JP6607380B2 - α-defensin secretion promoter and antibacterial agent - Google Patents

Description

The present invention relates to an α-defensin secretion promoter and an antibacterial agent.

In general, antimicrobial peptides are small molecules consisting of about 10-50 amino acid residues that can kill microorganisms such as bacteria, fungi, viruses by passing through cell membranes. There are a wide variety of organisms and types of antibacterial peptides, and hundreds of molecules have been identified from organisms such as plants, insects and mammals.

Defensin is known as a kind of antibacterial peptide. Defensins have a common structure with six cysteines that constitute three intramolecular disulfide bonds. There are roughly two types of defensins, α-defensins and β-defensins. The amount of α-defensin is known to be present in neutrophils and small intestinal crypt panel cells. β-defensins have a longer structure than α-defensins and are present in epithelial cells such as mucous membranes, skin, and trachea. It is known.

α-Defensin is also called kryptin and is an antibacterial peptide having a strong bactericidal ability. α-defensins play an important role in intestinal immunity and exert an infection defense function against bacteria and viruses.

By the way, barley young leaves are naturally derived products with high safety to living bodies. In addition, barley young leaves have been reported to have various pharmacological actions. For example, it is known that the juice components of barley young leaves have an antiviral action and an immune cell enhancing action (Patent Document 1 and 2).

Japanese Patent Laid-Open No. 3-169823 Japanese Patent Laid-Open No. 4-074128

However, Patent Documents 1 and 2 do not describe the relationship between the squeezed component of barley young leaves and α-defensin. In particular, these documents do not describe any antiviral action or immune cell enhancing action on barley young leaf powder. In addition, a substance having an action of increasing the amount of α-defensin in the living body has not been known so far, although it is a naturally derived product having high safety for living bodies such as young barley leaves.

Therefore, the present invention provides an α-defensin secretion promoter and an antibacterial agent comprising a natural-derived product that is highly safe for a living body and has an action of increasing the amount of antibacterial α-defensin in the living body as an active ingredient. It is a problem to be solved by the invention.

In order to solve the above problems, the present inventors have conducted extensive studies on various substances. Surprisingly, when barley leaves and / or stems are ingested, secretion of α-defensin in the intestine is promoted. Furthermore, it has been found that a pathogenic Salmonella bacterium that can be localized in the intestine can be killed by α-defensin whose secretion has been promoted. The inventors have succeeded in creating an α-defensin secretion promoter and an antibacterial agent containing barley leaves, stems and / or processed products thereof as active ingredients based on the obtained knowledge. . The present invention has been completed based on such knowledge and success.

Therefore, according to the present invention, there is provided an α-defensin secretion promoter containing barley leaves, stems and / or processed products thereof.

Preferably, in the present invention, the α-defensin secretion promoter is a gastrointestinal α-defensin secretion promoter.

Preferably, in the present invention, the α-defensin secretion promoter is a small intestinal α-defensin secretion promoter.

According to another aspect of the present invention, an antibacterial agent containing barley leaves, stems and / or processed products thereof is provided.

Preferably, in the present invention, the antibacterial agent is an antibacterial agent against bacteria localized in the digestive tract.

Preferably, in the present invention, the antibacterial agent is an antibacterial agent against Salmonella bacteria.

According to the present invention, by containing processed products such as barley foliage and powder thereof, it is possible to promote secretion of α-defensin in vivo and kill pathogenic bacteria in vivo. A defensin secretion promoter and an antibacterial agent are obtained. In addition, the α-defensin secretion promoter and antibacterial agent of the present invention characterized by containing barley foliage and processed products thereof as active ingredients are highly safe.

The α-defensin secretion promoter and antibacterial agent of the present invention are related to gastrointestinal tissue abnormalities such as gastrointestinal immunity deficiency, generation of harmful microorganism metabolites and harmful enzymes in the gastrointestinal tract by α-defensin secretion promoting action and antibacterial action. It can be expected to improve, alleviate, recover, treat and prevent various diseases and abnormalities.

1 is a graph showing the measurement results of the amount of α-defensins in feces of the control group, cellulose group and barley young leaf group described in Example 1. FIG. The numerical value represents an average value. FIG. 2 is a graph showing the measurement results of antibacterial activity of PBS, barley young leaf powder and rCrp4 described in Example 2 against defensin-sensitive strains of Salmonella typhi. The numerical value represents an average value. FIG. 3 is a diagram showing the measurement results of the amount of α-defensin in PBS and barley young leaf powder described in Example 3. The numerical value represents an average value.

Details of the present invention will be described below.
[1. α-Defensin Secretion Promoter]
The α-defensin secretion promoter of the present invention is an active ingredient having an α-defensin secretion-promoting action. Or one or more of them, that is, barley leaves, stems and / or processed products thereof. In the present specification, barley leaves and / or stems may be collectively referred to as stems and leaves. The α-defensin secretion promoter of the present invention may contain the above-mentioned active ingredient, and may contain other parts of barley such as barley root in addition to these.

The α-defensin secretion promoting action means an action to increase the abundance of endogenous α-defensin in the living body, and the mechanism is not particularly limited. For example, it has the effect of promoting α-defensin secretion from small intestinal crypt panel cells. Therefore, the α-defensin secretion promoter of the present invention is preferably an α-defensin secretion promoter for the digestive tract, and more preferably an α-defensin secretion promoter for the small intestine. In addition, the effect of increasing the amount of α defensin in feces accompanying the secretion of α defensin in vivo, the effect of maintaining the increased amount of α defensin in feces, and the effect of suppressing the decrease in the amount of increased α defensin in feces It is included in the secretagogue action.

Barley ( Hordeum vulgare L. ) is native to Central Asia and is an annual or perennial herb belonging to the family Gramineae, and is broadly classified into Nijo barley, Rojo barley and the like, depending on the panicle shape. Barley used in the present invention is not particularly limited, and may be any of the above-mentioned varieties, wild species, hybrid species, and the like.

The barley foliage may be harvested at any harvest time, and is not particularly limited. For example, it is preferably harvested before the maturity period, that is, from the beginning of splitting to the early stage of heading.

Barley foliage is preferably processed immediately after harvesting. When time is required for the treatment, it is preferably stored by storage means commonly used by those skilled in the art, such as low-temperature storage, in order to prevent deterioration of barley foliage.

The processed barley foliage is not particularly limited as long as it is obtained by subjecting the harvested barley foliage to one or more processing. Processed products of barley foliage include, for example, dry powder of barley foliage, crushed barley foliage and dry powder thereof (hereinafter, the dry powder of the pulverized product is also referred to as “ground powder”), barley foliage Shredded product and dried powder thereof (hereinafter, the dried powder of the shredded product is also referred to as “striped powder”), barley foliage and dried powder (hereinafter referred to as dried powder of juice) Are also referred to as “squeezed powder”), barley stalk and leaf extract and dried powder thereof (hereinafter, the extract dry powder is also referred to as “extract powder”), and the like, but is not limited thereto. However, it is preferably a dry powder of barley foliage from the viewpoint of ease of processing, storage, transportation, etc. and versatility of usage forms. Examples of dry powder of barley foliage include dry powder of barley foliage, crushed powder of barley foliage, fragmented powder of barley foliage, squeezed powder of barley foliage, extract powder of barley foliage . In addition, in this invention, powder shall include the thing granulated or tableted by adding an excipient | filler etc. as needed.

For example, barley stalks and leaves can be made into dry powder by combining a drying treatment and a pulverization treatment. At this time, the drying process and the pulverization process may be performed simultaneously, or either may be performed first, but it is preferable to perform the pulverization process after the drying process is performed first. When barley stalks and leaves are made into dry powder, if necessary, one or more treatments selected from treatments such as blanching treatment and sterilization treatment may be combined. Moreover, the frequency | count of performing a grinding | pulverization process is not specifically limited, You may implement as a process of 1 time or 2 times or more like performing the fine grinding process which grind | pulverizes more finely after performing a rough grinding process.

The blanching treatment is a treatment for keeping the barley stalks and leaves green vividly. Examples of the blanching treatment include hot water treatment and steaming treatment.

Examples of the hot water treatment include a method of treating barley foliage in hot water or steam at 70 to 100 ° C., preferably 80 to 100 ° C. for 60 to 240 seconds, preferably 90 to 180 seconds. Further, in the hot water treatment, it is preferable to use a carbonate such as magnesium carbonate or a hydrogen carbonate such as sodium hydrogen carbonate. By dissolving the hydrogen carbonate salt in hot water, the green color of the barley foliage is more vivid. can do.

The steaming treatment is preferably an intermittent steaming treatment in which barley stems and leaves are steamed and steamed under normal pressure or pressure. In the intermittent steaming process, the steaming process is performed, for example, for 20 to 40 seconds, preferably 30 seconds. The cooling treatment after the steaming treatment is preferably performed immediately, and the method is not particularly limited, but it is immersed in cold water, refrigerated, cooling with cold air, evaporative cooling with hot air, evaporative cooling combining hot air and cold air Etc. are used. Among these, evaporative cooling combining hot air and cold air is preferable. Such cooling treatment is performed such that the barley foliage has an article temperature of preferably 60 ° C. or lower, more preferably 50 ° C. or lower, and even more preferably 40 ° C. or lower. In order to produce barley stalk and leaf powder rich in nutrients such as vitamins, minerals, and chlorophyll, it is preferable to repeat the intermittent steaming treatment 2 to 5 times.

The sterilization treatment is not particularly limited as long as it is a sterilization treatment generally known to those skilled in the art. For example, it is a treatment that physically or chemically kills microorganisms using temperature, pressure, electromagnetic waves, chemicals, and the like. Can do.

When the branching process is performed in addition to the drying process and the pulverization process, the branching process is preferably performed before the drying process. Moreover, when performing a sterilization process in addition to the drying process and the pulverization process, the sterilization process is preferably performed after the drying process or before or after the pulverization process.

Although a drying process is not specifically limited, For example, the process dried so that the moisture content of barley foliage may be 10 wt% or less, Preferably it is 5 wt% or less. The drying treatment is performed by any method known to those skilled in the art, such as a method of drying in the sun, hot air drying using a dryer, high pressure steam drying, spray drying, vacuum drying, fluidized drying, electromagnetic wave drying, freeze drying, and the like. Can be done. Drying by heating can be performed, for example, at a temperature and a time at which barley foliage is not discolored by heating at 40 to 140 ° C., preferably 80 to 130 ° C.

The pulverization process is not particularly limited, and examples thereof include a process of pulverizing a plant body by an arbitrary method commonly used by those skilled in the art using a crusher, a mill, a blender, a millstone, or the like. The crushed barley foliage is sieved as necessary. For example, it is preferable to use a barley foliage powder that passes through 30 to 250 mesh. By making the particle size below 250 mesh, the barley foliage powder becomes easier to handle during further processing, and by making the particle size above 30 mesh, the barley foliage powder and other materials Uniform mixing with is easy.

In the pulverization process, in the coarse pulverization step, barley stalks and leaves using any coarse pulverization device or instrument commonly used by those skilled in the art, such as a cutter, slicer, dicer, etc., for example, the major diameter of barley shoots is about 20 mm or less, Preferably, it is crushed to be about 0.1 to 10 mm. In the fine pulverization step, barley foliage is finely pulverized so that 90 wt% or more passes through the 200 mesh section using any fine-powder-equipment equipment or tools commonly used by those skilled in the art, such as crushers, mills, blenders, and stone mills. It is preferable.

As a specific method for producing a pulverized powder, for example, after cutting barley foliage, it is subjected to a blanching treatment, and then dried so that the water content is 10% by mass or less, preferably 5% by mass or less. The method of grind | pulverizing is mentioned (refer Unexamined-Japanese-Patent No. 2004-000210). In addition to this, for example, after cutting barley stalks and leaves, a branching treatment is performed, followed by twisting, and then drying and pulverization (see Japanese Patent Application Laid-Open No. 2002-0665204 and Japanese Patent No. 3428956). A method in which barley foliage is dried and coarsely pulverized, heated at 110 ° C. or higher, and further finely pulverized (see Japanese Patent Application Laid-Open Nos. 2003-033151 and 3277181).

Although the method of fragmenting barley stalks and leaves is not particularly limited, for example, methods commonly used by those skilled in the art, such as slicing, crushing, and chopping, to fragment a plant can be used. As an example of fragmentation, a slurry may be used. Slurry is performed by putting barley stalks and leaves in a mixer, juicer, blender, mascolloider, etc., and forming barley stalks and leaves in a crumpled shape (suspension of liquid and solid). By slurrying in this way, 80% by mass or more of the foliage of the foliage is preferably 1 mm or less, more preferably 0.5 mm or less, even more preferably 0.1 mm or less, and still more preferably 0.8% or less. It is fragmented to be 05 mm or less, and has fluidity.

As a method for drying barley stalks and leaves, a method commonly used by those skilled in the art may be used, and a conventionally known method may be used. For example, the above-mentioned barley stalks and leaves can be dried.

The method of squeezing barley foliage is not particularly limited, and examples thereof include a method of squeezing barley foliage or a fragmented product thereof, and a method of centrifuging or filtering a barley foliage fragmented product. As a specific example of the squeezing method, the squeezed juice is obtained by squeezing with a mechanical crushing means such as a mixer or a juicer, and if necessary, removing the crude solid content by means such as sieving or filtration. A method is mentioned. More specifically, the methods described in JP-A-08-245408, JP-A-09-047252, JP-A-5-7471, JP-A-4-341153 and the like can be mentioned. A person skilled in the art can appropriately select and carry out the method. In addition, you may concentrate a squeezed product as needed.

As a method for dry-pulverizing the barley foliage, the juice may be dry-powdered by a method commonly used by those skilled in the art, and a conventionally known method may be used. For example, the juice may be dried and powdered as it is, or an appropriate binder or excipient may be added to the juice and dried to be powdered. Moreover, an excipient | filler etc. may be added as needed to the squeezed product or the dried powder of the squeezed product, and it may shape | mold and granulate by the granulation method of well-known wet and dry methods.

As a method for extracting barley stalks and leaves, a method commonly used by those skilled in the art may be used, and a conventionally known method may be used. For example, a method of adding an extraction solvent usually used by those skilled in the art, such as ethanol, water, hydrous ethanol, etc. to barley foliage or a fragmented product thereof, and heating and extracting as necessary. In addition, you may concentrate an extract as needed. In order to increase the concentration of the specific component, the extract may be fractionated by a method commonly used by those skilled in the art, or the specific component may be purified by a method commonly used by those skilled in the art. Moreover, you may use what dried the extraction residue which remains after extracting from barley foliage as a powder of barley foliage.

As a method for dry-pulverizing the barley foliage extract, the extract may be dry-powdered by a method commonly used by those skilled in the art, and a conventionally known method may be used. For example, the extract may be dried to be powdered, or an appropriate binder or excipient may be added to the extract and then dried to be powdered. Moreover, an excipient | filler etc. may be added to an extract or the dry powder of an extract as needed, and it may shape | mold and pulverize by a granulation method of well-known wet and dry methods.

Of the processed barley foliage, it is particularly preferable to use a dried powder of barley foliage because the α-defensin secretion promoter and antibacterial agent of the present invention can be made more excellent in appearance. The characteristics of the dry powder of barley foliage are not particularly limited. For example, the angle of repose is preferably 20 ° to 80 °, more preferably 30 ° to 70 °, still more preferably 40 ° to 60 °, and more preferably 45 ° to Even more preferred is 55 °. The angle of repose is measured using an angle of repose measuring instrument (As One, ASK-01) by dropping about 50 g of sample into the center of a petri dish having an altitude of 12 cm to a radius of 4.3 cm, and then depositing the sample in a mountain shape. After measuring the height, the angle of repose can be calculated by the following formula from the radius of the petri dish and the height of the deposited sample.
Angle of repose = tan ⁻¹ (b / a) × 180 ÷ π (where a = the radius of the petri dish, b = the height of the deposited sample)

When barley foliage is a pulverized powder, its characteristics are not particularly limited. For example, the median diameter is 16 μm or more, the aspect ratio is 2.0 or less, and the median diameter is 20 μm or more. preferable. By using the pulverized powder of barley stalks and leaves having a specific particle size and aspect ratio combination, the palatability can be improved.

Further, as another characteristic of the barley stalks and leaves, for example, the median diameter is less than 20 μm, the distribution constant n by the rosin-Rammler equation is 1.6 or more, and the median diameter is less than 16 μm. Is preferred. By using a pulverized powder of barley stalks and leaves having a specific particle size and a distribution constant n according to the Rosin-Rammler equation, the palatability can be improved.

Here, the distribution constant n by the Rosin-Rammler equation is a constant n in the Rosin-Ramler equation relating to the particle size distribution, and is a value representing the uniformity of the particle size.

The Rosin-Rammler formula is represented by the following formula (1).
R = 100exp (−βDn) (1)
In the formula, D represents a particle size, R represents a percentage (%) of particles larger than D (particle size) to all particles, β represents a particle size characteristic coefficient, and n represents a distribution constant.

Here, assuming that β = 1 / Den, the above formula (1) is R = 100exp {− (D / De) n} (2)
Can be rewritten as De is a particle size characteristic coefficient.

Furthermore, when the above equation (2) is rewritten, the following equation (3) is obtained.
log {log (100 / R)} = nlogD + C (3)
However, C = log (log) -nlogDe.

From the above equation (3), when the relationship is plotted on a Rosin-Rammler (RR) diagram with log D on the x-axis and log {log (100 / R)} on the y-axis, it becomes a substantially straight line. The slope of the straight line (distribution constant n) represents the degree of uniformity of particle size. The larger the value of distribution constant n, the narrower the particle size range and the uniform particle size. It can be evaluated as excellent.

The dry powder of barley stover used in the present invention may contain water-insoluble dietary fiber. The water-insoluble dietary fiber contained in the dry powder is contained in an amount of 20% by mass or more, preferably 30% by mass or more, in terms of dry mass, preferably 20 to 80% by mass, preferably 30 to 70% by mass, more preferably 35%. It is more preferable to contain -60 mass%.

The α-defensin secretion promoter of the present invention can exhibit an α-defensin secretion promoting action by containing the barley foliage and processed product thereof. The α-defensin secretion-promoting action of the α-defensin secretion-promoting agent of the present invention is, for example, in the first week from the start of the test when the amount of α-defensin in feces measured by the method described in the examples described later is used as an index. The effect of maintaining or increasing the amount of fecal α-defensin in the second week, preferably the amount of fecal α-defensin in the first week from the start of the test is 150% or more, more preferably 200% or more, More preferably, it is a certain effect at 300% or more; or, preferably, the fecal α-defensin level is 200% or more, more preferably 300% or more, more preferably 400% or more, compared to the control in the second week from the start of the test. It is a certain effect.

The α-defensin secretion promoter of the present invention can be used in various forms for the purpose of obtaining an α-defensin secretion promoting action. For example, it may be used for various purposes without adding special processing. For example, it is used as a pharmaceutical, a quasi-drug, etc. or blended with these.

In the solid content of the α-defensin secretion promoter of the present invention, the content of barley foliage is dry mass, and the lower limit is preferably 0.1% by mass or more, more preferably 0.5% by mass or more. More preferably 10% by weight or more, still more preferably 20% by weight or more, and the upper limit is preferably 99.9% by weight or less, more preferably 90% by weight or less, and 80% by weight or less. Is more preferable. When the barley foliage content is less than 0.1% by mass, the α-defensin secretion promoting action may not be sufficiently exhibited.

The daily use amount of the α-defensin secretion promoter of the present invention is not particularly limited, and can be appropriately set according to the use mode, the use contents of the user, the intestinal flora, etc. For example, the mass of barley foliage In conversion, it is 1 to 6000 mg / kg, preferably 1 to 4000 mg / kg, more preferably 10 to 3000 mg / kg, still more preferably 10 to 2000 mg / kg, based on the weight of the user. . Similarly, the single use amount of the α-defensin secretion promoter of the present invention is not particularly limited, and is, for example, 0.5 to 3000 mg / kg based on the weight of the user in terms of mass of barley foliage. , Preferably it is 1-2000 mg / kg, More preferably, it is 5-1000 mg / kg.

Moreover, the usage-amount of the alpha defensin secretion promoter of this invention is 0.01-30 g in conversion of the mass of barley foliage, for example, Preferably it is 0.05-20 g, More preferably, it is 0.1-10 g. More preferably, it can be 0.3-7g. Similarly, the daily use amount of the α-defensin secretion promoter of the present invention is, for example, 0.01-100 g, preferably 0.05-70 g, more preferably 0.5- 50 g, more preferably 1 to 30 g.

The α-defensin secretion promoter of the present invention can be used in various forms for the purpose of obtaining an α-defensin secretion promoting action. The α-defensin secretion promoter of the present invention can be, for example, an oral or parenteral α-defensin secretion promoter. When the α-defensin secretion promoter of the present invention is used for parenteral use, the dosage form is not particularly limited, and examples thereof include external preparations and cosmetics used by applying to the skin, scalp, hair and the like. When the α-defensin secretion promoter of the present invention is used for oral use, the dosage form is not particularly limited. For example, pharmaceuticals; quasi-drugs; foods capable of displaying functionalities other than general foods (for example, functionally-labeled foods) Insurance functional foods such as foods for specified health use). Similar to drugs and quasi-drugs, foods can be broadly classified into foods that can be displayed with functionality and foods that cannot display with functionality. The α-defensin secretion promoter of the present invention is preferably used as a food capable of displaying a function or is blended in a food capable of displaying a function.

The form of the α-defensin secretion promoter of the present invention is not particularly limited and can be any form. As the form of the oral α-defensin secretion promoter, for example, a form suitable for oral use, specifically, powder, granule, granule, tablet, rod, plate, block, solid , Round shape, liquid shape, candy shape, paste shape, cream shape, capsule shape such as hard capsule and soft capsule, caplet shape, gel shape, chewable shape, syrup shape and the like.

The packaging form of the α-defensin secretion promoter of the present invention is not particularly limited, and can be appropriately selected depending on the dosage form, for example, blister pack such as PTP; strip packaging; heat seal; aluminum pouch; Film packaging using glass; glass containers such as vials; plastic containers such as ampoules.

The α-defensin secretion promoter of the present invention is in a powder form (powder, granule, etc.) and is a form in which a mixture mixed with water is used orally to prevent spoilage and be suitable for long-term storage. To preferred. In addition, when the α-defensin secretion promoter of the present invention is in a solid form such as powder or tablet, it can be mixed with water to form a liquid and can be used orally as described above. Depending on the user's preference, it may be used orally as a solid.

The α-defensin secretion promoter of the present invention may contain only barley foliage powder or may contain other components in addition to barley foliage powder. Examples of other components include various excipients, binders, lubricants, stabilizers, diluents, extenders, thickeners, emulsifiers, colorants, fragrances, additives, and the like. The content of other components can be appropriately selected depending on the usage form of the α-defensin secretion promoter of the present invention.

It is preferable that the other component blended in the α-defensin secretion promoter of the present invention is an ingredient that exhibits an α-defensin secretion promoting effect in addition to or synergistically with the action of barley foliage.

The target of use of the α-defensin secretion promoter of the present invention is not particularly limited, and examples include humans such as infants, infants, children, boys, and adults, and mammals such as cats, dogs, cows, and horses. Also good. However, it is preferable that the α-defensin secretion promoter of the present invention is used for a use subject who benefits from the physiological activity of α-defensin by having an α-defensin secretion promoting action. Specifically, the α-defensin secretion enhancer of the present invention is effective for patients with and at risk of gastrointestinal tract immunodeficiency, gastrointestinal tissue abnormalities such as generation of harmful microbial metabolites and harmful enzymes in the gastrointestinal tract. It is preferable to use it. Furthermore, it is more preferable that the α-defensin secretion promoter of the present invention is used for middle-aged and elderly people and those who lack exercise.

Improvement, mitigation, recovery, treatment, and prevention when using the present invention are used in the meaning commonly known to those skilled in the art, for example, onset of abnormalities, diseases, and symptoms in an individual. Prevention and delay; reduction of the risk of developing abnormalities, diseases and symptoms; improvement of abnormalities, diseases, symptoms and conditions; prevention and delay of deterioration of abnormalities, diseases, symptoms and conditions; abnormalities, diseases and symptoms Includes reversal of progress, prevention and delay.

As another aspect, the α-defensin secretion promoter of the present invention can be used as an antibacterial agent, a gastrointestinal immunity deficiency improving agent, a small intestine immune activator, etc., which will be described later, and further used for producing these agents. be able to.

The usage mode of the α-defensin secretion promoter of the present invention is not particularly limited. For example, when the α-defensin secretion promoter of the present invention is used by being administered to humans or animals, it may be a therapeutic use or a non-therapeutic use. Non-therapeutic use is use that does not include so-called medical practice, that is, use that does not include a method for operating, treating, or diagnosing a human.

[2. Antibacterial agent]
The antibacterial agent of the present invention is characterized by containing barley leaves, stems and / or processed products thereof as an active ingredient having an antibacterial action, like the α-defensin secretion promoter of the present invention.

In the present invention, the antibacterial action is an action of selectively attenuating specific microorganisms living in the digestive tract such as the small intestine in a living body, or selectively sterilizing or sterilizing the microorganisms (referred to as killing them together). The mechanism is not particularly limited. For example, the effect | action which reduces the number of pathogenic bacteria, such as Salmonella genus bacteria, Campylobacter genus bacteria, colon_bacillus | E._coli in the intestine, is mentioned. Therefore, the antibacterial agent of the present invention is preferably an antibacterial agent against bacteria localized in the digestive tract such as the small intestine, and more preferably an antibacterial agent against Salmonella bacteria.

The antibacterial action exhibited by the antibacterial agent of the present invention is, for example, 1/0 compared to the PBS group when the number of viable bacteria of a highly sensitive strain of Salmonella typhi determined by the method described in Examples below is used as an index. The effect is such that the number of viable bacteria is 10, preferably 1/100.

The antibacterial agent of the present invention may contain only barley foliage powder, or may contain other components in addition to barley foliage powder. Examples of other components include various excipients, binders, lubricants, stabilizers, diluents, extenders, thickeners, emulsifiers, colorants, fragrances, additives, and the like. The content of other components can be appropriately selected according to the use form of the antibacterial agent of the present invention.

It is preferable that the other component mix | blended with the antibacterial agent of this invention is a component which shows an antibacterial effect additively or synergistically with the effect | action which barley foliage has, or auxiliary. Examples thereof include useful microorganisms such as lactic acid bacteria, substances that increase or activate useful microorganisms such as lactic acid bacteria, and other antibacterial components.

As long as the antibacterial agent of the present invention has an antibacterial action, other items correspond appropriately [1. [alpha defensin secretion promoter] can be referred to.

Hereinafter, the present invention will be described in more detail with reference to examples. However, the present invention is not limited to these examples, and the present invention can take various modes as long as the problems of the present invention can be solved. .

It was evaluated as follows that barley stalks and leaves show an α-defensin secretion promoting action and an antibacterial activity.

[Example 1. Promotion of α-defensin secretion by milled powder of barley stover]
1. As a test substance example, barley young leaf powder (manufactured by Toyo Shinyaku Co., Ltd.), which is a pulverized powder of barley stalks and leaves, was used. Moreover, in order to verify the difference in absorbability depending on the type of dietary fiber, a white powder of cellulose (manufactured by Oriental Yeast Co., Ltd.) was used as a comparative example.

2. Preparation of test feed As shown in Table 1 below, each ingredient constituting a non-fiber high-fat diet (NFHF) based on young barley powder and AIN-76 standard feed is fed using a pestle and mortar. It mixed and prepared so that each component in it might become uniform, and the barley young leaf powder containing NFHF powder feed was obtained. Moreover, the cellulose containing NFHF powder feed was obtained by mix | blending a cellulose. Furthermore, an NFHF powder feed to which no test substance was added was used as a control. The barley young leaf powder-containing NFHF powder feed and the cellulose-containing NFHF powder feed were prepared so that the total dietary fiber content was 5 wt%. The numerical values in Table 1 indicate wt%.

3. Test animals 4-week-old male ICR mice (Nippon Charles River Co., Ltd.) were acclimated for 5 days or more. As a rearing environment, the lighting time is 12 hours, and for 6 days from the start of habituation, the cage is a flat bottom cage made of polycarbonate (W220 × D320 × H135mm; Toyo) with a wood chip (soft chip; Nippon SLC Co., Ltd.) as the floor. Riko Co., Ltd.) was used for the next 9 days, and a stainless steel 5-series cage (W150 × D210 × H150 mm; Tokiwa Corporation) was used. The number of accommodated individuals was 2 to 3 per cage in a flat bottom cage, and one per compartment in a 5-cage cage. The five-cage cage was used for stool collection.

During the habituation period, the feeding method was generally free intake. The feed for 6 days from the start of habituation was MF solid feed (Oriental Yeast Co., Ltd.), then the normal feed of Table 2 below was used for 9 days, and tap water was used for drinking water.

After the habituation period, healthy test animals were divided into groups so that their body weights were almost uniform, and were subjected to the test at 6 weeks of age. However, no difference was observed between the groups in the weight of wet feces collected on the fourth to −1 day from the start of the test.

4). For test animals of 6 weeks of age grouped after the habituation period of the test procedure, each test feed (barley leaf powder-containing NFHF powder feed, cellulose-containing NFHF powder feed and NFHF powder feed) from the test start date (day 0) Ad libitum for 16 days. As a breeding environment, the lighting time was 12 hours, and a stainless steel 5-unit cage was used. The number of individuals accommodated was one per division. As the feeder, Roden Cafe (Oriental Yeast Co., Ltd.) was used in order to make it easy to measure the amount of food intake. Table 3 summarizes the group composition.

During the test period, body weight and food intake were measured once a week. From the start of the test to the 4th day to the -1 day (week 0), from the 6th to 9th day (1st week) and from the 13th to 16th day (2nd week) Feces were collected. The collected stool was placed on a plastic tray, dried overnight at room temperature, and stored frozen at −30 ° C. until the end of the test.

5. Measurement of α-defensin secretion amount The amount of α-defensin (cryptptin-4) in feces was measured using the feces collected in “4. Test procedure” above. The details of the test method were carried out according to the method described in the section “Sandwich ELISA procedure” of Nakamura et al. (Analytical Biochemistry, 443 (2013) 124-131). The recovered stool was prepared as follows. That is, stool was pulverized with a pulverizer (Beads Crusher μT-12; manufactured by Taitec Co., Ltd.), and 100 μl of PBS (−) was added to 10 mg of stool and suspended with a vortex mixer. The obtained suspension was centrifuged at 4 ° C., 20,400 × g for 10 minutes. The supernatant obtained by centrifugation was collected and used as an ELISA measurement sample.

6). Results The results of measurement of the amount of alpha defensin in feces measured using feces at 0 weeks, 1 week and 2 weeks after the start of the test are shown in FIG.

The amount of α-defensin in feces on the first week of the test was higher in the cellulose group and the barley young leaf group than in the control group. Surprisingly, it was found that the amount of α-defensin in the stool was significantly increased in the barley young leaf group as compared with the cellulose group.

The fecal α-defensin amount in the second week after the start of the test also had a similar tendency. Surprisingly, however, the amount of alpha defensin in the feces decreased from the first week of the test in the control group and the cellulose group, whereas the amount of alpha defensin in the feces was further increased from the first week of the test in the barley young leaf group. increased. This result shows that in the barley young leaf group, the decrease in the amount of α-defensin in feces was suppressed from the first week to the second week of the test, and rather the increase or maintenance of the amount of fecal α-defensins was induced.

From these results, the barley young leaf powder has a markedly good α-defensin secretion promoting effect on cellulose powder with the same dietary fiber content, and is not obtained with cellulose powder. It was clarified that it exhibits an inhibitory effect on α-defensin secretion decrease and an α-defensin secretion promoting and maintaining effect.

[Example 2. Measurement of antibacterial activity by panel cell secretion supplemented with young barley powder]
The antibacterial activity was measured by the panel cell secretion supplemented with barley young leaf powder.
1. As a test substance example, barley young leaf powder (manufactured by Toyo Shinyaku Co., Ltd.), which is a pulverized powder of barley stalks and leaves, was used.

2. Test Method Details of the test method were performed according to the method described in Japanese Patent No. 5088911. Male ICR mice (Nippon Charles River Co., Ltd.) were dissected under anesthesia, and the small intestine tissue was collected. Subsequently, a suspension rich in crypt (crypt suspension) was prepared. A panel cell group suspension containing live panel cells in PBS (-) was prepared from this crypto suspension. Barley young leaf powder (final concentration 100 μg / mL) was added to 100 crypts, followed by incubation at 37 ° C. for 30 minutes. In addition, the thing which does not add barley young leaf powder was set as control (PBS). After culturing, the supernatant was collected, and after acetic acid was added to the collected supernatant to 30% (v / v), acetic acid extraction was performed by stirring for 30 minutes at room temperature. Obtained 5 mL of acetic acid extract. Next, a 0.1% (v / v) acetic acid aqueous solution was used as an external solution and dialyzed with a membrane filter to recover about 1.5 mL of the internal solution. Then, it freeze-dried with the freeze dryer (FreeZone; Labconco), and the freeze-dried material of each sample was obtained. Each lyophilized product was dissolved in 1 mL of sterilized distilled water to obtain a panel cell cell secretion aqueous solution.

3. Measurement of antibacterial activity 10 μL each of the two aqueous solution of panetocyte secretions (barley young leaf powder, PBS) and reduced kryptin (rCrp4) (final concentration 5 μg / mL), and Salmonella serovar Typhimurium phoP- ) Was added to 40 μL of Tryptic Soy Broth (TSB) medium containing 1 × 10 ⁶ CFU and incubated at 37 ° C. for 60 minutes. The cultured Salmonella-containing solution was applied onto a nutrient agar plate prepared using Triptic soy agar (Difco), cultured at 37 ° C. overnight, and then the number of viable bacteria was measured.

4). Results The results of measuring the antibacterial activity of the test barley young leaf powder, PBS, and reduced kryptin (rCrp4) against the test bacteria are shown in FIG.

As shown in FIG. 2, it was confirmed that the panet cell secretion added with the barley young leaf powder showed excellent antibacterial activity against the test bacteria. The antibacterial activity when rCrp4 was used was below the detection limit. From this result, it was clarified that barley stem powder promotes the secretion of antibacterial substances from the panel cells.

[Example 3. Promotion of α-defensin secretion by milled powder of barley stover]
The panetocyte secretion aqueous solution obtained in “2. Test method” of Example 2 was collected, and the amount of α-defensin secretion was measured. The measurement method was performed in accordance with “5. Measurement of α-defensin secretion amount” in Example 1. Note that the panelet cell secretion aqueous solution was directly used as an ELISA measurement sample. The results are shown in FIG.

As shown in FIG. 3, it was confirmed that the barley young leaf powder showed an excellent α-defensin secretion promoting action. Moreover, it was suggested that the secretion mechanism promotes the secretion of α-defensin by acting directly on the panel cells.

Since the α-defensin secretion promoter and antibacterial agent of the present invention contain natural products that are highly safe and have an action that increases the amount of antibacterial α-defensin in the living body, Expected to be used to improve, alleviate, recover, treat and prevent various diseases and abnormalities related to gastrointestinal tissue abnormalities such as gastrointestinal immune deficiency, generation of harmful microorganism metabolites and harmful enzymes in the gastrointestinal tract It can contribute to the health and welfare of those suffering from these diseases and abnormalities.

Claims (3)

An α-defensin secretion promoter containing barley leaves, stems and / or pulverized powder thereof. The α-defensin secretion promoter according to claim 1, wherein the α-defensin secretion promoter is a gastrointestinal α-defensin secretion promoter. The α-defensin secretion promoter according to claim 1, wherein the α-defensin secretion promoter is a small intestinal α-defensin secretion promoter.