WO2012063553A1 - Agent for reducing risk of disease onset resulting from non-dipper type intra-day blood pressure fluctuation - Google Patents

Abstract

Provided is an agent that reduces the risk of disease onset resulting from non-dipper type intra-day blood pressure fluctuations and that can reduce the onset of disease resulting from intra-day blood pressure fluctuation risk and prone to occur towards the morning when early morning systolic blood pressure significantly decreases from that of the night time in humans having a particular non-dipper type intra-day blood pressure fluctuation pattern, particularly healthy individuals exhibiting normal SBP and DBP values. The agent is characterized by containing: a hydrolysate containing VPP and IPP and obtained by hydrolyzing animal milk protein; or a concentrate thereof.

Description

Non-dipper type blood pressure diurnal agent

The present invention has a low blood pressure drop rate from night to early morning, and is generally a non-dipper, particularly a normal blood pressure person whose average blood pressure for 24 hours is normal. The present invention relates to an agent capable of reducing the risk of developing various diseases caused by blood vessel troubles such as cardiovascular and cerebrovascular due to fluctuations.

It has been conventionally known that hypertension is an important risk factor for many diseases such as cerebral infarction, myocardial infarction, myocardial ischemia, atherosclerosis, heart failure, stroke, coronary artery disease, peripheral vascular disease and the like.
Therefore, many components having a blood pressure lowering effect have been found and used for hypertensive patients. For example, tripeptides such as Val Pro Pro and Ile Pro Pro, and casein hydrolysates containing these have angiotensin I converting enzyme inhibitory activity (ACEI activity) and are known to exhibit blood pressure lowering activity ( (See Patent Documents 1 and 2).
The definition of such a hypertensive patient is not necessarily clear and varies depending on the measuring device, environment, etc. In general, in the measurement using a portable 24-hour automatic sphygmomanometer that has recently become widespread, the average of systolic blood pressure (SBP) is generally In many cases, the target is 130 mmHg or more and the average diastolic blood pressure (DBP) is 80 mmHg or more (see Non-Patent Document 1).

With the spread of the portable 24-hour automatic blood pressure monitor, the pattern of blood pressure diurnal variation by the ABPM (ambulatory blood pressure monitoring) method has been measured in recent years, and risk factors for various diseases have been classified (non-patent literature). 2).
Circadian blood pressure fluctuations generally tend to be higher during the day and lower from night to early morning. Among such blood pressure fluctuation patterns, there is a method of classifying a human whose fall rate of the average systolic blood pressure at night is 10% or more lower than the average systolic blood pressure during the day as a Dipper type and a human who is lower than 10% as a non-dipper type. Although known (Non-Patent Document 3), there is no provision for clear classification criteria of the Dipper type and Non-dipper type at present. In any case, non-dipper type human blood pressure diurnal variation is more cerebral infarction, myocardial infarction, myocardial ischemia, atherosclerosis, heart failure, stroke, coronary artery disease, peripheral vascular disease than Dipper type human Have been reported to tend to develop in the morning.
It has been found that such a tendency is not only seen in the hypertension patient group, but also in the healthy person group whose normal blood pressure for 24 hours is normal.
In general, various types of antihypertensive agents to be administered to hypertensive patients are known, such as those having an action of lowering daytime blood pressure, and those that lower the overall daily blood pressure having a long-lasting effect. However, in the above normal blood pressure healthy people who have not previously required the use of antihypertensive agents, it has the effect of bringing the average systolic blood pressure from the night to early morning of humans classified as non-dipper type closer to Dipper type. Little is known about the agents indicated.

Japanese Patent Laid-Open No. 6-40994 Japanese Patent Laid-Open No. 3-120225

"High Blood Pressure Treatment Guidelines 2009" (Chapter 2, Blood Pressure Measurement and Clinical Evaluation, p8-17, (2009) Japan Hypertension Society) Japanese Circulation Journal Vol. 64, Suppl. V, 2000 (Guidelines for the use of 24-hour sphygmomanometer (ABPM) standards, p1207-1237) THE JOURNAL OF CLINICAL HYPERTENSION (The Impact of Lactotripeptides on Blood Pressure in Stage 1 and Stage 2 Hypertensives, p1-7)

An object of the present invention is to provide a blood pressure that is likely to develop in the morning for a specific non-dipper, particularly a normal blood pressure person whose average blood pressure is normal for 24 hours, with a low blood pressure drop rate from night to early morning. The object is to provide an agent that can reduce the risk of developing diseases due to diurnal variation.

According to the present invention, the brachial blood pressure was measured every 30 minutes with a portable 24-hour automatic sphygmomanometer, and the mean systolic blood pressure of 0:00 to 5:00 to the mean systolic blood pressure of 8:00 to 21:00. Non-dipper type humans with a decrease rate of less than 10%, especially systolic blood pressure (SBP) measured by a portable 24-hour automatic sphygmomanometer every 30 minutes and the average systolic blood pressure (SBP) is less than 130 mmHg, diastolic blood pressure (DBP) is an agent that can be taken orally by humans with an average of less than 80 mmHg, and can reduce the risk of developing diseases due to blood pressure diurnal variation,
Non-dipper-type blood pressure diurnal risk-reducing agent containing hydrolyzate containing Ile Pro Pro (IPP) and Val Pro Pro (VPP) or its concentrate obtained by hydrolyzing animal milk protein ( Hereinafter, it may be abbreviated as the mitigating agent of the present invention).
According to the present invention, the upper systolic blood pressure was measured every 30 minutes with a portable 24-hour automatic blood pressure monitor, and the average systolic period of 0:00 to 5:00 with respect to the average systolic blood pressure of 8:00 to 21:00. Non-dipper type humans whose blood pressure decrease rate is less than 10%, in particular, the upper arm blood pressure measured every 30 minutes with a portable 24-hour automatic sphygmomanometer is less than 130 mmHg, and the average DBP is less than 80 mmHg Use of a hydrolyzate containing IPP and VPP obtained by hydrolyzing animal milk protein or a concentrate thereof to produce an agent that can be taken orally by humans and that can reduce the risk of developing disease due to diurnal fluctuations in blood pressure Provided.
Furthermore, according to the present invention, the mean systolic period of 0:00 to 5:00 is measured with respect to the mean systolic blood pressure of 8:00 to 21:00, where the brachial blood pressure is measured every 30 minutes by a portable 24-hour automatic sphygmomanometer. Non-dipper type humans whose blood pressure decrease rate is less than 10%, in particular, the upper arm blood pressure measured every 30 minutes with a portable 24-hour automatic sphygmomanometer is less than 130 mmHg, and the average DBP is less than 80 mmHg Provided is a method for reducing the risk of developing disease due to blood pressure diurnal fluctuation, by allowing a human to orally ingest a hydrolyzate containing IPP and VPP or a concentrate thereof obtained by hydrolyzing animal milk protein.

The alleviating agent of the present invention effectively reduces systolic blood pressure from night to early morning for humans having a specific non-dipper type blood pressure diurnal variation pattern, particularly healthy individuals with normal values of SBP and DBP. The risk of developing many diseases such as cerebral infarction, myocardial infarction, myocardial ischemia, atherosclerosis, heart failure, stroke, coronary artery disease, peripheral vascular disease, etc. Can be expected to reduce.

It is a graph for comparing SBP before and after the measurement of the disease onset risk reduction effect by the Dipper type blood pressure diurnal variation performed in Example 1. 2 is a graph for comparing SBPs before and after measurement of a disease onset risk reduction effect by non-dipper blood pressure diurnal variation performed in Example 1. FIG.

Hereinafter, the present invention will be described in more detail.
The alleviation agent of the present invention has an average systolic period of 0:00 to 5:00 with respect to an average systolic blood pressure of 8:00 to 21:00, in which the upper arm blood pressure is measured every 30 minutes by a portable 24-hour automatic blood pressure monitor. Non-dipper type humans whose blood pressure decrease rate is less than 10%, in particular, the upper arm blood pressure measured every 30 minutes with a portable 24-hour automatic sphygmomanometer is less than 130 mmHg, and the average DBP is less than 80 mmHg Targeting humans. In particular, the latter human is a human who is not normally recognized as a hypertensive patient and does not require administration of a hypotensive agent even if it is a non-dipper type. However, recently, it has been found that the non-dipper type group having a low blood pressure decrease rate at night or early morning has a higher risk of developing disease due to diurnal fluctuations in blood pressure than the Dipper type human group. In the present invention, it is possible to reduce the blood pressure decrease rate at night or early morning in humans who have such a high risk, and to reduce the risk.

The mitigating agent of the present invention contains a hydrolyzate containing IPP and VPP or a concentrate thereof obtained by hydrolyzing animal milk protein.
Examples of animal milk proteins include cow milk, horse milk, sheep milk, goat milk, and skim milk, reduced milk, powdered milk, condensed milk, which are processed milk thereof, and milk or processed milk thereof is particularly preferable. It is already known that proteins contained in these include IPP sequences and VPP sequences.
The solid content concentration of animal milk is not particularly limited. For example, when skim milk is used, the non-fat milk solid content concentration is usually about 3 to 15% by mass, and 6 to 15% by weight is preferable in terms of productivity. .

The hydrolyzate used in the mitigating agent of the present invention preferably has a VPP content of 3 μg / ml or more, particularly 30 μg / ml to 60 μg / ml, so that IPP and VPP can be obtained from animal milk protein. Is obtained by hydrolysis so that the content of IPP is 30 μg / ml to 40 μg / ml and IPP content is 2 μg / ml or more, in particular 20 μg / ml to 40 μg / ml, more preferably 20 μg / ml to 30 μg / ml. It is.
Examples of the hydrolysis method include a method of fermenting a raw material containing animal milk protein with lactic acid bacteria to obtain a fermented product (A), a method of enzymatically decomposing the raw material containing animal milk protein with an enzyme (B), and the above There is a method of obtaining a hydrolyzate by combining (A) and (B).

In the method (A), examples of lactic acid bacteria include lactic acid bacteria belonging to the genus Streptococcus, Lactococcus, Lactbacillus, Bifidobacteria, etc., and the Lactbacillus genus is preferred. Specific examples include Lactobacillus bulgaricus, Lactobacillus helveticus, Lactobacillus casei, Lactobacillus acidophilus, and Lactobacillus fermentum. In particular, Lactobacillus helveticus excellent in IPP and VPP production efficiency can be preferably used.
More specifically, Lactobacillus helveticus ATCC 15009, Lactobacillus helveticus ATCC 521, Lactobacillus helveticus CM4 strain (National Institute of Advanced Industrial Science and Technology, Patent Biological Deposit Center, 1-1-1 Higashi 1-1-1, Tsukuba City, Ibaraki Prefecture, Japan: FERM BP-6060, date of deposit 1997.15.15) (hereinafter referred to as CM4 strain). This CM4 strain is registered under the above-mentioned deposit number in the Budapest Treaty concerning the international recognition of microbial deposits in patent procedures, and this strain has already been patented.

The lactic acid bacteria are preferably used as a sufficiently high starter that has been pre-cultured in advance. The initial bacterial count is preferably about 10 ⁵ to 10 ⁹ cells / ml.
In order to improve the flavor of the obtained fermented product and improve the palatability, yeast can be used in combination during the fermentation. The strain of yeast is not particularly limited, and preferred examples include yeasts of the genus Saccharomyces such as Saccharomyces cerevisiae. The content rate of yeast can be suitably selected according to the purpose.

In the fermentation method, one or more of the lactic acid bacteria are cultured in a medium, or one or more of the lactic acid bacteria and one or more of the yeast are mixed and cultured in a medium. Can be performed.
As the medium, raw material medium containing the animal milk protein, or yeast extract, vitamins such as ascorbic acid, amino acids such as cysteine, salts such as sodium chloride, glucose, sucrose, raffinose, stachyose as secondary components. Etc., a medium to which a stabilizer such as gelatin, a stabilizer such as gelatin, and a flavor are appropriately added can be used.
Fermentation can be performed at a fermentation temperature of 25 to 50 ° C., preferably 30 to 45 ° C., usually by standing or stirring culture so that the IPP and VPP can be obtained, and the fermentation time is 6 to 30 hours. It is preferably 10 to 24 hours. It can be performed by a method of stopping the culture when the number of bacteria reaches 10 ⁷ cells / ml or more and pH 5.0 or less under conditions such as the initial pH of fermentation of 6.0 to 7.0. Moreover, the animal milk protein before fermentation may be subjected to high-temperature heat sterilization or the like.

As the enzyme used in the method (B), for example, a peptidase capable of cleaving the Pro Xaa sequence at the carboxy terminus of the Xaa Pro Xaa sequence or Xaa Pro Pro Xaa sequence (Xaa represents an arbitrary amino acid) in animal milk protein. Preferred examples include enzymes.
The enzyme preferably contains a serine type proteinase having serine at the active center or a metal proteinase having metal at the active center. Examples of metal proteinases include neutral protease I, neutral protease II, leucine aminopeptidase, and the like, and further including at least one of these can efficiently produce a desired hydrolyzate in a short time, This is preferable because it can be obtained by a one-step reaction. The peptidase that can cleave the Pro Xaa sequence is preferably an enzyme having an isoelectric point in the acidic region.

Examples of the enzyme include an enzyme group derived from Aspergillus oryzae and other koji molds. Such an enzyme group includes an enzyme group obtained by culturing cells in an appropriate medium and extracting the produced enzyme with water, and in particular, the isoelectric point of the enzyme group derived from Aspergillus oryzae has an acidic region. The enzyme group which shows is preferably mentioned.
As the enzyme group derived from Aspergillus oryzae, commercially available products can be used. For example, Sumiteam FP, LP or MP (above, registered trademark, manufactured by Shin Nippon Chemical Co., Ltd.), Ummamizyme (registered trademark, Amano Enzyme) ), Sternzyme B11024, PROHIDROXY AMPL (trade name, manufactured by Higuchi Shokai Co., Ltd.), Orientase ONS (registered trademark, manufactured by Hankyu Bioindustry Co., Ltd.), Denateam AP (registered trademark, Nagase Biochemical Co., Ltd.) In particular, the use of Sumiteam FP (registered trademark, manufactured by Shin Nippon Chemical Co., Ltd.) is preferable.
When these commercially available enzyme groups are used, the optimum conditions are usually set. However, for example, the amount of enzyme used and the reaction time are determined according to the enzyme group used so that the hydrolyzate can be obtained. Changes can be made as appropriate.

The amount of the enzyme added is, for example, an enzyme / animal milk protein in an aqueous solution in which animal milk protein is dissolved in a mass ratio of 1/1000 or more, preferably 1/1000 to 1/10, particularly preferably 1/100 to 1. / 10, more preferably in an amount of 1/40 to 1/10.
The reaction conditions can be appropriately selected depending on the enzyme so that the desired hydrolyzate can be obtained, but the temperature is usually 25 to 60 ° C., preferably 45 to 55 ° C., and the pH is usually 3 to 10, preferably 5 To 9, particularly preferably 5 to 8. The enzyme reaction time is usually 2 to 48 hours, preferably 7 to 15 hours.
The enzyme reaction can be terminated by inactivating the enzyme. Usually, the enzyme can be inactivated at 60 to 110 ° C. to stop the reaction.

The hydrolyzate obtained by the method (A), (B), and the method combining (A) and (B) can remove the precipitate by centrifugation or various filter treatments as necessary. it can.
Moreover, the peptide which has a bitter taste and a odor can also be removed from the obtained hydrolyzate as needed. Removal of such bitter components and odor components can be performed using activated carbon, hydrophobic resin, or the like. For example, activated carbon can be added to the hydrolyzate obtained in an amount of 1 to 20% by mass based on the amount of animal milk protein used and reacted for 1 to 10 hours. The used activated carbon can be removed by a known method such as centrifugation or membrane treatment.

The obtained hydrolyzate can be used as it is as the reducing agent of the present invention. Moreover, in order to improve the versatility of the hydrolyzate, it can be dried and made into a powder form after concentration.
Such powders may contain various auxiliary additives in order to improve nutritional balance, flavor, and the like. For example, various carbohydrates, lipids, vitamins, minerals, sweeteners, fragrances, pigments, texture improving agents and the like can be mentioned.

The dosage of the reducing agent of the present invention is preferably about 1 mg to 30 g, particularly about 20 mg to 20 g in terms of solid matter, and preferably divided into several times a day for the subject human subject per day. May be taken.
The administration period is usually 1 day or longer, preferably 20 days or longer, and it is desirable to take ingestion continuously or continuously. The administration method is oral intake.

Examples of the form of the reducing agent of the present invention include tablets, pills, hard capsules, soft capsules, microcapsules, powders, granules, liquids and the like.
The formulation is carried out, for example, as necessary, as a pharmaceutically acceptable carrier, adjuvant, excipient, excipient, preservative, stabilizer, binder, pH adjuster, buffer, thickener. , Gelling agents, preservatives, antioxidants, etc., can be manufactured in unit dosage forms required for generally accepted formulation practice.
In formulating the above reducing agent of the present invention, it is possible to use known antihypertensive agents in combination in order to improve the effect of the present invention. In addition, the effect of the present invention is expected to be enhanced by using in combination with other drugs known to exhibit a normalizing action of Non-dipper such as olmesartan and altinolol arotinolol.

In addition to the hydrolyzate, the mitigation agent of the present invention includes components used in foods and drinks as necessary, such as sugars, proteins, lipids, stabilizers, vitamins, minerals, flavors, or mixtures thereof. Can also be added.

EXAMPLES Hereinafter, although an Example demonstrates this invention still in detail, this invention is not limited to these.
Example 1
<Preparation of CM4 fermented milk>
Commercial skim milk powder is dissolved in distilled water to a solid content of 9% (w / w), sterilized by heating at 105 ° C. for 10 minutes in an autoclave, cooled to room temperature, CM4 strain starter fermentation solution (bacteria Several 5 × 10 ⁸ pieces / ml) was inoculated with 3% (v / w) and fermented at 37 ° C. for 24 hours in a stationary state to obtain CM4 fermented milk.

<Measurement of IPP and VPP content in CM4 fermented milk>
First, 1 ml of CM4 fermented milk was directly centrifuged at 15000 rpm for 10 minutes, and the supernatant (whey) was recovered. 0.3 ml of this whey was adsorbed on a Sep-Pak Cartridge (manufactured by Waters) and washed with distilled water. It was eluted with 5 ml of methanol and dried under reduced pressure under centrifugal treatment. The dried product was dissolved in 0.3 ml of an aqueous solution of 0.05% trifluoroacetic acid and subjected to HPLC analysis under the following conditions.
Model used: Hitachi L4000 UV detector (detected at 215 nm), L6200 intelligent pump L5030 column oven (35 ° C.), separation conditions: flow rate 0.5 ml / min, eluent: 0.3 M NaCl, 0.05% trifluoroacetic acid in water Column: Trade name Asahipak GS320 (Φ3.9 × 600mm).
As a result, the amount of VPP in CM4 fermented milk whey was 38.5 μg / ml, and the amount of IPP was 23.5 μg / ml.

<Preparation of fermented dairy products for subjects>
To the obtained CM4 fermented milk, stabilizers, sweeteners, flavors, and water were added, homogenized, filled into glass bottles 160 ml at a time, sterilized at 85 ° C, and used as a fermented milk product for subjects. . Table 1 shows the component analysis results of the fermented milk products for subjects. The amount of VPP in the subject day's fermented milk product for subjects was 2.53 mg and the amount of IPP was 1.52 mg.

<Selection of normal blood pressure Dipper and non-dipper subjects>
For 12 subjects aged 55 to 71 years who are not currently taking antihypertensive drugs and have no experience of severe hypertension, allergic asthma, severe liver or kidney disease, stroke, or myocardial infarction, The following blood pressure measurements were performed.
Each subject was asked to wear a portable 24-hour automatic sphygmomanometer (model ES-H531, manufactured by Terumo Corporation) on the upper arm that was not the toe arm, and SBP, DBP and heart rate were measured every 30 minutes for 24 hours.
As a result, 6 subjects who had an average SBP for 24 hours of less than 130 mmHg, an average DBP for 24 hours of 80 mmHg or less, and no abnormal heart rate were selected.
Also, from the blood pressure measurement results of the selected six subjects, the average SBP reduction rate at 0:00 to 5:00 relative to the average SBP at 8:00 to 21:00 was calculated, and the reduction rate was 10% or more. Subjects were classified as Dipper type, and less than 10% of subjects were classified as Non-dipper type.
As a result, there were three normal blood pressure Dipper subjects (subjects 4, 6, 10) and three non-dipper subjects with normal blood pressure (subjects 3, 8, 9). Of the above-mentioned data of each selected subject, the DBP type subject's SBP data is shown in Table 2, the DBP data is shown in Table 3, the non-dipper type subject's SBP data is shown in Table 4, and the DBP data is shown in Table 5. Note that no change was observed in the heart rate of the subject during the measurement.

<Measurement of disease risk reduction effect by diurnal blood pressure>
After the normal blood pressure Dipper and Non-dipper subjects selected above did not change their lifestyle and do not take antihypertensive drugs for 4 weeks, they woke up the fermented dairy products for the subjects for one month. Later I took 160ml every morning.
The day after the last intake, each subject was asked to wear a portable 24-hour automatic blood pressure monitor (model ES-H531, Terumo Corp.) on the upper arm, not the upper arm. Every minute, SBP, DBP and heart rate were measured for 24 hours.
Among these results, 0:00 to 7:00 was set as the blood pressure diurnal variation time zone in the risk of developing the disease from night to morning, and the SBP values before ingesting the fermented dairy products for subjects were compared. The results are shown in FIG. 1 for Dipper-type subjects and in FIG. 2 for non-dipper-type subjects.

From the results of FIGS. 1 and 2, no significant difference was observed in the Dipper group before and after the intake of the fermented dairy product for the subject in the SBP from 0:00 to 7:00, but a significant difference (P < 0.05, paired t-test). Accordingly, the fermented dairy product for test subject containing the hydrolyzate of animal milk protein according to the present invention is a healthy person having normal values of SBP and DBP defined in the present invention, and a specific non-dipper blood pressure daily It was found that systolic blood pressure from night to early morning can be effectively reduced for humans with fluctuations, and the risk of developing diseases due to blood pressure diurnal fluctuations that are likely to develop in the morning can be reduced. Note that no change was observed in the heart rate of the subject during the measurement.

Claims (8)

Upper arm blood pressure was measured every 30 minutes with a portable 24-hour automatic sphygmomanometer, and the rate of decrease in average systolic blood pressure from 8:00 to 5:00 was 10% with respect to average systolic blood pressure from 8:00 to 21:00 An agent that can be taken orally by a non-dipper type human being less than the risk of developing disease due to blood pressure diurnal variation,
A non-dipper type blood pressure diurnal risk-reducing agent containing hydrolyzate containing Ile Pro Pro and Val Pro Pro or a concentrate thereof obtained by hydrolyzing animal milk protein. In the non-dipper type human, the average of systolic blood pressure (SBP) measured by a portable 24-hour automatic sphygmomanometer every 30 minutes was less than 130 mmHg, and the average of diastolic blood pressure (DBP) was less than 80 mmHg. The alleviating agent according to claim 1. The hydrolyzate according to claim 1 or 2, wherein the hydrolyzate is hydrolyzed under the condition that the content of Val Pro Pro is 3 µg / ml or more and the content of Ile Pro Pro is 2 µg / ml or more. Relief agent. The mitigating agent according to any one of claims 1 to 3, wherein the hydrolyzate is a fermented material obtained by fermenting a raw material containing animal milk protein with lactic acid bacteria. The mitigating agent according to claim 4, wherein the lactic acid bacterium comprises Lactobacillus helveticus. 6. The alleviating agent according to claim 5, wherein the Lactobacillus helveticus is Lactobacillus helveticus having a deposit number of FERM BP-6060. The mitigating agent according to any one of claims 1 to 3, wherein the hydrolyzate is a degradation product obtained by enzymatic degradation of a raw material containing animal milk protein with an enzyme derived from Aspergillus. The mitigating agent according to claim 7, wherein the enzyme derived from Aspergillus is an enzyme derived from Aspergillus soyzae.

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