JP7149053B2 - Liquids for prevention or improvement of diabetes - Google Patents

Description

TECHNICAL FIELD The present invention relates to a liquid useful for prevention or improvement of diabetes, particularly suppression of postprandial hyperglycemia or suppression of fasting hyperglycemia.

Diabetes is an endocrine disease in which blood sugar levels are pathologically elevated. The number of people with diabetes worldwide has exceeded 400 million and continues to increase. Diabetes is roughly classified into type 1 and type 2. More than 95% of adult diabetics in Japan have type 2 diabetes. Type 2 diabetes is attributed to a combination of genetic predisposition and environmental factors such as aging, overnutrition, and lack of exercise. The pathology of type 2 diabetes is characterized by impaired insulin secretion and insulin resistance. Diabetes is known to cause complications such as nephropathy, retinopathy, neuropathy, and arteriosclerosis. Therefore, prevention and treatment of diabetes are important.

In type 2 diabetes, rapid postprandial hyperglycemia (glucose spike) has also attracted attention as a major problem. In recent years, due to insufficient intake of dietary fiber, easier digestion of carbohydrates due to refined staple foods, and increased intake of rapidly absorbed carbohydrates such as sugar and isomerized liquid sugar blended in soft drinks, etc., glucose spikes have occurred. Easy to happen. Larger postprandial glucose spikes adversely affect vascular endothelium and are aggravating factors for diabetes and its complications (nephropathy, retinopathy, neuropathy, etc.). It is also known that a glucose spike causes hypersecretion of insulin, resulting in accumulation of visceral fat, insulin resistance, and a decrease in insulin secretory ability, resulting in poor glycemic control. Persistence of hyperglycemia causes protein glycation, which is greatly involved in the development of complications. Therefore, suppression of postprandial hyperglycemia is thought to help prevent diabetes and vascular disease.

Various antidiabetic agents, such as insulin secretagogues (sulfonylureas) and insulin sensitizers (pioglitazone, etc.), are known for treating diabetes. Also, carbohydrate absorption delaying agents such as water-soluble dietary fiber and indigestible dextrin, and α-glucosidase inhibitors such as acarbose and voglibose are known to suppress glucose spikes.

It is known that mixing air bubbles in a liquid enhances the cleansing effect and provides effects such as stimulating the skin (Patent Documents 1 to 3). The applicant of the present application also discloses an apparatus for generating microbubbles in a liquid (Patent Document 4), and also discloses that such microbubble-containing liquid can be used for various cleaning purposes.

JP 2010-7315 A JP 2009-274026 A JP 2009-78140 A Japanese Patent No. 4999996

As described above, conventionally, diabetes has been prevented and improved by administration of chemical substances such as insulin secretagogues. In the method using such chemical substances, continuous intake is not easy and the economic burden is large. In addition, the ingredients currently used to suppress glucose spikes (such as acarbose), due to their nature, must be taken immediately before meals, limiting the timing of administration. In addition, acarbose and the like must be taken at a specific time just before meals, and forgetting to take them is likely to occur. If you keep forgetting to drink, you will not be able to suppress the glucose spike, and it will be difficult to control your blood sugar level.

An object of the present invention is to provide an air bubble-containing liquid that can be easily and continuously ingested and is useful for preventing or improving diabetes, particularly for suppressing rapid postprandial blood glucose elevation or fasting hyperglycemia. and

The liquid for preventing or improving diabetes of the present invention contains bubbles having an average diameter of 500 nm or less in the liquid medium.

The liquid substance of the present invention is easier to take continuously than administration of chemical substances. In addition, unlike conventional α-glucosidase inhibitors, there are no restrictions on when to take the drug, so forgetting to take it can be prevented. Furthermore, in addition to suppressing glucose spikes, hyperglycemia can be corrected by improving insulin resistance and the like. Therefore, the liquid product of the present invention is useful for preventing and improving diabetes.

FIG. 2 is a diagram showing measurement results of particle size distribution of bubbles in ultrapure water, FB1, and FB2 used in Test Examples 1 to 3; 1 is a diagram showing the results of an oral glucose tolerance test (OGTT) in Test Example 1. FIG. (A) shows the results after 5 weeks of administration, and (B) shows the results after 12 weeks of administration. 2 is a diagram showing the results of an oral glucose tolerance test (OGTT) in Test Example 2. FIG. (A) shows the results after 4 weeks of administration, and (B) shows the results after 6 weeks of administration. FIG. 10 shows the results of an oral glucose tolerance test (OGTT) in Test Example 3; (A) shows the results after 12 weeks of administration, and (B) shows the results after 18 weeks of administration. FIG. 10 is a diagram showing the results of measuring the amount of C-peptide in Test Example 3; FIG. 11 shows the results of an insulin tolerance test (ITT) in Test Example 3. FIG. FIG. 10 is a diagram showing the measurement results of HbA1c in Test Example 3;

An embodiment of the present invention will be described below. This embodiment is an example of implementing the present invention, and the present invention is not limited to this embodiment.

The liquid substance for preventing or improving diabetes of the present invention (hereinafter referred to as "this liquid substance") contains bubbles having an average diameter of 500 nm or less in a liquid medium.

The gas contained in the bubbles is not particularly limited as long as it does not interfere with the preventive or ameliorating effect of diabetes. One kind of the gas may be used, or a mixed gas of two or more kinds may be used. Examples of the gas include air, oxygen, hydrogen, nitrogen, argon, carbon dioxide, and mixtures thereof.

The "average diameter" of the bubbles means the number average of hydrodynamic diameters (diameters of particles in liquid). Specifically, the average diameter is a value measured by nanoparticle tracking analysis of Nanosite Nanoparticle Analysis System. The analysis can be performed using, for example, "NS500" and "LM10" manufactured by Nanosite.

The average diameter is 500 nm or less. The upper limit of the average diameter can be selected from the group consisting of 500 nm, 450 nm, 400 nm, 350 nm, 300 nm, 250 nm and 200 nm. The lower limit of the average diameter can be selected from the group consisting of 10 nm, 20 nm, 30 nm, 40 nm, 50 nm, 60 nm, 70 nm, 80 nm, 85 nm, 90 nm, 95 nm, 100 nm, 105 nm, 110 nm, 115 nm, and 120 nm. A preferable range of the average diameter can be a combination of the upper limit and the lower limit. For example, the average diameter is 1 to 500 nm, 10 to 500 nm, 20 to 500 nm, 30 to 500 nm, 40 to 500 nm, 50 to 500 nm, 60 to 500 nm, 70 to 500 nm, 80 to 500 nm, 80 to 400 nm, 80 to 300 nm. , 90-200 nm, 95-200 nm. When the average diameter is within the above range, it is possible to suppress diabetes prevention and improvement, particularly glucose spike or fasting hyperglycemia.

The particle size of the air bubbles can be appropriately determined according to need. For example, the D ₁₀ of the bubbles can be 80-500 nm, 80-400 nm, 80-300 nm, 80-200 nm, 85-200 nm. Also, the _D50 of the bubbles may be 100-500 nm, 100-400 nm, 100-300 nm, 100-200 nm, and 120-200 nm. "D10" means the particle size of air bubbles in which ₁₀ % of the particles (number/ml) have a size smaller than the specified _D10 value. " _D50 " means the particle size of air bubbles in which 50% of the particles (number/ml) have a size smaller than the specified _D50 value.

The concentration (number concentration) of the bubbles can be appropriately determined as necessary, as long as it does not inhibit the diabetes prevention or amelioration effect. The concentration of the bubbles is preferably 1×10 ⁷ /ml or more, preferably 1.5×10 ⁷ /ml or more, more preferably 2×10 ⁷ /ml or more, more preferably 2.5×10 ⁷ . pcs/ml or more. When the concentration of the bubbles is within the above range, it is preferable because it is possible to prevent and improve diabetes mellitus, particularly suppress postprandial increase in blood sugar level or fasting hyperglycemia. The number concentration of bubbles can be measured by the nanoparticle tracking analysis described above.

The liquid medium is ingestible by animals such as humans, and is not particularly limited as long as it does not interfere with the preventive or ameliorating effect of diabetes. Water is usually used as the solvent. Distilled water, ultrapure water, highly pure water, tap water, ion-exchanged water, filtered water, electrolyzed water, and natural water, for example, can be used as the water.

The liquid may contain components other than the bubbles as long as they do not interfere with the prevention or improvement of diabetes. For example, the present liquid may contain other substances or naturally-derived ingredients that have an effect of preventing or improving diabetes. In addition, the present liquid may contain a flavoring agent, a preservative, or the like for the purpose of flavor correction, aroma correction, deterioration prevention, and the like.

The aforementioned “prevention” includes not only maintaining normal blood sugar levels but also suppressing the development of diabetes from a state in which blood sugar levels tend to be high even when diabetes has not been diagnosed. The "improvement" includes not only treating diabetes, but also lowering blood glucose levels from a state in which blood glucose levels tend to be high even when diabetes is not diagnosed. In addition, both the above-mentioned "prevention" and "improvement" include suppression of postprandial blood glucose elevation (glucose spike) or fasting hyperglycemia.

There is no particular limitation on the method for preparing the present liquid. The present liquid can be prepared, for example, by blowing a gas into the liquid medium at room temperature or under pressure, and generating microbubbles by stirring, shearing, or the like. The average diameter and/or number concentration of the bubbles can be adjusted by appropriately setting conditions such as the amount of gas blown, stirring, and the tip. Also, the present liquid can be prepared by attaching a bubble generator described in Japanese Patent No. 4999996 to a water faucet and passing tap water through the bubble generator. According to this method, the present liquid can be prepared and ingested on a daily basis. As a result, it is possible to appropriately control the blood sugar level over a long period of time and easily prevent or improve diabetes.

There is no particular limitation on the mode of utilization of the present liquid. As described above, a suitable bubble generator may be attached to a tap and the liquid prepared by passing tap water through the bubble generator may be ingested. When the stability of the air bubbles in water is extremely high, the liquid can be used as a drink (e.g., tea, mineral water), particularly as a functional drink aimed at preventing or improving diabetes. The present liquid is excellent in suppressing postprandial hyperglycemia or fasting hyperglycemia. Therefore, the present liquid can be used to suppress postprandial hyperglycemia or fasting hyperglycemia. For example, the present liquid can be used as a functional drink that suppresses postprandial hyperglycemia or fasting hyperglycemia.

EXAMPLES The present invention will be specifically described below with reference to examples. It should be noted that the present invention is not limited to the forms shown in the examples. The embodiments of the present invention can be variously modified within the scope of the present invention depending on the purpose, application, and the like.

(1) Preparation and Analysis of Water Containing Nanobubbles Water supply (Water supply for well water in the field of animal experiments at the Life Science Research Support Center, Gifu University. In the examples, "tap water" means this well water.) A bubble generator (manufactured by Tanaka Kinzoku Seisakusho) described in Japanese Patent No. 4999996 was installed in the faucet. Nanobubble-containing water 1 (FB1) was obtained by passing tap water at a water pressure of 0.3 MPa through the generator. In addition, nanobubble-containing water 2 (FB2) was obtained in the same manner as for obtaining nanobubble-containing water 1, except that the orifice diameter in the generator was changed from φ3.5 to φ2.5.

For ultrapure water (control), FB1, and FB2, using "LM10V-HS" (manufactured by Nanosite) (CMOS camera manufactured by Malvern, UK, violet laser (405 nm, <60 mW), analysis software: NTA3.2 ), and measured the particle size distribution and concentration of air bubbles in water. The results are shown in Table 1 and FIG.

(2) Test example 1
(A) Experimental Animals and Breeding Method As experimental animals, Goto-Kakizaki (GK) rats (12 males, 5 weeks old) were used. The rat is used as a Japanese type 2 diabetes (non-obesity type; impaired insulin secretion) model. GK rats and their origin, Wistar rats (6 males), were tested in individual gauges under the conditions of a 12-hour light-dark cycle (lights on at 8:00 a.m., lights off at 8:00 p.m.), humidity of 60%, and temperature of 22°C ± 2°C. bred. Feeding and watering were performed three times a week.

GK rats and Wistar rats were acclimatized for one week by ad libitum intake of a basic sample for rearing breeding ("CE-2" manufactured by Clea Japan) and tap water. After acclimation, GK rats were divided into two groups of 6 rats each. For 13 weeks from 6 to 19 weeks of age, GK rats and Wistar rats were allowed to freely ingest a high-fat diet "Quick Fat" (manufactured by Clea Japan; 30 kcal% fat) for research on type 2 diabetes and obesity. , Tap water or FB1 was given ad libitum as a drink. Table 2 shows the animals, number of animals, drink, and feed in each group. In Table 2, "W" is tap water and "QF" represents high fat diet.

(B) Oral glucose tolerance test (OGTT)
OGTT was performed 5 weeks (11 weeks old) and 12 weeks (18 weeks old) after the start of intake of the beverage (tap water or nanobubble-containing water 1). After an overnight fast, rats were weighed and orally administered glucose using a gastric tube. The oral dose of the glucose (D(+) glucose; manufactured by Wako Pure Chemical Industries, Ltd.) is 2 g glucose/4 ml distilled water/kg body weight. Blood was collected from the tail vein at 0, 30, 60, 120 and 180 minutes after administration, and the blood glucose level was measured at each elapsed time. Blood glucose levels were measured using a blood glucose self-monitoring device "Nipro Freestyle Freedom Light" (manufactured by Nipro). The results are shown in FIG.

(C) Evaluation As shown in FIG. 2, in all groups at 5 weeks and 12 weeks of administration, the blood glucose peaked at 30 to 60 minutes and then decreased over time. Compared to group 1-1 (normal rats + tap water), both group 1-2 (GK rats + tap water) and group 1-3 (GK rats + FB1) showed a rapid increase in blood glucose level. . This indicates that a rapid postprandial hyperglycemia (glucose spike) occurred in diabetes.

On the other hand, in the 5th week and 12th week of administration, at the stage of 60 minutes after administration, the 1-3 group ingesting FB1 significantly reduced the increase in blood sugar level compared to the 1-2 group ingesting tap water. did. Moreover, at the stage of 120 to 180 minutes after administration in the 5th week of administration, group 1-3 showed a tendency to decrease the increase in blood glucose level as compared with group 1-2. Similarly, at the stage of 30 to 180 minutes after administration on the 12th week of administration, group 1-3 showed a tendency to decrease the increase in blood glucose level compared to group 1-2. These results show that ingestion of FB1 containing nanobubbles can suppress rapid postprandial hyperglycemia in diabetic patients with decreased insulin secretion.

(3) Test Example 2
(A) Experimental Animals and Raising Method As experimental animals, Zucker Diabetic Fatty (ZDF) rats (18 males, 5 weeks old) were used. The rat develops a pathological condition similar to type 2 diabetes in adult humans, and is considered to be a Western type (obesity/severe) type 2 diabetes model. ZDF rats and Lean rats (6 males) were housed in individual cages under conditions of a 12 hour light/dark cycle (lights on at 8:00 am, lights off at 8:00 pm), humidity of 60%, and temperature of 22°C ± 2°C. Feeding and watering were performed three times a week.

ZDF rats and lean rats were acclimatized for one week by ingesting a normal diet ("CE-2" manufactured by Clea Japan) and tap water ad libitum. After habituation, ZDF rats were divided into 3 groups of 6 rats each. For 14 weeks from 6 weeks to 20 weeks of age, ZDF rats and Lean rats were allowed to freely ingest a high-fat diet "Quick Fat" (manufactured by Clea Japan; 30 kcal% fat) for research on type 2 diabetes and obesity. , Tap water, FB1, and FB2 were given ad libitum as drinks. Table 3 shows the animals, number of animals, drink, and diet of each group. In Table 3, "W" is tap water and "QF" represents high fat diet.

(B) Oral glucose tolerance test (OGTT)
OGTT was performed 4 weeks (10 weeks old) and 6 weeks (12 weeks old) after the initiation of intake of beverages (tap water or FB1). OGTT was performed by the same method as in Test Example 1. The OGTT results are shown in FIG.

(C) Evaluation As shown in FIG. 3, the blood glucose level peaked at 30 to 60 minutes and then decreased over time in all groups on the 4th and 6th weeks of administration. A sharp rise in blood glucose was observed in groups 2-2 to 4 (ZDF rats) as compared to group 2-1 (normal rats). Similar to Test Example 1, this indicates that a rapid postprandial hyperglycemia (glucose spike) occurred in diabetics.

On the other hand, in the 4th week of administration, at the stage of 60 to 120 minutes after administration, in the 2-3 group and 2-4 group ingesting FB1 and FB2, compared to the 2-2 group ingesting tap water, the blood glucose level It showed a trend of declining ascent. In addition, in the 6th week of administration, at the stage of 30 minutes after administration, in the 2-3 group and 2-4 group that ingested FB1 and FB2, compared to the 2-2 group that ingested tap water, the blood sugar level was significant. In addition, even after 60 minutes, the increase in blood sugar level tended to decrease. From these results, as in Test Example 1, it can be seen that rapid postprandial hyperglycemia can be suppressed by ingesting FB1 and FB2 containing nanobubbles.

(4) Test Example 3
(A) Experimental Animals and Breeding Method As experimental animals, C57BL/6J mice (42 5-week-old males) were used. The mice are induced obese type 2 diabetes by high fat diet. The mice were housed in individual cages under conditions of a 12 hour light/dark cycle (lights on at 8:00 am, lights off at 8:00 pm), humidity of 60%, and temperature of 22°C ± 2°C. Feeding and watering were carried out three times a week.

The mice were given a normal diet (“CE-2” manufactured by Clea Japan, Inc.) and tap water ad libitum and acclimatized for one week. After habituation, the mice were divided into four groups. For 13 weeks from 6 weeks to 19 weeks of age, the mice were given a high-fat diet "D12451" (manufactured by Research Diets, Inc; fat 45 kcal%) for type 2 diabetes/obesity research or the normal diet ad libitum. and freely ingested tap water, FB1, and FB2 as beverages. Table 4 shows the animals, number of animals, drink, and diet of each group. In Table 4, "W" is tap water, "CD" represents normal diet, and "FD" represents high fat diet.

(C) Oral glucose tolerance test (OGTT)
OGTTs were performed 12 and 18 weeks after initiation of drinking (tap water and FB1). OGTT was performed by the same method as in Test Example 1. The results are shown in FIG.

(D) Measurement of C-peptide Amount The amount of C-peptide was measured. Using "Morinaga Mouse C-Peptide Measurement Kit" (Morinaga Institute of Biological Science) and using "GE Healthcare, Ultraspec Visible Plate Reader II 96" as a measurement device, OGTT plasma at 18 weeks (0 minutes, 30 minutes , and 60 min) were measured. The results are shown in FIG.

(E) insulin tolerance test (ITT)
ITT was performed 17 weeks after the initiation of drinking (tap water, FB1 or FB2). After fasting overnight, the rats were weighed and intraperitoneally administered an insulin preparation (100 units of "Humarin R" manufactured by Eli Lilly Japan; 10 µL/g body weight). After that, blood was collected from the tail vein at 0, 30, 60, 120 and 180 minutes, and the blood glucose level was measured at each elapsed time. The blood glucose level was measured using a blood glucose self-measuring device "Nipro Freestyle Freedom Light" (manufactured by Nipro). The results are shown in FIG.

(F) Measurement of HbA1c After 19 weeks from the start of ingestion of beverages (tap water, FB1 or FB2), cardiac blood sampling and heparin anticoagulation were performed under somnopentyl (50 mg/kg body weight) anesthesia after overnight fasting. Using the obtained blood, using whole blood as a sample, using an automatic glycohemoglobin analyzer "HLC-723GHbV" (Tosoh Corporation), by measuring the ratio (%) of glycohemoglobin in total hemoglobin, HbA1c was measured. The results are shown in FIG.

(G) Evaluation As shown in FIG. 4, the blood glucose level peaked at 30 to 60 minutes and then decreased over time in all groups at 12 weeks and 18 weeks of administration. At week 12 of administration, there was no difference in the decrease in blood glucose level between the 3-2 group (high-fat diet + tap water) and the 3-3 and 3-4 groups (high-fat diet + FB1 or FB2). . However, at week 18 of administration, blood glucose levels decreased more rapidly in groups 3-3 and 3-4 (high-fat diet + FB1 or FB2) than in group 3-2 (high-fat diet + tap water). The results showed that the change in blood glucose level was close to that of normal mice (Group 3-1). From these results, it can be seen that continuous ingestion of FB1 and FB2 can suppress postprandial increase in blood sugar level. In addition, unlike existing α-glucosidase inhibitors that suppress the postprandial rise in blood sugar level, it can be seen that the postprandial rise in blood sugar level can be suppressed even if the drug is not taken immediately before a meal. This result indicates that suppression of postprandial blood glucose level elevation is not necessarily due to α-glucosidase inhibitory action.

C-peptide is a substance generated by the breakdown of proinsulin. C-peptide circulates in the blood almost undegraded and is excreted with urine. Therefore, C-peptide is an indicator of insulin secretion from the pancreas.

As shown in FIG. 5, no significant change in the amount of C-peptide was observed in group 3-1, whereas the amount of C-peptide increased significantly in group 3-2 20 minutes after the glucose challenge. This is thought to be due to increased insulin resistance in tissues and cells due to diabetes, and increased insulin secretion to counteract this. On the other hand, in the 3-3 group and the 3-4 group (FB1 and FB2 intake), the increase in the amount of C-peptide 20 minutes after the glucose loading was suppressed compared to the 3-2 group (tap water). From these results, it can be seen that ingestion of FB1 and FB2 can improve tissue and cell insulin resistance, and is effective in preventing or improving diabetes.

As shown in FIG. 6, in the insulin tolerance test, blood glucose levels were significantly lower in groups 3-3 and 3-4 (FB1 and FB2 intake) than in group 3-2 (tap water). It can be seen that the 3-3 group and the 3-4 group show the effect of insulin as compared to the 3-2 group.

HbA1c is a glycated protein in which glucose is bound to the end of the hemoglobin β chain. A hyperglycemic state continues for a long time, and it is formed by combining extra glucose and hemoglobin in blood vessels. Therefore, HbA1c is an index of blood sugar level in the past January to February.

As can be seen from FIG. 7, HbA1c is higher in groups 3-2 to 4 than in group 3-1, which is in a normal state, indicating that they are diabetic. On the other hand, compared to Group 3-2 (tap water intake), HbA1c was significantly decreased in Group 3-3 (FB1 intake), and HbA1c tended to decrease in Group 3-4 (FB2 intake). was accepted. From these results, it can be seen that the intake of FB1 and FB2 can suppress the elevation of blood sugar level for a long period of time.

From the results of the OGTT (Fig. 4), the mechanism of suppression of blood glucose elevation by ingestion of FB1 and FB2 is presumed to be glucose tolerance improving action other than digestion/absorption inhibition and incretin secretion. Considering this speculation and the results of ITT (Fig. 6), it is suggested that the mechanism of suppression of blood glucose elevation by ingestion of FB1 and FB2 is insulin resistance improving action. Therefore, it is thought that the liquid substance of the present invention can suppress the postprandial increase in blood sugar without taking it immediately before a meal, unlike the existing α-glucosidase inhibitors such as acarbose (experimental example The discussion in the section merely indicates the individual discussion of the inventor, and is not a statement that defines the present invention, nor does it have any intention of defining the present invention.).

Claims (6)

A liquid for preventing or improving diabetes, containing air bubbles having an average diameter of 500 nm or less in a liquid medium, for suppressing postprandial elevation of blood sugar. 2. The liquid for preventing or improving diabetes according to claim 1, wherein said liquid medium is water. 3. The liquid for preventing or improving diabetes according to claim 1, wherein the average diameter of said bubbles is 30 to 500 nm. The liquid for preventing or improving diabetes according to any one of claims 1 to ₃ , wherein D10 of said bubbles is 80 to 500 nm. The liquid for preventing or improving diabetes according to any one of claims 1 to 4, wherein the bubbles have a _D50 of 100 to 500 nm. The liquid for preventing or improving diabetes according to any one of claims 1 to 5, wherein the concentration of said bubbles is 1 x ¹⁰⁷ cells/ml or more.

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