JP2002335926A - Composition and drink comprising the same and skin liniment - Google Patents

Abstract

PROBLEM TO BE SOLVED: To obtain a composition preventing pulrefaction and oxidation in storing the composition including a β-1, 3-1, 6-glucan, and capable of increasing antioxidative property and antiallergic action in the body, and useful for a drink and a skin liniment. SOLUTION: This composition includes the β-1, 3-1, 6-glucan and an extract of an apple, preferably includes the β-1, 3-1, 6-glucan obtained by the culture of a microorganism belonging to the genus Aureobacidium and the apple extract obtained from the immature fruits. The composition is useful for drinks especially for healthy drinks preferably by formulating Vitamin C and adjusting pH at 4-6. The composition is also useful for the skin liniment and preferably includes horse oil.

Description

DETAILED DESCRIPTION OF THE INVENTION

[0001]

[0001] The present invention relates to β-1,3-1,
The present invention relates to a composition containing 6-glucan and an apple extract, and a beverage and a skin coating composition comprising the composition.

[0002]

2. Description of the Related Art β-1,3-1,6-glucan is β-3-1,6-glucan.
It is a polysaccharide in which a large number of glucoses are mutually linked by a 1,3 glucose bond and a β-1,6 glucose bond. It is known to be a substance having various effects as follows, and is a substance useful for maintaining and promoting human health. (1) Immunity-enhancing action: enhances the function of lymphocytes and improves immune function. (2) Anti-tumor activity: After cancer cells have already developed and become ill, they attack the cancer cells. (3) Cancer cell growth inhibitory action: suppresses the growth of cancer cells after they have occurred. (4) Antiallergic action: suppresses type I allergy such as atopic dermatitis. (5) Anti-inflammatory action: Improves the protective function by improving the immune system. (6) Cholesterol lowering action: lowers cholesterol in blood. (7) Dietary fiber effect: adsorbs carcinogens and discharges them out of the body. (8) Antithrombotic action: Prevents the inside of blood vessels from narrowing or clogging. (9) Blood pressure lowering action: When blood sent from the heart becomes abnormal due to the pressure applied to the artery wall, it returns to a normal state. (10) Hypoglycemic action: lowers excess glucose in blood. (11) Liver hyperactivity: Improves the detoxification ability for liver function.

[0003] β-1,3-1,6-glucan is contained in mushrooms such as awatake mushroom, shiitake mushroom and shirohirotake mushroom, and can be taken by eating these mushrooms and extracted therefrom. It is also possible to ingest what you have done. However, if you eat it as it is,
The absorption of 1,3-1,6-glucan into the body is insufficient and storage is not easy. Further, β-1,3-1,6-glucan can be extracted from these mushrooms, but long-term cooking from a large amount of mushrooms is required. There is a need.

Japanese Patent Application Laid-Open No. 57-149301 discloses that a microorganism belonging to the genus Aureobacidium of the black-and-white fungus family, Aureobacidium, has a non-reducing terminal having a non-reducing terminal of β from a main chain in which glucose is linked to β-1,3 glucose. It describes that a polymer polysaccharide having a structure branched by a -1,6 glucose bond and a phosphate group bonded to glucose is produced. The microorganism used here is the deposit number of No. 4257 (F.
ERM-P. 4257).

Japanese Patent Application Laid-Open No. 6-340701 discloses Aureobacidium pullulans.
pullulans) by culture of the IFO4466 strain, β-
A 1,3-linked glucose residue is used as a main chain, and β-
It is described that a β-glucan having a branched chain of 1,6-linked glucose residues as many side chains is produced.

[0006] In addition, the fact that a microorganism of the genus Aureobasidium produces β-1,3-1,6-glucan has been reported by Acta.
Chemical Scandinavia 17, 1351-1356 (1963), Agri
c. Biol. Chem. 47 (6), 1167-1172 (1983), Chem. Pha
rm. Bull., 40, 2215 (1992).

[0007] As described above, a method of efficiently obtaining β-1,3-1,6-glucan by culturing a specific microorganism has been reported.

[0008]

However, β-
When a composition containing 1,3-1,6-glucan is used for direct use by a general consumer, such as a health drink or a skin coating, there is a problem in that it easily rots due to propagation of various bacteria during storage. . In ordinary households, no special countermeasures against falling bacteria are taken, so even if the composition is once sterilized, the bacteria often adhere during use. In particular, in the case of directly drinking or applying a composition containing a culture solution of aureobasidium, it is difficult to prevent decay over a long period of time because the composition is a liquid having a composition in which microorganisms can easily propagate in the first place. there were. However, the use of chemically synthesized bacteriostatic agents has a great resistance to consumers who are naturally oriented to use them as health drinks or skin coatings.

Further, β-1,3-1,6-glucan is easily oxidized, and it is necessary to prevent oxidative deterioration for long-term storage. However, the use of chemically synthesized antioxidants still has a great resistance to natural-minded consumers.

The present invention solves these problems and provides a composition capable of enhancing antioxidant and antiallergic effects in the body, and is particularly useful as a beverage or a skin coating agent. The present invention provides a novel composition.

[0011]

The object of the present invention is to provide a β-1,3
This is achieved by providing a composition containing -1,6-glucan and apple extract. The apple extract, which is a natural product, prevents the decay and oxidation of the β-1,3-1,6-glucan-containing composition, and further enhances the antioxidant and antiallergic effects in the body. At this time, β-1,
When 3-1, 6-glucan is obtained by culturing aureobasidium, the effect of blending the apple extract is remarkable. As the apple extract, those extracted from immature fruits of apples are preferable. The composition is suitably used for applications applied to the human body as a medical composition or a pharmaceutical composition.

The composition of the present invention is useful as a beverage, particularly as a health drink, and can have various effects such as antitumor activity by drinking. At this time, add vitamin C
Is preferably adjusted to 4 to 6. Further, the composition of the present invention is also useful as a skin coating agent, and when applied to the skin, an antipruritic effect of atopic dermatitis can be obtained. At this time, a skin coating composition further containing horse oil is preferred.

[0013]

BEST MODE FOR CARRYING OUT THE INVENTION The composition of the present invention comprises β-1,3-
Contains 1,6-glucan. Here, β-1,3-
1,6-glucan is a polysaccharide in which a large number of glucoses are mutually linked by β-1,3 glucose bonds and β-1,6 glucose bonds. Glucose is β-
Those having a structure in which glucose is branched by β-1,6 bonds from a 1,3-bonded main chain (β-1,6-branched-β-
1,3-glucan) is preferred. Having such a branched structure increases the physiological activity.

The β-1,3-1,6-glucan of the present invention contains glucose as a main component, but may contain a small amount of other constituent sugars. The ratio of glucose in all constituent sugars is usually preferably 90% or more. In addition, the β-
1,3-1,6-glucan may have another functional group. In particular, having an acid group as another functional group enhances biological activity. The acid group contained is preferably a phosphoric acid group, and the content of the phosphoric acid group in this case is preferably at least 1 mol% based on glucose units.

The molecular weight of the β-1,3-1,6-glucan of the present invention is not particularly limited, and usually has a repeating structure of 100 or more glucose. Preferably, the number average molecular weight in the molecular weight measurement by osmometry (using a Zimm-Myerson type osmometer, measured using a cellophane semipermeable membrane) is 50,000 to 500,000
It is about.

Β-1,3-1,6- in the composition of the present invention
The content of glucan is appropriately adjusted depending on the purpose of use, and a content of usually 0.001 to 5% by weight is exemplified. However, considering the balance between manufacturing cost and effectiveness,
It is preferably 0.5% by weight, more preferably 0.02 to 0.3% by weight.

Β-1,3-1,6- used in the present invention
As glucan, Aureobacidiu (Aureobacidiu)
Those obtained by culturing microorganisms of the genus m) are preferred. The microorganisms of the genus Aureobasidium that can be used are not particularly limited, but they can be used with a deposit of No. 4257 (FERM-P.4).
257) and IFO4466 bacteria. Above all, deposit number 4257 (FERM-P.425)
The bacterium 7) is preferable from the viewpoint of bioactivity.

Β-1,3-extracted and purified from a culture solution
Although 1,6-glucan can be used, it is better to mix the culture solution into the composition as it is and to meet the needs of naturalists who want to avoid unnecessary processing. It is also preferable in that the cost required for purification can be saved. At this time, the content of the culture solution in the composition is preferably 10 to 99.999% by weight.

Further, the composition of the present invention contains an apple extract. Apples contain various polyphenols that have various effects such as antioxidant effect, bacteriostatic effect, antiallergic effect, prevent decay and oxidation of the composition, and also have antioxidant and antiallergic properties in the body. It enhances the action.

In the case of a composition containing only β-1,3-1,6-glucan and no apple extract, it is difficult to prevent rot over a long period of time in ordinary households. . This is greatly improved by further containing the apple extract. This is supported by the fact that the addition suppresses the growth of staphylococci and Escherichia coli in the examples described below.

Tea is well known as a polyphenol-containing food, but the apple extract is more than 10 times more soluble in water than the tea extract. It is preferable because a stable aqueous solution can be obtained. In addition, compared to a tea extract having a strong bitterness, an apple extract has a smaller effect on taste, and is suitable for use in eating and drinking. Further, since the tea extract is dark brown and the color tone of the product becomes brown, there is a possibility that consumers may avoid using the tea extract when it is used for eating, drinking, coating or the like.

More preferably, the apple extract used is extracted from immature apple fruits. Here, the immature fruit is one that is cut at a time when the weight per one is about 5 to 20 g, and one that is cut at a time when the polyphenol is most contained. Apple immature fruits contain about 10 times as many polyphenols as mature fruits.
The polyphenols contained are apple condensed tannin, (+)-catechin, (−)-epicatechin, chlorogenic acid, dihydrochalcone, quercetin glycoside, p-coumarate and the like. The extract of immature apple fruit is available from Nikka Whiskey Co., Ltd.
(Registered trademark) ".

The amount of the apple extract is appropriately adjusted depending on the purpose of use, and is usually 0.001 to
A content of 5% by weight is exemplified. However, considering the balance between production cost and effectiveness, 0.01%
To 1% by weight, more preferably 0.02 to
0.5% by weight. If it exceeds 5% by weight, it becomes bitter when used as a beverage, and tends to be colored when subjected to heat sterilization. On the other hand, when the amount is less than 0.001% by weight, the effect of the compounding is often insufficient.

The composition of the present invention comprises the above β-1,3-1,
It contains 6-glucan and an apple extract as essential components, and is usually used as an aqueous solution or a water-containing paste. If necessary, other components may be further added.

The components to be blended are not particularly limited. Cordyceps sinensis, bear bamboo extract, matcha or kelp, which are plant-derived nutrients for the purpose of complementing macronutrients such as vitamins, minerals, chlorophyll and protein in a well-balanced manner; vitamin B1 or vitamin B6 for complementing neurotransmitter proliferation; Glucose, chlorella, bulk powder, vitamin B12 or folic acid for supplementing hematopoietic regulation; alginic acid, catechin, licorice extract, glycyrrhizin, chlorella, tannin, vitamin C or propolis for supplementing antiviral function; Ginkgo biloba extract, γ-oryzanol, catechin, sesaminol, soybean saponin, tannin, vitamin E, vitamin B2 or vitamin C, etc. for the purpose of complementing the lipid-suppressing function; isomaltooligosaccharides, galactooligosaccharides, for the purpose of complementing the bifidus activity Kisil sucrose, ki Roorigo sugar, chitosan,
Glycomacropeptide, wheat fiber, corn fiber, soy oligosaccharide, hemicellulose or soy peptide, etc .; inositol, casein phos peptide or vitamin D, etc. for the purpose of supplementing calcium absorption promotion; Vitamin C or heme iron; opioid peptide, gymnema, wheat fiber, corn fiber, tannin, maltooligosaccharide, hemicellulose, etc. may be added for the purpose of supplementing digestive absorption control.

In addition, aloe extract, aloe vera extract, rose extract, amacha extract, emisoso, oegonia, ogon, oakaku, hypericum perforatum, seaweed, wormwood, licorice, kikyo, kumasasa, brown sugar, gennoshoko, burdock , Chlorella, Shiitake, Shikon, Shiso, Peony, Peony, Iris, Senkyo, Assemblage, Souhakuhi, Chimp, Carrot, Touki, Loquat, Bukuryu,
Peach, saxifrage, yuzu, reishi, royal jelly, orange, chamomile, birch, parsley, butterbur, dandelion, mulberry,
Extracts such as fennel, garlic, lemon, and Tochu tea, vitamins, ginkgo biloba extract, chitosan, heme iron, vitamin D, blueberry extract, mucopolysaccharide protein, nucleic acid extract, collagen, konjac and the like may be added.

[0027] The method of using the composition of the present invention is not particularly limited, but suitable uses include food and drink and external preparations.

First, the case of using as a food or drink, particularly a beverage will be described. By drinking the composition of the present invention, a tumor-suppressing effect and an I-type allergic reaction-suppressing effect are obtained, and the storage stability is also improved, so that the composition is suitable as a health drink.

The beverage of the present invention contains β-1,3-1,6-glucan and an apple extract. At this time, when the apple extract is not contained but contains only β-1,3-1,6-glucan, the preservability is not sufficient, and various bacteria are liable to propagate and rot or discolor. .
Moreover, when taken, there is a high incidence of side effects such as allergic rash and vomiting.
Furthermore, the period until the effect of drinking tends to be long tends to be long.

When the composition containing β-1,3-1,6-glucan and apple extract of the present invention is used as a beverage, vitamin C may be further added to the composition of the present invention. It is suitable.

Vitamin C is a water-soluble vitamin also called L-ascorbic acid, but cannot be synthesized in the body and must be taken from foods and the like every day. It is said that it has an antioxidant action, an antiviral action, a detoxifying action, and an anticancer action.
It is particularly preferable to mix 1,6-glucan in view of the ability to enhance the tumor-suppressing action and the ability to enhance the antioxidant action of the apple extract. Vitamin C is easily oxidized during storage, but the combined effect of vitamin C can be maintained by using applephenone in combination.
The amount of vitamin C is preferably 0.01 to 5% by weight, more preferably 0.02 to 2% by weight.

When the composition of the present invention is used as a beverage, the pH is preferably adjusted to 3 to 5. More preferably, the pH is 3.5 to 4.8. Usually, the pH of the aureobasidium culture solution is about 5.2, but if it is kept as it is, the preservability may be poor. By adjusting the pH to a slightly acidic side, the storage stability is improved. In addition, by doing so, a moderate sourness can be exhibited, and it becomes suitable for drinking also in terms of palatability. At this time, it is particularly preferable to adjust the pH by adding the above-mentioned vitamin C, since the effect of adding vitamin C is also obtained. In addition, the beverage of the present invention can also contain other sweeteners, flavors, and the like.

The intake of the beverage of the present invention when it is used as a health drink should be appropriately adjusted depending on the health condition, disease symptoms, etc., and the amount of β-1,3-1,6-glucan is 0.18. If you drink a beverage containing 3% by weight,
It is standard to drink a weight of / 10,000 to 8/10000.
That is, it is standard for a person weighing 60 kg to drink about 18 to 42 ml per day.

Since the beverage of the present invention has the effect of restoring and adjusting the immunity inherent in humans, it is expected to have the effect of improving various symptoms. Such effects are listed below, and various effects can be expected. (1) The effect of reducing various allergic symptoms such as atopic dermatitis, hay fever and asthma. (2) Improvement effect on immunological disorders such as collagen disease, rheumatism and HIV. (3) Suppressive effect on various cancers such as stomach cancer, lung cancer, colon cancer, uterine cancer, breast cancer. (4) Improvement effect on vascular disorders such as myocardial infarction and cerebral infarction. (5) Improvement effect on viral diseases such as hepatitis B and hepatitis C. (6) Improvement effect on urinary system diseases such as diabetes and dysuria. (7) The effect of improving digestive diseases such as constipation, diarrhea, stomatitis, hemorrhoids, anorexia, and hangover.

Next, a case where the composition is used as an external preparation, particularly as a skin coating agent, will be described. Β-1,3-1,6 of the present invention
-A composition containing glucan and apple extract has an antipruritic effect on atopic dermatitis, artificial dialysis pruritus, senile pruritus, xerosis, etc., and is applied to the skin for those who have these symptoms. Useful as an agent.

In recent years, atopic dermatitis has become a serious problem since it has been widely affected from infants to adults. In particular, many patients are suffering from severe itching, and there is an eager need for a method to stop itching.
Atopic dermatitis is caused by diet, housing, chemicals, clothing,
Due to the wide variety of causes, such as stress, it is not easy for experts to determine the cause and take a definitive solution, and it is unavoidable that only palliative treatment must be performed. Often.

From the viewpoint of dermatology, the inventor of the present invention has intensively studied to obtain a skin coating agent which can be easily applied to the skin and does not cause itching due to rash. As a result, they have found that the composition containing β-1,3-1,6-glucan and apple extract of the present invention can achieve the above object. That is, by applying a skin coating composition comprising the composition of the present invention, a kind of protective film is formed on the skin of the affected part, and by maintaining the protection state for a long time, invasion and propagation of various bacteria into the affected part are prevented. ,
It promotes the original regenerative power, and as a result, itching is reduced.

In addition to β-1,3-1,6-glucan,
By containing the apple extract, rot during storage can be prevented as described above, and long-term storage is easy. In addition, when only the β-1,3-1,6-glucan was contained without containing the apple extract, the itch was not sufficiently controlled.

The skin coating composition of the present invention preferably further contains horse oil. Horse oil has long been known to be effective in treating dermatitis, burns, cuts, abrasions and the like. Horse oil is a component extracted and purified from the subcutaneous fat of horses, has a moisturizing effect, an oil content adjusting effect, and can enhance the effect of preventing itching. Further, the absorption of the active ingredient can be improved. The preferred amount is 2 to 20% by weight, more preferably 3 to 15% by weight. If it exceeds 20% by weight, stickiness may occur after use, and if it is less than 2% by weight, the effect of the compounding is insufficient.

Horse oil blended in the skin coating composition of the present invention comprises:
It is usually deproteinized, blood removed, deodorized and decolorized.
Among them, those having a melting point of room temperature or lower, particularly 10 ° C. or lower, and being liquid at room temperature are preferable because they can improve the permeability of horse oil to the skin. If the melting point exceeds 10 ° C., it may take time for the protective agent to dissolve on the skin, or the usability such as elongation to the skin may be reduced. Examples of a method for adjusting the melting point include a method of adding hydrogen and a method of blending a high melting point wax such as beeswax.

When the skin coating composition of the present invention contains horse oil and is used as an emulsion, it preferably contains a surfactant. This is because moisture and oil derived from horse oil can maintain a stable emulsified state. The preferred content of surfactant is 0.2 to 15% by weight.

As the above-mentioned surfactant, a nonionic surfactant is suitable, and it is particularly preferable to mix and use both a nonionic surfactant having an HLB of 8 or more and a nonionic surfactant having an HLB of less than 8. The preferred content ratio of the former is 0.1 to 10% by weight, and the latter is 0.1 to 10% by weight.
It is.

Nonionic surfactant having an HLB of 8 or more and H
Both nonionic surfactants having an LB of less than 8 are used to balance O / W emulsification and W / O emulsification. Examples of the nonionic surfactant include a polyhydric alcohol ester and an ethylene (propylene) oxide block copolymer. Examples of the nonionic surfactant having an HLB of 8 or more include polyethylene glycol monostearate, and examples of the nonionic surfactant having an HLB of 8 or less include glyceryl monostearate. At least one of the agents is preferably of a self-emulsifying type. This is because if both nonionic surfactants are other than the self-emulsifying type, the temperature stability is reduced, it becomes difficult to prepare a uniform skin protective agent, and separation often occurs.

The skin coating composition of the present invention preferably contains a water-soluble polymer. Water-soluble polymers are those that are formulated to form gels and are based on polysaccharides (guar gum, xanthan gum, hyaluronic acid, pectin,
Carrageenan, etc.), cellulosic (methylcellulose, ethylcellulose, hydroxyethylcellulose, etc.), starch (soluble starch, etc.), algin (sodium alginate, etc.) or synthetic polymer (polyvinyl alcohol, sodium polyacrylate, carboxyvinyl polymer, etc.)
Are exemplified. The water-soluble polymer is usually blended in the form of a solution. The amount of the polymer itself in the skin protective agent is, for example, 0.1 to 5% by weight, and 0.2 to 2% by weight.
% By weight is preferred.

The skin coating composition of the present invention preferably contains a tea extract. By blending the tea extract, it is possible to enhance effects such as an antiallergic effect, an antibacterial effect, and an active oxygen suppressing effect. Main components in the tea extract include green tea catechins. As green tea catechin,
(+)-Catechin, (-)-epicatechin, (+)-gallocatechin, (-)-epigallocatechin, (-)-epicatechin gallate, and (-)-epigallocatechin gallate are exemplified. The content of the tea extract is preferably 0.01 to 1% by weight, more preferably 0.02 to 0.5% by weight.

Further, the skin coating composition of the present invention preferably contains a licorice extract. An anti-inflammatory effect can be obtained by blending the licorice extract. Glycyrrhizin is a main component of the licorice extract, and glycyrrhizic acid or a salt thereof obtained by hydrolyzing glycyrrhizin can also be used. For example, dipotassium glycyrrhizinate and the like can be used. The content of licorice extract is preferably 0.01
To 1% by weight, more preferably 0.02 to 0.5% by weight.

Further, it is preferable that the skin coating composition of the present invention contains allantoin. By containing allantoin, it is possible to reduce skin irritation and allergic reactions, and also has an effect of healing rough skin and wounds.
The content of allantoin is preferably from 0.01 to 1% by weight, more preferably from 0.02 to 0.5% by weight.

The skin application preparation of the present invention may contain a vegetable extract having skin astringency. As a plant extract, rose extract, aloe extract, arnica extract, horseshoe extract, horsetail extract, hypericum extract,
Sage extract, Achillea millefolium extract, Altea extract, Chamomile extract, Calendula extract and the like can be exemplified. It is particularly preferable to incorporate a rose extract, and an aqueous solution extracted from rose petals can be incorporated.

The skin coating composition of the present invention preferably has a pH of 5 to 8 because it causes less skin irritation to sensitive skin.
A more preferred pH is between 5.5 and 7.5. The pH is adjusted by adding an acid or an alkali.However, since the pH of an aureobasidium culture solution is usually about 5.2, the pH is adjusted to an alkaline side with an alkali such as potassium hydroxide or sodium hydroxide. Often do.

[0050]

The composition of the present invention will be specifically described below with reference to examples. First, an example in which the composition of the present invention is used for a health drink will be described.

[Example 1] A microorganism belonging to the genus Aureobacidium of the incomplete fungus black bacterium [Deposit No. 4257 (FERM)] according to the method described in JP-A-57-149301. -P.4257)]. 0.18% by weight of a polymer polysaccharide having a structure in which a non-reducing end is branched by a β-1,6 glucose bond from a main chain in which glucose is β-1,3 glucose bonded, and a phosphate group is bonded to glucose. The resulting culture broth was obtained. The pH of the main culture was 5.2.

To 500 kg of the main culture, 500 g of “Applephenone (registered trademark)” manufactured by Nikka Whiskey Co., Ltd., which is an extract of apple immature fruits, and 750 g of vitamin C
They were mixed and stirred in a kneading pot for 30 minutes. The obtained liquid was transferred to a filling tank, poured into a glass bottle having a capacity of 500 ml, stoppered, and then sterilized with steam at 85 ° C. for 30 minutes to obtain a health drink. The contents of β-1,3-1,6-glucan, apple extract and vitamin C in the health beverage are respectively
0.18%, 0.1% and 0.15% by weight of the liquid
H was 4.2.

[Tumor Inhibition Test 1 (Administration to Mice)] Sample Preparation Method The health drink of Example 1 was sonicated with an ultrasonic crusher (Ultrasonic Dielectric).
sonication treatment (200W / 20mi) using sruptor UR-200P)
n.) After that, a supernatant was obtained by ultracentrifugation (35000 rpm / 30 min.), and this was diluted 10-fold with a HEPES Good buffer (ph7.4, μ0.15) to obtain a test material. As a comparative sample, a powder obtained by drying a fruit body of Agaricus mushroom was suspended in HEPES Good buffer (ph7.4, μ0.15) to a concentration of 10% (w / v), and further, an ultrasonic crusher (Ultrasonic Disruptor). (UR-200P)
, And a hot water extraction operation was performed for 2 hours after sonication treatment (200 W / 10 min.). Then, ultracentrifuge operation (35000rpm / 30m
in. ) To obtain a supernatant, which was converted to a 1% (w / v) concentration based on the weight of the starting material to obtain a HEPES Good buffer (ph7.
4, μ0.15) to obtain a test material.

Test method (1) Experimental animal: BALB / c mice (female, 6 weeks old) were used in a group of 5 mice. (2) Transplantation of Sarcoma 180 solid tumor into mice: Sarcoma (sarcoma) 180 cell line was cultured in a 37C / 5% C02 incubator at 10% FCS.
/ RPMI-1640 culture solution. Logarithmic growth phase (Full
After removing the cells from the culture flask using Trypsin-EDTA, wash with RPMI-1640 culture solution and count the number of cells.
The one adjusted to 5,000,000 cells / mL was used. This 0.1 ml was implanted subcutaneously at the back of each test mouse. All operations were performed aseptically. (3) Method of Administration of Extraction Components From 24 hours after transplantation of Sarcoma180 solid cancer cells into mice, the test material sterile-filtered with a 0.22 μm filter was applied once a day (0.1 ml) for 14 days each. Intraperitoneal (ip)
Was administered.

Test Results Mice were sacrificed three weeks after tumor cell transplantation, the engrafted tumor weight (g) was measured after transplantation, and the results of calculating the tumor suppression rate (%) as compared with the control group (control) are shown in the table. It is shown in FIG. The control group used here was Na-Phosphate Buffered Sa
Line (PBS) was administered.

[0056]

[Table 1]

As is clear from the results, when the health food according to the present invention was administered, the tumor suppression rate was clearly higher than that in the control group. In addition, when the health drink according to the present invention is used, compared with Agaricus mushroom which has been conventionally recognized as having an antitumor effect, a small amount (about 1/30 of the amount based on the amount of money) has the same tumor suppression effect. Was obtained.

[Tumor Inhibition Test 2 (Inhibition of Hematological Tumor Cell Proliferation)] Preparation of Cells The following cell lines (1) to (6) were cultured in a 37C / 5% CO ₂ incubator in RPMI1640 containing 10% FCS. The suspension growth culture was performed in a 25 cm ² culture flask under a liquid environment. When all cells were in the logarithmic growth phase, they were washed and replaced in an RPMI1640 culture medium environment to adjust the cell number to 400,000-800,000 cells / mL.

(1) CCRF-CEM cells: Acute lymphoblastic
leukemia-T lymphocyte (Human-lympoblast) (acute lymphocytic leukemia) (2) K562 cells: Chronic myelogenous leukemia-bone
marrow (Human-lymphoblast) (chronic osteomyelitis) (3) MOLT-4CL # 8 cell: Acute lymphoblastic leukemia
-T lymphoblast (Human-lymphoblast) (Acute lymphocytic leukemia) (4) U937 cells: Histocytic lymphoma-macrophage, h
istiocyte (Human-monocyte) (tissue lymphoma) (5) Raji cells: Burkitt lymphoma? EBV infected B lym
phocyte (Human-lymphoblast) (Burkitt lymphoma) (6) YAC-1 cells: Natural killer cell-sensitive lym
phoma (Mouse-lymphoblast) (NK cells)

Reaction between tumor cells and test material Starting from the test material obtained from the healthy drink of Example 1 of the present application used in the above-described mouse antitumor experiment, a 10-fold diluted solution (1% stock solution) A 20-fold, 100-fold diluted solution was prepared with Na-Phosphate Buffered Saline (PBS), and this was sterile-filtered with a 0.22 μm filter to obtain a test material. Further, each of the prepared cell suspensions (1) to (6) was placed in a 96-well cell culture plate for 10 times.
The test material was added in equal amounts of 0 μl each, and an equal amount of the test material was added thereto, mixed well, and then subjected to reaction culture again in a 37C / 5% CO ₂ incubator for 36 hours. Next, each of these cultured cells was stained with 0.5% Trypan blue, the number of viable cells after the reaction was counted, and compared with Control (PBS) to obtain an antitumor (growth inhibitory) activity against each tumor cell. Calculated.

Test Results Table 2 shows the results of comparing the number of living cells and the ratio of living cells for each test material at each concentration to each tumor cell with the control (Control).
Shown in As is clear from the results, the health food according to the present invention has an antitumor effect on blood tumor cells.

[0062]

[Table 2]

[Type I Allergic Reaction Inhibition Test] Test Feed The amount of the health drink obtained in Example 1 corresponding to the daily intake (40 ml for a body weight of 60 kg) of humans and mice was determined.
(Approximately 20 g). About 25-30 of this calculated amount
After evenly mixing in normal mouse breeding powder so that the doubled amount becomes the intake concentration (oral administration experiment setting concentration),
It was used as a test feed after solid stamping and radiation sterilization. The substantial concentration in the mouse diet is about 5%. Control (con
trol) group received only normal mouse sterilized solid feed.

Experimental animals Spontaneous atopic dermatitis mice (NC / Nga mouse, clea
n, CV; obtained at 4 weeks of age from SLC Japan)
The test was performed in a group of 10 males and 10 females each, and the test feed administration group and the control group described above were performed in a total of 4 groups. All the experimental animals were pre-bred for one week after arrival, and the observation period was from the 5th week to the 16th age.

Test method Rearing conditions are 24 ± 1 ° C., relative humidity 55 ± 5%, dark and light for 12 hours each
(Lighting time: 7:00 am to 7:00 pm), the number of ventilations per hour with fresh air sterilized by a hepafilter was 12 per hour.
The animals were bred in a barrier system animal breeding room (mouse, rat room) that was set more than once. For breeding, 4 to 6 animals were bred in a sterilized plastic cage having a sterile bedding. The cage is changed once a week, and the feed is solid feed (Oriental Yeast Co., Ltd., CFR-1) in the feeder,
As for drinking water, tap water was put in a sterilized water supply bottle, and each was freely taken and bred.

The blood IgE level was measured at 1 week (6 weeks), 4 weeks (9 weeks), 7 weeks (12 weeks) and
After 11 weeks (16 weeks of age), blood was collected from the vein layer of the fundus of the mouse four times, and the change over time in the total amount of IgE in the obtained serum was determined by the sandwich ELISA method using a specific antibody against mouse IgE (enzyme antibody). Method). In addition, gross skin findings were compared and observed 11 weeks after administration (16 weeks of age).

Test Results Table 3 shows the variation of blood IgE concentration with age in male and female mice. Although the total amount of blood IgE concentration tended to increase with the growth of solids, the results showed that the test feed intake group showed a significant suppression of blood IgE production compared to the control group. Was. In particular, 11 weeks after administration (1
As for the blood IgE value at 6 weeks of age, a statistically significant t-test was performed in both male and female groups.

[0068]

[Table 3]

In the gross skin findings at 16 weeks of age (usually allergic spontaneous age), the control group showed dermatological symptoms, while the test feed group showed normal appearance. As compared with the control group, the tendency of suppressing skin allergic symptoms was clearly observed in the test feed intake group.

From these results, it was revealed that the health drink of the present invention is effective for suppressing the onset of type I allergic reaction in mice, and that it is also useful for suppressing the onset of allergic reaction when humans take equivalent components. Was done. Although this result is a result of drinking, it is also suggested that it may be useful for suppressing the onset of allergic reaction even in the use of a skin application as described below.

[Antibacterial activity test] Specimen Specimen 1; Aleobasidium culture solution cultured in the same manner as in Example 1 and 0.1% by weight of “Applephenone (registered trademark)” manufactured by Nikka Whiskey Co., Ltd. Specimen 2; Only Aureobasidium culture solution cultured as in Example 1

Test bacterium Test bacterium 1; Staphylococcus aureus IFO 12732 Test bacterium 2; Klebsiella pneumoniae IFO 13277

Test Method To each of 10 ml of each of the specimens 1 and 2, the bacterial solution of each of the above-mentioned test bacteria 1 and 2 was added at about 100,000 cells / ml and stored at 35 ° C. After and 24 hours later, the number of viable bacteria in the test solution was measured. In addition, as a control, only the bacterial solution was similarly stored, and the viable cell count was measured.

Test results Table 4 summarizes the test results. As is clear from the table, the specimen 1 containing β-1,3-1,6-glucan,
2 shows antibacterial activity, indicating that it has an antibacterial effect against any test bacteria. Moreover, it is clear from the viable cell count at the stage after 6 hours that the sample 1 containing the apple extract further has a stronger antibacterial activity.

[0075]

[Table 4]

As described above, it was confirmed that the combination of the apple extract was effective in preventing the composition containing β-1,3-1,6-glucan from decay due to propagation of various bacteria. is there. From these results, it can be inferred that the combination of the apple extract has a putrefaction-preventing effect in both beverage use and skin application.

Example 2 Next, an example in which the composition of the present invention is used as a skin coating agent will be described. The ratio of the blended raw materials is as follows.・ 60% by weight of the same culture solution as in Example 1 ・ Immature apple extract 0.1% by weight “Applephenone (registered trademark)” manufactured by Nikka Whiskey Co., Ltd. ・ 5% by weight of horse oil (melting point 8 ° C.) “Liquid” manufactured by Ikko Chemical Co., Ltd. Horse oil W "-Green tea extract (main component; green tea catechin) 0.05% by weight" Sanphenone (registered trademark) 100S "manufactured by Taiyo Chemical Co., Ltd.-Dipotassium glycyrrhizinate 0.1% by weight-Allantoin 0.1% by weight-Rose extract 20% by weight -Methyl paraoxybenzoate 0.2% by weight-Ethyl paraoxybenzoate 0.05% by weight-Carboxyvinyl polymer 0.8% by weight F. "Carbopol ULTRES-10" manufactured by Goodrich Chemical Co., Ltd.-Potassium hydroxide 0.2% by weight-Glycerin 5 parts by weight-Purified water 8.4% by weight

To the purified water, the same culture solution as in Example 1, dipotassium glycyrrhizinate, allantoin, methyl parahydroxybenzoate, ethyl paraoxybenzoate and glycerin were added and dissolved by heating to 80 ° C. This was once cooled, and a carboxyvinyl polymer was added. Further, potassium hydroxide was added and the mixture was stirred to gel. Next, the apple immature extract and green tea extract dissolved in a rose extract were added and mixed. Finally, horse oil was added and mixed well, and then filled into a container. The pH of the obtained skin application agent was 7.

[Application Test] An application test of the skin application agent of the present invention was conducted in cooperation with 10 patients with atopic dermatitis. The patient is a patient with mild and moderate atopic dermatitis and a pruritus score described later of 2 (mild) or more.

The affected area was directly rubbed with an appropriate amount of the skin application preparation of Example 2 once to several times a day. At this time, rubbing after bathing or before going to bed was essential. The administration period is 4 weeks, 2
Each week, clinical symptoms and the presence or absence of adverse events were confirmed and evaluated. However, three out of ten persons were not evaluated after four weeks. Regarding clinical symptoms, a score was evaluated for each item on a 5-point scale based on the following criteria.

Itching It was judged according to the following criteria by self-report of the patient. 0; no itching during the day. No itching at night. 1: It is not necessary to be patient during the day. Sleeps well at night but is slightly itchy. 2: During the day, sometimes scratching does not matter. At night, itching can be a bit irritated and subsides. 3: Irritated and constantly scratching during the day. At night, I wake up or sleep and scratch. 4: Can't stand during the day. At night, itchy and unable to sleep.

Flushing 0; none 1: slight (only slight redness) 2: mild (slightly reddish with slight edema) 3; moderate (clear reddish with slight edema) 4; advanced (presents a bright red or purple-red color with noticeable edema)

Papules 0; none 1: slight (slight papules) 2; mild (small papules) 3; moderate (between mild and altitude) 4; Collectively)

Dry roughening (keratin keratinization) 0; None 1: Minor (between none and mild) 2; Mild (slightly roughened skin, with slight keratinization consistent with pores) 3; Moderate (between mild and altitude) 4; Altitude (very rough skin, conspicuous keratinization of pores)

· Skin peeling (scratch, scratch marks) 0; none 1; slight 2: mild 3; moderate 4: high

Scale: 0; None 1: Minor (slight scale is observed) 2; Mild (skin is slightly rough and pathological keratin is slightly flattened from the skin surface) 3; Moderate (whole skin) 4) Altitude (state in which the whole skin is dry, and the pathological keratin rises from the skin surface and peels off)

Table 5 shows the test results. As is evident from the test results, the scores of the evaluation scores decreased in the items of pruritus, flushing, papules, dry roughening, and scales, and were flat in the exfoliation. As an adverse event, only one (C) complained of dry skin. Thus, it was confirmed that the skin application preparation of the present invention was effective in improving various symptoms of atopic dermatitis.

[0088]

[Table 5]

[Preservation test] The skin coating preparation obtained in Example 2 was stored in a sealed thermostat (dark place) at 40 ° C.
The appearance of the contents was observed every month. As a result, 1
There was no significant change in appearance and no separation was observed at any of the three months, two months and three months later. 40 ℃
1 month is equivalent to about 3 months at room temperature.
It was found that the skin application obtained in Example 2 had sufficient storage stability.

[0090]

EFFECTS OF THE INVENTION By blending an apple extract with a composition containing β-1,3-1,6-glucan, rot and oxidation during storage can be prevented, and antioxidant and antioxidant properties in the body can be prevented. Allergic effects can also be enhanced. A composition useful for a beverage or a skin application can be provided.

──────────────────────────────────────────────────続 き Continued on the front page (51) Int.Cl. ⁷ Identification symbol FI Theme coat ゛ (Reference) A23L 2/38 A61K 9/06 4C088 A61K 9/06 9/08 9/08 31/375 31/375 31 / 716 31/716 35/12 35/12 35/66 35/66 35/78 H 35/78 A61P 3/10 A61P 3/10 9/10 9/10 103 103 103 11/06 11/06 13/02 13 / 02 17/00 17/00 19/02 19/02 29/00 101 29/00 101 31/12 31/12 31/18 31/18 35/00 35/00 37/00 37/00 37/08 37 / 08 A23L 2/00 F (72) Inventor Taku Harada 2438-74 Nishi-Oshima, Kasaoka City, Okayama Prefecture (72) Inventor Nashiro Harada 2438-74 Nishi-Oshima, Kasaoka City, Okayama Prefecture F-term (reference) 4B017 LC03 LG01 LK13 LK16 4B018 LB08 MD25 MD33 MD52 ME06 ME07 ME13 4C076 AA06 AA11 BB01 BB31 CC21 CC22 CC40 FF68 4C086 AA01 AA02 BA18 EA20 MA02 MA03 MA04 MA17 MA28 MA52 MA63 NA05 NA10 NA14 ZA36 ZA40 ZA59 ZA66 ZA83 Z A89 ZB13 ZB15 ZB26 ZB33 ZC35 ZC55 4C087 AA01 AA02 BC05 MA02 MA17 MA28 NA05 NA10 ZA36 ZA40 ZA59 ZA66 ZA81 ZA83 ZA89 ZB13 ZB15 ZB26 ZB33 ZC35 ZC55 4C088 AB51 BA12 CA03 MA03 Z03A17 ZB26 ZB33 ZC35 ZC55

Claims (7)

[Claims] 1. A composition comprising β-1,3-1,6-glucan and an apple extract. 2. The composition according to claim 1, wherein the β-1,3-1,6-glucan is obtained by culturing a microorganism of the genus Aureobasidium. 3. The composition according to claim 1, wherein the apple extract is extracted from immature fruits of apples. 4. A beverage comprising the composition according to claim 1. 5. The composition further contains vitamin C to adjust the pH to 4 to 6.
The beverage according to claim 4, wherein the beverage is adjusted to: 6. A skin application comprising the composition according to claim 1. 7. The skin application according to claim 6, further comprising horse oil.