JP2020198791A - Sleep-improving composition, and food, drug and feed containing the composition - Google Patents

Abstract

To provide a sleep-improving composition capable of increasing non-REM sleep time and excellent in stability and safety, and food, food compositions, drug, drug compositions, and feed containing the sleep-improving composition.SOLUTION: The sleep-improving composition includes whey protein hydrolysate as an active ingredient; in particular, the whey protein hydrolysate is characterized in having a molecular weight distribution of a width of 10 kDa or less and a main peak molecular weight of 200 Da to 3 kDa, APL (average peptide length) of 2 to 8, a free amino acid content of 20% or less, and an antigenicity of 1/10,000 or less of that of β-lactoglobulin. There are also provided sleep-improving food, sleep-improving food compositions, sleep-improving drug, sleep-improving drug compositions, or sleep-improving feed containing the sleep-improving composition.SELECTED DRAWING: Figure 1

Description

The present invention relates to a sleep-improving composition that improves sleep and is excellent in stability and safety. The present invention also relates to foods, food compositions, pharmaceuticals, pharmaceutical compositions, and feeds containing the sleep-improving composition.

In recent years, changes in lifestyle have disturbed the rhythm of life, and the prevalence of sleep disorders such as insomnia has increased. According to a survey by the Ministry of Health, Labor and Welfare, one in five people has some sort of sleep problem (Non-Patent Document 1). It has been clarified that sleep disorders are closely related not only to the decrease in labor productivity due to drowsiness, but also to mental illnesses such as dementia and depression, and lifestyle-related diseases such as obesity and diabetes. The economic loss due to this lack of sleep is estimated to be about 3.5 trillion yen (Non-Patent Document 2).

For people with sleep disorders such as insomnia, the use of sleeping pills, which are medicines, may be considered. As the hypnotic, barbituric acid hypnotics, benzodiazepine hypnotics, and non-benzodiazepine hypnotics are mainly used. However, barbituric acid hypnotics have strong side effects such as dependence, and sleep with benzodiazepine hypnotics induces sleep showing an abnormal frequency range called benzodiazepine hypnotics, so it should not lead to physiological and good quality sleep. It has been known. In addition, it is known that both benzodiazepine and non-benzodiazepine hypnotics may cause side effects such as light-headedness, headache, dizziness, and symptoms such as not remembering the events leading up to falling asleep or the events during awakening. ing.

For this reason, the development of materials that can be taken on a daily basis instead of pharmaceuticals and that can induce physiological sleep is being actively carried out, and various natural ingredients and food ingredients have been proposed. ing. For example, there are a sleep disorder improving agent using glycine, which is an amino acid (Patent Document 1), and a sleep promoting composition using theanine (Patent Document 2).

Milk and dairy products have been eaten since ancient times, and it is said that ingesting warmed milk before falling asleep improves sleep, such as improving sleep. It has been reported that α-lactalbumin actually contained in whey has a sleep improving effect (Non-Patent Document 3). In addition, a patent has been reported that can treat anxiety, sleep disorders, and epilepsy caused by peptides derived from milk casein (Patent Document 3).

However, in order to obtain the sleep improving effect by taking the above-mentioned α-lactalbumin, it is necessary to take 40 g per day, which is difficult to take on a daily basis. In addition, the patent on milk casein-derived peptides only evaluates animal behavior for anxiety-like behavior by visual inspection or videography, and it is not clear or demonstrated whether the peptide can actually treat sleep disorders. The effects of this peptide are limited to anxiety.

Japanese Unexamined Patent Publication No. 2013-121961 International Publication WO 01/074352 Special Table 2011-524163

2014 National Health and Nutrition Survey Makoto Uchiyama, Japan Psychiatric Hospital Association Magazine 31 (11), 1163-1169, 2012-11 Markus, C.R. et. Al .: Am. J. Clin. Nutr., 81,1026-1033,2005

An object of the present invention is to provide a sleep improving composition which improves sleep and is excellent in stability and safety. Another object of the present invention is to provide a food, a food composition, a pharmaceutical product, a pharmaceutical composition, and a feed containing the sleep improving composition.

As a result of diligent studies to solve the above problems, the present inventors have found that whey protein hydrolyzate improves sleep, and its effect is significantly stronger than that of whey protein before hydrolysis.
That is, the present invention includes the following aspects.
(1) A composition for improving sleep containing a whey protein hydrolyzate as an active ingredient.
(2) The sleep improving composition is one or more selected from the group consisting of a non-REM sleep time increasing composition, a sleep time increasing composition, and a sleep induction promoting composition. , (1). The composition for improving sleep.
(3) The sleep improving composition according to any one of (1) and (2), wherein the decomposition rate of the whey protein hydrolyzate is 25% or more.
(4) The sleep-improving composition according to any one of (1) to (3), wherein the whey protein hydrolyzate has the following characteristics.
(A) The molecular weight distribution is 10 kDa or less, and the main peak is 200 Da to 3 kDa.
(B) APL (mean peptide chain length) is 2-8.
(C) The free amino acid content is 20% or less.
(D) The antigenicity is 1 / 10,000 or less of the antigenicity of β-lactoglobulin.
(5) The whey protein hydrolyzate enzymatically decomposes whey protein at pH 6 to 10 and 50 to 70 ° C. while heat-denaturing it with a heat-resistant protein hydrolase, and heats it to inactivate the enzyme. The composition for improving sleep according to any one of (1) to (4), which is obtained.
(6) The whey protein hydrolyzate enzymatically decomposes whey protein at pH 6 to 10, 20 to 55 ° C. using a protein hydrolyzate enzyme, raises the temperature to 50 to 70 ° C., and pH 6 to 10, 50. It is characterized in that it is obtained by enzymatically decomposing undegraded whey protein using a heat-resistant protein hydrolyzate at ~ 70 ° C while heat-denaturing it, and inactivating the enzyme by heating (1). To the composition for improving sleep according to any one of (4).
(7) A sleep-improving food, a sleep-improving food composition, a sleep-improving drug, a sleep-improving drug composition, which comprises the sleep-improving composition according to any one of (1) to (6). Things or feed for improving sleep.

The present invention provides a novel composition for improving sleep, and the term "sleep improvement" as used herein is one of an increase in non-REM sleep time, an increase in sleep time per day, or a reduction in sleep onset latency. It points to the above.

A comparison between the day before administration and the day of administration is shown for the non-REM sleep time of 12 hours after administration of whey protein hydrolyzate (left) or whey protein concentrate (right). A comparison between the day before administration and the day of administration is shown for the awakening time of 12 hours after administration of whey protein hydrolyzate (left) or whey protein concentrate (right). A comparison between the day before administration and the day of administration is shown for the non-REM sleep time of 1 hour after administration of whey protein hydrolyzate (left) or whey protein concentrate (right).

(Sleep improvement composition)
The sleep improving composition of the present application will be described.
The sleep-improving composition of the present application is a hydrolyzed whey protein.
More specifically
The decomposition rate of whey protein hydrolyzate is 25% or more,
The molecular weight distribution is 10 kDa or less, the main peak is 200 Da to 3 kDa, and
APL (mean peptide chain length) is 2-8,
Free amino acid content is 20% or less
Antigenicity is 1 / 10,000 or less of the antigenicity of β-lactoglobulin,
It has the feature.
The whey protein hydrolyzate used in the sleep-improving composition of the present application does not substantially contain full-length α-lactalbumin (based on the analysis results of the whey protein hydrolyzate by HPLC. No data is shown. .). More specifically, the total amount of α-lactalbumin contained in the whey protein hydrolyzate is 10% or less, preferably 5% or less, of the amount of α-lactalbumin contained in the whey protein before hydrolysis. 2% or less, 1% or less, 0.5% or less, 0.2% or less, or 0.1% or less.
The decomposition rate of the whey protein hydrolyzate of the sleep improving composition can be measured by the orthophthalaldehyde (OPA) method or the like, which measures by modifying a free transamination group. The molecular weight distribution of whey protein hydrolyzate can be measured by a method such as High performance size exclusion chromatography (HPSEC) method. The APL (average peptide chain length) can be measured by a method such as the TNBS (2,4,6-trinitrobenzenesulfonic acid) method. The free amino acid content can be measured by extracting free amino acids with 75% ethanol or the like and using an amino acid analyzer or the like. Antigenicity can be evaluated by the inhibition ELISA method.

(Intake of sleep improving composition)
Since the composition for improving sleep of the present invention contains a plurality of peptides as described above, it is possible to exhibit a sleep improving effect at an intake of 0.01 g / day or more due to the additive / synergistic effect of two or more peptides. On the other hand, since the sleep-improving composition of the present invention can be safely ingested with an antigenicity of 1 / 10,000 or less of β-lactoglobulin, the intake amount is 0.025 g / day or more depending on the administration subject and the form of food. 0.1g / day or more, 0.25g / day or more, 1g / day or more, or 2.5g / day or more, and 30g / day or less, 20g / day or less, 10g / day or less, 5g / day or less, 2.5g / day It can be less than or equal to 1 g / day or less.
The sleep-improving composition of the present invention can be added to foods, food compositions, pharmaceuticals, pharmaceutical compositions, and feeds as described below, and the amount of the sleep-improving composition added at that time is It may be adjusted appropriately so that the above-mentioned effective amount can be ingested.

(Evaluation method of sleep improving effect of sleep improving composition)
The sleep-improving effect of the sleep-improving composition of the present invention can be confirmed by the animal test on rats described in Examples.

(Manufacturing method of sleep improving composition)
The whey protein hydrolyzate, which is the active ingredient of the sleep improving composition of the present invention, can be obtained, for example, by the method reported in JP-A-4-112753. In this method, whey protein is adjusted to pH 6-10, 50-70 ° C, and heat-resistant protein hydrolase is added to it for enzymatic decomposition while heat denaturation, and this is heated to inactivate the enzyme. Be done. Before performing the above enzymatic decomposition, whey protein is enzymatically decomposed at pH 6 to 10 and 20 to 55 ° C. using a protein hydrolase, and the whey protein is immediately enzymatically decomposed under the above conditions without cooling to further increase the yield. Can be enhanced.

Further, the whey protein hydrolyzate prepared as described above is subjected to an ultrafiltration (UF) membrane having a molecular weight cut off of 1 kDa to 20 kDa, preferably 2 kDa to 10 kDa, and / or a molecular weight cut off from 100 Da to 500 Da, preferably 150 Da to. It is also possible to concentrate by a method selected from 300 Da nanofiltration (NF) membranes. By such a membrane treatment, the average molecular weight of the whey protein hydrolyzate is set to 300 to 500 Da, so that the bitterness can be further reduced and the transparency can be improved.

The whey protein in the present invention refers to whey prepared from the milk of mammals such as cows, goats, sheep and humans, its aggregates, powders, or purified proteins, and is used in an aqueous state when it is enzymatically reacted.

When preparing a whey protein hydrochloride by the method reported in JP-A-4-112753, the pH of the above solution is adjusted to 6 to 10, but the pH of whey protein is usually in this range, so it is exceptional. It is not necessary to adjust the pH, but if necessary, use an acid solution such as hydrochloric acid, citric acid and lactic acid or an alkaline solution such as caustic soda, calcium hydroxide and sodium phosphate to adjust the pH to 6-10. Although heating is carried out at 50 to 70 ° C., it is preferable to add the thermostable protein hydrolase before heating and carry out enzymatic decomposition rather than adding it at this temperature in terms of yield.

In addition, the optimum temperature of a general protease is 40 ° C. or lower, but the optimum temperature of a heat-resistant protein hydrolase is 45 ° C. or higher. Anything known as a heat-resistant protein hydrolase having an optimum temperature can be used without particular limitation. Examples of such heat-resistant protein hydrolases include papain, thermolysin, thermolysin S, protease S (trade name), proleather (trade name), samoase (trade name), alcalase (trade name), and protein A (trade name). Name) etc. can be exemplified. The thermostable protein hydrolase is preferably heated at 80 ° C. for 30 minutes to have a residual activity of about 10% or more. In addition, it is more effective to use a plurality of enzymes in combination than to use them alone. The reaction is preferably carried out for about 30 minutes to 10 hours.
Finally, the reaction solution is heated to inactivate the enzyme. The inactivation of the enzyme can be performed by heating the reaction solution at 100 ° C. or higher for 10 seconds or longer.

Then, the reaction solution is centrifuged to collect the supernatant, and the supernatant is dried to obtain a powder product. Since the degree of allergenization of the precipitate formed during centrifugation is smaller than that of the supernatant, it is preferable to remove it, but of course, the reaction solution may be dried and used as it is.

The sleep-improving composition of the present invention may be used as it is, but since it can be used together with other raw materials usually contained in foods, pharmaceuticals, and feeds, powders, granules, tablets, capsules, according to a conventional method, It can also be used as a drink. It can also be added to foods and drinks such as yogurt, milk drinks and wafers, and feed.

Examples and test examples are shown below, and the present invention will be described in detail, but these are merely examples, and the present invention is not limited thereto.

(Example 1)
Papain 50U / g, whey protein and Proleather (manufactured by Amano Enzyme) 150U / g, whey protein are added to 1L of a 10% whey protein aqueous solution, adjusted to pH 8, and the enzyme is denatured at 55 ° C for 6 hours. It was disassembled. The reaction solution was heated at 100 ° C. for 15 seconds or longer to inactivate the enzyme, centrifuged to collect the supernatant, and dried to obtain a whey protein hydrolyzate (Example product 1).
The molecular weight distribution of the obtained whey protein hydrolyzate was 10 kDa or less, the main peak was 1.3 kDa, the APL was 7.2, and the free amino acid content for all the constituents was 18.9%.
The decrease in antigenicity to β-lactoglobulin was measured by the inhibition ELISA method and found to be 1 / 10,000 or less, the decomposition rate was 28%, and the yield (the enzyme reaction solution was centrifuged and the supernatant was dried based on the dry weight of the charged amount. The weight ratio (%)) was 80.3%.
The whey protein hydrolyzate thus obtained can be used as it is as the sleep improving composition of the present invention.

(Test Example 1)
Whey protein hydrolyzate was used to evaluate sleep-improving effects.
Purchased an 8-week-old male Wistar rat (SLC Japan), fed a commercial solid feed CRF-1 (manufactured by Oriental Yeast Co., Ltd.) under a light-dark cycle every 12 hours, and freely ingested feed and water. I let you. Rats were operated on to attach electrodes for measuring brain waves and myoelectric potentials, and the test was conducted with a recovery period of 1 week or longer.
Whey protein degradation products were dissolved in water and orally administered at 2.5 g per 1 kg of rat body weight (corresponding to about 30 g per day when converted to humans using FDA guidelines), and whey protein concentrate for comparison. (Proteo content 80%) was dissolved in water and 2.5 g per 1 kg of rat body weight was orally administered. Administration was performed from 17:00 to 18:00 (within 1 hour after the lights were turned off), and compared with the sleep / wake time on the day before administration.
EEG and myoelectric potential were classified into sleep stages of arousal, non-REM sleep, and REM sleep using the sleep analysis software "Sleep sign" (Sleep sign ver2.0, manufactured by Kissei Comtech).
The awakening time and non-REM sleep time 12 hours after administration were integrated and compared with the awakening time and non-REM sleep time on the day before administration. In addition, Student's t-test test was performed, and it was judged that there was a significant difference when p <0.05.

As shown in FIGS. 1 and 2, it was shown that the administration of whey protein hydrolyzate significantly increased the non-REM sleep time 12 hours after the administration and decreased the wakefulness time as compared with the day before the administration ( The day before administration is expressed as 100%. The same shall apply hereinafter). Furthermore, as shown in FIG. 3, the administration of whey protein hydrolyzate significantly increased the amount of sleep for 1 hour immediately after administration, suggesting that it has a sleep-inducing effect. Since the whey protein hydrolyzate did not contain full-length alpha-lactalbumin (based on the results of HPLC analysis of the whey protein hydrolyzate, no data shown), the observed effect was alpha-lactalbumin. Is not the effect of.
On the other hand, in the whey protein concentrate used as a comparison target, the non-REM sleep time of 12 hours after administration, the awakening time, and the non-REM sleep time of 1 hour immediately after administration were not different from those on the day before administration.
Similar results to FIGS. 1-3 were obtained when different proteases (cysteine endopeptidase, metalloendopeptidase, serine endopeptidase, aspartic protease) than papain and proleather were used to hydrolyze whey protein. (No data shown).

(Example product 2) Capsules for sleep improvement After mixing the raw materials with the formulation shown in Table 1, granulated by a conventional method and filled into capsules to produce capsules for sleep improvement.

(Example product 3) Beverage for improving sleep
10 g of Example Product 1 was added to 700 g of deionized water, heated to 50 ° C., and stirred and mixed at 9,500 rpm for 30 minutes.
After adding the raw materials shown in Table 2 to the mixture, the mixture was filled in a 100 ml glass bottle, sterilized at 95 ° C. for 15 seconds, and then sealed to prepare 10 sleep-improving beverages (containing 100 ml).

(Example product 4) Feed for improving sleep for dogs
5 kg of Example Product 1 was added to 95 kg of deionized water, heated to 50 ° C., and then stirred and mixed at 3,600 rpm for 40 minutes to obtain a solution containing 5 kg / 100 kg of Example Product 1. The whole amount of the raw materials shown in Table 3 was mixed with 20 kg of this solution and sterilized at 120 ° C. for 4 minutes to prepare 100 kg of the feed for improving sleep for dogs of the present invention.

The present invention provides a composition containing a whey protein hydrolyzate and a food containing the composition, a food composition, a pharmaceutical product, a pharmaceutical composition, and a feed as a new sleep-improving composition. Sleep can be improved by ingesting the composition of the present invention or the like.

Claims (7)

A sleep-improving composition containing a whey protein hydrolyzate as an active ingredient. The claim is characterized in that the sleep improving composition is one or more selected from the group consisting of a non-REM sleep time increasing composition, a sleep time increasing composition, and a sleep induction promoting composition. The sleep improving composition according to 1. The sleep-improving composition according to any one of claims 1 or 2, wherein the decomposition rate of the whey protein hydrolyzate is 25% or more. The sleep-improving composition according to any one of claims 1 to 3, wherein the whey protein hydrolyzate has the following characteristics.
(A) The molecular weight distribution is 10 kDa or less, and the main peak is 200 Da to 3 kDa.
(B) APL (mean peptide chain length) is 2-8.
(C) The free amino acid content is 20% or less.
(D) The antigenicity is 1 / 10,000 or less of the antigenicity of β-lactoglobulin. The whey protein hydrolyzate is obtained by enzymatically decomposing whey protein at pH 6-10 and 50-70 ° C using a heat-resistant protein hydrolase while thermally denaturing it, and heating to inactivate the enzyme. The sleep-improving composition according to any one of claims 1 to 4, wherein the composition is characterized by the above. The whey protein hydrolyzate enzymatically decomposes whey protein at pH 6 to 10, 20 to 55 ° C. using a protein hydrolyzate enzyme, raises the temperature to 50 to 70 ° C., and pH 6 to 10, 50 to 70 ° C. The present invention is characterized in that it is obtained by enzymatically decomposing an undecomposed whey protein using a heat-resistant protein hydrolyzate while heat-denaturing it, and inactivating the enzyme by heating. The composition for improving sleep according to any one of 4. A sleep-improving food, a sleep-improving food composition, a sleep-improving drug, a sleep-improving pharmaceutical composition, or a sleep-improving pharmaceutical composition, which comprises the sleep-improving composition according to any one of claims 1 to 6. Feed for improving sleep.

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