tract of mammals including humans, and is known to have the effect of inhibiting the growth of pathogenic enterobacteria in the intestinal tract.
Generally, the intestinal flora of a healthy infant is dominated by Bifidobacteria. On the other hand, with age, the number of Bifidobacteria gradually decreases, and putrefactive bacteria such as Escherichia coli and Clostridium bacteria increase, which has a negative impact on the health of the human host. Therefore, in order to maintain a healthy state, it is important to maintain an intestinal flora dominated by bifidobacteria.

GMP is a peptide corresponding to the C-terminal side (residue numbers 106-169) of κ-casein, which is excised by treating κ-casein with the milk-clotting enzyme chymosin. GMP is highly modified with O-type sugar chains, and the sugar chain structure accounts for about 10% of the weight of GMP. Approximately 50% of this sugar chain structure is sialic acid.

GMP has been reported to have various physiological activities, most of which are thought to be mediated by sugar chains (particularly sugar chains having sialic acid). Physiological activities of GMP include, for example, antibacterial activity, bifidobacteria growth effect, immunomodulatory function, and appetite regulating function.

Patent Document 1 discloses that GMP has a protective effect against infection. Patent Document 2 discloses that GMP has a calcium absorption promoting effect. Patent Document 3 discloses that GMP has an effect of promoting the increase of Akkermansia muciniphila.

Japanese Patent Application Laid-open No. 63-284133 Japanese Patent Application Publication No. 10-117728 JP2019-43866A

Although there are some bacteria that do not have or have a low ability to assimilate GMP, there are many bacteria that can be used as beneficial bacteria. Therefore, even probiotics that do not have or have a low ability to assimilate GMP are useful as compositions if they can be grown using GMP.

Therefore, the main purpose of the present technology is to provide a technology that makes it possible to promote the proliferation of probiotics that have no or low ability to assimilate GMP using GMP assimilation products.

In order to solve the above problem, the present inventors first investigated the ability of various Bifidobacteria to assimilate GMP. As shown in the Examples below, bacteria belonging to Bifidobacterium bifidum were found to have assimilation ability, but other bacteria were found to have no or low assimilation ability for GMP. However, if bacteria belonging to Bifidobacterium bifidum are cultured in a medium using GMP as a carbon source, and then other bacteria are cultured in that medium, even if the bacteria do not have or have a low ability to assimilate GMP. They discovered that there are bacteria that can multiply and completed this technology.

That is, the present technology first provides a composition containing a product of κ-casein glycomacropeptide assimilated by bacteria belonging to Bifidobacterium bifidum.
In the composition according to the present technology, the bacteria belonging to Bifidobacterium bifidum include Bifidobacterium bifidum (ATCC29521T), Bifidobacterium bifidum (NITE BP-02429), and Bifidobacterium bifidum. (NITE BP-01252), Bifidobacterium bifidum (NITE BP-02431), Bifidobacterium bifidum (NITE BP-02648), Bifidobacterium bifidum (NITE BP-02432), Bifidobacterium - One or more bacteria selected from Bifidum (NITE BP-02433) can be used.
The composition according to the present technology may further contain bacteria belonging to Bifidobacterium breve and/or bacteria belonging to Bifidobacterium infantis.
In the composition according to the present technology, the bacteria belonging to Bifidobacterium breve include Bifidobacterium breve (ATCC15700T), Bifidobacterium breve (NITE BP-02460), and Bifidobacterium breve ( One or more bacteria selected from Bifidobacterium breve (FERM BP-11175), Bifidobacterium breve (NITE BP-02622) can be used.
In addition, as the bacteria belonging to Bifidobacterium infantis, one or more selected from Bifidobacterium infantis (ATCC15697T) and Bifidobacterium infantis (NITE BP-02623). Bacteria can be used.
In the composition according to the present technology, the nutritive product has a mass-to-charge ratio of 324 when subjected to a mass spectrometer after being purified with a sugar refining solid phase extraction column and derivatized with 2-aminobenzamide. Alternatively, it may contain a substance detected as a 363 anion.

Next, the present technology provides a Bifidobacterium bacterial growth promoter comprising κ-casein glycomacropeptide.
The Bifidobacterium bacterial growth promoter according to the present technology can further contain carbohydrates.
In the Bifidobacterium bacterial growth promoter according to the present technology, the bacteria belonging to Bifidobacterium bifidum include Bifidobacterium bifidum (ATCC29521T) and Bifidobacterium bifidum (NITE BP-02429). , Bifidobacterium bifidum (NITE BP-01252), Bifidobacterium bifidum (NITE BP-02431), Bifidobacterium bifidum (NITE BP-02648), Bifidobacterium bifidum (NITE BP-) Bifidobacterium bifidum (NITE BP-02433).

This technology also uses bifidobacteria to produce a substance that is detected as an anion with a mass-to-charge ratio of 324 or 363 when subjected to a mass spectrometer after being derivatized with 2-aminobenzamide. Bacteria belonging to the genus Um bifidum can be used.

The present technology further includes a fermentation step of fermenting the κ-casein glycomacropeptide using bacteria belonging to Bifidobacterium bifidum.
Provided is a method for producing a substance that is detected as an anion with a mass-to-charge ratio of 324 or 363 when subjected to a mass spectrometer after being derivatized with 2-aminobenzamide.
In the production method according to the present technology, in the fermentation step, 0.5 to 5% of a culture solution of bacteria belonging to Bifidobacterium bifidum is added to a κ-casein glycomacropeptide solution with a concentration of 0.8% to 12%. It can be added and fermented.

The composition according to the present technology and the Bifidobacterium bacteria growth promoter according to the present technology can be used in food and drink compositions and formula milk.

According to the present technology, it is possible to promote the growth of probiotics that have no or low ability to assimilate GMP using GMP assimilation products. Note that the effects described here are not necessarily limited, and may be any of the effects described in the present technology.

3 is a graph substituted for a drawing showing a total ion chromatogram (TIC) in Experimental Example 3.

Hereinafter, preferred embodiments for implementing the present technology will be described. Note that the embodiment described below shows an example of a typical embodiment of the present disclosure, and the scope of the present technology should not be interpreted narrowly thereby. In addition, in this specification, when a numerical range is expressed as "lower limit to upper limit", the upper limit may be "less than or equal to" or "less than", and the lower limit may be "more than" or "more than". It may be "super". Further, in this specification, percentages are expressed by mass unless otherwise specified.

1. Composition, Bifidobacterium Bacterial Growth Promoter The composition according to the present technology contains at least a GMP assimilation product by bacteria belonging to Bifidobacterium bifidum. Moreover, the composition according to the present technology can further contain bacteria belonging to Bifidobacterium breve and bacteria belonging to Bifidobacterium infantis.

In the composition according to the present technology, the nutrient product is purified using a sugar refining solid-phase extraction column, and the resulting purified product is derivatized with 2-aminobenzamide and then transferred to a mass spectrometer. It can contain a substance that is detected as an anion with a mass-to-charge ratio of 324 or 363 when provided.

The composition for promoting the growth of Bifidobacterium bacteria according to the present technology contains κ-casein glycomacropeptide and has a specificity for bacteria belonging to Bifidobacterium bifidum compared to other Bifidobacterium bacteria. It is an agent that can be used to grow plants. Moreover, the Bifidobacterium bacterial growth promoter according to the present technology can further contain carbohydrates.

(1) Bacteria belonging to Bifidobacterium bifidum Bacteria belonging to Bifidobacterium bifidum that can be used in this technology include known or unknown Bifidobacterium bifidum as long as it does not impair the effectiveness of this technology. One or more types of bacteria belonging to the following can be freely selected and used. For example, Bifidobacterium bifidum (ATCC29521T), Bifidobacterium bifidum (NITE BP-02429), Bifidobacterium bifidum (NITE BP-01252), Bifidobacterium bifidum (NITE BP-02431) , Bifidobacterium bifidum (NITE BP-02648), Bifidobacterium bifidum (NITE BP-02432) and Bifidobacterium bifidum (NITE BP-02433).

Bifidobacterium bifidum ATCC 29521T is available from the American Type Culture Collection, 12301 Parklawn Drive, Rockville, Maryland 20852, United States of America.

Bifidobacterium bifidum (NITE BP-02429), Bifidobacterium bifidum (NITE BP-02431), Bifidobacterium bifidum (NITE BP-02432), Bifidobacterium bifidum (NITE BP-02433) ) was submitted to the National Institute of Technology and Evaluation (NITE) Patent Microorganisms Depositary (Room 122, 2-5-8 Kazusa Kamatari, Kisarazu City, Chiba Prefecture 292-0818) on February 21, 2017 based on the Budapest Treaty. International deposits have been made.

Bifidobacterium bifidum (NITE BP-01252) was deposited on February 23, 2012 at the National Institute of Technology and Product Evaluation (NITE) Patent Microorganism Depositary (2-5 Kazusa Kamatari, Kisarazu City, Chiba Prefecture, 292-0818). -8 Room 122) has been internationally deposited under the Budapest Treaty.

Bifidobacterium bifidum (NITE BP-02648) was deposited on February 26, 2018 at the National Institute of Technology and Product Evaluation (NITE) Patent Microorganism Depositary (2-5 Kazusa Kamatari, Kisarazu City, Chiba Prefecture, 292-0818). -8 Room 122).

In addition, the strains identified by the above-mentioned strain names are not limited to the strains themselves that have been deposited or registered with the designated institutions under the strain names (hereinafter also referred to as "deposited strains" for convenience of explanation); Strains substantially equivalent thereto (hereinafter also referred to as "derivative strains" or "derived strains") are also included. That is, for example, "Bifidobacterium bifidum ATCC 29521T" is not limited to the strain itself deposited with the above depository institution under the deposit number of Bifidobacterium bifidum ATCC 29521T, but also includes strains substantially equivalent thereto. be done.
A strain that is substantially equivalent to the deposited strain may be, for example, a derivative strain whose parent strain is the deposited strain. Derivative strains include strains bred from the deposited strain and naturally occurring strains from the deposited strain.

Derivative stocks include the following stocks:
(i) Strains determined to be the same strain by Randomly Amplified Polymorphic DNA method or Pulsed-field gel electrophoresis method (described in Probiotics in food/Health and nutritional properties and guidelines for evaluation 85 Page 43)
(ii) A strain that possesses only genes derived from the deposited strain, has no foreign genes, and has a DNA identity of 95% or more. (iii) A strain bred from the strain, or a strain that has been genetically modified or suddenly Strains with identical traits, including mutations or natural mutations

These bacteria belonging to Bifidobacterium bifidum have the ability to assimilate GMP. Bacteria belonging to Bifidobacterium bifidum produce assimilation products when metabolizing GMP. The produced assimilation products can be used by probiotics that do not have or have low GMP assimilation ability, such as bacteria belonging to Bifidobacterium breve and bacteria belonging to Bifidobacterium infantis, which will be described later. It is possible to grow probiotics that have no or low GMP assimilation ability.

In addition, in this technology, "no or low ability to assimilate GMP" means that when cultured in a medium containing sugars such as glucose and lactose, which are commonly assimilated by the probiotic Bifidobacterium, as a carbon source. It means a bacterium that has no or low ability to assimilate, metabolize, and proliferate sugar when cultured in a medium containing GMP as a carbon source.
For example, this can be confirmed by checking the turbidity of microorganisms and the specific growth rate of microorganisms in a medium containing only GMP as a carbon source.

The specific growth rate of a microorganism is defined as the increase in cells per unit time. If the ability to assimilate GMP is absent or low, the specific growth rate of microorganisms in a medium using GMP as a carbon source is usually 80% or less, preferably 70%, compared to a medium using glucose or lactose as a carbon source. Hereinafter, it means a decrease more preferably to 50% or less, still more preferably 10% or less.

In addition, probiotic bacteria that do not have the ability to assimilate GMP are those that are unable to utilize GMP at all, are partially deficient in metabolic pathways, and do not have transporters that take metabolized sugar into the body. It means a bacterial body.

In this technique, the amount of bacteria belonging to Bifidobacterium bifidum used for GMP assimilation can be freely set as long as the effect of this technique is not impaired. In this technique, in particular, the amount of bacteria belonging to Bifidobacterium bifidum used for assimilation of GMP can be set to 1×10 ³ to 1×10 ¹² CFU/mL or g relative to GMP.

In the composition according to the present technology, the bacteria belonging to Bifidobacterium bifidum used for assimilating GMP can also be removed after assimilating GMP. The method of removal is not particularly limited as long as it does not impair the effects of the present technology, and for example, removal methods such as filter removal and centrifugation can be freely selected and used. Furthermore, even if the composition according to the present technology contains bacteria belonging to Bifidobacterium bifidum without removing the bacteria belonging to Bifidobacterium bifidum, the effects of this technology can be achieved. It is.

(2) Bacteria belonging to Bifidobacterium breve Bacteria belonging to Bifidobacterium breve that can be used in this technology include known or unknown Bifidobacterium breve as long as it does not impair the effectiveness of this technology. One or more types of bacteria belonging to the following can be freely selected and used. Specifically, for example, Bifidobacterium breve (ATCC15700T), Bifidobacterium breve (NITE BP-02460), Bifidobacterium breve (FERM BP-11175), Bifidobacterium breve - Breve (NITE BP-02622), etc. can be mentioned.

Bifidobacterium breve (ATCC 15700T) is available from the ATCC American National Bacterial Culture Collection, 10801 University Boulevard, Manassas, VA 20110-2209, USA.

Bifidobacterium breve (NITE BP-02460) was acquired on April 24, 2017 by the National Institute of Technology and Evaluation, Patent Microorganisms Depositary (2-5-8 Kazusa Kamatari, Kisarazu City, Chiba Prefecture, 292-0818). It has been internationally deposited in Room 122) under the Budapest Treaty.

Bifidobacterium breve (FERM BP-11175) was discovered on August 25, 2009 at the National Institute of Advanced Industrial Science and Technology Patent Organism Depositary (1-1-1 Higashi, Tsukuba City, Ibaraki Prefecture, 305-8566, Japan). International deposit has been made in accordance with the Budapest Treaty at No. 6 (currently IPOD, National Institute of Technology and Evaluation, Patent Organism Depositary Center, Room 120, 2-5-8 Kazusa Kamatari, Kisarazu City, Chiba Prefecture, 292-0818 Japan). .

Bifidobacterium breve (NITE BP-02622) was acquired on January 26, 2018 by the National Institute of Technology and Evaluation, Patent Microorganisms Depositary (2-5-8 Kazusa Kamatari, Kisarazu City, Chiba Prefecture, 292-0818). It has been internationally deposited in Room 122) under the Budapest Treaty.

(3) Bacteria belonging to Bifidobacterium infantis Bacteria belonging to Bifidobacterium infantis that can be used in this technology include known or unknown bifids, as long as they do not impair the effectiveness of this technology. One or more types of bacteria belonging to Bacterium infantis can be freely selected and used. Specific examples include Bifidobacterium infantis (ATCC15697T) and Bifidobacterium infantis (NITE BP-02623).

Bifidobacterium infantis (ATCC 15697T) is available from the ATCC American National Bacterial Culture Collection, 10801 University Boulevard, Manassas, VA 20110-2209, USA.

Bifidobacterium infantis (NITE BP-02623) was transferred to the Patent Microorganisms Depositary Center, National Institute of Technology and Evaluation (2-5 Kazusa Kamatari, Kisarazu City, Chiba Prefecture, 292-0818) on January 26, 2018. -8 Room 122) has been internationally deposited under the Budapest Treaty.

(4) Carbohydrates As the carbohydrates that can be used in the present technology, one or more known or unknown carbohydrates can be freely selected and used as long as the effects of the present technology are not impaired. Specific examples include glucose, lactose, dextrin, sucrose, galactooligosaccharide, raffinose, lactulose, fructooligosaccharide, fucosylated oligosaccharide, acetylated oligosaccharide, and sialylated oligosaccharide.

2. Application of the composition according to the present technology By administering the composition of the present invention or the growth promoter of Bifidobacterium bacteria to humans, an effect of promoting the growth of probiotics acting on beneficial bacteria can be expected. That is, by proliferating probiotics in the intestines, the intestinal flora is improved. By improving the intestinal flora, diseases caused by the intestinal flora can be improved or prevented, making it possible to treat them.
Diseases caused by intestinal flora include inflammatory colitis, ulcerative colitis, irritable colitis, colon cancer, diabetes, and arteriosclerosis.
Furthermore, by administering the composition of the present invention or the growth promoter of Bifidobacterium bacteria, immune function is promoted. By promoting immune function, it becomes possible to prevent, improve, or treat allergies such as hay fever, asthma, and atopic dermatitis.
The composition or Bifidobacterium bacterial growth promoter according to the present technology can be suitably used in food and drink products, formula milk, pharmaceuticals, quasi-drugs, feeds, and the like.

Note that the present embodiment may be used for therapeutic purposes or non-therapeutic purposes.
"Non-therapeutic purpose" is a concept that does not include medical treatment, that is, treatment of the human body through treatment. Examples include health promotion and beauty practices.
"Amelioration" refers to improving a disease, symptom or condition; preventing or slowing the worsening of a disease, symptom or condition; reversing, preventing or slowing the progression of a disease or condition.
"Prevention" refers to preventing or delaying the onset of a disease or symptom in a target, or reducing the risk of a disease or symptom in a target.

In the composition, the probiotics, such as bacteria belonging to Bifidobacterium breve or Bifidobacterium infantis, are present in an amount of about 1 x 10 ³ to about 1 x 10 ¹² CFU/g per dosage unit. or mL, but higher values are also acceptable. When the composition is administered to an adult human (i.e., a person over 16 years of age), the probiotic is preferably at least 1 x 10 ⁶ CFU/g or mL or more, more preferably at least 1 x 10 ⁸ CFU/mL. Preferably, it is administered in g or mL or more. When the probiotic composition is administered to children (i.e., humans under the age of 16), the probiotic is preferably administered at a concentration of about 1×10 ⁶ to about 1×10 ¹¹ CFU/g or mL, more preferably about Preferably, doses are administered in the range of 1×10 ⁶ to about 1×10 ⁹ CFU/g or mL. Note that CFU means colony forming unit.

In the composition, the GMP assimilation product by bacteria belonging to Bifidobacterium bifidum is preferably administered in an amount of 0.05 to 20.0% by mass per dosage unit, but higher values are also acceptable. be done. When the composition is administered to an adult human, the assimilation product of GMP by bacteria belonging to Bifidobacterium bifidum is preferably administered in an amount of at least 0.05% by mass or more, more preferably at least 0.5% by mass. be done. When the composition is administered to children, the GMP assimilation product by bacteria belonging to Bifidobacterium bifidum is preferably administered in a range of 0.05 to 20.0% by mass.

In the Bifidobacterium genus bacterial growth promoter, GMP is preferably administered at 0.05 to 20.0% by mass per dosage unit, but higher values are also acceptable. When the Bifidobacterium bacterial growth promoter is administered to an adult human, GMP is preferably administered in an amount of at least 0.05% by mass or more, more preferably 0.5% by mass or more. When the Bifidobacterium bacterial growth promoter is administered to children, GMP is preferably administered in a range of 0.05 to 20.0% by mass.

The subjects to be administered according to the present invention are preferably infants.
Infants include infants and young children, more particularly infants, infants and newborns, and more particularly infants, infants, newborns, premature infants, preterm infants and low birth weight infants. In the present invention, the species of infant includes humans, unless otherwise specified. An infant refers to a child in its infancy period, and infancy refers to a period when the main source of nutrition is milk such as breast milk. In the case of humans, the period of infancy is usually less than one year old. Infants generally refer to children up to preschool age. A newborn baby refers to a child in the neonatal period, and the neonatal period refers to the period immediately after birth, and for humans, the neonatal period usually occurs within four weeks after birth.

Furthermore, in the present invention, prebiotics may also be used in addition to GMP assimilation products by bacteria belonging to Bifidobacterium bifidum. Prebiotics are indigestible food ingredients that have a beneficial effect on the host and improve its health by selectively altering the growth and activity of specific bacteria in the large intestine.
Prebiotics are not particularly limited as long as they can better exhibit the effects of the present invention when ingested together with the present bacteria or its culture, but include, for example, lactulose, raffinose, galactooligosaccharides, fructooligosaccharides, and soybean oligosaccharides. , milk-fruct oligosaccharide, xylooligosaccharide, isomooligosaccharide, coffee bean mannooligosaccharide, gluconic acid, polydextrose and inulin, with lactulose, raffinose and galactooligosaccharide being preferred, and lactulose, raffinose and galactooligosaccharide being more preferred.

<Food and beverages>
The composition or the Bifidobacterium genus bacterial growth promoter according to the present technology can also be prepared by adding it to a known food or drink. New food and drink products can also be produced by mixing the composition into raw materials for food and drink products.

Foods and beverages containing the composition or the Bifidobacterium bacterial growth promoter according to the present technology may be in the form of liquid, paste, solid, powder, etc., and may include tablets, liquid foods, etc., as well as wheat flour products. , instant foods, processed agricultural products, processed seafood products, processed livestock products, milk and dairy products, oils and fats, basic seasonings, compound seasonings and foods, frozen foods, confectionery, beverages, and other commercially available products. It will be done.

Examples of the flour products include bread, macaroni, spaghetti, noodles, cake mix, fried flour, and bread crumbs.
Examples of the instant foods include instant noodles, cup noodles, retort/cooked foods, cooked canned foods, microwave foods, instant soups/stews, instant miso soup/suimono, canned soups, freeze-dried foods, and other instant foods. It will be done.
Examples of the processed agricultural products include canned agricultural products, canned fruits, jams and marmalades, pickles, boiled beans, dried agricultural products, and cereals (processed grain products).
Examples of the processed seafood products include canned seafood, fish ham/sausage, fish paste products, seafood delicacies, and tsukudani.
Examples of the processed livestock products include canned livestock products, pastes, and livestock hams and sausages.
Examples of the milk/dairy products include fermented milk, processed milk, milk drinks, yogurts, lactic acid bacteria drinks, cheese, ice creams, formula milks, creams, and other dairy products.
Examples of the oils and fats include butter, margarines, and vegetable oils.
Examples of the basic seasoning include soy sauce, miso, sauces, processed tomato seasonings, mirin, and vinegar.
Examples of the complex seasonings/foods include cooking mixes, curry ingredients, sauces, dressings, noodle soups, spices, and other complex seasonings.
Examples of the frozen foods include raw frozen foods, semi-cooked frozen foods, and cooked frozen foods.
Examples of the confectionery include caramel, candy, chewing gum, chocolate, cookie, biscuit, cake, pie, snack, cracker, Japanese confectionery, rice confectionery, bean confectionery, dessert confectionery, and other confectionery.
Examples of the beverages include carbonated drinks, natural fruit juices, fruit juice drinks, soft drinks with fruit juice, pulp drinks, fruit drinks with fruit particles, vegetable drinks, soy milk, soy milk drinks, coffee drinks, tea drinks, powder drinks, and concentrated drinks. , sports drinks, nutritional drinks, alcoholic drinks and other recreational drinks.
Examples of commercially available foods other than those mentioned above include baby food, furikake, and ochazuke nori.

The content of GMP assimilation products by bacteria belonging to Bifidobacterium bifidum in the food and drink according to the present technology can be freely set as long as the effects of the present technology are not impaired. In this technology, in particular, the content of GMP assimilation products by bacteria belonging to Bifidobacterium bifidum in food and beverages is set to 0.05% to 20.0% of the final composition of food and beverages. can do.

The content of probiotics that do not have or have low GMP assimilation ability, such as bacteria belonging to Bifidobacterium breve or bacteria belonging to Bifidobacterium infantis, in foods and drinks related to this technology is It can be set freely as long as the effect is not impaired. In particular, with this technology, the content of probiotics that have a low ability to assimilate human milk oligosaccharides in foods and beverages is adjusted to 1×10 ³ to 1×10 12 CFU/mL or 1×10 3 to 1×10 ¹² CFU/mL or g.

[Food and beverages with functional claims]
Furthermore, the food and drink products defined by the present technology can also be provided and sold as food and drink products with specific uses (particularly health care uses) and functions displayed.
The act of "displaying" includes all acts to inform the consumer of the above-mentioned use, and if it is an expression that can recall or infer the above-mentioned use, the purpose of the display, the content of the display, the display Regardless of the object, medium, etc. used, all acts fall under the act of "display" under this technology.

Moreover, it is preferable that the "display" be performed using expressions that allow the consumer to directly recognize the above-mentioned use. Specifically, the act of transferring, handing over, exhibiting for transfer or delivery, or importing food and drink products or products with the above-mentioned usage written on their packaging, advertisements related to products, price lists, or transaction documents. Examples include actions such as displaying or distributing information with the above-mentioned uses written thereon, or writing the above-mentioned uses on information containing these items and providing it by electromagnetic (Internet, etc.) method.

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

In addition, "labeling" includes health foods, functional foods, foods for the sick, enteral nutritional foods, foods for special purposes, foods with health claims, foods for specified health uses, foods with functional claims, foods with nutritional function claims, and pharmaceutical products. It may also be indicated as a foreign product. Among these, in particular, labeling approved by the Consumer Affairs Agency, for example, labeling approved under the Food for Specified Health Use System, the Food with Function Claims System, and similar systems, etc. can be mentioned. More specifically, labeling as a food for specified health uses, labeling as a food for specified health uses with conditions, labeling as a food with functional claims, labeling that it affects the structure or function of the body, labeling to reduce disease risk, etc. can be mentioned. Among these, a typical example is labeling as a food for specified health uses (especially labeling for health purposes) as stipulated in the Health Promotion Law Enforcement Regulations (Ordinance No. 86 of the Ministry of Health, Labor and Welfare of Japan, April 30, 2003). , labeling as a food with functional claims stipulated in the Food Labeling Act (Act No. 70 of 2013), and similar labeling.

[Formulated milk]
It is particularly preferable that the food or drink containing the composition or the Bifidobacterium bacterial growth promoter according to the present technology is a formula milk. Formulas include infant formula, follow-up milk, low birth weight infant formula, school child formula, adult formula, elderly formula, allergy formula, and lactose intolerance formula. , formula milk for congenital metabolic abnormalities, etc. In particular, it is preferably used for infant formula, follow-up milk, low birth weight infant formula, and school child formula. Infant formula refers to infant formula for infants aged 0 to 12 months, follow-up milk for infants aged 6 to 9 months and younger children (up to 3 years of age), and We sell formula milk for low birth weight infants weighing less than 2,500 g (low birth weight infants), and various therapeutic milks used to treat infants with medical conditions such as cow's milk allergy and lactose intolerance. Point. Furthermore, the composition can be applied to foods with health claims and foods for patients. The food with health claims system was established to cover not only regular foods but also foods in the form of tablets, capsules, etc., based on domestic and international trends and consistency with the existing food for specified health use system. It consists of two types: food for specified health uses (individual permission type) and food with nutritional function functions (standard type).

By administering the composition of the present technology, bacteria belonging to Bifidobacterium breve and/or bacteria belonging to Bifidobacterium infantis proliferate in the intestine of the subject. Furthermore, by administering the Bifidobacterium bacterial growth promoter of the present technology, bacteria belonging to Bifidobacterium bifidum proliferate in the intestine of the subject. Proliferation of these probiotics in the intestines improves immunity and further improves intestinal flora. Particularly in infants, by administering the composition of the present invention or the growth promoter of Bifidobacterium bacteria, the number of bifidobacteria in the intestine is effectively increased and the intestinal environment is maintained. Furthermore, by incorporating bad bacteria and opportunistic bacteria along with bifidobacteria, immunity can be acquired, and the resulting intestinal flora and other intestinal environment can be carried over into adulthood.

As the method for producing formula milk, one or more known production methods can be used in any combination as long as the effects of the present technology are not impaired. Specifically, for example, the composition according to the present technology and various additives are mixed with raw milk, and the product can be manufactured through steps such as homogenization, sterilization, concentration, drying, granulation, and sieving.

Specifically, the formula milk of the present technology can be produced, for example, by the following method.

That is, the present technology utilizes the assimilation products of GMP by bacteria belonging to Bifidobacterium bifidum and, if necessary, the products belonging to bacteria belonging to Bifidobacterium breve and/or Bifidobacterium infantis. Provided is a method for producing formula milk, which involves mixing bacterial powder containing bacteria to obtain formula milk.
Specifically, a method for producing formula milk is provided, which includes, for example, the following steps A and B:
Step A: A step of culturing bacteria belonging to Bifidobacterium bifidum in a medium containing GMP to obtain a culture;
Step B: A step of mixing the culture, milk components, and, if necessary, bacteria belonging to Bifidobacterium breve and/or bacteria belonging to Bifidobacterium infantis to obtain prepared milk.

Moreover, the food composition of the present technology may be, for example, a supplement for infants. A supplement for infants can be produced, for example, by the following method.

That is, the present technology provides a method for producing a supplement, including, for example, the following steps C to E:
Step C: A step of culturing bacteria belonging to Bifidobacterium bifidum in a medium containing GMP to obtain a culture;
Step D: Mix the culture, milk components, excipients, and if necessary bacteria belonging to Bifidobacterium breve and/or bacteria belonging to Bifidobacterium infantis, and prepare the mixture. The process of obtaining;
Step E: Step of forming the mixture into a formulation by tableting or the like.

In any of the above manufacturing methods, components other than those mentioned in the above steps may be used in combination as appropriate.

The method of adding the composition according to the present technology is not particularly limited as long as it does not impair the effects of the present technology, but for example, in addition to adding the composition according to the present technology in liquid form during mixing as described above, It can be added later and mixed into powder for production.

The content of GMP assimilation products or GMP produced by bacteria belonging to Bifidobacterium bifidum in the infant formula according to the present technology can be freely set as long as the effects of the present technology are not impaired. In particular, with this technology, the content of GMP assimilation products or GMP by bacteria belonging to Bifidobacterium bifidum in formula milk is reduced from 0.05% to 20.0% with respect to the final composition of formula milk. Can be set to .

The content of probiotics with no or low ability to assimilate GMP, such as bacteria belonging to Bifidobacterium breve and bacteria belonging to Bifidobacterium infantis, in the formula according to this technology is It can be set freely as long as the effect is not impaired. In particular, with this technology, the content of probiotics that have a low ability to assimilate human milk oligosaccharides in formula milk is adjusted to 1 x 10 ³ to 1 x 10 ¹² CFU/mL or 1 x 10 12 CFU/mL to the final composition of formula milk. g.

<Drugs, quasi-drugs>
The composition or the Bifidobacterium genus bacterial growth promoter according to the present technology may be added to known pharmaceuticals or quasi-drugs (hereinafter also referred to as "medicines, etc."). New medicines, etc. can be manufactured by mixing the composition according to the present technology into raw materials for medicines, etc.

When a pharmaceutical product, etc. contains a composition according to the present technology or a Bifidobacterium bacterial growth promoter, the pharmaceutical product, etc. can be formulated into a desired dosage form as appropriate depending on the administration method, such as oral administration or parenteral administration. can do. The dosage form is not particularly limited, but for oral administration, for example, solid preparations such as powders, granules, tablets, troches, and capsules; liquid preparations such as solutions, syrups, suspensions, and emulsions. can be converted into For parenteral administration, it can be formulated into suppositories, sprays, inhalants, ointments, patches, and injections, for example. In the present technology, it is preferable to formulate the drug into a dosage form for oral administration.
In addition, formulation can be carried out by a known method as appropriate depending on the dosage form.

When formulating, a pharmaceutical carrier may be added as appropriate. In addition to the peptide of the present technology, components such as excipients, pH adjusters, coloring agents, and flavoring agents that are commonly used in formulation can be used. Furthermore, components having the effect of preventing, improving, and/or treating diseases and symptoms that are known or will be discovered in the future may be used in combination depending on the purpose.

As the pharmaceutical carrier, various organic or inorganic carriers can be used depending on the dosage form. Carriers for solid preparations include, for example, excipients, binders, disintegrants, lubricants, stabilizers, and flavorings.

Examples of the excipient include sugar derivatives such as lactose, sucrose, glucose, mannitol, and sorbitol; starch derivatives such as corn starch, potato starch, α-starch, dextrin, and carboxymethyl starch; crystalline cellulose, and hydroxypropyl cellulose. , cellulose derivatives such as hydroxypropylmethylcellulose, carboxymethylcellulose, and carboxymethylcellulose calcium; gum arabic; dextran; pullulan; silicate derivatives such as light anhydrous silicic acid, synthetic aluminum silicate, and magnesium aluminate metasilicate; phosphate derivatives such as calcium phosphate; Examples include carbonate derivatives such as calcium carbonate; and sulfate derivatives such as calcium sulfate.

Examples of the binder include, in addition to the excipients described above, gelatin, polyvinylpyrrolidone, and macrogol.

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

Examples of the lubricant include talc; stearic acid; stearic acid metal salts such as calcium stearate and magnesium stearate; colloidal silica; waxes such as pea gum and gay wax; boric acid; glycol; carboxylic acid such as fumaric acid and adipic acid. Acids; 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 acids such as silicic anhydride and silicic acid hydrate; and starch derivatives. Can be mentioned.

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

Examples of the flavoring agent include sweeteners, acidulants, and fragrances.
Note that carriers used in the case of liquid preparations for oral administration include, for example, solvents such as water, and flavoring agents.

The content of GMP assimilation products or GMP produced by bacteria belonging to Bifidobacterium bifidum in the pharmaceutical according to the present technology can be freely set as long as the effect of the present technology is not impaired. In particular, with this technology, the content of the GMP bacterial assimilation products belonging to Bifidobacterium bifidum in the drug is set to 0.05% to 20.0% with respect to the final composition of the drug. can do.

The content of probiotics with no or low ability to assimilate GMP, such as bacteria belonging to Bifidobacterium breve or Bifidobacterium infantis, in the pharmaceutical products related to this technology is It can be set freely as long as the effect is not impaired. In particular, with this technology, the content of probiotics that have a low ability to assimilate human milk oligosaccharides in pharmaceuticals can be reduced to 1×10 ³ to 1×10 ¹² CFU/mL or g in the final composition of pharmaceuticals. Can be set.

<Feed>
The composition according to the present technology or the Bifidobacterium bacterial growth promoter may be added to known feeds. Feed can be manufactured by mixing the composition into raw materials for feed.

When the feed contains the composition according to the present technology or the growth promoter of Bifidobacterium bacteria, the raw materials for the feed include, for example, cereals such as corn, wheat, barley, and rye; bran, wheat bran, rice bran, and defatted rice bran. Rice bran such as corn gluten meal, corn jam meal, etc.; Animal feed such as skim milk powder, whey, fish meal, bone meal; Yeast such as brewer's yeast; Mineral feed such as calcium phosphate, calcium carbonate; Examples include fats and oils; amino acids; and saccharides. Further, examples of the form of the feed include feed for pet animals such as pet food, livestock feed, and fish feed.

The content of GMP assimilation products or GMP produced by bacteria belonging to Bifidobacterium bifidum in the feed according to the present technology can be freely set as long as the effects of the present technology are not impaired. In this technology, in particular, the content of GMP, a synthetic product produced by bacteria belonging to Bifidobacterium bifidum, in the feed is set to 0.05% to 20.0% of the final composition of the feed. can do.

The content of probiotics such as bacteria belonging to Bifidobacterium breve or bacteria belonging to Bifidobacterium infantis that do not have or have low ability to assimilate GMP in the feed related to this technology is It can be set freely as long as the effect is not impaired. In particular, with this technology, the content of probiotics that have a low ability to assimilate human milk oligosaccharides in the feed is reduced to 1×10 ³ to 1×10 ¹² CFU/mL or g based on the final composition of the feed. Can be set.

3. Method for producing a substance that is detected as an anion with a mass-to-charge ratio of 324 or 363 when subjected to a mass spectrometer after being derivatized with 2-aminobenzamide As mentioned above, Bifidobacterium bifidum When the assimilated product of κ-casein glycomacropeptide by bacteria belonging to 2-aminobenzamide was derivatized with 2-aminobenzamide and then subjected to a mass spectrometer, it was detected as an anion with a mass-to-charge ratio of 324 or 363. Contains substances that That is, by assimilating κ-casein glycomacropeptide using bacteria belonging to Bifidobacterium bifidum, when it is derivatized with 2-aminobenzamide and then subjected to a mass spectrometer, its mass-to-charge ratio is It is possible to produce a substance that is detected as a 324 or 363 anion.

Specifically, by performing a fermentation process in which κ-casein glycomacropeptide is fermented using bacteria belonging to Bifidobacterium bifidum, it is derivatized with 2-aminobenzamide and then subjected to a mass spectrometer. When this is done, it is possible to produce a substance that is detected as an anion with a mass-to-charge ratio of 324 or 363.

In the fermentation step of the method for producing the substance according to the present technology, a general method can be used as a specific method as long as it does not impair the effects of the present invention. For example, fermentation can be carried out by adding a culture solution of bacteria belonging to Bifidobacterium bifidum to a κ-casein glycomacropeptide solution.

At this time, the concentration of the κ-casein glycomacropeptide solution is not limited as long as it does not impair the effects of the present technology, but in this technology, the concentration of the κ-casein glycomacropeptide solution is particularly set to 0.5 to 15%. It is preferably set to 0.8% to 12%, and more preferably set to 0.8% to 12%. By setting the concentration of the κ-casein glycomacropeptide solution within this range, inhibition of growth of bacteria belonging to Bifidobacterium bifidum can be prevented.

Furthermore, the amount of the culture solution of bacteria belonging to Bifidobacterium bifidum to be added is not limited as long as it does not impair the effects of the present technique, but in the present technique, it is particularly preferable to add 0.5 to 5%. By setting the amount of the culture solution to be added within this range, it is possible to suppress the influence of the culture medium brought into the culture solution.

Note that the present technology can also adopt the following configuration.
[1]
A composition containing a product of κ-casein glycomacropeptide assimilated by bacteria belonging to Bifidobacterium bifidum.
[2]
A food/beverage composition comprising a composition containing a product of κ-casein glycomacropeptide assimilated by bacteria belonging to Bifidobacterium bifidum.
[3]
A formula comprising a composition containing a product of κ-casein glycomacropeptide assimilated by bacteria belonging to Bifidobacterium bifidum.
[4]
A pharmaceutical or quasi-drug composition comprising a composition containing a product of κ-casein glycomacropeptide assimilated by bacteria belonging to Bifidobacterium bifidum.
[5]
A feed composition comprising a composition containing a product of κ-casein glycomacropeptide assimilated by bacteria belonging to Bifidobacterium bifidum.
[6]
Use of the assimilation products of κ-casein glycomacropeptide by bacteria belonging to Bifidobacterium bifidum for compositions.
[7]
Use of the assimilation product of κ-casein glycomacropeptide by bacteria belonging to Bifidobacterium bifidum for food and drink compositions.
[8]
Use of the assimilation products of κ-casein glycomacropeptide by bacteria belonging to Bifidobacterium bifidum for formula milk.
[9]
Use of the assimilation product of κ-casein glycomacropeptide by bacteria belonging to Bifidobacterium bifidum for pharmaceutical or quasi-drug compositions.
[10]
Use of the assimilation products of κ-casein glycomacropeptide by bacteria belonging to Bifidobacterium bifidum for feed compositions.
[11]
for the production of compositions,
Use of κ-casein glycomacropeptide assimilation products by bacteria belonging to Bifidobacterium bifidum.
[12]
For the production of food and drink compositions,
Use of κ-casein glycomacropeptide assimilation products by bacteria belonging to Bifidobacterium bifidum.
[13]
For the production of formula milk,
Use of κ-casein glycomacropeptide assimilation products by bacteria belonging to Bifidobacterium bifidum.
[14]
For the production of pharmaceutical or quasi-drug compositions,
Use of κ-casein glycomacropeptide assimilation products by bacteria belonging to Bifidobacterium bifidum.
[15]
for the production of feed compositions,
Use of κ-casein glycomacropeptide assimilation products by bacteria belonging to Bifidobacterium bifidum.
[16]
Assimilation products of κ-casein glycomacropeptide by bacteria belonging to Bifidobacterium bifidum;
Bacteria belonging to Bifidobacterium breve and/or bacteria belonging to Bifidobacterium infantis;
A composition containing.
[17]
Assimilation products of κ-casein glycomacropeptide by bacteria belonging to Bifidobacterium bifidum;
A food or drink composition containing a composition containing bacteria belonging to Bifidobacterium breve and/or bacteria belonging to Bifidobacterium infantis.
[18]
Assimilation products of κ-casein glycomacropeptide by bacteria belonging to Bifidobacterium bifidum;
A composition containing bacteria belonging to Bifidobacterium breve and/or bacteria belonging to Bifidobacterium infantis;
Formula containing.
[19]
Assimilation products of κ-casein glycomacropeptide by bacteria belonging to Bifidobacterium bifidum;
A composition containing bacteria belonging to Bifidobacterium breve and/or bacteria belonging to Bifidobacterium infantis;
A pharmaceutical or quasi-drug composition containing.
[20]
Assimilation products of κ-casein glycomacropeptide by bacteria belonging to Bifidobacterium bifidum;
A composition containing bacteria belonging to Bifidobacterium breve and/or bacteria belonging to Bifidobacterium infantis;
A feed composition containing.
[21]
for the composition,
Assimilation products of κ-casein glycomacropeptide by bacteria belonging to Bifidobacterium bifidum;
Bacteria belonging to Bifidobacterium breve and/or bacteria belonging to Bifidobacterium infantis;
Use of.
[22]
For food and drink compositions,
Assimilation products of κ-casein glycomacropeptide by bacteria belonging to Bifidobacterium bifidum;
Bacteria belonging to Bifidobacterium breve and/or bacteria belonging to Bifidobacterium infantis;
Use of.
[23]
for formula milk,
Assimilation products of κ-casein glycomacropeptide by bacteria belonging to Bifidobacterium bifidum;
Bacteria belonging to Bifidobacterium breve and/or bacteria belonging to Bifidobacterium infantis;
Use of.
[24]
For pharmaceutical or quasi-drug compositions,
Assimilation products of κ-casein glycomacropeptide by bacteria belonging to Bifidobacterium bifidum;
Bacteria belonging to Bifidobacterium breve and/or bacteria belonging to Bifidobacterium infantis;
Use of.
[25]
for feed compositions,
Assimilation products of κ-casein glycomacropeptide by bacteria belonging to Bifidobacterium bifidum;
Bacteria belonging to Bifidobacterium breve and/or bacteria belonging to Bifidobacterium infantis;
Use of.
[26]
for the production of compositions,
Assimilation products of κ-casein glycomacropeptide by bacteria belonging to Bifidobacterium bifidum;
Bacteria belonging to Bifidobacterium breve and/or bacteria belonging to Bifidobacterium infantis;
Use of.
[27]
For the production of food and drink compositions,
Assimilation products of κ-casein glycomacropeptide by bacteria belonging to Bifidobacterium bifidum;
Bacteria belonging to Bifidobacterium breve and/or bacteria belonging to Bifidobacterium infantis;
Use of.
[28]
For the production of formula milk,
Assimilation products of κ-casein glycomacropeptide by bacteria belonging to Bifidobacterium bifidum;
Bacteria belonging to Bifidobacterium breve and/or bacteria belonging to Bifidobacterium infantis;
Use of.
[29]
For the production of pharmaceutical or quasi-drug compositions,
Assimilation products of κ-casein glycomacropeptide by bacteria belonging to Bifidobacterium bifidum;
Bacteria belonging to Bifidobacterium breve and/or bacteria belonging to Bifidobacterium infantis;
Use of.
[30]
for the production of feed compositions,
Assimilation products of κ-casein glycomacropeptide by bacteria belonging to Bifidobacterium bifidum;
Bacteria belonging to Bifidobacterium breve and/or bacteria belonging to Bifidobacterium infantis;
Use of.
[31]
In [1] to [30],
The above-mentioned nutraceutical products are purified using a sugar refining solid-phase extraction column, derivatized with 2-aminobenzamide, and then subjected to a mass spectrometer, where they are detected as anions with a mass-to-charge ratio of 324 or 363. Contains substances that
[32]
A Bifidobacterium bacteria growth promoter containing κ-casein glycomacropeptide, which causes bacteria belonging to Bifidobacterium bifidum to specifically grow compared to other Bifidobacterium bacteria.
[33]
A preparation containing a Bifidobacterium growth promoter that specifically grows bacteria belonging to Bifidobacterium bifidum, including κ-casein glycomacropeptide, compared to other Bifidobacterium bacteria. milk.
[34]
A pharmaceutical product containing a Bifidobacterium growth promoter that specifically grows bacteria belonging to Bifidobacterium bifidum, including κ-casein glycomacropeptide, compared to other Bifidobacterium bacteria. or a quasi-drug composition.
[35]
Feed containing a Bifidobacterium growth promoter that specifically grows bacteria belonging to Bifidobacterium bifidum, including κ-casein glycomacropeptide, compared to other Bifidobacterium bacteria. Composition.
[36]
For a Bifidobacterium bacteria growth promoter that specifically grows bacteria belonging to Bifidobacterium Bifidum compared to other Bifidobacterium bacteria,
Use of κ-casein glycomacropeptide.
[37]
For a food and drink composition for promoting the growth of Bifidobacterium bacteria, which specifically grows bacteria belonging to Bifidobacterium Bifidum compared to other Bifidobacterium bacteria,
Use of κ-casein glycomacropeptide.
[38]
For formula milk for promoting the growth of Bifidobacterium bacteria, which specifically grows bacteria belonging to Bifidobacterium Bifidum compared to other Bifidobacterium bacteria.
Use of κ-casein glycomacropeptide.
[39]
For a pharmaceutical or quasi-drug composition for promoting the growth of Bifidobacterium bacteria that specifically grows bacteria belonging to Bifidobacterium Bifidum compared to other Bifidobacterium bacteria. ,
Use of κ-casein glycomacropeptide.
[40]
For a feed composition for promoting the growth of Bifidobacterium bacteria that specifically grows bacteria belonging to Bifidobacterium Bifidum compared to other Bifidobacterium bacteria,
Use of κ-casein glycomacropeptide.
[41]
For the production of a Bifidobacterium bacteria growth promoter that specifically grows bacteria belonging to Bifidobacterium Bifidum compared to other Bifidobacterium bacteria,
Use of κ-casein glycomacropeptide.
[42]
For producing a food and drink composition for promoting the growth of Bifidobacterium bacteria, which specifically grows bacteria belonging to Bifidobacterium Bifidum compared to other Bifidobacterium bacteria.
Use of κ-casein glycomacropeptide.
[43]
For the production of a formula for promoting the growth of Bifidobacterium bacteria, which specifically grows bacteria belonging to Bifidobacterium Bifidum compared to other Bifidobacterium bacteria,
Use of κ-casein glycomacropeptide.
[44]
Production of a pharmaceutical or quasi-drug composition for promoting the growth of Bifidobacterium bacteria that specifically grows Bifidobacterium Bifidum bacteria compared to other Bifidobacterium bacteria. for,
Use of κ-casein glycomacropeptide.
[45]
For producing a feed composition for promoting the growth of Bifidobacterium bacteria, which specifically grows bacteria belonging to Bifidobacterium Bifidum compared to other Bifidobacterium bacteria.
Use of κ-casein glycomacropeptide.
[46]
In [32] to [45],
The Bifidobacterium bacterial growth promoter, the food and drink composition, the formula milk, the pharmaceutical, the quasi-drug composition, or the feed composition further contains a carbohydrate.
[47]
In [1] to [45],
The bacteria belonging to Bifidobacterium bifidum include Bifidobacterium bifidum (ATCC29521T), Bifidobacterium bifidum (NITE BP-02429), Bifidobacterium bifidum (NITE BP-01252), and Bifidobacterium bifidum (NITE BP-01252). Bacterium bifidum (NITE BP-02431), Bifidobacterium bifidum (NITE BP-02648), Bifidobacterium bifidum (NITE BP-02432), Bifidobacterium bifidum (NITE BP-02433), One or more bacteria selected from.
[48]
In [16] to [31],
Bacteria belonging to Bifidobacterium breve include Bifidobacterium breve (ATCC15700T), Bifidobacterium breve (NITE BP-02460), Bifidobacterium breve (FERM BP-11175), and Bifidobacterium breve. um breve (NITE BP-02622).
[49]
In [16] to [32],
The bacteria belonging to Bifidobacterium infantis are one or more bacteria selected from Bifidobacterium infantis (ATCC15697T) and Bifidobacterium infantis (NITE BP-02623). .
[50]
A method for producing formula milk, including the following steps A and B.
Step A: A step of culturing bacteria belonging to Bifidobacterium bifidum in a medium containing GMP to obtain a culture;
Step B: A step of mixing the culture and milk components to obtain prepared milk.
[51]
A method for producing a supplement, including the following steps C to E.
Step C: A step of culturing bacteria belonging to Bifidobacterium bifidum in a medium containing GMP to obtain a culture;
Step D: mixing the culture, milk components, and excipients to obtain a mixture;
Step E: Step of forming the mixture into a formulation by tableting or the like.
[52]
A method for producing formula milk, comprising the following steps F and G.
Step F: Cultivating bacteria belonging to Bifidobacterium bifidum in a medium containing GMP to obtain a culture;
Step G: A step of mixing the culture, milk components, and bacteria belonging to Bifidobacterium breve and/or bacteria belonging to Bifidobacterium infantis to obtain prepared milk.
[53]
A method for producing a supplement, including the following steps H to J.
Step H: A step of culturing bacteria belonging to Bifidobacterium bifidum in a medium containing GMP to obtain a culture;
Step I: mixing the culture, milk components, excipients, and bacteria belonging to Bifidobacterium breve and/or bacteria belonging to Bifidobacterium infantis to obtain a mixture;
Step J: Step of forming the mixture into a formulation by tableting or the like.
[54]
In [50] to [53],
The milk component is milk protein.
[55]
In [54],
The milk protein is at least one component selected from the group consisting of whey, whey hydrolyzate, and casein.
[56]
In [50] to [55],
The bacteria belonging to Bifidobacterium bifidum include Bifidobacterium bifidum (ATCC29521T), Bifidobacterium bifidum (NITE BP-02429), Bifidobacterium bifidum (NITE BP-01252), and Bifidobacterium bifidum (NITE BP-01252). Bacterium bifidum (NITE BP-02431), Bifidobacterium bifidum (NITE BP-02648), Bifidobacterium bifidum (NITE BP-02432), Bifidobacterium bifidum (NITE BP-02433), One or more bacteria selected from.
[57]
In [52] to [56],
Bacteria belonging to Bifidobacterium breve include Bifidobacterium breve (ATCC15700T), Bifidobacterium breve (NITE BP-02460), Bifidobacterium breve (FERM BP-11175), and Bifidobacterium breve. um breve (NITE BP-02622).
[58]
In [52] to [57],
The bacteria belonging to Bifidobacterium infantis are one or more bacteria selected from Bifidobacterium infantis (ATCC15697T) and Bifidobacterium infantis (NITE BP-02623). .
[59]
Bacteria belonging to Bifidobacterium bifidum for producing a substance that is detected as an anion with a mass-to-charge ratio of 324 or 363 when subjected to a mass spectrometer after being derivatized with 2-aminobenzamide. Use of.
[60]
A fermentation process comprising fermenting κ-casein glycomacropeptide using bacteria belonging to Bifidobacterium bifidum.
A method for producing a substance that is detected as an anion with a mass-to-charge ratio of 324 or 363 when subjected to a mass spectrometer after being derivatized with 2-aminobenzamide.
[61]
In the fermentation step, 0.5 to 5% of a culture solution of bacteria belonging to Bifidobacterium bifidum is added to a κ-casein glycomacropeptide solution with a concentration of 0.8% to 12% and fermented [60 The manufacturing method described in ].

Hereinafter, the present technology will be described in more detail based on examples. Note that the embodiment described below shows one example of a typical embodiment of the present technology, and therefore the scope of the present technology should not be interpreted narrowly.

[Experiment example 1]
In Experimental Example 1, the ability of various Bifidobacteria to assimilate GMP was investigated.

<Experimental method>
(1) Preparation of sugar solution Glucose (manufactured by Nacalai Tesque Co., Ltd.) was dissolved in ion-exchanged water to a concentration of 1%. Further, GMP (manufactured by Sigma-Aldrich) was dissolved in ion-exchanged water to a concentration of 10%. Thereafter, the obtained glucose solution and the obtained GMP solution were sterile-filtered using a 0.22 μm filter (manufactured by Merck Millipore).

(2) Preparation of culture solution for each strain A liquid medium is prepared aseptically by adding L-cysteine hydrochloride (Kanto Kagaku) to Lactobacilli MRS medium (manufactured by Nippon Becton Dickinson) to a concentration of 0.05%. (hereinafter referred to as "MRS+Cys medium"). For the bacteria belonging to Bifidobacterium bifidum shown in Table 1 below and the strains of bifidobacteria other than Bifidobacterium bifidum shown in Table 2, the culture solution of each strain was maintained in MRS + Cys medium in advance. The cells were inoculated into a newly prepared MRS+Cys medium at an inoculum amount of 3%, and cultured anaerobically at 37°C for 16 hours.

The obtained culture solution of each strain was inoculated into a newly prepared MRS+Cys medium at an inoculum amount of 3%, and cultured anaerobically at 37° C. for 16 hours. The obtained culture solution of each strain was used in the following tests.

(3) Comparison of GMP assimilation ability between different Bifidobacteria strains A medium was prepared based on the composition of the MRS liquid medium, and the glucose prepared in (1) was added as a sugar source to 0.1%. was used as MRS+Glc medium. Furthermore, a medium was prepared in which the GMP concentration prepared in (1) was added as a sugar source to the MRS+Glc medium at 0.8%, and this was used as an MRS+Glc+GMP medium. The culture solution of each bacterial strain prepared in (2) was inoculated into these media so that the inoculum amount was 1%. After culturing the cells anaerobically at 37°C for 24 hours, OD600 was measured.

The ability of each strain to assimilate GMP was evaluated by the "growth ratio 1" calculated from the following formula (1). The evaluation results are shown in Table 3 below.

As shown in Table 3, the growth ratios 1 of the bacteria belonging to Bifidobacterium bifidum shown in Table 1 were all values of 1 or more, whereas the growth ratios of other Bifidobacteria shown in Table 2 were: It showed a value lower than 1. That is, it was found that the bacteria belonging to Bifidobacterium bifidum shown in Table 1 have the ability to assimilate GMP, whereas the other Bifidobacteria shown in Table 2 do not have the ability to assimilate GMP. Ta.

[Experiment example 2]
In Experimental Example 2, the utilization of assimilation products by bacteria belonging to GMP Bifidobacterium bifidum by other Bifidobacteria shown in Table 2 was investigated.

<Experimental method>
(1) Preparation of sugar solution A 10% GMP solution was prepared according to the method of Experimental Example 1 above.

(2) Preparation of culture solution for each strain (pre-culture/pre-culture)
Based on the method of Experimental Example 1, culture solutions of each Bifidobacterium strain listed in Tables 1 and 2 were prepared and used in the following tests.

(3) Utilization of assimilation products by bacteria belonging to GMP Bifidobacterium bifidum A medium in which the 10% GMP solution prepared in (1) was added to the MRS+Cys medium so that the GMP concentration was 0.8%. (hereinafter referred to as "MRS+GMP medium 1"). The culture solution of each strain listed in Table 1 prepared in (2) was inoculated into this medium at an inoculum amount of 1%, and cultured anaerobically at 37° C. for 24 hours.

Each culture solution was subjected to centrifugation, and the resulting culture supernatant was subjected to sterile filtration through a 0.22 μm filter. The culture solution of each strain listed in Table 2 prepared in (2) was inoculated into the sterilized culture supernatant at an inoculum amount of 1%. After culturing the cells anaerobically at 37°C for 24 hours, OD600 was measured. As a test control, MRS+GMP medium 1, which was not inoculated with bacteria belonging to Bifidobacterium bifidum, was also inoculated with the culture solution of each strain listed in Table 2 prepared in (2) and incubated anaerobically at 37°C for 24 hours. Culture was performed for a period of time, and OD600 was measured.

The utilization of assimilation products by bacteria belonging to Bifidobacterium bifidum in GMP by each strain was evaluated by "growth ratio 2" calculated from the following formula (2). The results are shown in Table 4 below.

As shown in Table 4, all of the bacteria belonging to Bifidobacterium breve and the bacteria belonging to Bifidobacterium infantis exhibited a growth ratio 2 of 1 or more. That is, it was considered that bacteria belonging to Bifidobacterium breve and bacteria belonging to Bifidobacterium infantis could utilize the assimilation products of GMP by bacteria belonging to Bifidobacterium bifidum. Moreover, its utilization was significantly higher in bacteria belonging to Bifidobacterium bifidum.

On the other hand, the bacteria belonging to Bifidobacterium lactis and the bacteria belonging to Bifidobacterium longum exhibited a growth ratio 2 value of approximately 1 or less than 1. That is, bacteria belonging to Bifidobacterium lactis and bacteria belonging to Bifidobacterium longum have low or no utilization of the products utilized by bacteria belonging to Bifidobacterium bifidum in GMP. It was considered.

[Experiment example 3]
In Experimental Example 3, an analysis was conducted to identify the assimilated products of bacteria belonging to Bifidobacterium bifidum under GMP.

<Experimental method>
(1) Preparation of sugar solution A 16% GMP solution was prepared according to the method of Experimental Example 1 above.

(2) Preparation of culture solution for each strain Based on the method of Experimental Example 1 above, culture solutions for Bifidobacterium bifidum (NITE BP-02648) and Bifidobacterium breve (ATCC15700T) were prepared. , they were used in the following tests.

(3) Preparation of assimilated products by bacteria belonging to Bifidobacterium bifidum using GMP A 16% GMP solution and MRS+Cys medium were mixed at a ratio of 1:3 (hereinafter referred to as "MRS+GMP medium 2"). The culture solution of each strain listed in Table 1 prepared in (2) above was inoculated into this medium at an inoculum amount of 1%, and cultured anaerobically at 37° C. for 24 hours. Each obtained culture solution was subjected to centrifugation, and the obtained culture supernatant was subjected to sterile filtration through a 0.22 μm filter. The culture solution of Bifidobacterium bifidum (NITE BP-02648) was aseptically filtered, then inoculated with Bifidobacterium breve (ATCC15700T) at an inoculum amount of 1%, and then incubated anaerobically at 37°C. The cells were cultured for 24 hours. The obtained culture solution was subjected to centrifugation, and the obtained culture supernatant was subjected to sterile filtration through a 0.22 μm filter.

For MRS+GMP medium 2 and the above culture supernatant, lipids were removed by the Folch method, and then proteins were removed by the ethanol precipitation method to concentrate the sugar fraction. That is, 1 mL of the sterile-filtered medium was mixed with 4 mL of a chloroform/methanol (2/1) mixture, and then subjected to centrifugation. After collecting only the upper layer of the solution divided into two layers, an equal amount of ethanol, which had been previously cooled at -30°C, was added. The resulting mixture was stored at -30°C for 1 hour to allow protein precipitation to proceed. Thereafter, the stored mixture was subjected to centrifugation, and only the sugar fraction, which was the supernatant, was collected. The collected sugar fraction was dried using a centrifugal concentrator and then redissolved in 100 μL of ultrapure water. The obtained sugar solution was applied to a sugar purification solid-phase extraction column GlycoBlot (manufactured by Sumitomo Bakelite), and sugar purification and sugar derivatization treatment with 2-aminobenzamide (2-AB) were performed according to the attached protocol. went.

(4) GMP liquid chromatograph-mass spectrometer (LC-MS) analysis of assimilation products by bacteria belonging to Bifidobacterium bifidum. 10 μL of the total sample volume of 100 μL obtained by the above method was Under the conditions shown in Table 5, the molecular weight of the assimilated products was determined by LC-MS analysis.

The total ion chromatogram (TIC) is shown in FIG. In the MRS + GMP2 medium, no significant peak was observed after a retention time (RT) of 11 minutes (Fig. 1A), whereas in the culture supernatant of Bifidobacterium bifidum (NITE BP-02648), Multiple peaks were observed at retention times (RT) of 11 to 25 minutes (Figure 1B). These peaks were not confirmed in the culture supernatant of Bifidobacterium breve (ATCC15700T) strain. When Bifidobacterium breve (ATCC15700T) strain was ingested and cultured in the culture supernatant of this Bifidobacterium bifidum (NITE BP-02648) strain, one of the peaks with a retention time (RT) of 12.6 minutes was observed. disappeared, and the peak at a retention time (RT) of 21.4 minutes attenuated (FIG. 1C). The mass-to-charge ratio of the ions detected at each retention time (RT) was 363 (Fig. 1D) at a retention time (RT) of 12.6 min and 324 (Fig. 1E) at a retention time (RT) of 21.4 min. there were.

As shown in Figure 1, the peaks observed at retention time (RT) of 21.4 minutes and retention time (RT) of 12.6 minutes indicate the assimilation of GMP by Bifidobacterium bifidum (NITE BP-02648) strain. It was found that the peak was derived from a product that promotes the growth of Bifidobacterium breve (ATCC 15700T) strain. The anion was detected with a mass-to-charge ratio of 324 or 363 when subjected to MS after the derivatization reaction with 2-AB.

[Manufacturing example]
Production examples of compositions, pharmaceutical compositions, fermented milk, and prepared milk are shown below.

<Manufacture example 1>
Bifidobacterium bifidum is added to 1:3 mL of MRS+GMP medium, and after anaerobic cultivation at 37° C. for 24 hours, the culture solution is concentrated and freeze-dried to obtain a freeze-dried powder. The obtained lyophilized powder, whey protein concentrate (WPC), lactulose, raffinose, and galactooligosaccharide are uniformly mixed to obtain a composition. 20 g of the obtained composition is dissolved in 200 g of water to obtain a composition.

<Manufacture example 2>
Bifidobacterium bifidum was added to 1:3 mL of MRS + GMP medium, and after anaerobic cultivation at 37°C for 24 hours, the culture supernatant from which Bifidobacterium bifidum was removed by filter was concentrated, freeze-dried, and frozen. Obtain a dry powder. This freeze-dried powder, dry powder of milk protein concentrate (MPC480, manufactured by Fonterra, protein content 80% by mass, casein protein: whey protein = approximately 8:2), lactulose, raffinose, and galactooligosaccharides were uniformly mixed. to obtain a composition. 20 g of the obtained composition is dissolved in 200 g of water to obtain a composition.

<Manufacture example 3>
Bifidobacterium bifidum was added to 1:3 mL of MRS + GMP medium, and after anaerobic cultivation at 37°C for 24 hours, the culture supernatant from which Bifidobacterium bifidum was removed by filter was concentrated, freeze-dried, and frozen. Obtain a dry powder. Next, the obtained freeze-dried powder, lactulose, raffinose, galactooligosaccharides, and crystalline cellulose are placed in a stirring granulator and mixed. Thereafter, purified water is added and granulated, and the granulated product is dried to obtain a granulated product (pharmaceutical composition).

<Manufacture example 4>
A method for producing fermented milk to which Bifidobacterium breve and/or Bifidobacterium infantis is added is shown below.
First, milk raw materials, water and other components as required are mixed, preferably homogenized, and heat sterilized. Homogenization treatment and heat sterilization treatment can be performed by conventional methods. A lactic acid bacteria starter is added (inoculated) to a sterilized milk preparation that has been sterilized by heating, and fermentation is carried out while maintaining a predetermined fermentation temperature to obtain a fermented product. Fermentation forms curd.

As the lactic acid bacteria starter, for example, lactic acid bacteria commonly used in yogurt production such as Lactobacillus bulgaricus, Lactococcus lactis, and Streptococcus thermophilus can be used. Once the pH reaches the target value, the formed curd is crushed by stirring and cooled to below 10° C. to obtain a fermented product. By cooling to 10° C. or lower, the activity of lactic acid bacteria can be reduced and acid production can be suppressed.

Next, the fermented product obtained in the fermentation step is heat-treated to obtain a heat-treated fermented product. By heating the fermented product appropriately, acid production by lactic acid bacteria in the fermented product after heat treatment can be suppressed. As a result, it is possible to suppress a decrease in pH during the subsequent production process and/or during storage of concentrated fermented milk containing bifidobacteria, and as a result, the survival of bifidobacteria can be improved.

On the other hand, Bifidobacterium bifidum is added to 1:3 mL of MRS+GMP medium, and after anaerobic culture at 37° C. for 24 hours, a culture supernatant from which Bifidobacterium bifidum is removed by a filter is obtained.

The obtained culture supernatant is added to the heated fermented product obtained in the heat treatment step together with Bifidobacterium breve and/or Bifidobacterium infantis. The amount of Bifidobacterium breve and/or Bifidobacterium infantis added is preferably 1×10 ⁷ to 1×10 ¹¹ CFU/mL, and 1×10 ⁸ to the fermented product after heat treatment. ˜1×10 ¹⁰ CFU/mL is more preferred. If Bifidobacterium breve and/or Bifidobacterium infantis are killed, CFU/mL can be replaced with individual cells/mL.

After adding the culture supernatant, Bifidobacterium breve and/or Bifidobacterium infantis to the fermented product after the heat treatment, concentration is performed. The concentration step can be performed using a known concentration method as appropriate. In the centrifugation method, the whey in the concentrate (heated fermented product to which bifidobacteria and prebiotics have been added) is removed to increase the solid concentration of Bifidobacterium breve and/or bifidobacteria. Concentrated fermented milk of Bacterium infantis is obtained. By ingesting the obtained fermented milk, it is possible to improve intestinal flora.

<Manufacture example 5>
A method for producing formula milk containing Bifidobacterium breve and/or Bifidobacterium infantis is shown below.
Bifidobacterium bifidum is added to 1:3 mL of MRS+GMP medium, and after anaerobic cultivation at 37° C. for 24 hours, the culture solution is concentrated and freeze-dried to obtain a freeze-dried powder. 500 g of the obtained freeze-dried powder, 10 kg of desalted milk whey protein powder (manufactured by Mirai), 6 kg of milk casein powder (manufactured by Fonterra), 48 kg of lactose (manufactured by Mirai), 920 g of mineral mixture (manufactured by Tomita Pharmaceutical), Dissolve 32 g of vitamin mixture (manufactured by Tanabe Pharmaceutical Co., Ltd.), 500 g of lactulose (manufactured by Morinaga Milk Industry Co., Ltd.), 500 g of raffinose (manufactured by Nippon Beet Sugar Co., Ltd.), and 900 g of galactooligosaccharide liquid sugar (manufactured by Yakult Pharmaceutical Co., Ltd.) in 300 kg of warm water, and The mixture was heated and melted at ℃ for 10 minutes, and 28 kg of prepared fat (manufactured by Taiyo Yushi Co., Ltd.) was added and homogenized. Thereafter, sterilization and concentration steps are performed, followed by spray drying to prepare approximately 95 kg of formula milk. To this is added 100 g of bacterial powder of Bifidobacterium breve and/or Bifidobacterium infantis dispersed in starch to prepare about 95 kg of prepared milk. The obtained milk formula is dissolved in water to obtain a milk formula having a total solids concentration of 14% (w/v), which is a standard milk formula concentration. The number of Bifidobacterium breve and/or Bifidobacterium infantis in the obtained milk preparation is 2.7×10 ⁹ CFU/100ml. By administering the formula obtained as described above, it becomes possible to promote the growth of probiotics in the intestines.

By administering the composition, pharmaceutical composition, fermented milk, or prepared milk produced in Production Example 1-5 above to humans, probiotics can utilize GMP and proliferate in the intestine, thereby increasing the intestinal Leads to improvement of inner flora.

Claims (13)

Promoting the growth of bacteria belonging to Bifidobacterium breve and/or bacteria belonging to Bifidobacterium infantis, containing the assimilation product of κ-casein glycomacropeptide by bacteria belonging to Bifidobacterium bifidum. Composition for . The bacteria belonging to Bifidobacterium bifidum include Bifidobacterium bifidum (ATCC29521T), Bifidobacterium bifidum (NITE BP-02429), Bifidobacterium bifidum (NITE BP-01252), and Bifidobacterium bifidum (NITE BP-01252). Bacterium bifidum (NITE BP-02431), Bifidobacterium bifidum (NITE BP-02648), Bifidobacterium bifidum (NITE BP-02432), Bifidobacterium bifidum (NITE BP-02433), The composition according to claim 1, wherein the composition is one or more bacteria selected from. The composition according to claim 1 or 2, further comprising bacteria belonging to Bifidobacterium breve and/or bacteria belonging to Bifidobacterium infantis. The bacteria belonging to Bifidobacterium breve include Bifidobacterium breve (ATCC15700T), Bifidobacterium breve (NITE BP-02460), Bifidobacterium breve (FERM BP-11175), and Bifidobacterium breve (FERM BP-11175). The composition according to claim 3, wherein the composition is one or more bacteria selected from Bacterium breve (NITE BP-02622). The bacteria belonging to Bifidobacterium infantis are one or more bacteria selected from Bifidobacterium infantis (ATCC15697T) and Bifidobacterium infantis (NITE BP-02623). 5. The composition of claim 3 or claim 4, wherein: The above-mentioned assimilation product is purified by a sugar refining solid-phase extraction column, and the resulting purified product is derivatized with 2-aminobenzamide and then subjected to a mass spectrometer. 6. The composition according to any one of claims 1 to 5, comprising a substance detected as a 324 or 363 anion. A growth promoter for bacteria belonging to Bifidobacterium breve and/or Bifidobacterium infantis, which contains a product of assimilation of κ-casein glycomacropeptide by bacteria belonging to Bifidobacterium bifidum. . The growth promoter according to claim 7, further comprising a carbohydrate. The bacteria belonging to Bifidobacterium bifidum include Bifidobacterium bifidum (ATCC29521T), Bifidobacterium bifidum (NITE BP-02429), Bifidobacterium bifidum (NITE BP-01252), and Bifidobacterium bifidum (NITE BP-01252). Bacterium bifidum (NITE BP-02431), Bifidobacterium bifidum (NITE BP-02648), Bifidobacterium bifidum (NITE BP-02432), Bifidobacterium bifidum (NITE BP-02433), The growth promoter according to claim 7 or 8, which is one or more bacteria selected from. A fermentation process comprising fermenting κ-casein glycomacropeptide using bacteria belonging to Bifidobacterium bifidum.
When subjected to a mass spectrometer after derivatization treatment with 2-aminobenzamide, it was detected as an anion with a mass-to-charge ratio of 324 or 363 , indicating that it was a bacterium belonging to Bifidobacterium breve and/or Bifidobacterium. - A method for producing a substance for promoting the growth of bacteria belonging to Infantis . In the fermentation step, 0.5 to 5% of a culture solution of bacteria belonging to Bifidobacterium bifidum is added to a κ-casein glycomacropeptide solution having a concentration of 0.8% to 12%, and fermentation is carried out. 10. The manufacturing method according to 10. A food or drink composition containing the composition according to any one of claims 1 to 6 or the growth promoter according to any one of claims 7 to 9. Formulated milk containing the composition according to any one of claims 1 to 6 or the growth promoter according to any one of claims 7 to 9.