JP2003210136A - Method for producing health nutritive food - Google Patents

Abstract

<P>PROBLEM TO BE SOLVED: To provide a method for producing liquid health nutritive food that can produce a liquid health nutritive food that can exactly develop health care action by selecting a strain that can efficiently assimilate a substrate as a fermentation microorganism. <P>SOLUTION: A saccharide prepared by hydrolyzing starch including corn starch with amylase is used as the substrate for a culture medium, to which vegetable juices and yeast as nitrogen sauces are added to prepare the culture medium for the fermentation. The culture medium is inoculated with a fermentation speed including Bacillus subtilis AK (FERM P-18291) (Bacillus natto) and a lactic acid bacterium, fermented and ripened. Then, the formed liquid component is separated as a food concentrate. The prescribed Bacillus natto efficiently assimilates saccharides and the nitrogen source in the prescribed fermentation medium to produce a variety of amino acids, lipoproteins, lipopolysaccharides, lipids, and the like, having a variety of activities. <P>COPYRIGHT: (C)2003,JPO

Description

Detailed Description of the Invention

[0001]

BACKGROUND OF THE INVENTION 1. Field of the Invention The present invention relates to a method for producing a health and nutrition food, which is a food that has a positive effect on health promotion.

[0002]

2. Description of the Related Art As a health and nutritional food for promoting human health, it enhances metabolic activity so that various components necessary for the living body can be efficiently produced in the body and at the same time, immunizes the immune function by blasting T lymphocytes. Enzamine (trade name) is well-known as a fermented metabolite extract which is considered to have the above-mentioned action, and its production method is disclosed in JP-A-54-160787.

[0003] According to the same publication, a beverage stock solution containing an in vivo metabolic regulation factor is prepared by sufficiently digesting yellow corn starch with amylase, then squeezing baker's yeast and vegetable squeezed juice under high pressure, and cooling the medium. Using this medium, inoculate the fermented bacteria such as cultivated Bacillus natto, koji mold, lactic acid bacteria, etc., ferment at constant temperature for 2 months or more, ripen for 6 months or more, and collect the clarified filtrate. Can be manufactured by.

[0004]

However, in the above-mentioned conventional method for producing a health and nutrition food, since there is no disclosure of fermentative fungi that efficiently assimilate the substrate, the production efficiency is sufficiently high. However, there is a problem in that a nutritional food that reliably exerts health effects cannot be efficiently produced.

Therefore, an object of the present invention is to solve the above-mentioned problems, to employ a strain that efficiently assimilates a base material as a fermentative fungus, and to provide a health and nutrition food that more reliably exerts its health action. It is a manufacturing method.

[0006]

In order to solve the above problems, according to the present invention, a saccharified product obtained by hydrolyzing starch containing corn starch with amylase is used as a medium substrate, and vegetable juice and yeast as a nitrogen source are added to the medium. The fermentation medium was adjusted by addition, and fermented bacteria containing Bacillus subtilis AK (FERM P-18291) and lactic acid bacteria, which are Bacillus natto, were inoculated into this medium, fermented and aged, and then the produced liquid component was collected. Then, a method for producing a health and nutrition food, which comprises preparing an undiluted solution for food, was adopted.

According to the production method of the present invention configured as described above, the amino acid having various activities as a fermentation product, in which the predetermined Bacillus natto efficiently assimilates sugar and nitrogen source in the predetermined fermentation medium, It produces lipoproteins, lipopolysaccharides (lipopolysaccharides), and lipids. The amino acids (short peptides) and the like thus obtained are considered to contain a large number of factors that are active for regulating cells in vivo, and as is clear from the experimental results described below, antidiabetic (Insulin secretagogue), hypertension suppressant (angiotensin converting enzyme inhibitor), immune enhancing (mucopolysaccharide activation of T lymphocytes), diet effect (sugar absorption suppressant and lipid metabolism promoting), antibacterial It is recognized that the sex is surely exhibited.

And Bacillus subtilis AK,
A starch-containing starch containing corn starch is hydrolyzed with amylase as a base material, and it grows efficiently in a fermentation medium in which yeast is added as a vegetable juice and nitrogen source. It is resistant to growth by X-ray irradiation, the presence of competing lactic acid bacteria, and even in severe environments and conditions such as high and low temperature environments, and it has the property of being seen as a mutant strain or subspecies.

In order to produce a health and nutrition food that more reliably exerts health effects, the fermentation conditions for fermenting bacteria containing Bacillus tin butyris AK and lactic acid bacteria are from pH 4.5 to
It is preferable to ferment under conditions of 6.5 and 28 to 32 ° C. for 2 months or more, and aging conditions are pH 4.0 to 6.0 and 13
It is preferable to age for 4 months or more under the condition of -17 ° C.

[0010]

BEST MODE FOR CARRYING OUT THE INVENTION The saccharified product used as a substrate for a medium is a starch containing corn starch hydrolyzed with amylase, and corn starch is a protein separated from corn (grain) and purified. More preferably, crude yellow corn starch is used. As the starch other than corn starch, soybean, rice bran and the like can be adopted.

The vegetable juice used in the present invention is not particularly limited to the kind of vegetable, but it is preferably a vegetable containing saponin or S-allyl-L-cysteine, if possible. Specific examples of such vegetables include Chinese cabbage, cabbage, carrots, medicinal carrots, parsley, celery, and onions. Vegetable juice is a juice whose main component is water squeezed out by squeezing these vegetables. For example, this is subjected to high-pressure sterilization treatment at about 120 ° C. for about 20 minutes before being used as a medium substrate. Added.

Yeast used as a nitrogen source is
Single cell (eukaryotic) that can be used for fermentation and brewing of food
It is a microorganism, and the genus Saccharomyces is a typical example. The yeast to be used as the nitrogen source may be dead cells, yeast extract or dry powder.

As described above, in addition to the saccharified product obtained by hydrolyzing starch with amylase, vegetable juice and yeast, proteins, inorganic salts and the like may be added to the substrate for the medium, if necessary. Such inorganic salts include calcium chloride,
Examples thereof include sodium chloride and sodium phosphate.
Examples of proteins include soybean protein and other vegetable proteins.

It is also preferable to further add sugar to the saccharified product obtained by hydrolyzing starch with amylase, for example, sucrose (granulated sugar), glucose (dextrose), starch syrup and the like.

The fermenting bacterium inoculated into the fermentation medium thus prepared is Bacillus subtilis which is a natto bacterium.
It is a fermenting bacterium containing AK and lactic acid bacteria.

[0016] Bacillus subtilis AK can be treated with ultraviolet rays, X, against Bacillus subtilis which is an ordinary natto bacterium.
Line, high / low temperature environment (100 ° C, 0 ° C), competition with lactic acid bacteria, medium that easily forms spores [meat extract 5.0, peptone 10.0, sodium chloride 5.0, agar 15.0, vegetables (cabbage) , Carrots, celery, parsley), and the ratios are all parts by weight], and resistant bacteria that can grow without dying even under these harsh environments and conditions are found, and subcultured. It was obtained by repeatedly selecting.

The mycological properties of the above strains are as follows. (a) Morphological properties Cell shape and size Bacteria 1.0 to 1.2 × 3.0 to 50 μm Presence or absence of polymorphism in cells Presence or absence of motility (peripheral flagella) Presence or absence of spores Oval cells Almost middle (b) Culture property Meat broth agar plate culture Round colony White turbid broth liquid culture Upper or lower microbial aggregate (c) Biochemical properties Gram stain Positive nitrate reduction Positive MR test Negative VP test Positive Indole formation Negative hydrogen sulfide Negative Use of citric acid Positive catalase Positive Growth range pH 5.5 to 7.0 Temperature 25 ° C to 40 ° C This strain was deposited at the National Institute of Advanced Industrial Science and Technology as "(deposit number) FERM P-18291" There is.

The lactic acid bacteria used in the present invention include Lactobacillus, which is a lactic acid bacillus, and Streptococcus, which is a lactic acid coccus, and it is needless to say that it is not limited to pathogenic fungi.

The fermentation conditions preferably used in the present invention are pH 4.5 to 6.5 and 28 to 32 ° C. for 2 months or more. It is presumed that the specified Bacillus natto used in the present invention does not efficiently utilize sugars and nitrogen sources even if fermented for 2 months or more under conditions of stronger acidity than the above and below a predetermined temperature, and the obtained foods have The desired effect cannot be obtained sufficiently. In the neutral or alkaline conditions above the above acidic range and above a predetermined temperature at a high temperature, fermentation is insufficient at a high temperature and the specified Bacillus natto does not efficiently assimilate sugar and nitrogen sources, and the obtained health food is Similarly, the desired effect cannot be obtained sufficiently.

The aging conditions preferably used in the present invention are pH 4.0 to 6.0 and 13 to 17 ° C. for 4 months or more. In the stronger acidic region and under the temperature lower than a predetermined temperature, amino acids, lipoproteins, lipopolysaccharides (lipopolysaccharides), lipids and the like having various activities as fermentation products have sufficiently low molecular weight even if they are aged for 4 months or more. It is presumed that it does not occur, and the desired effect cannot be obtained on the obtained food. It is presumed that the activity will decrease even if it is aged at a high temperature above a predetermined temperature under a neutral or alkaline condition exceeding the above acidic range, and the health and nutrition food obtained has sufficient expected effects as above. It will not be possible.

In order to separate the produced liquid component, well-known separation means such as filtration or centrifugation may be adopted.
The obtained undiluted solution for foods can be used as it is, but can be concentrated or diluted as needed before use.

By the way, the health food obtained by the production method of the present invention stimulates life phenomena to enhance and promote various functions in the living body, strongly maintains homeostasis of the living body, and promotes health. It is intended for the purpose of including the substance for facilitating in-vivo enzyme synthesis, that is, the fragment obtained by reversibly cleaving the enzyme obtained by fermentation into an active amino acid residue. ,
In addition, biocytin, mevalonic acid, lipoic acid, adenine,
It contains useful substances that can be utilized in vivo such as guanine, cytosine, thymine, uracil, ergosterin, and mannan (yeast mannan). Such useful substances include an anti-virus tissue-activating factor proposed by Besredka, a life source stimulant described by Filatov, which are combined by a biochemical reaction to act safely and effectively. It is considered to be the culture filtrate treated as above.

The enzymes produced by the fermentation process in the culture filtrate are listed below by type. Redox enzyme: glucose oxidase, catalase, peroxidase, cytochrome oxidase Hydrolase: pectinase, pectinesterase, tannase, phosphatase, lactase, invertase, α-amylase, β-amylase, cellulase, hemicellulase transferase: hexonase, aminotransferase , Transaldolase Peptide hydrolase: Exopeptidase, Endopeptidase, Aspergillopeptidase, Other proteases Isomerase: Phosphotriose isomerase, Phosphoriboisomerase, Glucose isomerase Synthase: Acetyl CoA ligase Liase: Pyruvate decarboxylase, Fumarase , Aspartase

[0024]

EXAMPLE First, 2.3 kg of yellow cornstarch,
Soybean peptone 0.5 kg, rice bran juice 0.5 kg, calcium chloride 80 g, salt 150 g, purified water 50 kg,
It was heated and dissolved. It is then cooled and amylase 4
0 g was added for sufficient saccharification. After saccharification, 1.5 kg of granulated sugar, 1.5 kg of glucose (glucose),
450 g of yeast extract, 1.5 kg of starch syrup, 80 g of sodium phosphate, 5 kg of vegetable juice (cabbage, carrot, celery, parsley) 5 kg, and purified water were added to bring the total amount to 150 kg.

Then, the pH is adjusted by adding sodium hydroxide.
Was adjusted within the range of 7.3 to 7.8, and this was put in a culture can and subjected to high-pressure sterilization at 120 ° C. for 20 minutes. After cooling this, Bacillus subtilis AK strain was inoculated at a temperature of 30
Fermentation is performed in a temperature-controlled room of ± 2 ° C at a pH of 4.5 to 6.5 for 60 days, and then in a temperature-controlled room of 15 ± 2 ° C to a pH of 4.0 to 6.
The culture was clarified by aging it for 180 days under the condition of 0. This was filtered through a filter to obtain 125 liters of liquid stock solution of health food and nutritional food (hereinafter referred to as culture filtrate).

The general analysis results in 100 g of the obtained health food and nutritional food concentrate are shown in Table 1 below. The symbol φ in the table
Indicates a trace amount below the detection limit.

[0027]

[Table 1]

With respect to the above culture filtrate, a column (TSKgel G2500PWXL) manufactured by Tosoh Corporation was used, and the mobile phase was 55: 45: 0.
High-performance liquid chromatogram for 1 mixture (Shodex:
Size exclusion chromatography (SE on GPC SYSTEM-21)
C) and the detector sensitivity at that time (UV spectrophotometer:
mV) is compared with the elution time of a standard with a known molecular weight, and a diagram of the molecular weight distribution is shown in FIG. 1, and the ratio (percentage) of the area of the molecular weight fraction in this diagram to the whole is shown in Table 2.
It was shown to.

[0029]

[Table 2]

Furthermore, in order to confirm the safety of the obtained culture filtrate, an oral acute toxicity test was conducted using Wistar rats. The results are as follows.

A 4 × concentrated solution is converted into a male salary 14.0, 4
Even if 2 ml / kg, female 14.0, 42 ml / kg (10 times or 30 times the usual amount) were administered, neither male nor female died, no intoxication symptoms were observed, and body weight was generally changed. The condition also remained clean during the test period, and the blood test area was in the normal area. The dose of 10 ml / kg is considered to be the highest dose for rats, and when the test sample is converted to the usual dose, the oral LD50 value is 4 for both males and females.
It was estimated to be 2.0 ml / kg or more.

Next, the antibacterial properties of the culture filtrate obtained as described above, the effects of administration on hypertensive rats, and the effects of administration on artificially induced diabetic rats were examined. The test methods and results are shown below.

[Antibacterial Properties of Culture Filtrate] The strains used and the medium for measurement thereof are as follows. (1) Escherichia coli (ATCC25922): DHL agar medium (2) Salmonella Enteritidis (ATCC25923): DH
L agar medium (3) Klebsiella pneumoniae (ATCC13883): BTB
Preparation of culture medium: 30 ml of tryptobroth and 100 preculture liquid
Add μl. The cells are cultivated with shaking at 35 ° C., stopped at an appropriate absorbance, and diluted with a phosphate buffer so as to obtain 10 ml of a bacterial suspension of 10 ⁵ CFU / ml. Sterilization operation: The prepared bacterial solution was centrifuged at 3000 rpm for 15 minutes,
After replacing the supernatant with an equal amount of the test substance, shake reaction is performed at 35 ° C. for 2 hours. As a control, a bacterial suspension containing nothing is placed under the same conditions. Application / culture / counting of sample: 100 μl of a sample serially diluted with a phosphate buffer was applied to 2 sheets of each measuring medium with a conradi stick and cultured at 35 ° C. for 48 hours, and grown colonies were counted. The results are shown in Table 3 below.

[0034]

[Table 3]

[Effects of Administration on Hypertensive Rats] A male spontaneously hypertensive rat (SHR) judged to be healthy after a quarantine / training period of 5 days was divided into 3 groups with 7 rats aged 6 weeks. , Test group-1 had an equal volume of the culture filtrate, test group-2 had a 10-fold volume, and the control group had distilled water in a water bottle and was allowed to freely ingest with a predetermined feed for 28 consecutive days. . Blood pressure was measured simultaneously at weekly (7 days) intervals, and the results are shown in Table 4.

[0036]

[Table 4]

As is clear from the results in Table 4, the culture filtrate-administered group showed suppression of increase after 2 weeks as compared with the control group.

[Effect of Administration on Artificially Induced Diabetic Rats] A male Wistar ST strain rat (6 weeks old) judged to be healthy after a quarantine / training period of 7 days was given 1 kg of streptozotocin. Inject 40 mg per tail into the tail vein, measure the blood glucose level from the next day, divide 5 animals into 2 groups so that the blood glucose level will be almost constant, and add the human culture filtrate for 14 days to the test group (a). About 10 times the dose was placed in a water bottle and continuously orally administered.

[0039]

[Table 5]

As is clear from the results in Table 5, the culture filtrate-administered group showed a slight tendency to suppress the increase from the 6th day. On the other hand, the control group (b) had an increase rate of about 3.5 times after 15 days, which was about 80% higher than that of the culture filtrate administration group. From these tendencies, it was suggested that the group to which the culture filtrate was administered showed a tendency to suppress the increase in blood glucose level, and had a good influence on the prevention of diabetes and the suppression of the progression.

Tables 6 to 9 show blending examples of health drinks, health foods in capsules, lotions, and nutrients for animals and plants produced by using the stock solutions of health and nutritional foods obtained below.

Health drinks were prepared by mixing 8 to 16% of the culture filtrate with other raw materials as shown in Table 6.

[0043]

[Table 6]

The capsule-shaped health food was prepared by concentrating the culture filtrate 20 times and blending other raw materials as shown in Table 7.

[0045]

[Table 7]

The lotion was prepared by blending 8% of the culture filtrate with other raw materials as shown in Table 8.

[0047]

[Table 8]

The nutrient for animals and plants was prepared by mixing 20% of the culture filtrate with the other raw materials shown in Table 9.

[0049]

[Table 9]

[0050]

As described above, the present invention efficiently utilizes sugar and nitrogen sources in a predetermined fermentation medium with a predetermined Bacillus natto, and ferments and ripens them under predetermined conditions to produce a health and nutrition food. It has an advantage that it can be produced efficiently, and in particular, a predetermined strain is adopted as a fermentative fungus, whereby a health action can be more reliably and efficiently provided as a liquid health and nutrition food.

[Brief description of drawings]

FIG. 1 is a diagram showing the relationship between detector sensitivity (mV) and elution time (minutes) in size exclusion chromatography, and the molecular weight distribution of culture filtrates analyzed from this.

Claims (3)

[Claims] 1. A saccharified product obtained by hydrolyzing starch containing corn starch with amylase is used as a base material for a medium, and a vegetable juice and yeast as a nitrogen source are added to the medium to prepare a fermentation medium. Bacillus subtilis
A method for producing a health and nutrition food, which comprises inoculating a fermenting bacterium containing AK (FERM P-18291) and lactic acid bacteria, fermenting and aging, and then collecting the produced liquid component to obtain a stock solution for food. 2. Fermentation of pH 4.5-6.5, 28-3
The method for producing a health and nutrition food according to claim 1, wherein the fermentation is carried out at 2 ° C for 2 months or more. 3. Aging is carried out at pH 4.0 to 6.0, 13 to 1
The method for producing a health and nutrition food according to claim 1 or 2, which is aging at 7 ° C for 4 months or more.