JP2018095594A - Immune function improving composition - Google Patents

Abstract

PROBLEM TO BE SOLVED: To provide immune function improving compositions based on the immune function improving action of Lauraceae, Lindera umbellata Thunb. extract by examining the pharmacological action of the extract.SOLUTION: Provided is an immune function improving composition characterized by containing as an active ingredient Lauraceae Lindera umbellata Thunb. extract extracted with a solvent, particularly water, lower alcohol, or a mixed solvent of a lower alcohol and water, the composition being useful for the preventive improvement and/or therapeutic improvement of immune function in a systemic immune system and/or gastrointestinal immune system caused by psychological stress.SELECTED DRAWING: Figure 1

Description

  The present invention relates to a composition for improving immune function, which comprises a solvent extract of a plant, in particular, an extract of Camphoraceae Chromoji.

Herb, which is a herbal medicine that dried the stem branch and root bark of the camphor tree, the camphor tree, is also referred to as a fishing rod. Application to mainly gastrointestinal symptoms such as diarrhea, etc., nausea, abdominal pain, abdominal tension, stagnation, etc. is described (Non-patent Document 1).
Research on the pharmacological action of sputum has been conducted for a long time. For example, antispasmodic action (Non-patent document 2), sedative action (Non-patent document 3), anti-ulcer action (Non-patent documents 4 and 5), or melanin synthesis Inhibitory action (Non-Patent Document 6) has been reported.

  In addition, proposals have been made in which an extract of black moji itself is applied to various uses. For example, anti-influenza virus agents containing the extract as an active ingredient, compositions containing the same, foods and drinks (Patent Document 1), blood pressure lowering Applications such as an agent (Patent Document 2), a blood sugar-lowering beverage (Patent Document 3), a protease inhibitor (Patent Document 4), and a skin external preparation (Patent Document 5) have been proposed.

Since the black moji is a camphor family, the leaves and bark are aromatic, and because of its specific aroma and bactericidal power, high-grade toothpicks that are added to Japanese sweets are made, and the black moji tree and leaves are distilled. Chromoji oil obtained in the past has been actively used as a fragrance.
Therefore, there are many reports on the essential oil component of black moji, and it has been reported as an essential oil component represented by linalool, geraniol and the like (Non-Patent Documents 7 to 9).
On the other hand, there are few reports regarding non-essential oil components, and only a few types of tannins, alkaloids, flavonoids, etc. have been reported (Non-Patent Documents 5 and 10).

In general, the plants collectively referred to as camphoraceae are: Linder umbellata Thunb., L. umbellata var. Membranacea (Maxim.) Momiyama, L. umbellata var. Lancea Momiyama ), Kemo moji (L. sericea (Sieb.et zucc.) Blume) and Usge black moji (L. sericea var. Glabrata Blume), which are deciduous shrubs widely distributed in the mountains from southern Hokkaido to Kyushu. It is common to use the trunk branches and root bark of the blackwood of the camphor family of blackwood as the moth. It may be used as
In addition, it is a plant different from the avian medicine (Uyaku) obtained by drying the roots of tendered yam (Lindera strychnifolia) (Non-patent Documents 11 and 12).

The inventor of the present invention has newly found that an extract of Chromomozi has an excellent immune function improving effect as a result of repeated studies on Chromomozi.
Kuromoji is a raw material for herbal medicines as cocoon, and its toxicity is extremely low (Non-patent Document 3), and it is also described as “non-toxic” in the Japanese-Wood Wooden Book (Non-Patent Document 1).
In addition, due to its excellent fragrance, it has been widely applied to food as a fragrance, and is extremely safe and can be eaten safely and safely on a daily basis.
Therefore, the extract of black moji can provide highly safe functional foods, pharmaceuticals, animal feeds, and feeds for fish farming as a material for the composition for improving immune function.

JP 2004-059463 A JP 2007-051120 A JP 2002-199864 A JP 2001-122728 A JP 07-277951 A

Japanese and Chinese medicine: 424-426, 1970, Ishiyaku Publishers Biopharmaceutical journal: 36 (2), 134-138 (1982) Biopharmaceutical journal: 36 (2), 139-144 (1982) Biopharmaceutical journal: 36 (4), 350-355 (1982) Planta Medica: 1, 34-38 (1985) Chem. Pharm. Bull.,: 43 (5), 893-895 (1995) The Flavor Industry ,: 405-406 (1970) Phytochemistry ,: 13, 2171-2174 (1974) Fragrance: 115, 31-40 (1976) Pharmaceutical Journal: 85 (5), 737-740 (1969) Plant World No.99, Shukan Asahi Encyclopedia, 66-71 (1997) Asahi Shimbun Primary Color Japanese Medicinal Plant Dictionary, No.2, 89-90 (1066) Sebundo Shinkosha Acta. Phytotax. Geobot., 161-167 (1987)

  Therefore, in view of the above-mentioned present situation, the present invention has an object to provide a composition for improving immune function based on the immune function improving action of the extract based on the study of the pharmacological action of the extract of Camphoraceae black moji.

  The present invention for solving this problem is a composition for improving immune function, characterized in that, as a basic aspect thereof, an extract of Camphoraceae chromophore is used as an active ingredient.

  Specifically, in the present invention, the extraction solvent is water, a lower alcohol or a mixed solvent of lower alcohol and water, particularly the lower alcohol is methanol or ethanol, and the obtained solvent extract, particularly the extract, The above-mentioned composition for improving immune function, which is dried.

  The present invention also relates to the above-mentioned composition for improving immune function, characterized in that a decrease in immune function is due to mental stress, and the decrease in immune function is a decrease in systemic immune system and / or intestinal immune system function. The above-mentioned composition for improving immune function, wherein the improvement of immune function is preventive improvement and / or therapeutic improvement of immune function.

  Furthermore, this invention is food / beverage etc. which contain the composition for immune function improvement mentioned above as another aspect, and is use as a functional display food, a pharmaceutical, an animal feed, and a feed for fish farming.

According to the present invention, a highly safe composition for improving immune function is provided.
In particular, the composition comprising an extract of the cinnamonaceae black momodi provided by the present invention as an active ingredient can prevent a decrease in immune function, and can further restore a decreased immune function. Because it is capable of improving the decline in immune function and is highly safe, foods and drinks that have a highly safe immune function improving effect that can be easily ingested in daily life, especially functional indication foods and pharmaceuticals Furthermore, it has the advantage of providing animal feed for pets or dairy farms, and feed for fish farming.

  That is, the present invention always improves immune function decline, in particular, suppresses systemic immunity and / or intestinal immunity function decline due to mental stress, and does not increase systemic immunity and / or intestinal immunity more than necessary. It exhibits an excellent effect as a composition for preventive improvement and / or therapeutic improvement of systemic immunity and / or intestinal immunity function having an effect of maintaining the immune system at a normal level.

It is the figure which showed the result of the thymocyte number in a preventive effect test. It is the figure which showed the result of the peripheral blood white blood cell count in a preventive effect test. It is the figure which showed the result of 11-OHCS (11-hydroxycorticosteroid) in serum in a preventive effect test. It is the figure which showed the result of the Peyer's patch cell number in a preventive effect test. It is the figure which showed the result of the apoptosis of the Peyer's patch cell by Annexin V (Annexin A) dyeing | staining in a preventive effect test. It is the figure which showed the result of the proliferative ability of Peyer's patch T cell by ConA (concanavalin A) stimulation in a preventive effect test. It is the figure which showed the result of the proliferative ability of Peyer's patch B cell by LPS (lipopolysaccharide) stimulation in a preventive effect test. It is the figure which showed the result of Th1 / Th2 balance of Peyer's patch helper T cell in a preventive effect test.

It is the figure which showed the result of the thymocyte number in the recovery promotion effect test. It is the figure which showed the result of the peripheral blood white blood cell count in a recovery promotion effect test. It is the figure which showed the result of the Peyer's patch cell number in a recovery promotion effect test. It is the figure which showed the result of the proliferative ability of Peyer's patch T cell by ConA (concanavalin A) stimulation in a recovery promotion effect test. It is the figure which showed the result of the proliferative ability of Peyer's patch B cell by LPS (lipopolysaccharide) stimulation in the recovery promotion effect test. It is the figure which showed the amount of IgA in an intestine washing | cleaning liquid in a recovery promotion effect test. It is the figure which showed the result of the amount of IgA in an intestinal tract tissue in a recovery promotion effect test.

In addition, Student's T test is used for the test between two groups, and Bonferroni's multiple comparison test is performed for the test between multiple groups.
#: There is a statistically significant difference with respect to the normal control group at p <0.05 or less,
##: There is a statistically significant difference with respect to the normal control group at p <0.01 or less,
*: Statistically significant with respect to the stress control group at p <0.05 or less,
**: p <0.01 or less and statistically significant difference from the stress control group,
Indicates.

As described above, the basic aspect of the present invention is a composition for improving immune function, characterized in that an active ingredient is an extract of Camphoraceae black moji, particularly a lyophilized solvent extract.
As the camphoraceae black moji used in the present invention, L. umbellata var. Lancea Momiyama, L. umbellata var. Lancea Momiyama, L. umbellata var. Membranacea (Maxim.) Momiyama, L. umbellata var. sericea (Sieb.et zucc.) Blume), Usgekuromoji (L. sericea var. glabrata Blume), etc., but are not limited to these, including all of the types described as “Kuromoji”. It is a waste.

In addition, there exists a shark (herb) as a crude drug which dried the trunk branch and root bark of black moji, and this shark is also called a fishing rod (butterfly).
Therefore, it should be noted that, in the present invention, this coral (fish) or fishing rod (butterfly) is also included in the camphor family.
In addition, it has not been known so far that a solvent extract such as kuromoji, which is a camphoraceae, has an immune function improving effect, and the present invention is specific in that respect.

In the present invention, in preparing the extract that is the active ingredient, as the extraction site of the black camphor that is the camphor family, for example, a dried product of each part such as trunk branches, root bark, leaves, flowers, fruits, Alternatively, it is a whole plant dried product including these, and a dried product of trunk branches and root barks is preferably used.
Moreover, the rod as a crude drug or a fishing rod can be used as it is.
Hereinafter, in the present invention, chromophores, buckwheat ceramus, cerebrum pheasant, cerebrum moth, usge chromophore, or rods or fishing rods may be collectively referred to as “Asteraceae chromophore”.

In the extraction, for example, it is preferable to carry out solvent extraction by appropriately cutting a trunk branch such as chromophore which is a camphoraceae, a dried root bark, or a rod or a fishing rod.
As a solvent used for extraction, water, a lower alcohol or a mixed solvent of lower alcohol and water is preferable, and methanol or ethanol is particularly preferably used as the lower alcohol.

  As a specific method of extraction, commonly used extraction means is adopted, for example, a stem branch such as black crab that is a camphor family, a dried root bark, or a rod or a fishing rod appropriately cut. Thereafter, 1 to 50 times, preferably 5 to 20 times the amount of water, lower alcohol or a mixed solvent of lower alcohol and water, preferably methanol or ethanol, and about 1 to 24 hours at room temperature. -Immersion / heat extraction may be performed within the range of the boiling point of the solvent used, and extraction may be performed under pressure as necessary. After extraction, it is filtered as necessary, and the resulting extract is concentrated under reduced pressure to remove the extraction solvent used and lyophilized to obtain the desired extract Can be prepared. The drying means may be vacuum drying or spray drying.

  Thus, it is possible to prepare an extract of the camphoraceae, which is an active ingredient of the present invention. As a result of the study by the present inventors, the obtained extract, in particular, a dried extract, has an excellent immune function improvement. It was found to have an effect.

By the way, the immune system is composed of an innate immune system represented by macrophages and neutrophils responsible for initial protection, and an acquired immune system represented by lymphocytes such as T cells and B cells that follow thereafter.
T cells in the acquired immune system are important lymphocytes that play a role in the control tower of the immune system. T cells are made in the bone marrow as progenitor cells, then move to the thymus, and after proliferating, differentiation and maturation while moving between thymic epithelial cells, acquire the ability to distinguish between self and non-self, It goes out of the thymus as T cells and is sent to the whole body such as lymph nodes. For this reason, the thymus is an important organ that plays a central role in the systemic immune system.

Furthermore, the intestinal tract immune system is an important defense mechanism for the body, along with the systemic immune system of blood, thymus and lymph nodes.
Since the digestive tract is an organ directly connected to the outside world, it is the place where microorganisms, bacteria, pathogens, etc. are most likely to enter. If there is no immune system in this area, it becomes a place where the immune system is most concentrated in the body because it becomes a burrow of pathogens and even lives are lost.
In the intestinal tract, there is an intestinal lymph node called Peyer's patch that is sandwiched between organs responsible for gastrointestinal absorption. When T cells are activated in Peyer's patch subjected to antigen stimulation, B cells proliferate actively and secretory immunoglobulin A (IgA) is produced. Since this IgA prevents the invasion of foreign enemies in the digestive tract, a decrease in intestinal immunity function decreases the production of IgA and easily causes infection.
Thus, Peyer's patches are a particularly important immune system for intestinal immunity.

Among T cells, helper T cells (Th) are cells that regulate the immune system, and Th1 that produces interleukin-2 (IL-2) and interferon γ (IFN-γ), IL-4, and IL-. There is Th2 which produces 5.
Th1 and Th2 differentiate and mature from naive T cells upon receiving antigen presentation from macrophages and the like. Th1 acts to regulate cellular immunity (reacts to cells and viruses), and Th2 acts to regulate humoral immunity (responsive to allergens such as mites and molds).

This balance between Th1 and Th2 is called Th1 / Th2 balance, and it works to suppress each other's action by cytokine secreted from each cell so that either one of the reactions does not become excessive. , Th1 / Th2 balance is maintained.
However, it has been found that when this balance is lost, a number of immune diseases are caused. When Th1 is dominant, cellular immunity becomes too strong and causes onset of autoimmune diseases, while Th2 is dominant. The humoral immunity becomes too strong and it is easy to cause allergic diseases such as infections and atopic dermatitis.

Abnormalities in the immune system are known to cause infections, onset of cancer, allergies such as hay fever and atopic dermatitis, and autoimmune diseases.
In addition to genetic factors, abnormalities in the immune system are said to be caused by aging, eating habits, lifestyle habits such as lack of exercise, environmental factors, and stress. It can be said that the modern society has many problems that cause abnormalities in immune functions, such as stress society and super-aging society.

When stressed, the brain releases CRH (Corticotropin Releasing Hormone) from the hypothalamus and promotes the release of ACTH (Adrenocorticotropic Hormone) from the pituitary gland.
ACTH released into the blood acts on the adrenal cortex to increase glucocorticoid production and suppress the immune system. In addition, immune cells are controlled by autonomic nerves, and the release of adrenaline and noradrenaline occurs due to the excitation of sympathetic nerves due to stress, affecting the immune system.

  In clinical trials, one of the spouses left behind by the spouse's death feels stressed, resulting in a decrease in lymphocyte responsiveness, and an increase in mental stress increases virus morbidity. It has been reported.

Also in livestock, weaning, restraint / isolation, group rearrangement / relocation, and narrow rearing space are stressed, causing a decline in immune function and tending to cause opportunistic infections.
In addition, many studies using experimental animals have been reported, such as atrophy of the thymus, a decrease in the number of peripheral blood leukocytes, and induction of apoptosis in Peyer's cells of the intestinal tract immune system due to restraint stress in mice, a model of human mental stress. . Thus, it is known that stress affects not only the systemic immune system such as thymus and peripheral blood leukocytes but also the intestinal tract immune system.

  Th1 / Th2 balance is also known to be affected by stress, and it is known that when it receives mental stress excessively, cellular immunity functions are lowered, and thus it tends to be inclined to Th2. Yes. It is said that the increase in the allergic population in modern society is caused by stress. For this reason, keeping the Th1 / Th2 balance normal is also the key to keeping the immune function normal.

  In this way, the decline in immune function due to stress leads to many diseases related to the immune system including infections, but it prevents the decline in the functions of multiple immune systems as well as some immune systems. Recovery is extremely important from the viewpoint of biological defense.

The inventors of the present invention experimentally confirmed that the solvent extract of black moji of the present invention improves the systemic immune function and intestinal immunity function reduced by restraint stress to mice in a preventive and therapeutic manner (described later). See the test to do).
From the above, the solvent extract of black moji can suppress the reduced immune function and reduce the reduced immune function in humans and animals (pets, livestock) with few side effects derived from natural products that can be widely applied to food, livestock, and pet food. It can be provided as a food composition.

  That is, the present invention improves the systemic immune function and / or intestinal immunity function reduced by the mental stress load, or suppresses the decrease in the systemic immune function and / or intestinal immunity function due to the mental stress load. Composition for preventive improvement and / or therapeutic improvement of systemic immune function and / or intestinal immunity function having an effect of always maintaining the immune system at a normal level by not enhancing systemic immunity function and / or intestinal immunity as described above It provides things.

The composition for improving immune function provided by the present invention is prepared as preparations of various forms by usual methods using the solvent extract or dried product as it is or using commonly used excipients, additives and the like. be able to.
For example, it can be processed into a form such as a solid, liquid, emulsified, pasty, jelly, etc. to produce food and drink.

That is, the composition for improving immune function provided by the present invention can be effectively applied as a food or drink.
The composition for improving immune function of the present invention can be provided as a food that can be immediately ingested as a food, a food that can be ingested after cooking, or a premixed material for food production.

  The food and drink containing the composition for improving immune function of the present invention may be in any form of solid, powder or granule. Specific examples include, but are not limited to, various confectionery such as biscuits, cookies, cakes, snacks, rice crackers, breads, powdered beverages (powdered coffee, powdered cocoa, etc.), rice cakes, caramels and the like. It is not a thing.

  Moreover, various drinks, such as juice, a carbonated drink, a lactic acid bacteria drink, can be mentioned as a liquid, an emulsion form, a paste form, and a jelly form, It is not limited to these. Among them, it is preferably a paste, jelly, or liquid

  In the composition for improving immune function of the present invention, the extract used as an active ingredient should be ingested about 1 mg to 50 g, preferably about 10 mg to 10 g per day, based on the immune function improving action as a standard. In general, the content of the active ingredient to be taken immediately before meals may be set 1 to 3 times a day, preferably 3 times a day, and a dosage form may be designed.

  When the solvent extract itself is contained, the solvent extract can be diluted with an appropriate solvent to prepare a liquid composition. Alternatively, the solvent extract itself can be mixed with an appropriate carrier, preferably fatty acid triglyceride, and filled in a soft capsule or the like in a liquid state.

  In the above formulation, commonly used excipients, binders, disintegrants, lubricants, stabilizers, surfactants, solubilizers, reducing agents, buffers, adsorbents, fluidizing agents, Select one or more kinds of formulation additives such as antistatic agents, antioxidants, sweeteners, flavoring agents, cooling agents, light-shielding agents, flavoring agents, fragrances, fragrances, coating agents, plasticizers, etc. Needless to say, it may be added.

  Specific examples of such formulation additives include crystalline cellulose, low-substituted hydroxypropylcellulose, hydroxypropylmethylcellulose, croscarmellose sodium, maltodextrin, ethylcellulose, lactose, sorbitol, silicic anhydride, and silicic acid. Magnesium, hydroxypropylcellulose, stearic acid, oleic acid, liquid paraffin, dicalcium phosphate, dibutyl sebacate, macrogol, propylene glycol, corn starch, starch, pregelatinized starch, gelatin, popidone, crospovidone, glycerin, polysorbate 80, citrus Acid, acesulfame potassium, aspartame, sodium carbonate, talc, magnesium stearate, calcium stearate, etc. That.

  In preparation of the composition for improving immune function of the present invention, other ingredients such as vitamins, minerals, etc. within a range of quality and quantity that do not impair the immune function improving action of the extract as an active ingredient. Can also be added.

Furthermore, although this invention is a composition for immune function improvement which contains the solvent extract prepared above as an active ingredient, this composition can be used as a foodstuff. That is, the composition for improving immune function prepared as described above can be directly mixed with food or other ingredients to form a functionally labeled food or pharmaceutical form.
Furthermore, it can be provided as food for livestock and pets.

  Hereinafter, the present invention will be described in detail by way of specific test examples, examples, and the like, but the scope of the present invention is not limited to these examples.

Example 1: Preparation of a solvent extract of Kuromoji 1 kg of dried trunk branches of Kuromoji was cut into small pieces, 10 L of 30% ethanol / 70% water-mixture was added thereto, and extraction was performed by heating at 60 ° C for 1 hour. . After the extraction treatment, the alcohol solution obtained by filtration was concentrated to dryness under reduced pressure, and then freeze-dried to obtain 44.5 g of a chromophore extract.

Example 2: Confirmation of preventive and therapeutic amelioration effect of chromophore extract on immune function decline in restraint stress-loaded mice (Test 1 and Test 2)

Experimental method 1) Animals used
Balb / c mice (9 weeks old, male) were preliminarily raised for 1 week and then used for experiments.
2) Test substance Obtained by adding 20 L of 30% methanol / 70% water-mixture to chromodi 6.5 kg, heating and extracting (60 ° C., 1 hour), concentrating and freeze-drying as in Example 1. A product (hereinafter referred to as “chromomy extract”) was suspended in a 0.5% CMC (carboxymethylcellulose: Wako Pure Chemical Industries, Ltd.) solution and used.
3) Schedule Stress was restrained for 16 hours (PM 5: 00 to the next day AM 9:00) by placing the mouse in a plastic 50 mL centrifuge tube.

Test 1: Examination and confirmation of prophylactic effect Chromoji extract was orally administered 1 hour before stress loading, and after applying 16 hours of restraint stress, treatment was performed, and each item was measured (n = 8-10) ).
Chromoji extract dosage:
2 doses of 0.5 g / kg and 1.0 g / kg In addition, a normal control group that receives 0.5% CMC solution as a subject and does not load restraint stress, and only 0.5% CMC solution that loads restraint stress And a stress control group.

Measurement item:
Thymocyte count Peripheral blood leukocytes Blood 11-OHCS
Peyer's patch cell number Peyer's patch apoptosis Peyer's patch proliferating Peyer's patch Th1 / Th2 balance

Test 2: Examination / confirmation of recovery-promoting effect Chromoji extract was orally administered immediately after release (AM9: 00) and day 1 of release (AM9: 00 after 24 hours) after 16 hours of restraint stress load, and day 2 of release (AM 9:00 after 48 hours) was treated, and measurements were made for each item (n = 7-8).
Chromoji extract dosage:
2 doses of 0.5 g / kg and 1.0 g / kg In addition, a normal control group that receives 0.5% CMC solution as a subject and does not load restraint stress, and only 0.5% CMC solution that loads restraint stress And a stress control group.

Measurement item:
Thymic cell count Peripheral blood leukocyte Peyer's patch cell number Peyer's patch proliferative capacity Intestinal IgA amount

The actual measurement will be described in detail below.
Mice were bled from the abdominal vena cava under anesthesia, and then the thymus and intestine were removed and weighed.
For peripheral blood, the number of blood cells was measured using Celltac (Nihon Kohden). In addition, serum was collected from peripheral blood after centrifugation, and 11-OHCS (11-hydroxycorticosteroid), which is a corticosteroid hormone, was measured by a fluorescence method.
For the intestinal tract, the small intestine from the duodenum to the terminal ileum is taken out, the intestinal contents are washed with 20 mL of phosphate buffered saline (PBS), and the collected material is stirred and centrifuged. Then, the supernatant is collected and washed. The amount of IgA was measured with BD OptEIA ELISA Sets (BD Pharmingen). The Peyer's patches were all removed from the remaining small intestine, and the intestinal tract tissue was homogenized with a 50-fold amount of PBS containing 0.05% Tween 20 and centrifuged. The supernatant was collected and the amount of IgA in the intestinal epithelial tissue was measured.

The extracted Peyer's plate is suspended in RPMI1640 medium containing 10% Fetal Bovine Serum (FBS: fetal bovine serum) that has been immobilized while grinding the cells with a stainless steel mesh, and using a flow cytometer, Population and apoptosis measurements were made. Apoptosis was measured using Annexin V-FITC kit (manufactured by Sigma). The remaining Peyer's patch cells were measured for the number of cells, adjusted to RPMI1640 medium containing 10% FBS (1 × 10 ⁶ / mL), and mitogen ConA (Concanavalin A: manufactured by Sigma). 2 μg / mL, or LPS (lipopolysaccharide: manufactured by Difco) 25 μg / mL was added, and the cells were subjected to stimulation culture under conditions of 5% CO ₂ and 37 ° C., and cell proliferation ability was measured after 5 days. The absorbance value at 450 nm was measured by the 8 method (manufactured by Dojindo).
The cell growth ability was expressed as a Stimulation Index (SI) value obtained by dividing the absorbance value for the stimulated culture (cultured with ConA and LPS added) by the absorbance value for the unstimulated culture.

Furthermore, the washed Dynabeads Mouse CD4 solution (Dynal) is added to the Peyer's plate cell suspension to react, and only the CD4 cells are separated with a magnetic separator, and the DETACHa BEAD Mouse CD4 (Dynal) solution is added to Dynabeads. PMA [phorbol 12-myristate 13-acetate] (50 ng / mL: manufactured by Wako Pure Chemical Industries, Ltd.) + ionomycin (1 μg / mL: manufactured by Sigma) was added to the cell suspension from which the (microbeads) were removed. _2. Stimulated culture was performed at 37 ° C., and the IL-4 and IFN-γ levels in the supernatant after 2 days of culture were quantified by BD OptEIA ELISA Sets (BD Pharmingen), and the Th1 / Th2 balance was It was expressed as a ratio (IL-4 / INF-γ) of INF-γ (ng / mL) which is a Th1 cytokine and IL-4 (pg / mL) which is a Th2 cytokine.

  The results based on the above measurements are described below.

Experimental Results <Preventive Effect Test: Results of Test 1>

1. Thymocyte count, peripheral blood leukocyte count and serum 11-OHCS
The results are shown in FIGS.
FIG. 1 shows the result of thymocyte count, FIG. 2 shows the result of peripheral blood leukocyte count, and FIG. 3 shows the result of serum 11-OHS.
As can be seen from the results shown in the figure, the thymocyte count and peripheral blood leukocyte count of the systemic immune system were significantly reduced by restraint stress load, but the chromodi extract (0.5, 1.0 g / kg) significantly suppressed this decrease. In addition, 11-OHCS in serum was remarkably increased by restraint stress, but administration of chromomy extract (0.5, 1.0 g / kg) did not affect this increase.

2. Peyer's patch cell count The results are shown in FIGS.
FIG. 4 shows the results of Peyer's patch cell count, and FIG. 5 shows the results of Peyer's patch cell apoptosis by Annexin V (Annexin A) staining.
As can be seen from the results shown in the figure, the number of Peyer's patch cells in the intestinal tract immune system was significantly reduced by the restraint stress load, and apoptosis (cell death) was increased.
The administration of chromomy extract tended to suppress the decrease in the number of Peyer's patch cells, and significantly suppressed the increase in apoptosis.

3. Peyer cell proliferation ability, Th1 / Th2 balance The results are shown in FIGS.
FIG. 6 shows the results of Peyer's patch T cell proliferation ability by stimulation with ConA (Concanavalin A), FIG. 7 shows the results of Peyer's patch B cell proliferation ability by stimulation with LPS (lipopolysaccharide), and FIG. 8 shows Th1. The result of / Th2 balance is shown.
As can be seen from the results shown in the figure, the proliferative ability of Peyer's patch T cells by stimulation with ConA (Concanavalin A) tends to be suppressed by restraint stress loading, and Peyer's patch by LPS (lipopolysaccharide) stimulation. The proliferation ability of B cells was remarkably suppressed.
Further, the ratio of cytokine production in Peyer's patch Th cells by PMA + Ionomycin stimulation (IL-4 / INF-γ) was dominated by IL-4 due to restraint stress load, and the balance between Th1 and Th2 was inclined to Th2.
The administration of Kuromoji showed a tendency to suppress the decrease in proliferation ability due to ConA stimulation, and significantly suppressed the decrease in proliferation ability due to LPS stimulation.
In addition, the bias toward Th2 in the balance between Th1 and Th2 was significantly suppressed.

<Recovery Acceleration Effect Test: Results of Test 2>
The results are shown in FIGS.
FIG. 9 shows the result of thymocyte count, FIG. 10 shows the result of peripheral blood leukocyte count, FIG. 11 shows the result of Peyer's patch cell count, and FIG. 12 shows Peyer's patch T cell by ConA (Concanavalin A) stimulation. FIG. 13 shows the results of Peyer's patch B cell proliferation by stimulation with LPS (lipopolysaccharide), FIG. 14 shows the amount of IgA in the intestinal lavage fluid, and FIG. 15 shows the results of intestinal tissue. It is the figure which showed the result of IgA amount.

  As can be seen from the results shown in the figure, thymocytes and peripheral blood leukocytes in the systemic immune system were significantly reduced even after 2 days of release of restraint stress. Administration of black moji extract (0.5, 1.0 g / kg) after release of stress load significantly restored this decrease.

Two days after releasing the restraint stress load, Peyer's patch cells in the intestinal tract immune system decreased, and a tendency to suppress the proliferation of Peyer's patch T cells by ConA stimulation was observed. Inhibition of proliferation ability was observed.
The administration of chromomy extract (0.5, 1.0 g / kg) significantly recovered the decrease in Peyer's patch B cell proliferative ability caused by LPS stimulation.

  In addition, two days after the release of the restraint stress load, the amount of IgA in the intestinal lavage fluid and in the intestinal tissue decreased, but administration of chromodi extract (0.5, 1.0 g / kg) The amount of IgA in the lavage fluid and intestinal tissue was restored to decrease.

  From these experiments, it was confirmed that the chromophore extract of the present invention has an effect of prophylactically and therapeutically improving immune function decline, particularly systemic immune function and intestinal immune function decline due to mental stress.

Example 3 Pharmaceutical (1) Tablet Containing Chromoji Solvent Extract Extract 55.0 g of Chromoji 30% ethanol extract (freeze-dried product) obtained in Example 1 was prepared by adding 249.5 g of lactose and magnesium stearate 0 A tablet with a diameter of 10 mm and a weight of 300 mg was produced by tableting with a single tableting machine together with .5 g.

(2) Powder:
Add 0.5 g of magnesium stearate to 100.0 g of 30% ethanol extract (freeze-dried product) of black moji obtained in Example 1 above, and compress, crush, granulate and sieve to give 20-50 mesh granules. Obtained.

Example 4: Various foods and drinks containing a solvent extract of Kuromoji 30% ethanol extract of chromomy obtained in Example 1 (freeze-dried product: hereinafter simply referred to as extract) in the composition shown below Various foods and beverages were manufactured.
(1) 飴:
(Composition) (Parts by weight)
Powdered sorbitol 89.70
Fragrance 0.25
Extracted extract 10.00
Sorbitol seed 0.05
Total amount 100.00

(2) Gum:
(Composition) (Parts by weight)
Gum base 20.00
Calcium carbonate 2.00
Stevioside 0.10
Extracted extract 10.00
Lactose 66.90
Perfume 1.00
Total amount 100.00

(3) Caramel:
(Composition) (Parts by weight)
Granulated sugar 32.00
Minamata 20.00
Milk powder 30.00
Hardened oil 4.00
Salt 0.60
Perfume 0.03
Water 3.37
Extracted extract 10.00
Total amount 100.00

(4) Carbonated drink:
(Composition) (Parts by weight)
Granulated sugar 8.00
Concentrated lemon juice 1.00
L-ascorbic acid 0.10
Citric acid 0.09
Sodium citrate 0.05
Coloring agent 0.05
Carbonated water 80.71
Extracted extract 10.00
Total amount 100.00

(5) Juice:
(Composition) (Parts by weight)
Frozen concentrated orange juice 5.00
Fructose glucose liquid sugar 1.00
Citric acid 0.10
L-ascorbic acid 0.09
Extracted extract 10.00
Fragrance 0.20
Dye 0.10
Water 83.51
Total amount 100.00

(6) Lactic acid bacteria beverage:
(Composition) (Parts by weight)
Milk solid 21% fermented milk 14.76
Fructose dextrose liquid sugar 13.31
Pectin 0.50
Citric acid 0.08
Fragrance 0.15
Water 61.20
Extracted extract 10.00
Total amount 100.00

(7) Alcoholic beverages:
(Composition) (Parts by weight)
50% ethanol 32.00
Sugar 8.60
Fruit juice 2.40
Extracted extract 10.00
Water 47.00
Total amount 100.00

  As described above, the present invention provides a composition for improving immune function comprising a plant extract, in particular, a solvent extract of Camphoraceae chromophore, as an active ingredient. Because the extract can be applied to provide food and drink using a highly safe composition for improving immune function that can be easily ingested in daily life, especially functional indication foods, and pets and livestock feed The industrial applicability is tremendous.

Claims (9)

  A composition for improving immune function, which comprises a solvent extract of Camphoraceae black moji as an active ingredient.   The composition for improving immune function according to claim 1, wherein the extraction solvent is water, a lower alcohol or a mixed solvent of lower alcohol and water.   The composition for improving immune function according to claim 2, wherein the lower alcohol is methanol or ethanol.   The composition for improving immune function according to any one of claims 1 to 3, wherein the solvent extract is in the form of a dried product.   The composition for improving immune function according to any one of claims 1 to 4, wherein the decrease in immune function is due to mental stress.   The composition for improving immune function according to any one of claims 1 to 5, wherein the decreased immune function is a decreased function of the intestinal tract immune system.   The composition for improving immune function according to any one of claims 1 to 5, wherein the decreased immune function is a decreased function of the systemic immune system.   The composition for improving immune function according to any one of claims 1 to 7, wherein the improvement of immune function is preventive improvement and / or therapeutic improvement of immune function.   The composition for improving immune function according to any one of claims 1 to 7, which is used as a functional display food, a pharmaceutical product, an animal feed, or a fish feed.

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