JPH03244367A - Food additive for enhancing immunological function - Google Patents

Abstract

PURPOSE:To prevent the infection to digestive system, respiratory system, etc., of human by preparing a food additive for enhancing immunological function by removing capsule of the cell of bacteria belonging to genus Bifidobacterium and using the treated cell as an active component and adding the additive to a food. CONSTITUTION:Cells of bacteria belonging to genus Bifidobacterium, Bacillus, Corynebacterium, etc., are subjected to a treatment with a surfactant or a stirring or shaking treatment without using yeast, etc., to obtain bacterial cells free from capsule. The obtained cell has an activity to enhance the immunologi cal function of human, especially infant. A food effective in enhancing immuno logical function and preventing infections diseases of digestive system and respi ratory system can be prepared by adding an active amount of the cell to a food. The amount of the agent administered to a person per day is usually about 10 mug to 1 mg per 1 kg body-weight.

Description

DETAILED DESCRIPTION OF THE INVENTION [Industrial Field of Application] The present invention relates to a non-toxic immune-enhancing food additive that has the effect of enhancing the immune function of humans, especially infants.

[Conventional technology]

In humans, especially in infancy and old age, immune organs and immune cells generally have weak functions, and infections of the digestive system or respiratory system, such as diarrhea, often occur.

Currently, various antibacterial agents such as antibiotics and sulfa drugs are being used to prevent and treat these diseases, but their effectiveness is not necessarily sufficient. There is a tendency to be limited by the persistence of drug-resistant bacteria and the emergence of drug-resistant bacteria.

Therefore, there is a demand for the development of drugs that are not antibacterial agents, such as antibacterial substances, that have the effect of preventing these infectious diseases.

Near hand, BCG (Bacillus Calmatte-
Cell wall components isolated from microorganisms such as Guerin and Nocardia were found to have an immune-enhancing effect, and that component was peptidoglycan ('Journal of National Ca.
ncer In5titude”, Volume 52, 1571
Page, 1974; “Cann, Vol. 69, 66.
Page 9, 1976: “Cancer Immuno
"logy and 1mmunotherapy" 4th
Afterwards, Durham-positive bacteria, which are widely distributed in nature and do not contain endotoxins, attracted attention, and peptidoglycan obtained from some Durum-positive bacteria has already been used in piglets. It has been found that it has an immune-enhancing effect and also has an effect on weight gain (“Japanese
se Journal of Veterinary
5science, vol. 49, p. 235, 1987).

[Problem to be solved by the invention]

Using the above-mentioned Durham-positive bacteria that are widely distributed in nature,
To provide an immunity-enhancing food additive that obtains a substance that enhances immunity in humans, especially infants, and adds this substance to food to enhance human immunity and prevent infections in the digestive system and respiratory system. This is the object of the present invention.

[Means to solve the problem]

The present inventor has previously discovered that the substance has an excellent weight-increasing effect and immune-enhancing effect on livestock and poultry in their infancy, and that the substance can be used only after removing the capsule of Bifidobacterium cells. Therefore, they found that the substance was sufficiently effective, and filed a patent application No. 160969/1983 for a feed containing the same substance.

The present inventor has conducted extensive research with the aim of further contributing to the promotion of human health by using the bacteria from which the capsule has been removed to strengthen the immunity of humans rather than livestock, and has found that Escherichia coli, We have confirmed that this capsulated bacterial cell exhibits protective activity and high immune-enhancing activity against highly pathogenic microorganisms such as Salmonella food-poisoning bacteria and Salmonella typhimurium, and we have obtained test results showing that it is non-toxic. We have completed a food additive that can be added to food and consumed by humans to maintain their health.

The present invention is an immunity-enhancing food additive containing decapsulated bacterial cells as an active ingredient.

The bacteria of the present invention may be any bacteria or yeast, but include Bifidobacterium infantes (Bifidobacterium infantes).
Bifidobacterium bacteria such as B. obacterium 1nfantis, Bacillus subtilis (B.
Bacillus bacteria such as Acillus 5ubtilis, Corynebacterium obis
Corynebacterium bacteria such as Acterium ovis, Actinomyces pyogenes (Acti
Actinomyces bacteria such as L. nomyces pyogenes, Lactobacillus acidophilus (L.
actobacillus acidophilus)
Lactobacillus bacteria such as Streptococcus
Ficaares (Strepzococcus facc)
Streptococcus lK [i and Staphylococcus conii] such as
Bacteria belonging to the genus Staphylococcus such as K. cus cohnii) as well as Kluyveromyces lactis (K.
luyversmyces 1aclis), Saccharomyces cerevisiae (Saccharomyces c
Yeasts such as S. ervisiae) are used. Particularly preferred bacteria are those of the genus Bifidobacterium.

The cells of the present invention from which the capsule has been removed can be obtained by treating the cells with a surfactant or by stirring or shaking the cells without using yeast or the like. Treatment with a surfactant can be carried out immediately by suspending the bacterial cells in a surfactant solution, or by suspending the bacterial cells in advance in water or a buffer solution and adding the surfactant. It can be carried out. In addition, when stirring or shaking, inactivate the autolytic enzyme before performing this operation.
It is advantageous to protect the cell wall of the fungus.

The bacterial cells obtained as described above have their capsules removed and their cell walls exposed.For example, the cell walls of Durham-positive bacteria are mainly composed of peptidoglycan, polysaccharides, or ticoic acid. It is a bag-like structure that is covalently bonded to a structure unique to the bacterium, and it circumscribes the cytoplasmic membrane.
It not only maintains the shape of the bacterial cell but also protects the cytoplasmic membrane and its functions from various external stimuli.

In particular, the peptidoglycan moiety is not only important as the center of the immune-enhancing effect, but also plays an important role in maintaining the shape of the entire bacterial cell. In other words, peptidoglycan has a three-dimensional structure in which glucan chains consisting of N-acetylglucosamine and N-acetylmuramic acid and peptide chains are bonded together in a network, and is water-insoluble as a whole and is physically extremely strong. Forms a giant polymer.

The bacterial cells obtained as described above, from which the capsule has been removed, may be used as they are, or they may be further made into a solid form by freeze-drying or the like and used as the active ingredient of the present invention.

The bacterial cells obtained in the above manner from which the capsule has been removed are made into a powder or fine granules with an edible filler such as starch, or
Alternatively, it can be made into an aqueous solution and used as the food additive for immune enhancement of the present invention.

This immune-enhancing food additive is used to enhance the immunity of infants by adding it to foods such as drinks, soft drinks, confectionery, infant beverages, infant sweets, and the like.

The amount to be used for humans is usually added by an appropriate method so that about 10 μg to 1 mg can be ingested per kg of body weight per day.

Next, examples of the present invention, as well as the immune-enhancing effect and toxicity of decapsulated bacteria, which are active ingredients, will be described.

〔Example〕

Bifidobacterium Infantes (Bifido)
bacterium 1nfantis) ATCC15
697 was cultured by a conventional method, and the obtained wet bacterial cells 10'O
g to 0.05M phosphate buffer (pH 7, O) 1.50
Suspended in water, heated at 70°C for 45 minutes to inactivate the autolytic enzyme possessed by the bacteria itself, cooled rapidly in running water, and heated to 37°C.
The mixture was stirred for 24 hours. Then 20 at 3.500 rpm
Collect the bacterial cells by centrifugation for 1 minute, suspend them once more in purified water, and
．． Cells were collected by centrifugal washing at 50 rpm for 15 minutes. This was freeze-dried to obtain 13 g of Bifidobacterium infantes cells as a dried product with the capsule removed.

The immune-enhancing substance hexosamine (main components are N-acetylglucosamine and N-acetylglucosamine and
-acetylmuramic acid) was analyzed by the Morgan-Elson method and the Dische-Bordenfreund method (indole/hydrochloric acid method), and found to be 2.89, respectively.
% and 3.98%.

The bacterial cells thus obtained were diluted with starch to obtain a food additive.

Using this bacterial cell, an immunity enhancement test against Escherichia coli, Salmonella food poisoning bacteria, and Salmonella typhimurium was conducted.

Test Example 1 The test animals were ICR mice (6 weeks old), and 0 mice were used in each administration group and non-administration group. The attacking microorganism is Escherichia coli (E
Scherichia coli septicemia), Salmonella eoteritd
is, food poisoning-derived strain), Salmon typhimurium (Salmon
ella typhimurium, ATCC131
1 share).

The groups to which the freeze-dried cells prepared in the examples were administered had a bacterial cell amount of 1000 μg, 500 μg, 100 μg, and 50 μg per animal.
There were 5 groups of 25 μg and 25 μg, and a non-administration group (control group).

Dried bacterial cells were administered intraperitoneally 7 days and 2 days before challenge with each microorganism. The amount of attacking bacteria is 2.1 x 1 E. coli.
08 CFU/fish, salmonella food poisoning bacteria 2.4X107
CFU/mouse, Salmonella typhimurium 2.9 x 10'C
Each mouse was inoculated intraperitoneally at FU/mouse. The effectiveness was determined using the survival rate as an index, and the survival rate was calculated using the following formula.

◎: (≧75%, extremely effective) Q: (50-75%, effective) Uni (25-50%, very effective) ×, (≦25%, ineffective) Survival rate = (dry bacterial cell administration Group survival area number 1 control survival group) 3
Number of mice in each group x 100 The results are shown in Table 1 below.

As is clear from the above test results in Table 1, each administration group was extremely effective against E. coli and Salmonella food poisoning bacteria (except for the effectiveness of the 25 μg administration group). For Salmonella Typhimurium ((IOCFU/mouse dies), 50μg
It was ineffective in the groups administered less than 100 μg, but it was quite effective in the groups administered more than 100 μg.

In addition, in the non-administered control group, all the animals died within 1 to 5 days after being challenged with each of the attacking microorganisms.

From the above test, the bacterial cells from which the capsule, which is the active ingredient of the present invention, was removed were found to be effective against highly pathogenic microorganisms such as Escherichia coli, Nano 5 Monella food poisoning bacteria, and Salmonella typhimurium in mouse tests.
It is clear that it has protective activity and high immune-enhancing activity.

Next, we will discuss toxicity tests.

Test Example 2 Male and female IRC mice (6 weeks old, weight 21±2 g) were used as test animals. The bacterial cells obtained in the examples were suspended in sterile physiological saline at concentrations of 5, 10, and 1 per kg body weight.
Each dose was administered daily for 10 days. 10 mice per group were observed for 1 month after administration, and the number of dead mice was determined.

The test results are shown in Table 2.

(Margins below) As is clear from the test results in Table 2 and above, no mice died at each dose, and anaphylaxis, a type of acute toxicity that occurs when antigenic substances are continuously administered, was observed. was also not observed.

〔Effect of the invention〕

INDUSTRIAL APPLICABILITY The present invention is an extremely useful invention that can enhance the immunity of humans, especially infants, without any toxicity, and prevent infectious diseases that they are susceptible to by adding it to foods.

Patent application Takeru Hiki (1 other person)

Claims (1)

[Scope of Claims] 1. An immunity-enhancing food additive containing decapsulated bacterial cells as an active ingredient. 2. The food additive for immunity enhancement according to claim 1, wherein the bacterial cells are bacterial cells belonging to the genus Bifidobacterium.