JP2024007998A - Composition for production promotion of adenylate including atp, adp, and amp and product containing composition for production promotion of adenylate including atp, adp, and amp - Google Patents

Abstract

To provide a new composition for production promotion of adenylate including ATP, ADP, and AMP.SOLUTION: A composition for production promotion of adenylate including ATP, ADP, and AMP includes ultrafine bubbles at a concentration of 1×108/mL or more as an active component.SELECTED DRAWING: Figure 2

Description

The present disclosure relates to a composition for promoting production of adenylate containing ATP, ADP, and AMP, and a product containing a composition for promoting production of adenylate containing ATP, ADP, and AMP.

Patent Document 1 discloses a composition for promoting AMPK activity containing a peptide derived from wakame as an active ingredient. AMPK (AMP-activated protein kinase) is a protein widely present in living organisms, and plays an important role as a regulator of cellular energy metabolism. It is known that when the intracellular ATP level decreases, AMPK activity increases, thereby promoting ATP synthesis and promoting energy consumption.

Japanese Patent Application Publication No. 2018-188404

Conventionally, there are various substances that promote intracellular ATP production, but there is concern that the molecules may cause unintended chemical reactions after their action, resulting in side effects. In addition, methods to improve the reaction efficiency of glycolysis, citric acid cycle, etc. include the use of amino acids, sugars, and enzymes, but all of these methods require expensive materials.

The present disclosure has been made in view of the above problems, and aims to provide a new composition for promoting the production of adenylate containing ATP, ADP, and AMP. The present disclosure can be realized as the following forms.

The composition for promoting production of adenylate containing ATP, ADP, and AMP of the present disclosure contains ultrafine bubbles having a concentration of 1×10 ⁸ bubbles/mL or more as an active ingredient.

According to the present disclosure, a new composition for promoting production of adenylate containing ATP, ADP, and AMP can be provided.

FIG. 1 is a diagram illustrating the presumed mechanism by which ATP production is promoted. FIG. 2 is a graph showing the relationship between UFB concentration and total adenylate amount in Test 1. FIG. 3 is a graph showing the relationship between UFB concentration and total adenylate amount in Test 2. FIG. 4 is a graph showing the relationship between culture time and total adenylate amount in Test 3. FIG. 5 is a graph showing the relationship between UFB concentration and total adenylate increase rate in Test 3.

Here, a desirable example of the present disclosure will be shown.
- The composition for promoting the production of adenylate containing ATP, ADP, and AMP is selected from the group consisting of yeast extract, glucose, sodium chloride, dipotassium phosphate, peptone, casein-sui digested peptone, and soybean-papain digested peptone. It is preferable to contain at least one kind of.
・A composition for promoting the production of adenylate containing ATP, ADP, and AMP can be used for impregnation, injection, dialysis, eye drops, ear drops, enema, drinking, gargling, oral mucosa, external skin use, and nasal absorption. The dosage form may be selected from the group consisting of , and inhalation.
・A composition for promoting the production of adenylate containing ATP, ADP, and AMP can improve blood flow, inner ear disorders, dizziness, tinnitus, hearing loss, ear fullness, swelling, digestive motility, digestive mucous secretion, and fatigue. It may be used to treat, prevent, or improve diseases, symptoms, or functions associated with the production of ATP selected from the following.
・A product containing a composition for promoting the production of adenylate containing ATP, ADP, and AMP is a product containing the above-mentioned composition for promoting the production of ATP, ADP, and AMP, and is suitable for foods, health foods, and beverages. , alcoholic beverages, feed, fodder, pet food, aquaculture water, plant cultivation water, culture solution, cosmetics, pharmaceuticals, and quasi-drugs.

The present disclosure will be described in detail below. In addition, in this specification, descriptions using "-" for numerical ranges include the lower limit value and the upper limit value, unless otherwise specified. For example, the description "10-20" includes both the lower limit value "10" and the upper limit value "20". That is, "10-20" has the same meaning as "10 or more and 20 or less".

1. Composition for promoting the production of adenylate containing ATP, ADP, and AMP The composition for promoting the production of ATP (adenosine triphosphate), ADP (adenosine diphosphate), and AMP (adenosine monophosphate) has a concentration of 1× Contains 10 ⁸ or more Ultra Fine Bubbles (UFB) as an active ingredient. Hereinafter, adenylate containing ATP, ADP, and AMP is also referred to as total adenylate, and a composition for promoting production of adenylate containing ATP, ADP, and AMP is also referred to as a composition for promoting total adenylate production.

(1) Ultra-fine bubbles Ultra-fine bubbles are bubbles with a diameter of less than 1 μm (ISO 20298-1). Ultra-fine bubbles can be prepared according to known methods for producing nano-sized microbubbles. For example, it can be produced by a micropore method, a gas-liquid mixing shear method, a static mixer method, a venturi method, a cavitation method, a vapor condensation method, an ultrasonic method, a swirling flow method, a pressurized melting method, or the like. Among these, from the viewpoint of the usefulness of the obtained composition for promoting total adenylate production, the microporous method is preferable, and the microporous method using a ceramic porous material selected from silica, alumina (γ-alumina), zeolite, etc. method is more preferable.

The gas in the ultra fine bubble is not particularly limited. The gas in the ultra-fine bubble is preferably selected from the group consisting of air, water vapor, hydrogen, tritium, rare gas, oxygen, carbon dioxide, nitrogen, and fluoride gas. Examples of the rare gas include helium (He), neon (Ne), argon (Ar), krypton (Kr), xenon (Xe), and radon (Rn). Examples of the fluoride gas include CF ₄ , C ₂ F ₆ , C ₃ F ₈ , C ₄ F ₈ , CHF ₃ , SF ₆ , NF ₃ and the like. The technology of the present disclosure can be said to be a highly versatile technology that can be applied regardless of the type of gas. The gas in the ultra-fine bubbles is preferably oxygen or water vapor from the viewpoint of the usefulness of the resulting composition for promoting total adenylate production.

The liquid used in the composition for promoting the production of ATP, ADP, and AMP is not particularly limited as long as it can maintain ultra-fine bubbles and is physiologically acceptable (less harmful to cells). Examples of such liquids include pure water, tap water, rainwater, fresh water, seawater, artificial seawater, physiological saline, cosmetics, and beverages, depending on the use of the composition for promoting production of ATP, ADP, and AMP. Liquids, seasonings, alcoholic beverages, culture fluids (glucose-containing fluids, albumin-containing fluids, antibody-containing fluids, vitamin-containing fluids, serum-containing fluids, plasma-containing fluids), infusion preparations (sugar solutions, hypotonic electrolyte solutions (starting solution (1) No. 3 solution), maintenance infusion (No. 3 solution), injection solution, extracellular fluid replenisher (lactated Ringer's solution, acetic Ringer's solution), artificial colloid solution, artificial blood, buffer solution (acetate buffer, phosphate buffer, citric acid) Buffer, citrate phosphate buffer, borate buffer, tartrate buffer, Tris buffer, phosphate buffered saline, McIlvaine buffer), etc. can be used.

The ultra-fine bubble concentration (hereinafter also referred to as UFB concentration) of the composition for promoting total adenylate production is not particularly limited. The UFB concentration of the composition for promoting total adenylate production is 1 x 10 ⁸ cells/mL or more, preferably 1 x 10 ⁸ cells/mL or more and 1 x 10 ¹⁴ cells/mL or less. UFB concentration can be measured using a nanoparticle analysis system. An example of a nanoparticle analysis system is NanoSight (manufactured by Spectris PLC, NS-300, hereinafter simply referred to as NanoSight).

(2) Application of composition for promoting total adenylate production In the present disclosure, promotion of total adenylate production refers to an effect of increasing the production amount of adenylate including ATP, ADP, and AMP per unit number of cells. The increase in the production amount of adenylate containing ATP, ADP, and AMP is thought to be due to the increase in the production amount of at least one of ATP, ADP, and AMP, and the increase in the production amount of ATP directly or indirectly. can contribute. ATP is widely present in living cells and serves as a substance that supplies the cells with the energy they need. ATP is synthesized in sugar metabolic pathways such as glycolysis and the citric acid cycle.
The technology of the present disclosure is mainly based on new findings obtained from tests using Escherichia coli and Bifidobacteria. In tests using Escherichia coli and Bifidobacterium, when the UFB concentration in the medium was 1 x 10 ⁸ cells/mL or more, the total adenylate concentration after a specified period of culture was lower than when it was less than 1 x 10 ⁸ cells/mL. It was confirmed that the amount of adenylate (including ATP, ADP, and AMP) increased. Furthermore, it is known that the amounts of ATP, ADP, and AMP produced by E. coli vary depending on the dissolved oxygen concentration. In this test, it was confirmed that even in a medium with a reduced dissolved oxygen concentration, when the UFB concentration increased, the total amount of adenylate increased. Bifidobacteria are obligate anaerobes and are known to normally lose their activity in the presence of oxygen. In this test, it was found that the rate of increase in the total amount of adenylate increased depending on the UFB concentration. From these findings, it was confirmed that the increase in the total amount of adenylate was not dependent on the dissolved oxygen concentration but was dependent on the UFB concentration. Based on these findings, the present inventors have discovered that ultrafine bubbles act through a mechanism different from that of dissolved gas in promoting total adenylate production in Escherichia coli and Bifidobacteria, and have developed the technology of the present disclosure. reached.
Escherichia coli and Bifidobacterium are known as model organisms having a glycolytic system and a citric acid cycle. The ATP production pathway and the basic structure of biological cell membranes (membrane transport proteins, liposomes, etc.) have many parts in common among many organisms. The technology of the present disclosure is applicable to microorganisms such as Escherichia coli and Bifidobacterium, and is also considered to be a highly versatile technology that can be applied to all organisms including cells that have an ATP production pathway.

The composition for promoting total adenylate production may be an ultra-fine bubble solution containing components other than ultra-fine bubbles. The composition for promoting total adenylate production is revolutionary in that it can be obtained by forming gas into fine bubbles, and that ultra-fine bubbles and other solutes can be used together.

Components other than ultra-fine bubbles in the composition for promoting total adenylate production are not particularly limited as long as they are physiologically acceptable components. The composition for promoting total adenylate production preferably contains components useful for the growth of the target organism from the viewpoint of promoting the production of adenylate. The composition for promoting total adenylate production contains at least one member selected from the group consisting of yeast extract, glucose, sodium chloride, dipotassium phosphate, peptone, casein-sui digested peptone, and soybean-papain digested peptone. Preferably.

In the composition for promoting total adenylate production, ultrafine bubbles may promote the uptake into cells of sugars such as glucose that are degraded in the ATP-producing pathway. From this point of view, it is preferable that the composition for promoting total adenylate production contains sugars that are decomposed in a pathway that produces ATP. Examples of sugars that are broken down in the pathway that produces ATP include glucose, galactose, fructose, mannose, and pentose. Note that even if the composition for promoting total adenylate production does not contain sugar, it is thought that ultrafine bubbles can act on sugar in the body and promote the production of total adenylate. That is, as long as the composition for promoting total adenylate production contains at least ultrafine bubbles, it does not necessarily need to contain sugars that are decomposed in the ATP producing pathway.

The composition for promoting total adenylate production may be administered to any organism that contains cells that have an ATP production pathway, such as microorganisms, animals, plants, and the like. When the subject to be administered is an animal, it may be a mammal or a human. The human subject to administration may be healthy or may be suffering from a disease that is improved by the action of ATP. When treatment of a disease is intended, the composition for promoting total adenylate production is typically administered to subjects suffering from or at risk of suffering from the disease.

The composition for promoting total adenylate production includes ATP, ADP, and It can be used to treat, prevent, or ameliorate diseases, symptoms, or functions associated with AMP production. ATP has a vasodilatory effect, increases blood flow in organs, activates tissue metabolism, and improves these functions. The composition for promoting total adenylate production, like ATP preparations, increases cerebral blood flow, coronary blood flow, gastric artery blood flow, vertebral/common carotid artery blood flow, expands inner ear microvessels, increases gastric motility, and increases gastrointestinal blood flow. It is thought that these effects can be expected to be effective in improving many diseases and symptoms, such as improving mucosal secretion and inner ear dysfunction. In addition, the composition for promoting total adenylate production, like ATP preparations, improves headaches, dizziness, etc. caused by after-effects of head trauma, improves swelling caused by heart failure, and improves eye symptoms caused by asthenopia. improvement of systemic symptoms such as headaches, fever, flickering, etc.), improvement of chronic gastritis (especially improvement of relatively mild symptoms), and associated symptoms such as dizziness, tinnitus, hearing loss, and aural fullness due to vertigo such as Meniere's disease. It is expected that it will be used in a wide variety of clinical settings, such as for the purpose of improving blood flow and diagnosing and treating arrhythmia.

The dosage form of the composition for promoting total adenylate production is not particularly limited. The composition for promoting total adenylate production can be expected to have an effect of promoting total adenylate production in cells by being administered in a manner that allows ultrafine bubbles to come into contact with cells. The composition for promoting total adenylate production is, for example, from the group consisting of impregnation, injection, dialysis, eye drops, ear drops, enema, drinking, gargling, oral mucosal use, skin external use, nasal absorption, and inhalation. The dosage form may be selected.
The state of the composition for promoting total adenylate production is not particularly limited. The state of the composition for promoting total adenylate production may be liquid, gel, or mist depending on the dosage form.

(3) Presumed mechanism The reason why the composition for promoting total adenylate production of the present disclosure can promote the production of adenylate including ATP, ADP, and AMP is not clear, but it is presumed as follows. Note that the technology of the present disclosure is not interpreted to be limited by this reason for speculation.
Membrane transport proteins with pore diameters of several tens of nanometers are present in the cell membranes of living organisms (see Figure 1). Glucose dispersed in extracellular fluid is mainly taken into cells by passive transport via membrane transport proteins. Ultrafine bubbles are charged to a negative zeta potential and are known to adsorb positively charged substances. On the other hand, glucose and liposomes, which are the main components of cell membranes, are known to be positively charged substances. Therefore, it is thought that the following steps promote the uptake of glucose into cells.
Step 1: Glucose adsorbs on the UFB surface.
Step 2: UFB wrapped in glucose adsorbs to the cell membrane surface.
Step 3: Glucose and UFB wrapped in glucose are taken into cells via membrane transport proteins.
Since sugar metabolism is a chemical reaction, it is presumed that when the amount of glucose uptake (substance amount) increases, consumption of glucose is accelerated and production of ATP is promoted.

In addition to the above-mentioned mechanism, ultra-fine bubbles are electrically charged, which causes nutrients to be electrically adsorbed to the bubble surface, causing nutrients to be locally condensed. The dispersibility in the liquid improves due to the influence of electrification, and the concentration distribution of nutrients becomes balanced. When cells take in nutrients through ion channels, they also take in air bubbles, and the air bubbles contribute to metabolic reactions. You can also think of an order.

2. Method for producing a composition for promoting total adenylate production A composition for promoting total adenylate production can be produced by generating ultra-fine bubbles to obtain a liquid containing ultra-fine bubbles at a concentration of 1 x 10 ⁸ bubbles/mL or more. . When the composition for promoting total adenylate production is a solution, it can be produced by mixing a liquid containing ultrafine bubbles at a predetermined concentration or higher with a solution containing other solutes.

The method for producing ultra-fine bubbles is not particularly limited. Ultra-fine bubbles can be generated using, for example, a device equipped with a porous structure (element). Examples of devices equipped with a porous structure (element) include the device described in JP2018-86632A, the device described in International Publication No. 2020/004653, the device described in JP2021-154262A, etc. , the contents of which are incorporated herein.

3. Field of Application The field of industrial application of the composition for promoting total adenylate production is not particularly limited. For example, when plants are to be administered, fields such as agriculture and forestry can be mentioned. When animals are to be administered, fields such as aquaculture and livestock farming can be mentioned. When microorganisms are to be administered, examples include fields such as alcoholic beverages, fermented foods, culture, and biopharmaceuticals. In addition, when the subject is to be administered to humans, fields such as medicine, healthcare, and sports can be mentioned. In addition, cultivation applications include fields such as fermentation processes for foods and alcoholic beverages, biopharmaceuticals, and bioprocesses for regenerative medicine. In addition, for marine applications, it can be applied to aquaculture (growth promotion) of various types of fish. For agricultural and fishery purposes, it can be applied to hydroponic cultivation (growth promotion), etc. Applications to humans include treatment, beauty, health, slimming, training, and other applications that benefit from promoting sugar metabolism. Since ultrafine bubbles can be produced at low cost, they are expected to be used in various fields as a composition for promoting total adenylate production.

The product containing the composition for promoting total adenylate production is not particularly limited. The composition for promoting total adenylate production has the effect of promoting the production of total adenylate in cells when used as a liquid that comes into contact with cells, such as blood, infusions, aquaculture tanks, agricultural water, and culture fluids, or when added to these liquids. You can expect it. Products containing the composition for promoting total adenylate production include foods, health foods, beverages, alcoholic beverages, feed, fodder, pet food, aquaculture water, plant growth water, culture solutions, cosmetics, pharmaceuticals, and quasi-drugs. You can choose from the following groups. Products containing the composition for promoting total adenylate production are expected to be used as products with the added value of promoting total adenylate production at low cost. In addition, the ultra-fine bubbles themselves are not thought to accumulate in the body, and are therefore promising as a product with fewer concerns about side effects.

The dosage form and effective dosage of pharmaceuticals and quasi-drugs using the composition for promoting total adenylate production of the present disclosure depend on the subject to be administered, the dosage form, the patient's condition, and the doctor's judgment. Yes, but not limited. For example, for an adult weighing 60 kg, 50 mL to 800 mL of ultra-fine bubble liquid can be administered per dose. The frequency of administration is preferably once every 8 hours to 5 days, for example.

1. Test 1
(1) Preparation of culture medium (culture solution) Using a device equipped with a porous ceramic element, ultra-fine bubbles of oxygen gas are generated in pure water using a micropore method, and water containing ultra-fine bubbles is was created.

A twice-concentrated TBS medium was prepared. The components of the TBS medium are as follows.
Components of TBS medium: casein-sui digested peptone 34.0g/L, soybean-papain digested peptone 6.0g/L, sodium chloride 10.0g/L, dipotassium phosphate 5.0g/L, glucose 5.0g/L L

Water containing ultra-fine bubbles and twice the concentration TBS medium were mixed in equal amounts to prepare media for samples 1-2 and 1-3. The culture media of Samples 1-2 and 1-3 are examples of compositions for promoting total adenylate production.

A medium for sample 1-1 was prepared in the same manner as for samples 1-2 and 1-3, except that pure water was used instead of water containing ultra-fine bubbles. The medium of Sample 1-1 is a comparative example (Blank).

Optical sterilization was performed on the culture medium of each sample 1-1, 1-2, and 1-3 using UV light (wavelength: 250 nm, output: 6 W).

(2) E. coli test At Kitasato Environmental Science Center, a test was conducted to culture E. coli using the above medium. The outline of the test is as follows.
Escherichia coli (NBRC331) was inoculated into the culture medium of each sample at a concentration of 1×10 ⁶ CFU/mL, and was statically cultured at 36±2° C. A closed type culture container without a vent in the lid was used. 2 hours, 3 hours, 4 hours, 5 hours, 6 hours, and 7 hours after the start of culture, the total adenylate amount (RLU) contained in the medium and E. coli, and the glucose concentration (mg/ dL) was measured.

2. Exam 2
(1) Preparation of medium (culture solution) The medium of sample 2-1 was prepared by aerating the medium obtained in the same manner as the medium of sample 1-1 with a diffuser tube as a treatment to increase the dissolved oxygen concentration. The medium of Sample 2-1 is a comparative example (Blank).

The medium for Sample 2-2 was prepared by vacuum degassing the medium obtained in the same manner as the medium for Samples 1-2 and 1-3 as a treatment to lower the dissolved oxygen concentration. The medium of sample 2-2 is an example of a composition for promoting total adenylate production.

The medium for sample 2-3 was prepared in the same manner as the medium for samples 1-2 and 1-3. That is, the culture medium of Sample 2-3 was not subjected to any treatment to adjust the dissolved oxygen concentration. The medium of Sample 2-3 is an example of a composition for promoting total adenylate production.

The culture medium of each sample 2-1, 2-2, and 2-3 was sterilized by filtration using a syringe filter (membrane type: polypropylene) with a pore size of 0.22 μm. The culture medium of each sample 2-1, 2-2, and 2-3 was sealed in an aluminum pouch container and brought to the testing institution in order to suppress fluctuations in dissolved gas.

(2) E. coli test At Kitasato Environmental Science Center, a test was conducted to culture E. coli using the above medium. The outline of the test is as follows.
Escherichia coli (NBRC331) was inoculated into the culture medium of each sample at a concentration of 1×10 ⁶ CFU/mL, and was statically cultured at 36±2° C. The culture container used was an open type with a vent in the lid. 2 hours, 4 hours, 5 hours, 6 hours, 7 hours, and 24 hours after the start of culture, the total adenylate amount (RLU) contained in the medium and E. coli, and the glucose concentration (mg/ dL) was measured.

3. Exam 3
(1) Preparation of culture medium (culture solution) Using a device equipped with a porous ceramic element, ultrafine bubbles of air gas are generated in pure water using a microporous method, and water containing ultrafine bubbles is produced. was created.

Culture media for samples 3-2 and 3-3 were prepared by mixing 100 mL of water containing ultra-fine bubbles and a liquid culture medium for general viable bacteria (10.5 mL of MTV001-01 manufactured by Biotheter). The culture media of Samples 3-2 and 3-3 are examples of compositions for promoting total adenylate production. The components of the liquid medium are as follows.
Components of liquid medium: Yeast extract 5.0mg/mL, peptone 10.0mg/mL, glucose 2.0mg/mL

A medium for sample 3-1 was prepared in the same manner as for samples 3-2 and 3-3, except that pure water was used instead of water containing ultra-fine bubbles. The medium of Sample 3-1 is a comparative example (Blank).

(2) Bifidobacterium test A bifidobacteria intestinal stabilizer (BF-13 tablet manufactured by Biofermin Pharmaceutical) containing 12 mg of bifidobacteria per tablet was dissolved in 100 mL of pure water. 1 mL of a solution containing a bifidobacterial intestinal stabilizer was added to the culture medium of each sample, and the mixture was statically cultured at 36±2°C. Three specimens were prepared for each sample. Measure the total amount of adenylate (RLU) contained in the medium and Bifidobacterium at any of the following culture times: 20 hours, 68 hours, 90 hours, 112 hours, 115 hours, or 119 hours after starting the culture. did.

4. Analysis (1) Dissolved oxygen concentration The dissolved oxygen concentration of the medium of each sample before the start of culture was measured. A dissolved oxygen meter (TOA DKK DO-31P) was used for the measurement. This dissolved oxygen meter is a device that detects the dissolved oxygen concentration in a solution using a diaphragm electrode method.
(2) UFB concentration The UFB concentration (UFB cells/ml) in the medium of each sample before the start of culture was measured. A NanoSight NS-300 was used for the measurement. The measurement conditions were as follows.
[Measurement condition]
Frequency: 405nm
Camera Level: 15
Number of captures: 5
Capture duration:60
Detection Threshold: 4

(3) Total Adenylate Amount After a predetermined time from the start of culture, the total adenylate amount contained in the medium and E. coli or Bifidobacterium was measured. The measurement was performed by ATP test (A3 method) using Kikkoman Lumitester PD-30. The ATP test (A3 method) is a method for detecting the total amount of ATP, ADP, and AMP. More specifically, after ADP and AMP are converted to ATP by enzymes, the total amount of ATP, ADP, and AMP contained in the sample is detected by the amount of light emitted when they react with luciferase (RLU, Relative Light Unit). do. Measurements were performed three times for each specimen.

(4) Glucose concentration After a predetermined time from the start of culture, the glucose concentration of the medium was measured. A glucose monitor (GlucCell manufactured by Wakenbee Tech) was used for the measurement. GlucCell is a device that detects the glucose concentration in a dropped liquid using an electrochemical biosensor (enzyme electrode method).

5. Results (1) Results of Test 1 Table 1 shows the dissolved oxygen concentration (mg/mL) and UFB concentration (units/mL) of the medium of each sample before the start of culture. The UFB concentration of sample 1-2 was 1.05×10 ⁸ cells/mL. The UFB concentration of sample 1-3 was 4.44×10 ⁸ cells/mL.

The total amount of adenylate was measured 2 hours, 3 hours, 4 hours, 5 hours, 6 hours, and 7 hours after the start of culture. After 7 hours, sample 1-2 and sample 1- The difference between the average value of Sample 3 and Sample 1-1 (Blank) was the largest. Table 1 shows the total amount of adenylate (RLU) and the turbidity of the medium (OD ₆₆₀ ) 7 hours after the start of culture.

FIG. 2 shows the relationship between UFB concentration and total adenylate amount 7 hours after the start of culture. The dotted line is a linear approximate curve obtained by the least squares method. The equation of the linear approximation curve is y=0.0002x+124069, and the correlation coefficient is R ² =0.995.

Seven hours after the start of culture, that is, at the time when the difference between the average value of samples 1-2 and 1-3 and the total amount of adenylate between sample 1-1 (Blank) and the total adenylate amount is maximum, the UFB concentration and total adenylate It was found that there is a correlation between the amount and In addition, since the turbidity (OD ₆₆₀ ) of the culture medium of Samples 1-1, 1-2, and 1-3 after 7 hours from the start of culture is the same, Samples 1-1, 1-2, and 1- It is thought that the number of E. coli in sample No. 3 is approximately the same. That is, during the period from the start of culture to 7 hours, the production of ATP, ADP, and AMP per unit number of E. coli in Samples 1-1 and 1-2 was as low as per unit number of E. coli in Sample 1-3. It is presumed that the production of ATP, ADP, and AMP was promoted.

The glucose concentration (mg/dL) of Sample 1-2 and Sample 1-3 7 hours after the start of culture was lower than the glucose concentration (mg/dL) of Sample 1-1 (Blank). It is presumed that the decrease in the glucose concentration of the medium is due to E. coli consuming glucose as an energy source and producing ATP.

Further, sample 1-3 has a higher UFB concentration than sample 1-2. The total adenylate amount of sample 1-3 was higher than that of sample 1-2. Since the dissolved oxygen concentrations of Samples 1-2 and 1-3 were equivalent, it was suggested that the total adenylate amount did not depend on the dissolved oxygen concentration but increased depending on the UFB concentration.

(2) Results of Test 2 Table 2 shows the dissolved oxygen concentration (mg/mL) and UFB concentration (units/mL) of the culture medium of each sample before the start of culture. The UFB concentration of sample 2-2 was 7.33×10 ⁸ cells/mL. The UFB concentration of sample 2-3 was 6.97×10 ⁸ cells/mL.

The total amount of adenylate was measured 2 hours, 4 hours, 5 hours, 6 hours, 7 hours, and 24 hours after the start of culture. After 7 hours, sample 2-2 and sample 2- The difference between the average value of Sample 3 and Sample 2-1 (Blank) was the largest. Table 2 shows the total amount of adenylate (RLU) and the turbidity of the medium (OD ₆₆₀ ) 7 hours after the start of culture.

The relationship between the UFB concentration and the total amount of adenylate 7 hours after the start of culture is shown in FIG. The dotted line is a linear approximate curve obtained by the least squares method. The equation of the linear approximation curve is y=8E-05x+93300, and the correlation coefficient is R ² =0.9354.

Seven hours after the start of culture, that is, at the time when the difference between the average value of Sample 2-2 and Sample 2-3 and the total amount of adenylate between Sample 2-1 (Blank) and the total amount is maximum, the UFB concentration and total adenylate were determined. It was found that there is a correlation between the amount and In addition, since the turbidity (OD ₆₆₀ ) of the culture medium of samples 2-1, 2-2, and 2-3 after 7 hours from the start of culture is the same, samples 2-1, 2-2, and 2- It is thought that the number of E. coli in sample No. 3 is approximately the same. That is, during the period from the start of culture to 7 hours, the production of ATP, ADP, and AMP per unit number of E. coli in Samples 2-1 and 2-2 was as low as per unit number of E. coli in Sample 2-3. It is presumed that the production of ATP, ADP, and AMP was promoted.

The glucose concentration (mg/dL) of Sample 2-2 and Sample 2-3 7 hours after the start of culture was lower than the glucose concentration (mg/dL) of Sample 2-1 (Blank). It is presumed that the decrease in the glucose concentration of the medium is due to E. coli consuming glucose as an energy source and producing ATP.

Further, sample 2-2 has a lower dissolved oxygen concentration and higher UFB than sample 2-3. The total adenylate amount of sample 2-2 was lower than the total adenylate amount of sample 2-3. This suggests that the total amount of adenylate does not depend on the dissolved oxygen concentration, but increases depending on the UFB concentration.

(3) Results of Test 3 Table 3 shows the UFB concentration (units/mL) in the medium of each sample before the start of culture. The UFB concentration of sample 3-2 was 2.44×10 ⁸ cells/mL. The UFB concentration of sample 3-3 was 2.95×10 ⁸ cells/mL.

Table 3 shows the total amount of adenylate at 20 hours, 68 hours, 90 hours, 112 hours, 115 hours, and 119 hours after the start of culture. Note that the total amount of adenylate was measured three times for each sample for three samples. The average of the nine measurements in total is shown in the "Average value of total adenylate amount" column of Table 3.

The relationship between the culture time and the average value of the total amount of adenylate is shown in FIG. The details of the graph are as follows.
[Sample 3-1]
Line graph: black dashed line, plot cross Linear approximate curve obtained by least squares method: gray dashed line Formula of linear approximate curve: y = 692.0x + 118993, R ² = 0.8882
[Sample 3-2]
Line graph: solid black line, plot circle Linear approximate curve obtained by least squares method: solid gray line Formula of linear approximate curve: y = 2152.8x + 135391
[Sample 3-3]
Line graph: Black dot-dash line, plot triangle Linear approximation curve obtained by least squares method: Gray dot-dash line Formula of linear approximation curve: y = 2538.8x + 142390, R ² = 0.9702

Regarding the relationship between the culture time and the average value of the total amount of adenylate, the slope of the linear approximation curve is also shown in Table 3 as the rate of increase in total adenylate (RLU/h). The relationship between UFB concentration and rate of increase in total adenylate is shown in FIG. 5. In the graph of FIG. 5, the dotted line is a linear approximate curve obtained by the least squares method. The equation of the linear approximation curve was y=7E-06x+520.43, and the correlation coefficient was R ² =0.9978.

These results revealed that there is a high correlation between the UFB concentration and the rate of increase in total adenylate. Since Bifidobacterium is an obligate anaerobic bacterium that does not require air or oxygen for cultivation, it can be seen that the ultra-fine bubble itself has an effect on metabolism and promotion of ATP production.

The above results suggested that in a medium containing ultrafine bubbles at a concentration of 1×10 ⁸ cells/mL or more, the production of ATP, ADP, and AMP is promoted depending on the UFB concentration.

4. Effects of Examples As described above, according to this example, it was possible to provide a new composition for promoting total adenylate production, which contains ultrafine bubbles with a concentration of 1×10 ⁸ bubbles/mL or more as an active ingredient.

It should be noted that the embodiments disclosed herein are illustrative in all respects and should not be considered restrictive. The scope of the present invention is not limited to the embodiments disclosed herein, and includes all modifications within the scope indicated by the claims or within the scope equivalent to the claims. is intended.

Claims (5)

A composition for promoting the production of adenylate containing ATP, ADP, and AMP, which contains ultrafine bubbles at a concentration of 1×10 ⁸ bubbles/mL or more as an active ingredient. ATP, ADP according to claim 1, containing at least one selected from the group consisting of yeast extract, glucose, sodium chloride, dipotassium phosphate, peptone, casein-sui digested peptone, and soybean-papain digested peptone. , and a composition for promoting production of adenylate containing AMP. Claim 1, wherein the administration form is selected from the group consisting of impregnation, injection, dialysis, eye drops, ear drops, enema, drinking, gargling, oral mucosal use, skin external use, nasal absorption, and inhalation. Or a composition for promoting production of adenylate containing ATP, ADP, and AMP according to claim 2. Treatment of diseases, symptoms, or functions related to ATP production selected from the group of blood flow, inner ear disorders, dizziness, tinnitus, hearing loss, ear fullness, swelling, gastrointestinal motility, gastrointestinal mucosal secretion, and fatigue. A composition for promoting the production of adenylate, comprising ATP, ADP, and AMP according to claim 1 or 2, which is used for preventing or improving the production of adenylate. A product containing the composition for promoting production of adenylate containing ATP, ADP, and AMP according to claim 1 or 2, which is suitable for food, health food, beverage, alcoholic beverage, feed, fodder, pet food, aquaculture. A product containing a composition for promoting production of adenylate containing ATP, ADP, and AMP, selected from the group consisting of water, plant growth water, culture solution, cosmetics, pharmaceuticals, and quasi-drugs.

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